Advances in Carbohydrate Chemistry, Volume 10

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ADVANCES XN CARBOHYDRATE CHEMISTRY VOLUME 10

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Carbohydrate Chemistry Editor MELVILLE L. WOLFROM Arsistant Editor R. STUART TIPSON Board of Advisors C. B. PURVES J. C. SOWDEN ROYL. WHISTLER

HERMANN 0. L. FISCHER

It. C. HOCKETT

W. W. PIGMAN

Board of Advisors for the British Isles J4. L. HIRST

STANLEY PEAT

MAURICESTACEY

Volume 10

1955

ACADEMIC PRESS INC., PUBLISHERS NEW YORK, N. Y.

Copyright, 1955, by ACADEMIC PRESS INC. 125 East 23rd Street New York 10, N. Y.

A12 Rights Reserved

No part of this book may be reproduced in any form by photostat, microfilm, or any other means without written permission from the publishers. Library of Congress Catalog Card Number: 45-11351

PRINTKD I N T H E UNITED STATES OF AMERICA

CONTRIBUTORS TO VOLUME 10 W. W. BINKLEY,Department of Chemistry, The Ohio State University, Columbus, Ohio*

G. P. ELLIS,Department of Chemistry, King's College, University of London, England A. B. FOSTER,Department of Chemistry, The University, Edgbaston, Birmingham, England

L. J. HAYNES,The University, Edinburgh, Scotland ,JOHN E. HODGE,Northern Utilization Research Branch, Agricultural Research Service, U . S . Department of Agriculture, Peoria, Illinois JOHN HONEYMAN, Department of Chemistry, King's College, University of London, England A. J. HUGGARD, Department Columbus, Ohio

0.f

Chemistry, The Ohio State University,

GEORGEG. MAHER,Research Laboratories, Clinton Foods Inc., Clinton, Iowa J. A. MILLS, Division of Biochemistry and General Nutrition, Commonwealth Scientific and Industrial Research Organization, Adelaide, Australia

F. H. NEWTH,The University, Cambridge, England W. J. POLGLASE, Department of Biochemistry, University of British Columbia, Vancouver, British Columbia, Canada 'Present address: The New York Sugar Trade Lahoratorg Inc., 113 Pearl Street, N e w York

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4 , N . 1'.

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PREFACE With this Volume, “Advances in Carbohydrate Chemistry” comes of age through the completion of a series of ten issues. Herein, modern conformational analysis is applied to the carbohydrate field by J. A. Mills (Adelaide), The ever-recurrent and always puzzling subject of nitrogen chemistry is elaborated in a discussion of the glycosylamines and their rearrangement products by G. P. Ellis with J. Honeyman (London) and by J. E. Hodge (Peoria). The preparation and reactivity of the useful glycosyl halides is presented by L. J. Haynes (Edinburgh) and F. H. Newth (Cambridge). W. W. Binkley (Columbus) summarizes the present status of column chromatographic technique as applied to the sugar group. Our series of chapters on the methyl ethers is augmented by G. G. Maher (Clinton, Iowa). Polysaccharide chemistry is represented by a chapter on the non-cellulosic components of wood from the pen of W. J. Polglase (Vancouver) and by one on the biochemically significant subject of heparin from A. B. Foster and A. J. Huggard (Birmingham and Columbus). These, together with an obituary of the late, esteemed Dr. E. G. V. Percival of Edinburgh, complete this Volume and are offered as a contribution to the summarizing of progress in the ever-growing subject of carbohydrate chemistry.

M. L. WOLFROM Columbus, Ohio

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CONTENTS CONTRIBUTORS TO VOLUME 1 0 . .. . . . . . . , , . . , . . . . . . . . , , . . . , . . . . . . . . . . . . . . . . . . . . .

v

PREFACE.. . . . . . . , . _ _, . . . . , , . . . . , , . . . . . . . . . . . , . . , , . . . . . , . . , , . . . . . . . . . . . . . . . . . vi EDMUNI) GEORGE VINCENT

!?ERrIVAL. . . . . . . . . .

. , . . , . . . . . . . . . . . . . . . . . . . . . . . . . . X ... lll

The Stereochemistry of Cyclic Derivatives of Carbohydrates BY J. A. MILLS,Department of Biochemistry and General Nutrition, Commonwealth Scientijic and Industrial Research Organization, Adelaide, Australia

I. Introduction . . . . . . . . . . . . . , . , . . , . _ _. ._ _ ,, . . . . . . . . . . . . . . . . . . . . . . _ _. . . . . . 2 11. Nomenclature and Methods of Illustration. . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3 111. Steric Requirements of Reactions. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6 IV. Fundamental Properties of Rings.. . . . . . , . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10 V. Cyclic Acetals . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 25 VI. Anhydro Compounds.. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 46 Column Chromatography of Sugars and Their Derivatives BY W. W. BINKLEY, Department of Chemistry, T h e Ohio Stale University, Columbus, Ohio

I. Introduction . . . . _ _, . . . . , . . . . . . . . . . . , . . . . . , . . , . . . . . . . . . . . . . . . . . . . . . . . . . 55 11. Adsorbents for the Column Chromatography of Sugars, Sugar Alcohols, 56 and Sugar Acids . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 111. Column Chromatography of Sugar Derivatives. . . . . . . . . . . . . . . . . , . . . . . . . 79 Glycosylamines BY G. P. ELLISAND JOHNHONEYMAN, Department of Chemisfry, King's College, University of London, England I. 11. 111. IV. V. VI. VII. VIII. IX.

Introduction ..... _ _. . . _ . . . . . . _ .. . . . . , , . . . . . . . . , . , . . . . . . . . . . . . . . . . . . . . Nomenclature . . . . . . . . . . . . . . . . . , . . . . . , . . . . _ .. _ _ _ . . . . _ .. _ . ._ _ _ . _ _. . .... Preparation . . . . , . . ,.. , . . . . . . . . . . . . , . . . . . . . . . . . . . . . , . . . , . . . . . . . . . . . . . . . . Physical Properties.. . . . . . . . . . , . . . . . . . . . , . , , . . . . . . , , , . . . . . . . . . . . . . . . . . . Structure. . . . _ .. , . . _ .. _ ., . . _ _ . _. .. . . . . . . . . . . . , . . . _ .. . . . . . _ . . . . . . . . _. . . Diglycosylamines . . , . . . . . . . . . . . . . . . . . . . . . . , . . . . . . . . . . . . . . . . . . . . . . . . . . . . Diitmide Derivatives of Aldoses.. . . , . . . . , , . . . . . . . . . . . . . . . . . . . . . . . . . . . . Uses of Glycosylamines.. . . . . . . . . . . . . . . , . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Tables of Properties of Glycosylamines.. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ,

95 96 97 101 102 120 121 124 125

The Amadori Rearrangement BY JOHN E. HODGE,Northern Utilization Research Branch, Agricultural Research Service, 7J. S . Department of Agriculture, Peoria, Illinois

I. Introduction... ............... . . ... ... ... . .......... . . . . . . . . . . . . . . . . . . . 169 ix

CONTENTS

X

I1 . Nomenclature . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 111. Scope of the Reaction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IV . Experimental Methods . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . V . Mechanism of the Reaction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VI . Physical Properties of Amadori Rearrangement Products . . . . . . . . . . . . . . . VII . Chemical Properties of Amadori Rearrangement Products . . . . . . . . . . . . . . . VIII . Proof of Structure of Amadori Rearrangement Products . . . . . . . . . . . . . . . . IX . Retrospect and the Future . . . . . . . ..................... .. X . Tables of Compounds . . . . . . . . . . . . ............................

172 173 175 178 185 187 199 201 203

The Glycosyl Halides and Their Derivatives BY 1, . J . HAYNES, l’he University, Edinburgh, Scotland A N D F . H . NEWTH, The l J n i v w s i t y , Cambridge, England I . Introduction., . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . I1 . Preparation of Glycosyl Halide Derivatives . . . . . . . . . . . . . . . . . . . . . . . . 111. Structure of Glycosyl Halides . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 229 IV . General . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 233 V . Reaction Mechanism and Effect of Structure on Reaction Rates . . . . . . . . 234 V I . Reactions of the Poly-0-acylglycosyl Halides . . . . . . . . . . . . . . . . . . . . . . . . . . . 239 VII . Tables of Properties of Some Glycosyl Halide Derivatives . . . . . . . . . . . . . . 246 The Methyl Ethers of the Aldopentoses and of Rhamnose and Fucose BY GEORGEG . MAHER,Research Laboratories, Clinton Foods I n c . , Clinton, Iowa Table I . ,. . . . . . . . . . . . . . . . . . . . . . . . . . ...................................... 257 Table I1 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 261 Table I11 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 262 Table IV . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 265 Table V . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 266 Table V I . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 266 Table V I I . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 268 Table VIII . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 269 The Methyl Ethers of D-Galactose BY GEORQEG . MAHER,Research Laboratories, Clinton Foods I n c . , Clinton, Iowa Table Table Table Table

I......................................................................

I1 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 111 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IV . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

273 274 277 279

Polysaccharides Associated with Wood Cellulose BY W . J . POLGLASE, Department of Biochemistry, University of British Columbia, Vancotcuer, British Columbia, Canada .................................... I . Introduction. . . . . . . . . . . . . . . . . I1 . Carbohydrate Constituents of Wood . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . I11 . Carbohydrates in Wood Cellulose Preparations . . . . . . . . . . . . . . . . . . . . . . . . . IV . Preparation and Composition of Wood Cellulose . . . . . . . . . . . . . . . . . . . . . . . . V . Fine Structure of Wood Cellulose and Associated Polysaccharides . . . . . . .

283 285 287 316 328

CONTENTS

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The Chemistry of Heparin BYA . B . FOSTER. Chemistry Department. T h e University. Birniingh.am. England A N D A . J . HUOGARD. Chemistry Department. T h e Ohio State University. Columbus. Ohio I . Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 336 I1. The Discovery of Heparin., . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 336 111. The IV . The V . The VI . The

Isolation and Purification of Heparin . . . . . . . . . . . . . . . . . . . . . . . . . . . 337 . . . . . . . . 348 Structure of Heparin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Anticoagulant Activity of Heparin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 359 Biosynthesis of Mucopolysaccharides . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 367

AUTHORINDEX .................................... SUBJECT

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EDMUND GEORGEVINCENTPERCIVAL 1907-1 951 The death of E . G. V. Percival a t the early age of 43, when he was a t the height of his powers as a scientist, was a severe blow to carbohydrate chemistry. He was born on 10th November, 1907, in the Midlands of England a t Hinckley in the County of Leicester, being the younger son of Albert Henry and Elizabeth Percival. As a boy, he attended the King Edward VII Grammar School in the neighboring town of Coalville. His school record showed great promise, and in 1925 he entered the University of Birmingham where he studied for the honours degree in chemistry. His undergraduate career was one of unusual brilliance, and it was a foregone conclusion that he would obtain a First Class Honours in Chemistry a t the end of the course in 1928. A research career was clearly indicated for him, but, before choosing a field in which to specialize, Percival wisely decided to gain as wide an experience as possible. I n the fall of 1928, he joined William Wardlaw’s group of workers in the Birmingham University laboratories and took part in work on the chemistry of metallic co-ordination compounds. A year later, he gained a Research Fellowship awarded by the Canadian Pulp and Paper Association, tenable a t McGill University, Montreal, and there, in Harold Hibbert’s laboratory, he investigated various physicochemical problems concerned with addition compounds of cellulose. Before returning to England, he made an extensive tour of research centers in Canada and the United States, gaining much experience which was of value to him in his later work, and making, as was the case wherever he went, many lasting friendships. The experience in Montreal had given him a deep interest in carbohydrate chemistry, and this became his main field of work 011 his return to Birmingham University in October, 1930, as senior research assistant to W. N. Haworth (later Sir Norman Haworth). This period was one of rapid and successful progress, and a t this stage he decided to seek a post which would enable him to pursue a career in the academic world. The opportunity came three years later (1933), when he was appointed to a lectureship in organic chemistry in the University of Edinburgh. Here, in the Department presided over by James P. Kendall, F.R.S., Percival settled happily, and during the next 18 years he built up a research school in carbohydrate chemistry which had an international reputation. He found conditions so congenial in Edinburgh that he made no serious attempt to gain xiii

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EDMUND GEORGE VINCENT PERCIVAL

promotion by moving elsewhere. In 1934, he married Ethel Elizabeth Kempson, herself a chemist, a graduate of the University of Birmingham where she had been a student in Sir Norman Haworth’s laboratories. After her arrival in Edinburgh, she took an active part in the development of Percival’s research activities, and assisted in the teaching of undergraduates. She and their two children survive him, and Mrs. Percival is COIItinuing, as a member of the University staff, some of the work initiated by her husband. Percival entered fully into the scientific and social life of both the University and the City of Edinburgh. He took a keen and sympathet,ic interest in student welfare and activities, and amongst his University duties was that of Director of Studies. He was elected t o membership in the Faculty of Science, and in 1948 was promoted to the status of Reader in the University. Shortly before his death he had been chosen to serve as a member of the Senatus Academicus. He gave his time and energy, with unselfish enthusiasm, to work for various scientific societies. For many years he had been interested in the work of the Pharmaceutical Societ,y and in British Chemical Abstracts. H e was concerned with work for the Institute of Brewing, he was one of the original pioneers in investigations on the chemistry of marine algal polysaccharides sponsored by the Scottish Seaweed Research Association, and he was an active member of this Association’s Chemical Advisory Committee. He served also on the local district committees of the Royal Institute of Chemistry and of the Society of Chemical Industry, and in addition acted for two years as Chairman of the Edinburgh and South East Scotland section of the Royal Institute. He was the local representative in Edinburgh of the Chemical Society (London), and at the time of his death was a member of the Society’s Council and of its Committee on Carbohydrate Nomenclature. His help was called for on all special occasions, and in 1948, when the Society of Chemical Industry held its annual general meeting in Edinburgh, and again in 1951, when he was Local Secretary for the Chemistry Section of the British Association for the Advancement of Science, his unselfish and unstinted efforts contributed in large part to the success of the meetings. It was characteristic of Percival that, to everything he undertook, he brought enthusiasm and intensity of purpose, whether it was in playing tennis or cricket, discussing painting (in which art he was deeply knowledgeable), walking on the Cheviot Hills, or in teaching and research. He had a brain which acted with lightning rapidity, and he had the power to carry through with accuracy (and with every attention to detail) a phenomenal amount of work in a very short time. He was a clear lecturer and expositor, and in consequence he was frequently invited to visit other universities and chemical societies. These visits, especially those which in-

EDMUND GEORGE VINCENT PERCIVAL

xv

volved traveling, he found highly congenial. He was a t his best as a teacher with advanced students of quick perception, who could follow without difficulty the logical but rapid development of his theme. Friendly, generous, sympathetic, and approachable, he was an inspiring leader for postgraduate research workers, who found him full of ideas and suggestions, and marveled a t his encyclopedic memory for details. All who had the privilege to work with him and enjoy his friendship will remember especially his kindliness, generosity, and unselfishness, his deep sense of loyalty, and his capacity to see and appreciate the humorous side of everyday happenings. Percival's outstanding ability was soon recognized, and awards came to him a t an early age-Fellowship of the Royal Institute of Chemistry in 1936, the D.Sc. degree of the University of Edinburgh in 1938, and, in 1941, election t o the Fellowship of the Royal Society of Edinburgh. Although Percival's main interest in chemistry lay in the carbohydrate field, his earliest research, carried out under William Wardlaw in Birmingham (1928-1929) was concerned with polynuclear co-ordination compounds of cobalt with amines.' During the following year, he held a Research Fellowship in Montreal, working with Hibbert on the constitution of sodacellulose, the hydrolysis of polysaccharides, and the absorption of aluminum ions on cellulose. It was demonstrated that a chemical compound is formed between the sodium hydroxide and the cellulose, containing about 15 % of alkali (or one molecule to each anhydro-D-glucose unit). Investigation of the rates of hydrolysis indicated that the lower rate for cellulose as compared wit,h that of starch is a reflection of the comparative rates of hydrolysis of cellobiose and maltose.2 I n the following three years (1930-33), working with Sir Norman Haworth, Percival began his studies in structural carbohydrate chemistry, and took part in the early work of the Birmingham School on the investigation of polysaccharides. Some of the most important results he obtained were concerned with the structures of starch and glycogen. Up to this time, strictly chemical evidence was lacking as to the presence of preformed maltose units in starch and glycogen, and concerning the nature of the ring present in the D-glucose residues. Percival developed a method of simultaneous deacetylation and methylation of the acetylated polysaccharides followed by acetolysis of the methyl derivatives. These yielded, amongst other products, a partially methylated maltose. The hexa-0methylbiose was oxidized to the bionic acid and subjected to further methylation. The resulting methyl octa-0-methylmaltobionate was then (1) E. G . V. Percival and W. Wardlaw, J . Chem. Soc., 1317, 1505, 2628 (1929). (2) A. C. Cuthbertson, H . Hibbert and E. G. V. Percival, J. Am. Chenz. SOC.,62, 3257, 3448 (1930); H. Hibbert and E. G . V. Percival, ibid., 62, 3995 (1930).

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hydrolyzed and the tetra-0-methyl-D-glucopyranose and tetra-0-methylD-glUCOniC 1,4-lactone were separated and identified, thus showing that the maltose structure must be present in starch and glycogen. Similar methods were applied to a di-0-methylxylan, in which the methoxyl groups were found to be attached to C2 and C3, showing that the linkage occurs a t C4 and that the D-xylose is present in the pyranose forrne3The methylation and hydrolysis technique was further applied to inulin, the methylated derivative of which gave rise to rather labile D-fructofuranose derivatives. In order to avoid unnecessary decomposition, the hydrolysis (after methylation) mas carried out using aqueous methanolic oxalic acid, and the yield of tetra-0-methyl-D-fructofuranose indicated the presence of one terminal nonreducing D-fructofuranose unit in 32 residues? One of Percival’s major contributions during his Birmingham period was the part he played, in collaboration with other members of the School, in determining the constitution of ascorbic acid (vitamin C). Oxidation studies were carried out which showed t,hat L-threonic acid is one of the products of oxidation, indicating ascorbic acid to be a derivative of L-gulose. The nature of the ring system was investigated by methylation studies, and vitamin C was proved to be the enolic form of 2-keto-~-gulonic1 , 4 - l a ~ t o n e . ~ Confirmation was obtained for this structure by the synthesis of vitamin C by the same group of workers. A t this point in his career, Percival moved to Edinburgh (1933), where he continued his work in the carbohydrate field. During the succeeding 18 years, his numerous publications covered a wide variety of subjects. From among the early work in Edinburgh may be ment,ioned the study of the compounds of alkali-metal hydroxides with sugars and polysaccharides, as a result of which he suggested that the alkali-suga,r complexes are definite compounds, and indicated how, by methylation, the points of attachment of the added hydroxides could be determined. Compounds of alkali-metal hydroxides with mono-, di-, and poly-saccharides were investigated.6 The methylation studies resulted in the production of partially methylated sugars whose identification required reference compounds, and Percival proceeded to study various mono-0-methylhexoses not previously charac(3) W. N. Haworth and E. G. V. Percival, J. Chem. SOC.,1342, 2850 (1931); 2277 (1932). (4) W. N. Hanorth, E. L. Hirst and E. G. V. Percival, J. Chew. SOC.,2384 (1932). ( 5 ) R. W. Herhert, E. L. Hirst, E. G. V . Percival, R. J. W. Reynolds and F. Smith, J. Chem. SOC.,1270 (1933) ; R . G. Ault,, D. I<. Baird, H . C. Carrington, W. N . Haworth, R. W . Herhert, E. L. Hirst, E. G. V. Percival, F. Smith and M. St,acey, ibid., 1419 (1933). 1160 (1934);648 (1935); E. G. V. Percival and (6) E. G. V. Percival, .J. Chem. SOC., G. G. Ritchie, i b i d . , 1765 (1936); W. J . Heddle and E. G. V. Percival, i b i d . , 1690 (1938).

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t,erized; for example, 4-O-methyl-~-glucoseand 2- and 6-O-methyl-~-galactose. He also devoted much attention to the structure of sugar osazones. By the methylation method, he decided that the previous assumption that osazones possess a straight-chain structure was erroneous, and that sugar osazones could exist in a ring form. I n the case of D-arabinohexose phenylosazone (“glucosazone”), the ring was shown to be of the 2 , 6 type. I n the course of these studies, the hitherto-unknown 5-0-methylD-arabino-hexose phenylosazone was characterized for the first time by a single methylation of D-arabino-hexose phenylosazone, and the work was also extended t o include the structures of several new anhydrohexose osazones of the Diels type.’ An entirely different approach to the structure of methylated and free sugars was made by the application of Boeseken’s method, involving measurements of the conductivity of sugars in boric acid solution. I n this way, the respect,ive configurations assigned to the hydroxyl groups on C l and C2 in a- and P-D-glucose were confirmed, the conductivity being considerably influenced by the cis or trans arrangement of the hydroxyl groups. The method was extended and applied to such methylated sugars as 3,4,6tri-0-methyl-a-D-mannose, where the hydroxyl groups on C1 and C2 were shown to have the trans arrangement.8 The main portion of Percival’s work in the years following 1936 was concerned with the structure of polysaccharides; in particular, of plant mucilages and seaweed polysaccharides. At this time, the polysaccharides of marine algae had been but little investigated; Percival was among the first in this field and became a recognized authority on the subject. One of the earliest studies he embarked on was that of the structure of agar, which presented a.n extremely difficult and complex problem. The results of the preliminary work on acetylation and methylation indicated that the basic structure consists of o-galactose units linked directly or indirectly through C l and C3,as shown by the isolation of the hitherto-unknown 2,4,6-tri0-methyl-a-D-galactose and of an unknown portion giving strong ketose reactions. Later work proved the presence also of 3,6-anhydro-L-galactose, but the complexity of the molecule precluded t,he solution of the detailed structure atj that t,ime. This study also entailed the synthesis of several anhydrogalactose derivatives used in the invest,igat>ionas reference compounds.$ At about the same time, work was commenced on other algal (7) Elizabeth E. Pereival and E. G. V. Percival, J . Chem. SOC.,1398 (1935); 1320 (1937); 750 (1941); F:. G. V. Percival, ibid., 1770 (1936); 1384 (1938); 783 (1945); J. R. Muir and E. ‘2. V. Percival, ibitl., 1479 (1940); W. J. Heddle arid E. G. V. Percival, ibid., 1511 (1940). (8) H. T . Macpherson and E. G . V. Percival, J . Chem. SOC.,1920 (1937). (9) €3. G. V. Percival and J. C. Somerville, J. Che,m. SOC., 1615 (1937); I. A. Forbes and E. G. V. Percival, ibid., 1844 (1939); T. L. Cottrell and E. G. V. Percival, ibid., 749 (1942).

xviii

EDMUND GEORGE VINCENT PERCIVAL

polysaccharides, notably those forming the mucilaginous extracts of Chondrus crispus and Gigartina slellata, collectively known as Irish moss. The polysaccharides from the two algae are essentially similar, in that both contain a considerable number of ester sulfate groups and consist of galactopyranose residues linked through C1 and C3, a portion of the galactose being in the L-form in the Chondrus polysaccharide. The complete elucidation of the structure proved difficult owing to the presence of the sulfate groups which interfere with methylation.1° In view of these difficulties and of the need for more knowledge of carbohydrate sulfuric esters, Percival then investigated the sulfates of D-glucose and D-galactose, their rates of hydrolysis with reference to the position of the sulfate group in the molecule, and the formation of 3,6- and 5,6-anhydro sugars on alkaline hydrolysis.]' With the collaboration of the newly-formed Scottish Seaweed Research Association, Percival commenced a further series of investigations in the marine algal field,lla the earlier work being concerned with the development of specific methods of analysis for the various constituents present in seaveed. Procedures were worked out for the estimation of D-mannitol, alginic acid, laminarin, and fucoidin.12 At the same time, structural studies were instituted on the last three polysaccharides.*3 Laminarinlla was shown to consist essent>iallyof (3 1) linked P-D-glucose units, and to occur in two forms, a soluble and an insoluble, depending on the species of seaweed used for extraction. I n fucoidin,lla the principal sugar is L-fucose, along with sulfate groups, and a branched structure was proposed in which 2 + 1a-L links predominate. Among other seaweed polysaccharides, the xylan from Rhodymenia palmata was found to be of unusual interest in that it was shown to contain both (3 --f 1) and (4--+ 1) linked D-XylOSe residues as part of the main structure of the polysaccharide. Percival had had previous experience with xylans while investigating the structures of the mucilages

-

(10) J. Buchanan, Elizabeth E. Percival and E. G. V. Percival, J . Chem. SOC., 51 (1943); E. T. Dewar and E. G. V. Percival, ibid., 1622 (1947); R . Johnston and E. G. V. Percival, ibid., 1994 (1950). (11) Elizabeth E. Percival and E. G. V . Percival, J . Chem. Soc., 1585 (1938); 874 (1945); E. G. V. Percival and T. H. Soutar, ibid., 1475 (1940); R. B. Duff and E. G. V. Percival, ibid., 830 (1941); 1675 (1947); E. G. V. Percival, ibid., 119 (1945); R. B. Duff, J . Chem. SOC.,1597 (1949). (Ila) See also, T. Mori, Advances in Carbohydrate Chem., 8, 315 (1953). (12) M. Christine Cameron, E. G. V. Percival and A. G. Ross, J . SOC.Chem. Znd. (London), 67, 161 (1948); E. G. V. Percival and A . G. Ross, ibid., 67, 420 (1948); W. A. P. Black, W. J . Cornhill, E. T. Dewar, E G. V. Percival and A. G . Ross, ibid., 69, 317 (1950). (13) E. G. V. Pereivsl a n d A. G . Ross, .I. Cl,eni. Sot-., 717 (1950); 720 (1951); J. Conchie and €2. G. V. Percival, itlid., 827 (1950) ; J . J. Connell, E. L. Hirst and E. G. V . Percival, ibid., 3494 (1950).

EDMUND GEORGE VINCENT PERCIVAL

xix

from t,he seeds of various meinhers of t'he plantain family (Plantago lanccolata, Plantago arenaria, and Plantago ov,uta). All t'hese sylans are complex in st,ructure and contain, in addit>ion t o D-xylose, ot,her pentoses, hexoses, and uronic acid residues; for example, the mucilage from Plantago arenaria contains D-xylose, L-arabinose, u-galactose, and wgalacturonic acid. The mode of' attachment of the uronic acid is not clear, and the struct,ure of the main D-xylopyranose chain includes various types of linkage and is highly branched. I n the mucilage from Plantago lanceolnta, for example, the following types of D-xylose residue were foundL4to occur as building unit,s: D-xylp 1, 3 D-xylp 1, 4 D-xylp 1, :D-xylp 1, iD-xylp 1, and :D-xylp ;. Further studies were carried out on the xylans from esparto and from pear cell-wall. I n the former, the xylan was shown to be free from L-arabinose and t o consist of a singly-branched molecule, the main chain containing about 75 D-xylopyranose units linked through the 4 position, the branching point occurring on C3. For the pear cell-wall polysaccharide, the general structure was found similar, but with the modification that t3he terminal residue consists of a D-glucuronic, acid unit and the main chain is rather longer (ca. 115 D-xylopyranose Percival also applied himself to the study of t.he polysaccharides from lichens, as, for example, tjhe complex product of alkaline extraction of Iceland moss (Cetraria islandica). As a result of this work, he concluded tJhat t,he polysaccharide consists of p-D-glucose residues united by various linkages: 1,2, 1,3, 1,4, and 1 , G , and includes also terminal D-galacto- and D-gluco-pyranose end groups. It was not possible to decide whether these linkages all occur in one polysaccharide.16 I n the later stages of his career, Percival published papers dealing with barley starch1? and wood starches,L8the former being found to be, in the main respects, similar to other cereal starches. Fructans also attracted his attention, and those from couch grass and perennial rye grass were particularly investigated; it was shown that the fructan from couch grass has both 2 , l and 2,G linkages in the molecule, whereas that from rye grass is essentJially a 2,G-linked, straight-chain p o l y s a ~ c h a r i d eFinally, .~~ mention may be made of the inulin from dahlia tubers, in which the presence of (14) J. Mullan and E. G. V . Percival, J . Chem. Soe., 1501 (1940); W . A. G. Nelson and E. G. V. Percival, i b i d . , 58 (1942); R. A. Laidlaw and E. G. V. Percival, ibid., 1600 (1949); 528 (1950); E. G. V. Percival and I. C. Willox, i b i J . , 1608 (1949). (15) S. K. Chanda, E. L. Hirst and E. G. V . Percival, J . Chem. SOC.,1240 (1951). (16) H. Granichstadten and E. G. V . Percival, J . Chem. SOC.,54 (1943). (17) I. C. MacWilliam and E. G. V . Percival, J . Chem. SOC.,2259 (1951). (18) W. G. Campbell, J. L. Frahn, E. L. Hirst, D. F . Pnckman and E. G . V. Perc i v d , J . Chem. SOC.,3489 (1951). (19) P. C . Arni and E. G. V . Percival, J . Chem. SOC.,1822 (1951).

xx

EDMUND GEORGE VINCENT PERCIVAL

u-glucose as an integral part of the molecule was demonstrated and possible structures were suggestedz0in which one n-glucose residue occurred as aii end group, linked to D-fructose by a sucrose type of link, whilst the second u-glucose residue was in the middle of the chain, linked through C1 and C3. Oiily a few days before his death, Percival took a prorninerit part in a conference 011 grass, sponsored by the Nutrition Society, a t which he read a p a p e P 011 the “Carbohydrate Coiistituerits of Herbage.” In this paper, he emphasized the importance of the fructaris as reserve carbohydrates in the metabolism of the plant, arid expressed the opinion that they would be found to play a considerable part, in the chemistry of the preservation of grass in the form of silage. E. L. HIRST A. G. Ross (20) E. I,. Hirst, D I. McGilvrag and E. G. V. Percival, J . Cheni. Soc., 1297 (1950). (21) E. G. V. Percivnl, Brit. J . Nutrztion, 6, 104 (1952).

THE STEREOCHEMISTRY OF CYCLIC DERIVATIVES OF CARBOHYDRATES* BY J . A . MILLS Division of Biochemistry and General Nutrition. Commonwealth Scientific and Industrial Research Organization. Adelaide. Australia

CONTENTS 1. 1ntroduct)ion. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . I1. Nomenclature and Methods of Illustration . . . . . . . . . . . . . . . . . . . . . . . . . . 1. Definitions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 . Structural Formulas . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 111 . Steric Requirements of Reactions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1. Reversible and Irreversible Reactions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 . Reactions with Specific Steric Requirements . . . . . . . . . . . . . . . . . . . . . I V . Fundamental Properties of Rings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 . Conformations . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 . Monocyclic Compounds . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . a . Five-membered Rings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b . Sis-membered Rings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c . Other Rings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .3. Structures Containing Fused Rings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . a . Two Five-membered Rings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b . Two Sis-membered Rings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c . One Five-membered and One Six-membered Ring . . . . . . . . . . . . . . . . . . . . d . Other Rings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4 . Bridged Rings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5 . The Course of Irreversible Ring Closures . . . . . . . . . . . . . . . . . . . . . . . . . . V . Cyclic Acetals . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1. General Remarks . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 . Monocyclic Acetals . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . a . Five-membered Rings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b . Six-membered Rings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3 . Acetals Containing Fused Rings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . a . Fused Five-membered Rings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b . Fused Six-membered Rings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . V I . Anhydro Compounds . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1. The 1,4:3,6-Dianhydrohesitols. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 . 1 , 6-Anhydro Derivatives of Pyranose Sugars . . . . . . . . . . . . . . . . . . . . . 3 . The Scission of Epoxide Rings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ~~~~~

~

~

~

*This review is based on literature available t o the author up t o July. 1954.

2 3 3 4 6 6 8 10 . 10 12 12 14 17 17 17 18 20 20 21 23 25 25 26 26 28 31 31

36 46 46 40 51

2

J. A . MILLS

I . INTRODUCTION The chief aim of this article is to make a comparison between carbohydrates and analogous alicyclic compounds, with emphasis on the stereochemistry of the ring systems involved. In alicyclic chemistry, notable additions have recently been made to our knowledge of the stereochemistry This new information of saturated rings, particularly six-membered already has had a considerable impact on carbohydrate chemistry, but has 6, The wider application attempted been restricted t o a few here will, it is hoped, assist in the coordination of considerable parts of the vohminous and diverse information available about carbohydrates, and may possibly lead to further development of stereochemical theory. I t is of interest to recall the development of the chemistry of terpenoids and steroids. Initially, these compounds were studied as problems in structural chemistry, but it was later realized14 that terpenoids of established structure provided first-class material for the study of the mechanisms of reactions, their availability in optically active forms being a notable advantage. Stereochemical theory has also greatly benefited in recent years from studies on steroids, largely because of the wider range of structural types available through the complexity of steroid molecules. Carbohydrates also possess the desirable features of optical activity, crystallinity, and availability, and display a diversity in structural types and reactions that cannot be matched in other fields. The writer, whose interests have lain mainly in the alicyclic field, has been greatly impressed by the scope that s t

(I) C. W. Beckett, K. S. Pitzer and R . Spitser, J . Am. Chem. Soc., 69,2488 (1947). (2) 0. Hassel and B. Ottar, dcta Chenr. Scand., 1, 929 (1947). (3) 0. Hassel, Research (London), 3, 504 (1950). (4) D. H. R. Barton, Experientia, 6, 316 (1950). (5) S. J. Angyal and J. A. Mills, Revs. Pure and A p p l . Chem. (Australia), 2, 185 (1952). (6) D. H. R. Barton, J. Cheni. Soc., 1027 (1953). (7) 0. Hassel, Quart. Revs. (London), 7, 221 (1953). (8) H. C. Brown, J. H. Brewster and H. Shechter, J . A m . Chern. Soc., 76, 467 (1954). (9) Further discussions are given by (a) R. B. Turner, in “Natural Products Related to Phenanthrene,” L. F. Fieser and Mary Fieser, eds., Reinhold Publishing Corp., New York, N. Y., 3rd Edition, 1949, p. 620; (b) V. Prelog, J . Chem. Soc., 420 (1950); (c) W. Klyne, in “Progress in Stereochemistry,” W. Klyne, ed., Academic Press Inc., New York, N. Y., Butterworths Publications Ltd., London, 1954, p. 36. (10) R . E. Reeves, J . A m . Chem. SOC.,72, 1499 (1950); and preceding papers in the series. (11) R. E. Reeves, Advances in Carbohydrate Chem., 6, 107 (1951). (12) R. J. Dimler, Advances in Carbohydrate Chem., 7, 37 (1952). (13) R. C. Cookson, Chemistry & Industry, 223 (1954). (14) See, for example, W. Huckel, Die Chemie, 66, 227 (1942).

CYCLIC DERIVATIVES OF CARBOHYDRATES

3

carbohydrate chemistry offers for elaborating and testing stereochemical hypotheses. Important contributions have, of cvursc, already come from this source, but usually as by-products of the development of carbohydrate chemistry, rather than through the deliberate choice of carbohydrate compounds as models for experimental study. To permit full advantage to be taken of the opportunities offered, a more detailed knowledge of the stereochemistry of cyclic carbohydrates is needed. The simple pyrariose and furanose sugars pose formidable problems of interactions between neighboring groups, but this difficulty largely disappears in their bicyclic and tricyclic derivatives. Accordingly, attentjion is confined mainly to compounds containing several rings, about which much information has been collected in recent reviews on cyclic acetalsI6 and anhydro I7 Research on these topics may seem to have lost some of its compounds,16~ initial impetus, but it will be seen that they still offer scope for further work of fundamental importance. Some of the conformational assignments are necessarily tentative a t present, and the opportunity is taken to point out directions in which more decisive evidence might be found. 11. NOMENCLATURE AND METHODS OF ILLUSTRATION 1 . Definitions

The convenient system of nomenclature for cyclic acetals introduced by Barker and Bourne15 will be used here. Rings formed by the engagement of hydroxyl groups attached to carbon atoms that are adjacent, separated by one atom, or separated by two atoms, are called a-,p-, or y-rings, respectively, and, if both hydroxyl groups are secondary, the designation C for two hydroxyl groups on t\he same side of the Fischer projection of the acyclic polyalcohol, or T for two groups on opposite sides, is added. Those parts of a polyol carbon chain not incorporated into an acetal ring, which appear as substituents in the ring, will be called residues. 2,4-O-Methylene-~-glucitolcontains a pC-ring, with the residues -CH20H and -CHOH-CH20H as substituents. The term fused rings wiH-bd used to denote the type of union found in decalin, where two rings have two atoms in common; the two atoms comprise a ring juncticn and are situated a t angular positions. Bridged rings are rings in which non-adjacent ring-atoms are joined by a bridge containing contains two one or more atoms. Methyl 3,6-anhydro-a-~-glucofuranoside (15) S.A. Barker and E. J. Bourne, Advances in Carbohydrate Chem., 7,137 (1952). (16) S.Peat, Advances in Carbohydrate Chem., 2, 37 (1946). (17) L.F.Wiggins, Advances in Carbohydrate Chem., 6, 191 (1950). (18) S. A. Barker and E. J. Bourne, J . Chem. SOC.,905 (1952).

4

J. A. MILLS

fused five-membered rings, whereas methyl 3 ,6-anhydro-a-n-glucopyranoside contains a bridged riug, and may be regarded as a six-membered ring bridged by two atoms or as a seven-membered ring bridged by one atom. 2. Structural Formulas

There is no settled convention for the representation of polyacetals and other complex cyclic derivatives of carbohydrate^.^^ To facilitate the comparison of polycyclic derivatives of carbohydrates with alicyclic compounds, the carbohydrate derivatives will be drawn in perspective, with single rings and fused rings placed in the plane of the paper, and with darkened or broken lines used to show the orientation of substituents and as is cusbridges above or below the plane of the rings, tomary for terpenes arid steroids. The convenience of the proposed usage is seen on comparing the Fischer projection (I) and the new representation (11) of 1,2:3,5-di-O-isopropylidene-a-D-glucofuranose; I1 shows clearly the nature of the ring-junctions and the relative positions of substituents, which are the factors controHing the stability and reactivity of the compound. The steroid type of formula is suitable for showing the configurations at new asymmetric centers in cyclic acetals, if these are known. Hydrogen atoms in the angular positions

I

CH,OH I

I1

111

a t ring-junctions must always be shown, but those a t points of substitution (as a t C5 in 11) may be omitted. The diagrams are more suitable for setting in type if drawn with bonds at angles of 45” and 90” only, but the true shape of the molecules is approached more closely by the representation 111. Fischer projections are still essential for showing genetic relations between carbohydrates, but are definitely unsuitable for precise stereochemical deductions; and reliance on them may have been responsible for some (19) W. W. Pigman and R. M. Goepp, J r . , “Chemistry of the Carbohydrates,” Academic Press Inc., New York, N. Y., 1948, p. 53. (20) J. A. Mills, Ph.D. Thesis, Cambridge, Engl., 1953. (21) J. A. Mills, Chemistry & Industry, 633 (1954).

CYCLIC DERIVATIVES OF CARBOHYDRATES

5

errors found in the literature. Formulas similar to I1 have already been used and for derivatives of D-glucofuranuronofor 1 4: 3 6-dianhydrohe~itols'~ r-lactone,22and the writer believes that they should be regularly used for depicting complex carbohydrate derivatives containing fused or bridged rings. Recourse to accurate models is necessary in studying intricate stereochemical problems, but with practice much information may be read from properly drawn diagrams of the type of 11. Because the plane of the paper is the plane of reference, the new formulas may be rotated in the plane of the paper without alteration of their significance, although this may not be true of all perspective formulas.23 Haworth perspective formulas24 are suitable for single pyranose and furanose rings and for some simple bridged structures, but when it is necessary to show conformations of six-membered rings, the non-planar "chair)' and "boat" forms will be drawn in perspective. A method has been devisedz5 for drawing chair rings with lines at angles of 45" and 90" only, suitable for setting in type; it is, however, not readily applicable to some bicyclic structures with cis ring-junctions. Two veiy useful rules for comparing Fischer projections with the new formulas may readily be deduced. (1) In a single ring, of any kind or size, resulting from closure a t two secondary hydroxyl groups on the same side. of a Fischer projection, the two residues will be in cis arrangement, whereas two hydroxyl groups on opposite sides of a Fischer projection will lead to a trans arrangement of residues. ( 2 ) When two adjacent carbon atoms of a Fischer projection become the ring-junction of a pair of fused rings, the ring-junction will be trans if the hydroxyl groups attached to the two carbon atoms are on the same side of the Fischer projection, but cis if the hydroxyl groups are on opposite sides. )

)

As an example, formula V will be transformed to a Fischer projection. Inspection

CH,OH

IV (22) (23) (24) (25)

1

V

K.-C. Tsou and A. M. Seligman, J. A m . Chem. Soc., 74, 5605 (1952). C . S . Hudson, Advances i n Carbohydrate Chent., 3, 1 (1948). H. D. K. Drew and W. N. Hnworth, .I. Chem. SOC.,2303 (1926). R . S. Cahn, J . Chem. Soc., 695, footnote (1952); 3520 (1951).

6

J. A . MILLS

of the steric arrangement a t C2 in V shows that i t is derived from a hydroxyl group on the left of a Fischer projection. Considering ring B only, it is seen t h a t the substituents C5 (in ring A) and C1 are trans; therefore the 2,4-ring is a OT-ring, and the hydroxyl group a t C4 is on the right of the Fischer projection. The ring-junction is cis, so the hydroxyl groups a t C3 and C4 are on opposite sides of the projection. By inspection, or by considering the substituents in ring A , the configuration a t C5 is (IV). Similar determined. Formula V represents 2,4:3,5-di-O-methylene-~-mannitol reasoning may be used t o derive steroid-type formulas from Fischer projections.

To assist comparisons between diagrams drawn according to different conventions, the ring-oxygen atom will, as far as is practicable, be placed at the top in furanose rings, and at the top right-hand corner in pyranose rings. The important class of bicyclic acetals represented by V will always be depicted with the orientation of oxygen atoms shown in V.

111. STERIC REQUIREMENTS OF REACTIONS I . Reversible and Irreversible Rbactions Carbohydrate chemistry provides many examples of reactions that could proceed by alternative pathways, to afford isomeric products that differ in configuration or structure. The effect of the configuration of the reactants on the course of such reactions may depend very largely on whether the reactions are reversible or irreversible. If all pathways are irreversible, the composition of the products depends on the relative speed along the pathways; it is kinetically controlled. These speeds are determined by entropy changes and by the stereochemistry of the traiisition states26concerned; the latter is often related to the stereochemistry of the reactants in a complex way, and has still to be elucidated for many reactions. On the other hand, if each pathway of reaction is reversible, and i f a true equilibrium is established, the composition of the products is quite independent of the mechanism of reaction, being determined solely by the relative thermodynamic stabilities of the constituents, even though the speed of the individual reartions may be very different. A study of such reversible reactions is therefore suitable for assessing the relative stabilities of isomeric compounds. Carbohydrate chemistry is especially rich in examples of ring structures formed by reversible reactions, such as g l y c ~ s i d e sand ~ ~ cyclic acetals,28 and provides a more convenient field for examining the factors governing the stability of rings than does alicyclic chemistry, in which ring closures are usually irreversible. Furthermore, the prominent reversible reactions (26)For reviews and references, see (a) E. D. Hughes, Quart. Revs. (London), 2, 107 (1948); (b) C. R. Ingold, “Structure and hfechanism in Organic Chemistry,” Cornell University Press, Ithsca, N . Y , 1953,pp. 40-49. (27)1’. A. Levene, A. I,. R:LJmond and It. T. Dillon, J . B i d . (‘hem ., 96, 699 (1932). (28) R. M. Hann and C.S. Hudson, J. Ant. C’hem. Soc., 66, 1909 (1944).

CYCLIC DERIVATIVES OF CARBOHYDRATES

7

of carbohydrates are nearly all catalyzed by acids, whereas the equilibria most studied in alicyclic chemistry, such as epimerizatioii of secondary hydroxyl groups4~ 2 9 , ao and of ring 29 are base-catalyzed. Epimerizations a t a positions in aldonic acids3' and 2 , 4 :3 ,&di-O-methylenehexaric acids5" are well established, but base-catalyzed epimerizatioii of secondary hydroxyl groups in the rings of g!ycosides and anhydro derivatives does not seem to have been investigated. Irreversible reactions of carbohydrates include the formation of glycosides and 1-deoxy-1-thioglycosidesfrom acylglycosyl halides33,34 and thioa c e t a l ~ , respectively, ~~-~~ esterifications by acyl and sulfonyl halides or anhydrides in ~ y r i d i n e ,40~displacements ~ of sulforiyloxy groups by various reagents,3Yand most examples of formation and scission of anhydro rings.l6V 17939 It is more difficult to define the influence of stereo effects on the course of these reactions; and some of the results obtained await explanation. I n using the results of reversible reactions in stereochemical analysis, a decision is often difficult as to whether a true equilibrium had in fact been established in experiments not specifically designed for quantitative studies. Recent syntheses of cyclic acetals are examples. When reducing sugars are condensed with acetone or acetaldehyde in the preseiice of strong acids, it is possible that decompositions caused by the catalyst will proceed more rapidly than the establishment of an equilibrium. This risk is less in reactions of alditols with benzaldehyde or formaldehyde, but again a true equilibrium may not result, because of the crystallization of insoluble acetals, or through interruption of the experiment before the rather difficultly hydrolyzable methylene acetals have come to equilibrium. The processing of reaction mixtures is often very incomplete. These uncertainties have been 1

(29) W. Huckel, Ahn., 633, 1 (1938). (30) W. von E. Doering and T. C. Aschner, J. Am. Chem. SOC.,71, 838 (1949), and references cited there. (31) E. Fischer, Ber., 23,799 (1890); W. N . Hnworth and C. W. Long, J . Chem. SOC., 345 (1929). (32) W. N. Haworth, W. G. M. Jones, M. Stncey and L. F. Wiggins, J . Chem. SOC., 61 (1944). (33) E. Pacsu, Advances in Carbohydrate Chem., 1, 77 (1945). (34) W. L. Evans, D. D. Reynolds and E. A. Talley, Advances in Carbohydrate Chem., 6, 27 (1951). (35) J. W. Green and E. Pacsu, J . Am. Cheni. SOC.,69, 1205, 2569 (1937). (36) E. Pacsu, J . Am. Chem. SOC.,61, 1671 (1939). (37) M. L. Wolfrom, D. I. Weisblat and A. R. Hanze, J . Am. Chem. Soc., 66, 2065 (1944). (38) A. L. Raymond, Advances in Carbohydrate Chem., 1, 129 (1945). (39) R. S. Tipson, Advances i n Carbohydrate Chem., 8, 107 (1953). (40) J. M. Sugihara, Advances in Carbohydrate Cheni.,8, 1 (1953).

8

J. A . MILLS

kept in mind when using data on cyclic acetals for the generalizations of Section V, and most weight has been assigned to the results of the experiments that seem to have more nearly approached the ideals of a true equilibrium and complete subsequent processing. 2. Reactions with Specific Steric Requirements

That the bimolecular displacement reaction (SN2) requires a collinear arrangement of three atomic centers is well known. Although few kinetic studies have been made, SN2reactions are probably fairly common in carbohydrate chemistry, especially intramolecular reactions leading to the establishment of anhydro rings. If the required steric arrangement is not attainable, the reaction may be entirely prohibited.16 The closure and scission of ethylene oxide rings is a special case of the intramolecular SN2reaction,16 and would seem to call for a tranx =-Cc,.- sition state with a specific arrangement (A) of four atomic ,. \o,.,, centers.l3. l 6 It is true that a trans arrangement of groups16 is necessary for the reaction, but this will not be sufficient A for ensuring reaction if molecular rigidity prevents the attainment of the transition state, as is shown by the following example. The reaction of 5,6-anhydro-l, 2-O-isopropylidene-cr-~-glucofuranose (VI) with bases is a fruitful source16r41 of D-glucose derivatives having a substituent a t C6. However, even when the opening of the anhydro ring of VI, by external attack of an anion, proceeds with difficulty, there is apparently little tendency for an intramolecular displacement by the free hydroxyl group a t C3; there is no mention4I of the formation of 3,6-a11hydro-1 ,2-O-isopropy~idene-cu-~-g~ucofuranose (VII), although this42 is a well-defined solid, and would have been readily recognized. On the other (VIII) reacts hand, 1,2-O-isopropylidene-5,6-di-O-tosyl-cr-D-glucofuranose

'

VI

VII

VIII

very readily with base, giving42the 5-0-tosyl derivative of VII. Scission of ethylene oxide rings through intramolecular attack by adjacent hydroxyl groups has often been observed,I6 and the failure of such a n attack to occur with VI must be ascribed to steric factors. Approach of C6 to the hydroxyl group a t C3 may occur with about equal ease for VI and VIII, but whereas (41) H. Ohle and K. Tessmar, Be?.., 71, 1843 (1938). (42) H. Ohle, L. von Vargha and H. Erlbach, Ber., 61, 1211 (1928).

CYCLIC DERIVATIVES O F CARBOHYDRATES

9

reaction of VIII involves only a simple displacement a t C6, reaction of VI will require a specific arrangement of C6, C5, and the attacking and displaced oxygen atoms, which apparently is unattainable. It is interesting to note that 1,4-anhydro-6-0-tosylsorbitol affords43I ,4:3,6-dianhydrosorbito1 on treatment with base; formation of the 5 ,6-anhydro compound may reasonably be postulated as an intermediate stage, and it seems that the greater flexibility of t,he single five-membered ring, compared with that of the bicyclic system of VI, permits the establishment of the transition state required for conversion of the 5,6-anhydro to the 3,6-anhydro compound. The bimolecular elimination reaction (E2) also requires a specific ar~ B, the groups X and Y are rangement (B) of four atomic ~ en ters.4In coplanar with the two carbon atoms and antiparallel (anti-trans, or “true

trans” relation). If the configuration of the reactant prevents the establishment of the transition state B, eliminat,ion by the E2 mechanism will not occur. It is doubtful whether this mode of elimination has been kinetically established for any carbohydrates, but for this, as for other stereospecific reactions, they provide a potentially useful testing ground. Some interesting results have recently been obtained45in studies on elimination reactions of esters of hydroxy acids. The mechanism is not fully established, but probably is of the bimolecular type. An especially interesting observation is that sodium iodide promotes the removal of two vicinal sulfonyloxy groups by a process of cis-elimination; a series of elimination reactions of this type is known in carbohydrate chemistry,39but apparently does not yet include an example from which the stereochemistry of the reaction could be deduced. The spontaneous decomposition of the diazonium ions obtained on treating non-aromatic primary amines wit,h nitrous acid in weakly acidic solutions (deamination reaction) is an irreversible reaction possessing great driving force under mild conditions. The steric consequences of the reaction in the aliphatic and alicyclic fields have proved to be quite diverse,4°-6n (43) (44) (45) (1953). (46)

V. G. Bashford and I,. F. Wiggins, J . Chem. Soe., 299 (1948). See ref. 26(b), p. 465. R. P. Linstead, L. N. Owen and R. F. Webb, J . Chem. SOC.,1211, 1218, 1225 Phyllis Brewster, F. Hiron, E. D. Hughes, C. K. Ingold and P. A. D. S. Rao,

10

J. A . MILLS

but amenable to classification. Its application to carbohydratesl6<4 3 , 62 has already afforded interesting results, including cyclizations and changes in the size of rings. Most of the results obtained so far can be interpreted with the aid of information already gained in other fields, but further use of this reaction in t,he carbohydrate field may well be interesting and profitable. 51e

IV. FUNDAMENTAL PROPERTIES OF RINGS The more important stereochemical features of small alicyclic rings will be mentioned in this Section, with a comparative summary of the properties of the corresponding het,erocyclic rings found in carbohydrates. The stereochemistry of selected carbohydrate derivatives will be discussed in detail in the following Sections, with some amplification of the simple theory outlined here. 1. Conformations

The full importance of the precise shape of molecules in determining stability and chemical reactivity has only recently been recognized. Calculations of the shape of various rings had previously been made, and pyranose and furanose rings had been examined t h e ~ r e t i c a l l yand ~ ~ by x-ray analys ~ s ,55~ but ~ , the significance of the calculated shapes, and methods for selecting the more probable shapes when alternatives were possible, could not be properly assessed until the importance of interactions between nonbonded atoms was realized. The classical theories of steric hindrance related largely to restrictions on rotation in rigid molecules containing double bonds or aromatic rings, and restrictions on rotation above single bonds caused by the collision of bulky groups, as in optically active biphenyls. It is now knownz6,6 6 that steric effects are equally important in cases where rotation about single bonds had previously been considered to be unrestricted. This was first proved Nature, 166, 179 (1950); but see also M. B. Watson and G. W. Youngson, J . Chem. Soc., 2145 (1954). (47) G. E. McCasland, J . Am. Chem. SOC.,73, 2293 (1951). (48) J. A. Mills, J . Chem. Soc., 260 (1953). (49) J. D. Roberts and R. H. Maeur, J . Am. Chem. Soc., 73, 2509 (1951). (50) R. J . W. Cremlyn, D. L. Garmaise and C. W. Shoppee, J . Chem. Soc., 1847 (1953). (51) V . G . Bashford and L. F. Wiggins, Xature, 166, 566 (1950). (52) A . B. Foster, E. F. Martlew and M. Stacey, Chemistry & Industry, 825 (1953). (53) H. S . Isbell, J . Research Natl. BUT.Standards, 18, 505 (1937). (54) C. A. Beevers and W. Cochran, Nature, 167, 872 (1946). (55) 0. L. Sponsler and W. H. Dore, Ann. Rev. Biochem., 6, 63 (1936). (56) J. C. McCoubrey and A. R. Ubbelohde, Quart. Revs. (London), 6,364 (1951).

CYCLIC DERIVATIVES OF CARBOHYDRATES

11

for simple aliphatic molecules; it has been demonstrated spec%roscopically, by electron diffraction measurements, and in other ways.66 For ethane, the most stable state is the staggered state, in which each C-H bond of one methyl group bisects the angle between two C-H bonds of the other group, when the molecule is viewed along the C-C axis; the least stable is the eclipsed state, in which C-H bonds of the two groups -’ -1e in pairs when viewed in this way. Rotation about the C-C bond in etriane requires a passage through successive eclipsed states, and there is a potential barrier opposing this rotation, amounting to about 3 kral./ mole.57The nature of the potential barrier is not fully understood, but seems56t o be due mainly to repulsions between the hydrogen atoms of the two methyl groups. Repulsion will be greatest when the molecule is in the eclipsed state, as this causes the closest approach of hydrogen atoms. When larger atoms or groups are substituted for hydrogen atoms in ethane, there may be several possible staggered states. I t is probable56 that the larger groups tend to take up posiLions as far apart as possible. For long polymethylcne chains a planar, zig-zag arrangement of the carbon “backbone” is particularly favorable. The term conformations is usually applied to those possible configurations of molecules that represent different stages in rotation about single bonds but do not involve significant departures from normal interatomic distances and valence angles. Individual conformations cannot be isolated and studied as pure species, except in the crystalline state. Kevertheless, it is unlikely that an assembly of molecules in the unexcited state will a t any instant contain more than a very small fraction of its membership in highly unfavorable confcpations, and the physical and chemical properties of the assembly will correspond largely to those to be expected of the favorable conformations. The aims of conformational analysis are to devise a set of reliable rules for assessing the relative stabilities of different conformations of molecules in which rotation or oscillation about single bonds is possible and to correlate conformations with chemical and physical properties. It must be emphasized that a molecule need not react in a particular conformation, even though this conformation is far more stabIe than others. If the steric requirements of a reaction demand an unfavorable conformation of the reactant, the reaction will usually still occur (perhaps very sluggishly), especially if the driving force of the reaction is great. For cyclopentane and larger saturated alicyclic rings, which do not require distortion of the carbon valence angles, several conformations are possible, but far fewer than for the acyclic analogs. The potential barriers between stable conformations in cyclohexane and the slightly larger rings (57) J. G. Aston, S. Isserow, G. J. Sanse and R. M. Kennedy, J. Chem. Phys., 12, 336 (1944).

12

J . A . MILLS

will be greater than in acyclic compounds, because change from one conformation to another requires distortion of valence angles as ~ las lpassage through eclipsed states. The factors to be considered'-s in assessing conformational stability in rings are the same as for acyclic compounds: the number of pairs of eclipsed bonds should be as small as possible, and the spatial separation of groups, especially bulky groups, should be as great as possible. Barker, Bourne and Whiffen58have shown that, if the preferred conformation of glycitols is that with the planar, zig-zag arrangement of carbon atoms characteristic of long-chain polymethyleiie compounds, it is 18 of cyclicpossible to explain the main features of the observed patternxb# acetal formation. Brown has drawn attentions to the existence of a potential barrier to the rotation of the methyl groups in dimethyl ether which is comparable in magnitude to the barrier observed for propane.b7He has suggested that this may be attributed to provision by the unshared pairs of electrons, on the oxygen atom, of a source of repulsive force similar to that of a C-H bond, and may have an important bearing on the conformations of heterocyclic rings containing oxygen atoms.8 Further examples of this effect are considered subsequently; it seems to be real, but less in magnitude than the repulsions due t o C-H bonds. 2. Monocyclic Compounds a. Five-membered Rings.-If cyclopentane were perfectly planar, the carbon valence angles would be distorted from 109.46' to log", and all C-H bonds would be in eclipsed positions. The ring is known to be puckered,69 to an extent that seems to diminish non-bonded interactions a t the expense of introducing a slight, valence-angle strain. It has been found that trans1,2-dimethylcyclopentane is rather more stable than the cis isomer, and cis-1 ,3-dimethylcyclopentane slightly more stable than the trans isomer; HaresnapeGohas shown that these orders of stability are to be expected from lion-bonded interactions and the nature of the puckering in the ring. The same order for the 1 ,Zdimethyl derivatives, but not necessarily for the 1,3-, would be predicted for planar cyclopentane rings. For many purposes, such as formation of cyclic derivatives of glycols,61-62 viciiial cis groups attached to the cyclopeiitane ring may be considered to (58) S. A. Barker, E. J. Bourne and D. H. WhifTen, J . Chem. SOC.,3865 (1952). (59) J. E. Kilpatrick, K. S. Pitzer and R. Spitzer, J . Am. Chem. SOC.,69, 2483 (1947); F. A. Miller and R. C. Inskeep, J . Chem. Phys., 18, 1519 (1950); seealso Ref. 26(b), pp. 52-56. (60) J . N . Haresnape, Chemistry & Industry, 1091 (1953). (61) V. Prelog, K. Schenker and H. H. Ciinthard, Helv. Chim. Acta, 36, 1598 (1952). (62) B. Englund, J . prakt. Chem., 122, 121 (1929).

CYCLIC DERIVATIVES OF CARBOHYDRATES

13

be eclipsed, and vicinal trans groups to subtend a projected angle63of 120". An angle of 180" seems to be readily attained with vicinal trans groups, as elimination reactions and epoxide formation proceed normally; only a slight distortion beyond that of the normal puckering is required. The precise shape of tetrahydrofuran is less well known, but the ring is probably puckered, although to a lesser degree than that of cyclopentane. It has fewer eclipsed C-H bonds than has cyclopentane, arid the magnitude of the repulsive forces tending to distort the ring from a plane will depend on the unknown value of the effects contributed by the unshared electrons on the oxygen atom. An x-ray investigation of sucrose54showed a furanose ring markedly distorted from a plane, but part of this distortion may be due t o forces in the crystal lattice and part to the bulky nature of the substituents. For most purposes, the tetrahydrofumn ring may be regarded a8 a plane, fairly readily distortable to meet, the needs of reactiorp: A trans arrangement of vicinal substituents should be more stable than a cis arrangement, but for non-vicinal substituents the order of stability is not readily predictable. The 1 ,S-dioxolane ring (IX), important in cyclic acetals, is probably nearly planar; some small changes in valence angles or interatomic distances are needed on closing the ring.58There are only two pairs of eclipsed C-H bonds, but here, also, the contributions from the unshared electrons on oxygen to the repulsions tending to force the ring out of a plane cannot be accurately assessed. The occurrence of cis isomers, only, in fusions of this ring to five- or six-membered rings is evidence for a preferred planar conformation of the I ,3- IX dioxolane ring. I n cyclic acetals, IX may be substituted at C2, C4, and C5. A trans arrangement of substituents a t C4 and C5 will be more stable than a cis arrangement, as is clearly shown by the quantitative m e a s ~ r e r n e n t son ~~ the cyclic 0-isopropylidene derivatives of erythro- and threo-1 ,2-diphenyl1,2-ethaaediol, and the qualitative evidence given in Section V. The order of stability for isomeric derivatives with substituents at C2 and C4 (or C5) cannot be predicted by the method used60 for the 1,3-dimethylcyclopentanes, since that takes into account the puckering of the cyclopentane ring. Unsaturation in a five-membered ring alters the position slightly. A double bond exocyclic to a cyclopentane ring, as in cyclopentanone, represents a highly stable arrangement, whereas a similar arrangement in a cyclohexane ring introduces an element of instability.s If this is also true of heterocyclic rings containing oxygen atoms, it explainsS why aldonic acids

c-

(63) Defined in ref. 11. (64) P. H. Hermans, Z . p h y s i k . Chem., 113, 337 (1924).

14

J. A . MILLS

tend to form 1,4- rather than l15-lactones. A carbon atom of a cyclopentane ring passes more readily t80the trigoiial state (formation of a carboriium ion) than an atom in a cyclohexane ring, and if this also holds for t)he heterocyclic rings of sugars, it may8 explain why furanosides are formed arid hydrolyzed more rapidly than pyranosides. b. Six-membered Rings.-The stereochemistry of the cyclohexane ring is known in great detail.‘-8*65 Of the possible conformations, the chair form (X)66 is invariably encountered as the predominant species, unless the ring

X

XI

is constrained, by bridging, to assume the boat form (XI).66 Valence-angle strain is absent in both forms, but, whereas in the chair the adjacent C-H bonds are all in fully staggered arrangement, the boat has four sets of eclipsed bonds, as well as a very close approach of two atoms or groups (A and Q1 in XI) a t the ends of the boat. Bonds to substituents in the chair ring are geometrically of two sorts. Six bonds, three on each side, are parallel and normal to the mean plane of the ring, and are now called6’ “axial” ( a ) bonds (formerly termed E or “polar”). The other six, “equatorial” (e) bonds (formerly designated K ) , are directed outward, making angles of 60” and 120” alternately with the vertical, axial bonds. Each equatorial bond is parallel to two sides of the ring. For the majority of substituted cyclohexanes, two alternative chair conformations are possible: all equatorial bonds in the first become axial bonds in the second, and vice versa. It has beeii found that substituents tend to occupy equatorial rather than axial positions, and that, if there are several substituents, the greater number of substituents, or the larger substituents, will be equatorial. The chief reason for the greater stability of conformations with equatorial substituents is the repulsion between an axial substituent and the axial hydrogen atoms or other groups on the same side of the ring. A conformation with two axial groups larger than hydrogen atoms on the (65) D. H. R. Barton, Chemistry & Industry, 664 (1953). (66) Hydrogen atoms are omitted. The diagrams are true for parallelism of bonds, but not necessarily for interatomic distances. The positions for substituent groups were chosen purely for convenience of exposition. (67) D. H. R. Barton, 0. Hassel, K. S. Pitzer and V. Prelog, Nature, 172, 1096 (1953).

CYCLIC DERIVATIVES O F CARBOHYDRATES

15

same side of the ring (such as B and Qz in X) is very unfavorable, as the van der Waals barriers of the groups will overlap. Substituents in the cis-l,3 relation will therefore be equatorial if possible. Two groups in le, 2a relation (cis) are exactly the same distance apart in the chair ring as the same two groups in le, 2e relation (trans): in X, group P is equidistant from the two groups Q', Q2. The IP, 212 arrangement is, however, the more stable. If the requirements of a reaction force two adjacent bonds t o coplanarity, it is easier for this to occur with le, 2a bonds (cis) than with l e , 2e bonds (trans), because the latter deformation tends to crowd together the axial hydrogen atoms.2*68 Vicinal, cis-glycol groups in six-membered rings afford cyclic acetals, whereas the trans isomers normally do not. An alternative conformation for vicinal trans groups is la, 2a (groups A and B in X), which is unstable relative to the le, 2e arrangement, but has the groups antiparallel, and is therefore a favorable conformatioil for reactions requiring this specific geometry. It may be noted that a complete conversion of the ring is not needed in order to change conformation le, 2e to l a , 2a: conversion to the appropriate boat form is sufficient (compare X and X I for groups P, Ql), and this may happen in relatively rigid structures for which complete conversion of the ring is impossible. Even when cpmplete interconversion of chair conformations occurs, a boat form is probably5 an intermediate stage. Because of the differing degrees of hindrance by neighboring atoms or groups, groups in equatorial positions are more accessible than those in axial positions, and this accounts for the observed order of reactivity of axial and equatorial substituents in such reactions as esterification, hydrolysis, and oxidation of alcohols to ketone^.^^ Barton has shown6 66 that an approximate calculation of the relative accessibility at various positions in the steroid nucleus is possible, and the calculations agree with experimental values. Saturated heterocyclic rings containing oxygen, nitrogen, or sulfur atoms also exist preferentially in stable, chair conformations resembling that of cyclohexane. Physical measurements, such as those of electron diffraction and dipole moments, have shown this conformation for p - d i ~ x a n e , ~ ~71. s - t r i ~ x a n e ,71~ ~~*a r a l d e h y d e ,73~ ~~ - t r i t h i a n e ,and ~ ~ 2,4,6-trimethyl-s-tri9

(68) S. J. Angyal and C. G. Macdonald, J . Chem. SOC.,686 (1952). (69) 0. Hassel and H. Viervoll, Acta Chem. Scand., 1, 149 (1947). (70) K.E. Calderbank and R . J. W. Le FBvre, J . Chem. SOC.,199 (1949). (71) M.Kimura and K. Aoki, J . Chem. SOC.Japan, Pure Chem. Sect., 72, 169 (1951);Chem. Abstracts, 46,3341 (1952). (72) R. J. W. Le FBvre, Joan W. Mulley and B. M. Smythe, J . Chem. Soc., 290

(1950). (73) E. Lippert and R. Mecke, Z , Elektrochem., 66, 366 (1951).

16

J. A . MILLS

thiane.69A chair ring is consistent with the chemical properties of 2,6disubstituted p - d i o x a n e ~ The . ~ ~ stereochemistry of alkaloids and related compounds has been extensively d i s c u ~ s e d , ~and ~ - ~the ~ experimental evidence is indicative of a stable chair form for the piperidine ring. In view of the evidence adduced for these diverse ring systems, it is reasonable to assume that a chair form is preferred for the m-dioxane ring (XII) of cyclic acetals, and this agrees with a calculations0 from infrared absorption frequencies. The pyranose ring is always encountered in a chair form unless constrained to a boat form by bridges.2*6 * l1 This should hold for tetrahydropyran rings in general. X-ray crysXII tallographic data54s 5 5 on the pyranose ring have been confirmed by the evidence from formation of complexes with cuprammonium ions.l0.l1 Hassel and Ottar2 had recognized the probability that the pyranose ring would be conformationally analogous to cyclohexane, and they showed that this assumption explained several features of carbohydrate chemistry. Reeves extended the theoretical interpretation of pyranose ring conformations, and has listed101 11 “instability factors” to be used in assessing the relative stability of alternative chair conformations for pyranose rings. His treatment of the subject differs somewhat from that given to derivatives of cyclohexane by other workers. Following Hassel and Ottar? he regarded as unfavorable the simultaneous occurrence of the terminal primary alcoholic group and a hydroxyl group (at C1 or C3) in axial positions on the same side of the ring, yet he did not assign an unfavorable weighting to the presence of two axial hydroxyl groups (at C2 and C4, or Cl and C3) on the same side of a ring. The repulsions between two hydroxyl groups should be quite strong, although not as great as that between -OH and -CH20H, if size of group is the most important factor involved. Information about the conformation of methyl a-D-talopyranoside (or, better, 1 ,5-anhydro-~talitol), in which the two effects are in opposition, might settle this point. Reeves assigned a heavy unfavorable weighting to the close approach of three oxygen atoms (&‘A2effect”)‘O, l1 encountered in methyl 0-D-mannopyranoside. His weighting may be excessive, when it is considered that no

(0

(74) R. I<. Summerbell and J. R . Stephens, J. Ant. Chem. Soc., 76, 731 (1954). (75) R. C. Cookson, Chemistry & Industry, 337 (1953). (76) Asima Chatterjee, A. K. Bose and S. Pakrashi, Chemistry & Industry, 491 (1954). (77) J. McKenna, Chemistry & Industry, 406 (1954). (78) K. Alder and H. A. Dortmann, Chem. Ber., 86, 1544 (1953). (79) I. G . M. Campbell, Ann. Repts. on Progr. Chem. (Chem. SOC.London), 60, 164 (1953). (80)H. Voetter and F. Wollrab, Acta Phys. Austriaca, 6, 529 (1952):

CYCLIC DERIVATIVES OF CARBOHYDRATES

17

marked instability seems to be associated with comparably close approaches of tjhree or even four oxygen atoms in other compounds, such as ort,hoesters and the 2,4:3,5-diacetals of iditol (p. 39). It must be emphasized that the exact shape of these heterocyclic chairform rings is not known. The different interatomic distances (C-C, 1.54; C-0, 1.43; C-S, 1.8 K.) in the rings, and uncertainty about valence angles, may cause some departure from the nice parallelism of bonds found in cyclohexane, but probably not enough to render invalid arguments developed for cyclohexane. It may, howeve?, be more difficult to draw fine distinctions between conformations. Replacement of carbon atoms by oxygen decreases the possibilities for eclipsed C-H bonds (to zero for s-trioxane), and the persistence of the chair ring as the favored conformation is good evidence for the postulated conformational effect8of unshared pairs of electrons. Replacement of carbon by oxygen also reduces the possibilities for interactions between substituents and axial hydrogen atoms; this is discussed in detail in the Section on cyclic acetals (p. 29). c. Other rings.-Three- and four-membered rings are so strained that they may be considered to be planar for all purposes. The largest ring of importance in carbohydrate chemistry is seven-membered, and little is known of the fine structure of the alicyclic analog, cycloheptane; most probably it resembles cyclohexane in general shape, but is more flexible and therefore imposes less restraint on reactions requiring a specific alignment of groups. 3. Structures Containing Fused Rings a. Two Five-membered Rings.-The cis (XIII) and trans forms of bicyclo[S.S.O]octane are known, but combustion data8' show that the trans isomer is the less stable. If the ring junction is labilized by an adjacent carbonyl group, derivatives of the cis form only are obtainable.82Evidently the trans ring junction requires a distortion of the rings beyond that normally attainable. Any factors tending to increase the planarity of the rings in a pair of fused five-membered rings should further increase the stability of the cis isomer relative to the trans. In agreement with this, systems of two fpgmembered rings in which tetrahydrofuran or 1,S-dioxolane is one of the components have so far been encountered only as cis isomers. This should not be taken as proof that the trans forms are incapable of existence; it might be possible to prepare, for example, 1 , 4 : 3,G-dianhydrodulcitol, if a suitable method were chosen. All molecules (in this class) which have cis ring junctions may be re(81) J. W . Barrett and R. P. Linstead, J . Chem. Soc., 611 (1936). (82) A. H. Cook and R. P. Linstead, J . Chem. Soc., 946 (1934).

IS

J. A. MILLS

garded as being composed of t,wo planar rings meeting a t an angle of about 120", in a broad V-shape. This causes an important type of isomerism: A

H

H

0 H

H

B

XIV

XI11

substituents a t other than angular positions may be either inside the T' (endo) or outside (em). An ezo substituent is necessarily far apart from any substituent in the other ring, but an endo substituent may be fairly close to an endo substituent in the other ring. The closest approach is found with two endo substituents located at the points X and Y in XIV, but the endo substituents A and B are also fairly close to each other. (A model of XIV requires that the V be below the plane of the paper, rather than above the plane, in a projection onto paper.) The lesser interactions with other atoms or groups should render compounds with e m substituents relatively more stable than their endo isomers. Furthermore, the faces of the rings inside the V are less accessible than the faces outside, so ex0 and endo isomers should exhibit differences in reactivity in a direction determined by the steric requirements of the reaction in question. 6. TWOSix-membered Rings.-The isomeric decalins had been studied for many yearszgbefore final details of their stereochemistry were elucidated. I t is now known4' 6 9 f 8* that the component rings of both trans-decalin (XV) and cis-decalin (XVI) exist preferentially in the chair form, as in

XV

XVI

cyclohexane itself. trans-Decalin is rigid and essentially flat, with the hydrogen atoms at the angular positions held in axial relationship to both rings. cis-Decalin is L-shaped and flexible; each angular hydrogen atom is axial to one ring and equatorial to the other, and, by a transformation that converts every axial bond to an equatorial bond and vice versa, there is (83) 0. Bastiansen and 0. Hassel, Nature, 167, 765 (1946). (84) R.B.Turner, J . A m . Ghem. SOC.,74, 2118 (1952).

19

CYCLIC DERIVATIVES OF CARBOHYDRATES

derivablea3an alternative stable conformation which is the mirror image of XVI but is ,otherwise stereochemically equivalent to it. In substituted trans-decalins, one of a pair of epimeric substitution products is necessarily axial and the other equatorial. In cis-decalins, the possibility of inversion of the ring conformation permits an equatorial arrangement for each of a pair of epimers.48, 6 0 , a5 If two or more substituents are present in cis-decalin, there will be a t least one axial group in some of the stereoisomers, and it then is necessary, when selecting the most stable conformation, to avoid conformations that show the effect seen in XVII, where an axial group is inside the L-shape and very close to three axial hydrogen atoms. That trans-decalin is more stable than cis-decalin has been shown in various ways,2g a7 the reason for this beings4repulsions between the four hydrogen atoms inside the L-shape (shown in XVI). In trans-decalin, the axial H-H distances are the same as in cyclohexane. Fusions of two m-dioxane rings are found in the 1 , 3 :2,4- and 2,4:3,5diacetals of glycitols, and fusions of a m-diosane and a pyranose ring in the 4,6-acetals of glycopyranosides. If the constituent rings retain the chair forms, the bicyclic systems with trans junctions will be rigid and completely analogous to trans-decalin. However, an important new feature arises in those with cis junctions. Formulas XVIII and X I X show two possible I

XVII

XVIII

x IX

conformations of 1,3:2,4-di-O-methylene-~-threitol,the simplest of the acetals containing two m-dioxane rings with a cis junction. They are interconvertible by the operation described for cis-decalin, but are not stereochemically equivalent: XIX, which will be referred to as “H-inside,” has a close approach of four axial hydrogen atoms, as for cis-decalin; whereas in XVIII, “0-inside,” axial hydrogen atoms do not interfere, but there is a fairly close approach of four oxygen atoms. “H-Inside” and “0-inside” conformations are also possible for fusions of a m-dioxane ring and a pyranose ring with a cis junction, but only three oxygen atoms are then involved. The available evidence strongly indicates that “0-inside” is t,he (85) I<. I,. Elirl, E.ipericnfLn,9, 91 (1953). (S6) C;. F. Davies and 15. C. Gilbert, J . *tm. (‘hem. Soc., 63, 1585 (1941). (87) N. D. Zelinsky and M . B. Turowa-Pollak, Ber., 66,1299 (1932).

20

J. A. MILLS

more stable arrangement. This might have been expected: the unshared electrons on the oxygen atoms may have an effects in promoting chair conformations for the rings, but their close approach is not likely to be as unfavorable as the close approach of the van der Waals barriers of four hydrogen atoms. c. One Five-membered and One Six-rnembered Ring.-For the hydrindans, the heats of combustion29 and the epimerizations effected by aluminum bromide**point to the trans form (XX) as the more stable of the hydrocarbons, but epimerization of 1-hydrindanone by base89 shows that the cis form is the more stable. With cyclic acetals containing an analogous fusion of rings, only cis isomers are stable: this is well known for cyclohexanol, the cyclitols, and H pyranose rings, which never afford acetals engaging trans glycol groupings unless6*the six-membered ring is considerably sx distorted by other fused rings. These results are attributable to the differing shapes of the five-membered ring component. I n cyclopentane, the C-H bond interactions force the ring out of a plane; therefore, the distortions required for the formation of trans-hydrindan tend to stabilize the cyclopentane component, although they destabilize the cyclohesane component by an unfavorable crowding of axial hydrogen 68 On balance, these opposed effects seem to confer greater stability on the trans isomer than on the cis, in which iiiteractions from eclipsed C-H bonds are important. However, if the five-membered component becomes more nearly planar, as in ryclopentanone, tetrahydrofuran, or I ,3-dioxolane, the greater crowding of axial hydrogen atoms in the six-membered ring (required for a 2rans junction) becomes the dominant factor, and the cis junction is the more stable arrangement. d. Other Rings.-A cis junction, only, is possible when three-membered or four-membered rings are fused to other small or medium-sized rings. and 1,3-anhydro-2,43,5-Anhydro-l,2-O-isopropylidene-~-xylofuranose~~ of fusion of a four-membered @ m e t hyl e ne - ~ ~ -x y lito are l~ representatives ~ ring to a five-membered ring and to a six-membered ring, respectively, and other combinations of four-membered rings undoubtedly could be prepared. Anhydro sugars containing an ethylene oxide ring fused to a pyranose ring are an important class. Their conformations are probably the same as

03

(88) N . D. Zelinskg and M. B Turowa-Pollak, Rer , 62, 1658 (1929). (89) W. Huckel, Margarethe Sachs, J. Yantschulewitscli and F. Nerdel, Ann., 618, 155 (1935). (90) 1’ A. 1,evene and A I,. R a y m n n d , J . Bzol. (‘hen, , 102, 331 (1933). (91) R. M IInnn, N Ii Richtmycr, H. W. Dkhl and C. S. Hudson, d A m . Chem. Soc., 72, 561 (1950).

CYCLIC DERIVATIVES OF CARBOHYDRATE8

21

those of cyclohexene oxide and cyclohexene, which contain a chair ring somewhat modifiedg2by the requirement that four atoms in the sis-membered ring be coplanar. Both isomers of 1,2-~ycloheptanediolaffordg3 cyclic 0-isopropylidene derivatives; therefore, both cis and trans forms should be stable in any fusions of seven-membered rings to five-membered or larger rings.

4. Brzdged Rings Bridging of six-membered rings, only, will be considered, as other types do not a t present seem to be important. Alicyclic compounds related to OicycZo[d.d.llheptane (XXI) arc very numerous, but no carbohydrate analogs of proved structure seem to be known. They will of necessity be confined to 1,4-aiihydroglycopyranoses (1,5-anhydroglycofurarioses)and 2,5-anhydroglycofuranosides.Some rep95 but the structures resentatives of the former may have been prepared,34* await confirmation. I n X X I and its analogs, the six-membered ring is necessarily in a boat form, with eclipsed bonds arid also some valence-angle strain. The 1,5-anhydroglycofuranoses are, therefore, likely to be fairly reactive. Dimler has suggestedI2 that they could profitably be examined, in vieu. of the peculiar properties of 1,G-anhydroglycofuranoses, but the latter probably have an entirely different conformation (see below). BicycZo[2.2.d]octane(XXII) also is a rigid structure containing a boat

XXI

XXII

ring, but differs from XXI in being free of valence-angle strain, The numerous eclipsed bonds nevertheless make it rather unstable (see below). Methyl 2,6-anhydro-~~-~-altropyranoside~~ is the sole carbohydrate representative; it is hydrolyzed by acids much more readily than are normal pyranosides. BicycZo[3.2.l]octane(XXIII) has not been studied in detail, but the prop7 8 , 7 9 are entirely consistent with the conerties of the tropane alkaloids75* formation XXIV for nortropane. The 1 ,6-anhydroglycopyranoses,such as (92) D . H. R. Barton, R. C. Cookson, W. Klyne arid C. W. Shoppee, Chemistry & Indiistry, 21 (1954). (93) J. Boeseken and H. G. Derx, Rec. trav. chim., 40, 529 (1921). (94) K. Freudenberg and E. Braun, Ann., 460, 288 (1928); Ber., 66, 780 (1933). (95) P. A. Levene and E. T. Stiller, J. Biol. Chewi., 102, 187 (1933). (96) D. A. Rosenfeld, N. K. Richtmyer and C. S. Hudson, J . A m . Chem. Soc., 70, 2201 (1948).

22

J. A. MILLS

I

SSIII

OH

0 I3

XSIV

OH XXVI

XSV

GH Ho

2

OH

H' XXVII

1,6-anhydro-P-~-glucopyranose (XXV), are knownlO* I1 to contain the chair ring, and presence of a chair ring (XXVI) is consistent'O with some propIt is therefore reasonable erties of methyl 3,6-anhydro-a-~-glucopyranoside. to expect that X XIII and other heterocyclic analogs of i t have chair rings. This will include the 1,6-anhydroglycofuranoses, such as 1 ,6-anhydro-P-~glucofuranose (XXVII), and two other carbohydrate groups, the 2,6anhydroglycofuranosides and 2,5-aiihydroglycoseptanosides, of which no representatives are yet known. Bridging a chair ring in this way causes some distortion of valence angles but, in compensation, non-bonded interactions are small. Evidence t,hat reduction in non-bonded interactions is more important than the introduction of some valence-angle strain is found in the rearrangement of bicyclo[S.S.O]octune (XIII) to X X I I I on treatment with aluminum chloride,*l and in rearrangementsg7~ of derivatives of bicgcZo[2.d.b]octune to derivatives of XX I I I during displacement reactions. A difference between alicyclic and carbohydrate chemistry is found in the rearrangement of methyl 3,6anhydroglycopyranosides of suitable configuration to methyl 3,6-anhydroglycofuranosides under non-hydrolytic conditions of acid catalysid6; the change in ring structure is in the opposite sense to the change of XI11 to XX I I I . The smaller interaction between eclipsed bonds in the tetrahydrofuran rings is the main factor favoring the system of two fused five-membered rings in the carbohydrate derivatives, but the difference in bond lengths in the two classes of compound cannot entirely be neglected. The action of bases on 6-0-tosyl derivatives of methyl a-D-mannopyrano(97) W. von E. Doering and M. Farber, J . Am. Chem. Soc., 71, 1514 (1949). (98) W. C . Wildman and D. R. Saunders, J. Am. Chem. Soe., 76, 946 (1954).

CYCLIC DERIVATIVES OF CARBOHYDRATES

23

affords in each case the 3 ,&anhydro sideg9and of 1,5-anhydro-~-mannitol~~~ derivative rather than the 2,6-anhydro derivative. This selectivity is undoubtedly a conformational effect, but the reactions are irreversible, and probably are not to be taken as conclusive evidence for the relative stability of the two types of anhydro compound. A chair ring may be bridged unsymmetrically by three atoms, and one conformation of the structure has two interlocking chair rings (XXVIII); it is free of valence-angle strain, but there is a close approach of the two hydrogen atoms shown. Some heterocyclic compounds containing this fused-ring system are kno~vn,7~*lot and it is possible that cis-l,3-glycol groupings in chair rings may, under some circumstances, give acetals with this ring structure.lol* Cuprammoiiium complexes of certain cis-l,3-glycols seem to have been obXXVIII tained." 78v

5. The Course of Irreversible Ring Closures Many examples are known of ring closures that afford a five-membered ring when a six-membered ring could be formed and would be the more stable structure. The action of mercuric chloride on thioacetals in neutral 16, ~ 38 ~and 1 ,4-anhydro-hexitols result solution affords g ly co fu ran ~ sid es,~ Other examples from the deamination of I-amino-l-de~xy-hexitols.~~~ may be found in the reviews by PeatI6 and Wiggins.I7It has also been found that, whereas heating ~-mannitolwith hydrochloric acid for a short time affords 1,4-anhydro-~-mannitol,longer heating affords 1 ,5-anhydro-~mannitol.102-*03 Furthermore, PeatI6 has pointed out that formation of a11 ethylene oxide ring seems to take precedence over that of a tetrahydrofuran ring, but his example may not be strictly comparable to the above, as it concerned anhydride formation from 1 ,2-O-isopropylidene-6-O-tosyl-~g l u c o f u r a n ~ s eNevertheless, .~~~ it might plausibly be assumed that, in some at least of the other examples quoted above, the formation of the fivemembered ring was preceded by formation of an epoxide that subsequently (99) W.T.Haskins, R. M. Hann and C. S. Hudson, J . Am. Chem. Soc., 68, 628 (1946). (100) R . C. Hockett and Elizabeth L. Sheffield, J. A m . Chem. Soc., 68, 937 (1946). (101) G.A. Haggis and L . N . Owen, J . Chem. SOC.,399 (1953). [S.B. Baker, Can. J . Chem., (10la) 1,5:3,6-Dianhydro-2,4-0-methylene-~-glucitol 32, 628 (1954))is an example. (102) A. B. Foster and W. G. Overend, J . Cherri. SOC.,680 (1951). (103) H. G.Fletcher, Jr., and H. W. Diehl, J. Am. Chem. Soc., 74, 3175 (1952). (104) H. Ohle and L. von Vargha, Ber., 62, 2435 (1929).

24

J. A . MILLS

suffered intramolecular attack by the hydroxyl group found to be engaged in the product. The above results are understandable when the influence of changes in entropy on rates of reactionlos is taken into account. If the preferred conformation of a glycitol is that8with the planar, zig-zag arrangement of the carbon backbone,58the probability of finding the chain in the configuration favorable for ring closure becomes progressively smaller as the size of the ring is increased; work is required in order to rotate the chain to the favorable configuration, and appears in the rate equation as a large entropy of activation. It has been found106 that the rate of formation of ethylene oxide is over six times that of tetrahydrofuran when the appropriate chlorohydrins are treated with a base, yet the relative heats of activation are strongly in favor of tetrahydrofuran. The controlling factor is the very much greater change in entropy required in the formation of tetrahydrofuran, which quite overshadows the effect of the heat of activation and leads to a slower rate of formation for the less-strained ring. If entropy differences, as well as the inherent strain in the rings, are taken into account, it is reasonable to assume that the ease of formation of anhydro rings will decrease in the order three- > five- > six-membered rings. The position of four-membered rings in the series is difficult to assess; in practice, they are rarely found. If an ethylene oxide ring is formed as an intermediate stage, its enlargement to a four-membered ring through intramolecular displacement mould appear to be highly improbable. The relative ease of scission of anhydro rings, three- >> four- > five- > six-membered, is more nearly related to the degree of strain of the rings. The carbohydrate field is unique in the opportutiities it offers for the study of factors controlling the formation of rings. For example, on treatment of a 1-0-tosylhexitol with base it is possible to observe, in competitive reactions, the relative rates of formation of anhydrides with from three to six atoms in the ring, all resulting from closure a t a secondary hydroxyl group. Derivatives of glycitols up to a t least the octitol stage are accessible,Ia7and the configuration of the carbon chain and the position of the reactive group may be varied. Techniques such as filter-paper chromat ~ g r a p h y or ’ ~ ionophoresis ~~ now make possible the detailed examination of the mixtures of products likely to be formed. Digressing, we may note an interesting application of the postulate58 of a planar, zig-zag arrangement of the carbon chain in polyhydric alcohols. I n such chains, the 0-0 distances have been calculated as 2.51 A. for pC(105) A. E. Remick, “Electronic Interpretations of Organic Chemistry,” John Wiley and Sons, Inc., New York, N. Y., 1943, pp. 196,206. (106) H. W. Heine and W. Siegfried, J . Am. Chem. Soc., 76, 489 (1954). (107) C. S. Hudson, Advances in Carbohydrate Chem., 1, 1 (1945). (107a) G. N. Kowkabany, Advances in Carbohydrate Cham., 9, 303 (1954).

CYCLIC DERIVATIVES OF CARBOHYDRATES

25

groups and 3.43 A. for /?T-g ro ~ p s.~ Now, * in the synthesis of higher aldoses by the cyanohydrin method, an interesting regularity has beeii notedlo*;the predominant product almost, always is the aldonic acid in which the hydroxyl group a t the new asymmetric cetiter is in the pT-relationship to the hydroxyl group that had been located a t C3 in the original aldose, aiid this is not affected by the steric arrangement a t C2 in the aldose. This means that the preferred epimer is the one with the more favorable 0-0 separation, if the aldonic acids also have the planar, zig-zag arrangement). Some examples are known where the formation of alternative, anhydroring structures is possible, and the deformation of the molecule required in the alternative reactions is approximately the same, so that entropy changes may be largely neglected. In such cases the ethylene oxide ring is not favored. The action of bases on methyl 3,4-di-O-acety1-2,6-di-O-tosyl-p-Dglucopyranoside10g and on methyl 2,6-di-O-mesyl-a-~-galactopyranoside~~~ affords predominantly the 3,6-anhydro-2-0-sulfonyl derivative in each case. The above arguments do not apply to the formation of cyclic acetals. In the more stable classes of six-membered acetal rings, the shape of the carbon chain is not greatly distorted from that of the free glycitols,68aiid it is not necessary to postulate that formation of a five-membered acetal ring is intermediate to the formation of six-membered acetals. V. CYCLICACETALS 1. General Remarks

The properties of ring systems discussed above may be used to answer two questions raised by the use of cyclic-acetal groupings for the selective masking of hydroxyl groups in carbohydrates. What is the most, probable ring structure of the acetals obtainable from a particular polyhydroxy compound ? For a given acetal, how many diastereoisomers are likely to be stable ? The first question has been largely answered for acetalation of alditols by the development of empirical rules by H a m , Ness and Hudson,2** and by the extension of these rules by Barker and Bourne.16' However, the empirical approach becomes very cumbersome when attempts are made to cover all the complexities of multi-ring compounds, and its success is entirely dependent on the range and quality of published work available. (108) See ref. 107, p. 26, for a summary of the evidence, and also C. S.Hudson, J . Am. Chenz. SOC.,73,4498 (1951), for variations in the ratio of epimers under certain conditions. (109) E. Hardegger, R. M. Montavon and 0. Jucker, Helu. Chim. Acta, 31, 2247 (1948). (110) A . B. Foster, W. G. Overend, M. Stacey and 1,. F. Wiggins, J. Chem. SOC., 2542 (1949). (111) A. T. Ness, R. M. Hann and C. S.Hudson, J. Am. Chern. Soc., 70, 765 (1948).

26

9. A . MILLS

There is now enough evideuce available to permit an analysis of' the coilformational factors concerned in the formation of cyclic acetals from all classes of polyhydrosy compounds likely to be encountered, and to allow the recognition of structural types for which more evidence is needed. I t will be found that the empirical rules are all sound in principle and may be used to assess quickly the types of acetals likely to be encountered in a given situation, but that a study of conformations is necessary where alternative structures seem to be possible from the empirical rules.21 Barker, Bourne and WhiffenS8concluded that the empirical rules were soundly based by examining the consequences of a planar, zig-zag conformation of the carbon chain in glycitols. The writer feels that a study of the end-products of reaction is the safer approach to the problem of acetal formation, and of other reversible reactions, because deductions based on the conformations of the reactants will only be sound if these conformations, and the mechanism of the reactions, are well established. If all factors concerned could be accurately assessed, the two approaches would give identical answers. Formation of cyclic acetals seems to be the only instance in which both approaches to the problem of preferred ring structure are possible. The answer to the second question above is of considerable practical importance: if several of the possible stereoisomers of an acetal are present in the crude product, the separation of a pure compound may be very difficult. The analysis of structure of acetal rings shows the few cases where mixtures of stereoisomers of comparable stability are likely to be encountered; measures to avoid these mixtures may then be taken. When the analysis indicates that only one stereoisomer will be stable, it defines the configuration at the new asymmetric centers in the acetal rings. The analysis presented below relates, in the main, to acetal formation in the approximate temperature range of 15" to 100". There is evidence that relative orders of stability may sometimes be altered at higher temperatures, five-membered rings becoming increasingly favored.112The change from six-membered to five-membered rings may be accompanied by a disIi4 ruption of fused-ring 2. Monocyclic Acetals

a. Five-membered Rings.-The possibilities for isomerism are generalized in formulas X X I X and XXX. Type XXIX, with an aT-ring, is knownI6' 18 (112) S. M. Trister and H. Hibbert, Can. . I Research, . 14B. 415 (1936). (113) P. A. Levene and A. L. Raymond, Ber., 66, 384 (1933). (114) C. I,. Mehltretter, R. L. Mellies and C. E. Rist, J. Am. Chem. Soc., 70, 1064 (1948).

CYCLIC DERIVATIVES OF CARBOHYDRATES

XXIX

27

xxx

to be favored over type XXX, with an &-ring, as would be expected from interaction between groups R1 and R2.The same order follows from the rotations needed68in order to form the rings from a zig-zag chain. This order of stability for aT- and aC-rings will hold for any carbonyl component. If no other type of ring is possible, formation of aC-rings may occur, even in the decidedly unfavorable case of erythro-1 2-diphenyl-l}2-ethanedi01.~ From theoretical considerations,68 as well as from the general experience that a more symmetrical substitution of a ring tends to increase stability, the aT-ring should be more stable than the terminal a-ring, in which R1 or R2is hydrogen. The best experimental evidence for this is the rearrangeto 2,3:4,5-di-Oment115 of 1, 2:4 , 5 -di-O-isopropylidene-~~,-galactitol isopropylidenedulcitol, catalyzed by pyridinium chloride or quinolinium chloride. Stereoisomerism a t the acetal carbon atom is, of course, impossible if the I n an aT-ring, it is impossible carbonyl component is symmetrical (R3= R4). when R' = RZ,whatever the nature of R3 and R4.In all other instances, it is possible; and no information is available to show whether one of the stereoisomers will be markedly more stable than the other. If R' and R2 are - closely similar, the two isomers of XXIX will almost certainly be obtained in comparable amounts. To avoid the formation of mixtures, aTacetal rings should therefore be prepared from symmetrical carbonyl compounds. Two isomers are known1I6of' 1 ,6-di-O-benzoy1-2,3 )4,5-di-O-benzylidenedulcitol, and, since only the 2,3:4,5-type of acetal, with two aT-rings, is likely t o be formed, the known compounds are two of the three stereoisomers possible for the 2 , 3 :4,5-di-O-benxylidene derivative. The possibility of stereoisomerism a t the acetal carbon atom in a-rings is of considerable practical importance when they are prepared from aldehydes other than formaldehyde. For the cis and trans isomers XXXI and XXXII i t is impossible to predict whether, in any given case, one will be (115) R . M. H a n n , W. D. Mnclay and C. S. Hudson, J . A m . Cheni. SOC.,61, 2432 (1939). (116) W. T.Haskins, R. M. Hitnn and C. S. Hudson, J . A n ! . Cheni. Soc., 64, 136 (1942).

28

J. A. MILLS

Xi11

XXXII

so much more stable than the other as to be virtually the sole product.

Stereoisomerish in the five-membered ring probably occasions the difficulty encountered117-11gin preparing crystalline 1, 3 :2,4:5,6-tri-O-ethylidene-~glucitol, and accounts for the variable melting point quoted120,lz1 for 1,3:2,4:5,6-tri-O-benzylidene-~-glucitol; it will be shown that stereoisomerism in the 1,3:2,4-portion of these acetals is unlikely. The pendent rings of 1,2:5 , 6 and 2 , 3 :5 , 6 diacetals of furanose sugars may also provide diastereoisomers, unless symmetrical carbonyl compounds are used in acetal formation. Two stereoisomers of 2,3-S-benzylidene-2,3-dideoxy-2,3dithio-DL-glyceritol have been obtained.122 b. Six-membered Rings.-Consideration of the generalized, six-membered, cyclic-acetal structures X X X I I I and XXXIV shows that, if the ring is a chair form, the possibilit,ies for stereoisomerism will in practice be more restricted than for five-membered rings. In the pC-ring X X X III, R1and R2 may both be equatorial, whereas in the pT-ring XXXIV one must be axial and one equatorial. This is the reason for the observed precedence16,l8 of PC- over pT-rings. Furt,her, once R1 and R2are assigned equatorial positions in XXXIII, it is clear t>hatthe most stable arrangement a t the acetal carbon atom is that which has the larger group in the equatorial position R3. Since formation of six-membered acetal rings is practically confined to aldehydes, the stable forms of the acetals will be represented by X X X I I I with R4= H. The rule for the selection of the most stable configuration of a six-membered, cyclic-acetal ring is, therefore, that the two residues and the substituent at the acetal carbon atom will all be on the same side of the ring. However, if the groups R3and R4in a DT-ring (XXXIV) are different, it will in general be impossible to decide which will be t,he more stable arrangement a t the acetal carbon atom, unless, for example, Rl is far more bulky than R2, and R3 than R4,which would then require an equatorial arrangement. of Rl and R3.Therefore, if a pT-ring is to be established, formaldehyde is the reagent of choice. (117) (118) (119) (120) (121) (122)

H . Appel, J . Chewi. Soc., 425 (1935). W. R. Sullivan, .J. Awl. (.'hem. Soc., 67, 837 (1945). E. J. Bourne and L. F. Wiggins, J . Cheitr. SOC.,1933 (1948). S. J . Arlgyal and J. V. I,awler, J . A I N .V / m r l . Soc., 66, 837 (1944). 1'. Iiarrer and J. Riichi, H e h . C/iini. .Ictn, 20, 86 (1937).

I,. W . C. Miles and L. K. Owen, J . C h e n i . Soc., 2938 (1950).

CYCLIC DERIVATIVES OF CARBOHYDRATES

20

xxsv A more detailed analysis of the chair-form pC-ring yields some interesting results. The method followed is that used by Barton65in assessing the relative stability of substituents a t various positions in highly substituted derivatives of cyclohexane. In generalized formula XXXV, R5and R6 represent the hydrogen atom and hydroxyl group that will be present a t this point. Consider first the acetal carbon atom. Group R4 (axial) is close to the two axial hydrogen atoms, but group R3 (equatorial) has no hydrogen atoms close enough for interaction. On the reasonable assumption that the repulsive forces from unshared electrons on the ring oxygen atoms are considerably less than those from a hydrogen atom, the difference in stability, in favor of the equatorial group, is much greater than for two similar groups attached to a cyclohexane ring. Therefore, it is highly unlikely that the isomer of a PC-acetal having the substituent a t the acetal carbon atom in axial arrangement will ever be stable enough to be isolated. It may likewise be shown that groups R1 and R2 are strongly constrained to equatorial positions, and that PT-rings are, in consequence, particularly unfavorable structures. The preferred configuration is very clearly defined, even for terminal acetal rings. The situation of the groups R5 and R6 is even more interesting. R6will suffer repulsions from R', R2, and the two axial hydrogen atoms, but R6, although subjected to the same repulsion from R1 and R2as for RE,has no adjacent hydrogen atoms. The repulsion between R6and the oxygen atoms cannot be accurately assessed, but it may well be that the axial group R6 is more stable than the equatorial group R5. There is, therefore, no barrier to the occurrence of axial hydroxyl groups in BC-acetnls of the general formula and XXXIII. 2,4-O-Methylene-~-glucitol,1, 3 :4,6-di-O-mcthylenedulcitol, the related et,hylidene and benzylidene derivativesI5 are stable acetals with axial, ring-hydroxyl groups. The benzylidenation of arabitol has so

30

J. A. MILLS

far afforded only one 0-benzylidenearabitol (D and L forms k n o ~ n ) , ~ * ~ 9 proved124lZ6to be the l13-acetal; this must have the ring-hydroxyl group in axial position. The other possible six-membered acetal of arabitol, the 3 ,5-acetal, would have an equatorial hydroxyl group in the ring, and it is most interesting that formation of the isomer with the axial hydroxyl group seems t o be favored. 1, 3:4,g-Diacetals of mannitol are readily obtainable,16 so an equatorial position for the hydroxyl group is quite permissible. An attempt t o introduce one tosyl group into 1,3-0-benzylidene-~arabitol'26showed that the ring hydroxyl group is unusually reactive, and attempted unimolecular tosylation of 1-deoxy-2,4-O-methylene-~-xylitol~~ afforded a t least 14% of the 3,5-di-O-tosyl derivative. Such ready esterification of axial hydroxyl groups is attributable to the absence of axial hydrogen atoms, which normally hinder the reactions of axial groups. This should be kept in mind in future attempts at selective esterification of hydroxyl groups in compounds containing six-membered acetal rings. Two isomers of 1,3-O-benzylideneglyceritolhave been prepared, but the configurations are not yet known.lZ8The conformations will be represented by XXXV, with R1 = R2 = R4 = H, R3 = Ph, and with the free hydroxyl group at R6in one isomer and a t RRin the other. It will be seen that no sound conclusions could be drawn from the results of any experiments designed to establish the relative reactivity of the hydroxyl groups in the epimers by esterification, or the relative stability by epimerization reactions, and other methods will have to be used for the configurational assignments. The writer knows of no other simple, cyclic, secondary alcohol for which these established methods of configurational determination would be in-. effective. According to the above analysis, the isomerism of the 1,3-O-benzylideneglyceritols is not to be regarded as an example of isomerism a t the acetal carbon atom, as the phenyl group will be equatorial in each isomer. Indeed, true isomerism at the acetal carbon atom in simple, six-membered rings [other than the rare pT-rings) is most improbable. It has been claimed that 1,3 :5,7-di-O-benzylidene-~and -L-perseitol are obtainable in stereoisomeric forms,ll1 but the two forms had identical rotations, and it is difficult to see how the reported interconversion of the isomers could be effected in the basic solvent used. The two substances are probably dimorphic modificaa

(123) E. Fischer, Ber., 27, 1524 (1894); C. A. Lobry de Bruyn and W. Alberda van Ekenstein, Rec. trav. chim., 18, 150 (1899). (124) Marguerite Steiger and T. Reichstein, Helv. C'him. Acta, 19, 1016 (1936). (125) W. T. Hnskins, R. hl. Hanil a n d C. 8. Hudson, J . Am. ('hem. Soc , 66, 1663 (1943). (126) R Grewe and €1. Pnchaly, ('heui. Ber., 87, 46 (1954). (127) E. Zissis and N . K. Richtmyer, J . Am. Cheni. Soc., 76, 129 (1953). (128) P. E. Verknde and J. D. van Roon. Rec. trav. chim., 61, 831 (1942).

CYCLIC DERIVATIVES OF CARBOHYDRATES

31

ttione, and the extremely large difference in melting poiiits (153-155" and 280 f 2 O ) becomes less surprising on noting that the molecule consists of several planar groups separated by bonds about which free rotation is possible, and thereby has several possible modes of packing into a crystal lattice. The demon~tration6~ that the two known isomers of 2,4,6-trimethyl-strithiane have the chair conformation, and that one therefore has an axial methyl group, might be thought to disprove the reasoning used above to show that substituents at acetal carbon atoms in PC-rings are strongly constrained t o an equatorial position. However, in the s-trithiane ring the distance from an axial group t)o adjacent, axial, hydrogen atoms is about 25 % greater than in m-dioxane (if the axial bonds are truly parallel in each), and the repulsive interactions will consequently be considerably reduced. A reason for the marked tendency of acetone and other ketones to afford five-membered rather than six-membered acetal rings is not readily apparent when the process of acetal formation is examined.68It has now been suggested129that formation of a six-membered 0-isopropylidene derivative is not favored because it would require one of the methyl groups to be axial. This view is supported by the demonstration above that such a group would be conformationally unstable, but it is possible that slight alterations in bond lengths or valence angles may be part of the reason for the marked preference for five-membered rings with the gem-dimethyl group. More information about the relative stability of five- and six-membered isopropylidene acetals might be obtained by acetonation of derivatives of allitol or ribitol in which the terminal hydroxyl groups are masked; the possibilities are the formation of the unfavorable &-ring, or the pC-ring with an axial methyl group.21 3. Acetals Containing Fused Rings a. Fused Five-membered Rings.-The only members of this class requiring detailed discussion are the acetals resulting from reaction at adjacent hydroxyl groups in reducing sugars. The course of acetal formation from these sugars is harder to classify than that from glycitols, and, although sufficient evidence is already available to define some of the main features of the reactions, much more is needed. The commonest type of acetal in this class has the I ,3-dioxolane ring fused cis to a furanose ring. Formation of 1,2-acetals of a-D-glucofuranose has been proved for reactions of D-glucose with acetone,130cycl~hexanone,'~~ (129) See ref. 8, footnote 46. (130) C. G. Anderson, W. Charlton and W. N. Haworth, J. Chem. SOC.,1329 (1929). (131) R. C. Hockett, R. E. Miller and A. Scattergood, J. Am. Chem. SOC.,71, 3072 (1949).

32

J. A. MILLS

be n~a l de hyde ,'~ ~ and f0rma1dehyde.l~~ Other examples of acetals with two fused five-membered rings are 2,3: 5,6-di-O-isopropylidene-Dm a n n o f u r a n ~ s e , ' 2,3-0-isopropylidene ~~'~~~ derivatives of D-ribofuranose,9 5 ' 138 and 1 , 2 :3 ,5-di-O-isopropylidene-~-xylofuranose.~~~-~~~ The second acetal grouping in the diacetals may be combined in different ways, but on partial hydrolysis of several diacetals of glucose,'31 137 mansystem of two five-membered rings nose,137and x y l ~ s e141, ~the ~ ~bicyclic ~ always is the more stable portion. The stable, bicyclic monoacetals are of two kinds, containing one or other of the isomeric structures XXXVI (glucose, xylose) and XXXVTI (mannose, ribose) ;since both structures are 1

reasonably stable, the stability is due to steric rather than electronic factors, the most important being the decrease in interaction of non-bonded groups compared to the alicyclic analogs. An entirely different arrangement, in which two five-membered acetal rings are fused to a pyranose ring, is found in the 1,2:3,4-di-O-isopropylidene derivatives of 143 and a r a b i n 0 ~ e . l14b ~ ~Altrose . is reported146t o afford a mixture of 1 ,2 :3,4-and I ,2 :5,6-di-O-isopropylidene derivatives. The one formula XXXVIII shows the stereochemistry of the 142q

(132) J. C. Sowden and Dorothy J. Kuenne, J . A m . Chem. Soc., 74, 686 (1952); B. Helferich and A. Porck, Ann., 682, 233 (1953). (133) Anne White and R. M. Hixon, J . Am. Chem. SOC.,56, 2438 (1933). (134) 0. T. Schmidt, A . Distelmaier and H . Reinhard, Chem. Ber., 86, 741 (1953). (135) K. Freudenberg and A. Wolf, Ber., 68, 300 (1925). (136) H. Ohle and Gertrud Bercnd, Ber., 68, 2590 (1925). (137) K . Freudenberg, W Durr and H. von Hochstetter, Ber., 61, 1735 (1928). (138) P. A. Levene and E. T. Stiller, J . B i d . Chem., 104, 299 (1934). (139) 0. Svanberg and R.Sjoberg, Be?-., 56, 863 (1923). (140) W. N . Haworth and C. R. Porter, J . Chem. SOC.,611 (1928). (141) P. A. Levene and A. 1,. Raymond, J . Biol. Chem., 102, 317 (1933). (142) K. Freudenberg, A. Noe and E. Knopf, Be?-., 60, 238 (1927). (143) P. A. Levene and G. M. Meyer, J . B i d . Chem., 64,473 (1925); 92, 257 (1931). (144) K. Freudenberg and 0. Svanberg, Ber., 65, 3239 (1922). (145) P. A. Levene and J. Compton, J. Biol. Chenz., 116, 189 (1936). (146) F. H . Newth and L. F. Wiggins, J . Chem. SOC.,1734 (1950).

33

CYCLIC DERIVATIVES OF CARBOHYDRATES

Go Ra ,R1

O

3

R2,R1

2

M e l k O HH XXXVIII

)Me*

M e . L O HH 0

Me2

XXXIX

1,2:3,4-di-O-isopropylideiiederivatives of L-arabinose (Rl = R2 = H), D-galactose (R' = €1, R2 = CH20H) and L-altrose (R' = CH20H, R2 = H). The significant feature is the fusion of the two five-membered rings on opposite sides of the pyranose rings: in the nomericlature normally used for alicyclic compoundsg'") the steric arrangement is cis-anti-cis. This involves much smaller repulsions between substituents than the isomeric cis-syn-cis arrangement XXXIX. It could not have been predicted that, for galactose and arabinose, the acetals with ring structure XXXVIII would be more stable than the alternatives containing the structure XXXVI, but it is clear that ribose should show no tendency to form a 1,2:3,4-di-O-isopropylidene derivative of structure X X X I X (R' = R2 = H). From the evidence available147148 the most stable isopropylideiie derivative of fructose seems to be the 2 , 3 :4,5-diacetal, which also has a cis-anti-cis arrangement of rings, similar to XXXVIII. The preference for the formation of the structure XXXVIII in the examples quoted could possibly be due to the use of acetone as the carbonyl reagent : in I ,2-O-isopropylidene-a-D-galactofuranose (XL) one methyl group and the two-carbon "tail" would be in endo positions a t the points where maximum interference between endo groups is possible (p. 18) ; the same would be true of analogous derivatives of arabinose, altrose, and fructose. This explanation probably has to be rejected, because 2,343benzylidene- and 2 , 3 :4 ,5-di-O-benzylidene-~-fructopyranoseare readily obtainable directly from fructose,149 rather than 2,3-0-benzylidene-~fructofuranose, although the last has no endo substituent a t the acetal carbon atom. D-Galactose affords 1 , 2 :3,4-di-O-methylene-~-gaIactopyran0se,lS0although in rather poor yield. The present indications are that the cis-anii-cis arrangement XXXVIII is produced because it is inherently a stable structdre, but more work is desirable on the four sugars concerned. The reason for the formation of two different diacetals from D-altrose is not apparent. 9

(147) H. Ohle and Ilse Koller, Ber., 67, 1566 (1924); M. L. Wolfrom, W. L. Shilling and W. W. Binkley, J. Am. ('hen]. SOC., 72, 4544 (1950). (148) W. L. Glen, G. S. Myers and G . A. Grant, J . Chem. Soc., 2568 (1951). (149) P. Brig1 and 0. Widmaier, Ber., 69, 1219 (1936). (150) L. Hough, J. K. N. Jones and M. S. Magson, J . Chem. Soc., 1525 (1952).

34

J. -4. MILLS

a0)m XL

H O C H ~ . ~ ~ . ; H

HOCHi

9

2

HO XLI

0

HocH H

O V 0

O V 0

Me XLII

XLIII

hle2

The relative stability of exo and endo substituents in two fused fivemembered rings decides the course of acetal formation from aldoses that could afford either 1 , 2 or 2 , 3 acetals of the furanose form.z0D-Ribose is an example. Acetals of the pyranose form may be disregarded for the reasons given above, but acetonation could still afford three different acetals : 1,20-isopropylidene-a-D-ribofuranose (XLI) and the CY and 0 agomers (XLII the latter pair being and XLIII) of 2,3-O-isopropylidene-~-ribofuranose, interconvertible under conditions permitting mutarotation. The 1,2-acetal (XLI) has two endo substituents (a methyl group, and the hydroxyl group a t C3), but the 2,3-acetal has only one endo substituent (methyl) in the &form XLIII. The relatively greater stability of compounds with ex0 substituents therefore*Ois the factor determining the observed formation of the 2 , 3 - a ~ e t a lSimilar .~~ arguments explain the observed formation of the 2 , 3 :5 , 6 - , not the 1,2: 5,6-,di-O-isopropylidene derivative from mannose, and it may be predictedz0 that allose, gulose, and talose will be found to afford 2,3:5,6-diacetals. An interesting problem in the chemistry of ribose may now be solved. Levene and Stillerg6attempted a monotosylation of their preparation of 2,3-O-isopropylidene-~-ribofuranose, but instead obtained a small yield of a di-0-tosyl derivative in which only one tosyloxy group was replaceable by iodine; the resulting monoiodo-mono-0-tosyl derivative was resistant to alkaline hydrolysis. They tentatively suggested that the tosylation had given the 1,5-di-0-tosyl derivative XLIV, and that the inert tosyloxy group was that in position 1. This opinion has been endorsed,16' but is inadmissible, since glycosyl p-toluenesulfonates are highly r e a ~ tiv e ,3and ~ it is doubtful whether a compound of structure XLIV would have survived the conditions (151) R. W. Jeanloz asd H, G. Fletcher, Jr., Advances in Carbohydrate Chem., 6, 435 (1951).

CYCLIC DERIVATIVES OF CARBOHYDRATES

35

of isolation. Levene and Stiller's '2 3,0-isopropylidene-ribose had not been purified through a crystalline derivative, and the presence in it of a small amount of the Il2-acetal XLI might reasonably be expected, and would not have been detected by the methods usedg6for structural assignment. The subsequent isolation of the 3,5-di-O-tosyl derivative of this minor isomer accountsz0for all of the facts mentioned above. Isomerism a t the acetal carbon atom is theoretically possible in fivemembered acetal rings formed when pyranose or furanose sugars are condensed with aldehydes. For furanose derivatives, the isomerism is exo-endo, and the superior stability of exo isomers ensures that one isomer only will be obtainable, and this of the exo configuration.20 1 ,2-0-Benzylidene-a~-glucofuranose'~~ therefore has the complete configuration XLV. If the

H Tso"';Q~Ts

HOCHt-CH OHy' $4 3o y b

O V 0

Mel XLIV

KO XLV

five-membered acetal ring is fused to a pyranose ring, no convincing arguments in favor of a preferred configuration a t the acetal carbon atom can be devised, and a mixture of diastereoisomeric acetals may be encountered. Configurations differing only at C* in XLVI have been assigned2' to the isomeric di-0-benzylidene derivatives of methyl a-D-mannopyranoside,lb2 and the pair of isomers of 1 ,5-anhydro-di-0-benzylidene-~-mannit0~~~~ is of the same type. A compound believed to be 1 6-di-0-benzoyl-3 4-0-benzylidene-2 5-0metlhylene-D-mannitol (XLVII) has been prepared from 1 6-di-0-benzoyl-

I P11 XLVI

XLVII

2 5-0-methylene- and 1 6-di-O-benzoyl-S,4-0-benzylidene-~-mannito1,'6~ (152) G.J. Robertson, J . Chem. SOC.,330 (1934). (153) Y.Asahina and H. Takimoto, Ber., 64, 1803 (1931). (154) A.T. Ness, R . M. Hann and C. S.Hudson, J . Am. Chem. Soc., 66,2215 (1943).

36

J. A. MILLS

but the structure is not rigidly proved, as a rearrangement of acetal groups could have occurred during the preparations. Such a fusion of five- and seven-membered rings with a h m s junction is stereochemically feasible.5s b. Fused Sixmembered Rings.-The fusion (cis junction) of a six-membered acetal ring to a furanose ring is known in several diacetals, but is in general less stable than the five-membered ring also present in the diacetals. A t low temperatures, 1,2-0-isopropyhdene-a-~-g~ucofura1ioseaffords a 3 ,5-0-benzylidene deri~ative,"~and 1 , 2 :3 ,5-di-O-methylene-a-~-gluco15" A st,ructural examination of other difuranose is readily 0btainab1e.I~~) ~~ be made, to determine whether 1,2:3,5acetals of ~ - g l u c o s e 'should diacetals are the favored type in condensations of D-glucose with aldehydes. It will be noted that the arrangement of rings is cis-anti-cis. Fusions of the six-membered 0-isopropylidene ring to the furanose ring are known : D-xylose affords 1,2: 3 5-di-O-isopropylidene-a-~-xylofuran0se139-14i and, if position 6 is masked, derivatives of 1,2:3,5-di-O-iso157~are 6 ~ obtainable from D-glucose. propylidene LY ~-g lu co fu ran o ~ e* Interesting conformational effects are possible with the six-membered 0-isopropylidene ring. It has been found158that formation of 6-deoxy1,2 :3 ,5-d~-~-~sopropyl~dene-~-n~tro-a-~-g~ucofuranose (XLVIII) occurs much much less readily than formation of the corresponding derivative (XLIX) of L-idofuranose when the 1 2-0-isopropylidene derivatives are )

)

6

CHzNOz

CHzIiOz

Me2

hIez

Me

Me XLVIII

XLIX

condensed with acetone; the 1 ,Zmonoacetals may be separated by preferential acetonation of the idose derivative. The inferencelK9that this selectivity is due to a difference in distance between the relevant hydroxyl groups in the monoacetals is unsound; the distance is the same, but there is a conformational difference. The 3,5-acetal rings may be somewhat distorted from the true chair form, but probably not enough to prevent identification of equatorial and axial groups; therefore, in XLVIII, there must be (155) R. L. Mellies, C . L. Mehltretter and C. E. Rist, J . Am. Chem. Soc., 73, 294 (1951). (156) H. Ohle and L. von Vnrghe, Ber., 61, 1203 (1928). (157) K. Freudenberg, H. Toepffer and C. C . Andersen, Ber., 61, 1750 (1928). (158) J. M. Grosheinta and H. 0. L. Fischer, J . Am. Chem. SOC.,70, 1476 (1948). (159) H. G. Fletcher, Jr., Aduances in Carbohydrate Chem., 3, 45 (1948).

CYCLIC DERIVATIVES OF CARBOHYDRATES

37

two axial groups on one side of the ring in either of the possible conformations (in one, the groups are the terminal carbon atom and the methyl group below the plane of the rings, and in the other, C 2 and the other methyl group), whereas the idose derivative XLIX can exist in a conformation with no interfering axial groups. Examples of six-membered acetal rings fused to another six-membered ring are very numerous, and discussion will be restricted to the two cases in which the second ring is a pyranose ring or another acetal ring. Compounds of this type with a trans ring junction present no problems. The bicyclic system is comparable to trans-decalin, and is therefore rigid, with sharply defined equatorial and axial positions a t every point of substitution. The substituents at acetal carbon atoms will be equatorial. Apparently the first definite assignment of a chair form to a six-membered, cyclic-acetal ring was made5 in order to explain the occurrence of only one stable isomer of methyl 4,6-0-benzylidenc-~-~-glucopyranoside, which will have the conformation L.15Qa The stable forms of 4,6-O-benzylidene derivatives of methyl P-D-mannopyranoside, methyl P-D-allopyranoside, and methyl p-D-altropyranoside will differ from L only in the configurations a t C2 and C3.30 The pattern of formation of fused-ring diacetals of glycitols having a trans junction of the rings conforms to expectations,21the known stable acetalsi5 of this type all having the residues in equatorial positions. 2,4:3,5-Di-O-methyleneallit01~~~ (LI, R = H) is a representative of the most stable of all such acetals, those with two equatorial residues. Substituents a t acetal carbon atoms will have the configurations shown. The trans bicyclic system with one equatorial residue (LII) will occur in 1 ,3:2,4-

Ph L

LI

LII

(159:~)I n Ref. 5 , the mirror image of this was inadvertently reproduced. (160) M. I,. Wolfrom, B, W. Lew and R. h1. Goepp, J r . , J . A m . ('hem. Soc., 68,1443 (1946).

38

J. A. MILLS

diacetals of ribitol, 1,3:2,4-diacetals and 1,3:2,4 :5,6-triacetals of allitol, and 1,3:2,4-diacetals and 1,3:2,4:5,6-triacetals of D-talitol. No triacetals of allitol have been described, and this may be taken as evidence that the symmetrical substitution in LI is more stable than the unsymmetrical substitution in LII. The precise reason for this difference is not known; the argument adduced for the superior ease of formation of $3-rings relative to P-ringsKsmay be relevant, but is based on rate of reaction rather than stability. The acetalation of allitol under forcing conditions would be of has been prepared,’e’ but interest. 1,3 :2,4: 5,6-Tri-O-methylene-~-talitol in poor yield, and the 1,3:2,4-diacetal was not detected. A tri-0-benzylidene-n-talito1 has, however, been made by several workers,1Kand undoubtedly is the 1,3 :2,4: 5,6-triacetal. A more exhaustive study of acetals of talitol would be useful, because another type of acetal theoretically obtainable from it is the 3 , 5 :4,6-diacetal, which would be of the “H-inside” type discussed below. The reaction of 1,6-disubstituted allitols with acetone would be of interest, as there is a possibility of obtaining the 2,4:3,5-di-O-isopropylidene derivative with two axial methyl groups instead of the 2 , 3 :4,5-diacetal with two crC-rings.21 Bicyclic trans diacetals with one axial residue will be considerably less stable than the acetals with equatorial residues; this is shown by the absence of 3 , 5:4,6-diacetals of glucitol, 1 ,3 :2,4-diacetals of mannitol and 2,4:3,5-diacetals of arabitol from the products of acetalation of the free glycitols. However, by suitable masking of hydroxyl groups, some trans diacetals with axial substituents should be obtainable. The most interesting will be 2,4: 3,5-di-O-methylene-~-talitol(LIII), in which one terminal group is axial and the other equatorial (see p. 43). It would seem impossible to prepare trans bicyclic acetals with both residues axial, as this is a very unfavorable conformation, and reaction under any conditions could afford other, more stable types of acetal. For example, in attempts to prepare 2,4:3,5-diacetals of dulcitol, which would have two axial groups, there would be no means of preventing the formation of the stable 2 , 3:4,5-diacetals with two a?’-rings. Discussion of the conformation and relative stabilities of bicyclic diacetals with cis ring junctions requires consideration of the possibilities of “Hinside” and “0-inside” conformations of the flexible ring system (XVIII and XIX) as well as of axial and equatorial positions of residues. One possibility that may be excluded is an “inside” axial position for a substituent of the type XVII shown for cis-decalin, as this would be extremely unstable compared to other ronformations. (161) It. M. IIanri, W . T.Haskiris arid C. 6 . IIudson, J . h i . Chew. S o c . , 69, 624 (1947).

CYCLIC DERIVATIVES OF CARBOHYDRATES

30

Diacetals of iditol therefore present no problem. The general formula for 2,4:3 ,j-diacetals of L-iditol is LIV, with “0-inside” ring system and

LIII

LIV

equatorial positions for residues and substituents a t the acetal carbon atoms. The conformation is relatively rigid, and retains the planar zig-sag arrangement6s of the carbon chain of iditol. The formation of the 2,4:3,5diacetals occurs very readily.68The same ring conformation LIV, but with only one substituent (equatorial) will be contained in the 1,3:2 ,bdiacetals of iditol, glucitol, and xylitol, and in 1,3:2,4:5,6-triacetals of iditol and glucitol. The triacetals of glucitol are readily obtainable,I6 but an attempt to prepare a tri-0-methyleneiditol failed.’@ The occurrence of a tri-0benzylidene-L-iditol has been proved,163 and it undoubtedly is the 1,3:2,4: 5,6-triacetal, for which the complete configuration may be defined a t all points except the acetal carbon atom in the 5,B-ring. The 2,4:3,5-diacetals of glucitol and its derivatives are of great interest and importance, and masking of the hydroxyl group a t C1 is sufficient to ensure the synthesis of these diacetals. 2,4:3 ,ti-Di-O-rnethylene deriva166 D-gluconic 167 and D-glucaric tives are known for ~-‘glucitol,’~~1 6 5 * 16’, In these acetals, one residue must be axial and one equatorial for any combination of chair rings, and it is easy to see that the preferred conformation is LV, with “0-inside” rings and the terminal group at C6 axial; the alternative “H-inside” arrangement is excluded because itt would have the terminal group at C1 in the “inside” axial position, analogous t o XVII. The formation of the 2,4:3,5- rather than the 3,5:4,6diacetal from D-glUCOniC acid shows that “0-inside” diacetals with cis junction and one axial group are more stable than diacetals with a trans junction and one axial group. Chemical evidence supporting the conforma(162) R. M. Hann and C. S. Hudson, J . Am. Chem. SOC., 67, 602 (1945). (163) J . SicB, J . Am. Chem. SOC.,76, 1661 (1954). (164) R. M. Hann, J . K. Wolfe and C. S.Hudson, J . Am. Chem. Soc., 66, 1898 (1944). (165) W. N. Haworth and L. F. Wiggins, J . Chem. SOC.,58 (1944). (166) 0. T. Schmidt and H. Heiss, Chem. Ber., 82, 7 (1949). (167) C. L. Mehltretter, R. L. Mellies, C. E. Rist and G. E. Hilbert, J . Am. Chem. SOC.,69, 2130 (1947). (168) W. G. M. Jones and L. F. Wiggins, J . Chem. SOC., 364 (1944).

40

J. A. MILLS

tion LV for 2,4:3,5-diacetals with the D-glucitol configuration is found in the acetolysis of 2 , 4 :3, 5-di-0-methylene-D-glucitol; the 3 5-ring, with the axial group, is cleaved preferentially.”j4 Diacetals of maniiitol with the groups disposed 2 , 3 , 4 , 5 present several problems. The structure of only one has been established beyond doubt: 2,4:3 ,5-di-O-methylene-~-mannitoP”168 has been converted to 2 ,4 :3,5-di0-methylene-L-idaric acid by oxidation and epimerization. Two conformations for this diacetal are possible, the “H-inside” with two equatorial, terminal groups (LVI) or “0-inside” with two axial groups (LVII). It is )

0

1

R LV

LVI

LVII

reasonable to select LVI as the more stablelZ1but this is by no means proved, and it is possible that the diacetal is a typical example of conformational instability, showing some properties of each of the two conformations LVI and 1,VII. There is evidence from other sources suggesting that t,he “H-inside” conformation with equatorial substituents is not very stable. Dulcitol could afford 1, 3 :2 4-diacetals with this conformation, but only 1 , 3 :4 ,6diacetals have been obtained from condensations of dulcitol with aldehydes; consequently, the 1 3 :2 ,4-structure is less st,able than an isolated @-ring. The @ring is less stable than the pC-ring,‘5 and the pC-ring is less stable than the “0-inside” conformation with two equatorial substituents (shown by the formation of diacetals, not monoacetals, from idito1162),therefore the “H-inside” conformation with one equatorial residue is considerably less stable than the “0-inside” conformation with two equatorial residues. Arabitol seems to show considerable reluctance to give 1, 3 :2,4-diacetals, which are conformationally analogous to the 2,4:3,5-diacetals of mannitol, the possibilities being “0-inside” with one axial residue or “H-inside” with )

)

CYCLIC DERIVATIVES O F CARBOHYDRATES

41

one equatorial residue. However, 1,3 :2 ,4-di-O-methylene-~-arabitol has recently been obtained,168athereby fulfilling the predictionz1that it would be more stable than the 2 ,4 :3 ,5-diacetal. The conformational flexibility postulated for the 2 ,4: 3 ,5-diacetals of mannitol would disappear in acetals prepared from higher aldehydes and therefore possessing substituents at the acetal carbon atoms, but the status of such acetals is a t present obscure. -Two isomers of 1,6-dichloro-l ,6dideoxy-2,3 ,4 ,5-di-O-ethylidene-~-mannitolare known,lBgbut the structures have not been determined. Two stereoisomers with the 2,4:3 ,5structure and related t,o the “H-inside” and “0-insidel’ conformations LVI and LVII, respectively, are clearly possible, but the structures 2,3:4,5 and 2 ,5 :3 ,4 must also be considered. There is considerable scope for more work 011 2, 3 ,4,5-diacetals of mannitol, and confirmation of the structure XLVII advanced by Ness, Hann and Hudson164for their mono-0-benzylidenemono-0-methylene derivative is a necessary step in the assessment of the relative stability of the different structural types. The situation becomes even more complex on noting that two isomeric di-0-isopropylidene derivatives have been obtained from 1 ,6-dichloro-1 ,6dideoxy-~-mannitol.‘~~ These must be structural isomers, and of the three possibilities, 2,4:3,5, 2,5:3,4 and 2,3:4,5, the first is found to be conformationally very unfavorable, although none of the three may be excluded from consideration. A diacetal could not be obtained”O from the reaction of acetone with 1,6-di-0-benzoyl-D-mannitol. Two other approaches to the problem presented by the diacetals of mannitol are available. The first is through the base-catalyzed epimerizaacids. It is known3z.1 6 6 - 168 that the tion of 2,4:3,5-di-O-methylenehexaric derivatives with L-idaric, D-glucaric, and D-mannaric configurations (derived by oxidation of LIV, LV and LVI, respectively) are interconvertible in basic solutions, and, although this is not proved, an equilibrium probably is established. The composition a t equilibrium is not accurately known, but the available evidence shows that the L-idaric derivative is the most stable, and the D-mannaric derivative by far the least stable. At present it may only be said that the two possible conformations for 2,4:3 ,5-di-0methylene-D-mannaric acid (corresponding to LVI and LVII) are both unfavorable, but if similar epimerizations could be achieved with ethylidene and benzylidene derivatives it might be possible to assigri structures and configurations to some 2 ,4: 3,5-diacetals of mannitol. The second possible approach is an examination of the reactivity of sub(168a) E. Zissis and N. I<. Richtmyer, J . A m . Chenb. Soc., 76, 5515 (1954); Dr. Richtmyer kindly provided this information in advance of publication. (169) L. F. Wiggins, J. Chem. Soc., 384 (1946). (170) A. Muller, B e y . , 66, 1055 (1932).

42

J. A . MILLS

stituents at the terminal carbon atoms: axial and equatorial groups differ in degree of accessibility, and therefore in reactivity, and this difference is a recognized aid to conformational studies. Of the acetals under consideration, 2)4:3,5-diacetals of glucitol will provide the basic information required, because it is known (p. 39) that C1 is equatorial and C6 axial. A difference in reactivity a t the two centers has been observed several times, but in most cases it is not definitely known whether C1 or C6 was the more reactive center. A di-0-methylenehexonic acid was isolated166as an intermediate in the oxidation of 2 4 :3 ,5-di-O-methylene-~-glucitolto 2 , 4 :3 ,5-di-o-methyleneD-glucaric acid, but its structure is not known; if oxidation occurs preferentially a t the equatorial group in LV, the acid will be the D-gluconic 3,5-di-0derivative. One chlorine atom in 1,6-dichloro-l,6-dideoxy-2,4: methylene-D-glucitol is more reactive than the other in replacement by iodine or elimination by treatment with metallic sodium,171and one bromine atom in 2 4: 3,5-di-O-benzylidene-l 6-dibromo-l,6-dideoxy-~-glucito1 is replaceable by iodine more readily than the 0ther.1~~ The precise course of these reactions is unknown, and information on this point would be of great importance in establishing the mechanism of the reactions. The sugges172 that the (terminal) group a t C6 is the more reactive site is based ti0nl~~0 only on a comparison of the reactivities of terminal groups in the 2,4:3 )5diacetals of mannitol and iditol, without any knowledge of the conformations of the acetals. In fact, in the series of 2,4:3,5-di-O-methylene derivatives of iditol, mannitol, and glucitol, the (terminal) group a t C6 in the glucitol derivative is different from all the other terminal groups in being strongly constrained to an axial position. I n one instance, the course of a selective reaction a t one terminal group has been established, with a very surprising result. The tosyloxy group a t C6 in 2 , 4 :3 5-di-O-methylene-l , 6-di-O-tosyl-~-glucitol is replaced by iodine more readily than that a t C1 when the di-0-tosyl derivative is heated with sodium iodide in acet0ne.17~Preferential reaction a t the axial group would not have been expected in a reaction that seems to be39 rather strongly retarded by increase in the size of substituents in the vicinity of tosyloxy groups. The observed result is, however, in agreement with other results obtained during replacements of terminal tosyloxy groups in bicyclic diacetals with cis ring junctions. Replacement by iodine of the tosyloxy groups in 2 , 4 : 3,5-di-O-methylene-l,6-di-O-tosyl-~-iditol,which are both equatorial, is very slow in acetone at 100°,162 whereas both groups in the )

(171) I,. F. Wiggins and D. J. C. Wood, J . C h e m . Soc., 1180 (1951). (172) W. G. Overend, R . Montgomery and L. F. Wiggins, J . Chem. SOC.,2201 (1948). (173) A. T.Ness, R. M, Hann and C. S.Hudson, J . Am. Chem. Soc., 86,1901 (1944).

CYCLIC DERIVATIVES OF CARBOHYDRATES

43

corresponding derivative of D-mannitol, which may he axial (see LVII), are readily replaceable under the same condition^.'^^ Replacement by iodine is also slow for the equatorial tosyloxy groups in 2 ,4 : 3 ,5-di-O-methylene-lO- t os yl - ~~- xy litoand l’~ ~ 2,4: 3 ,5-di-0-henzylidene-l-0-tosyl-n~-xylitol.~7~ The extension of the reaction with sodium iodide tlo tosyl derivatives of other acetals of iditol, glucitol, and mannitol could provide evidence for the conformation of 2,4:3,5-diacetals of mannitol and, more important, lead to a clarification of the steric requirements of this reaction. The importance of 2,4:3,5-diacetals of talitol, a t present unknown, is now clear: they will have one axial and one equatorial terminal group (see LIII), and the trans ring junction imposes a rigidity of conformation greater than that of any of the diaeetals with cis junctions. Attention may be drawn to a new reagent, chromium trioxide in pyridine, that is said177to oxidize primary alcohols to aldehydes. The potential importance of its application to bicyclic diacetals with free terminal groups is clear, both as a preparative method for aldoses and as a means of assigning conformations. The above evidence seems to show that the “H-inside” conformation is not favored for bicyclic diacetals, and it might therefore be expected that the “0-inside” conformation will be the more stable in fused ring systems made up from a pyranose ring and a six-membered acetal ring, although only three oxygen atoms are present in the rings. Methyl 4,6-O-benzylidene-a-D-galactopyranoside will then have the conformation LVIII, rather than the alternative “H-inside” conformation, and the ,f3 anomer will also have the same conformation and the same configuration a t the acetal carbon atom. The behavior of the two glycosides in cuprammonium solution178 shows that the assigned conformations are correct, a t least for the pyranose ring portion. The position is not so clear for methyl 4,6-O-benzylidene-a-~-idopyranoside. This was made”9 from the galactoside LVIII under conditions not likely to invert the configuration a t the acetal carbon atom, and therefore should also have the “0-inside” conformation (LIX), yet the behavior in cuprammonium solution suggests1O that the pyranose ring has the conformation found in the “H-inside” conformation L X for the bicyclic system. (174) W. T. Haskins, R. M. Hann and C . S. Hudson, J . A m . Chem. SOC.,66, 67 (1943). (175) R . M. Hann, A. T. Ness and C. S. Hudson, J. A m . Chem. SOC.,66,670 (1944) (176) M. L. Wolfrom, W. J. Burke and E. A. Metcalf, J . A m . Chem. SOC.,69, 1667 (1947); A. T. Ness, R. M. Hann and C. S. Hudson, ibid., 76, 132 (1953). (177) G. I. Poos, G. E. Arth, R. E. Beyler and I,. H. Sarett, J . A m . Chem. SOC., 76,422 (1953). (178) R . E. Reeves, J . A m . Chem. SOC.,71, 1737 (1949). (179) E. Sorkin and T. Reichstein, Helv. Chim. Acta, 28, 1 (1945).

44

J. A . MILLS

LIX

LVIII

LX

Thc conformation LIX has a highly unfavorable arrangement of three axial groups in the pyranose ring, and it is possible that the acetal is an example of conformational instability, showing some properties of each of the conformations LIX and LX, or boat forms related to them; the configuration a t the acetal carbon atom is probably that shown in LIX. A preparation of a 4,6-O-benzylidene derivative directly from methyl CFDidopyranoside would be helpful in settling the problem of the configuration and conformation of the stable form. The question is of considerable importance, because this is the first example of conflict between information (on conformation) derived by the use of cuprammonium solution and that derived by other means. The possibility that different workers may not have used the same stereoisomer of methyl 4,6-O-benzylidene-a-D-galactopyrano~ide"~" adds to the difficulty. Parallel arguments apply to methyl 4,6-O-benzylidene-P-~-idopyranoside: the conformation of the pyranose ring observedLoin cuprammonium solution is in conflict with that deduced from the observed conformation of methyl 4,6-O-benzylidene-~-~-galactopyranoside.~~~ Six-membered acetal rings fused to seven-membered rings are found in acetals formed directly from mannitol and certain of its derivatives. The 1 , 3 :2 , 5 :4,6-structure has been proved for the tri-0-methylene164and triO-ethylidenelso derivatives of mannitol, and the 1 ,3:2,5-structure for the di-0-methylene derivative of 6-deoxy-~-mannitol.~~~ These acetals are markedly more stable than the 2,4:3,5-diacetals discussed above. Their , ~ ~is stability has been related to the probable mechanism of f ~ r m a t i o nbut also understandable on conformational grounds.21 The ring junctions are trans, and the triacetals LXI have the trans-anti-trans configuration, which (179a) H. G. Fletcher, Jr., H. W. Diehl and R. K. New, J . A m . Chenz. Soc., 76. 3029 (1954). (180) E. J. Bourne, G. T. Bruce and L. F. Wiggins, J . Chetn. SOC.,2708 (1951). (181) W. T. Haskins, R. M. Hann and C. S. Hudson, J . Am. Chem. SOC.,67, 1800 (1945).

45

CYCLIC DERIVATIVES OF CARBOHYDRATES

is kiiuwns(a) to confer marked stability on the analogous arrangement of three cyclohexane rings. Only glycitols with the nianno configuration can afford triacetals of the trans-anti-trans type; this is a consequence of the “all-trans” disposition of substituents in 2,5-acetals (LXII) of maunitol.

CHzOH

4-J: LXI

CH,OH LXII

2,5-O-Methylene-~-mannitol (LXII, R = H) is the only acetal containing a single seven-membered ring to be isolated so far, and it probably is the most stable of acetals of its class. In the acetals LXI and LXII the sevenmembered acetal ring seems to be conformationally equivalent to a cyclohexane ring. Axial and equatorial positions a t the acetal carbon atoms in the six-membered rings of LXI are clearly defined because of the trans ring junctions; therefore, when R = Me, LXI represents the stable configuration of I ,3 :2 ,5 :4 ,B-tri-O-ethylidene-~-mannitol,of which only one diastereoisomer is known, in agreement with theory. Triacetals of mannitol prepared from other aldehydes will probably also have the configuration LXI. It will be interesting to see whether a similar combination of seven- and six-membered acetal rings is obtainable from other glycitols for which they are theoretically possible. 1,4:3 ,5-Di-O-methylene-~-arabitolwill be conformationally analogous to 6-deoxy-l,3: 2 ,5-di-O-methylene-~-rnannitol, but it lacks the terminal equatorial group; the present indications are that the absence of this group renders the 1,4:3 ,5-diacetal less stable than the 1,3 :2 ,4-diacetal of arabitol. D-ghJcero-D-talo-Heptitol (volemitol) could afford the 2 , 4 :3 , 6 :5 ,7-tri-0-methylene derivative (LXIII) in which the free terminal group is and this might be expected to exceed the triacetal of mannitol (LXI, R = H) H°CHz in stability. An alternative reaction of volemitol would be formation of the 1 , 3 :2,4-di-O-methylene derivative, with two six-membered rings, trans junction, and one 0 ,, 5 equatorial substituent; a careful study of the acetalation of volemitol may therefore establish the true order of stability of the tricyclic system LXI relative to the more 0 common bicyclic acetals. LXIII (182) W. S.Johnson, J . Am. Chem. Soe., 76, 1498 (1953).

c20

46

J. A . MIIALS

The above discussioii of acetals of glycitols shows that the present empirical rules relating to acetal formatioiiIb have a sound theorctic~1.1basis, aiid that, by an esamination of ring conformations, it is possible to detect further subtle differences in stability of acetals. However, even for glycitols, it is impossible at present to draw up a complete order of stahility of the different classes of cyclic acetal, as some of the experimentztl tests of stability will have to be carried out with heptitols, octitols, and iionitols. For esample, it would be possible to compare the relative stability of bicyelic diacetals having a trans ring junction arid one equatorial residue with that, of those having an “0inside” cis ring junction and one equatorial residue by examining the products from D-ylgcero-D-yluco-heptitol, the two structures named being contained in 4 , G :5,7- and 1 , 3 :2,4-diacetals of this heptitol. It must be pointed out once more that the discussion is based on thermodynamic stability, not rate of formation, of ring structures. It is quite conceivable that the most stable acetals will not be those formed most rapidly in a reaction misture, and adequate time for equilibrium to be established must be allowed when attempting to assess small differences in stability.

VI. ANHYDRO COMPOUNDS 1. The 1 ,4 :3,6-Dianhydrohexitols

The very interesting proper tie^'^, of I , 4 :3,G-dianhydro-D-mannitol (LXIV), 1 ,4 :3 ,6-dianhydro-D-glucitol (LXV), and 1 ,4 :3,G-dianhydro-Liditol (LXVI) have iiot yet, been fully explained, but an explanatioii of most,

LXIV

LXV

LXVI

of them may be deduced from the stereochemistry of the three anhydrides. In the iditol derivative, LXVI, both hvdroxyl groups are e m ; in the mannito1 derivative LXIV, both are endo; aiid in the glucitol derivative LXV, that a t C2 is exo and that a t C5 is endo. The configurations LXIV and LXVI are interconvertible by simultaneous inversion of configuration a t both free hydroxyl groups, whereas such aii operation on 1,XV reproduces the original configuration. The order of stahility of the three dianhydrides, when deduced from ease

CYCLIC DERIVATIVES OF CARBOHYDRATES

47

of formation'sY.la4, and e p i m e r i ~ a t i o nunder ~ ~ ~ conditions of catalytic hydrogenation-dehydrogenation, is LXVI' > LXV > 1,XIV. In LXV, t,he 3 ,6-anhydro ring is established less readily,186 and opened by acids more readily,Is7 than is t.he I ,4-anhydro ring. These differences show that the presence of an exo substit>uentincreases the stability of the ring to which 3,is attached, by virtue of the diminished opportunities for interaction between non-bonded subst,ituent,s. The two endo hydroxyl groups in LXIV are sufficiently close together to afford cyclic acetals, and 1 , 4:3 ,6-dianhydro-2, 5-O-metfhylene-D-mannitol is readily obtainable.188~ lag The formation of cyclic acetals from LXV and LXVI appears to be impossible because of the wide separation of t'he hydroxyl groups, mid the substai1c.e reported as 1 , 4 : 3 ,&dianhydro-2,5-0methyleae-L-iditol'aa must have some other structure. The most interesting behavior of the three diaiihydrides is found in the displacement reactions of t>heir0-sulfonyl derivat,ives, and at present it seems that the report>ed results may be covered by three general rules. (I) Displacement of sulfonyloxy groups from the 0-sulfonates may be achieved under certain conditions, and occurs with inversion of configuratJion a t t,he point, of displacement. (2) Ammonia and halide ions displace sulfonyloxy groups, but alkali hydroxides (and perhaps alkoxides) merely hydrolyze the su1fonat)eesters by 0-sulfonyl fission, with ret,ention of configuration. (3) Displacements by halide ions are possible only for endo sulfoiiyloxy groups. These rules are only tent,ative, and may be modified when more sulfonyloxy compounds, and t,heir behavior to a wider variety of anions, have been studied. Rules (1) and (3) are extensions of the arguments used by Matheson and Angyallgoto explain the behavior of the di-0-tosyl derivatives of the three dianhydrides t,oward sodium iodide. These authors found that, in the Dmannitol derivative, both tosyloxy groups are readily replaceable by iodine; but in the D-glucitol derivative only one, and in the L-iditol derivative neither, tosyloxy group was replaceable. Wiggiiis and Wood171had obtained a parallel series of results from react,ions of the di-0-tosyl and di-0-mesyl (183) It. Montgomery and L. F. Wiggins, J. Chem. Sac., 390 (1946). (184) R. Montgomery and 1,. F. Wiggins, J. Cherri. Sac., 433 (1947). (185) H. G. Fletcher, J r . , and It. M. Goepp, J r . , J . A m . Chem. Sac., 67, 1042 (1945); 68, 939 (1046). (186) S.Solt,zberg, R . M. Goepp, J r . , and W. Freudenherg, J . A n / . Cherri. Soc., 68, 919 (1946). (187) R . Montgomery and I,. F. Wiggins, .I. ( ' h e m . Sac., 237 (1938). (188) S. B. Baker, C a n . J . Phent., 31, 821 (1953). (159) J. A. Mills, unpublished observation. (190) N . K. Matheson and S. J. Angyal, J . C h e m . SOC.,1133 (1952).

48

J. A. MILLS

derivatives with sodium iodide and lithium chloride, respectively. The resistance to displacement by halide ions is very marked for the ex0 sulfonyloxy groups.171These results show1g0that the approach of large anions to the faces of the rings on the inside of the V-shaped molecules, required to displace ezo groups with inversion, is strongly hindered. It follows that the product of a successful displacement of sulfonyloxy groups by halide ions will always have the configuratioii of 1,4:3 ,G-dianhydro-~-iditol;for example, a disulfonate of 1 , 4 :3,G-dianhydro-D-glucitol will react with sodium iodide to afford the sulfonate of I ,4:3,6-diankydrod-deoxy-5-iodoL-iditol. The esception noted for hydroxyl ions in rule ( 2 ) is well established for other sulfoiiic esters of carbohydrate^,^^ which therefore differ from simple of hydrolysis of an ezo sulfonyloxy group is alkyl s u l f o n a t e ~An . ~ ~example ~ provided by the compound described as “methanesulfonyl-monochloro1 ,4:3 ,6-dianhydrosorbit 01’’ 171 n-hich must now be designated 1 ,4 :3,6dianhydro- 5 - chloro - 5 - deoxy -2- 0-mesyl- L-iditol: reaction with sodium hydroxide affords the free, hydroxy compound of the same c~nfiguration.~~’ Examples of the hydrolysis of endo sulfonyloxy groups are not available in the dianhydrohexitols, but the endo tosyloxy group in the stereochemically equivalent 3,G-anhydro-1 ,2-O-isopropylidene-5-O-tosyl-~-glucofuraiiose (LXVII) is hydrolyzed hy sodium hydroxide to the corresponding hydroxyl

LXVII

H LXVIII

compound (VII) without any evidence of inversion a t C5.42This is a surprising result, as the conditions are seemingly ideal for displacement of the tosyloxy group with inversion of configuration to afford an exo substituent. The reactions of tosyl derivatives of the three dianhydrohexitols with ammonia1g”* l g 3 are explicable if it is assumed that ammonia, by virtue of its small size, is able to approach either inside or outside faces of the rings and effect displacement with inversion. The product from 1 , 4 : 3 ,G-dianhydro2,5-di-0-tosyl-~-mannitol~~~ will then be 2,5-diamino-l ,4: 3 ,G-dianhydro2,5-dideoxy-~-iditol,and that from 1 ,4:3 ,G-dianhydro-2,5-di-O-tosyl-Dg l ~ c i t o lwill ~ ~ be ~ 2,5-diamino-1 ,4:3.G-dianhydro-2, 5-dideosy-D-glucito1, inversion at C2 and C5 occurring in each case. The formation of 1,4:3,G(LXVIII) from 1,4: 3 , Gdianhydro-2 ,5-dideoxy-2,5-imino-~-mannitol (191) See ref. 26(b)! p. 341. (192) R. Montgomery and L. F. Wiggins, J . Chcni. SOC.,303 (1946). (193) V. G. Bashford and L. F. Wiggins, J . Cheni. SOC.,371 (1050).

CYCLIC DERIVATIVES OF CARBOHYDRATES

49

dianliydro-2 ;5-di-O-tosyl-~-idit,01~93 conforms to the same pattern: initial displacement of one of the e m tosyloxy groups, with inversion of configuration, is followed by attack of the resulting endo amino group on the other tosyloxy group, with a second inversion of configuration. The deamination of the 2,5-diamino-l 4: 3 6-dianhydro-2,5-dideoxy derivatives of L-iditol and of D-glucitol has been observed'g3to yield some 1 4 :3 6-dianhydro-L-iditol in each case. The reactions evidently follow the s ~ mechanism 1 often observed in deaminations of simple aliphatic amines,46 and afford the product of t,he most stable type, with two P X O hydroxyl groups. Analogies drawn between the behavior of the dianhydrohexitols and the 2,4:3,5-diacetals of hexaric acids17'171 may be somewhat misleading, because the behavior of the latter is determined solely by thermodynamic stability, whereas in the former the steric requirements of reactions become very prominent. The two classes are in some ways complementary, the dianhydrohexitols affording opportunities for studying the relations between steric arrangement and course of reaction at secondary carbon atoms, and the diacetals providing similar opportunities for studying these relations a t terminal groups. 2. 1 6-Anhydro Derivatives of Pyranose Sugars The most interesting feature of the chemistry of these anhydro compounds is the spontaneous formation of certain isomers from the free sugars under conditions that completely hydrolyze other isomers back to the free sugars. The present position is as follows. Spontaneous conversion to a 1 ,6-anhydroaldopyrariose (or equivalent derivative of a ketose) occurs in l g 4 . lg6 acidic solutions of several aldoses and ketoses having the a l t r ~ , 'lg7 ~ ~or . g ~ configurations. l ~ ~ The ~anhydrides ~ are present in equi.librium with the free sugar, and the reported conversions to anhydride range from 43 to 85 %. On the other hand,. it is well known that the 1,6anhydro derivatives of glucopyranose, mannopyranose, and galactopyranose are apparently almost. completely hydrolyzed to tlhe free aldoses in dilute acid solutions. (However, levoglucosan has been isolated'98a as a (194) L.F . Wiggins, J . Chem. SOC.,1590 (1949). 74, 2210 (195) J. W. Pratt, N. K. Richtmyer and C. S. Hudson, J . Am. Chem. SOC., (1952) ; 76, 4503 (1953). 62, 961 (1940). (196) N. K. Richtmyer and C. S.Hudson, J . .4m.Chem. SOC., (197) J. W. Pratt, N. K. Richtmyer and C. S.Hudson, J . Am. Chem. Sac., 74,2200 (1952). (198) Laura C. Stewart, N. K. Richtmyer and C. S. Hudson, J . Am. Chem. SOC., 74, 2206 (1952). (198a) A. Thompson, Kimiko Anno, M. L. Wolfrom and M. Inatome, J. Am. Chem. SOC.,76, 1309 (1954).

50

J. A . MILLS

component of the products formed by the action of dilute hydrochloric acid 011 D-glucose.) Reeves’O has suggested that the spontaneous formation of 1 ,G-anhydro derivatives of idoppanose and altropyranose may he due to the conformational behavior of the aldoses: his “instability factors” show that p-Didose and P-D-altrose will exist partly in the conformations favorable for closure of the 1,g-anhydro ring. This reasoning could lead to the further conclusions that D-talOSe, but probably not D-gulose, should fairly readily afford a 1,G-anhydro derivative of the pyranose form. Since the formation of the anhydrides in acid solution iiivolves an equilibrium, it is more correct to examine the conformations of the end-products of the reaction. An explanation is then apparent. I n 1 ,G-anhydro-P-Didopyranose (LXIX), the three free hydroxyl groups are all equatorial in the chair ring’” known to be present in these anhydrides, and two equatorial (LXX) and hydroxyl groups are present in I ,6-anhydro-P-D-altropyranose (LXXI). The known stable types of in 1,6-anhydro-p-~-gulopyranose

HO

OH

OH LSS

LXIS

LXXI

1,6-anhydropyranoses are therefore relatively free of interactions between non-bonded substituents. Of the three known unstable anhydrides, 1 ,Ganhydro-P-D-glucopyranose(XXV) has all three hydroxyl groups axial, and it will be seen that the derivatives of mannose and galactose each have two axial hydroxyl groups, one group being on the same side of the ring as the bridge. Non-bonded interactions in these are therefore very considerable. If’ steric strain due to repulsions between substituents in the chair ring is the chief factor deciding the stability of 1 ,G-anhydroaldopyranoses, it is possible to calculate that, as the configuration of the nldose is varied, the amount of anhydride present at, eyuilihrium in dilute acidic solutions should decrease in the following or er of configurations,

4

ido

> altro, gulo > talo > a110 > galacto, manno > gluco.

The latest work, lgaa. 19*b however, appears to place allo before talo, and shows that the glum configuration permits some formation of anhydride. (198b) J. W. P r a t t and N. K. Richtmyer, Abstracts Papers Am . Chem. Soc., 126, 22D (1954); L. D. Ough and R. G. Rohwer, ibid., 126, 16D (1954).

CYCLIC DERIVATIVES OF CARBOHYDRATES

51

It is possible that, in t,he allo and gluco anhydrides, hydrogen bonding between a xkl hydroxyl groups may promote stability. A similar order should hold for ketoses, adthough the additional equatorial group (at the anomeric center) may increase the stability of individual isomers, and the diketose dianhydrides are actually always formed. 3. The Scission of Epoxide Rings Since t.his topic was last reviewed in this serieslL6much additional information about the scission of epoxy derivatives of pyranose sugars has become pvailable. Only this t)ype of epoxide will be discussed here. Several w o r k e r ~ ' ~ have.cornpiled 9-~~~ lists of t,he available evidence. There is general agreement that scission of the epoxide ring to provide two free substituenh always occurs with inversion of configuration16a t the point of attack by the reagent, and the chief problem is the predic~tioiiof t,he direct,ion of opening of the ring when two alternatives are possible, which is always the case for 2,3- and 3,4-anhydro derivatives of pyranose rings. For example, a 2,3-anhydride with the D-manno configuration may afford products with the D-gkKO configuration (attack a t C2), or with the D-altro configurat'ion (attack a t C3); products with the D-gluco and D-altro configurat'ions may also result from a 2,3-anhydride with the D-a,llo configuration, but in this case by attack a t C3 and C2, respectively. Some interesting regularities were noted. Derivatives with the D-altro configuration predominate in the products of scission, by bases, of methyl and methyl 2,3-an2,3-anhydro-4,G-0-benzylidene-a-D-allopyranoside hydro-4,6-O-benzylidene-a-~-mannopyranoside.~~~~ 204 Scission of methyl 2,3-anhydro-4,6-O-benzylidene-a(and P)-D-talopyranoside and methyl 2,3-anhydro-4,6-O-benzylidene-a(and @)-D-gulopyranosideaffords, almost ~ ~The ~ * action of ammoniazo6or sodium entirely, derivatives of ~ - i d o s e .*06 gives derivatives of methoxidez06 on 1 ,6: 2,3-dianhydro-p-~-talopyranose 1 ,6-anhydro-p-~-galactose,and t,he same reagentszo6with 1 ,6:3,4-dianhydro-P-D-talopyranose yield derivatives of I ,B-anhydro-P-~-manqo(199) F. H. Newth, W. C. Overend and L. F. Wiggins, J. C h e w . Soc., 10 (1917). (200) S. Mukherjee and H. C. Srivastilva, Proc. I n d i a n A c a d . Sci.,36A, 178 (1952). (201) A. I(.Bose, D. K . It. Chaudhuri and A. K. Bhattacharyya, Chemistry & Industry, 869 (1953). (202) F. H. Newtjh, Chenristry & I n d u s t r y , 1257 (1953). 1193 (1935); G. J. Robert(203) G . ,J. Robertson and C. F. Griffith, J . Chem. SOC., son and W. Whitehead, ibitl., 319 (1940); W. €I. Myers and G. J. Robertson, J . A m . ('hem. Soc., 66, 8 (1943). (204) N. K. Richtmyer and C . S. Hudson, J . A m . Chevr. Soc., 63, 1727 (1941); N . K. Richt,myer, ,~dva7iccsZ'TL ('arbohydrafe ('/ienr,, 1, 37 (1945). (205) L. F. Wiggins, J . f'hem. Soc., 522 (1944). (206) Syllil 1'. .James, F. Sinit!, M. Stacey and I,. F. Wiggins, J . <:hem. Soc., 625 (1946).

52

J. A . MILLS

pyranose; very little of the alternative product with the D - i d 0 configuration is formed in any of these reactions. Other epoxy derivatives of pyranose sugars do not show any regularity in the direction of scission. Methyl 2,3-anhydro-4,6-di-O-methyl-@-~mannopyranoside reacts with ammonia to afford predominantly the product with the D-altro configuration, but with sodium methoxide equal amounts of D-altro and D-gluco isomers are obtained.*07No useful rules could be formulated to predict the direction of opening of epoxides by acidic reagents.Ig9.2oo The writer pointed out20,208 that, in cases where the conformations of the products of scission of epoxides are readily deducible, the major product of scission is that with the two resulting groups in axial position; such carbohydrate epoxides therefore appear to conform to the rule formulated in the steroid field,209namely, that scission of epoxide rings occurs in the direction affording two axial groups in the products. The conformation of methyl 4,6-O-benzylidene-a-D-altropyranoside is rigidly fixed as LXXII, with two axial hydroxyl groups, by virtue of the trans ring junction ; therefore the predominance of products with the altro configuration in the scission of 2,3-anhydro-4,6-O-benzyliPI. OMe dene derivatives of the met,hyl mannopyOH ranosides and methyl a l l o p y r a n o ~ i d e s204~ ~ ~ ~ LXXII conforms to the rule. The preferred direction of scission for 1,6:2,3- and 1,6:3,4dianhydro-/3-D-talose206also provides two axial groups in the products. The preponderance of D-id0 derivatives in the products of scission of the and -Depimeric methyl 2,3-anhydro-4,6-O-benzylidene-~-talopyranosides gulopyran~sides'~~* *05 will also conform to the rule if methyl 4,6-0-benzylidene-a-D-idopyranoside has the "0-inside" conformation L I X discussed on p. 44. The above rule is empirical, and was intended to be used only with epoxides that will afford products having well defined conformations. It was restricted to reactions with basic reagents, because of the likelihood that acids would destroy the rigidity of the conformations by removal of henzylidene groups or cleavage of stabilizing anhydro rings before effecting 208 scission of the epoxide C o o k ~ o n has ' ~ examined the mechanism of the scission of rarbohydrate epoxides in more detail. He considers that the initial product of scission by

k : ?

(207) W. IT. G. Lake and S. Peat, J. Cheni. SOC.,1417 (1938); W. H G. Lake, W. N. Haworth and S. Peat, ibid., 271 (1939). (208) See F. H . N m t h and R. F. Homer, J . ('hem. SOC.,989 (1953). (209) A. Fiirst and P. A. Plattner, 12th Intern. Congr. Pure and A p p l . ( ' h e m . , New I-ork, 1951; Abstracts, p. 409.

CYCLIC DERIVATIVES OF CARBOHYDRATES

53

acids or bases mill always have the two groups in axial positions (antiparallel arrangement), as would be anticipated from the transition state of the reaction (p. S), and that the direction of scission will depend largely on the preferred conformation of the epoxide, which may be deduced with some confidence in many cases.13~92 In rigid ring systems, the epoxide will have a rigid conformation, arid will open in one direction; the initial arrangement will be retained] and the major product has two axial groups. For epoxides of monocyclic pyranosides, the conformation will not be so rigidly fixed, and there is more possibility for scission in each of the two possible directions. The initial product of opening, in either direction, will have axial groups, but may then pass into a more stable conformation with two equatorial groups. Cookson has also postulatedL3a directing electronic effect from the ring-oxygen atom in the opening of non-rigid epoxides by acidic reagents] but more work on this type of scission would be desirable. At present, it may safely be stated that scission, by basic reagents, of epoxide rings in rigid ring structures will afford predominantly the product with two axial groups. This is likely to be true for acidic reagents, too, if the conditions of reaction are such that benzylidene or other stabilizing groups are not removed prior to scission of the epoxide.208The scission of epoxides by Grignard reagents does not conform to the rule.210An attempt to apply conformational rules to the opening of 1 ,2-anhydro derivatives of pyranose sugarszoLis premature, as it has never been shown that epoxides of the glycosan type can suffer scission a t a point other than the anomeric center. (210) A. B. Foster, W. G. Overend, M. Stacey and G. Vaughan,

3308 (1953).

J. Chem. Soc.,

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COLUMN CHROMATOGRAPHY OF SUGARS AND THEIR DERIVATIVES

BY W. W. BINKLEY Depnrlwient [$ Chemistry, T h e Ohio State University,(hlunibus, Oh,io

CONTESTS 1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11. Adsorbents for the Column Chromatography of Sugars, Sugar Alcohols, and Sugar Acids . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1. Carbon . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. Fuller’s Earth Clay . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. Carbohydrates . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. Ion-exchange Resins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5. Calcium Acid Silicate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6. Other Adsorbents . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 111. Column Chromatography of Sugar Derivatives . . . . . . . . . . . . . . . . . . . . . . . . . . 1. p-Phenylazobenzoyl Esters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. Acetate Esters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. Methyl Ethers.. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. Other Derivatives. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

55 56 56 64 70 75 77 79 79 79 83 88 92

I. INTRODUCTION The stunning complexity of some of the sugar mixtures from natural sources and from synthetic studies presents a formidable deterrent to many investigators. New hope now exists. The serious application, beginning scarcely more than a decade ago, of column chromatography to carbohydrate investigations has produced resolutions rivaling in magic that of Aladdin’s lamp. Column Chromatography is a specialized manipulation of the general technique of selective adsorption with powdered materials.’ These adsorbents are molded vertically in the shape of cylindrical columns. Three techniques are usually employed in the formation of such chromatographic columns: dusting dry powder into an empty dry tube, slowly adding dry adsorbent to a tube containing liquid, and pouring a thin slurry of adsorbent and developer into an empty tube. Other useful techniques closely or remotely related to column chromatography involve use of the (1) Column chromatography is the separation, by selective adsorption, of a mixture of substances into zones or bands, on a column of powdered materials, after the passage of proper solvents.

55

56

w. w.

BINKLEY

chromatopile,? the ~hromatopack,~ and blocks of cellulo~e.~ The removal, with a powdered adsorbent, of some of the components of a solution of a mixture of sugars has been achieved5’6 ; but this important process is not chromatography. The recent flood of reviews and books 011 chromatography refleck in part the iiicreasing popularity of this versatile tool.?

11. ADSO~~BENTS FOR THE COLUMN CHROMATOGRAPHY OF SUGARS, SUGARALCOHOLS,AND SUGAEACIDS 1. Carbon

The use of carbou columns or beds for t,he purification of sugar mixtures by selective adsorption* preceded the advent of modern chromatography by nearly a c e i ~ t u r yOnly . ~ little more than a decade ago, the potent)ialities of this adsorbeut for the resolution of sugar mixtures were recognized.1° A (2) H. I<. Mitchell and F. A. Haskins, Science, 110, 278 (1949). (3) W. 1,. Porter, A n a l . Chwn., 23, 412 (1951). (4) L. S. Cuendet, R. Montgomery and F. Smith, J . Atti. Che.nz. Soc., 76, 2764 (1953). ( 5 ) F. Hnyashi, J . Biocheni. (Japan), 16, 1 (1932); Chem. Abstracts, 27, 8 (1933). (6) Edna M. Montgomery, H. F. Conway and F. B. Weakley, U. S. P a t . 2,610,931 (1952); Chcni. Abstracts, 47, 2523 (1953). (7) (a) H . G . Cassidy and others, “Conference on Chromatography,” Ann. N . Y . Acad. Sci.,49, 141-325 (1948); (b) E. L. Hirst and J . K. N . Jones, Discussions FaraXo. 7, 268 (1949); (c) H. H. Strain, A n a l . Chein., 22, 41 (1950); (d) T. I. d a y SOC., Williams, Discovery, 396 (1950) ; (e) H. H . Strain, “Chromatographic Adsorption Analysis,” Interscience Publishers, Inc., New York, N . Y ., 1945 (revised reprint) ; ( f ) T . I. Williams, “An Introduction t o Chromatography,” Blackie & Son, Ltd., London, 1946; (g) W. W. Binkley and M . L. Wolfrom, “Chromatography of Sugars and Related Substances,” Sugar Research Foundation, N . Y . , Sci.Rept. S e r . , No. 10 (1948); (h) E. Lederer, “ProgrBs de la Chromatographie, 1. Chimie Organique e t Biologiqae,” Hermann & Cie., Paris, 1949; (i) L. Zechmeister, “Progress in Chromatography, 1938-1947,” John Wiley and Sons, Inc., New York, N. Y., 1950; (j) H. G . Cassidy, “Adsorption and Chromatography,” Vol. V of “Technique of Organic Chemistry,” A. Weissberger, ed., Interscience Publishers, Inc., New York, N . Y., 1951; (k) 6 . N . Balstori and B. E. Talbot, “A Guide to Filter Paper and Cellulose Powder Chromatography,” H. Reeve Angel and Co., Ltd., London, 1952; (1) R . C. Brimley and F. C. Barrett, “Practical Chromatography,” Reinhold Publishing Carp., New York, N . Y., 1953; (m) L. Hough, “Analysis of Mixtures of Sugars by Paper and Cellulose Column Chromatography,” Vol. I of “Methods of Biochemical Analysis,” D. Glick, ed., Interscience Publishers, Inc., New York, N . Y., 1954; (11) R . J . Dimler, T r a n s . A7n. Assoc. Cereal Chemists, 12, 1 (1954); ( 0 ) T. I. Williams and H. Weil, A r k i v Kemi, 6, 283 (1953). (8) E. P. Barrett, Advar~cesin Curbohydrafe Chew., 6, 205 (1951). (9) NI. Tswett, Ber. deut. botan. Ges., 24, 316 (1906). (10) 4 . Tiselius, A r k i v Kemi, Mineral. Geol., 14B, No. 22, 4 (1940).

57

COLUMN CHROMATOGRAPHY OF SUGARS

versatile mcthod for the chromatography of colorless substances 11 as developed.ll Sugar solutions and developers, under pressures of up to one atmosphere, were forced u p through an adsorbent bed packed in a metal tube (5-20 mm. high by 20 mm. inside diameter). This tube was connected to the bottom of a rectangular glass (or Lucite) tube (a cuvette, 19 by 94 mm. cross-section by 150 mm. height) which facilitated observation of the refractive index and photographing of the interfaces12 in the effluent from the chromatogram. The entire chromatographic unit was immersed in a thermostatically controlled, clear bath. The utility of carbon for the separation of mono- and di-saccharides was apparent a t the outset (see Fig. (a)

-

L

Meniscus

Sodium chloride

:I 3

v

.Q .4-

Lactose

U

I

c 0

EL

/

increasing volume of developer

FIG. 1.-(a) Sketch of Interfaces Formed in the Effluent from the Chromatogmm, on Carbon (Kahlbaum Active Carbon), of Sodium Chloride, D-Glucose and Lactose; (b) Graph of the Refractive Index u s . Volume of the Effluent of the Same Chromatogram.ll

and separations were extended quickly t o a trisaccharide (raffi(11) A. Tiselius, Arkiv Kemi,Mineral. Geol., 14B. No. 32 (1941); Science, 94, 145 (1941); R. J. P. Williams, L. Hagdahl and A. Tiselius, Arkiv Kemi, 7, 1 (1954).

(12) Method reviewed by H. Schardin, “Das Toeplersche Schlierenverfahren,” VDI-Verlag, Berlin, 1934.

58

W. W . BINKLEY

iiose).ll, l3 The sensitivity and flexibility of this chromatographic method were improved markedly by allowing t)he sugars and developers to pass, under pressure, down through the adsorbent into a very small cuvette (80.0 mm. in length by 1.4 mm. inside diameter, volume 0.13 ml.; see Fig. 2 ) , Pressure applied

Refractive index observed

~Light source

Windows

FIG.2.-Sketch of a Chromatographic Assembly for Interferometric Measurements 011 the Column Effluent.'*

which was connected to a frartion collector outside the temperaturecontrolled bath.I4 The cuvette was aligned with a sensitive interfer~meter'~ which permitted rapid rec3gnition of composition changes in the effluent. The effluent turnover was quick in this small cell; thus, the opportunity for the intermixing of zones was greatly reduced. The combination of this (13) A. Tiselius, Advances in Colloid Sci.,1, 89 (1942); Kahlbaum active carbon, Carboraffin, Suprsnorit, Merck animal carbon, and Merck blood charcoal were found useful in these separations. Norit P-3, Carboraffin Supra, and Darco G-60 were used subsequently. (14) A. Tiselius and S. Claesson, A r k i v K e m i , Mineral. Geol., 16B,No. 18,l (1942); Claesson, Ann. N . Y . Acad. Sci., 49, 183 (1948). T) F. Haber and F. Liiwe, Z . angew. Cheni.,23, 1393 (1910).

COLUMN CHROMATOGRAPHY O F SUGARS

59

excellent method together with an established scale of certain sugars in the order of their increasing affinity to carbon (see Table 1)16 provided a powerful tool for the resolution of sugar mixtures. Three general analytical procedures have employed this method ; they are identified by the mode of development used, namely, frontal, elution, and displacement analysis.l6,l 7 An aqueous solution containing D-glucose, sucrose, and raffinose is allowed to pass through a carbon column; no developer is used. Pure D-glucose (the least tightly held, see Table I) appears first in the effluent, followed by a mixture of D-glucose and sucrose, and finally by all the components of the original solution. Each change in effluent composition produces a new vertical front or step in the graph TABLEI T h e Order o j Increasing A f i n i t y to Carbon (Norit P - 3 ) of Certain Sugars, Phenol, and Ephedrine f r o m Their Aqurous Solictions'O L-Rhamnose L-Arabinose o-Fructose D-Xylose D-Glucose D-Galactose D-Mannose Sucrose Lactose Maltose Raffinose Phenol Ephedrine

showing the refractive index as a function of effluent volume (see Fig. 3).18 This has been designated a fronta>lanalysis; t,he yuant,itative value of such a diagram is limited. An aqueous solution of D-glucose, sucrose, and raffinose can be adsorbed a t the top of a carbon column and developed with wat)er and ethanol-water mixtures until all of t,he sugars have passed through the column. Separately, D-glucose, sucrose, and finally raffinose will be recovered, and each component will produce a peak in the interferometric change of the column effluent with volume as portrayed in Fig. 4. This is (16) A . Tiselius, Kolloid-Z., 106, 101 (1943). (17) A . Tiselius, Arkiv K e m i , Mineral. Geol., 16A, No. 18, 1 (1943). (18) T,. Hagdahl, Acta C h e w . S c a d . , 2. 574 (1948). The definition of the diagrams of the chromatograms, by t'rontrtl analysis arid displacement development, was improved markedly by the use of n. series of t.hree small columns connected t o a micromiser (a capillary filled with qunrte sand) which led directly t o the cuvette for the interferometric readings.

60

W . W. BINKLEY

an elution analysis. The amount of each component in the mixture can be estimated from the area under its respective peak (see Fig. 4). Complete recovery of strongly held substances by elution chromatography is slow

I

I I Raffinose, I

X '0 QI

.-E 5 .-a

c 0

2

L

a

E

Ic

Increasing effluent volume

FIG.3.-Frontal Analysis on Carbon (Carboraffin Supra) of a Mixture of D - G ~ u cose, Sucrose, and Raffinose (Refractive Index us. Volume of EBuent). Developer

-+ Water Jr.-

5%Ethanol

+-15%Ethanol-

Increasing effluent volume

FIG.4.--Elution Analysis, on Carbon, of a Mixture of o-Glucose, Sucrose, and Raffinose; Developed with Water a.nd Ethanol-Water Mixtures (Refractive Index us. Volume of Effluent).

and often difficult. These obstarles are largely eliminated by addition to the developer of a substance more strongly adsorbed than any of the other adsorbates. Soon, the adsorbent is saturated with this substance which has displaced all of the less strongly adsorbed materials; the order of their entry into the effluent is not altered (see Fig. 5). This has been designated a displacement analysis. The concentration of an individual component in a mixture can be estimated from the length of its respective plateau (with

61

COLUMN CHROMATOGRAPHY OF SUGARS

the aid of proper calibration diagrams). The applicationig of this displacement technique to a corn-starch, limit dextrin fractionz0of molecular weight, 975 showed the presence of tri-, tetra-, penta-, and hexa-saccharide comI I I

I

I

Increasing effluent volume

FIG.5.-Chromatography on Carbon (Carboraffin Supra) of a Mixture of D-Glucose, Sucrose, and Raffinose by Displacement Development with 0.5% Ephedrine in Water (Refractive Index us. Volume of Developer) .I *

I

I I

Saccharides

I

I

I

1

I

I

I

I

L

Increasing effluent volume

FIG.6.-Chromatogram on Carbon (Carboraffin Pretreated with 0.15% Ephedrine) of a Corn-starch Limit Dextrin (MW 975), by Displacement Development with 0.5% Ephedrine (Refractive Index us. Volume of Effluent). 1 9

ponents (see Fig. G). The resolution of gram quantities of sugars was found to be sharper with a series of successively smaller carbon rolumns (see Fig. 7)'l than with a single large o m . (19) C. Weibull and A. Tiselius, A i k ~ Kemz, v M i n e r a l . G ~ o l .19A. , NO.19,22 (1945). (20) B. Ortenblad and K. Myrback, HLochCwL. Z , 303, 335 (1940). (21) S. Claesson, A r k z v Kemz, Mzneral. Geol., 24A, No. 16, 3 (1947). The displace-

ment developer was 4% phenol in water.

62

W. W. BINKLEY

Developer

columns

Effluent

FIG.7.-A

Series of Carbon Columns Arr:mged for Chromatography of Fairly Large Quantities of Sugars.18, 21

Recently, the versatility of carbon (Darco G-GO) as an adsorbcnt for the segregation of sugars into molecular-weight groups was emphasizedz2using the simple chromatographic apparatus of Tswet.t.9In a model experiment, a mixt,ure of a mono-, a di-, and a tri-saccharide and limit dextrin was adsorbed on t8hecarbon column ; the monosaccharide was developed off the column with water, and the disaccharide with 5 % ethanol. The operation of this flowiiig chromatogram was facilitated by use of an automatic supplier of developer and a large, laboratory-scale, automat,ic effluent>collector.23n'umerous effluent fractionators have been proposed for column c h r o m a t ~ g r a p h y .24~ ~The . successful application of these devices was (22) R . I,. Whistler and D. F. Durso, J . Anz. Chem. SOC., 72, 677 (1950); 74, 5140 (1952). (23) D. F. Durso, E. D. Schall and R. L. Whistler, Anal. Chem., 23, 425 (1951); J . L. Hickson and R. I,. Whistler, ibid., 26, 1425 (1953). (24) I,. Hough, J. K. N . Jones a n d W. H. Wadinan, . J . Chwm. SOC.,2511 (1949); D. M. P. Phillips, Natctrc, 164, 545 (1949); S. 9. Itnndall arid A . J. 1'. Martin, Biochem. d . (I,ondon), 44, €'roc.. ii (1941)); A . R. Gilson, C h e n i i s f r g Le. I r ~ d z r s l l y29, , 155 (1951); R . A . Grcirit a n d S. R.. Stitch, ibitl., 230 (19.51); J . 0 , Harris, ibid., 255 (1951); I,. A. Boggs, I,. S. Caendct, M. Dulmis and F. Sniitli, Annl. Chem., 24, 1148 (1952); J.Etlelnian and It. V. Martin, Bioch,ern. J . (London), 60, Proc. xxi (1952); A . T.Carlander and S. Gardell, Arkiv Kemi, 4, 461 (1952); J . E. Varner and W. A . Bulen, J . Chem.

63

COLUMN CHROMATOGRAPHY O F SUGARS

exemplilied with a chromatogram of an acid hydrolyzate of an alpha Schardinger cleutrin, using gradient development with ethanol in water (the concentration of the ethanol in water was changed gradually and continuously from 0 to 20 % during the course of the development,; see Fig. 8).2b

0.4-

0

50

10

80

Fraction no.

FIG.8.-Chromatography of a Mixture of Mono- t o Hexa-saccharides from an Acid Hydrolyzate of a Schardinger Dextrin. [The Adsorbent was Carbon (Darco G-60) Pretreated with 1% Ethanolic Stearic Acid; the Developer was Gradually Changed from 0% to 15% Ethanol.]

The unusual utility of this adsorbent has been demonstrated by its role in the preliminary isolation of isomaltose,26(a)maltotriose,26(a)maltotetraose,26(b)and panose26(n) from the hydrolytic products of starch; in the preparation of D-xylo-biose, -triose, -tetraose, -pentaose, -hexaose, and -heptaose from a partially hydrolyzed xylanZ7;in the resolution of bimolecular dianhydrides of L-sorbose28(a) and D-fructose2s(b) ; in an improved procedure for the preparation of s t a c h y ~ s e and ~ ~ ; in the preparation of a series of maltodextrins (as high as maltoheptaose) from the partial, acid Educ., 29,625 (1952) ; H. J. Huisman and S. A. Krans, Chent. Weekblad, 48,1007 (1952) ; R. J . Dimler, J. W. Van Cleve, Edna M. Montgomery, L. R. Bair, F. J. Castle and J. A. Whithead, Anal. Chenz., 26, 1428 (1953); E. S. Sandcrson, ibid., 26, 944 (1954); D. Fraser, ibid., 26, 1858 (1954). (25) A. Tiselius, Endeavour, 11, No. 41, 13 (1952); R. S. Alm, R. J. P. Williams and A. Tiselius, Acta Chem. Scand., 6, 826 (1952); R . S. Alm, i b i d . , 6, 1186 (1952). (26) (a) M. L. Wolfrom, A. Thompson, A . N . O’Neill and T . T . Galltowski, J . Am. Chem. SOC.,74, 1062 (1952); A. Thompson and M. L. Wolfrorn, i b i d . , 74, 3612 (1952); 73, 5849 (1951); (b) R. L . Whistler and J. L . Hickson, zbid., 76, 1671 (1954); Edna M. Montgomery and F. B. Weakley, J . Assoc. O j i c . Agr. Chemists, 36, 1096 (1953). (27) R. L . Whistler and C.-C. T u , J . A m . Chem. SOC.,73, 1389 (1951); 74, 3609 (1952); 76,645 (1953); R. L. Whistler, J. Bachrach and C.-C. Tu, i b i d . , 74,3059 (1952). (28) (a) M. 1,. Wolfrom and H. W. Hilton, J . A m . Chem. Soc., 74, 5334 (1952); (b) B. Wickberg, A d a Chem. Scand., 6, 961 (1952). (29) M. L. Wolfrom, R C. Burrell, A. Thompson and S. S. Furst, J . A m . Chern. Soc., 74, 6299 (1952); R. A. Laidlaw and Clare B. Wylam, J . Chem. SOC., 567 (1953).

64

W. W. RINKLEY

hydrolysis of pot,ato amylase."" Carbon-column chromatograms of the sugars of Cmbilicaria pustulnta, when developed wit>h 1, 15, and 50% aqueous ethanol, yielded D-arabitol, D-marinitol, umbilicin [3-0-(@-~galactopyranosy1)-o-arabit,ol],a ,a-trehalose, and sucrose.31 The carboncolumn analysis of the acid hydrolyzates of the hemicelluloses of Scots pine and black spruce showed the presence of D-xylose, ~-arabinose,D-mannose, D-galactose, L-rhamnose, L-fucose, D-galacturonic acid, ~-xylose-(2-+ I ) 4-~-methyl-~-g~ucosiduronic acid, and 4-O-methyl-~-g~ucuronic acid.32 A crystalline nitrogen-containing tetrasaccharide was obtained from human milk by carbon c h r ~ m a t o g r a p h yUse . ~ ~ of this adsorbent made possible the isolation of certain oligosaccharides from enzymic transglycosidations using sucrose in the substrate; some examples are k e ~ t o s earid ~ ~n e o k e ~ t o s ewith ~~ yeast invertase, a new trisaccharide with T a k a d i a ~ t a s e and , ~ ~ a reducing O-(D-fructosyl)-D-glucoseamong the products from sucrose, D-ghcose, and yeast invertase.37 The salient chromatographic advantages of carbon are general availability, low cost, keen selectivity, and good capacity. Less attractive features of this adsorbent are that it is disagreeable to handle and that it usually requires some pretreatment before use. Furthermore, the rate of solvent flow through columns of finely divided carbon is too low unless up to an equal weight of an inert diluent38is added; a corresponding decrease in column capacity results from this addition. The last objection is met by employing columns composed only of carbon capable of passing through a 40- or 60-mesh screen but retained by ail 80-mesh screen.39 2. Fuller's Earth Clay

Zones formed in the chromatography of colorless substances on columns of colorless or light-colored adsorbents may be detected by streaking the (30) W. J. Whelan, J . M. Bailey and P. J. P. Roberts, J . Chem. SOC.,1293 (1953). (31) B. Lindberg and B. Wickberg, Acla Chem. Scand., 7 , 140 (1953). (32) A. R . N. Gorrod and J . K . N . Jones, J . Chem. Soc., 2522 (1954). The acidic disaccharide was also resolved chromatographically from an acid hydrolyzate of hemicellulose-B from corn cob; R. I,. Whistler and L. Hough, 6 .A m . Chem. Soc., 76, 4918 (1953); R. L . Whistler, H . E. Conrad and L. Hough, ibid., 76, 1668 (1954). (33) R . IZuhn, Adeline Gauhe and H . H. Baer, Chem. BeT., 86, 827 (1953). (34) N . Albon, D. J . Bell, P. H. Blanchard, D. Gross and J. T . Rundell, J . Chem. SOC.,24 (1953). (35) D. Gross, P. H. Blanchard and D. J . Bell, J . Chem. Soc., 1727 (1954). (36) J . S. D. Bacon and D. J . Bell, J . Chem. SOC.,2528 (1953). (37) D. J. Bell and J. Edelman, J . Chem. S O C .4652 , (1954). (38) Celite No. 535, a diatomaceous filter aid, is a product of the Johns-Manviile Co., New York, N. Y . , and is often used for this purpose. (39) R . L. Whistler, Science, 26, 899 (1954).

COLUMN CHROMATOGRAPHY OF SUGAHS

65

extruded columii with a suitable indicator (streak reagent).40The potent'ialities of this method are great. The rapid evaluation and selection of adsorbeiit,s and developers are made possible, using only milligram quantities of adsorbate. Aqueous, alkaline, potassium permanganate4' is often employed as a reagent for the detection of sugars and their colorless derivatives. The st,reaked adsorbent column is cut into sections as indicated by the reagent. The streak is scraped off the column section, and the zone is then eluted with a solvent mixture containing 10 to 20% more water than t.he developer employed in the chromatogram. The recovered adsorbate is oft,en obtained in crysballiiie form. The suitability of fuller's eart>h for the chromatography of carbohydrates was demonstrated by the establishment of an adsorpt,ioii series which included nearly 100 sugar subst,ances (see Table 11).43 The developers mere aqueous 2-propanol and aqueous et,hanol; the water content of these mixtures was gradually increased in order to develop more st'rongly adsorbed carbohydrates, particularly sugar Here, the developing effect of aqueous ethanol was the reverse of its role in the chromatography of sugars 011 carbon.45 In general, the adsorption of sugars on clay did not present a definite pattern. The adsorptive bond of the members of the D-glucose-sucrose-raffinose series did become stronger with an increase of molecular weight ; glycerol, erythritol, ribitol, and allit,ol behaved similarly. Glycosides are held more (30) 12. Zechmeister, L. de Cholnoky and (Mlle.) E. Ujhelyi, Bicll. soc. chim. bid., 18, 1885 (1936). (41) W. H. McNeely, W. W. Binkley and M. L. Wolfrom, J . A m . C'hem. Suc., 67, 527 (1945). (42) These clays are hydrated magnesium aluminum silicates containing some iron and calcium. (43) B. W. Lew, M. L. Wolfrom and R . M. Goepp, Jr., J . A m . Chem. SOC.,68,1449 (1946). A suitable fuller's earth clay was Flores XXX; it was produced by the Floridin Co., Warren, Pa. Other selective clays were Floridin XXX (Floridin Co., Warren, Pa.), Types A and AA Attapulgus Clays (Attapulgus Clay Co., Attapulgus, Ga.), Bleaching Clay 260 (Industrial Minerals and Chemical Co., Berkeley, Calif.), and J. Neutrol (Filtrol Corp., Los Angeles, Calif.). (44) Often, sugar acids can be detected with Congo Red indicator, and ascorbic acid with 2,6-dichlorophenolindophenol. (45) The detection of sugar alcohols in chromatograms developed with aqueous ethanol was difficult because the streak reagent, alkaline potassium permanganate, was rapidly decolorized. This difficulty was largely overcome in small exploratory chromatograms by the use of aqueous tert-butyl alcohol as the developer. The rate of flow of these solvents through fuller's earth clay was very slow and this developer was unsatisfactory for larger columns of this adsorbent. These columns were developed with aqueous ethanol, and extruded. They were wrapped with aluminum foil, leaving exposed t o the air an area, 1 cm. wide, from the top t o the bottom of the adsorbent column. Sufficient solvent had evaporated from the exposed surface in 18 hours to permit the use of alkaline permanganate.

66

W . W . BINKLEY

TABLE I1 Chroinatoyraphzc Adsoi ptzon Series,u on Fzrllci 's E a r t h Clay, oJ Curbohydrulcs and Some Related Substances43 Class I (10 cc. of 70% ethyl alcoholb) Potassium acid D-glwarate, D-glucosamine hydrochloride, 1)-galactosamine (chondrosaniine) hydrochloride (dertro)-Tartaric acid D-Gluconic acid, ammonium wgluconnte, sodium D-gluconate D-Arabonic acid, L-arabonic acid, potassium D-arabonate, citric acid D-Mannonic acid a-D-Galacturonic acid Class ZI {lo cc. of 90% ethyl alcohol) Lactitol, melibiitol L-Iditol, inyo-inositol (m. p. 225"),stitchyose, Schardinger a-dextrin Lactose D-Glucitol, dulcitol, L-perseulose, D-perseulose, Schardinger 8-dextrin, (Zeao)malic acid Raffinose Xylitol D-Mannitol, u-talitol, gentiobiose Class IZI (5 cc. of 90% ethyl alcohol) D-Gulose Melibiose, D-mannose, wribose, 3,6-anhydro-~-glucitol,u-psicose Cellobiose l-Deoxp-o-glucitol Ascorbic acid D-ArabitoI Class Ik' (4 cc of 95% ethyl alcohol) Maltose, u-galactose, D-fructose, melezitose, u-)r2unno-heptrIlose, trehalose, D glycero-D-yulo-heptose L-Fucitol, turanose, 1,5-anhydro-u-niannitol, 1,4-anhydro-~-mannito1, D-lyxose D-Rhamnitol, L-rhamnitol, 2-deoxy-~-"glucitol,"D-gluco-heptdose L-Arabinose, D-arabinose, ribitol, quercitol, diethylene glycol (2,2'-oxydiethanol), dipentaerythritol Sucrose, erythritol, L-altrose ~-G~ucurono-2,6-lactone L-Sorbose, succinic acid Anhydroenneahepti to1 n-Glucose, glycerol Class 1' (5 cc. of 90% isopropyl alcohol) L-Allose 1,4:3,6-Dianhydro-1)-mannitol, amygdalin ~-D-Ga~ncto-metasitccharinic acid 3-Hydroxypropyl 8-o-glucopyranoside rJ-Fucose

BT

COLUMN CHROMATOGRAPHS O F SUGARS

TABLEI1 (Contznued) - ______-____. ~-

_

_

~

Class VZ (6 cc of 97% isopropyl nlcoliol)

Methyl cu-i,-glucopyranosidc, methyl ~-u-glucopyranoside Methyl a-D-mannopyranoside, D-xylose, D-glucono-l,5-lactone 1,4-Anhydro-o-gluc~itol,dipropylene glycol, pentaerythritol, 1,5:3,6-dianhytIrou-mannitol, 1,4:3,6-dianhydro-~-glucitol D-Clucono-l , 4-lactone Class V I I (4 c c . of 97% isopropyl alcohol) L-Rliamnose, salicin Ethylene glycol (1,2-ethancdiol) Propylene glycol (I ,2-propnnediol) 1’11lori x i n 1 , 4 : 3 ,A-Dianhydro-~-idi to1 a Arranged in decreasing order of adsorptive strength. Adsorbent: 1.68 g. of Florex XXX43/Cclite38 (5/1). Dimensions of adsorbent column: 0.9 X 6 cm. of adsorbent. Adsorbate solution: I nig. in 0.5 cc. of developer shown. Developer: noted following the class heading. Prepared by adding 30 cc. of water t o 70 cc. of absolute ethanol. Other dilutions were made similarly, except, t h a t 95% ethyl alcohol was the axeotrope.

loosely than sugars, sugar acids more tightly. Columns of d a y are difficult, to extrude from the usual, straight, glass tubes; they are removed more readily from metal tubes or from glass tubes tapered uniformly and gradually outward from the bottom to the top.46The rate of developer flow through clay columns was slow; the rate was improved by the admixing of a filter aid.38The flow rate can be increased &fold when the clay is air-blown (to remove the finer particles) and the developer is subjected to a pressure of 120 Ibs. per sq. in.47The keen selectivity of clay was exemplified by the separation and isolation in gram quantities (by the extrusion method) of two new di-D-fructose dianhydrides, D-fructopyranose-Dfructofuranose 1,2’:2, l’-diaiihydride4* and a second of unestablished ~ t r u c t u r e The . ~ ~ interconversion, by heat, of D-fructose tlo D-glucose was discovered with the aid of extrusion, clay chromatography. bo 2,3-0-Isopropylideue-D-fructose was isolat,ed chromatographically from partially (46) Ref. 7(g), p. 7 . Tapered glass tubes are produced by the Scientific Class Co., Bloomfield, N . J. (47) D. F. Mowery, Jr., J . Am. C h ~ m SOC.,73, 5047 (1951). Heavy-walled Pyrex tubes were needed for these chromatograms. (48) M. L. Wolfrom and M. Grace Blair, J . Am. Chem. SOC.,70, 2046 (1948); M. I,. Wolfrom, W. W. Binkley, W. I,. Shilling and H . W. Hilton, ibid., 73, 3553 (1951). (49) M. L. Wolfrom, H . W. Hilton and W. W. Binkley, d Am. Chern. SOC.,74, 2867 (1952). (50) M. 1,. Wolfrom and W. L. Shilling, J . Am. Chem. SOC.,73, 3557 (1951).

-

68

W. W. BINKLEY

hydrolyzed “ij-diacetoiie-D-fru~tose,” showing the structure of thc latter to be 2,3:4,5-di-~-isopropylidene-~-fructopyraiiose.~~ Clay has been used effectively in the flowing or elutJioiichromatography of sugars. The residual sucrose was removed almost quantitatively from beet molasses, arid in 73 % yield from cane, blackstrap molasses.52Subscucose

0.19

0.16

u) u)

-

2

.+.

0

@h

2 L

g o.oe u

0.04

0.01

t

I

I

15

39

54

89

Fraction no.

FIG.9.-Chromatography on Fuller’s Earth Clay of a Fraction from Cuban, Blackstrap Molasses. [The Developer was Azeotropic Ethanol from X t o Y , and Ethanol/Water (90/10) from Y t o Z.]

quently, crystalline D-glucose and myo-inositol, and zones containing Dfructose and D-mannitol, were isolated from cane molasses (see Fig. 9); a similar chromatogram of its yeast-fermentation residue revealed the presence of D-glucose, sucrose, erythritol, D-fructose, D-arabitol, and (51) M. L. Wolfrom, W. L. Shilling and W. W. Binkley, J . Am. Chem. Soc., 74, 4544 (1950). (52) W. W Binkley and M. L. Wolfrom, J. Am. Chem. Soc., 69, 664 (1947); M. L. Wolfrom and W. W. Binkley, U. S. Pat. 2,504,169 (1950); Chem. Abstracts, 44, 6180 (1950).

69

COLUMN CHROMATOGRAPHY OF SUGSRS

D-mannitol (see Fig. 10).53 Crystalline D-fructose was prepared chromatographically from invert sugar (acid-hydrolyzed sucrose) .64 Fuller's earth clay is capable of some unusual and difficult separations of sugars, and because of its comparatively light color this adsorbent is adaptable to extrusion, streak-reagent chromatography; furthermore, it is cheap. Most organic solvents pass rapidly through columns of clay. The

D-Frucrose D-Arabitot

o . 0 6 ~, x Erythritol

5

10 13

D-Mannitol

19

29

35

49

Fraction no.

FIG.10.-Chromatography, on Fuller's Earth Clay, of a Fraction from the NonFermented (by Yeast) Residue of Cuban, Blackstrap Molasses. [The Developer was Aeeotropic Pzthanol from X t o Y, and EthanollWater (90/10) from Y to Z.]

addition of wat,er to these solvents markedly decreases tjhe rate of their percolation, and so the admixture of 15 t,o 20 % of filt,eraid t30t,he adsorbent is desirable. The capacity of fuller's eartjh clay is low, and some pretreat'ment is required when t>heclay is t,o be used for isolation work. Whereas the adsorptive propert,ies of clays from different areas are variable, this adsorbent from any particular geographical locatJion is except.ionally uniform. The poor extrusion propert.ies of the makrial are partially overcome by the (53) W. W .Binkley and hl. L. Wolfroni, J . A m . Chevr. Soc., 72, 4775 (1950). (54) D. F. Mowery, Jr., J . h ~ Cheni. . SOC.,73, 5049 (1951); D. F. Mowery, Jr., and G. R . Ferrante, i b i d . , 76, 4103 (1954), for the separation of methyl u-galactoside anomers.

70

W. W. BINKLEY

use of metal, or tapered-glass, chromatographic tubes. Certain inorganic substances are leached from the adsorbent during the elution process, particularly when water-rich solvents are employed, and these form troublesome suspensions. They are removed from an aqueous solution of the adsorbate by treatment with decolorizing carbon and passage through acid-washed asbestos under diminished pressure, followed by gravity filtrat,ion t,hrough hardened filter paper.49

3. Carbohydrates T s v ~ e t t himself, ,~~ recognized the potentialities of powdered sucrose as a chromatographic adsorbent, and employed t,his sugar in his investigations on chlorophyll. Soon after,56the branched form (smylopectin) and the h e a r form (amylose) of potato starch were separated on fibrous celliilose (washed cotton) ; corn starch has also been so f r a c t i ~ n a t e d Interestjingly, .~~ the linear form is the more st,roiigly adsorbed although its molecular weight is lower. Amylose may be able to form with t,he cellulose, a t best,, some t3ype of weak inclusion complex.58 Thus, the removal of amylose from amylopectin is not complete,59and some modifications of this adsorption procedure likewise were unsuccessful.60 The behavior of cellulose nitrate and of non-carbohydrate polymers, on carbon, is similar.6‘ The advent of modern, filt>er-paperchromatography,62and its extension to ~ u g a r s , 6have ~ produced a powerful, qualitative, analytical method. Often, only microamounts of adsorbat,es are handled by this procedure; but larger yuantities of resolved materials were needed for some quant,itative separations, isolations, arid proper identifications, and columns of tightly packed, powdered cellulose were found to be 64 A mixture containing 50 mg. each of L-rhamnose, D-ribose, L-arabinose, and D-galaCtOSe was adsorbed on a 12 by 1.25 in. (diam.) column of powdered cellulose7~ prewetted with t,he (55) RI. Tswett, Y i p o g r . Warshawskago iitsch,ebnago Okrwgn, Warsaw, 1910 (in Russian). (56) C. Tariret,, C o w p t . rend., 168, 1353 (1914). (57) F:.Pacsu :tnd J. W. Mullen, 11, J . A m . Cheni. Soc., 63, 1168 (1941). stsJlizat,ion,” i i i “Technique of Organic Chemistry,” A. (58) R. S. Tipson, ‘‘ Weissherger, ed., Interscience Publishers, I n c . , New York, N. Y., Vol. 111,2nd Edition, i n press. (59) T. J . Schoch, Arlaances in Corhohydrate Chenf,.,1, 247 (1945). (60) G . A. Gilbert, C. T. Greenwood and F. J. H y h r t , J. Chew. Soc., 4454 (1954). (61) 8. Claesson, A r h i v K e m i , Jlineral. Geol., 26A, No. 24 (1948). (62) R . Corisden, A . H. Gordon nnd A. J. P. Martin, Bioche,ni. .I. (London), 38, 224 (1048); A . J . P. Martin, P r i x N o b r l , 110 (1952). (63) S. M . Part,ritlge, .Vatitre, 168, 270 (1946). See G. 1;.Komkahany, Adzinnces in Cnrhoh!/drnta C h e n ~ .9, , 303 (19.54). (64) I,. Hough, J . K . N. Jones iind W. I€.Wildman, ;Vnfur.e,162,448 (1048). (65) A commodity of H. Reeve Angel and Co., Inc., 52 I3uane St., New Yorlr 7 , N.T.

COLUMN CHROMATOGRAPHY O F SUGARS

71

developer, 1-butaiiol saturated with water coiitaiiiiiig 1 % of ammoilia. The flowing chromatogram was employed, and the resolved, cystalline sugars were recovered in 95 % yield from the column effluent. Columns of wet, powdered cellulose are extruded readily i t ith compressed air, hut no suitable streaking reagent is available at present. Fortunately, the chromatographic behavior of powdered cellulose is identical with that of f lterpaper strips and sheets. The flowing chromatogram of this adsorbent, using an automatic, effluent-fraction collectorz4 coordinated with paper-strip analysis, has produced a strong tool. ITsually, the rate of' solveiit percolation through this adsorbent is sufficiently rapid that neither the application of pressure (or vacuum) nor the addition of an inert dilueut (filter aid) is required. The method is also very effective in the resolution of TABLE111 Flowing Chromatogram, on Powdered Cellulose, or 1.6 g. of the Sugars from the Incomplete Hydrolysis or the Gum of the Sterculia setigera Trees@(*) Cofiiponrnls

Yield, mg

I I1 111 IV V

15 413 145 373 640

a ketose L-rhamnose wtagatose and an aldose wtagatose, wgalactose, and an aldose 11-galactose

Total, 1,586 a

The developer was 1-butanol, saturated with water.

mixtures of methylated sugars (which is discussed in detail in a later Section). The broad applicability of this method is already apparent in the successful elucidation of the structure of certain naturally occurring, plant polysaccharides. Powdered-cellulose chromatography of the sugars from the incomplete, acid hydrolysis of t,he gum from the Sterculia setigera tree yielded pure, crystalline L-rhamnose hydrate and D-galactose, as well as D-tagatose (also crystalline, see Table III)66(a); other methods showed the presence of D-galacturonic acid residues. Pure crystals of' L-rhamnose and D-xylose were obtained by powdered-cellulose chromatography of a hydro(66) (a) E. L. Hirst, L. Hough and J. K. N. Jones, Nature, 163, 177 (1949); J . Chem. Soc., 3145 (1949); (b) P. Andrews, D. H. Ball and J . K. N . Jones, ibid., 4090 (1953),

for peach and cherry gums; (c) P. Andrews and J. K. N . Jones, ibid.,1724,4134 (1954), for lemon and golden apple gums; (d) E. L. Hirst and A . S. Perlin, ibid., 2622 (1954), fov the gum of Acacia pycn.antha.

72

W. W. BINKLEY

lyzed mucilage from flax seed; L-arabinose was also 67 The acidhydrolyzed mucilage of the slippery elm (Ulmus ,fuZva,) was found to contain ~-rhamnose-(2+ 1) D-galactosiduronic acid and a mixture of sugars, resolved by this chromatographic method int>o it,s components, D-galactose, L-rhamnose, and 3-O-methyl-~-galactose.~~ The application of the method, using a hydrocellulose, to the acid hydrolyzates of several galactomannans (from clover,'j9 and fenugreek seed70) gave crystalline D-mannose and D-galactose; n-glucose was obtained, in addition to these sugars, from certain Iris The residues of D-galactose, Larabinose, and 4-O-methyl-~-glucuronic acid (isolated by another procedure) were shown, using powdered-cellulose columns, to be present in gum myrrh72;and D-xylose and D-galactose residues were found to be components of the polysaccharide of tJhefresh water alga, N o s l o ~An . ~ ~L-arab. inopyranose disaccharide [3-O-(P-~-arabinopyranosyl)-~-arabinose] was separated from the hydrolytic products of larch g gal act an.^^ Chromatography, on powdered hydrocellulose, of the sugars from the hydrolyzed, animal polysaccharide of frog-spawn mucin led to the identification of L-fucose, D-mannose, D-galactose, D-glucosamine, and D-galactosamine among the components (see Table IV).75 Further evidence of the diversity of chromatographic applicability of hydrocellulose was shown in its ability t o separate a hexitol from a hexose; crystalline D-arabitol and D-galactose were isolated from acid-hydrolyzed umbilicin of Umbilicaria p ~ s f u l a t a . ~ ~ The powdered-cellulose column has performed valuable service in the study of sugar synthesis. DL-Xylose was resolved from the products of the action of heat and alkali on paraformaldehyde; DL-xylose and DL-arabinose were isolated after the action of alkali a t 20" on a mixture of glyceraldehyde and gly~olaldehyde~';subsequently, DL-ribose (identified as the p-tolylsulfonylhydrazone7*) was obtained79 using a column operating a t 60". (67) D. G. Easterby and J . K. N. Jones, Nature, 166, 614 (1950). (68) L. Hough, J. K. N . Jones and E . L. Hirst, Nature, 166, 34 (1950); E.L. Hirst, I,. Hough and J . K. W. Jones, J . Cheni. SOC.,323 (1951); for okra mucilage, R . L. Whistler and H. E. Conrad, J . A m . Chem. SOC.,76, 1673, 3544 (1954). (69) P. Andrews, L:Hough and J. K. N. Jones, J . A m . Chem. SOC.,74, 4029 (1952). (70) P. Andrews, L. Hough and J. K. N. Jones, J . Chem. SOC.,2744 (1952). (71) P. Andrews, L . Hough and J. K . N . Jones, J . Chem. SOC.,1186 (1953). (72) L. Hough, J . I(. N. Jones and W. H. Wadman, J . Chhem. SOC.,796 (1952). (73) L. Hough, J . I<. N. Jones and W. H. Wadman, J . Cham. Soc., 3393 (1952). (74) J. K. N. Jones, J . Chem. S O C . 1672 , (1953); Chemistry & Industry, 954 (1952). 2136 (1950). (75) B. F. Folkes, R. A . Grant and J. K. N. Jones, J . Chem. SO C., (76) B. Lindberg, C. A . Wachmeister and B. Wickberg, Acta Chem. Scand., 6, 1052 (1952). (77) L. Hough and J . K. N . Jones, J . Chem. SOC.,1122 (1951). (78) L. Hough and J . K. N. Jones, J . Chem. Soc., 3191 (1951). (79) J. N . Counsell, L. Hough and W. H. Wadmzn, Reseaich (London), 4,143 (1951).

COLUMN CHROMATOGRAPHY OF SUGARY

73

n-threo-Pentulose (D-xylulose, tentatively identified as the osazone) was found chromatographically to be among the products of the action of a pea enzyme on glycolaldehyde and triose phosphate (prepared from D-fructose 1 ,G-diphosphate)*O;similarly, 6-deoxy-D-fructose and ti-deoxyL-sorbose resulted from m-lactaldehyde,BOsedoheptulose from D-eryt>hrose,B" 5-deosy-~-threo-pentulosefrom acetaldehyde and a triose phosphate,81 and D-ido-heptulosan from D-xylose.82 The powdered-c*ellulose, flowing chromatogram of an unfermentable residue from the mixture obtained by the action of heat, on D-fructose yielded a sirup containing a ketose having a mobility greater than that of D-fructose; i t may be ~-psicose.*~ D-Psicose (identified as the phenylosazone TABLEIV 17lowing Chrornntograrn on Powdered Cellulose of the Sugars from the Hydrolysis of 1.082 y. of Frog-spawn Mucin7$ Ej4uent fractiona

Yield, mg.

I I1

59 16

111 IV V VI

15 148 108 -

Component

L-fucose o-mannose (identified as the phenylhydrazone) amino sugar o-galactose D-glucosaminc o-galactosamine (chondrosamine)

1

Physicel form o//raclion

crystalline sirup sirup crystalline crystalline hydrochloride crystalline hydrochloride

~

Total, 346 0

The developer was 1-butanol, half saturated with water.

and as 1,2 : 3 ,4-di-O-isopropylide11e-~-psicofuranose)was found among the products of the action, a t 37") of ammonia on ~ - g l u c o s e . ~Chromato~(~) graphic assay of the products formed by the action of this base on maltose and lactose revealed their partial conversion to m a l t u l o ~ e ~and ~ ( ~lactu) 1 0 ~ e , ~respectively; ~(~) ammoniacal melibiose yielded melibiulose, ti-O-(c~-~galactopyranosy1)-p-D-mannose, D-tagatose, and ~ - g a l a c t o s e . ~ ~ ( ~ ) A fine example of the rapacity and selectivity of powdered cellulose is (80) L. Houyh and J . K. N Jones, J . Chenr. Sac., 4047 (1952); 4052 (1952); 342 (1953). (81) P. A ,J. Gorin, L. Ho~ighand J . K. N . Jones, d . Chem. Sac ,2140 (1953). (82) J. K. N. Jones, J . Chern. Sac., 3643 (1954). (83) L. Sattler, F. W. Zerhan, G . I,. Clark, C.-C. Chu, N . Albon, D. Gross and H. C. S. de Whalley, Ind. Eng. Chem., 44, 1127 (1952). (84) (a) L . Hough, J . K. K.Jones and E. L. Richards, J . Chem. Sac., 2005 (1953); (b) 295 (1954).

74

W. W. BINIiLEY

the quantitative determination of raffinose in raw-beet sugars.85h chromatogram of 20 g. of the raw sugar, on a 30 by 7.5 cm. (diam.) column of adsorbent, sharply separated 0.5 % of raffinose (as the pentahydrate) from the very large amount of sucrose and extremely small quantity of stachyose; the developer was 2-propanol/l-butanol/water(7/1 /2).a6 Starch columns were found suitable for the separation of certain monos a c c h a r i d e ~These . ~ ~ columns, pretreated with aqueous I -butanol and the developer 1-butanol/l-propanol/water (4/1/1), and developed under

2oot

a

c

c

3

L

150-

L L

f

-

E

e

l

100P aJ

: 5 Fn

x

50 -

f.n

01

I

I

I

20

50

100

Effluent volume, mi.

FIG.11.-Chromatogram, on a 20 x 0.9 cm. Potato-Starch Column, of (a) L-Rhamnose, (b) L-Fucose, (c) D-Ribose, (d) D-Xylose, (e) D-Mannose, ( f ) D-Glucose, and (g) o-Galactose. [The Developer was 1-Butanol/l-Propanol/Water(4/1/1).]

pressure, \wre capable of sharp separations (see Fig. 11). The capacity of starch was found to be lorn; the maximum weight-ratio of adsorbate to adsorbent was ca. 1:1000. The coordinated use of powdered-cellulose and starch chromatography has far-reaching potentialities. Cellulose powder has moderately good capacity, and the rate of flow of developer through its columns is good if slight pressure is applied. Filterpaper strips and the cellulose powder have identical adsorptive properties, (85) D. Gross and N . Albon, Arinlyst, 78, 191 (1953). (86) Unless otherwise noted, solvent ratios will be expressed on a volume basis. (87) S. Gardell, Acta Chem. Scand., 7, 201 (1953). Potato starch was conditioned by suspension in a mixture of 1-butanol and water; the total moisture in the system was in the weight ratio of waterlanhydrous starch:3/10.

COLUMN CHROMATOGRAPHY OF RTJGARS

75

and they form a useful chromatographic partnership. The progress of colored and of fluorescent (iii ultraviolet light) suhstaiices, moving on cellulose, is readily followed. Chromatographic cellulose is expensive, and it usually requires some treatment before use. Starch is capable of separating closely related sugars. I t is inexpensive, but t,he rate of flow of developer through starch columns under pressure is low, and the adsorbate capacity of starch is very low. This carbohydrate must be conditioned and prewashed before it is suitable for chromatography. 4. Ion-Pxchange Resins

Serious thought and a considerable amount of effort have been devoted to the utilization of ion-exchange resins in the sugar industry.88Some success has been achieved in the beet industry, but the situation is much less certain in cane-sugar production. The principal objectives are the retention by the resins of the ionic (organic and inorganic) impurities and the maximum recovery of sucrose. Quite different is their use in the separation of sugars. A column of an anion-exchange resin (Amberlite IR-400,s9 regenerated with sodium hydroxide) retained, in general, reducing sugars when developed with water. Sugar alcohols and methyl a-D-glucopyranoside were not adsorbed,g0 and sucrose was only partially retained. The sugars were completely eluted by 10 % sodium chloride. The chromatographic separation of sugar mixtures was not attempted. The polyhydroxy functions of sugars and related substances permit the formation of complexes with the borate ion of boric acid and borate salts.g1 These complexes are readily formed in low concentrations of borate and display different adsorptive strengths when added to a column of an anionexchange resin (Dowex-la2) regenerated to the borate form.93 The chromatogram was developed first with 0.018 M sodium borate; D-fructose and D-galactose, in succession, appeared in the effluent, followed by D-glucose, which required 0.03 M sodium borate. The adsorptive strength of nonreducing sugars is dependent upon their molecular weights, stachyose being more strongly bonded than raffinose, and raffinose more tightly held than (88) R . Kunin and R . J. Myers, “Ion Exchange Resins,” John Wiley and Sons, Inc., New York, N . Y., 1950; R . Kunin, Chem. Eng. News, 32, 3046 (1954). (89) A product of the Resinous Products Division of Rohm and Haas Co., Philadelphia, Pa. (90) S. Roseman, R . H. Abeles and A . Dorfman, Arch. Biochem. and Biophys., 36, 232 (1952). (91) J. Boeseken, Advances i n Carbohydrate Chem., 4, 189 (1949). (92) A product of the Dow Chemical Co., Midland, Mich. (93) J. X. Khym and L. P. Zill, J. A m . Chem. Soc., 73, 2399 (1951).

76

W. W. RINKLET

The effect of a functioiial group was even greater, the adsorptive strength of D-fructose being higher than that of the oligosaccharides. A mixture of the pentoses, D-ribose, D-arabiiiose, and D-xylose, was resolved .~~ by Dowex-1 when developed with 0.015 M potassium t e t r a b ~ r a t e The fine selectivity of this method was established by its successful application to a series of disaccharides (see Table V). The progress of these chromatograms was followed quantitatively (for total sugar) with anthrone963 y7 and orcinol r e a ge nt ~ .~The 8 individual sugars were identified by paper chromatography after the removal of potassium ions with a cation-exchange resin. The isolation of pure sugars was completed after the formation, and removal under diminished pressure, of the boric acid as its methyl ester.99 This method is also applicable to sugar alcohols, as exemplified by the TABLE V Chromatography; on an Anion-exchange Resin ( D o w e x - I ) , of Certain DisaccharidesQs Disaccharide (order of appearance in efluenl)

Volume required lo produce maximum concentralion in eBuent, 1nl.b

Sucrose Trehalose Cellobiose Maltose Lactose

175 370 660 750 X1@ ~

The developer was 0.005 M K2B,07 . 6 An amount of 10-25 mg. of each sugar waa added t o an 11 by 1 cm. (diam.) column of resin. a

separation of D-glucitol, galactitol and D-mannitol; to sugar acids in the resolution of D-galacturonic and D-glucuronic acids on Dowex-1,92 acetate or formate form, with aqueous acetic or formic acid as the developerloo; and to amino sugars in the chromatography of D-glucosamine and D-galactosSedoheptulosan is readily sepamine (as hydrochlorides) lol on D0wex-50.~~ arated from the more strongly adsorbed s e d o h e p t u l o ~ e . ~ ~ The chief disadvantage of the method is the unusually large volume of (94) G. R. Noggle and L. P. Zill, Arch. Biochem. and Biophys., 41, 21 (1952). (95) J. X. Khym and L. P. Zill, J . Am. Chem. Soc., 74, 2090 (1952). (96) R. Dreywood, Ind. Eng. Chem., Anal. Ed., 18, 499 (1946). (97) D. L. Morris, Science, 107, 254 (1948). (98) A. H. Brown, Arch. Biochem., 11, 269 (1946). (99) L. P. Zill, J. X. Khym and G. M. Cheniae, d . A m . Chenz. Soc., 76, 1339 (1953). (100) J. X. Khym and D . G. Doherty, J . A m . Chem. SOC.,74, 3199 (1952); J. X. Khym and W. E. Cohn, ibid., 76, 1818 (1954); J. X. Khym, D. G. Doherty and W. E. Cohn, ibid., 76, 5523 (1954) for the chromatography of o-ribose phosphates. (101) S. Gardell, Acta Chem. Scand., 7,207 (1953); S . Gardell and S. Rastgeldi, ibid., 8, 362 (1954); E. Drake and S. Gardell, Arkiw Kemi, 4 , 469 (1952).

COLUMN CHROMATOGRAPHY OF S U G A R S

77

developer required in order to achieve the desired separations. The durutiori of the experiments is long, and the recovery of small amounts of separated sugars from large volumes of solution is tedious. TABLEV I Chromatographic Adsorption Series,= on Calcium Acid Silicutc, of Some Sugars and Related S r ~ b s t u n c e s ~ ~ ~ CInss Z (10 cc. of 90% dioxane) a-D-Galacturonic acid Lactose monohydrate 1,actitol Drilcitol Melezitose, raffinose pentahydrate, gentiobiose, D-glyccro-D-gubheptose Sucrose, maltose monohydrate, cellobiose, D-glucitol (sorbitol) D-Galactose, D-mannitol D -Glucose, o -fructose, I) -mannose, L-sorbose L-Fucitol L-Arabinose Class IZ (5 cc. of 90% dioxane) L - Fu cose o-Xylose, L-rhamnose monohydrate Methyl a-D-ghcopyranoside Class I11 (10 cc. of 90% diosane) D-glycero-D-gulo-Heptonamide o-Galactonamide, D-glnconamide Class ZV (5 cc. of 90% dioxane) D-Lyxonamide D-Ribonamide L-Fuconamide a Arranged in decreasing order of adsorptive strength. Adsorbent: 1.8 g . of Silene EF10Z/Celite38 (5/1) (wt. ratio). Dimensions of adsorbent column: 0.9 X 11 cm. Adsorbate solution: 2 mg. in 0.2 cc. of 90% dioxane. Developer: noted following the class heading.

5 . Calcium Acid Silicate In addition to fuller's earth clays,43certain synthetic silicates are useful in the column chromatography of sugars. Calcium acid silicate (Silene EF102) was, in its original e v a l ~ a t i o n , found ' ~ ~ to be suitable for the separation of such groups of sugars as pentoses from hexoses, and mono- from oligo-saccharides, following the general pattern of increased adsorptive strength with molecular weight (see Table VI) . The influence of molecular (102) A product of the Columbia Chemical Division, Pittsburgh Plate Glass Co., Barberton, Ohio. (103) I,. W. Georges, R . S. Bower and M. L. Wolfrom, J . Am. Chem. Soc., 68, 2169 (1946).

78

W. W. BINKLEY

weight was as usual overshadowed by that of the carhosyl group of the sugar arid ; in the series studied, tlic 1,oud het~\veeua-D-galacturouic acid and this silicate adsorbent was the st,rongest. C'ertain properties of this adsorbent, preseiit, a st,roiig rase for its c-hromatographic~use. ITnlikcfuller's eart'h clays, t,his syiit,hetic: silicate is readily estjrricled from st,rsight, glass tubes. The adsorlieiit is white, arid if it is used with t,he proper dcvelopers (aqueous dioxilrie or aqueous lert-butyl alcohol), dctectioii of slowly reduring sugar substances is possible with the permangallate-streak reagent,. Separate lot,s of t,his silicate varied greatly in adsorbent properties when test'ed under st,aiidard conditions with il mixture of D-glucaose and maltose."'" The adsorbents were gradually deact,ivated by continuous, water washing; they were reactivated by sodium hydroxide and certain salts. The pH of an aqueous susperisioii of the alkali-reactivat,ed adsorbent is 10-1 2; and so TABLE VII T h e pH of th,e Supernatant Liquors froni the M i x t u r e of 1 9. of Various Silicate Adsorbents with 16 ml. of Water'"" Adsorbent*

- ~ _ _ ~ _ _ _ _

I

__-_~

Hgdrnt,etl c:ilciiim :wid silic:ite, lot. K O . 1" Hytlr:Lt,ed c:ilciuu x i d silicate, lot, N o . 2" Hydrat.erI calcirini acid silicate, lot, N o . 3" Hydrated magnesium acid silicate,, Fuller's earth clay"

PfJ

10.3 10.4 10.6 8.3 8.6

a The adsorbents were mixed with :t filter aid in the weight-rtit,ios of 5/1. EF.'a2 c Magnesol.'o5 d Florex XSX.43

* Silene

the adsorbent is then unsuitable for the resolution of alkali-sensitive sugars. The alka1iiiit.y of untreated calcium acid si1icat.e was greater t,hsn that, of some other si1icat.e adsorbents (see Table 1'11). Sufficient. colloidal and soluble iiiorganir substances are roiitributed to the eluat,e by the adsorbent, during zoiie elution, t,hat a tedious procedure must, be followecl hi order t o recover the resolved adsorbate. These difficulties are not. encountered in t,he chromat,ography of sugar derivatives which require much less polar developers. The principal advantages of t,his adsorbentJ are it,s low cost, its \vhite color, the fairly high rate of solveiit percolation through it, and t,he ease of its ext,rusioii. (104) D . 0 . IIotTrnaii (with h l . I,. Wolftom), P1i.D. Ilissertation, The Ohio St,:it,e Universit,y, 1048. A n mioiirit of 0.2 ml. of :L solut>ioriof r)-glucosc (100 mg.) anti malt,ose (100 ing.) in a Inixcture of I nil. of w:iter :mtl $1 ml. of dioxane was added a t the t q of a 120 X 0 mm. (tlinm.) colu~niiof Silenc E;F/filter aid (.5/1, \ v t . rat,ios). The clirornnt,ograin was developed wit,h 8 ml. of :L soliition prel)*.red by mixing 1 i d . of vi:tt,er wit,h 10 ml. of diossrie.

COLUMN CHROMATOGRAPHY O F SITGARS

79

6. Other Adsorbents Certain adsorbents have been selected for specific, sugar separations, without consideration of t>heirpossible general applicability t,o carbohydrates. Mixt'ures of C14-labeledD-glucose and D-fructose were resolved on a column of hydrated magnesium acid silicate,lo6employing 05 % ethanol as the developer."J6The acid-treated silicate (essentially silicic acid) was useful as an adsorbent in the investigation of the structure of sapote gum.107 Alumina,'O* one of the most popular chromatographic adsorbents, has rarely been used for uncombined sugars. A sharp separation of the branched (upper zone) from t,he linear (lower zone) constituent of starch was stated to be effected on a column in which the upper section of the alumina was adjusted to pII 4.5 and t,he lower section tJopH 8.2; t'he developer was 0.0025 N iodine in pot)assium iodide.logA multiplicity of zoiies resulted on chromatographing animal glycogens on freshly precipitated calcium carbonate; aqueous iodine was employed as the developer and zone indicator.l10 The adsorption on silica gel of the polysaccharides of Mycobacterium tuberculosis and successive development with hexane, chloroform, methanol, and water produced 21 distinct fractions.11*

111. COLUMN CHROMATOGHAPHY O F SUGAR DEarvATIVEs I . p-Phenylaxobenxoyl Esters

The epic of the column chromatography of sugars began a little more than a decade The sugars D-glucose and D-fructose, being colorless, were esterified with a colored acyl halide, to yield colored sugar derivat,ives. The acylating agent chosen was p-phenylazobenzoyl ("azoyl") chloride, and red-colored, crystalline pentaazoates were obtained. A mixture of these esters was adsorbed on a column of silica, and developed under pressure with a 1/1 mixture of benzene/petroleum ether (b. p. SO-SO"). The progress of the resolution was followed visually; a typical Tswett chromatogram was (105) hlagnesol, it hydrated magnesium acid silicate produced by the Food Machinery and Chemical Corporation, Westvaco Chemical Division, South Charleston, W. Virginia. This adsorbent has been useful in the chromatography of several types of acyl sugar derivative. (106) G. R. Noggle and R. A . Bolomey, Plant Physiol., 26, 174 (1951). (107) E.V. White, J . A m . C h m . SOC.,76,4906 (1954). (108) H. Brockmann and H. Schodder, Ber., 74, 73 (1941). (109) pvl. Ulmnnn, Makrorrrol. Chem., 9,76,97 (1952); 10, 147,221 (1953); Die Sturke, 4, 73 (1952); Pharmazie, 7, 787 (1952). (110) V . V. Koval'skil, Doklady A k a d . Nauk S. S . S. R., 68, 1083 (1947); Chem. Abstracts, 44, 7902 (1950). (111) B. Siege], G. A . Candela and R. M. Howard, J . Am. Ch.e,m. SOC.,76, 1311 (1954). (112) W. S. Reich, Compt. rend., 208, 589, 748 (1939); Biochem. J . (London), 33, 1000 (1939).

80

W . W. BINKLEY

produced. A red-colored zone containing the more strongly adsorbed D-fructose ester remained near the top of the column; below it was an orange-colored interzone, and lower still on the column was a second redcolored zone containing the D-glucose ester. This classical demonstration initiated the modern, column chromatography of sugars and led to the preparation of a new and interesting series of sugar esters (see Table VIII) In addition to alumina and silica, &lagneso1106 and silicic were found to be useful adsorbents for the separation of sugar azoates; the TABLE VIII Sugar Azoates Suitable f o r Clhromatograph?/‘13 Azoafe o/

a-D-Glucose 0-D-Glucose a-D-Galactose p-D-Galactose 0-D-Fructose a-D-XylOSe (3-D-Arabinose p - L - Arabinose Sucrose a ,a-Trehalose (3,fl-Trehalose a-Lactose @-Lactose a-Gentiobiose p-Maltose &Cellobiose p-Melibiose Meleeitose

Sinfers at, “C.

265

268

.Welling poinf, “C. (mi-.)

265-266: 252-253 275-276 255-255.5 124.5-125.5 15C-157 261.5-262 262-262.5 125-125.5 134-134.5 328-329 287-288 199-204 232-233 274275 272-273 279.5-280

127-130 143-145

223

-50

436 170 -440 244 -755 755 35 210 17 320 167 62 2 105 172 188 146

selectioii of the proper developers is critical in the evaluation of these adsorbeiits (see Table IX) . l I 3 “Azoyl chromatography” of sugars, employing Magnesol, separated mono- from di-saccharides, such as D-galactose from lactose, as well as the two disaccharides, sucrose from lactose.115Application of this method, with silicic acid as the adsorbent, resolved these binary mixtures of sugar azoates: D-galactose from D-fructose, D-glucose from melezitose, and a- from P-D-galactose; and was extended successfully to ternary and quaternary (113) G. H. Coleman end C. M. McCloskey, J . Am. Chern. Soc., 66, 1588 (1943). (114) Reagent grade, Merck and Go., Rahway, N. J. (115) G. H. Coleman, 8.G. Farnhani and A . Miller, J . Am. Chem. SOC., 64, 1501 (1942).

COLUMN CHROMATOGRAPHY OF SUGARS

81

Several factors have limited the general acceptability of this method. Completely azoylated sugars may contain up to 85% of the azoyl group. In addition to the tedious procedure for the preparation of these esters, coiisiderahle quantities of the comparatively rare axoyl chloride are (YJI~sumed. Reconversion of the axoates isolated t80 the free sugars is uiiTABLE IX InJ?iiPnce of the Dciirloprr on the Bvuluatzon of Adsodents for the Separation p-o-Gltirose Penfaazonfe and p-Cellohzosr Clctanzoatc'13 Adsnrbenl

I

/)eveloper

:

(ij

Re.solulioir

Magnesol and Dicalitee Silicic acid

I,igroine/benzene/chloroforni 1/1/1 (Developer A)

Magnesol and Dicalite Silicic acid Magnesol and Dicalite Silicic acid Silicic acid

A containing 0.4% of ethanol"

Good; two zones; cellobiose zone remained a t top of column Good; two zones

A Containing 0.4% of ethanol A containing ]-lo% of ethanol

Fair; two zones Poor t o none

A containing ]-lo% of cthanol Chloroform

Poor t o none Adequate; two zones; cellobiose zone remained a t top of column Adequate; two zones; cellobiose zone remained at top of column Adequate; two zones; cellobiose zone moved down somewhat from top of column Poor

A

Silicic acid

Chloroform with 0 1% of ethanol

Silicic acid

Chloroform with 0.2% of ethanol

Silicic acid

Chloroform with more than 0 2% of ethanol

None

A diatomaceous filter-aid produced by the Dicalite Co., New York, N . Y. cent,

* Volume per

satisfactory. However, the idea of combination of a colored group with sugars to be chromatographed was fuiidameritally sound, arid some of these objections have now been eliminated. Partially methylated D-glucoses were azoylated and then chromatographed 011 alumina, using chloroform as the developer,L16and 011 silica (prepared from sodium silicate) with chloroform/beneene/petroleum ether (b. p. 65-1 10"):l / l / l . I L 7 The color (116) K. Myrback and C. 0. Tsmm, Svensk Kern. l'idskr., 63,441 (1941). (117) J. K. Mertzweiller, D . M. Carney and F. F . Farley, J . Am. Chem. Soc., 66, 2367 (1943).

82

w.

U'. %INKLEY

iiitroduced into a hexose molecule I)y a single azoyl group is suficient to permit visual observat,ion of t.he progress of t,he adsorbat>edown the column. The adsorpt>ivebonds become weaker with increase in number of azoyl groups and with devrease in number of methoxyl groups. The yields in the formation of this type of compound were 95 ?4 or better, and the recovery of resolved substances was nearly quantitative. Hardly believable though it may seem, a mixture of methylated sugars had been separated, virtually quantitat,ively, at room temperature with very simple apparatus. A new and powerful tool for the elucidation of structure of polysaccharides had been forged; but, until recently, it was rarely used. This method was further refined"* before actual use in the structural studies of polysaccharides. Partially methylated D-glucoses were reduced t o their corresponding hexitols, which were converted to their azoyl esters, thus avoiding the complication of anomers. Markedly simplified chromatograms were obtained with alumina.118- The utility of this method was demonstrated by its successful application120to the hydrolyzate from fully methylated lichenin. Elution of the zones from the chromatogram on alumina led to the recovery of 68 f 4 % of 1,4,5-tri-0-azoyl-2,3,6-tri-O-methy~-D-glucitol (denoting 1 -+ 4 links) and of 32 f 4 % of 1,3,5-tri-O-azoyl-2,4,6-tri-O-rnethyl-~glucitol (denoting 1 -+ 3 links), which confirmed the results obt,ained by a different chromatographic method.121 Another interesting variation of the chromatography of sugar derivatives containing both colorless and colored substit,uent groups is the use of the acetates of p-phenylazophenyl glycosides.lz2These compounds are prepared from interaction of poly-0-acetylglycosyl chlorides (formed more easily than methylated sugars) with p-hydroxyazobenzene (more readily obtained than azoyl chloride). They were adsorbed on silica gel'23and then developed with chloroform or benzene, or with a mixture of these solvents with petroleum ether. Typical separations were D-xylose and D-glucose; (118) R. A. Boissonnas, Erperienlia, 3,238 (1947) ;Helv. Chim. Acta, 30, 1689 (1947). (119) The alumina must be exactly neutralized with acid prior t o use as an adsorbent for sugar esters since these are decomposed by alkali. R. W . Jeanloz, H . G. Fletcher, Jr., and C. S. Hudson, J . Ant. Chern. SOC.,70,4054 (1948). H . E. Stavely and J. Fried, ibid., 71, 137 (1949). (120) R. A. Boissonnas. Helv. Chim. Acta, 30, 1703 (1947). (121) K. H . Meyer and P. Gurtler, Helv. Chim. Acta, 30, 751 (1947). (122) C. D. Hurd and R. P. Zelinski, J . Am. Chem. Soc., 69, 243 (1947). (123) The silica gel was prepared by slowly adding 200 ml. of 3 N hydrochloric acid t o a mechanically agitated solution of 100 g. of water-glass in 200 ml. of water. The gel was broken up; it was heated on a steam bath in a current of air t o remove most of the excess water and acid. The friable residue was crushed, washed with water until neutral t o litmus, and dried a t 70" for 3-4 days. It was ground t o 40-60 mesh before use.

COLUMN CHROMATOGRAPHY O F SUGARS

83

D-galactose, D-glucose, and maltose; lactose and maltose; and D-xylose, D-glucose, and lactose, all as their acetylated p-phenylazophenyl glycosides. 2. Acetate Esters

The introduction of the “brush” or “streak” technique40for the locating of zones of colorless substances added vital impetus t’o the progress of modern, column chromatography. Colorless compounds are adsorbed on columns of white or light-colored adsorbents; the chromatogram is developed, and the moist column of adsorbent is extruded. The column is then brushed or streakedI24lengthwise with a reagent sufficiently sensitive to undergo a color change in the presence of the adsorbates. The resolved substances are located; the column is cut into appropriate sect’ions,and the portion of the adsorbent wetted by the streaking reagent is removed by gentle scraping. These sections are then eluted, and the individual materials are recovered. This method has been extended to sugars and to several of their colorless derivatives. Its first and most extensive app1icat)ion has been to sugar acetates. Several factors influenced the selection of this group of compounds. They are a simple type of sugar derivative, are easily prepared, and are soluble in many organic solvents. Often, sugar acetates crystallize readily, and they are most extensively described in the sugar literature. Further, the individuality of the sugars is masked far less in the acetates than in their benzoate and azoate esters, and so sharper chromatographic separations can be expected. Directive acetylation was employed in order to encourage the formation of one anomer of each sugar; thus, each resolved sugar would be represented by a single zone. Hydrated magnesium and calcium acid silicates (MagnesolIo5and Silene EF,lo2respectively) are often suitable for sugar-acetate chromatography. The original extension of the brush technique to sugar acetates employed Magnesol,4* and this adsorbent has since been generally preferred. A major factor in the subsequent success of the method was t,he discovery, during its adaptat,ion to sugar acetates, of an unusually versatile txush or streak reagent., namely, alkaline ~erma1iganate.l~~ The chromatographic selectivity of Magnesol is notably dependent upon its moisture content.Iz6 The octaacetates of p-gentiobiose and p-maltose are st,rongly adsorbed and (124) This may he accomplished with a medicine dropper the tip of which is drawn out fine. (125) The streak reagent was prepared by dissolving 50 mg. of potassium permanganate in 5 ml. of 2.5 N sodium hydroxide. It was most, sensitive immediately after preparation. (126) M . L. Wolfrom, A . Thompson, T . T . Galkowski and E:. J. Quinn, Annl. Chcm., 24, 1670 (1952). This interesting eti’ect is not uniisual with adsorbeIit,s.

84

\V. W. BINKLEY

are only slightly separated at low moisture contents; zones of these substances are diffuse and widely separated when this adsorbent possesses over 20 % of moisture. The best results (based on the consequent separation of these acetates) mere obtained in the moisture-content range of 12-18 %. A benzeneltert-butyl alcohol (125/1) developer was used in these evaluations, and i t has frequently been preferred to benzene-ethanol mixtures because of its greater stability to alkaline ~ e r m a n g a n a t e . ~ ~

~ZZZZ-

a-Cellohexaose eicosaacetate

a-Celiotriose hendecaacetate

a-D-Glucopyranose pentaacetate FIG.12.-Chromatograms of Cellulose Acetolyzate on (a) Hydrated Magnesium Acid Silicate arid (b) Hydrated Calcium Acid Silicate. [The Chromatograms were Developed with Benzene/Ethanol (100/1) .] See Addendum, p. 94.

Mono-, di-, and tri-saccharide acet,ates were easily separated, using benzene -ethanol developer, into groups whose composition reflected the influence of molecular weight (see Fig. 12) .Iz7 The influence of t.he position of (see Fig. l3),lz6.128 and t,he loss of polarit,y of, a funct,ional group (see Fig. 14)Iz9on the adsorbabilit,y of acet,ates was interesting. Transformat,ion of t,he carbosyl group of pentn-O-acet,yl-D-gluconic acid t,o cert,ain nitro(127) E . E. Dickey and M. I,. Wolfrom, J . A m , Chptn. Soc., 71, 825 (1949); M. I,. Wolfrom and J . C . Dacons, ibl'tE.,'74, 5331 (1952); J. G . Leech, T a p p i , 36, 249 (1952). (128) W. W. Binkley and hf. 1,. Wolfrom, ./. Ant. Chevt.SOC.,68, 1720 (1946). (129) Ref. 7 ( g ) , p. 28.

85

COLUMN CHROMATOGRAPHS OF SUGARS

p-Gentiobiose octaacetate

@-Maltose octaacetate

-a-D-Glucopyranose pentaacetate a-D-Glucopyranose pentaacetate

~

~

FIG.13 -Chromatograms, on Hydrated Magnesium Acid Silicate, of (a) 01- and p-i>-Glucopyr:tnose Pentaaretntcs and of (b) p-Gentiobiose Octaacctate and p-Maltose 0 c t aa ce t at e .

pentaacetate

+ ’L’//,’///,/////:

fi-D.Glucopyranose __f

2,3,4,64etraacetate

OAC

H @.D.Glucopyranose

,’, / ,/

pentaacetate

A

c--- c‘’

’

‘OR‘

I

FIG.14.-Relationship Between the Adsorptive Strength and the Polarity of C1 of Certniri D-Glucose Acetates Chromatographed on Hydrated Magnesium Acid Silicate.

8G

W. W. BINRLET

genous and other derivatives produred, 011 a silicic acid adsorbeiit’”Owith henzenc-acetic acid developers, the following series (in order of derreasing adsorptive strength) : amides, phenylhydrazides, acids, lactones, and nitri1es.la1 Kumerous publit-atious i n the last decade have testified to the soundness of sugar-acetate chromatography on Magnesol as a separative procedure. The potentialities of the method are demonstrated in the elucidationz6, of the structure of the trisaccharide, p a n 0 ~ e . lThe ~ ~ acetate of its sugar alcohol (panitol dodecaacetate) was partially hydrolyzed and then chromatographed. The five components of the hydrolyzate were separated, recovered as crystalline materials, and properly identified. They were acetates of D-glucitol, p-D-glUCOpyranOSe, 0-isomaltose, maltitol, and unreacted panitol; thus, the structure of the trisaccharide was unequivocally established as 4-0-(cr-isomaltopyranosyl)-~-glucose.The presence in amylopectin of glycosidic 1 -+ 6 links (indicating branched structures) was proved by the chromatographic isolation of the acetates of p-isomaltose [~-O-(LY-Dglucopyranosyl)-p-~-glucose]~~~ 135 and panose.26(a) Maltotriose [4-0-(0(maltopyranosyl)-~-glucose]~~6 was found1” among the hydrolytic products of starch, and the structure of this triose was confirmed26(a)by “acetate chromatography.” Application of the method to glycogen revealed the .~~~ existence of glycosidic 1 + 6 links in this animal p o l y ~ a c c h a r i d eCrystalline aldehydo-D-galactose heptaacetate and aldehydo-D-xylose hexaacetate were separated chromatographically from the acetolyzates of guaran acetate and xylan acetate, respe~tive1y.l~~ 1

(130) A product of the Mallinckrodt Chemical Works, St. Louis, Mo. The adsorbent mixture was silicic acid/Celite No. 535 (3/1). (131) J. F. Haskins and M. J. Hogsed, J. Org. Chem., 16, 1275 (1950). (132) M. L. Wolfrom, A . Thompson and T . T . Galkowski, J . A m . Chem. SOC.,73, 4093 (1951); A . Thompson, M. L. Wolfrom and E. J . Quinn, ibid., 76, 3003 (1953). (133) S. C. Pan, A . A. Andreasen and P. Kolachov, Science, 112, 115 (1950); S. C. Pan, L. W. Nicholson and P. Kolachov, J . Am. Chem. SOC.,73,2547 (1951). (134) M. L . Wolfrom, J. T . Tyree, T. T . Galkowski and A . N . O’Neill, J . Am. Chem. Soc.,73,4927 (1951). (135) The first isolation of crystalline p-isomaltose octaacetate was accomplished by means of Magnesol chromatography; I,. W. Georges, I . L. Miller and M. L. Wolfrom, J . Am. Chem. SOC.,69, 473 (1947); M. L. Wolfrom, L. W. Georges and I. L. Miller, ibid., 71, 125 11949). (136) J. M. Sugihara and M. L. Wolfrom, J . A m . Chem. Soc., 71,3357 (1949). (137) M . L. Wolfrom, L. W. Georges, A . Thompson and I. L. Miller, J . Am. Chem. Soc .,71,2873 (1949). (138) M. L. Wolfrom, E. N. Lassettre and A. N . O’Neill, J . Am. Chem. Soc., 74, 3162 (1952). (139) R . 1,. Whistler, Eileen Heyne and J. Bachrach, J . A m . Chem. Soc., 71, 1476 (1949).

COLUMN CHROMATOGRAPHY OF SUGARS

87

In addition to its use in carbohydrate structural studies, sugar-acetate chromatography has been employed as an analytical tool. The first isolation of D-glucose and D-fructose (as their crystalline acetates) from cane juice was achieved by its aid.128D-Glucose, n-fructose, D-mannitol, and myoinositol were found in cane, blackstrap molasses (see Fig. 9),53.140 and, in addition to these, D-arabitol and erythritol in its fermentation residue (see Fig. The interconversion, by heat, of D-fructose to n-glucose was column chroreadily predicted on the basis of enolizatiori matography supplied the proof.50hiialysis of the acid reversion products of D-glucose by a combination of carbon chromatography and magnesium acid silicate chromatography led to the isolat,iori of gent.iobiose, isomaltose, maltose, cellobiose, sophorose, P ,P-trehalose, and 1evoglClcosan (all as acetates), and of the acetate of an unknown disaccharide, temporarily designated y - a ~ e t a t e . ' ~ ~ Sugar-acetate chromatography was instrumental in the preparation of crystalline, key-intermediate compounds in some intricate syntheses of sugars, such as keto-D-glycero-D-gulo-octulose heptaacetate in the format,ion of ~-glycero-~-gulo-octulose.~~~ Sugar diazo compounds have often been purified by this technique; some derivatives responding were l-deoxy-ldiazo-keto-L-galacto-heptulose p e n t a a ~ e t a t e ,1-deoxy-1 '~~ -diazo-keto-D-glyceroD-galacto-octulose h e ~ a a c e t a t e , ' ~and ~ 1-deoxy-1-diazo-keto-L-manno-heptulose p e n t a a ~ e t a t e . The ' ~ ~ anomeric a- and @-D-glucosaminepentaacetates A chromatogram of an acet'ylated, residual were resolved on Magnes01.l~~ sirup from tjhe preparation of N-methyl-L-glucosaminic acid nitrile separated the acet,ate thereof from its epimer, penta-0-acetyl-N-methyl-Lmannosaminic a~id.1~8 Magnesol chromatography was one of the keystones in the actual isolation of sucrose (as its octaacetate) after its first chemical synthesis.149This (140) W. W. Binkley, NI. Grace Blair and M. L. Wolfrom, J . A,tn. Chem. SOC., 67, 1789 (1945). L:Wolfrom I. and W. L. Lewis, J . Am. Che,m. Suc., 60, 837 (l(328). (141) & (142) A. Thompson, Kiiniko Anno, M. 1,. Wolfrom and M. Inatome, J . Atti. Chma. SOC.,76, 1309 (1951) ; the y-acetate is now known t o be 3 - O - ( o l - ~ - g l u c o p y r a n o sy l ) - ~ - ~ glucopyranose octaacetate. (143) M. L. Wolfrom and A . Thompson, J . Am. Chem. SOC., 68, 1453 (1946). (144) M. L . Wolfrom, J. M. Berkebile and A . Thompson, J . Am. Chem. SOC., 71, 2360 (1949). (145) RI. I,. Wolfrom and 1'. W. Cooper, J . A m . Chein. SOC., 72, 1345 (1950). (146) M. I,. Wolfrom and H. B. Wood, J . A m . Chem. SOC.,73, 730 (1951). 71, 2870 (147) M. I,. Wolfrom, R . U. Lemieux and S. M. Olin, J . A m . Cheni. SOC., (1949). (148) M. L. Wolfrom and A. Thompson, J . h a . Chem. SOC.,69, 1847 (1947). 76, 4118 (1953). (149) R . U. Lemieux and G. Huber, J . Am. Chem. SOC.,

88

W. W. BINKLEY

adsorbent can distinguish the difference between the glycosidic bonds of sucrose octaacetate and isosucrose octaacet at.e.150 A chromatographic series showing t.he adsorptive strength between certaiii sugar acetates and a hydrated calcium acid silicate (Silene EF) has been prepared (see Table X) .lo3 151 Silene EF is a weaker acetate adsorbent than is Magnesol; thus, the former has greater utility in the separation of the acetates of t,he larger oligosaccharides, such as those cf a-cellotetraose, a-cellopentaose, a-cellohexaose, and a-cellohept)aose (see Fig. 12).lZ7 T h e 9

TABLE

Chroriialoqraphic Adsorption Series; on Calcium Acid Silicate, of Some Sugar Derivatives'~3 J

Class I[I5 cc. of benzenelethanol (250/l)b] Raffinose hendecaacctate 8-R.lelibiose octaacetate, sucrose octaacetate 8-Maltopyranose octaacetate Class ZZ 115 cc. of benzene/ethanol (500/1)] keto-wFructose pentnacetate D-Glucitol (sorbitol) hexaacetate, D-mannitol hexaacetate 6-D-Glueopyranose pentaacetate ~ - - ~ - A m b i r i o p y m n otetraacetate se CIoss I I Z [15 cc. of benzenc/ethanol (lOOO/l)] WL-FUCO t>et,raacetate S~ Class I T T (15 cc. of benzenelethanol ( l O O / l ) ] 2,3-Di-O-methyl-~-glucose 2 , 3 ,B-Tri-O-methyl-n-glucose Class V [12.5 cc. of benzene/ethanol (250/1)] 2,3,4,6-Tetra-O-methyl-~-glucose a Arranged in decreasing order of adsorptive strength. Adsorbent: 1.8 g. of Silene EF1°2/Celite" (5/1) ( a t . ratio). Dimensions of adsorbent column: 0.9 X 11 cm. Adsorbate solution: 2 mg. in 0.5 cc. of absolute benzene or chloroform. Developer: noted in class heading. b The benzene was thiophene-free, and the ethanol was absolute.

increased adsorptive strength of this series reflected the increase in their molecular weights. 3. Methyl Ethers Nearly all polysaccharides are st,ableto alkalis, aiid often they are met,hylat,ed almost completely by ail appropriate, single t>reatment with methyl su1fat)e aiid a strong hase.15' Vsually, succwsive t'reat,ments with this or (150) W . W . I h k l e y atid hI. I,. Wolfroni, J . A v i , C h n . Soc.., 68, 2171 (1946). (151) The variability in adsorptive strength of various lots of Silene EF for sugaracetate chromatography was small, and its relatively large content of alkali and other water-soluble material was not troublesome. (152) W. N. Haworth, J . Chenr. Soc., 107, 8 (1915).

COLUMN CHROMATOGRAPHY O F STJG.4 RR

89

other methylatiiig s g e ~ l t s produce l~~ fully methylated products which may he used a.s at.art,iiig materials for t.hc elucidation of the structure of the polysaccharides. Formerly, t,licse et,heritied subst,aiices wcrc hydrolyzed, and the rt:sulting partiadly rnethylated proclucts were subjected to frart>ional distillatiou under diminished pressure. Rarely is this rectification completely satisfactory because of the inaccuracies introduced by demethylation, pyrolysis, arid incomplet,e recovery of the methylated sugars. These dificult.ies have tjo a great extent been eliminat'ed by chromat,ography. The colored azoyl esters of partially methylat,cd sugars were prepared, and the conditions established for t.heir resolution cn and silica.123Separat.ion of the methyl glycosides of 2 , 3 ,6-tri-O-met.hyl- and tetra-0-methyl-D-glucopyranose was accomplished with a flowing rhromatogram on The completely methylated hesoside moved rapidly on development with an ether-petroleum ether mixture, and was recovered in 94% yield from a portion of the column effluent,. The more strongly adsorbed methyl tri-0-methyl-P-D-glucoside appeared next in the effluent; however, development n-ith methanol was necessary in order to release the a - anomer; ~ the total yield of the anomers was 88%. These recoveries were good, but not quantitative. The magnitude of t.he repeat,ing units in rice starch and banana starch was found by this method to be 33 and 26 D-glucose residues, respectively; these values confirmed the rcsult,s obtained by fractional-distillation 156 The ariomeric met,hyl glycosides of fully methylated D-glucose and D-galactose have been partially separated on alumina, with chloroform as the developer.157The formation of anomers during the preparation of these met,hyl glycosides is a complication; often, two zones represent a single component of the original mixture. Polymerization and demethylation may also occur during glycoside formation. These object,ioris have for the most part been met by using a combination of (a) extraction with part,ition solvents and (b) chromatography on silica-water columns, for the resolution of di-, t,ri-, and tetra-0-methyl-Dg l ~ c o s e s The .~~~ Molisch reagent was used for detecting the zones in the extruded chromatograms of these substances. EssentJially the same separa1

(153) T. Purdie and J. C. Irvine, 6.Chem. Soc., 83, 1021 (1903). (154) J. K. N. Jones, J . Chern. Soc., 333 (1944). (155) E. L. Hirst and G. T. Young, J . Chem. Soc., 1471 (1939). (156) E. G . E. Hawkins, J . I<.N. Jones and G. T. Young, J . Chem. Soc., 390 (1940). (157) F. Brown and J . K. N . Jones, J . Chem. Soc., 1344 (1947). (158) D. J. Bell, J . Chem. Soc., 473 (1944); the silica-water adsorbent was prepared exactly as described by A. H. Gordon, A . J . P. Martin and R. I,. M. Synge, Biochem. J . (London), 37, 79, 86, 92 (1943); this adsorbent was employed in the elucidation of the structure of a dextran produced by Betacoccus arabinosaceotcs with a sucrose-mineral substrate [S.A . Barker, E. J. Bourne, G. T. Bruce, W. B. Neely and M. Stacey, J . Chew,. SOC., 2395 (1954)] arid of certain bacterial levans [D. J. Bell and R. Dedonder, ibin!., 2866 (1954)l.

90

W. Mi. BINKLEY

tions occur on filwous alumina, developed witJh an acetone-benzene mixture, followed by rechromatography of the effluent sugar mixture using only benzene as t,he deve10per.I~~ The combinattion procedure (extraction and silica-water column chromatography) has been extended to partially met)hylated f ruc toses . I 6o The adsorptive strengths of powdered cellulose and filter paper for methylated sugars are closely parallel. Filter-paper strip and sheet chromatography provide an essential service in the evaluation of the effluent from powdered-cellulose chromatograms. This adsorbent has recently been popular in end-group assay st,udies, more as a device for the preparation and isolat,ion of crystalline methylated sugars t.han as an analytical tool.64 Suitable micro-methods applicable to filter-paper sheets have been developed for t,he quantitative estimation of t,he resolved substances with alkaline hypoioditel6I and, colorimetrically, with aniline phthalate in methanol.162The conversion (with 85 % phosphoric acid) of powdered cellulose to hydrocellulose produces an adsorbent with a greater selectivity and capacity for methylated sugars.163Examples of the utility of this method are numerous. Equimolar quantities of 2,3,5-tri-O-methyl-, 2,3-di-Omethyl-, and 2-O-methyl-~-arabinosecrystals were obtained from a chromatogram of hydrolyzed, methylated, sugar-beet araban.64 The method was employed successfully in structural studies on : the galactomaimam of lucerne and clover fenugreek seeds,'O and certain iris seeds71;the xylans of esparto grasP4(a)and of wheat s t r a ~ 1 6 ~alginic ( ~ ) ; acidIss; Karaya gum166;degraded cherry and wood saponinsI68; an animal polysaccharide (from the intestine of horse and the rumen of sheep) which re(159) Ethelda J . Norberg, I . Auerbach and R. M. Hixon, J . Am. Chem. Soc., 67, 342 (1945). (160) D . J. Bell and Anne Palmer, Nature, 163, 846 (1949); J . Chein. Soc., 2522 (1940) ; 3763 (1952). (161) E. L. Hirst, L. Hough and J . K. N . Jones, J . Chem. Soc., 829 (1949). (162) (Miss) J. K. Bartlett, L. Hough and J . K. N. Jones, Chwnistr2/ & Industry, 76 (1951). (163) J. D . Geerdes, Bertha A. Lewis, R . Montgomery and F. Smith, Anal. Chem., 29, 264 (1954); P. A. Rebers and F. Smith, J . ,4m. Chem. Soc., 76,6097 (1954). (164) (a) S. K. Chanda, E. I,. Hirst, J . K. K.Jones and E. G. V. Percival, J . C h e m . Soc., 1289 (1950); G. 0. Aspinall, E . L. Hirst, R . W. Moody and E. G. V. Percival, ibid., 1631 (1953); (b) G. 0. Aspinall and R. S. Mahomed, i b i d . , 1731 (1954); I . Ehrenthal, R. Montgomery and F. Smith, J . Am. Chem. Soc., 76, 5509 (1954). (165) S. K. Chanda, E. L. Hirst, E. G. V. Percival and A. G. Ross, J . Ch.eni. Soc., 1833 (1952). (166) E. L. Hirst and Sonia Dunstan, J . Chem. Soc., 2332 (1953). (167) J . K. N. Jones, J . Chem. Soc., 3141 (1949). (168) R . A. Laidlaw, J . Ch,eirr. Soc., 752 (1954).

COLUMN CHROMATOGRAPHY O F SUGARS

91

sembles amyl~pectinl~g; and the di- and tri-galacturonic acids formed during the enzymic degradation of pectic a ~ i d . 1 ~ ~ The powdered-cellulose, chromatographic assay of t’he hydrolyzate of methylated stachyose has furnished important structural data.2g1 , 3 ,4, 6Tetra-0-methyl-D-fructose, 2 , 3 , 4 ,6-tetra-O-methyl-~-galactose,2,3,4-tri0-methyl-D-galactose, and 2 , 3 ,4-tri-O-methyl-~-glucose were recovered in approximat,e!y equimolar quantities; the glycosidic 1 + 6 bond for the D-glucose residue and for one of the D-galactose residues in stachyose is clearly indicated. Powdered-cellulose, chromatographic, end-group assay has also been applied to fructans. The fructose polysaccharides from rye grass (LoZium perenne)”l and leafy cocksfoot grass (Dactylis g l o r n e r a t ~ ) ’were ~ ~ shown to possess a linear structure, whereas a fructan from the rhizomes of couch grass (Triticum repens I,.) was highly branched (see Table XI).17aThe D-glucose in inulin hydrolyzates was found to originate from, D-glucose residues bound in this fructan.”* The application of this end-group assay to a glucan from barley grain revealed that it consisted of unbranched chains of P-D-glucopyranose units linked about equally 1 3 and 1 4 4,176 whereas a polysaccharide (“nigeran”) synthesized from sucrose by Aspergillus niger was found to contain linkages in the same positions, but the units were a - ~-g lu co p y ran o se.~ ~ A ~tentative structure for beechwood hemicellulose-A, a straight chain of approximately 70 &D-xylopyrnose units (I + 4) with a 4-0-methyl-~-glucopyranuronicacid residue linked (1 + 2) to every tenth unit,’” was based on a similar analysis. The first crystals of 2,3-di-O-methy1-a-~-xylose ever isolated were j

(169) G. Forsyth, E. L. Hirst and A. E. Oxford, J . Chem. SOC., 2030 (1953); G. Forsyth and E. L. Hirst, ibid.,2132 (1953). (170) J. K . N . Jones and W. W. Reid, Chemistry & Industry, 303 (1953); J . Chem. SOC., 1361 (1954). (171) R. A. Laidlaw and S.G. Reid, J . Chem. SOC.,1830 (1951); V. D. Harwood, R. A. Laidlaw and R. G . J. Telfer, ibid., 2364 (1954). (172) G . 0. Aspinall, E. 1,. Hirst, E. G. V. Percival and R. G. J. Telfer, J . Chern. SOC.,337 (1953); G. 0 . Aspinall and It. G. J. Telfer, Chemistry & Tndustry, 1244 (1952). (173) P. C. Arni and E. G. V. Percival, J . Chem. SOC.,1822 (1951). 1297 (1950). (174) E. 1,. Hirst, n. I. McGilvray and E. G. V. Percival, J . Chem. SOC., (175) G. 0. Aspinall and R. G. J. Telfer, J . Chem. SOC.,3519 (1954). (176) S. A. Barker, E. J. Bourne and M. Stacey, J . Chem. SOC., 3084 (1953); S. A. Barker and T. R. Carrington, ibid., 3588 (1953); S.A. Barker, E. J. Bourne and T. R. Carrington, i b i d . , 2125 (1954), oligosaccharides from sucrose and Aspergillus niger. (177) G . 0. Aspinall, E. 1,. Hirst a n d Ti. 8. Mahorned, J . Chem, SOC.,1734 (1954).

92

W. W. BINKLEY

formed in a chromatographically homogeneous sirup of this sugar obtained from a powdered cellulose column.178

4.Other Derivatives Aluminai79is an excellent adsorbent for the chromatography of certain sugar derivatives. It sharply differentiates between anomers, yielding crystalline substances from both sirupy arid crystallirie mixtures. Frequently, the p-D aiiomer moves down the adsorbent column more rapidly than does the (Y-D form; 1,2,4-tri-o-acctyl-G-deoxy-3-O-methyl-P-~-gluTABLE XI Powdcrcd-Cellulose Chromnlography of the Hydrolyzutes of Some Methyluted Fru~tans'~~ Per cent

0.f

coniponenl in j r u i t a n

Coinponenl

Rhizomes of couch grass

R y e grass

Inulin

-I 4 93.3 0.7a

42 11 4 39

-

-

-

-

99.8

98.0

9G

98

97 3

4 1,3,4,6-Tetra-O-methyl-~-fructose 1,3,4-Tri-0-methpl-~-fructose 93 3,4,6-Tri-0-methyl-D-fructose 3,4-Di-O-metliyl-wfructose 2.8 2 , 3, ~ , 6 - T e t r a - O - m e t h ~ ~ ~ - ~ - g 1 ~ 1 c o s eMixture of tri-0-methyl-D-glucoses

Total

2 _

_

_

_

-

2.2 3.7 100.1

~

Identification was incomplete. fructan. a

3.2

91

Concentration in hydrolgzate from untreated

cose,I8O methyl 2,~i-d~deoxy-~i-~odo-3-~-mcthy~-4-0-tosy~-~-~-g~i~opy side,'*I and met hgl2,6-dideoxy-3,4-di-0-tosyl-~-~-zyZohesopyrano8ide~~~ a,ll precede their momers. The order of deparhre, from t,he adsorbent column, of the anomers of 1 ,2,4-tri-O-acet~.l-~-dig.italose ( I ,2,4-tri-O-acetyl-6(178) S. K . Chanda, Elizabeth E. Percival and E. G. V. Percival, J . Chenz. SOC., 260 (1952). (179) The developers for thesc chromatograms on nlumina vary from petrolcum ether containing small amouiits of beneciie up t o 100% benzene (for \venkly held substances) nnd from benzerie w i t h smitll t o large porportions of ethyl ether, ethanol, or acetour for modrrately and st,rongly held :tdsorbxtev. (180) E. Vischer and T . Iieiclistriii, Ele[ci. Chi,w. 4 c t u , 27, 1332 (1944). (181) €1. Haueristeiri and T. Reichsteiii, Hclzi. Chi,t>.Actu, 33, 446 (1050). (182) H. R. Bolliger and T. Reichstein, Helv. Chiin. A c f a , 36, 302 (1953).

COLUMN CHROMATOGRAPHY OF SUGARS

93

deoxy-3-O-methyl-~-galactose)is, however, the reverse.183 Alumina is capable of sorting out the derivatives of certain hexoses according to the number of acyl groups they possess. The order of appearance in the effluent of a flowing chromatogram was methyl 4,6-O-benzylidene-2,3-di-O-tosy1-cr-~-g1ucopyranoside followed by methyl 4 ,6-0-benzylidene-2-0-tosyl-a-~-glucopyranosideIs4;that is, the adsorpt’ive bond between the free hydroxyl group (at C3) and alumina was greater than that from its more massive replacement, the tosyl ester. This adsorbent was discerning with respect to t’he position of the tosyl group, setting apart the preceding 2-0-tosyl derivative from its isomer having the acyloxy group at C3.lS4Alumina differentiated between such anhydrohexosides as methyl 2,3-anhydro-4,6-0-benzylidenea-D-talopyranoside and methyl 2,3-anhydro-4,6-O-benzylidene-c~-~-gulop y r a n o ~ i d e . ’Anhydro ~~ and deoxy compounds are readily separated, from for example, methyl 2,3-anhydro-4,6-di-O-tosyl-a-~-allopyranoside Outstanding success methyl 2-deoxy-4,6-di-O-tosyl-a-~-allopyranoside.~~~ has been experienced in the isolation of crystalline sugar derivatives from crystalline and sirupy mixt,ures.I87 Naturally occurring flavonoid glycosides form anot’her interesting group of sugar derivatives responsive to column chromatography. These glycosides, dissolved in acetone, were adsorbed on Magnesolio6and were resolved by development with aqueous ethyl acetate. Five glycosylquercetins (3,3’,4’, 5,7-pentahydroxyflavone) have been isolated from huckleberry (Vuccinium my~tiZZus)leaves,’**and quercitriii was obtained from a crude extract of oak bark (“lemonflavin”) .189 Cert,ain prepared mixt’ures of flavonoid substances were separable on Magnesol; they included quercitrin(183) C. Tamm, Helv. Chim. Acta, 32, 163 (1949). (184) H . R. Bolliger and D. A. Prins, H e h . Chini. Acta, 28, 465 (1945); 29, 1116 (1046); F. Reber and T. Reichstein, ibid., 28, 1164 (1945); for benzoyl esters, M. Gyr and T. Reichstein, ibid., 28, 226 (1945). (185) H. Huber and T. Rcichstein, Helv. Chhinz. Arta, 31, 1645 (1948). (186) H. R . Bolligcr and M. Thiirkauf, Helv. Chim. Acta, 36, 1426 (1952). (187) C. A. Grob and D. A . Prins, Helv. C h h . Acta, 28, 840 (1945); D. A . Prins, ibid., 29, 1 (1046); A . S. Meyer and T. Reiehstein, ibid.,29,152 (1946); F. Reber and T . Reichstein, ibid., 29, 343 (1946); R.W. Jeanloz, D. A . Prins and T. Reichstein, i b i d . , 29, 371 (1946); A . C. Maehly and T . Reichstein, ibid., 30, 496 (1947); M. Gut, D. A . Prins and T . Reichstein, ibid., 30, 743 (1947) ; C. Tamm and T . Reichstein, ibid., 31, 1630 (1948); F. Blindenbacher and T. Reichstein, ibid., 31, 1669 (1948); employing silicic acid as the adsorbent, R . W. Jeanloz, J . A m . Chent. Soe., 76, 555, 558 (1954); 1’. J. Stoffjrn and R. W. Jeanloz, i b i d . , 76, 561,563 (1954). (188) C. H.Ice arid S. H. Wender, J . An,. C‘hvu. Soc., 76, 50 (1953). (189) C. H . Ice a r i d S . H. Wender, Arch. Biochcm.a n d Bioph?/s.,38, 185 (1952). This adsorbent was successfully employed in similar assays on grapes [B. I,. Wil1i:tms and S. H. Weiider, J . A m . Clte7tr. Sor., 74, 4372 (1952)], apricots [Arch. Biochwti. cvnd R i o p h y s . , 43,310 (1953)], and black currants [ U . I,. Williams, C. H. Ice and S. H. Wcader, J . Am. Chem. Xoc., 74, 4566 (1952)l.

94

W. W . BINKLEY

rutin-quercetrin, xanthorhamnin-quercitrin-quercetin, and xanthorhamnin-rutin-quercitrin.'90 Magnesol chromatography proved essential for successful isolation of the synthetic isoquercitrin from the reaction, in liquid ammonia, between 2,3,4,6-tetra-O-acetyl-~-ghcosyl bromide and the monopotassium salt of quercetrin.lgl This method was applied successfully in the preparation of the rutin, hesperidin, and naringin glucosides from their rhamnoglucosides.192A glucofrangulin (as an octaacetate) was obtained from an extract of Rhamnus frangula bark by flowing chromatographylg3on Floridin XXX43and silicic acid. A similar investigation of the glycosides in certain seeds has been made, employing aluminalg4 l g 6 and magnesium silicate/Celite (2/l).1g4 Amides are useful sugar derivatives, and some aldonamides have responded to chromatography on calcium acid silicate, using aqueous dioxane as the developer (Table VI).lg6 Some degree of resolution of uronic acids and of their low molecularweight polymers has been achieved.66,l o o 9

(190) C. H. Ice and S. H. Wender, Anal. Chem., 24, 1616 (1952). (191) C . H. Ice and S. H. Wender, J . A m . Chem. Soc., 74,4606 (1952). (192) D. W. Fox, W. L. Savage and S. H . Wender, J . A m . Chem. Soc., 76, 2504 (1953). (193) E. Seebeck and 0. Schindler, Helv. Chim. Acta, 29, 317 (1946). (194) J. C. Hess, A . Hunger and T. Reichstein, Helv. Chim.Acta, 36, 2202 (1952). (195) 0.Schindler and T. Reichstein, Helv. Chim. Acta, 36, 921,1007 (1953). (196) M. L. Wolfroni, R. S. Bower and C . G. Maher, J . A m . Chent. Sac., 73, 875 (1951).

ADDENDUM In Fig. 12 (p. 841, the middle zone of column a is an artifact and represents a non-cellulosic disaccharide from the wood cellulose (filter paper) employed. It was absent when cotton cellulose was used.

GLYCOSYLAMINES

BY G. P. ELLISAND JOHNHONEYMAN* Deparfnienl o j C h e n i i s f r y , King’s (’ollegr, I ’ n i z ~ s i f oy j London, EnglantI

CONTENTS I. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11. Nomenclature . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 111. Preparation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IV. Physical Properties.. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . V. Structure . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1. Introduction.. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. Mutarotation and Hydrolysis.. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. Methylation. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4 . Acetylation.. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5. Benzoylation. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6. T r i t y l a t i o n . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7. Periodate Oxidation.. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8. Conclusion.. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VI. Diglgcosylamiiies . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VII. Diamide Derivatives of Aldoses . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VIII. TJses of Glycosylamines . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IX. Tables of Properties of Glycosylamines.. . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

95 96 97 101 102 102 104 109 110 115 116 116 120 120 121 124 125

I . INTRODUCTION

A large number of important, naturally occurring, carbohydrate compounds contain nitrogen. These include nucleic acids, coenzymes, polysaccharides, some virus components, and some of the members of the vitamin B group. Furthermore, compounds made up by union of proteins with carbohydrates are of possible wide occurrence and major biological importance. Reducing sugars react readily, under mild conditions, with compounds containing the primary amino group, notably with ammonia, hydroxylaniine, and a wide variety of amines, hydrazines, and amino acids. A number of second’ary amines give similar products, although with many of them reaction is more sluggish. In the main, the present article will be limited to discussion of compounds resulting from the condensation of one molecule of a reducing sugar with one molecule of ammonia or of a primary or secondary aliphatic, alicylic. or aromatic amine. Although, in recent years, con-

* Harold Hibbert Memorial Fellow a t McGill University, Montreal, Canada, during the period when this article was completed. 95

96

G. P. E L L I S AND J O H N HONEYMAN

siderahle progress has been made in determining the structure, react ions, and importance of these compounds, much reinnins to he done and worb is in progress in many laboratories. For this reason, this review will I P a report on selected parts of thc wide field. Information about nucleosidcs, nucleotides, and nucleic acids is contained in other reviews in this Series.' Derivatives of amino acids will not be discussed. Compounds containing two amide groups attached through their respective nitrogen atoms to C 1 of a monosaccharide will be mentioned incidentally. Since the special nature and properties of the products obtained by reaction of the sugars with o-phenylenedisniine have been reviewed recently by IZicht,myer,? these, also, will not be considered here. In addition, one very important reaction, known as the Aniadori rearrangeinelit, is given separate treatment in an accompanying Chapter.2a The importance, in the non-enzymic browning of foods, of compounds of glycosylamine nature is also outside the scope of the present artirle, but readers interested in the topic are referred to a recent review hy Danehy and Pig~man.~

11. NOMENCLATURE Three representatives of the kind of compound here considered result from the coiidensation of D-glUCOSe with aniline, or ammonia, or piperidine in equimolar amounts. That obtained from D-glucose arid aniline, and assumed for the present to be I, has been called D-glucose anilide, aniline D-glucoside, aniline N-D-glucoside, and D-glucosidoatiilinc, but none of these names can be justified on systematic grounds. The nomenc~lature adopted here is that agreed on between British and American committee^.^ Compound 11, prepared from D-glucose and ammonia, is named D-glUCOSy1amine, because it is ammonia with one of its hydrogen atoms replaced by

I

CHNII,

-

HOCH HCOH

HCO

I

CHzOH I

I I HOCII I HCOH I IICOII

HkOH

HCO-

IICO

I

CHZOH I1

I

CHIOH 111

(1) R. S. Tipson, Advances in Curbohlidrate Chenz., 1, 193 (1945); R.W. Jeanloz and H. G. Fletcher, J r . , ibid., 6, 136 (1951); G. R. Barker, ibid., forthcomingVolunie. (2) N . K. Richtmyer, Advances i n Carbohydrate Chem., 6, 175 (1951). (2a) J. E. Hodge, Advances in Carbohydrate Chenz., 10, 169 (1955). (3) J. P. Danehy and W. W. Pigman, Advances i 7 ~Food Research, 3, 241 (1951). (4) Editorial Report on Nomenclature, ,I. Chem. SOC.,5108 (1952); Chem. Eng. N e w s , 31, 1776 (1953).

GLYCOSYLAMINES

07

the D-glucosyl radical. For t,he purposes of nomenclature, compounds of simple amiiies are regarded as being derived from such glycosylamines; hence, I is N-phenyl-D-glucosylamine or, more fully (to show the ring size), N-phenyl-D-ghcopyranosylamine. I n an alternative acceptable nomenclature, I is D-glUCOpyranOSylalliline, but this type of name is not used unless it is more convenient. Thus, the analogous compound from piperidine, 111, is best named D-glucopyranosylpiperidirie. The name “glycosylamine” used throughout this article is to be considered the general name of the compounds resulting from the condensation of aldoses and ketoses with ammonia and amines. D-Glucosylamine is the name for the compounds obtained from D-glucose and ammonia.

111. PREPARATION The first synthetic cornpounds of this t’ype were prepared by Schiff , 6 who obtained glasslike products by heating aniline or p-toluidine with dry D-glucose. By using a lower reaction temperature, Sorokin6isolated crystaltine N-phenyl-D-glucosylamine, -D-galactosyhmine, and -D-fructosylamine; he found also that further improvement in quality results from using a temperature of 130-135” and from adding hot ethanol after the reaction. This method has riot been widely used, part’icularlybecause Amadori found t’hat, in certain cases; “abnormal” products are obt,ained. Amadori studied t.he react,ion of D-glUCOSe with p-phenetidine,7 p-anisidine,* and p - t o l ~ i d i n e , ~ and suggested’O that these “abnormal” products have the open-chain, Schiff-base structure. Subsequent workza showed this t’o be incorrect and revealed t,heir true nature. The most generally used method of preparation, also described by Sorokin,6 consists in heating the amiiie and the reducing sugar in boiling methanol or ethanol, containing up to 10% of water. Small amounts of acids have been used successfully to catalyze the condensations. The product crystallizes on cooling, or after evaporation, if necessary, of some of the solvent, foilowed by the addition of ether. In this way, Sorokin obtained N-pheiiyl-D-glucosylaniirie and -D-galactosylamine in good yield and the -Dductosylamine in low yield. A large number of successful examples of the use of this method are described in t’he literature. Sometimes, con(5) H. Schiff, A,nn. Chen2. Phamn., 140, 123 (1866); 164, 30 (1870). (6) B. Sorokin, Ber., 19, 513 (1886), see J . Ch.em. SOC.,60, 526 (1886); J. Russ. Phys.-Chern. Soc., Chem. Pt., 1, 377 (1887), see J . Chew. SOC.,64,807 (1887) and Bet-., 20, (Rrfcrala), 783 (1887); J. prakt. Chern., [2] 37, 291 (1888). (This author is also referred t o as W. or v. Sorokin in some pubtications.) (7) M. Amadori, Atti reale accad. nazl. Lincei, [6] 2 , 237 (1925). (8) M. Amadori, Atti reale accad. nazl. Lincei, [6] 9, 226 (1929). (9) M. Amadori, Atti reale accad. nazl. Lincei, [6] 13, 72 (1931). (10) M. Amadori, Atti reale accad. nazb. Lincei, [6]9, 68 (1929).

98

G . P. ELLIS A N D JOHN HONEYMAN

densation proceeds in these solvents a t room temperature, with the product crystallizing out as i t is formed. This method was used by Irvine aiid Gilmour" * I2 t o prepare N-phenyl-D- and N-o-carboxyphenyl-D-glucosylamine. The method was extended by Irvirie and Hynd13 for preparing the crystalline N-o-carboxyphenylglycosylamines from D-galactose, D-mannose, L-rhamnose, and maltose, using cold aqueous ethanol (60% for maltose, 75% for rhamnose, and 80% for D-galac,tose and D-mannose), and evaporating the solvent, if necessary. Eerger and Lee14'l 5 used this met,hod with conspicuous success in preparing N-aryl-D-ribosylamines and -Larabinosylamines, and following t,his, Ellis and Honeyman16devised a general method for preparing, as characteristic derivatives, t,he crystalline N-p-tolylglycosylamines of the common aldoses (except L-arabinose). Two isonieric N-pheiiyl-D-ribosylaniines are known. Ellis and HoneymanI6 showed that one is obtained under anhydrous reactmioilconditions and the other when even small amounts of water are present. This discovery was extended to other sugars, and, by using anhydrous conditions, new, labile isomers of N-p-tolyl-D-glucosylamine and -D-galactosylamine were obtained. Kuhn and Strobele" were able to cause such weakly basic amiiies as o-nitroaniline to react with sugars by adding a 0.05 molar proportion of ammonium chloride and, in a similar manner, Frkrejacque18 used methanol containing 4 % of aqueous acetic acid. Hydrogen chloride was employed by Kuhn aiid Birkoferig t o induce reaction betJweenp-phenetidine and D-fructose, L-sorbose, and D-mannose. Weygand20introduced a simple general method, attractive fur large-scale preparations, which consists of dissolving the sugar in the minimum amount of hot water and heating with the arylamine for a few minutes longer than is required for achievement of complete miscibility; the product crystallizes on adding alcohol and cooling. Several N-alkyl-D-glucosylamines were prepared by Mitts and Hixon21by heating D-glucose with the amine containing a small amount of 0.5 N hydrochloric acid. (11) (12) (13) (14) (15) (16) (17) (18) (19) (20) (21)

J . C. Irvinc arid It. Gilmour, J . Chem. Soc., 93, 1429 (1908). J. C. Irvine arid R. Gilmour, J . Chein. SOC.,96, 1545 (1909). ,J. C. Irvine and A. Hynd, J . Ch.ein. SOC.,99, 161 (1911). L. Berger and J. Lee, J. Org. Chem., 11, 75 (1946). L. Berger and J. Lee, J . Org. Chein., 11, 84 (1946). G. P. Ellis and J. Honeyman, J . Chem. SOC.,1490 (1952). R. Kuhn and R. Strobele, Ber., 70, 773 (1937). M. Frbrejacque, C o m p t . r e n d . , 207, 638 (1938). R. Kuhn and I,. Birkofer, BeT., 71, 621 (1938). F . Weygand, B e r . , 72, 1663 (1939).

Eleanor Mitts and R. M. Hixon, J . AWLChem. SOC.,66, 483 (1944).

GLSCOSYLAMINES

99

recent paperz2gives the results of a study of tJhe formation of N - p sulfopheiiyl-D-glycosylami~iesin 0.01 M aqueous solutions of different pH values. React'ion proceeds best using excess of sugars a t pH 3 to 4, slowly at pH 7, and riot at all in alkaline solutions. Ammonium chloride shows no

catalytic: effcct under these conditions, but acids, particularly acetic avid, act favorably. Indirect methods have also been used. One, introduced by S a b a l i t ~ c h k a , ~ ~ consists of heating the amine with an O-acetylglycosyl halide and subsequently deacet,ylating the product. Other indirect methods for preparing aretates of glycosylainines are outlined on p. 112. A large number of N-su bst,ituted aldosylamines have been synthesized by the above methods, but only a few N-substituted ketosylamines have yet been made. These have been found difficult to prepare, and the yields have been poor. Recently, however, Barry and H ~ n e y r n a nhave ~ ~ described an .improved method for preparing N-phenyl- and N-p-tOlyl-D-frUctOSylamines, in which the amine and D-fructose are heated in anhydrous ethanol with a catalytic quantity of amine hydrochloride. Yields of up to 70% (based on the unrecovered sugar) result when the duration of reaction is short (seven minutes), and well characterized compounds have been obtained for the first t'iine. I n a similar manner, Helferich and PortzZ5have isolated N-p-hydroxyphenyl- and N-p-methoxyphenyl-D-fructosylamine in yields of about 20%. A study of the Tables at the end of this review will reveal that a wide range of amines has been used to prepare compounds of the type under discussion. They include ammonia, and primary and secondary aliphatic, alicylic, and aromatic amines, as well as many heterocyclic amines. The methods used for preparing arylhydrazones of sugars (and the properties of these compounds) have many similarities with those for glycosylamines. These hydrazones have been surveyed by PercivaLZ6 Under certain conditions, aldosylamines undergo a transglycosylation, in which the amine residue is replaced by another amine residue. Kuhn and DansiZ7 obtained N-p-tolyl-D-glucosylamine by reaction of N - ( 4 , 5 dimethyl-2-nitrophenyl)-~-glucosylamine with p-toluidine. Further exIr.

(22) Y . Inoue, K. Onodera, J. Shishiyama and S. Kitaoka, J . Agr. Chem. Soe. J a p a n , 26, 329 (1952); Chem. Abstracts, 48,2003 (1954). (23) T. Sabalitschka, Ber. deut. pharm. Ges., 31, 439 (1921); Chem. Abstracts, 16, 2009 (1922). (24) C . P. Barry and J. Honeyman, J . Che7n. Soc., 4147 (1952). (25) B. Helferich and W. Ports, Chem. Ber., 86, 604 (1953). (26). E. G . V. Percival, Advances in Carbohydrate Ckem., 3, 23 (1948). (27) R . Kuhn and A. Dansi, Ber., 69, 1745 (1936).

100

G. P. ELLfS AND J O H N HONEYMAN

amples are described by Iiioue and Onodera,28 and Bogn&r and N B n h ~ i ? ~ have recently investigated the reaction more fully. They found that the solubilities of the reactants and of possible products play an important part in the change, which is a true transglycosylation (and not hydrolysis followed by another condensation). The rate of reaction is increased by raising the temperature and by the presence of acid; good yields then result. In certain reactions, it has been found that the sugar can react with more than an equimolar amount of amine. For example, Mitts and HixonZ1 noted that the crystalline product they obtained from D-glucose and cyclohexylamine has the analysis required for a conipound derived from one mole of D-glucose and two moles of the amine. Recently E r i ~ k s o nin , ~con~ nection with the reaction of sugars with aliphatic amines, observed that ketoses condense more readily than do aldoses; that D-fructose readily gives with octadecylamine a white solid in the formation of which two amine molecules have reacted with one of sugar; and that solids are obtainable showing that four or five molecules of amine have reacted with one of D-glucose or of L-sorbose respectively. No information is available regarding the structure of these compounds (which are, therefore, not discussed further in this review). From the reaction between 2,3,4,5-tetra-O-acetyl-aldehydo-~-ribose and aniline in ethanol it was possible31to isolate a crystalline compound which, on elemental analysis, appeared t o be of the Schiff-base type (IV) with one molecule of ethanol of crystallization. Similarly, when the condensation was performed in methanolic solution the product corresponded to a compound of the same type with methanol of crystallization. CH=NCeH,

I I HCOAc I HCOAc I HCOAc

CHIOAC IV

,OMe(Et) CH, I NHC'Hs HCOAc 1 HCOAc

I

HCOAc

I

CH~OAC

v

The infrared absorption spectra of these compounds showed, however, that the compounds do not have the structure of type IV, but that the structure is in fact of the aldehyde-amine, addition type (V); that is, the compounds are analogous to aldehyde-ammonia compounds. The expected (28) (29) (30) (31)

Y. Inoue a n d K. Onodera, J . A g r . Chem. SOC.Japan, 22, 119 (1948). R . Bognhr a n d P. NhnAsi, Nature, 171, 475 (1953). J. G. Erickson, J . Am. Chem. Soc., 76,2784 (1953). P r i v a t e communication from Professor M. Stacey.

101

GLYCOSYLAMINES

absorption peaks were a strong band at 1745 cm.-' (caused by the acetyl groups), a strong band a t 1610 cm.-' for the

\

C====N.C~HE, grouping

/

(compare the strong band a t 1603 cm.-l in the spectrum of benzaldehyde phenylhydrazorie), and a weaker band a t 1600 cm.? (for a benzene frequency). The compounds prepared were found to absorb strongly a t 1745 cm.-' and less strongly a t 1600 em.-', but showed no absorption for a

\

C=N-

group. I n this region, the spectrum was the same as that for

/

acetylated N-pheriyl-D-ribopyranosylamine.

IV. PHYSICAL PROPERTLES The vast majority of glycosylamines are crystahine, but a few have still only been prepared in the amorphous state. A doubtful case is N-phenyl-Dglucosylamine, which Sorokin6 and Irvine and Giliourll claim to have isolated in crystalline condition, whereas Honeyman a n d TatchelP2 obtained it as a glass. An unusually large number of the compounds have been isolated with solvent, of crystallization; thus, B e n e and H 0 1 m s ~ ~ describe N-na-, N-o-, and N-p-tolyl-D-glucosylamines as aemihydrates. By using different solvents, Irvine and GilmourIl prepared N-p-tolyl-D-glucosylamine as, respectively, anhydrous, hemihydrate, arid moriohydrate crystals. Indeed,. the possibility exists that, in some compounds at least,, an extra molecule of water may be present which is not water of crystallization but which arises because water was not eliminated during the preparation. This means that the compound concerned may he an aldehyde-amine addition compound, analogous to aldehyde-ammonia compounds. This possibility requires investigation. * Nearly all glycosylamines are colorless, although t,he compounds made from nitroarylamines are yellow or orange.l6s 17' 3 4 , 35 Some are bitter to the taste.Ig Their stabilities differ considerably. The N-arylaldosylamines of aniline a.nd of the toluidines tJurn brown on normal storage a t room .tempera.ture for a month or so, and eventually decompose complctely to hlack, tarry subst,anc,es.Similar decomposition products from N-D-gluaosylpiperidine have been shown by Hodge and R i ~ t,o t contain, ~ ~ among other compounds, t,he Amadori rearrangement isomer and reductones produced (32) J. Honeymarl s n d A . R . Ttttchell, J. Chenr. Soc., 967 (1950).. (33) S. Bapne and W . H. Holms, J . C h e w . SOC.,3247 (1952). * T h e authors are indebted to John E. Hodge for t,his int.eresting suggest,ion. (34) P. Rfalnalis, V . Petrow arid B. Sturgeon, J . Pharnr. Pharmucol., 2, 491 (1950). (35) F. Weygnntl, W. Perkow and P. Kuhner, Chciri. B e r . , 84, 594 (1051). (36) J . E. Hodge and C. E. Rist, J . h r . Chew. Soc., 76, 316 (1953).

102

G. P. ELLIS AND JOHN HONEYMAN

therefrom. The stability of N-aryl-D-glucosylamines increases in this order: N-phenyl, N-p-tolyl, N-m-tolyl, N-0-t0lyl.33 Weygand20 states that decomposition is prevented by keeping the compounds in a desiccator containing a trace of ammonia. In striking contrast, the N-phenyl- and N-p-tOlyl-Dfructo~ylamines~~ and the N-nitrophenylaldosylamines can be stored for years without decomposition. Where decomposition is rapid, complete drying is difficult, and melting points are not necessarily of much value as criteria of purity; in fact, melting is generally preceded or accompanied by browning.6, 13, 14, 15 1 6 , 32 Most of the compounds are readily soluble in wat’er and in pyridine, slightly soluble.in methanol and in ethanol (which are the usual recrystallizing solvents), and sparingly soluble in ether, acetone. light. petroleum, chloroform, benzene, and ethyl acetate. N-Phenyl- and N-p-tolyl-D-mannosylamine, like D-mannose phenylhydrazone, have unusually low solubilities. The specific rotations of the compounds are important. The majority mutarotate in water and in alcohols, in striking contrast to the nonmutarotation of other sugar derivatives in which the reducing center is substituted. I

V. STRUCTURE 1. Introduction

When SchifP first condensed D-glUcOSe and aniline he naturally regarded the product as a Schiff base, with the acyclic structure VJ. Sorokin,6 applying Tollens’s suggestion that D-glucose is cyclic, envisaged a cyclic molecule (VII). Von Miller and PlO~ h l,~ having ? established that hydrogen cyanide adds to N-phenyl-D-glucosylamine, -D-galactosylamine, and -D-fructosylamine, considered that a double bond must be present and that these compounds are accordingly to be represented by formulas like VI. Marchlewski’s suggestion38that hydrogen cyanide could add to VII, also giving VIII, was later subst,antiated by VotoEek and W i ~ h t e r l e ,who ~ ~ showed that hydrogen cyanide adds not only to compounds like those examined by von Miller and Plochl, but also to D-glucopyranosylpiperidine (111),which cannot possibly have the Schiff-base type of structure. It has been shown by infrared spectroscopy that the

\

C-N-

linkage is absent in N-phenyl-

/

D-ribosylamine and 2-deoxy-N-phenyl-~-glucosylamine.~~ Isbell and Frush41 ( 3 7 ) W. yon Miller and J. l’lochl (with R. Straws), Ber., 27, 1281 (1894). (38) L. Marchlewski, .I. p r a k t . Chem., [2] 60, 95 (1894). (39) E. Votoitek and 0. Wichterle, Collection Czechoslov. Chem. C c m m m s . , 9, 109 (1937). (40) I<. Butler, S. Laland, W. G . Overend anti M. Stncey, J . Chenr. Sac., 1433 (1950); F. Smith. K. Butler, W. G. Overend and M. Stsce.v, Chemistry & Z n d u s t / y , 351 (1949).

103

GLYCOSYLAMINES

,_--___

“H

;o-.H

z j ~ ~ s ~ ,

HCOH

I HOCH I

HCOH

i I

HCOH CH,OH

CII=NC$I.,

I I

HCOH

-

I

HOCII

HFoH HCOH I

CHlOH VI

p

I

IHCN

I

CHNHGH~

I

HCOH I

H~OH

I

CHLOH 1.111

HCOH

I

CH:OH VII

have suggested that such additions take place through the intermediate formation of an imonium ion, the concentration of which determines the rate of addition. Such ions, discussed in the next Section of this Chapter, are obtainable from primary, secondary, and tertiary glycosylamines. The infrared spectra of solid N-2-naphthyl- and N-o-tolyl-D-glucosylamine are considered by Legay42 as giving evidence for these compounds’ having a Schiff-base structure. Both give a peak at 6.05 I / , usually caused by

\

C=N-.

Unfortunately, simultaneous demonstration of absence of the

/ \

NH group is not possible, so the spectroscopic evidence requires further

/

support. This peak is not found in the spectra of some other similar compounds, including N-p-nitrophenyl- and N-p-tolyl-D-glucosylamine. From the structural point of view, catalytic hydrogenation might be expected to give results similar to those obtained by the addition of hydro(41) H. S. Isbell and Harriet L. Frush, J . Research N u l l . B w . Standards, 46, 132 (1951). (42) F. Legay, Compt. rend., 234, 1612 (1952).

104

G . P. ELLIS AND JOHN HONEYMAN

gen cyanide to the glycosylamint. Iiarrer and c o ~ o r k e r have s ~ ~ shown that hydrogwation of N-alkyl- and N-aryl-D-gliIcosylamirles gives the corresponding N-subhtituted 1-amino-1-deoxy-D-glucitol (or “glucaniine”). They also discovered that N-p-tolyl-D-glucosylaniine tetraacetate can be hydrogenated, whereas, under the same conditions, N-methyl-N-phenyl-D-glucosylamiiie tetraacetate, which cannot exist in the Schiff-base form, was recovered unchanged. However, in the light of the information given in the succeeding Section, this reaction is of little value ill struc%ural studies, particularly as Kuhn and B i r k ~ f ehave ~ ~ found ~ that D-gliicosylpiperiditle can tie hydrogenated. 2 . Mutarotation and Hydrolysis

By analogy with the unsubstituted sugars, it has been suggested by various workers, notably Irviiie and Giltnour” and Kuhn and Birkofer,l9 44 that mutarotation is attributable t o an equilibrium involving partjial conversion of the compound into its anomer through the intermediate production of the acyrlic Schiff base.

I

HCOH

I

CJI,N=CH I HCOH I

~

-

1

HCNHChHi

HCOH

I

This possible mechanism is applicable only t o compounds having a hydrogen atom attached t o the nitrogen atom, yet Kuhn and Birkofer found that mutarotation occurs in pyridine solutioiis of derivatives of secondary amines, namely, D-glucosylpiperidiiie and N ,N-dibenzyl-D-glucosylamine, where 110 such hydrogen atom is available. To explain mutarotation in such cases they postulated the formation of an acyclic imoiiium ion by expulsion of a proton from a substituted arnmonium ion.

(CsHsCII,)zNCH HCOH

8

( CGH.,CH 2 ) jNCH

I

HCOH I

-

Hodge and R i ~ have t ~ ~fourid that carefully purified ~-glucosylpiperidii~e does not inutarotate in dry pyridine, and that the compound believed by Kuhn and Birkofer t o be N ,N-dibenzyl-D-glucosylamine had un~

(43) P. Karrer,.H. Salomon, R . Kunz and A . Seebach, Helv. Chim. Acta, 18, 1338 (1935); 20, 83, 1016 (1937). (44) R . Kuhri and I,. Birkofer, B(,r.,71, 1535 (1958). (45) J. E. Hodgc and C. E. Rist, , J . An2 Cheni. Soc., 74, 1494 (1952).

GLYCOSYLAMINES

105

dergone a11 Aniadori rearrangement and is, in fact, N ,N-dibenzyl-(1amino-1-deoxy-D-fructose). However, mutarotation does occur in authentic compounds of this type, for Hodge and Rist observed a mutarotation of both D-mannosyl- and D-galactosyl-piperidhe in dry pyridine. Since, in this solvent, the production of the cation by the mechanism postulated above cannot be expected, they suggested that an intramolecular rearrangement may have taken place. Pigman and c o l l e a g ~ e shave ~ ~ likewise shown that the stability of glycosylamines varies with the parent amiiie. Their results for simple cornpounds are in general agreement with the principle of Mitts and I-Iixon2Lwhich states that, except for D-glucosylamine itself, the ease of hydrolysis of glucosylamiries parallels the basicity of the amine. These studies included D-glucaosylamiiie,N - (n-buty1)-,N - (n-decy1)-, N-(n-hexyl) -,N - (n-octy1)-, and N-phenyl-D-glucosylamine, as well as amorphous N-(n-dodecy1)maltosylamiiie and impure D-ghcOSylglycine. By following the change in specific rotation and, where appropriate, the proportion of free amine, the amount of hydrolysis was determined. Acetic acid (3 N ) was found to be a general and very effective hydrolyzing agent, but 0.5 N hydrochloric acid was less effective. The effect of 0.01 N sodium hydroxide is especially dependent on the particular aryl or alkyl radical, for in that solvent the decornposition of N-phenyl-D-glucosylamine is arrested, whereas N-(n-butyl)-D-glucosylamine is hydrolyzed more rapidly than in 0.5 N hydrochloric acid. In general, in aqueous solution, a change in specific rotation usually occurs, to give an equilibrium a t which hydrolysis is not necessarily complete. For hydrolysis by acid, the products have been shown to be the ainine and sugar, but with alkali, although amine is released, the fate of the sugar has not been established. The sugar may also be obtained by decomposing the glycosylamine with benzaldehyde. Another examination into the stability of glycosylamines was carried out by H a n a ~ k awho , ~ ~ studied polarimetrically the rates of hydrolysis by dilute sulfuric acid of a number of N-aryl-D-glucosylamines, and then extended the work to such derivatives of other sugars. He found that whereas a hydroxyl, methoxyl, ethoxyl, or methyl group attached to the benzene ring reduces the stability, the preseuce of a chlorine atom or a carboxyl group increases the resistance of the compound to hydrolysis. A number of N-arylglycosylamines have been found to be hydrolyzed more rapidly than are glycosides, and N-aryl-deoxyglycosylanlines are even more readily h y d r ~ l y z e d . ~ " The change in rotation of aqueous solutions of N-p-tolyl-D-galactosyl(46) W. W. Piynian, E. A. Clevelaiid, D. H. Couch and J. H. Cleveland, J. A m . Chciti SOC., 73, 1976 (1951). (47) K. Hanaoka, J. Bzochetn. (Japan), 28, 109 (1938)

106

G . P. ELLIS AND JOHN HONEYMAN

amine aiid of '2-deoxy-N-p-tolyl-D-galactosylamine has been shown by Barclay, Foster and Overend4*to be due, a t least in part, to hydrolysis, since chromatography of the equilibrium solution showed the presence of free sugar and free p-toluidine when aqueous butanol was used to develop the chromatogram. With anhydrous solvent, however, no decomposition occurred. Paper chromatography of the products of reaction of reducing sugars with ammonia, using solvents containing ammonia for developing, shows the presence of a number of compounds which have been provisioiially identified as hexose-ammonia, glycosylamine and diglyc~sylamine.~~ Isbell and Frush4I carefully investigated the behavior of L-arabinosylamine in aqueous solution, and demonstrated the great importance of the pH of the solution. Mutarotation (without decomposition) and hydrolysis take place, the rate of each being dependent on pH. Mutarotation without decomposition, which becomes immeasurably rapid a t a pH below 7.8, is catalyzed by the hydronium ion but hardly a t all by the hydroxyl ion. The graph of the rate of hydrolysis of L-arabinosylamine against pH is a catenary, with the maximum rate a t pH 5 and virtually no hydrolysis below pH 1.5 or above 9. Isbell and Frush carried out a large number of determinations of the rates of mutarotation aiid of hydrolysis a t different pH values, and propounded mechanisms to account for their observations. In doing so, they made use of the ideas on the mutarotation of sugars put forward by Fredenhagen and Boiihoeff er50 and adapted for glycosylamines by Howard, Kenner, Lythgoe and Todd.51 Mutarotation of L-arabinosylamine is acid-catalyzed and therefore differs from that of sugars, which is catalyzed by both acids and bases. Isbell and Frush compared the mechanism of the acid-catalyzed mutarotations, dealing first with that of a sugar.

H

q

+

IIA e f I y T

I

HCO

I

Sugar Acid (any cyclic form)

HCO I HA

="y-""~-""q

= " oBCO q + HA

HCoHA*

I

H'ioHA

I

11

All open-chain forins

(48) J. L. Barclay, A . B. Foster and W. G. Overend, Chemistry & Induslry, 462 (1953) . (49) I . D. Raacke-Fels, Arch. Biocheni. and Biophys., 43, 289 (1953); Chem. A b stracts, 48, 4445 (1954). (50) H . Fredenhagen and K . F . Bonhoeffer, Z . p h y s i k . Chenf., A181, 392 (1938). (51) G. A . Howard, G. W. Keaner, B. Lythgoe and A . R . Todd, J . Chem. Soc., 855 (1946).

107

CLYCOSYLAMINRS

The first step is the addition of acid (catalyst) to the oxygen atom of the ring. This is followed by the formation of an open-chain form, through an electron shift in which an unshared pair of electrons of the oxygen atom of the hydroxyl group on C1 helps the release of electrons to the ring oxygen atom. All other acyclic forms arc quickly formed from this intermediate. The reverse process of c+yclizationalso proceeds and may involve C4 or C5, with the LY-D or 0-D configuration of C l . Consequently, a n equilibrium mixture is obtained. The rase of L-arabinosylamine and analogs will now be ronsidered. tl

czil t

r

,

HINCH -

HCO Glycosylamine

HC

J ' -+2H-

;

7

It

Acid

7-

I

4

I

I HCOHA' I

4

I HCO-

HCO I+

+ HA

I

All open-chain forms

This is exactly analogous t o the mechanism for the mutarotation of the sugars; the shift of the lone pair of electrons from the amino nitrogen atom results in the formation of the imonium ion,

\

C=NH*+, from which the

/

various cyclic and acyclic forms possibly present in the equilibrium mixture are produced. This mechanism, generally applicable to all glycosylamines (including those which are secondary, or even tertiary, amines), accounts for the sensitivity of the mutarotation t o acid and for the mutarotation of those compounds of this type which are tertiary amines. In truth, the imonium ion shown above is the same as that postulated by Kuhn and Birkofer (see p. 104), although the mode of its production is different. The mechanism for the base-catalyzed mutarotation of sugars will next be considered.

r

+ HB *

I HCO

HCO

I

Sugar

(cyclic)

I

HCO

I

@

B~HOCH I

i

HCO-

I

r

HOCH

--

!

;

1 HCO-

+ BQ

I

Base All open-cliain forms

Here, the first step is the combination of the anion (hydroxyl ion) of the base with the hydrogen of the hydroxyl group on C1, The resulting complex decomposes, with production of an acyclic form from which the aldehydo

108

G. P. ELLIS AND JOHN HONEYMAN

form of the sugar results by addition of a proton a t C4 or C5 and from which the various anoineric, cyclic forms are readily produced. Although a similar mechanism is possible for glycosylamines, bases have little efHc=N1’ fect on their mutarotations because the nitrogen atom has ! much less tendency than has the oxygen atom t o release a proton t o give the necessary ion shown on left. A similar step is I not called for in the mechanism proposed for the acid-catalyzed HCO@ mutarotation. I n suggesting a mechanism to explain the rapid hydrolysis of L-arabinosylamine within a limited pH range only (ca. 3 to 7), and the absence of hydrolysis in strongly acidic and basic solutions, Isbell and F r ~ s again h ~ ~ assume the acid-catalyzed formation of the imonium ion as the first step, accompanied by a side reaction leading to an ammonium ion. The next step, requiring hydroxyl ions, leads to the unstable aldehyde-amnionia compound, which decomposes to L-arabinose and ammonia. This allows the reaction to go to completion, because the ammonia is removed as ammonium ion. This sequence is presented here in a simplified form; for full details the reader is referred to the original paper.41

’9

HCNH,

H

HC=NH?

-

HCO I

I

L-Arabinosylamine

HB

OH

:

HCOH i

I

Imonium ion

I

HOCNH,

!j

-L-Arabinose

+ NHs

HCOH i

I

Aldehyde-ammonia compound

Hydrolysis does not take place in concentrated acid because of the low concentration of hydroxyl ions, nor in concentrated alkali because of the lack of imonium ions. These kinetic studies have been extended by Isbell arid FrushS2to the D-galactosylamines, where similar results were obtained. It is likely that similar mechanisms operate during the mutarotation and hydrolysis of other compounds of this type. Isbell’s work largely follows the earlier kinetic work carried out on the general acid-catalyzed condensations of nitrogen bases and carbonyl compounds, the original references to which are given by Hammett .53 There is no evidence to show that glycosylamines are hydrolyzed in such (52) Harriet L. Frush and I€. S.Isbell, J. Research Natl. BUT.Standards, 47, 239 (1951). (53) L. P. Hanimett, “Physical Organic Chemistry,” McGraw-Hill Rook Company, Inc , Kcw York, N.Y., and London, 1940, p. 329.

109

GLYCOSYLAMINES

non-aqueous solvents as alcohols and pyridine; in these, change of rotation is usually occasioned by true mutarotation. On long standing, some decomposition, accompanied by the development of color, does occur in such solutions, but Ellis and Honeyman16 have recovered unchanged N-phenylD-ribosylamine from the alcoholic solution a t equilibrium. The fact that mutarotation does take place in solution is a complicating factor in structural studies. Crystalline ‘Lethersand esters of glycosylamines” have been prepared, but there is always the danger that they are actually derivatives, not of the original compound, but of an isomer produced during mutarotation.

3.Meth ylation Early evidence that the open-chain formula is, in itself, inadequate to explain the behavior of the glycosylamines emerged from the methylation experiments of Irvine and his coworkers.” l2 64 N-Phenyl-D-glucosylamine was methylated to afford a crystalline tetramethyl ether which was hyCondensation of * drolyzed by acid to 2,3,4,6-tetra-O-methyl-~-glucose. the latter compound with aniline afforded the original ether, which consequently must be 2,3,4,6-tetra-O-methyl-N-phenyl-~-glucosylamine. CH20CH3 CHJ Ag?O

H

OH N-Phenyl-

D-glucopyranosylsmine

H OCHa 2,3,4,G-Tetra-O-methyl-N-phenylD-glucosyhmine CsHsNHl

hydrolysis

CHzOCHs

H

OCH,

2,3,4,G-Tetra-0-metliyl-D-glucose

Methylation thus points to N-phenyl-D-glucosylamine’s reacting in the cyclic pyranose structure and certainly lends no support for the existence of the acyclic form (VI) which would yield. after methylation and hydrolysis, penta-0-methyl-aldehgdo-D-glucose. lrvine and his colleagues conducted thcir methylation experiments under extremely mild condi(54) J. C. Irvine arid D. McNicoll, J. Chem. S o c . , 97, 1449 (1910).

I10

G . P. ELLIS AND JOHN HONEYMAN

tions, hut, even so, the yield, although unstated, was probaldy low. Ellis and Honeymans5 have recently converted N-phenyl-~-gluc~osylamille to the banie pyranose tet tamcthyl ether, rtsing gentle treatment ivith bodium hydroxide and dimcthyl sulfate. The yield was only 23 % ' of the theoretiral, SO that all that van be said is that some of the N-phenyl-~-gluvosylaiiiiiie reacts in the pyranose form, but the presence of other forms is not excluded. A siniilar conclusion is reached from the work of Kuhn and D a n ~ i 'who ~ obtained 2 , 3 , 4 , G-tetra-O-methyl-N-p-tolyl-~-glucosj7lan~irie by met hylation of N-p-tolyl-~-glucosylamine.The methylation method has proved to be of little use in this field for, a t best, the yields are low and, in several cases (such as the attempted methylation of the ~ - p h e n y ~ - ~ - r i b o s y ~ a m i i i e s by Berger and LeeI4), no identified product has resulted. The fully methylated derivatives, when obtained, are stable, sharp-melting, crystalline solids which mutarotate in methanol55 and in methanol containing a trace of but not in acetone nor in chloroform. During such mutarotations, the possibility of the pyranose ring's changing to furanose is excluded, because this change would have to be preceded by demethylation. The preparation of crystalline methyl ethers of N-phenyl- or N-p-tolylaldosylamine by condensation of aniline or p-toluidine with aldose methyl ethers proceeds, in many cases, readily arid quantitatively in methanolic or ethanolic solution, either a t room or reflux temperature. For this reason, such derivatives are frequently used to characterize aldose methyl ethers isolated during constitutional studies on polysaccharides. For D-glucose, for example, crystalline 2-O-methyl-, 6-0-methyl-,57 3 ,4-di-0-methyl-,b* 2,3,4-tri-0-niethyl-,592,4,6-tri-0-methyl-,60and 2 , 3 , 4 , B-tetra-O-methylN-phenyl-D-glucosylamine" have been prepared in this way. Although anomeric forms of N-arylaldosylamine methyl ethers are possible, such anomeric pairs have not been isolated. No similar derivatives have been obtained from ketose methyl ethers. 4. Acetylation

The acetates are the best known derivatives of the glycosylamines. Direct acetylation by methods involving hot acetic anhydride is often unsuccessful, leading only to discolored oils. The most generally useful method consists in treating the glycosylamine with escess acetic anhydride in pyridine solut,ion a t 0" for periods varying from several hours to two days. (55) G. P. Ellis and J . Honeyman, J . Cheni. Soc., 2053 (1952). 60, 837 (1928). (56) M. L . Wolfrom and W. L. Lewis, ,I. A m . Chem. SOC., (57) K. Hanaoka, J . Biochenz. (Japan), 31, 95 (1940). (58) D. J. Bell and G . D. Greville, J. Chem. Soc., 1902 (1950). (59) S. Peat, Elsa Schluchterer arid M. Stacey, J . Chem. Soc., 581 (1939). (60) S. A . Barker, E. J. Bourne and ILI. Stacey, Cheinistry & Industry, 7.56 (1952).

GLYCOSYLAMINXS

111

The solution is then poured into iced water, and after several hours the solid acetate is collected and recrystallized, often from ethanol or ethanollight pet,roleum. The yield of pure compound is usually high. In this way, all the hydroxyl groups are acetylated, but where there is only one hydrogen atom (on the nitrogen atom) it is not replared; that is, the 2,3,4,6-tJetra0-acetyl derivative is obt,ained from such compounds as N-p-tOlyl-6-Dglucosylamine. Under more drastic conditions, however, the hydrogen at,om on the nitrogen atom can also be substituted, as demonstrated by Fr&rejacque,61 who obtained the N-acetyl-t,etra-O-ac.etyl derivat,ives from N-p-bromophenyl-, N-phenyl-, arid N-p-tjolyl-D-glucosylaniine by heating t,hese a t 120" witth acetic anhydride containing sodium aretate. When the compound is a primary amine, one of the hydrogen at,oms att'ached to t,he nitrogen atom is also easily replaced by the acetyl group. Frush and Isbel15? found t,hat the acetylatioii of 0-D-galactosylamine for three days a t ice-salt. temperature, followed by one day at room temperature, leads to production of N-acetyl-6-D-galactosylaniine 2 , 3 , 4 ,6-tjetraacetate in high yield. The a anonier reacted similarly, but t,he product was isolated in low yield. Isbell and Frush4' showed t.hat L-arabinosylamine also behaves in this way. Although mild conditions have been used in the successful acetylations, a mixture of anomers is somet.imes obtained. Honeymari and T a t ~ h e l l ~ ~ obtained from N-phenyl-n-glucosylamine a mixture of tetraacetates which was separated by fractional recrystallization into the N-phenyl-a- bnd N-phenyl-P-D-glucopyranosylamine tetraacetates. An alternative and general met.hod of separation of such acetates was developed by Frbrej a c q ~ e61. ~In~this, ~ the mixture of aiiomeric acetates is dissolved in boiling tiarbon t.etrachloride, from which the 6-D compound separates as a cryst'alline complex with carbon tetrachloride. Evaporation of the mother liquor leaves the a-Danomer. A mixture of acetates is to be expected, since the initial, amorphous N-phenyl-D-glucosylamine is probably a mixture. Nevertheless, even when the starting compound is homogeneous, mixtures of anomers are sometimes obtained. Butler, Smith and Stacey62acetylated crystalline N-pheiiyl-D-galactosylamiiie and obtained a mixture of N phenyl-a- and N-phenyl-P-D-galactopyra~iosylaiiiiilebetraacetates although, when the theoretical amount of acetic anhydride was used, one anomer predominated. The converse has also been observed, for although Berger and Lee14 considered t>hatacetylation of each of their two distinct crystalline N-phenyl-D-ribosylarniiies gave a different glass-like triacetate, Todd and coworkersb1present evidence for believing t,hat these amorphous triacetates arc, in fact,, one compound. Thus, hydrolysis of t,hc aniline residue, followed by ncetylatioii of the result,ing triacet,at,e,gives, from each, D-ribose (61) M. FrBrejacque, C o r q t . rend., 202, 1190 (1936). (62) K. Butler, F. Smith and M. Stacey, J . Chem. SOC.,3371 (1949).

112

G. P. ELLIS AND JOHN HONEYMAN

1,2,3,4-tetraacetate. Consequently, when such changes in structure may accompany acetylation, caut,ion is necessary in applying the results of acetylation in structural studies. If mutarotation precedes acetylation, the structure of the derived acetate has little relevance to the problem of determining the structure of the unest,erified glycosylaniine. Indirect methods for preparing such acetates include the condensation of 0-acetyl-aldosyl bromides or aldohexose 2 , 3 , 4 , G-tetraacetates with the appropriate amine, usually in chloroform or ether solution. Another method, introduced by Frkrejacque,61consists in heating an aldohexose 1,2 ,3 ,4 ,6 pentaacetate in aqueous, ethanolic, acet,ic acid and adding the amine to the cooled solution. These methods lead to the isolation of the compound with the 0-D-pyranose structure. Frhrejacque examined some of these acetates and showed that, by keeping the compound a t its melting point for an hour, a mixture of the original compound and its anomer is obtained. These were separated by fractional recryst,allization, so making available several anomers previously unobtained. From the specific rotation then det,ermined, F r k r e j a ~ q u ereported ~~ that8 such acet'ates do not conform to Hudson's rules of isorotation, but berth^^^ has shown that the acetates of primary glycosylamines do conform t,o these rules, the contribution of C1 being 16,000. Helferich and PortzZ5found trhat addition of benzylamine t o a- or 0-D-glucopyranose peutaacetate removes the acetyl group on C 1, giving an addition compound of D-glucose 2 ,3 ,4 ,G-tetraacetat,e with benzylamine, which, on heat>ing, yields fv-benzyl-D-ghcosylamine t,et)raacetate. Another method for preparing this compound is of interest. Helferich and Mitrowsky6$found that D-glucosylamine tetraacetate readily gives Schiff bases on reaction with aromatic aldehydes. They prepared in this way N-benzylidene-D-glucosylamine tetraacetate which was readily hydrogenated in the presence of Raney nickel to N-benzyl-D-glucosylamine t,etraacetate. When a- or P-D-glucopyranose pentaacetate is treated with piperidine (t,hreemolecular proport,ions are best), deacetylation occurs on C1 and C2, whose and the product is N-(3,4, G-tri-O-acet,yl-D-glucosyl)piperidine,66 structure was proved by conventional methods, including its conversion into 2-O-methyl-~-g~ucose. The same triacetate is obtained when piperichloride or with D-glucose dine reacts witah 3,4,G-tri-O-acet~yl-~-glucosyl 2 , 3 , 4 ,G-tet,raacet'ate. Loss of t,he acetyl group on C2 must accompany or

+

(63) M. Frhrejacque, Conipt. r e n d . , 204, 1480 (1937). (64) A. Bertho, A . m . , 662, 229 (1949). (65) B. Helferich and A . Mitrowsky, Che,nz. Ber., 86, 1 (1952). (66) J. E. Hodge and C. E. Rist, J. A m . Chem. SOC.,74, 1498 (1952).

GLYCOSTLAMINES

113

precede formation of the cart,on-to-iiitrogeii bond, since N-(%,3,4,&tetrao-acet,yl-D-glu(,osyl)piperiditieis unaffected by piperidiiie. Deacetylat'ioii of the esters, which proceeds normally, has hecn successfully carried out with ammonia, sodium methoxide, or harium methoxide in methanol. In this way, for example, N-p-tolyl-p-D-glucosylamiiie 2 , 3 , 4 , (i-tetraacetate is converted into N-p-tolyl-p-D-glucosylamine, and N-acetyl-p-D-galactosylaniiiie tetraacetate into N-acetyl-p-D-galactOSylamine.5z All tfhe known crystalline acetJat#esof the glycosylamines, whether prcpared directly or indirect,ly, have the pyranose ring st,ructure. One reaction sequence carried out by Kuhn and DansiZ7is, from the s h c t u r a l point of view, typical of the method used to determine structure. Condensation of p-toluidine with D-glucose 2 , 3 , 4 ,6-tetraacetate or with 2,3,4,6-tetra-0acetyl-a-D-glucosyl bromide leads to the same compound, which is, without, doubt, N-p-tolyl-/3-D-glucosylamine 2,3,4,G-tetraacetate. The same rompound is obtained by the acetylation of N-p-tolyl-p-D-glucosylamine. In addition, Ellis and H ~ n e y m a nhave ~ ~ successfully used acid hydrolysis of N-p-t801yl-p-D-ghicosylaminetetraacetate (prepared from N-p-tolyl-p-Dglucosylamine) as a method for preparing D-glucose 2 , 3 , 4 ,&tetraacetate. Incidentally, t,his makes an att,ractivepreparative method for such acetates, alternative to that involving the 0-acetylaldosyl bromide. The various stages that have been achieved are shown on p. 114. A somewhat similar reaction sequence was carried out by Butler, Smith and StaceyfiZfor N phenyl-D-galactosylamine. The method has been extended to the ketose series by Barry and H o ~ i e y m a nshowing ,~~ that the crystalline tetraacetates, readily obtainable from N-phenyl- and N-p-tolyl-D-fructosylamine by acetylation in pyridine, are also pyranose. These tetraacetates are hydrolyzed to D-fructose ll3,4,5-tetraacetate. This tetraacetate does not condense with aniline, but 1 , 3 , 4,5-tetra-O-acet~yl-~-fructosyl chloride does, to give the above N-phenyl-D-fructosylamine 1,3,4 ,&tetraacetate as the only recognized product, in 10 % yield. One interesting and, at first, surprising instance was described by Zemp1611, Csuros and B r u c k i ~ e r who , ~ ~ found that trimethylamine reacts with the heptaacetates of cellobiosyl bromide and of lactosyl bromide, respectively. Later work by Zempl6n and BruckneP showed that the product of such reactions is the same as that obtained when dimethylamine is used. Thus, the cellobiose reaction product is N ,N-dimethylcellobiosylamine (67) G. ZemplBu, Z . Csiirijs and Z.Bruckner, Ber., 61, 927 (1928). (68) G . Zempi6n and Z . Bruckner, B e r . , 61, 2481 (1928).

114

G . P. ELLIS AND JOHN HONEYMAN

H OH iV-p-Tolgl-P-D-glucopyranosyhmine

>

CHiOAc

c-

H

H < AcO

H ,OAc

H

OAc

A’-p-Tolyl-P-Dglucopyi anosylamlne tetraacetate

OAc

D-ClucoW 1,2,3,4,6pcnt aacetate

tl H OAc ~ ~ - G l u c o sbromide yl 2,3,4.6-tetraacetate

H OAc D-Glucose 2,3,4,6tetraacetate

FIG.1

heptaacetate. This reaction appears comparable with that between tertiary amines and alkyl halides,69as exemplified by the following. 2 CsH5N(CHJ)2f C6HllBr + CsH6N(CHR)J3r f C6H5K(CI-T3)C.5H~1

The reaction betweeu tertiary amines and “acetobromo” sugars was further studied by F. Micheel and Hertha M i ~ h e e 71I ~who ~ ~ found that trimethylamine forms quaternary ammonium salts with some acetylated glycosyl bromides (for example, those of D-glucose and D-galactose) but not with others (for example, those of D-mannose and L-rhamnose). This was attributed t o the C2 hydroxyl group’s being on the same side as the oxygen hridge to C l in D-glucose and D-galactose, but 011 the opposite side (69) A C l a w and P. Rnutenberg, Ber., 14, 620 (1881). (70) F Michcel and Hertha Micheel, Be? , 63, 386 (1!)30) (71) F hIichce1 arid Herthn Michcel, Ber , 63, 2862 (1030).

GLYCOSYLAMINES

115

in D-mannose and L-rhamnose. The formation or nonformation of a quaternary ammonium salt is thus useful for determining whether a sugar has the Q or ,B configuration. Most glycosylamine acetates are readily obtainable in crystalline condition. Their stability is excellent; they can be stored indefinitely without occurrence of any decomposition. Since their melting points are sharp and reproducible, and their chloroform solutions do not mutarotate, they are derivatives suitable for identifying sugars and glycosylamines.

5. Benzoylation

The preferred method for the preparation of crystalline benzoates of g l y c ~ s y l a m i n e sis~ ~to treat the glycosylamine in pyridine with benzoyl chloride a t 0" for periods of up to four hours; longer reaction times are detrimental. I n one instance, reaction with benzoyl chloride in aqueous sodium hydroxide solution was used successfully, but the yield was lower than by the pyridine method. By these methods, all of the hydroxyl groups were benzoylated and the resultant ester was obtained by pouring the reaction solution into water and, if necessary, extracting with ehloroform and purifying and evaporating the chloroform solution. In this way, the tetrabeiizoates of N-p-tolyl-D-glucosylamine and N-p-tolyl-D-mannosylamine (and N-p-tolyl-n-xylosylainine tribenzoate) were obtained in high yield. From certain N-phenylaldosylamines treated in this way, benzanilide was the only crystalline product isolated. Similar treatment of N-phenyl-Df r ~ c t o s y l a m i n egives ~ ~ the tetrabenzoate in high yield, but a comparatively low yield (30%) was obtained from the N-p-tolyl derivative. The benzoates are more resistant to acid hydrolysis than are the corresponding acaetates,but by boiling with hydrochloric acid in acetone the amine radical is removed, giving good yields of D-glucose 2,3,4,6-tetrabenzoate, ~ - i n a i i n o x2,3,4,&tetrabenzoate, D-xylose 2 , 3 , -k-tiit)enzoate, and D-fructose 1,3,4,thtetrabenzoate, compounds of known structure, previously prepared and c h a r a c t e r i ~ e d . ~ ? - ~ ~ Benzoylation thus resembles acetylation in that the glycosylamine rcacts in the pyranose form. However, only one anomer of each benzoate has heen isolated. Again, this is an attractive method, alternative to the use of 0-bcnzoylaldosyl bromides, for preparing aldose benzoates with C1 unsuhtituted. (72) 5:. Fisclin :mcI €I. N o t h , Bcr., 61, 332 (1918). (73) R.I<, Nrss, H. G . Fletcher, J r . , and C . 8. Hutlson, J . A7n. C h ~ n r Soc., . 72, 3200 ( 1 9.50). (74) It. T.R1:Ljor :tiid I(;. W. Cook, J . .4m.C'ltc?,r. S o r . , 68, 2333 (1036). (75) P. Brig1 and R.Schinle, Ber., 66, 325 (1933).

11G

G . P. ELLIS AND JOHN HONEYMAN

6. ~ r i ~ ~ l ~ ~ i o n Attempts have been made to use the reaction of glycosylamines with triphenylmethyl chloride in pyridine7‘jfor elucidating structures. Kuhn and Strobele” claim to have obtained a trityl derivative in high yield from N-(4,5-din1ethyl-2-nitrophenyl)-~-ribosylamine and from the corresponding L-arabinose derivative. Unfortunately, no practical details or constants are given in their paper. The conclusion drawn was that, because trityl derivatives are produced, these pentosylamiiies are furanose compounds having a free primary alcohol group. For reasons given earlier in this review, this conclusion cannot be sustained and, indeed, it has been criticized elsewhere.32*62 Another important weakness in the use of tritylation for purposes of structural elucidation has been established by Zeile and K r ~ c k e n b e r g , ~ ~ who showed that, under the usual reaction conditions, the Amadori rearrangement takes place. Thus, the product may not be derived from the glycosylamine a t all. Nevertheless, Berger and Lee14attempted to use tritylation for establishing the structure of their two N-phenyl-D-ribosylamines, but their conclusions cannot be justified. Although of no structural value, the conversion of N-p-nitrophenyl-D-glucosylamine to the 6-trityl ether by Weygand, Perkow and K ~ h n e is r ~of~ interest because this glycosylamine does not undergo the Amadori transformation.

7. Periodate Oxidation The use of oxidation by periodic acid, sodium periodate, or lead tetraacetate in structural studies on carbohydrates is well known. These reagents act on a-glycols with the production of carbonyl groups and the consumption of one molecule of oxidant for each such glycol group. Where there is a primary amino group in the position a to a hydroxyl, similar oxidation 1

I

c=o

c=o I -C€I(OII)

I

--CH(OH)

[o]

H

+ I C=O I

H

I

+

I120

-CHNH2

I

-CH(OH)

[o]

+ I

+ NH,

c=o I

H

occurs. Although the reaction has not been so widely investigated in compounds having a secondary amine group adjacent to a hydroxyl group, Nicolet and Shinn78J9and Van Slyke and colleagues*”have shown that such (76) B. Helferich, A d v a n c ~ si n Carbohydrafe Chem., 3, 45 (1948). (77) K. Zeile and W. Iiruckenberg, Ber., 76, 1127 (1942). (78) B . H . Nicolet and L. A . Shinn, J. Am. Chem. SOC.,61, 1615 (1939).

117

GLYCOSYLAMINES

compounds undergo similar oxidative cleavage. By contrast, acyl derivatives of seririe are oxidized extremely slowly, suggesting that -CH(OH)82 CH.NH.CO.Ris not affected by glycol-fission These findings are substantially confirmed by quantitative studies on the periodate oxidation of glycosylamines. Niemann and Hayss3 prepared N-acetyl-D-glucosylamine, by the action of ketene on D-glucosylamine, and investigated its oxidation. Each mole of this N-acetyl-D-glucosylamine, known t o be cyclic because it gives a tetraacetate, consumed two moles of periodate. The product of this oxidation, when further oxidized with bromine water, gave a dibasic acid isolated as the barium and crystalline brucine salts, which, on hydrolysis, gave positive response to a qualitative test for D-glyceronic acid. The reaction scheme shows that both compounds

7

CHNHCOCH,

I I

HCOH HOCH

I I HCO I

HCOH

-

CHO

2 HI04

VHO

H$O

CHzOH N-Acetyl-Dglucopyranosylamine

HCO

I

HCOH I CHzOH

N-Acetyl-Dglucofuranosylamine

OBre

COzH HFO I

CHzOH

CHO HOCH

2

-1

7

I CHNHCOCHa I

CHNHCOCH,

HIO, CHO

HCO

I

C HO

CH~OH Dibasic acid

1OB~O

COzH

I

-

COzH

1

HFOH

~ H ~ O H D-Glyceronic acid

COzH

I I

HCOH COzH

COzH

Tribasic acid

D-Glyccraric acid

(the furanose and the pyranose) require two molar proportions of oxidant, but that only the pyranose form will lead to a dibasic acid which can give (79) L. A. Shinn and B. H . Nicolet, J . Biol. Chem., 138, 91 (1941). (80) D. D. Van Slyke, A. Hiller, D. A. MacFadyen, A. B. Hastings and F. W. Klemperer, J . Biol.Chem., 133,287 (1940). (81) B. H . Nicolet and L. A . Shinn, J. Biol. Chem., 142, 139 (1942). (82) T. Posternak and H. Pollaczek, Helv. Chim. Acta, 24, 1190 (1941). (83) C. Niemann and J. T. Hays, J. A m . Chem. SOC.,62, 2960 (1940).

118

G . P. ELLIS AND JOHN HONETMAN

D-glyceronic acid. Frush and Isbellbz performed similar oxidations on N-acetyl-a and N-ac~etyl-P-D-galactosylamine,arid found that each has a pyraiiose ring. They followed the periodate oxidations polarimetrically and noted that the final readings, when cdculated for the dialdehyde products, correspond to [ a ] D +70° for the a-D ariomer aiid [a]D -112' for the p-D anomer. These products result froin the Oxidation of any N acetyl-D-aldohexosylamine (and their eiiaiitiomorphs from the L series) and hence the above specific rotations can be used for correlating structures. In a similar oxidation of N-acetyl-L-arabinosylamine, two moles of oxidant were required, showing directly t,hat the pyranose, not the furanose, ring is present.

I

CHNHCOCHa

I

CHO CHOH

CHO

CHOH

I

CHi0-p N-Ace tyl-Larabopyranosylamine

CHNHCOCHa ;€€OH

I CHOH I CHO I

1

CHzOH

CHNHCOCHa CHO CHO

I

CHzOH

N-Acetyl-Larabofuranosylamine

Niemanii and Hayss4 later showed that the oxidation of one mole of N-acetyl-D-glucosylamine by lead tetraacetate involves the slow consumption of slightly more than two moles of oxidant, so confirming the result of periodate oxidation. Hockett arid ChandleF prepared an isomeric ~-acety~-D-g~ucosy~amirle, also cyclic since it gives a tetraacetate, by the action of 29 % aqueous ammonia on aldehydo-D-glucose pentaacetate or on hexa-o-acetyl-"a"-D-glucoheptonic nitrile. The oxidation of this isomer produced formaldehyde (isolated as its 2,4-dinitrophenylhydrazone),but (84) C . N i e m a n n and J. T. Hays, J. A m . Chem. Soc., 67, 1302 (1945). (85) R C. Hockett and JJ B. Chandler, J . A m . Chutr. Soc., 66, 957 (1944).

GLYCOSYLAMINES

119

the consumption of lead tetraacetate was greater than expected. From the graph published by Hockett and Chandler a t least 4.4 moles of oxidant were consumed. From the isolation of formaldehyde, they concluded that their isomer is N-acetyl-D-glucofuranosylamine, although the consumption of oxidant was clearly excessive for this compound. Niemann and Hayss4 prepared the new isomer by the action of methanolic ammonia on p-Dglucose pentaacetate, and found that 3.38 moles of lead tetraacetate per mole were consumed in oxidation. They believed that this observation could be explained by the actual oxidation using the expected two moles, followed by an additional reaction involving another mole of lead tetraacetate. However, in oxidation by periodate, five moles of oxidant were required, the reaction becoming slow in the later stages. Kiemaiiii and Hays concluded that the two N-acetyl-D-glucosylamines are certainly not anomers and that, possibly, the second is indeed a furanose compound. The opinion of the reviewers is that, in this second isomer, the N-acetyl group has failed to stabilize the ring, so that, although 011 acetylation the ring system is apparently retained, complete oxidation with lead tetraacetate or periodate takes place through an acyclic form. The oxidation of this N-acetyl-Dglucosylamine appears similar to that of N-arylglycosylamines described next. Howard, Kenner, Lythgoe and ToddE6applied periodate and lead tetraacetate oxidation to a number of N-arylglyrosylamines. They found that complete oxidation takes place, usually requiring four or five moles of oxidant, and presumably giving arylamine, carbon dioxide, formaldehyde, and formic acid. The method is, therefore, of no value for determining the structure of such compounds. By contrast, periodate oxidation of certain purine and pyrimidine nucleosides proceeds in the manner normal for a stable sugar ring, and so is helpful in determining the structures of these types of corn pound^.^^ A recent paper by Kawashiro,88the original of which is not available to the reviewers, gives details of the oxidation of N-m- and N-p-nitrophenylD-glucosylamines by periodic acid under the usual conditions. The degree of hydrolysis of the glucosylamines is less than 30 %, and exactly five moles of oxidant are consumed, with the production of the quantity of formic acid which would result from the similar oxidation of glucose. The reaction is given as follows. (86) G. A. Howard, G. W. Kenner, B. Lythgoe and A . R. Todd, J . Chem. Soc., 861 (1946). (87) J. Davoll, B. Lythgoe and A. R. Todd, J . Citem. Soc., 833 (1946). (88) I. Kawashiro, J . Pharnz. Soc. Japan, 73, 892 (1953); Chenz. Abstracts, 47, 12758 (1953).

(;.

1'LO

P. ELLIS AND J O H N HON EY M SN

RNHz HCO

+ (n+

2) HCOzH

+ CHzO

I

CHZOH

8. Conclusion Glycosylamiries and sugars are similar in that they mutarotate in solution. For the sugars, this behavior is explained by the establishing of an equilibrium mixture containing the various cyclic and open-chain structures. Derivatives of sugars are known having furanose, pyranose, and even septanose rings, as well as acyclic structures. For this reason, it is incorrect to refer to D-glucose, for example, as a pyranose compound, without specifying the conditions. Ordinary crystalline D-glucose is in the a-D-pyranose form, but D-glucose in aqueous solution consists of all the possible forms in equilibrium; it can react in any of these forms, and may react exclusively as any one form in any particular reaction. The work of Isbell and Frush on the mutarotation of aldosylamines shows clearly that in solution there is an equilibrium mixture of the possible cyclic and acyclic forms; this situation, with little doubt, applies to all types of glycosylamines which mutarotate. Thus, the aldosylamines may be expected to react to give derivatives which may be open-chain or have pyranose or furatlose rings, just like the sugars. The types of reaction which glycosylamines can undergo without decomposition are limited, so comparatively few crystalline derivatives haye been prepared. All that are crystalline and adequately characterized have been shown to be pyranoid. The conclusion is that in these reactions the glycosylaniines have reacted in the pyranose form. It is erroneous t o state that this means that the glycosylamines are exclusively pyranoid and will always react as such. Primary, secondary, and tertiary glycosylamines can all mutarotate, and can, therefore, all exist in the various forms. Where mutarotation is possible, periodate oxidation is expected to lead to complete decomposition; just as in the sugars, oxidation may proceed through the acyclic form. The N-acetylaldosylamines which retain their ring structure on oxidation (with periodate) do not mutarotate. Apparently, in these compounds the acyl group is able to stabilize the pyranose ring.

VI. DIGLYCOSYLAMINES By boiling a solution of D-glucosylamine in methanol for several hours, Sjollemasg obtained a hygroscopic precipitate from which he isolated crys~ +lo" (in tals, Cl?H?3NOlo.H@,having m. p. 132-134", [ a ] -20.75 Tvater). This compound, hydrolyzed by aqueous acid to D-glucose and (S9) B. Sjollema, Rec. trav. chim., 18, 292 (1899).

121

GLYCOEYLAMINES

ammonia, is di-D-ghCOsylamine in which two D-glucosyl radirals have each replaced a hydrogen atom of ammonia. The bame compound is obtained as a hyproduct from the deacetylation of a- or 8-D-glucose pentaacetate by ammonia in ~ t h e r , and ~ ” from the reaction of ammonia with tetra-0-acetylD-glUCOSyl bromide. Acetylation of D-gluqosylamine gives a small amount of a di-D-glucosylamine octaacetategl which is deacetylated to another di-D-glucosylamine, designated “a,” having m. p. 167-168”, [a]D -k 85.1” (in water). By following Sjollema’s method, Brigl and Keppler obtained the dihydrate of the “P” isomer, having m. p. 102-103”, [ a ] ~ 20” (in water). Treatment of tetra-0-acetyl-a-D-glucosyl thiocyanate with methanolic ammonia gives di-(tetra-O-acetyl-D-glucosyl)amine in 11 % yield.g2 Greatly improved yields (about 70%) of this octaacetate have been obtained by Helferich aiid MitrowskyC6by treating D-glucosylamine tetraacetate with 2,4,G-trinitrobenzaldehydein methanol or with n-butanesulfoiiyl chloride in pyridine. No further information is available as to the structure of the isomeric di-D-glucosylamines. The importance of formation of glycosylamines and diglycosylamines during the drying process in the detection of sugars by paper chromatography, when ammonium salts of weak acids are in the solvent, has been demonstrated by Bayly, Bourne and S t a ~ e y . ~Occurrence 3 of certain unusual “back spots” was traced to formation of glycosylamines. The ease of conversion of D-glucosylamine into di-D-glucosylamine was demonstrated, and an improved method was provided for preparing the “a”-di-D-glucosylamine octaacetate of Brigl and K e p ~ l e r . 9These ~ observations have some significance in the chromatographic detection of sugars in such biological fluids as urine.

+

VII. DIAMIDE DERIVATIVES OF ALDOSES Deacylat,ion of sugar acetaks and benzoates by alcoholic ammonia has frequently been observed to give, in addition to reasonable yields of the sugar, a small proportion of a product having two amide residues attached to C1 of the aldose. For example, the action of methanolic ammonia on aldehydo-D-glucose pentabenzoate was found to give n-glucose and some D-ghcOSe d i b e n ~ a m i d eSimilarly, .~~ Deulofeug5obtained L-erythrose diacetamide from L-erythrose triacetate. The reaction and its mechanism were (90) (91) (92) (93) (94) (95)

J. C. Irvine, R . T . Thompson and C. S. Garrett, J . Chem. Soc., 103,238 (1913). P. Brigl and H. Keppler, Hoppe-Seyler’s 2.physiol. Chem., 180, 38 (1929). A. Miiller and Adrienne Wilhelms, Ber.. 74, 698 (1941). R. T. Bayly, E. J. Bourne aiid M. Stacey, Nature, 169, 876 (1952). P. Brigl, H. Muhlschlegel and R. Schinle, Ber., 64, 2921 (1931). V. Deulofeu, J . Chem. SOC.,2973 (1932).

122

G . P. ELLIS A N D JOHN HONEYMAN

closely studied by Ishell and Frush,!6 who obtaiiicd L-arabinosc diawtamide (in 53% yield) from the action of ethanolic ammonia on aZdehydo-Larabinose tetraacetate. They compared this behavior with the conversion by similar means of 1 , 3 ,-k ,(i-tet,ra-O-acetyl-D-glucosamille hydrochloride into N-acetyl-D-glucosamine (2-acetamido-2-deoxy-D-glucose), in which an acetyl group is removed from an oxygen atom and has become attached to the nitrogen atom.97 The mechanism proposed by Isbell and Frush involves, first, the addition of ammonia to the aldehyde group, followed by the illustrated migration of an acet,yl group to the nitrogen atom through the intermediate formation of a cyclic orthoester. The monoacetamido OH

OH

IICNH

HCNH

0

It

HC

I

I

+ NH3

I

HCOCCH3

IH HCOCCH,

18

18

A

I\ II

HCOCCHl

OH

aldehydo- L-Arabinosc tetraacetate

Cyclic orthoeqter 4

OH

OH

I

I

HCNHCCH,

HCNHCCHI

I1

I

0

.PH

__t

__c

cH3vH

-

l oI1

CH,COCH I

hlonoacetamido derivative

H I

H I CSHCCHI

CH~CNHCNHCCH, II

II

CH,COH

I

0 -

HCOH

I

+

I

HOCH

CH

1

I

Cyclic orthoester

HCOH

”

L-Arabinose diacetarnide”

(96) H. S. Isbell and Harriet L. Frush, J . Am. C h e m . SOC.,71, 1579 (1949). (97) T. White, J. Chem. SOC.,1498 (1938).

123

GLYCOSYLAMINES

derivative now reacts further, with the addition of another molecule of ammonia to an 0-acetyl group. The amino group so produced replaces the hydroxyl on C1, with inversion of configuration there, to give a cyclic orthoester which rearranges to the diacetamido derivative actually obtained. In some instances, the reaction may stop a t the monoacetamido stage; this is an explanation for the isolation of N-acetyl-D-ghosylamine (the cyclic form of “D-glucose monoacetamide”), which has already been discussed (see p. 118). Free acetamide is not involved in this mechanism and, indeed, as cited by Hockett, Deulofeu and Deferrari,g* acetamide does not react with sugars or derivatives even when the latter are in the aldehydo form. Diacetamido derivatives of sugars are also important because they are produced in the Wohlg9degradation of an aldose to the next lower carbon analog. I n this method, the oxime of the aldose is converted by acetic anhydride to the acetate of the aldononitrile, which is then degraded by ammonia to the diacetamido derivative of the next lower aldose. Silver ions were originally used together with ammonia, but later work has shown x that ammonia alone is sufficient, although, if silver ions are included, a rapid precipitation of silver cyanide shows that the first stage of the reaction is the removal of the cyanide group. An example of this degradation is the following. ’

CN

I

HC04c

I

2,3,4,5,6-Pcnta-O-acct,yID-glucononitrile

-

HC<

NHAc‘.

1 NHAc

“

+ NH,CN + 3 AcNHa

D-Arabinose diacctamide”

Hockett, Deulofeu and Deferrarig8 have obtained valuable confirmatory evidence for Isbell and Frush’s mechanism by treating tetra-0-acetyl-Larabononitrile with ethanolic ammonia in which some of the nitrogen atoms were NI5, in the presence of excess acetamide having normal nitrogen atoms only. The proportion of N15 in the resulting L-erythrose diacetamide was practically the same as in the original ammonia. Had any condensation with acetamide occurred, a lower proportion of N16 would have been present in the product. Later work by Deulofeu and Deferrariloofurther supports this mechanism; they showed that D-glucose dibenzamide results from the respective action of ammonia on the pentabenzoates of a-D-glucopyranose, p-D-glUC0(98) R. C. Hockett, V. Deulofeu and J. 0. Deferrari, J . A m . Chem. SOC., 72, 1840 (1950). (99) A . Wohl, Ber., 26, 730 (1893). See also, V. Denlofeu, Advances in Carhohydrate Chern., 4, 119 (1949). (100) V. Deulofeu and J . 0. Deferrari, Nature, 167, 42 (1951); J . Org. Chem., 17,

1087 (1952).

124

G. P. ELLIS AND JOHN HONEYMAN

pyranose, and aldehydo-D-glucose. I n addition, no alteration in the yield of D-glucose dibenzamide from a-D-glucopyranose pentabenzoate is caused by the presence of a large amount of acetamide, and an acetamido-sugar compound is not obtained. Similar results are obtained for D-inannose and D-galactose esters,"J' although, in the debenzoylation of a- and p-D-mannopyranose pentabenzoates, equal quantities of D-mannOSe mono- and dibenzamide derivatives are obtained. These authors also pointed out that the over-all results, and the yields obtained, are the consequence of a series of competitive reactions, including ammonolysis of the acyl groups and the availability of a free aldehyde group on t,he sugar. In this context, it is noteworthy that aldehydo-D-galactose pentaacetate gives the diacetamido compound in a yield of 49 %, which is unusually high. An interesting variety of compound of this type has been obtained by treating N-methylsulfonyl-D-glucosylamine tetraacetate with methanolic ammonia.65The compounds are shown in acyclic form. OH

I I

HCNHS02CHa HCOCOCHX

I

VIII.

-

H

I I HCOH I

CHXONHCNHSOxC'Ha

USES O F GLYCOSYLAMINES

Oiie important use of glycosylamines, for the characterization of methyl ethers of sugars obtained during structural studies on polysaccharides, has already been mentioned (see p. 110). The condensation of D-ribose with an amine for synthesis of riboflavin is well known. In this connection, Berger and colleagues used glycosylamines in a practicable method for the separation of D-ribose from admixtures with D-arabinose. They found that there separates (in 87.5% yield) from Dribose plus aniline and sodium sulfate a complex salt of N-phenyl-D-ribosylamine with sodium sulfate; from this complex, N-phenyl-D-ribosylamine may he extracted (in 98 ?4 yield) with boiling ethanol., Similar complexes are obtained from D-mannose and D-lyxose, but not from D- and L-arabinose, D-galactose, D-xylose, D-fructose, and L-sorbose. A cis pair of hydroxyl groups on C2 and C3 of the aldose is probably necessary for complex formation. D-Ribose is readily separated from D-arabinose in this way and the free sugar, sufficiently pure for use, can be obtained after steam-distilling the complex Scdt in tlhe presenre of acetic SatoI03 used a similar (101) J. 0.Deferrari arid V. Deulofeu, J . Org. Chcm., 17, 1093, 1097 (1952).

(102) I,. Berger, U. V. Solmssen, F. Leonard, E. Wrriis and J. Lee, J . O i g . C h ~ n i . , 11, 91 (1946). (103) T. Sato, J . Cheni. SOC.Japan, 70, 312 (1949).

GLYCOSYLAMINES

125

method for separating epimers. He found that only the aldoses with cis hydroxyls on C2 and C3 give precipitates when the N-phenyl- or N-tolylaldosylamine is added to boric acid. I n the preparation of D-fructose by the hydrolysis of sucrose, Weygand, Perkow and K ~ h n e made r ~ ~ use of the ready reaction between aldoses and amines. Addition of m- or p-nitroaniline to the mixture of D-glucose and D-fructose gives a precipitate of the crystalline N-D-glucosylamine.'Removal of this, and of excess amine, leaves a solution from which crystalline Dfructose is easily obtained. The glucosylamines derived from several compounds of therapeutic interest have been prepared and studied. The sodium has the same in salt of N-(4-carboxy-3-hydroxyphenyl)-~-glucosylamine vitro tuberculostatic activity as has the parent 4-aminosalicylic acid.lo4 Water-soluble derivatives of therapeutic interest have been made from sulfapyridine by condensing it with D- or L-arabinose, L-rhamnose, D-xylose, D-galactose, or D-glucose.1O6The claim is made that the Schiff-base forms of glycosylamines are antioxidants,lO6 and plastics incorporating glycosylamines have also been claimed.lo7

IX. TABLES OF PROPERTIES OF GLYCOSYLAMINES The following tables summarize characteristic properties of glycosylamines (and their derivatives) that are the subject of this article. (104) G. Haberland, Arzneimittel-Forsch., 1, 298 (1951). (105) H. A . Shonle, U.S.Pat. 2,268,780 (June 6, 1942); Chem. Abstracts, 30, 3010 (1942). (106) I. Gubelmann, U.S. Pat. 2,396,097 (March 5, 1946); Chem. Abstracts, 40, 3471 (1946). (107) Marie B. Rousset and A. V. Keller, British Pat. 475,686 (Nov. 24, 1937); Chem. Abstracts, 32, 3514 (1938); The Challenge Adhesives Ltd., British Pat. 476,903 (Dec. 17, 1937); Chern. Abstracts, 32, 3864 (1938); W.P . Ericks and M. C. Whitaker, U.S. Pat. 2,389,723 (Nov. 27, 1945); Chem. Abstracts, 40, 1353 (1946); S. Sussman, U.S. Pat. 2,392,105 (Jan. 1, 1946); Chem. Abstracts, 40, 1256 (1946).

126

G . P. ELLIS AND J O H N HONEYMAN

TABLEI D-Arabinos(llat~iinesand T h e i r Ilcrivalives Compound

D-Arabinose-1-diacetamide n-Arabinosylamine N- (4,5-dimethyl-2-nitrophenyl) monohydrate triacetate (furanose ?) 2,3,4-triacetate N-3,4-dimethylphenylN-2-naphthylN -pheiiyl-

rW elting poini, "C.

130 N-p-phenylazophenyleneN-o-Arabinosylpiperidine

203 103-104

ences

-9.5

187 111 181-182 166 212 137-138 125-126 168-170 130 130

Refer-

[a]",degrees

-72 -20 -137 -1s

f 2

f3 f 5 f2

, -108.7 -34.0 4 +2.5 +8.9 4 -13.2 +68 --t -4.3 +27.4 + -8.0 +82 +1.5

-20

99

4

-5

MeOH C5H5N MeOH C 5H 5N MeOH

* References 108 to 238 appear in a separate list rtarting on page 165.

17 108* 17 17 108 109 110 57 15 15 15 15 111 57

TABLEI 1 L-ArabinosJjlamines and Their Derivatives ~

Com9ound

Melling goinl, "C.

Rqierences

Rotalion solvent

[elD, degrees

~

L-Arabinose-1-diacetamide L-Arabinosylamine N-acetyltriacetate N-carbamoyl-

triacetate

N - (4,5-dimethyl-2-nitrophenyl) -, I 1 triacetate

+

+150.9 +10.5 -140.8 +26 3~ 3 -7.8 +137 f 5 +142 $76 it 1 +71

+24 -90

+83.6 +146.5 - 10 -22.6

H,O H,O H,O CHClj H20 HrO

C sH jS MeOH-EtOH

CHClz CHCI, CHCIz C sH J CsHSN RleOAc CHC1, CjH5N CjH,?;

96

112 41 41 113 111 113 115 115 116 116 116 31

17 31 17

34 li 31

CHCIz CsHjN

17 34 17 117

CjH,jN CHCl, CjH5I; CHC1,

34 34 31 34

I27

iV-3,4-dimethylphenylN-methylN-(4-methyl-2-nitrophenyl)-, I triacetate N-(4-methyl-2-nitrophenyl)-, I1 triacetate

+9.8 83 +69.1 +89.6 +51.5 +51.9 +45.9 -10.9

GLYCOSYLAMINES

triacetate N-p-carbethoxyphenyl&p-carboxyphenylN-(4-chloro-2-nitrophenyl)triacetate, I I1 N - (3,4-dimethyl-2-nitrophenyl) -, triacetate N - (4,5-dimethyl-2-nitrophenyl)-, I

189-191 124 222-224 177-178 193 (dec.) 192 212 180-181 15C158 110 194 117 167 111 108-109 212 213-214 186 187.5 213-214 143-144 139 (dec.) 101 133-135 194 141-142 77-79

128

TABLE I1 ( C o n t i n i d ) Co inpound

N-2-naphthylN-o-nitrophenyl-, I triacetate

N-phenyl3-methyl ether 2,3-dimethyl ether 2,4-dimethyl ether 3,5-dimethyl ether N-p-sulf amylphenyl-

N-p-sulfophenylN-p-tolylL- Arabinosylpiperidine N , N'-Di-L-arabinosylbenzidine N,N'-Di-L-arabinosylurea hexabenzoate

164-1 66 159-161 151 154 173-175 90-9 1 202

103.&106 (dec .) 126 117 139 sirup 145;126 118 194 191 197 >300 124-126 104 86 (dec.) ca. 227 260-261 (dec.)

[ul,,, degrees

Roialion solvenl

CjHjN CjHjN

-109.1 $55.5 +133.8 f 1 f159 +21.1 -23 -140.6 -156 4 -150 -132 4 -78.6

CHCI, CsHjN CHC1, C 5H jN C5HjN CjHjN

+2.5

MeOH

-37.0

+

MeOAc

+ 10% HzO + 10% AcOH

H20 -28.8

H2O

+42.7 +I2

CjHjN-HyO H2O

+14

MeOH

-15.4

CsHjN

+62.1 +I63

HZ0 C jHjN

Rejerences

110 34 17 34 34 34 35 35 35 118 57 119 120 121 122 123 19 124 125 125 126 77 127 113 113

G. P. ELLIS A N D JOHN HONEYMAN

N-o-nitrophenyl-, I1 triacetate iV-p-nitropheny1-a-

Melling poinl, "C.

129

GLYCOSYLAMINES

TABLE I11 Gellobiosylanaines and Their Deriva.1ive.s Compound

Cellobiose-1-diacetamide heptaacetate Cellobios ylamine N-acetylheptaacetate heptaacetate N,N-dimethylheptaacetate hydrobromide N-ethylN - (N-ethylthiocarbamoyl) heptaacetate

Melting point, "C.

Rolnfion soluanl

Refer-

degrsks

196 180-182 (dec.) 246 188 196 200

-20 -3.3 20 -20.3 -7.5 -8.4 -9.5

+

HzO CHC13 H20 H20 CHCls CHCIs CHC13

129 129 128 129 128 129 64

198-199 (dec.) 20.5206 (dec.) 206 148-149 110-112 (tlec.)

-10.5 -11.1 -10.7 -7.5 -7.8

CHCla CHCI, CHCIa CHC13 HzO

68 67 130 67 128

198 196- 198

+30.7

N - (N-methylthiocarbamoyl) heptaacetate N-phenylheptnacetate N-p-sulfamylphenyl-, tetrahydrate

207-208 133-135 (dec.) 218 215-216

heptaacetate N-p-tolylheptaacetate

271-275 (dec.) 122-123 (dec.) 215

I)icellobiosylamine

170 200-210 (dec.)

+12.8 -62.3 - 16 -81 -88 -31.4 -65.8

ences

92 131

CHC1, H2O CHCI:g CSH5X H*O CsHSN H,O

131 128 61 124 124 124 128 61 128 128 128

7

+4.5

HrO

-

TABLE IV L-E:r.!jlhr.os,ylrLt?iineDerivatives Co nz pound

p itfelfrng o inf, .,c.

I,-Erythrose-l-diacetamide triacetate

210 147

[a]., degrees

Reference $~~~~~ --

+31.7

H?O

95 95

TABLE V D-Fructos!jlamines and Their Derivatives Compound

D-Fructosylamine N-p-ethoxyphenylN-p-hydrosyphenylN-phenyl-

felting poinl, “C.

[a],, , degrees

Rotation solvent

Reler-

144 152 (dec.) 151 147 (dec.)

- 187 -165 + -118 - 132 -109 -104.3 -185.5 -215.7 -181.1 -203.6 4 -161.0 -209.3 4 -203.8 -209.4 + -164.1 -220.6 -+ -202.5 -206.8 + -175.6 -149.6 -132.0

C jH 5N CjHbN CSH,N EtOH 90% E t O H 90% E t O H MeOH EtOH EtOH C5H5N 90% E t O H MeOH CHCls CHCls

-168 -88 -207.7 -176.9 -200.4 ---t -167.3 -141.0 -131.4 -50.62 -+ -4.34

EtOH C5HsN CHCIS CHCls HZO

19 25 25 6 6 6 6 24 24 24 24 24 24 24 125 126 24 24 24 24 132

1 50 149 (dec.)

1,3,4,5-tetraacetate 1,3,4,5-tetrabenzoate N-p-sulfophenylN-p-tolyl-

1,3,4,5-tetraacetate 1,3,4,5-tetrabenzoate N-D-Fructosplguanidine

151 100-102 >300 154 138 (dec.) 120 167.5 90 (dec.)

---f

-+

?

ences

TABLE \’I o-Ficcosylwnine Derivative Conzpound

o-Fucosylamine N-pheny-, 2,3,4-trimethyl ether

WeltiTg p o d , C.

degrees

[a],, ,

Rotnlion solvent

Reference

133-135

+76

EtOH

133

TABLE VII L-Fucosylamines and Their Derivatives Cont pound

L-Fucosylamine N-n-butylN-n-heptylN-n-hexylN-methylN-l-pentylN -phen yl2,3,4-trimethyl ether N-n-propyl-

Y e 1ling

point, “C

88-89 84-85 109-110 125 750.5 150-151 133-1 34 124-125

130

[a]., degrees

+lo2

+

-77

+49

Rotation Jolvcnl

MeOH EtOH

References

117 117 117 117 117 117 133 117

TABLE VIII D-Gdactosylawiines and Their Dsrivntivrs Compound

D-Galactose-1-diacetamide

pentaacetate

D-Galactose-1-dibenzamide D-Galactosylamine iV, N-diacetyl-2,3,4,5,6-peuta-O-acetyla-D-Galactosylamine ammonia complex N-acetgltetraacetate 8-D-Galactosylamine tetraacetate N-acetyltetraacetate D-Galactosylamine N-n-butyliV-p-(p-butylaminobiphenylsu1fone)N-carbamoylN-o-carbomethoxyphenylAV-(4-carboxy-3-hydroxyphenyl) N-o-carboxyphenyl-

N -p-carboxypheny1X - (3,4-dimethyl-2-nitrophenyl) -, tetraacetate N- (4,5-dimethpl-2-nitrophenyl) -, tetraacetate

Melting point, “C.

195-197 184 200 183a 203-204a 207 141 (dec.) 205’

,degrees

[&

Rotation solvent

+7.1 +9.3 -38.3 -149.6 -33.6 -6.2 f64.1 4 +58.1 -38.9

HfO HrO CHCl, CHC1, CHCL(?) CjH&

(?) +138 +194.9 +117.4 +62.2 -+ (?) +26.7

H,O H,O CHCl, H,O MeOH HnO CHC1,

0) CHC1,

References

101 134 101 134 134 101

112 135 F

--f

179-180 172-173 139 233 173

+9.8

+34.7

82-83 amorphous

152 180 (dec.)

152 154-1 56 19G-197 180

-11.5 +15 - 102 134 -17.3 4 + 4 . 3

+

- 16 -110 -149.2 -23.4

(?I H,O KtOH C SH jN FtOH FtOH XtOH CHC1, CHC1,

d

52

d

52 52 52 136a

5r:

.72

3 P 4 F

> 5 P

52

117 136b 113 137 124 13 137 115 31

34

r

53 1

TABLEV I I I (Continued) Comjound

N - p - (p-dodecylaminobiphenylsulfone)N-(N-n-dodecylcarbamoyl) N - p -ethoxyphen yl-

N - p - (p-ethylaniinobiphenylsulfone) N-n-heptylA - p - (p-hexylaminobiphenylsulfone) N-1- (2-hydroxynaphthyl) -N-phenylmethylN - p - (p-isobutylaminobiphenylsulfone) N-p-(p-isopentylaminobiphenylsulfone). N - (4-methyl-2-nitrophenyl) N-2-naphthylA-o-nitrophenyl-, tetraacetate N-p-nitrophenyl-p-

Melfing point, "C.

-13.0 165-168 165 155 (dec.) 140

- 102

1

Rofation solvenf

Peferences

(?I

136b 138 139 19 20 136b 117 136b 140 136b 136b 34 34 110 34 35 35 35 18 18 18 136b 117 136b 111 40 6 6 54 64 20

C 5H 5N

-20.0

0)

-11.5

0)

-15.0 -11.5

(?) (?)

-21.6 -130.7 -34.6 -187 -+ -248 -224 -+ -250 -206 + -244 -202 -73 +120 -12.0

CHC13 C sH sN CHCL, CsHsN CsH 5T\'-Hz0 C jH5N -AcOH CsH5N CHCls CHClz

-11.5

(?I

-+ -53 -31 -33 -88.5 + -7 -76.7 + -31.6

MeOH-HsS04 EtOH-H?O MeOH EtOH MeOH

99-100 206 (dec.)

204 (dec.) 190 138-1 40 178 219 203

tetraacetate N-acetyl-tetra-O-acetylN - p - (p-octylaminobiphenylsulf one) N-1-pentylN - p - (p-pentylaminobiphenylsulfone)N-4-phenylazophenylene2-deosy-hr-phenyIN-phenyl-8-

[ale, degrees

140

(?)

I10 205 134-135 (dec.) 147 (dec.) 151 144

-116

~-Galnctosy1amine,N-phenyl-6- (Continued)

PIIeaCO MezCO C jHsK

62 62 62 40 16 62 62 141 142 143 144 145 146 141 146 147 146 144 54 146 148 145 149 144

Me&O

150

151-153 -114.7

2,3,4,6-tetraacetate 2-methyl ether 4-methyl ether 2,3-dimethyl ether 2,4-dimethyl ether 2,6-dimethyl ether 4,6-dimethyl ether 2,3,4-trimethyl ether 6-trityl ether 2,4,6-trimethyl ether tetramethyl ether 2,3,4,6-tetramethyl ether

157-159 (dec.) 146148 (dec.) 157-158 (dec.) 120-121 165 168 130-131 216 121-122 207 169 170 152 179 170 192 197 195; 202 193-194 188 192 192-193

--t

-56

-92 -+ -37 -92 4 -43.5 -31.4 -58

CjH$ MeOH MeOH CHC1, C ,H jN

+119.4

EtOH

+I5 - 174

EtOH CjHjS

-68

MenCO

-92 -77.1

-+

+38

Me&O

-+

+37.7

MezCO

-80 -79 + +39 - 141 (initial value) -80 4 +44

8* 0

8 s+E

3

u,

c-’

W W

TABLE V I I I (Continued) Conrpound

Melting pain/, "C.

[alD, degrees

Rolation solvenl

Reference!

~

N-phenyl-a2,3,4,6-tetraacetate

141-143 17.5176

N-n-propglN-p- (p-propylaniinobiphenylsrilfone) N-p -sulfamylphenyl-

127-128

monohydrate N-p-sulf ophenylN-o-tolyl2-deoxy-.VV-ptolylN-p-tolyl-

aldeh?~do-D-Galactosylaniline2,3,4,5,6-pentaacetate N-D-Galactosglpiperidine

-22.1 --t -40.3 +202 t-238 -18.0

sirup 171-174 17GIi5

>300 11k-116 142-1 43 139 (dec.) 154-155 161-162 162-163 160-1 62 135-137 84-86 129 123-1 29

-41.0 --t -18.0 - 149 -34 -49.5 --t +10.5 -80 4 +8.8 -33 4 -14.1 -46 4 +10 -56.0 ---* -27.5

+60

-1

N-D-Galact,osylsulfapyridine a

Two isomers obtained in t h e same reaction.

155; 197

4

---* 4

4

0) C jH J C jH jiY H?O

-110 -97 -90

+10.0 +14.4 -10.2 +I6

MeOH CHCls C gH 6K

-3.5 -3.6 -20.7 -7 -25

MeOH C gH SIC MeOH EtOH-H20 McOH MeOH MeO1-I

MeOH CHCly R4eOH

MeOH CjHjX MeOH CgHjK

16 62 62 Ili 136b 125 124 124 124 125 33 151 6 20 16 16 126 33 152a 40 57 45 45 36 36 152h

GLYCOSYLAMINRS

L-Galactosylamine 2,3,4,6-tetrnmethyl ether

Compound

Gentiobiosy lamine N-p-tiit,rophenyl-P-

heptaacetate

~

197

Melting poinl, "C.

~

+70

[-I,, , degrees

Rolation solvrnl

RrJprrnrr

MczCO

152c

Rotation solsent

ReJerencr

35 35 35 35

154 141 136 215

N-6-Deoxy-~-glucosyltrimethyl~mmonium bromide

i 1 162

1-14.8

i

KtOH

I

153

TABLEXI1 o-Glucosylamines and Their Derivatives Co 111poibnd

N,~r-“~”-Di-(D-glucosyl)aniine octsace tate

N , N-“p”-Di- (D-glucosyl)amine, dihydrate octaacetate N,N-Di-(D-glucosj.l)amine nonaacetate N,hr’-Di-(D-glucosyl)benzidine N , N’-Di-(D-Rlucosylguanyl)-L-cystine N , N’-Di-(D-glucosylguanyl) -L-histidine N , N’-Di- (D-glUCoSylgUanyl)-oL-lysine N2, Ns-Di-(o-glucosyl) -DL-lysine N,N’-Di- (D-glucosyl)urea 2.5 water octaacetate octabenzoate D-Glucose-1-dibenzamide pentaacetate D-Glucose-1-di-(cyclohexylamine) N-D-Glucosvl-DL-alanine

D-Glucosylamine

Melling poinl, “C.

167-1 68 216-217 214-215 213-214 132-134 125-1 26 190-192 192 127 (dec.) 220 (dec.) 160 (dec.) 178 (dec.)

[&,

, degrees

+85.1 +87 +86.2 86 -20.8 + + l o 20 +7.6 -9.2

+ +

-34.5 230 (dec.) 235245 (dec.) 164 ca. 150 202 (dec.) 201-202 (dec.) 196 97-98 114

122-123 127-128 127-128

-35.8 -3.7 +19.9 +l.5 +l.3 -39.7 -23.5 + -11.6 +45.7 +24.6 (after 24 hr. a t No) +22.7 +19.4 +20.3 +22.1 +21.2

CHC13

91

H?O (?) H2O H20 CHCl3 CHCI,

100 89 9i 91 91 127 154 154 154 154a 155 113 155 113 94 100 100 21 13 13

Hi0 C sH jN CHCl3 C jH jS C5H5N CsH5N CHCl3 EtOH

H>O H,O HZO H2O H20 H?O MeOH

156 157 158 90

? a M

r

; *

3 2d 2,

X

0

z

4

FZ

128-129

N-acetyl- (furanose?) Kacetyl-

-..

(furanose?) (furanose?) tetraacetate

t,etraacetate hydrochloride N-p-acetoxyphenyl-, tetraacetate N-acetyl-N-p-bromophenyl-, 2,3,4,6-tetraacetat N-(2-amino-3,4-dimethylphenyl) -, tetraacetate N-(2-amino-4,5-dimethylphenyl) -, tetraacetate N- (2-amino-4-methylphenyl)-, tetraacetate N-o-aminophen yl-, 2,3,4,6-tetraacetate N - (2-benzisothiazolin-3-one-l , 1-dioxide)-, 2,3,4,6-tetraacetate [tetra-0-acetyl-D-glucosyl saccharin] N-benzyltetraacetate N-benzylidene-, tetraacetate n~-(N-bensyl-N-rnethyl)-, 2,3,4,6-tetraacetate D isomer L isomer

201-202 255 257 (dec.) 189- 191 193-194 121-121.5 82.5-84.5 160-161 159-160 126 127 ca. 170 133 134 128-129 128-129 130-131 182 154 81.5 110 162-1 63 98 93 98

+21.8

CbHaN

+18.8

aq. NaOH

+20.2 +17.4 4 +25.1 f 2 4 . 2 4 f53.5 +86.2 -22.4 -22 3 -23 +86.7 4 +85.8 f86.3 +37.2 +32.7 +17.7 +16.2 +11.1

HzO CHCIa CHCI 3 CHCl3 CHCI, MeOH

+48.4 -36.4 -30.6 -47.0 -89.5 -40.3

CHC1, CHCI, CHCI, CHCI, CHCli CHCl,

HzO aq. HC1 AcOH H20

HLO HzO H20

-22.7

MeOH

-29.1 +67.1 +9.6 4 +44.9 +25.2 -+ -9.7

AcOEt CHC13 MeOH MeOH

-42.7

3

90

46 46 46 46 100 112 91 85 84 84 85 84 91 64,136a 65 136a 65 61 34 34 34 116 159

160 65 65 16aa 160a 160a

TABLEXI1 (Continued)

F

02

[a].,degrees

Compound

~ - G h c o s y l a m i n e(Continued) N-4-biphenylN-p-bromophenyl-or-, 2,3,4,6-tetraacetate N-p-bromophenyl-8-, 2,3,4,6-tetraacetate 2,3,4,6-tetramethyl ether

N-n-butyl-

N-p-butoxycarbonylphenyldihydrate tetraacetate N-carhamoyl-

N-acetyl-tetra-0-acetyltetraacetate N-henzoyltetrabenzoate N-(4-carbet hoxy-3-hydroxpphenyl)N-o-carbethoigphen3.1N-p-car~ethox).pheii~l

118-120 (dec.) 134 160 154

97-98 96-97 88-89 88-90

125-128 85-86 127-130 207 (dcc.) 21S-217 200 CCI. 100 211-212 117 191 1S7 (dcc.) 137 174- 1’78

-93 -48.5 +240 +239 +259 -22 -24.2 +18.9 -9.5 -9.3 -2.9

Rolation solvent

+

$59.4 +59.4 +95.4 +139 +142

4

-7.8

EtOH

+7.7 +6.4 +31.2 +35.1 -51.5

H?O aq. NaOH nq. HCl a q . AcOH AcOH

4

--t --t --t

--f --f

--t

--t --t

CHCls-A4~OH CHClrAcOH AcOH-AcOEt AcOEt-EttO AcOEt-EtxO

-96.T -121.8 -92.0 -23.5 -23.4 -15.9 -6.9 -29.7 -128.5 - 136 +5.0 -23.2

MeOH C5H5K MeOH

IStOH

References

46 63 63, 161 162 162 162 117 21, 163 45 46 46 46 46 46 164 164 164 164 165 166 113 I13 113 165 103 124 57 I64

00

? c @ t3

E u: L-

3 4

0

5:

Z

EZ m

*z2

N-o-carbomethoxyphenyl-

178 181-182 112-1 15 192 187-189 (dec.) 145-146

tetraacetate N-p-carbomethoxyphenyl-

152 151-152 180-1 8 1

tetraacetate iV-(4-carbomethoxy-3-hydroxyplienyl)-

monohydrate phenyl ester sodium salt pentaacetate 2,3,4,6-tetraacetate

118-120 I52 140-142 (dec.) 142 (dec.) 117-120 135-137 (dec.) 185-186 (dec.)

123-127

-63.2 -90.6 - 54 -56.2 -101.9 -153.6 -98.2

- I43 -126.9 -72.3 - 133

131-132 (dec.) 137-138 180-182 181 80-85 184-186 122-125

-51.8 -99.0 -70.0 +87 4 -14.5 -13.4 -100.5 +65 + -9.5 +68 - 90 -63.9 +77

-50.6

c 5H jN NeOH CjHjN EtOH CSHjN KtOH CaHjN EtOH C5HJX c sH 6 ; v CjHSN CjHjS

164

CjK,N

164 10: 167n 167a 1673

H10 -4cOEt

167)) 101 1673

HJ)

c

51% 5

N

CHC'I, H,O (?) EtOH C sH SS EtOH-HnO EtOH-H,O CzHsN AcOEt EtOH EtOH

124 124 12 164 164 3:3 137 164 168 168 137 164

139

N-scetyl-, ammonium salt tetraacetate" N-m-carboy-phenpl-

- 145

-112

N-o-carboxyphenyl-

tetraacetate

+22

- 127

164 164 115 152n 161 104 124 164 164 13i 164 164 GLTCOSTLAMINES

tetraacetate N - (4-carbosy-3-hydroxyphenyl) hemi hydrate

+11.4 -32.7 -25.7

s

TABLEXI1 (Continued) Co inpound

H e l l z a g puinf, “C

0

,DI.[

degrees

Rolulion solvenl

Rejerences

D-Glucosylamine, N-m-carboxyphenyl- (Continued)

tetraacetate ,V-p-carboxyphenyl-

t,etraacet,ate 2-diethylaminoethyl ester, monohydrochloride (“Procaineh‘-n-gIucoside” or “Novocaini N-D-glucoside”) monohydrochloride, monohydrate

N-o-carboxyphenylsulfonylsodium salt N-2-chloroethylosycarbonyl-, tetraacetate N- (4-chloro-2-nitrophenyl)tetraacetate, I tetraacetate, I1 W-o-chlorophengl-

N-m-chlorophenyl-

109 122-123 (clec. 175-177 127 132-134 134-135 132 132-133 (dec. 155157 I 7 6 1 78

-96.9 +6 t 1 8 . 0 + -41.5 -89.5 -112 + -51.5 -65.6 -194 + +30 -68.6 -105 -+ -48.5 -107.7

CsHsN EtOH EtOH-HtO C SH sN NeOH EtOH MeOH EtOH-HyO EtOH-HLO CsH&

164 137 33 164 57 164 169 115 33 164 1703.

? ’d i?

E t

3 LI

2

130-1 31 140-141 142-144 (amorphous) 119 (dec.) 114 185 150 126 137 139-1 40 116-117

-100.7 -90.7 --* -74.0 (5 hrs.) -90 +2.8 -14.9 -11.6 +7.96 -63.4 +115.8 -50.5 4 +5.0 -72.0 -121.0

H2O H?O

170b,c 170d,e

82:

HyO HyO

170a 159 159 155 116 116 116 57 33 33

FZ

H20

Et.0 CsHt CHCI, CHC13 hfeOH MeOH MeOH

2

N-p-chlorophenyl2,3,4,64etraacetate 2,3,4,6-tetramethyI ether N-n-decylN , N-diethylN ,N-di-(2-hydroxyethyl) hexaacetate N , N-dimethyl2,3,6-trimethyl ether

126 120-1 22 147 141

N - (4,5-dimethyI -2-nitropheny1)-

(amorphous) 128 71.5 sirup b. p., 109"/0.1 mm.

157 157 213 214

tetraacetate tetraacetate, I tetraacetate, I1 N-2,3-dimethylphenyl-

+

-30

-22.5

+268 -+ +100.5 +266 + +155 +270 --* +146

MeOH MeOH AcOH-AcOEt EtONa-EtnO EtONa-Et20

103--104

hydrochloride hydriodide

N-(N,N-dimethylcarbamoy1)N-(3,4-dimethyl-2-nitrophenyl)-,tetraacetate

-40

132-135 168-169 150-151 154-1 55 105-106 95-97 84-102 110-111 145

t11.5 -22.5 -48 -7.2 t18.6 -9.4 -41.2 -33 -172.5 t-54.6 +11.7 $61.5 f 0.5 -64.3 - 62 +111

-104 -+ -46 -88 4 -30 -101 -+ -45 -95 + -25 -102.5 -+ -45 -103.5 -+ -25 - 107 -92.5 + -41.5 -79 + -29.5 -102.5 -+ -44

MeOH CsH5N CHCl3 MeOH H20 H20 H2O H*O CHCla EtOH-H2O CBH~N AcOMe CsHsN CHCls CHCI3 MeOH EtOH MeOH EtOH MeOH EtOH CsHsN MeOH EtOH MeOH

57 33 161 162 162 162 46 90 45 45 90 171 171 171 171 165 31 27 17 17 172 34 34 47 47 47 47 47 47 19 47 47 47

3 e 4 d

0 m r; d

>

5

3m

+

e

In]",degrees

R~iere,rre>

-

~

-96 4 , s diniethyl-l,2 AT,N'-phen!.lenedi-, octaacetatt [ ~ ~ , N ' - d i - ( t e t r a - O - a c e t ~ l - u - g l u c -4,5os~l) dimeth?.l-o-pheri3.lenedisnii1ie] N-1 -dodecylS-(3-etho.tyethylideiieamino-4 niethylphenyl)-, 2,3,4,6-tetraacetate ,\J-2-(l-ethoxyeth~4idenea1nino)phenyl-, 2,3,4,6tetraacetate .V-o-ethosyphenylS-p-ethougphenyl-

tetraacetate A-ethylA-(Ar-ethylthiocarbamoy1)-, tetraacetate Xl-heptylN-l-hexadecylN-1 -hexyl-

AV-(2-hydroxy-3,5-dinitrophenyl)4,6-O-ethylidene-2,3-oxidoethylideneS-2-hydroxyethyl.V- (2-hydroxv-5-nitrobenzylidene)

IS1

--t -30 -19.4

47 34

105.5 138

-65.7

4G 116

189

-91.7

116

157 110-120 (clec.) 11s 115-116 92 132 107- 105 157-1 59 97 97-98 105105 79-80 93-95

-96.1

4

-35.3

c

-80.5 -22 -28.6

+

-27.4

-12.5 -12.3 +15.4

-13

+

4

+

-7

4-10.1

+

t29.5

-25.3

4

13.5

165

200 11S116 116 (dec.) 170

173 12 20 25 PO, l i l 90 90 92 117 21 21 117 46 65 175 163 45

65

N-o-hydroxyphenylN-p-hydroxyphenyltetraacetate pentaacetate N -3-hydroxy-4-propoxycarbon ylphenylN-(4-iodo-2-nitropheny1)tetraacetate N -0-methoxyphenylN-p-methoxyphenyl2,3,4,6-tetraacetate 2 , s ,4,6-tetramethyl ether

N - (4-methyl-2-nitrophenyl). tetraacetate, I tetraacetate, I1 N-methyl-N-phenyl-, 2,3,4,64etraacetate N-methylsulfonyl-, tetraacetate N-(N-methylthiocarbamoyl)-, tetraacetate N-[2-methyl-3- (0-tolylazo)phenyl]N-l-naphthylN-2-naphthyl-

-+

-+

- 126 +8.6 -69.8 -38

+297 f259 f257

---t -+ -+

+lo7 +I57 +145

AcOEt EtONa-EtsO EtONa-Et?O

78-80 207 215 (dec.) 120-128 1so 129 102 160-162 182-1 84 135 92 117 113-120 (dec.) 172.$1 73 70-75 184 172-186 178

-30.3 -31.8 -51.7 +107.7 49 $51 +10.3 $13.1

+

-86.5

-+

-111

-+

-58.0 -48.1 -133.5

-136.3 -114 +26.1 -75.2 -+

-171

CHCI, CHCl p AcOEt AcOEt-HCI CHCl3 CHCI, RleOH AleOH C&H,N (?)

C5HbN AcOPvIe

19 65 25 65 124 116 116 173 8 161 162 I62 162 117 176 165 113 34 34 34 161 161 65 92 177 57 12 178 178 17 17, 34 39 35

143

2,3,4,6-tetraacetate N-o-nitrophenyltetraacetate N-m-ni trophenyl-

+I2 -68 -12.4

-88 -89.4

CLY COSYLAMINES

N-methylN- (N-methylcarbamoyl) -

148-150 148-149 140 133 135-137 (dec.) 136 (dec.) 135 146 86 129 110

TABLEXI1 (Continued) Con+ound

Melting point, "C.

[a],, , degrees

n-Glucosylamine, A-m-nitrophenyl- (Continued) -169 tetraacetate N-p-nitrophenyldihydrate

2,3 ,4-triacetate 6-trityl ether tetraacetate N-acetyl3-methyl ether, monohgdrate

N-(N-1-octadecylcarbainoyl). N-l-octadecylN-l-octglN-l-pentylWphenyl-

175 136 175 184

219 80-140 180 155 161 182

181-189 (dec.) 103.5 102 9697 crc. 147 (dec.)

147 (dec.) 110-150 (dec.) 140

-1

-81.5 -212 -192 --t -202 -150 + -215 -125 --t -161

CjHJ-AcOH CjH5N CHClr CsH5N CjHj?; CjH 5SHrO C jHjS-AcOH

- 120 - 101 +lo0 -319 --t -192.5 -198.5 4 -196.7 -215 --t -198

CjHjX CHCI. CHClz C jH 5N C5HjX-HjO CjHjN-AcOH

--t

- 132

-100

Rolalion soloenl

Reievences

35 18 18 18 35 35 35 35 35 35 35 18 35 35 35 135 46 46

-22

--t

-44 -47

-8

4-10.2 --t -52.3 -15 + -52 f11.25 + -51.9 +14.8 + -14.3 +37.2 -+ -85.6 +36.9 +27.7

EtOH EtOH MeOH MeOH MeOH MeOH H20 NaOH-H?O HCl-HJO HCI-HrO

21, 117 6 6 11

23 32 46 46 46 46

l-

r. e

134-135

tetraacetate N-acetyl2,3,4,6-tetraacetate, a 2,3,4,6-tetraacetate, p-

2-methyl ether 3-methyl ether 4-methyl ether 6-methyl ether 2,4-dimethyl ether 3,4-dimethyl ether 2,3,4-trimethyl ether 2,3,6-trimethyl ether 2,4,6-trimethyl ether 2,3,1,6-tetramethyl ether

131-136 9696 118 100 143 149-1 50 98 97 97-98 161 154-155 182-1 83 130 196 2 19-220 177-178 145-116 140 sirup 160-162 163-165 135 135

137-139 13s 133-135 134-135 N-1-phenJ-lztzophenyleneN - (I\i-plienylcarbamoyl)2-deosy -h-phenyl-

199 223 (dec.) 193-194

+53

+10.5

+

-20.6

+76.1 +180 + +41.6 +180 -73.7 -54.8 + t 4 1 . 6 -57 -106.6 +38.3 -108.5 + -50.3 -84.0 4 -39

+30 f 3 - 166

-79 -81 +238.4 +224 4 +17 +285 + +lo6 +266.6 + +162.9 +290 + +160 +228.7 + +58.9 +266 + +58.3 +227.8 + f 5 7 . 5 240 +238.5 -114

.--)

+

--t

CHCl, CHCl,-ScOH CHCI, EtOH CHCl1-AcOH CHCl, AIeOH SIeOH MeOH

MeOH EtOH

MeOH MeOH AIe2CO% MeOH AcOH-AcOEt EtOSa-EtiO EtOSn-Et~O AIeOH-HC1 MeOH-HCI MeOH ;LIe2C0 CHClj

H,O

-55

-138

16 33

-52.2 -50.9 --*

-106

c 5H 6N

23 179 161 63 32 161 63 32 57 1so 1803 57 181 lROa 58 59 152 141 183 183 184 11 162 162 162 185 56 55 55 55 111 165 156

0

2

0

% 2

* 3P

F

5

Y

?P iil

[elD, degrees

Compound

o-Glucosylamine, A-phenyl-2-deosy- (Continued) 3,5,6-trimethyl ether N-2-phenylethyl-, monohydrate N - (N-phenylthiocarbamoyl)-, tetrascetate N-p-sulfacetamidophenyl2,3,4,6-tetraacetate :V-p-s:ilfamylphenyl-

18&189 135-136 92-93 110 (dec.) 159 197-198 (dec.) 195 204 210 204

tetraacetate hexaacetate N-p-sulfamylphenyl-or-,2,3,4,6-tetraacetate N-p-sulfamylphenyl-~-,2,3,4,6-tetraacetate N-p-sulfophenylN-thiocarbamoylN’-benzoyl-, tetrabenzoate N-o-tolpl-

N-m-tolyl-

189 115 204-205 (dec.) 204

>300 210-212 (dec.) 215-216 205 101 97-98 97 95-96 117

106-107

-153 -64

-102 -60 -19.2 -25 ---t +2 - 22 - 56 +12.8 4

---t

- 123 - 125 - 122 -119.6 +29.7 -86 77 203 +197 -81

+

+

- 36 -35.7 +45 -99 --t -51 -103 4 -22

-79 -102.9 -102 -97.5

+

+

+ +

-50 -50.3 -32 -49.5

Rolalioa solvenl

CsH5Y IIeOH EtOH CsHsN CHCl3 H,O

CsH,?j HLO aq. SstlCOa

H?O H?O aq. HC1 CjH5N C5HsS C sH s N CHCI, C5HsS

H2O H,O C .H sN XeOH EtOH

MeOH MeOH EtOH hIeOH

40 40 187 36 65 125 121 19 19 19 125 188 188 19 124 124 124 124 125 92 166 113 47 47 20 173 33 47 47 33

t, Y

t 2:

3

N-p-tolyl-

N -p-tolyl-cutetraacetate N-p-tolyl-Bhemihydrate monohydrate

135-136 125 117-119 (dec.)

-94.6 + -21.1 --t -97.6 + -92.5 ---t -101.2 +

-82 -85

3,4,6-triacetate 2,3,4,6-tetraacetate

,V-acetyl2,3,4,6-tetrabenzoate

133-134 147 143-146 148 143-1 44 144-145 145-146 141-1 42 147-1 48 146 142 209

2-methyl ether 2,3-dimet,hyl ether

15G151 151

-47.3 -45 -45.2 -35.5 -45.8

-47.6

MeOH EtOH MeOH MeOH CHC13-AcOH MeOH MeOH MeOH EtOH MeOH

-40 -44.5

HzO EtOH CHCl,

-57.5 + +34.2

AcOMe CHClrAcOH

-33.3 -34.2 -35.0

CHC1, CHC13 CHClz

---t

-26.6 +64.2 t14.1 +50.0 EtOH

12 12 12 27 32 20 9 9 189 161 27 63 20 32 32 55 179 189 61 55 55 21 135

147

+26.6 (equilibrium value)

6 6 12 16 63

GLYCOSYLAMINES

115-120 (dec.) 114-115 117-1 18 112-1 13 115

-43.9 -38.8 +l81.9 + -45 $208.9 + -44.6 4-119 + +34.2

TABLEXI1 (Continued) c Rololioia solvenl

n-Glucosylamine, N-p-tolyl-P- (Continued) 2,3,4,6-tetramethyl ether

I44 147-150 151

151-152 192 (dec.)

.v-p-tolylsulfonyl2,3,4-triacetate, 6-p-toluenesulfonate tetraacetate 6-p-toluenesulfonate N-u-Glucosylhenznmide, 2-thioet hyl ether N-n-Glucosylcarhnmic acid, ethyl ester meth?l ester pentyl ester N-~-Glucosyl-3-carbanioyl-l,2(or 1,6)-dihydropyridiiiium bronlide 6’-tetraacetate 2’, 3’ ,4’, i~-~-Glucosyl-3-carbamoylppridinium bromide

N-n-Glucosyl-3-carhethoxy-l, 2-dihydropyridine, 2’, 3’, 4’,6’-tetraacetate ~-n-Gliicosyl-3-cyano-l, 2-dihydropyridinium bromide, 2‘, 3’,4‘, 6’4etraacetate N - ~ - G l ~ c o ~ y2-dihydropyridine, l-l, 2‘,3‘,4’,6‘tetraacetate

I9i 174-176 163 132 186-189 (dec.) 66-72 75-8 1 88

203-205 157-158 192-200 151-152 (dec.) 146.5

156 154156

+I63 --+ ? f272 -+ +I10 +249 + f153 +220.5 -+ +157.5 $221.4 --+ +60.0 +207.0 -67.5 --+ +10 -67.5 -+ $10 + +42.5

MeOH AcOEt-AcOH

EtONa-EtZO EtONa-EtzO MeOH CHCl,

EtOH-H20 EtOH-HCl

+33.5 f29.6 -5.5

CsHsS

-24.5 -13.7 -6.6 +4 .9

HrO H.?0 Et,O H2 0

-11.1 -18.3

+28.6

CHCl, CHClx

Refeerniaces

13 27 162 162 162 55 55 27 27

hb

00

? ’j M F

$ 65 65 65 65 94 189a 159a

155 190 190 190 190Q 190

190 190

* 15 cl

Z

m

0

e:

5z

ethyl ester sodium salt N-n-Glucosylguanidine N-D-Ghcosylguan ylgl ycine N-D-Glucosylguanylglyc ylglycine N-~-Glucosylhydantoicacid ethyl ester, tetraacetate potassium salt N -n -Glucosylhydantoin n-Glucosyl isocyanate, tetraacetate D - G ~ U C isothiocyanate OS~~ tetraacetate

N-n-Glucosyllysine N-n-Glucosylpiperidine

hydrochloride 2,3,4,6-tetraacetate 3,4,6-triacetate 2-carbanilate 2-p-nitrobenzoate 2-p-toluenesulf onate 2-methyl ether

+ +18.2 +24.8 -5 -8 + +20 4-31.96 + -0.57 -115

-6.6

N-D-Glucosylglycine, barium salt 108 94 (dec.) 208 (dec.) 180 (dec.) 169.5170 149-149.5 134-137 (dec.) 27G271 117-1 18 112-114 111.&113 100 75 (dec.) 115 129-130 130 (dec.) 144 150-151 123; 136 125 (dec.) 164 158-159 (dec.) 154 (dec.) 113

-25.8 -3

+5 -7.5 -12.8 +6.02 -8.5 -16.9 -43 + -13 +8.5 +3.0 +4 4 +33.5 -1 + $33 +31.6 +37 f52.3 17 +23

+

46 46 190b 154a 132 154 154 191 191 191 191 166 189a 191 166 166 192 19 20 45 45 45 45 193 66 66 66 66 66

d

84 F

>

z

M

TA

TABLE XI1 (Continued) Compound

Herling point, "C

[ a l,degrees ~

Kolalion S d U e n l

____

N-o-Glucosylpiperidine (Continued) 2-carbanilate 2-p-toluenesulfonate 2-methyl ether 3-methyl ether tetramethyl ether N-o-Glucosylpyridinium bromide, tetraacetate N-D-Glucosylpyrrole N-D-Glucosyl-DL-serine N-D-Glucosylsuccinamide tetraacetate N-o-Clucosylsuccinimide dihydrate N-o-CIucosylsulfapS.ridine N-D-Glucosylthiocarbamicacid, ethyl ester, tetraacetate N-D-Glucosylthiohydantoicacid ethyl ester ethanolate tetraacetate potassium salt

N-~-Glucosyl-2-thiohydantoin N-D-Glucosyltrimethylammoniunibromide 6-trityl ether N-D-Glucosyltrimethylammonium chloride, tetraacetate N-D-Glucosyltrimethylammoniumchloroplatinate, tetraacetate N-D-Glucosyltrimethylammoniuniiodide N-D-Glucosyltrimethylammoniumpicrate tetraacetate N-D-Glucosyltrimethylarnmoniumperchlorate, tetraacetate 0

3

Ll

0

152 (dec.) 111-112 (dec.) 114 13Q-131 74 174 190

+63 3-4

-2.5

-15 203-204 192 88-90 103-104 159-160

4 -6.0 +8.8 +91.7 -6.43

+19 +12.8 -17.4 --t

+11.5

CjHsN CsHsS C,HsS C sH sS MeOH

HLO

H,O (CHC1,)i

HZO

(CHCI?)*

66 66 66 36 60 194 195 154a 166 166 166 152b

z

113

>

3

? 3 F F

2:

3

2 152-1 55 119-1 21 151-152.5 137-141 (dec.) 224-225 161-162 183-185 173

-31.3

+a +22.8

+5.0 +14.8 4-6.26

EtOH

191 191 191 191 191 196 153

HZO

196

HzO CHCI,

H?O HrO

209-210

196

162-163 144 133 190

196 196 196 196

The elementary analysis given by the authors does not accord with this formulation.

2

2,

z

2 3

5z

151

GLYCOSYLAMINES

TABLEXIIA D-Idosylamine Derivative Con,pound ~~

D-Idosylamine 6-deoxy-N-phenyl-, 3-methyl ether

1

1

62-63

I

~

196a

TABLEXI11 Lactosylaniines and T h e i r Derivatives Compound

f e l l i n g poinf, "C

degrees

Iefer:nces

(?I

-98.3

+

+24

CHCla-AcOH

192

-14.3

-+

+24

CHC13-AcOH

63

HL' H,O CHCli MeOH

HZO

165

-89

H,O

I04 167b

-21.4

C6H6

130

+2.1

30-240 (dec.

208-21 2 154 17-110 (dec. 139 (dec.) 197 152

N-p-sulfamylphenyl trihydrate

190 210-212

N-p-tolyl-a-, heptaacetate N-p-tolyl-@-,heptaacetate

189 202 208 169-170

Lnctosyl isothiocyanate, heptaacetate N-Lactosylthiocarbamic acid ethyl ester, heptaacetate

Rololion solvenl

44-148 (dec. 209

246-248 142-146

+39.5 +38.5 +1.5 f 0.5 4-2.7 +7.4

-26 +90.2 $21 +lo1 -31 -31 4 +21 --f

N -phen y1-p-, hep twncctate

-

12 128 1961) 196b 196b 46 63

Lactosylamine N-acetyl-, dihydrate heptaacetate, monohydrate N-4-biphenylN-p-bromophenyl-a-, heptaacetate N-p-bromophenyl-p-, heptaacetnt e N -carbarnoylN-(4-cnrboxy-3-hydroxyphenyl) sodium salt phenyl ester, tetrahydrate N , N-dimethyl-, heptaacetate N-l-dodecylN-p-ethoxyphenylN-phen yl -a- , heptnacet at e

,

'[a],

119

-69 -79 182.3 + +24

-29

-+

+24.

46 19 61 63 61 63 152b 124 C 6H 6N H?O 124 CHCls-AcOH 63 61 CHCla-AcOH 63 114

C,H,N CHCl2 CHCla-AcOH CHCl3 CIIC~:S-ACOII

114

-

152

G . P. ELLIS AND JOHN HONEYMAN

TABLE XIV 1)-Lyxosylamines and Their Derivatives ~

Melting

Compound

point, “C.

[a]” , degrees

Rotation soluenl

References

______

D-Lysosylamine iV-3,4-dimethyIphenylN- (1,5-dimethyl-2-nitrophenyl) 2‘,3 ‘ , 4’-triacetate N-p-nitrophenyl-

142-143 146-147 198-199 190-191 143-145

-109 f 3 -154 f 2

C5H5X CsHsN

197 197a 108 108 198

TABLEXV L-Lyxosylartziiie Derivatives Compound

Melting

poinf,v-,

L-Lyxosylamine N-(4,5-dimethyl-2-nitrophenyl)- 195196 2’, 3’, 4’-triacetate 184-185

[=ID, degrees

Rotatiotz sotuenf

Refer-

+112 f 2 +155 f 2

C5H5N C5H5N

108 108

ence

TABLEXVI Maltosylamines and Their Derivatives j f e l l i n g point, "C.

N , N'-Dimaltosylbenzidine Maltosylamine heptaacetate N - (4-carbomethoxy-3-hydroxypheny1)N - (4-carboxy-3-hydroxyphenyl)N -0-carboxyphenylN-l-dodecyl-

175 ( d e c . ) ca. 165 ( d e c . ) 191-191.5 235-240 13&145 153-155 4&54

Rotation solvent

Rejerences

+I3 +I18 +73.7 -35.7 -42.4 t48.9 -+ +68 +67 +73 +72 -+ +86 +68 +65 +97.4 +92 +93.4

HzO Hz0 CHC1, C 5H sN C sH sN MeOH Ha0 HCl-H*O AcOH-H~O AcOH Hz0 HCL-HzO

127

+76.5 +37.5 +92.5 +36.9 - 186

H2 0 CHC13-AcOH CHCla CjHsN

+39 +94.4 +39.3

CHCls-BcOH CHC13

[&,degrees

-+

N-l-hexylN-l-octadecylN-phenylheptaacetate N -p-sulf amylphenylN-p-tolyl-, heptaacetate

-+

8&115 90-100 205 212-214 236 182

-+

-+

112 64 104 104 13 46 46 46 46 46 46 46 57 63 61 124 199 63 61

0

3 g 4 t?

k

z

M

m

TABLE XVII D-Vannosylnmines Coiiipound

N,N’-Di-(D-mannosyl)urea D-Mannose-1-diacetamide D-Mannose-1 dibenzamide pentabenzoate D-Mannosylamine N-benzoyltetraacetate N-n-butylN-p-carbethoxyphenylN-o-carbomethoxyphenylN - (4-carboxy-3-hydroxyphenyl). sodium salt phenyl ester N-o-carboxyphenylN-m-carhoxyphenylN-p-carboxyphenylN-(3,4-dimethyl-2-nitrophenyl) tetraacetate N - (4,5-dimethyl-2-nitrophenyl)triacetate, 6-trityl ether tetraacetate N-3, C-dimethylphenylN-p-ethoxyphen yl-

(irirl

T h e i r Derivatives

Melting p o i n f , “C.

[aID, degrees

188 219 21&219 226 (dec.) 22.5226 140-142

-45.8

H,O

-13.8 +3.6 t2.8 +25.3

HZO CsH5N C 5H 5N

+6.4 -28.8

CjH5N CHCl,

+7.2 -54 - 191 -97.5

EtOH C 5H 5 s

ca. 254 253-254 135-136 71-72 179-180 177-1 78 140-240 (dec.) 191.5-193 126 136 182 215-216 (dec.) 154-155 213 (dec.) 130 218 18&185 157 (dec.)

-29.4

+ -21.1 - 10 t35.7 -28.5 258 -35.0 -41.1

-

--t

CHC1,

EtOH

H,O

hIeOH EtOH EtOH CHCl, CHCla C 5H 5N C sH 5N AcOMe

-93.8

- 174 -155

Rolafion solvent

-145

C 5H 5N C 5H 5N

Pejerences

165 94 iai 94 101 101 94 101 101 117 115 137 104 104 167h 13 13i 115 34 34 34 17 17 I7 17 15

N-l-hexylN-1- (2-hydrosgnaplithyl) -M-phenylethylN-(4-niethy-2-nitrophenyl)tetraacetate N-2-naphthglN-o-nit rophenyLj3tetraacetate N-rn-nitrophenyl-pN-p-nitrophenyLj3dihydrate

tetraacetate N-l-pentyl-

N-4-phen ylazophenylene N-phenyl-

2,3-dimethyl ether 2,3,6-t rimethyl ether

2,4,6-trimethyl ether 3,4,6-trimethyl ether 2,3,1,6-tetramethyl ether

75 207-208 (dec .) 205-206 (dec.) 144-145 195-196 196 214-21 5 126 127-128 199 209 219

184 70-71 247 181 (dec.) 180-1 81 181 sirup 127-128 131 234 134 140- 143 112-143

144

-97.8 -224 -22.5

- 107 - 103 - 187 -336 -406 -+ -325 -306 4 -282 -333 3 -322 - 150

-178.5

4

-81.5

-179.3 -101.4 -+ -45

-155

4

-39

-150 -+ +8 +154.5 +55.5 -87.9 3 - 8 . 3 -95.5 ---t -38.9 -8 (final value)

CHCli CJHJN CbH5N CHCL CHCl, C5H5N C5Hhh’ C )H sN CjHJ”H3O C~ H ~ N - A c O H CHCL,

CsHjS C 5H 5 s MeOH

MeOH MeOH MeOH MeOH Me&O MeOH-HCl

117 140 34 34 110 18 34 18 34 18 18 35 35 35 18 117 111 54 20 16 16 144 200 20 1 144 202 202 54 54

56 20 1

156

TABLE XVII (Continued) Compound

didring point, "C.

[LII],,,degrees

Rolalion solvent

Yeferences

A-p-sulf amylphenyl-

monohydrste N-p-sulfophenylN-p-tolyl-

tetrascetate 2,3,4,6-tetrabenzoste N-D-Mannos ylguanidine N-D-nIsnnosylpiperidine

14G-145 145-146.5 202 204 194 >300 184 183-184 175 183 165-167 133-134 80 (dec.) 11G-117 11.5116 (dec.)

-84.0

4

-7.5

MeOH

- 163 - 103 - 186

C jH S I X B,O C jH sN

- 181

CsHjN -12 -45.0

MeOH MeOH

-125.6 +10.03 4 -0.72 -17.5 4 +12 -21.6 4 +13.3 -27.7 + +24.2

CHClz HzO MeOH MeOH C SH sN

-44 -101.4

+ --*

202 203 19 125 124 125 204 20 126 16 126 16 132

60 45 45

G . P. ELLIS AND JOHN HONEYMAN

D-Alannosylamine, N-phenyl-,2,3,4,6, - tetramethyl ether (Cont i m e d )

TABLE XVIII L-Rhanmosulamines and Their Derivatives Coin pound

L-Rhamnosylamine 0.5 MeOH 0.5 EtOH N-n-butylN-p-carbethoxyphenylN-o-carboxyphenyl-

N-p-carboxyphenylN , N-dimethyl-, 0-isopropylidene acetal N - (4,5-dimethyl-2-nitrophenyl) triacetate, I triacetate, I1 N -ethyl N-l-heptylN-l -hexylN-l-(2-hydroxynaphthyl)-N-phenylmethylN-methylN- (4-methyl-2-nitrophenyl) triacetate N-o-nitrophenyltriacetate N-m-nitrophenylN-p-nitrophenyltriacetate N -1-pen tyl-

Melfing p o d ,

T.

116 (dec.) 80 136-137 194 167-1 68

165-166 169-170 b. p. 82-84 a t 1 mm. 169 106-107 141-142 138 132-133 192 (dec.) 126-127 215 (dec.) 161-162 225 (dec.) 185 150 208 209 139-140

Rolafion solven

+38 +28 f14 +66.4

$51.2 +42.9 +148.8 + f100.2 f 52 -28.6 -20.2 --t

Hz0 H20 EtOH MeOH EtOH CsH& EtOH EtOH Hz0

References

112 112 126 115 13 13 13 137 115 205

g 4 4

+100.8

CHC13

+104.9

CHCh

f117.6 +191 +320 123

CHCla CsHsN C5H;N CHCl,

+

;:

117 117 117 140 117 34 34 34 34 18 18 l8 117

+

P

B

+ u1 ~

TABLEXVIII (Continued) Comjound

N-phenyl-

2,3-dimethyl ether

2,4-dimethyl ether 2,3,4-trimethyl ether

N -4-phenylazophenyleneN-n-propylN-p-sulfamylphenylN-p-sulf ophenylN-p-tolyl-

'otalion soloenl

Melting point, OC.

118 121-127 (dec.) 144 138-139 135-136 141-142.5 141-142 111-1 13 111 227 145 208-210 (dec.) 181 >300 151-154

-50.4 +77.1 +136.9 --f

EtOH EtOH

206 118 54

207 208 +147.8 +128.5 +110 +138.5 +138.3

+42.8 +5.6 4 +7 4 +16.9 +46.9 +

+

--f

+lo1

+92

+

+80

EtOH EtOH EtOH EtOH Me&O

H20

MeOH

209

m 196a 54 54 210 111 117 211 125 125 126

TABLEXIX o-Ribosylamines and Their Derivatives Compound

n-Ribosylamine N- (2-amino-4,5-dimethylphenyl) -, 2,3-di-O-acetyl5-0-tritylN-p-carboxyphenyl-a-, (pyranose) N-o-chlorophenyl-a-, (pyranose) N- (4,5-diethyl-2-nitrophenyl)-, (pyranose) N- (4,5-dimethyl-2-nitrophenyl) -

Melling p o i n f , "C.

N - (4,5-dimethyl-2-thioformamidophenyl)-, 2', 3' ,4'triacetate N- (4-ethyl-2-nitropheny1)N-ethyl-N-2-nitrophenylN- (3-hydroxy-4-methylphenyl)-,(pyranose), 2 EtOH N-4-methoxyphenyl-a-, (pyranose) N-1-naphthyla-, (pyranose) N-2-naphthyl-, (pyranose) N-o-nitrophenylisomer A isomer B N-phenyl-, Ac

Rotalion solvenf

References

C sH jN

212 172

137-138 (dec.) f10.7 129-130 (dec.) 152-153 171-177 164

2,3-di-O-acetyl-5-0-trityltriacetate 5-trityl ether N-3,4-dimethylphenylisomer Am isomer B*

[a]., degrees

163 118 (dec.) 128-130 (dec.) 110-112 (dec.) 9&98 190.5194.5 130-134 (dec.) 133-135 (dec.) 109-110 (dec.) 146-147 (dec.) 119-120 183-185 (dec.) 167-168 193-194 138-140 (dec.) 126-127 123-124

+231 +136

---* ---*

+70.2 +125

-85 +go 68 160 24 +172 +171.7 4 +56.5 +94.5 4 +53.0

+ + +

-107.5 +116 4 +32.4 +I22 + +40.8 +I22 4 +29.2 +96.6 - 109 -122.5 -109.1 +176.5 -+ +156.6 +180 +180 4 +161 +I82 .+ +52.3

C sH sN C gH 5X C gH gN C5H5N C 5H jN AcOMe C 5H :N

CsH5N C 5H 5 3 C 5H 5N C gH jN

15 15 213 17 172 17 172 17 14 14 172 214 212 15 15 15 15 15 16 16 14 51 51 40

0

r

2

3s

k 3

;R

CL

cn a

TABLEXIX (Continued) Melting point. "C.

Compound

[4,, degrees +135 4-12 +176.4 +134.9 -+ +14.1 +63.4 f48.6 $60 60 +48.8 -+

133-134 (dec.)

Bd, hemihydrate

125-127 (dec.) 119 114-116

-+

+

-+

+62 --* +50 +23 +13 +60.2 +23.8 +8.3 32 ---f

112-114 (dec.)

-+

N-p-tolyl-

resinous 56.5 165-166 168 172-173 169-170 1712172 130 (dec.)

2 EtOH 0.5 HzO

102-103 (dec.) 123 (dec.)

triacetate 2,3,5-trimetJ1iylether 2-deoxy -N-phenyl-

+

+19.5 +17.4 f141 -+ +47 +164 -+ +64 +136.2 -+ +12.5 +178.2 +76.0 f53.2 +58.6 +22.1 -+ +11.7 -+

2~deoxy-N-p-tolylN-~-Ribosyl-3-carbamoylpyridinium bromide

167-168 141-142 (dec.) 147 (dec.) 170 142

N'-acetylN-~-Ribosyl-3-carbamoylpyridine 0 Claimed t o be furanose. some authors.

b

Claimed t o be pyranose.

c

Claimed t o be furanose by some authors.

Rolalion solocnl

References

MeOH C 5H sN MeOH C 5H sN C6Ht.N CSH sN-HZ0 C sH sN MeOH C 5H 5N MeOH CHCI,

40 16 16 14 51 51 40 40 16 16 51 215 216 217 218 219, 220 219 16 16 15 16 16 218 221 19Oa 19Oa 190a

EtOH EtOH C5HsN C sH sN MeOH C sH sN C sH 5N C 5H 5N MeOH HzO HzO H?O HzO d

Claimed t o bepyranose by

161

GLYCOSYLAMINES

TABLE XX L-Ribosylamine Derivative Compound

L-Ribouylamine 2-deoxy-N-phenyl-

Compound

L-Sorbosy laniine N -p-ethoxyphenyl-

217 40

169.b170.5 172-173

p$$:!t. 160

[u]., degrees

Rofalion soloenl

Reference

- 191

C 6H 6PvT

17

compound

N ,N’-Di-D-xylosylurea D-Xylosylamine triacetate N-o-aminophenyl-, triaeetat e N-n-butylN-p-carbethoxyphenylN-o-carbomethoxyphenylN-o-carboxyphenylN -p-carboxyphenylN- (4-chloro-2-nitrophenyl) triacetate, I triacetate, I1 N-(4,5-dimethyl-2-nitrophenyl) triacetate N-o-(1-ethoxyethy1idenearnino)phenyl-, triacetate N-l-hexylN-(4-methyl-2-nitrophenyl) triacetate, I triacetate, I1 N-o-nitrophenyltriacetate N-p-nitrophenyl-a-

N -p-nitrophenyl-p-

Mclfing poinf, ‘C.

230-235 (dec.) 130 101-102 179 81-82 116-117 170 167 172 180 162 117 212-213 (dec.) 168-169 139 87 ca. 170 183 13&132 172-176 149 192

101; 109

dcgrecs

-20 -18.1 -15.9 -22.4 -50

- 28 - 22

Rotation solucnt

H2 0

0) H20 CHCL CHCI,

-11 +61.6 -59.6 -90.1 +96.7,

EtOH EtOH EtOH EtOH C5HsN CHCl, CHCls

-62.1

CHCli

-87.3 f7.9

CHCla CHCla

-109.5 +292.5 +260 4 $280.4 f260 --t +285 -95.6

AcOMe C5H5N C,H,N-H,O C~H~S-ACOH CsHjN

References

113 112 222

6.4 116 117 115 137 137 115 116 116 116 34 34 116 117 34 32 34 34

17 35 35 35 3.5

N -phenyl-

triacetate 2-methyl ether

3-methyl ether 2,3-dimethyl ether

2,4-dimethyl ether

148 140-141 142 142-144 143-144 151 125-126 128 123-124 138 136 146 145

-94.3 -79.6

2,3,4-trimethyI ether

2-deoxy -N -phenyl-

-70.8 -24

-87

+214 +23.7 +77

AcOEt

-+

-48 -21.9

+185 190 +118 + +75

+

-40

-84

-97 -37.4

C~H~X-ACOH MeOH CsHsN MeOH MeOH CHCli AcOEt AcOEt

4

+25

120-122 122-123 123 128 174 155-157 170 121 126 98-100 100-101 102 104 97-98 137

+

-90 -84.1

-7 3,4-dimethyl ether

3

+87 -82 +

AcOEt .ScOEt AcOEt-AcOH

AcOEt AcOEt -AcOH Dioxane

-+

+47

EtOH

+

+32.8 -20

MeOH H2 0

-+

35 39 20 51 40 16 51 223, 224, 225 224a 226 223 227 228 229 229 230 226 227 231 148(a) 232 232 233 234 235 223 225 236 122 224a 237

L3

F

r!

n 0 7l 4

F

>

5

3

n

L

3

w

TABLEXXII (Continued) Compound

Melting point, "C.

[aID, degrees

Rotation solvent

I

References

D-Xylosylamine (Continued) N-p-sulfacetamidophenylN-p-sulf amylphenylN-p-sulfophenylN-p-tolyl-

trihenzoate N-D-Xylosylpiperidine, hydrochloride N-D-Xylosyltrimethylammoniumbromide, 2,3,4-triacetate

139 121 168-169 157 >300 124-125 122-124 124-125 180-181 125 181

-58

-62.3 -48 -41.5 -44-t -12 -76.6 + + 21.2 -59.0 +48.3 +9.6 -20.8

238 1 25 124 125 125 20

126 16 16 55 77 153

GLYCOSYLAMINES

165

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1GG

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v.

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THE AMADORI REARRANGEMENT

BY JOHN E . HODGE Northern Utilization Research Branch. Agricultural Research Service. U . S . Department of Agriculture. Peoria. Illinois

CONTENTS I . Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . I1. Nomenclature. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . I11 . Scope of the Reaction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

IV . ExperimentnlMethods

.............................................

RIechanism of the Reaction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1. F:xperinierital Basis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 . Proposed Mechanisms. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3 . Comparison with the Lobry de Bruyn-Alberda van Ekenstein Transformation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . V I . Physical Properties of Amadori Rearrangement Products . . . . . . . . . . . . . . 1. Color and Taste . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 . Crystallization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3 . Melting . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. 0ptic:ilRotntiorl . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5. Light Absorption . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VII . Chemical Properties of Amadori Rearrangement Products . . . . . . . . . . . . . . . 1. Enolizatio~iand Oxidation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 . Hydrogenation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3 . Dehydration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . a . 2-Furaldehyde Formation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b . Reductone Formation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4. Fission . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5 . Derivatives . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . a . Salts . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b . Esters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c . Ethers . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . d . Oximes and Hydrazones . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . e . Osazones . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11.

169 172 173 175 178 178 181

184 185 185 185 185 186 186 187 187 191 192 193 194 104 195 195 195 196 196 197 ................................................. 198 Amadori Rearrangement Products . . . . . . . . . . . . . . . 199 I S . Retrospect and the Future . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 201 ......................... 202 S . Tables of Compounds . . . . . . . . . . . . . . . . . . . .

I . INTRODUCTION The reaction to be discusked here is the isomerieation of an aldosylamine to a 1-amino-1-deoxy-2-ketose. This “rearrangement” was named after 169

170

JOHN E. HODGE

Amadori’ by Kuhn and Weygand,2for Amadori was the first to demonstrate that condensation of ,D-glucose with an aromatic amine (p-~henetidine,~. * p-a~iisidine,~ or p-toluidiid) would yield, according to experimental condit,ions, two sttruct,urallydifferent isomers which are not members of an a,/3 anomeric pair. Amadori did not realize that an isomerization (“rearrangement”) from an aldose t.o a ketose configuration had occurred. However, he did discern (by change of optical rotation in acid solution) that one isomer is much more labile t h m t8he other toward hydrolysis and more suscept,ible to decomposition on standing in the solid state in air. He recognized correctly that the labile isomer is the N-substit,ut,ed glucosylamine. but he niistakeiily t,hought t,hat the stable isomer was a compound of the Schiff-base t,ype4-5 , The fact that his supposed Schiff base resists acid hydrolysis was evident.ly overlooked. Kuhn and Dansi7 determined that Amadori’s stable isojner is riot a Schiff base but t,he product, of a molecwlar rearrangeinent. They also proved t.hat t,he labile isomer is in fact, the N-substituted glucosylamine. However, they assumed incorrect,ly (on the basis of C-methyl analyses) t>hatt’hesugar moiety of the &able isomer contained a branched-carbon skelet’on. I n 1937, the true st,ruct>ureof the rearranged product was reported by Kuhn and Weygand.2 hmadori’s stable isomers were shown to be unbranched N-subst,it,ut,ed1-amino-1-deoxy-Zketoses. Therefore, in a strict sense, the reaction named by Kuhn and Weygand the Amadori renrranyemerit is the complete conversion of a N-substituted aldosylamine to a N-subst’ituted 1-amino-1-deoxy-2-ketose. On this basis, simple enolization of an aldosylamine or ketosylamine to t,he int,ermediat.e 1,2-enolic form common t.0 both should not be construed as the Amadori rearrangement. Also, the recently demonstrated conversion of a ketosylamine to a 2-amino2-deoxyaldoseg does not come under the definition in spite of an obvious similarity to the Amadori rearrangement. (1) The author is indebted t o Dr. Alfred0 Dansi, Milan, Italy, for the following biography. Mario Amadori (1886-1941) studied chemistry a t the University of Padua, Italy. After graduation in 1908, he remained a t the University until 1926 as an assista n t of Professor Bruni in the Institute for General Chemistry. In 1926, he was appointed professor in the Pharmaceutical Institute of the University of Modena and was elected Dean of the faculty in 1940. He died suddenly on August 1, 1941. His work was devoted mostly to general and inorganic chemistry. He was the author of textbooks on general, inorganic, and pharmaceutical chemistry, and published a number of papers throughout the period 1909-1935. (2) R . Kuhn arid F. Weygand, B e r . , 70, 769 (1937). (3) M. Amadori, Atti reule nccnd. n a d . Liricei, [6],2, 337 (1925). (4) M. Amadori, Alti renle accnd. nu.zZ. Lincei, [6]9, 68 (1929). (5) M. Amadori, Atti renle nccad. nazl. Lincei, [6] 9, 226 (1929). (6) M. Amadori, Atti ieale accad. nazl. Lincei, [6] 13, 72 (1931). (7) R. Kuhn and A. Dansi, Ber.., 69, 1745 (1936): (8) K. Heyns and K. Meinecke, Chem. Ber.., 86, 1453 (1953).

AMADORI REARRANGEMENT

171

Although Amadori published his findings in the years 1026-1931, it is probath that the first crystalline product of the rearrangement hearing his name had been prepared hy 1897. In that year, Claw and RBe9 ohtained a German patent on the preparation of (.ondensation products of glucose arid galactose, respectively, with p-phenetidine. Since the g1uc:ose-p-phenetidiHe compound melted a t 160" (on rapid heating) and easily reduced silver salts in aqueous solution, this compound doubtless was the rearrangement product, m.p. 155", first investigated by Amadori13rather than N-p-phenetylD-glucosylamine, which melts3 at 118". This example serves as a warning that properties of all the sugar-amine condensation products reported in the older literature need careful scrutiny in order that one may determine whether the products represent the aldosylamine or the isomeric l-aminol-deoxy-2-ketose derivatives. The toxicity of the condensation product obtained from D-glucose and p-phenetidine was a matter of dispute during the period 1897-1913.9. l o , 11, 12 The differences may have arisen through investigation of the easily hydrolyzed aldose derivative in some experiments and of the ketose derivative, which is stable toward mild acid hydrolysis, in others. The Amadori rearrangement was at first thought to apply exclusively to N-substituted derivatives of ~-glucosylamine.~~ Also, until recently, it was believed to apply only to glycosyl derivatives of primary aromatic amines.l4. l5 The scope of the reaction has been extended in recent years, so that the Amadori rearrangement is now recognized to be a general reaction that might occur with virtually all classes of aldoses and amines. The usefulness of the AmadGi rearrangement in chemical synthesis was demonstrated by WeygandlGwho showed how D-arabinose, instead of the more expensive D-ribose, could be used in the preparation of riboflavin. Condensation of D-arabinose with 3,4-dimethylaniline in the presence of an acid catalyst gave the Amadori rearrangement product, l-deoxy-l(3,4-dimethylanilino)-~-erythro-pentulose (not isolated) I which could be catalytically hydrogenated in alkaline solution to produce the desired Also, Amadori reintermediate, l-deoxy-l-(3,4-dimethylanilino)-~-rib~tol. (9) W. H. C l a w and A. R6e, German P a t 97,736 (Sept. 25, 1897); Chem.-Ztg., 22, 545 (1898). (10) S. Mostowski, Bull. intern. acad. sci. Cracovie, 641 (1909); Chem. Abstracts, 4, 2321 (1910). (11) J. C. Irvine and R. Gilmour, J . Chem. S O C .96, , 1545 (1909). (12) J. A. Hewitt, Biochem. J. (London), 7 , 207 (1913). (13) R. Kuhn and L. Birkofer, Ber., 71, 621 (1938). (14) W. W. Pigman and R. M. Goepp, Jr., "Chemistry of the Carbohydrates," Academic Press Inc., New York, N. Y., 1948, p. 386. (15) L. I. Smith and R . H. Anderson, J . Ory. Chem., 16, 963 (1951). (16) F. Weygand, Ber., 73, 1259 (1940); German P a t . 727,402 (Oct. 1, 1942); IJ. S. P a t . 2,354,846 (Aug. 1, 1944).

172

JOHN E. HODGF,

arrangement products have been used to advantage in the preparation of o s a z o n ~ s ,quiiiosaliiics,lR ~~ osone-like compounds,18*l o and in a chemical synthesis of folic Recently, the significaiice of the Amadori rearrangement i l l two broad fields of current research has heen adduced. CrottJsrhalk2l 22 published evidence that the carbohydrate-amine linkage in the physiologically important mucoproteins is that of the 1-amino-1-deoxy-%ketoses. HodgeZ3 has indicated the importance of the Amadori rearrangement in the group of chemical reactions (the Maillard reaction) which is considered t o represent, a type of browning in foods and feeds. These concepts explain, for example, tJhedifficulty of removing saccharides from mucoproteins by acid hydrolysis and the loss of sugar that accompanies the hydrolytic process. I n foods, and in all living cells where sugars and amines coexist, thcrc are carbonyl-amino additions and condensations which provide the chalice for occurrence of the Amadori rearrangement. Once the rearrangement, has occurred, the sugar-amino linkage is no longer readily susceptible to hydrolysis; however, the ketose sugar moiety is more readily decomposed by dehydration and fission tJhanis the original a l d o ~ e Such . ~ ~ L'amine-catalyzed" sugar decompositions probably have important consequences for biological syntheses and transformations.

11. NOMENCLATURE Emil Fischer??was the first t o synthesize and name the aminodeoxyketose shown below as I. Because of its isomeric relationship to D-glucosamine

CILNH~ I

c=o I

HOCH

I I HCOH I

HCOH CHZOH I

CHO

I

HCN HP

I

HCOH I HCOH

I

CHzOH I1

CHzNH-CsHa-CHa

I I HOCH I

c=o

HCOH

I

HCOH

I

CHaOH 111

(17) F. Weygand, Ber., 73, 1284 (1940). (18) F. Weygand and Annemarie Bergmann, Chem. Ber., 80, 255 (1947). (19) Y. Inoue, I<. Onodera and 1. Karasawa, J . A g r . Chem Soc. J a p a n , 26, 75 (1951). (20) F. Weygand, A. Wacker and V. Schmied-Kowarzik, Chem Ber., 82,25 (1949). (21) A. Gottschalk, Nature, 167, 845 (1951). (22) A. Gottschallr, Biochem. J . (London), 62, 455 (1952). (23) J. E. Hodge, J . A g r . Food Chem., 1, 928 (1953). (24) E. Fischer, Ber., 19, 1920 (1886).

AMADORI REARRANGEMENT

173

(11), he called it isoglucosamine. (The formulas are written arbitrarily in the open-chain form.) Kuhn and Weygand2 began the custom of naming the Amadori rearrangement products as derivatives of I; for example, 111 is N-p-tolyl-D-isoglucosamiiie in their terminology. Glucosamine and isoglucosamiiie are ambiguous names, hence the American-British Committee on Carbohydrate Nomenclature has tentatively recommended the name for II.25 1-amino-] -deoxy-D-fructose for I, and 2-amino-2-deoxy-~-g~ucose It follows that an Amadori rearrangement product should be named as a 1-amino-1-deoxy-2-ketose. This might be done in two different ways. For example, I11 might be called (a) N-p-tolyl-1-amino-1-deoxy-D-fructose or ( b ) 1-deoxy-1-p-toluidino-D-fructose. The system used for ( a ) presents difficulties in naming the N-substituted l-amino-1-deoxy-2-ketoses derived from heterocyclic amines and also those from amino acids. For example, N ,N-pentamethylene-1-amino-1-deoxy-D-fructoseand N ,N-3-oxapentamethylene-1-amino-1-deoxy-D-fructose would be the names required for the piperidino and morpholino derivatives, respectively, under this system. Because of its simplicity and the avoidance of unwieldy radical names, the system used for (b) is recommended for Amadori rearrangement products and is used throughout this Chapter. The radical names anilino, anisidino, benzidino, cumidino, dimethylamino, guanidino, morpholino, piperidino, and toluidino are approved according t o the nomenclature rules of the International Union of Pure and Applied Chemistry126but analogous names for amino acid radicals (for example, glycino, alanino, leucino) are not specifically approved. The American-British Committee on Carbohydrate Nomenclature also recommends an alphabetical order of the prefixes, including the deoxy prefix. According to the Rules, complete names for two hypothetical Amadori rearrangement products would be 1-deoxy-1-piperidino-a-D-fructofuranose ‘and l-benzylamino-l-deoxy-/3-D-tagatopyranose. 111. SCOPEOF THE REACTION Authentic crystalline products of the Amadori rearrangement have so far been obtained only from D-glucose, D-mannose, and li-O-trityl-~-xyloseas the sugar components. Occurrence of the rearrangement (or a t least 1,2enolization of the N-substituted glycosylamine) was demonstrated indirectly, however, by isolation and characterization of crystalline epimeric hydrogenation products derived from D- and L-arabinose and also from D-xylose. For example, 1-amino-1-deoxyribitols were obtained from the arabinoses, and a 1-amino-1-deoxy-D-lyxitol was given by D-xylose after conditions known to produce the Amadori rearrangement were applied.16 (25) Chem. Eng. News, 31, 1776 (1953); J . Chem. Soe., 5108 (1952). (26) J. Chem. Soc., 5064 (1952).

174

JOHN E. HODGE

Although hydrogenatioii of the reaction mixtures derived from D-galactose and from L-rhamnose gave crystalline aniinohexitols, they were identified as derivatives of galactitol and rhamnitol, respectively, instead of those of the epimeric compounds. Therefore, proof for occurrence of the rearrangement was lacking in these two cases. However, from the initial crystallization of the N-substituted galactosyl- and rhamnosyl-amines after a short heating period, and their subsequent disappearance from the darkened reaction mixture on longer heating, it appears that rearrangement or 1,2-enolization of the glycosylamine did occur.16 From the experiments of Sttrain and Spoehr27 on the rearrangement of glyceraldehyde to pyruvaldehyde in the presence of amines, and also from the more recent work of Smith and Anderson15*28 and of Anderson29on the rearrangement of 3-C-phenylglyceraldehyde,it appears that aldotrioses also might undergo the Amadori rearrangement. Glyceraldehyde (or 3-Cphenylglyceraldehyde), when treated with m-nitrobenzhydrazide in dilute acetic acid solution, formed the osazone of pyruvaldehyde (or 3-C-phenylpyruvaldehyde) only if a primary aromatic amine or an aliphatic diamine was added. The addition of a strongly basic, primary amine, instead, did not produce the m-nitrobenzoylosazone in solutions having a high content of water; but, on the other hand, in dioxane-acetic acid-water solutions containing less than 10 % of water, the osazone was produced in the presence of strongly basic a m i n e ~ ?Strain ~ and Spoehr, and also Smith and Anderson, interpreted the formation of the m-nitrobenzoylosazone as an indication that free pyruvaldehyde (or 3-C-phenylpyruvaldehyde) was present in the reaction mixtures; neither was actually isolated. Because Weygand has shown that osazone formation proceeds directly from Amadori rearrangement products in high yields,17 it is not necessary to assume that a nonnitrogenous osone exists in the reaction mixtures. The hydrazono radical could have replaced the amino radical of the pyruvaldehyde derivative directly, to form the osazone by the mechanism proposed by Weygand (see Section VII, 5e). Since it has been shown that acidic conditions are requisite for osazone formation from Amadori rearrangement products,17 the osazone would not be formed in aqueous solutions of strongly basic amines under Anderson's condition~.~9 An Amadori rearrangement product apparently was formed in the reaction of 3-C-phenylglyceraldehydewith N-methylaniline in dry ethanolacetic acid solution as conducted by Smith and Anderson.2s A white, crystalline l-deoxy-l-(N-methylanilino)-3-C-phenylketose,C6H,(CH3)N. C&02+C& , was isolated. Whether the carbonyl group (the. compound (27) H. H. Strain and H. A. Spoehr, J . B i d . Chem., 89, 527 (1930). (28) L. I. Smith and R. H. Anderson, J. Org. Chem., 16, 972 (1951). (29) R. H. Anderson, J . Org. Chem., 19, 1238 (1954).

AMADORI RBARRANGBMENT

175

forms a semicarbazone) is at C2 or C3 was not established; nevertheless, the compound reduces Fehling solution and Methylene Blue in alkaline solution a t room temperature in the manner of an Amadori rearrangement product. As for the amine moiety, the rearrangement has been demonstrated (although not unequivocally in all cases) for mono-N-substituted glycosylamines derived from aromatic amines and aralkyl- and alkyl-amines; and for di-N-substituted glycosylamines containing dialkyl, alkyl-aryl, aralkyl, isocyclic, and heterocyclic alkyl radicals. The rearrangement has been reported for N-glucosyl derivatives of amino acids,22'3 * c f . , 29 but not as yet for any member of the group of nucleosides. A list of well-substantiated crystalline rearrangement products is given in Table I . Those reported but not well substantiated, or not isolated in the crystalline state, are given in Table 11. The rearrangement of N-substituted ketosylamines to 2-amino-2-deoxyaldoses is deserving of study. Already, Heyns and Meineckes have reported the isomerization of unsubstituted D-fructosylamme to D-glucosamine (2-amino-2-deoxy-~-glucose).Also, ketols such as acetoin, benzoin, D-fructose, and L-sorbose have been condensed with amines, whereupon a carbonyl group was apparently generated in place of an a-hydroxyl g r o ~ p . 3' 0~~31~ IV. EXPERIMENTAL METHODS Amadori's original method for conducting the rearrangement consisted in fusing eyuiniolar proportions of the reactants (D-glucose and p-pheneti* p-anisidine,S or p-toluidinec) in the dry state and holding the solidified melt a t 70-80" for two hours. After the residual, brown mass had been dissolved in hot ethanol, the ketose derivative crystallized on cooling. One recrystallization from alcohol gave the rearrangement product in nearly pure form. Although Amadori reported a yield of about 70% of theory for the p-phenetidino compound, other workers have been unable to duplicate that yield by his method. For example, only a 13% yield of 1deoxy- 1-p-toluidino-D-fructose was obtained by Kuhn and D a n ~ iFurther.~ more, a series of experiments performed in Amadori's way by Weygand,1,6 using various aromatic amines'with D-glucose, gave yields of only 15-30 %. Amadori also isolated the N-substituted glucosylamine before heating it in the dry state, but no advantage was gained t h e r e b ~ For . ~ example, Inoue and Onodera32 held pure N-p-tolyl-D-glucosylamine just above the melting point a t 120' and obtained only a 17% yield of the Amadori rearrangement product. However, when they added 1 millimole of aretic (30) J. F.Carson, J. A m . Cheni. SOC., 76,4337 (1953);77, 1881 (1955). (31)J. G.Erickson, J . Am. Chem. SOC.,76, 2784 (1953);77, 2839 (1955). (32) Y. Inoue and K. Onodera, J. Agr. Chem. SOC.Japan, 22, 120 (1948).

176

JOHN E. HODGE

acid per mole of the glucosylamine, the yield was doubled; and, with the addition of 2 millimoles of acetic acid, the yield was nearly tripled (47 %). Hence, Amadori's method of heating in the dry way may be used preparatively if an acid catalyst is added in appropriate amounts. When sugar and amine are heated together in the dry way, the concentration of acid catalyst is a critical factor. Recent experiments by Inoue and O n ~ d e r agave ~ ~ 1-deoxy-1-o-toluidino-D-fructose in 23 % yield after two recrystallizations, when D-glucose (1 mole) and o-toluidine (1.5 moles) were heated for 30 minutes at 100" with 0.04 mole of glacial acetic acid. However, in parallel experiments, when less (0.004 mole) or more (0.08 mole) acetic acid was added, the rearranged product could not be isolated. In another was isolated in 18% yield, set of experiments, 1-anilino-1-deoxy-D-fructose with 0.06 mole of glacial acetic acid per mole of D-glucose present in the reaction mixture. But, with smaller concentrations of acetic acid, none of the fructose derivative could be isolated. A distinct improvement in the preparative method was made by Weygand,16 who was the first to demonstrate that the rearrangement is catalyzed by acids. Besides resulting in a faster and more reliable reaction, purer products in much improved yields (50-60%) were obtained. A mixture of 1.0 mole of aldose, 1.1 to 1.4 moles of aromatic amine, 2.5 to 3.0 moles of water, and 0.002 to 0.02 moles of an acid catalyst was heated for 10 to 30 minutes on a boiling-water bath. The darkened liquid was diluted with hot alcohol, affording the crystalline ketose derivative in nearly pure state. Mineral or organic acids, or acid-reacting salts, were employed as the catalyst. The foregoing experimental procedures were successful only with primary aromatic amines. Weygand added water to his reaction mixtures, but such practice cannot be recommended with strongly basic amines because N-glycosy1 derivatives of the latter are easily hydrolyzed.29* 35 Hydrolysis in strongly basic, aqueous media results in alkaline degradation of the sugar.36 Occasionally, the Amadori rearrangement has been accomplished by heating the sugar and amine components, or the N-substituted glycosylamine, in alcohol without the addition of an acid c a t a l y ~ t .9~, ,l 3 However, acidic catalysts are probably generated in these instances by the accompanying decomposition reactions. Heating in alcohol without an added catalyst does not provide a reliable preparative method. The yields are low, the products 34v

(33) K. Onodera, letter t o the author (1953). (34) Eleanor Mitts and It. M. Hison, J . Am. Cl~enz.Soc., 66, 483 (1944). (35) M. L. Wolfrom, R . D. Schuetz and L. F. Cnvalieri, J . Avi. Chem. SOC.,71, 3518 (1949). (36) L. F. Cavalieri and M. L. Wolfrom, J . Am, Che~ia.SOC., 68, 2022 (1946).

AMADORI REARRANGEMENT

177

are impure, and the desired ketose derivative is not alwayb ohtainedsoinetimes the N-substituted aldosylamine is recovered.’G The rearrangement in alcoholic solution may be consistently acromplished if a compound containing an artivated methylene group (=C-CH2-C=) is added in relatively large proportion together with a c:ttalytic proportion of a secondary a n i i ~ i e .38~ ~This . discovery made possihle the isolation of Amadori rearrangement products from N-glycosyl derivatives of non-aromatic amines, whereas previous attempts,l3,34 except for the special rase of dibenzylamine plus D-glucose,la 37 had failed. Compounds containing a(*tivated methylene groups which were employed successfully were ethyl malonate, 2,4-pentanedione (acetylacetone), phenylacetone, diphenylmethane, and malonic acid. Ethyl acetoacetate apparently gave a different reaction that prevented isolation of the desired ketose derivative. This method of IIodge and Rist allowed the isolation of N-substituted l-aminoI-deoxyfructoses from all classes of amines, in yields which ranged from 35 to 89% of theory.38 A critical factor limiting the yield of ketose derivative is the decomposition which takes place concurrently with the rearrangement, Decomposition is shown by the formation of a deep-red or red-brown color. Too much acid, too long a heating period, and too high a temperature, all result in destruction of the sugar moiety. The nature of this decomposition is discussed in Sections V and VII. The highest yield of an Amadori rearrangement product yet reported (89 %) was obtained with D-glucose plus dibenzylamine by the method of Hodge and R i s q w h e r e i n none of the deep-red color was produced. Recent work by Hodge and Fisher39has shown that the Amadori rearrangement of D-glucose and some secondary amines of intermediate basicity (morpholine, di-n-butylamine, dibenzylamine and N-methylaniline) can be accomplished satisfactorily with glacial acetic acid as the c a t d j s t . One mole of D-glucose and 1.3 nioles of the secondary amine were stirred with absolute ethanol a t 70-80”, and then the organic acid (1.0-1.3 moles) was dropped into the reaction mixture. Color formation was carefully watched and the reaction was stopped a t the deep-amber to bright orange stage. Yields of the crystalliiie ketose derivatives were 50-60 %. Three attempts to isolate the corresponding piperidino derivative in this way were fruitless. However, when part of the acetic acid was replaced by 3-mercaptopropionic acid (0.2 mole; total acid, 1.O mole) t o delay deep-color formation, l-deoxy-l-piperidino-D-fructose was isolated in 22 % yield. (37) J. E. Hodge and C. E. Rist, J. A m . Chem. Soc., 74, 1494 (1952). (38) J. E. Hodge and C. E. Rist, J . Am. Chem. Soc , 76, 316 (1953) (39) J. E. Ilodge and B. E. Fisher, forthcoming publication.

178

JOHN E. HODGE

V. ME:CIIANISM OF

REACTION No estensive study of the reaction mechanism of the r2inaclori rearrangement has been made. As the isonierization is usually cwnducted, about half of the amount of aldose or N-substituted aldosylamirie originally present is not accounted for at the end of the reaction. The intermediates and byproducts of the decomposition have not been identified or extensively investigated. consequently, any conclusions now drawn on the ciourse and meehanisni of the reaction are speculative. In this Sertion, the limited number of positive and negative experimental results which bear upon the reaction mechanism will be presented first, and then their implications for the reaction merhanism will be discussed. THE

1. Experimental Basis

Positive experimental results indicate, but do not prove, that, starting from the sugar and amine, sugar-amine condensation is the first step in the Amadori rearrangement. When an acidic catalyst is not presentl3'l a , 40 or if the reaction time is shortened in the presence of an acidic ratalyst,I6 3 4 , 3* the N-substituted aldosylamine crystallizes from the reaction mixture in 85-95 % yields. Yet, further heating has, in both instances, produced the Amadori rearrangement product. In addition, the rearrangement has been demonstrated in dry media, starting with the preformed N-substituted glycosylamine.6' 38 Although there is no positive evidence in support of it, a plausible alternative to the above reaction sequence is eiiolization of the aldose before condensation with the amine. In such a sequence, the N-substituted glycosylamine would not be a precursor of the aminodeoxyketose. This postulate merits investigation because, in spite of attempts to isolate N , Ndibenzyl-D-glucosylamine, this compound could not be found in the reaction product of D-glucose with dibenzylamiiie; only D-glucose or the Amadori rearrangement product was isolated.37 It is well established that addition of an active proton donor (mineral acid, organic acid, acid-reacting salt, or compound containing an activated methylene group) results in a faster transformation of the aldose to the crystalline ketose derivative (see Section IV). Although the rearrangement is catalyzed by acids, the reaction medium i s basic. For example, in the procedure recommended by Weygand,l6 only a few hundredths or thousandths of a mole of acid is used, and this minute amount exists in the presence of 0.1 t o 0.4 mole of amine in excess of that required for complete condensation with the sugar. In the procedure recommended by Hodge and R i ~ t rearrangement ,~~ occurs in absolute alcohol upon adding a relatively (40) F. Weygand, Ber., 73. 1663 (1939).

179

AMADORI REARRANGEMENT

large proportion of an active methylene compound as the proton donor; nevertheless, the medium remains basic because of the large proportion of basic N-substituted glycosylamine present. N-Glycosyl derivatives of the more acidic nitroanilines are apparently not basic enough to accept protons from the acid catalyst, for rearrangement does not occur under the usual preparative condition^.^^. 42 Nor does rearrangement occur for the weakly basic N-acetyl-~-glucosylamine.~~ Weygand added piperidine acetate and other amine salts, instead of an acid, in attempts t o catalyze the rearrangement of pure N-p-tolyl-D-glucosylamine in both alcoholic and aqueous solutions. Little, if any, rearrangement occurred, presumably because the strongly basic piperidine acetate, rather than the weakly basic, aromatic glycosylamine, accepted the catalyzing protons.16Apparently, a necessary condition for occurrence of the Amadori rearrangement is that the N-substituted glycosylamine must be sufficiently basic to accept a proton from the surrounding medium. When a compound containing an activated methylene group is the proton-donating catalyst, the presence of a secondary amine is required for easy isolation of the desired ketose derivative in crystalline form.38The conditions resemble those requisite for a Knoevenagel reaction.43’ 44 A condensation of the Knoevenagel type evidently does not occur, as the dehydrated product would be too stable for practical reversibility. However, the addition compound which is an intermediate in the Knoevenagel reaction,44 such as V, could be formed from IV. Splitting of V to yield the ketose deR-N H -CHz

R-NH-CHz

I

I c=o

HOG-CH(C0R‘)z

I

HOCH

I HCOH I HCOH I

CHSOH IV

+CHz(COR’h

I I HCOH I HCOH I

HOCH

CHiOH V

rivative in a cyclic form would complete the rearrangement. The only evidence for the existence of V is the initial precipitation (upon adding ether or acetone i n the method of I-Todge and Rist3*)of a viscous gum which does not crystallize upon seeding wit8hthe expectredN-substitded l-aminol-deosy-2-ket80se. However, after long standing, addition of acid, or re(11) R.Kuhn and It. StrGbele, B e r . , 70, 773 (1937). (42) F. Wepgaritt, W. Pe?kovl- and 1’. Iiuhner, Chem. B e r . , 84, 594 (1951). (43) 15. Knoevenngcl, Be,., 31, 730, 2585, 2596 (1898). (-44)A. C. Cope, ./. A m . Chela. Soc., 69, 2327 (1937).

180

JOHN

E.

HODGE

heating of the gum in alcohol, the expected crystalline ketose derivative is finally obtained. The compound containing the active methylene group probably acts in a specific way to aid isolation of the crystalline rearrange; ment product, but further investigation is necessary for determination of its actual role. Another important experimental observation is the decomposition with color formation (browning) that occurs along with the rearrangement. Yellow, orange, or deep-red byproducts have been found in every case so far reported. The results of WeygandlG and of Rosen, Johnson and Pigman45 show that addit>ionof increasing amounts of hydrochloric acid to N-aryl-Dglucosylamines, in quantities up to one mole per mole of the glucosylamine, greatly increases the rate of color formation. When one equivalent of acid has been added, the browning is a t a maximum and no ketose derivative can be isolated. When more than one equivalent of acid is added, the reaction is apparently of a different type. There is no longer an induction period preceding the browning, and less color is Strong acid is not required for decomposition to occur; the browning discoloration also 4 6 , 4 7 , 48 takes place quite rapidly in the presence of glacial acetic As to the nature of the decomposition, both dehydration and fission of the 4 7 , 49 but exact knowledge of the rethe sugar are known to occur 2 2 , actions is lacking. Two a1ternat.e pathways for the production of tarry decomposition products from glycosylamines were considered by Bayne and H 01m s. ~~ According to their speculations, tar could be formed either directly from the Aniadori rearrangement product or from “unsaturated intermediates” which could be produced from the glycosylamine and would be in equilibrium with the ketose derivat,ive. GottschalkZ2has proposed t,hat it is the 1,2-enol, which may be formed from the glycosylamine and also from the ketose, that leads to the formation of humin,like substances -presumably t,hrough 5-hydroxymethyl-2-furaldehyde or a Schiff base thereof. Decomposition of the 1,2-enol (which should be considered to be the ammono analog of the 1,2-enediol form of unsubstituted sugars), would channel the course of the reaction through that form without allowing the production of much ketose derivative. The following scheme shows the 23v

38s

(45) L. Rosen, K. C. Johnson and W. W. Pigman, J. Am. Cheni. SOC.,76, 3460 (1953). (46) C. N. Cameron, J . Ant. Chent. SOC.,48, 2233, 2737 (1926). (47) C. N. Cameron, J . A m . Cheni. SOC.,49, 1759 (1927). (48) W. W. Pigman, €3. A . Cleveland, D. H. Couch and J. H. Cleveland, J . Am. Chem. Soc., 73, 1976 (1951). (49) A. Gottschalk and S.h1. Partridge, N u t w e , 166, 684 (1950). (50) S. Bnyne and W. H. Holins, J. Chela. SOC., 3247 (1952).

AMADORI REARRANGEMENT

181

chief stages of the rearrangement for aldohexoses and the known decomposition reactions (see Section VII) which are believed to arise from the 1,2eno1.22, 2 3 , 49, 60 Aldohesose+ arnine

11-

Ha0

A'-Substituted glycosylan~~ne (VI)

5-Hydroxymeth yl2-furaldehyde

amino-reductones

I Highly colored melanoidins 1 2. Proposed Mechanisms

From the experiment,al evidence presented in the preceding Section on the proton acceptor-proton donor relationship of glycosylamine and acid catalyst, the Amadori rearrangement must be classed as another of the many examples of acid-base ~ a t a l y s i sPresumably, .~~ the reaction involves merely the acceptance of a proton by the glycosylamine base, prototropic shifts, and the subsequent discharge of a proton from the cation originally formed. Thus, the rearrangement is pictured as the ammono analog of aldose enolization when the latter is catalyzed by acids. For example, Petuely62 gave the following representation of sugar enolieation by acid catalysis, for 6 3 * 64 which there is now definite experimental (51) (52) (53) (54)

W. F. Luder and S. Zuffanti, Chem. Revs., 34, 345 (1944). F. Petuely, Monatsh., 84, 298 (1953). M. L. Wolfrom and W. L. Shilling, J. A m . Chem. Soc., 73, 3557 (1951). E. Berner and S. Sandlie, Chemistry & Industry, 1221 (1952).

182

JOHN E. HODGE

HC=O

+ H+ ,

I

HC=O+H

HCOH

- H+

I

II

CHZOH

CHzOH

- I{+

I - 1 C=O+H I

C=O

I

The ammono analog of the above reaction mechanism would be as follows. HC-N-R HC=N+H-R

I

+ H+

-

HCOH

- H1 +

I

IIC-NF-R

II COH I

H+

-

+ H+

HCOH

P

I

CHZNH-R

- +I H+

P

- H+

- H+

C=O+H

' +H+

I

CHzNH-R

' I

c=o I

This is the mechanism that was proposed originally by Kuhn and Weygand.2*l6 The nitrogen of the N-substituted glycosylamine (VI, R' = H or a radical) accepts a proton from the acid catalyst, to form the ammonium ion (VII) which presumably is in equilibrium with the cation of the Schiff base (VIII). Isbe11b6then explains the shift of the double bond on the basis e R-NH-R'

fi!!l I

El

HC=NRR'

I

HCOH

HOCH

-

+ HIQ

HCOH

-

HCOH

HCOH

HFO

HFOH

I

HCO

I

I

R-N-R'

-

CH2 HOCH I

I HCOH I HCOH I

CHiOH X

~

I

HOYH HCOH

I I

HCOH

CHzOH VIII

~~~~

HCOH I HCO

CHzOH IX R-N-R'

I

CHI

I

-

- H@

I

R-N-R' I CH1

I

I c=o

-

I

CH2OH VII

CHzOH VI

7

I HOCH

L

HCNRR' II COH

-

I

CHiOH XI

(55) H. S. Isbell, Ann. Rev. Bioehem., 12, 206 (1943).

I I

HOC -

H°FH HCOH I I

HCOH HzCOXI1

183

AMADORI XEAHHANGEMENT

of flow of electrons toward the positive nitrogen atom, which makes C1 transiently positive. The secondary flow of electrons from C2 to C1 then weakens the C-H bond on C2, causing the expulsion of that proton to give the enol form of the ketose derivative (IX). The tautomeric shift to the keto form (X) would presumably be driven by the strong tendency of X to go over into one of the stable ring forms (XI or XII). The foregoing mechanism is also applicable to N-glycosyl derivatives of secondary amines (R’ = R or another radical, or R’ N . R’ = a heterocyclic, secondary amine). I n the mechanism proposed by Gottschalk,22 the catalyzing proton is presumed to add to the ring oxygen of VI to give XI11 (R’ = H or a radical). This mode of proton addition has also been suggested by Isbell and Frush6‘j and by Petuely67 to explain the mutarotation of sugars and glycosylamines. Opening of the ring of the cation XI11 would leave C1 with a transient positive charge (as shown by XIV) to which an electron pair from C2 could be attracted to form the enol (XV) directly, without formation of a Schiff-base intermediate.52 A similar mechanism was proposed by GottschalkZ2for conversion of the 1-amino-1-deoxy-2-ketose to the labile 1,2-en01 (XV).

I

CHrOH XI11

I

CHzOH XIV

&H~OH

xv

Whether or not enolization of a sugar or glycosylamine can occur without the formation of an intermediate carbonyl or azomethine group, respectively, is a moot question, discussion of which is beyond the domain of this it is difficult to imagine in the first Chapter (see P e t ~ e l y57). ~ ~However, . place how the weakly basic oxygen atom of the sugar ring could preferentially take on the catalyzing proton in face of the more strongly basic nitrogen atom which is present in the same molecule. Moreover, on present evidence, the Amadori rearrangement does not Occur unless the nitrogen atom is sufficiently basic to accept a proton; hence, any reaction mechanism which ignores t,he primary role apparently played by the amino-nitrogen atom is to be questioned ad this time. (56) H.S. Isbell and Harriet L. Frush, J. Research Natl. Bur. Standwds, 46, 132 (1951). (57) F. Petuely, Angew. Chem., 66, 268 (1953).

184

JOHN E. HODGE

3. Comparison with the Lobry de Bruyn-Alberda van Ekenstein Transformation The Aniadori rearrangement has some features of the Lobry de BruyiiAlberda van Ekenstein transformation,68as can be seen from the ammono analogy to sugar enolization formulat>edin Part 2 of this Section. 130th reactions occur in basic media, and each doubtless involves 1,2-enolization of the sugar. However, the Amadori rearrangement proceeds by acceptance of a proton from the acid catalyst, whereas the Lobry de Bruyii-Alberda van Ekeiistein transformation proceeds by delivery of a proton to the base catalyst. Aside from what may be argued as to the enolizatiori mechanism, there are other important differences. -As no demonstration of reversibility or of a quasi-equilibrium has been made for the Amadori rearrangement. Either epimer of an aldosylamine (e.g., either N-substitut,ed D-glucosylaniine or N-substituted D-mannosylamine) may be used as starting material, with production of the ketose derivative in good 3 7 , 38 But, despite their excellent crystallizing properties, N-substituted ~-mannosylamineshave not been found during the isomerization of N-substituted D-glucosylamines. Nor have both the N-substituted glycosylamine and the Amadori rearrangement product been isolated from the same reaction mixture. By the Lobry de RruynAlberda van Ekeiistein transformation, both D-glucose and D-mannose can be produced, starting with D-fructose. But neither epimer of a N-substituted aldosylamine has been obtained from a N-substituted l-amino-l-deoxy-2ketose. The reason for these differences probably lies in the greater lability of the intermediate eneamiriol (XV), supposedly formed in solution during the Amadori rearrangement. Decomposition accompanied by browning occurs in both the Lobry de Bruyn-Alberda van Ekensteiri t r a n s f ~ r m a t i o n and ~ ~ the Ainadori rearrangement. In aqueous solutions under conditions of equal p H , browning is more severe in the presence of amines than in the presence of strong alkali.38This can be explained by the different nature of the decomposition products in the two reactions. Decomposition in aqueous alkali arises mainly from sugar fission and saccharinic acid rearrarigemeiit~.~~ 5 9 Sugar acids, including lactic acid and fatty acids, are known to be relatively stable toward further decomposition with browning.23On the other hand, in the Amadori rearrangement, the main decomposition reactions are sugar fission and sugar dehydration, both of which produce very reactive intermediates (for example, a-dicarbonyl compounds, a ,b-unsaturated aldehydes, and reductones) which continue to react, in the absence of high (58) L. Sattler, Advances zn Carbohydrate Cheni., 3, 113 (1948). (59) J , B. Gottfried and D. G. Benjamin, Z 7 d . Eng. Chem., 44, 141 (1952).

AMADORI REARRANGEMENT

185

concentrations of hydroxyl ions, to form colored by product^.'^ Therefore, reversihi1it)y and estahlishnient of a quasi-equilibrium during the Amadori rearrangement cannot be expected, unless sugar decomposition is blocked.

VI. PHYSICAL PROPXRTIES OF AMADORI REARRANGEMENT PIEODUCTS 1. Color and Taste

Like the N-suhstitut,ed glycosylamines, the N-substituted 1-amino-1deoxy-2-ketoses are colorless when the parent amine is colorless. 1-DeoxyI -p-toluidiiio-D-fructose is t n ~ t ~ e l e sbut s , ~ derivatives of the more strongly basic. amiiies are bitter. 2. Crystallization The rearrangement products derived from aromatic and non-aromatic heterocyclic amines crystallize readily from the lower alcohols. Unlike those of many of the N-substituted glycosylamines, the crystals are not solvated. On the other hand, the ketose derivatives of aralkyl- and alkyl-amines, such as 2-phenylethylamine, ethanolamine, diethanolamine, glycine ethyl ester, and phenylalanine (see Table II), are hydrated or alcoholated, or both, and are difficult to isolate in pure crystalline form. The crystals which have been isolated were hygroscopic.Z2 38 Alcohols, aqueous alcohols, and water are the most commonly used solvents for crystallization. Acetone, ether, or benzene have been added to the alcoholic media in order to increase the yield of crystalline compound. The use of solvents that contain peroxides promotes decomposition of the crystals during storage.60 The crystalline form of the 1-amino-1-deoxy-2-ketoses varies. Most commonly, acicular needles or lamellar plates are formed, but columnar prisms are also known. Amadori3' reported that, a t different times, his stable isomers crystallized from the same solvent as either needles or flakes. 9

3. Melting Amadori rearrangement products, like the N-substituted glycosylamines, generally melt with decomposition. I n conformity with the greater stability of the rearrangement products as compared with that of the glycosylamines, the former usually melt and decompose a t higher temperatures, or with slower decomposition a t a given temperature. I n the aromatic series, virtually all of the currently known rearrangement products melt a t some 30 (or more) degrees above the melting points of the corresponding glycosylamines. With strongly basic piperidine derivatives, there is little difference in the decomposition temperatures. However, the ketose derivative does decompose more slowly than the aldose derivative a t temperatures below the melting point.37*38

186

JOHN E. HODGE

Kuhn and Birkofer13 reported that aromatic N-substituted glyrosylamines generally foam upon melting, whereas the Amadori rearrangement products melt and decompose without foaming. 4. Optical Rotation

All the Amadori rearrangement products so far isolated in crystalline form belong to the D-series. As all but one of them are D-fructose derivatives, they are rather strongly levorotatory. Mutarotation of the rearrangement products is commonly observed in absolute methyl alcohol or ethyl alcohol solut,ions, and also in dry pyridine (which is the best solvent)).The change in rotation is generally slow, and the rotation decreases from a large negative value to a smaller, nearly constant, negative value. On the other hand, solutions of these products in water or dilute acid display a constant rotation over a period of several hours. Unfortunately, most of the aminodeoxyketoses derived from aromatic amines are not appreciably soluble in water. They are, however, soluble in dilute acids; hence, it, is recommended that water containing at least one equivalent of hydrochloric acid be used in determining the optical rotations of Amadori rearrangement products. A small excess of acid does not alter the rotation. Aqueous solutions of the hydrochloride salts of aminodeoxyketoses, and also of the aminodeoxyalditols, generally exhibit an optical rotation of higher magnitude than that of the parent basic a m i n e ~ .3~7 *. 3 * , 6n Another advantage is gained by the use of dilute hydrochloric acid: because the isomeric N-substituted glycosylamines are easily hydrolyzed-to give, finally, the rotation of the unsubstituted sugar-measurement of the rotation in acid solution affords a check on the identity of the compound. The optical rotations of the known 1-amino-1-deoxy-2-ketoses are recorded in Table I (page 203). 5. Light Absorptian According to ultraviolet-absorption measurements made by Kuhn and Dansi,l the spectrum of 1-deoxy-1-p-toluidino-D-fructosein alcohol is insignificantly different from that of N-p-tolyl-D-glucosylamine (or of pure p-toluidine61*'j2).Because an unsaturated linkage in conjugation with those of the aromatic nucleus is not indicated by the spectra, neither N-p-tolylD-glucosylamine nor the rearrangement product shows the structure of a Schiff base in alcohol. L e g a ~ came ~ ~ to the same conclusion on the basis of infrared measurements on the same two compounds in the solid state. No absorption a t 6.05 p, corresponding to a -C=Nlinkage, was found. (60) (61) (62) (63)

K . Zeile and W. Kruckenberg, B e r . , 76, 1127 (1942). R . G . Cooke and A . K . Macbeth, J. Chem. Soc., 1595 (1937). F. Pruckner and B. Witkop, Ann., 664, 132 (1943). F. Legay, Conzpt. rend., 234, 1612 (1952).

AMADORI

REARRANGEMENT

187

However, he did find an absorption band a t 6.05 p for solid N-o-tolyl-Dglucosylamine, and also for N-2-naphthyl-~-glucosylamine,indicating for the first time the existence of a Schiff-base structure for N-substituted glucosylamines in the solid state. For the other N-substituted D-glucosylamines. tested (N-phenyl, N-p-nitrophenyl, and N-rn-nitro-p-tolyl), no Schiff-base structure was indicated. All compounds exhibited a band a t 2.88 p which could have been given by either an NH or an OH grouping. VII. CHEMICAL PROPERTIES OF AMADORI REARRANGEMENT PRODUCTS 1. Enolization and Oxidation

Ketoses are more easily dehydrogenated than aldoses, to yield the mutual osone. For example, aldoses are not attacked by hot selenious acid whereas, under the same conditions, D-fructose reduces the reagent to selenium.64The oxidation product, at least in part, is D-glucosone.66Hydrazine also dehydrogenates D-fructose, but not D-glucose, in weakly acid or weakly alkaline solution at 100’. That the chief product is D-glucosone (or an imino analog of D-glucosone) was shown by Weygandand Bergmann18 through the quick formation of the phenylosazone or tetrahydroxybutylquinoxaline derivatives a t low temperatures. Furthermore, D-fructose and * 68 or D-threo-pentulose are oxidized by cupric salts,@ o-dinitr~benzene,~~ 2 , 6-dichlorophenolindophenol,69in alkaline solution a t ordinary temperatures, much more quickly than are the corresponding aldoses. The ketotriose, 1,3-dihydroxy-2-propanone(dihydroxyacetone), is dehydrogenated or 2 , 6-dichlorophenolindopheno169in alkali, by either o-dinitr~benzene~o and the reaction is quantitative for one mole of enediol, using the latter reagent .71 If N-substituted I-amino-1-deoxy-D-fructoses form ammono analogs of enediols with the same ease that D-fructose is transformed to an enediol, then dehydrogenating agents would be expected to distinguish between N-substituted aldosylamines and the corresponding Amadori rearrangement products. Such has been found to be the case. The distinction between (64) H. L. Riley, J. F . Morley and N. A. C. Friend, J . Chem. Soc., 1875 (1932). (65) K. C. Dixon and K. Harrison, Biochem. J. (London), 26, 1954 (1932). (66) 0. T. Schmidt and R . Treiber, Ber., 66, 1765 (1933). (67) H. L. J. Chavassieu and A. Morel, Compt. rend., 143, 966 (1906). (68) R . Kuhn and F. Weygand, Ber., 69, 1969 (1936). (69) H. von Euler, H . Hasselquist and G. Hanshoff, Arkiu Kemi, 6, 471 (1954). (70) W. R. Fearon and E. Kawerau, Biochern. J . (London), 37,326 (1943). (71) H. von Euler and H. Hasselquist, “Reduktone,” Sammlung Chemischer und Chemisch-technischer Vortriige, No. 50, Ferdinand Enke, Stuttgart, Germany, 1950, p. 6.

188

JOHN E. HODGE

the two is quite sharp when use is made of such oxidation-reduction indicators as o-dinitrobenzene,z' 1 3 , 2 2 , 38 Methylene Blue,l3,2 2 ~ 2 8 38 , and 2,6-dichlorophenolindophen~l~~~ 3 7 , 38 in alkali a t 25". If the test with o-dinitrobenzene is carried out according to Fearon and Kawerau's direction^,^^ one may distinguish between true reductones (that is, stable enediol-acarbonyl compounds, such as ascorbic acid, triose-reductone, and reductic acid, which produce the purple color immediately), Amadori rearrangement products (which form the color after about 1 minute), and N-substituted glycosylamines (which give no color within 15 minutes or longer).38Other oxidizing agents, such as permanga~iate,~ Benedict,2" Fehling,l3. and toll en^'^ solutions, also show a faster rate of reaction with the aminodeoxyketose derivative, but the distinction is not as marked as with the use of enediol reagents. A strongly reducing enediol(s) is formed from an Amadori rearrangement product only in strongly alkaline solution. For example, in methanolpyridine-water, N-substituted 1-amino-1-deoxy-2-ketoses give a negative test with titanium trichloride by the method of Weygand and C s e n d e ~ , ' ~ indicating the absence of an a-enediol gr~uping.'~ The explanation for the lack of a definite end-point upon titration of Amadori rearrangement products in alkali with Methylene Blue or 2 , 6-dichlorophenolindopheno113 may lie in the finding of P e t ~ e l that y ~ ~D-glucosone reacts with alkali t o form one or more reductones (a-enediol-a-carbonyl compounds which are strongly reducing, even in acid solution). Because the phenylosazone and tetrahydroxybutylquinoxaliiie derivatives of D-glucosone have been isolated from 1-deoxy-1-p-toluidino-D-fructose in high yields,l8' l9 1,2-enolization of the aminodeoxyketose must predominate over 2,3-enolization. No proof for 2,3-enolization is known a t present,. The sequence of reactions (in alkali) which lead from tbe aminodeoxyketose (XVI) through the 1,2-enediol (XVIII) to the glycosone (XIX) are thought to be as follows: f

XVI

XVII

XVIII

Hydrolysis of the eneaminol (XVII) must precede the dehydrogenation in (72) F. Weygand and E. Csendes, Chem. Bw.,86, 45 (1952). (73) J. E. Hodge, unpublished observations. (74) F. Petuely, Monatsh., 83, 765 (1952).

189

AMADORI REARRANGEMENT

alkali. Although it is true that, if R‘ = H in XVII, dehydrogenation of the eneaminol could occur before hydrolysis, such would not be t,he case when -NRR’ is a secondary amino radical-yet secondary amino rearrangement products are as readily dehydrogenated in alkali as are those derived from primary a m i n e ~ .38~ It ~ , has been shown that the anilino derivative of a grouping such as XVII (R = phenyl, R,’ = H) will reduce o-dinitrobenzene in alkali, whereas the N-methylanilino derivative (R = phenyl, R’ = CH3) will 76 The initial reversibility of the enolization (XVI XVII) was demonstrated by Kuhn and D a n ~ i Dissolution .~ of 1-deoxy-1-p-toluidino-D-fructose in alcoholic sodium hydroxide caused a lowering of the specific rotation from -31.5 to -3.Go, and acidification immediately thereafter restored the initial rotation. Furthermore, addition of sodium hydroxide did not cause displacement of the wavelengths of maximum absorption in the ultraviolet region, showing that D-glucosone or a stable reductone is not formed immediately. Nevertheless, alkaline solutions of aminodeoxyketoses soon turn golden yellow 011 standing, arid later become reddish and brown in the known manner of osone preparations.4’ 5 . 6 , 7 , 38 Amadori4v 5 , showed by diazotization reactions that the amine actually is liberated from the rearrangement products on stJanding in aqueous or alcoholic alkali, and GottschalkZ2recovered nearly all of the amino acid after heating l-deoxy-lphenylalanino-D-fructose in dilute sodium carbonate solution. I n the latter reaction, none of the starting carbohydrate could be detected chromatographically after the alkali treatment. In contrast to the Amadori rearrangement products, the N-aryl-D-glucosylamines are relatively stable in alkali. They do not readily liberate the amine, for their solutions give a constant optical rotation while remaining 6 , 6 ’ However, the N-alkyl-Dcolorless in alkali over extended glucosylamines are much more labile than the N-aryl derivatives in alkali.34 35 36 37 Alkaline solutions of Amadori rearrangement products give an isonitrile odor when heated,7 indicating fission between CI and C2 of the sugar radical. Weygand and B e r g ~ n a n nproduced ~~ this fission oxidatively by shaking the aminoketose with oxygen and a platinum-on-charcoal catalyst in ammonium hydroxide. From the ammonium salt of tjhe enol form (XX, R = CH3, OCH3 , or OC&), the ammonium salt of D-arabonic acid (XXI)

+

l

I

(75) W. Cocker, R . A . Q. O’Meara, J. C. P. Schwarz and E. R . Stuart, J . C h m . SOC.,2052 (1950). (76) W. Cocker, I). S. .Jerikinson a n d P. Scliwarz, .I. Cheiu. SOC.,1628 (1953). (77) F. Weygand and Annemarie Bergmann, Chrm. B e i . , 80, 261 (1947).

190

JOHN E. HODGE

HC-NH-C6H4-R

HC-NH-CsH4-R II c.00 NHe@

I

HOCH I H~OH HCOH I

I

II

0

--c Ha0

R-C,H,-NH,

+ COz -I- H 8

0

9/1

XXII

<

0

II

C-Oo

I HOCH I HCOH I HCOH I

CHiOH

xx

NH,@

CHZOH XXI

was isolated in 53% of the theoretical yield. The free amine was also isolated. N-Formyl-p-toluidine (XXII, R = CH3), when oxidized under the same conditions, took up 48% of the theoretical amount of oxygen and produced p-toluidine together with unchanged N-formyl-p-toluidine. An analogous oxidative fission occurred spontaneously when crystalline acetonylaniline (XXIII, P, = H) was allowed to stand in light and air a t room temperature; it deliquesced rapidly, and produced a strong isonitrile odor together with an acidic, tarry residue. Acetic acid was the only acid detectable in the residue by paper chromatography?8 Since no %carbon acid was found in the residue, the main reaction can be represented by the following equation (R = H).

0

0

CgWs-NH-CHR-C-CH3 XXIII

II

+

0 2

-+

C&NH-C-R

II

+ CH3COOH

XXIV

Likewise, ,Julia11 and succeeded in separating benzanilide (XXIV, R = phenyl) from the oxidative decomposition products of XXIII, R = phenyl. This reaction also occurred spontaneously, and rapidly, in air. Four moles of periodate were required for the oxidation of l-deoxy-lpiperidino-D-fructose and 1-deoxy-1-morpholino-D-fructose a t pH 4.5.38As indicated, one mole of these compounds would be expected to consume four moles of oxidant to produce a-piperidino- or a-morpholino-acetic acid, three moles of formic acid, and one mole of formaldehyde. Although the acid titration showed that three moles of acid were formed, the presence (78) V Wolf, ,inn , 678, 83 (1952) (79) Cited by 1' I, J u l i n n , E W Meyer, A Magnani i t r i d W Cole, J A m . Ch(n2. SOC, 67, 1203 (1945)

191

AMADORI REARRANGEMENT

of the amphoteric ainiiio acid together nith one mole of hydrochloric ticid (used for neutralization) made this result somewhat lower than the espected amount. HZC’

CH %H,

HzC,

,CH* N-He Cle

I

I

I CHz I C=O I

HOCH

I I HCOH I

HCOH

CHzOH

C,H

€1104

HI04

HIO, HI04

*

*

I

-CH&OOH

H@ c1”

+ + HCOOH + HCOOH + CHLO HCOOH

2. Hydrogenation Amadori rearrangement products can be hydrogenated readily in water or in aqueous alcohols in the presence of a platinum7*1 3 , or Raney nickel catalyst.38Sodium amalgam has also been used, but the alkali-lability of the 1-amino-1-deoxy-2-ketoses makes this reagent less desirable.I6 Weygandl6 observed an uptake of three moles of hydrogen by one mole of 1-deoxy-1-p-toluidino-D-fructose in acid solution (Adams platinum catalyst); this led him to assume that only the aromatic nucleus was hydrogenated. The hydrogenated acid solution was then made alkaline, and one additional mole of hydrogen was consumed, presumably to hydrogenate the enol form of the ketose. Weygand’s experiments were conducted a t 20” and atmospheric pressure; and the hydrogenation products were not isolated or identified. Since it is now known that Amadori rearrangement products are very easily dehydrated in acid solution to form furfural derivatives (see Section VII, Part 3), the chance that such reactions might occur during hydrogenation, particularly at elevated temperatures, should not be overlooked. I n weakly alkaline solution, hydrogenation with a platinum catalyst ( Z O O , atmospheric pressure) is limited to the sugar radical, and l-arylamino1-deoxy-2-ketoses are then readily converted to the two epimeric l-arylamino-1-deoxy-glycitols. However, both epimeric aminoglycitols have actually been isolated in only one experiment. According to the experimental conditions, one of the two possible isomers is usually formed to the almost complete exclusion of the other reaction product. As Weygand explained,80 such behavior is to be expected, siwe the en01 form of the aminoketose can (80) F. Weygand, B e r . , 73, 1278 (1940).

192

JOHN E. IIODGE

exist in either the cis (XXV) or t r a m (XXVI) arrangement, depending on IiNH

\ /

I1

C II

c / \

HOCH

I

HNIL

I I c=c I HOCH I CH1

Oo Na@

xxv

HNIl

\ /

H

c

II c o/ \

Na@ 0 HOCH

XXVI

the experimental conditions. As an illustration, l-dcoxy-l-(3,4-dimethylaniliiio)-L-cr!~thro-pentulose,OH hydrogenation a t 10" in alcoholic solution in the presence of the acid catalyst used for the rearrangement, gave oiily the 1-deoxy-l-(3,4-dimethylanilino)-~-arabitol; whereas, in alcoholic solution made alkaline with sodium hydroxide a t 20", only the epimeric L-ribitol derivative was formed.'6 For the hydrogenation products of 1-arylamino- l-deoxy-2-ketoses, Kuhn and Birkofer13 and WeygandB0have developed the following rule for determining the direction of optical rotation from the structural formula. When the hydroxyl group on C2 of the polyhydroxyalkyl chain is on the right (in the customary form of Fischer's projection formula), the optical rotation in pyridine will be negetive, and vice versa. I n other words, the configuration of the second of the first two carbon atoms is determinative when they adjoin a conjugated, unsaturated system in the aromatic aniine; the configuration of the remainder of the sugar radical accounts for only a minor fraction of the total rotation.80This rule holds for the arylamine derivatives only. For example, the two possible hydrogeiiatioii products of I-deoxy-1-piperidino-D-fructose (the amino radical of which contains a saturated ring) do not conform t o this rule; both give negative rotations in pyridine. Also, 1-deoxy-l-piperidino-D-glucitol with the second hydroxyl 38 group on the right gives the less negative A list of the dihydro compounds that have been derived from Amadori rearrangement products (see Table 111, page 205) shows that the l-arylamino-I-deoxy-D-fructoses have given rise preferentially to the mannitol derivatives in every known case. The rearrangement products from galactosyl- and rhamnosyl-arylamines have, on the contrary, given dihydro compounds of the same configuration as the parent sugar. The rearranged aldopentosylamiiies have produced either or both epimers, depending upon the experimental conditions.

3. Dehydration The dehydration of Amadori rearrangement products can occur in a t least two different ways. Under acidic conditions in aqueous media, 5-hydroxymethyl-2-furaldehyde is easily formed from the aniinodeoxyketo-

AMADORI REARRANGEMENT

193

hexoses. I n the presence of amiiie salts and in the dry state (wherein amirie salts are generated upon heating or long storage), six-carbon reductones are formed. a. %Furaldehyde Formation.-Iiuhii and Dam? reported that, hot, dilute hydrochloric acid causcs the decomposition of l-deoxy-l-p-toluidinoD-fructose with formation of much insoluble humin. Also, a strong discoloration was produced by reaction in a solution saturated with hydrogen chloride, after standing a t - 15" overnight. Got,tschalk22demonstrated chromatographically that 5-hydroxymethyl-2-furaldehydeis probably one of the main products of such acidic reactions and also that it is formed under quite mild conditions. For example, D-fructose was heated for 2 hours a t 100" in 2 N acetic acid without producing enough 5-hydroxymethyl-2furaldehyde to be detectable by paper chromatography. Under the same conditions, 1-deoxy-1-p-toluidino-D-fructose gave 36 %, and l-deoxy-lphei~ylalariiiio-D-fructose gave 54 %, of the theoretical amount of 5-hydroxymethyl-%furaldehyde. Concurrently, 51 and 52 % of the amine present was released, and the formation of melanoidins was also observed. Although the corresponding N-substituted aldosylamine derivative also yielded 5-hydroxymethyl-2-furaldehyde (29 %) under these conditioiis (with acetic acid), the result, was quite different in 1 N hydrochloric acid. In this mineral acid, 100% of the amine, 90% of the aldose, and only 3 % of 5-hydroxymethyl-2-furaldehyde mere formed. Under the same conditions, the two rearrangement products gave I 4 to 18 % of 5-hydroxymethyl-2-furaldehyde, 51 and 52% of the amine, and no sugar. It must be pointed out that 5-hydroxymethyl-2-furaldehyde mas not positively identified in Gottschalk's chromatographic experiments, and the analytical procedure was only semiquantitative. The significance of these findings for an explanation of non-enzymic browning of the sugar-amine (Maillard) type has been pointed out e l s e ~ h e r e . ~ ~ After the foregoing demonstration, one might suppose that ketose reagents that, for a positive color reaction, are said to depend on the formation of 5-hydroxymethyl-2-furaldehyde might be suitable for distinguishing betweeii Amadori rearrangement products and the isomeric N-substituted glycosylamines. When such tests were carried out, however, l-deoxy-l-ptoluidiiio-D-fructose gave a negative test with the customary Seliwaiioff reagent (resorcinol in hot 4 N hydrochloric acid).22'33 However, the test was positive when performed in ethanol saturated with hydrogen chloride.22 Tests made a t the same time on D-fructose and N-p-tolyl-D-fructosylamine with the aqueous reagent were positive.33H ~ d g e 'has ~ been unable to distinguish between Amadori rearrangement products and the corresponding N-substituted glucosylamines with three different ketose reagents: Tauber,81 (81) H. Tauber,

J. Biol. Chem., 182, 605 (1950).

104

JOHN E. HODGE

Ihl-Pechmann (according to Theis and Kallinichs2), and Balde6n.83Each of these reagents gave a sharp distinction between D-fructose and D-glucose. 1-Deoxy-1-piperidino-D-fructose gave no characteristic color with any of the three reagents. Some other N-substituted 1-amino-1-deoxy-D-fructoses gave positive tests with the latter two reagents, but so did the glucosylamine derivatives. Tauber's reagent (aminoguanidine in concentrated sulfuric acidsL)gave no color with any of the aldosylamines, nor was the color given with aminodeoxyfructoses that which is characteristic for ketosesfaint brownish-yellow and pale-blue colors were formed instead. An explanation for the negative tests with ketose reagents may lie in the interference presented by the amine of the aminodeoxyketose under test. However, another point to be considered is that, in concentrated acids, the 1-amino-1-deoxy-2-ketoses may not form much of the 1,2-enol, and, from it, 5-hydroxymethyl-2-furaldehyde. For example, if the 2,3-enol were formed, 2-hydroxyacetylfuran instead of 5-hydroxymethyl-2-furaldehyde would result, according to the reaction scheme of P e t ~ e l y . ~ ~ b . Reductone Formation.-Hodge and Rist3*heated l-deoxy-l-piperidinoD-fructose in the dry state under diminished pressure and observed the formation of a t least two reductones. One was distilled, under diminished pressure, from the residue as a fragrant oil, and the other was left in crystalline form after extraction of the red-brown residue with methanol and acetone, The crystalline reductone possessed all the reducing properties of ascorbic acid, was optically inactive, and gave analyses corresponding to the formula CllH17N03.The net effect of the reactmionon one mole was, therefore, a dehydration with the loss of two moles of water from the sugar radical and without the loss of the piperidino radical. Fission and recombination of fragments is a possible mode of formation, as well as direct dehydration of the sugar moiety. The structure of this reductone, and its derivation from Amadori rearrangement products, have not yet been determined. Since the same reductone, CllH17N03, was formed spontaneously on allowing pure N-D-galactosylpiperidine to stand and undergo browning decomposition a t 25" during more than 2 years, and also with a shorter reaction time under conditions known to produce the Amadori rearrangement,38this dehydration reaction appears to have significance for the elucidation of non-enzymic browning reactions.23 4. Fission

Fission of the sugar radical in Amadori rearrangement products in the l-deoxyabsence of air was demonstrated by Hodge and R i ~ tOn . ~heating ~ (82) H. Theis and G. Kallinich, Chem. Ber., 86, 438 (1952). (83) E. R . Baldebn, Actas y trabujos congr. peruano quim., 3rd Congr. (Lima, Peru), 2, 448 (1949); Chem. Abstracts, 46, 6119 (1951).

AMADORI REARRANGEMENT

195

I-piperidino-D-fructose in the dry state for 5 hours a t 110" under diminished pressurc, piperidinc acetate sublimed arid was isolated in 20 % yield; the formation thereof indicates that the sugar radical rearranged to yield acetic acid and that piperidine was split from the compound. Dehydration of the sugar radical occurred at the same time (as described in the preceding paragraph). [How the sugar rearranged in order to produce acetic acid is undetermined (for further discussion on this point, see Ref. 23).] This result is not entirely without precedent; Sowden and Schaffers4have shown that the chief volatile acid formed in the Lobry de Bruyn-Alberda van Ekenstein transformation, when this is conducted in the absence of air, is acetic acid. 47 At a time when the rearrangement was as yet unknown, was unwittingly studying the Amadori rearrangement of N-aryl- and N-benzyl-D-glucosylamines and the accompanying decomposition reactions. N-Benzyl-D-glucosylamine, after standing in alcohol with acetic acid, gave a positive test for pyruvaldehyde with sodium nitroprusside. In the absence of acetic acid, the test was negative. D-Glucose (1 mole), benzylamine (possibly 3 moles), and acetic acid (3 moles) were allowed to stand in alcoholic solution at room temperature for 2 days, whereafter the presence of pyruvaldehyde (or acetol-hydroxy-2-propanone) was demonstrated by adding p-nitrophenylhydrazine and isolating the crystalline bishydrazone of pyruvaldehyde (or acetol). Oxidative fission is discussed under Part 1 of this Section. 5. Derivatives a. Salts.-The solubilizing effect of acids on water-insoluble Amadori rearrangement products4 7 indicates salt formation. In a few instances, the 6o 85 These salts hydrochlorides have been isolated in crystalline are stable, in contrast to those formed from N-substituted aldosylamines. The latter dissociate into the sugar and the amine hydrochloride on standing in a moist atmo~phere.~' b. Esters.-Kuhn and Dansi7 prepared a tetra-0-benzoyl derivative of 1-deoxy-1-p-toluidino-D-fructose by the Schotten-Baumann procedure. The yield was not stated but was probably low, since decomposition of the ketose, with liberation of the amine, is known to occur in alkali. The virtual absence of active hydrogen in the tetrabenzoate was proved by analysis, hence only four esterifiable hydroxyl groups were shown to exist in the original compound. Although Helferich and PortzS5 successfully acetylated 1-deoxy-1-(phydroxyani1ino)-D-fructose in pyridine-acetic anhydride a t room temperature, and obtained a penta-0-acetyl derivative (by acetylation of one (84) J. C. Sowden and R . Schaffer, J. A m . Chem. SOC., 74, 499 (1952). (85) B. Helferich and W. Portz, Chem. Ber., 86, 604 (1953).

196

JOHN E. HODGE

phenolic and four sugar hydroxyl groups), Hodge and R i ~ encountered t ~ ~ difficulties in acetylating the 1-piperidino, 1-morpholino, and l-dibenzylamino derivatives of 1-deoxy-D-fructose in this way. Even a t a reaction temperature of 0", dark, red-brown, water-soluble sirups were obtained. (The dihenzylamino derivative did yield38 a small amount of colorless, crystalline triacetyl compound.) The miscarriage of these acetylations, with concomitant color formation, can be attributed to the ease with which the aniinodeoxyf ructoses are dehydrated. c. Ethers.-In view of the lability of Amadori rearrangement products i n alkali, it is not surprising that the methyl ether derivatives arc unknown. Kuhn and Dansi7 conducted an attempted methylation of l-deoxy-l-ptoluidino-D-fructose with methyl iodide and silver oxide, only to find the methylated fission product, N-methyl-p-toluidine, as the sole crystalline product. The 5-trityl ether of 1-deoxy-1-piperidino-D-threo-pentulose was formed when Zeile and KruckenbergGoattempted the tritylation of N-D-xylosylpiperidine hydrochloride in pyridine. In acetone containing 1% of hydrogen chloride, the corresponding monoisopropylidene ketal was readily formed3sfrom l-deoxy-l-piperidinoD-fructose. The crystalline product, which separated as the hydrochloride, still retained the free reducing group. Upon oxidation of one mole of the hydrochloride with sodium metaperiodate, one mole of the oxidant was consumed and 0.66 mole of acid was formed. Assuming that a single acetone molecule would react preferentially with cis hydroxyl groups on contiguous carbon atoms, and that the low acid titer was caused by the formation of piperidinoacetic acid hydrochloride, the isopropylidene compound is most probably the 4,5-O-isopropylidene pyranose derivative (XXVII).

Hk0 XXVII

d. O x i m e s and Hydraxones.-Crystalline oxiiiies of l-deoxy-l-p-toluidino-

AMADORI REARRANGEMENT

197

and 1-deoxy-1-piperidino-D-fructose were formed by heating these compounds in alcohol with a small excess of hydroxylamine.2*38 When hydroxylnmine hydrochloride was used, the oxime derivative could be isolated as the hydr~chloride.~~ Colorless phenylhydrazones of 1-deoxy-1-piperidino- and l-deoxy-l. ~ ~ a small morphoho-D-fructose were prepared by Hodge and R i ~ t Only excess of phenylhydrazine, without any acetic acid, was used (in order to prevent osazone formation, because osazones are known to be readily formed from Amadori rearrangement products in the presence of acids).17.86 Neither the oximes nor the phenylhydrazones reduced 2,6-dichlorophenolindophenol in aqueous or alcoholic alkali, and hence the presence of a carbonyl or hemiacetal reducing group in the original compounds was demonstrated. e. 0sazones.-Phenylosazones of the sugars can be made more quickly, and in much higher yields (72-SO%), by starting from l-arylamino-ldeoxy-2-ketoses instead of from the corresponding aldose sugars, as was shown by Weygand.17The arylamino radical is replaced quantitatively by the phenylhydrazono radical during the reaction. It is not necessary to start with the Amadori rearrangement product; an arylamine may be added to the usual reaction mixture in order to increase the yield.17r87 The addition of aniline to reaction mixtures containing an aldopentose or aldohexose plus phenylhydrazine and acetic acid increased the rate of osazone formation many times over that of the control. Thus, aldoses could be distinguished from ketoses by noting the increased velocity of osazone formation after adding aniline to the reaction mixtures to promote rearrangement to the k e t o ~ e Ketoses .~~ do not exhibit this acceleration, because, unlike aldoses, they can be directly dehydrogenated to the osone (or imino analog) by hydrazines.'s Weygand has suggested two possible mechanisms for osazone formation which depend on the occurrence of an Amadori rearrangement after initial condensation of the hydrazine with the carbonyl gr0up.1~Both routes were presented and reviewed by Percivals8 in Volume 3 of this Series. Since that time, Weygand's theory has remained valid. Criticism by Ruggli and Zellersg was satisfactorily answered by Weygand and R e c k h a u ~ , ~ who o showed that (86) See V. Wolf, Chem. Ber., 86, 840 (1953). (87) F. Weygand a i d Margaret Reckhaus, Chein. Ber., 82, 442 (1949). (88) E. G . V. Percival, Advances 712 Cuibohydrafe Cheni., 3, 42 (1948). (89) P. Ruggli and P. Zeller, Hrlv.Chiirr. Actn, 28, 747 (1945). (90) F. Weygand and Margaret Reckhaus, Ckeni. Ber., 82, 438 (1949).

198

JOHN E. HODGE

the mechanism suggested by Ruggli and Zeller was incorrect. Furthermore, the experiments of Wolfa6with model systems representing l-arylamino-ldeoxy-Zketoses have upheld Weygand's theory. Wolf showed, through quantitative determinations of the yield of each reaction product, that acetonylarylamines readily form the p-nitrophenylosazone of pyruvaldehyde according to the following mechanism (which is the same as that proposed for arylamino-1-deoxy-%ketoses by Weygand"). The quantitative relationships were the same, regardless of the arylamine substituent R (which was varied from H to CH3 t o COOGH6).

HC-NH-CsH

II

a-

It

c-NH-NH-C~H~-NO~

I CHI

HC=N -CrH t-R

I I

C=NH

+ NO2 -C~HI-NH,

CH,

.f. QuinozaZines.-It is possible to prepare 2-(~-arabo-tetrahydroxybutyl)yuinoxaline in 90% yield by starting with one mole of l-deoxy-l-p-toluidino-D-fructose and adding about two moles of hydrazine in addition to two moles of o-phenylenediamine. I n a manner reminiscent of the osazoneforming reaction, the o-phenylenediamine replaces the p-toluidino radical, with high yields. The advantage that hydrazine gives in the quinoxaline preparation was demonstrated by Weygarid and 13ergmann,18who found that it dehydrogenates the aminoketose to D-glucosone or one of its imino analogs, although it does not dehydrogenate D-glucose. The dehydrogenation, therefore, takes place on an intermediate in the synthesis, such as the hydrazone of the Amadori rearrangement product (XXVIII).'8 The prototropic shift t o X X I X and fission of X X I X to give the imino compound (XXX) would then follow as in Weygand's proposed niechaiiism for osazoiie formation,17 to yield finally 3-(D-arabo-tet,rahydroxybutyl)quillosaliiie (XXXI); see also, Ohle and Kriiyff .O1 (91) H. Ohle and J. J. liluyff, Ber., 77, 507 (1944).

199

AMADORI REARRANGEMENT

NH

NHz

If11

I HOCH I HCOH I HCOH I

CH20H XXVIII

N

I

I I

II

1

A

II C-NH-NHz I

HOCH I HCOH

I

HCOH

I

CHiOIl XXlS

-

Q R NII,

CH

Cli

CHz CzN-NHZ

NII:

I C=NH I

- i y ~ ~

I

HOCH

HCOH

I

HCUH I CHzOH XXX

N N \ / HC-C

- i y ~ ~

I 1 HCOH I HCOII I

HOCH

CIIZOH XXXI

VIII. P R O O F OF STRUCTURE OF AMADORIREARRANGEMENT PRODUCTS Amadori believed that his three “stable” compounds were all Schiff bases, although he recorded no steps taken to prove their structure. Kuhn and Dansi7 worked with the “stable” isomer Amadori had derived from D-glucose plus p-toluidine, and showed that it could not be a Schiff base. On catalytic hydrogenation, an unknown dihydro compound of m. p. 195” was isolated; whereas, if the LLstable” isomer had been a Schiff base, the known 1-deoxy-1-p-toluidino-D-glucitol, of m. p. 143”, would have been formed. Kuhn and Weygandz next showed that the dihydro compound melting for they made this compound at 195” is 1-deoxy-1-p-toluidino-D-mannitol, by hydrogenating N-p-tolyl-D-mannosylamine and proved its identity with Kuhn and Dansi’s hydrogenation p r o d ~ c t .It~ was then evident that a transition from a D-glucose radical to a D-mannose radical had been effected. Since the original ‘Lstable”isomer was not split by mild, acid hydrolysis and gave the strong reducing behavior toward o-dinitrobenzene in alkali characteristic of ketoses, Kuhn and Weygand pronounced the LLstable” isomer to be N-p-tolyl-D-isoglucosamine (111).They submitted confirmatory evidence in that the compound formed an oxime and therefore possessed a carbonyl group or a hemiacetal ring structure derived from it.2 The ring form(s) of any one of the various Amadori rearrangement products still has not been determined-nor have the anomeric pairs been isolated. The Amadori rearrangement products are thought to have the p-D configuration, merely because all the known crystalline compounds (which, with one exception, are D-fructose derivatives) show moderate to large levorotations and mutarotate toward the right. Since the structural forms which are involved in the mutarotation are unknown, any formulas now drawn to represent Amadori rearrangement products are purely arbitrary. As yet,, no-one has produced D-fructose from a N-substituted l-amino-l-

200

JOHN E. HODGE

deoxy-D-fructose, nor has anyone started with D-fructose (or any other ketose) and synthesized an dmadori rearrangement product.92Nevertheless, various other experiments have made the 1-amino-1-deoxy-2-ketose structure certain. By separately reacting the same amine with each of two epimeric sugars, D-glucose and D-mannose, the same rearrangement product, a D-fructose derivative in every case, has been formed. This has been accomplished with p-toluidine,I6 d i b e n ~ y l a m i n ep, ~i ~~e r i d i n eand , ~ ~ m o r p h ~ l i n eSince . ~ ~ the two epimeric sugars differ only in the configuration on C2, the location of the keto group can be assumed, with but little room for doubt, to be C3. Any doubt is dispelled after referring to the results of hydrogenation experiments. Hydrogenation of the rearrangement products has given either, or both, epimeric l-amino-l-deoxyglycitols,’3~ 16, 38 differing only in the configuration of the hydroxyl groups on C2. Moreover, l-arylamino-l-deoxyhexitols and -pentitols have been oxidized back to the corresponding aldose and 2-ketose sugars with bromine in aqueous solution.93Because the l-amino-1-deoxyglycitols prepared by hydrogenation of the rearrangement products have also been prepared by hydrogenation of the N-substituted glycosylamines, the location of the amine radical on C l of the ketose is established beyond doubt. Catalytic oxidation of three arylamino-1-deoxy-D-fructoses has produced the ammonium salt of D-arabonic acid (identified as the phenylhydrazide) in 50 % yields. Thus, the warabo configuration of the tetrahydroxybutyl “tail” is -fixed, and confirmatory evidence for placement of the carbonyl group on C2 is given. The preparation of D-glUCOSe phenylosazoiie and 2-(~-arabo-tetrahydroxybutyl)quinoxalinefrom the rearrangement products leads to the same concl~sion.’~ The tetrabenzoyl derivative made by Kuhn and Dansi’ contained no active hydrogen, and hence only four free hydroxyl groups are present. Periodate oxidation data38indicated that the four hydroxyl groups are contiguous. Finally, the formation of oxbes2s 38 and phenylhydra~ones~~ confirms the presence of a carbonyl group or a hemiacetal hydroxyl group derived therefrom. Apparently, all that remains t o be done is to determine the various anomer, ring, and enol forms that exist under a given set of conditions.

-

(92) This probably could be done from tetra-0-acetyl-l-chloro-l-deoxy-keto-nfructose, which has been synthesized, starting with tetra-O-acetyl-D-arabonylchloride and diazomethane, by M. L. Wolfrom, S. W. Waisbrot and R. L. Brown, J. Am. Chem. SOC.,64, 1701 (1942). Merely adding the proper amine t o the chloro compound a t low temperatures, followed by deacetylation, should produce the desired N-substituted 1-amino-1-deoxy-D-fructose. (93) F. Weygand and G. Schaefer, Chem. Ber., 84, 603 (1951).

AMADORI REARRANGEMENT

20 1

IX. RETIEOSPECT AND THEFUTURE A survey of the literature on the condensation reactions of sugars and amines, particularly that published before the Amadori rearrangement was known, indicates that “rearrangement” may occasionally have gone unnoticed. For example, the studies of Cameron46*47 on the color-producing, decomposition reactions of D-glucose with aniline, the toluidines, and benzylamine were made under conditions which are now known to produce the Amadori rearrangement. Cameron observed positive, Seliwanoff tests for ketose, as well as the more rapid reduction of Methylene Blue and permanganate for the reactions that occurred in the presence of acetic acid, indicating that the rearrangement had occurred. VotoEek and Wichterleg4investigated the reaction of N-substituted glycosylamines with hydrocyanic acid, but they did not start with the isolated glycosylamine. The sugar and amine were heated in alcohol, the hydrocyanic arid was added, and the mixture was allowed to cool. The structure of the crystalline nitrile isolated was not determined. Because the hydrogen cyanide could have added to the carbonyl group of the ketose after an Amadori rearrangement had occurred, the constitution of their products is open t o question. I n other work, VotoEek and Valenting5investigated the reaction of methyl or ethyl acetoacetate, and of 2,4-pentanedione (acetylacetone), with rhamnose or mannose in alcoholic ammonia. Owing to the presence of active methylene compounds and ammonia, the conditions were favorable for the occurrence of an Amadori type of rearrangement.38 The proportions of one mole of sugar reacting with one mole of ammonia and one mole of the active methylene compound were firmly established, but the structures proposed for the crystalline reaction products were not proved. The structures of the crude, tan-colored compounds that Hodge and Rist3*isolated after heating D-glucosylamine or N-(2-hydroxyethyl)-~-glucosylamine in the presence of 2,4-pentanedione or ethyl malonate were also not determined, but alkaline solutions of the products reduced 2,6-dichlorophenolindophenol rapidly and extensively a t 25” in the manner of Amadori rearrangement products. Hence, this is another subject for future investigation. In several other studies reported in the literature, acids and acidic salts have been used in order to catalyze the condensation of sugars with amino compounds, without subsequent testing of the product to see if an Amadyri rearrangement had occurred. Kuhn and BirkoferI3 thus missed observing (94) E. VotoEek and 0 . Wichterle, Collection Czechoslov. Cheqn. Communs., 9, 109 (1937). (95) E. VotoEek and F. Valentin, Collection Czechoslov. Cheni. Cotiimuns., 7, 290 (1935).

202

JOHN E. HODGE

the occurrence of an Amadori rearrangement in the reaction of D-glucose with dibenzylamine in alcohol containing ammonium chloride. In a reinvestigation of the reaction, Hodge and Rist38used alcoholic alkali in makmg the distinguishing reducing test on the water-insoluble product, Therefore, ii is advisable, in every .cme of a sugar-amine condensation, to make the simple reducing test with o-dinitrobenzene, Methylene Blue, or 2,(+dichlorophenolindophenol, in order to determine whether or not a rearrangement has occurred. The Amadori rearrangement has not received from chemists attention commensurate with its importance. Since the range of the reaction is now broadened, intensive and far-reaching studies of the reaction are needed. In particular, the precise course of the reaction of sugars with primary aliphatic amines arid with amino acids is uncertain. The hydrogen atom remaining on the nitrogen atom in secondary glycosylamines apparently allows further condensations to proceed ; these eventually produce amorphous, polymeric, melanoidin-like substances instead of crystalline ketose derivative^.^^ Those dehydration and fission reactions of sugars that occur so easily, presumably after enolization of the glycosylamines, also need study. More research on these reactions would contribute t o our knowledge of the whole complex of reactions which cause the non-enzymic browning of foods and feeds. The flavor-producing reactions that occur during the ripening and cooking of a host of foods are also involved. Nature’s mode of synthesis of such organic compounds as the polyphenols, tannins, pyrones, flavones, anthocyanins, lignin, mucins, humic acids, etc.., has ever been a challenge to the chemist. It is not inconceivable that many such naturally occurring substances arise through ramifications of one primary reaction-the condensation of sugars with amines. Such reactions as the Amadori rearrangement, sugar dehydration, and sugar fission no doubt play leading roles in the dramatic production that follows.

The following Tables summarize characteristic properties of some Amadori rearrangement products and of certain dihydro derivatives thereof.

X.TABLESOF COMPOUNDS TABLEI Authentic Crgstalline A w m d o r i Rearrangement Products Compound

Parent sugar

Rotalion solwenl

Rejerences

20 20 20 21 24 20 20 20 17 13 20 20 20 17 20 20 20

HzO MeOH C bH gN HCI CHCli CaH& HCl CsHsN MeOH MeOH HCl MeOH CsHd MeOH H2O HCI CsHsN

5, 5 3, 5 5 6 3, 3, 0 7, 3, 4 2, 0 7 7 2 7

22 16 25 20

MeOH H2O CsHsN CdsN

18 18 87, 38 3

25 25 25 25 22 25 25 25

C SHsN H2O CsHbN C sH sN HzO

88

19 19

EtOH EtOH

__-

final

__

__._

1-Deoxy-D-fructose

derivative 1-Arylarnino p-anisidino 3,4-dimethylanilino p-hydroxyanilino pentaacetate 2-nnphthylamino p-phenetidino 0-toluidino p-toluidino

oxime tetrahenzoate 1-Aralkglamino benzylamino 4,6-O-benzylidene acetal hioxalate di benzylamino

1-Alkylamino morpholino phenylhydrnzone piperidino oxime hydrochloride phenylhydrazone 1 -Deoxg-D-threo-pentrrlose derivative I-Alkylamino piperidino 5-trityl ether hydrochloride

G, L , n

G G , Man

G , Mttn G, Man

4&141 -28 -40 61-162 -61 -41. 108 +15. 134 48-150 -21 54-155 -50 -63 26-128 -19. 23-125 -34. 53-154 -50 -45 -63. -23. -42 -56 35-136 -21 19-120

130 -67. -6 170 62-163 -89 59-160 -88 -117 -65. 164 -58. 127 -115 -57. 41-142 -31 65-166 -44 175 -41, 147

68 148

-5 -42

const. -25 -2s -37.4 const. const. -20 -4.0 const. const. -25 -21.5 -10.0

const. -35 -40 -50.8 const. -50.0

- 14 const. const.

HzO H& CaHsN

16 1G

4 16 33 16 38

18

8 8 18 88 8 18

io 10

(96) H. Lehr and H. Erlenmeyer, Helv. Chim. Ada, 29, 66 (1946). (97) Y. Inoue and K. Onodera, Bull. Inst. Chem. Research, Kyoto Univ., Commem. VoE. Silver Jubilee, 139 (1951). (98) B. Helferich and A. Porck, Ann., 682, 233 (1953).

203

204

JOHN E. HODGE

TABLEI1 Non-crustalline, or Incomplete, Authenticated, Amadori Rearrangement Products Camfiaund

Parenl sugar

[a]", degrees

Melting (decamp.) poinl, "C.

inilial

final

~

1 -Deoxy-D,fructose derivative I-anilino 1-diethanolamino 1-glycino, ethyl ester 1-DL-phenylalanino 1 - (2-phenylethylamino) 1 - Deoxy -9- C-pheny l triulose derivative 1-N-methylanilino hydrochloride semicarbazone a

G G G G G

127-130 sirup 60-75 sirup 75-80

Catatia,

C.

Rotation solvent

References

97, 33 38 38 22 38

~

-37.8

const.

13

MeOH

- 13

-8

25

EtOH

- 10

const.

25

CsHaN

103-104 117-119 145-1 46

Dimeric 3-C-phenylglyceraldehyde was the parent sugar.

28 28 28

205

AMADORI REARRANGEMENT

TABLEI11 Dih,ydro Derivatives Parent ketose

Amadori Rearrangement Products flexitol. (I-amino-I-demy-

dleltrng point, "C.

Rejerences

1-L)eoxy-~ f ructose dei ivative 1-Arylaniino

anilino p-anisidino 3,4-dimethylanilino p-phenetidino p-toluidino 1-Alkylarriino piperidino I -Aryhmino-l-deOXy-D-tUgUtOSe derivative p-toluidino 1- A r y l a m i n o - l , 6 - d i d e o x y - ~ - a r a b o hexulose derivative p-toluidino 1-Arylamino-l -deoxy-Derythro-pent dose derivative 3,4-dimethylanilino

175-1 76 191-192 182 185-186 187-1 88 19bl96

+37.4 +27.8 +I4 +21.4 +22 +28.8

19 20 23 19 21 21

16 16 13 16 13 2, 16

116

-17.2

25

38

u-galacti to1

180-181

- 13

20

16

L-rhamnitol

183-184

-19.7

20

16

D -ambit01 o-ribitol

142

+I2 -31.4

21

30 16

L-arabitol L-ribitol L-arabitol L-ribitol

138-139 143 178-1 79 140-141

-12.3 +31 -7.1 $31

21 21 18 19

16 16 16

156-1 58

+26

19

16

wmannitol D -manni to1 D -mannit01 o-manni to1 D -manni to1

D-ylucitol

1-Arylumino-l -deoxy-L-erythro-

pentulose derivative 3,4-dimethylanilino p-toluidino

16

1 -Arylamino-l -deoxy-D-threopentulose derivative p-toluidino a

The rotation solvent was pyridine.

-

This Page Intentionally Left Blank

THE GLYCOSYL HALIDES AND THEIR DERIVATIVES

BY L . J . HAYNES

F . H . NEWTH

AND

T h e University. Edinburgh. Scotland

The University. Cambridge. England

CONTENTS I . Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 207 I1 . Preparation of Glycosyl Halide Derivatives . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 209 1 . By Replacement of an Acyloxyl, Especially an Acetoxyl, Group a t the 210 Potent.ially Reducing Carbon Atom . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . a . By Treatment with Liquid Hydrogen Halide . . . . . . . . . . . . . . . . . . . . . . . . 210 1. . By Treatment with Dry Hydrogen Halide in Dry Ether . . . . . . . . . . . . 212 c . By Treatment wit.h Hydrogen Halide in Glacial AcetJic A d . By Treatment with Phosphorus Halides under Various C e . By Treatment with Titanium Tetrachloride (or Bromide) in Chloroform . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 218 219 2 . By Exchange of Halogen . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . a . Iodides . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 219 b . Halides of the “p-Series” . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 219 3 . By Replacement of the Hydroxyl Group at the Reducing Carbon Atom . . . 221 4. By Replacement of Alkoxyl and Related Groups a t the Potentially Reducing Carbon Atom . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 222 a . Of Glycosides . . . . . . . .......... . . . . . . 222 b . Of Oligosaccharides ....................... 222 c . Of Isopropylidene Compounds . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 225 d . Of Orthoesters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 227 5 . From Glycosans . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 227 6 . By Addition of Halogen and Hydrogen Halide t o Unsaturated Compounds . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 228 111. Structure of Glycosyl Halides . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 229 IV . General . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 233 V . Reaction Mechanism and Effect of Structure on Reaction Rates . . . . . . . . . . 234 VI . Reactions of the Poly-0-acylglycosyl Halides . . . . . . . . . . . . . . . . . . . . . . . . . . . . 230 1 . Formation of Glycosides . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 240 2 . Formation of Glycosylamines (“N-Glycosides”) . . . . . . . . . . . . . . . . . . . . . . 243 3 . Miscellaneous Reactions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 245 VII . Tables of Properties of Some Glycosyl Halide Derivatives . . . . . . . . . . . . . . . . 246

I . INTRODUCTION The glycosyl halides are compounds in which the hydroxyl group a t the reducing carbon atom of a sugar is replaced by halogen . As such, the glycosyl halides are rare, only a few fluorides being well authenticated. However, the fully acylated glycosyl halides, particularly the acetylated 207

208

L. J. HAYNES AND F. H . NEWTH

compounds (sometimes referred to as “acetohalogeno sugars” or “acetohalogenoses”), are very well known and this article will be mainly concerned with them. The most familiar of these is 2,3,4,6-tetra-O-acetyl-ao-glucosyl bromide (“acetobromo-a-D-ghcose”) (I). Some partially aCy1-

“I HCOAc

Aco(7H HCOAc HCO

AcO

I

H

CHIOAC

OAc

I

ated and a few alkylated glycosyl halides are known, but are less important than the fully acylated compounds. The poly-0-acyl derivatives of the glycosyl halides are among the most important intermediates for synthesis in carbohydrate chemistry; moreover, their chemistry has considerable intrinsic interest. I n spite of this, no major review has yet appeared in which this group is discussed as a whole, although various aspects of the formation, chemistry, and synthetic uses of members of the group have received consideration, particularly in earlier volumes of Advances in Carbohydrate Chemistry. Among these are descriptions of the use of glycosyl halide derivatives in the preparation of carbohydrate orthoesters,’ thio- and seleno-sugars,2 anhydro sugar^,^ oligosa~charides,~ and g l y ~ a l s .The ~ preparation and properties of the glycosyl halide derivatives of altrose,6 ribose,’ and fructose8 have also been reviewed in articles on these sugars and their derivatives. Lemieux has recently discussed the implications in carbohydrate chemistry of theories relating to the mechanisms of replacement reaction^,^ and so treatment of reaction mechanisms in this article has been limited to very recent work. I n synthetic carbohydrate chemistry, the poly-0-acetylglycosyl halides first came into prominence as intermediates in the preparation of glycosides (1) E. Pacsu, Advances in Carbohydrate Chem., 1, 78 (1945). (2) A. L. Raymond, Advances i n Carbohydrate Chem., 1, 129 (1945). (3) S. Peat, Advances in Carbohydrate Chem., 2, 38 (1946). (4) (a) G. ZemplBn, Fortschr. Chem. o i g . N a t u r s t o f e , 2, 160 (1939); (b) W. L. Evans, D. D. Reynolds and E. A. Talley, Advances in Carbohydrate Chem., 6.27 (1951). (5) B. Helferich, Advances in Carbohydrate Chem., 7, 210 (1952). (6) N. K. Richtmyer, Advances i n Carbohydrate Chein., 1, 37 (1945). (7) R. W. Jeanloe and H. G. Fletcher, Jr., Advances in Carbohydrafe Chem., 6, 135 (1951). (8) C. P. Barry and J. Honeyman, Advances in Carbohydrate Chem., 7, 84 (1952). (9) R. U. Lernieux, Advances in Carbohydrate Chem., 9, 1 (1954).

GLYCOSYL HALIDES AND THEIR DERIVATIVES

200

(see Section VI, 1). Many natural products occur as glycosides, for example the anthocyanins, the cardiac glycosides, the saponins, and the hydroxyanthraquinone glycosides,I0 and, following the development of Koenigs and Iinorr’s glycoside synthesis, a very large number of compounds of this type have been synthesized using these halides. I n recent years, more attention has been paid to the synthesis of glycosylamines (“N-glycosides”), particularly the biologically important nucleosides and coenzymes (see Section VI, 2). The intrinsic interest in the chemistry of glycosyl halide derivatives will no doubt center on their use in determining the influence of the conformation of the molecule, and of the positions in space of the various substituents, on the reactivity of the halogen atom. Already, there exists a large amount of data, but the task of organizing these data to produce a coherent account of the chemistry of the glycosyl halide derivatives is only begun. 11. PREPARATION OF GLYCOSYL HALIDEDERIVATIVES Most of the methods for the preparation of glycosyl halide derivatives depend on the replacement by halogen of a n acyloxyl (nearly always acetoxyl) group a t the reducing carbon atom. Hence, the methods for the synthesis of a large number of the glycosyl halide derivatives can be exemplified by description of those used for convert,ing fully acetylated sugars into 0-acetylglycosyl halides. As might be expected, treatment for prolonged periods often results in replacement of a second acetoxyl group. Other methods available for the synthesis of glycosyl halide derivatives involve the replacement of hydroxyl and alkoxyl by halogen, the opening of anhydro rings with hydrogen halides or equivalent reagents, and the addition of halogen or hydrogen halide t o certain unsaturated sugar derivatives. Glycosyl iodide derivatives are usually prepared from the corresponding bromides by reaction with sodium iodide. It will be appreciated that, in the same way as the simple glycosides can exist in anomeric forms, the glycosyl halides can exist in two forms. This is exemplified in the two forms, a (11) and p (111), of tetra-0-acetyl-Dglucopyranosyl chloride. With the halides, however, one form is usually CH~OAC

H&g CHzOAc

Ac AcO

I

I

H

OAc I1

I

I

H

OAc

I11

(10) For a brief review, see W. W. Pigman and R. M. Goepp, Jr., “Chemistry of the Carbohydrates,” Academic Press Inc., New York, N. Y., 1948, pp. 459-474.

210

L. J. HAYNES AND F. H. NEWTH

considerably niorc stable than the other. In most of the halides investigated, the stable form appears to be the a ;the most notable exceptions are the derivatives of arabinopyranose, ribopyranose, arid fructopyranose (see Section 111). For this reason, the less stable forms are often referred t o collectively as “glycosyl halides of the P-(D or L) series.” These halides of the P-(D or L) series are very unstable and are characterized by their marked tendency to invert to the stable form, especially in the presence of hydroxylic compounds. Most of the preparative methods to be described give the stable form of the poly-0-acylglycosyl halide as the final product, irrespective of the anonieric configuration of the starting material. There is, however, some basis for supposing that the initial product may be either the unstable or the stable form of the halide, depending on the configuration of the starting material, and that the unstable form inverts either immediately or during the processing of the reaction mixture. The most widely used method for the formation and isolation of the unstable forms of the poly-0-acylglycosyl halides is that of Schlubach,” in which the stable form of a poly-0-acylglycosyl bromide is caused to react with freshly prepared, “activated” silver chloride; a metathetical reaction occurs, accompanied by a Walden inversion to give the unstable form of the chloride. When a method which gives only the stable form of the halide is employed, the yields from either of the anomeric acetates are usually the same. However, Stacey and his coworkers1*have found that crystalline tetra-0-acetyl-a-L-arabinopyranose, on treatment with hydrogen bromide in glacial acetic acid, gives tri-0-acetyl-p-L-arabinopyranosyl bromide in 84 % yield, whereas the sirupy tetra-0-acetyl-p-L-arabinopyranose under identical conditions affords only a 12% yield. 1. B y Replacement of a n Acyloxyl, Especially a n Acetoxyl, Group at the

Potentially Reducing Carbon A t o m a. B y Treatment W i t h Liquid Hydrogen H~lide.’~-This is the only important method for the preparation of poly-0-acylglycosyl fluoride deIt has not been widely used for the preparation of derivatives rivative~.’~ of the other halides, although it has some advantages in the preparation of glycosyl halide derivatives which are so reactive that the use of water during isolation has to be avoided. An example is the preparation of crude tri-0-acetyl-D-ribofuranosyl bromide (IV) which may be obtained in good (11) H. H. Schlubach, Ber., 69, 840 (1926), and subsequent papers. (12) R. E. Deriaz, W. G. Overend, M. Stacey, Ethel G . Teece and 1,. F. Wiggins, J . Chem. Soc., 1879 (1949). (13) E. Fischer and E. F. Armstrong, Ber., 34, 2885 (1901). (14) D . H. Brauns, J. A m . Chem. Soc., 46, 833, 2388 (1923).

GLYCOSYL HALIDES AND THEIR DERIVATIVES

21 1

CHnOAc

OAc

OAc IV

yield15 by treatment of tetra-0-acetyl-D-ribofuranose with liquid hydrogen bromide in a sealed tube at room temperature for 10 minutes. The hydrogen bromide is then evaporated and the last traces of hydrogen bromide and acetic acid are removed by Levene and Tipson’s rnethod,l5&namely, by evaporation with toluene and then with benzene under diminished pressure. This compound (IV) is probably formed on treating the tetraacetate with hydrogen bromide in glacial acetic acid, but it cannot be isolated by the usual means (diluting with chloroform and washing out acetic acid and excess hydrogen bromide with iced water) because of its great reactivity. This method has also been used in the preparation of the bromide.Is very reactive tetra-0-acetyl-0-D-galactofuranosyl Fischer and Armstrong’3 found that treatment of penta-0-acetyl-a-Dglucopyranose with liquid hydrogen chloride gives impure tetra-o-acetylP-D-glucopyranosyl chloride (the unstable form), whereas the P-penta~ This claim was subsequently acetate affords the (stable) a - chloride. withdrawn by Fischer” on the grounds that the first experiment could not be reproduced. However, Schlubach and his coworkers,18 making use of their greater knowledge of the extreme lability of this type of compound, have confirmed that tetra-0-acetyl-P-D-glucopyranosylchloride can be by the action of liquid prepared from penta-0-acetyl-a-D-glucopyranose hydrogen chloride. Prolonged action (for example, for one week) of liquid hydrogen bromide on a poly-0-acetylhexopyranose is the method most used for the preparation of the corresponding 1,6-dibromo-l,6-dideoxy compounds. Thus, penta-0acetyl-0-D-glucopyranose gives 2,3,4-tri-O-acetyl-6-bromo-6-deoxy-a-1~glucosyl bromide (“acetodibromoglucose”) ,19 and penta-0-acetyl-D-galactopyranose givesz0the corresponding “acetodibromogalactose.” This method is quite mild, and nearly always gives the expected product. (15) G. A . Howard, B. Lythgoe and A. R. Todd, ,I. Chent. Soc., 1052 (1947). (15a) P. A. Levene and R. S. Tipson, J . Biol. C h e m . , 90,89 (1931). (16) H. H. Schlubach and E. Wagenitz, Hoppc-Seyler’s Z . physial. C h e m . , 213, 87 (1932). (17) E. Fischer, B e r . , 44, 1898 (1911). (18) €1. H. Hchlubach, P. Stadler and Irene Wolf, B e r . , 61, 287 (1928). (19) E. Fischer and E. F. Armstrong, B e r . , 36, 836 (1902). (20)H. H. Schlubach and E. Wagenitz, Ber., 66, 304 (1932).

212

L. J. HAYNES AND F. H. NEWTH

Two examples in which unexpected products have been obtained are recorded by Brauns in his studies on the preparation of poly-0-acylglycosyl fluorides. Octa-0-acetylcellobiose, on treatment with liquid hydrogen fluoride during t,hirty minutes, gives a moderate yield of the expected hepta-0-acetylcellobiosyl fluoride (V) ,14 but if the reaction time is extended to five hours the main product is 3,6-di-O-acety1-4-0-(2,3,4,6-tetra-Oacetyl-P-D-glucosy1)-a-D-mannosyl fluoride (VI) ,21 in which the acetyl group next t o one potentially reducing carbon atom has been eliminated, the elimination being accompanied at some stage by a n inversion A similar I

HCF

I I

HCOAc AcOCH

I

HCOK

I I

HCO CH~OAC V

I I

HCF

HOCH

' I

AcOCH

I

HY°K HCOI

CH~OAC VI

I

H7°\ HCO/C\CHI I

AcOCH I HCOR

I

HCO

I

CHZOAC VII

reaction occurs when penta-0-acetyl-P-D-fructose is treated for 26 hours with liquid hydrogen fluoride, save that no inversion occurs, the product fluoride.22 being 3,4,5-tri-O-acetyl-~-~-fructosyl b. By Treaiment with Dry Hydrogen Halide in f ) r g Ether.-This method is closely related to the preceding, and is probably one of the mildest procedures for the preparation of glycosyl halide derivatives. Used almost exclusively for the synthesis of the chlorides, the method merely involves dissolving the corresponding acetate in saturated, dry, ethereal, hydrogen chloride solution a t 0" and allowing the solution to stand until there is no further change in optical rotation (2 to 3 days). The solution is then evaporated t o dryness under diminished pressure and the last traces of hydrogen chloride are removed by a similar evaporation with benzene.23 Again, this method has the advantage that there need be no contact with water during the isolation, and so it can be used for the preparation of very reactive compounds. Illustrative of this is its use in the preparation of crude tri-0-acetyl-D-ribofuranosyl chloride from tetra-O-acetyl-D-ribofuran0 8 8 , ~26~ and of tetra-0-acetyl-P-D-glucopyranosyl chloride (111) (in 65 % yield) from penta-O-acetyl-a-~-glucopyranose.2~ I

(21) (22) (23) (24) (25) (26)

D. H . Brauns, J . Am Chem. SOC.,48, 2776 (1926). D. H. Brauns and Harriet L. Frush, Bur. Standards J . Research, 6,449 (1931). Z. H. Skraup and R. Kremann, Monntsh., 22. 375 (1901). J. Davoll, B. Lythgoe and A. R. Todd, J . Chew!. Soe , 967 (1948). H. Zinner, Chem. Ber., 83, 153 (1950). J. J. Fox and I. Goodman, J . Am. C'lierr~.SOC.,73, 3256 (1951).

GLYCOSYL HALIDES AND THEIR DERIVATIVES

213

A product of unexpected structure was obtained by the action of dry hydrogen chloride in ether on a benzene solution of octa-0-acetyl-@-malt o ~ e . ~The ’ - ~reaction ~ mixture was allowed to stand a t - 10”until a crystalline material began to separate and was then kept a t 0” for fifteen hours. There resulted a good yield of a compound isomeric with the hepta-0acetyl-a-maltosyl chloride obtained by the action of liquid hydrogen This new compound was extremely chloride on octa-O-a~etyl-@-maltose.~~ reactive and, in the presence of traces of moisture, decomposed to hepta0-acetylmaltose. With silver acetate, it gave an “octaacetylmaltose” different from the known octa-0-acetyl-a- and -p-maltoses, and, with silver carbonate and methanol, it afforded a “methyl hepta-0-acetylmaltoside,” having properties similar to those of the sugar orthoacetates.’ From this evidence, the chloride has been formulated as a hexa-O-acetyl-l , 2-0-( 1chloroethy1idene)maltose (VII). Oiily one other compound of this type is known.31 33-This c . By Treatment W i t h Hydrogen Halide in Glacial Acetic is the method most widely used for the preparation of glycosyl bromide derivatives, and it may also be employed for the formation of the chlorides and iodides. For the bromide, the acetate is kept a t room temperature with a solution (32 t o 40%) of hydrogen bromide in glacial acetic acid for some 30 t o GO minutes after dissolution is complete; the total time employed varies, but is usually 2 t o 3 hours (For some of the disaccharide acetates and monosaccharide benzoates, which dissolve but slowly, addition of alcohol-free chloroform or methylene dichloride t o the reaction mixture is a d v a n t a g e ~ u s .3~5 .~36) The reaction mixture is then diluted with alcohol-free chloroform and poured onto ice. The chloroform extract is separated, washed with ice-cold water until the washings are no longer acid t o Congo Red, dried over anhydrous sodium sulfate, and evaporated t o dryness under diminished pressure. The resulting sirup is then induced t o crystallize by treatment with appropriate solvents, usually anhydrous ether and light petroleum. The method is convenient, as it involves no special apparatus and the hydrogen bromide-acetic acid reagent may be stored indefinitely; it usually gives excellent yields, and the expected product is nearly always obtained. 1

It has been suggested37that hydrogen bromide in acetic anhydride has (27) (28) (29) (30) (31) (32) (33) (34) (35) (36) (37)

K. Freudenberg and 0. Ivers, Ber., 66, 941 (1922). K. Freudenberg, H. von Hochstetter and H. Engels, Ber., 68, 666 (1925). K. Freudenberg, W. Durr and H. von Hochstetter, Ber., 61, 1740 (1928). K. Freudenberg and H. Schole, Ber., 63, 1969 (1930). B. Helferich and H . Jochinke, Ber., 74, 719 (1941). See also Section 11, 4c. A. Bodart, Monatsh., 23, 1 (1902). E. Fischer and H. Fischer, Ber., 43, 2521 (1910). G. ZemplBn, Ber., 63, 996 (1920). K. Freudenberg and W. Nagai, Ann., 494, 63 (1932). H. G. Fletcher, Jr., and C. S. Hudson, J . A m . Chem. SOC., 72, 4173 (1950). W. W. Pigman and R. M. Goepp, Jr., Ref. 10, p. 160.

214

L. J. HAYNES AND F. H. NEWTH

some advantages over hydrogen bromide in glacial acetic acid as the reagent, in that the solution has a lower vapor pressure and freezing point and that a higher concentration of hydrogen bromide may be obtained. Occasionally, combination of the operations of acetylation and treatment with hydrogen hromide is advantageous when a sugar is to be converted to its “acetobromo” derivative. Full details for one such conversion, in which D-glucose is acetylated with acetic anhydride using a few drops of sulfuric acid as catalyst, and hydrogen bromide is passed into the acetyla.~~ tion mixture, are given in Organic S y n t h e ~ e s Tetra-0-acetyl-a-D-glucopyranosyl bromide is obtained from D-glucose in a yield of 80 to 87% of the theoretical. Nicholas and Smith39 have shown that perchloric acid is an efficient catalyst for the acetylation of sugars with acetic anhydride, and have prepared tetra-0-acetyl-cu-D-glucopyranosyl bromide and hepta-0-acetylmaltosyl bromide in good yield by passing hydrogen bromide into the reaction mixture after completion of the acetylation. A variation on this, in which the hydrogen bromide is generated in situ, has been published.40The sugar is added portionwise to acetic anhydride containing a little perchloric acid, the temperature being kept a t 30 to 40”. When acetylation is complete, amorphous phosphorus is added, the mixture is cooled to 0”, and bromine, followed by 90 % of the stoichiometric amount of water, is added with stirring. After 90 minutes, the mixture is diluted with chloroform and processed in the usual way. The method is convenient, and over-all yields are excellent. By this method, L-arabinose gives a 50 % yield of tri-0-acetyl-P-L-arabinopyranosylbromide, whereas Stacey and his coworkers12were able to obtain only a 30 % yield of this compound by passing hydrogen bromide into a suspension of L-arabinose in acetic anhy~~ a 36 % yield on treating a dride,41s42 and Gehrke and A i ~ h n e risolated sirupy mixture of the a! and forms of tetra-0-acetyl-L-arabinose with hydrogen bromide in glacial acetic acid. It is noteworthy that Stacey and associates12 have found that acetylation of D-arabinose by the perchloric acid method gives a crystalline product, different from both the a! and P forms of tetra-0-acetyl-D-arabinopyranose, which they were unable to identify. Another method for the direct conversion of a sugar to its “acetobromo” (38) C . E. Redemann and C. Niemann, Org. Syntheses, 22, 1 (1942). (39) S. D. Nicholas and F. Smith, Nature, 161,349 (1948). (40) M. BBrczai-Martos and F . Korosy, Nature, 166, 369 (1950). See also, P. G. Scheurer and F . Smith, J. Am. Chem. SOC.,76,3224 (1954). (41) J. Meisenheimer and H. Jung, Ber., 60, 1462 (1927). (42) G. E. Felton and W. Freudenberg, J. A m . Chem. Soc., 67, 1637 (1935). (43) M. Gehrke and F. X. Aichner, Ber., 60, 918 (1927).

GLYCOSYL HALIDES AND THEIR DERIVATIVES

215

or “acetochloro” derivative is treatment with the appropriate acctyl halide4*,4 5 , 4 6 ; it is of historical interest in that it was the method by which tetra-0-acetyl-D-glucopyranosyl chloride, albeit in an impure state, was first prepared.44The procedure has been applied by later worker^,^' 4 8 , 4 9 but is now little used. Bernoulli and StaufferS0have shownsoa that an amorphous 2 , 3 ,4,G-tetra-O-p-tolylsulfonyl-a-~-glucosyl chloride can be prepared by reaction of D-glucose with p-toluenesulfonyl chloride in pyridine a t room temperature for five days. Poly-0-acetylglycosyl chlorides and bromides may also be prepared by reaction of the sugar with a saturated solution of hydrogen halide in acetic anhydride containing acetic 5 2 * 53 The reaction is rather violent, and better yields are obtainable by other methods. d. By Treatment with Phosphorus Halides under Various Conditions.-A method which was a t one time widely used for the preparation of poly-0acetylglycosyl chlorides involves treatment of the corresponding acetate with a mixture of phosphorus pentachloride and aluminum chloride in somc inert solvent, usually either chloroform or acetic anhydride.54,5 5 , 56 For preparat,ive work, this method suffers from the disadvantage that it causes the isornerization of some sugars. For example, octa-0-acetyllactose, by this method, yields the expected hepta-0-acetyllactosyl chloride (VIII,

I I AcOCH I HCOAc I HCOR I HCO I HCCl

HCOAc AcOCH HCOR HCO

I

CHzOAc VIII

CHzOAc IX

(44) A. Colley, Ann. chim. p h y s . , [4] 21. 363 (1870); Conrpt. rend., 70, 401 (1870). (45) H. Ryan, J. Chem. SOC.,76, 1055 (1899). (46) M. van Charante, Rec. trav. chim., 21, 42 (1902). (47) C. S. Hudson and J. M. Johnson, J . A m . Chem. SOC.,37,2748 (1015). (48) C. S. Hudson and F. P. Phelps, J . A m . Chenz. SOC.,46, 2591 (1924). (49) D. H. Brauns, J . A m . Chem. SOC.,46, 1484 (1924). (50) A. L. Bernoulli and H. Stauffer, Helv. Chinz. Acta, 23, 615 (1940). (50a) See also, R. S. Tipson, Advances in Carbohydrate Chem., 8, 107 (1953). (51) J. K. Dale, J . Am. Chem. Soc., 37,2745 (1915); 38, 2187 (1916). (52) C. S. Hudson and R. Sayre, J. A m . Chem. SOC.,38,1867 (1916). 40, 992 (1918). (53) C. S. Hudson and J . K . Dale, J . Am. Chenr. SOC., (54) F. von Arlt, Monatsh., 22, 144 (1901). (55) Z. H. Skraup and R. Kremann, Monatsh., 22, 375 (1901). (56) W. A. Bonner, Advances in Carbohydrate Chem., 6, 251 (1951).

216

L. J. HAYNES AND F. H. NEWTH

R = 2,3,4,6-tetra-O-acetyl-P-D-galactosyl) plus up to 45 % of hepta-0acetylneolactosyl chloride (IX, R = 2,3,4,G-tetra-O-acetyl-P-D-galactosyl) in which the D-glucose residue of the lactose molecule has been isomerixed to D-altrose.‘j 57-60 Similarly, octa-0-acetylcellobiose gives rise to hepta-0acetylceltrobiosyl chloride (IX, R = 2,3,4,6-tetra-O-acetyl-P-D-ghcosyl) as well as to hepta-0-acetylcellobiosyl chloride (VIII) 62 Although treatment of penta-0-acetyl-D-glucose with these reagents gives tetra-0acetyl-D-glucosyl chloride as the main product, D-altrose and D-mannose derivatives are also formed, presumably by a rearrangement similar to that outlined above.63The mechanism and scope of this striking rearraiigemeiit are unknown. With octa-0-acetyllactose, hepta-0-acetylneolactosyl chloride is formed in 4 % yield if only aluminum chloride is used, but the yield is increased to 33-45% in the presence of phosphorus pentachloride. The substitution of thionyl chloride, phosphorus trichloride, and phosphorus oxychloride for phosphorus pentachloride produces yields of 3, IG, and 23 %, respectively. Phosphorus pentachloride, alone, causes no “chlorination.” Hepta-0-acetyllactosyl chloride is not rearranged by the joint action of aluminum and phosphorus chlorides, so that this compound cannot be the primary reaction product.60 When either 1,3,4,5-tetra-O-acetyl-~-fructopyranose or penta-o-acetylketo-D-fructose is treated with a mixture of phosphorus pentachloride and aluminum chloride in acetic anhydride, the product is 1,3,4,5-tetra-0acetyl-P-wfructopyranosyl chloride (“P-acetochloro-D-fructose”). If, however, l ,3,4,5-tetra-O-acetyl-~-fructoseis treated with phosphorus pentachloride in acetic anhydride, without the addition of aluminum chloride, an isomer is formed.64This second product is very much more stable than the first, and B r a ~ n noted s ~ ~ that, during the quantitative estimation of acetyl, only very small amounts of chlorine were split off by alkali a t 0”. Brauns suggested that this compound is an isomeric tetra-0-acetyl-D-fructosyl chloride, and it was considered by Hudson65to be “a-acetochloro1

.‘jl’

(57) (58) (59) (60) (61) (62) (63) (64) (65)

C. S. Hudson and A. Kunz, J . Ani. Chem. SOC., 47, 2052 (1925). A. Kunz and C. 9. Hudson, J . Am. C h e n ~SOC., . 48, 1978 (1926). A. I h n z and C. S. Hudson, J . Am. Chein. SOC.,48, 2435 (1926). N. I<. Richtmyer and C. S. Hudson, J . Am. Chem. SOC.,57, 1716 (1935). C. S. Hudson, J . Am. Chem. SOC.,48, 2002 (1926). N . I<. Richtmyer and C. S. Hudson, J . A m . (‘hem. Soc., 58, 2534 (1936). N. I<. Richtmyrr and C. S. Hudson, unpublished work cited in Ref. 6 , p. 46. D. H. Brauns, J . A m . (Ihem. SOC.,42, 1846 (1920). C. S. Hudson, J . A m . Cheni. SOC., 46, 477 (1924).

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217

fructose." Later work by Pacsu and Rich66showed that the second product does not react with silver acetate in acetic anhydride and, on catalytic reduction, gives a product nhich still contains a stable chlorine atom. This and other evidence suggested that the so-called "a-acetochlorofructose" is not a stereoisomer of tetra-0-acetyl-p-D-fructopyranosyl chloride but a structural isomer, 1,3,4,5-tetra-O-acetyl-6-chloro-6-deoxy-keto-~-fructose, derived from the open-chain form of fructose.* Phosphorus pentachloride and penta-0-acetyl-0-D-glucopyranose, heated together a t 100" for two hours, react to give 3,4,6-tri-O-acetyl-2-O-trichloroacetyl-0-wglucosyl chloride (X).67 68 This very interesting compound is remarkable in that it is a stable derivative of 0-D-glucopyranosyl chloride. The chlorine atom a t C1 is not very reactive; qualitatively, it appears less reactive than the bromine atom in tetra-0- AcOCH acetyl-a-D-glucopyranosyl bromide, and, in fact, the HCOAC I trichloroacetyl group can be removed, leaving this HCO I chlorine atom in place, by careful treatment with a CH~OAC dry, ethereal solution of ammonia. Similar compounds X have been prepared from maltose,69 6-deoxy-~-gluco~ y r a n o s e , ~~-galactopyranose,69~ ~" and ~ - m a n n o p y r a n o s e . ~ ~ ~ This reaction has been studied by Abramovitch,7° who suggested that the phosphorus pentachloride reacts by two mechanisms : first, replacement of the 1-0-acetyl group by chlorine; and second, chlorinatioii of the 2-0-acetyl group, the chlorine arising from the dissociation of the pentachloride, which cannot therefore be successfully replaced by phosphorus trichloride or thionyl chloride. Penta-0-acetyl-P-D-glucopyranose was recovered unchanged after refluxing it for four hours with phosphorus trichloride. Lemieuxg*71 has suggested that the fact that only the C2-acetoxy group is chlorinated can be accommodated in the following reaction sequence involving cyclic, carbonium-ion intermediates. I

q

(66) E. Pacsu and F. V. Rich, J . Am. Chent. SOG.,64, 1697 (1932); 66, 3018 (1933). (67) P. Brig], Hoppe-Seyler's 2. physiol. Chem., 116, 1 (1921). (68) W. J. Hickinbottom, J . Chem. SOC.,1676 (1929). (69) P . Brig1 and P. Mistele, Hoppe-Seyler's 2. physiol. Chem., 126, 120 (1923). (694 E. Hardegger and R. M. Montavon, Helv.C h i m . Acta, 30, 632 (1947). (69b) A. M. Gakhokidze and N . D. Kutidze, Zhur. Obshched Khinz., 22,139 (1952). (69c) A. M. Gakhokidze and N. D. Kutidce, Zhur. Obshche'l Khim., 22,247 (1952). (70) R . A. A. Abramovitch, J. Chem. Soc., 2996 (1951). (71) R. U. Lemieux, Can. J. Chem., 29, 1079 (1951).

218

t. J. HAYNES AND F. H. NERTH

If this mechanism is correct, preparation of compounds of a similar type from any 1,2-trans acetylated sugar should be possible. e. By Treatment with Titanium Tetrachloride (or Bromide) in Chloroform.-The most convenient method for the preparation of poly-o-acetylglycosyl chlorides is that of Pacsu,72 in which the corresponding acetyl compound is refluxed with titanium tetrachloride in chloroform. A deeply colored complex is formed ; this yields the chloride on decomposition witah iced water. The reagent apparently does not cause rearrangements of the type observed with phosphorus pentachloride and aluminum chloride. However, it is an efficient catalyst in the interconversion of a- and p-glycosides (transglycosidation), and, when used to prepare acetylglycosyl halides derived from disaccharides, transglycosidation sometimes occurs. For example, L~ndberg?~ has shown th at treatment of octa-0-acetyl-6-0-(~-~-galactopyranosy1)-0-D-glucopyranose with titanium tetrachloride in chloroform gives a mixture of hepta-0-acetyl chlorides from which, after reaction with mercuric acetate in acetic acid, can be obtained a small yield of octa-0acetylmelibiose [octa-~-acety~-6-~-(a-~-ga~actopyranosy~)-~-~-g~ucopyran ose]. 2,3,4-Tri-O-acetyl-~-rhamnosyl bromide has been prepared from tetra0-acetyl-L-rhamnose by the action of titanium tetrabromide in chl~roform,’~ (72) E. Pacsu, Ber., 61, 1508 (1928). (73) B. Lindberg, Acta Chem. Scand., 6, 340 (1951). (74) G. Zempl6n and A. Gerecs, Ber., 67, 2049 (1934).

GLYCOSYL HALIDES AND THEIR DERIVATIVES

219

but this method appears to offer no advantages over the hydrogen bromideacetic acid method.?6’7 6 . 7 6 a Silicon tetrachloride is completely inactive both in catalyzing the transglycosidation reaction and in the formation of poly-0-acetylglycosyl chlorides. Stannic chloride is a weak catalyst for the transglycosidation reaction and can be used to convert penta-0-acetyl-P-D-glucopyranose into penta-0-acetyl-a-D-glucopyranose ; only by prolonged reaction a t elevated temperature is the a-o-pentaacetate slowly converted to tetra-0-acetyl-a-Dglucopyranosyl ~ h lo rid e.~Sublimed ? ferric chloride has no action on acetylated disaccharides in pure chloroform, but if the chloroform contains a trace of alcohol the corresponding acetylated ethyl glycoside is formed.’? I n a study of the reaction of the anomeric forms of penta-0-acetyl-Dglucopyranose with titanium chloride in chloroform solution, Lemieux and have found that, at 40°,the a-Danomer reacts but slowly, whereas the p-D anomer reacts extremely rapidly, with the initial formation of (unstable) tetra-0-acetyl-P-D-glucopyranosyl chloride which then rearranges relatively slowly to (stable) tetra-0-acetyl-a-D-glucopyranosyl chloride. 2. By Exchange of Halogen

a. Iodides.-Although acetyl- and benzoyl-glycosyl iodides can be prepared by the action of hydrogen iodide in glacial acetic acid on the correanother widely used sponding acetyl or benzoyl corn pound^,^^^ 4 9 , method is that in which the corresponding bromide is treated with a solution of sodium iodide in dry acetone.81Sodium bromide is precipitated, with the formation of the glycosyl iodide derivative. The iodides are unstable and have not been employed in synthetic work. b. Halides of the “p-Series.”-Reaction of tetra-0-acetyl-a-D-glucopyranosyl bromide with freshly prepared “active” silver chloride in an inert solvent for a short time causes replacement of bromine by chlorine, the reaction being accompanied by an inversion, so that the product is tetra-0acetyl-p-D-glucopyranosyl chloride. This method, one of the most important 79v

(75) E. Fischer, M. Bergmann and 4.Rabe, Ber., 63, 2362 (1920). (76) W. N. Haworth, E. L. Hirst and E. J. Miller, J . Chem. Soc., 2469 (1929). (76a) R . S. Tipson, C. C. Christman and P. A . Levene, J . B i d . Chem., 128, 609 (1939). (77) G. Z e m p l h , Z. Bruckner and A. Gerecs, Ber., 62,895 (1929). (78) R . U. Lemicux and Carol Brice, Can. J. Chern., 30, 295 (1952). (79) D. H . Brauns, Bur. Sfandards J . Research, 7, 573 (1931). (80) R. K. Ness, H. G. Fletchrr, Jr., n i i d C. S. Hudson, J . A m . Chem. SOC.,72, 2200 (1950). (81) H. Helfcrich and R. Gootz, Ber., 62, 2791 (1929).

220

L. J. HAYNES AND F. H. NEWTH

for obtaining glycosyl halides of the non-usual configuration, was discovered by Schlubach and his coworkers.l1' 82 It is essential that the silver chloride be freshly prepared and in the active condition described by Schlubach; even then, the reaction is by no means reliable. Use of aged silver chloride gives replacement of bromide by chloride without inversion.83 Both tetra-0-acetyl-a-D-mannopyranosyl bromidelsa' 84 and tri-o-benzoyl8-D-ribopyranosyl bromides5 give chlorides without inversion. These may be true results or may be reflections of the erratic nature of the reaction. Ness, Fletcher and Hudsons5 have shown that tri-0-benzoyl-a-u-ribopyranosyl chloride is formed in small quantity, together with the p-D compound, during the preparation of the latter by the action of titanium tetrachloride in chloroform on tetra-0-benzoyl-p-D-ribopyranose, and found that the a-D compound is relatively stable. Lemieuxg has given a detailed account of a possible mechanism for this reaction. It is suggested that absorption of a 1 ,2-cis-poly-0-acetylglycosyl bromide on the silver chloride facilitates the formation of carbonium ions which are stabilized (immediately they are formed) by assuming a I ,2cyclic structure. Reaction of this cyclic intermediate with chloride ion then yields the 1 ,2-truns-poly-0-acetylglycosyl chloride.

HcFl-k AgC1,

AgC1,BrO

HCOAc

+

HCOAc

It CldH I HCOAc

I

HA0 +

@

I >C-CH

HCO

I

This mechanism implies that 1 ,2-cis-acet ylglycosyl chlorides cannot be formed from acetylglycosyl bromides for which the 1,2-truns anomer is the stable form; this is true of the above mannose and ribose examples. Howards6 has proposed a similar mechanism for reactions of this type, but points out that a 2,3-cis-diacetoxy groupingS6a is also a feature of those sugar derivatives which react without inversion, and suggests that the (82) H. H. Schlubach and R . Gilbert, Ber., 63, 2295 (1930). (83) W. N. Haworth, E. L. Hirst and M. Stncey, J . Chena. Soc., 2864 (1931). (84) F. Micheel and Herthn Micheel, B e r . , 63, 386 (1930). (85) R. K. Ness, H. G . Fletcher, Jr., and C. S. Hudson, J . A m . Cheni. Soc., 73,959 (1951). (86) G. A . Howard, J. Chew. Soc., 1045 (1950). (86a) R. S. Tipson, J . Biol. Cheni., 130, 55 (1939).

GLYCOSYL HALIDES AND THEIR DERIVATIVES

22 1

function of the 3-acetoxyl group is to restrict the movement of the carbonyl group in the 2-acetoxyl group to the neighborhood of the chlorine atom. Presumably, the correctness of this suggestion could be established by a study of the reaction of tetra-0-acetyl-a-D-altropyranosyl bromide with activated silver chloride. The reaction of a glycosyl halide with silver salts has also been applied t o the preparation of fluoro compounds. Helferich and Gooten’ have prepared tetra-0-acetyl-0-D-glucopyranosyl fluoride by reaction of tetra-0acetyl-a-D-glucopyranosyl bromide with silver fluoride in acetonitrile. The successful use of this highly polar solvent, in which tetra-0-acetyl-P-Dglucopyranosyl chloride rapidly rearranges, is possible because of the low reactivity of the glycosyl fluorides. It may be noted here that Zemplh and coworkersn7* have claimed that tetra-0-acetyl-0-D-glucopyranosyl chloride may be prepared from penta-0-acetyl-0-D-glucopyranose by treatment with aluminum chloride in cold chloroform, isolation being effected through treatment with ice and water. They claim that the structure of this compound is best represented as an orthoacetyl chloride, but the evidence for this is not unequivocal.

3. Replacement of the Hydroxyl Group at the Reducing Carbon Atom This method is not widely used for the preparation of poly-0-acylglycosyl halides because the partially acylated sugar required as starting material is usually prepared by hydrolysis of the corresponding halide, itself derived from a fully acylated sugar. However, acetylation of u-fructose under the usual conditions gives 1 3 4,5-tetra-O-acetyl-~-fructopyranose, and this material is used in the preparation of tetra-0-acetyl-0-D-fructopyranosyl chloride. Similarly, benzoylation of D-fructose gives 1,3,4,5-tetra-0benzoyl-D-fructose, which can be converted to 1,3 4,5-tetra-O-benzoyl-PD-fructopyranosyl bromide in good yield by the action of hydrogen bromide in acetic acid.88 2 3 :5 , 6-Di-0-isopropylidene-D-mannosyl chloridess has been prepared from di-0-isopropylidene-n-mannose by the action of phosphorus pentachloride in light petroleum in the presence of anhydrous sodium arbo on ate.^^ Hepta-0-methylcellobiose and tetra-0-methyl-D-glucopyranose have been converted to the corresponding glycosyl chlorides by the same method.35 7

(87) B. Helferich and R. Gootz, Ber., 62, 2505 (1929). (87a) G. ZemplBn, L. Mester and E. Eckhart, Acla Chirn. Acad. Sci. Hung., 4, 73 (1954). (88) R. I<.Ness and 13. G. Fletcher, Jr., J . Am. Chenc. Soc ,76, 2619 (1953). (89) I<. Freudenbcrg and A. Wolf, Ber., 60, 232 (1927). (90) K. Freudenberg, A . Wolf, E. Knopf anti S. II. Zahcer, Ber., 61, 1743 (1928). (91) J . I3. Allison and R. M. Hixon, J . Ant. Chem. SOC.,48, 406 (1926).

222

L. J. HAYNES AND F. H . NEWTH

4. Replacement of Alkoxyl and Related Groups at the Potentially Reducing Carbon Atom a. Of G1ycosides.-Although the glycosidic linkage in disaccharide acetates seems perfectly stable t o the reagents commonly used in the preparation of poly-0-acetylglycosyl halides, the glycosidic linkage in the simple glycosides is quite readily ruptured when the more acidic reagents are used. Zemp16n34 92 has shown that hepta-0-acetylcellobiosyl bromide can be obtained in fairly good yield by the action of hydrogen bromide in glacial acetic acid on a number of alkyl and aryl a- and P-cellobioside acetates, although tetra-0-acetyl-a-n-glucopyranosyl bromide could not be obtained from methyl tetra-0-acetyl-8-D-glucopyranoside. Schlubach, Rauchenberger and which Schultzeg3have found that “penta-O-acetylmonoacetoneglucose,”g4 they formulate as XI, on treatment with liquid hydrogen chloride for some hours a t room temperature, yields a n unstable crystalline AcOC(CH& compound which is probably tetra-0-acetyl-D-glucofuranoI syl chloride. 0 I As mentioned previously, titanium tetrachloride in chloroform is a useful catalyst for the transglycosidation of alkyl glycosides, and replacement of the alkoxyl group by halogen AcOCH is not normally observed. However, methyl tetra-o-acetylI a-D-fructopyranoside on treatment with this reagent yields HCO I tetra-0-acetyl-P-D-fructopyranosyl chloride, and not the exHCOAc and pected methyl tetra-0-acetyl-~-~-fructopyranoside,~~ I CH~OAC the 2,3,4,6-tetra-O-acetyl-fl-~-glucoside of glycolic ester XI yields tetra-0-acetyl-a-D-glucopyranosyl chloride.g6 b. Of Oligosaccharides and Po1ysaccharides.-The glycosidic linkage in the acetates of the reducing disaccharides is reasonably stable t o most of the reagents commonly used to prepare poly-o-acetylglycosyl halides, so that poly-0-acetyl halide derivatives of disaccharides can usually be prepared in good yield by the standard methods. However, if the normal reaction period is prolonged, particularly when the more acidic reagents are employed, the disaccharide is often degraded. Thus, although maltose can be converted into hepta-0-acetylmaltosyl bromide in good yield by the action of acetyl bromide,33when treated with acetic anhydride containing hydrogen bromide it undergoes degradation and the final product is tetra-0-acetyl-a-D-glucopyranosyl 9

(92) (93) (94) (95) (96)

G . Zempl h, BcT.,62, 985 (1929). H. H . Schlubach, W. Rauchenherger and A . Schultze, BBT.,66, 1248 (1933). P. Hrigl and W. Zerrwick, Rer., 66,936 (1933). E. Pacsu and F. B. Cramer, J . k l m . Chem. SOC.,69, 1059 (1937). Thelma M. Reynolds, J . PTOC. Roy. SOC.N . S . Wales, 66, 167 (1932).

GLYCOSYL HAL1I)ElS A N D T’HEfI? DERIVATIVES

223

bromide.”, Ox Similar studies have 1)ccn reported for cellobiosc9g and gentiobiose.lnOConsequently, to ensure obtaining the optimum yield of poly-0-acet,yl-“disaccharyl” halide, it is advisable to follow the rcaction polarin~etrically.98 The glycosidic linkage in the higher oligosaccharides and polysaccharides is less stable than that in disaccharides, and the breakdown of polysaccharides, especially starch and cellulose, to give simple glycosyl halide derivatives has been much investigated in the hope that it might throw some light on polysaccharide st,ruetures. Bergmann, Karrer, Freudenberg, and Hess and their coworkers have all been prominent in this field. The products isolated from starch and cellulose seem t o depend, among other things, on the acidity of the reagents used, the more acidic reagents causing breakdown to D-glucose derivatives, whereas the milder reagents give derivatives of maltose from starch and of cellobiose from cellulose. For example, air-dried, rice starch reacts with acetic anhydride saturated with hydrogen bromide to give tetra-0-acetyl-a-D-glucopyranosyl bromide in more than 85 % yield,97whereas dry, potato starch on reaction with acetyl bromide in the presence of a trace of moisture gives hepta-0-acetylmaltosyl bromide as the main product.’O’! 102 Tri-0-acetylcellulose and tri-0-acetyllichenin are broken down to give hepta-0-acetylcellobiosyl bromide by the 1°3- 104 action of hydrogen bromide in acetic This breakdown of di- and poly-saccharide acetates to poly-o-acetylglycosyl bromides (“acetobrominolysis”) has been studied by Jeanes, Wilham and Hilbert.98 These workers used as the degrading reagent a mixture of equal volumes of acetyl bromide and hydrogen bromide-acetic acid, and allowed the degradation to proceed until there was no further change in the optical rotation of the reaction mixture. It was first shown that tetra-0-acetyl-a-D-glucopyranosyl bromide is unchanged by the reagent and that, from rotational measurements, the yields of tetra-0acetyl-a-D-glucopyranosyl bromide from penta-0-aeetyl-a-D-glucopyranose and tri-0-acetyllevoglucosan are quantitative at 25”. The behavior of diand poly-saccharide acetates under the same conditions was then studied. (97) M. Bergmann and F. Beck, Ber., 64, 1574 (1921). (98) Allene Jeanes, C. A. Wilham arid G. E. Hilbert, J. Am. Chem. SOC.,76, 3667 (1953). See also, Allene Jeanes, C. A. Wilham, R. W. Jones, H. M. Tsuchiya arid C. E. Rist, ibid., 76, 5911 (1953). (99) F. Micheel, Ann., 466, 69 (1927). (100) D. H. Brauns, J. Am. Chem. SOC.,49, 3170 (192i). (101) P. Karrer and C. Nageli, Helv. Chim. Acta, 4,263 (1921); P. Karrer, C. Nageli, 0. Hurwitz and A. Walti, ibid., 4, 678 (1921); 6, 402 (1923). (102) K. Freudenberg and K. Soff, Ber., 69, 1252 (1936). (103) E. Fischer and G. ZemplBn, Ber., 43, 2536 (1910); Ann., 372, 254 (1909). (104) 1’. Karrer and F. Widmer, Helv. Chirn. Acta, 4, 700 (1921).

224

L . J. HAYNES AND F. 13. NEWT13

It was found that the acetates of maltose or of polysaccharides containing only 4 -+ 1-a- or 3 -+ 1-a-D-glucopyranosidic linkages break down, virtually quantitatively, t o tetra-0-acetyl-a-D-glucopyranosyl bromide, and that acetates of 1,6-linked di- and poly-saccharides apparently produce no tetra-0-acetyl-a-D-glucopyranosyl bromide but form substances of low optical rotation and undetermined structure which contain both active and stable bromine. Triacetates of polysaccharides containing both 4 -+ l - a - ~ and 6 + ~ - L Ylinkages -D give final optical rotations of magnitude in direct proportion t o the relative numbers of these linkages, that is, t o the degree of branching in the molecule. This work shows that it is possible to rationalize the results obtained with polysaccharides of D-glucose, and application of the reaction to other polysaccharides should prove of considerable interest. This degradation has been applied by Hess and his coworkers t o give a direct demonstration50a of the 1,4-linkage in starch derivatives. For example tri-0-tosylstarch breaks down, on treatment with hydrogen bromide in acetic acid, to give 4-0-acetyl-2,3,6-tri-O-tosyl-a-~-glucosyl bromide. Treated with sodium iodide in acetone a t 100" for three days, tri-0-tosylstarch gives 6-deoxy-6-iodo-2,3-di-O-tosylstarch, which breaks down, with the above reagent, to give 4-0-acetyl-6-deoxy-6-iodo-2,3-di-0-tosyl-a-~glucosyl bromide. The production of 4-O-acetyl-~-glucosy1bromides in these reactions shows the presence of 1,4-linkages in the original starch derivatives,105-107 Degradation to glycosyl halide derivatives has also been used in structural studies on the methylated polysaccharides. Haworth and Percivallo8 have shown that tri-0-methylamylose is broken down to a maltosyl bromide derivative on treatment with acetyl bromide in chloroform, and, by degradation of the product, demonstrated that the original amylose contained 4 -+ 1 D-glucopyranosidic linkages. Peat and Whetstone1oghave used this reagent in a method for the rapid liberation of the end group in methylated starch. I n these experiments, the glucosyl bromide derivatives were not isolated but were immediately converted t o the corresponding methyl glucosides. However, SchlubachllO has isolated 2,3,6-tri-O-methyl-D-glucopyranosyl chloride by the treatment of tri-0-methylstarch with liquid hydrogen chloride. The reaction of liquid hydrogen chloride with unsubstituted D-glUcOSe (105) (106) (107) (108) (109) (110)

K. Hess, 0. Littmann and R. Pfleger, A n n . , 607, 55 (1933). K. Hess and 0. Littmann, Ber., 67, 465 (1934). K. Hess and W. Eveking, Ber., 67, 1908 (1934). W. N. Haworth and E. G. V. Percivsl, J . Chem. Soc., 1342 (1931). S. Peat and J. Whetstone, J. Cheni. SOC., 276 (1940). H . H. Schlubach, Angew. Chein., 47, 132 (1934).

GLYCOSYL HALIDES AND THEIR DERIVATIVES

225

polysaccharides possibly gives free D-glucosyl chloride as an initial product, but the final product is a p o l y - ~ - g l u c o s a n .Anhydrous ~ ~ ~ - ~ ~ ~ hydrogen fluoride affords a similar final c. Of Isopropylidene Cornpo~nds.~~~--When an isopropylidene derivative of a sugar is treated with hydrogen bromide in acetic acid, it is to be expected that the isopropylidene group will be removed, and if one valence of the isopropylidene group is attached to the oxygen atom a t C1 the product should be a glycosyl halide derivative, the other hydroxyl group liberated becoming acetylated. This mas first shown to be the case by Freudenberg and I v e r P who prepared the crystalline 3-0-benzoyl and 3-0tosyl derivatives50a of 2,4,6-tri-O-acetyl-a-D-glucosyl bromide by the action of hydrogen bromide in acetic acid on the 3-0-benzoyl and 3-0-tosyl derivatives, respectively, of 1, 2 :5,G-di-O-isopropylidene-D-ghcose. In an extension of this work, Ohle and his showed that 6-0-benzoyl1,2-O-isopropylidene-~-glucofuranose gives a sirupy bromo compound which yields methyl 2 , 3 ,4-tri-0-acetyl-6-0-benzoyl-~-~-glucoside on treatment with methanol and silver carbonate. I ,2-O-Isopropylidene-3-0tosyl-D-glucofuranose116also reacts in the expected manner to yield 2 ,4 ,6 tri-0-acetyl-3-0-tosyl-a-~-glucosylbromide. However, I ,2-O-isopropylidene-6-0-tosyl-D-glucofuranose affords an unexpected product, a bromo compound which reacts with methanol in the presence of silver carbonate to give a compound first considered to be methyl 2,3,4-tri-O-acety1-6-0tosyl-/3-~-glucoside,~~~ but later to be different from a compound of this structure synthesized by an unequivocal route.lLsThe structure of the product remains unknown, but, since it is isomeric with the expected methyl glucoside derivative, it may have an orthoester structure. It should be noted that, in nearly all of these reactions, the crude reaction product also contained bromo compounds in which the bromine atom could not be replaced by methoxyl on treating with methanol in presence of silver carbonate, even on boiling. A characteristic play of colors is observed when some, though not all, of these isopropylidene compounds are treated with hydrogen bromide in acetic acid. I n these reactions to give derivatives of D-glucopyranosyl bromide, the original glucofuranose molecule has rearranged to the pyranose form, pre(111) H . H. Schlubach, H. Elsner and I'ilma Prochownick, Angew. Cheni., 46, 245 (1932). (112) K. Fredenhagen and G. Cadenbach, Angew. Chem., 46, 113 (1933). (113) H. H . Schlubach and Vilma Prochownick, Angew. Chem., 47, 132 (1934). (114) 13. Helferirh arid 8 . Bottgrr, Ann., 476, 150 (1929). (115) II. Ohle and K.Spcncker, Ber., 69, 1836 (1926). (116) H. Ohle and H. Erlbarh, Ber., 61, 1870 (1928). (117) H. Ohle and L. von Vargha, Ber., 61, 1203 (1928). (118) B. Helferich, H. Bredereck and A. Schneidmiiller, Ann., 468, 111 (1927).

226

L. J. HAYNES AND F. H . NEWTH

suinably simultaneously with detachment of the isopropylidene groups. Treatment with hydrogen bromide (in glacial acetic acid) of those derivafrom which the formation tives of 1,2-O-isopropy~idene-~-glucofuranose of pyranose derivatives would not be possible should then be a convenient route to derivatives of D-glucofuranosyl bromide; only partial success has been attained. 3 5,G-Tri-O-acetyl-l , 2-O-isopropylidene-~-glucose, on trcatment with hydrogen bromide in glacial acetic acid,11ggives the play of colors mentioned above and, after the reaction mixture has been processed in the usual way, yields a sirupy product which contains reactive and unreactive bromine and in which no trace of tetra-0-acetyl-a-D-glucopyranosyl bromide is detectable. However, when I , 2-0-isopropylidene-3 ,5 G-tri-0tosyl-D-glucose was allowed to stand for eighteen hours with hydrogen bromide in glacial acetic acid, a crystalline solid which exhibited the analysis required by 0-acetyl-tri-0-tosyl-D-glucosyl bromide separated from the reaction mixture in 77 % yield. 5,G-Di-0-acetyl-1 ,2-0-isopropylidene-3-O-tosyl-~-glucosesimilarly gave a D-glUCOfUranOSyl bromide derivative, although in poor yield. 5 , G-Di-0-benzoyl-1 ,2-0-isopropylidene-3-0tosyl-u-glucofuranose afforded a product which crystallized directly from the reaction mixture.*20This product, which was formed quantitatively and in analytically pure state, was extremely reactive and decomposed on attempted recrystallization. Surprisingly, it mas strongly levorotatory, [ a l Z o-~101", and it was therefore considered to be 2-0-acetyl-5,G-di-Obenzoy~-3-0-tosy~-~-~-glucosyl bromide. If the crystals were dissolved in chloroform and the solution was shaken with aqueous sodium bicarbonate solution, the material did not crystallize on evaporation of the chloroform. Ohle and Wilcke suggested that this was because of partial rearrangement to the CY-D form, for, whereas the crystalline product gave only a 5 % yield of the corresponding methyl D-glucofuranoside derivative on treatment with methanol and silver carbonate, the sirupy product from the chloroform solution gave a 30% yield. Results rather different from the above have been obtained by Helferich and his coworkers in a study of the action of hydrogen bromide in acetic acid on methanesulfonates of mono- and di-0-isopropylidene-D-glucofuranose. Treatment of 1 , 2:5,G-di-O-isopropylidene-3-O-methylsulfonylD-glucose with the reagent gives a 25% yield of crystalline 2,G-di-Oacetyl-3-0-mesyl-a-~-glucopyranosyl bromide.121The production of only a di-0-acetyl derivative instead of the expected triacetate is noteworthy. Similar treatment of 5 6-di-O-acetyl-3-O-mesyl-~-glucose gives a 50 % (119) H . Ohle, H. Erlbach and K . Vogl, Ber., 61, 1875 (1928). (120) H. Ohle and H. Wilcke, Ber., 71, 2316 (1938). (121) B. Helferich, H. Dressler and R. Griebel, J. prakt. Chem., 163, 285 (1939).

GLYCOSYL HALIDES AND THEIR DERIVATIVES

227

yield of a crystalline compound which was first formulated as the expected 2 ,5 ,6-tri-O-acetyl-3-0-mesyl-a-~-g~ucosyl bromide. However, later work3’ showed that the compound, on treatment with methanol in pyridine, gives a “methyl glucoside” derivative which possesses an acetyl group stable to alkali, a feature characteristic of carbohydrate orthoacetates.‘ Since the of the bromo compound indicates high positive rotation, [ c Y ] ~ ~191.5”, D that i t is not a @-D derivative, Helferich and Jochinke have suggested that i t should be formulated as 5,6-di-0-acetyl-1 ,2-O-(l-bromoethylidene)3-O-mesyl-~-glucose(XII) . If this view is correct, t,his is only the second compound known which possesses this type of structure, the other being the maltose derivative 7 HCO, ,Br of Freudenberg and Ivers (see Section 11, l b ). 1 ,c, The difference in properties of the two comHCO CH, pounds is remarkable. Freudenberg and Ivers’ CH3S02--OCH I compound is very reactive and unstable, whereas I HCO the D-glucofuranose derivative is quite stable. I HCOAc By reaction with silver acetate it gives an acetyl I derivative from which it can be regenerated by CH~OAC the action of hydrogen bromide in acetic acid, XI1 under conditions in which ordinary orthoacetates are extremely labile. Possibly, similar structures may also apply to some of the tosyl derivatives described by Ohle and his coworkers, and it is obvious that further work is needed in this field. d. Of 0rthoesters.-Carbohydrate alkyl orthoacetates are readily converted to glycosyl halide derivatives on treatment with hydrogen halide in glacial acetic acid or chloroform, and many examples of this reaction, which has been reviewed by Pacsu,l have been reported.97~lZ2-126 The conversion can also be effected by prolonged heating of the orthoester with titanium t e t r a ~ h l o r i d e .IZ2 ~~

+

5. From Glycosans I n early work on 1,6-anhydro-~-~-glucopyranose (levoglucosan), Karrer and Smirnoff lZ7 showed that 2 ,3 ,4-tri-0-acetyl-1 ,6-anhydro-/3-~-glucoseis (122) H.S. Isbell, Bur. Standards J. Research, 7, 1115 (1931). (123) W.W.Pigman and H. S. Isbell, J. Research Natl. Bur. Standards, 19, 189 (1937). (124) C.W.Klingensmith and W. L. Evans, J. Am. Chem. SOC.,60,2559 (1938). (125) Harriet L. Frush and H. S. Isbell, J. Research Natl. Bur. Standards, 27,413 (1941). (126) E.A. Talley, D. D. Reynolds and W. L. Evans, J. A m . Chem. SOC.,66,575 (1943). (127) P.Karrer and A. P. Smirnoff, Helv. Chim. Acta, 6, 124 (1922).

228

L. J. HAYNES AND F. H . NEWTH

converted into 2,3,4-tri-0-acetyl-6-bromo-6-deoxy-c~-~-glucosyl bromide by treatment with liquid hydrogen bromide a t room temperature for three days or by heating with phosphorus pentabroniide a t 100” for fifteen minutes. This latter reaction is probably the most convenient for the preparation of “acetodibromoglucose.” On reaction with hydrogen bromide forms in glacial acetic acid, tri-0-acetyl-1 ,6-anhydro-P-~-glucopyranose tetra-0-acetyl-a-D-glucopyranosyl bromide.lD2lo* This reaction seems fairly yields 6-O-acetylgeneral ; 1,6-anhydro-tri-O-benzoyl-/3-~-glucopyranose 2,3,4-tri-O-benzoyl-a-~-g1ucosyl bromide (isolated as the corresponding methyl D-glucoside)lZx lZ9 and 2,4-di-O-acetyl-l , 6-anhydro-3-0-tosyl-P-~glucose gives a good yield of 2,4,6-tri-0-acety~-3-O-tosy~-a-~-glucosy~ bromide.130 On treatment with titanium tetrachloride in chloroform, tri-O-acetyl-l , 6anhydro-0-D-glucopyranosereacts smoothly to give 2 , 3 ,4-tri-O-acetyla-D-ghCOSyl chloride in good yield.13*Titanium tetrabromide affords the corresponding bromide.132This reaction has been used for the preparation chloride from 2 ,3,4-tri-O-acetyl-l , 6of 2,3,4-tri-O-acetyl-a-~-galactosyl anhydro-/3-~-galactose,’~~ but is not of completely general application. 1,6-Anhydro-tri-O-benzoyl-~-~-glucopyranose does not react with titanium tetrachloride in chloroform, 1,6-anhydro-tri-O-rnethyl-P-u-glucopyranose gives amorphous products containing only small amounts of halogen, and 2 ,3,4-tri-O-acetyl-1 ,6-anhydro-D-mannose does not give tri-0-acetyl-Dmannosyl ch10ride.I~~ The galactosyl and glucosyl halides with an unprotected hydroxyl group a t C6 have been of value in the synthesis of disaccharide~.~

6. By Addition of Halogen and Hydrogen Halide to Unsaturated Compounds The addition of halogens and halogen acids to the glycals has recently been reviewed by H e l f e r i ~ h .Since ~ that review was completed, Davoll and L y t h g ~ e have ‘ ~ ~ re-examined the addition of hydrogen bromide and of hydrogen chloride to tri-0-acetyl-D-glucal (first studied by Fischer, Bergmann and Schotte136) and have found that the product from the reaction bromide or in benzene solution is 3,4,6-tri-O-acetyl-2-deoxy-~-glucosyl chloride, since it condensed with theophylline silver to give, after deacet(128) M. Bergmann and F. K . V. Koch, Ber., 62, 311 (1929). (129) K . Josephson, Ber., 62, 317 (1929). (130) G . ZemplBn, Z. Csuros and S. J. Angyal, Ber., 7 0 , 1848 (1937). (131) G . ZemplBn and Z. Csuros, Ber., 62, 993 (1929). (132) G. Z e m p l h and A. Gerecs, Ber., 64, 1545 (1931). (133) G. Z e m p l h , A. Gerecs and H. Flesch, Ber., 71, 774 (1938). (134) G. ZemplBn, A. Gerecs and Theodora Valatin, Ber., 73, 575 (1940). (135) J. Davoll and B. Lythgoe, J . Chem. Soc., 2526 (1949). (136) E. Fischcr, M. Bergmsnn and H. Schotte, Ber., 63,517 (1920).

229

GLYCOSYL HALIDES AND THEIR DERIVATIVES

ylation, (2-deoxy-~-glucopyranosyl)theophylline. They have also prepared (2-deoxy-~-ribopyranosyl) theophylline by reaction of silver theophylline with the product from the addition of hydrogen chloride to di-0-acetyl-Darabinal in benzene. This method appears more satisfactory than any other yet devised for the preparation of 2-deoxyglycosyl halides. The addition of halogens and halogen acids t o I , 2-glycoseen derivatives has been less studied than the addition to glycals. The products from the addition of halogen are uncrystallizable mixtures of stereoisomers. The 2 (or the coraddition product from 2,3,4,6-tetra-O-acetyl-~-glucoseen-l, responding D-galactose derivative) and chlorine has been used136. in a very interesting synthesis of the mold metabolite kojic a ~ i d . ~ ~ ~ - ~ ~ ~ 111. STRUCTURE OF GLYCOSYL HALIDES That the poly-0-acetylglycosyl halides are sugar acetates in which the acetoxyl group a t the reducing carbon atom has been replaced by halogen was realized a t an early date, the main evidence being their ready conversion into aryl or alkyl glycoside derivatives. Once the ring structures of the methyl glycosides had been established, those of the glycosyl halides followed automatically. The problem of the configuration of the group a t C1 was not so easily solved. t ~name ~ ~ forms of The prefixes a-, p-, and y- were first used by T a i ~ e to D-glucose which had differing rotatory powers; a-glucose had [ O ~ ] ~ O D 106", y-glucose had [ ~ ] * O D +22.5", and these two formsmutarotated to give P-glucose with [a]"D +52.5". It was soon realized that "p-glucose" is a n equilibrium mixture of the a- and y-glucoses, so the name p-D-glucose was given to Tanret's "y-glucose." The two forms (a- and p-) of D-glucose were related to two known isomers of methyl D-glucoside by A r m ~ t r o n g , 'and ~ ~ to two known forms of penta-0-acetyl-D-glucose by Behrend and R ~ t h . '143~ ~ Tanret discovered new forms of several sugars and, in all cases, named the more strongly dextrorotatory form the 01 form. Hudson144pointed out a number of objections to this system of nomenclature and made the pro-

+

8

(136a) M. Grace Blair, Advances in Carbohydrate Chem., 9, 97 (1954). (137) K. Maurer, Ber., 63, 25 (1930). (138) K. Maurer and A. Muller, Ber., 63, 2069 (1930). (139) I<. Maurer and W. Petsch, Ber., 64, 2011 (1931). (140) C. Tanret, Compt. rend., 120, 1060 (1895). For a general account, see W. W. Pigman and R . M. Goepp, Jr., Ref. 10, pp. 42-56. (141) E. F. Armstrong, J . Chem. SOC.,83, 1305 (1903). For an interesting commeutary on Armstrong's work, see C. E. Ballou, S. Roseman and K. P. Link, J . Am. Cheni. SOC.,73, 1140 (1951). (142) R . Behrentl ant1 P. Roth, Ann., 331, 359 (1904); R. 13ehrent1, ibid., 353, 109 (1907). (143) C. S. Hudson a i d J. I<. Dnk, J . Am. Chem. Soc., 37, 1264 (1915). (144) C. S. Hudson, J . Am. Chem. Soc., 31, 66 (1909).

230

L. J. HAYNES AND F. H. NEWTH

posal, subsequently adopted, that “The names of the a- and &forms of the derivative of a sugar should be so selected that the difference of their rotations (a - 0) is equal t o and of the same sign as the difference for the two forms of the similar derivative of that glucose (D- or L-) to which the first sugar is genetically related.” Hudson’s rule could not be directly applied to assigning of a configuration to tetra-0-acetyl-D-glucopyranosyl bromide because the anomeric form was not then known. However, as tetra-0-acetyl-D-glucopyranosyl bromide yields P-D-glucopyranosides or p-D-acetates when the bromine atom is replaced, it was named a /3 compound. Emil Fische+ had recognized that this basis for naming tetra-0acetyl-D-glucopyranosyl bromide as a p-D compound is unsound owing to the possibility of a Walden inversion’s taking place during the replacement, and noted that “it should not be overlooked that the acetylglucosyl halides rotate strongly t o the right in opposite sign to the rotations of the p-glucosides that may be obtained from them.” Nevertheless, the position remained unchanged until 1924, when Hudson published the first paper146 in his series entitled “Relations between Rotatory Power and Structure in the Sugar Group.)) Applying his a d a p t a t i ~ n ’of~ ~Van’t Hoff’s principle of optical superposition, Hudson first showed that the rotation of the (potentially reducing) end asymmetric carbon atom of an O-acetyl-D-glycosyl halide (A,) is an approximately constant quantity for the various aldoses, each halogen (X) giving rise to the respective values: A , = 9,800; - 1 ~ 1= 39,800; ABr = 59,300; A , = 85,400. had prepared a second form of “acetoSome years previously, chloro-D-galactose” (which we now know t o be a furanose derivative148) and, by comparison with the a and /3 forms of the corresponding pentaacetates, showed that this second “acetochloro-D-ga1actose” gave A ~1 = -32,300. This value agrees with the average value for Acl given above, but differs in sign. Since the above galactose compound was of the D-series, the negative sign was not a result of the convention used in naming the sugar derivatives (as is the case with O-acetyl-L-rhamnosyl bromide). The results could be explained on the hypothesis that the molecular rotation of the second acetochloro-D-galactose was not (B’gal ACl), but rather (Blpa1A c l ) . This meant that the second acetochloro-D-galactose had a configuration a t C l opposite to that of the first acetochloro-D-galactose (which gave a positive value for A c l ) . This meant that, if the system of nomenclature proposed by Hudson were adopted, the second acetochloro compound had to be designated a 0-D derivative (since A c l had a negative value) and the

+

(145) (146) (147) (148)

E. Fischer, Ber., 44, 1898 (1911). C. S. Hudson, J . A w . (’hem. Soc., 46, 462 (1924). C. S. Hudson and J. M. Johnson, J . Am. Chem. Soc., 38, 1223 (1916). H. H. Schlubach and Vilmtt Prochownick, Ber., 63, 2298 (1930).

GLYCOSYL HALIDES AND THEIR DERIVATIVES

23 1

first an a-D derivative. If the first acetochloro-u-galactosc were all a compound, it followed that the poly-0-acetyl halides of D-glucose, I)-sylose, lactose, maltose, and cellobiose had to be renamed as a compounds, hecause they also give positive values for A , . The above work classifies sugar derivatives into two groups which have similar rotational properties. Thus in a-u-glucose, methyl a-D-glucoside, and tetra-0-acetyl-a-D-glucopyranosyl bromide, the groups at C1 all have a similar effect on the rotation of the whole molecule, and hence are presumed to have a similar configuration. The next task is to relate these configurations to actual structures. From measurements of the conductivity that of sugars freshly dissolved in boric acid solution, B o e ~ e k e n suggested '~~ a-D-glucose has a cis pair of hydroxyl groups a t C1 and C2, and P-D-glucose a trans pair. Other evidence, and especially the determination of the crystal structure of a-D-glucopyranose by x-ray methods16" has confirmed this view, so that, if the rotational evidence is valid, a-acetobromo-D-glucose may be written in the Haworth formula XIII. This structure is commonly accepted, and there can be little doubt as to its correctness, although no direct chemical proofs have been advanced. It has been found that acetohalogeno sugars which have a cis configuration of the halogen and acetoxyl a t C1 and C2, AcO when written in the form shown react normally with H OAc methanol and silver carbonate to give the correspondXI11 ing methyl glycosides (with inversion of configuration a t C l ) , whereas those with a trans configuration react to give the glycoside plus varying amounts of orthoester derivatives. Insofar as this and similar reactions agree with current ideas as to the mechanism of replacement reaction^,^ they provide some confirmation of the structures given. Nevertheless, some direct proof-and here x-ray evidence would again be especially valuable-of the structures commonly accepted seems desirable. Another numerical relationship involving the optical rotations of the poly-0-acetylglycosyl halides has been deduced by Brauns, who has sum~ n a r i z e d the l ~ ~ results obtained after many years of precise observation on carefully purified materials. Brauns has shown that, in a number of cases, the differences of the specific rotations of the acetohalogeno derivatives of a sugar are directly proportional to the differences in the atomic radii of the halogen atoms. That is, for a given sugar the differences of the specific rotations (acetochloro - acetofluoro), (acetobromo - acetochloro), (ace-

Hq$r

(149) J . Boeseken and H. Couvert, Rec. trav. chim., 40, 354 (1921). (150) T. R . R . Macdonald and C. A. Beevers, Acta Crysf., 6, 654 (1952) (151) D. H . Brauns, Rec. trav. chim., 69, 1175 (1950).

232

L.

J . HAYNER AND F. H. NEWTH

toiodo - acetobromo) are in the ratio 41:17:21. This relationship holds for the poly-0-acetyl halides of D-glucosc, D-arabinose, D-XylOSe, melihiose, arid maltose, but, for D-mannose, the ratio can be expressed either as 25 :17 :21 or as 41 :25 :35. The acetohalogeno derivatives of 4-O-~-glucosylD-mannose, cellobiose, and gentiobiose also give anomalous values. At this point, i t should be noted that the acetohalogeno derivatives of some sugars, notably D-mannose, L-rhamnose, and D-lyxose, give values for -4, which are appreciably different from the approximately constant values quoted above. Since the divergence from the normal value is still relatJively small, it may be argued that any major structural change is unlikely, and, indeed, the chemical reactions of these compounds can be fully accommodated on the basis of the usually accepted structures. A full discussion of Hudson's and Brauns' rules is not within the scope of this article. It should, however, be noted that these rules depend on the initial assumptions that the optical rotation of a poly-0-acetylglycosyl halide may be split into two independent parts under the arbitrary conditions (rotations measured in chloroform solution a t 20") stipulated, and also that the conformations of the pyranose ring in the a! and p forms of the sugars are the same.152-155 There can be no doubt that Hudson's assignment of the stable tetra-0acetyl-D-glucopyranosyl bromide to the a! series is correct, but whether the initial assumption is too broad to make divergences from what are regarded as normal values of A , of any significance remains a matter for further research. As mentioned in the introductory part of Section 11, one anomer of an acetohalogeno sugar is usually much more stable than the other. D-G~UCOpyranose, D-mannopyranose, n-xylopyranose, and D-sorbopyranose give stable a-acetohalogeno derivatives, whereas D-galactofuranose, D-ribopyranose, D-arabinopyranose, and D-fructofuranose give stable p derivatives. At first sight, it appears impossible to predict which will be the stable form for any given acetohalogeno sugar. Closer examination shows that a prediction can be made, using two general rules which may be expressed as follows. (1) I n the stable forms of the polyacetates of the aldohexopyranosyl and aldohexofuranosyl halides, the halogen atom a t C l is trans to the group a t the ring junction when the structural formulas are written according t o the Haworth convention, (XIV) to (XVII). (2) I n the stable forms of the 0-acetylaldopentopyranosyl halides and the (152) (153) (154) (155)

E. Pacsu, J . A m . Chem. SOC.,61, 2669 (1939). C . S. Hudson, J . Am. Chem. SOC.,61, 2972 (1939). 0. Hassel and B. Ottar, Acta Chem. S c a d . , 1, 929 (1947). R . E. Reeves, Advances i n Carbohydrate Cherri., 6, 107 (1951)

233

GLYCOSYL HALIDES AND THEIR DERIVATIVES

XIV

xv

XVI

ACOCH, XVII

0-acetylketohexopyranosyl halides, the halogen atom a t C1 is trans to the acetoxyl group a t C3 when the structural formulas are written according to the Haworth convention, (XVIII) to (XXI). Since both D-allose and D-allomethylose (6-deoxy-~-allose)form stable a-acetohalogeno derivatives,

c~>~ G">: qoy OAc

< oOAc y x ~ 2 ~ ~ c

CHzOAc

XVIII

OAc XIX

OAc

xx

XXI

rule 1must outweigh rule 2. It seems a reasonable prediction that the stable forms of the acetohalogeno-D-aldopentofuranoseswill all be (Y compounds, as are the stable forms of the 0-acetyl-D-aldohexopyranosyl halides. A hypothesis t o explain the observations with the pyranose derivatives, on the basis of theories concerning the preferred conformations of hexose rings, has been advanced by Hassel and Ottarls4and has been discussed by Lernie~x.~ IV. GENERAL The majority of the stable 0-acetylglycosyl halides are beautifully crystalline, colorless compounds, soluble in most organic solvents with the exception of light petroleum, and may usually be crystallized from ether plus light petroleum. The stabilities are in the order fluoride > chloride > bromide > iodide. The fluorides are extremely stable, and may even be deacetylated without loss of fluorine, t o give the corresponding glycosyl fluorides. The iodides, on the other hand, are unstable compounds which, even in favorable cases, decompose at room temperature within two weeks. The bromides show a reasonable balance between reactivity and instability, and have been by far the most widely used halides for synthetic work. Of recent years, the use of poly-0-benzoylglycosyl bromides has become increasingly popular in studies in this group; they are more stable than the acetates but more reactive than poly-0-acetylglycosyl chlorides.

234

L. J. HAYNES AND F. H. NEWTH

Oiily a few poly-0-acetylglycofuranosyl halides have been prepared; of chloride. They these the best known is tetra-0-acetyl-P-D-galactofuranosyl are all much less stable than the corresponding pyranosyl halides. The unstable f o r m of the poly-0-acetylglycosyl halides, that is, those of the “@-series,” have in many cases been obtained crystalline. They are characterized by their extremely easy rearrangement to the stable forms; most of them, with the exception of the fluorides, undergo rearrangement when dissolved in polar solvents. Thus, tetra-0-acetyl-@-D-glucopyranosyl chloride rearranges rapidly, even in pure chloroform. The mechanism of this rearrangement has been discussed by Hassel and Ottar154and by Lemieux .g

v. REACTION MECiX4NISM AND EFFECT OF STRUCTURE ON REACTION RATES The course of the reaction of the poly-0-acylglycosyl halides with nucleophilic reagents has recently received attention. Thus, Lemieuxg has related the theories of the mechanism of replacement reactions to some reactions in carbohydrate chemistry, and has summarized our knowledge concerning the mechanism of reaction of the poly-0-acylglycosyl halides. Hence, only those aspects of the subject which have become clarified by recent work need be discussed here. In this series of sugar derivatives, if the halogen atom is cis t o an 0-acetyl group a t C 2 , reaction with a iiucleophilic reagent results in complete inversion of coilfiguration a t C1. On the other hand, a trans disposition of the halogen with respect to the neighboring acetyl group leads to the formation of a considerable amount of the 1,2-orthoacetate, together with the anomeric a- and @-glycosides.’I n all previous discussions, it has been assumed that solvolysis of the cis-halides follows a unimolecular course (S,l), whereas formation of an orthoester from the trans-halides has been described as involving a bimolecular mechanism (SN2).’9 125, 156 However, kinetic order in a solvolysis gives no indication of the molecularity of the process, and a well-known analysis is necessary for its dete1-minati0n.I~~ Oiily recently have the solvolytic reactions of tetra-0-acetyl-a-D-glucopyranosyl bromide in hydroxylic media been studied with a view to establishing the mechanism of reaction.l58 The stereochemical course of a S N l reaction usually permits racemization, whereas, in the bimolecular process, every act of substitution must lead to an inversion of configuration. I n the alcoholysis of the cis-poly-0-acetylglycosyl halides, complete inversion occurs; this would seem, a t first sight, a case for expecting a SN2mechanism. The reaction has now been established as unimolecular, in the ab(156) Harriet 1,. Frush and H. S. Isbell, J . Research N a t l . B u r . Standards, 36, 111 (1945); 43, 161 (1949). (157) E. D. Hughes, Trans. Paraday SOC.,37, 611 (1941). (158) F. H. Newth and G. 0. Phillips, J. Chern. Soc., 2896 (1953).

GLYCOSYLSHALIDES A N D ~ T H E I R DERIVATIVES

235

sence of a n acid-acceptor. The evidence for this decision lies in the increase in first-order solvolysis rate in more polar mcdia, and in the fact that] thc rate is not affected by the presence of alkali. There is also an indication, from measurements of optical rotation, that racemixation occurs in aqueous acetone. It has been pointed that the high reactivity of the poly-0-acylglycosyl halides is typical of the a-halogeno ethers. I n this class of compound, the inductive polarization of the C-X bond is increased by an electromeric release of electrons from the oxygen atom and this promotes a unimolecular (rather than a bimolecular) reaction. Comparison'6o of the rate of methanolysis of 2,3-dichlorotetrahydropyran(XXII) with those of 3,4,6-tri-O-acety1-2-chloro-2-deoxy-a-~-glucosyl chloride (XXIII) and CHZOAC

XXII

XXIII

XSIV

tetra-0-acetyl-a-D-glucopyranosylbromide showed that the tetrahydropyran derivative reacts 800 to 1000 times as fast as do the two sugar derivatives. Although there will be a neighboring-group effect between the reacting halogen and its neighboring group, this effect is insufficient in magnitude t o explain such a large difference in rate for these compounds (where the lactol oxygen atom can facilitate reaction a t the a-carbon atom). The higher velocity of reaction of the tetrahydropyran derivative must be attributed to the absence of large substituent groups (which impede reaction a t the a-carbon atom in the sugar derivatives). The slower reaction of 2,3-dichlorotetrahydrothiapyran(XXIV), as compared with XXII, illustrates the part played by the hetero atom in the nucleophilic displacement of the halogen, since, as is well known, the electromeric effect of sulfur is less than that of oxygen. Furthermore, compounds in which the halogen is attached t o C2 or C3 of the sugar molecule are completely resistant to alcoholysis. By extending the study outlined above, relation of the solvolytic reactivity of a number of poly-0-acetylglycosyl halides to their structure has been possible.l61 The importance of steric factors was demonstrated by a comparison of reactivity of 3 ,4,6-tri-0-acetyl-2-0-trichloroacetyl-/3-~(159) (a) W. W. Pigman and R. M. Goepp, Jr., Ref. 10, p. 160. (b) C. D. Hurd and R. P. Holysz, J. Am. Chem. Soc., 7 2 , 2005 (1950). (160) F. H. Newth and G. 0. Phillips, J . Chem. SOC.,2900 (1953). (161) F. H. Newth and G. 0. Phillips, J . Chem. SOC.,2904 (1953).

236

L. J. HAYNES AND F. H. NEWTH

glucosyl chloride (XXV) and 3 ,4,6-tri-O-acetyl-P-D-glucosyl chloride (XXVI). Bot,h compounds undergo unimolecular solvolysis, and the methanolysis a t 21" of XXVI is over 20 times as fast as that of XXV. This difference in rate can only be due to a decrease in steric hindrance a t C1 in the latter compound, which lacks the bulky trichloroacetyl group a t C2. The situation of this group in the molecule is such that the possibility that its inductive effect could be transmitted to the Cl-X bond (and in this way be responsible for the difference in reactivity) is unlikely. Included in Table I are the rate constant,s for the methanolysis of the tetra-0-acetyl-a-glycosyl TABLEI Methanolysis Rates for Several Poly-0-acylglycopyranosyl Halides'&' 105 k (sec.7)

Compound

remp.,

Relative k

"C.

~~

3,4,6-Tri-O-acetyl-2-O-trichloroacetyl-~-~-g1ucopyranosy1 1.18 chloride (XXV) chloride (XXVI) 66.6 3,4,6-Tri-O-acetyl-p-~-glucopyranosyl Tri-O-acetyl-a-~-xylopyranosyl bromide (XXX) 50 139 Tetra-0-acetyl-a-u-mannopyranosyl bromide (XXVIII) 30.0 10 12.4 Tetra-0-acetyl-a-n-galactopyranosyl bromide (XXIX) 4.5 Tetra-0-acetyl-a-D-glucopyranosyl bromide (XXVII) 1 2.8 12.1 Hepta-0-acetyl-a-cellobiosyl bromide 11.1 Hepta-0-acetyl-a-gentiobiosyl bromide 29.8

21.2 21.2 21.2 21.2 21.2 21.2 35 35 35

bromides of D-glUCOpyranOSe (XXVII), D-mannopyranose (XXVIII) , D-galactopyranose (XXIX), and D-xylopyranose (XXX). This series may be regarded, with reference to D-glucose, as representing a change in configuration a t C2 and C4, and removal of the 5-C-acetoxymethyl group. The CH~OAC

CHzOAc

H$o-?i

H
H

H

OCOCCI,

H

ACO

xxv CH~OAC H

C

Q cH +~ :

AcO

OAc Br

AcO

H OAc XXVII

H OH XXVI

F

AclCP OAc

Br

Br

H H XXVIII

H

CHZOAC

OAc AcO

AcO

CHzOAc

H

OAc XXIX

H

OAcC H

F Br

AcO

H

OAc

xxx

GLYCOSYL HALIDES AND THEIIl I)EHIVATIVES

235

order of their reactivity is XXX > XXVIII > XXIX > XXVII, and the explanation of this difference must lie in the spatial arrangement of the acetyl groups in relation to C1. Accurate models of the four compounds were constructed on the basis of Reeves' C1 conformation,156and the groups were positioned in such a way that their fractional charges would represent a structure with minimum potential energy. It was found that the steric hindrance or amount of "crowding" of groups a t C1 is inversely proportional to the rate of methanolysis. (The effect of the acetyl group a t C3 could not be evaluated, since the appropriate derivatives of D-allose were not available, but Howards6had suggested that, in a cis-2 ,3-diacetoxy system, the 3-acetoxyl group would restrict movement of the 2-acetoxyl group t o the neighborhood of Cl.) Further support for the view that variation in rate of unimolecular alcoholysis is a function of hindrance a t C1 was given by a study of hepta-0-acetyl-a-cellobiosyl bromide and hepta-0acetyl-a-gentiobiosyl bromide. Their models showed that there should be little difference in rate from that of tetra-0-acetyl-a-D-glucopyranosyl bromide, and this is borne out by their rate constants, given in Table I ; the higher value for the gentiobiose derivative is accounted for by the p-r)-biose linkage. The discussion so far has been concerned primarily with the mechanism of solvolysis of the cis halides. There is no reason to suppose that the trans halides react by a different mechanism, and indeed, tetra-0-acetyl-a-1,mannosyl bromide has been shown to undergo unimolecular solvolysis. On the basis of an SN1mechanism as promoted by the lactol ring oxygen, and of the steric influence of all the groups in the molecule, it should be possible to interpret the reactions of both the cis and the trans halides and to explain the nature of the products from each. The neighboring-group concept of Winstein has been used to explain the difference in reactivity exhibited by anomeric pairs of halides.a6 8K This suggestion is by no means improbable, and would explain the facts that tri-0-benzoyl-P-D-ribopyranosyl bromide is 19 times more reactive a t 20" in 1:9 dioxane-methanol than is the a anomer, and that the rate for the trans-0 chloride shows an 85-fold increase over that of the cis-a chloride. However, in view of the properties of the series XXVII to XXX, steric effects from the whole molecule may be equally important, although the disadvantage of this concept is that hindrance as assessed from models is difficult t o define precisely. It is of interest to compare the rates of methanolysis of tri-0-benzoyl-a-~-xylopyranosyl bromide36and of tri-0-acetyla-D-xylopyranosyl bromideL6'[105k200= 51 (sec-l) in 1:9 dioxane-methanol and 105k210= 139(sec-l) in 100 % methanol, respectively]. Although the value of k for the former compound would he higher a t 21" in 100 % meth-

238

L. J. HAYNES AND F. H . NEWTH

anol, a satisfactory explanation would, in the absence of directly coinparable figures, seem to be in terms of greater hindrance by the benzoyl groups. The extensive work of H. G. Fletcher, Jr., arid the late C. S. Hudson and their c o l l e a g ~ e *Os ~ 8~5 -~8x 162 on the reactions of the poly-0-benzoylglycosyl halides has shown that glycoside formation occurs with the expected inversion when the halogen atom is cis to the benzoyl group a t C2, but proceeds with retention of configuration when the groups are trans. Although the cationic, cyclic intermediate XXXI has been postulated in order to account for the latter reaction, the fact that no 1,2-orthobenzoate in the pyranose series has been isolated (although its presence was sought) may be significant. It was found, however, that the frans halides do give rise to a small amount of cis-glycoside (together with the main product). The manner in which these compounds and the corresponding acetates react now seems clear.

Q7

Q 0-c:?

0-c? \

XXXI

‘CHa

Ph

XXXII

The reactions of both the cis-acetohalogeno sugars and cis-benzoylhalogeno sugars involve inversion of configuration. The initial ionization of the C1-X bond followed by the departure of the halide ion on the same side as the neighboring acyl g o u p which can protect the C1 cation, leaves only the opposite side open for attack by solvent molecules, and so inversion is complete. When, however, the halogen atom is trans to an acetyl group, its departure, which may well be facilitated by this situation, is followed by a competitive, nucleophilic attack from the opposite side by solvent (ROH) molecules and by the oxygen of the polarizable carbonyl group. Thus, both the cyclic cation XXXII and the cis-glycoside can be formed. Compound XX X I I can then itself undergo further unimolecular solvolysis, to give the trans-glycoside or react per se to form the orthoacetate. This mechanism is essentially the same as that formulated by Frush and Isbe11lS6and by Pacsu,l except that the carbonyl oxygen of the neighboring acyl group is not required t o participate directly in an initial bimolecular transition state as pictured by these authors, and it thus explains the formation of some of the cis-glycoside. The failure of 3,4,6-tri-0-acetyl-2-0-trichloroacetyl-/3-D-glucosyl chloride to give a 1,2-0rthotrichloroacetate is impor(162) (a) R . W. Jeanlor, H. G. Fletcher, Jr., and C. S. Hudson, J . Am . Chem. SOC.,70,4055 (1948); (b) R. K. Ness, H. G. Fletcher, Jr., and C. S. Hudson, ibid., 73, 296,3698 (1951); (c) R. K. Ness and H. G. Fletcher, Jr., ibid., 74,5344 (1952).

(:LYCOSYL HALIDES .1ND THEIR DEIilVa\'171VES

239

tant, and may be explained on the grounds that the inductive effect of the trichloromethyl group so diminishes the polarization of the carbonyl group that the nucleophilic activity of the carbonyl oxygen is insufficient to permit formation of the cyclic, orthoester cation. In such a trans system the steric effect of the trichloroacetyl group will still be operative and will permit the formation of both the a- and the p-glycoside.6RAlthough acid strength is satisfactory in explaining the properties of the trichloroacetyl group, it does not account for the difference between acetyl and benzoyl derivatives in the pyranose series, where the latter do not form orthobenzoates. If acid strength were the only consideration, the orthobenzoate structure should be formed as readily as is the orthoacetate. The present picture is further complicated by the recent discovery that hydrolysis of tri-0-benzoyl-Dribofuranosyl bromide gives u-ribofuranose 1,2-(orthobenzoate) 3,5-dibenzoate as a major product of the reaction, and that this compound is converted t o 2 ,3 ,5-tri-O-benzoyl-p-~-ribose through t,he action of pyridine.163 The kinetics of the reactions of poly-0-acylglycosyl bromides with amines With such in acetone have been described in a recent comrnunicati~n.'~~ strongly nucleophilic, secondary amines as piperidine, di-n-butylamine, and diethylamine, biniolecular reactions are undergone by tetra-o-acetyland tri-0-acetyl-Da-D-glucopyranosyl, tetra-0-acetyl-D-galactopyranosyl, xylopyranosyl bromides, and these show second-order kinetics in pure acetone, whereas tetra-0-acetyl-a-D-mannosyl bromide exhibits the incursion of some unimolecular reaction. With the weakly nucleophilic reagents N-methylaniline, pyridine, and 3-picoline1 unimolecular reactions bromide; and, with were observed for tetra-0-acetyl-a-D-mannopyranosyl tetra-0-acetyl-a-D-glucopyranosyl bromide, the reactions pursued a mixed mechanism. The influence of neighboring acyl groups on the rate of reaction appears to be smaller than that observed in the unimolecular solvolytic reactions. 61 VI. REACTIONS OF THE POLY-O-ACYLGLYCOSYL HALIDES I n the preceding Section, the poly-0-acylglycosyl halides have been discussed with particular reference to the effect of their structure on the reactivity of the halogen atom and on the configuration of the product resulting from the unimolecular, nucleophilic substitution. I n the present Section is given a general account of the reactions of the halides which refers more to the type of product which may be obtained from these compounds. (163) R. K. Ness, H. W. Diehl arid H. G. Fletcher, Jr., J . A m . Chem. Soc., 76,763 (1954); R. K. Ness and H. G. Fletcher, Jr., ibid., 76,1663 (1954). (164) N. B. Chapman and W. E. Laird, Chemistry & Industry, 20 (1954).

240

L. J. HAYKSS AND F. H .

Nmvm

1. Formation of Glycosides Synthesis in this very large group of natural products165 has been achieved mainly through the reaction of poly-0-acylglycosyl halides with hydroxylic compounds. Since t’he early formation of the phenolic glycosides, arbutin and methylarbutin, by Michael,166most of the naturally occurring p-1)glycosides have been synthesized. The procedure is straightforward, and when, for example, tetra-0-acetyl-a-D-glucopyraliosyl bromide in an inert solvent is allowed t o react with a hydroxylic compound in the presence of a suitable base, the tetra-O-acetyl-P-D-ghcopyranoside is formed in adequate yield. It is unnecessary to enumerate all the glycosides made in this way, but two examples of interest may be given. I n 1924, amygdalin, one of the earliest-known natural glycosides, was synthesized independently by Haworth, by Zemplh, and by I h h 1 1 I ~and ~ their coworkers by condensing hepta-0-acetyl-a-gentiobiosyl bromide with ethyl m-mandelate in the presence of silver oxide. Resolution of the amide, followed by dehydration and deacetylation, provided marldelonitrile 0-gentiobioside identical with the natural product. Ailot,hcr important application was made in 1932, when Robinson and Todd’6s synthesized the anthocyanins peonin, pelargonin, malvin, and cyanin chlorides. The intermediate 2-(tetra-O-acetyl-P-~glucopyranosy1)phloroglucinaldehyde was obtained by condensing tetra-0acetyl-a-D-glucopyranosyl bromide with phloroClo OCH, glucinaldehyde in acetonitrile containing aqueous alkali, but in acetone the yield was higher HO OAc and less “diglucoside~’was formed. The other OG(OAc)4 intermediate, t,he substituted w-hydroxyacetoOG(0Ac)r phenone D-glucopyranoside, was prepared using XXXIII the Koenigs-Knorr conditions.16g When the two components were condensed in the presence of acid, the acet>ylated 3,5-bis(~-glucopyranoside) (XXXIII) of peonin was formed. The oligosaccharides represent a most important group of glycosides. Synthesis ill this field has been extensive and has been reviewed by Evans, Reynolds and T a l l e ~ . ~ ‘The “ ) formation of bhe p-biose linkage presents no difficulty when the 2-0-acetyl group of the poly-0-acylglycosyl halide is (165) E. F. Armstrong and K. F. Armstrong, “The Glycosides,” Longmans, Green and Company, New York, N . Y., 1931. J. Honeyman, “The Plant Glycosides,” Arnold, London, 1949. (166) A. Michael, Ber., 14, 2097 (1881). (167) R.Campbell and W. N. Haworth, ,I. Chei>t.SOC.,125,1337 (1924). G. Zempl6n and A. Kunx, B e T . , 67, 1357 (1924). R. Kuhn and €1. Sobotka, ibid., 57, 1767 (1924). (168) R. Robinson and A. R . Todd, J. Cherii. SOC.,2290, 2488 (1932). (169) W. Koenigs and 13. Knorr, Ber., 34, 957 (1901).

GLYCOSPL HALIDES AND THEIR DEKIVATIVES

241

cis to the halogen atom, as in tetra-0-acetyl-a-D-glucopyranosyl bromide. There is then no neighboring-group interference which could lead to an orthoester, and the p-glycosidic linkage is smoothly formed. Several refinements have been incorporated in the original Koenigs-Knorr condition^,^(^) among which is the introduction of iodine as a catalyst into the reaction mixture.I7OThis catalytic effect may well be due to an increase in the polarity of the solvent chloroform by the iodine, an effect which would enable the SN1reaction of the halide to proceed more smoothly. With all thenknown refinements, Reynolds and Evans171 described a practical method of obtaining octa-0-acetyl-P-gentiobiose in 74 % yield. A yield comparable to this has been recorded when tetra-0-acetyliu-D-glucopyranosylbromide was allowed to react with the melt obtained by dissolving sodium in molten 1,2,3,4-tetra-O-acetyl-P-~-glucose. Sodium bromide is eliminated, and an 80 % yield of octa-0-acetyl-0-gentiobiose is obtained.172The synthesis of octa-0-acetyl-P-cellobiose was also achieved in this way, using 1,2 , 3 , Gtetra-O-acetyl-@-D-glucopyranose, but the yield was only about 40 %. The direct formation of the a-D-glucosyl linkage has been a subject of study for a number of years. The problem is still of great interest, particularly in connection with the synthesis of maltose, isomaltose, and sucrose. A successful, chemical synthesis of sucrose has been described by Lemieux and H ~ b e r ,who ~ ~ ~heated 3,4,6-tri-O-acetyl-l , 2-anhydro-a-~-glucose and, after (Brigl’s anhydride) with 1,3 ,4,6-tetra-O-acety~-D-fructofuranose acetylating the product, obtained octa-0-acetylsucrose in small yield. Although this long-outstanding objective in sugar chemistry has been achieved, the method employed does not settle unequivocally the a , @ nature of the biose linkage, since the D-fructose derivative was not of a particular configuration and the reactions of Brigl’s anhydride usually lead to the formation of @-I,-glucosides.The problem as t o how a-D-oligosaccharides may be formed through the use of the 0-acylglycosyl halides remains. In view of the discussion in the previous Section, it would seem that the formation of an a-D-glycosyl linkage by employing the &halide must be difficult, since orthoester formation will be concomitant in the reaction. Indeed, in the reaction product of tetra-0-acetyl-a-u-mannopyranosyl bromide with 1 , 2 , 3,4-tetra-O-acetyl-@-~-glucose,Reynolds, Evans and Talley126found two isomeric disaccharide orthoesters. The normal biose, of undefined configuration, was also formed and its yield was greater in the presence of iodine. The compound likely t o be of greatest use in the forma(170) B. Helferich, E. Bohn and S. Winkler, Ber., 63, 989 (1030). (171) D. D. Reynolds and W. 12. Evans, J . A m . Chem. SOC.,60,2559 (1938). (172) Violet E. Gilbert, F. Smith and M. Stacey, J. Cliem. Soc., 622 (1946). (173) R. U. Lemieux and G. Huber, Abstracts Papers Am. CItcni. SOC.,134, 181) (1953); J. A w . Chem. Soc., 76,1118 (1953).

242

L . J. HAPNES AND F. H. NEWTH

tion of a-D-glucosyl disaccharides is Brigl's 3 ,4 ,6-tri-O-acetyl-@-~-glucosyl ~hloride.6~6* I t s reactivity with nucleophilic reagents is much higher than that of many other halidePo (see Table I), and the absence of a 2-0-acyl group obviates any danger of forming a 1 ,2-orthoester during the reaction of the @-halogenatom. Inversion of configuration would give primarily the a-u isomer, although a certain amount of @-D form would be produced because of the unimolecular character of the reaction. Further investigation may well show that condensation of a poly-0-acylglycosyl halide with a hydroxylic molecule proceeds more readily in a polar solvent (such as acetonitrile or dimethylformamide) than in the conventional chloroform. was The synthesis of melibiose [6-0-(a-~-galactopyranosyl)-~-glucose] ~ ~ quinoline as a condescribed in 1928 by Helferich and B r e d e r e ~ k . 'Using densing agent in the reaction between tetra-0-acetyl-a-D-galactopyranosyl bromide and 1,2 ,3 ,4-tetra-O-acetyl-@-~-glucose,they were able to isolate a small amount of octa-0-acetylmelibiose. I n the presence of silver oxide, only the 0-Dlinkage was formed.I76 The reaction of the halides with nitrogen compounds is discussed later in this Section, but a possible function of the quinoline may be mentioned here. Reaction of the a-D-galactosyl bromide with quinoline could give a quaternary ion having the 0-D configuration, and reaction of this with the 6-hydroxyl group of the tetra-0-acetyln-glucose would then give the a-D-biose linkage. I n view of the small yield, however, the full course of the reaction is likely to be more complicated. The difficulties encountered in the synthesis of a-Dlinked oligosaccharides are not so apparent in the formation of a-D-glycosides from simple alcohols. Zemplen and his coworkers176examined the reaction of hepta-0-acetyl-acellobiosyl bromide with methanol, and found that, in the presence of mercuric acetate, both methyl a- and P-cellobioside are formed. Solvolysis by alcohol in the presence of mercuric acetate gives mainly the @ form, whereas with a molar quantity of alcohol in benzene the a form is obtained. With silver salts, only the /3 form is obtained. Then, Helferich and Wedem e ~ e r "examined ~ the effect of several metallic oxides and salts on the methanolysis of tetra-0-acetyl-a-D-glucopyranosylbromide. They found that although the yield of methyl tetra-0-acetyl-P-D-glucopyranoside is lorn in the presence of zinc, cadmium, and mercuric oxides, much higher yields of glucoside result with zinc acetate or mercuric cyanide. A comparably high yield of benzyl tetra-0-acetyl-@-D-glucopyranoside was ob(174) B. Helferich and H. Bredereck, A m . , 466, 166 (1928). (175) B.Helferich and H . Rsuch, Ber., 69,2655 (1926). R . Hclferich and G . Sparniberg, ibid., 66, 806 (1933). (176) G.ZemplBn, Fortschr. Chem. org. Naturstoffe, 1, 1 (1938). This review covers work carried out between 1929 and 1932. (177) J3. Helfcrich and K. F. Wedemeyer, A n n . , 663, 139 (1949).

GLYCOSYL HALIDES AND T H E I R DERIVATIVES

243

tained in the presence of zinc oxide. The homogeneous reaction in the presence of aryl mercuric acetate gave only P - ~ - g h c o s i d e sand ~ ~ the ~ kinetic analysis is c~mplicated.’~~ The factors governing this reaction (and all those which are heterogeneous) are by no means yet understood, since surface effects and complex formation make interpretation difficult. 2. Formation of Glycosylamines ( “ N - G l y c o ~ i d e s ” ) ~ ~ ~ ~

Xucleophilic replacement of the halogen atom in the poly-0-acylglycosyl halides occurs with nitrogenous compounds and a “N-glycoside” is formed. In general, the stereochemistry follows the same course as in the formation of glycosides, and inversion occurs. Different types of product may, however, be obtained, depending on whether the amine is primary, secondary, or tertiary. Furthermore, as was pointed out in the preceding Section, the reactions are non-solvolytic. There is evidence that they are b i m o l e ~ u l a r , ~ ~ ~ arid the finer steric considerations will not necessarily be the same as those which have been evaluated for the solvolytic reactions. An example of the formation of a “N-glycoside” is the reaction of tetra-0acetyl-a-D-galactopyranosyl bromide with aniline, when tetra-0-acetyl-P1)-galactopyranosylanilineresults in good yield With secondary amines, dehydrohalogenation occurs, and a 1,2-glycoseen is obtained.181Quaternization is the first step in the reaction with tertiary amines, and treatment of the quaternary ammonium salt with barium hydroxide leads to the formation of a 1,6-anhydro ring. In this way, 1,6-anhydro-P-~-galactohave been prepared. The pyranosela2and 1,Ci-anhydro-P-~-glucopyranose~~~ formation of 3,4,6-tri-O-acetyl-l,2-anhydro-a-D-glucose by treating 3 ,4 , ti-tri-0-acetyl-P-D-glucosyl chloride with ammonia in benzene solution may also be mentioned.67 An important use of the poly-0-acylglycosyl halides in synthesis of “iV-glycosides” has been in their coupling with heterocyclic bases to form the naturally occurring nucleosides. Early synthetic experiments on the formation of pyrimidine riucleosides by treating a diethoxypyrimidine with a poly-0-acetylglycosyl halide, to give the analogs of cytidine and uridine, have been reviewed by LythgoelB4;by using tri-0-acetyl-a-u-ribofuranosyl hromide, this method has led to the successful synthesis of uridine and cytidine.15 The thymine nucleosides have been synthesized in a similar (178) B. Helferich and I<. F. Wedemeyer, (’hem. Ber., 83, 538 (1950). (179) K. F. Wedemeyer and W. Haus, Cheni. Ber., 83, 541 (1950). (1798) See also, G. 1’. Ellis and J. Honeyman, this volume, p. 95. (180) K. Butler, F. Smith and M. Stacey, J. Chew. Soc., 3371 (1949). (181) K. Maurer, Ber., 62, 332 (1929). (182) F. Micheel, Ber., 62, 687 (1929). (183) P. Karrer and A. P. Smirnoff, Helv. Chim. Acta, 4, 817 (1921). (184) B. Lythgoe, Ann. ZZepts. on Progr. Chem. (Chem. SOC.London), 41,200 (1944).

244

L . J. HAYNES AND F. H. NEWTH

way,185and Kenner1X6has reviewed advances in the field of synthesis of nucleosides, some of which are the work of Todd and his collaborators. I n general, the purines and pyrimidines are weak bases, and it is necessary to employ the silver salt in order to obtain the required “N-glycoside.” One of the earliest examples of this use was given by Fischer and Helferich,ls7 who condensed tetra-0-acetyl-a-D-glucopyranosyl bromide with the silver salt of 2,8-dichloroadenine. They obtained an adenine or guanine glucoside, and it was subsequently shown that the alkylation occurs a t CO and not C7 of the purine. Syntheses of both adenosine and guanosine have been accomplished in this way, using tri-O-acetyl-a-D-ribofuranosyl bromide,24 and the D-xylofuranosyl and D-arabinofuranosyl derivatives of theophylline and adenine have been obtained similarly.1x8The use of the silver salt of adenine itself in such reactions is not satisfactory, since the purine is sufficiently basic t o remove hydrogen halide from the sugar derivative by q u a t e r n i ~ a t i o n ,but ~ ~ ~this difficulty was somewhat overcome by nullifying the basic character of the amino group by acylation. A further modification was the use of the chloromercuric salt of the purine, which is more satisfactory than the silver salt since the reaction mixture is then homogeneous. I n this way, adenosine, guanosine, and crotonoside have been synthesized,lgOand the reaction has been applied to the preparation of some benzimidazole “nucleosides.”lgl The discovery‘ of 5,g-dimethyl-l(a-D-ribofuranosy1)benzimidazole as a degradation product of vitamin BIZ has stimulated the investigation of glycosylbenzimidazoles. The reaction between the silver salt of 5,B-dimethylbenzimidazoleand tetra-o-acetyla-D-ghCOpyranOSylbromide gave only a very small yield of the 8-D “N-glu~ o s i d e . Further ’ ~ ~ ~ ~ study confirmed Davoll and Lowy’s finding that the chloromercuric derivatives are more satisfactory, but only the p-D isomer was formed. However, condensation of (excess) free benzimidazole base with the poly-0-acetylglycosyl halides gives a mixture of a and p anomers which are separable as their picrates, and 5,6-dimethyl-l-(a-~-ribofuranosyl)benzimidazole was synthesized by this method.lg3 The formation of a- and p-D “N-glycosides” was also observed by Davoll and L y t h g ~ e , ’ ~ ~ (185) D . W. Visser, I. Goodman and IZ.Dittmer, J . A m . Chem. Soc., 70,1926 (1948). N. Z.Newmark, I. Goodman and I<. Dittmer, ibid., 71.3847 (1949). (186) G. W. Kenner, Forfschr. Chem. org. Naturstofle, 8. 96 (1951). (187) E. Fischer and B. Helferich, Ber., 47, 210 (1914). (188) P. Chang and R . Lythgoe, J , C‘herri. Soc., 1992 (1950). N . W. Bristow anti €3. Lythgoe, ibid., 2306 (1949). (189) J. Davoll and B. A. Lowy, J . Am. Cherri. Soc., 73, 1650 (1951). (190) J. Davoll and B. A. Loay, J . Am. Chem. Soc., 73, 5174 (1951). (191) J. Davoll and G. B. Brown, J . Am. Chem. Soc., 73, 5781 (1951). (192) J. G. Buchanan, A. W. Johnson, J. A. Mills and A. R. Todd, J. Chen7. Soc., . 2845 (1950). (193) A. W. Johnson, G . W. Miller, J. A. Mills and A . R. Todd, J. Chem. Soc., 3061 (1953).

245

GLYCOSYL HALIDES AND THEIR DERIVATIVES

who found that condensation of 3 ,4-di-0-acetyl-2-deoxy-~-ribopyranosyl chloride with silver theophylline gives a mixture of 7-(3 4-di-O-acetyl-2deoxy-a ,0-u-ribopyranosyl) theophyllines. Finally, mention must be made of the nicotinainide “nucleosides.” Fischer and Raskelg4 observed the quaternization of tetra-0-acetyl-a-Dglucopyranosyl bromide with pyridine, and Karrer and coworkers extended this reaction to the synthesis of quaternary, nicotinamide n ~ c l e o s i d e s . ~ ~ ~ The condensation has been improved by Haynes and Todd,lg6who used acetonitrile as solvent. With tri-0-acetyl-a-u-ribofuranosyl bromide, n-ribofuranosylnicotinamide-a structural unit of coenzyme I (DPN)-was obtained, and this was reduced with sodium dithionite to the dihydro derivative. )

3. Miscellaneous Reactions

A number of important reactions of the poly-0-acylglycosyl halides have heen reviewed already in Advances in Carbohydrate Chemistry and there is no need to discuss further the formation of glycals5and of thio- and selenosugars2,lg7 or the Grignard processes.66 The reduction of poly-0-acylglycosyl halides with lithium aluminum hydride has been studied. The lactol ring remains intact, and halogen is replaced by hydrogen. Deacylation simultaneously occurs, and this method of forming anhydroglycitols is more convenient than that of the reductive desulfurization of I-deoxy-1-thioglycosides or of glycosyl x a ~ ith a te s .’~ ~ Treatment of tetra-0-acetyl-a-D-glucopyranosyl bromide with lithium aluminum hydride gives 1,5-aiihydro-~-glucitol(polygalitol) , and 1 5-anhydro-D-mannitol (styracitol) is formed from tetra-0-acetyl-a-u-mannopyranosyl bromide.lg8 Tetra-0-acetyl-u-galactofuranosyl chloride is similarly reduced t o l,4-anhydro-u-galactitol.1gg Reduction of tetra-0-benzoyl-@-Dfructopyranosyl bromidc with lithium aluminum hydride gives a very small amount of 1,5-anhydro-u-mannitol, and the major product of the reaction is 1 5-anhydro-L-gulitol, the latter being produced by inversion a t C2 on replacement of the bromine by hydrogen.200 )

(194) E. Fischer and I<. ltaske, Rer , 43, 1750 (1910). (195) P. Karrer, €3. H. Ringier, J. Biichi, H. Fritzsche and U. V. Solmsscn, Helv. Chin?. Acta, 20, 55 (1937). (196) L. J. Haynes and A. R. Todd, J . hen^. SOC.,303 (1950). (197) H. G. Fletcher, Jr., and N. N. Richtmyer, Advances in Carbohydrate Chew., b, 1 (1950). (198) R . K. Ness, H. G. Fletcher, J r . , and C. S. Hudson, J . Am. Pheni. Soc., 72, 4547 (1950). (199) R. N. Ness, H. G. Fletcher, Jr., and C. S. Hudson, J . Am. Cheni. Soc., 73, 3742 (1951). (200) R. K. Ness and H. G. Fletcher, Jr., J . Am. (’hem. SOC.,76, 2610 (1953).

246

L. J. HAYNES .4ND F. H. NEWTH

VII.

TABLES OF PROPERTIES OF SOME

GLYCOSYL HALIDEDERIVATIVES”

TABLE I1 Properties of Some Glycosyl Fluorides Fluoride

I n water.

Temp.,

Melting poinl, “C.

Monosneeharides P-L-Arabinopyranosyl, tri-0-ncetyl117-118 a-D-Fructopyranosyl 1,3,4,5-tetra-O-acetyl112 3,4,5-tri-O-acetyl134-135 1,4,5-tri-O-acetyl -3-0-methyl 113.4 3 , -I 5-tri-0-acetyl-1-0-methyl, 94 a-D-(flucopyranosyl 118-125 2,3,4,6-tetra-O-acetylI08 2,3,4-tri-O-acetyl-6-chloro-6-deoxy- 151-152 2,3,4-tri-O-acetyl-6-O-trit4.1147-148 2,3,4,6-tetra-O-benzoyl110-112 2,3,4-tri-O-benzoyl-6-0-t ritylnmorph. 6-chloro-6-deosy138 (dec.) 6-0-trityl140 P-~-Glucopyranosyl,tetra-o-acetyl98 a-n-Mannopyrnnosyl, tetra-0-acetyl68-69 a-r)-Sylopyranosyl, tri-0-acetyl87 Disacehnrides Cellobiosyl, hepta-0-acetyl187 Epicellobiosyl, hepta-0-acetyl155-156 Gentiobiosyl 215-220 (dec.) heptn-0-acetyl168-169 2’, 3’, 4’, 6’-Cetra-O-acetyl-2,3,4-tri O-benzoy195-196 4-0-(8-D-Glucopyranosyl) -a-o-maIinosy1 hepta-0-acetyl155-156 3,6,2’, 3’,4’,6’-hcsn-O-acetyl145 Lactosyl 180-195 (drc.) hepta-0-ncetylnmorph. 174-175 Maltosyl, hepta-0-acetylMelibiosyl, hepta-0-acetyl135

“C.

f138.2

20

-90.4 -128.8 -88.7 -116.3 +96. ia

20 20 20 20 18 20 20 20 22 18 20 14

References

49

20 20

14, 22 22 22 22 201, 202 14 203 201 20 1 201 203 201 87, 204 79 14

f30.6 +13.6 +33.5“ +43.8

20 20 20 20

14 21 201 100

+I5

20

201

$13.6

20 20 15

21 21 87 87 205 205

f90.1

+lo7 +ll9.6 1lo* +75.1b +88. So +58. 4b +21.9 f21.5 +67.2

+

+20.7 +83. 2a +111.1 +149.7

18

-

20 20

I n pyridine.

* In the following Tables, the rotation solvent. employed WRS chloroform, except where otherwise indicated. (201) B. Helferich, K. Bauerlein and F. Wiegand, Ann., 447, 32 (1926). (202) B. Helferich and 0. Peters, A n n . , 494, 101 (1932). (203) B. Helferich and H. Bredereck, Ber., 60, 1995 (1927). (204) Violet E. Sharp and M. Stacey, J . Chem. Soc., 285 (1951). (205) I). H . Brauns, J . Am. Chetti. Soc., 61, 1820 (1929).

247

GLYCOSYL H A L I D E S AND THEIR DERIVATIVES

TABLE 111 Properties of Some Glycosyl Chlorides Temp

"C.

a-D-Altropyranosyl, tetra-O-acetyl P-D-Arabinopyranosyl, tri-O-acet,yl P - L - Arabinopyranosyl 2,3,4-tri-O-acetyl3 , 4-di-O-acety1-2-chloro-2-~eo~y~ L-gZ~ce~o-r~-gaZa-Heptopyranosyl, 2,3,4,6-tetra-O-acet,\ll-7-d~oxyu-Ribopyranosyl, 3,4-di-O-acct yI -

2-deoxy8-1~-Fructopyranosyl, tetrs-0:1cetylI)-Fructofuranosyl,tetra-O-aceiyl8-i)-Galaetofuranosyl, tet.ra-0:tcctyla-D-Galactopyranosyl 2,3,4,6-tetra-O-acet?-I2,3,4-tri-O-acetyl3,1,6-tri-O-acetyl-2-cilloro-3deoxy2,3,4-t~ri-O-acetyl-6-0-tos~l-

Rejerences

101-102 151-152

+110 -244

20 20

206 26, 48

146-147 100-101

$244.4 f166

20 18

208

106-107

-208.3

20

209

sirup

-

83 sirup

-160.9 -

20

67 (dec.) 75-76;82-83 132

-79.1 +212.3 +207.8

20 20 21

147, 148 211 19, 55 133

105 120 108

+188.7 +134.5 +136

19 19

208 212

20"

213

+5.8

20

82

-79.5

I9

!14

+46.5 -113.5

20 19

93 !15

amorph.

2,4,6-tri-O-methyl-3-O-tosg/l8-u-G:tlactopyranosy1, t.ct.ra-0ncc t.yl93-94 (dec.) 13-1) -Galactosept a n osy 1, 2,3 -1 , 5 tetra-0-acety sirup ~)-Glucofuranosyl 2,3,5,6-tetra-O-acctyl5-0-benzoyl-2,3,6-tri-O-niet hy I 122-123 a-~-Glucofuranosyluronic acid 2,5-di-O-acetyl-, 6,3-lactonr 107.5-108.5 methyl ester 150.5-151.5 a-D-Glucopyranosyl 2,3,4,6-tetra-O-acetyl75-76 2,3,4-tri-O-acetyl121-125 2,3,4-tri-O-acetyl-6-c hloro-tideoxy156 3 , 4 , 6-tri-O-acetyl-2-chloro-2deoxy02-94 3,4,6-tri-O-acety1-2-0-tos~l121-122 4-0-acetyl-2,3,6-tri-O-met hylb.p., 143-148/0.04 mm. 4-0-acetyI-2,3,6-tri-O-tosyl 173-174

-

-

49, 207

135 64 210

~

t166.2 +191.5

10

19

116, 217 !17,218 !19 72, 220 31

tl96.8

! 0

!03

200b t134.8 t146.9

+-

.5 !O

36 121 122

+80.7

!O

123

+95.5 -16.7 -

22 11 -

17

248

L. J. HAYNES AND F. H . NEWTH

TABLEI11 (Continued) Chloride

-

Meltind point, "C.,

Monosaccharides (continued) a-u-Glucopyranosyl (continued) 2,3,4,6-tetra-O-benzoyl116-118 109 2,3,4,6-tetra-O-mesyl168-169 +110" 2,3,4,6-tetra-O-methyIsirup en. +154 2,3,6-tri-O-methyl2,3,4,6-tetra-O-propionylsirup +130.6 2,3,4,6-tetra-O-tosyl+61.9 78-80 2,3,4,6-tetrasulfate 8-u-Glucopyranosyl 2,3,4,6-tetra-O-acetyl98-99 -18.6d 3,4,6-tri-O-acetyl158 (dec.) +19.1c 3,4,6-tri-O-acetyl-2-0-trichloroacetyl142 +3 .Oe

+

"emp., "C.

References

20 20 20 25 20 -

80 224 30, 225 110, 226 227 50 228, 229

23 20

11, 2, 6 67, 230

14

67

20

83

20

83

21

89

20 20

72 80

a-D-g~ycero-D-gu~o-Heptopyraiiosy~,

2,3,4,6,7-pcnta-O-acetyl-

97

+95

0-D-glycero-D-gulo-Heptopyranosyl,

2,3,4,6,7-penta-O-acetyl125 +I1 a-D-Mannofuranosyl,2,3:5,6-di-0isopropylideneb.p.,119/0.1 mm +85.7 a-a-Mannopyranosyl 2,3,4,6-tetra-O-acetyl81 +90.6 2,3,4,6-tetra-O-benzoylamorph. -30.5 a-L-Rhamnopyrnnosyl 2,3,4-tri-O-acetgl73 - 127 2,3,4-tri-O-benzoj-l165-166 +136 a-D-Ribofuranosyl, tri-0-acetylsirup cu. +40 a-u-Ribopyranosyl, tri-0-henzoyl203-204 +60 P-o-Ribopyranosyl 2,3,4tri-0-acetyl95 -169.6 2,3,4-tri-O-beuzoyl162-163 - 147 a-L-Sorbopyranosyl, 1,3,4,5-tetra O-acetyl-83.3 67 a-o-Sylopyranosyl 2,3,4-tri-O-acetj.l+171.2 105 3,l-di-O-benzoyl-2-chloro-2deoxy178-180 +I10 @-D-~~1Opy1'anOSyI, tri-0-acctvl1 12-113 -131.0" wGalactopyranosy1 3,4,6-tri-O-acetyl159-167 +29.11' 3,4,6-tri-0-acetyl-2-O-trichloroacetyl150-156 +10.4 a-u-Glucopyranosyl 3,4-di-O-acetyI-6-deosy119 +222 2,3,4-tri-O-acetyl-6-deoxy136 +204

20 20 22 20

207 162(b) 24, 25 85

22 20

25 85

20

152

20

220

232 20 23.5 82 20

6911

20

6%

-

69a

-

69:1

249

GLTCOSYL HALIDES AND THEIR DERIVATIVES

TABLEI11 (Continued) Chloride

Tern?. “C.

Yelling Point, “C.,

ReJerences

. Monosaccharides (continued: B-i)-Glucopyranosyl 3,4-di-O-acety1-6-deoxyRt4-di-0-acetyl-6-deoxg2 O-trichloroacetylI ) -hInnnopyranosyl 3 ,4,6-tri-O-acetyl3,4,6-tri-O-acetyl-2-0-trichloroacet ~ 1 -

-

ca. 68

-

69a

-

69a

148

$7

151-152

+17.1

20

69c

134-136

+11.7

20

69c

Di-and Oligo-saccharides a-Crllobiosyl hept a-o-acptylhepta-0-methyl6-0-(/3-Cellobiosyl)-a-~-g~u~opyranosyl, deca-0-acetyla-Celtrobiosyl, hepta-0-acetyla-Epicellobiosyl, hepta-0-acetyl a-Grntiobiosyl, hepta-0-acetyl6-0- (/3-o-Glucopyranosyl)-a-o-glucopyranosyl, hepta-0-acetyl4-O-(B-o-Glucopyranosyl) -a-D-mannopyranosyl, hepta-0-acetpla-Lactosyl, hepta-0-acetyla-Maltosyl, hepta-0-acetyli3-Maltosy1, hexa-0-acetyl-2-0-tri . chloroacetyla-lllelibiosyl, hepta-0-acctyla-Neolsctosyl, hepta-0-acet yla-Primeverosyl, hexa-0-acetyl6-O-(B-~-Rhamnopyranosyl)-a-w galactopyranosyl, hexa-0acetyla-Robinobiosyl, hexa-0-acetyla-Rutinosyl, hexa-0-acetyl@-Turanosyl,hepta-0-acetylI n acetone. bon tetrachloride. a

(206) (207) (208) (209)

b

200-201 sirup 223-224 (dec.)

+73.0 +48.4

20

141-142 172 142-143

+64.2 +51.2 +89.2

20 20 20

136.5-137

+82.8

18

132

172 120-121 I25

+51.1 +83 .!) +159.5

20 20 20

21 57, 72 205

132-133

20

127 182 (dec.) 190-192

+58.7 tc +80e +192.5 +71.2 +70.8

20 25 20

205 6, 58 233

166.5-167.5 180 (dec.) 150.5-151.5 165

+67.6 -5.05 f65.9 -0.44

21 26 20 20

133 232 133 235, 236

I n acetylene tetrachloride. 8 I n benzene.

-

18

-

I n ethyl acetate.

21 35 132 62 21 72, 100, 132

69

I n car-

N. K. Richtmyer and C . s. Hudson, J . Am. Chem. Soc., 63, 1727 (1941). H. Ohle, W. Marecek and W. Bourjau, Ber., 62, 833 (1929). A. M. Gakhokidze, Zhur. Obshchet Khim., 10, 507 (1940). E. L. Jackson and C. S. Hudson, J . Am. Cheni. SOC.,76, 3000 (1953).

250

L . J. HAYNES AND F. H . NEWTH

REFERENCES FOR TABLE 111 (CoritirLued) (210) J. C. Irvine, J . W. H . Oldham and A. F. Skinner, J. A m . Cheiri. Soe., 61, 1279 (1929). (211) A. Pictet and If. Vel.net, Helv. C h i m . Aela, 6 , 444 (1922). . (212) H . Ohle and H. Thiel, Ber., 66, 525 (1933). (213) Elizabeth E . Percival and E. G. V. Percival, J. Chenz. Soc., 1585 (19.78). (214) F. Mieheel and F. Suckfull, Ann., 607, 138 (1933). (215) K. Hess and F. Micheel, Ann., 466, 100 (1928). (216) W. F. Goebel and F. H. Babers, J . Biol. Chem., 101, 173 (1933). (217) W. F. Goebel and F. H . Babers, J. Biol. Chenz., 106, 63 (1934). 74,5605 (1952). (218) K.-C. Tsou and A. M. Seligman, J . -4ni. C h e m . SOC., (219) A. Pictet and P. Castan, Helv. Chim. A d a , 4,319 (1921). (220) D. H. Brauns, J . Am. Chewt. SOC.,47, 1280 (1925). (221) Thelma M. Reynolds, J . Chem. SOC.,2626 (1931). (222) F. Micheel and K. Hess, Ber., 60, 1898 (1927). (223) K. Hess and L. Kinze, Ber., 70, 1139 (1937). (224) B. Helferich and A. Gniichtel, Ber., 71, 712 (1938). 93, 105 (1908). (225) J . C. Irvine and Agnes M. Moodie, J . (Them. SOC., (226) K. Freudenberg and E. Braun, Ann., 460, 288 (1928); Ber., 66, 780 (1933). (227) W. A. Bonner, C. D. Hurd and S. M . Cantor, J . Am. Chem. Soc., 69, 1816 (1947).

(228) (229) (230) (231) (232) (233) (234) (235) (236)

E . Gebauer-Fullneg, W. H. Stevens and E. Krug, Monatsh., 60, 324 (1928). P. Claesson, J . prakt. Chem., [2] 20, 18 (1879). A. M. Gakhokidze, Zhur. Obshche'i Khim., 11, 117 (1941). D. H. Brauns, J. Am. Chem. SOC.,44, 401 (1922). R. T. Major and E. W. Cook, J. Am. Chem. SOC.,68,2333 (1936). G. Zemplen and R. Bognhr, Ber., 72, 47 (1939). G. ZempMn and A. Gerecs, Ber., 68, 2054 (1930). E. Pacsu, J. Am. C I w x . Soc., 64, 3649 (1932). E. Pacsu, E. J. Wilson, Jr., and L. Graf, .I. A m . Chem. SOC.,61,2675 (1939).

25 1

GLTCOSYL HALIDES AND THEIR DERIVATIVES

TABLEIV Properties of Some Glycosyl Bromides Brornide

Melting Point, "C.

[a1n

Teml "C.

Referelrccs

-

____

D-Arabinofuranosyl,tri-0-ncetyl-

-

sirup

-

-

237

B-D-Arabinopyranosyl

tri-0-acetyltri- O-benzoyl8-L-Arabinopyranosyl,tri-0-acetyl a-D-Allopyranosyl,tri-O-acetyl-6deoxycu-D-Ghcopyranosyl

139 147-148 139

-283.4 -353.3 +283.6

sirup

-

3,4,6-tri-O-acetyl-2-deoxy3,4,6-tri-O-benzogl-2-deoxy2,3,4-tri-O-acetyI-6-deoxya-L-glycero-L-gala-Heptopyranosyl,

sirup 139 (dec.) 150-152

+121.4# +246.6

tetra-0-acetyl-7-deoxy8-D-Fructopyranosyl 1,3,4,5-tetra-O-acetyl1,3,4,5-tetra-O-benzoyl-[+0.25 CHaCOOH]

113-113.5 65

1,4,5-tri-O-acetyl-3-0-mesyl-

8-D-Galactofuranosyl 2,3,5,6-tetra-O-acetyl-

22 20 20

43 36 43

-

238

-

26

135 239, 240 241, 242

-249.0

20

209

-188.3

20

243

-179.5 -178.4

20 20

200 31

85-86

- 122.7' 18

16

83-S4.5

- 116.2' 20

148

+248

25

218, 244

+217

20

207

+203 +157.3

20 20

212

20

208

106-110 119

-

16

2,3,5-tri-O-acetyld-bromo-6-

deoxya-D-Galactofuranosyluronic acid,

2,3,5-tri-O-acetyl-, methyl ester or-u-Galactopyranosyl 2,3,4,6-tetra-O-acetyl2,3,4-tri-O-acety1-6-bromo-6deoxy2,3,4-tri-O-acety1-6-O-tosyl3,4,6-tri-O-acety1-2-bromo-2deoxy-

129.5-130.5 84-85 100

146-147

+17.8

-

20

252

L. J. HAYNES AND F. H. NEWTH

Rronride _______

Temp., "C.

Melling Pornl, "C. _____

-

a-D-Glucofuranosyl 2,5,6-tri-O-acetyl-3-O-mesyl(?) 123-123.5 140 (dcc.) 2,5,6-tri-O-acetyl-3-O-tosyl2-O-acetyl-5,6-di-O-benzoyl-3-0123-135 tosyl-(?) 2-0-acetyl-6-O-benzoyl-3,5-tli-0159-160 tosyl121-125 2-0-acetyl-3,5,6-tri-O-tosyla-u-Glucofuranosyluronicarid,2,5di-0-acetyl-, 6,3-lact one a-D-Glucopyranosyl 2,3,4,6-tetra-O-acetyl88-89 126-127 2,3,4-tri-O-acetyl52-53 6-0benzoyl6-bromo-6-deosy176.5 (dec.) 140-141 6-0-p-bromophenyl6-chloro-6-deosy166 (dec.) 127-128 6-deoxy -6-fluoro168-177 ((I??.) 6-deosy-6-iodo6-0-mesyl91-93 6-0-methylsirup 141 6-0-(2-naphthyl) 93-94 6-0-phenyl6-0-diphcnylphosphoryl74-75, 87-88 160 6-deoxy -6-thiocyano88-80 6-O-to~yl2,3,6-tri-O-acet3.1-4-O-tosyl2,4,6-tri-O-acetyl3-0-benzoyl3-0-mesy l3-0-tosyl3,4,6-tri-O-acetyl-2-amino-2-

6-deoxy-6-iodo-2-0-tosyl2,6-di-O-acetyl-3-0-mesyI4-0-acetyl-2,3,6-tri-O-tosyl-

+

160 +198.9

19 20

31, 121 116

- 101

20

120

+156.6 +124.5

20 19

119 119

-.

~

+197.8 +217.4 +180.6 +191.4 fl69.7 +200 +231 +178.9 +189

20 19 22

245 17, 220 132 240 10, 127 247 203 248 249 224 250 247 247 251 252 115, 117, 240 253

+212.1 +166.1

20 20 21 23 18 20 20 26 20 26

171

+141

20

152 150-151

+162. 51L +160 19 +164.P

154-155 149-150 113-114 136 (dec.)

1501 20 +148.4< +170 20 +lOl.S 20

254 255, 256 257 258

+176.4 +155+1( +154 +170 +113.1

246 259 257 121 105

-

deoxy-

hydrobromide hydrochloride 3,4,6-tri-O-acetyl-2-O-tosyl2,4-di-O-acety1-3,6-di-O-(tri-O acetylgalloyl)3,4-di-O-acetyl2,6-di-O-benzoyl2,6di-0-tosyl-

References ~~

oil 143 1'43 136-137 (dec.) 162

-

+177 +201 $149

+

22 20 20 19 20 ~

27 121 27

253

GLYCOSYL HALIDES AND THEIR DERIVATIVES

Bromide

_

_

if elting Point, "C. ~

Temp.

Re/erences

20

105 128 130 80,260 227

"C.

a-D-Glucopyranosyl(continued) 4-0-acetyl-6-deoxy-6-iodo-2,3-di-

O-tosyl6-0-acetyl-2,3,4-tri-O-benzoyl6-O-acetyl-2,3,4-tri-O-benzyl-

tt,tra-0-benzoyltetra-0-propionyl-

-

+114.7

-

-

-

-

-

129-130 11G

+123.6 +141

20 20

147-148 88

+258 -192.7

20.5 26I 17.5 262

Ris (2,3,4-tri-o-aCetyl-a-D-glUCO-

pyranosyl bromide), 6,6-carbonate a-r-Glucopyranosyl, tetra-0-acetyl ~-Dr-~lucopyranosyl, tetra-0acetyl-

85

-

-

262

a-D-glycero-D-gala-Heptopyranosyl,

2,3,4,6,7-penta-O-acetyl-

112

+208

20

111

+187

20

114

+143

20

156

139-140 sirup

- 124 -

20

125 265

62 amorph.

+131.6 +11.7

20 20

79 80

71-72 163-164

- 169

+64.8

20 20

74, 266 162(b)

96 151-153 164-166 sirup 94.5-95.5 84-85.5

-209.3 - 199 +78

25 20 20

+224.8 +165.6

23 20

267 162(a) 85 15 268 123

101-102 134-135

+211.9 +117

20 20

220 36, 232

102 134-135

-211.6 - 116

23 20

269 232

263

a-D-glycero-D-gulo-Heptopyranosyl,

2,3,4,6,7-penta-O-acetyl-

83, 264

,

a -D-glyccro-D-id0 - Heptopyranosyl

2,3,4,6,7-penta-O-acetyla-D-glycero-c-talo-Heptopyranosyl,

2,3,4,6,7-penta-O-acetyla-D-Lyxopyranosyl, tri-0-acetyla-D-Mannopyranosyl 2,3,4,6-tetra-O-acetyI2,3,4,6-tetra-O-benzoyl-

a-L-Rhamnopyranosyl 2,3,4-tri-O-acetyl2,3,4-tri-O-benzoylP-D-Ri bopyranosyl 2,3,4-tri-O-acetyl2,3,4-tri-O-benzoyla-D-Ribopyranosyl, tri-0-benzoylD-Ribofuranosyl,tri-0-acetylr,-Ribopyranosyl,tri-0-acetylD-Talopyranosyl,tetra-0-acetylD-Xylopyranosyl 2,3,4-tri-O-acetyl2,3,4-tri-O-benzoylL-Xylopyranosyl 2,3,4-tri-O-acetyl2,3,4tri-0-benzoyl-

-

-

-

D i - and Oligo-saccharides

or-Cellobiosyl, hepta-0-acetylhexa-0-acetyl-2-bromo-2-deoxy-

183 165-166 (dec.)

+95.8 20 t57.6" 20 -

14, 35 270

254

L. J. HAYNES AND F. H. NEWTH

Melting Point, “c‘

B~oiikide

-

6-O-@-Ccllobiosyl)-a-D-glUCOpyranosyl, deca-0-acetylDiarabinosyl, O-acetyla-Epicellobiosyl, hepta-0-acetyla-Gentiobiosyl hepta-0-acetylhexa-O-acetyl-6’-bromo-6’-deoxy6-O-@-Gentiobiosyl)-a-~-glucopyranos yi, deca-0-acetyl2-O-@-~-Glucopyranosyl) -a-D-glucopyranosyl, hepta-0-acetyl3-O-(~-~-Glucopyranosyl) -n-glucopyranosyl, hepta-o-acetyl4-O-(/3-~-Glucopyranosyl-) -a-D-man nopyranosyl, hepta-0-acetyl6-0-(a-D-Giucopyranosyl)-a-D-glucopyranosyl, hepta-0-acetyl6-O-(8-~-Glucosyluronicacid)-a-ngalactopyranosyl, hexa-0acctyl-, methyl ester 4-0- (~-n-G~ucosyluronic acid) - a - ~ glucopyranosyl, hexa-0acetyl-, methyl ester a-Lactosyl, hepta-0-acetyl[a-Laminaribiose, see 3-O-(p-nGlucopyranosyl) -D-glUCOpyranose] a-Maltosyl, hepta-0-acetyla -Meli biosyl, hept a-0-acetyl5-Turanosy1, hepta-0-acetyla

I n acetylene tetrachloride.

b

I

Rejerencea

209 (dec.) 104-105 168-169

+69.0 +207.6 +77.9

20 18 20

132 43 21

144 193 (der.) 193-194

+101.1 +108.1 +63.3

20 18 17

21, 100 249 271

194 (dec.)

+95.6

20

272

+85

18

275

+77.9

20

180.5-181.5 168-169

21, 128

131-133

+202

24

273

201-202

+194.7

23

271

200

+99.4

24

274

145 (dec.)

+108.7

23

112-113 116 133-134

+180.1 +209.9 -30.5

20 20 20

I n carbon tetrachloride.

33, 57

205 205 236

I n acetonc.

(237) N. W. Bristow and B. Lythgoe, J . Chem. Soc., 2306 (1949). (238) P. A. Levene and J. Compton, J . B i d . Chem., 117. 37 (1937). (239) M. Bergmann, H. Schotte and W. Lechinsky, Ber., 66, 158 (1922); 66, 1052 (1923). (240) P. A . Levene and F. Cortese, J . Biol.Chem., 92, 53 (1931). (241) F. Micheel, Ber., 63, 755 (1930). (242) J. Compton, J . A m . Chem. Soc., 60, 395 (1938). (243) D. H. Braunr, J . A m . Chem. Soc., 46,2381 (1923). (244) S . Morell, L. Baur and K. P. Link, J . Bio2. Chem., 110, 719 (1935). (245) C. Neuberg and W. Neimann, Hoppe-Seyler’s 2. physiol. Cheni., 44, 114 (1905). (246) P. Brig1 and H. Gruner, Ann., 496, 60 (1932). (247) H. Ohle, E. Euler and R. VoulliBme, Ber., 71, 2250 (1938).

GLYCOSYL HALIDES AND T H E I R DERIVATIVES REFERENCES FOR

.255

TABLEIV (Continued)

(248) B. Hclferich and A. Gnuchtel, Ber., 74, 1035 (1941). (249) B. Helferich and H . Collatz, Ber., 61, 1640 (1928). (250) B. Hclferich and E. Gunther, Ber., 64, 1276 (1931). (251) T. Posternak, J. Biol. Chem., 180, 1269 (1949). (252) A. Muller and Adrienne Wilhelms, Ber., 74, 698 (1941). (253) B. Helferich and A. Muller, Rer., 63, 2142 (1930). (254) E. L. May and E. Mosettig, J. Org. Chem., 16,890 (1950). (255) J. C. Irvine, D. McNicoll and A. Hynd, J. Chem. SOC.,99,250 (1911). (256) J . C. Irvine and J . C. Earl, J. Chem. Soc., 121, 2370 (1922). (257) E. Hardegger, 0. Jucker and R. M. Montavon, Helv. Chim. Actu, 31, 2247 (1948). (258) 0 . T. Schmidt, S. Berg and H. H. Beer, Ann., 571, 19 (1951). (259) E. Hardegger, R. M. Montavon and 0. Jucker, Helv. Chim. Actu, 31, 1863 (1948). (260) L. W. Muzzeno, Jr., J . Anz. Chem. SOC., 72, 1039 (1950). (261) D. D. Reynolds and W. 0. Kenyon, J . Am. Chem. SOC.,64, 1110 (1942). (262) P. Barrer, C. Nageli and A. P. Smirnoff, Helv. Chim. Actu, 5, 141 (1922). 64,247 (1942). (263) Edna M. Montgomery and C. S. Hudson, J . A m . Chem. SOC., (264) Edna M. Montgomery, N. K. Richtmyer and C. S. Hudson, J . Am. Chem. SOC., 65, 1848 (1943). (265) P. A. Levene and M. I,. Wolfrom, J . Biol. Chem., 7 8 , 525 (1928). (266) E. Fischer, M. Bergmann and A. Rabe, Ber., 63,2362 (1920). (267) P. A. Levene and E. P . Clark, J. Biol. Chenz., 46, 19 (1921). 61, 3012 (1939). (268) C. W. Klingensmith and W. L. Evans, J . Am. Cheni. SOC., (269) L. C. Kreider and W. L. Evans, J . Am. Chem. Soc., 68, 797 (1936). (270) M. Bergrnann and H . Schotte, Ber., 64,1570 (1921). (271) B. Helferich and R. Gootz, Ber., 64, 109 (1931). (272) K. Freudenberg and K . Soff, Ber., 69, 1245 (1936). (273) M. L. Wolfrom, L. W. Georges and I. L. Miller, J A m . Chem. Soc., 71, 125 (1949). (274) W. F. Goebel, R. E. Reeves and R . D. Hotchkiss, J . A m . Chem. SOC.,69, 2745 (1937). (275) P. Biichli and E. G. V. Percival, J. Chem. SOC., 1243 (1952).

256

L. J. HAYNES AND F. H . NEWTH

TABLEV Properties of Some Glycosyl Iodides Temp.,

Iodide

I

I

Monosaccharides P-L-Arabinopyranosyl, tri-0-acetyl- I a-n-Glucopyranosyl 2,3,4,6-tetra-O-acetyl109-110 2,3,4-tri-O-acet,yl-6-deoxy-6-iodo-ca. 150 (dec.) 2,3,4,6-tetra-O-benzoyI141-142 a-u-LVannopyranosyl 05 2,3,4,6-tetra-0-acetyl2,3,4 ,6-tetra-O-benzoylamorph. a-r,-Rhamnopyranosyl, t ri-0- benzoyl- 143-144 (dec.)

~

References

I$339.1

1

20

49

+23l .Oa f205.9 +139.5

20 21 20

33, 87 249 80

+190.5 +45 -27.1

20 20 20

70 80 162b

TABLE V (Continued) References

Disaccharides a-Cellobiosyl, hepta-0-acetyla-Epicellobiosyl, hepta-0-acetyla-Gentiobiosyl, hepta-0-acetylo-(P-D-Glucopyranosyl)-a-n-mannopyranosyl, hepta-0-acetyla-Lactosyl, hepta-0-acetyla-Melibiosyl, hepta-0-acetylTuranosyl, hepta-0-acetyla

I n acetylene tetrachloride.

160-170 (dec.) 140 (dec.) 134 (dec.) 140 (dec.) 145 (dec.) -

105-106

I

I

+125.7 +111.5 f126.1 +111.5

20 20 20 20

+136.9

23 20

-

-54.2

21 21 21, 100 21

57 205 236

THE METHYL ETHERS OF THE ALDOPENTOSES AND OF RHAMNOSE AND FUCOSE

BY GEORGEG. MAHER Research Laboratories, Clinton Foods Inc., Clinton, Iowa The following revised set of tables of the physical characteristics of the methyl ethers of the aldopentoses and rhamnose and fucose (and their derivatives) replaces those of R. A. Laidlaw and E. G. V. Percival, Advances i n Carbohydrate Chem., 7 , 1 (1952), and brings the subject up to date.

TABLEI The Methyl Ethers of D-Xylose Melting point, "C.

Substance

2-O-Methyl-8-~-xylose

1 132-133 1 134 I

I

131 135-137 137-138

triacetate 95 2-O-Methyl-N-phenyl-~-xylopyran125-126 osylamine 128 Methyl 2-0-methyl-8-n-xylopyran- 111-112 oside 3,4-diacetate 78-79 3,4-di-p-toluenesulfonate 123 Z-O-Methyl-~ -xylonolactonc 66-68 2-O-Methyl-~-xylonamide 3-O-Methyl-~-xylofuranose 1,2-O-isopropylidene acetal 1,2-0-isopropylidene acetal, 5-ptoluenesulf onate

t All references

96-98 98.101 sirup 114

[ a ] degrees ~,

Re{erencest ~.

-23.9 + +35.9 -22 -+ +26 (3 days) -21 -+ +23.8 (1 day) -23 -+ 4-35 -21 -+ +34 (2 hrs.) -2.2 +212, +214

HzO

1,2,

HzO

5

EtOH

17

H20 HzO

4, 6 109

CHCla EtOAc

1 3, 4,

+23.7 -67.7, -70

EtOAc CHCla

17

-38 - 16

CHC13 CHCla Hz0

1 1 6

H20

6

CHCls CHC13

7 7 8

3

5, 6

$101 + +74 (504 hrs.) +52.5

-64 -27.2

appear in a separate list starting on page 269. 257

Rolalion solvenf

1, 17

GEORGE G. M-kHER

.If ellifkg polnl, c.

phenylosaeone p-bromophenylosazonc

degvecs

LO8-104 36-88 15 )5-97 )&lo1 172

f 5 5 -> $17 tl9.5 f 4 5 + +19

L53-155

f G -+

3-O-Methyl-N-phenyl-a-~-sylopy- I37 ranosylamine

3-O-Methyl-~-xylonolactone

14

4-O-Methyl-~-xylose phenylosazone

sirup 158-158.5

160-161 Methyl 4-O-methyl-p-~-sylopyran-35 oside sirup 5-0-Methyl-D-xylose 170-171 p-bromophenylosazone

1,2-0-isopropylidene acetal, 3 - p toluenesulfonate Methyl 5-0-methyl -a-D-xyloside, 3p-toluenesulfonate Methyl 5-O-methyl-p-~-xyloside,3p-toluenesulfonate 2,3-Di-O-methyl-a-~-xylose

[..ID,

Rotalion Aolzrent

H1 0 K O HzO

J, 11

12 10 109 13 B

-14

+80 +77

+76 + t-40 (820 hrs.) +9 f 2 +25 + 0

ChH5NEtOII EtOAc EtOAr HzO

HzO C5H5NEtOH

9 10, 11, 12 14 10

15 16

15 15

-69

HzO

+32.8 --+ +36 -50 + -30

9 9

81-82

-31.8

HzO CbH5NEtOH CHCl,

sirup

+44.5

CHC1,

8

89

-51.7

CHCl,

8

79-80

+70 + f 2 3 (1 H,O day)

Methyl 2,3-di-O-methyl-a-~-xylo-sirup side Methyl 2,3-di-O-methyl-p-o-xylo- sirup side 63 56-59 4-p-toluenesulfonate 2,3-Di-O-methyl-N-phenyl-~-xylo-145-146 pyranosylamine 121-123, 126 131-132

8

+61.8

CIIIOH

18, 19, 20 I 21 18

-5.8

CHC13

1

-47.3 -8.8 +182.5 f 2 . 5

H2O CHCla EtOAc

+180

EtOAc

12 1 17, 18, 22 3, 22, 23 24

250

METHYL ETHERS OF PENTOSES AND 6-DEOXYHEXOSES

TABLEI (Conlinued) ~~

Substance

Melting

[a]~ degrees ,

point, "C.

2,3-Di-O-methyl-~-xylono-~-~acsirup tone

132-134

2,3-Di-O-methyl-~-xylonic acid, phenylhydrazide p-bromophenylhydrazide 2,4-Di-O-methyl-p-~-xylose

107-108 114-115 150-151 108 108 111 116-118 2,4-Di-O-methyl-N -phenyl-~-xylo- 170 155-157 sylamine Methyl 2,4-di-O-methyl-p-~-xylo-77.5-78.5 side 60-61 3-p-toluenesulfonate 88 75-76 2,4-Di-O-methyl-~-xylono~actone sirup

Rolafion solvent

ReJerences

H?O

18 17 24

$46

HzO 95% EtOH H2 0

+30

EtOH

+97 +69 (400 hrs.) 87 +94.3 --f

+

HzO CHCla dioxane EtOAc CHClz CHCI, CHCI, CHC13 HzO

17, 23, 25 18 23 18 26 109 4 , 10 16 26 16 16 1 1 16 3, 26

H,O H9 0 EtOH

16 16 16

+36 +16.4 +33.7

H,O EtOE-I CHCl,

8 8 8

-49.9 $24.9 -+ +20.5 +5.3 f40, f 3 0 $13 +31 f 5 -82 -33 -71 -34.8 -56 -+ -27 (65 hrs.)

CHCI,

8 1, 13

-30

--f

$22

-13 4 +23 -26 - 82 -40 - 70 -82.4 -58.0 +28.8 -15 --t +30 (3

HI0

days)

2,4-Di-O-methyl-~-xylo~amide 2,4-Di-O-rnethyl-r,-xylonic acid, phenylhydrazide 2,5-Di-O-methyl-~-xylose

98-100 143.5144.5 sirup

Methyl 2,5-di-O-methyl-a-~-xylo-sirup side, 3-p-toluenesulfonate p anomer sirup 3,4-Di-@methy1-D-xyl ose sirup

Methyl 3,4-di-O-methyl-p-~-xylo- 89-90 side 2-p-toluenesulfonate 105 3,4-Di-O-met,h~~l-1~-x~Ionolacto1ie 68

-13 +51 +47

--f

+29.5

H 2 0

CHCI3 CHCla MeOH MeOH CHCla CHCI, HP O CHCI? r-I .o

1 28 15 27 1 5 15 1 13, 15, 21,

27, 20, 30

260

GEORGE G. MAHER

TABLEI (Continued) Substance

3,4-Di-O-methyl-~-xylonic acid, phenylhydrazide 3,5-Di-O-methyl-o-xylose

1,2-0-isopropylidene acetsl p-bromophenylosazone

Melting point, "C.

15

sirup sirup 107-108

+11 +25 -60 -46 + -30 +72 + +41 (48 days) +75 -+ +27 (33 days) ~ 0 1 1 6 7 8 0 +81.5 + +39 (49 days)

95-96

+6

91-92

+64

+

+18

Methyl 2,3,4-tri-O-methyl-a-~-xysirup loside

+86 +49.5

Methyl 2,3,4-tri-O-methyl-8-~-xy49-50 loside 2,3,4-Tri-O-methyl-N-phenyl-o-xy-102 losylamine 97-98 2,3,4-Tri-O-methyl-~-xylonolac- 54,55,56 tone 2,3,4-Tri-O-methyl-u-sylonic acid, 138 phenylhydrazide 2,3,5-Tri-O-methyl-~-sylose sirup

-73 -66 -84 + +47

Methyl 2,3,5-tri-O-rnethyl-u-xylo-sirup side 2,3,5-Tri-0-methyl-I-xylonolac t,one

sirup

2,3,5-Tri-O-meth~.l-o-sylonamide 84-85 2,3,5-Tri-O-methyl-u-sylonic acid, 89

phenylhydrazide

References

132

sirup

3,5-Di-O-methyl-~-xylonic acid, phenylhydrazide 2,3,4-Tri-0-methyl-o-xylose

-

[ a ] degrees ~,

CHC13

7

HzO

7

CHCl, 7 CsHsN- 9 EtOH HzO

7

H20

5

H2O

9,31

EtOH

9

Hz0

4,11, 25, 29, 32, 33I 34, 35, 50 32 32

MeOH HC1EtOH CHCl, MeOH EtOH

16 15,35 10,14 17,22 11,16, 50 36

-97 + +32.8 -4 + +21 (120 hrs.)

MeOH

4-24.7+ +29.5 f32 $114 +134 +74 -+ +61.4 (504 hrs.) [alsrtI1 +I00

H,O

37

MeOH MeOH HzO

37 5 5 31,38

Hz0

H20

H?O

18 39 18

METHYL ETHERS OF PENTOSES AND

G-DEOXYHEXOSES

2G I

TABLE I1 The M e t h y l Et rs of D-) ubin ose Subslance

2-O-Methyl-n-arabinose

&felling

poin6, "C.

sirup

143 (dec.) phenylhydrazone 113 Methyl 2-0-mcthyl-p-~-nrsbinopyiano- 48 side 62-63 Methyl 2-O-niethyl-~-arnbiriosicle 2-O-Methyl-~-ara\~onolactorie 87 p-tolylsulfonylhydraxone

2-0-Me thy1-D -arabonamide hydrate 2-O-Methyl-~-arabonic acid,phenylhydrazide Ammonium 2-O-methyl-~-arabonate 3-0-Methyl-D-arabinosz

131 9697 158-159 (dcc.) 146 sirup

3-O-Methyl-~-arabonolactone

81

3-0-Methyl-D-arabonamide 2,4-Di-O-methyl-~-arabinose

132 sirup

2,4-Di-O-methyl-N-phengl-~-ar~ _inosy1amine 2,5-Di-O-niethyl-~-arabinose 2,5-Di-O-methyl-~-arabonolactone

142-143

2,5-Di-0-methyl-D-arabonamide

sirup sirup 59-60 131-132 30.5-131 166-167

2,5-Di-O-methyl-u-arabonic acid, phenylhydraaide 3,5-Di-O-methyl-~-arabinose sirup 3,5-Di-O-methyl-N-phenyl-~-arabino-118 sylamine 3,5-Di-O-niethyl-~-arabonolactone 74-75

3,5-Di-O-methyl-~-arabonamide 144 3,5-Di-O-methyl-o-arabonic acid, 144-145 phenylhydrazide Methyl 2,3,4-tri-O-methyl-o-arabi110-sirup side

[ a ] degrecs ~.

Rolnlion solvcnl

- 102

HzO

- 17

HzO

-205 -15.4 +52.7 4 +47.4 (90 hrs.) -53.2

Re/-

E P e nc E s

40,41, 42 41

41 41 M e O H 42 40 HI0 1% 2 0

40

H20

40 40

HzO

40

-27.7 HzO -90 + -43 (3 H 2 0 days) f99 -+ +-75 Hso (22 days) (passes through a max.)

40 5

-23

5

5

43 MeOH 43 43

-30.8 -37.8

HzO

+23 -51 -+ -32 (20 days) +62.2

HsO HzO

HzO

103 4

103 4

-34

HzO

103 4

447

1120 HzO

5 44

H2

0

5

H2

0

44

-28.8

$85 -+ +57 (22 days) -11

5

-248

H2

0

45

-

GEORGE G . MAHER

TABLEI1 (Continued) Melting poinl, “C

Substance

2,3,4-Tri-O-methyl-~-araboni~mide 96 2,3,5-Tri-O-methyl-~-arabinose sirup Methyl 2,3,5-tri-o-methyl-~-arabino- sirup

[.ID,

degrees

-25 $40 80

+

side 2,3,5-Tri-O-methyl-~-arabonolactone 30 2,3,5-Tri-O-niethyl-~-arahonnmide 134-135 -25 137-138 - 19 135-136 - 14 137-139 -16.5 sirup +18.4 144 -35.2

Rololion solvent

MeOH 47 MeOH 46 46 HzO 103

EtOH 44 MeOH 47 H?O

HzO H?O EtOH

TABLEI11 The M e t h y l Ethers of L-Arabinose Subslance

Melting boint, “C.

[a]~ degrees ,

Rejerences

4,44, 46 103 46 47

Rototion solvent

References

~~

2-O-Methyl-~-arabinose

sirup

+100

pheriylhydrazone

114, 116 p-tolylsulfonylhydrazone 145-145 Methyl 2-O-methyl-p-~-arabinopyrano- 63-65 side hydrate 46-47 2-O-Methyl-~-arabonolactone sirup 2-O-Methyl-~-arabonamide 131 3-0-Me thyl-L-arabinose sirup phenylosazone 163 3-O-Methyl-N-phenyl-~-arabinopyrano-117 sylamine 3-O-Methyl-~-arabonolactone 78 3-O-Methyl-~-arabonamide 132 2,3-Di-O-methyl-~-arabinose sirup

+208 -44 + -40 +52 $110

-74

+

107 +101

2,3-Di-O-methyl-N-phenyl-~-arabinopy138,

ranosylamine

139 35 30

-38 + -25 (12 days) -38 + --30 (7 days) -33

41,48, 49 41,49 50 48 48 49 49 51 52 51

51 51 49,52 53 49,53, 547 52 52 53

95

263

METHYL ETHERS O F PENTOSES AND 6-DEOXYHEXOSES

TABLE I11 Continued) Substance

2,3-Di-O-me thyl-L-arabonamide

Melting point, "C

162 156, 160 sirup

2,4-Di-O-methyl-N-phenyl-~-arabino- 145sylamine 146 126 2,4-Di-O-methyl-~-arabonolactone sirup

158 sirup Methyl 2,5-di-O-rnethyl-~-arabinoside sirup 2,5-Di-O-methyl-~-arabonolactone 60 131, 132 2,5-Di-O-methyl-~-arabonic acid

phenylhydrazide 3,4-Di-O-methyl-~-arabinose

[a]D,degrees

+17 +30. 85 +37.8= +I18 +129 i 4

+55 + +27 (14.5hrs.) +99 + +39 (17 hrs.)

+60

-2 -60 -60 -+ -44.8 (320 hrs.) - 19 +38

+I16 +lo4 +125

phenylosazone 132 Methyl 3,4-di-O-methyl-p-~-arabinosidesirup 2-benzoate sirup 2-p-toluenesulfonate 111112 3,4-Di-O-methyl-~-arabonolactone sirup 3,4-Di-O-methyl-~-ara bonamide 133 3,5-Di-O-methyl-~-arabinose phenylosa- 170 zone

Re/eTences

49, 52 23,53, 65, 95 43 43 53 55 53, 55 53 43 55 43 57

56 56 43,56, 57 50 4, 50, 56 43, 57 56, 43 2

+25.8 + -16 (120hrs.)

162, 163 sirup

Rulalion solvent

50,56, 43 58 53 111 53

+210.6 +143.5

58

+44 + -1 (6 hrs.) +28.2

111

58 58

111 59

264

GEORGE G. MAIlER

TABLE 111 (('ontini~erl) Subslance

3,5-Di-O-methyl-~.nrabonic acid, phenylhydrazide 2 , 3 , 4-Tri-0-methyl-L-arabillosc

M elling Poinl, "C.

[ a ] d~p , grees

-84

HpO

60

73 78 145 144 114

-83 - 43

1120 CHCI, Hp0

51 50 59,60 51

sirup

+l20, +122, 127

H,O

+46.2b

1120 MeOH HzO MeOH HIO

sirup

+10

51

+

+24b

+250b +223b +I45 + +22.4 hrs.)

+24

+45

62 75, 63 75 43, 63

MeOH

43 43, 53,

63, 64 53, 55

159, 160 0

H20

-43

-45 4 -24 (50 hrs.) -47, -44

H.0

138138

+20, +21

I120

138

+I6

H2

137 130

+17.4 +21, +24

Hp0

28,30 20, 31

62

EtOH HpO

156, 157

2,3,5-Tri-O-mcthyl-1,-arabinose sirup Methyl 2,3,5-tri-O-mrth~l-~-arabirioside sirup 2,3,5-Tri-O-niethyl-~-araborlolactone 33

53,55, 61, 62 62 62

(24

+I36 107, 103 2,3,4-Tri-O-meth.vl-r, arabonic arid, phenyl hydrazide

erences

75

Methyl 2,3, 4-tri-O-methyl-~-r,-:trabino- 46-48 side B anomcr 44-46

-+ -69 (28 days)

Ref-

Rolnlion solvent

HpO

57 60 43 05, 43 40, 50, 65 14,23, 59 43 49,60, 65, 66 104 17, 66 9

0

EtOH

These rotations wcre deduced from the rotations of the isomeric D compounds. * T h e rotations of the anomers have been reversed from those in the original paper, in view of Hudson's rule. a

T A B L EIV*

-

'I'he Methyl Ethers of D-Ribose Subslance

Meltiag poinl, "C.

Methyl 2,3-3nhq'dro-~-O-nict2lyl-p- 75-77 D-rihoside sirup 5-0-Met hyl -D -ribose 161-162 p-bromophenylosazone 2,3-Di-O-methyl-D-rihose

3,5-Di-O-methyl-~-ribosc phrnylosazone 2,3,4-Tri -0-methyl-D-ribose

35-86 Jaioo

2,3,4-Tri-O-methyl-~-ribonic acid 2,3,4-Tri-O-methyl-~-rihono-6-lac- sirup tone

sirup

2,3,5-Tri-O-meth?.l-N-phenyl-~ri- 55.5, 56.5 bosj~lamine sirup Methyl 2,3,5-tri-O-methyl-~-riboside 2,3,5-Tri-O-rnethyl-o-ribonicacid

2,3,5-Tri-O-methyl-u-ribono-r-lac-18.5-19 tone

phenylhydrazide

-~

~-

-7

€I20

-48

CsHhN8:to13

108.5109.5

Re/erenm

15 67 67 68 60

sirup 161

sirup

2,3,5-Tri-O-methyl-~-ribonic acid,

[a]D,degrees

Rolalron solvenl

-51.7

4

-40

I3rO

70 105 72 70

-26.7 -35

MeOH HIO

-24.2 f34.0 +69.3

MeOH CHCI,

70 70 70

f85.4 $114.1 -4.4++17.1 (191 hrs.) +46.2 f41.4

C 611 6 EtzO Hz0

70 70 70

MeOH MeOH

+40.6 f51.6

HzO HzO

72 69 7 106 67 72, 107 69, 71

$59.1

MeOH

69

+40.9 -+ $11.9 (448 hrs .) -20.2 3 -10.6 (141 hrs.) -18.9 3 $7.6 (703 hrs.) $55.9 +124.0 f83.1

Hz0

72

HzO

67, 69

Hz0

72

CHCls EtzO C6H6

72 72 72 60

HIO

* For additional material see G. R. Barker, T . M Noone, D. C. C. Smith and J. W Spoors, J . Chem. SOC.,1327 (1055); and G. R. Barker and D. C. C . Smith, .I. Chem. Soc., 1323 (1955). 265

TABLEV The Methyl Ethers of o-Lyxose

Melting oint, "C

Subslance

-

[u]D.

Rolnlion solvenl

degrees

+60

Methyl 2,3-anhydro-5-0-methyl-cu-u-lyxo- 43 side 2,3,4-Tri-O-methyl-~-lyxose 79 Methyl 2,3,4-tri-O-methyl-o-lyxoside sirup

-10

5 73 73 73 73

-22 +lo

--f

[a16461

+37.3 +35.5--9.3 (66.5 hrs.)

2,3,4-Tri-O-methyl -~-lyxonolactone

sirup

2,3,4-Tri-O-methyl-o-lyxonic acid, phenylhydrazide 2,3,5-Tri-O-methyl-o-lyxosc 2,3,5-Tri-O-methy~-~-lyxonolactorle

180181 sirup +39 41 +82.5 4 +56.5 (1000 hrs.) sirup -20.8 --L +25.6 (500 hrs.) 140 142

2,3,5-Tri-O-methy~-~-lyxonic acid phenylhydrazide

References

36, 73 74 74

74 74 36

TABLEVI* The Methyl Ethers oj" L-Rhamrkose Substance

Mellinx point, "C.

sirup 113-114 2-0-Methyl-N-phenyl-L-rhamnosyl-152 amine Methyl 2-0-methyl-~-rhamnopy138-140 ranoside 2-O-Methyl-~-rhamnono-y-lactone 116-1 17

2-O-Methyl-~-rhamnonamide 3-O-Methyl-~-rhamnose phenylosazone hydrate 4-O-Methyl-~-rhamnose

phenylhydrazone phenylosazone

117-118 113 128-130 118 (dec. 125-126 122

176 (dec. 162-163

triacetate

[a]D,

Rotirlion solvent

+31, +24

H,O

+43

CsHGN

References

76,110 77 110 76

-62 4 -64 (117 hrs.) +57 (17 hrs.)

HzO?

CfiHsNEtjOH

110 110 28 78 78

+13

-12.9

+26 $25.8 -12.2

266

degrees

-+

f14.3

MeOH MeOHNH3 H20

79 80

CfiH6NEtO H MeOH

79

110 80

80

TABLE VI (Continued) Subslance

Melling

Methyl 4-O-methyl-a-~-rhamnosidesirup j3 anomer sirup 4-O-Methyl-~-rhamnono-6-lactone82 80- 81

5-0-Methyl-L-rhamnose

[a]~ degrees ,

)aid, "C.

pheny lhydrazone

102-103 162-163

phenylosazone

123-124

triacetate 2,3-methylene acetal

115-116 77-79

Methyl 5-0-methyl-a-~-rhamnoside 59-60 5-O-Methyl-~-rhamnono-~-lactone 164-166 2,3-Di-O-methyl-~-rhamnose sirup 1,5-di-benzyl ether

119 2,3-Di-O-methyl-N-phenyl-~-rham138 136-137 nopyranosylamine

-50.2 -13.9 -141+ -115 (14 hrs.) -140 + -112 (18 hrs.) -4.3 -18.4 + +8.1 (7 days) $65.3 -+ +44.4 (3 days) -76.3 $6.4 + +4.5 (46 hrs.) -89.2 -36 f 4 $47.6, +42.5 +71.7 +147.8+ +42.8 (70 hrs.) -6, -14

Methyl 2,3-di-O-methyl-a-~-rhamsirup noside Methyl 5-0-benzy1-2,3-di-O-methyl93 -72 L-rhamnoside 2,4-Di-O-methyl-~-rhamnose 01-93 +10.6 2,4-Di-O-methyl-N-phenyl-~-rham+128.5 + f 5 . 6 141nosylamine 142.5 +136 + +4 3,4-Di-O-methyl-~-rhamnose -10 + +18.6 91-92 +18.2 (equilib94-95 rium) $24 + +18.5 98-99 19 102-103 3,4-Di-O-methyl-~-rhamnose, 1,2- 67 +36 (methyl orthoacetate) +40.6 67-68 3,4-Di-O-methyl-~-rhamnonic acid -15.9+ -118.1 3,4-Di-O-methyl-~-rhamnonolac- 76-78 -158.5 + tone -116.6 -154 + -116 78-79 (48 hrs.) -148 -+ -117 (72 hrs.) -153 --t -119 66-68 (150 hrs.) 152155, 154150 sirup

+

267

Rotation solvent

HnO

References

H20 HzO

79 79 81

HzO?

110

H*O C6H5N5

79 79

CsHbNEtOH MeOH

79

HzO? H2 0 HzO?

79 110

EtOH

79 110 78, 53 78 82, 83 108

H?O

82, 83

Me&O

78

1320

lOX(1)) 108(a) 108(b) 14, 84 85

H20 MenCO

EtOH EtOH H?O

HzO

H2

H rO HZO H?O H2 0

86 4 87 84 87 81 85, 88

HtO

89

H*0

4

Hz0

84

0

H20

84, 88

H2O

%90, 91, 92 ___

TABLE VI (Continued)

-

~

Melting )oant, "C.

Subslance

[WID,

2,3,4-Tri-O-methyl-N-phenyl-~- 111,112

Rotation solvent

degrees

References

44, 56

+127

rhamnosylarnine 110 119-120 sirup

hfethyl 2,3,4-tri-o-methyl-a-~rhamrioside 53-54 ,3 anomer 40-41 2,3,4-Tri-O-rnethyl-~-rharnnonolactone 2,3,4-Tri-O-methyl-~-rhamrionic 177 acid, phenylhydrazide 2,3,5-Tri-O-methyl-~-rhamnonic 160 acid, phenylhydrazide

+130 (14 hrs.)

Me&O

-15.1

HzO

+106 -130

84 94

H20 +

-78

93 92 91

HzO

56, 95 56

a No ethanol was used in the rotation solvent, according t o the authors. *;For additional material see G. R. Barker, T. M. Noone, D. C. C. Smith and J. W. Spoors, J . Chent. Soc., 1327 (1955); and G. R. Barker and D. C . C. Smith, J . Chem. Soc., 1323 (1955).

TABLE VII The Methyl Ethers of o-Fucose Melting boinl, "C.

Subslance

[..ID,degrees

Rotation solvent

5 ) O

$5 -

2-0-Methyl-n-fucose

155161 Methyl 3,4-0-isopropylidene-2-O-mcthyl-98-100 n-fucoside sirup 3-O-Methyl-~-fucose 106, 119 178, phenylosazone 179 170180 Methyl 3-0-met.h~l-cu-~-fucopyrarioside sirup 97-99 0 anomer 3-0-Methyl-u-fuconolactorie 136137 137138 2,3,4-Tri-O-methyl-~ -fucose hydrate

sirup 65

1332,3,4-Tri-O-meth~-l-S-p1ieriyl-~-fucosyl135 amine hlcthyl 2,3,4-tri-0-methyl-~-~-fucosidc03-98 268

+73

+

+87

H20

76 76

+ +

[I2 0 HzO e10

103 +110 106

96 98 97 99

-> +18 (14 hrs.) +124.4

C,H&

-75.3 f 4

h20

01 98 98

-92.5 + -74.9 (16 (lays) 106 +183 +128.8 (cnlc'd. as nnhydrous) +76

H20

00

H?O H2 0

13 00

EtOII

13

+11.2

HzO

f0.5

+

---f

EtOH MezCO

00

13

-

METHYL ETHERS OF PENTOSES AND G-DEOXYHEXOSES

269

TABLEVIII The Methyl Ethers of L-Fucose

-

~

M.eZfing p o d , "C.

Substance

[.ID,

L:

degrees

p

-

Me thy1 3,4-0-isopropylidene-2-0-methylB-L-fucoside 3-0-Methyl -L-fucose p heny 1osaz one Methyl 3-O-methyl-a-r,-fucopyranoside 3-O-Methyl-~-fuconolactone

88-92

-10.9

100

sirup 172-176 130-132 sirup

-94

102 102 102 102

3-O-Methyl-~-fuconamide 176180 2,3-Di-O-methyl-~-fucose sirup Methyl 2,3-di-0-methyl-a-~-fucopyranoside49-51 2,3 -Di-0-methyl-r. fuconolactone sirup 2,3-Di-O-methyl-~-fuconamide 3,4-Di-O-methyl-~-fucose Methyl 3,4-di-O-methyl-a-~-fucoside 2,3,4-Tri-O-methyl-~-fucose

hydrate

78-79 76, 82 100

36-37

65

2,3,4-Tri-O-methyl-N-phenyl-~-fucosyl- 133-134 amine 97-98 Methyl 2,3,4-tri-O-methyl-01-L-fucoside 85-92 p anomer 101.5102.5 2,3,4-Tri-O-methyl-~-fuconolactone sirup 102

- 173 +25 -+ +75 (62 hrs.) +16.4 +4.6 - 190 f 9 4 +47 (22 hrs.) +30.2 - 118 -213 -184 -+ - 128 -111 -169 -+ -118 (24 hrs.) -77 -209 - 196 -21.1

-1384 -36 (48 hrs.) -35

102 102 102 102 102 82 82 100

13 100

13

100 13 100 13 13

-

References (1) G. J . Robertson arid T. H. Speedie, J. Cheni. Soc., 824 (1934). (2) R. J. McIlroy, J . Cheiiz. SOC.,100 (1946). ( 3 ) G. 0. Aspinall and R. S. Mahomed, J . Chem. Soc., 1731 (1954) (4) E. L. Hirst,, E. G. V. Percival and Clare B. Wylam, J . Chem. SOC.,189 (1954). (5) Elizabct,h E. Percival and R. Zohrist, J . Chem. SOC.,564 (1953). (6) E. G. V. Percival arid I. C. Willos, J . ('hem. Soc., 1608 (1949). (7) R. A . Laidlaw, J . ( ' h w i . SOC.,2941 (1952). (8) G. J. Robertsoii and D. Gall, J . Chein. Soc., 1600 (1937). (I)) 1'. A . 1,evene and A . I,. Raymond, .I. B i d . ('hem., 102, 331 (1933). (10) R. A . 1,aidl:tw and IS. G. V . Percival, .I. P h t . SOC.,528 (1950). (11) E. V. White, J . Am. Chem. Soc., 76, 257, 4692 (1953).

270

GEORGE G. MAHER

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METHYL ETHERS OF PENTOSES AND

(52) (53) (54) (55) (56) (57) (58) (59) (60) (61) (62) (63) (64) (65) (66) (67) (68)

G-DROXYHEXOSER

27 1

F. Smith, J . (:hem. Soc., 753 (1939). P. Andrews, D. H . Ball and J. K. N . Jonw, J . C'hcttz,. SOC.,4090 (1953).

P. Andrews and J . K. N. Jones, J . Chem. SOC.,1724 (1054). J . K. N . ,Jones, J . Chcm. Soc., 1672 (1953). F. Smit,h, J . Clrcm. Soc., 1035 (1940). J . Ti. N . Jones, J . (?hem. SOC.,1055 (1947). J. Honeyman, J. Chem. SOC.,990 (1946). E. V. White, J . Am. Chem. Soc., 68, 272 (1946). J. I. Cunneen and F. Smith, J . Clwm. Soc., 1146 (1948). T. Purdie and R. E. Rose, J. Chem,. Soc., 89, 1204 (1906). E. L. Hirst and G. J . Robertson, J . Chem. Soc., 127, 358 (1925). E. V. White, J . A m . Chem. Soc., 74, 3966 (1952): 76, 257 (1953). J. K. N . Jones, Chemistry & Industry, 954 (1052). F. Brown, E. L. Hirst and J. K. N. Jones, J . Chem. Soc., 1761 (1949). E. L. Hirst and J. K. N . Jones, J . Chena. SOC., 506 (1946). P. A. Levene and E. T. Stiller, J . B i d . Chem., 102, 187 (1933). G. R . Barker and M. V. Lock, J . Chem. Soc., 23 (1950); R. W. JBanloz, G. R. Barker and M. V. Lock, Nature, 167,42 (1951). (69) G. R . Barker, J . Chem. SOC., 2035 (1948). (70) P. A. Levene and R. S. Tipson, J . B i d . Chem., 93,623 (1931). (71) A. S. Anderson, G. R. Barker, J . M. Gulland and M. V. Lock, J . Chem. Soc., 369 (1952). (72) P. A. Levene and R. S. Tipson, J . Biol Chem., 94, 809 (1932). (73) E. L. Hirst and J . A . B. Smith, J . ('hein. SOC.,3147 (1928). (74) 11. G. Bott, E. L. Hirst and J. A. B. Smith, J . Chem. Soc., 658 (1930). (75) J . Pryde, E . L. Hirst and R. W . Humphreys, J . Chem. Soc., 127, 348 (1925). (76) H. B. MacPhillamy and R. C. Elderfield, J . Org. Cheni., 4, 150 (1939). (77) F. G. Young and R. C. Elderfield, J . Org. Chem., 7, 241 (1942). (78) 0. T . Schmidt, E. Plankenhorn, and F. Kiibler, Ber., 76,579 (1942). (79) P. A. Levene and J. Compton, J . Biol. Chem., 114, 9 (1936). (80) P. A. Levene and I. E. Muskat, J . B i d . Chem., 106, 431 (1934). (81) R. E. Gill, E. L. Hirst and J. K . N. Jones, J . Chern. SOC.,1025 (1946). (82) Elizabeth E. Percival and E. G. V. Percival, J . Chcm. Soc., 690 (1950). (83) F. Brown, L. Hough and J. K. N. Jones, J . Chem. Soc., 1125 (1950). (84) W. N. Haworth, E. L. Hirst and E. J. Miller, J . Cheni. SOC.,2469 (1929). (85) R. S. Tipson, C. C. Christman and P. A . Levene, J . Biol. Chem., 128, 609 (1939). (86) E. L. Hirst, L. Hough and J . K. N . Jones, J. Chem. SOC.,3145 (1949). (87) R. S.Tipson and P. A. Levene, J . Biol.Chem., 130,235 (1939). (88) E. L. Hirst and Sonia Dunstan, J . Chem. Soe., 2332 (1953). (89) R. L. Whistler and H. E. Conrad, J . Am. Chem. Soc., 76, 3544 (1054). (90) E. L. Hirst and A. K. Macbeth, J . Chem. SOC.,22 (1926). (91) T. Purdie and C. R . Young, J . Chem. Soc., 89, 1194 (1906). (92) W. Baker, R. Hemmingand W. D. Ollis, J . Chem. SOC.,691 (1951). (93) Joyce W. E. Brading, Millicent M. T. Georg-Plant and Doreen M. Hardy, J . Chem. Soc., 319 (1954). (94) J. Avery and E. L. Hirst, J. Chem. Soe., 2466 (1929). (95) G. H. Beaven, E. L. Hirst and J . K. N. Jones, J . Chem. SOC.,1865 (1939). (96) 0. T. Schmidt and E. Wemiclre, Ann., 668, 70 (1947). (97) I. D . Lamb and S. Smith, J. Chem. Soc., 442 (1936).

272

G E O R G E G. M A H E R

(98) F. Rebcr a i d T. Iteichstcio, H e l v . (‘him. Acta, 29, 343 (1946). (99) 0. T. Schmidt, and E. Wernickc, Ann., 666, 179 (1944). (100) 0. T. Schmidt, W. &layer and A . Distelniaier, Ann., 666, 26 (1943). (101) C.Tainni, Helv. ( ’ h i m . Acta, 32, 163 (1949). (102) J. Conchie and 15. G. V. I’ercival, 1.(:hem. Soc., 827 (1950). (103) J . Fried and Doris E. Walz, J . Btn. (,’hetu. SOC.,74, 5468 (1952). (104) J. Fried and H. E. Stavely, J . A m . Chett,. Soc., 74, 5461 (1952). (105) P. A. Levene and J . Cornpton, J . Biol. Cheni., 116, 169 (1936). (106) P. A. Levene and R. S.Tipson, J . B i d . Chem., 97, 491 (1932). (107) P. A. Levene and R. S. Tipson, Science, 74, 521 (1931). (108) (a) I(.Butler, P. E’. Lloyd and M. Stacey, Chemistry ck Zrrdiastry, 107 (1954); (1)) J . Cheni. SOC.,1531 (1955). (109) R. A . Laitflaw, J . Cheni. Soc., 752 (1954). (110) P. Andrews, L. Hough and J. K. N . Jones, J . A m . Cheni. Soc., 77, 125 (1955). (111) R. L. Whistler and D. I. McGilvrsy, J . Atti. Cheni. Soc., 77, 1884 (1955).

THE METHYL ETHERS OF D-GALACTOSE

BY GEORGEG. MAHER Rescorch Laboratories, Clinton Foods I n c . , Clinton, Iowa

The following revised set of tables of the physical characteristics of the methyl ethers of mgalactose and their derivatives replaces those of D . J . Bell, Advances in C'arbohzJdrate C'hem., 6, 11 (1951), and brings the subject u p t o date.

TABLE I The Mono-0-niethyl-o-galactoses Substance

.&felling poinf, "C.

[ulo, degrees

Rotation solvent

Referncesi ~

147-149 145-148 147

1,6-Anhydr0-3,4-0-isopropylidene-20-methyl-D-galactose 2-O-Methyl-N-phenyl-~-galactopy- 165 ranosylamine Methyl 2-O-methy1-01-n-galactopysirup ranoside Methyl 3,4-0-isopropylidene-2-077-78 methyl -01 -D -galactoside 6-p-toluenesulfonate 90 Methyl 2-O-methyl-p-~-galactopy- 131-132 ranoside Methyl 3,4-0-isopropylidene-2-075-76 methyl-p-D-galactoside Methyl 4,6-0-beiizylidene-2-0-methyl- 152 01 -1) -galactoside 3 -p-toluenesulf onate 145 Methyl 4,6-0-benzyliderie-2-0160 mathyl-8-D-galactoside 169-170 3-p-toluenesul fonate 126 2-O-h~ethyl-~-galactonolactone sirup 2-0-Methyl-~-galactonamide sirup 2-0-Methylgalactaric acid, diamide 195

+53 -+ +82.6 +52 --+ +94 +49 --t +80 -84.5

H*0

h20 EtOH

1 2 3 2

2

+180

McOH

+157.4

4

4 4

+90.9 +1.69

1120

1, 4

+7.16

CHCls

1

S131.6

4, 7

+158.4 -32.8

4

-59.4 +38.4 -27 -+ 27

+

4

chc13 -24

t All references appear in a separate list starting 011 page 281 273

H20

HzO H20

1 4 3 3 6

-

Me[tins pDin!, "c.

Subslance

[WID,

Rotation solvent

degrees

!cfer!nces ~

3-0-Me thyl-or-D-galactose

144-147 +150.6 + +108.6 178-184 C17.2 176-179 +63.5 +31.9 sirup

phenylosazone

H2O

7

McOH

7

C gH sN H,O

Methyl 3-o-methyl-&~-galactopyranoside Methyl 4,6-di-O-benzyl-3-O-methyl-a174-175 +188.6 f 2 D-galactoside 9 , anomer 210 $33.0 f 2 4-0-Methyl-8-D-galactose 207 +62 + +92

CHC13

CHC13 H?O 218-221 +61+ f83 f 2 H?O

phenylosazone

150 148-150 4-O-Methyl-N-phenyl-~-galactosyl- 168 167-168 -84 + -39 f 1 amine 182-183 128 [ale7so +I44 +77 113-114 +137.2 + +77 +120.0 -+ +70.0 118 122-123 +112.0 + +66.0 +22.4 + +13.6 117.5 phenylhydrazone +24.4 -+ f14.4 179 +23.5 + +14.8 182-183 [alsw~f14.5 +141.0+ +91.8 200 phenylosazone 200,201 +144, +141 201-203 20.1-206 Methyl 6-O-methyl-8-~-galactofuranosirup side 6-O-Methyl-~-galactonicacid 6-0-Methyl-o-galactonolactone 6-O-Methyl-~-galactonicacid, phenylhydrazine salt

1, 8 9 7 7

7 3 10 3,11 12 3

MeOH

10

€120

10 13

H2O HzO H2O CsHsN C SH5N CsHsN CsH5N C sH SN C5HsN

CeHsN HzO

9 14 14 9

14 14 13 9 14 24 13

9

(pure?

-5.5 + -40.2 -43 4 -40 158-159 tor15780 4.7 156

sirup

+

HzO H2O

13

HzO

13

14

-

If heating is started a t 175",the higher mdting point is obtained.

TABLEI1 The Di-0-methyl-D-galactoses Rotation solvent

Substawe

~

sirup sirup 2,3-Di-O-methyl-N-phenyl-~-galactos130-1 ylamine 128-9

_

+64.7 4 +80.9 +17.2;+11.3

H2O CHCI,

+11B

H?O

$119.4

EtOH

154-5 -56.8 -+ +12.1 +173.7 $167

EtOH CHCl3 CHCla

Methyl 2,3-di-O-methyl-oru-~-galacto- sirup sirup pyranoside

274

References

_ 9 9, 8

77 8 9 77 8 77

_

-

TABLE I1 (Continued) Subslance

Veiling >in/,"C

Methyl 4,6-0-benzylidene-2,3-di-O- 123-4 128-0 methyl-a-D-galactoside

[a]~ degrees ,

+173.7

126.5- t170

Rolnlion solvenl

Reference.s

3 7

77

127 /3 anomer 2,4-Di-O-methyl-~~-~-galactose monohydrate

+18.0 148 18-00 +130 4 +85 +122 + t85.6 103 +113 -+ +85 105108 f 2

100-

103 104106 33-96 104, 105 2,4-Di-O-methyl-N-phenyl-~-galac 198 tosylamine

f85.0

75

204,

165166 162 +5 113 +162.2 + +52.6 +105 ---t +46.8 163165 165 +59 167 183 106108 119120 119-

10

19

- 180 216 219- +30 f 3 220 218228 4-142 Methyl 2,4-di-O-methyl-a-~-galacto-105

2,4-Di-O-methyl-~-galactonic acid, phenylhydrazide 2,6-Di -@methyl -O-D -galactose

15 11

t85.7

207 209210 216, 214217

side 0 anomer

1

16 3,

6, 17 17 18, 3 16

11, 19, 20 21, 35 75 10 7

22 11 11 3

11 19 16 19 11 11 23

24 15

120

128130 275

1

TABLE I1 (Continued) Substance

monohydrate

Melfing Noinf, “C

[a]~ degrees ,

Rolalion solvenl

128, 134 90,

Re/erences

25, 18 25, 18

98-

100 2,6-Di -0-methyl-N-phenyl-D-galacto- 121pyranosylamine 122 Methyl 2,6-di-O-methy1-8-o-galacto- 73-75 pyranoside 72 67-69 4546.: Methyl 3,4-O-isopropylidene-2,6-di-Omethyl - a --galac ~ t oside 8 anomer 5657, 55 2,6-Di-O -methyl -D -gal ac t onic acid 2,6-Di-O-methyl-~-galactonolactone sirup 2,6-Di-O-methyl-D-galactonamide 154155 2,6-Di-O-methyl-~-galactonic acid, 140 phenylhydrazide 140 3,4-Di-O-methyl,B-o-galactose 164166 Methyl 3,4-di-O-methyl-8-~-galacto- 102side 103 3,4-Di-O-methyl-o-galactonolactone sirup 3,4-Di-O-methyl-D-galactonamide 172174 4,6-Di-O-methyl -a-D -galact ose 131133 hydrate 105 phenylosazone 160162 153 159160 158 4,6-Di-O-methyl-N-phenyl-~-galac- 207 tosylamine Methyl 4,6-di-O-methyl-@-~-galacto- 140 side Methyl 2,3-di-O-benzyl-4,6-di-O68-69 methyl-8-n-galactoside Methyl 4,6-di -0-methyl-2,3-di-Osirup tosyl-p-D-galactoside 4,6-Di-O-methyl-~-galactonolactone

+15

EtO H

24

-24 +2 -22 -27 -23.3

CHClx HzO CHC13 CHC13 CHC13

23 23 24 26 1

+155

Hz0

4

-4.46

chc13

1, 24

HzO Hz0 H20

24 24 24

EtO H HzO

23 24 27

chc13

27

HzO

27 27

HzO

28

HzO EtO H

5 28

EtO H

5 4

-25 - 174

EtO H c6H 6N

29 5

-41.5

chc13

28

4,6-Di-O-methyl-~-gaIactonamide, hydrate

f54

164

276

+26 -49 +46

-24

-+

-44.8 +95

4

+117

-9.1 $89

+

$133

+7

---t

+76.9

+123 + +82 +51 -21 -+

+60

---t

-24

(?I CHCI,

f-3.05

28

(?)

CHC13

+5, +6

28

(?)

+91

+78

-+

+45

HzO MeOH Hz0

5 5 5

277

METHYL ETHERS OF D-GALACTOSE

TABLEI11 The Tri-0-niethyl-r-galacloses Substance

Yelling poinl, "C.

75-80 85 78-79 82-83 86 83 monohydrate 80 80 75 2,3,4-Tri-O-methyl-N-phenyl-r 164,166 165-167 galactosy lamine 167 167-168 169 1,6-Anhytlr0-2,3,4-tri-O-methyl-61 8-r-galactose 2,3,4-Tri-O-rnethyl-6-O-trityl-r-72-75 galactose 1-acetate 93 Methyl 2,3,4-tri-O-rncthyl-~u-u- sirup galactoside sirup 30

Methyl 2,3,4-tri-O-methyl-6-0trityl-r-galactoside 2,3,4-Tri-O-methyl-r-galactonic 107-108 acid phenylhydrazide 166-167

[ a ] degrees ~,

Rolation solvenl

+150.2 4-121.0 +154.07 + +122.02 +147 + +120.4 +156.0 + +119.1

IIzO H?O

30 30a

HzO

f114.5 $135 +152 + +114 +150 + +I14

H,O

-65

MeOII

31 32 2 33 11 2 33 23, 34 33 11 21, 37 2 2

--f

--f

+43

+

HzO HzO H?O

-69.2

EtOH

+33.3

EtOH

f132.2 +112.5 f198.4 f160.8 f150.1 +113.6 +44

MeOH H?O HzO MeOH

f29.8

HlO

4-29.5

slcohol

H,O alcohol

CHCI,

165-167 175-176 2,3,4-Tri-O-mcthyl- ~ - g a l n c t o n o 1ac t one

2,3,4-Tri 0-methyl-r-gnlactonam- 165 itle 2,3,4-Tri-0 methl lgalartaiic acid 100-101 Dimeth? 1 2,3,4-tii-0-mrthylgn98 1:tct:Llnte 100- 101 102-103

Rejtrences

+80 +92

f134 +135 +32 +42 +35 +28 +36

+

$1'3

--f

---f

+24 +25

H,O 1-120

HZO H,O H?O

\fe&O H,O [I20

[LO

37a, 38a 38 30 37h 32 32 37n 37a 11 37a, 38n 37a, 38a 39 2, 11 39 37a, 38a 11 2 11 74 39 40, 7 t 2

278

GEORGE G. M A H E H

TABLE 111 (Continued) Subslunce

Melting point, “C.

[=ID,

degrees

Re/erences

~

2,3,5-Tri-O-methyl-~-galactose sirup -5, -8 Methyl 2,3,5-tri-O-methyl-~-ga-sirup -55 lactoside 2,3,5-Tri-O-methyl-~-galactono- 90 -37 4 -32 91-02 lactone - 35 2,3,5-Tri-O-methyl -D-galactonam- 152 +3 ide 162-163 +5 2,3,5-Tri-O-methyl-~-galactonic 144 18 acid,phenylhydrazide 2,3,6-Tri-O-methyl-~-galactose sirup 87 +77.3, +78.0 +35.4, +35.7

+ +

2,3,6-Tri-O-methyl-~-galactonic 07-98, acid 98

08 2,3,6-Tri-0-methyl-D-galactono- 101 lactone

+80

4

-37

07-98 07-98 2,3,6-Tri-O-meth~l-u-galacto1~~tm135 ide 129-130 2,3,6-Tri-O-methyl-r-galactonic 175 -62.4 acid,phenylhydrazide 2,4,6-Tri-O-methyl-~-~-galactose 102-105 +121 +00.4 104-105 +124 +93 2,4,6-Tri-O-methyl-N-phenyl-~-179 -92 + +38 galactosylamine 180 -96.1 4 f40

-

--f

idc

41

42 41 43 44 44

45 48 47

09

172, 175, 177, 179 170 165-166 Methyl 2,4,6-tri-O-methyl-c-~-73-74 galactoside 62-61 112 3-p-toluenesulfonate hydrate 37 hqcthyl 2,4,6-tri-O-rncthyl-l-1)- 111-112 galactoside 102 83-85 hemihytlratc 130 3-p-tolu~1icsulfonntc 2,4,6-Tri-0-methyl-~-gazlactono-sirup lactone 2,4,6-Tri-O-methyl-~-gal~tctonam167

41 42

45 25, 46

-40 -44

41, 42 41

43 49 15 47 15 47 ‘4

29 5 75 17, 18

+107 +150 +101

50 51 35 16 4 29 4 29

-40.9

4

+ 18

3

9

+163.!)

3 +20.4 +152

+74

4

+50

20 29

279

METHYL ETHERS OF D-G.4LACTOSE

I

Suhsluiice ~~

[mlu. degrees ~

Rnla/ioii sohenl

Rrjerencm

~

3,4, 6-Tri-O-methyl-u-galactose sirup 1,2-0-Isopropylidene-3,4,6-tri-O- sirup methyl-D-galactose 3,4,6-Tri-O-methyl-~-galactono-sirup lactone

-4.3 -31.4 +46.8 +104.8

+

$24.8

[nlo, degrees

Substance

2,3,4,6-Tetra-O-methyl-a-~-galac-71-73 tose 72 69 70-72 70-72

72

75 2,3,4,6-Tetra-O-methyl-p-~-galac-sirup tose

f149.4 -+ +116.9 +142 -+ +118 +150 -+ +114 +142 -+ f117 +162 + +138 +117.3 +118 +110, +112 (pure?) +lo2 + +83 +102.2 -+ t109.5 +58.3 + +62.6 +73 -+ +90.0

2,3,4,6-Tetra-O-methyl-N-phenyl- 186-188 D-galactosylamine 18% 190, 190 -68 +41 191 -80 +44 191 -75, -76, 192 -77.1 + +37.7 -83.3 +40.7 192 192-194 -64 + +36, +41.5 -80, -77 193194, 194 -84 197 193, 194, -+

-+

--f

RIeO I1 McOII

52 52

HrO HrO

52 53

Rolelion

Rejerences

solvenl

73 56 57 40

58 55 59, 68 60 58 11 54 54 54 51 15,60, 61 G2 62 29,37, 50 63 64 21, 40 7

24, 76 57 5, 16, 18,

TABLEI V (Continued) ~~

.Welting point, T.

Substance

[=ID,

degrees

Rolation solvent

195, 106, 107

hIethgl 2,3,4,6-trtra-0-methvl-a 1)galactoside Me thy1 2,3,4,6-trtra-0-met hgl -p gals ct osicle

D-

2,3,4,6-T~tra-O-rneth\.l-n-g,zlnctonic acid 2,3,4,6 Tetra 0 - m r t 1ivl-D -gal,zrt onolactonr

2,3,4,6-Tetra-O-meth~l-1-gal~rtonamide 2,3,4,6-Tetra-0-methyl-I-galactonic acid, phenylhydrazide 2,3,5,6-Tetra-O-methy1 -o-galactose Methyl 2,3,5,6-tetra-O methyl-ngnlactoside 2,3,5,6 Tetra-0-metli?.l-u-galnctoriolart one

2,3,5,6-Tetra-O-methyl D-galactonamide

190 192, 193, 197 202 sirup

48-49 48-48.5

84

sirup

160-161 121 120 119-120 135-137 sirup sirup sirup

153

280

- 141

C SHsN C,€I sN

-110, -141

t188.5 t 190 t148 t18.7 tl9.6 t4.4 -24.4 f22.6 + +26.6

EL0

f155 + +27.4 t160.7 + +47.6 +157 + +26.1 +166.5 + +26.2 +153 +106.7 -+ t16.8 +lo1 f96 128

H?O

HzO MeOH IIzO H2O MeOH

H?O

HsO

Re/erenres

19, 22, 259 59 9 73 35 36, 57, 58 56 72 55 72 55 56 56 72 66 66 55

Hz0

68 69

H2O HzO

70 71

+

CHCl, EtsO C6H6

+35.7 +37.9

Me2C0

70 70 70 53 55 67 53 68

-21.2 -45.2 -46.3 -29.5 -27.1 -34 - 13 - 11 - 11 +6.53 +5.79

MezCO

H2O

HzO +

--+

-26.9 -25.2

EtOH H2O HzO H20 CHCl,

EtzO C6H6 Ha0 MezCO

66 66 66 71 66 70 70 70 70 67 67

METHYL ETHERS O F D-GALACTOSK

28 1

References (1) J . W. H. Oldham and D. J . Bell, J . Am. C'hern. Soc., 60, 323 (1938). (2) D. McCreath, F. Smith, E. G. Cox and A . I. Wagstaff, J. C'heni. Soc., 387 (1939). (3) E. L. Ilirst and J. I(. N . Jones, J. (Them. Soc., 506 (1946). (4) D . J . Bell and S. Williamson, J . Chenc. Soc., 1196 (1938). (5) E. L. Hirst, and J. I(. N. Jones, J. Cheni. Soc., 1482 (1939). (6) F. Brown, b;. I,. Hirst and J . K. N . Jones, J . Cheni. Soc., 1761 (1949). (7) F. Reber anti T. Reichstein, Helv. Chink. Acta, 28, 1164 (1945). (8) G. J. Robertson and R . A. Lamb, b . Chem. Soc., 1321 (1934). (9) E. P:tcsu and 6 . M. Trister, J . Am. Chem. Soc., 62, 2301 (1940). (10) R. W. .Jeanloz, J . Ani. Chenr. Soc., 76, 5654 (1954). (11) F. Smith, J. Chent. Soc., 1724 (1939). (12) E. G. V . Percival and G. G. Ritchie, J . Cheni. SOC.,1765 (1936). (13) K. Freudeiiberg and K. Smeykal, Ber., 69, 100 (1926). (14) J . Munro and E. G. V. Percival, J . Chem. Soc., 640 (1936). (15) P. Andrew, 1,. Hough and J . K. N. Jones, J. Cheni. Soc., 806 (1954). (16) G. 0. Aspinall, E. L. Hirst, R. W. Moody and E. G. V. Percival, J . Cheni. Soc., 1631 (1953). (17) F. Brown, E. 1,. Hirst and J . K. N. Jones, J . Chenz. Soc., 1757 (1949). (18) J. K. N. Jones, J. Chenk. soc., 3141 (1949). (19) E. Baldwin and D. J. Bell, J . Chena. Soc., 1461 (1938). (20) E. V. White, J. A m . Cheni. Soc., 68, 272 (1946). (21) M. 1,. Wolfrorn, G. Sutherland and M. Schlamowitz, J . Am. ('hem. Soc., 74, 4883 (1952). (22) W. H. Wadman, A . 1%.Anderson and W. Z. Hassid, J . A m . (:hem. Soc., 76, 4097 (1954). (23) D. J. Bell, J. Cheni. Soc., 692 (1945). (24) E. T. Dewar and E. G. V. Percival, J. (Them. SOC.,1622 (1947). (25) L. Hough and J . K. N . Jones, J. Cheni. Soc., 1199 (1950). (26) R. E. Reeves, J . Am. Chem. Soc., 71, 1737 (1949). (27) J . S. D. Bacon and D . J. Bell, J . Chem. SOC.,1869 (1939). (28) J. S. D. Bacon, D. J. Bell and J . Lorber, J . Chem. SOC.,1147 (1930). (29) E. G . V . Percival and J . C. Somerville, 1.Chen2. Soc., 1615 (1937). (30) P . A. Levene and L. C. Kreider, J . Biol. Chem., 121, 155 (1937). (30a) M. Onuki, Proc. I m p . Acad (Tokyo), 8,496 (1933), as quoted by Levene and &eider (30). (31) P. A. Levene, R. S.Tipson and L. C. Kreider, Science, 86, 332 (1937). (32) .'1 A. Levene, R. S. Tipson and L. C. Kreider, J. Biol.('hem., 122, 199 (1937). (33) It. A. Laidlaw and Clare B. Wylarn, J. Chem. Soc., 567 (1953). (34) M. Abdcl-Akher, F. Smith and D. Spriesterbach, J. Chem. Soc., 3637 (1952). (35) I. Ehrenthal, M. C. Rafique and F. Smith, J . Am. Che7)i. Soc., 74,1341 (1952). (36) Z. F. Ahmed and R. L. Whistler, J . Am. Chent. Soc., 72, 2524 (1950). (37) R . I,. Whistler and H. E. Conrad, J . Am. Chem,. Soc., 76, 3544 (1954). (37a) M. Onuki, Brill. Inst. PhzJs. Cheni. Research (Tokyo), 12,, 614 (1933); Sci. Papers Insf. Phys. ('hem. Research (Tokyo), 20, 234 (1932-33). (37b) M. Oiiuki, Bull. Inst. Phys. Che7n. Research (Tokyo), 12, 614 (1933), as quoted by Levene and Kreider (30). (38) M. Onuki, J. Agr. Chem. Soc. Japan, 9, 90 (1933); Chem. Abstracts, 27, 2138 (1933).

282

GEORGE G . MAHER

(38a) M. Onuki, J. Agr. Chem. SOC.Japan, 9, 90 (1933), as quoted by H . Elsner, "Kurzes Handbuch der Kohlenhydrate," Johann Barth, Leipzig, 1935, p. 356. (39) S. W. Challinor, W. N. Haworth and E. L. Hirst, .I. Chem. SOC., 258 (1931). 189 (1954). (40) E. L. Hirst, E. G. V. Percival and Clare B. Wylam, J . Chem. SOC., (41) Sybil P. Luckett and F. Smith, J . Chem. SOC.,1114 (1940). (42) B. H . Alesander, R. .J. Dimler and C. I,. Mehltretter, J . Am. Cheni. SOC.,73, 4658 (1951). (43) W. N. Haworth, H. Raistriek and M. S t a c y , Biocheni. J . (London), 31, 640 (1937). (44) S. Murakami, Acta Phylochinc. (Japan), 11, 201 (1940); Chem. Abstracts, 36, 1384 (1941). (45) J. J. Connell, Ruth M. Hainsworth, E. L. Hirst and J. K.N. Jones, J . Chem. SOC., 1696 (1950). (46) 15. 1,. Hirst, I,. Hough and J. K. N. Jones, J . Chem. Soc., 3145 (1949). (47) W. K.IIaworth, H . Raistrick and M. Stacey, Biochem. J . (London), 29,2668 (1935). (48) W. N . Haworth, E. L.Hirst and M. St>acey,J. Chem. SOC.,2481 (1932). (49) E. Pacsu, S. M. Trister and J . W. Green, J . Am. Chem. Soc., 61, 2444 (1939). (50) E. G. V . Percival and I. C. Willos, J . C h e n i . SOC.,1608 (1949). (51) R . Johnston and E. G. V. Percival, J. Chem. Soc., 1994 (1950). (52) P. A. Levene and G. M. Meyer, J. B i d . Chenz., 92, 257 (1931). (53) A. M. Gakhokidee and h'.D. Kutidze, Zhur. OhshcheZ K h i m . , 22, 139 (1952); Chenz. Abstracts, 46, 11116 (1952). (54) J. C. Irviiie arid A. Cameron, J . Chein. Soc., 86, 1071 (1904). (55) J. Pryde, E. I,. Hirst and R . W. Humphreys, J . Chem. SOC.,127, 348 (1925). (56) H . H. Schlubach and K. Moog, Ber., 66, 1957 (1923). (57) E. W. Putman and W. Z. Hassid, J . Am. Chem. Soc., 76, 2221 (1954). (58) M. C. Rafique and F. Smith, J . A m . Chem. Soc., 72, 4634 (1950). 76, 2617 (1953). (59) E. V. White and P. S. Rao, J . Am. Chem. SOC., 74,4029 (1952). (60) P. Andrew, L. Hough and J . K. N. Jones, J . Ant. Chem. SOC., (61) P. Andrew, L. Hough and J . K . N. Jones, J . Chem. SOC.,2744 (1952). (62) G. J. Lawson and M. Stacey, J . Chem. Soc., 1925 (1954). 97, 1449 (1910). (63) J. C. Irvine and D. McNicoll, J . Chem. SOC., (64) W. N. Haworth and Grace C. Leitch, J . Chem. SOC., 113, 188 (1918). (65) D. J . Bell, J . C'hem. SOC.,1543 (1940). (66) W. N . Haworth, D . A. Rue11 and G. C. Westgarth, J . Chem. Soc., 126, 2468 (1924). (67) It. W. Humphreys, J . Pryde and €3. T. Waters, J . Chem. Soc., 1298 (193i). (68) W. N . Haworth, E. L. Hirst arid D. I. Jones, J . Chem. Soc., 2428 (1927). (69) H. D. I(. Drew, E. H. Goodyear and W.N. Haw-orth,J. Chem.Soc., 1237 (1927). (70) W. N . Haworth, E. L. Hirst and J. A. B. Smith, J . Chem. SOC.,2659 (1930). (71) J . Pryde, J. C h e m . SOC.,123, 1808 (1923). (72) F. Micheel and 0. Littmann, A n n . , 466. 115 (1928). (73) W. Charlton, W. N . Hawort,h arid W. J. Hickinbottom, .I. Chem. SOC.,1527 (1927). (74) P. A..I,evene and L. C. Kreider, J . B i d . C'hem., 120, 597 (1937). (75) D. J. Bell and E. Baldwin, J. Chem. SOC.,125 (1941). (76) R. A. Laidlaw, d . ('hem. Soc., 752 (1954). (77) D. J. Bell and '2. D. Greville, J . Chem. Soc., 1136 (1955). For the 1,4-lactone of 2,3-di -O-methyl-wgalacttonic acid, these authors give [& -48.8 -36.2' (18 mins. -+ 270 hrs.; HSO); for the amide, m. p. 140"; [elDf13.1' (HzO). -+

POLYSACCHARIDES ASSOCIATED WITH WOOD CELLULOSE BY W . J . POLGLASE Department of Biochemistry. University of B r i t i s h Colrritthia. Vrrncoirvrr. Brilis h Colunihia Canutln

CONTEXTS I . Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . I1. Carbohydrate Constituents of Wood. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 111. Carbohydrates in Wood Cellulose Preparations . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 . General . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 . Mannan . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . a . Structure of Mannans . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b . Determination of Mannan., . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c Properties of Mannan in Wood Cellulose and Derivatives . . . . . . . . . . . 3 . Xylan . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . a . Structure of Xylans . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b . Determination of Xylan . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c . Properties of Xylan in Wood Cellulose and Derivat.ives. . . . . . . . . . . . 4 . Carbohydrates Containing Carboxyl Groups . . . . . . . . . . . . . . . . . . . . . . . . . . . a Structure . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b . Determination of Carboxyl Groups . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c . Properties of Carboxyl Groups in Wood Cellulose and Derivatives . . . . I V . Preparation and Composition of Wood Cellulose ...................... 1. Holocellulose . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 . Commercial Wood Cellulose . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . a . General . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b Sulfite Pulps . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . c . Alkaline Pulps . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . V . Fine Structure of Wood Cellulose and Associated Polysaccharides . . . . . . . . 1. Alpha-, Beta-, and Gamma-cellulose . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 . Accessibility of Cellulose . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

.

.

.

283 285 287 287 288 288 290 295 299 209 303 307 312 312 314 315 316 316 321 321 322 325 328 328 330

I . INTRODUCTION Cellulose, the chief constituent of all living plants, is composed of Dglucose anhydride units, linked by 4 -+ 1 p-D-glucosidic bonds . Chemists generally agree that the individual molecule is unbranched and is homogeneous with respect to the monomeric unit . Within a single preparation, or in the native state, cellulose is a polymolecular material, and its individual molecules form a polymer-homologous series . Except for this variation in molecular size, cellulose is considered to be a uniform chemical substance . The cellulosic material from which these characteristics have 283

284

W. J. POLGLASI?

beeii deduced is puritied cellulose from cot toil linters.’ Ccllulosc preparations from wood can be shown hy many chemical methods to he lcss homogeneous than cellulose from cotton linters. This is due, in part, to iiicomplete removal (from the wood cellulose preparation) of ligniii, ash, or wood cstractives. Morc important than t hese, in the quantitative sense, are the associated polysaccharides, sylaii and ma1inaii. I n addition, preparations of wood cellulose generally have a higher carboxyl-group content than has linters cellulose, and some of these carboxyl groups are believed to bc present eitjher as polyuronides or as occasiorial uronic anhydride moieties ill the cellulose chain or in associated polysaccharides. In certain preparations of ccllulose, minor amounts of araban and galactan have been detected. Neither in wood nor in any wood cellulose preparation has it been established that the polysaccharides are homogeneous. The use of the terms “xylati,” “mannan,” etc. in wood chemistry, although convenient, does not imply homogeneous polymers of anhydroxylose, anhydromannose, etc., but indicates only that the respective sugars or derivatives thereof may be detected after complete hydrolysis to monomeric units. The polysaccharides associated with cellulose in wood, or wood pulps, are often referred to as “polyoses,” a term first suggested by Staudinger.2 Because of greater homogeneity, both chemical and physical, cottonlinters cellulose is superior to wood cellulose in yielding viscose rayon and acetate yarns of high quality. However, the greater availability of wood cellulose, together with technological advances resulting from constant research, have placed wood cellulose in a strong position as a chemical raw material. Certain inferior properties of yarns from wood cellulose have, from time to time, been ascribed to the presence of polyoses. For example, part of the haziness of wood cellulose acetate may be due to the presence of xylan in the wood pulp. Concurrently with the development of improved types of chemical pulp has appeared the need for increasingly sensitive methods of analysis, in order to permit determination of the purity of a preparation. Physicochemical methods of analysis which yield information concerning degree of polymerization (D. P.), chain-length distribution, and accessibility of wood cellulose have been brought to a high degree of precision and have proved their value. Chromatographic methods are now yielding detailed information regarding the organic chemistry of wood cellulose. It is anticipated that a combination of these methods will eventually provide important data 011 the molecular weight and distribution of the associated polysarcharides, as well as on the nature of the associative forces. (1) E. F. Hinner, in “Cellulose and Cellulose Derivatives,” E. Ott, ed , Interscience Publishers, Inc., New York, N. Y., 1943, p. 519. (2) H. Staudinger and F Reinecke, HoZz Roh- u . Worksto$, 2, 321 (1939).

285

POLYRACCFIARIDER ASSOCIATED WITH WOOD CELLULOSE

I1 is proposed to consider, herein, the carbohydrate coinpositlion of numerous wood-cellulose preparations. The structure of the associated polysaccharides mill be reviewed, as well as current, speculations on the nature of the association between the various components of wood cellulose.

Ir.

CAHUOHYDRATE CONSTITUENTS

OF WOOD

Chemical analyses of wood hydrolyzates have shown that the following simple sugars are present in wood: D-glucose, manno nose, n-galactose, nsylose, and L-arabinosc. Although it has been knomn for some time that each of these sugars o c c w s in wood i n polymeric form, quantitative determination is a recent development3 resulting from the discovery of paper chromatography4 and its application to sugar analysis by P a r t r i d ~ e . ~ , TABLE I Analgsis of Composite Wood Samples? Sfiecies

Glucan,=

Douglas fir Western hemlock Loblollg pine Black spruce Western red cedar Southern red oak

48.3 44.5 46.6 45.6 47.5 43.7

7"

Mannan,h

Xylan,'

?6

?6

5.4 4.1 4.7 8.0 5.1

6.2 7.3 10.1 10.5 8.1 20.0

a Alpha-cellulose corrected for mannan, xylan, and uronic anhydride. ylhydraeone method.8 c Pentosan corrected for uronic anhydride.

Uronic anhydride,

2.8 5.0 3.8 4.1 4.2 4.5

* Phen-

Heretofore] most standard analyses for the carbohydrate components of wood have been performed by a combination of such less specific methods as those for pentosan, hemicelluloses, alpha-cellulose, etc. The carbohydrate composition of a number of wood species is listed in Tables I and 11. It will be noted that the chromatographic procedure of Gustafsson and coworkers3 (see Table 11) gives values for five sugars, but not for uroiiic anhydride. Those species classed as softwoods (coniferous) generally show a higher mannan content and a lower xylan content than do those classed as hard(3) C. Gustafsson, S. Pettersson and T. Lindh, P a p e r i j a P u u , 33, 300 (1951). (4) R. Consden, A . H. Gordon and A. J. P. Martin, Biochem. J . (London), 38, 224 (1944). (5) S. M. Partridge, Nature, 168, 270 (1946); Biochem. J . (London), 42, 238 (1948). (6) C. N. Kowkabany, Advances in Carbohydrate Chem., 9, 303 (1954). (7) L. E. Wise and E. K. Ratliff, A n a l . Chem., 19, 459 (1947). (8) E. Hagglund and 1,. C. Bratt, Papier-Fabr., 34, 100 (1936).

286

W. J. POLGLASK

Cluian,

Specrer

__

5

Spruce (I’zcca excclsa) Pine (Pinus silvestris) Cedar ( J u n i p e r i s cornrnunis) Birch (Betula verrucosa) I3irch (Betula pubescens) Oak ( Q u c w u s robur) Ash ( F r a z i n u s excelsior)

~

j

dlannan,

/n

C‘

-~

.

~

65.5 65.0 61 . 0 58.5 55.0 68.5 60.0

16.0 12.5 14.0 0.5 2.5 2.0 2.5

Sylun, c,;

__

9 .0 13.0 11.o 30.0 30.0 2G. 0 32.0

Galnrlan,

“6

6.0 6.0 13.5 1.5 1.0 2.5 3.0

Araban,

7%

____

3.5 3.5 0.5 0.5 2.5 1.0 2.5

a Determitied by hyclrolysis and paper chromatography ; results expressed as percentages of total sugar.

woods (deciduous). The softwoods contain more galactan than do the hardwoods, and, in both classes of wood, the araban content is relatively small. All woods contain uronic arid anhydride moieties which may be preseut either i n the hemicelluloses or in the pectic substances of the wood. [Jronic acids, or polyuronides, are usually determined by measuring the carbon dioxide evolved when the sample is heated with 12 % hydrochloric acid under standardized condition^.^ By this method, only the total quantity of uronic anhydride in the wood has been determined, with few studies being made on the structures of the monomers. However, itJ has been reported’” that aspen-wood hemicellulose contains both D-galacturonic and 4-O-methyl-~-gliicuroiiic acid residues. This positive identification of 4-0methyl-D-glucuroiiic acid in aspen wood is in agreement with the earlier studies of Sands and Gary“ on mesquite wood. The hemicellulose from this wood contained uronic anhydride which could not be converted to galactaric or D-glucaric acid derivatives by hydrolysis and nitric acid oxidation. Sands and Gary suggested that the uronic acid contained a methoxyl group, by analogy with the previous observations of Anderson and Kinsman.’* The latter workers had been able to crystallize small amounts of potassium acid D-glucarate from mesquite gum after hydrolysis and oxidation with nitric acid. The mother liquors from the crystallization contained methanol, and these facts seemed in accord with the idea of a methoxyl-containing, uronic acid moiety. The hemicelluloses studied by Sands and Gary” and by Anderson and (9) 13. 1,. Browning, in “Wood Chemistry,” I,. E. Wise and E. C. Jahn, eds., Reinhold Publishing Corp., Xew York, N. Y., 2nd Edition, 1952, p. 1206. (10) (a) J . K. N . Jones and I,. E. Wise, J . Chem. Soc., 2750 (1952); (b) ibid., 3389 (1052). (11) Lila Sands and W. Y. Gary, J . Biol. Chem., 101, 573 (1933). (12) E. Anderson and S. Kinsman, J . Biol. Chem., 94,39 (1931).

POLYSACCHARIDES ASSOCIATED WITH WOOD CELLULOSE

287

OtisL3contained substantial amounts of a “body X” or “substance X,” which resisted hydrolysis by acid and was obtained as an acid-insoluble residue. Norman14suggests that this substance may have been lignin associated with the extracted hemicellulose, although the possibility that it is a refractive, carbohydrate substance has not been excluded. The “mcthylpentose” L-rhamnose has been isolated from aspen wood by Jones and S c h ~ e t t l e r who , ~ ~ state that it occurs to the extent of less than 1 %. This sugar was detected by a chromatographic procedure applied to a dilute-acid hydrolyzate of aspen-wood meal. Extraction of several paper chromatograms then yielded sufficient L-rhamnose for identification hy optical rotation arid by characterization as the benzoylhydrazone. I n future studies, small amounts of other sugars or sugar derimtives may be found in wood, as a result of the application of increasingly sensitive techniques of analysis. I t is probable that most of these mill be found in readily accessible, easily hydrolyzed polysaccharides or in glycosides of the plant tissue, and will not, normally, be constituents of an extractive-free, delignified, mood-cellulose preparation. Pectic substances are found only i n relatively small amounts in wood, and are probably removed by most woodpulping processes. The sugar residues of glycosides, similarly, would be expected to be hydrolyzed off and dissolved during pulping. Indeed, most processes for preparing wood cellulose may be regarded as severe chemical treatments, of relatively low specificity, which remove by a variety of reactions the most reactive and soluble constituents of the wood, leaving behind the inert, insoluble, fibrous wood cellulose. I N WOODCELLULOSE PREPARATIONS 111. CARBOHYDRATES

1. General

The carbohydrate cornposition of a preparation of wood cellulose depends, first, on the composition of the wood, and second, on the process used to remove lignin. It is improbable that processes for preparing wood cellulose affect the internal monomeric units of the polysaccharides, with the possible esception that random oxidation of carbohydrates may occur to some extent. The final carbohydrate composition of wood cellulose will, then, be the sum of the carbohydrates of the wood less the amount of carbohydrate dissolved or degraded during delignification. i\ completely specific delignification process should leave a residue containing all of the wood polysaccharides. Most delignification procedures are, however, not specific, arid the more soluble or more easily hydrolyzed polyoscs are affected to (13) E. Anderson arid Louise Otis, .I. Am. (‘hem. Soc., 62, 44G1 (1930). (14) A. G . Norman, “Biochemistry of Cellulose, the Polyuronides, Lignin, rtc.,” Clareiidon Press, Oxford, 1937, p. 49. (15) J. K N. Jones and J. R. Schoettler, Tappi, 35, 102A (1952).

288

W. J. POLGLASE

some extent. In commercial processes, extensive dissolution or degradation occurs, and only the more stable polysaccharides are retained in the pulp. Whereas holocellulose preparations contain most of the wood carbohydrates, commercial wood-cellulose preparations contain only xylati, mannan, and carboxyl-contaiiiitig polyoses in addition to the cellulose compoiient. Certain less-refilled ~~--ood-cellulose preparations may contain trace amouuts of araban and galactan. There is also the possibility that n-glucose polymers other than cellulose will be present in some wood pulps. Most of our knowledge of the chemistry of the polyoses in wood-cellulose preparations comes from studies made on fractions of the hemicelluloses obtained from alkaline extracts of n.ood, agricultural residues, or other plant material. In the following, some of these studies will be considered as a starting point, and then the effect of pulping procedures on the amount aud nature of the associated polysaccharides will be discussed. It is not the purpose of this article to treat the chemistry of plant polysaccharides in detail, but rather to consider only those aspects which, in the opinion of the author, are pertinent to an understanding of the factors affecting the composition of wood-cellulose preparations. 2. Mannan

a. Strirctrtre of Mannans.-The chemistry of marinaris has been reviewea recently by Whistler and Smart .16 The mannan obtained from vegetable ivory by extraction with dilute sodium hydroxide solutioii yields two fractions on alcohol precipitation, mannan-h and mannan-B.’7 From methylation studies,Is x-ray examination of the native crystalline m a r i i i a ~ i , ~ ~ solubility characteristics, and viscosity and film properties of nitrated mannan, Meyer and Mark suggestz0for the structure of ivory-nut mantian, a linear chain of D-mannose units linked 4 + 1 P-D. Klages’ methylation studiesz1provide evidence for a polymer consisting of D-mannopyranose units linked 4 -+ 1 for both mannan-A and maiinan-€3, although Klages suggestszLthat both a and P linkages occur in mannan-A. The I). P.s of ivory-nut mannan and of salep maiinanzzappear to be in the range of 70 to SO, from the amount of tetra-0-methy~-D-matinosefound on hydrolysis of the methylated polysaccharides. (16) R 1,. Whistler and C. L. Smart, “Polysaccharide Chemistry,” Academic Press Inr , Ncw York, N. Y., 1953, p 152. (17) K Hess and M Ludtke, A n n , 466, 201 (1027). (18) J Patterson, J (‘hem. Soc., 123, 1139 (1923) (19) R 0 Hrlizog and TI W Gonell, Naliiiwzssenschaften, 12, 1153 (1924). (20) K. 11. Mever a n d €1. &Park, “Der Aufbnu der Hochpolymeren organischen Naturstoffe,” Hirschualdsche Buchhandlung, Berlin, 1930, p. 168. (21) F. Klages, Ann., 609, 159 (1934); 612, 185 (1934). (22) F. Klages and R. Niemann, Ann., 623, 224 (1936).

POLYSACCIIAIZIDER ASSOCIATED WITH WOOD CELLULOSE

289

r 3

i he structure of ivory-nut maniiaii lists hecii reinvestigated fi~annan-A,[cY]'~D - 46" (c, 0.7 in N NaOH) yielded on hydrolysis D-mannose (97.6 %), D-galactose (1.8 %), and D-glucose (0.8%). From methylation studies, it was deducedz3that two types of molecule mere present, both comprising an average of 10 to 13 residues, oiie terminated a t the nonreducing end by a D-mannopyraiiose residue and the other by a D-galactopyranose residue. The majority of D-mannopyranose residues were found to be linked through C1 and C4 but some 6 + 1 linkages were also present.23 Mannan-Ij, [a]I6D -226" (c, 0.8 i n anhydrous formic acid) was found to be similar chemically l o mannan-A, but of greater molecular size (38 to 40 residues). Yeast mannan is an amorphous, homogeneous polysaccharide with a D. P. of about 500.24The proposed s t r u ~ t u r 25 e ~of~ this ~ mannan is a long chain, with radiating side-chains of one or two D-mannopyranose units. In this mannan, 2 -+ 1, 3 + 1, and 6 -+ 1 linkages have been found.26The mannan of the seaweed Porphyra umbilicalis contains chiefly 4 -+ 1 p-D linkages,26and is of the branched-chain type with about one branch for each 12 D-mannose residues. Polysaccharides have been isolated which contain both D-glucose and D-mannose, for example, the glucomannan of Amorphophallus k0njac.2~ From this Konjac mannan ( [ c Y ] ~ ~-42.8") D has been isolated a disaccharide composed of D-glucose and D-mannose units.28The polysaccharide from the corms of Amorphophallus cncophyllus is a fibrous substarice which contains 49 % of anhydroglucose, 41 % of anhydromannose, and 4 % of uronic an. ~ ~ substance is an excellent beater hydride, and yields 2 % of p e n t o ~ a n sThis additive when used in paper manufacture. Jones and coworkers,30in studies on mannose-containing polysaccharides, have isolated, from seeds of I r i s ochroleuca and I r i s sibirica, heterogeneous polysaccharides yielding on hydrolysis D-glucose and D-mannose in approximately equal amounts, as well as D-galactose (ca. 3 %). These workers30 consider the possibility that the polysaccharides are mixtures of glucomannan with galactomannan, but, because of the similarity (in composition) between the two iris polysaccharides, tend to the view that they are (23) G. 0. Aspinall, E. L. Hirst, E. G. V. Percival and I. R. Williamson, J . Chem. Soc., 3184 (1953).

(24) W. N. Haworth, E. L. Hirst and F. A. Isherwood, J . Cheni. Soc., 784 (1937). (25) W. N. Haworth, R. L. Heath and S. Peat, J . Chern. SOC.,833 (1941). (26) J . K. N. Jones, J . Chem. Soc ,3292 (1950). (27) Ref. 16, p. 302. (28) K. Nishida and H. Hashima, J. Dept. Agr. Kyushu I m p . liniv., 2 , 2 7 7 (1930); Chenr. Abstracts, 26, 498 (1931). (29) L. E. Wise, Arch. Biochem., 23, 127 (1949). (30) P. Andrews, L. Hough and J. K. N. Jones, J . Cheni. Sac., 1186 (1953).

290

15'.

J. POLGLASE

single polysadiaritles cwntaining all three sugars. Methylation studies indicated that D-galactose forms the majority of the end-groups, with a lesser number of D-glucose and D-mannose end-groups. The polysaccharide of I . ocliroleiica was helieved to contain 150 to 175 hcxose units, with a11 average of five or six branches per molecule, whereas the polysaccharide of I . sibirica appeared to be linear, or infrequently branched, and of smaller molecular weight. The structure of the manilan, or mannans, in wood cannot be described with certainty. Hess and Ludtkejl isolated a mannan from pine sulfite pulp identical with ivory-nut mannan-A in optical rotation and x-ray diagram. H ~ s e m a n t iprepared, ~~ from spruce hemicellulose, fractions having rotational values in agreement with those for the mannan from pine sulfite TABLEI11 Ovtical Rotations of Mannans Soloent

Malerial

I

[a]D, degrees

1

References

~

Ivory-nut mannan Ivory-nut mannan-A Ivory-nut mannan-B Salep rnannan Pine mannan Spruce mannan Slash-pine mannan Porphyra unibilicalis mannan Yeast mannan

NaOH N NaOH 15% NaOH water (or iV NaOII) K NaOH N NaOH 0% NaOH anhydrous formic acid water

-44.1 -44.7 -3s.2 -35 -44 -40 tjo -44 -41 -41 89

+

34 21 21 22 31 32 33 26 24

pulp.31Of special interest is the crystallization of a mannan fraction from slash-pine holocellulose."J Despite an optical rotation of the same order of magnitude as that of the homogeneous ivory-nut mannan (see Table 111), this crystalline fraction contained only 50% of mannan after repeated recry~tallization.~~ The other sugars presumed to be present in the polysaccharide were not identified. b. Determination of Mannun.-Three methods are now available for the determination of mannan in wood or in wood-cellulose preparations. None of these methods actually determines mannan as a polysaccharide but, rather, determines the quantity of anhydromannose units. The method of determination of mannan in woods, devised by S ~ h o r g e rdepends ,~~ upon (31) (32) (33) (34) (35)

K. Hess and M. Liidtke, Ann., 466, 18 (1928). E. Husemann, J . prakt. Chem., 166, 13 (1940). A. P. Yundt, T a p p i , 3 4 . 9 4 (1951). J. L. Baker and T. H. Pope, J. Chem. Soc., 77, 696 (1900). A . W. Schorger, Znd. Eng. Cheni., 9, 748 (1917).

POLYSACCHARIDES ASSOCIATED W I T H WOOD CELLULOSE

29 1

the hydrolysis of mannan to mannose by means of 5 % hydrochloric acid. The acid solution is neutralized with sodium carbonate and concentrated, and the mannose is precipitated as the phenylhydrazone, according t>othe original method of Bourquelot and HBri~sey.~6 This method was subsequently found to be unsatisfactory for wood, since a n appreciable part of the mannan resists hydrolysis by 5 % hydrochloric acid.37 Hagglund and overcame this difficulty by complete hydrolysis of the polysaccharides, prior t o precipitation of mannose phenylhydrazone. Their method has been modified somewhat by Wise, Ratliff and Browning.39 Complete hydrolysis is accomplished in two steps, by first dissolving the sample in 72 % sulfuric acid and allowing the solution to stand for 3 or 4 hours a t 20”. Then, the mixture is diluted with water, and the hydrolysis is completed by boiling for 4 hours. The solution is next neutralized with barium carbonate, and an aliquot of the filtrate is concentrated to a suitable volume, after acidification with acetic acid. After addition of a known amount of mannose, t o ensure quantitative Precipitation, phenylhydrazine acetate is added, and the precipitated phenylhydrazone is dried and weighed. The determination of mannan is an exacting procedure, and, even when all evident precautions are taken, the results do not possess a satisfying precisi0n.4~ There are two major problems to be considered in mannan determination-first, quantitative coiwersion of mannan to mannose, and second, determination of mannose. It has been reported that, whereas dilut,e acids do not hydrolyze all of the mannan, more concentrated acids, acting for prolonged periods of time, destroy part of the r n a ~ i n o s e .Furthermore, ~~ when the cellulosic sample is completely hydrolyzed, the total amount of sugars is large, and in the case of purified wood pulps, the mannose content may be as litOle as 1 % or less of the total sugars. In such circumstances, it is essential that the method be quite specific for mannose. The manipulations of quantitative filtrat,ion and concentration are troublesome because of the large amounts of barium carbonate required for neutralizing the excess acid. Wise and co~orl<ers3~ recommend the use of a large aliquot of the neutralized hydrolyzate-as much as can be obtained by thorough draining of the barium sulfate precipitate on a Buchner funnel. This aliquot must then be Concentrated (after acidification with acctic acid) to about (36) E. Bourquelot a n d 13. HBrisseg, Compt. rend., 129, 339 (1899). (37) E. Hagglund and F. W. Klingstctlt,, “Hdzchemie,” Akatlemischo Vorlagsges., Leipzig, 1st Edition, 1925. (38) E. IIagglund and L. C. Brat,t, Sverisk Kerri. Tidskr., 48, 125 (1936). (39) 1,. E. Wise, E. I<. Ratliff and 13. I,. Drowiiing. Antrl. C‘hc~vi.,20, 825 (104S). (40) See B. I,. Drowning, Ref. 0 , p . 1183. (41) H. Koch, Pnpier-Fnhr., 39, No. 8, 46 (1941).

292

W. J. POLGLASE

one tenth of its original volume, either on a steam bath or under diminished pressure. Although there is apparently no interference by glucose in the precipitation of mannose phenylhydrazone, xylose does lower the yield,42 so that, particularly in the case of hardwoods and hardwood pulps (which contain large amounts of xylan and only small amounts of mannan), the mannan values are likely to be low. Wise and coworkers3gwash the mannose phenylhydrazone precipitate with a saturated, alcoholic solution of man nose phenylhydrazone, to avoid losses occasioned by the slight but definite solubility of this compound. It is claimed39that the more thorough washing permitted by this technique removes any co-precipitated glucose phenylosazone. To ensure quantitative precipitation, the mannose concentration should not be less than 1 %, and sufficient mannose should be added (in the case of samples low in maiinan) to attain this minimum concentration. With this modified technique, Wise and demonstrated the absence of mannan in cotton linters, and the presence of small amounts of mannan in certain hardwoods for which mannan had been found to be absent by the original Hagg1und-BratV8 method. The use of quantitative, paper chromatography is discussed in this Section, since one of the most important factors in its application to woodcellulose analysis is that it permits a satisfactory determination of mannan. Reasonably reliable methods for the determination of xylan have been available for some time, but maiinan determinations, for the reasons discussed above, have been less satisfactory. The fact that (on chromatograms) uylose, as well as other carbohydrates, can be determined simultaneously with mannose is an added attraction in the use of this technique. Paper chromatography4 as a means of detecting the monomeric units of the polysaccharides of wood and wood cellulose, was first used by Wise, Green and R i t t e ~ i h o u s efollowing ,~~ the techniques developed by Partridges and others6 for the analysis of sugars. These demonstrated that xylose and mannose ran be detected in wood and mood-cellulose hydrolyzates by this technique. In later work, Wise and demonstrated the presence of galactose and arabinose in dilute-acid hydrolyzates of unbleached, sulfate pulps from Western hemlock and Douglas fir. The first quantitative determinations of sugars in wood-pulp hydrolyzates were made by Sundman, Saarnio and G u ~ t a f s s o n . ~ ~ perFor the preparation of wood-cellulose hydrolyzates, these form the hydrolysis essentially as described by Wise and coworkers,39but, (42) H. Eguchi and H Wnda, J . Soc Texhle and Celldose Znd. J a p a ~7,92 , (1951); Cheni ilbstracls, 46, 5810 (1952) (13) L E Wise, ,J W Green aiid R C Rittenhousr, T a p p z , 32, 335 (19-19) (11) I, I< Wise, R C Rittenhouse ;iricl C Garcia, T u p p z , 34, 15 (1951) (15) .J. Sundman, J Pnnrnio nnd C Custafsson, l'aperi J U Put/, B33, 115 (1951)

POLYSACCHARIDES ASSOCIATED WIT11 WOOD CELLULOSE

29.3

in place of barium carboiiatc, use aii anion-exchange column to remove sulfuric acid from the hydrolyzates. The column is theii thoroughly washed with water, and the resulting solution is concentrated, preparatory to analysis by paper chromatography. One disadvantage to this method of preparation of hydrolyzates is the necessity for thorough washing of the ion-exchange resin, so that the hydrolyzed sample is very much diluted and rather large amounts of' water must be removed in subsequent concentration. Thus, for 1 g. of pulp, 10 ml. of 72 % sulfuric acid is added to dissolve the sample; 320 ml. of' water is added prior to refluxing, and GOO ml. of watcr is used to wash the anion-exchange resin. This total (930 ml.) must then be concentrated to 10 ml. before chromatography is undertaken. The hydrolyzates (prepared as described above) are chromatographed, a t a concentration corresponding to 10 % of the air-dried pulp, on Whatman No. 1 filter paper, and are quantitatively estimated by a modification of the method of Flood, Hirst and Jones46and Hawthorne.47This method requires that the separated sugars be extracted from the filter paper and estimated by a reducing-sugar method. The location of the individual, separated sugars is established by reference to a parallel chromatogram on the same strip of filter paper. This parallel chromatogram, or marker strip, is treated with a reagent such as aniline phthalate14*which develops a brown color with hexoses and a pink or red color with pentoses. The corresponding areas of the untreated chromatogram are cut out and extracted, for determination of reducing sugar. Both xylose and mannose can be determined from a single chromatogram, although in the case of xylose it was found necessary45 to add about 10% of the actual value obtained, to correct for losses during the preparation of hydrolyzates. The method of elution of chromatograms was later49 replaced by a photometric method, applied directly to the chromatogram after the development of color with aniline phthalate. I n this method, the pulp or wood hydrolyzates, together with standard solutions containing known amounts of sugars, are chromatographed on Whatman No. 1 filter paper, the solvent employed and the duration of the run depending on the types of sugars present. For the determination of glucose, galactose, and xylose, butanol saturated with water was found to give satisfactory separations on chromatograms run for 8 days. For determinations where mannose, arabinose, and xylose were simultaneously present, 18-day chromatograms with a solvent mixture of n-peiityl alcohol, acetic acid, and water in the ratio 40:10:50 were preferred. It was found convenient to add the phthalic acid (subsequently required (46) (47) (48) (49)

A. E. Flood, E. L. Hirst and J. K. N. Jones, Nature, 160,86 (1947). J. R.Hawthorne, Nature, 160, 714 (1947). S . M. Partridge, Nature, 164, 443 (1949). C. Gustafsson, J. Sundmnn and T. Lindh, Paperi l a Puu, B33, 1 (1951)

294

W. J. POLGLASE

for color development) to the solvents, since it does not interfere. with sugar separation a t the concentration used (2.5 %). The chromatograms are removed after the prescribed time, dried, drawn through ether coiitaining 2% of aniline, and heated a t 105" for color development. A uniform color drvclopmeiit results from this tcchniqur of application of the anilinc phthalat e. The chromatograms are then photographed a t a reduction of 17.5 times, and the spot density on the film is evaluated with a Zeiss Schnell-photometer. I t was found that the amount of sugar is a straightline function of the integrated spot density, the slope of the line being determined in each Case by the two standards run simultaneously. The originators of this method49elaim that 5 to 150 microg. of xylose can be determined with a probable error of 3 to 5 %, in the presence of galactose, glucose, or mannose, although arabinose interferes if its concentration is more than 25 times that of the xylose. Arabinose (more than 5 microg.) can be determined in the presence of a maximum of 25 times as much xylose or mannose, arid mannose (more than 20 microg.) in the presence of a maximum of 15 times as much arabinose, both with a probable error of 5%. When glucose and galactose are present in approximately equal amounts, the error in determiiiatioii is about l o % , and if one exceeds the other by a factor of 6, glucose must first be removed by fermentation with Saccharomyces bayanus. Mannose can be determined with a probable error of 5 %. A method for the determination of mannan, proposed recently,60is based on the observation61 that lead tetraacetate, in glacial acetic acid solution, oxidizes glycols to pairs of carbonyl groups a t rates that are greater for cis than trans isomers and that are dependent on the particular glycol exIt -was ami~ied.~O ~ ~ postulated60 that the rate difference might be useful in distinguishing between the cis-glycol grouping a t C2 and C3 in mannan and the trans-glycol grouping in cellulose. If the difference in rates of reaction of these two isomeric groupings is sufficiently great, samples of cellulose which contain high percentages of mannan should show an initial, rapid reaction with lead tetraacetate, which would then decrease in rate as the mannan is oxidized and the reaction with cellulose becomes predominant. Steinmann and coworkers60 used the following procedure for determination of consumption of lead tetraacetate by cellulose samples. (50) B. B. White, H. W. Steinmann and R. W. Work, X I I I t h International Congress of Pure and Applied Chemistry, Stockholm, 1953; H. W. Steinmann and B. B. White, T u p p i , 37, 225 (1954). (51) R. Criegee, Ber., 64, 260 (1931); 66, 1770 (1932); Ann., 607, 159 (1933). (52) C. C. Price and H. J. Kroll, J. A m . Chem. Soc., 60, 2726 (1938). (53) C. C. Price and M. Knell, J. Am. Chem. Soc., 64, 552 (1942). (54) R. C. Hockett, M. L. Dienes and H. E. Ramsden, 6.Am. Chem. Soc., 66,1474 (1943).

-

POLYSACCHARIDES ASSOCIATED WITH WOOD CELLULOSE

295

A suspension of approximately 0.01 mole of pulp in 100 ml. of purified acetic acid was stirred for 2 hr. in a water bath at 30°C. Exactly 25 ml. of a n approximately 0.1 N solution of lead tetraacetate in glacial acetic acid was then added, the mixture being maintained a t a temperature of 30°C. Ten-ml. samples were pipetted out a t 15min. intervals, discharged into 20 ml. of potassium iodide-sodium acetate solution, and titrated with 0.01 N sodium thiosulfate solution, using starch as an indicator. A blank consisting of 100 ml. of purified glacial acetic acid and 25 ml. of lead tetraacetate solution was prepared, and sampled and titrated as previously. The difference between the value for the blank and that for the sample gave the amount of lead tetraacetate consumed by the pulp. When these data were plotted,60 curves were obtained which indicated an initial, rapid reaction which approached a linear rate after about the first hour. If the break in the curve was considered t o mark the end of the reaction with the cis-glycol grouping of mannan, and the subsequent slower reaction was assigned t o the trans hydroxyl groups of cellulose, extrapolation of the straight-line portion t o zero time would then give a measure of the mannan content of the pulp.

In order to equate the consumption of lead tetraacetate to the mannan content of the pulp, Steinmann and coworkerss0analyzed a series of pulps for mannan, by hydrolysis and subsequent precipitation of mannose phenylhydrazone. A satisfactory correlation was observed between the two methods. Since this work was restricted to a study of acetylation-grade pulps, which are generally low in mannan, the correlation between consumption of lead tetraacetate and mannan content did not extend beyond 2 % of mannan. Further studies on this method, to determine whether or not it can be applied to less refined pulps, would be of great interest. c. Properties of Mannan in Wood Cellulose and Derivatives.-The technique of acetolysis and chromatography, which Wolfrom and coworkersss' s6 applied to cotton-linters cellulose for the purpose of isolating acetates of oligosaccharides, has been used by Leechs7in a study of slash-pine alphacellulose. The alpha-cellulose was prepared from holocellulose by successive extraction with 5 % and 24 % potassium hydroxide. This residual alphacellulose contained 0.8 % of lignin, 0.6 % of ash, and 10.4 % of mannan. No uronic acid or arabinose, and only the slightest trace of xylose, could be detected in this alpha-cellulose, after hydrolysis followed by paper chromatography. The products of acetolysis were isolated by a chromatographic procedure,58 59 and were t.hen deacetylated, and chromatographed on filter paper. I n this way, a disaccharide of mannose was found, and also a second disaccharide, which, on hydrolysis, gave both glucose and m a n n o ~ e I. t~ ~ 7

(55) E. E. Dickey and M. I,. Wolfrom, J . A m . Cheni. SOC.,71,825 (1949). (56) M. L. Wolfrom and J. C. Dacons, J. Am. Cheni. SOC.,74,5331 (1952). (57) J. G. Leech, Tappi, 36, 249 (1952). (58) W. H. McNeely, W. W. Binkley and M. L. Wolfrom, J . Avti. Cheni. SOC.,71, 825 (1949). (59) W. W. Binkley and M. L. Wolfrom, "Chromatography of Sugars and Related Substances," Sugar Research Foundation, 2v. Y . , Sci. Rept. Series, No. 10, 1948.

296

W. J. POLGLASE

was c*oncluded that the latter disaccharide is an 0-glucosyl-mannose, although the data supplied57are not adequate to affirm this conclusion. Acetolysis experiments designed to establish whether or not synthesis (reversion) had occurred did not yield a glucose-mannose d i ~ a c c h a r i d e .It~ ~ was concluded57that these data implied a chemical linkage between glucose and maniiose units in the alpha-cellulose of slash pine. This co~ic~lusion is not contrary to other recognized facts relating to the mannan of wood. Everi the most severely refined, softwood pulps still retain a certain amount of mannan. Wise and Ratliff 6o extracted slash-pine holocellulose with I6 (3% potassium hydroxide, and treated the residue repeatedly with fresh charges of boiling 5 % sulfuric acid. A large part of the origina TABLE IV hdannan Content- of Coniferous Wood Samples Mannan, o/o Concentralton of polasiuiir hydronde, %

Sample

Slash pine

“I-Iemicellulose- A” “Hemicellulose-B” “Hemicellulose-C” Alpha-cellulose

5 16 24

Summation Original wood Per cent, based

011

1.3 0.7 0.8 6.1

1

Black spruce

1.2 0.9 0.4 5.0

-_

__

8.9 10.0

8.4 10.3

the original, unextracted wood.60

mannan remained in the undissolved residue. Extraction of slash-pine and black-spruce holocelluloses with aqueous potassium hydroxide left alphacellulose residues containing most of the mannan (see Table IV). Evidence for a chemical linkage between cellulose and maiinan, a t least in softwood pulps, continues to accumulate. In this connection, Steiiimann and coworkers50 havc observed that, during the normal preparation of cellulose acetate, a considerable part of the xylan may be lost (30 to 60 %), whereas the mannan content is not greatly affected. These authors suggest that this may indicate that the matinaii is bound to the anhydroglucose chain. Arcording to these workers, certain differences in properties of cellulose acetat,e prepared from different, samples of cellulose can be explained if a manriaii-to-cellulose bond actually does exist. However, it must he realized that no definitive proof has been preseiited of a mannati-to-cellulose linkage in any preparation of wood cellulose. (60) L. E. Wise and E. K. Ratliff, Arch. Biocherri., 19, 292 (1948).

POLYSACCIIARIDES ASSOCIATED WITH WOOD CELLULOSE

297

The part played by mannan in contributing to the properties of products derived from wood cellulose is receiving considerable attention. Although earlier methods of determination of niannan as the mannose phcnylhydrazone made detailed studies of correlations between maiinan content and product properties a tedious task, Dorr‘jl was able to show a correlation between tensile strength of viscose rayon and the amount of xylan and mannari in the pulp. Those pulps showing the lowest proportion of xylan plus mannan yielded rayon yarns with the greatest strength.61From recent studies,5nit seems probable that the polyoses, especially mannan, in cdlulose acetate have an effect on rheological properties. In spinning cellulose acetate from acetone solutions, it is necessary t o maintain the viscosity of the spiuning solution constant, within relatively narrow limits, in order to permit control of yarn quality. The variability of the viscosity of the spinning solution is dependent on concentration, which can be rigidly controlled, and on the uniformity of the acetate used. Assuming a product of constant degree of substitution, or acetyl value, the viscosity of solutions of cellulose acetate should be related to molecular weight. This can be approximated by determination of the relative viscosity of a dilute solution of the acetate in a solvent such as acetone, from which the intrinsic viscosity may be calculated by the use of suitable equations. When the viscosity of a concentrated, acetone solution of a woodcellulose acetate is compared with that of a cotton-linters acetate a t the same concentration, the wood-cellulose acetate will generally be found to have a higher viscosity than the cotton-linters acetate, even though both acetates have identical intrinsic viscosities. This “anomalous viscosity” effect in mood-cellulose acetate was studied by White, Steirimann and Work,5owho observed that, in the concentration region of 5 % to 30% cellulose acetate in acetone, a graph of log (viscosity) versus concentration gave a straight line for a number of cellulose-acetate preparations. The ratio of viscosities measured a t two convenient concentrations was thus a constant for a given cellulose acetate preparation, and could be used as a measure of anomalous viscosity. Steinmann and coworkers50used 27 % and 7 % acetone solutions for viscosity measurements] and defined the viscosity ratio as log (27 % viscosity) divided by log (7 % viscosity) times 1 P . When the mannan content of wood pulps, as determined by lead tetraacetate oxidation (see page 294), was plotted against the viscosity ratio, the relationship in Fig. 1 was obtained. The correlation coefficient of the regression line was calculated to be 0.81, which is significant a t the 1% level. Xylan and carboxyl groups were considered to play a less important role in anomalous viscosity, since it was observed that pulps having similar (61) R. E. Dorr, Reichsamt Wirtschajtsausbau Pruf-Nr. 64 (PB62006), 29 (1940) ; Chem. Abstracts, 41, 5301 (1947).

298

1%'.

J. POLGLAAE

00 80 70 0

2 60 m

6

50

-

40

m

aJ

g

30

0 +.

m

2 "

20

0

>

0

10

0.4

0

1.6

0.8 1.2 Percentage of mannan

:0

FIG.1.-Relationship Between the Mannan Content of the Wood Pulps and the Viscosity Ratios of Corresponding Cellulose Acetates.60

contents of t,hese could differ widely in viscosity ratio (see Table V). Further, it was observed that the decrease ih viscosity ratio obtained when cellulose acetate was heated under pressure in acetic acid solution was accompanied by a decrease in mannan but not in xylan (see Table VI). The conclusion reached by t'hese workerss0 is that mannan in cellulose TABLEV Typical Analysis of Cellulose Used for Cellulose Acetate ManuJacture6Q

I I

Source

Cotton linters Pulp A Pulp B

Pulp

1 la

2 3

j

I

AL~:,

98.0

Mannan, calcd. on pulp. Y, Pulp

CA

1.83 1.83 1.66 1.85

1.73 0.81 1.28 1.53

Loss i n mannan in C A prep.# %

0.10 1.02 0.38 0.32

I

Pentosans, cellulose ncefate

0.5

1.41

X y l a n , calcd. on pulp, Y, Pdp

CA

1.33 1.33 1.31 1.58

0.73 0.75 0.62 0.92

11.5

Loss in xylan i n C A prep., %

Viscosily ratios of C A

0.60 0.58 0.69 0.66

41.0 14.5 45.0 60.0

POLYSACCHARIDES ASSOCIATED WITH WOOD CELLULOSE

299

acetate is the predominant, pulp constituent which causes high viscosity ratios in cellulose acetate, with xylan possibly a minor contributor. The mechanism by which mannan could affect cellulose acetate viscosity is not known, but it was suggested60that if mannan were attached to cellulose as a branch or cross-link it could account for the observed viscosity phenomena. 3. Xylan a. Structure of Xy1ans.-The chemistry of xylan has been reviewed recently by WhistlerG2and by Whistler and Smart,63and only certain aspects of this subject will be considered here. These 63 should be consulted for details regarding isolation, purification, and structure of xylan. TABLEV I I Properties of Crystalline Xylan6a*6 6 Properlies

1

From Barley Straw

[ a ]in~ 6%

NaOH in 6% NaOH Pentosan, % Uronic anhydride, % Degree of polymerization Ash, %

-106" (c, 4 . 3 ) -204 ' 99.5 0.4 39 0.34

[cY]4360

1

From Paper Birch

-92" (c, 3 . 3 ) - 196" 98.4 2.40 35 0.40

Xylan from wood has received somewhat less attention from carbohydrate chemists than has the xylan from such rich sources as wheat straw, corn cobs, and esparto grass. However, specific optical rotations of the xylans obtained from the aforementioned sources are of the same general order of magnitude as those of some xylaiis obtained from wood.63Yundt crystallized xylans from paper birchG4and from barley straw66as spherocrystals, and compared their properties (see Table VII). From methylation data and molecular-weight determinations, a structure for esparto-grass xylan ([CX]~~D-92", in aqueous NaOH)66has been suggested by Hirst and associates.66The molecule consists of about 75 P-D-xylopyranose units linked 4 1, with a single branch point (3 -+1) linkage.66A similar structure has been proposed for a xylan isolated from

-

R . I,. Whistler, ildvances in Carbohydrate Chert/.,6, 269 (1950). R . 1,. Whistler arid C . I,. Smart, Ref. 16, p. 131. A . P. Yundt, .I. .4m.~ ' h r r i SOC., . 71, 757 (1949); T n p p i , 34, 91 (1951). A. P. Yundt, T a p p i , 34, 89 (1951). (66) S. I<. Chnntla, E. I,. Hi&, J . K. N.,Jones a n d 13. G . V. l'crcival, J . P h e w . SOC.,

(62) (63) (64) (65)

1289 (1950).

300

W. J. POLGLASE

pears, except that this xylan contains a terminal D-glucuronic acid residue.67 Xylan from corn cobs has been converted by graded, acid hydrolysis to a homologous series of crystalline D-xylo-oligosaccharides, which were further characterized as their 62 63 This continuous series extended from D-xylobiose to D-xylohexaose. Methylation studies on the D-xylobiose established the presence of the 4 --z 1 glycosidic linkage. Evidence from the optical rotation of xylans, the rotational shift,64* 6g and the increase in rotation on acid hydrolysis suggests that the P-D-glycosidic linkage predominates. D. P.s ranging from 35 to 150 have been reported for various purified preparations of xylan.62*63 I n the foregoing brief discussion, the chemistry of a few, carefully purified preparations of xylan from various sources has been considered. In general, these preparations have been found to have a high xylan content and to contain only small amounts of other carbohydrate units. Through studies of these substances, it has been established that xylans contain predominantly 4 1 P-D-glycosidic bonds, with little branching. That not all of the xylans of wood are homogeneous polymers of anhydroD-xylose units has been shown by the classical studies of O’Dwyer (1923 to 1940) on hemicelluloses of American white oak. O’Dwyer prepared a hemicellulose fraction from water-extracted, oakwood sawdust by extraction for two days with 4 % aqueous sodium hydroxide solution. The polysaccharide material was obtained, after acidification, by the addition of ethanol. The product ([a],, -75” in 1% sodium hydroxide), contained 70% of pentosan, and yielded D-xylose, D-mannose, D-galactose, and ~ - a r a b i n o s eon ~ ~hydrolysis. Recognizing the need for a more uniform, polysaccharide material for detailed constitutional shdies, O’Dwyer prepared two hemicellulose fractions from beechwood previously extract>edwith water and 0.2 % sodium hydroxide. As in previous ~ o r kthe , ~hemicelluloses ~ were extracted from beechwood sawdust with 4 % aqueous sodium hydroxide. Pectic materials were precipitated with lime-water, and a fraction designat,ed “hemicellulose-A” was obtained by acidifying the solution wit,h acetic acid. T o t8he filt,rat,c,from hemicellulose-A, was added 95 % ethanol to precipitate “hemicellulose-B.” After further purificat’ion, both of these hemicelluloses were subjected tto chemical study. Hemicellulose-A7’ ([a],,- 107” in 1 % sodium hydroxide) was observed to be soluble in cold water when freshly prepared, but it was insoluble after being dried. The preparation did not reduce

-

(67) (68) (69) (70) (71)

S. I<. Chanda, E. L.Hirst and E. G. V. Percival, J . Chetn. Soc., 1240 (1951). R. I,. Whistler and C.-C. T u , J . A m . C’he77i. Soc., 73, 1389 (1951). R . E. Reeves, Advauces in Carbohydrate Chewi., 6, 107 (1951). Margaret € I . O’Dwyer, Riochettc. J . (London), 17, 501 (1923). Margaret €1. O’Dwyer, Riochetti. J . (London), 20, 656 (1926).

30 1

POLYSACCHARIDES ASSOCIATED WITH WOOD CELLULOSE

Fehling solution. It contained anhydro-o-xylose units and 11 % of it was uronic acid anhydride. D-Glucuronic acid was identified in hydrolyzates, ~ in but a test for D-galactose was negative. Hemicellulose-B ( [ a ]-120" 0.5 % sodium hydroxide) yielded 4.8% of galactose (galactaric acid test), an undetermined amount of L-arabinose, and 64 % of uronic acid identified as galacturonic a ~ i d . 7 ~ ~~ hemiReturning to studies on oak hemicelluloses, O ' D ~ y e robtained celluloses-A and -B from bot,h heartwood and sapwood of oak, by techniques similar to those applied to beechwood. Not only were hemicelluloses-A and -B different in physical and chemical properties, but differences TABLEV I I I Analyses of Oak Hernicellulose-A72

1

Green sapwood Green heartwood

1 :::1

- 62" -82"

1 it:: 1

10.16 10.71

8:;3

~

04.16 97.24

TABLEI X Analyses of Oak Hewiicellulose-B7~

_

Green sapwood Green heartwood

~

+25.5' -52.3"

~

3.64 1.75

16.81 16.87

~

50.53 82.03

1

_

_

2.02 2.24

~ ~

76.34 98.9

were also rioted which depended on whether the source was sapwood or heartwood (see Tables VIII and IX). Further differences in the extracted hemicelluloses were encountered as a result of variations in conditions of drying the The analyses of heartwood and sapwood hemicellulose-A (Tablc VIII) were similar, and for both there was found a ratio of 11 anhydro-D-xylose units to 1 0-methylhexuronic acid unit. However, sapwood hemicellulose-A gave a blue color with iodine, but heartwood hemicellulose-A did not. Furthermore, hemicellulose-A from sapwood was less stable toward acid hydrolysis than was that from heartwood?? Treatment of hemicellulose-A from sstpwood with Takadiastase, for 44 hours, ciaused the blue color (with iodine) to disappear, and yielded D -~ I u (72) Margaret H. O'Dwyer, Biocheni. J . (I,ondon), 28, 2116 (1034).

302

W. J. POLGLASE

cose in s0lution.7~The residual material, and a purified, ash-free hemicellulose-A from heartwood, then had identical rotations ([(Y]D -97.5' in 2% sodium hydroxide). A protracted treatment (210 hours) with Takadiastase, of either of these preparations, caused dissolution of most of the hemicellulosic material. The aqueous solution then contained 3 parts of D-xylose to 2 parts of a water-soluble polysaccharide ([a]D-51.2", c 2 in water). This polysaccharide contained amounts of uronic anhydride, methoxyl group, and xylan in agreement with those calculated for a theoretical molecule consisting of 6 anhydro-D-xylose units and 1 O-methylhexuronic acid ~ n i t . Acid 7 ~ hydrolysis of 1 mole of this material yielded 5 moles of D-xylose and 1 mole of an O-methylaldobiouronic acid ([(Y]D 69", in water).74 Hemicellulose-B(Tab1e V) from sapwood gave a blue color with iodine, which disappeared after treatment with Takadiastase under carefully controlled condition^.^^ The solution was found to contain D-glucose, and its reducing value was equivalent to 24.6% of hexosan. On the basis of analytical data, heartwood hemicellulose-B appeared to be similar in constitution to the polysaccharide (6 anhydro-D-xylose units and 1 O-methylhexuronic acid per structural unit) isolated. after prolonged enzymic hydrolysis of heartwood hemicellulose-A. These extensive studies by O'Dwyer have shown that part of the D-xylose-yielding materials of wood exists in heterogeneous polysaccharides of considerable complexity. Furthermore, the polysaccharides in heartwood and in sapwood differ in composition, although the fundamental structural building unit may be the same in all cases. Further studies on hemicelluloses of hardwoods have shown that these may exhibit a considerable range in the content of xylan and O-methyl77 In a recent review of hemicelluloses, Wise78 hexuronic anhydride.76* suggests that hardwood hemicelluloses may have a rather simple, architectural pattern, consisting predominantly of D-xylose units and smaller amounts of O-methyluronic acid units. Softwood hemicelluloses also contain this fundamental structural unit consisting of a mono-0-methyluronic acid in combination with D-xylose units.79In the case of pine, this fundamental unit may be associated with a part of the m a i i i i a ~ i . ~ ~ The structure of wood xylari has been studied further by Gustafsson and (73) Margaret H. O'Dwyer, Biochem. J. (London), 31, 254 (1937). (74) Margaret H. O'Dwyer, Biochem. J . (London), 33, 713 (1939). (75) Margaret H. O'Dwyer, Biochem. J. (London), 34, 149 (1940). (76) E. Anderson, M. G. Seeley, W. T. Stewart, J. C. Redd and D. Westerbeke, J . B i d . Chetti., 136, 189 (1940). (77) E. Anderson, R . B. Kaster and M. C . Seeley, .I. R i d . Chert/.,144, 767 (1942). (78) I,. E. Wise, Ref. 9 , p. 369. (79) E. Anderson, J. Kesselman arid E. C. Bennett, J . Biol. Chcm., 140,563 (1941).

POLYSACCHARIDER ASSOCIATED WITH WOOD CELLIJLOSE

I

COOII

;I

FIG.2.-Structure

303

dli

of an Acidic Xylan from Birch.80

associates.80These workers isolated, from birch-wood holocellulose, a xylan which consists of about 20 xylose units and one 4-O-methyl-~-glucuronic acid unit per molecule. It has been noted earlier in this article (p. 286) that 4-O-methyl-~-ghcuronicacid was isolated by Jones and Wiselo(”)from aspen wood. These workers‘O(b)also obtained, from aspen wood, an aldobiouronic acidsoawhich was identified as ~-xylose-(2-+ 1) 4-O-methyl-a-~-glucosiduronic acid. Gustafsson and coworkersBowere able to obtain this same aldobiouronic acid by partial hydrolysis of the xylan which they isolated from birch holocellulose. The above data, together with the results of periodate oxidation and a viscometric chain-length determination, were in accord with the formula given in Fig. 2. b. Determination of XyZan.-Until the recent development of quantitative, chromatographic methods, no specific method for the determination of anhydroxylose units in plant materials had been available. However, in wood pulps, the major part of the pentosan is xylan, and it is probable that the conventional pentosan analysis8’ gives a reasonably accurate value for xylan. The determination of pentosans is based on their conversion, through acid hydrolysis, into pentose sugars which, under the conditions of the analysis,s’ are converted immediately into furfural (2-furaldehyde). The furfural may be estimated, after distillation, by one of a number of gravimetric, volumetric, or colorimetric methods. The large volume of research on pentosan estimation has been reviewed by Dor6e,s2 by Browne and Zerban,a and, more recently, by Browning.81 (80) J. Saarnio, Kristina W a t h h and C. Gustafsson, Acta Chem. Scand., 8, 825 (1954). (80a) See also, G. 0. Aspinall, Advances in Carbohydrate Chem., 9, 131 (1954). (81) See B. L. Browning, Ref. 6, p. 1162. (82) C. DorBe, “The Methods of Cellulose Chemistry,” D. Van Nostrand Co., Inc., New York, N. Y., 1933, p. 354. (83) C. A. Browne and F. W. Zerban, “Physical and Chemical Methods of Sugar Analysis,” John Wiley and Sons, Inc., New York, N. Y., 1941, p. 904.

304

W. J. POLGLASE

The researches have dealt with each phase of the pentosail dctermiiiat#ioii. Many modifications have been int]roduced in t,he distillation procedure, i i i attempts to obtain maximal yields of furfural. For example, some workersfi4 prefer to distil with 23 % hydrohromic acid, rather than with the convcntional 12 % hydrochloric acid. Otherss6 distil in the presence of added sodium chloride, 60 avoid changes in acid concentration. Steam dist,illat,ion has been used by a number of workers, who claim theoretical yields of furfural from peiitoses,86 but Launer and Wilsons7 found no advant'a,ge either in salts or in steam in the analysis of pulps and papers. Int'erferiiig substances are of two types: materials other than pentosans which form furfural in the pentosari analysis, and substarices which yield products which may be determined as furfural. Uronic acids arid polyuroriides yield furfural, although not quantitatively, and, in the case of materials containing appreciable quantities of these substances, i t is usual to make a correction. The value of the correction to be applied has been determined experimentally by several workers, with somewhat differing result,s. Norris and Reschgs determined furfural yields from uronic acids, under the conditions normally used for pentosari determination, and obt,ained about 37 to 44% of the theoretical yield of furfural. I n later w ~ r k t,he ,~~ effect of various furfural-yielding substances on one another was ascertained, arid a standardized procedure was developed for analysis of t8he types of mixtures commonly met with in cell-wall chemistry. Apparent,ly, the yield of furfural from uronic acids and polyuronides varies with the proportion of these substances present, and with their ratio to other sugars. From oxycellulose, Uiiruh and Kenyongoobtained furfural yields of 18 to 20 % of the theoretical on the basis of carboxyl content. Wise and Ratliff ,i however, in summative analysis of wood, used a correction based on t,he assumption that the uroiiic acids present evolve 35 % pf the theoretical amount of furfural. It is questionable if a truly valid correctioii for uroiiic anhydride can be applied, in view of the results obtained by Sarkar, Mazumdar and Palg1on the hemicelluloses of jute fiber. These workers isolated a methoxyl-contjain(84) G. Jayme and P. Sarten, Biochem. Z., 308, 109 (1941); 310, 1 (1941). (85) C. Kullgren and H. TydBn, Zng. Vetenskaps Akad. Handl., 9 4 , 3 (1929); Cherti. Abstracts, 24, 1316 (1930). (86) N. C . Pervier and R. A. Gortner, I n d . Eng. Chem., 16, 1167, 1255 (1923). (87) H. F. Launer and W. K. Wilson, J . Research Natl. Bur. Standards, 22, 471 (1939). (88) F. W. Norris and C. E. Resch, Biochena. J . (London), 29, 1590 (1935). (89) S. Angell, F. W. Norris and C. E. Resch, Biochern. J . (London), 30,2146 (1936). (90) C. C. IJnruh and W . 0. Kengon, J . Am. Chem. SOC.,64, 127 (1942). (91) P. I3. Sarkar, A. K. hlazumdar and K. B. Pal, Textile Research J . , 22, 529 (1952).

POLYSAC!CHARIDES ASSOCIATED WITH WOOD CELLULOSE

305

itig “xylo-aldobiouronic acid” from jute hemicellulose. From t)ot,h t,he aldobiouronic acid and the purified, jute hemicellulose, tthe furfural yield was equivalent to the D-xylose content. Furthermore, methanol could not be detected i n the distillate from 12 % hydrochloric acid, nor could methoxyl be detected in the furfural-phloroglucinol precipitate. Sarkar and coworkers are of the opinion that mono-0-methyluronic acid is incapable of yielding , ~found ~ that the carbon dioxide evolufurfural. In this same W V O ~it ~was tion from the methoxyl-containing aldobiouronic acid was quantitat>ive.It is, therefore, doubtful if the accuracy of the pent,osan determination is improved by a correction which assumes that t,he entire uronic-anhydride cont,ent of a platit. material will yield a charact’erist,icamount of furfural. Those substances which may be determined as furfural include hydroxymethylfurfural from hexose units, methylfurfural from 6-deoxyhexose units, and small amounts of formaldehyde, acetone, and levulinic acid. The gravimetric determinat>ionof furfural requires the formation of an insoluble derivative through reaction with one of a number of reagents, for example, 2,4-dinitrophenylhydra~ine,9~ pheny1hydrazine,g2 p-~itrophenylhydrazine,~~ phlorogluci~iol,~~ barbituric and t’hiobarbituric Of these reagents, phloroglucinol and barbituric acid are the most specific. Phloroglucinol forms precipit,ates wit,h methylfurfural and with hydroxymet.hy1furfural, but these are somewhat more soluble than the furfural-phloroglucinol. Rarbituric acid does not precipitate hydroxymethylfurfural if the latter is present in small a r n ~ u n t ~ sThe . ~ *reaction of furfural to form furfuralphloroglucinol is not st,oichiometric, and t’he precipitat>ionis not complete. After extensive experimentation, Kroberg9standardized a procedure and prepared tables for converttirig the weight of furfural-phloroglucinol to the equivalent weight of furfural. Krober’s tablesa3 were constructed from the results of distillation (with 12% hydrochloric acid) of L-arabinose and of D-xylose, and, since the yield of furfural from each of these pentoses differs, accurate calculations are not possible when the relative concentrat,ion of these pentoses is unknown. In wood pulps, t’here is seldom an appreciable quaiit’ityof arabinose units, so that the figures for converting the weight of furfural-phloroglucinol to xylan are used. Volumetric methods for determination of furfural from pentosans in(92) (93) (94) (95) (96) (97) (98) (99)

A . Gfinther and B. Tollens, Ber., 23, 175 (1890). L. Maaskant, Rec. trav. chim., 66, 1068 (1936). E. Simon, Biochevn. Z., 247, 171 (1932). C. Councler, Che7ti.- Z t g . , 18, 966 (1894). R. Jbgcr and E. Unyer, Ber., 36, 4440 (1902). A. W. Dox and G. 1’. Plaisance, J . Am. (:hem. Soc., 38,215G (1916). W. Gierisch, Cellulosechenzie, 6, 61 (1925). E. Krober, J . Landwirtsch., 48,357 (1900); 49, 7 (1901).

306

W . J. POLGLASE

clude: titration with the bromine formed by acidification of a bromidebromate mixture,"JO titration (with iodine) of excess bisulfite, after reaction lo2 oxidation to furoic acid by hypoiodites of the latter with the furfural,iO1in strongly alkaline and determination of the acid liberated after reaction of the furfural with hydroxylamine hydroch10ride.l~~ A number of reagents give, with furfural, colored compounds which are useful for quantitative colorimetry.105The reaction with aniline has been studied by Stillings and Browning,'06 who were able to determine furfural a t low concentrations even in the presence of appreciable amounts of methylfurfural and hydroxymethylfurfural. For further details on the determination of pentosans, the reader should consult the recent review by Browning.81 It is apparent from the foregoing that a more specific method for the determination of xylan would be desirable. To this end, the determination of xylose, after acid hydrolysis of the polysaccharide material, has been attempted. Xylose may be oxidized to xylonic acid which can be precipitated with cadmium bromide as the double salt,g3 but the precipitation is not quantitative. Xylose forms an insoluble, crystalline di-0-benzylidene dimethyl acetal which permits identification in the presence of other sugars,1o7 but the necessity for anhydrous reaction conditions precludes the adaptation of this method to ordinary analysis. Wise arid Ratliff lo8 prepared this derivative of both D- and L-xylose, as well as analogous derivatives from other aromatic aldehydes, and concluded that, with either the di-0-benzylidene or the di-0-(p-isopropylbenzylidene)dimethyl acetal, ail excellent, highly specific, qualitative test was available for D- or L-xylose. Biological methods have been studied which employ yeasts to remove the hexoses, and specific organisms have been used for the fermentation of D-xylose (Lactobacillus ga yonii) , or L-arabinose (Lactobacillus mannitopoeus) .lo9 The determination of D-xylose in mixtures, by an appropriate combination of fermentation and chemical methods, has been accomplished by Wise arid Appling.'lo (100) W. J . Powell and H. Whittakcr, J . SOC.f'heni. I n d . (London), 43,35T (1924). (101) A. Jolles, Ber., 39, 96 (1906). (102) A. P. Dunlop and F. Trimble, Ind. Eng. Chem. Anal. E d . , 11, 602 (1939). (103) H. R. Rogers, Ind. Eng. Chem. Anal. Ed., 16,319 (1944). (104) A. No11 and W. Belz, Papier-Fabr., 29.33 (1931). (105) F. D. Snell and Cornelia T. Snell, "Colorimetric Methods of Analysis," D. Van Nostrand Co., Inc., New York, N. Y., 3rd Edition, 1953, Vol. 3, p. 185. (106) R. A. Stillings and B. I,. Browning, Znd. Eng. Chern. Anal. Ed., 12,499 (1940). 738 (1945). (107) I,. J. Breddy and J. K. N. Jones, J. Cheni. SOC., (108) I,. E. Wise and E. K. Ratliff, Anal. Chenz., 19, 694 (1947). (109) E. W. Hopkins, W. H. Peterson and E. B. Fred, J . Am. C h m . SOC.,62, 3659 (1930). (110) L. E. Wise and J. W. Appling, Ind. Eng. Chon&.Anal. Ed., 17, 182 (1045).

POLYSACCHARIDES ASSOCIATED WITH WOOD CELLULOSE

307

The most promising of currently available methods for the rapid, routine determination of xylose is quantitative, partition chromatography6 on filter paper, which permits simultaneous determination of maimose and other sugars (see p. 292). c. Propertics 01Xylan in Wood Cellulose and Derivatives.-Several properties of xylan are of importance in determining the extent of its removal in a mood-pulping process. For example, xylan is soluble in alkali, and a t least part of it can be extracted from wood, together with other hemicelluloses, by dilute, sodium hydroxide solutions. This solubility in alkali probably is ascribable, in part, to the relatively low molecular weight of xylaii and, in part, to the presence of uronic acid groupings in the xylan molecule. I t is, therefore, logical to expect that, in alkaline pulping, a certain amount of the xylan will be dissolved. Indeed, xylan has been found in cooking liquors during the early stages of a sulfate cook.111 Another property of xylan which is important in wood pulping is its ease of hydrolysis by acids, as a result of which, under acidic pulping conditions, xylose and soluble xylo-oligosaccharides are formed and dissolved. That xylan is very susceptible to hydrolytic attack by sulfite cooking liquors has been shown by Sundman.112He sampled cooking liquors during the course of sulfite cooking of spruce, pine, birch, and aspen, and found that xylose appears in the pulping liquors a t an early stage. An additional factor of importance in determining the effect of the pulping operation is the nature of the association between xylan and cellulose. Those preparations of xylan which have been studied have exhibited a low degree of branching, which, together with the similarity to cellulose in ring structure (pyranose) and linkage (4 -+ 1 p-D) would permit a close physical association of xylan with cellulose. It has been suggested1I3 that xylan molecules in the fibers actually participate in the micelles (crystallites) and are oriented in the same direction as is the cellulose chain, as a sort of mixed crystallization. Thus, the dissolving of xylan during a pulping operation would depend t o a considerable degree on the ability of the cooking liquors to penetrate the micelles and degrade or elute the xylan. Finally, there is the possibility that a chemical linkage, presumably glycosidic, might exist between cellulose and ~ y 1 a n . Such l ~ ~ a linkage would protect the xylan from the dissolving and degradative action of alkaline solutions, without decreasing its susceptibility to acid attack. No direct evidence for such a linkage has been obtained, but the resistance of part of the xylan toward alkaline treatment (see below) suggests this p ~ s s ib ility .~ '~ (111) (112) (113) (114)

J. Saarnio and C . Gustafsson, Paperija Puu,36, 65 (1953). J. Sundman, Paperija Puu,32B, 267 (1950). W. T. Astbury, R . D. Preston and A. G. Norman, Nature, 136, 391 (1935). A. G. Norman, Ref. 14, p. 28.

308

W. J. POLGLASE

Wheii wood pulps are extracted with cold 17.5 % sodium hydroxide sohition, pentosan is dissolved to an extent dependent on the species of wood and the pulping procedure. A portion of the pentosan remains in the alkaliinsoluble (alpha-cellulose) fraction, and may bc determined therein by the usual analytical procedure. This alkali-insoluble peritosan has been called “resistant pentosan” or “resistant xylan.” Sulfite pulps from softwoods may be quite low in resistant, xylan (0.2 % or less), whereas alkaline pulps from hardwoods represent the other extreme, with as high as 8 to 9 % of resistant xylan. Other factors besides wood species and pulping procedure affect resistant xylan. It has been reported by Meller”6 that the drying of pulps a t 105” increased the coiitent of xylan resistant to the dissolving action of 7 % sodium hydroxide, whereas beating in the LampBn mill had the opposite effect. Norman116 compared the behavior, toward 4 % sodium hydroxide, of wheat-straw Cross-and-Bevan cellulose before and after regeneration from cuprammonium solutions and from viscose. The regenerated samples retained only 0.4 and 1.1%, respectively, of xylan, whereas the original sample retained 8.3 %. This was considered to be evidence that a major part of the xylan resistant to 4 % alkali is retained by secondary valence forces in the organized cellulose structure. After dissolution and regeneration of the cellulose, these organizing forces are no longer in effect, and the xylan can be readily removed. S ~ h o e t t l e has r ~ ~studied ~ the alkali resistance of the xylan in pulps made from aspen wood. Verifying and extending previous work by March,l18hell7 observed a higher content of xylan resistant to 17.5 % sodium hydroxide in aspen alkaline pulps (sulfate or soda process) than in aspen, chlorite holocellulose. Further chlorite treatment of an alkaline pulp did not reduce its xylan or resistant-xylan content. Alkaline (sulfate) cooking of the holocellulose did not increase the resistant-xylan content. These results are summarized in Table X. Bleached, soda pulp contained about the same amount of resistant xylan as bleached, sulfate pulp, and a bleached, sulfite pulp contained only a small it was observed that the resistantamount (Table X). In this same st~dy,~17 xylan content was greater in an air-dried or oven-dried pulp than in the same pulp before drying (see Table XI). In agreement with the work of NIeller,116a reduction in resistant xylan resulted from mechanical beating or grinding. S ~ h o e t t l e also r ~ ~studied ~ the effect on xylari of different alkalis a t several (115) A. Meller, Paper Trade J . , 126,57 (1947). (116) A. G. Norman, Biochen~.J. (London), 30, 2054 (1936). (117) J. R. Schoettler, Ph.D. Thesis, The Institute of Paper Chemistry, Appleton, Wisconsin, June 1952; T a p p i , 37, 686 (1954). (118) R. E. March, Paper Trade J . , 127. No. 17,51 (1948).

309

POLYSACCHARIDES ASSOCIATED WITH WOOD CELLULOSE

TABLEX X u l a n and Resistant-xvlan Content o.f A s p e n Pulps"' Pulp

Chlorite holocellulose" Bleached* sulfate Overbleachedb sulfate Kraft-cooked holocellulosc Bleached * soda Bleached* sulfite

X ylan,

Yo

X ylan in alpha-cellulose (Resistant xylan).

21.6 16.2 16.4 17.8 14.1 5.5

a Prepared by the method of Wise and cow0rkers.1~~ hypochlorite.

7;

2.0 8.1 8.0 1.6 7.7 0.2

* Bleached

with sodium

TABLEX I Effect of Drying on Resistant-xylan Content of A s p e n Bleached Kraft Pulp"' Drying conditions

Moisture, %

Xylan in alphacellulose (Resistant zylan) , %

Undried Air-dried, 70"F., 50% relative humidity Air-dried, lOO"F., 20% relative humidity Oven-dried, 50"C., in vucuo

7.1 4.3 0.5

5.2 8.8 8.7 9.2

different concentrations. A bleached, sulfate pulp which contained 9.2 % of ~ y l a nresistant to 17.5 % sodium hydroxide (5.20 Jl) contained only 2.6 % of xylan resistant to24 % potassium hydroxide (5.20 M ) . Six per cent sodium hydroxide removed more xylan from aspen alkaline pulps than did 17.5% sodium hydroxide, and less xylan was removed a t a concentration of 24% sodium hydroxide than a t 17.5 %. It was discovered117that resistant xylan can be isolated by extraction of alpha-cellulose with 24% potassium hydroxide, followed by acidification with acetic acid and precipitation with dcohol. Most of the above results are in accord with the idea that sylaii and cellulose co-exist in a sort of mixed crystallization11dheld together by secondary valence forces (such as hydrogen bonds). Mechanical treatment would be expected to disrupt such bonds, thus decreasing the resistant xylan, and the removal of water from a wet pulp would allow neighboring chains t o come into contact, resulting in augmentation of the number of secondary bonds and thus increasing the resistant xylan. (119) 1,. 15 Wise, h4 Murphy and A . A . D'Addicco, Paper Trade J . , 122, No 2, 35 (1946).

310

W. J. POLGLASE

The low proportion of resistant xylan in sulfite pulps can be explained partly as a hydrolytic attack, by the acidic sulfite pulping liquor on the acid-labile xylan, resulting in depolymerization to small, easily soluble molecules, and partly as a random hydrolytic attack on the cellulose, occasioning fissures in the micellar structure. The resulting pulp can be penetrated readily by alkaline solutions, and the xylan (because of extensive depolymerization) can be eluted. In an alkaline pulping process, there is 110 hydrolysis of glycosidic bonds but only an attack on reducing ends of polysaccharides which has little effect on molecular size. The structure of the fibers is preserved, and the resulting pulp cannot be easily penetrated by strong alkali. The net result for alkaline pulps would, therefore, be a high level of resistant xylan. Schoettler"' has proposed a special mechanism to explain the behavior of aspen chlorite holocellulose, which contains relatively small amounts of resistant xylan. He has suggested that the chlorite reagent splits ligninxylan bonds with resulting oxidation of the xylan a t these points of cleavage, possibly with the formation of carboxyl groups on the xylan. This oxidative cleavage may be accompanied by depolymerization, the combined effects increasing the solubility of the xylan in alkali. In contrast, it was suggested"? that alkaline pretreatment or alkaline pulping splits the ligninxylan bond by alkaline hydrolysis, without depolymerization or formation of free carboxyl groups. The recent studies of the Finnish groups00 l 2 0 further emphasize the complexity of the xylan in wood cellulose. These workers have shown that the properties of the xylan in wood pulps depend not only on the method of pulping but also on the type of raw material. It was observedlZ0that, when pulps are prepared by the (acidic) sulfite process from the softwoods, pine and spruce, a substantial part of the xylan is of the acidic type (see Fig. 2, p. 303). Even when these sulfite pulps are refined further with hot alkali, some acidic xylan remains. However, when pulps are prepared from pine and spruce by the (alkaline) sulfate process, the acidic xylan is removed and only neutral xylan remains. On the other hand, when the hardwoods, birch and aspen, are pulpcd by either the sulfate process or the sulfite process, a substantial proportion of the xylan remaining in the pulp is of the acidic type.lZ0 Many excellent articles have been written concerning the benejicial effect of hemicelluloses on the properties of pulps to be used for paper-making'21-'2hSrveral authors have commented on thc tr'etrimmtnl effects of (120) J . Saarnio, Kristina WathCn andC. GuRtafssort, P n p e f i j a Picit, 36,209 (1954). (121) I,. E. Wise, Ref. 9, p. 399. (122) I,. G . Cottrall, T n p p i , 33, 471 (1950); P u l p & P n p e v M a g . ('on., 61, No. 10,

135 (1950).

POLYSACCHARIDES ASSOC!IATED WITH WOOD CELLlJLOST",

31 1

liemicelluloses in pulps to be used for chemical conversion.L21126 These subjects will, therefore, not be discussed in detail in this article. Wise notes121that hydration capacity and tensile strength of pulps are more nearly related to hemicellulosic xylari (removable with aqueous alkali) than to total xylan. The addition of xylan to hemicellulose-free, rag pulps decreased the hydration time, and increased the folding and tear resistance as well as increasing the tensile strength of the pulp. A s the xylan content was increased further (to about 6.5%), the tear and folding resistance usually dropped.121 127 In a discussion of wood pulps for use in the manufacture of viscose rayon, Haas and coworkersLzsstate that the wood polyoses, including xylan and Inannan, do not contribute to the strength of the rayon because of their low D. P. Further, the wood polyoses lessen the laundering resistance of According to these same authe rayon because of their alkali thors,l2'j the wood polyoses prevent a uniform reaction in the viscose process, giving rise t o filtration difficulties. They suggest126that the polyoses, partly because of their location in the outer layer of the secondary cellwall, and partly because of their high reactivity, react early in the viscose process and consume more carbon disulfide than corresponds to the average which would be at their disposal in the case of a uniform reaction. A s a result, insufficient carbon disulfide is left for complete reaction of the cellulose. These undissolved, or partly dissolved, fibers then impede viscose filtration.126 D o r F also has commented on difficulties in viscose filtration caused by the presence of xylan, and has noted, further, a correlation between viscose-yarn strength and the total of xylan and mannan in pulps from which the yarns were made. I n this case, the strongest yarns had the lowest, totalpolyose content.61It should be noted that, since the viscose process involves steeping in about 18% caustic solution, the xylan carried through to the final yarn will be chiefly "resistant xylan" (see page 308). WhistlerB2has reviewed the behavior of xylan on acetylation, and notes that dry xylan is acetylated with difficulty. Under reaction conditions which avoid drying, xylan is acetylated readily, to yield a diacetate which is insoluble in most solvents. Because of its insolubility, xylan diacetate, especially if the D. P. is relatively high, can cause filtration difficulties and haziness in commercial cellulose acetates. 0

(123) D. A. Sitch and H. B. Marshall, C a n . J. Research, F26. 376 (1950). (124) H. W. Giertz, Svensk Papperstidn., 63, 673 (1950); 64, 267 (1951); 64, 769 (1951); 66, 72 (1952). (125) L. E. Wise, Paper I n d . and Paper World, 29, 825 (1947). (126) H. Haas, E. Battenberg and D. Teves, T a p p i , 36, 116 (1952). (127) H. Obermanns, Tech. Assoc. Papers, 2 0 , 4 2 9 (1935).

312

3%’.

J. POLGLASE

4. Carbokgdrafes Coritainiriy Carboxill Groups

a. Stritctwe.-Wood pulps, titrated with alkali or examined by ot’hcr procedures*28designed to estimatje free carbosyl groups, are always found to be more acidic than cotton-linters cellulose. These acidic groups may be associated directly with the “true” cellulose, in which case they are probably a t the C6 position arid a t the terminal C l positioii. The acidic groups may exist in tbe cellulose prior to isolation, or t,hey may be int,roduced as a result, of t’he technique used for isolation or purification. There are several known types of 0xycellulose,1~9alt,hough tjhe reactions involved in the oxidation of rellulose by various reagents are far from being understood. “Glucoiiic” acid oxycellulose is obt,airied when the terminal aldehyde groups in a hydrocellulose are oxidized t,o gluconic acid uiiit,s under conditions which result in oxidat’ion of aldoses to aldoiiic acids.130Oxidation of cellulose with periodate cleaves the glucose units a t the 2,3-glycol grouping, with the formation of aldehyde groups.131When this periodatetreated cellulose is oxidized wit,h alkaline hypoioditje,132bromine in aqueous sodium bicarbonate solution,133* 134 or chlorous a ~ i d , ~ t,he 3 ~expected carboxylic acid is formed; t,his yields, on hydrolysis, subst,antial amounts of glyoxylic and D-erythronic acids. A t,hird type of carhoxyl-containing oxycellu136 The chief lose is obtained by oxidizing cellulose with riit>rogen action of t,his reagent is t,o convert t,he primary alcoholic groups in cellulose to acidic groups, to yield “celluronic acids”12gof varying degrees of oxidation. A number of other reagents, such as alkaline, neut,ral, and acid hypochlorite, hypobromite, hydrogen peroxide, ozone, permanganate, oxygen and alkali, etc., oxidize cellulose with production of acidic groups.125 Pectic material exists in all woods and often appears in holocellulose preparation^,'^^ alt,hough the amount is small in completely delignified holorellulose. Andersonrd7 prepared holocellulose contJaining 2 % of lignin (by t,he method of Wise138)from Douglas fir and Western hemlock, and extracted the pect,ic substances wit’h dilut>eammonium hydroxide. Thc (128) B. L. Browning, Ref. 9, p. 1249. (129) C. B. Purves, Ref. 9, p. 166. (130) A. Meller, T a p p i , 34, 171 (1951). (131) E. L. Jackson and C. R. Hudson, J . A m . C h e m . Soc., 69,2049 (1937); 60,989 (1938). (132) H. A. ltutherford, F. W. Minor, A . R. Mart,in and M. Harris, J . Research N a t l . B u r . Standards, 29, 131 (1942). (133) G. Jayine and S. Maris, Ber., 77, 383 (1944). (134) E. Pacsu, Textile Research J . , 16, 354 (1945). (135) E. C. Yackel and W. 0. Kenyon, J . Am. Cheni. Soc., 64, 121 (1942). (136) E. Anderson, J . Biol. Chein., 112, 531 (1936). (137) E. Anderson, J. Biol. Chem., 166, 233 (1946). (138) I,. E. Wise, Znd. Eng. Cheni. Anal. E d . , 17, 63 (1945).

POLYSACCIIARIDES ASSOCIATED WITH WOOD CELLULOSE

313

pectic material had then to be separated from lignin and hemicelluloses which were simultaneously dissolved by the ammonium hydroxide. About 0.5 % of the wood was obtained as purified pectic acid, in the case of Western hemlock. The somewhat elaborate procedure necessary for isolation and purification precluded a satisfactory quantitative estimation. Galacturonic acid was identified by conversion to galactaric acid by the method of Heidelberger and G0ebe1.l~~ Pectic materials were isolated, by the same technique, from other moods, including Western red cedar, loblolly pine, and black spruce. The hemicelluloses of wood78 contain uronic acid residues which frequently bear methyl ether groups. Hydrolysis of the hemicelluloses yields aldobiouronic and aldotriouronic acids. In researches rcferred to previously ~~-~~ the hemicelluloses from oakwood. (page 300), O ’ D n ~ y e r investigated She isolated an aldobiouronic acid as a barium salt after combined enzymic and acidic hydrolysis. Analyses of this compound were in agreement with the empirical formula for a barium salt of an O-methyl-xylo-aldobiouronic acid. This compound was isolated from a polysaccharide material which contained xylose (G parts), uronic acid (1 part), and methosyl (1 part). Studies by 77 on the hemicelluloses of a number of hardwoods also showed that O-methylhexuronic acid residues are present in glycosidic union with xylose, in ratios extending from 8 to 19 xylose units per uronic acid unit. The work of Jones and Wiselo (p. 286) and subsequent work by Gustafsson and coworkersE0 120 (p. 303) has established that 4-0-methyl-D-glucuronic acid is present in an “acidic xylan” which has been obtained from certain holocellulose preparations.80’ l20 Indeed, it has been stated120that 4-0-methyl-~-glucuronicacid is the predominant uronic acid in technical pulp. The polyuronides of pulp can, therefore, be regarded as acidic xylans (see Fig. 2, p. 303) which are originally present in the wood.120 Studies on the mannan-containing hemicelluloses of the conifers have failed to yield evidence that uronic acid and mannose units exist together in a single polysaccharide molecule. Anderson aiid coworkers79isolated an aldotriouronic acid from white-pine hemicellulose, but this acid contained only xylose units in addition to the uronic acid. Wise states140that the fractions of hemicelluloses from black spruce and slash pine which arc least soluble i n alkali contain the greatest proportions of mannose units. Similarly, Anderson found that the least soluble hardwood hemicellulose fractions have the highest xylan contents. The solubility of the hemicelluloses is undoubtedly related to their content of acidic groups. Bjorkqvist arid Jorgensen found that the uronic acid content of holocellulose from 3

(139) M. Heidelberger and W. F. Goebel, J . Biol. Chem., 74, 613 (1927). (140) L. E. Wise, Ref. 9, p. 379.

314

W. J. POLGLASE

spruce (Picea excelsa) was 5.1 %, and for birch (Betula verrucosa) a value of 6 % was obtained.14' After extraction of the alkali-soluble hemicelluloses, the remaining materials contained, respectively, 0.01 and 0.14 % of uronic acids, showing that the majority of the carboxyl groups in these preparations belong to the hemicellulose fraction. It is reasonable t o conclude that the carboxyl groups of wood cellulose are associated with the hemicellulose fraction, or with small amounts of pectic materials, or with the cellulose itself (in which case they probably arise as a result of oxidation by the reagents used in preparation of the cellulose). b. Determination of Carboxyl Groups.-The total amount of uronic anhydride present in a plant material or in wood cellulose is generally determined by boiling the substance with 12 % hydrochloric acid and measuring the carbon dioxide The estimation of uronic acid in mood and cellulose has been reviewed recently by B r o i ~ n i n g . 'A~ ~method specifically applicable to cellulosic material has been developed by Whistler, Martin and Harris.144Kenyon and a ~ s o c i a t e shave l ~ ~ reported on a kinetic study of the evolution of carbon dioxide from celluronic acids prepared by oxidation of cellulose with nitrogen tetroxide. The higher the titratable-carboxyl value (calcium acetate method), the more closely did the gas-evolution curves approach that of D-glucuronic acid. Substances of non-uronic structure, which might be expected to arise by oxidation at various points in the anhydro-D-glucose units, were also investigated. These compounds, tartaric, D-gluconic, oxalic, and glyoxylic acids, potassium acid D-glucarate, D-glucono-y-lactone, cellobiose, D-glucose, and D-xylose, yielded only small amounts of carbon dioxide, as a linear function of time, in contrast to the evolution of carbon dioxide from uronic structures, which followed an exponential function. Kenyon and associates146have compared the uronic acid values obtained by the above method with the carboxyl values obtained by the calcium acetate rnethod.l47 In the latter procedure, a calcium acetate solution is added to cellulose which contains free carboxyl groups. A replacement reaction takes place, and the acetic acid liberated is titrated with sodium (141) C. Bjorkqvist and L. Jorgensen, Acta Cheni. Scand., 6,978 (1951). (142) K. U. Lefhvre and B. Tollens, Ber., 40,4513 (1907). (143) B. 11. Browning, Ref. 9, p. 1206. (144) R. L. Whistler, A. R. Martin and M. Harris, J . Research Natl. B u r . Standards, 24, 13 (1940). (145) E. W. Taylor, W. F. Fowler, Jr., P. A . McGee and W. 0. Kenyon, J . Am. Cheni. Soc., 69, 342 (1947). (146) C . C . IJnruh, P. A. McGee, W. F. Fowler, Jr., and W. 0. Kenyon, J. An7. Chem. SOC.,69, 349 (1947). (147) M. Ludtke, Angew. Chem., 48, 650 (1935); Biochem. Z., 286, 78 (1936).

POLYRACCFTARIDER ASSOCIATED WITH WOOD CEI~LULOSI?

31 5

hydroxide. Keiiyo11~~~ found lower values by this method than by the uronic acid method. It was suggested'46 that,, whereas the rigorous hydrolytic conditions of the uronic acid method decompose all rarboxyl groups in uronic acid uiiits (whether free or bound), the calcium avctatc method measures only free varboxyl groups. Other methods for drtcrminiiig carboxyl groups iii cellulose have been reviewed recently by 13rowriiiig.'2s A new method, which appears capable of great precision, has been developed by Ant-Wuorinen for determining the carboxyl content of cellulose.i48Improvements in techniques were later proposed by Ant-Wuorinen and V i ~ a p a a . In ' ~ ~this method, which requires specially designed equipme~i t , ' (the ~ carboxyl groups of the cellulose sample are liberated by a prolonged, automatic, washing operation with water saturated with carbon dioxide. The cellulose sample (freed from carbon dioxide) is allowed to react, under a nitrogen atmosphere, with sodium chloride solution. An ionexchange reaction takes place and the acid, liberated by the carboxyl groups of the cellulose, is titrated with sodium hydroxide solution. During the titration, the pH is followed by means of a pH-meter connected to a recording potentiometer. Special precautions are taken to eliminate interference by carbon dioxide and bicarbonates.149 c. Properties of Carboxyl Groups in Wood Cellulose and Derivatives.-According to Meller,130 the oxidation of the terminal aldehyde group of cellulose to yield "gluconic acid cellulose" increases the carboxyl value, and yields a product of high stability toward hot alkali. Meller postulatesi3n that chlorine bleaching agents, when employed under certain conditions, may oxidize the end groups, thus influencing the response of the cellulosic material to subsequent, alkaline purification. Kenyon and coworkersls0 found that celluronic acids are degraded in alkaline solution to generate acidity, and suggested that this alkali-lability is due to the presence of small proportions of ketonic groups in these products. They postulated15nthat the ketone groups enolize in alkali, the enediols then split, and adjacent glycosidic links hydrolyze in such a way as to initiate an extensive alkaline degradation. Jayme and von Koppen have stressed the importance, for paper-making, of the carboxyl groups in wood pulp.I6*Saarnio has shown a direct relationship between hydrophilicity, rate of beating, and carboxyl content of wood pulps.'52 (148) 0. Ant-Wuorinen, Paperi j a Puu, 33B, 105, 174 (1951). (149) 0. Ant-Wuorinen and A. Visapaa, Paperija Pzcu, 36,233 (1954). (150) P. A. McGee, W. F. Fowler, Jr., C. C . Unruh and W. 0. Kenyon, J . A m . Chena. Soc., 70, 2700 (1948). (151) G. Jayme and A. von Koppen, Das Papier, 4, 373, 415, 455 (1950). (152) J. Saarnio, Paperija Puu, 36, 217 (1953).

316

W. J. POLGLASE

Evans and S p ~ i r l i n have ’ ~ ~ studied the effect of carboxyl groups present in O-ethylcellulose on its viscosity in dilute solution. Although free carboxyl groups do not affect viscosity, neutralization with metal ions increases the viscosity, especially in non-polar solvents. According to these workers, the effect of bound, metal ions is to produce large electrostatic forces of attraction between the chains to which they are attached.

IV. PREPARATION AND COMPOSITION OF WOODCELLULOSE 1. Holocellulose

The name “holocellulose” mas assigned by Ritter and KurthIS4to the residue obtained after delignification of wood by alternate treatments with TABLE XI1 Analysis o j Maple Wood and Holocelluloseo 1

2 Percenl o j original i n (3)

Properties

Yield, % Lignin, % ’ Methoxyl, OCH, , % C02 (from uronic anhydride), % Acetyl, CH3C0, % Pentosan, % Ash, % a From d a t a of Ritter and Kurth.lK4 hot water.

100 22.8 6.10 1.09 3.59 19.6 0.30 b

78.0 1.9 1.42 1.43 4.45 24.5

76.2 0 1.24 1.37 4.68 24.9 0.29

15.6 96 100 96.9

Extracted with alcohol-benzene, and

chlorine and alcohol-pyridine. As implied by the term, holocellulose, t>his residue was believed154to contain all of the carbohydrates of the plant tissue. I n the delignification process, chlorine reacts with lignin to form chlorolignin, which is then dissolved by the treatment with alcohol-pyridine. Some of the analyses reported by Ritter and Kurth for maple wood are given in Table XII. I n later work, Van Beckum and RitterlS5found that 3 % ethanolamiiie in 95 % ethanol is more effective than alcohol-pyridine for the cxtractioii of chlorolignin. An outline of the method follows. The wood extractives are removed by treating the wood mpal (60 t o 80 mesh) with 95% alcohol for 4 hours, then with alcohol-benzene, and finally with water (in a boil(153) E. F. Evans and H. M. Spurlin, J . A m . Chena. Soc., 7 2 , 4750 (1950). (154) G. J. Ritter and E. F. Kurth, Ind. Eng. Chem., 26, 1250 (1933). (155) W. G. Van Beckum and G. J. Ritter, Paper Trade J., 106, No. 18, 127 (1937).

POLYSACCHARIDES ASSOCIATED WITH WOOD CELLULOSE

317

iiig wat>cr bath for 3 hours). Thc extractive-free wood is air-dricd, treated wit,h chlorine gas for 5 niiiiutes, washed n i t h alcohol, and then t rcatcd with hot, alcoholic itionoet llanolamine for tlvo successive 2-minute intervals. The residue is next washed n-ith alcohol and water. The chlorination and extract,ion treatments are repeatcd unt i l a white product is obtained; t,his is washed thoroughly with alcohol, cold water, :tlcohol, and cther, and is thcii ovcn-dried.

If the holocellulose so obtained contained the entire carbohydrate fraction of the wood, the sum of holocellulose and lignin for t'he extractive-free ~ - ' was ~ ~ found t,o he wood should equal 100%;in a number of ~ t u d i e s , ' ~this true. In later work, however, it was no-ted that part of the ethanolamiiie 159~Comused to extract the chloroligniii is retained by the h o lo c e ll~ lo s e .~ *~ pensating for this increase in weight,, there was rioted to be a slight, loss in polyuronide-containing material during the holocellulose isolation.i59 A number of investigators have observed that there is also a small decrease in peritosan during the isolation of holocellulosesi60-i62by this procedure. In another method for isolating the carbohydrate fraction of wood, chlorine dioxide is used, instead of chlorine, and the lignin derivatives formed are ext,racted out with pyridine-water. This method was first, reported by Schmidt and G r a ~ m a i i n Several . ~ ~ ~ years later, J a ~ m e ' ~ ~modified , the procedure by using sodium chlorite and acetic acid in place of the explosive, chlorine dioxide gas. Jayme's method was subsequently modified by Wise and coworkers,7. I L 9 , and, in this form, has been used extensively in the characterization of wood. The procedure may be applied directly to airdried wood without previous extraction, except in the case of resinous woods, where a preliminary treatment with alcohol and ether is desirable. I n the method, a 5-gram sample of wood meal (40 t o 60 mesh) is heated at 70 to 80°C. with 160 ml. df water containing 1.5 grams of sodium chlorite and 0.5 ml. of glacial acetic acid. After 1 hour, fresh portions of acetic acid (0.5 ml.) and sodium chlorite (1.5 grams) arc added, and the heating is continued. This treatment is rcpeated until a white product, containing2 to4% of lignin, is obtained. For hardwoods, three treatments are generally sufficient, whereas four treatments are usually rcquired for coniferous woods.

As in the Van Beckman and Ritter chlorination procedure, there is also (156) (157) (158) (159) (1945). (160) (161) (162) (163) (164) (165)

R. D. Freeman and F. C. Peterson, Znd. Eng. Chem. A n d . E d . , 13,803 (1941). A. B. Anderson, Znd. Eng. Chem., 36, 662 (1944). J. E. Atchison, Paper Trade J . , 116, No. 22, 23 (1943). 13. B. Thomas, Paper Znd. and Paper W o r l d , 26, No. 10, 1281; 27, No. 3, 374 G. Jayme, K. Storch, E. Kerler and G . Schwab, Papier-Fabr., 37,57 (1939). C. V. Holmberg and E. C. Jahn, Paper Trade J.,111, No. I, 33 (1940). T . E. Time11 and E. C . Jahn, Svensk Papperstidn., 64, 831 (1951). E. Schmidt and E. Graumann, Ber., 64, 1860 (1921). G. Jayme, Cellulosechemie, 20, 43 (1942). G. Jayme and G. Schwab, Papier-Fabr., 40, 147 (1942).

318

W. J. POLGLASE

some loss of carbohydrate material in the chlorite procedure for prepariiig holocellulose. I t has heen shown162that, for paper-birch wood, the loss of pentosaii is greater a t the same ligiiin content than in the chloriiiation procedure. An investigation166on liquors obtained after treatment of spruce wood by the chlori tc procedure showed the presence of polysaccharides. This material could be precipitated by the addition of alcohol, and upon hydrolysis and paper chromatography was found to contain mannose, galactose, and uronic acids, with smaller quantities of xylose and arabinose.166The preparation of holocellulose by the above methods has been It has been re-investigated in order to compare carbohydrate 10sses.l~~ stated that the chlorine-ethaiiolamine met hod results in less carbohydrate loss than does the chlorite procedure.167 Nevertheless, the holocellulose fraction of mood (as prepared by these relatively specific deligiiificatioii methods) undoubtedly represents the least degraded material currently available for a study of the carbohydrates associated with wood cellulose. Investigations have shown that the methods of holocellulose preparation may also be applied to non-woody plant material.l6*,169 Holocellulose appears to have certain advantages over whole wood as a raw material for the study of hemicelluloses. When wood is used as the starting material, a considerable part of the hemicellulose is not extractable by dilute alkali, except after removal of the lignin. On the other hand, a series of hemicellulose fractions may be obtained quite readily from holocelluloses.7 When wood is extracted with alkali, that part of the lignin that is removed simultaneously with the hemicellulose causes difficulties in the recovery of the hemicellulose fraction. In alkaline extractions of holocellulose, however, this difficulty is not encountered. Wise140has suggested that it may be a disadvantage to start with holocellulose, in studies on uronic acid fractions, since there may be the complication of aldonic acid formation because of the oxidizing action of the delignifying agents. Wise and Ratliff7 extracted slash pine (Pinus caribaea) chlorite holocellulose a t 20" with increasing concentrations of aqueous potassium hydroxide. In the range of 2 to 10 % potassium hydroxide, the larger part of the hemicelluloses dissolved. Thereafter, less material dissolved, and above 16 % potassium hydroxide, the weight of residue remained constant. The hemicelluloses could be recovered quantitatively by acidification of the alkaline extracts with acetic acid, followed by precipitation with an excess of ethanol. For the practical purposes of analysis, Wise and Ratliff chose to divide the (166) (167) (168) (169)

W. J. Bublitz, T a p p i , 34, 427 (1951). B. I,. Browning and I,. 0. Bublitz, Tnppz, 36, 452 (1953). E Bennett, Anal ("hem , 19, 215 (1947). G. A. Adams and A. E. Castagnc, Can. .I. Research, B26,325 (1948)

319

POLYSACCHARIDES ASSOCIAT’ED WITH WOOD C E I J L ~ J L O S E

hemicelluloses isolated from holocellulose into two fractions: A , the fraction removed by 5 % potassium hydroxide; and B , the fraction removed from the residue by 24 % potassium hydroxide. The data as presented by Wise and Ratliff do not include carbohydrate aiialyses 011 holocellulose preparations. However, analyses for xylan (pentosan method), mannan (phenylhydrazone met>hod),and uronic anhydride were performed on the original wood samples aiid, if it be assumed that these substances were retained quantitatively in the holocellulose fraction, the approximate carbohydrate composition of the holocellulose may be calculated (see Table XIII). TABLE XI11 Carbohydrate Content of Holocelliilose~ Source

______

Slash pine Douglas fir Western hemlock Loblolly pine Black spruce Western red cedar Southern red oak Overcup oak

67.8 71.3 67.1 70.3 68.9 67.4 60.0 67.2

12.2

8.7 10.9 14.4 15.2 12.1 29.0 27.1

11.5 7.6 6.1 6.7 11.6 7.6

6.8 3.9 7.5 5.4 5.9 6.2 6.6 8.6

67.1 67.7 66.4 66.2 66.1 70.5 63.4 60.4

a Calculated from the d a t a of Wise and Ratliff.’ Pentosan corrected for c Phenylhydrazone method. d Method of Burkhart, Raur uronic anhydride. and Link.”* 6 Residue after extraction of holocellulose with 24% potassium hydroxide, corrected for residual lignin, xylan, mannan, and uronic anhydride.

Analyses performed’ on the alpha-cellulose residue, obtained after extraction of the holocellulose with 5 70and 24 % potassium hydroxide, showed that a large part of the maiman (50 % or more) aiid a part of the xylaii (less than 20%) and uronic anhydride (about 30%) were retained in the socalled alpha-cellulose. Corrections were, therefore, applied in order to establish the true cellulose content of the wood (see Table XIV). I n one case, the alpha-cellulose residue from a holocellulose preparation has been studied by methods which yield information concerning structure (see page 295). It is to bc hoped that work of this kind will be continued and, furt>her, that the techniques of structural Carbohydrate chemistry mill continue to be applied to the elucidation of the nature of the hemiccllulosic fractions which may be obtained so readily from holocellulose prepara(170) B. Burkhart, L. Baur and K. 1.’ Link, J .

Biol. Chem., 104, 171 (1934).

320

W. J. POLGLASE TABLE c'cii

~*an"an,%

Il'ood

Slash pine Douglas fir Western hemlock Loblollg pine Black spruce Western red cedar a

x v

bohtldrate Analilsis of Alpha-cellulose P r e p a r e d f r o m Holocelltclose

2.6 1.7 1.8 2.8 1.7 1.5

10.4 10, 9a 8, 9" 9, 9a 7.1 6, 7"

Uronic

anhydride,

o/n

Re/erences

2.6 2.5 1.7 2.0 2.1 1.3

7 171 171 171 7 171

Determinations on separately isolated alpha-ccllulose preparation^.'^'

tions. The recent work of the Finnish groupSO. illustrat,es the importance of such studies. The investigations of Mitchell and R i t t e P on holocellulose isolat'ed from sugar maple are of interest,. These workers isolated 25 % of the holocellulose in four, levorotatory, soluble fractions by successive breatments with hot water, cold 2 % sodium carbonate, cold 4% sodium hydroxide, and hot 10 % sodium hydroxide. The fraction extracted with hot water contained acet'yl groups. All fractions contained xylan, and uronic anhydride and methoxyl were found in them in equivalent proport.ions. Wise has recently suggested173t>hatfundamental studies on holocellulose should now be undertaken. He proposes that holocellulose prepared by both the chlorit,e procedure and the chlorine-alcohol-ethanolamine technique should be methylated, and t'he products fractionated and methanolyzed according to t>heestablished techniques of carb0hydrat.e chemistry. Ligiiin can be removed by a number of other procedures, but these, in general, result in incomplete delignification or in serious loss or degradation of polysaccharides. The classical procedure of Cross and Bevanli4 employs extraction with 1 % sodium hydroxide, followed by repeated chlorinations and extractions with hot 2 % sodium sulfite solution. The yields of cellulosic material obtained in this procedure are considerably lower t'han those obtained by the holocellulose procedure. Van Beckum and R i t t ~ showed ' ~ ~ that hydrolysis of holocellulose nit'h hot 1.3 % sulfuric acid for 2 hours yields residues which are equal in and similar in chemical properties,176to Cross-and-Revan cellulose obtained from the same wood. (171) H. F. Lewis, T a p p i , 33, 299 (1950). (172) R. I,. Mitchell and G. J. Rit,ter, J . A m . Chent. Soc., 62, 1958 (1940). (173) L. E. Wise, Ref. 9, p. 408. (174) E. J. Bevan and C . F. Cross, J . Chem. SOC.,38, 666 (1880); C. F. Cross, E. J. Bevan and C. Beadle, ''Cellulose," Longmans, Green and Co., London, 1895. (175) W. G. Van Beckum and G . J. Ritter, Paper Trade J., 108, No. 7, 27 (1939).

POLYSACCJIARIDES ASSOCIATED WIT11 WOOD CELLUT,OGE

32 I

h iium\m of other deligiiificatioii procedures have heen devised in which the primary objective has been to obtain, for various purposes, a measure of the total-carbohydrate fraction. These procedures have employed as the active delignifying agents such materials as nitric acid,li6 monoethanola* l i 8 atid other amines,17* neutral h y p o ~ h l o r i t eavid , ~ ~ ~hypochlorite,’sO etc. These methods have recently been discussed by RrowningLS1and by .Jahn.L82Although such procedures are of general interest, their application has not yielded much information that could be considered fundamental to an understanding of the chemistry of wood cellulose. Neither have these procedures herome important in the commerrial production of wood cellulose. 2 . Commercial Wood Cellulose

a. General.-Technical wood pulp is prepared by one of the following three pulping procedures: the sulfite, the sulfate, and the soda processes. In t’he first of these, the cooking liquor is acidic; in the other two, it is alkaline. The scope for variation of reaction conditions is enormous. Some of the more obvious variables are: type of raw material, dimensions of wood chips, concentration of active ingredients of pulping liquor, liquor-to-wood ratio, time-temperature relationship, washing and screening procedures, etc. Added to this are the possibilities for endless variations of methods for bleaching and purification, or for treatment of wood chips prior to pulping. The selection of specific conditions depends on the u1t)imate use for which the cellulosic material is intended. For a full understanding of pulping processes,18~a knowledge of the anatomy of ~ 0 0 d ~is~important, ~ - ~ 8 ~since marly of the problems of pulping are associated with such factors as ensuring penetration of wood chips by cooking liquors and avoiding mechanical damage to fibers. The nature of the reaction of pulping reagents with lignin is also of prime importance to (176) G. dayme arid P. Schorning, Papier-Fabr., 36, No. 21/25, 235; No. 37, 393 (1938). (177) L. E. Wise, F. C. Peterson and W. M. Harlow, Ind. Eng. Chem. A n d . E d . , 11, 18 (1939). (178) P . Bloom and E. C. Jahn, Tech. Assoc. Papers, 24, 127 (1941). (179) A. G. Norman and S. H. Jenkins, Biochem. J . (London), 27, 818 (1033). (180) J . H. Ross, A. L. Davidson and E. 0. Houghton, Pulp & Paper Mag. Can., 27, No. 25, 925 (1929). (181) B. L. Browning, Ref. 9, p. 1153. (182) E. C. Jahn, Ref. 9, p. 1021. (183) J. N. Stephenson, ed., “Pulp and Paper Manufacture,” McGraw-Hill Book Co., Inc., New York, N . Y., 1950. (184) H. P. Brown, Ref. 9, p. 7. (185) W. M. Harlow, Ref. 9, p. 99. (186) G. J . Ritter, Ref. 1, p. 286.

322

W. J. POLGLARE

an understanding of pulping processcs. Such aspects will receive oiily cursory att,ention in the following discussion. The chief object'ive will be to outline the apparent effect of certain chemical pulping processes on the carbohydrate composition of wood pulps. 0. Sulfcte Pulps.-The process for making white pulp from wood by treatment with sulfurous acid or calcium bisulfite under pressure was discovered by Tilghman.ls7,lS8 In spite of many difficulties, the process eventually permitted product,ion of pulp a t a profit, and, t,oday, it is still one of t'he most important processes for the preparation of wood cellulose. The chemistry of delignification in the sulfit,e process has been discussed recentJy by Hagglu1id~8~ and by E r d t m a i ~ 'Depolymerization ~~ of polysaccharides occurs to an extent dependent, on the acidit,y of t8hecooking liquors, the reaction time, and the temperature of reaction. This has been shown by reducing-sugar analysis on liquors during t3he sulfite cook.1g1Evidently, complex sugars are liberated during the early stages of cooking, since acid hydrolysis increases t,he reducing value of sampled cooking liquors. According t,o H a g g l ~ n dthe , ~ ~final, ~ wast,e, sulfite liquors contain, chiefly, monomeric sugars and aldonic acids (formed as the result of the oxidation of sugars by bisulfite ions). Both D-mannonic and n-xylonic acids have been isolated from sulfite, waste liquors.18g The order of appearance of different sugars in sulfite cooking liquors has been studied with filter-paper chromatographic techniques by Sundman.112 The liquors were sampled a t various stages during cooks of spruce, pine, birch, and aspen. Arabinose appeared in the cooking liquors before the temperature had reached loo", and increased rapidly ill amount, eventually decreasing toward the end of the cook. Xylose and galactose appeared a t the same time, and a little later than arabinose, but their proportion did not decrease on prolonged cooking. Mannose was detected when t,he ternperature of digest>ionreached I 30°, and glucose at 140". The quantitative data obtained by Isbell and Frush1g2on hydrolysis by dilute hydrochloric acid of met,hyl glycosides are of interest, in this connection. From these studies,lg2 it may be deduced that the rat,es of hydrolysis of the methyl 0-D-pyranosides of t,he wood sugars arc i n the following order: D-xylOSe = n-galactose > n-mannose > D-glucose. The ext,reme acid(187) B. C. Tilghman, U. s. P a t . 70,483 (1867). (188) For a history and an outline of the sulfite process, see G . C . McGregor, Ref. 183, Vol. 1, p. 252. (189) E. Ragglund, "Chemist,ry of Wood," Academic Press Inc., NPWYork, N . Y., 1951. (190) H. Erdtman, Ref. 9, p . 999. (191) E. Hiiggluntl, Ref. 189, p. 430. (102) H. S. Isbell and Harriet I,. Frush, J . Research N a t l . Bur. Slandards, 24, 125 (1940).

POLYSACCHARIDES ASSOCIATED W I T I I WOOD CELLULOSE

Analysis

I

Spruce Spruce Pine Aspen Aspen Birch Birch

3.75 6.50 6.50 2.50 5.25 2.75 5.50

I

0.f

323

TABLE XV Unbleached, Sulfite Pulps46

-_

58.3 52.1 54.0 55.6 51.5 54.6 48.9

87.1 88.1 85.6 83.3 86.3 77.1 81.6

_____

89.5 90.1 86.2 88.0 90.4 83.4 87.5

6.3 6.0 7.7 2.1 2.0 1.4 1.5

4.2 3.9 6.1 9 .9 7.6 15.2 11.0

Cooking liquor: total 802 , 71.1%; CaO, 1.3%. Time t o 60°, 0.5; time from 60' t o 105", 3; time from 105" to 130°, 0.5 hours. Maximum temperature, 130".

lability of L-arabinose units in polysaccharides is well known and is related to the furanose-ring form usually exhibited by this sugar in polysaccharidcs. Thus, the order in which these sugars appear in sulfite cooking liquors is consistent with the known stability, toward acids, of the glycosidic bonds of model substances, The final liquors from pine and spruce mere rich in mannose and galactose, whereas birch and aspen liquors were high i n xylose.l I 2 I n further studies on the sulfite pulping process, Sundman, Saarnio and G u s t a f s ~ o nanalyzed ~~ a number of sulfite pulps by hydrolysis, followed by paper chromatography. Some of their results are shown in Table XV. Although galactose and arabinose had been detected in complete hydrolyzates of wood, by chromatographic methods (see Table XV), neither of these sugars could be detected in the hydrolyzates of sulfite pulps. After chlorination and hypochlorite bleaching of these sulfite pulps, the mannan and xylan content did not change a p p r e ~ i a b l y . ~ ~ For the purpose of conversion to textile fibers, dispersions or solutions of cellulose or its derivatives are achieved by various means, in order to make possible the extrusion of the fiber-forming material through the small orifices of the spinning jets. Wood pulps prepared for these and similar uses (such as the manufacture of cellophane) are known as "dissolving pulps.'7 The manufacture of dissolving pulps is a highly developed art, with processes protected by or, more effectively, within company files. The purification of siilfite pulps to a degree suitable for manufacturc of textile rayon (YO to 9.1%) alpha-cellulose), tire cord (94 to 95% alpha-cellulose), ' alpha-rellulosc) requires some kind of and cellulose ncetnte (95 to 96 % (193) Sce G. rJ:Lyme, Paper Trade J . , 106, NO. 21, 37 (1938).

324

W. J. POLGLASE

alkaline treatment,Ig4in addition to treatments for removal of residual lignin. Such pulps are frequently termed "high-alpha pulps." According to Hisey,Ig4the increase in alpha-cellulose content results from the removal of hemicelluloses and short-chain cellulose by dispersal in alkali, which leaves the longer cellulose molecules intact. Sodium hydroxide appears to be the most important alkali employed in commercial operations, a.nd two general methods for its use are of importance. One method employs dilute, sodium hydroxide solutions a t elevated temperatures, and t,he other uses concentrated solut'ioiis a t low temperatures. In the hot, alkaline refining process, temperature, time, concentration of alkali, and consistency of pulp may be varied over wide limits. The literature contains only a limited amount of data showing the effect of alkaline treatment on sulfite-pulp composition. However, from the available data, certain principles are apparent. Norman116has studied the effect on xylan of hot, alkaline extraction of cellulose prepared from oak, wheat, barley, and jute by the hypochlorite method of Korman and ,Jenkins.179It was observed'I6 that the xylan content of t,hese cellulose preparations can be diminished by boiling with various concentrations of alkali, but, in agreement with previous work by Heuser and Haug,Ig5an appreciable percentage of the original xylan could not be removed. Furthermore, it was noted that continued boiling with alkali removes hexosan (of undetermined configuration) a t a greater rate than xylan. Examples of the diminution of th.e pentosan of wood pulps by alkaline purification may be taken from the work of Richter.Ig6In a series of papers dealing chiefly with the pulping of Richter showed that either hot, dilute, sodium hydroxide or more concentrated sodium hydroxide a t ordinary temperatures removes a substantial part of the pentosan from sulfite pulps. Some of Richter's data are shown in Table XVI. In attaining a high alpha-cellulose with simultaneous removal of pentosail, there is a considerable loss of other material, presumably hexosan in nature. Although higher purity can be attained by treat'ment with cold, concentrated sodium hydroxide, this results in mercerization of the cellulose. Mercerized cellulose is of no use in the acetylation process unless it is specially treated i n order t,o avoid inactivation by The effect of alkaline treatment, on the mannan content of wood pulps (194) W. 0. Hisey, R.ef. 9, p. 1055; M. Martin, Can. Pulp and Paper, 6. No. 6, 8 (1952); L. K. Bickell, ibid., 6, No. 3, i 2 (1953). (195) E. Heuser and A . Haug, 2 . angew. Cheni., 31, 166 (1918). (196) G . A. Richter, Znd. Eng. ( ' h e m . , 33, 1518 (1941). (197) G . A. Itichter, Znd. Eng. ('hem., 33, 75, 532 (1941). (198) F. Olsen, Znd. E,ng. C h w i . , 30, 524 (1938).

POLYSACCIIARIDES ASSOCIATED WIT11 WOOD CELLULOSE

325

TABLEXVI Reniooal of Pentosans from Unbleuched, Suljife P u l p s l g ~ Pulp

'".*

h'aOIl added

Yield, %

Alphacellulose,

%

%

Penlosan,

~~~

Softwood

Mixed hardwoods

White birch

0 10% (on pulp)" 15% (solution concentration)

125 25

82.2 86.5

87.7 94.6 06.9

4.5 2.5 0.93

0 10% (on pulp)"

125

84

87.8 94.0

7.2 3.1

83.6

86.8 96.3

8.4 1.9

0 18% (solution concentration)

30

a At a consistency of lo%, this is equivalent t o a 1% solution concentration of sodium hydroxide.

has not been reported. However, Sundman and coworkers45state that a pulp having low manna11 content ( % unspecified) was obtained when a spruce, sulfite pulp was digested by a mild, sulfate cook. Presumably, the mannan content of sulfite pulps also is reduced by refining in alkaline solutions. It has been noted (page 319) that mannan is not as readily removed from softwood holocellulose by cold, concentrated alkalis as is xylan. It would appear, then, that the effect of the sulfite process on wood polysaccharides is primarily a depolymerization which leads in part to soluble sugars (predominantly from the more readily hydrolyzed, non-glucose polymers). The sulfite pulp, as normally obtained, contains a certain amount of low molecular-weight xylan, mannan, and glucan. Subsequent alkaline treatments disperse the main part of this short-chain material, to yield high-alpha cellulose containing residual amounts of inaccessible or high D. P., non-glucose polyoses. It has been shown120 that pulps prepared by the sulfite process contain uronic acid anhydride residues (apparently, 4-O-methyl-~-glucuronic acid anhydridelZ0). The uronic acid content of such pulps can be diminished by hot alkaline refining.lZ0 c. Alkaline Pulps.-Two general processes for pulping wood with alkali are important. In the soda process,1ggthe active component of the cooking liquor is sodium hydroxide. The stability of the glycosidic bond, together with the insolubility of cellulose, permits the use of high temperatures and high concentrations of alkali in effecting delignification. According to the theory of Koon,200lignin is dissolved as sodium lignate, and part of the hemicelluloses are removed by a combination of dissolution, colloidal dis(199) c. Watt and H. Burgess, U. S. Pats. 1,448 (1854); 1,449 (1854). (200) C. M. Koon, Ref. 1, p. 502.

32G

W . S. POLGLASE

persion, and degradation. Koon200 discusses the work of I3rauris and Grimes,'O' who had concluded that about half of the total alkali ronsumcd is used up by unknown mechanisms, t)o dissolve the carbohydrates. HagglundZo2 found lactones and hydrosy-acids (saccharinic, lactic), 1o the extent of 18.2 % of the wood, in the waste liquor ohtained from the alkaline pulping of spruce."? These are undoubtedly the products of alkaline degradation of the hemicelluloses removed from the wood. The sulfate process or kraft process has the advantage of higher pulp yield than is afforded by the soda process. The so-called "sulfate liquor" is a mixture of two components, sodium hydroside arid sodium sulfide. I n industrial practice, sodium sulfide is formed by the reduction of added sodium sulfate when the organic matter from a previous pulping operation is burned for chemical recovery. I n a review of alkaline processes, HolzerZo4 cites experimental work which shows that the higher pulp yield in the sulfate process is ascribable to the rate of dissolution of lignin, which is higher than in the soda cook, whereas the rate of dissolution of carbohydrate is the same in the two procedures. This results in a shorter reaction time for delignification, which, according to H a g g l ~ n d is , ~the ~ ~ most important reason for the favorable effect of sulfide. In a recent study on the dissolution and destruction of carbohydrates during the sulfate cook, Saarnio and Gustafssonlll reported a higher rate of destruction of dissolved hexosans than of dissolved pentosans. These workers"' isolated polysaccharide fractions from samples (taken hourly) of sulfate cooking liquor, and analyzed the fractioiis (after hydrolysis to monosaccharides) by quantitative, paper c h r ~ m a t o g r a p h y .They ~~ observed"' an increase in the proportion of polysaccharides in the cooking liquor in the early stages of the cook, when dissolution predominates. Soon after reaching the maximum temperature, however, dissolution decreases and destruction of polysaccharides becomes predominant. On comparing data on the carbohydrate composition of wood3* 206-208 with those on sulfate pulps, Sundman and coworkers45concluded that the galactans and arabans of wood are almost completely dissolved during the sulfate cook. About 70 % of the maiinan is dissolved from pine wood, as is about 60% of the mannan from birch. From pine, about 50 to 70% of the (201) F. E. Brauns and W. S. Grimes, Paper Trade J . , 108, No 11, 40 (1939). (202) E. Hagglund, Cellulosechemie, 6, 81 (1924). (203) Ref. 189, p. 467. (204) W. F. Holzer, Ref. 9, p. 975. (205) E. Hagglund, Ref. 189, p. 476. (206) C. Gustafsson, P. I. Ollinman and J. Saarnio, Acta C'hem. Scand., 6, 1299 (1952). (207) J. Larinkari, Finnzsh Paper Tinrber J . , 27, 143 (1945). (208) W. Jensen, Finnish Paper Timbe, J . , 31, No. 7A, 20 (1949).

POLYSAC(:IIhHIDES

327

ASSOCIATED WITH WOOD CELLULOSE

TABLEXVII ('urbohydrate Composition, of Unbleached, Sulfate Pulps" lannan, 7" Xylan, ?4

Araban.

76

~~

Spruce Pine Pinee Pine * Birch Birch"

47.7 44.2 39.9 37.8 57.3 48.7

89.3 90.2 90.9 92.9 81.4 86.1 81.7

84.4 84.4 88.3 92.8 74.5 76.4 82.8

4.7 5.6 5.9 2.8 1.2 1.2 0.5

9.9 9.0 5.3 4.4 23.8 22.4 16.8

a Cooked a t higher chemical concentration than the previous sample. Pre-hydrolyzed (2 hours a t 140'C.). hydrolyzed (2 hours a t 150°C.).

1 1 0.5 0 0.5

+ 0

* Pre-

xylan is removed, and from birch about 50 to 6075, depending on the degree of cooking. About 15% of the glucan in pine wood, but practically none of the glucan of birch wood, is dissolved in the cooking liquor.111 This glucan was determined"' as a yield loss, not as glucan in the alkaline cooking liquors. Since definite evidence for presence of glucan in the cooking liquors could not be obtained, it was suggested that pine may contain an easily destroyed glucan, either of low molecular weight or of a constitution other than that of a poly-(4 -+ 1 p-D-ghcoside). The carbohydrate analyses reported by Sundman, Saarnio and Gustafsson for sulfate pulps are listed in Table XVII. These results were obtained by quantitative, paper c h r ~ m a t o g r a p h yon ~ ~completely hydrolyzed samples. I n another study,'*O the uronic acid anhydride content of some wood pulps was determined. It was notedLz0that sulfate pulps prepared from hardwoods retain a considerable proportion of uronic acid, whereas sulfate pulps from softwoods may be entirely free from polyuronides. Some comparisons between the composition of sulfate and sulfite pulps are of interest (see Tables XV and XVII). The alpha-cellulose content of sulfate pulps is generally higher than that of sulfite pulps from the same species, but the glucan content is higher for the sulfite pulps. In every case, escept for prehydrolysed samples, the alpha-cellulose content of sulfate pulps is higher than the total, true cellulose (glucan) content, and, for sulfite pulps, the alpha-cellulose content is always lower than the true cellulose content. In the sulfate pulps, some non-cellulose polysaccharide is retained in the alpha-cellulose, whereas in sulfite pulps, at least some of the glucan must enter the beta- or gamma-fractions or both. When yields are considered in terms of the anhydro-D-glucose content of the pulp, the sulfite process is superior. The pre-hydrolyzed pine and birch sulfate pulps (see Table XVII) war-

328

W. J . POLGLASE

rant special comment, since the preparation of commercial, wood pulps by processes involving steam, water, or acid pre-hydrolysis is becoming increasingly popular. I n the case of pine, pre-hydrolysis yields a sulfate pulp containing about half as much xylaii and mannan as does the nonprehydrolyzed pulp. Similarly, for birch, a reduction in non-cellulose polyoses may result from pre-hydrolysis. When certain softwoods are pulped by the sulfite process, the resulting pulp usually contains a considerable proportion of uronic acid anhydride which cannot be removed entirely in subsequent alkaline refining. When the sulfate process is used 011 softwoods, a pulp \vhich is free from uronic acid anhydride may be obtained. This important difference between the sulfite and sulfate pulps from softwoods does not apply to hardwoods, which, when pulped by either process, still retain a considerable proportion of uronic acid anhydride in the final product.120

V. FINESTRUCTURE OF WOODCELLULOSE A N D ASSOCIATED POLYSACCHARIDES 1. Alpha-, Beta-, and Gamma-cellulose

.4 number of methods of analysis are important in wood-cellulose chemistry. One of the most important is the determination of solubility in sodium hydroside solution a t room temperature. This distinguishes between alphaccllulose (not dissolved by 17.5 % sodium hydroxide), beta-cellulose (dissolved by 17.5 % sodium hydroxide, but precipitated when acidified with acetic acid), and gamma-cellulose (dissolved by 17.5 % sodium hydroxide, and not precipitated on acidification). The empirical nature of this fractionation may be appreciated by comparing the composition of a number of different preparations of alphacellulose. The alpha-cellulose from such coniferous woods as spruce, pine, and hemlock may contain appreciable amounts of mannan, although the xylan content is usually low. Similarly, the alpha-cellulose from hardwoods may contain appreciable quantities of non-cellulosic polyoses, although, in this case, the chief polyose is xylan, not mannan. On the other hand, certain types of wood pulp, prepared by sequences involving both acidic and alkaline treatments, show a linear relationship between gamma-cellulose and non-cellulosic polyoses. This may be illusstrated from the studies of White, Steinman and Workb0on acetylation pulps. Acetylation-grade pulps are usually prepared from softwoods by sulfite cooking followed by hot-caustic refining. When the total polyose content of a number of such pulps was compared with their gamma-cellulose content, a linear relationship was obtained (see Fig. 3 ) . In spite of this excellent correlation, it cannot be assumed that all of the non-cellulosic polyoses are contained in the gamma fraction of these pulps (see below).

POLYSACCHARIDES ASSOCIATED W I T H WOOD CELLULOSE

:329

u) W

0 J

5.0

J

W 4 0 0

a

$ 3.0 a

0

0 lL 2.0

:: 1.0 W

b-

z :0 ~

g

t o o

10

2 .O

3 0

PERCENTAGE O F M A N N A N

PLUS XYLAN

FIQ.3.-Relationship Between the Gamma-cellulose Content and the Mannan Plus Xylan Cont.ent of Wood Pulp.’13

The limitations of the alpha-cellulose determination have become more apparent as the need for pulps of greater purity has increased. For certain uses, particularly in the manufacture of cellulose-acetate rayon, it has been deemed desirable to prepare a “high-alpha” pulp, since this was believed to be synonymous with high purity. This is not surprising in view of the high-alpha content of purified, cotton linters (99 %). High-alpha, cellulose pulps can be most readily obtained by a modified, alkaline (kraft), pulpiiig process. Yet, despite their lower alpha-cellulose content, hot-caustic refined, sulfite pulps produce superior acetates. Current studies on woodcellulose composition point to the non-cellulosic polyoses as of importance to the properties of the product. In certain cases, the higher-alpha pulps may actually have a lower, true-cellulose content than have pulps with lower alpha-content. The physical characteristics of alpha-, beta-, and gamma-cellulose fractions have been studied with the electron microscope and x-ray camera, by Rhby.2°9 The investigations were made on commercial, softwood pulps prepared both by the sulfite and the sulfate processes. The electron-microscope investigation showed that alpha- and gamma-celluloses are two separate phases; the alpha consists of micellar strings, and the gamma is a dispersed phase without defined structural elements. The beta-cellulose could not be classified directly from the electron micrographs, but the x-ray diagrams were similar t o those of the alpha-cellulose, implying that the betacellulose from softwoods consists of disordered and broken micelle strings, containing only short-chain cellulose. This assumption receives support from the recognized fact that the beta fraction increases as a result of various different types of degradation of technical, wood pulps. (209) I3. G. Rbnby, Svensk Papperstidn

, 66,

115 (1952).

330

W. J. POLGLASE

These coiiclusions are in agreement with Mitchell’s studies210on the chain length of nitrated, cellulosic constituents of wood. In these studies,210 only two distinct fractioiis were obtained from nitrated, Western-hemlock wood, namely, hemicellulose nitrate which had a D. P. of about 70 and amounted to 30 % of the total, and alpha-cellulose nitrate with an average D. P. of about 2000 to 2500, representing the remaining 70%. Mitchell suggests that his resultsindicate that the beta portion of an industrial, wood cellulose is probably made in the pulping and bleaching operations, and that it consists of short-chain fragments resulting from cleavage of the long-chain, alpha-cellulose component. Mitchell speculates, further, “that wood cellulose may consist of only two portions-one that is water-insoluble, alpha, and one that is potentially water-soluble, gamma. The difference in solubility would be due not only to differences in chain length but, perhaps primarily, to differences in chain unit, end groups and side groups. Gammacellulose may be held in the fiber by cross links or secondary bonds only to become soluble when liberated-for example, by the swelling action of sodium hydroxide.” Simmons211has studied the carbohydrate composition of the alpha-, beta-, and gamma-fractions of bleached, coniferous, chemical wood-pulps. He concluded211that the gamma fraction consists largely of mannan and xylan. The alpha and beta fractions may contain appreciable proportions of mannan and xylan. However, the mannan and xylan in the beta fraction are occluded during precipitation of the beta-cellulose. These results211are in agreement with the ideas expressed above that alpha-cellulose and betacellulose are chemically similar, and differ only in D. P. 2 . Accessibility of Cellulose

An analytical method for the determination of accessibility of cellulose was developed by Nickerson212in 1941. Since that time, the procedure, or a modification 214 has been used extensively in the examination of cellulosic materials.216 The original method212was based on the observation that a boiling mixture of ferric chloride and hydrochloric acid causes relatively rapid evolution of carbon dioxide from D-glucose. When cellulose is heated with these reagents, D-glucose is formed by hydrolysis, and is then oxidized to yield (210) R. L. Mitchell, I n d . Eng. Chem., 38,843 (1946). (211) J. R. Simmons, B r i t . Paper and Board Makers’ Assoc., Proc. Tech. Sect., 33, 513 (1952). (212) R . F. Nickerson, I n d . Eng. Chem. A n a l . E d . , 13, 423 (1941). (213) C. C. Conrad and A. G . Scroggie, I n d . Eng. Chem., 37,592 (1945). (214) E. L . Love11 and 0 . Goldsrhmid, I n d . Eng. Chem., 38, 811 (1946). (215) R . F. Nickerson, Advances in Carbohydrate Chem., 6 , 103 (1950).

33 1

POLYSACCHARIDES ASSOCIATED W I T H WOOD CELLULOSE

carbon dioxide, which may be collected in absorption tubes. Since, under controlled conditions, the rate of evolution of carbon dioxide is proportional to the concentration of D-glucose, the course of the hydrolysis of cellulose may be followed by measuring the evolution of carbon dioxide. For similar determinations on wood pulps, Love11 and G ~ l d s c h m i d ,recognizing ~~~ that hemicelluloses in mood pulps might not give a satisfactory rate of evolut,ion of carbon dioxide, proposed that the sample be weighed after given times of hydrolysis. By either method, and for a large number of cellulose samples, it has been found that an initially rapid rate of hydrolysis is followed by a slower, relatively constant rate of hydrolysis. The iniTABLEXVIII A m o u n t of Easily Accessible Material, and Hydrolysis Rate Constant, k , for the Dificultly Accessible Fraction, f o r Diferent Pulps217 Ftber material

Easily accessible, 7/0

k

Cotton Hot-alkali refined linters Flax pulp Holocellulose Spruce sulfite, strong Spruce sulfite, rayon Spruce sulfite, hot-alkali refined Pine sulfate Pre-hydrolyzed sulfate Aspen sulfite Birch sulfate Viscose rayon

11.9 6.2 10.0 35.6 15.7 10.0 6.6 16.8 10.2 11.3 28.0 35.9

0.0062 0.0065 0.0066 0.014 0.012 0.013 0.013 0.012 0.011 0.012 0.017 0.039

tial, rapid reaction is presumed to denote hydrolysis of amorphous cellulose, whereas the later, slower reaction is believed to result from the hydrolysis of crystallites. During the first 30 minutes of the rapid reaction, thc ciiprammonium viscosity falls to a value which remains nearly constant for several hours,216 despite continuing hydrolysis. A possible explanatioiF is that disordered cellulose lying between crystallites is first attacked and that, thereafter, attack occurs mainly on the lateral surfaces of crystallites. If this is true, the length of the crystallites is indicated by the minimum viscosity attained. This crystallite length is usually expressed as a ch.ainlength “limiting 1). P.” 111 this andyticaal method, wood pulps in general differ from cotton linters in showing a greater acctessibility, as shown in Table XVIII (from the data (21G) R F Nickchrson ant1 .J. A Hatbile, I n d . Eny ( ’ / / e n / , 39, 1507 (1947). (217) W. €1 Algar, H W Giritz and A M. Custafsson, Ssensk Pappershdn., 64, 335 (1051).

332

W. J. POLGLASE

TABLE XIX Removal o j Carbohvdrates During Sul-fite Cookin Yield of bleached Pulp, %

Pulp -.

Wood Holocellulose Extra strong Greaseproof Medium Soft Rayon Acetylation pulp"

%

___

67.9 50.2 49.7 48.8 46.6 44.6 33.4

Cellulose, % '

Easily accessible

Beta-

Gamma-

77.3 86.3 86.6 87.2 88.9 90.1 95.0

traces tracef 1.4 1.6 2.9 2. 9

20.0 12.8 11.0 9.8 8.8 6.9 2.1

61.2

0. 4

34.2b

Alpha-

17

AlphaDificultl y ceiklose, accessible the wood) 96

IA

~

34.5 35.6 17.4 16.0 15.3 13.8 11.3 6.6

Holocellulose (alpha-, beta-, garnma.de. termination with hydrolytic pretreatment)

1

.o

43.4 43.4 41.5 41.6 41.4 40.3 40.1 31.1

52.5 43.3 43.0 42.6 41.4 40.2 31.7 42.6

Hot alkali refined, 10% NaOH, 120°C. b This value includes the amount of material dissolved during the short, hydrolytic pretreatment.

of Algar, Giertz and G u s t a f s s o ~ i ~These ~ ~ ) . workers217note a very good correlation between the gamma-cellulose fraction and the easily hydrolyzable fraction of wood-cellulose. As a result, the fiber is picturedZ17as consisting of a well-ordered, well-defined micelle-string structure embedded in amorphous hemicellulose. Giertz and coworkers217prepared a chlorite holocellulose, and a series of sulfite pulps representing a yield range of 68 t o 33%. On these cellulose preparations they performed accessibility estimations by the method of Nickerson,212as modified by Love11 and Gold~chmid,2~~ as well as conventional alpha-, beta-, and gamma-cellulose determinations. Their results are given in Table XIX. I n the range of sulfite pulps from the extra strong pulp to the rayon pulp, the variation in alpha-cellulose is relatively small, whereas both the gamma content and the accessibility decrease appreciably. The gamma content of holocellulose is low, compared to the percentage of accessible holocellulose. Giertz and associates217suggest that this is ascribable to the structural strength of the holocellulose fibers, which do notJswell sufficiently to allow outward diffusion of the interfibrillar substance. After a short,, vigorous hydrolysis of the holocellulose, the gamma-cellulose increased to a value consistent with the accessibility value, and the alpha-cellulose decreased proportionately. Since there exists a close connection hetween easily hydrolyzable material and gamma-cellulose, Giertz and c o ~ ~ o r l i ~point r s ~ ~ out 7 that therc must also be n relationship between difficultly hydrolyzable material and alpha- plus beta-cellulose. This means that the beta-cellulose is, from the hydrolysis viewpoint, of the

POLYSACCHARIDES ASSOCIATED WITH WOOD CELLULOSE

333

same character as the alpha-cellulose, and consequently is difficultly accessible. Just as Mitchell Giertz and associate^^'^ believe that the low beta-cellulose values of holocellulose and strong-sulfite pulp indicate that there is no beta-cellulose in wood, but that it is an artifact formed during the refining operation. Furthermore, it is that the betacellulose is formed, during pulping, from the difficultly accessible parts of the fiber, and is actually degraded alpha-cellulose. Thus, from nitrate studies, chemical analyses, electron-microscopic and x-ray diffraction work, and accessibility studies, come the same conclusions-namely, that wood cellulose consists of two phases: an ordered, cellulosic phase of high molecular weight, and an amorphous, accessible, hemicellulosic phase of low molecular weight. The cellulosic phase is alkaliinsoluble (alpha-cellulose) except for the short chains formed as a result of attack by chemical reagents during pulping (beta-cellulose). The hemicellulosic phase (gamma-cellulose) is water-soluble when once it has been liberated from the cellulose matrix by the swelling action of alkali. Accumulated evidence indicates that part of the non-cellulose polyoses are in the cellulosic phase (the “cdlulosans” of NormanI4) and part are in the hemicellulosic phase.

This Page Intentionally Left Blank

THE CHEMISTRY OF HEPARIN

BY A . B . FOSTER* A N D A . J . HIJGGARD f l r pmistry Departments, T h e Ilniaersity. Birmingham. lhgland. nnd The Ohio Sfate University. Columbus. Ohio

CONTENTS I . Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 336 I1. The Discovery of Heparin. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 336 111. The Isolation and Purification of Heparin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 337 1 . Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 337 2 . Methods of Isolation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 337 a . Early Procedures . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 337 b . Renzidine Salt Procedures . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 339 3 . The Homogeneity of Heparin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 344 4 . The Action of Enzymes, Alkali, and Acid on Heparin . . . . . . . . . . . . . . . . . 345 a . Enzymes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 345 b . Alkali . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 346 c . Acid . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 346 348 5 . Other Methods of Extractlion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . I V . The Structure of Heparin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 348 1 . Acidic Hydrolysis of Heparin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 348 a . Total, Acidic Hydrolysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 351 b . Hexosamine and Hexuronic Acid Content . . . . . . . . . . . . . . . . . . . . . . . . 351 c . Sulfur and Nitrogen Content.. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 352 ............................ 352 d . Partial, Acidic Hydrolysis . . . . 2 . Acetylative Desulfation Procedures . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 356 3 . Other Methods of Degradation of Heparin . . . . . . . . . . . . . . . . . . . . . . . . . . . . 357 a . Action of Nitrous Acid on Derivatives of 2-Amino-2-deo.uy-1~-glucose and Heparin., . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 357 b . Oxidation in the Presence of Ascorbic Acid . . . . . . . . . . . . . . . . . . . . . . . . 35'3 V . The Anticoagulant Activity of Hcparin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 359 I . The Biological Activity of Heparin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 359 2 . The Structure and Anticoagulant Activit.y of Heparin . . . . . . . . . . . . . . 360 a . Degree of Sulfation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 361 b . The N-Sulfate Group. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 363 c . Distribution of the Sulfate Groups . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 363 d . Molecular Weight of Heparin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 363

* Fellow of the Rockefeller Foundation at The Ohio State University. 1953.54 . The authors wish t o thank Professor M . L . Wolfrom for access t o much unpublished data on heparin during the preparation of this Chapter. and for valuable criticism and advice . 335

336

A.

B. FOSTER A N D A . J . HUGGARD

3. The R.Iolccula,r Shape of Heparin.. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . a . Sulfat,ion of Heparin.. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . b. Inactivation of Heparin by Dilut,e Acid. . . . . . . . . . . . . . . . . . . . . . . . . . . . c. Ot,her Considerations... . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . V I . The Biosynthcsis of Muco1)olgsaccharidcs. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

364 364 365 366 367

I . INTRODUCTION Heparin,* the blood anticoagulant present in circulatory tissue is now recognized to be an important and chemically unique polysaccharide of considerable biological significance. Since its discovery almost forty years ago, heparin has been intensively investigated by numerous workers. Much of its chemistry has consequently been evaluated and it is rather surprising that no comprehensive chemical review has recently appeared. In his excellent book, Jorpesl has described in some detail the history and early chemistry of heparin and the development of its medical applications. This Chapter is intended to present an account of the more recent developments in the chemistry of heparin together with the significant features of the earlier work.

11. THE DISCOVERY OF HEPARIN Heparin was discovered in 1916 by McLean2 who was working under the direction of Howell a t Johns Hopkins University. I n order to ascertain the origin of its blood-coagulating properties, crude cephalin was submitted to a careful fractionation. Fractions were obtained which unexpectedly inhibited the coagulation of oxalated, dog serum. This was a significant discovery since no anticoagulant had previously been found in mammalian tissue. Because of its abundance in liver, the anticoagulant material was named h e ~ a r i n Howell .~ was quick to recognize the potential therapeutic value of heparin, and in his Harvey Lecture; in 1917, he expressed the hope that the substance would find “suitable application . . . in the therapeutic treatment of disorders of coagulation.” Initially, heparin was thought to be a phosphatide because of its closely similar behavior to cephalin in the extraction procedure, but it is now recognized to be a highly sulfated mucopolysaccharide.

* The term heparin has not been used consistently since its introduction; it usually implies the sodium salt of the sulfated polysaccharide, but many other salts are known. Perhaps a more logical name would be heparinic acid; the sodium salt would then be termed sodium heparinate, and the naturally occurring complex which contains the anticoagulant, protein heparinate. In this Chapter, theterm heparinsignifies the sodium salt, unless otherwise qualified. (1) (J.) E. Jorpes, “Heparin,”Oxford University Press, London, 2nd Edition, 1946. (2) J. McLean, Am. J . Physiol., 41,250 (1916). (3) W. H. Howell and E. Holt, Am. J. Physiol., 47, 328 (1918). (4) W. H. Howell, Harvey Lectures, Ser. 12 (1916-17).

THE CHEMISTRY O F HEPARIN

337

111. THEISOLATION AND PURIFICATION OF HEPARIN 1. Introduction

I n the chemical literature prior to 1940, the majority of papers published on heparin were devoted to the isolation and purification of the polysaccharide. The enrlicst workers were concerned with the isolation of highly active, non-toxic preparations. Later, as attention was focused on the problem of heparin structure, many further attempts were made to isolate the pure substance, and extensive data on the activity and analytical composition of the various preparations have consequently accumulated. During the early period, the different research laboratories each claimed to have isolated pure heparin, but it is only within the last decade, with the application of physicochemical methods, that relatively homogeneous preparations have become available. 2 . Methods of Isolation a. Early Procedures.-Some details of the original procedures for the extraction of heparin will be given, since they form the basis for subsequent methods. Heparin does not exist in the free state in the body under normal conditions, but is invariably linked with a protein which in one case (in mast cells, see page 360) has been shown to be a l i p ~ p r o t e i nI. n~ certain abnormal conditions, free heparin may appear in the body. Thus, heparin is released from the liver into the blood stream in dogs in anaphylactic shock.6 In the isolation of heparin from normal tissue, a stepwise purification procedure is required. The earliest method for the isolation of heparin, developed by H ~ w e l l , ~ involved the treatment of dried and minced dog liver with boiling methanol, and subsequent extraction with physiological saline solution from which the active principle was precipitated by acetone. Glycogen was then removed by digestion with Takadiastase and the proteins were split off with cadmium chloride. As a result of this work, heparin was made available commercially. Subsequently, Howell8 improved the separation of heparin from protein by introducing the use of Lloyd's reagent (aluminum silicate) in acetic acid. The final stage then involved the precipitation of the heparin nith excess barium hydroxide. Howell claimeds that his heparin preparation con(5) 0. Snellman, B. S y l v h and Christina JulBn, Biochim. et Biophys. Acta, 7, 98 (1951). (6) L. B. Jaques and E. T. Waters, A m . J . Physiol., 129, 389 (1940); J. Physiol. (London), 99, 454 (1941). (7) W. H. Howell, Am. J. Physiol., 63, 434 (1922-23); 71, 553 (1924-25). (8) W. H. Howell, Bull. Johns Hopkins Hosp., 42, 199 (1928).

338

A. B . POSTKR A N D ,4. J. HUGGARD

tained 110 nitrogen (undoubtedly because of the insensitive test used), phosphorus, or sulfur, in spite of the fact that inorganic sulfate had been identified amongst the products after hydrolysis with hydrochloric acid. He thought that heparin was consequently not a protein or phosphatide, and he showed it to be thermostable, insensitive to bacteria, and resistant to the common enzymes. Howell suggested that heparin was carbohydrate in nature and contained uronic acid. I n 1933, Fischer and Schmitz9 first claimed to have isolated a pure heparin preparation. Their lengthy extraction and isolation procedure culminated in the isolation of a “microcrystalline” hruciiie salt 32 times as active as the starting material. KO evidence other than that of appearance was quoted in support of the claimed crystallinity of the product. It was concludedg that heparin was a carbohydrate (Molisch test) and contained a uronic acid. Fischer and Schmitzlowere also interested in the interaction of heparin with proteins, such as casein and serum albumin. They reported that heparin appeared to shift the isoelectric point of the proteins to the acid side, probably by the formation of a molecular complex, and that combination with the protein occurred only near the isoelectric point and on the acid side.” The complex was reversibly dissociated by the addition of alkali. These results are of irnportance in selecting conditions for the purification of crude heparin preparations, and constitute the basis for the extraction procedure developed by Charles and Scott,12which has subsequently been used extensively. Charles and Scott employed the cheaper and more readily available beef liver’? and beef as the source material. Beef liver (or lung) a a s minced and then autolyzed for twenty-four hours before extraction with an alkaline solution saturated with ammonium sulfate. Protein was precipitated by warming the extract, and the heparin-protein complex was precipitated from the supernatant liquor on acidification. Extraction of the complex with ethanol removed fatty material, and tryptic digestion removed most of the protein. The heparin u a s precipitated with ethanol, redissolved in h a r m alkaline solution to destroy trqpsin, and reprecipitated \titti acetone. This material, “crude heparin,’’ a a s isolated in a lield of 15-50 g. per 100 Ib. of animal tissue. In a later paper,14 the purification of crude heparin by fractionation successively n i t h Lloyd’s reagent, cadmium chloride, and acetone, mas described. The “purified heparin” was 100 times as active as the crude material. Scott and Charles“ reported the presence of nitrogen (9) A . Fischer and A Gchmitz, Hoppe-Sqilet’s Z . physzol. (’hem., 216, 264 (1933). (10) A. Fischer, ~vnlirrwissenfichnSten,19, 965 (1931); A . Fischer and A. Schmit7 tbid, 20, 471 (1932); B?orhein. Z , 269, 61 (1933) (11) See I,. R . Jnques, Biochew. . J . (London), 37, 189 (1043). (12) A F. Charles and D. A . Scott, J. Biol. (‘hem., 102, 425 (1933) (13) A . F. Charles and D. A. Scott, Trans. Roy. Sac. Can., B, 28, 55 (1935). (14) D. A. Scott and A. F. Charles, J. B i d . Chem., 102, 437 (1933).

THE CHEMISTRY OF HEPARIN

339

and inorganic ash, but failed t o obtain a positive naphthoresorcinol test for uronic acid.

A few years later, Jorpes confirmed much of the work done by Charles and Scott. Heparin was purified by their procedure and, after several successive treatments with Lloyd’s reagent, the product ivas found to coiitairi 1.64 % of nitrogen. The presence of sulfur (as both bound and free sulfate), inorganic ash, uronic acid, hexosamine, and acetic acid was reported. Electrodialysis of the purified heparin removed free sulfate, and afforded a highly active preparation which could be precipitated as an amorphous brucine salt.16 Lipmann and Fischer’G later confirmed the presence of nitrogen and sulfur in heparin isolated by the procedure of Charles and Scott.l2-l4 There was now fairly good agreement between the various laboratories regarding the qualitative composition of heparin. Jorpes and Berg~trorn’~ offered an explanation for the failure by Charles and ScottI4 to obtain a positive naphthoresorcinol test for uronic acid when they showed that highly active heparin is resistant to depolymerization by acid. It was well known, a t this time, that heparin occurs widely distributed in mammalian tissues. Its presence in dog liver,3 lymph glands; and blood ve ~se l s ,in ~ uterine mucous membrane,18 and in extracts of milk, kidney, lung, heart, arid other had already been demonstrated. Charles and Scottz0 determined the quantitative distribution in various tissues, organs, and fluids, as recorded in Table I . Heparin is now thought to originate in the mast cells. These cells are generally located in the connective tissue, in the vicinity of the capillaries, and in the walls of the blood vessels. Mast cells are especially plentiful in liver and lung tissue, which are consequently the best sources of heparin; and they give a purple stain with Toluidiiie Blue, owing to the interaction of the dye with heparin. A detailed account of the histological demonstration of heparin has been published elsewhere.’ b. Benzidine Salt Procedures.-In 1936, Charles and Scottz1described a method for the purification of heparin (through the benzidirie salt) which was a significant development in heparin chemistry. When “purified” heparin was treated with benzidine hydrochloride, a salt was obtained which was (15) (a) (J.) E. Jorpes, Nalurwissenschuften, 23, 196 (1935) ; (b) Biochern. J . (London), 29, 1817 (1935); (c) Acta Med. S c a d . , 88, 427 (1936). (16) F. Lipmann and A. Fischer, Hoppe-Seyler’s 2. physiol. Chem., 237,273 (1935). (17) (J.) I%. Jorpes and S. Bergstrom, Hoppe-Seyler’s 2. physiol. Chem., 244. 253 (1936). (18) J. I,. King, Am. J. Physiol., 67, 444 (1921). Igakkni Zasshi, 3, 61 (1926). (19) 0. Kashiwamura and R. Katsuki, K?~mamoto (20) A. F. Charles and D. A. Scott, J . B i d . Chein., 102.431 (1933). (21) A. F. Charles and D. A. Scott, Biochem. J. (London), 30, 1927 (1936).

340

A . R . FOSTER AND A . J. HUGGARD

readily decomposed with dilute ammonium hydroxide to afford the aniinoniuni salt of a highly active heparin which contained only 0.7 % of ash. The benzidine salt was readily convertible into metallic salts; and, when treated with barium acetate in acetic acid, a barium acid salt of heparin was ohtained whose photomicrograph showed it to be crystalline. Charles arid Toddz2re-examined this material and stated that “the crystals N ere tvv small and heavily twinned for an x-ray diagram”; the crystals displayed Most workers from 1936 onward regarded the only low birefringen~e.~~ barium acid salt as a good starting point for chemical investigations on heparin. The analysis of the barium acid salt, originally made by Charles and Distribution

TABLE I Heparin i n the Bodyz0 .

0.f .

Heparin,

n,g./kg. of’ tissue

Location

Reef liver spleen heart blood thymus serum lung muscle Hog liver Dog liver

“Crude”

“Parijied”

780 1000 200 260 640 24 840 2000 1400 900

190 230 54 66 310 230 600 340 330

Scott,21was performed on material that was not anhydrous; likewise, Meyer and S m ~ t analyzed h ~ ~ an incompletely dried sample obtained from Charles (see Table 11, column 1). Astrup and J e n ~ e nconverted ~~ the crystalline barium acid salt to the sodium salt of heparin, which they subjected to elemental analysis. Results in good agreement with those of Charles and Scottz1were claimed when allowance was made for the “hydration.” Reinert and WintersteinZ6also prepared heparin by the benzidine-salt procedure and obtained 25-50 mg. (22) A. F. Charles and A. R. Todd, Biochem. J . (London), 34, 112 (1940). (23) M. L. Wolfrom, D. I. Weisblat, J. V. Karabinos, W. H. McNeely and J. McLean, J . Am. Chem. Soc., 66, 2077 (1943). (24) K. Meyer, Cold Spring Harbor Symposia Quant. Biol.,6, 97 (1938). (25) T. Astrup and H. B. Jensen, J . Biol. Chem., 124, 309 (1938). (26) M. Reinert and A. Winterstein, Arch. intern. pharmacodynamie, 62,47 (1939).

34 1

T H E CHEMISTRY OF H E P A R I N

of heparin per kg. of liver. The highest sulfur content (sodium salt basis) recorded by these workers was 12 %. Charles and Toddzzreported that the barium acid salt and the ammonium salt were found to be readily interconvertible, which indicated that both were definite compounds. Analytical data (see Table 11,column 2 ) reported for incompletely dried material were in agreement with those of earlier workers when calculated on a “hydrated” basis. Masamune, Suzuki and Kondohz7 also reported analytical data on an incompletely dried sample (degree of hydration not stated) of the barium (acid) salt (see Table 11, column 3). No significant chemical differences were found in samples of

Authors

l+D

I

degrees

C

H

Ba

Conimenls

_ _ _ - _ ~ _ _ - ~ - Meyer and Smyth24 Charles and Todd21

+53 +5W

17.8 3.1 1.9 20.2 3.5 2 . 2

Masamune and coworkers25 Wolfromandcoworkers23

9.8 2.2

2.36

+36

2.1

+47.5 20.2 2 . 9 1.9

0.0

9.6 1.32.9 11.0

16.3

20.1 33.3 33.4

Incompletely dried sample 19.7 Moistnre content, 12.1% 22.5 Recalculated

to anhydrous basis Degree of hydra0.0 8 . 7 0 . 0 26.4 28.7 27.7 tion got stated 0.3*11.3 0 . 0 0 . 0 1 7 f 1 18 40.4’ 23.8 Anliydrous sample

heparin (barium acid salt) isolated from dog, beef, swine, and sheep tissues.z8 29 I n 1943, Wolfrom and coworker^?^ published the results of a thorough investigation of the elemental composition of the barium acid salt of heparin, and special attention was given to the preparation of anhydrous material. The results are quoted in Table 11, column 4. Summation of the analytical results apparently accounted for 88.6 % of the material present. However, there was by no means unanirrious agreement that the barium acid salt of heparin was the most pure and active material prepared. In particular] J o r p e ~ who , ~ ~ had been very critical of the evidence for the crystallinity of the barium acid salt, continued t o purify his heparin prepa(27) H. Masamune, ill. Susuki and P.Kondoh, J. Bioehem. (Japan), 31,343 (1940). (28) 1,. B. Jaques, Scienre, 92, 488 (1940). (29) I,. B. Jaqutw, 13. T. Waters and A . F. Charles, .I. Riol. C h ~ m . 144, , 229 (1942) (30) (J.)E.Jolprs, Riochsni. .I. (T,oridon), 36,203 (19-12); Hoppr-Scylcr’s Z . physiol. Chew/., 278, 7 (1943).

342

A . B. FOSTER AND A. J. HUGGARD

rations by fractionation of the brucine salts. It was m a i n t a i i ~ e dthat ~ ~ further purification could be effected by this procedure, but this claim was specifically denied by Charles and Todd.22Jorpes’ procedureL5involved electrodialysis of the sodium salt (purified through the benzidine salt) against distilled water. The acidic, cathodic solution was treated with brucine sulfate, and the resultant salt was fractionated by repeated freezing and thawing of its aqueous solution. Jorpes30 associated activity with degree of sulfation (see page 361), and he reported that his amorphous preparations had a sulfur content of 13.5 % (sodium salt basis). The fractionation procedures (brucine salt) were performed on heparins obtained from various tissues:O and it was postulated that the heparin material was not a definite compound but was a mixture of “mucoitin polysulfuric acid esters.” Fractions corresponding in their sulfur content t o mono-, di-, and tri-sulfates (per disaccharide period) were obtained, together with sulfur-free fractions32(see also Marbet and Winterstein33).The most active material had a sulfur content of 13.8%, corresponding t o three sulfate groups per disaccharide unit. In 1943, Kuizenga and Spaulding3* advanced more evidence that a barium acid salt of heparin that they had prepared was not homogeneous. A fractionation mas achieved using aqueous acetone, to give two products xvhich, after conversion to the sodium salts, showed different activities and sulfur contents. This result would appear to support the contention that heparin is a mixture of closely related substance^.^^ It should be emphasized that the barium acid salt described by Kuizenga and S p a ~ l d i n gwas ~~ obtained by a procedure different from that by which the barium acid salt described by Charles and Scott2‘ and by Wolfrom and coworkers23was obtained. The latter salt was reported to be electrophoretically homogene~us.~~ More recently, it has been reported by Wolfrom and coworkers36that a relatively homogeneous polymer fraction with a high degree of sulfation may he obtained from commercial heparin, after fractionation as the (31) S. Bcrgstrom, (J.)E. Jorpes a i d 0. Wilander, Skand. Arch. PhysioE., 76. 175 (1937); (J.) 13. Jorpes and S. Bergstr6m, Biochern. J. (London), 33, 47 (1939). (32) (J.) E. Jorpes and S . Gardell, J . Biol. Chem.,176,267 (1948). (33) R.Marbet and A . Winterstein, Helv. Chim. Acta, 34, 2311 (1951). (31) M. H. Kuizenga and L. B. Spaulding, J . B i d . Chem., 148, 641 (1943). (35) AT. I,. Wolfrom, R. K. Madison and M. J. Cron, J . Am. Chem. SOC.,74, 1491 (1952). This paper reportrtl a correction t o the earlier work of M. L. Wolfrom and F.A 11. Rice, J . A n , . Chein. Soc., 69,2918 (1947). What was originally suspected as a second component in heparin was shown to be an abnormally large boundary anomaly. (36) Af. I,. Wolfrom, R. Montgomery, ,J. V. I<ar:it)inos and P. Rathget), .I. A M . (‘hem. SOC., 7 2 , 5796 (1950).

THE CHEMISTRY OF HEPARIN

343

neutral barium salt with subsequent purification using cold acetic acid.3GJ Charles and Scott'2*l4 recognized that heparin could be precipitated as the barium salt, and fractions of low sulfate content were also obtained by its use.39The neutral barium salt is also prepared through the henzidiiie salt. Other derivatives of heparin have been reported to have a crystalline appearance; for example, the salts formed with piperidine, n-pentylamine, and i~opentylamine.~~ D o d e ~ y l a m i n eand ~ ~ de~amethylenediamine~~ have also been used in the preparation of heparin. It is clear from the preceding account that much of the confusion which arose in the earlier work on heparin was due primarily to insufficient purification of the preparations. In actual fact, no satisfactory criterion of purity had been established. Most workers strove to obtain preparations of high anticoagulant activity, and the situation was complicated by the fact that, during this period, several different methods for the assay of anticoagulant activity were in use. Jorpes and his associates had associated activity with sulfur content, and the latter was used as a criterion of purity. This is currently thought to be an over-simplification (see page 360). Bell and IG-antz40 studied the ultraviolet absorption of a series of heparin preparations, and found no correlation between absorption and activity. Some of the highly active preparations showed no absorption in the range 230-300 mp, whilst others showed a maximum at 265-292 m p and a minimum a t 240-260 mp. These results would appear to indicate the presence of impurities, since, on re-investigation of the ultraviolet absorption of heparin, no absorption in this region was This inference is supported by the results of a chromatographic study of heparin.42The movement of heparin on paper, under the influence of a 1-propanol-water solvent system, showed two components (RF0.00, 0.57). The original heparin preparation showed absorption a t 270 mp and, after chromatography, the more rapidly moving component, which had all of the anticoagulant activity, showed no absorption a t 270 mp. (364 M. L. Wolfrom and P. Rathgeb, unpublished results. (37) D. A. Scott, A. F. Charles and A. M. Fischer, Trans. Roy. SOC.Pan., V , 36, 49 (1942). (38) A. E. O'Keeffe and J. A. Shannon, U. S. P a t . 2,552,507 (1951); Chem. Abstracts, 46, 6806 (1951). (39) J. Lee and L. Berger, U. S. Pat. 2,561,384 (1951); Chem. Abstracts, 46, 10515 (1951). (40) F. K. Bell and J. C. Krantz, J . Am. Pharm. Assoc., S c i . Ed., 39, 94 (1950). (41) A. B. Foster and E. F. Martlew, unpublished results. (42) D. Molho and L u c k Molho-Lacroix, Compt. rend., 236, 523 (1952).

344

A.

R. FOSTER AND A. J. IIUGGARD

3. The Homogeneity of Heparin

In the last decade, physicochemical methods have been applied in assessing the homogeneity of heparin. The first recorded electrophoretic study of heparin was made by Wilander,4"awho demonstrated the presence of two components, one of which was thought to be protein. Chargaff, Ziff and Moore43studied Roche heparin44and concluded that it was homogeneous, but this result was not confirmed by later workers. Jensen, Snellman and S y l ~ 6 nreported ~~ that both Roche and V i t r ~ mheparin ~~ contained two fractions. The faster-moving component contained 75 % of the biological activity and had the higher sulfur content (9.3 %) ; it was apparently homogeneous. The slower-moving fraction (S, 7.3 %) probably contained more than one substance. These findings were confirmed by Meyer and S c h ~ v a r t zin~ a~ later study of Roche heparin. The metachromatic activity of heparin has also been used for demonIt has been known for some time that heparin strating its i~ihornogeneity.~~ (and other sulfated polysaccharides) gives a staining reaction with certain basic dyes such as Toluidine Blue,@and this reaction has been used extensively for investigating the distribution of heparin in body tissue.' When commercial heparin preparations were submitted to fractionation using alcohol, dioxane, or acetone, the biological activity and metachromasia of the fractions showed a marked discrepancy, indicating the precipitation of more than one substance, although no separation could be effected. The use of adsorption-front analysis49also indicated the inhomogeneity of commercial heparin. Various unspecified samples of the sodium salt of heparin have been submitted to Craig counter-current distribution between an aqueous buffer solution and n-pentyl Three fractions were obtained, two of which showed anticoagulant activity. (42a) 0. Wilander, Skand. Arch. Phusiol., Suppi!. 16, 89 (1939). (43) E. Chargaff, M. Ziff and D. H. Moore, J. B i d . Chem., 139, 383 (1941). (44) Commercial samples of heparin referred t o in this Chapter are Roche (United States) and Vitrum (Sweden). (45) R. Jensen, 0. Snellman and B. SylvBn, J. Biol. Chem., 174, 265 (1948). (46) K. H. Meyer and D. E. Schwarta, Helv. Chim. Acta, 33, 1651 (1950). (47) L. B. Jaques, Margaret B. Mitford and Ann G. Ricker, Rev. can. biol., 6, 740 (1947). (48) A spectrophotometric study of the metachromatic reaction of Toluidine Blue with sulfated polysaccharides has recently been made; J . Ball and D. S. Jackson, Stain Technol., 28, 33 (1953). (49) 0. Snellman, R. Jensen and B. Sylven, Acta. Chem. Scand., 3, 589 (1949). (50) A. E. O'Keeffe, F. M. Russo-Alesi, M. A. Dolliver and C. J. Stiller, J . Am. Chem. Soc., 71, 1517 (1949).

THE CHEMISTRY OF HEPARIN

345

The behavior of heparin in filter-paper electrophoresis has also been studied.61 More recently,36Wolfrom and coworkers have obtained a heparin preparation (by fractionation of the neutral barium salt) which appeared to be homogeneous in both electrophoresis and counter-current distribution studies. Consequently, it would seem that this material is the best available for structural studies. There can be little doubt that the isolation of a homogeneous heparin preparation is difficult. In fact, Jorpes considered that heparin is not a definite compound but a mixture of inucoitin polysulfates. He suggested32 that the isolation of a range of heparins with increasing sulfur content could be explained in two mays; firstly, it is possible that heparin is elaborated and broken down in the body in such a way that heparins with different levels of activity exist side by side; secondly, the purest form of heparin may undergo chemical modification during the isolation procedure, t o yield less active materials. Jorpes favored the former alternative. 4. The Action of Enzymes, Alkali, and Acid on Heparin

At this point, it may be worthwhile to consider the various stages in the extraction and isolation of heparin in terms of present knowledge of the stability and activity of the molecule. During the process of isolation, heparin is treated with enzymes, alkali, and acid; these stages will be considered in turn. a. Enzymes.-Very little appears to be known about the enzymes, in animal tissue, which are capable of modifying heparin. Jacques and cow o r k e r ~53 ~have ~ ~ reported the inactivation of heparin by an enzyme “heparinase” isolated from rabbit liver. The optimum conditions for the autolysis of minced liver in the initial stages of heparin isolation have been is added to freshly investigated by Kuizenga and S p a ~ l d i n g If . ~ water ~ minced liver and incubation is allowed to proceed a t 35” for 30 minutes and a t room temperature for a day, heparin of twice the purity and in twice the yield (over earlier procedures) is isolated. Jorpes30reported that the degree of autolysis had little effect on the activity and sulfur content of the heparin subsequently extracted, but no experimental data were cited. From the available evidence it would appear that the autolysis procedure increases in efficiency when made slightly more drastic, and hence it is not probable that degradation of the heparin occurs. Neither is there any evidence that tryptic digestion of crude heparin causes any modification of the molecule. (51) K. G . Rienits, Biochern. J . (London), 63,79 (1953). (52) I,. B. Jnques, J . Biol. Cheni., 133, 445 (1940). (53) L. B. Jaques and Eve Kuri-Szantie, Can. J. M e d . Sci., 30, 353 (1952).

346

A . B. FOSTER AND A . J. HUGGARV

Jorpes30showed that a highly active sodium salt of heparin was not inactivated under the conditions of tryptic digestion. b. Alkali.-Much more is known of the stability of heparin toward alkali (and acid). Charles arid Scott12investigated the inactivation of impure heparin, and found that treatment with 0.25 N alkali a t 80" caused loss of half the activity in two hours. J ~ r p e reported s~~ that a highly active sodium salt of heparin was not inactivated in 10 N alkali and 50% amit was reported that heparin monium sulfate a t 80" for one hour. dissolved in N sodium hydroxide a t 100" lost 75% of its activity in five minutes. During Charles and Scott's alkaline extraction procedurez0 of minced tissues, the crude material was maintained in 0.5 N sodium hydroxide, saturated with ammonium sulfate, a t 50" for 30-60 minutes. It would seem highly probable that some modification of the heparin molecule occurs a t this stage. Certain recent results are of interest in this respect. It has been pointed outs5that the results of Gilbert and may have some bearing on the isolation of heparin. These workers demonstrated that starch isolated from potatoes by alkaline treatment under anaerobic conditions was much more highly polymerized than starch similarly isolated under normal conditions. Further, it was shown that highly polymeric starch was but little degraded by the action of 0.2 N sodium hydroxide at 100' in the absence of oxygen, but was rapidly depolymerized when oxygen was introduced. Cellulose is also degraded by alkali and oxygen.6saRelated results were reported by Blix and Snellmans7 who found that hyaluronic acid and chondroitinsulfuric acid, chemical relatives of heparin, were more highly polymerized when isolated under anaerobic conditions in the absence of alkali. The possibility that the action of alkali on heparin, during the isolation procedure, may lead to fragmentation of the molecule, must be borne in mind when considering the data on the homogeneity of heparin. It would be of interest to isolate heparin under anaerobic conditions. c. Acid.-The acid-hydrolytic characteristics of heparin have received close attention, especially in recent years. Schmitz and Fischer6*noted that heparin lost 70% of its activity in one minute when treated with 0.1 N hydrochloric acid at 70", arid Charles and Scott14reported that, a t pH 2.5, crude heparin lost half its potency in two hours a t 80". Recent studies36 (54) (J.) E. Jorpes, R. Bostrcm and V. Mutt, J . B i d . Cheni., 183,607 (1950). (55) .4. B. Foster, E. F. kfartlen arid M. Stacey, Chemistry & Zndiistry, 899 (1953). (56) R. J Bottle, C, A Gitlwrt, C. T. Greenwood arid N. K. Saad, Chemistry d Zndiistry, 541 (1953). (5th) E. Heuser, "Ctieniistry of Cellulos~,"John Wiley arid Sons, Inc., New York, N. Y., 1941. (57) G . Blix arid 0. Snellman, Nature, 163, 587 (1944). (58) A. Schinitz and 8 . Fischer, Hoppe-Seyler's 2. physiol. Cherra., 216, 264 (1933).

THE CHEMISTRY O F HEPARIN

347

have shown that heparin is inactivated rapidly in warm, dilute, acetic acid (see p. 365). During the extraction and purification procedures, heparin is treated in acidic conditions a number of times: (1) heparin is precipitated along with protein from an alkaline extract of liver by acidification, ( 2 ) the precipitate is washed n ith warm acid, (3) coagulation of inert material is effected by warming the solution of heparin a t p1-I 4.1 in the presence of ammonium sulfate to SO”, and (4) removal of protein-like impurities is carried out with precipitating and absorbing agents in acidic solution. Charles and Scott’2. 21 assayed the biological activity of their heparin samples a t different stages in the extraction and isolation procedure, and thus calculated the “total activity” at each point. However, it should be emphasized that, especially in the early stages of separation from protein, assay of “total activity” does not give any indication of chemical modification of the heparin molecule. Astrup and J e n ~ e nexamined ~~ the total activity of heparin samples at various stages during the isolation, and found that a t certain steps, such as deproteinization, a n increase in total activity occurred. These workers noted that binding with protein may actually mask the total potential activity of a crude heparin sample; this may be especially true of in vitro assays. Treatment of the crude heparin-protein complex may increase the total activity by liberation of protein, and decrease the potency by modification of the heparin molecule itself. Jacpes52 believes that if the affinity of heparin for proteins (not concerned with the coagulation system) is sufficiently great, linkage with such proteins might inhibit heparin activity in uiuo. To assay, accurately, the effect of a certain chemical procedure on crude heparin, the conditions must be simulated on a purified sample. That some inactivation of heparin will occur when it is treated with warm acid solutions is to be expected. However, many of the stages outlined above involve acidic treatment, either a t room temperature or when the heparin is precipitated from solution and a two-phase system exists (so that inactivation may not be extensive). When heparin is purified as the barium acid salt by repeated recrystallization from warm, dilute acetic acid, some inactivation is to be expected. Inactivation a t this stage has been cited as the cause of so-called species specificity of heparin. Jaques and Waters,Z9 in 1940, claimed that the use of Charles and Scott’s procedure12r21 for the isolation of heparin (as the barium acid salt) from tissues of different animals gave preparations of quite different activity. Thus heparin preparations from dog, beef, swine, and sheep lungs had relative potencies of 10, 5, 2, and 1. Wolfrom and coworkers60noted that Jaques had used slightly different conditions to crystallize the barium acid salt of (59) T. Astrup and H. B. Jensen, Skand. Arch. Physiol., 79, 290 (1938). (60) M. I,. Wolfrom, J. V . Karabinos, C. S. Smith, P. H. Ohliger, J . JPC m i l 0. Keller, J . A m . Chem. Soc., 67, 1624 (1945).

348

A . B. FOSTER AND A. J. HUGGARD

heparin from each species, alld suggested that this might, he respoiisible for the different levels of activity. When the procedure was standardized,6" heparin from dog and beef liver showed the same activity. Later, it, was confirmedG1that the barium acid salt of heparin isolated from pig liver and beef liver had the same potency. The isolation of heparin as thc rieutral barium salt36,36u. itivolves no warm-acid treatment. 5 . Other Methods of Extraction

It appears that chemical modification of heparin is possible during the most commonly used extraction and isolation procedures. In recent years, milder techniques have been applied to the isolation of heparin. Snellman, Jensen and SylvBnG2 circutnvented the use of alkali extraction by employing solutions of potassium thiocyanate; these rapidly and efficiently extract the mast-cell, granular substance from liver and skin. Subsequent removal of the thiocyanate by dialysis led to the precipitation of a heparin-protein cotnplex which was carried forward to the tryptic digestion stage as before. It is of interest that the calcium chloride extraction procedure, introduced and used extensively by Meyer and S n i ~ t in h ~the ~ extraction of tissue mucopolysaccharides, did not extract the heparin present in the granular substance of tissue mast cells. This is rather surprising in view of the fact that Howell' effected extraction by the use of sodium chloride solutions. For the separation of heparin from heparin-protein complexes, Homan and Lens64have developed a method which avoids the use of acid media. The heparin-protein complex is dissolved in aqueous solution a t pH 7.5 and is extracted with phenol (which removes most of the protein). The method also facilitates the removal of colored impurities which are normally difficult to eliminate. Extraction of the heparin-protein-octylamine complex with phenol has been studied.'jS I n conclusion, satisfactory methods are now available for the extraction and purification of heparin; these have been developed in the light of modern knowledge of heparin activity. The homogeneity of heparin preparations may also be ascertained before structural studies are begun. IV. THE STRUCTURE OF HEPARIN 1. Acidic Hydrolysis of Heparin Considerable information concerning the structure of heparin has been obtained from hydrolytic studies. Controlled, acid hydrolysis is a valuable W. C . Risser, J. Am. Chem. S o c . , 68, 341 (1946). 0. Snellman, R. Jensen and B . SylvBn, Nature, 161, 639 (1948). K. Meyer and Elizabeth M. Smyth, J. Biol. Chem., 119,507 (1937). J. D. H. Homan and J. Lens, Biochim. et Biophys. Acta, 2,333 (1948); J. Lens and J. D . H . Homan, Dutch Pat. 62,276 (1949); Chem. Abstracts, 43, 4817 (1949). (65) F. C. Monkhouse and L. B. Jaques, J . Lab. Clin. Med., 36.782 (1950). (61) (62) (63) (64)

THE CHEMISTRY OF HEPARIN

349

technique currently in extensive use in the study of polysaccharide structure. Complete hydrolysis of a polysaccharide to the nionosaccharide stage, and analysis of the hydrolyzate, serves to identify the sugar components present in the polymer. Graded, acid hydrolysis yields fragments, the identification of which may throw light on the structure of the parent polysaccharide with respect to (1) the sequence of sugar units aloiig the polysaccharide chain, ( 2 ) branching of chains, and (3) position and configuration of the glycosidic linkages. Thus, partial, acid hydrolysis of amylopectintic yields maltose and isonialtose, thereby confirming the presence of a-w(l+ 4) and a-r)-(l+ 6) linkages previously inferred from methylation studies. A general account, a t this point, of the acidic hydrolysis of heparin will provide a perspective for later, detailed discussion. The application of controlled, acid hydrolysis to heparin has been only partially successful owing to the unique, chemical structure of the mucopolysaccharide. If heparin is represented by the partial formula (I) (see page 357 for the detailed structure), on acidic hydrolysis the first stage will involve rapid cleavage of the labile, sulfamic acid groupsti7 [together with slower hydrolysis of the 0-sulfate groups, not shown in I] to yield “$-heparin” (11). Subsequent hydrolysis of I1 will be controlled to a large exteiit by the positive charges of free amino groups in acid solution which will provide an effective electrostatic shield around the adjacent hexosamiiiidic linkages b (and also, t o a much smaller extent, around the linkages a). Hydrions, the effective hydrolyzing agents, will be repelled from the molecular locality of the hexosaminidic linkages, and hydrolytic scission will tend t o occur predominantly a t the D-ghcosiduronic linkages a. The establishment of -NH,+ groups along the heparin molecules in the early stages of acidic hydrolysis accounts for the greater general stability of heparin over that of other mucopolysaccharides, such as hyaluronic acid and chondroitinsulfuric acid, which have acetylated amino groups.32 This type of electrostatic shielding has been studied in simple derivatives of 2-amino-2-deoxy-~-g~ucose(D-glucosamine) .68 Thus, methyl 2-amino-2-deoxy-a-~-glucopyranoside is much more resistant to acidic hydrolysis a t the glycosidic center than is the N acetyl derivative, which is hydrolyzed a t approximately the same rate as methyl a-D-glucopyranoside. A similar effect has been observed with the 1-phosphates of 2-amino-2-deoxy-~-glucose~~ and 2-amino-2-deoxy(66) M. L. Wolfrom, J . T. Tyree, T. T. Galkowski and A. N. O’Neill, J . A m . Chem. SOC.,72, 1427 (1950) ; 73. 4927 (1951); see A. Thompson and M. L. Wolfrom, ibid., 73, 5849 (1951). (67) A. B. Foster, E. F. Martlew and M. Stacey, Abstracts Papers Am. Chem. SOC.,126, 6D (1954). (68) (a) R. C. G. Moggridge and A. Neuberger, J . Chem. Soc., 745 (1938); (b) A. B. Foster and M. Stacey, Advances in Carbohydrate Chem., 7, 271 (1952). (69) D. H. Brown, J . B i d . Chem., 204, 877 (1953).

350

A. B . FOSTER AND A. J. HUGGARD

COOH

CHZOH

CHiOH

CHiOH

COOH

CHZOH

COOH

-0 NHSOIOH

NHSOIOH

i

warm dilute acid

a

a

CHiOH

n

U

COOH

I

TO I I

NH? ;

NH? I

b

b

NH?

I

I

L

I1 $-Heparin

+

other oligosaccharide fragments

I11

/

CHjOH

0 +

C01 and other uronic acid

decomposition products

NH?

IV

D-galactose (D-galactosamine, chondrosamine) which are more stable toward acidic hydrolysis than are the corresponding derivatives of D-glucose and D-galactose. Acidic hydrolysis of +-heparin (11) will result in predominant fragmentation of the chain a t the D-glucosiduronic linkages a to give a series of oligosaccharides, the smallest of which will be the disaccharide 111. Further hydrolysis leads to the destruction of the D-glucuronic acid moiety, leaving 2-amino-2-deoxy-~-glucose(IV) as the main final product of hydrolysis. (70) C . E. Cardini and Id.F. Leloir, A r c h . Biochem. and R i o p h y s . , 46, 55 (1953).

THE CHEMISTRY OF HEPARIN

35 I

a. Total, Acidic Hydrolysis.-Complete, acid hydrolysis was the first hydrolytic procedure applied to heparin, and 2-amino-2-deoxy-a-~-glucose hydrochloride was subsequently identified.'7 Isolation and identification of the D-glucuronic acid moiety of heparin presented some difficulty. Qualitative indications of the presence of uronic acid were early obtained8 and were followed by quantitative studie~.~5,*8 Wolfrom and Karabinos,?l in a study of the hydrolytic characteristics of heparin showed that the rates of liberation of 2-amino-2-deoxy-~-glucose and destruction of u-glucuronic acid were roughly parallel. It was suggested by these authors that decarboxylation of the uronic acid might occur whilst it was glycosidically bound in the hydrolytic fragments of the polysaccharide. The isolation of D-glucuronic acid following direct acid hydrolysis of heparin would appear to be precluded. The evidence for the presence of D-glucuronic acid was put on a definitive crystalline basis by Wolfrom and Degradation of heparin using a bromine-sulfuric acid mixture resulted in oxidation of the glycosidic carbon atom of the D-glucuronic acid moiety following hydrolysis of the D-glucosiduronic linkages. The D-glucaric (saccharic) acid so formed was stable under the hydrolytic conditions employed. It was isolated as the crystalline potassium acid salt which had a specific optical rotation identical with that of authentic material. Potassium acid D-glucarate also has a characteristic x-ray diffraction pattern.73 2-Amino-2-deoxy-~-gluconic acid was also isolated following oxidative hydrolysis. At this point, it is pertinent to review the information that may be gained from analytical data concerning the structure of heparin. Since 1935, many analytical data on heparin have been accumulated, but, as noted earlier, much of the work was performed on inhomogeneous or incompletely dried samples. The most thorough quantitative examination of the composition of heparin was made by Wolfrom and coworkers23on the barium acid salt. 0. Hexosamine and Hcxuronic Acid Content.-,Jorpe~~~ has estimated the presence in heparin of 17-19 % of hexuronic acid and the amount of hexosamine t o be that calculated for one molecule of hexosamine per molecule of hexuroriic acid. The hexuronic acid was estimated as carbon dioxide, foland the lowing acid hydrolysis by a modified Tolleiis-LefBvre hexosamine by the Elson-Morgan colorimetric method.75 (71) M. I,. Wolfrom and J. V. Karabinos, J . A m . Chetn. SOC.,67, 679 (1945). (72) M . L. Wolfrom and F. A . H. Rice, J . Am. C h e m . Soc., 68, 532 (1946). (73) M. I,. Wolfrom and W . B. Neeleg, J . A m . f'herr/. Soc., 76, 2778 (1953). (74) A D. Dickson, H. Otterson and K. P. Link, J. A m . C h e m . SOC., 62,775 (1930). (75) L. A. Elson and W. T. J. Morgan, Biochem. J . (London), 27, 1824 (1933); see Ref. 68(b), p. 257.

352

A. B. FOSTER AND A. J . HUGGARI)

MeyerZ4and Wolfrom and their detected the uronic acid in the barium acid salt of heparin both qualitatively and quantitatively, and considered it to be the source of the acidity in the acid salt. Amino sugar was also estimated. These results, which cannot be said to be as precise as the elemental data, indicate a ratio of 1: 1 for amino sugar and hexuronic acid. c. Sulfur and Nitrogen Contmt.-Jorpeslb recognized that the high ash content of heparin was due to the presence of ester sulfate, and in his early 3 0 ' 31 he reported that heparin contained 2.5 sulfate groups per disaccharide period. Later, by fractionation with brucine, fractions n ere obtained which, according to sulfur analysis, had 3 sulfate groups per disaccharide unit, and in one case30 an even higher sulfate content. The homogeneity of these samples has not been assessed. Electrometric titration of free heparin obtained by electrodialysis showed both weak acid (uronic acid) and strong acid (ester sulfate) There appeared to be three sulfate groups per uronic acid. Wolfrom and coworkers23have summated their analytical data on the barium acid salt of heparin, and have expressed the data in terms of a tetrasaccharide unit which comprises two molecular proportions each of 2-amino-2-deoxy-~-glucose residue and D-glucuronic acid residue, and 5 (rather than 6) ester sulfate groups, in an K :S ratio of 2 :5. The barium :sulfur ratio was 1:2, which indicated that the sulfur is essentially present as ester sulfate, and hence the acidity must be due t o the carboxyl group, in accordance with the shape of the titration curve.42a d. Partial, Acidic Hydrolysis.-As indicated previously, the first step in the hydrolysis of heparin is the cleavage of the N-sulfate groups to give +heparin (11).Although this step is now well established, its elucidation was a problem of considerable difficulty. Confusion was created by the fact that heparin, although containing an amino sugar, does not contain any N-acetyl groups. It had been suggested76that all mucopolysaccharides have the amino sugar N-acetylated, and, in this respect, heparin is unique within the class of mucopolysaccharides. (Mucopolysaccharides have been classified by M e y d 7 and S t a ~ e y . ~It * )had been known for some time that the nitrogen in heparin is not present as the free amino group,21-y 2 , 587 79 since only small amounts of nitrogen are formed from heparin on treatment with nitrous acid (Van Slyke). Charles and Scott" noted that the action of nitrous acid completely inactivates heparin, and Charles and Todd2*observed that the amino-nitrogen value increases slowly in the Van Slyke esti(76) (77) (78) (79)

0. Furth, H. Hwmnnn and R. Scholl, Biochem. Z . , 271, 395 (1934). K. Meyer, Advances in Protein C ' h e ? ~ .2, , 249 (1945). M. Stacey, Advances zn Carbohydrate C h e m . , 2, 161 (1946) (J.) E. Jorpes and S. Bergstrom, J . B i d . Cherri., 118, 447 (1937).

THE CHEMISTRY OF HEP.4RIN

353

illation uiitil, aftcr 1ti hours, all the iiitrogeii is accouiited for. 2-AcetJaniido2-deosy-D-glucose (N-ncetyl-D-glucosatnine) gives no nitrogen under these condit'ions. All the nitrogen was present, in the amino sugar,?:!and evidcnre tJhatJthe amino group is substituted had been advanced by Wilander,"S;L who could detect no sign of buffering between pH 7 and 10 in the electrometric titration of free heparin. Heparin was assayed for N-acetyl by various workers, a.nd a range of N-acetyl contents was reported, t,he value being invariably lcss than t,he theoretical amount for acetylation of all the amino groups.15 2z Complet,e absence of N-acet,yl groups was claimed by other workers.23'z 5 , 27 In 1950, Meyer and Schwartz46 demonstrated that commercial heparin which had a low acetyl content was split, on electrophoresis, into two components. The more rapidly moving component contained all of the anticoagulant activity but had zero acetyl content. The slower moving component had an acetyl content corresponding to that, of a polymer of 2-acetamido-2-deoxy-~-glucose. It has been suggestedz3 8o that the small quantities of acetic acid in heparin may originate in the purification procedure when acetic acid is used. Heparin appears to have a definite capacity for the absorptive retention of acetic acid. A l t e r n a t i ~ e l y , ~ ~ t,he acetyl content may arise from an impurity which accompanies heparin in the isolation procedure and which is difficult to remove. 56 it has been shown by interaction of heparin with nitrous acid and l-fluor0-2,4-dinitrobenzene that 6 to 10% of tthe amino groups are present as -NH2 . Various suggestions as to the mode of linkage of the remaining amino groups have been made by several workers. It was shownz3 t,hat the residue which blocks the amino group is easily cleaved by acid, and I

hence it could not be a

\

N-CH3

grouping. The amino group is not in-

/

volved in a glycosidic linkage since, on release of the amino groups under mild, acid hydrolytic conditions, there is not a corresponding increase in the reducing power.54.81 The presence of a linkage involving the carboxyl group of the uronic acid and the amino group was also pre~luded,4~ since the potentiometric tit,ration of heparin indicated the presence of one carboxyl group per nitrogen atom and showed that t,he only acidic groups in the polysaccharide are -C02H and -S020H. Masamune, Suzuki and Kondohz7 were the first to suggest a sulfuric group in heparin in the form of bridges, -C-0-S02-NH-C-, but they presented no experimental evidence therefor. This suggestion was criticized by Jorpes, Bostrom and who pointed out that W i l a ~ i d e r ~ ~ * had shown that all the sulfate groups in heparin (acid form) are free and (SO) M. L . Wolfrom, Ohio State Univ. Eng. Espt. Sta. News, 26, No. 5, 22 (1953). 67, 748 (1945). (81) M. L. Wolfrom and W. H. McNeely, 6.Am. Chem. SOC.,

354

A . B. FOSTER AND A. J. HTJGGARD

titratable. The presence of a sulfamic acid groupiiig, -NI-I-S02011, was again suggested by Wolfrom and McNeely8*on the basis of the results of inactivation studies (see p. 365). The properties of heparin with respect to the presence of a sulfamic acid group accord with the known general chemistry of substituted sulfamic acids.82The stability of substituted sulfamic acids is related to the basicity of the parent aminesSs3Thus, an amine of low basicity gives a substituted sulfamic acid (arylsulfamic acid) which is very unstable in acid solution. Alkylsulfamic acids are comparatively stable in cold aqueous solution. It would appear that the acidic hydrolysis of alkylsulfamates has a high, temperature coefficient. Cyclohexylsulfamic acid, which is describeds4 as a “fairly strong acid,” is hydrolyzed in hot water. Since the basicity of 2-amino-2-deoxy-~-glucoseis intermediate between that of methylamine (and cyclohexylamine) and aniline,s5the properties of “N-sulfate” might be expected to be intermediate between those of the two classes of substituted sulfamic acids. Recent on the hydrolysis of the “N-sulfate” group in heparin showed that, when a 1% solution of the sodium salt in 0.04 N hydrochloric acid is heated a t loo”, hydrolysis of the “N-sulfate” is complete in 90 minutes. Meyer and in a study of model compounds structurally related to heparin, showed that the hydrolytic release of sulfate under acid conditions from 2-amino-2-deoxy-~-ghcoseN-sulfate is more rapid than from D-glucose 6-0-sulfate. These workers also obtained a biologically inactive, nitroso derivative of heparin, and offered this as evidence for a sulfamic acid linkage in the molecule. Wolfrom, Shen and Summerss6prepared dibarium methyl 2-am~no-2-deoxy-N-sulfo-tr~-0-sulfo-~-~-g~ucopyranos~de salt, and found that, on heating a 3 x loT4M solution in 0.004 N hydrochloric acid, the “N-sulfate” is lost in 20 minutes and the 0-sulfate after 12 hours. Some difficulty was experienced in establishing a strict correlation between liberation of amino group and sulfate release. For example, Wolfrom and McNeely,81and Jorpes, Bostrom and did not obtain correspondence between these when heparin was treated with hot, dilute acid. It was suggestedb4that retention of the barium sulfate in the colloidal state (peptization) by the heparin might be responsible for inaccurate results in sulfate analysis. If, however, heparin was first treated with hot, dilute alkali, then, on subsequent acid treatment, correspondence between release

<

(82) L. F. Audrieth, M. Sveda, H. H. Sisler and M. Josetta Butler, Chem. Revs., 26, 49 (1940). 69, 2113 (1947). (83) C . D. Hurd and N. Kharasch, J . Am. Chem. SOC., (84) L. F. Audrieth and M. Sveda, J . Org. Chem., 9,89 (1944). 60, 168 (1928). (85) J. B. Allison and R . M. Hixon, J . A m . Chem. SOC., (86) M. I,. Wolfrom, (Miss) T. M. Shen and C. G. Summers, J . A m . Chem. SOC., 76, 1519 (1953).

T H E CHEMISTRY O F HE P A R I N

355

of sulfate ions and of amino groups was obtained.64 Recently, Foster, Martlew and S t a ~ e have y ~ ~shown that there is a strict correlation between amino-group and sulfate-ion release on direct treatment of heparin with dilute acid. As the hydrolysis proceeded, aliquots were treated with l-fluoro2,4-dinitroben~ene,~~ which reacts only with the free amino groups to give a N-(2,4-dinitrophenyl) derivative. The substituted heparin could be isolated, after dialysis, by freeze-drying. The N-(2,$-dinitrophenyl) group has a characteristic, peak absorption a t Xssao, and the value of ernax.a t this wavelength is a measure of the N-(2,4-dinitrophenyl) groupsB8introduced and, hence, of the amino groups liberated. Elemental analysis on the N-(2,4dinitropheny1)heparins gave the loss in sulfur. A plot of S-loss against NHz-liberation showed a closely similar rate and, after correction for the low rate of sulfate hydrolysis,46a 1: 1 correspondence on complete hydrolysis of the “N-sulfate.” On the grounds that carbohydrate 0-sulfates are known to be relatively stable toward acid hydrolytic and from the preceding it would appear that the suggestion t’hatsulfate release from heparin under acidic hydrolytic conditions proceeds by a sulfate migration36, is unlikely to be correct. Further work would appear to be necessary in order to establish that an unexpected “neighboring-group effect” is not operating during the acid hydrolytic cleavage of the “N-sulfate” group in heparin. Berglundgoshowed that sulfamic acid derivatives are relatively stable under alkaline conditions which cause hydrolysis of N-acetamido groups. Jorpes, Bostrom and M a n n s ~ n utilizing ,~~ these data, found that less than 10% of the total amino groups is liberated when a 0.2% solution of heparin in N sodium hydroxide is heated a t 100”for 2 hours, whereas 55 % of that in chondroitinsulfate is released. Little work has been published concerning the isolation of oligosaccharides following graded, acidic hydrolysis of heparin. Wolfrom and cow ~ r k . e r have s ~ ~ studied the partial, acid hydrolysis of a carefully purified heparin. (Purification was effected through the benzidine salt and the neutral barium salt was fractionally precipitated from water. The lesssoluble fraction, after conversion to the bariuni acid salt, had a X:S ratio of 2:5 and 5 sulfate groups per tetrasaccharide period, consumed 1 mole of periodate per tetrasaccharide period, and was shown to be homogeneous by e1ect)rophoresis and by countercurrent distribution. The molecular 8

(87) F. Sanger, Biochon. J . (London), 39,507 (1945); 44, 126 (1949); 46, 563 (1949). (88) E. I?. Annison, A . T. James and W. T. J . Morgan, Biocheni. .I. (London), 48, 477 (1951). (89) E. G. V. Percival, Quart. Revs. (London), 3, 369 (1949); E. G . V. Percival and T . 13. S:out,ar,.I. 1‘hevr. S o c . , 1475 (1940). (90) E. Berglund, J. Chern. Soc., 34, 643 (1878). (91) (J.) E. Jorpes, H. Bostrom and B. Mannson, Acta (‘hem. Scand., 6, 797 (1052).

356

A. B. FOSTER AND A. J. HUGGARD

weight was ca. 20,000.) From the hydrolyzate was isolated, in 20% yield, a reducing disaccharide, heparosinsulfuric acid. Periodate oxidation, under conditions which minimized formate hydrolysis, showed an uptake of 3 moles of oxidant per mole, with the formation of 1 mole of formic acid but no formaldehyde. Both the hexosamine and the uronic acid moieties were destroyed in the oxidation. The N-acettyl derivative of heparosinsulfuric acid took up 2 moles of oxidant per mole, and gave 1 mole of formic acid but no formaldehyde. Together with analytical data and the demonstration that the uronic acid moiety constitutes the reducing end of the disaccharide, the structure of heparosinsulfuric acid is most probably V. The WD configuration of the glucosaminidic linkage was inferred from the high positive rotation of heparin itself.

$-poq>H,oH CHzOH

COOH

~OSOZO

H

NH,'

H

OH

V

2 . Acetylative Desulfation Procedures

The application of the classical niethylation t,echiiiyues to hepariii has not yet been s ~ c c e s s f u l .The ~ ~ presence of a high percentage of sulfate groups in the heparin molecule is undoubtedly one of the complicating factors. Thus, a satisfactory method of desulfation would be of value in furthering structural studies of heparin. Considerable progress toward this end was made with the introduction of a chemical method of desulfation The by Wolfrom and M o n t g ~ i n e r y .94~ ~ , t,echnique involves the use of absolute sulfuric acid and acetic anhydride. It is most probable that the act,ivc desulfating agent is the acetylium ion, CH3-CO+, which is known to be produced when strong acids are added to acetic anhydride.9b A possible mechanism of desulfatioii is as follows:

(CH,--CO)zO CHa-CO+ HO-SOz+

+ HzS04 + R-0-SOzOH + HSO,

--+

+ HSO4- + CH;r-COzH R-O-COCH3 + +SOz-OH

+

HzSz07

CH3-C0+

(92) h1. Stacey, private communication; hl. L. Wolfrom, private communication. (93) M. L. Wolfrom and R. hlontgomery, J . A m . Chevi. SOC.,72, 2859 (1950). (94) No use has yet been reported of enzymic desulfation in t,he study of heparin. Enzymes are known which will effect, the hydrolysis of mucopolysaccharitle sulfate groups; see K. S. Dodgson and B. Spencer, Biochem. J . (London), 63, iv (1953). (95) H. Burton and P. F. G. Praill, Quavt. Revs. (London), 6, 302 (1952).

357

T H E CHEMISTRY O F HEPARIN

The use of absolute sulfuric acid tends to create experimental difficulties, and may cause dehydration and degradation. It would be of interest to examine other sources of acetylium ions as potential desulfating agents with respect t o thc obviation of degradation. The effect of the acetic anhydridesulfuric acid reagent on a wide range of simple carbohydrates was studied,93 and permitted evaluation of the stability of glycosidic linkages under reaction conditions. Acetylative desulfation of D-glucose 3-0-sulfate proceeded without Walden inversion, thus suggesting the possible mechanism of desulfation shown below. R-O--SO~-OH

t CHSCO

- @1

R-0-SO,-OH --.---I

R-0-CO-CH~

co

I CHI

@

+

HO-SOz@

Application of the procedure to heparin gave a degraded product which was sulfur-free. I n work described in a later paper,36this material was de-0acetylated and then was found to consume 2 moles of periodate per tetrasaccharide unit, without the formation of formic acid or formaldehyde. The original heparin (see p. 355) consumed 1 mole of periodate, and contained 5 sulfate groups, per tetrasaccharide unit. From these data, together with those reported for heparosinsulfuric acid (see p. 356), Wolfrom and coworkers36have postulated VI as a probable structure for the barium acid CH,OH

p-ko0

COOH

-0 Q0QO

OH

Ba@3020 H

hHSO,OBa(

H

coon

CH,OH

OH

Ba~OSO~O H VI

H

OH

NHS020Ba,

H

0-

OSOIOBat

salt of heparin. Certain structural features of heparin are not yet known or have not been proved unequivocally. They are: (1) the distribution of the 0-sulfate groups, (2) the identity of the sugar unit a t the reducing and nonreducing ends of the chains, (3) the configuration of the D-glucosiduronic linkage, and (4) the presence or absence of branching in the chains. The determination of these points may be important, especially in connection with the relationship of the anticoagulant activity of heparin to its molecular architecture.

3. Other Methods of Degradation of Heparin a. Action of Nitrous Acid on Derivatives of 2-Amino-2-deoxy-~-glucoseand Heparin.-The effect of the action of nitrous acid on 2-amino-%deoxy-~glucose or chitosan (de-N-acetylated chitin) has been known for many

358

A. B . FOSTER AND A. J. HUGGARD

years; 2,5-anhydro-u-mannose (VII) is aff ~ r d e d A. ~monomolecular ~ substance, later recognized as VII,96& was obtained very readily by the action of nitrous acid upon chit0san.~6~ Substance VII is also formed when methyl 2-amino-2-deoxy-a- or 0-D-glucopyranoside (VIII) is treated with nitrous CHiOH

qH,oH

HO H(-+Ho

H

H VII

H

hHg VIII

acid.g7I n both cases, the glycosidic methyl group is split off, but the rate of reaction is much more rapid for the 0-D anomer. A study of treatment of $-heparin (11) and chitosan with nitrous acid showed that the rates of deamination of the polymers closely follows those of methyl 2-amino-2deoxy-a- and -0-D-glucopyranoside (VIII), respectively. The linkages in chitosan are known to be 0-D,and thus there is strong indication that the hexosaminidic linkages in heparin are a-Din configuration, in agreement with the heparin formula (VI) suggested by Wolfrom and The nitrous acid treatment of $-heparin, in addition to providing a correlation between the rate of deamination and the configuration a t the glycosidic center, constitutes a new method of degradation of heparin. Although little has yet been reported on the structure of the products of the deamination, the method is of interest since it leads to the selective cleavage of the hexosaminidic linkages (which are those most stable in acidic hydrolysis). A study of the infrared absorption spectra of D-glucopyranose and its derivativesg8has shown that definite absorption bands may be associated with a-D or fl-D configuration a t the glycosidic center. The absorption is characteristic in mono-, oligo- and poly-mglucose derivatives and in deUnfortunately, the infrared rivatives of 2-amino-2-deoxy-~-g~ucopyranose. spectrum of heparin appears to be considerably influenced by the high percentage of sulfate ester groups present and, a t the present time, information on the configuration of the glycosidic links in heparin cannot be gained thus.99 The infrared absorption spectra of certain mucopolysac(96) For a detailed account of the deamination of 2-amino-2-dcoxy-~-glucose, see S. Peat, Advances i n Carbohydrate Cheni., 2 , 37 (1946). (96a) Personal communication from the late Prof. K. H. Meyer. (96b) K. H. Meyer and H. Wehrli, Helv. Chim. Acta, 20, 361 (1937). (97) A. B. Foster, E. F . Martlew and M. Stacey, Chemistry J1- Industry, 825 (1953). (98) S. A. Barker, E. J. Bourne, M. Stacey and D. H. Whiffen, J. Chem. Soc., 171 (1954). (99) A. B. Foster and M. Stacey, unpublished data.

THE CHEMISTRY OF HEPARIN

359

charides have been studied with respect to -C02H, -NHCOCH, , and -S020H groups.'Oo b. Oxidation in the Presence of Ascorbic Acid.-Skanse and Sundblad'O' have reported an interesting degradation of polysaccharides. They have shown that heparin, hyaluronic acid, starch, and cellulose are degraded in the presence of ascorbic acid and oxygen, with Cu++or hydrogen peroxide also present. The products formed are nonreducing but are acidic and dialyzable. I n the case of hyaluronic acid, the product appeared to be a disaccharide in which the D-glucosiduronic linkage was intact. Few details mere given on the products of heparin degradation.

V. THE ANTICOAGULANT ACTIVITYOF HEPARIN 1. The Biological Activity of Heparin The most familiar property of heparin is its activity as a blood anticoagulant. The coagulation of blood is an efficient and complicated process, and, although the mechanism has not been completely elucidated, the broad outlines are known and may be represented in the scheme shown below.1o2* The effective step in the sequence is the conversion of fibrinogen, a soluble protein present in blood, into its insoluble form, fibrin. The fibrillar network of the fibrin is responsible for the conversion of the liquid phase of normal blood to the solid phase of the clot. The conversion of fibrinogen to fibrin is controlled by the protein thrombin, the formation of which from prothrombin is catalyzed by thrombokinase. Heparin, together with Caff and a plasma co-factor, controls the level of thrombokinase in the blood under normal conditions. In the event of mechanical or chemical injury to tissue or blood platelets, the thrombokinase contained therein is released a t the site of the injury, counteracts the action of heparin, and initiates the coagulation process. I n the condition of thrombosis, where blood clots develop in the veins and arteries, the injection of heparin may cause dispersion of the clots.' Thrombokinase

i -

_ _ _ _ _ - - _ --_- - --Heparin,

Prothrombin

Thrombin and a plasma co-factor

Fibrinogen (100) (101) (102) (103) (1955).

Ca++

I

___c

Fibrin

S. F. D. Orr, R. 6.C. Harris a n d B. Sylven, Nature, 169, 544 (1952). R . Skaiise and L. Sundblad, Acta Physiol. Scarid., 6. 37 (1943). K. H. Meyer, R . P. Piroue and M. E. Odier, Helv. Chirn. Acta, 36,574 (1952). T . Astrup, Advances i n Enzynrol., 10, 1 (1950); W. € 3 . Seegers, ibid., 16, 23

360

A. B. FOSTER AND A. J . HUGGARD

Snellnian, S y l v h and Ju16n5have isolated what appears to be the plasma co-factor from tissue, mast-cell cytoplasm. The co-factor was a lipoprotein containing lecithin, cholesterol, and neutral fats, together with a low molecular-weight polypeptide. Only the six amino acids cysteine, threonine, tyrosine, glycine, leueine, and tryptophan were present in the polypeptide. I n addition to its role in the blood-coagulation process, heparin shows other types of biological activity, such as its action in clearing fat globules from the blood stream (alimentary lipemia)In4and its use in the treatment of frost-bite.' For ail extensive account of the development and use of heparin in medical and surgical practice, the reader is referred to Jorpes' book.' Heparin also possesses activity toward certain strains of bacteria; for example, in a protein-free medium, heparin was found to be bacteriostatic toward Micrococcus pyogenes a t 100 p.p.rn.ln6 At 100,000 p.p.m., heparin was riot bactericidal but caused the development of a high percentage of mutants, smooth + rough, which did not revert. The mutagenic action of heparin is probably related to its ability to displace ribo- and deoxyribonucleic acid from cells.106Heparin can also function as an inhibitor for pancreatic r i b o n u c l e a ~ eA . ~series ~ ~ of derivatives of +heparin (11)in which the amino group was substituted by various organic residues, such as nicotinyl, isonicotinyl, and trifluoromethylphenyl, showed little activity against several species of b a ~ t e r i a . ~The ' bacteriostatic activity of heparin in certain cases appears to be due to the prevention of gelation of cytoplasmic material, which is an essential preliminary to cell division (mitosis).loSThe effect is reversible. 2 . The Structure and Anticoagulant Activity of Heparin

The mechanism whereby heparin functions as an anticoagulant, and the architectural features of the molecule which are associated with the activity, have not been fully elucidated. In the following Sections, an attempt has been made to integrate and evaluate the work done to date on the structure of heparin in relation to its activity. The suggestions subsequently made are not necessarily the oiily interpretations of the known data. It would appear that the high anticoagulant activity of heparin is due to the concerted effect of a series of molecular features which include (1) degree of sulfation (and distribution of the sulfate groups), ( 2 ) molecular size, and (3) molecular shape. Many attempts have been made to imitate (104) (105) (106) (107) (108)

E. A. Nikkila and E. Haahti, Acla Cheni. Scund., 8, 363 (1954). J. R. Warren and F. Graham, J . Bacteriol., 60, 171 (1950). N. G. Anderson and I(.M. Wilbur, Federation Proc , 9, 254 (1950). J . S. Roth, Arch. Biochem. and Biophys., 44, 265 (1953). L. V. Heilbrunnand W. L. Wilson, Proc. Soc. E x p t l . Biol. Med., 70,179 (1949).

THE CHEMISTRY O F HEPARIN

361

oiie or more of these features in the preparatioii of synthetic, sulfated polysaccharides. These studies derived their impetus from the need for a cheap, synthetic anticoagulant to replace the expeiisive natural product. I n view of the iiicompletcness of our knowledge of the structure of heparin in relation to its activity, the preparation of sulfated polysaccharides as synthetic anticoagulants has been undertaken in the past on a somewhat empirical basis. a. Degree of Sulfation.-The recognition of a high sulfate-ester content in heparin led to the examination of synthetic, highly sulfated polysaccharides as potential anticoagulants. Many polysaccharides have been sulfated and studied,1ogincluding chondroitinsulfuric acid, cellulose, inulin, xylan, chitin, chitosan, alginic acid, and dextran. All of the products were found to have anticoagulant activity, but of an order much lower than that of heparin. Further, the toxicity of many of the synthetic, sulfated polysaccharides was such as to contraindicate their therapeutic use. Meyer and coworkers102have studied in detail the sulfation of chondroitinsulfuric acid, since, a t the time, this appeared to be the most active of all the synthetic products. Various methods are available for the sulfation of polysaccharides.ln2Most commonly used is the mixture pyridine-chlorosulfonic acid, but chlorosulfonic acid and the dioxane-sulfur trioxide system have also been employed ; these reagents lead to extensive degradation of the polysaccharide. Meyer and coworkers102used sulfur dioxide as the liquid phase and sulfur trioxide or chlorosulfonic acid as the sulfating agent, and claimed that these systems caused very little degradation of the polysaccharide. Recently,llo formamide and N ,N-dimethylformamide have been used as the liquid phase, with sulfur trioxide as the sulfating reagent. This system, which facilitates a homogeneous reaction, also leads to some degradation of the polysaccharide. Using chondroitinsulfate (molecular weight 30,000) from hog nasal cartilage, and sulfating with the sulfur dioxide and sulfur trioxide system, Meyer and coworkers102obtained a high yield of product; this was little degraded and contained 12-14 % of sulfur (the sodium salt of heparin with a N :S ratio of 2 :5 has 12 % of sulfur). The activity of this product, measured by in vitro methods, was 10-25% of that of standard heparin,"' but in vivo methods (injection into rabbits) indicated 30-45% of the activity (109) See Reference 102 and the references given therein. (110) M. L. Wolfrom, (Mrs.) T. M. Shen Han and T. Y. Shen, private rommunication. (111) A preparation of heparin from beef liver was selected in 1942 by the Department of Biological Standards of the National Institute of Medical Research in London to serve as the provisional international standard. Quoted by (J.) E. Jorpes and S. Gardell, Ref. 32.

362

A. B. FOSTER AND A . J. HUGGARD

of standard heparin. Wolfroni, Shen and Summers86 also reported a low i n vitro activity for sulfated chondroitinsu1fat)e. The variety of methods that have been used for assaying anticoagulant activity makes difficult a comparison of the results of different workers."' It is claimed that the most satisfactory, although not the most convenient, method involves animal tests. By the use of three groups of animals, for control and for injection of standard heparin and of synthetic products, the effect of the latter group is thought to be most accurately assessed. Subsequent references to the activity of sulfated polysaccharides will be limited to comparisons with that of the heparin preparations used by individual authors and not with that of a common standard. Recently, a detailed study of dextran sulfate has been made.113Dextran ( polyglucose elaborated by Leuconostoc is essentially an a - ~ -1+6)-linked mesenteroides. The molecules as synthesized by the micro-organism are gigantic, but they may be degraded to molecules of specific molecular weights by various methods. Two variables were studied in dextran sulfate: (1) degree of sulfation, and (2) molecular weight. It was found that a degree of sulfation higher than 5.2 sulfate groups per tetrasaccharide period, compared to the 5 sulfate groups in heparin, is necessary in order to give appreciable activity, and this was ca. 10 % of the activity of heparin. Dextran sulfates with a high molecular weight (30,000 and 300,000) had a high toxicity. As the molecular weight was reduced, the toxicity fell, and this effect appeared to parallel a decreasing ability of the dextran sulfate to precipitate fibrinogen. It was suggested that, on injection, the high molecular-weight dextran sulfates cause precipitation of fibrinogen, with subsequent agglutination of the platelets around the precipitate and consequent toxic effects. A dextran sulfate of molecular weight of 7,000-8,000 was found to have a low toxicity and to be satisfactory for clinical use. The administration of synthetic, polysaccharide sulfates in large doses may lead to death from internal bleeding.lI4 The preceding data suggest that the activity of heparin is not simply a function of the degree of sulfation, since heparin preparations of high activity contain fewer sulfate groups than the more weakly acting, synthetic products. Karrer and associates116also point out that the relatively rapid disappearance of heparin activity in vivo, compared to the slow disappearance of activity of synthetic, sulfated polysaccharides, may be due to a different mode of action. (112) See B. Blombiick, Margarita Blomback, E. V. Corneliusson and (J.) E. Jorpes, J. Pharm. Pharmacol., 6, 1031 (1953). (113) C. R. Ricketts, Biochem. J. (London), 61,129 (1952); C. R. Ricketts and K . W. Walton, Chemistry & Industry, 869 (1952). (114) T. Astrup and J. Piper, Acta Physiol. Scand., 9, 28 (1945); 11, 1211 (1946). (115) P. Karrer, H. Koenig and E. Usteri, Helv. Chim. Acta, 26, 1296 (1943).

THE CHEMISTRY OF HEPARIN

363

b. The N-Sulfate Group.-The presence of the “N-sulfate” group in heparin and the physiological properties of the polysaccharide, which are unique within the class of mucopolysaccharides, suggest that there may be a relationship. This possibility has been examined recently, following the synthesis of sulfated polysaccharides which contain the “N-sulfate” grouping. I n a study of chitosan sulfate (obtained by the sulfation of de-Nacetylated chitin) , Doczi, Fischman and King116reported obtaining a product with a n activity ca. 50% of that of heparin. No details of the method of sulfation were given, but they indicated that the activity was assayed by an in vivo method. Similar results were simultaneously reported by Wolfrom, Shen and Summers,86who sulfated chitosan with the pyridinechlorosulfonic acid system and isolated a product with ca. 50% of the activity of heparin but with greater toxicity (because of high molecular weight). It was found that the product obtained by sulfation of de-Nacetylated chondroitinsulfate also had an activity ca. 50% of that of heparin. However, direct sulfation of chondroitinsulfate gave a product with < l o % of the activity of heparin. Rickettsll’ did not succeed in obtaining a chitosan sulfate of high activity. The fact that synthetic polysaccharide sulfates have been obtained which have “N-sulfate” groups present and which display activity considerably higher than when these groups are absent would seem to indicate that the “N-sulfate” group per se may be associated in an important way with the activity. Heparin preparations may, however, have a high percentage of “N-sulfate” groups but a low Perhaps significant is the observation that the sulfur content of the active chitosan sulfates (13-17 % for the sodium salts) is considerably higher than that of active, heparin preparations (12%). It would be of interest to compare the activities of preparations of heparin and chitosan sulfate which have similar sulfur content and molecular weight. c. Distribution of the Sulfate Groups.-The distribution of the 0-sulfate groups in heparin has not been fully established; more hydroxyl groups than sulfate ester groups are present in the molecule, and hence several distributions are possible. Enzymic sulfation of the natural heparin polysaccharide most probably leads to a regular distribution of the sulfate groups, whereas the laboratory synthesis of polysaccharide sulfates probably gives a random distribution. The heterogeneity of the sulfation procedures used most extensively to date may also tend to give an uneven distribution of sulfate groups in the final product. d. Molccidar I’C’eighi of Heparin.-Several measurements have been made on the molecular weight of heparin, and the values obtained approximate (116) J. Doczi, A . Fischman and J. A. King, J . Am. Chem. Soc., 76, 1512 (1953). (117) C . R . Ricketts, Research (London), 6, 17-S (1953).

364

A. B. FOSTER AND A.

J. HUGGARD

to 20,000. It is possible that the results may be influenced by the potent polyelectrolyte character of heparin. The values are as follows: 17,00018,000 by the reducing value46;17,000,4520,000,36and 15,00011s from diffusion measurements. 3. The Molecular Shape of Heparin Recent 1vorkS5indicates that heparin preparations may have a low activity and yet have a high percentage of “N-sulfate” groups. It was suggested5*that this and other observations might be explained by postulating the presence of a small number of intramolecular, sulfate bridges of the type IX. The presence of such bridges in sniall amount could hold the heparin in a shape, specific for activity, which would be irreversibly changed by cleavage with acid or alkali, as shown (IX). Detection of these bridges on the basis of analytical data would be difficult, acid ; since the consequent reduction in cationic content I I would be within the normal variation of experiC H - K H - ~ O ~4-0-CH 1 ; ; 1 mental error. The report by Wilander42athat all of the sulfate groups in heparin were free and i bise titratable is also subject t o the same limitation IX (S analysis). There is also some doubt concerning the purity of the heparin preparation used by Wilander, since only recently have methods been established which permit the isolation of (chemically) well defined and (physically) relatively honiogeneous heparin preparations. It is of interest to consider some of the known facts about heparin in the light of the above suggestion of the presence of sulfate bridges. a. Sulfation of Heparin.-The sulfation of a highly active heparin preparation (S, 12 % for the sodium salt) with the pyridirie-chlorosulfonic acid system, used in the preparation of chitosan sulfates,E6causess1’86 an increase in the sulfate content (S, 14.4%) but a reduction in activity of more than 50%. I n fact, the activity of the sulfated heparin was closely similar to that of chitosan sulfate (S, 15.3%) and of sulfated, de-N-acetylated chondroitinsulfate. Possibly, in the process of sulfation, sulfate bridges, if present, would be cleaved and the increase in sulfate content would then give a molecule structurally related to the sulfated, de-N-acetylated chondroitinsulfate. That, in the sulfation, the heparin is degraded below a critical, molecular size would appear unlikely, since the product has appreciable activity, and toxicity (rather than activity) varies with molecular weight.13 It appears that some degree of chemical modification of heparin may he #

I

(118) K. H. Meyer, “Xatural and Synthetic High Polymers,” Intersckncc Pub lishers, New York, N. Y., 2nd Edition, 1050, p. 456.

THE CHEMISTRY OF HEPARIN

365

achieved without decreasing the activity. Thus, Bell and J a q ~ e s reported I~~ that acetylation of active heparin with ketene introduced 4.02 % of acetyl groups without reducing the activity. Deacetylation, again without loss in activity, could be effected by heating in neutral solution. The ease with which the acetyl groups were removed makes difficult any suggestion as to the mode of t,heir attachment to the heparin molecule. That the acetyl content is due t o absorbed acetic acid is not precluded. b. Inactivation of Heparin by Dilute Acid.-Heparin rapidly loses its acThe inactivation has tivity in warm, dilute, acid solution.g*1 6 * 2 3 , 5 9 , been studied in detail by Wolfrom and McNeely,81who found that a 2 % solution of the barium acid salt in 11 % acetic acid, a t 68 f 2", lost most of its activity in 48 hours. Several significant observations were made. (1) The inactivation was solely a function of hydrions; (2) there was no significant change in the low reducing value; (3) there was no appreciable change in the optical rotation; (4)the inactivated product still gave the Toluidine Blue test characteristic of sulfated p o ly s a c ~ h a rid e s ~ lZ0; ~ '~and ) ( 5 ) the inactivated product could be precipitated from aqueous solution by means of excess acetic acid or ethanol. These observations would appear to indicate that there was no marked change in molecular size during the inactivation. Release of amino groups was also observed to occur during the inactivation, although most of the activity had disappeared when an appreciable portion of the amino groups were still combined. Only a small apparent loss of sulfate groups occurred during inactivation (S 12.9% + 11.8 %). Thus, during the inactivation process, the heparin molecule would appear to have undergone little change in elemental content. Wolfrom and McNeelylsl commenting on the result of Charles and Todd,22point out that the action of dilute, methanolic, hydrogen chloride on heparin leads to a much greater rate of inactivation than of sulfate release. It is interesting to note that Jaques and associate^^^ have reported the isolation of an enzyme preparation which inactivated heparin without causing loss of sulfate. The suggested presence of sulfamic bridges in heparin would not be a t variance with the preceding results. Charles and Scott21 reported that heparin was inactivated by dilute nitrous acid. Heparin is known to contain a small percentage (&lo%) of 81 The action of nitrous acid would be expected to free amino cleave the polysaccharide chains wherever a free amino group occurs (see p. 357) and, if these groups are randomly distributed, fragmentation and consequent inactivation would follow. 7 9 8

(119) H. J . Bell and L. B. Jaques, Can. J . Research, 26B,472 (1917). (120) I,. Lison, Conipt. rend. soc. biol., 118, 821 (1935); (J.) E. Jorpes, Actu Med. Scand., 88, 427 (1936).

366

A. B. FOSTER AND A. J. HUGGARD

c. Other Considerations.-Jensen, Snellman and S y l ~ B nhave ~ ~ studied the change in physical properties of heparin during the inactivation caused by recrystallization from warm, dilute acetic They found no decrease in the sulfate content during the inactivation (few sulfur analyses were reported in the paper). The sulfur content of the heparin studied was low and its absolute activity was not stated; the fall in activity was expressed as a percentage of the original activity. As the activity fell (100 + 44 %) a continuous increase in the sedimentation rate occurred (8202.07 -+ 2.7), and the ultracentrifugation diagrams indicated an increased degree of polydispersion. It was suggested that a “structural rearrangement” had occurred, such as a change in shape or hydration. That a large change in molecular weight had not occurred was indicated by the fact that the molecular weight, as determined from the diffusion constant, was 17,000 for the original heparin and 16,600 after inactivation. During inactivation, the frictional ratio fell from 2.5 to 1.81. Assuming that the original heparin molecule was an ellipsoid, these figures would indicate a change in axial ratio of 35:l .--f 16:l. The above data further substantiate the hypothesis of a change in shape rather than in size during inactivation. The shape of the heparin molecule may also be controlled to some extent by the sequence and configuration of the glycosidic linkages. In general, an Q-D-( 1 .--f 4)-linked, unbranched poly-D-glucopyranose tends to be helical in structure, as is evidenced by the classical example of amylose.121When the a linear molecule results; this has fibrous characteristics configuration is 0-D, if the molecular weight is sufficiently high (compare, cellulose). The mucopolysaccharides chondroitinsulfate and hyaluronic acid contain predominantly p-D linkages and, whilst the shape of the former is probably influenced by the D-galactose configuration of the amino sugar, the fibrous nature of the latter is well known. Heparin, however, appears to have predominantly a-D-glycosidic linkages, again rather unusual in the class of mucopolysaccharides. A helical structure somewhat distorted from that of -D amylose could result from the alternating sequence of (3 -+ ~ ) - ( Y and (4 l ) - a - linkages ~ in heparin. The presence of sulfate bridges which could endow the molecule with a certain degree of rigidity, and the possibility of a subtle change in shape on cleavage of these bridges, mould not be difficult to envisage in such a structure. An alternating sequence of (4 1 ) a - i ) - and (3 + I)-a-D-glucosidic links has also been shown to occur in a polysacncharide synthesized by Aspergillus niger,122but little appears to be known about the shape of this molecule. -+

.--)

(121) “Chemistry and Industry of Starch,” R . W. Kerr, ed., Academic Press Inc., New I7ork, N. Y.,2nd Edition, 1950,pp. 170, 185. (122) S. A. Barker, R. J. Bourne and M. Stacey, J . Vltem. Soc., 3084 (1953).

THE CHEMISTRY O F HEPARIN

307

An interesting sulfated polysaccharide which has anticoagulant activity has been described by Marbet and Winterstein.33 They reported the isolation of “@-heparin”from the byproducts obtained in the preparation of heparin from beef lung. It was found that &heparin had (1) a molecular weight of ca. 16,000 (from viscosity measurements) , (2) predominantly p-D-glucosidic linkages, (3) uronic acid and 2-acetamido-2-deoxy-~-galactose (N-acetylchondrosamine) in 1 : l ratio, and (4) 2 sulfate ester groups per tetrasaccharide unit. Chemically, 8-heparin is closely related to chondroitinsulfate, but it has a much higher activity (ca. 25 % of that of heparin). The facts that the &heparin molecule is of approximately the same size as that of heparin and that it has a low sulfate content suggest that its appreciable activity may well be a function of the shape of the molecule. Summarizing, it may be stated that the combination of size, shape, degree of sulfation, and distribution of sulfate groups endows heparin with a specific, anticoagulant activity of a type which may be different from that shown by synthetic, sulfated polysaccharides,l16 and that inactivation appears to take place with relatively little chemical change in the molecule. The sulfation of heparin may well result in its conversion to the class of synthetic, sulfated mucopolysaccharides.

VI. THEBIOSYNTHESIS OF MUCOPOLYSACCHARIDES I n conclusion, it may be of interest to summarize briefly our present knowledge of the biosynthesis of mucopolysaccharides, including heparin. This would appear a t first sight to be more complex than the biosynthesis of poly-~-glucoses~~3 (which is fairly well understood), since more than one sugar species is involved. Hyaluronic acid, one of the mucopolysaccharides, has been the most studied with respect to its biosynthesis, since it is elaborated by certain micro-organisms and is thus readily accessible. Probably, the biosynthesis of individual mucopolysaccharides follows the same broad outline, especially with respect to the elaboration of the amino sugar and uronic acid, so that observations on one mucopolysaccharide may have a general relevance. Some studies have been made on the biosynthesis of chondroitinsulfate in cartilage tissue slices, and it is probably only a matter of time before heparin will be studied in a similar manner. Both D-glucuronic acid and 2-amino-2-deoxy-~-glucoseappear to be incorporated into the mucopolysaccharides through utilization of intact D-glucose, with prior or subsequent oxidation or amination, respectively. This has followed from tracer studies, using ~-glucose-6-C’*and D-glucose(123) S. A. Barker and E. J. Bourne, Quart. Revs. (London), 7,56 (1953); M. Stacey, Advances i n Enzymol., 16,301 (1954).

368

A . B . FOSTER AND A. J. HUGGARD

l-C14, made by Roseman and coworkers,124and by Topper and L i p t ~ n . ~ ? ~ Topper and L i p t ~ n found ~ ? ~ that 2-aniino-2-deoxy-D-glucose and 2-acet%mido-2-deoxy-D-glucose,but not D-glucuronic acid, are incorporated into microbial hyaluronic acid. Thus, D-glucose may be oxidized after its incorporation into the mucopolysaccharide. D-Glucosone may be involved in the reaction sequence which leads to 2-amino-2-deoxy-~-glucose.~~~ Iz6 Thus, D-ghCOSe-l-C14and ~-glueosone-l-C'~ are converted to serum %amino-2-deoxy-~-glucosein rats, but ~ - g l u c o s o n e - l - Cis~ ~the more effective precursor. Lowther and Rogers127have showii that L-glutamine may be involved in the synthesis of 2-amino-2-deoxy-~-glucose in microbial hyaluronic acid. A partially purified, enzyme extract from the mold Neurospora crassa has been found to catalyze the conversion of hexose 6-phosphate to 2-amino2-deoxy-~-glucosephosphate in the presence of L-glutamine.128Phosphorylation of 2-amino-2-deoxy-~-glucoseto give the 6-phosphate has been effected with brain extractlZ9and with crystalline yeast h e ~ o k i n a s e . ~Phospho~" glucomutase from rabbit muscle converts 2-amino-2-deoxy-~-glucose 6-phosphate to the l - p h o ~ p h a t e , ~and 3 ~ 2-amino-2-deoxy-~-glucose 1 ,6-diphosphate may function as a coenzyme in this process, analogously to D-glucose 1,6-diphosphate in the D-glucose series. Similar results have been reported with 2-amino-2-deoxy-~-galactosc.~~ None of these phosphate derivatives have been synthesized by chemical methods. The exchange of acetyl and the uptake of S3504"in chondroitinin slices of cartilage tissue have been studied. The mechanism whereby the backbone chains of the mucopolysaccharides are elaborated is a t present unknown. At least two possible sequences in the biosynthesis could involve stepwise addition of either (1) (alternately) amino sugar and uronic acid or ( 2 ) initially formed disaccharide units which may contain a hexuronic or hexosaminic linkage. I t is, however, clear that, a t this point, a tremendous gap in our knowledge of heparin and other mucopolysaccharides exists. 7

(124) S. Roseman, J. Ludowieg, Frances E. Moses and A. Dorfman, Arch. Biochem. and Biophys., 42, 472 (1953); J. Biol. Chern., 206, 665 (1954); S. Roseman, Frances E. Moses, J. Ludowieg and A. Dorfman, ibid., 203, 213 (1953). (125) Y. J. Topper andM. M. Lipton, J . Biol.Chem., 203, 135 (1953); see also J. F. Douglas and C. G. King, ibid., 202, 865 (1953). (126) C. E. Becker and H. G. Day, J . B i d . Chem., 201, 795 (1953). (127) D. A. Lowther and H. J. Rogers, Biochem. J . (London), 63, xxxix (1953). (128) L. F . Leloir and C . E. Cardini, Biochim. et Biophys. A c t a , 12, 15 (1953). (129) R. P. Harper and J. H. Quastel, N a t u r e , 164, 693 (1949). (130) D. H. Brown, Biochim. et Biophys. Acta, 7, 487 (1951). (131) D . H. Brown, J . B i d . Chem., 204, 877 (1953). (132) H. Bostrom and B. Mansson, Acta Chem. Scand., 6, 1559 (1952). (133) H. Bostrijm and S. Aqvist, Acta C h e w Scand., 6, 1557 (1952) and references cited therein.

Author Index Numbers in parentheses are footnote numbem. They are inserted to indicate the reference when an author's work is cited but his name is not mentioned on the page.

A Abdel-Akher, M., 277(34) Abeles, R.H., 75 Abramovitch, R.A. A,, 217 Adams, G. A., 318 Adler, O., 128(127), 136(127), 153(127) Ahmed, Z . F., 280(36) Aichner, F. X., 214,251(43), 254(43) Alberda van Ekenstein, W., 30 Albon, N., 64,73,74 Alder, K., 16,21(78) Alesander, B.H., 278(42) Algar, W. H., 331,332(217), 333(217) Allerton, R.,131(135), 147(135) Allison, J. B.,221,354 Alm, R. S.,63 Amadori, M.,97, 142(7, l73), l43(8),

146(173), 147(9), 170,171,175,176(3), 178(3' 5)' lS5' 189(4' 5' 6)' 195(4)1 203(3, 4, 5) Anand, N., 131(136b), 132(136b), 134 (136b) Andersen, C. C., 36 Anderson, A. B.,275(22), 280(22), 317 Anderson, A. S.,265(71) Anderson, C. G., 31 Anderson, E.,286,287,302,312,313(76, 77,79) Anderson, N. G., 360 Anderson, R. H., 171, 174, 175(15, 29) 176(29), I88(28), 204(28) Andreasen, A. A,, 86 Andrews, P., 71, 72, 90(69, 70, 71), 128 (122), 133(149), 163(122), 262(53), 263(53), 264(53), 266(110), 267(110), 275(15), 278(15), 279(15, 60,61), 289

Annison, E. F., 355 Anno, K., 40, 50(198a), 87 Antalti, H., 127(116), 137(116), 140(116), I42 (116), I43(1 16), 162(116) Ant-Wuorinen, O., 315

% ;:! &',;.

v., 159(214)

Appel, H., 28 Appling, J. W . , 306 Aqvist, S.,368 Armstrong, E.F., 210, 211,213(13), 229, 240, 247(19), 252(19) Armstrong, K' F'*240 Arni, 1'. C., 91 Arth, G. E., 43 Asahina, Y., 35 Asrhner, T. C., 7 91, 163(226,2277 230)~ Aspina11,G . O',

257(3), 258(3, 23, 12), 259(23), 263 (23), 264(23), 275(16), 278(16), 279 (16),289,303 Astbury, W. T., 307,309(113), 329(113) Aston, J. G., 11, 12(57) Astrupy T'y 3403 3479353(25)9 3597 362* 365(59) Atchison, J. E., 317 AudriethJ L' F'l 354 Auerbach, I., 90 Avery, J., 268(94)

B Babers, F. H., 247(216,217) Bachrach, J.,63,86,260(39) Bacon, J. S.D., 64,276(27, 28) Bachli, P., 254(275) Baer, H.H., 64,252(258) Bailev, J. M., 64 Angel], S.,304 Angyal, S. J., 2, 12(5), 14(5), 15, 16(5), Bair, I,. R., 63,70(24), 71(24) 20(68), 28, 37(5), 47, 48(190), 228, Baker, J. L.,290 253(130) Baker, J. W., 138(161, 162), 141(161,

369

370

A4UTHOR INDEX

162), 143(161, 162), 145(161, 162), 147(161), 148(162), 149(193) Baker, S.B., 23, 47 Baker, W., 267(92), 268(92) B a l d e h , E. R., 194 Baldwin, E., 275(19, 75), 278(75), 280(19) Ball, D. H., 71, 128(122), 163(122), 262 (53), 263 (53), 264 (53) Ball, J., 344 Ballou, C. E., 229 Balston, J. N., 56, 70(7) Barclay, J. L., 106 B&rczai-Martos,M., 214 Barker, C . C., 163(233), 259(26) Barker, G. R., 160(215), 265(68, 69, 71) Barker, S. A , , 3, 12, 13(58), 24(58), 25 (15, 18), 26,27(58), 28(15, 18), 29(15), 30(15), 31(58), 36(58), 37(15), 38(15, 58), 39(15, 58), 40(15), 44(58), 46(15) 89,91, 110, 145(183), 150(60), 156(60), 358, 366, 367 Barrett, E. P., 56 Barrett, F. C . , 56, 70(7) Barrett, J. W., 17, 22(81) Barry, C. P., 99, 102(24), 113, 115(24), 130(24), 208, 217(8) Bartlett, J. K., 90 Barton, D. H. R., 2, 7(4), 12(4), 14, 15 (1,6, 65), 18(4), 21, 29, 53 (92) Bashford, V. G., 9, 10, 23(43, 51), 48, 49 (193) Bastiansen, O., 18, 19(83) Battenberg, E., 311 Bauerlein, K., 246(201) Baur, L., 251(244), 319 Bayly, R. T., 121 Bayne, S., 101, 102(33), 134(33), 139(33), 140(33), 141(33), 145(33), 146(38), 180, 181(50), 185(50), 203(50) Beadle, C., 320 Beaven, G. H., 262(95), 263(95), 264(95), 268 (95) Beck, F., 223, 227(97) Becker, C. E., 368 Beckett, C . W., 2, 12(1), 14(1) Beevers, C. A., 10, 13(54), 16(54), 231 Behrend, R., 229 Bell, D. J., 64, 89, 90, 110, 145(58, 181), 273(1, 4), 274(1, 77), 275(1, 19, 23, 75, 77), 276(1, 4, 23, 27, 28), 277(23), 278(4, 75), 280(19, 65)

Bell, F. K., 343 Bell, H. J., 365 Bellars, A. E., 130(132), 149(132), 156 (132) Bels, W., 306 Benjamin, D. G., 184 Bennett, E., 318 Bennett, E. C., 302,313(79) Berend, G., 32 BerezovskiI, V. M., 152(197a) Berg, S., 252(258) Berger, L., 98, 102(14, 15), 110, 111, 116, 124, 126(15), 159(14, 15), 160(14, 15), 343 Berglund, E., 355 Bergmann, A., 172, 187, 188(18), 189, 197 (18), 198, 200(1S) Bergmann, M., 219, 223, 227(97), 228, 247 (136), 251(239), 253(128, 266, 270) Bergmann, W., 127(114), 136(155), 140 (155), 1481155), 151(114) Bergstrom, S., 339, 342, 351(17), 352, 365 (79) Berkebile, J. M., 87 Berlenbach, W., 136(154), 149(154) Berner, E., 181 Bernoulli, A. L., 215,248(50) Bertho, A , , 112, 129(64), 131(136a), 137 (64, 136a), 153(64), 162(64) Betti, M., 132(140), 155(140), 157040) Bevan, E. J., 320 Beyler, R. E., 43 Bhattacharyya, A. K., 51, 53(201) Bickell, L. K., 324 Binkley, W. W., 33, 56, 65, 67, 68, 69, 70 (7, 49), 83(41), 84,87, 88, 295 Birkofer, L., 98, 101(19), 104, 128(19), 130(19, 132(19), 141(19), 143(19), 146(19), 149(19), 151(In), 156(19), 171, 176(13), 177(13), 186, 188(13), 191(13), 192, 200(13), 201, 203(13), 205(13) Bjorkqvist, C., 314 Blair, M. G., 67, 87, 229 Blanchard, P. H., 64 Blindenbacher, F., 93 Blix, G., 346 Bloch, H., 134(152a), 139(152a) Blomback, B., 362 Blombiick, M., 362 Bloom, P., 321

37 1

AUTHOR INDEX

Bodnrt, A , , 213 Boeseken, J., 21, 75, 231 Bottger, S., 225 Boggs, L. A., 62, 70(24), 71(24) B o g d r , R., 100, 128(124), 129(124), 131 (124), 134(124), 138(124), 139(124), 143(124), 146(124), 151(124), 153(124), 156(124), I64 (124), 249(233) Bohn, E., 241 Boissonnas, R. A , , 82 Bolliger, H. R., 92, 93 Bolomey, R. A,, 79 Bonhoeffer, K. F., 106 Bonner, W. A , , 215,245(56), 248(227), 253 (227) Bose, A. K., 16,51,53(201) Bostrom, H., 346, 353(54), 354, 355, 364 (54), 368 Bott, H. G., 266(74) Bottle, R. J., 346 Bourjau, W., 247(207), 248(207), 251(207) Bourne, E. J., 3, 12, 13(58), 24(58), 25(15, 18), 26, 27(58), 28, 29(15), 30(15), 31 (58), 36(58), 37(15), 38(15, 581, 39 (15,58), 40(15), 44,46(15), 89,91,110, 121, 145(183), 150(60), 156(60), 358, 366, 367 Bourquelot, E., 291 Bower, R. S., 77,88(103), 94 Brading, J. W. E., 268(93) Bratt, L. C., 285, 291, 292(38) Braun, C. E., 146(188) Braun, E., 21, 141(171), 248(226) Brauns, D. H., 210,212, 215,216,219,223, 231, 246(14, 21, 22, 49, 79, 100, 205), 247(49,64,219), 248(220,231), 249(21, 100, 205), 251(243), 253(14, 79, 220), 254(21, 100, 205), 256(21, 49, 79, 100, 205) Brauns, F. E., 326 Breddy, I,. J., 306 Bredereck, H., 225, 242, 246(203), 247 (203) Brewster, J. H., 2, 12(8), 13(8), 14(8), 17 (8)I 20 (8) Brewster, P., 9,49(46) Brice, C., 219 Brigl, P.,33,115,121,136(91,94),137(91), 148(94), 154(94), 217, 219(67), 222, 242 (67), 243 (67), 248 (67), 249 (69), 252 (246)

Brimley, R. C., 56,70(7) Bristow, N . W., 244, 251 (237) Brockmann, H., 79 Bromund, W. H., 148(189a), 149(189a) Brown, A. H., 76 Brown, D. H., 349, 368 Brown, F., 89, 158(208), 260(50), 262(50), 263(65), 264 (50, 65), 267 (83), 273(6), 275(6, 17), 278(17) Brown, G. B., 244 Brown, H. C., 2,12(8), 13(8), 14(8), 17(8), 20 (8) Brown, H. P., 321 Brown, R. L., 200 Browne, C. A., 303, 305(83), 306(83) Browning, B. I,., 286, 291, 292(39), 303, 306, 312,314,315, 318,321 Bruce, G. T., 44,89 Bruckner, Z., 113, 129(67, 68), 219 Bublitz, L. O., 318 Bublita, W. J., 318 Buchanan, J. G., 244 Biichi, J., 28,148(190), 245 Bulen, W. A., 62, 70(24), 71(24) Burgess, H., 325 Burke, W. J., 43 Burkhart, B., 319 Burrell, R. C., 63,91(29) Burton, H., 356 Butler, K., 102, 105(40), 111, 116(62), 132 (40), 133(40, 62), 134(40, 62), 146(40), 158(209), 159(40), 160(40), 161(40), 163(40), 243, 267(108) Butler, M. J., 354 Bywater, R. A. S., 259(25), 260(25) C

Cadenbach, G., 225 Cahill, J. J., 141(172), 159(172) Cahn, R. S., 5 Calderbank, K. E., 15 Cameron, A., 279(54) Cameron, C. N., 137(160), 180, 195(46, 47), 201 Campbell, I. G. M., 16,21(79) Campbell, R., 240 Campbell, W. G., 133(148) Candela, G. A., 79 Cannell, J. S., 140(170d,e) Cantor, S. M., 248(227), 253(227) Cardini, C. E., 350, 368 Carlander, A. T., 62, 70(24), 71(24)

372

AUTHOR INDEX

Carney, D . M., 81 Carrington, T. R., 91 Carruthers, A. E., 160(32) Carson, J. F., 175 Cassidy, H. G., 56,70(7) Castagne, A. E., 318 Castan, P., 247(219) Castle, F. J., 63, 70(24), 71 (24) Cavalieri, L. F., 149(19Oh), 176, 189(35, 36) Cnvallini, G., 134(1521)), 150(152b), 151 (152b) Challinor, S. W., 277(39) Chanda, S. K., 90, 92, 163(229), 258(19, 20), 260(33, 34), 299,300 Chandler, L. B., 118, 137(85) Chang, P., 244 Chapman, N. B., 239, 243(164) Charalambous, G., 151(196a), 158(196a) Chargaff, E., 344 Charles, A. F., 338,339,340,341,342,343, 346, 347, 352, 353(22), 364(13), 365 Charlton, W., 31, 279(73), 280(73) Chase, E. C., 126(108), 152(108) Chatterjee, A., 16 Chaudhuri, D . K. R., 51,53(201) Chavassieu, H. L. J., 187 Cheniae, G. M., 76 Christman, C . C., 219, 267(85) Chu, C.-C., 73 Claesson, P., 248(229) Claesson, S., 58, 61, 63(21), 70 Clark, E. P., 253(267) Clark, G. L., 73 Claus, A., 114 Claus, W. H., 132(139), 171, 176(9) Cleveland, E. A , , 105, 137(46), 138(46), 141(46), 142(46), 144(46), 149(46), 151 (46), 153(46), 180 Cleveland, J. H., 105, 137(46), 138(46), 141(46), 142(46), 144(46), 149(46), 151 (46), 153(46), 180 Cochran, W., 10, 13(54), 16(54) Cocker, W., 189 Cohn, W. E., 76 Cole, W., 190 Coleman, G. H., 80 Collatz, H., 252(249), 254(249), 256(249) Colley, A , , 215 Compton, J., 32, 251(238, 242), 265(105), 266(79), 267(79)

Conchie, J., 260(102) Connell, J. J., 145(183), 278(45) Conrad, C. C., 330 Conrad, H. E., 64, 72, 259(30), 267(89), 277(37), 279(37) Consden, R., 70, 285, 292(4) Conway, H. F., 56 Cook, A. H., 17 Cook, E. W., 115, 218(232), 249(232), 253 (232) Cooke, R. G., 186 Cookson, R. C . , 2,8(13), 16,21,52(13), 53 Cooper, P. W., 87 Cope, A. C., 179 Corneliusson, E. V., 362 Cortese, F., 251 (240), 252(240) Cottrall, L. G., 310 Couch, D. H., 105, 137(46), 138(46), 141 (46), 142(46), 144(46), 149(46), 151 (46), 153(46), 180 Councler, C., 305 Counsell, J. N., 72 Couvert, H., 231 Cox, E. G., 273(2), 277(2) Cramer, F. B., 222,227(95) Cremlyn, R. J. W., 10, 19(50) Criegee, R., 294 Cron, M. J., 342 Cross, C. F., 320 Csendes, E., 188 Csuros, Z., 113, 129(67), 228,247(131), 253 (130) Cuculescu, V., 143(178) Cuendet, L. S., 56, 62, 70(24), 71(24) Cunneen, J. I., 264(60)

D Dacons, J. C., 84, 88(127), 295 D’Addieco, A. A , , 309,317(119) Dale, J. K., 215,229,247(135) Danehy, J. P., 96 Dansi, A., 99, 110, 113, 140(169), 141(27), 147(27), 148(27), 170, 175, 185(7), 186, 188(7), 189, 191(7), 193, 195, 196, 199, 200, 203(7) Davidson, A. L., 321 Davies, G. F., 19 Davoll, J., 119,212,228,244,245,248(24), 251 (135) Day, H. G., 368 de Cholnoky, L., 65, 83(40)

373

AUTHOR INDEX

Dedonder, R., 89 Deferrari, J. O., 123, 124, 131(101), 136 (loo), 137(100), 154(101) Deriae, R. E . , 160(217), 161(217), 210, 214(12) Derx, H. G., 21 Deulofeu, V., 121, 123, 124, 126(99), 131 (101), 136(100), 137(100), 154(101) Dewar, E . T., 133(145), 274(24), 275(24), 276(24), 279(24) d e Whalley, H . C. S., 73 Dhat, &I. L., 131(136b), 132(136b), 134 (l36b) Dickey, E. E., 84, 88(127), 295 Dickson, A. D., 351 Diehl, H. W., 20,23, 44, 239 Dienes, M. I,., 294 Dillon, R. T., 6 Dimler, R. J., 2, 21(12), 56, 63, 70(7, 24), 71 (24), 278 (42) Distelmaier, A., 32, 36(134), 268(100), 269(100) Dittmer, K., 244 Dixon, K . C., 187 Doczi, J., 363 Dodgson, K. S., 356 Doering, W. von I<., 7, 22 Dorr, R . E., 297, 311 Doherty, D . G., 76 Dolliver, M. A , 344 Dore, W. H., 10, 16(55) DorBe, C., 303 Dorfman, A , , 75, 368 Dortmann, H. A , 16,21(78) Douglas, J. F., 368 Dou, A . W., 305 Drulte, E., 7G Dressier, H., 226, 352(121) Drew, H D . K., 5, 260(38), 280(69) Dreyuood, R , 7 6 Dubois, M., 62, 70(24), 71(24) D urr, W , 32, 213 Dnnlop, A . l’., 306 Dunstan, S . , 90,267(88) Durso, D . F., 62

E Earl, J. C , 252(256) ICasterby, D . G., 72 Eckhart, E , 221 Edelman, J., 62, 64, 70(24), 71(24)

Eguchi, H., 292 Ehrenthal, I., 90, 133(144), 145(144), 155 (144), 163(224a), 257(17), 258(17, 22), 259(17), 260(17, 22), 264(17), 275(35), 278(35), 280(35) Elderfield, R. C., 266(76, 77), 268(76) Eliel, E. L., 19 Ellis, G. P., 98, 101(16), 102(16), 109, 110, 113, 115(55), 133(16), 134(16), 145 (16, 55), 147(16, 55), 148(55), 155(16), 156(l6), 159(16), 160(16), 163(16), 164 (16, 55), 243 Elsner, H., 225, 277(38a) Elson, L. A , , 351 Emerson, G. A., 126(108), 152(108) Engels, H., 213 Englund, B., 12 Erdtman, H., 322 Ericks, W. P., 125 Erickson, J. G., 100, 132(138), 144(138), 175 Erlbach, H., 8, 225, 226, 252(116, 119) Erlenmeyer, H., 126(110), 128(110), 132 (110) , 134(152a), 139(152a), 155(110), 203 Ettel, V., 133(177) Euler, E., 252(247) Evans, E. F., 316 Evans, W. I,., 7, 208,227,228(4), 240,241, 253(268, 269) Eveking, W., 224

F Farber, M., 22 Farley, F. F., 81 Farnham, A . G , , 80 Fearon, W. R., 187, 188(70) Felton, G. E., 214 Ferrsnte, G. It., 69 Fischer, A , , 338, 339, 346, 365(!1, 16) Fischer, A. M., 343 Fischer, E., 7, 30, 115, 138(166), 146(166), 149(166), 150(166, 191), 172, 210, 211, 213, 219, 222(33), 223, 228, 230, 244, 245, 247(19, 136), 252(17, 19), 253 (266), 254 (33), 256(33) Fischer, H., 213,219(33), 222(33), 254(33), 256(33), Fischer, H. 0. L., 36 Fischman, A , , 363 Fisher, B. E., 177, 180(39)

374

AUTHOR INDEX

Garcia, C., 292 Gardell, S., 62, 70(24), 71(21), 74, 76, 342 343 (32), 349(32), 361 Garmaise, D. L., 10, 19(50) Garrett, C. S., 121, 136(90), 137(90), 141 (go), 142(90) Gary, W. Y., 286 Gauhe, A , , 64 Gebauer-Fullneg, E., 248(22S) Geerdes, J. D., 90 Gehrke, M., 214, 251(43), 254(43) Georges, L. W., 77, 86, 88(103), 254(273) Georg-Plant, M. M. T., 268(93) Gerecs, A., 218, 219, 228, 247 (133), 249 (132,133,234), 252(132), 254(132) Gierisch, W., 305 Giertz, H. W., 310,331,332(217), 333 Gilbert, E. C., 19 Gilbert, G. A , , 70, 346 Gilbert, R., 220, 247(82), 248(82) Gilbert, V. E., 241 Gill, R. E., 267(81) Gilmour, R., 98,101, 104, 109(11, 12), 110 ( l l ) , 139(12), 142(12), 143(12), 144 ( l l ) , 145(11), 147(12), 151(12), 171 Gilson, A. R., 62, 70(24), 71(24) Glen, W. L., 33 Gniichtel, A , , 248(224), 252(248) Godfredson, W. O., 139(167b), 15l(l67b), 154(l67b) Goebel, W. F., 247(216,217), 254(274), 313 Goepp, R. M., Jr., 4,37,47,65,66(43), 67 (43), 77(43), 78(43), 171,209, 213,229, 235 Goldschmid, O., 330,331,332 Gonell, H. W., 288 Goodman, I., 212, 244, 247(26) Goodyear, E. H., 260(38), 280(69) Gootz, R., 219, 221, 246(87), 254(271), 256 (87) Gordon, A. H., 70, 89, 285, 292(4) Gorin, P. A. J., 73,160(219) Gorrod, A. R. N., 64, 259(28), 266(28) Gortner, R. A,, 304 Gottfried, J . B., IS4 G Gottschalk, A , , 172, 175(22), 180, 181(22, Gakhokidze, A. M., 217, 247(208), 248 49), 183, 185(22), lSS(22), 189, 193, 204(22) (69h, 230), 249(69r), 251(208), 279 Gmf, I,., 249(236), 254(236), 256(236) (53),280(53) Galkoaski, T. T., 63, 83, 84(126), 86, 349 Graham, F., 360 Grant, G. A., 33 Gall, D., 257(8), 258(8), 259(8)

Flesch, H., 228, 247(133), 249(133), 251 (200) Fletcher, H. G., Jr., 23, 34, 36, 44,47, 82, 89(119), 96, 115, 208, 213, 219, 220, 221, 237(36, 85), 238, 239, 245, 246, 248(80, 85, 162b), 251 (36), 253(36, 80, 85, 162a,b), 256(80, 162b) Flood, A. E., 293 Folkers, K., 126(108), 141(172), 152(108), 159(172) Folkes, B. F., 72, 73(75) Forsyth, G., 91 Foster, A. B., 10, 23, 25, 53, 106, 343, 346, 349, 351(68b), 353(55), 355, 358, 360 (67), 363 (55), 364 (55), 365(55) Fowler, W. F., Jr., 314, 315 Fox, D. W., 94 Fox, J. J., 212, 247(26) Fraser, D., 63, 70(24), 71(24) Fred, E. B., 306 Fredenhagen, H., 106 Fredenhagen, K., 225 Freeman, R. D., 317 Frhrejacque, M., 98, 111, 112, 129(61), 132 (18), 137(61), 138(63), 144(18), 145 (61, 63), 147(61, 63), 151(61, 63), 153 (61, 63), 155(18), 157(18) Freudenberg, K., 21, 32, 36, 141(171), 157 (205), 213, 221, 223, 225, 228(102), 248 (30, 226), 249 (35), 252 (27), 253 (35), 254(272), 274(13) Freudenberg, W., 47, 214, 248(89) Fried, J., 82, 89(119), 261(103), 262(103), 264(104) Friend, N. A. C., 187 Frier, R., 129(128), 151(128) Fritzsche, H., 148(190), 245 Frush, H. L., 103, 106, 108, 111, 113(52), 118,122,127(41,96), 131(52), 183,212, 227, 234, 238, 246(22), 253(125, 156), 322 Fiirst, A , , 52 Furth, O., 352 Furst, S. S., 63, 91(29)

A'IJTHOR INDEX

375

Grant, R. A , , 62, 70(24), 71(24), 72,73(75) 142(17), 143(.57),115(57), 146(47), 153 Grasshof, H., 140(170a) (57) Grauniann, E., 317 Hann, R. RI., 6, 20, 23, 25, 27, 30, 35, 38, 39, 10(162, 164), 41, 42, 43, 44 Green, .J. W., 7, 23(35), 278(49) 292 Greene, R. D., 145(185) Hanshoff, G ., 187 Greenwood, C. T., 70, 346 Hanze, A. R., 7 Greville, G. D., 110, 145(58), 274(77), 275 Hardegger, E., 25, 217, 248(69a), 249 (69a), 252 (257, 259) (77) Grewe, R., 30 Hardy, D. M., 268(93) Griehel, R., 226, 252(121) Haresnape, J. N., 12, 13(60) Griffith, C . F., 51, 52(203) Haring, I<. M., 149(191), 150(191) Grimes, W. S., 326 Harloff, ,J. C. , 129(130), 151 (130) Grob, C . A., 93 Harlow, W. M., 321 Grosheintz, J. M., 36 Harper, R. P., 368 Gross, D., 64,73,74 Harris, J. O., 62, 70(24), 71 (24) Griiner, H., 252(246) Harris, M., 312,314 Harris, R. J. C., 359 Gubelmann, I., 125 Gunthard, H . H., 12 Harrison, K., 187 Giinther, A , , 305 Harwood, V. D., 91 Hashima, H., 289 Gunther, E., 252(250) Gcirtler, P., 82 Haskins, F. A., 56 Guilhot, J., 126(111), 132(111), 145(111), Haskins, J. F., 86 155(111), 158(111) Haskins, W. T., 23,27, 30,38,43,44 Hassel, O., 2, 12(2,3,7), 14, 15, 16(2, 69), Guillot-Urbain, G., 140(170c) 18, 19(83), 20(2), 31(69), 232, 233, 234 Gullsnd, J. M., 265(71) Gustafsson, A. M., 331,332(217), 333(217) Hasselquist, H., 187 Gustafsson, C., 285, 286(3), 292, 293, 294 Hassid, W. Z.,275(22), 270(57), 280(22, (49), 303, 307, 310, 313(80, 120), 320 57) (80, 120), 323, 325(45, 120), 326, 327 Hastings, A. B., 117 Hauenstein, H., 92 (45, 111, 120), 328(120) Gut, M., 93 Haug, A., 324 Haus, W., 243 Gyr, M., 93 Hawkins, E. G. E., 89 H Haworth, W. N., 5, 7, 31, 32, 36(140), 39, 40(32), 41(32, 165), 42 (165), 52, 88, Haahti, E., 360 128(123), 155(200,202), 156(202), 163 Haas, H., 311 (228), 219, 220,224, 228(108), 240, 248 Haber, F., 58 (83), 253(83) 258(18), 259(18,25), 260 Haberland, G., 125, 139(104), 151(104), (18,25,31,36,37,38),261 (44), 262(44, 153(104), 154(104) 46), 266(36), 267(84), 268(44, 84), 277 Habrle, J. A , , 331 (39), 278(43, 47, 48), 279(64, 68, 73), Hagglund, E., 285, 291, 292(38), 322, 326 280(66, 68, 69, 70, 73), 289,290(24) Hagdahl, L., 57,58(11), 59,61(18), 63(18) Hawthorne, J. R., 293 Haggis, G. A , , 23 Hayashi, F., 56 Hainsworth, R. M., 278(45) Haynes, L. J., 148(190), 245 Halliburton, G . J., 261(42) Hays, J. T., 117, 118, 119, 137(84) Hammett, 1,. P., 108 Heath, R. L., 155(202), 156(202), 289 Hampton, H . A , , 163(228), 258(18), 259 HebkL, J., 143(177) (18), 260(18) Heidelberger, M., 313 Hanaoka, K., 105, 110, 126(57), 128(57), Heilbrunn, L. V., 360 134(57), 138(57), 140(57), 141(47, 57), Heine, H . W., 24

376

AUTHOR INDEX

l38(21), 142(21), 144(21),147(21), 176, Heiss, H., 39 177(34), 178(34), 179(34),189(33),221, Helferich, B., 32, 35(132), 99, 112, 116, 354 121, 124(65), 127(113), 128(113), 130 (25), 131(113, 134), 136(113), 137(65, Hochreut er, It., 148(190a), 160(19O:t, 221 ) 160a), 138(113), 142(25, 65, 175), 143 Ilockett, R . C., 23, 31, 32(131), 118, 123, 137(85), 294 (25, 65, 113), 146(65, 113), 148(65), 149(25), 150(113), 162(113), 195, 203, Hodge, J . E., 96,97(2a), 101, 104, 112, 134 (36,45), 138(45),141(45), 142(45),146 208, 213, 219, 221, 225, 226, 227(31), (36), 149(45, 66), 150(36, 66), 156(45), 228(5), 241, 242, 243, 244, 245(5), 246 172,175(38), 177, 178(37,38), 179(38), (87,201,202,203), 247(203), 248(224), 180 (23, 38, 39), 181(23), 184(23, 37, 251(31), 252(31, 121, 248, 249, 250, 38), 185(23, 37, 38), 186(37, 38), 188, 253), 254(249, 271), 256(87, 249) 189(37, 38), 190(38), 191(38), 192(37, Hemming, R., 267(92), 268(92) 38), 193(23), 194(23, 38), 195(23, 37, Herbst, R. hf., 148(189a), 149(189a) 38), 196, 200(37, 38), 201, 202, 203(37, Hkrissey, H . , 291 38), 204(38), 205(38) Hermann, H . , 352 Hoffman, D. O., 78 Hermann, P., 128(118), 158(118) Hogg, T. P., 145(180) Hermans, P. H . , 13, 27 Hogsed, M. J., 86 Herzog, R. O., 288 Holly, F. W., 141(172), 159(172) Hess, J. C., 94 Hess, K., 224,247(215,222,223), 252(105), Holmberg, C. V., 317 Holms, W. H., 101, 102(33), 139(33), 140 253(105), 288, 290 (33), 141(33), 145(33), 146(33), 180, Heuser, E., 324, 346 181(50), 185(50), 203 Hewitt, J. A., 171 Holt, E., 336, 339(3) Heyl, D., 126(108), 152(108) Holysz, R. P., 235 Heyne, E., 86 Holser, W. F., 326 Heyns, K., 170, 175 Homan, J. D. H., 348 Hihbert, H., 26 Hickinbottom, W. J., 217, 239(68), 242 Homer, R. F., 52, 53(208) Honeyman, J., 98,99, 101, 102(16, 24,32), (68), 279(73), 280(73) 109, 110, 113, 115(24, 55), 116(32), 130 Hickson, J. L., 62, 63, 86(26) (24), 133(16), 134(16), 144(32), 145(16, Hilbert, G. E., 39, 223 32, 55), 147(16, 32, 55), 148(55), 155 Hiller, A , , 117, 129(128), 151(128) (16), 156(16), 159(16), 160(16), 163 Hilton, H. W., 63,67,70(49) (16), 164(16,55), 208,217(8), 240,243, Hinner, E. F., 284 262 (48), 263 (58) Hiron, F., 9, 49(46) Hirst,, E. L., 56, 70(7), 71, 72, 89, 90, 91, Hopkins, E. W., 306 128(119), 133(142, 146, 148), 145(183), Hotchkiss, R. D., 254(274) 155(2CO), 163(227, 228, 229, 230, 233), Hough, L., 33, 36(150), 56, 62, 64, 70, 71, 72, 73, 90, 133(149), 158(208), 160 219, 220, 248(83), 253(83), 257(4), 258 (220), 163(235), 258(15, 21), 259(15, (12, 18, 23), 259(4, 18, 23, 25, 26), 260 21, 30), 260(15), 265(15), 266(110), (4, 18, 25, 32, 33, 34, 50), 261(4), 262 267(83, 86, 110), 275(15), 276(25), 278 (4, 49, 50, 51, 95) 263(4,23, 49, 50, 65, (15, 25, 46), 279(15, 60, 61), 280(25), 95), 264(23, 49, 50, 51, 62, 65, 66, 75, 289 95), 266(73, 74), 267(4, 81, 84, 86, 88, 90), 268(84, 94, 95), 273(3, 6 ) , 274(3), Houghton, E. O., 321 275(3, 6, 17), 276(5), 277(39, 40), 278 H o w a r d , G . A . , 106, 111(51), 119, 159(51), 160(51), 163(51), 211, 220, 237, 243 (3, 5, 16, 17, 45, 46, 481, 279(5, 16, 40, (15), 253(15) 55, 68), 280(55, 68, 70), 289, 290(24), Howard, R. M., 79 293,299, 300 Howell, W . H., 336,337,339(3), 348 Hisey, W . O., 324 Hiuon, R. M., 32,90,98, 100,105, 136(21), Huber, G., 87, 241

377

AUTHOR INDEX

Huber, H . , 93 Hudson, C . S., 5, 6, 20, 21, 23, 24, 25, 27, 30, 35, 38, 39, 40(162, 164), 41, 42, 43, 44, 49, 51, 52(204), 82, 89(119), 115, 213, 215, 216, 219, 220, 229, 230, 232, 237(36,85), 238,245, 247(48,147, 206, 209), 248(80,85,162b), 249(57,58,62), 251(36, 209), 253(36, 80, 85, 162a,b, 263, 264), 254(57), 256(57, 80, 162b), 261 (45), 312 Huckel, W., 2, 7, 18(29), 19(29), 20 Hughes, E. D., 6, 9, 10(26), 49(46), 234 Hughes, I. W., 146(187) Huisman, H . J., 63,70(24), 71(24) Huniphreys, R. W., 264(75), 279(55), 280 (55, 67) Hunger, A , , 94 Hurd, C. D., 82, 235, 248(227), 253(227), 354 Hurwitz, O., 223 Husemann, E., 290 Hybart, F. J . , 70 Hynd, A , , 98, 102(13), 131(13), 136(13), 143(176), 148(13), 153(13), 154(13), 157(13), 252(255)

Isherwood, F. A , , 289, 290(24) Isserow, S., 11, 12(57) Ivers, O., 213, 225, 252(27)

J

Jackson, D. S., 344 Jackson, E. L., 247(209), 251(209), 312 Jager, R., 305 Jahn, E. C., 317, 318(162), 321 James, A. T., 355 James, S. P., 51, 52(206), 130(133), 258 (13), 259(13), 268(13), 269(13) Jaques, L. B., 337, 338, 341, 344, 345, 347, 348, 353 (52), 365 Jayme, G., 304, 312, 315, 317, 321, 323 Jeanes, A., 223 Jeanloz, R. W., 34, 82, 89(119), 93, 96, 145(180a), 208, 238, 253(162a), 265 (68), 274(10), 275(10) Jenkins, S. H., 321, 324 Jenkinson, D. S., 189 Jensen, H . B., 340, 347,353(25), 365(59) Jensen, R., 344, 348, 364(45), 366(45) Jensen, W., 326 Jochinke, H., 213, 227(31), 251(31), 252 (31) I Johnson, A. W., 244 Johnson, J. M., 215, 230, 247(147) Ice, C . H., 93,94 Johnson, K. C., 180 Inatome, M., 49, 50(198a), 87 Ingold, C. K., 6,9, 10(26), 32(261) see 59), Johnson, T. B., 127(114), 136(155), 140 (155), 148(155), 149(191), 150(191), 48(26b see 191), 49(46) 151(114) Inoue, Y., 99, 100, 127(115), 128(125, 126), 130(125, 126), 131(115, 137), 134 Johnson, W. S., 45 (125, 126), 139(115, 137), l40(115, Johnston, R., 278(51), 279(51) 137), 145(179), 146(125), 147(179,189), Jolles, A., 306 154(115, 137), 156(125, 126), 157(115, Jones, D. I. 279(68), 280(68) 126, 137), 158(125, 1261, 162(115, 137), Jones, J. K. N., 33, 36(150), 56,62, 64,70, 71,72,73,89,90,91, 128(119,122), 133 164(125, 126), 172, 175, 178(32), 188 (142, 146, 148, 149), 158(208), 160 (19), 203, 204(97) (219), 163(122, 148a, 229, 233, 234, Inskeep, R. G., 12 235, 236), 258(15), 259(15, 26, 27, 28), Irvine, J . C., 89,98, 101,102(13), 104, 109, 260(15, 33, 50), 261(41), 262(41, 49, 110(11), 121, 131(13), 132(54), 133 50, 51, 53, 95), 263(49, 50, 53, 55, 57, (54), 136(13, 90), 137(90), 139(12), 65, 95), 264(49, 50, 51, 53, 55, 57, 64, 141(90), 142(12, go), 143(12), 144 ( l l ) , 65, 66, 95), 265(15), 266(28, 110), 267 145(11, 180, 184), 147(12), 148(13), (81, 83, 86, IlO), 268(95), 273(3, 6), 151(12), 153(13), 154(13), 155(54), 274(3), 275(3,6, 15, 17, 18), 276(5, 18, 157(13), 158(54), 171, 247(210), 248 25), 278(3, 5, 15, 17, 18, 25, 45, 461, (225), 252(255, 256), 279(54, 63) 279(5, 15, 18, 60, 61), 280(25), 286, Isbell, H . S., 10, 103, 106, 108, 111, 113 287, 289, 290(26), 293, 299, 303, (lo), (52), 118, 122, 127(41, 96), 131(52), 306, 313(10) 182, 183, 227, 234, 238, 253(123, 125, Jones, R. W., 223 156), 254(123), 322

378

AUTHOR INDEX

Jones, W. G. M., 7,39,40(32,168), 41(32, 168) Jorgensen, JA., 314 Jorpes, (J.) E., 336,339,341,342, 343(32), 344(1), 345, 346, 349(32), 351, ,352, 353(15, 54), 354, 355, 359(1), 360(1), 361,362, 364 (54), 365 Josephson, K., 137(159), 140(159), 228 Jucker, O., 25, 252(257, 259) Juldn, C., 337, 360 Julian, P. L., 190 Jung, H., 214

K Kallinich, G., 194 Karabinos, J. V., 340, 341(23), 342, 345 (36), 347, 348(36, 60), 351, 352(23), 353(23),355(36), 357(36), 358(36), 364 (36), 365(23), 366(23) Karasawa, I., 147(189), 172, 188(19) Karrer, P., 28, 104, 129(130), 139(168), 148(190,19Oa), 150(196), 151(130), 160 (190a, 221), 223, 227, 243, 245, 252 (127), 253(262), 362, 367(115) Kashiwamura, O., 339 Kaster, R. B., 302, 313(77) Katsuki, R., 339 Kawashiro, I., 119 Kawerau, E., 187, 188(70) Keller, A. V., 125 Keller, O., 347, 348(60) Kennedy, R. M., 11, 12(57) Kenner, G. W., 106, 111(51), 119, 159(51), 160(51), 163(51), 244 Kent, P. W., 128(123), 160(216), 261(41, 44), 262(4l, 44, 47), 268(44) Kenyon, W. O., 253(261), 304, 312, 314, 315 Keppler, H., 121, 136(91), 137(91) Keps, J. E., 132(138), 144(138) Kerler, E., 317 Kesselman, J . , 302, 313(79) Kharasch, N., 354 Khym, J. X., 75, 76 Kilpatrick, J . E., 12 Kimura, M., 15 King, C. G., 368 King, J . A , , 363 King, J. L., 339,351 (18) Kinsman, S., 286 Kinxe, L., 247(223)

.

Kitaoka, S., 99, 127(115), 128(125), 130 (125), 131(115, 137), 134(125), 139 (115, 137), 140(115, 137), 146(125), 154(115, 137), 156(125), 157(115, 137), 158(125), 162(115, 137), 164(125) Klages, F., 288, 290(21, 22) Iclemer, A , , 136(154a), 149(154a), 150 (154a) Klemperer, F. W., 117 Klingel, H., 153(199) Klingensmith, C. W . , 227, 253(268) Klingsberg, A , , 163(232), 258(16), 259 (16), 260(16) Klingstedt, F. W., 291 Klyne, W., 2, 21,53(92) Knell, M., 294 Knoevenagel, E., 179 Knopf, E., 32, 221 Knorr, E., 240 Koch, F. K. V., 228, 253(128) Koch, H., 291 Koenig, H., 362, 367(115) Koenigs, W., 240 Kolachov, P., 86 Koller, I., 33 Kondoh, Y., 341,353 Kon’kova, V. A , , 126(109) Konoshin, O., 127(115), 131(115), 139 (115), 140(115), 154(115), 157(115), 162(115) Koon, C. M., 325 Korosy, F., 214 Kosche, W., 127(113), 128(113), 131(113), 136(113), 138(113), 143(113), 146(113), 150(113), 162(113) Koval’sltil, V. V., 79 Kowkabany, G. PIT., 24, 70,285,307(6) Krans, S. A , , 63, 70(24), 71(24) Krantz, J . C., 343 Kreider, L. C., 253(269), 277(30, 31, 32, 74) Kremann, R., 212, 215,247(55) Kroher, E., 305 Kroll, H. J . , 294 Kruckenberg, W., 116, 128(77), 164(77), 186, 195(60), 196, 203(60) Kriiger, G., 151(196b) Krug, E., 248(228) Kruis, J., 158(206) Kruyff, J. J., 198 Kiibler, F., 266(78), 267(78)

AUTHOR INDEX

Kuenne, D. .J., 32,35(132) Iiuhn, R . , 64, 98, 99, lOl(17, 19), 104,110, 113, 116, 126(17), 127(17), 128(17, 191, 130(19), 132(19), 141(17, 19, 27), 143 (17, 19), 146(19), 147(27), 148(27), 149(19), 151(19, 196h), 154(17), 156 (19, 204), 159(17), 161(17), 162(17), 170, 171, 173, 175, 176(13), 177(13), 179,182,185(7), 186,187,188(2,7,13), 189, 191(7, 13), 192, 193, 195, 196, 197, 199, 200(2, 13), 201, 203(2, 17, 13), 205(2, 13), 240 Kuhner, P., 101, 116, 125, 128(35), 132 (35), 135(35), 143(35), 144(35), 155 (35), 162(35), 163(35), 179 Kuiaenga, M. H., 342,345 Kullgren, C., 304 Kunin, R., 75 Kunz, A , , 216, 240, 249(57, 58), 254(57) 256 (57) Kunz, R., 104 Kuri-Szantie, E., 345 Kurth, E. F., 316, 317(154) Kutidze, N . D., 217, 248(69b), 249(69c), 279(53), 280(53)

L LaForge, F. B., 152(197), 159(212) Laidlaw, R. A., 63, 90, 91, 145(182), 163 (223, 231), 257(7, log), 258(10, 14, log), 259(10, log), 260(7, 10, 14), 264 (14), 267(14), 277(33), 279(76) Laird, W. E., 239,243(164) Lake, W. H. C., 52 Laland, S., 102, 105(40), 132(40), 133(40), 134(40), 146(40), 159(40), 160(40), 161 (4O), 163(401, Lamb, I. O., 268(97) Lamb, R. A , , 133(143), 274(8), 275(8) Lambooy, J. P., 159(213,214) Lapin, H., 139(167a) Larinkari, J., 326 Lassettre, E. N . , 86 Launer, H. F., 304 Lavigne, J. B., 141(172), 1591172) Lawler, J. V., 28 Lawson, G. J.: 133(150), 279(62) Lechinsky, W., 251 (239) 'Lederer, E., 56,70(7) Lee, J., 98, 102(14, 15), 110, 111, 116, 124,

379

126(15), 159(14, 15), 160(14, 151, 343, 347, 348(60) Leech, J . C., 84, 88(127), 295, 296(57) Lefbvre, K . U . , 314 Le FBvre, R. J. W., 15 Legay, F., 103, 186 Lehr, H., 126(110), 128(110), 132(110), 134(152a), 139(152a), 155(110), 203 Leitch, G. C., 279(64) Leloir, I,. F . , 350, 368 Lemieux, It. U., 87,208,217,219,220,231 (9), 233, 234, 241 Lens, J., 348 Leonard, F., 124 Levene, P. A., 6, 20, 21,26,32, 34, 35, 36 (113, 141), 152(197), 159(212), 162 (222), 211, 219, 220(15a), 251(238, 240), 252 (240), 253 (265, 267), 258 (91, 260(9), 265(67, 70, 72, 105, 106, 1071, 266(79, 80), 267(79, 85, 87), 277(30, 31, 32, 74), 279(52) Lew, B. W., 37, 65, 66(43), 67(43), 77(43), 78(43) Lewis, B. A., 90 Lewis, H. F., 320 Lewis, W. L., 87, 110, 145(56, 185), 155 (56) Lindberg, B., 64, 72, 218 Lindh, T., 285, 286(3), 293, 294(49), 326 (49) Ling, A. R., 136(158) Link, K. P., 229,251(244), 310,351 Linstead, R. P., 9, 17, 22(81) Lipmann, F., 339,365(16) Lippert, E., 15 Lippold, U., 140(170a) Lipton, M. M., 368 Lison, L., 365 Littmann, O., 224, 252(105), 253(105), 280(72) Lloyd, P. F., 158(209), 267(108) Lobry de Bruyn, C. A., 30, 127(112), 131 (112), 136(157), 137(112), 153(112), 157(112), 162(112) I,ock,M. V., 265(68,71) Lowe, F., 58 Lowenfeld, R., 152(198) Long, C. W., 7, 260(36), 266(36) Lorber, J., 276(28) Lovell, E. L., 330, 331, 332 Lowther, D. A., 368

380

AUTHOR INDEX

Manners, D. J., 145(181) Mannson, B.,355,368 Marecek, W., 247(207),248(207), 251(207) Marbet, R . , 342,367 March, R.E., 308 Marchlewski, L., 102 160(51), 163(51), 211, 212, 228, 243, Maris, S., 312 288 244, 245, 247(135), 248(24), 251(135, Mark, H., Marshall, H.B., 310 237), 253(15) Martin, A. J. P., 62,70, 71(24), 89,285, M 292(4)

Lowy, B. A , , 244 Luckett, S.P., 278(41) Luder, W.F., 181 Ludowieg, J., 368 Lfidtke, M., 288,290,314 Lythyoe, U., 106, 111(51), 119, 159(51),

Maaskant, L., 305 Macbeth, A. K., 186,%7(90) McCasland, G. E., 10 McCloskey, C. M., 80 McCoubrey, J. C., 10,ll(56) McCreath, D., 133(141), 273(2), 277(2) Macdonald, C. G., 15,20(68) Macdonald, T.R. R., 231 MacFadyen, D.A., 117 McGee, P. A . , 314,315 McGilvray, D.I.,91,263(111) McGregor, G. G., 322 Mcllroy, R . J., 257(2),261(42) McKenna, J., 16 Maclay, W.D., 27 McLean, J., 336,340,341(23),342(23), 351

Martin, A. R., 312,314 Martin, M., 324 Martin, R. V . , 62,70(24), 71(24) Martlew, E.F., 10,343,346,349,353(55),

355, 358, 360(67), 363(55), 364(55), 365(55) Masamune, H., 341,353 Matheson, N.K., 47,48(190) Maurer, K., 229,243 May, E. L., 252(254) Mayer, W., 268(100), 269(100) Mazumdar, A. K., 304,305(91) Mazur, R. H., 10 Maszeno, L. W., Jr., 253(260) Mecke, R., 15 Mehltretter, C. L.,26,36,39,278(42) 170,175 (23), 352(23), 353(23), 365(23), 366 Meinecke, K., Meisenheimer, J., 214 (23) Meller, A , , 308,312,315 MacLennan, W. C., 153(199) McNeely, W. H., 65,83(41),295,340,341 Mellies, R . L., 26,36,39 81 (23, Sl), 342(23), 351(23), 352(23), Mertzweiller, J. K., 353,354,364(81), 365(23, 81), 366(23) Mester, L., 221 McNicoll, D., 109,132(54), 133(54), 155 Metcalf, E. A., 43 Meyer, A. S., 93 (54),158(54), 252(255), 279(63) Meyer, E.W., 190 MacPhillamy, H.B., 266(76), 268(76) Meyer, G. M., 32,279(52) Madison, R.K.? 342 Meyer, K., 340,348,352 Maehly, A. C., 93 Meyer, K.H., 82,288,344,353(46), 354 Magnani, A , , 190 355(46), 358,359,361,364 hIagson, M. S., 33,36(150) Maher, G . G., 94 Michael, A , , 240 Mahomed, R.S., 90,91,163(226,227),257 Micheel, F . , 114,129(128), 135(153), 136 (154,154a), 149(154a), 150(153, 154n) (3),258(3,12) 151(128), 164(153), 220, 223,243,247 Maier, J . , 131(136a), 137(136a) (214,215,222), 251(241),280(72) Major, R . T., 115,248(232), 249(232), 253 Micheel, H., 114, 135(153), 150(153), 164 (232) (153),220 Mamaeva, V. V . , 149(192) Mamalis, P . , 101, 127(34), 128(34), 131 Miles, L. W. C., 28 (34), 132(34), 137(34), 141(34), 142 Miller, A . , 80 (34), 143(34), 154(34), 155(34), 157 Miller, E.J., 219,267(84), 268(84) Miller, F.A . , 12 (341,162(34)

381

AUTIIOli INDEX

lkfillcr, C. W., 244 Miller, I . I,., SG, 254(273) hliller, It. IS., 31, 32(131) AIills, J . A . , 2, 4, 10, 12(5), 11(5), 15(5), 16(5), 19(48), 26(21), 31(21), 34120), 35(20, 21), 3 7 ( 5 , 20, 21), 35(21), 40 (21),41c21),44(21),45?21),47,52,244 Minor, F. W., 312 Mist,ele, P., 217, 249(69) Mitchell, H. R., 56 IRIitc~hell,II. I,., 320, 330, 333 AIitford, M. B., 344 Mitrowsky, A , , 112, 121, 124(65), 137(65), 142(65), 143(65), 146(65), 148(65) Mitts, E., 98, 100, 105, 136(21), 138(21), 142(21), 144(21), 147(21), 176, 177 (34), 178(34), 179(34), 189(34) Moggridge, R. C. G., 349 Mohammnd, A , , 138(163), 142(163) Molho, D., 343 Molho-I,acroix, L., 344 Monlchouse, F. C., 348 Montavon, It. M., 25, 217, 248(69a), 240 (60a), 252(257, 259) Montgomery, E. M., 56, 63, 70(24), 71 (24), 86(26), 253(263, 264) 47, 48, 56, 90, 163 Montgomery, R., (224a), 257(17), 258(17), 259(17), 260 (17), 264(17), 342, 345(36), 348(36), 355(36), 356, 357(36, 93), 358(36), 364 (36) Moodie, A . M., 145(184), 248(225) Moody, R. W., 90, 163(230), 258(23), 259 (23), 263(23), 2G4(23), 275(16), 278 (16), 279(16) Moog, K., 279(56), 280(50) Moore, D. H., 344 Moore, M. U., 126(108), 152(108) Morel, A , , 187 Morell, S., 251(244) Morgan, W. T . J., 351, 355 Morley, J. F., 187 Morrell, R. S., 130(132), 149(332), 156 (132) Morris, D. I,., 76 Moses, F. E., 368 Mosettig, E., 252(254) Mostowski, S., 142(174), 171 Mowery, D. F., J r . , 67,69 Miihlschlegel, H., 121, 136(94), 145(94), 154 (94)

a,

Miiller, A . , 42, 121, 120(02), 136(W), 142 (02), 153(92), 1-16(02), 229, 252(252, 2.53) hlukherjee, S., 51, 521200) Rlullen, d . W., 11, 70 Mulley, J . W . , 15 Munro, J., 274(14) Murakanii, S.,278(44) Murphy, M., 309, 317(119) Muskat, I. I<., 266(80) MI1lt, V . , 346, 353(51), 354, 355(5J), 3 G 4 (54) Myers, G. S., 34 Myers, R. J., 75 Myers, W. II., 51 Myrhack, K., 61, 81 N Nageli, C., 139(168), 223, 253(262) Nagai, W., 213, 221(35), 240(35), 253(35) NiLniLsi, P., 100, 128(124), 129(124), 131 (124), 184(124), 138(124), 139( I 24), 143(124), 146(124), 151(124), 153(124), 156(124), 164(124) Kanji, D. R., 136(158) Neely, W. B., 89, 351 Neimann, W., 252(245) Nerdel, F., 20 Ness, A. T., 25, 30(111), 35, 41, 42, 43 Ness, R. K., 44, 115, 219, 220, 221, 237 (85), 238, 239, 245, 246, 248(80, 85, 162b), 251(200), 253(80, 85, 162b), 256(80, 162b) Neuberg, C., 252(245) Neuberger, A., 349 Newmark, N . Z., 244 Newth, F. H., 32, 51,52, 53(208), 234,235 236 (161), 237 (161), 239 (1 61), 242(160) Nicholas, S. D., 214 Nichols, S. H., Jr., 146(188) Nicholson, L.W., 86 Nickerson, R. F., 330,331,332 Nicolet, B. H., 116, 117 Nielsen, E. J., 139(167b), 151(167b), 164 (167b) Niemann, C., 117, 118, 119, 137(84), 214 Niemann, R., 288, 290(22) Nikkila, E. A , , 360 Nishida, K., 280 Noe, A,, 32

382

AUTHOR INDEX

Noggle, G. R., 76,79 Noll, A., 306 Norberg, E. J . , 90 Norman, A. G., 287, 307, 308, 300(113), 321, 324, 333 Norris, F.W., 304 Noth, H., 115

0 Obermanns, H., 311 Odier, M. E., 359, 361 (102) O’Dwyer, M.H., 300,301,302,313 Ohle, H., 8,23,32,33,36,48(42),198,225, 226, 247(207, 212), 248(207), 251(207, 212), 252(115, 116, 117, 119, 120 247) Ohliger, P . H., 347, 348(60) O’Keeffe, A. E., 343, 344 Olcott, H. S., 138(163), 142(163) Oldham, M. A , , 262(48) Oldham, J. W. H., 247(210), 273(1), 274 (I), 275111, 276U) O h , S. M., 87 Ollinman, P. I., 326 Ollis, W. D., 267(92), 268(92) Olsen, F., 324 O’Meara, R. A. Q., 189 O’Neill, A. N., 63, 86, 349 Onodera, K., 99, 100, 128(125, 126), 130 (125, 126), 131(137), 134(125, 126), 139(137), 140(137), 145(179), 146(125), 147(179, 189), 154(137), 156(125, l26), 157(126, 137), 158(125, 126), 162(137), 164(125, 126), 172, 175, 176, 178, (32), 188(19), 193(33), 203, 204(33, 97) Onuki, M., 277(30a, 37a,t>,38, 38s) Orr, S. F. D., 359 Ortenblad, B., 61 Otis, L., 287 Ottar, B., 2, 12(2), 14(2), 15(2), 16(2), 20 (2), 232, 233, 234 Otterson, H., 351 Ough, L. D., 50 Overend, W. G., 23, 25,42,48(171), 51,52 (199), 53, 102, 105(40), 106, 131(135), 132(40), 133(40), 134(40, 151), 145 (186), 146(40, 187), 147(135), 159(40), 160(40, 217), 161(40, 217), 163(40, 237), 210, 214(12) Owen, L. N., 9,23,28 Oxford, A. E., 91

P Pachaly, €I., 30 Pacsn, E., 7, 23(35, X ) , 70, SOX, 217, 218 222, 227, 232, 234(1), 238(1), 247(72), 248172, 152), 249(72, 235, 236), 254 (2361, 256(236), 274(0), 278(49), 312 Pakrashi, S., I t i Pal, K. B., 304, 305(91) Palmer, A,, 90 Pan, S. C., 86 Partridge, S. M., 70, 180, 181(40), 285, 293 Patterson, J., 288 h a t , S., 3, 8(16), 10(16), 22(16), 23(16), 51, 52, 110, 145(59), 155(202), 156 (202), 208, 224, 259(25), 260(25), 262 (46), 358 Peel, E. W., 159(172) Percival, E. E., 92, 151(196a), 158(196a, 207), 163(224), 247(213), 257(5), 258 (20), 259(5), 260(5), 261(5), 266(5), 267 (82), 269 (82) Percival, E. G. V., 90,91,92,99,133(145), 135(152c), 145(183), 158(207), 163 (223,225,229,230,231,236), 197,224, 228(108), 247(213), 254(275), 257(4, 6), 258(6, 10, 14, 19,20,23), 259(4, 10, 23), 260(4, 10, 14, 33, 34), 261(4), 262 (4), 263(4, 23), 264(14, 23), 267(4, 14, 82), 269(82, 102), 274(12, 14), 275 (16, 24), 276(24, 29), 277(40), 278(16, 29, 50, 51), 279(16, 24,29,40, 50, 51), 289 299, 300, 355 Perkow, W., 101, 116, 125, 128(35), 132 (35), 135(35), 143(35), 144(35), 155 (35), 162(35), 163(35), 170 Perlin, A. S., 71 Pervier, N . C., 304 Peters, 0.. 246(202) Peterson, F. C., 317, 321 Peterson, W. H., 306 Petrow, V., 101, 127(34, 116), 128(34), 131 (34), 132(34), 137(34, 116), 140(116), 141(34), 142(34, l l 6 ) , 143(34, 116), 154(34), 155(34), 157(34), 162(34, 116) Petsch, W., 229 Pettersson, S., 285,286(3) Petuely, F., 181, 183, 188, 194 PReger, R., 224,252(105), 253(105)

A4UTHOR INDEX

383

(144), 258(22), 260(22), 275(35), 278 (35), 279(58), 280(35, 58) Raistrick, H., 278(43, 47) Ramsden, H. E., 294 Ranby, B. G., 329 Randall, S. S., 62,70(24), 71(24) Rao, P. A. D. S., 9,49(46) Rao, P . S., 259(29), 260(29), 279(59), 280 (59) Raske, K., 150(194), 245 Rastgeldi, S., 76 Rathgeb, P., 342, 343, 345(36), 348(36, 36a), 355(36), 357(36), 358(36), 364 (36) Ratliff, E. K., 285,291,292(39), 296,304, 306, 317(7), 318(7), 319 Rauch, H., 242 Rauchenberger, W., 222, 247(93) Rautenberg, P., 114 Rayman, B., 158(206) Raymond, A. L., 6, 7, 20, 23(38), 26, 32, 36(113, 141), 208, 245(2), 248(2), 258 (g), 260(9) Reber, F., 93, 268(98), 273(7), 274(7), 275 (7 1 Rebers, P . A , , 90 Reckhaus, M., 197 Redd, J . C., 302,313(76) Redernann, C. E., 214 RBe, A., 132(139), 171, 176(9) Reeves, R . E., 2,13(11 see 63), 16 (10, l l ) , 22(10, l l ) , 23(11), 25(11), 43, 44(10, 178), 50, 232, 237, 254(274), 276(26), 300 Reich, W. S., 79 Reichstein, T., 30, 43, 51(179), 52(179), 92, 93, 94, 268(98), 273(7), 274(7), 275 (7) Reid, S. G., 91 Reid, W. W., 91 Reinecke, F., 284 Reinert, M., 340 Reinhard, H., 32, 36(134) Q Reiter, R., 139(167b), 151(167b), 154 (167b) Quastel, J. H., 368 Remick, A. E., 24 Quinn, E. J., 83, 84(126), 86 Resch, C. E., 304 R Reynolds, D. D., 7, 208, 227, 228(4), 240, 241, 253(261) Raacke-Fels, I. D., 106 Reynolds, T. M., 222,247(221) Rsbe, A, 219,253(266) Rafique, M. C., 133(144), 145(144), 155 Rice, F. A. H., 342,351

l’helps, 17. P., 215,247(48), 260(35) Phillips, D. M. P., 62, 70(24), 71(24) Phillips, G. O., 234, 235, 236(161), 237 (161), 239(161), 242(160) Pictet, A , , 247(211, 219) Pigman, W. W., 4, 96, 105, 137(46), 138 (46), 141(46), 142(46), 144(46), 149 (46), 151(46), 153(46), 171, 180, 209, 213, 227,229, 235, 253(123), 254(123) Pinczesi, I., 129(129) Piper, J., 362 PirouB, R . P., 359,361 (102) Pitzer, I(. S., 2, 12, 14 Plnisance, G. P., 305 Plankenhorn, E., 266(78), 267(78) Plate, E., 129(128), 151(128) Plattner, P. A , , 52 Plisov, A. K., 150(195) Plochl, J., 102 Pollaczek, H., 117 Poos, G. I., 43 Pope, T. H., 290 Porck, A., 32,35(132), 142(175), 203 Porter, C. R., 32,36(140), 260(31) Porter, W. L., 56 Portz, W., 99, 112, 130(25), 137(160a), 142 (25), 143(25), 149(25), 195,203(85) I’osternak, T., 117,252(251) Powell, W. J., 306 Praill, P. F. G., 356 P rat t , J. W., 49, 50, 261(45) Prelog, V., 2, 12, 14 Preston, R . D., 307,309(113) Price, C. C., 294 Prim, D. A., 93 Prochownick, V., 225, 230, 247(148), 251 (148) Pruckner, F., 186 Pryde, J., 264(75), 279(55), 280(55,67,71) Purdie, T., 89, 264(61), 267(91), 268(91) Purves, C . B., 260(35), 312 Putman, E. W., 279(57), 280(57)

384

.\UTIIOlt

Rich, F. V . , 217 Richards, li:. I,., 73 Richards, G. N., 160(218) Richter, G. A . , 324 Richtniyer, N. I<.,20, 21, 30, 41, 49, 50, 51, 52(204), 96, 208, 216, 245, 247 (206), 248(6), 249(6, 62), 253(264), 261 (45) Ricker, A . G., 344 Ricketts, C. R., 362, 363 Rienits, K . G., 345 Riley, H. I,., 187 Ringier, B. H., 118(100),215 Itisser, W . C., 348 Rist, C. E., 26, 36, 39, 101, 104, 112, 134 (36, 45), 138(15), 141(45), 142(45),146 (36), 149(45, 66), 150(36, 66), 156 (U),175(38), 177,178(37,38),179(38), 1SO(38), 184(37, 3 8 ) , 185(37, 38), 186 (37, 3 8 ) , lSS(37, 3 8 ) , 189(37, 38), 190 (38), 191(38), 192(37, 38), 194(38), 195 (37,38), 196,200(37, 38), 201, 202, 203 (37, 38), 204(38), 205(38), 223 Rit,chie, 6. G., 274(12) Rittenhouse, R. C., 292 Ritter, G. J . , 316, 317(154, 155), 320, 321 Rol)ert,s,-1. D., 10 Roherh, 1’. ,J. I’., 64 Robertson, G. J., 35, 51, 52(203), 133 ( l a ) , 257(1, 8 ) , 358(1, S), 259(1, 81, 264(62), 274(8), 275(8) Robinson, R., 240 Rodionova, E. P., 152(197a) Rogers, H. J., 368 Rogers, H. R., 306 Rohner, R. G., 50 Rose, K.E., 264(61) Roseman, S., 75,229, 36s Rosen, I,., 180 liosenfeld, D. A , , 21 Ross, A. G., 90 Ross, J. H., 321 Rot,h, J. S., 360 Roth, P., 220 Rousset, bl.B., 125 Itiiell, 1). A , , 280(GO) Ruggli, P., 197 Rundell, J. T., G4 Russo-hlesi, F. %‘I., 344 Rutherford, H. A . , 312

INDEX

Ryan, H., 215

S s:L:td, N . rc., 346 Saarnio, J.,292,293(45),303,307,310,313 (80, 120), 315, 320(80, 120), 323, 325 (45, 120), 326, 327(15, 111, 120), 328 (120) * Snbalitsc,hkn, T., 99, 144(23), 145(23) Saccarello, A , , 134(152b), 150(152t)), 151 (152,) Sachs, M., 20 Salomon, H., 104 Sanderson, E. S., 63,70(24), 71(24) Sandlie, S., 181 Sands, I,., 286 Sanger, F., 355 SanniB, C., 138(164), 139(164, 167a), 140 (164, 170h,r) Sarett, I,. H., 43 Sarkar, P. B., 304,305(91) Sarten, P., 304 Sato, T., 124, 138(10:1) Sattler, I,., 73, 184 S:tunders, D. R., 22 Savage, W. I,., 94 Sayre, R., 215 Scattergood, A., 31, 32(131) Schaefcr, G . , 200 Schaffer, R., l9%5 Schall, E. D., 62 Schardin, H., 57 Schenker, K., 12 Scheurer, 1’. G., 214 Schiff, H., 97, 102 Schindler, O . , 94 Schinle, R,, 115, 121, 13G(O-1-), 148(94), 154 (94) Schirp, H., 131(134) Schlamowitz, M., 275(21), 277(21), 279 (21) Schlutmch, H. H., 2:0, 211, 220, 222, 224 225, 230, 247(82, 03, 148), 248(11, 52, 110),251(16,20, 148),279(56),280(56) Schliichterer, E., 110, 145(59) Schmidt,, I<., 317 32, 36(134), 39, 187, 252 Schmidt, 0. T., (258), 261 (40), 266 (78), 267 (78), 268 (96, 99, loo), 269(100) Schmied-Kowarzik, V., 172

AUTHOR INDEX

Schmitz, A , , 33S,348, 365(9) Schnritlmiiller, A , 225 sciloctl, rr. . J . , 70 Schodcler, H , 79 Schvinfeltlt, I< , 139(1G7h), 151(167t)), 151 (167t)) Schoettler, J. It., 287,289(16 see 27), 308, 309( 117) Srholl, R., 352 Scholz, H., 213, 248(30) Schoorl, N , 138(165), 14l(lG5), 14R(IG5), 145(165), 151 (1G5), 154(105) Sctiotger, A. W., 290 Sehorning, P., 321 Schotte, H., 228, 247(136), 251(239), 253 (270) Schuetz, R. D., 149(190b), 176, 188(35) Schultze, A , , 222, 247(93) Schwat), G., 317 Schwartz, D. E., 344, 353(46), 354, 355 (46), 364 (46) Schwarz, ,J. C. P , 189 Schwarz, I’., 189 Scott, D. A , , 338, 339, 340, 342, 343, 346, 347, 352, 364(13), 365 Scroggie, A. G., 380 Seebach, A , , I04 Seebeck, E., 94 Seegers, W H., 359 Seeley. G. M , 302, 313(76,77) Seligman, A. M , 5, 247(218), 251(218) Shafizadeh, F., 134(151), 163(237) Shannon, J. A , , 343 Sharp, V. E., 246(204) Shechter, H., 2, 12(8), 13(8), 14(S), 17 (81, 20(8) Sheffield, E. L., 23 Shen, T M , 354, 355(86), 362, 363, 364 (86) Slien, T . Y., 361 Shen Han, T. M., 361 Shilling, W. L., 33, 67, 68, 87(50), 181 Shiiiri, 1). A , , 116, 117 Shishiyama, J., 99, 128(125), 130(125), 134(125), 146(125), 156(125), 158(125), 164(125) Shonle, H. A , , 125 Shoppee, C. W , 10, 19(50), 21, 53(92) Shurik, C. H , 126(108), 141(172), 152 (108), 159(152) SicB, J., 39

385

Yiegel, B., 79 Siegfried, IV., 24 Simmons, J . I{ ., :K30 Simon, 8., 261(40) Simoii, E., :105 Sisler, 13. H., 354 Sitch, D. A., 310 Sjoberg, K., 32, 36(139) Sjollema, B., 120, 136(89) Skanse, B., 359 Skinner, A. F., 247(210) Skraup, Z. H., 212, 215, 247(55) Smart, C . L., 288, 299 Smeykal, K., 274(13) Smirnoff, A. P.,227, 243, 252(127), 253 (262) Smith, C. S., 347, 348(60) Smith, F., 51, 52(206), 56, 62, 70(24), 71 (24), 90, 102, 105(40), 111, 116(62), 128(120, 121), 130(133), 132(40), 133 (40, 62, 141, 144), 134(40, 62), 145 (144), 146(40), 155(144,201), 158(210), 159(40) 160(40),161(40), 163(40,224a), 214,241,243, 257(17), 258(13, 17, 221, 259(13, 17), 260(17, 22), 261(43), 262 (52), 263(43, 52, 56), 264(17, 43, G O ) , 2G8(13, 56), 269(13), 273(2), 274(11), 275(11, 35),277(2, 31,34),278(35,41), 279(11, 58), 280(35, 58) Smith, J. A. B., 266(73,74), 280(70) Smith, 1., I., 171, 174, 175(15), 188(28), 204 (28) Smith, S., 268(97) Smyth, E. M., 348 Smythe, B. M., 15 Snell, C. T., 306 Snell, F. D . , 306 Snellman, O., 337, 344, 346, 348, 360, 364 (45), 366 (45) Sobotka, H., 240 Soff, K., 223, 228(102), 254(272) Solmssen, U. V., 124, 148(190), 245 Soltzberg, S., 47 Somerville, J. C., 276(29), 278(29), 278 (29) Sorkin, E., 43, 51(179), 52(179) Sorokin, B., 97, 101, 102(6), 130(6), 132 (6), 134(6), 144(6), 147(6) Soutar, T. H., 355 Sowden, J. C., 32, 35(132), 195 Sparmherg, G., 242

386

AUTHOR INDEX

Spaultling, 1,. B., 342, 345 Speedie, T . II., 257(1), 258(1), 250(1) Spencer, B , 356 Spencker, K . , 225, 252(115) Spitzer, It., 2, 12, 14(1) Spoehr, €1. A , , 174 Sponsler, 0. L., 10, 16(55) Spriesterbach, D., 277(34) Spurlin, H. M., 316 Srivastava, H. C., 51,52(200) Starey, M., 7, 10, 25, 39(32), 40(32, 168), 41 (32), 51,52(206), 53,89,91,100,102, 105(40), 110, 111, 116(62), 121, 128 (123), 132(40), 133(40, 62, 150), 134 (40,62,151), 145(59, 183,186), 146(40, 187), 150(60), 156(60), 158(209), 159 (40), 160(40, 216, 217), 161(40, 217), 163(40, 237), 210, 214, 220, 241, 243, 246(204), 248(83), 253(83), 261 (41, 44), 262(41, 44), 267(108), 268 (44), 278(43,47,48), 279(62), 346,349, 351(68h), 352, 353(55), 355, 356, 358, 360(67), 363(55), 364(55),365(55), 366, 367 Stadler, P., 211, 220(18) StanEk, J., 145(186) Staudinger, H., 284 Stauffer, H., 215, 248(50) Stavely, H. E., 82, 89(119), 264(104) Steengaard, I., 139(167b), 151(167b), 154 (167b) Steiger, M., 30 Stein, J. Z., 156(203) Steinmann, H. W., 294, 295(50), 296(50), 297 (50), 298(50), 299 (50), 328 Stepanov, A. V., 149(192) Stephens, J. R., 16 Stephenson, J. N., 321 Stevens, W. H., 248(228) Stewart, I,. C., 49 Stewart, W. T . , 302, 313(76) Stiller, C. J., 344 Stiller, E. T . , 21, 32, 34, 35, 265(67) Stillings, R. A., 306 Stitch, S. R., 62, 70(24), 71(24) Stoffyn, P. J., 93 Stone, W. E., 136(156) Storch, K., 317 Strain, H. H., 56, 70(7), 174 Straws, R., 102 Streight, H. R. L., 155(200)

Std’chrlIlLts, I,. I., 152(107;1) Strohele, I < , ,98, 101(17), 116, 126(17),127 (17), 128(17), 141(17), 143(17), 154 (17), 150(17), 161(17), 162(17), 179 Stu:irt, E. R . , 189 Sturgeon, B., 101, 127(34), 128(34), 131 (34), 132(34), 137(34), 141(134), 142 (34), 143(34), 154(34), 155(34), 157 (34), 162(34) Suckfull, F., 247(214) Sugihara, .J. M., 7, 88 Sullivan, W . R., 28 Summerhell, R. K., 16 Summers, C. G., 354, 355(8(3), 362, 363, 364 (86) Sundblad, L., 359 Sundman, J., 292, 293, 204(49), 307, 322 (112), 323, 325, 326, 327(45) Sussman, S., 125 Sutherland, G., 275(21), 277(21), 279(21) Srizuki, M., 341, 353 Svanberg, O., 32, 36(139) Sveda, M., 354 SylvBn, B., 337, 344, 348, 359, 360, 364 (45), 366(45) Synge, R . L. M., 89 Szasz, G . J., 11, 12(57)

T Tachi, J., 258(24), 259(24) Takimoto, H., 35 Talbot, B. E., 56, 70(7) Talley, E. A , , 7, 208, 227, 228(4), 240, 241 Tamm, C., 93, 268(101) Tamm, C. O., 81 Tanret, C., 70, 229 Tatchell, A. R., 101, 102(32), 116(32), 144 (32), 145(32), 147(32) Tauber, H., 193, 194(81) Taylor, E. W., 314 Teece, E. G., 160(217), 210, 214(12) Telfer, R. G. J., 91 ter Kuile, J., 150(196) Tessmar, K . , 8 Teves, D., 311 Theis, H., 194 Thiel, H., 247(212), 251 (212) Thomas, B. B., 317 Thomas, T . G. H., 135(152c) Thompson, A., 49,50(198a), 63,83,84(126), 86, 87, 91(29), 349

387

AUTHOR INDEX

Thompson, R. T . , 121, 136(90), 137(90), 141(go), 142(90) Thurkauf, M., 93 Tilghman, B. C., 322 Timell, T . E., 317, 318(162) Tipson, R. S., 7, 9(39). 34(39), 42(39), 48(39), 70, 96, 211, 215, 219, 220, 224 (50a), 225(50a), 265(70, 72, 106, 107), 261 (85,87), 277(31,32) Tiselius, A,, 56, 57, 58,59, 61, 63 Todd, A. R., 106, 111, 119, 148(190), 159 (51), 160(51), 163(51), 211, 212, 240, 243(15), 244, 245, 248(24), 253(15), 340, 341, 342, 352(22), 353(22), 365 Toepffer, H., 36 Tollens, B., 305, 314 Topper, Y. J., 368 T6th, G., 129(129) Towle, J. L., 146(188) Treiber, R . , 187 Trimble, F., 306 Trister, S. M., 26, 274(9), 278(49) Tsou, K.-C., 5, 247(218), 251(218) Tsuchiya, H. M., 223 Tswett, M., 56,70 T u , C.-C., 63, 260(39), 300 Turner, R. B., 2, 7(9a), 18, 19(84), 33 ( 9 4 , 45(9a) Turowa-Pollak, M. B., 19, 20 TydBn, H., 304 Tyree, J. T., 86, 349

U Uhhelohde, A. R., 10, ll(56) Ujhelyi, E., 65, 83(40) Ulmann, M., 79 Unger, E., 305 linnih, C. C., 304, 312(90), 311, 315 Ustcri, E., 3G2, 367(115) V Valatin, T . , 228 Valentin, F., 127(117), 130(117), 131 (117), 132(117), l:34(ll7), 138(117), 142(177), 143(117), 144(117), 154(117), 155(117), 157(117), 158(117), 162(117), 201 Van Beckum, W . G., 316, 317(155), 320 van Ch:trniife, If.,215 Van Clcve, ,J. W., G 3 , 70(21), 71 (21) van Leent, F. H., 127(112), 131(112), 137 (112), 153(112), 157(112), 162(112)

van Roon, J. D., 30 Van Slyke, D. D., 117 Varner, J. E., 62, 70(24), 71(24) Vaughan, G., 53 Verkade, P . E., 30 Vernet, H., 247(211) Viervoll, H., 15, 16(69), 18(69), 31(69) Vincent, M., 140(170b) Visapaa, A., 315 Vischer, E., 92 Viscontini, M., 148(19Oa), 160(190a, 221) Visser, D. W., 244 Voetter, H., 16 Vogl, K . , 226,252(119) von Arlt, F., 215 von Euler, H., 187 von Hochstetter, H., 32, 213 von Kiippen, A., 315 von Miller, W., 102 von Vargha, I,., 8, 23, 36, 225, 252(117) Votohek, E., 102, 127(117), 130(117), 131 (117), 132(117), 134(117), 138(117), 142(117), 143(39, 117), 144(117), 154 (117), 155(117), 157(117), 158(117), 162(117), 163(39), 201 VoulliEme, It., 252(247) Vyas, G. N., 131(13611), 132(136b), 134 (136b)

W Wachmeister, C. A., 72 Wacker, A , , 172 Wada, H., 292 Wadman, W. H., 62, 70, 71(24), 72, 90 (64), 275(22), 280(22) W l l t i , A,, 223 Wagenitz, E., 211, 251(16, 20) Wagstaff, A. I., 273(2), 277(2) Waisbrot, S. W., 200 Walton, K. W., 362 Walz, D. E., 261(103), 262(103) 360 Warren, J. R.., Waters, E. T . , 280(67), 337, 341, 347 Wathln, I<., 303, 310, 313(80, 120), 320 (SO, 120), 325(120), 327(120), 328(120) Watson, M. B., 10 Watt, C., 32.5 Wmkley, 17. n., 56, 83, 88(2G) W e l h , It. I?., 9 Wedemeyer, K. F., 242,243 Wehrli, H., 358 1

388

AUTHOlt INDEX

Weibull, C., 61 Weidmann, H., 139(168) Weil, H., 56, 70(7) Weisblat, D. I., 7, 340, 341(23), 342(23), 351(23j, 352(23j, 353(23), 365(23), 366(23) Wender, S. H., 93,94 Wenis, E., 124 Wernicke, E., 268(96, 99) Westerheke, D., 302, 313(76) Westgarth, G. C., 260(37), 280(66) Weygand, F., 98, 101, 102(20), 116, 128 (35), 132(20, 35), 134(20), 135(35), 142(20), I43(35), 144(35), 146 (20), 147(20), 149(20), 152(198), 155(20, 35), 156(20,204), 162(35), 163(20, 3 5 ) , 164(20, 238), 170, 171, 172, 173, 174 (16, 17j, 175, 176, 177(16), 178, 179, 150, 182, 184(16), 187, 185, 189, 191, 192(16, 801, 197, 198, 199, 200, 203(2, 16), 205(2, 16, 80) Whelan, W. J., 64 Whetstone, J., 224,262(46) Whiffen, D. H., 12, 13(58), 24(58), 26, 27(58), 31(58), 36(58), 38(58), 39 (58), 44(58), 358 Whistler, R. L., 62, 63, 64, 72, 86, 156 (203), 258(21), 259(21, 30), 260(39), 263(111), 267(89), 277(3i), 279(37 j , 280(36), 258, 299, 300, 311, 314 Whitaker, h l . C., 125 White, A . , 32 Whit,e, B. B., 294, 295(50), 296(50), 297 (SO), 2% (SO), 290 (50), 328 Whit,e, E. V., 79, 133(147), 258(11), 250 (29), 260(11, 29), 263(59), 264(.59, 63), 275(20), 270(59), 280(59) White, T., 122 Whitehead, W., 51, 52(203) Whitehonse, M. W., 262(47) Whithead, J. A , , 63, 70(24), 71(24) Whittaker, II., 306 Wichterle, O., 102, 143(39), l63(39), 201 Wic!kl)erg, B., O?,, 61, 72

Widmaier, 0 . , - : 3 3 Widmer, F., 223 K’ieKaiicI, F , , 246(201) \Viggiris, I,. I<’., 3, 5 ( l i i , 7, 9, 10, 23(17, 4 3 , 51), 25, 28,32, 30, 40(32), 41, 42, 44, 46(17, 171), 47, 48,49, 51, 52(199,

205, 206), 160(216, 217), 161 (217), 210, 214(12) Wilander, O., 342, 344, 352(31, 42a), 353. 364 Wilbur, K. M., 360 Wilcke, H., 226, 252(120) Wildman, W. C., 22 Wilham, C. A . , 223 Wilhelms, A , , 121, 129(92, 131), 136(92), 142(92), 143(92), 146(92), 252(252) Williams, B. I,.,93 Williams, E. M. I,., 128(119), 262(51), 264 (51) Williams, R. J . I’.,57, 58(11), 63 Williams, T. I.,56, 70(7) Williamson, I R., 289 Williamson, S., 273(4), 276(4), 278(4) Willox, I C., 163(225), 257(6), 258(6), 278 (50), 279 (50) Wilson, E. J., Jr., 249(236), 254(236), 256 (236)

Wilson, W. K., 304 Wilson, W. L., 360 Winkler, S , 241 Winterstein, A , , 340,342,367 Wintersteiner, O., 163(232), 258(16), 259 (16), 260(16)

Wise, L. E., 163(234), 259(27), 285, 286, 289, 291, 292, 296, 302, 303(10), 304, 306, 309, 310, 311, 312, 313, 317(7, 119, 138), 318(7, 140), 319, 320, 321 Witkop, B., 186 Wohl, A , , 123, 126(99) Wolf, A , , 32, 157(205), 221, 24Y(89) Wolf, I., 211, 220(18) Wolf, V., 190, 197, 198 Wolfe, J. K., 39, 40(164) Wolfrom, M. L ,7,33,37,43,41),50(198a), 56, 63, 65, 66 (43), 67, 68, 69, 70 (7, 49), 77, 78, 83, 84, 86, X7, 88, 91(29), !’4, 110, 145(56), 149(190b), 155(56), 176, 181, 180(35, 36), 200, 253(2&,5), 254 (273), 275121), 277(21), 279(21i, 2115, 330, 3 I I , ;M2, N : 3 , 345, 3 17, 34X(;3(i, 3ti:i, G O ) , 340, fl.51, 352, 353, 3.51, 355, 356, 357(36, 93), :35S(36), 361,3(i2, 363, 364(3G, SI, Ni), 31i5(2:1, XI), XX(23) WnIlr:it), F., 16 \Volz, € I. , 164(238) Wood, D. J . C., 42, 46(171), 47, 49(171) Wood, H. B., 87

AUTHOR INDEX

Work, R. W., 294, 295(50), 206(50), 207 (50), 208 (501, 299 (50), 328 Wylani, C. B., 63, 91(29), 145(182), 257(4), 259(4), 260(4), 261(4), 262(4), 263(4), 267 (4), 277 (33,40), 270 (-10)

Y Yackel, E. C., 312 Yamamori, N., 258(24), 250(24) Yantschulewitsch, J., 20 Young, C. R., 267(91), 268(91) Young, F. G., 266(77), Young, G. T., 89 Yonngson, G. W., 10 Yundt, A. I’., 290, 299, 300(64)

Z Zaheer, S. H., 221

389

Zechmeister, L., 56, 65, 70(7), 83(40), 129(129) Zeile, K., 116, 128(77), 164(77), 186, 195 (60), 196, 203 (60) Zelinski, R. P., 82 Zelinsky, N. D., 10,20 Zeller, P., 197 Ze m plh, G., 113,129(67, 68), 208,213,218, 219, 221, 222, 223, 228, 240, 242, 247 (131, 133), 249(132,133, 233, 234), 252 (132), 253(130), 254(132) Zerban, F. W., 73, 303,305(83), 306(83) Zerrwick, W., 222 Ziff, M., 344 Zill, L. P., 75, 76 Zinner, H., 212, 248(25) Zissis, E., 30,41 Zobrist, R., 163(224), 257(5), 259(5), 260 (5), 261(5), 266(5) Zuffanti, S., 181

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Subject Index A Acacia pycnantha, gum, 71 Acetaldehyde, 73 Acetic acid, or-morpholino-, 190 -, a-piperidino-, 190, 196 Acetoacetic acid, ethyl ester, 177, 201 methyl ester, 201 "Acetobr ominolysis ," 223 Acetoin. See 2-Butanone, 3-hydroxy.. Acetol. See 2-Propanone, hydroxy-. Acetolysis, 295 Acetone, acetyl-. See 2,4-Pentanedione. -, dihydroxy-. See 2-Propanone, 1,3dihydroxy-. -, phenyl-. See 2-Propanone, 1-phenyl-. Acetonitrile, 221, 245 Acetylation, of glycosylamines, 110 Acetyl groups, exchange in cartilage tissue, 368 Adenine, chloromercuric salt, 244 silver salt, 244 -, Ne-acy-, 244 -, o-arabinofuranosyl-, 244 -, 2,8-dichloro-, silver salt, 244 -, o-xylofuranosyl-, 244 Adenosine, synthesis of, 244 Adsorbents, for column chromatography, 56 Alanine, N-D-glucosyl-DL-, 136 -, phenyl-, 185 Alcohols, primary, oxidation to aldehydes, 43 Aldehydes, preparation from primary alcohols, 43 Alditols, 24 acetslation of, 25 Aldobiouronic acid, from aspen wood, 303 from wood, 313 xylo-, of jute hemicellulose, 305 -, 0-methyl-, from oak hemicellulose-A, 302 xylo-, barium salt, 313

Aldohexofuranosyl halides, poly-0acetyl-, stable forms of, 232 Aldohexopyranosyl halides, poly-0acetyl-, stable forms of, 232, 233 Aldohexosylamine, N-ace tyl -D -, per iodate oxidation of, 118 Aldonic acids, column chromatography of amides, 94 epimerization of, 7 lactone formation, 14 synthesis of, 25 Aldopentopyranosyl halides, poly-0acetyl-, stable forms of, 232 Aldopentoses, methyl ethers of. See the tables on pages 267 to 266. Aldoses, 2-amino-2-deoxy derivatives, formation of, 170 condensation withaliphatic amines, 100 diamide derivatives of, 121 differentiation from ketoses, 197 methyl ethers, characterization of, 110 Aldosylamines, isomerization of, 169,170 Wsubstituted, preparation of, 99 -, N-nitrophenyl-, 102 -, N-phenyl-, methyl ethers, 110 -, N-p-tolyl-, methyl ethers, 110 Aldotrioses, 174 Aldotriouronic acids, from hemicellulose of white pine, 313 from wood, 313 Alginic acid, 90 sulfated, 361 Allitol, 66 -, 2,4: 3,5-di-O-methylene-, 37 Allose, L - , 66 Alloside, methyl 2,3-anhydro-4,6-0benzylidene- a-D-,51, 52 -, methyl 2,3-anhydro-4,6-di-O-tosylor-D-, 93 -, methyl 2-deoxy-4,6-di-O-tosyl- (Y-D 93

391

392

SUBJECT INDEX

Allosyl t)roniicle, 2,3,4-tri-O-ac deosy- L U - I ) - , 251 Allosyl halides, tetra-0-acetyl- a - I ) - 233 , -, tri-0-acetyl-6-deoxy-~-r-,233 Altropyranose, 1,6-anhydro-P-r-, 50 Altropyranoside, methyl 2,6-anhydroL Y - I - , 21 Altrose, I ) - , derivatives, 216 L - , 66 -, 1,2:3,4-di-O-isopropylidene-~-, 32, 33 -, 1 , 2:5,6-di-O-isopropylidene-, 32 Altrosyl chloride, 2,3,4,6-tetra-Oacetyl-a-o-, 247 Altrosyl halides, poly-O-acyl-, 208 Alumina, for column chromatography, 81, 89,92 fibrous, for column chromatography, 90 Aluminum lithium hydride, 245 Amadori rearrangement, 101, 105, 169205 during attempted tritylation, 116 experimental methods, 175 mechanism of, 177, 178 products of, chemical properties of, 187 color of, 185 cryst,allizntion of, 185 dehydration of, 192 derivat,ives of, 195 dihydro derivatives of, 20.5 enolization and oxidation of, 187 fission of, 194 2-fiiraldehyde forniation from, 193 hydrogenation of, 191 light absorption of, 186 melting of, 185 optical rotation of, 186 redwtone formation I)?., lB-1 structure of, 199 t,aste of, 185 scope of, 173 use of, in chemical syntheses, 171 Amberlite IR-400, 75 Amines, acetonylaryl-, 198 -, galactosylaryl-, 192 -, rhamnosylary-, 192 Amino acids, N-glucosyl derivatives, 175 Ammonia, reaction with reducing sugars, 106

Amnioriiiim I)romide, N -(6 -d e o x y -~ glucosy 1) t rime thy1-, 135 -, Ar-l,-glucosyltrimethgl-, 150 6-tritaylether, 150 -, N - ( 2 , 3 , ~-tri-O-,zcct~1-1~-xylosyl) trimet,hyl-, 164 hmmonium chloride, A-(tet,ra-0-acetyli)~glucosyl)trimetli?il-,150 Ammonium chloroplatinate, N - ( t e t r a O-acet~I-i)-glucosyl)trimethyl-, 150 Ammonium iodide, N-r-glucosyltrimethyl-, 150 Ammonium perchlorate, K-(tet,ra-Oacetyl-D-glucosyl)trimethyl-,150 Ammonium picrate, ~V-u-glucosyli.riinethyl-, 150 tetraacetat,e, 150 A i n o r p h o p h a l l u s konjuc, 289 Aniorphophallus oncophyllus, polysaccharide from corms of, 289 Amygdalin, 66 synthesis of, 240 Amylopectin, separation from amylose, 70 structure of, 86, 349 Amylose, 366 inclusion comples with cellulose, 70 pot.ato, acid hydrolysis of, 64 separation from amylopectin, 70 trimethyl ether, 224 Aniline, action on 2,3,4,5-tetra-Oacetyl-aldehijdo-u-ril)ose, 100 basicity of, 354 condensation with aldose methyl ethers, 110 contiensation with wglucose, 97, 102 phthalnte, 90, 293 reaction with 2-furaldchytie, 306 --, acetmyl-, I90 -, 3,4-dimethyl-. See 3,4-Xylidine. -, uldelr.ydo-o-galactosyl-, 2,3,4,5,6pentlaacetate, 134 -, r-gluropyranosyl-, 97 -, wglucosido-. See Glucosylamine, N phengl -D-. -, N-met.hy1-, 174, 177, 239 -, ?ri-nit,ro-, 125 -, o-nitro-, reaction with sugars, 98 -, p-nitro-, 125

SUBJECT INDEX

-, tet,ra-O-acct,gl-8-u-g:tlactop).ranosgl-, 243 p-Anisicline, conclcnsation n-it,h wglucose, 97, 170, 175 Anomer, t.he term, 230 Anomeric prefixes, history of, 229 Anthocyanins, 209, 240 Anthrone reagent, 76 Anticoagulant suhstances, ass:ty of n r tivity of, 361, 362 synthetic, 361 Apple, gum of golden, 71 Apricots, 93 Arabans, 284 of sugar bect, 90 of unbleached, sulfate pulps, 327 of wood, 326 Arahinal, di-0-acetyl-o-, 229 Arabinonic acid, D-, 66, 180 D-, ammonium salt, 200 D-, phenylhydrazide, 200 I ) - , potassium salt, 66 I,-, 66 L-, chloride, tetraacetate, 200 L - , nitrile, tetraacetate, 123 -, 2,3-di-O-methyl-~-,amide, 263 lactone, 262 -, 2,4-di-O-methyl-~-,amide, 263 lactone, 263 -, 2,5-d-O-methyl-r-, amide, 261 lactone, 261 phenylhydrazide, 261 -, 2,5-di-O-methyl-~-,263 amide, 263 lactone, 263 phenylhydrazide, 263 -, 3,4-di-O-methyl-~-,amide, 263 lactone, 263 -, 3,5-di-0-methyl-o-, amide, 261 lactone, 261 phenylhydrazide, 261 -, 3,5-di-O-methyl-~-,amide, 264 lactone, 264 phenylhydrazide, 264 -, 2-O-methyl-~-,amide, 261 hydrate, 261 ammonium salt, 261 lactone, 261 phenylhydrazide, 261

-, 2-O-nicthyl-~-,gniidc, 262 lactone, 262

.’

-, 3-O-met,hyl-1)-,aniidc, 261 lact.one, 261

-, 3-O-met~hyl-x.-,ainide, 262 lactone, 262 2,3,4-tri-O-methyl-~-, amide, 262 2,3,4-tri-O-methyl-~-,amide, 264 lactone, 264 phenylhydrazide, 264 -, 2 , 3 , 5 - t r i - f ~ - m e t h y l - ~nmide, -, 262 lactone, 262 -, 2,3,5-tri-O-methy-r,-, amitle, 264 lactone, 264 -, 2,4,5-tri-O-methyl-r,-,amicte, 262 Arahinopyrnnoside, methyl 2-0-methylp-u-, 261 -,methyl 2-O-methyl-p-~-,262 hydrate, 262 Arabinopyranosylamine, N-acetyl-L-, periodate oxidation of, 118 -, 2,3-di-O-methyl-N-phenyl-~-, 262 -, 3-O-methyl-N-phenyl-~-,262 Arabinose, 318,322 D-, 66, 173 D-, condensation with 3,4-dimet,hylaniline, 171 D - , 1-diacetamide derivative, 123, 126 D-, separation from 11-ribose, 124 DL-, 72 L-, 64, 66, 77, 173 L-, 1-diacetamide derivative, 122, 127 L-, fermentation of, 306 L - , 2-furaldehyde from, 305 L-, in polysaccharides, 323 -; 3-0-(8-L-arabinopJ.ranosyl)-L-, 72 -, 1,2:3,4-di-O-isopropylidene-~-, 32, 33 -, 2,3-di-O-methyl-~-,90, 262 -, 2,4-di-O-methyl-o-, 261 -, 2,4-di-O-methyl-~-,263 -, 2,5-di-O-methyl-~-,261 -, 2,5-di-O-methyl-~-,263 -, 3,4-di-O-methyl-~-,263 phenylosazone, 263 -, 3,5-di-O-rnethyl-o-, 261 -, 3,5-di-O-methyl-~-, phenylosnzone, 263 -, 2-O-niethyl-~-,261 phenylhydrazone, 261 p-tolylsulfon~~lhydrazone, 261

-, -,

394

-,

SUBJECT INDEX

form 11, 127 2-0-methyl-~-,90, 262 triacetate, 127 phenylhydrazone, 262 -, N - (3,4-dimethylphenyl)-u-, 126 p-tolylsulfonylhydrazone, 262 -, N-(3,4-dimethylphenyl)-~-, 127 -, 3-0-methyl-u-,261 -, N-methyl-L-, 127 -, 3-O-methyl-~-,262 -, N-(4-methyl-2-nitrophenyl)-L-,forms phenylosazone, 262 I and 11, 127 -, 1,2,3,4-tetra-O-acetyl-a-~-, 88 triacetate, forms I and 11, 127 -, tetra-0-acetyl-aldehydo-L-, action of -, N-2-naphthyl-o-, 126 ethanolic ammonia on, 122 -, N-P-naphthyl-~-,128 -, tetra-0-acetyl-L-, 214 -, N-o-nitrophenyl-I,-, forms I and 11, -, I ,2,3,4-tetra-O-acetyl-a-~-, 210 128 p anomer, 210 triacetate, forms I and 11, 128 -, tetra-0-azoyl-&D-, 80 -, N-p-nitrophenyla-L-, 128 -, tetra-0-szoyl-p-L-, 80 -, N-phenyl-D-, 124, 126 -, 2,3,4-tri-O-methyl-~-,264 2,4-dimethyl ether, 261 -, 2,3,5-tri-o-rnethyl-o-, 262 3,5-dimethyl ether, 261 -, 2,3,5-tri-O-methyl-r.-, 90, 264 -, N-phenyl-L-, 124, 128 -, 2,4,5-tri-O-methyl-o-, 262 2,3-dimethyl ether, 128 Arabinoside, methyl 2,5-di-O-methyl-r~-, 2,4-dimethyl ether, 128, 263 263 3,5-dimethyl ether, 128 -, methyl 3,4-di-O-methyl-p-r.-, 263 3-methyl ether, 128 2-henzoate, 263 -, N-p-phenylazophmylene-D-, 126 2-tosylate, 263 -, N-p-sulfamylphenyl-L-, 128 -, methyl 2-O-methyl-~-,261 -, N-p-sulfophenyl-r,-, 128 -, methyl 2,3,4-tri-O-methyl-~-,261 -, N-P-tolyl-L-, 128 -, methyl 2,3,4-tri-O-methyl-a-~-, 264 Arabinosyl bromide, 2,3,4-tri-O-acetyl0 anomer, 264 B-D-, 251 -, methyl 2,3,5-tri-O-rnethyl-~-,262 -, 2,3,4-tri-O-acetyl-p-~-, 210, 251 -, methyl 2,3,5-tri-O-methyl-~-,264 preparation of, 214 Arabinosylamine, L-, 106, 107, 108, 127 -, 2,3,5-tri-O-acetyl-o-, 251 acetylation of, 111 -, 2,3,4-tri-O-benzoyl-p-D-, 251 N-aryl derivatives of, 98 Arabinosyl chloride, 3,4-di-O-acetyl-2-, N-acetyl-L-, 127 chloro-2-deoxy-@-~-, 247 -, N-acetyl-tri-0-acetyl-L-, 127 -, 2,3,4-tri-O-acetyl-p-D-, 247 -, N-carbamoyl-L-, 127 -, 2,3,4-tri-O-acetyl-p-~-, 247 triacetate, 127 Arabinosyl fluoride, 2,3,4-tri-O-acetyl-, N-p-carbethoxyphenyl-L-, 127 &L-, 246 -, N-p-carboxyphenyl-L-, 127 Arabinosyl halides, tri-0-acetyl-D-, 232 -, N-(4-chloro-2-nitrophenyl)-~-, 127 Arabinosyl iodide, 2,3,4-tri-O-acetyl-ptriacetate, forms I and 11, 127 L - , 256 -, N- (3,4-dimethyl-2-nitrophenyl) -L-, Arabitol, D-, 64, 66, 72, 87 triacetate, 127 -, N-(4,5-dimethyl-2-nitrophenyl)-u-, -, 1-amino-1-deoxy-D-, 205 -, 1-amino-1-deoxy-L-, 205 126 -, 1,3-0-benzylidene-~-,30 monohydrate, 126 -, 1,3-0-benzylidene-~-,30 triacetate (furanose?),126 tosylation of, 30 2,3,4-triacetate, 126 -, N-(4,5-dimethyl-2-nitrophenyl)-~-,-, 1-deoxy -1- (3,4-dimethylanilino) +, 192 form I, 127 -, 1,3:2,4-di-O-rnet,hylene-o-,41 triacetate, 127

SUBJECT INDEX

-,

395

Benzimidazole, 5,6-dimethyl-, chloromercuric salt, 244 silver salt, 244 -, 5,6-dimethyl-1- @-~-glucopyranosyl)-, 244 -, 5,6-dimethyl-l-(a-D-ribofuranosy1)-, 244 B anomer, 244 Benzimidazoles, glyeosyl-, 244 Benzoic acid, n m i t r o - , hydrazide, 174 Benzoin, 175 Benzoyl chloride, p-phenylazo-, 79 Benzylamine, compound with 2,3,4,6tetra-0-acetyl-D-glucose,112 Betacoccus arabinosaceous, 89 Betula pubescens, 286 Betula verrucosa, 286, 314 Bicyclo[2.2.l]heptane. See Norcamphane. Bicyclo[2.2.2]octane, 21, 22 Bicyclo[3.2.l]octane, 21 Bicyclo[3.3.0]octane. See Pentalene, ocB tahydro-. Barbituric acid, 305 Birch, acidic xylan from, 303 -, thio-, 305 glucan of, 327 Barley, 324 holocellulose of, 303,314 grain, 91 mannan of, 326 xylan from straw of, 290 pulp from, 327 Beater additive, for paper manufacture, sulfate pulp from, 310, 331 sulfite cooking of, 307, 322 289 sulfite pulp from, 310, 323, 325 Beech wood, hemicellulose-A of, 91 xylan of, 327 hemicellulose fractions from, 300 Black spruce. See Spruce. Beets. See Sugar Beets. Bleaching agents, effect on cellulose, 315 Benzaldehyde, phenylhydrazone, 101 -, 2-(tetra-O-acetyl-~-u-glucopyrano- Blood, coagulation of, 359 heparin, as anticoagulant for, 359 syloxy) -4,6-dihydroxy-, 240 plasma, co-factor in, 359, 360 -, 2,4,6-trihydroxg-, 240 “Body X,” from hemicelluloses, 287 -, 2,4,6-trinitro-, 121 Boric acid, complexes of, with aldosylBenzamide, N-u-glucosyl-, 2-thioethyl amines, 125 ether, 148 with sugars, 231 Benzanilide, 190 Brauns’ rule, for optical rotations of Benzene, 0-dinitro-, 188, 199,202 glycosyl halides, 231, 232 -, l-fluoro-2,4-dinitro-, 353, 355 Brigl’s anhydride, 241 Benzhydrazide, nt-nitro-. See Benzoic Browning, 102, 172, 180, 184 acid, ~n-nitro-,hydrazicle. of N-D-galactosylpiperidine, 194 Benzidine, N ,N‘-di-I.-arabinosyI-, 138 non-enzymic, 96, 193, 191, 202 -, N,N’-di-D-glucosyl-, 136 1 -But:tnesuIfonyl chloride, 121 N,N’-dimaltosyl-, 153 2-Butanone, 3-hydroxy-, 175 3-0-(P-D-gi~laCtOpyranOSyl)-D-,64, 72 Arabonic acid. See Arabinonic acid. Arbutin, 240 -, methyl-, 240 Arylamines. See Amines, aryl-. Ascorbic acid, 66, 188 Aspen, bleached kraft pulp from, 309 sulfate pulp from, 310 sulfite cooking of, 307,322 sulfite pulp from, 310, 323, 331 wood, 287, 303 pulps from, 308, 309 Aspergillus niger, 91, 366 Azobenzene, p-hydroxy-. See Phenol, p-phenylazo-. Azoyl chloride. See Benzoyl chloride, p phenylaeo-. Azoyl chromatography, of sugars. See Column chromatography, use of sugar azoates.

-.

396

SUBJECT INDEX

AT- (N-et hylthiocarbamoyl) -, 129 heptaacetate, 129 Cane juice, analysis of, 87 -, N-(N-methylthiocarl,amoyl)-, 129 Carbamic acid, Ak-glucosyl-, ethyl heptaacetate, 129 ester, 148 -, N-phenyl-, 129 methyl ester, 148 heptaacetate, 129 pentgl ester, 148 -, N-p-sulf amylphenyl-, heptaacet#ate, -, N-(hepta-0-acetyllactosy1)thio-, 129 ethyl ester, 151 tetrahydrate, 129 Carbohydrates, chromatogrnphic ad-, N-p-tolyl-, 129 sorption series, 66 heptaacet.ate, 129 containing carboxyl groups, 312 Cellobiosyl bromide, hepta-O-acet,yl-, relative adsorption on fuller’s earth 222,223 clay, 66 01 anomer, 236, 237, 253 . of ~ o o d285,326 , reaction with methanol, 242 in wood-cellulose preparations, 287 hexa-O-aeet~yl-2-hromo-2-deoxy-a-,253 Carbon, affinity of sugars for, 59 Cellohiosyl chloride, liepta-0-acetyl-, beds or columns of, for sugar purifica216 tion, 56 a anomer, 249 for column chromatography, 62 hepta-0-methyl-, 221 for resolution of sugar mixtures, 56 a anomer, 249 Carboxyl groups, of cellulose, 315 Cellobiosyl fluoride, hepta-0 acetyl-, determination of, in carbohydrates of 212,246 wood, 314 Cellobiosyl halides, poly-0-acetyl-, 232 of 0-ethylcellulose, 316 Cellobiosyl iodide, hept,a-O-acetyl-a-, of wood cellulose, 315 256 Cedar, Western red, 319, 320 Cellohept,aose, a - ,88 pectic material from, 313 Cellohexaose, a - , 88 Celite, 67, 86 Cellopentaose, a-, 88 No. 535, use in column chromatoCellophane, 323 graphy, 64 Cellotetraose, 01.) 88 Cellobiose, 66, 76, 77, 87, 223, 314 Cellulosans, 333 8-, azoatc, 80 Cellulose, 283,366 1-diacetamide derivative, 129 accessibility of, 330 heptnncetate, 129 acet,nte, 296, 323 -, hepta-0-methyl-, 221 comniercirtl, 311 -, octa-0-acetyl-, 212, 216 cffect, of polyoses on, 207 p anomer, 241 viscosity o f solrit,ioiis of, 297 -, octa-O-aeoyl-8-, 81 viscosit,y rat>ioof, 298 Celloliiosides, alkyl and aryl p o 1 ~ -0from wood cellulose, 281 acetyl-a-(and p - ) , reaction u i t h hyyarns, 284 drogen bromide, 222 analyses of, 298 Cellot)iosylamine, 129 heptaacetate, 129 :issorint,ion with sylan, in wood, 307, -, A-acetyl-, 129 309 heptaacetate, 129 1)loclis of, for chrom:it,ogr:tphy, 56 -, Ar,Aklimethyl-,129 enrliosyl-groiip coiitcnl. o f , 284 heptanect:ite, 113, 129 rnmniercinl, from wood, 321 hydroliromide, 329 conipoiintl with ni:innan, 296 -, K-ethyl-, 129 from cotton linters, 284

C

-,

SUBJECT INDEX

acetolysis and chromatography of, 295 analysis of, 298 carboxylic acids in, 312 Cross -and -Bevan, 320 degradation of, 223 by alkali plus oxygen, 346 determination of carboxyl groups in, 315 dispersions of, 323 inclusion complex with amylose, 70 mercerization of, 324 nitrate, 70 oxidation of, 312 in presence of ascorbic acid, 359 with nitrogen tetroxide, 314 powder, for column chromatography, 74,90 reaction with lead tetraacetate, 294 short-chain, 324 solutions of, 323 sulfated, 361 triacetate, 223 wheat-straw, Cross-and-Bevnn, 308 wood, 283 composition of, 333 fine strncture of, 325 preparation of, 316 xylan -con taining , 324 -, alpha-, 328, 329 from holocellulose, 320 from sulfite pulps, 323 in snlfate pulps, 327 in sulfite pulps, 327 nitrate, 330 of black spruce, 296, 320 of Douglas fir, 320 of loblolly pine, 320 of slash pine, 295, 296, 320 of unbleached, sulfate pulps, 327 of unhleached, sulfite pulps, 325 of Western hemlork, 320 of Western red cedar, 320 of woods, 285 so-called, 319 -, beta-, 328, 329, 332, 333 origin of, 330 -, 0 - e t h v l - , c ~ a r l m \1~grnups in, 316 -, gamma-, 32X, 329, 332 -, hydro-, 312

397

preparation of, 90 use in column chromatography, 72 -, OXY-,312 formation of 2-furaldehyde from, 304 of gIuconic acid type, 312, 315 Celluronic acids, 312, 314 degradation of, 315 Celtrobiosyl chloride, hepta-0-acetyl-, 216 cy anomer, 249 Cherry gum, 71 degraded, 90 Chitin, de-N-acetylated, 357 sulfated, 361 Chitosan, 357 action of nitrous acid on, 357 sulfate, 361,363,364 Chlorophyll, 70 Cholesterol, 360 Chondroitinsulfuric acid, 346, 349, 355, 366,368 biosynthesis of, 367 de-A-acetylated and sulfated, 363, 364 sulfated, 361, 362, 363 Chondrosamine. See Galactose, 2-amino2-deoxg-r)-. Chromatography, 106, 287, 295 books on, 56 of colorless substances, 57 column. See Column chromatography. paper. See Paper chromatography. reviews on, 56 Chromatopack, 56 Chromatopile, 56 Citric acid, 66 Clays, for column chroma1ogr:tphj , 65 fuller’s earth, for column chromatography, 69 Clover, galacto~nannrtnof, 72 galactomannan of seeds, 90 Coenzyme I (Codehydrogenasc I, Ill”), 245 Column chromatography, adsorbents for, 57, 58 rllert of devcloprr on ev:du:Ltion of, 81 affinity of sugars for rartmn in, 59 alktiline prrni:tng:irrntr iis lirnsh or streak rr:igent, 83 analytical procedures in, 59

398

SUBJECT INDEX

apparatus for, 57, 58, 62 with interferometry of effluent, 58 detection of sugar acids, 65 of zones, 64, 65 developers, 65,84,90,92 effect on adsorbent evaluation, 81 rate of flow of, 67 for sugar azoates, 80, 81 supplier for, 62 displacement analysis in, 59, 60 do>\nward, 58 effect of molecular weight, 84 effluent fractionators for, 62 elution analysis in, 59, 60 elution method, 68 extrusion of column, 67 flowing chromatogram, 62 flowing method, 68 for analysis, 87 for fractionation of starch, 79 for separation, of amino sugars, 76 of di- from mono-saccharides, 57 of sugar acids, 76 of sugar alcohols, 76 of sugars, 75 formation of columns for, 55 frontal analysis in, -59 metal tribes for, 67 of aldonic acid amides, 04 of methylated sugars, 88 of penta-0-acetyl-D-gluconic acid and derivatives, 84,86 of sugar acetates, 83 of sugar acids, 56 of sugar alcohols, 56 of sugar p-phenylazot)erizoates, 79 of sugars, 55, 56 of uronic acids and polymers, 04 on alumina, 79, 80, 81, 80, 0 2 , 94 on calcium acid silicate, 77, 94 on calcium carbonate, 79 on carbohydrates, 70 on carbon, 64 on cellulose poFvder, 70, 74, 90 on cotton, 70 on fibrous alumina, 90 on fuller’s earth clny, 69 on hydrorellrilosc, 72 nn ion exchange resins, 75 on magnesium acid silicate, i 0 on magnesium silicate, 91

on Magnesol, 80, 87, 93 for sugar acetates, 86 on Silene EF, 88 on silica, 79, 89 on silica gel, 79, 82 on silicates, 78 on silica-water, 89, 90 on silicic acid, 79, 80, 86, 93 on starch, 74, 75 on sucrose, 70 solvent developers for, 74 tapered glass tubes for, 67 t.he term, 55 upward, 57 use of carbon (Darco G-60), 58, 62 (Kahlbaum, active), 57,58 (Merck, animal), 58 of Carboraffin, 58 of Carboraffin Supra, 58 of Celite No. 535, 64 of charcoal (Merck, blood), 58 of Congo Red, 65 of Florex XXX, 65 of Floridin XXX, 65 of fuller’s eart,h clay, 64 of heated column, 72 of Norit 1’-3, 58 of series of columns, 61, F2 of streak reagents, 65 of sugar azoates, 79 of Supranorit, 58 zone location in, by brush or streak method. 83 Complexes, of ammonia with a-ogalactosylamine, 131 of amylose with cellulose, 70 of lioric acid with glycosylamines, 125 of carbon tetrachloride with o-glucosylaniine derivative, 111 of sodium sulfate with glycosylamines, 124 Conductivity, electric, of solutions of sugar-horat,e rompleses, 231 Conformations, determination of, 44 Congo Red, use in r.oliimn rhromatography, 65 Conifers, hemicelliiloses of, 313 Corn (mtize) cnl)s, acid hydrolyznte of hemicellulose-B of, 64 sylan from, 300

399

SUBJECT INDEX

Cotton, cellulose from, 284 linters, 292, 329, 331 C,’rot,onoside.S e e Isoguaiiosine. Ciipraiiimonium soliitions, for deterniination of conformations, 43, 44 Currants, Mack, 93 Cyanin chloride, 240 Cyanohydrin reaction, 25 1,2-CycIoheptanediol, 21 Cylohexane, stereochemistry of t.he ring, 14 CQclohexanesulfnnii(. acid, 354 Cyclohexene, 21 oside. See 7-Oxal)icyclo[4.1 .O]heptane. Cyclohexylamine, 354 reaction with o-glucose, 100 Cyclopentane, 12 -, cis-l,2-dimethgl-, 12 trans isomer, 12 -, cis-1,3-dimethyl-, 12 trans isomer, 12 Cyclopentanone, 13 Cysteine, 360 Cystine, N,N’-di-(u-glucosylgusnyl) -I,-, 136 Cytidine, synthesis of, 243

D Dactylis glonaerata, 91 Decalin. See Kapht,halene, decahgdro-. Delignification, 287,318,320,321,324,326 Cross-and-Bevan method, 320 soda process for, 325 sulfite process for, 322 of wood, 316 Dextran, 89,362 partially degraded, 362 sulfate, 361, 362 for clinical use, 362 Dext,rin, corn-starch limit, fractionation of, 61 Schardinger a-,66 acid hydrolyzate of, 63 Schardinger (3-, 66 Diarabinosyl bromide, acetate, 254 Dibenz ylamine, 200 reaction with D-glucose, 177, 178 Dibutylamine, 177, 239 Dicalite, 81 Dicellobiosylamine, 129

I)iet,h:tnol:tmine. See Ethanol, 2,2’iminodi-. Diethylamine, 230 Uiethylene glycol, 66 Digitalose, u-. See Galactose, 6-deoxy3-0-methyl-u-. Di-u-glucosylamine, 121 nonaacetate, 136 “a” form, 121, 136 octaacetate, 121, 136 “(3” form, dihydrate, 121, 136 octaacetate, 136 ortaacetnte, 121, 136 Diglycosylamines, 106,120 p-Dioxane, 15 1,3-Dioxolane ring, 13 Dipentnerythritol, 66 Dipropylene glycol, 67 Disaccharides, 1 4 6 linked, 224 reducing, conversion of acetates t o acetylated ethyl glycosides, 219 poly-0-acetyl halides of, 222 synthesis of, 228 Douglas fir, 292, 319, 320 holocellulose from, 312 Dowes-I, 75 Dowex-50, 76 D . P., the term, 284 DPN. See Coenzyme I . Dulcitol, 66,77 1,3:4,6-diacetals of, 40 -, l16-di-O-benzoyl-2,3,4,5-di-O-l)enzylidene-, 27 -, 2,3:4,5-di-O-isopropylidene-, 27 -, 1,3:4,6-di-O-methylene-, 29

E Enneaheptitol, anhydro-, 66 Ephedrine, relative affinity for carbon, 59 Epicellobiosyl bromide, hepta-0-acetyla-,254 Epicellobiosyl chloride, hepta-0-acetyla-,249 Epicellobiosyl fluoride, hepta-0-acetyl-, 246 Epicellobiosyl iodide, hepta-0-acetyla - , 256 Epirhamnosylamine. See Glucosylamine, 6-deosy-. Erythritol, 66, 87

400

SUBJECT INDEX

Fructose, I ) - , 66, 77, 175 action of hydrazine on, 187 action of selenioris acid on, I87 himolecular dianhydrides o f , 63 CL4-lahelcd,79 conversion to 11-glucose,67, 87 1,6-diphosphate, 73 oxidation of, by cupric salts, 187 by 2,6-dichlorophenolindophenol, 187 by o-dinitrobensene, 187 partially methylnted, separation of isomers, 90 pentaacetate, of keto-u-, 88 pentaazoate, 79 p anomer, 80 preparation from sucrose, 125 F reaction with p-phenetidine, 98 Fenugreek, galactomannan of seeds of, -, 1-alkylamino-1-deoxy-D-, 203 72,90 -, 1-amino-1-deoxy-n-, 173 Ferric chloride, 219 -, 1-anilino-1-deoxy-o-, 176, 204, 205 Fibers, for textiles, 323 -, 1-(p-anisidino)-l-deoxy-D-, 203, 205 Fibrin, 359 -, 1-aralkylamino-1-deoxy-D-, 203 Fibrinogen, 359, 362 -, 1-arylamino-1-deoxy-D-, 203 Fir, Douglas. See Douglas fir. catalytic oxidation of, 200 Flavone, 3,3‘,4‘, 5,7-pentahydrosy-. See -, 1-benzylamino-l-deoxy-D-,203 Quercetin. 4,B-O-benzylidene acetal, 203 Flavonoids, chromatographic separnbioxalate, 203 tion of, 93 -, 1-chloro-1-deoxy-keto-n-,tetraaceFlax, mucilage from seed of, 72 tate, 200 pulp, 331 -, 6-deoxy-~-,73 Florex XXX, 65, 67, 78 -, 1-deoxy-1-dibenzylamino-u-, 105, 203 Floridin XXX, 94 triacetate, 196 use in column chromatography, 65 -, 1-deoxy-1-diethanolamino-u-, 204 Folic acid, synthesis of, 172 -, l-deoxy-l-(3,4-dimethylanilino) -D-, Formaldehyde, 305 203,205 2,4-dinitrophenylhydrazone, 118 -, 1-deoxy-1-glycino-u-,ethyl ester, 204 Formamide, 361 -, 1-deoxy-1 -(p-hydroxyani1ino)-D-, 203 -, N,N-dimethyl-, 361 pentaacetate, 195, 203 Frangulin, gluco-, oct>aacetate,94 -, 1-deoxy -1-morpholino-n-, 203 Fraxinus excelsior, 286 periodate oxidation of, 196 Frog spawn, mucin of, 72, 73 phenylhydraxone, 197, 203 Frostbite, treatment of, 360 -, l-deoxy-l-(2-naphthylamino) - D - , 203 Fructans, 91, 92 -, 1-deoxy-l-IN,N - (3-oxapentamethylof leafy cocksfoot grass, 91,92 ene)amino]-D-, 173 of rye grass, 91, 92 -, 1-deoxy-1-(N,N-pentamethyleneFructofuranose-D-fructopyranose, D-, amino)-D-, 173 1,2’:2,1’-dianhydride, 67 -, 1-deoxy-1- (p-phenetidino)-D-, 203, Fructopyranose, 2 , 3 - 0 - b e n z ~ l i d e n e - ~ - , 205 66 -, 1-deoxy-1-phenylalanino-u-, 189, 193

Erythronic acid, D-, 312 Erythrose, 11-, 73 L - , 1-diacetamide derivative, 121, 123, 120 t,riace t.:tt,e, 12!) -, tri-O-acct.yl-r,-, 121 Esparto grass, xylan of, 299 1,2-Ethanediol, 1,2-diphenyl-erythro-, 13 -, 1,2-diphenyl-threo-, 13,27 Ethanol, 2-amino-, 185, 311, 316, 317, 318 Ethanol, 2,2’-iminodi-, 185 P:t,hanolamine. See Ethanol, 2-amino-. Et,hylamine, 2-phenylL. See I’henet,hylamine. Ethylene glycol, 67 Ethylene oxide, 24

-

SUBJECT INDEX

-,

40 1

stability of, 102 l-deo~y-l-(i)L-phen).lalariillo)-i)-,204 1,3,4,5-tetraacetate, 113, 130 -, 1-deoxy-l-(2-phenylethylamino) -D-, 1,3,4,5-tetrahenxoate, 115, 130 204 -, 1-deoxy-1-piperidino-o-, 177,194,203, -, N - (p-sulfophenyl) -o-, 130 -, N-(p-tolyl)-D-, 99, 130, 193 205 stability of, 102 action of heat on, 194 1,3,4,5-tetraacetate, 113, 130 fission of, 195 1,3,4,5-tetrabenzoate, 115, 130 hydrogenation products from, 192 4,5-0-isopropylidene acetal, hydro- Fructosyl bromide, I ,3,4,5-tetrit-Oacetyl-p-D-, 251 chloride, 196 -, 1,3,4,5-tetra-O-benzoyl-p-~-, 221, osime, 197,203 251 hydrochloride, 197, 203 reduction of, with lithium aluminum periodate oxidation of, 190 hydride, 245 phenylhydrazone, 197, 203 1,4,5-tri-O-acety1-3-O-mes~.l-p-i~-, -, 1-deoxy-l-(o-toluidino)-D-, 176, 203 -, 251 -, 1-deoxy-l- (p-toluidino)-o-, 173, 185, Fructosyl chloride, 1,3,4,5-tetra-O189, 191, 193, 198, 203, 205 acetyl-D-, 113 oxime, 196, 197, 203 p anomer, 216, 221, 222, 247 hydrochloride, 196, 197 -, 1,3,4,6-tetra-O-acetyI-~-, 247 preparation of, 175 Fructosyl fluoride, 1,3,-2,5-tetra-Otetrahenxoate, 195, 203 acetyl-p-w, 246 ultraviolet absorption spectrum of, 186 212, 246 -, 0-diacetone-D-. See Fructose, 2,3:4,5- -, 3,4,5-tri-O-acetyl-p-u-, -, 1,4,5-tri-O-acetyl-3-0-rnethyl-p-~-, di-0-isopropylidene-o-. 2-26 -, 2,3:1,5-di-O-benzylidene-~-, 33 -, 2, 3:4,5-di-O-isopropylidene-o-,33, -, 3,4,5-tri-O-acet~~l-l-O-methyl-p-~-, 246 68 Fructosyl halides, poly-0-acyl-, 208 -, 3,4-di-O-methyl-~-,92 Fucitol, L-, 66, 77 -, 2,3-O-isopropylidene-~-, 67 Fuconic acid, L-, amide, 77 -, penta-0-acetyl-p-w, 212 -, 2,3-di-O-methyl-~-,amide, 269 -, penta-0-acetyl-keto-o-, 216 lactone, 269 -, 1,3,4,5-tetra-O-acetyl-o-, 113, 216, -, 3-O-methyl-~-,lactone, 268 22 1 -, 3-O-methyl-~-,amide, 269 -, I ,3,4,6-tetra-O-acetyI-~-, 241 lactone, 269 -, 1,3,4,5-tetra-O-acetyl-6-chloro-6de-, 2,3,4-tri-O-methyl-r,-, aniide, 269 O X Y - k e t O - D - , 217 lactone, 269 -, 1,3,4,5-tetra-O-benzogI-, 221 Fucopyranosidc, methyl 2,3-di-O-, 1,3,4,6-tetra-0-benzo3.1-D-, 115 methyl-a-L-, 269 -, 1,3,3,6-tetra O-methyI-i)-, 91 -, methyl 3-0-methyl-a-i)-, 268 -, 1,3,4-tri-O-methyl-~-, 92 p anomer, 268 -, 3,4,6-tri-O-methyl-o-, 92 Fructoside, methyl 1,3,4,5-tetra-O- -, methyl S-O-methyl-a-~-,269 Fucose, I>-, 64, 66,72, 73,77 acetyl-a-D-, 222 -, 2,3-di-O-methyl-~-,269 Fructosylamine, D - , 175 -, 3,4-di-O-methyl-~-,269 -, N - ( p-ethoxyphenyl)-I>-, 130 -, 2-O-methyl-~-,268 -, N-(p-hydroxypheny1)-D-, 99, 130 -, 3-O-methyi-o-, 268 -, N-(p-methoxypheny1)-D-, 99 phenylosazone, 268 -, N-phenyl-D-, 99, 124, 130 -, S-O-methyl-~-,269 preparation of, 97 phenylosaxone, 269 reaction with hydrogen cyanide, 102

402 -, tetra-0-acetyl-a-L-,88 -, 2,3,4-tri-O-methyl-~-,268

SUBJECT INDEX

G

Galactan, 284 hydrate, 268 of larch, 72 -, 2,3,4-tri-O-methyl-~-,269 of wood, 326 hydrate, 269 Galactaric acid, 313 Fucoside, methyl 3,4-di-O-rnethyl-a-~-, -, 2-O-methyl-, diamide, 273 -, 2,3,4-tri-O-methyl-, 277 269 - , methyl 3,4-0-isopropylidene-2-0- dimethyl ester, 277 Galactitol, 76, 174 methyl-I)-, 268 -, methyl 3,4-0-isopropylidene-2-0- -, I-amino-1-deoxy-u-, 205 -, 1,4-anhydro-~-,245 methyl-O-L-, 269 27 -, methyl 2,3,4-tri-O-methyl-8-~-, 268 -, 1,2:4,5-di-O-isopropylidene-~~-, -, methyl 2,3,4-tri-O-methyl-a-~-, 269 Galactofuranose, 1,2-O-isopropylideneD - , 33 p anomer, 269 Galactofuranoside, methyl 6-0-methylFucosylamine, A-n-butyl-L-, 130 @-I-, 274 -, N-n-heptyl-L-, 130 Galactomannans, 72, 90 -, A'-n-hexyl-L-, 130 Galactonic acid, D - , amide, 77 -, N-methyl-L-, 130 -, 2,4-di-O-methyl-~-,amide, 275 -, N-1-pentyl-L-, 130 lactone, 275 -, N-phenyl-D-, 2,3,4-trimethyl ether, phenylhydrazide, 275 130,268 -, 2,6-di-O-methyl-~-,276 -, N-phenyl-L-, 130 amide, 276 lactone, 276 2,3,4-trimethjd ether, 130, 269 phenylhydrazide, 276 -, N-n-propyl-I,-, 130 Fuller's earth clay, adsorption of carbo- -, 3,4-di-O-rnethy-o-,amide, 276 lactone, 276 hydrates on, 66 -, 4,6-di-O-niethyl-1-, amide, hydrate, use in column chromatography, 64 276 2-Furaldehyde, compounds resembling, lactone, 276 305 -, 2-O-methyl-u-, amide, 273 conversion tables, for phloroglucinol lactone, 273 compound, 305 -, B-O-methyl-~-,274 determination of, 305 lactone, 274 colorimetric methods, 306 phenylhydrazine salt, 274 gravimetric methods, 305 -, 2,3,4,6-tetra-O-methyl-~-, 280 reagents for, 305 amide, 280 volumetric methods, 305 lactone, 280 from oxycellulose, 304 phenylhydrazide, 280 from pentosans, 303 -, 2,3,5,6-tetra-o-methyl-~-, amide, 280 from nronic acids, 304 lactone, 280 -, 5-hydroxymethyl-, 180, 192, 193, 305, -, 2,3,4-tri-O-methyl-~-, 277 306 amide, 277 -, methyl-, 305,306 lactone, 277 Furan, 2-hydroxyacetyl-, 191 phenylhydruzide, 277 -, tetrahydro-, 13, 24 -, 2,3,5-tri-O-methyl-~-,amide, 278 Furfuraldehyde. See 2-Furaldehyde. lactone, 278 Puroic acid, 306 phenylhydrazide, 278

SUBJECT INDEX

403

-, 1,6-anhydr0-2,3,4-tri-O-meth3.l-p-i)-, amide, 278 277 Iactone, 278 -, I , 2 : 3 , 4 - d i - O - i s o ~ ~ r o ~ ~ y l i d e32, n e 33 -~-, phenylhydrazitle, 278 -, 2,3-di-O-met,hyl-p-i)-, 274 -, 2,4,6-t,ri-O-met,hyl-~-,amide, 278 -, 2,4-di-O-methyl-ou-u-,275 lactone, 278 nionohydrate, 275 -, 3,4,6-tri-O-methyl-u-, Iactone, 279 -, 2,6-di-O-methyl-p-~-,275 Galactopyrunose, 1,6-anhydro-&~-,243 monohydrate, 276 322 Galactopyranoside, methyl @-I-, -, 3,4-di-O-methyl-P-i~-,276 -, methyl 2,6-di-O-rnesyl-a-~-,25 -, 4,6-di -0-methyl-01- I) -,276 -, methyl 2,3-di-O-methyl-a-~-,274 hydrate, 276 4,6-O-benzylidene acetal, 275 phenylosazone, 276 p anomer, 275 -, 1,2:3,4-di-O-methylene-n-, 33 -, methyl 2,6-di-O-methyl-P-~-,276 -, hept,a-O-acetyl-alde~i~(lv-i~-, 86 3,4-O-isopropylidene acetal, 276 -, 2-O-methyl-p-~-,273 01 anomer, 276 -, 3-O-methyl-o-, 72 -, methyl 2-O-methyl-01-~-, 273 01 anomer, 274 4,6-0-benzylidene acetal, 273 phenylosaeone, 274 3-tosylate, 273 -, 4-O-methyl-P-o-, 274 3,4-0-isopropylidene acetal, 273 phenylosazone, 274 6-tosylate, 273 -, 6-0-methyl-cr-o-, 274 -, methyl 2-O-methyl-p-o-, 273 phenylhydrazone, 274 4,6-0-benzylidene acetal, 273 phenylosazone, 274 3-tosylate, 273 -, penta-0-acetyl-D-, a! and P anomers, 3,4-O-isopropylidene acetal, 273 230 -, methyl 3-O-methyl-p-r)-, 274 -, 1,2,3,4,6-penta-O-acetyl-u-, 21 1 4,6-dibenzyl ether, 274 -, penta-0-acetyl-aldehydo-v-, 124 a anomer, 274 -, 2,3,4,6-tetra-o-methyl-v-, 91 Galac topyranosylamine, 2,6-di-001 anomer, 279 methyl-N-phenyl-D-, 276 @ anomer, 279 -, 2-0-methyl-N-phenyl-o-, 273 -, 2,3,5,6-tetra-O-methyl-~-, 280 Galactopyranosyl chloride, W D - , 247 -, 2,3,4-tri-O-acetyl-l,6-anhydro-D-o-, -, 6-0-(~-z-rhamnopyranosyl)-cr-o-, 228 hexaacetate, 249 -, 1,2,4-tri-O-acety1-6-deoxy-3-0Galactosamine, I>-, See Galactose, 2methyl-D-, a! and p anomers, 92, 93 amino-2-deosy-~-. -, 2,3,4-tri-O-methyl-1-, 91 Galactose, v-, 64, 66, 77, 174, 318, 322 01 anomer, 277 I-diacetamide derivative, 124, 131 monohydrate, 277 pentaacetate, 131 -, 2,3,4-tri-O-methy1-6-O-trityl-r-, 277 1-dibenzamide derivative, 131 1-acetate, 277 methyl ethers of. See tables on pages -, 2,3,5-tri-O-methyl-~-,278 679 to 680. -, 2,3, g-tri-o-methyl-~-, 278 pentaazoate, 01 and @ anomers, 80 -, 2,4,6-tri-O-methyl-o-~-, 278 -, 2-acetarnido-2-deoxy-o-, 367 -, 3,4,6-tri-O-methyl-~-,279 1,2-0-isopropylidene acetal, 279 -, 2-amino-2-deoxy-o-, 72, 73, 350, 368 Galactoseen-l,2, 2,3,4,6-tetra-Ohydrochloride, 66, 76 acetyl-D-, 229 -, 1,6-anhydro-B-~-,51 -, 1,6-anhydr0-3,4-0-isopropylidene-2-Calactoseptanosyl chloride, 2,3,4,5tetra-0-acetyl-P-D-, 247 0-methyl-D-, 273 -, 2,3,6-tri-O-methyl-o-, 278

403-

SUBJECT INDEX

Galactoside, methyl 3,G-anhydro-2-0- -, N-o -carbomet hoxyphen yl -D -, 131 iiieyyl-a-u-, 25 -, N-(.l-carboxy-3-hydroxy~lie1i~~l)-u-, -, methyl 4,G ~ - b e l l Z ~ ~ i d e l l e - a -43, D -44 , 131 B anomer, 43, 44 -, N-o-cait~oxyphenyl-~-, 98, 131 -, methyl 2 , 4-di -0-methyl -01 -u -, 275 -, N-p-carhoxyphenyl-D-,131 anomer, 275 -, 2 deoxy-N-phenyl-u-, 132 -, methyl 3,4-di-O-iiiethyl-p-u-,276 -, 2-deoxy-N-p-tolyl-u-, l O G , 134 -, methyl 4,6-di-O-methyl-p-~-,276 -, N , N-diacetyI-2,3,4,5,6-penta-O2,3-dibenzyl ether, 276 acetyl-1,-, 131 2,3-ditosylate, 276 -, N - (3,4-dimethyl-2-nitrophenyl) -I)-, -, methyl tetra-0-methyl-D-, 89 tetraacetate, 131 -, methyl 2,3,4,G-tet rn-0-met hyl-a-r)-, -, N - (4,5-dimethyl-2-nitiophenyl) -w, 280 tetraacetate, 131 nnomer, 280 -, 2,3-di-O-methyl-N-l,henyl-a-, 274 -, met hy1 2,3,5,G-t etra-0-methyl-w, p anomer, 133 280 -, 2,4-di -0-methyl-N-yhenyl-u-, 275 -, methyl 2,3,4-tri-O-methyl-a-~-,277 p anomer, 133 -, methyl 2,3,5-tri-O-methyl-~-,278 -, 2,6-di -0m e t h y l -N-phenyl-p-u -, 133 -, methyl 2,4,6-tri-O-rnethyl-a-~-, 278 -, 4,G-di-O-methyl-N-phenyl-~-, 276 hydrate, 278 0 anomer, 133 3-tosylate, 278 -, N-p-(p-dodecylaminobiphenylsul-, methyl 2,4,6-tri-O-methyI-B-~-, 278 fone)-D-, 132 hemihydrate, 278 -, N-(N-n-dodecylcarbarnoy1)-o-,132 3-tosylate, 278 -, N-p-ethoxyphenyl-n-, 132 -, methyl 2,3,4-tri-O-methyl-6-0-trityl-, N-p-(p-ethylaminobiphenylsu1fone)D-, 277 D-, 132 Galactosiduronic acid, ~-rhamnose-(2+ -, N-n-heptyl-D-, 132 1) D-, 72 -, N - p - (p-hexylaminobiphenylsulfone) Galactosiduronic acids, di- and tri-, 91 D-, 132 Galactosylamine, D - , 108, 131 -, N-1-(2-hydroxynaphthyl)-N-phenyla - ~ -ammonia , complex, 131 methyl-D-, 132 B-D-, 131 -, N-p-(p-isobutylaminobiphenylsulacetylation of, 111 fone)-D-, 132 tetraacetate, 131 -, N-p-(p-isopentylaminobiphenylsul-, N-acetyl-a-o-, 131 fone)-D-, 132 periodate oxidation of, 118 -, N-(4-methyl-2-nitrophenyl-o-, 132 tetraacetate, 131 -, 2-O-methyl-N-phenyl-B-~-,133 -, N-acetyl-p-D-, 113, 131 -, 4-O-methyl-N-phenyl-~-,274 periodate oxidation of, 118 p anomer, 133 tetraacetate, 131 -, N J - n a p h t h y l - ~ - 132 , -, N-acetyl-2,3,4,6-tetra-O-acetyl-a D-, -, N-o-nitrophenyl-D-, tetraacet ate, 132 111 -, N-p-nitrophenyl-p-D-, 132 p anomer, 111 tetraacetate, 132 deacetylation of, 113 -, N-p-(p-octy1aminobiphenylsulfone)-, N-acetyl-tetra-O-acetyl-N-p-nitroD-, 132 phenyl-pa-, 132 -, N-1-pentyl-D-, 132 -, N-n-butyl-D-, 131 -, N-p-(p-penty1aminobiphenylsulfone)-, N-p-(p-1iutylaminobiphmylsulfone)D - , 132 D-, 131 -, N - p h e n y h - , 113, 124, 134 -, N-carbamoyl-D-, 131 acetylation of, 111

SUBJECT INDEX

405

-, 3 , 4 , 6 - t r i - O - acet y l - ~ - 248 , p anomer, 132, 133 2 trichloroacetate, 248 preparat,ion of, 97 -, 3,4,6-t,ri-O-acet yl-2-chloro-2-deoxyreaction wit.h hydrogen cyanidt., 102 LY-D-, 247 -, N-(4-phenylaeophenVlene)-1~-,132 -, 2,4,6-tri-O-iiieth\~I-3-0-tosyl-a-~-, -, N-n,-propyl-D-, 134 247 -, N-p-(p-propylaminohiphenylsul- Galactosyl phosphate, I > - , 350 fone)-o-, 134 -, 2-amino-2-deoxy-~-,350 -, N-p-sulfamylpheiiyl-D-, 134 Galacturonic acid, 313 monohydrate, 134 D - , 64, 76 -, N-p-sulfopheiiyl-u-, 134 LY anomer, 66, 77, 78 -, 2,3,4,6-tet ra-O-acet).l-N-pheiiyI-Wu-, of aspen-wood hcmicellulose, 286 D - , 13-1 in beech wood, 301 p anomcr, 133 -, 2,3,5-tri-O-acetyl-l -hromo-1-deoxy-, 2,3,4,6-tet,ra-O-mcthyl-~-,135 LY-D-, methyl ester, 251 -, tetra-O-methyl-N-phenyl-p-n-, 133 Gentiobiose, 66, 77, 87, 223 -, 2 , 3 , 4 , 6 - t etra-O-met hyl-N-phengl-n-, LY , azoate, 80 279, 280 -, octa-0-acety-p-, 83 p anomer, 133 synthesis of, 241 -, N-f-tolJTl-D-,134 Gentiobioside, maridelonitrile p-, 240 -, N-p-tolJrl-D-, 98, 134 Gentiobiosylamine, N-p-nitrophenyl-8-, hydrolysis of, 105 135 -, 2 , 3 ,4-t,ri-o-methyl-N-p~iciiyl-o-,277 heptaacetate, 135 p anomer, 133 Centiobiosyl bromide, hc.pta-0-acetylL Y - , 236, 237, 240, 254 6-trityl ether, 133 -, 2,4,6-tri~O-met,hyl-N-phenyl~r)-, 278 -, hexn-O-acetyl-6’-bromo 6’-deoxy-a-, 6 anomer’, 133 254 Galact osyl bromide, 2,3,4,6-t et r a - 0 - Gentiobiosyl chloride, hepln-0-acetylacetyl-ol-D-, 242, 243, 251 a-,249 methitnolysis of, 236 Gentiobiosyl fluoride, 246 reaction of, with quinolime, 242 heptaacetat e , 246 with secondary amines, 239 -, 2‘, 3’, 4‘, 6’-tet ra-O-ilcetyl-2,3,4-triwith t.rimet,hylamine, 111 0-benxoyl-. 246 -, 2,3,5,6-t.et.ra-O-acet.yI-~-~-~ 211, 251 (:entiol)iosyl halidcs, poly-0-acetyl-, -, 2,3,4-t~ri-O-acetyl-6-brorno-6-~lcoxy- 232 a - ~211, , 251 (>entiobiospl iodidc, ~iept:L-O-acctyl-c-, -, 2,3,5-t ri-0-ace1 yl-6-bl.oino-6-ileosy256 @-I-, 251 Glucal, tri-O-acetyl-i)-, 228 -, 3,4,6-t ri-0-ace1 yl-2-bl.oriio-2-tlt:osy- Glucamiiie. Scr Glucitol, 1 arnino-la - ~251 , dcoxy-. -, 2,3,4-t ri-O-:tcet )I-6-0-t osyl -LY-D-, 251 (>Incam, from bntleu, !)1 G:tlartosyl chloride, 2,3,4,6-tetrit-0hiosynthesis of, 367 : i c e t y l - ~ - ~247 -, of I)irch, 327 p iinomer, 247 of pine, 327 -, 2 . 3 , s ,n-tetr:L-O-:~CPtyl-D-,230 in sulfate pulps, 327 p itnoini’r, 234, 247 in sulfite pulps, 323, 327 retlnrtion with lithirim alumininn h y the t e r m , 327 tlridr, 245 of M nods, 2% -, 2 , 3 , 4 - t t ~ i - O - n c ~ t y I - ~22S, - i ~ - 2/17 , Glno:iric arid, D-, 351 6-tosplate, 247 potassium :wid salt, 66, 286, 314, 351 a anomer, 134

406

-,

SUBJECT INDEX

Glucofuranosylamine, N-acetyl-o-, sup2,4:3,5-di-O-met hylene-I)-, 39 epimerization of, 41 posed, 119 Glucoheptonic acid, hexa-0-acetyl-"a"formation of, 42 D- , nitrile, reaction with ammonia, Glucitol, D-, 66, 76, 77 118 acetate, 86 Glucomannan, of Amorphophallus konhexaacetate, 88 1,3:2,4:5,6-triacetals, 39 j a c , 289 Gluconic acid, D-, 66, 314 -, 1-amino-1-deoxy-D-, 104, 205 amide, 77 N-substituted, 104 -, 1,4-anhydro-~-,67 ammonium salt, 66 1,4-lactone, 67, 314 6-tosylate, 9 1,5-lactone, 67 -, 1,5-anhydro-~-,245 sodium salt, 66 -, 3,6-anhydro-~-,66 -, %amino-2-deoxy-~-,351 -, 1-deoxy-o-, 66 -, 2-deoxy-~-,66 -, 2,4 :3,5-di-O-methylene-u-, 39 -, penta-0-acetyl-D-, 84 -, 1-deoxy-1-piperidino-u-,192 nitrile, 123 -, 1-deoxy-1-p-tohidino-D-, 199 -, 2,5-diamino-l,4:3,6-dianhydro-2,5-Glucopyranose, a-D-,120 W D- , crystal structure of, 231 dideoxy-o-, 48 -, 1,4:3,6-dianhydro-~-,9, 46, 67 0-D-, acetate, 86 D-, and derivatives, infrared absorp2,5-ditosylate, 47, 48 tion spectra of, 358 -, 1,5:3,6-dianhydro-2,4-O-methyleneD-, 23 -, 2-amino-2-deoxy-~-, infrared rtbsorption spectra of derivatives of, -, 2,4: 3,5-di-O-benzylidene-l,6-di358 h o m o - l ,B-dideoxy-~-, 42 -, 1,6-dichloro-l,6-dideoxy-2,4:3,5- -, 1,6-anhydro-p-u-, 22, 49, 51, 87, 227 preparation of, 243 di-0-methylene-o-, 42 triacetate, 223 -, 2,4 :3,5-di-O-methyIene-o-, 39 tribenzoate, 228 acetolysis of, 40 trimethyl ether, 228 oxidation of, 42 -, 6-O-(or-D-galactopyranosyl)-p-~-. See 1,6-ditosylate, 42 Melibiose. -, 2,4-0-methylene-~-,3, 29 oc-, 1,3,5-tri-O-azoyl-2,4,6-tri -0-methyl- -, 6-O-(p-D-galactopyranosyl)-p-~-, taacetate, 218 D - , 82 OCta-, 1,4,5-tri -0-azogl-2,3,6-tri-O-methyl--, 3-0-(or-D-glUCopyraKloSy1)-p-D-, acetate, 87 I)-, 82 -, 1 , 3 :2,4:5,6-tri -0-benzylidenc-u-, 2s -, tctra-0-acetyl-a-r-, configuration and structure of, 231 -, 1,3:2,4:5,6-tri-O-ethylidene-~-, 28 -, 1,2,3,6-tetra-O-acety1-8-1,-, 241 Glucofuranose, a - u - , I,a-acetals of, 31 -, tetra-0-methyl-D-, 221 -, 1,6-anhydro-p-~-,22 -, 6-O-henzoyl-l , 2-0-isopropylidene-I-> Glucopyranoside, (substituted) o-hydroxyacetophenone I>-, 240 225 -, 3-hydroxypropyl WD-, 66 -, 1,2-O-henzylidene-a-~-,35 -, methyl cu-u-, 67, 77 -, 1,2-O-isoprop~lidene-3-O-tosyl-r,-, p anomer, 67, 322 225 hydrolysis of, 349 -, 1,2-0-isopropylidrn~~ 6 O-tOS).l-D-, -, methyl 2-amino-2-deoxy-a-~-, 349. 23,225 358 Clurofurnnositlr, methyl 3,G-anhytlrop anomrr, 349, 358 a-u-, 3 -, methyl 3,g-anhydro a-D-,4, 22

SUBJECT INDEX

-,

407

methyl 3,6-suliydio-2-0-tosyl-~-i)-, from u-fructose. 67, 87 25 latbelcd with CP, 79 -, methyl 2,3,6-1ri-O-methyl-u-, 89 monoacetamide derivative, cyclic a arid p anomers, 89 form, 123 p:trtiallq methylated, azoylation of, 81 -, peoni11-3,5 his-(D-, octaacctate, 240 Glucopyranosylaniine, N -phenyl -u-, 97, “penta-0-acctyl-monoacetone” deriv109 ative, 222 pentaazoate, 79 Glucopyranosyl bromide, 6-0-(p-cellobiosyl)-a-D-, decaacetate, 254 a anomer, 80 -, 2,6 -di -0-acet yl-3-0-mesyl-a- D -, 226, p anomer, 80,81 252 phenylosazone, 200 reaction of, with aliphatic amines, 100 -, 6-0- (8-gent iohiosy1)-a-a-, drcawith aniline, 97 acetate, 254 -, 2-O-(P-~-glucopyranosyl)-a-~-, heptawith p-anisidine, 97 with dibeneylamine, 200 acetate, 254 with morpholine, 200 -, 3-O-(p-~-glucopyranosyl)-o-, heptaacetate, 254 with p-phenetidine, 97 with piperidine, 200 -, 6-O-(a-D-glUCOpyranOSyl)-a-D-,heptaacetate, 254 with p-toluidine, 97, 200 separation di-, tetra-, and tri-methyl Glucopyranosyl chloride, 6-O-(p-cellobiosyI)-a-o-, decaacetate, 249 ethers of, 89 -, 6-O-(P-~-glucopyranosyl)-cx-~-, hep3-sulfate, simultaneous acetylation taacetate, 249 and desulfation of, 357 -, 2,3,6-tri-O-methyl-a-, 224 6-sulfate, 354 a anomer, 248 -, 2-acetamido-2-deoxy-n-, 122, 353, Glucopyranosyl fluoride, a - ~ -246 , 368 -, 6-chloro-6-deoxy-a-1,-, 246 diethyl thioacetal, 87 -, 6-O-trityl-a-~-,246 polymer of, 353 triacetate, 246 -, 2,4-di-O-acetyl-1,6-anhydro-3-0tribenzoate, 246 tOSyl-fi-D-, 228 Glucosamine, D-. See Glucose, 2-amino-2- -, 5,6-di-0-acetyl-l,2-O-(l-bromodeoxy-D-. et hylidene)-3-0-mesyl-~-,227 Glucosaminic acid, N-methyl-L-, nitrile, -, 5,6-di-O-acetyl-l,2-0-isopropyli87 dene-3-0-tosyl-~-,226 Glucosan, POIY-D-,225 -, 5,6-di-0-acetyl-3-0-mesyl-~-, 226 -, 2-amino-2-deoxy-~-,72, 73, 122, 173, Glucose, D - , 66, 77, 173, 314, 322 a,p , “a,” and (‘7’’ forms, 229 175, 349, 368 acid reversion products from, 87 action of nitrous acid on derivatives of, action of ammonia on, 73 357 anilide. See Glucosylamine, N-phenylbasicity of, 354 D-. 1,6-diphosphate, 368 condensation with aniline, 102 electrostatic shielding, in derivatives condensation with aromatic amines, of, 349 170,175 hydrochloride, 66, 76, 351 configuration of a-D-,231 incorporation into mucopolysaccha1-dibenzamide derivative, 121, 123, rides, 367 136 pentaacetate, WD-, 87 pentaacetate, 136 0 anomer, 87 1-di-(cyclohexylamine) derivative, 136 6-phosphate, 368 1,6-diphosphate, 368 N-sulfate, 354

408

SUBJECT INDEX

-, 5 , ~ - : ~ 1 i h y d r o - 1 , 2 - ~ - i s o ~ ~ r o ~ ~ y-,i i ~ peilta-O-be~izoyl-alrleh!/do-u-, c11~-~action 8 of met hanolic itinnionia on, 121, 124 3,6-anhydro-l,2-~-isopropylidene--, 1,2,3,4-tetra-O-acetyl-p-~-, 241 5-0-tosyl-m-~-,8, 48 -, 2,3,4,6-tet ra-0-acet y1-n-, compound -, 3,5-O-heiizylidene-l,2-O-isopropyl-- with benzylamine, 112 idene-a-D-, 36 prepamt.ion of, 113 -, 6-deoxy-l , 2:3,5-di-O-isopropylidenc- -, 1,3,4,6-tetra-O-acctyl-2-amino-2-~~6-nitro -a-D -, 36 OXY-D-, hydrochloride, 122 -, 5,B-di-O-benzoy-l , 2-0-isopropyl- -, 2,3,4,6-tetra-O-benzo,1~l-n-, 115 idene-3-O-tosyl-o-, 226 -, 2,3,4,6-tet ra-O-met~hyl-o-, 88, 92, -, 1,2 :3,5-di-O-isopropylidene-ff~n - , 36 109 graphic reprcsent,at,ion of, 4 -, 2,3,4-t,ri-O-acetyI-l,G-anhydro-p-~-, -, 1,2:5,6-di-O-isoprop~Iideiie-n-, 225 227,228 3-benzoate, 225 -, 3,4,6-t~ri-O-acetyl-l,2-nnhydro-~-n-, 3-mesylate, 226 241,243 3-tosylate, 225 , 1,2,4-tri-O-acetyl-6-deoxy-3-O-, 2,3-di-O-methyl-~-,88 m e t h y h - , OL and 6 anomers, 92 -, 1,2:3,5-di-O-methylene-a-~-, 36 -, 3,5,6-tri-O-acetyl-l, 2-0-isopropyli-, O - ( D - f r U ctO S J'l) - D -, 64 dene-o-, 226 -, 6-O-(a-D-galactopyranosyl)-~-. See -, tri-O-methyl-D-, isomers of, 92 Melibiose. -, 2,3,4-t,ri-O-met,hyl-i,-, 91 -, 6-O-(ay-o-glucopyranosyl)-p-~-.See -, 2 , 3 , 6-tri-0-methyl-D-,88 Isomaltose. Glucose-l-C14, I)-, 368 -, .l-O-(or-isornaltopyranosyl)-o-. See Glucose-6-Cl4, D-, 367 Panose. Glucoseen-l,2, 2,3,4,6-tetra-O-acelyl-, 1,2-O-isopropylidene-5, 6-d-O-t,osylD - , 229 ff-D-, 8 Glucoside, aniline D - . See Glucosyl-, 1,2-0-isopropylidene-3,5,6-tri-O- itmine, N-phenyi-o-. toSyl-D-, 226 -, aniline N-D-.See Glucosylamine, N -, 4-0 - (or-maltopyranosyl) -D-. See Malt ophenyl-D-. triose. -, benzyl 2,3,4,6-tjetra-0-acetyl-p-u-, -, 2-0-1nethy1-~-,112 242 -, penta-O-acet,yl-D-, 216 -, hesperidin, 94 01 and p anomers, 229 -, methyl W D - , 229 action of ammonia on, 121 anomer, 229 -, 1,2,3,4,6-penta-O-acetyl-a-~-, 21 1, con6gurat)ion of, 231 212,219,223 -, methyl 2-amino-2-deosy-N-sulfop anomer, 88, 211, 217 3,4,6-tri-O-sulfo-p-~-, diharium salt, conversion t o 01 anomer, 219 354 react.ion wit>h aluminiim chloride, -, methyl 4,6-0-benzylidene-p-o-, 37 221 -, methyl 4,6-0-benzylidene-2,3-di-Oanomers, reaction of, with benzyltoSyl-a-D-, 93 amine, 112 -, methyl 4,6-0-benzylidene-2-O-tosylwith piperidine, 112 a - D - , 93 with titanium tetrachloride, 219 -, methyl 4,6-0-benzylidene-3-0-tosyl-, penta-O-acetyl-aldeh?/do-D-, action a - n - , 93 of ammonia on, 118 -, methyl 3,4-di-O-acety-2,6-di-OtoSyl-p-D-, 25 -, 1,2,3,4,6-penta-O-berizoyl-o-, 01 and p anomers, action of ammonia on, -, methyl 2,3,4,6-tetrn-O - s c e t g l - p - ~ - , 123 222,242 D-,

,

-

SUBJECT I N D E X

-,

me thy1 2,3,4,6-tet rit -0-mct hyl -u - , 89 inethyl 2,3,4-t,ri-0-acetyl-6-O-benzoyl-b-n-, 225 -, methyl 2,3,4-t ri-O-acet.yl-6-O-tosy1@I>-, 225 -, naringin, 94 -, rutin, 94 Glucosidurouic acid, 1-l)romo-l-drosy01-u-galactopyr~nose-(6+ 1) 0-D-, hesaacetat,e, methyl ester, 254 -, 1-bromo-1-deoxy-ol-o-glucoDyranosc(4 + 1) b-u-, hesascetate, met.hgl ester, 254 -, u-xylose-(P + 1) 4-0-mct,hyl-u-, 64 01 auomer, 303 Glucosone, D-, 187, 188, 198, 368 imino analog, 187 phenylosazone, 187,188 tetrahydroxybutylquitioxali~iederivative, 187, 188 G l u c ~ s o n e - l - CD ~ -~, ,368 Glucosylamine, I>-, 96, 136, 137 acet.ylation of, 121 N-acet,ylation of, 117 hydrolysis of, 105 reaction of, with ethyl m:tlonat.e, 201 with 2,4-pent,anedione, 201 N-suhstituted derivatives of, 171, 184 Schiff-base structure of, 187 -, N-p-acetox?iphenyl-D-, tet,raacetate, 137 -, N-itcctyl-D-, 123, 137, 179 furanose form, supposed, 137 oxidation of, with lead tetraacetate, 118 with pcriodate, 117 prcparation of, 117 second isomer of, 118 t.etraacet.ate, 117, 137 -, N - acctyl - N - p - bromophenyl-i)-, 2 , 3 , 4 , 6 - t d r a a ~ e t a t ~111, e , 137 -, N - (2 -amino - 3,4 - dimethylpheny1)D - , t e t r u c e t a t e , 137 -, N - (2 -amino - 4,5 - dimet,hylphenyl)I ) . , tetraacetate, 137 -, N - (2 - amino - 4 - methylphenyl)-o-, t,etraacetate, 137 -, N - (o-nniinophenyl)-D-, 2,3,4, 6-tetra+cetat.e, 137

-,

40'3

-, N - (2 - I)euzisothiazoliii 3-0nc-1,l-dioxide) -D-,2,3,4,6-teti aacetat e, 137

-, N-henzyl-o-, 137 tetraacetate, 112,137 preparation of, 112 -, N-bcnzylidmc I)-, tetraacetate, 112, 137 -, D - (N-benzyl - N-met h yl) -1) -,t et raacr tntc, 137 DL-, 137 I.-, 137 -, N-4-biphenyl-1,-, 138 -, N-p-hromophen?ll-ol-u-, 2,3,4,6-te t raacetate, 138 p anomer, 2,3,4,6-tetraacetate, 138 2,3,4,6-tetramethyl ether, 138 -, N-n-bUtJrl-D-,138 hydrolysis of, 105 -, N-carbamoyl-u-, 138 N-acetyl-tetra-0-acetyl derivative, 138 N'- benzoyl - tetra - 0 - acetyl derivative, 138 tetraacetate, 138 tetrabenzoate, 138 -, N - (4 - carbethoxy - 3 - hydroxyphenyl)-D-, 138 -, Wo-carbet hosyphenyl-D-, 138 -, N-p-carbethoxyphenyl-u-, 138, 139 tetraacetate, 139 -, W ( 4 - carbomethosy - 3 - hydroxyphenyl)-o-, 139 -, N-o-carbomethoxyphenyl-o-,139 tetraacetate, 139 -, N-p-carbomethoxyphetiyl-o-, 139 tetraacetate, 139 -, N-(4-carboxy-3-hyd1oxyphenyl)-n-, 139 hemihydrate, 139 monohydiate, 139 pentaacetate, 139 phenyl ester, 139 sodium salt, 139 tuherculostatic activity of, 125 2,3,4,6-tet raaret ate, 139 -, N-m-carhoxypheny~-~-, 139, 140 tetraacetate,l40 -, N-o-carboxyplienyl-u-, 98, 139 N-acetyl derivative, sodium salt, 139 tetraacetate, 139 tetraacetate, 139

410

-, N-p-carboxyphenyl-u-, 1-40

SUBJECT INDEX

-, N - 2 - (1 - ethoxyethylideneamino)butyl ester, 138 phenyl-D-, 2,3,4 ,g-tetraacetate, 142 dihydrate, 138 -, N-o-ethoxyphenyl-D-, 142 tetraacetate, 138 -, N-p-ethoxyphenyl-o-, 142 2-diethylaminoethyl ester, monohytetraacetate, 142 drochloride, 140 -, N-ethyl-u-, 142 monohydrate, monohydrochloride, -, N - ( N - ethylthiocarbamoyl) - D - , 140 tetraacetate, 142 tetraacetate, 140 -, N-1-heptyl-D-, 142 -, N-o-carboxyphenylsulfonyl-u-, 140 -, N-1-hexadecyl-D-, 142 sodium salt, 140 -, N-1-hexyl-D-, 142 -, N-2-chloroethgloxycarbonyl-~-,tethydrolysis of, 105 raacetate, 140 -, N-(2-hydroxy-3,5-dinitrophenyl) -u-, -, N-(4-chloro-2-nitrophenyl) -D-, 140 142 tetraacetate, forms I and 11, 140 4,6 - 0 - ethylidene - 2 , 3 - oxidoethyli-, N - n f-chlorophenyl-o-, 140 dene metal, 142 -, N-o-chlorophenyl-u-, 140 -, N-2-hydroxgethyl-1-, 142, 201 -, N-p-chlorophenyl-o-, 141 -, N-(2-hydroxy-5-nitrobenzylidene) -D2,3,4 ,6-tetraacetate, 141 142 2,3,4,6-tet ramethyl ether, 141 -, N-o-hydroxyphenyl-o-, 143 -, N-n-decyl-D-, 141 -, AT-p-hydroxyphenyl-D-, 143 hydrolysis of, 105 pentaacetate, 143 -, 2-deosy-N-phenyl-o-, 145, 146 t etraacetate, 143 infrared absorption spectrum of, 102 -, N-(3 - hpdroxy - 4 - propoxycarbonyl3,5,6-trimethyl ether, 146 phenyl)-o-, 143 -, 2-deoxy-N-p-tolyl-o-, 148 -, N-(4-iodo-2-nitrophenyl)-~-, 143 -, N , N-dibenzyl-D-, supposed, 104 tetraacetate, 143 -, N , N-diethyl-D-, 141 -, N - o -methox ypyen yl -D -, 143 -, N , N-di-(2-hydroxyethyl)- D - , 141 -, N-p-methoxyphenyl-D-, 143 hexaacetate, 141 2,3,4, &tetraacetate, 143 -, N,N-dimethyl-D-, 141 2,3,4,6-tetrarnethyl ether, 143 2,3,6-trimethyl ether, 141 -, N-methyl-D-, 143 hydrochloride, 141 -, N-(N-methylcarbamoyl)-~-, 143 hydriodide, 141 -, N-(4-methyl-2-nitrophenyl)-~-, 143 -, N-(N,N-dimethylcarbamoyl)-o-,141 tetraacetate, forms I and 11, 143 -, N-(3,4-dimethyl-2-nitrophenyl) -D-, -, N-meth yl -N-phenyl -D -, t e t raacet ate, tetraacetate, 141 104 -, N-(4,5-dimethyl-2-nitroplie1iyl)-~-, 2,3,4, &tetraacetate, 143 99, 141 -, N-methylsulfonyl-o-, tetraacetate, tetraacetate, 141 124, 143 forms I and 11,141 -, N-(N-methylthiocarbamoy1)-D-,t e t -, N-2,3-dimethylphenyl-~-, 141 raacetate, 143 -, N-2,4-dimethylphenyl-o-,141 -, N-[2 - methyl-3-(0 - tolylazo)phenyl]-, N-2,5-dimethylphenyl-~-, 141 D - , 143 -, N-3,4-dimethylphenyl-o-,141 -, N-1-naphthyl-o-, 143 -, N-3,5-dimethylphenyl-~-, 141, 142 -, N-2-naphthyl-D-, 143 -, N-1-dodecyl-D-, 142 infrared absorption spectrum of, 103, -, N-(3-ethoxyethylideneamino-4-meth187 ylphenyl)-D-, 2,3,4,6-tetraacetate, 2,3,4 ,&tetraacetate, 143 142 -, N-rn-nitrophenyl-D-, 125, 143, 144

SUBJECT INDEX

periodate oxidation of, 119 tetraacetate, 144 -, N-o-nitroplieiiyl-u-, 1Y3 tetraacetate, 143 -, N - p nitrophenyl-o-, 125, 141, 187 dihydrate, 144 infrared absorption slmh-uni, 103 3-methyl ether, monohydrate, 144 periodate oxidation of, 119 tetraacetate, 144 N-acetyl derivative, 144 2,3,4-t riacetate, 144 6-trityl ether, 144 6-trityl ether, 116 -, N-(3-nitro-p-tolyl) - D - , 187 -, N-1-octadecyl-u-, 144 -, N - (N-1-octadecylcarbamoyl) -w, 144 -, N-l-OCtyl-D-, 144 hydrolysis of, 105 -, N-1-pentyl-o-, 144 -, N-p-phenetyl-D-, 171 -, N-phenyl-D-, 96, 97, 101, 144, 145, 187 N-acetyl - tetra - 0 - acetyl derivative, 111, 145 2,4-dimethyl ether, 145 3,4-dimethyl ether, 110, 145 hydrolysis of, 105 methylation of, 109 2-methyl ether, 110, 145 3-methyl ether, 145 4-methyl ether, 145 6-methyl ether, 110, 145 preparation of, 97 reaction with hydrogen cyanide, 102 relative stability of, 102 tetraacetate, 145 2,3,4,6-tetraacet ate, a anomer, 111, 145 B anomer, 111,145 complex with carbon tetrachloride, 111

2,3,4,6-tetramethylether, 109,110,145 2,3,4-trimethyl ether, 110,145 2,3,6-trimethyl ether, 145 2,4,6-trimethyl ether, 110, 145 -, N-4-phenylazophenylene-~-, 145 -, N-(N-pheny1carbamoy.I)-a-, 145 -, NS-phenylethyl-~-, monohydrate, 146

41 1

-, N-(Npheriylthiocarbamoyl)-i)-,tet raacetate, 146

-, N-p-sulfacetamidol,henyl-D-, 146 2,3,4,6-tet raacetat e, 146 -, N-p-sulfainylphenyl-D-, 146 hexaacetate, 146 tetraacetate, 146 2,3,4,64etrititcetate, CY anomer, 146 /3 anomer, 146 -, N-p-sulfophenyl-D-, 146 -, tetra-0-acetyl-u-, 121, 1.71 hydrochloride, 137 reaction with aromatic aldehydrs, 112 -, tetra-0-acetyl-N-mesyl-o-, 124, 143 -, N-thiocarbamoyl-D-, 146 N’-benzoyl -tet r:t-0-benzoyl dcrivat i ve, 146 -, N-m-tolyl-u-, 146 hemihydrate, 101 relative stability of, 102 -, N-0-tolyl-D-, 146 hemihydrate, 101 infrared absorption spectrum, 103, 187 relative stability of, 102 -, N-p-tOlyl-D-, 98, 99, 101, 147, 175, 179 N-acetyl-tetra-0-acetyl derivative, 111 LY anomer, 147 tetraacetate, 147 B anomer, 113 acetylation of, 113 N-acety1-2,3,4,6-tetra-O-acetyl derivative, 147 2,3-dimethyl ether, 147 hemihydrate, 147 2-methyl ether, 147 monohydrate, 147 2,3,4,6-tetraacetate, 111, 113, 147 deacetylation of, 113 2,3,4,6-tetrabenzoate, 147 2,3,4,64etramethyl ether, 148 3,4,6-triacetate, 147 hemihydrate, 101, 147 infrared absorption spectrum, 103, 186 methylation of, 110 monohydrate, 101, 147 relative stability of, 102 tetraacetate, hydrogenation of, 04 tetrabenzoate, 115 2,3,4,6-tetramethyl ether, 110 ultraviolet absorption spectrum 186

412

SUBJECT INDEX

-, ~ V - p - t o l ~ l s u l f o I ~ ~ ~148 l-I,-,

reaction o f , with active silver chloride, 21!) tet rxtcet at e , 145 with silver fluoride, 221 6-p-toluenesulfon:~tte,1-18 with secondary ainines, 239 2.3,4-t.riacetat e , 1-18 %-ith t~rin~cthylamine, 114 Glucosylamines, iV-alkyl-w, !IS, 189 rcduction with lithium aluniinuin hy1Lydtogen:ttion of, 104 dricle, 245 -, N-aryl-u-, 180, lS!), 1!J5 from rice starch, 223 hydrogenat,ion of, 104 solvolytic reactions of, 234 hydrolysis of, by sulfuric acid, 105 for synthesis of glycosides, 240 order of stability of, 102 -, 2,3,4,6-tet ra-O-ncet,yl-a-o~-,253 -, !V-benzyl-i)-, 195 Glricosyl bromide, 2-0-ac.etylB-O-t)rn- -, 2,3,4,6-t'etra-O-acetyl-cu-r,-, 253 -, 2,B,4,6-tetr:~-O-l,enzoyl-~u-l,-,253 my1 -3,5-di -0-tosyl- a - i ) - , 252 -, 2,3,4,6-tetra-0-propionyl-cu-u-, 253 -, 4-O-aeetyl-6-deoxg-6-iodo-2,3-di-0-, 2,3,4-tri-O-acetyl-cu-1)-, 228, 252 tosyl-a-u-, 224, 253 6-benzoatej 252 -, 2-0-acetyl - 5,6-di-O-benzoyl - 3 - 0 6-bromo-6-deoxy derivative, 211, 228, tospl-a-u-, 252 252 p anomer, supposed, 226 -, 6-0-acetyl-2,3,4-t,ri~O-l)enzoyl-a-~-, 6-p-bromophenyl ether, 252 6-chloro-6-deoxy derivat,ive, 252 228, 253 6-deoxy-6-fluoro derivative, 252 -, 6 -0-:tcet.yl-2,3,4- t ri -0-l)enzyl-cu - I)., 6-deoxy-6-iodo derivative, 252 253 6-deoxy-6-t,hiocyano derivative, 252 -, 0-acetyl-tri-0-tospl-u-, 226 6-di phen yl p h ~ s p l i a t ~252 e, -, 2-0-acet,yl-3,5,6-tri-0-t osyl-a -D-, 252 6-mesylate, 252 -, 4-0-acet y1-2,3,6-tri-O-t~osyl-a-r-,224, 6-methyl ether, 252 252 6-(2-naphthyl) ether, 252 bis-(2,3,4-tri-O-acet y l - a - ~ - 6,6-car, 6-phenyl ether, 252 bonate, 253 3,4-di-0-acetyl-6-deoxy-6-iodo-2-0- 6-tosylate, 252 -, 2,4,6-tri-O-acetyl-a-~-, 225 toSyl-a-D-, 252 3,4-di-O-acetyl-2,6-di-O-bensoyl-a-3-benzoat,e, 225, 252 3-mesylate, 252 D - , 252 3-tosylate, 252 3,4-di-O-acetyl-2,6-di-0-tosyl-a-~-, -, 3,4,6-tri-O-acetyl-2-amino-2-deosy252 CPD-, hydrobromide, 252 2,4-di-O-acety1-3,6-di-0-(tri-0-acehydrochloride, 252 tyigailoyl) -a-D-,252 tetra-0-acetyl-a-o-, quaternization -, 2,4,6-tri-0-acet,yl-3-0-henzoyl-a-~-, 252 mith pyridine, 245 reaction with ammonia, 121 -, 2,3,4,6-tetra-O-acetyl-~-, 94 naming of anomers of, 230 stable form of, 232 -, 2,3,4, 6-tetjra-O-acetyl-cY-D-, 208, 222, 223, 224,237, 240, 241, 244, 252 configuration of, 231 from levoglucosan, 228 methanolysis of, 236, 242 rate of, 235 preparation of, 214

-,

2,3,4-tri-O-acetyl-6-bromo-6-deoxy-

WD-,

252

-, 2,3,4-tri-o-acetyl-6-deoxy-a-o-, 251 -, 3,4,6-tri-o-acety1-2-deoxyu-, 228 anomer, 251 2 , 5 , 6 - tri - 0 - acet,yl-3-O-mesyl - a I>-, supposed, 227, 252 -, 2,3,6-tri - 0 - acetyl - 4-04osyl - LY D-, 252 -, 2,4,6-tri-O-acetyl-3-O-tosyl-ol-o-, 225, 228 -, 2,5,6-t,ri-O-acety1-3-O-tosyl-, 252 01

-,

SUBJECT INDEX

413

-, 3,4,6-tri-O-acety1-2-O-tosyl-a-u-, 252 Glucosyl isocyanate, tetra-0-acetyl-D-, -, 3,4,6-tri-O-benzop1-2-deo~g-a-i~-, 251 149 Glucosyl chloride, 4-0-acetyl-2,3,6-tri- Glucosyl isothiocyanute, u-, 14‘3 @met hyl-a-D-, 247 tetraacetate, 149 -, 4-0-acetyl-2,3,6-tri-O-tosyl-ol-u-,247 Glucosyl phosphate, u-, 350 -, 5 -0-henzoyl-2,3,6- t r i - f ~ - i n e t h y Dl -- , -, 2-aniino-2-dcoxy-~-,340, 368 G lucosylpyridi ni uni bromide tleriva247 -, 3,4-di-0-acetyl-6 -deoxy -a-D -,248 tives. See un.tler l’yridinium bromide. p anomer, 240 Glucosyl thiocyanute, t,etra-O-scetyl-a-, 3,4-di-0-acetyl-6-deosy-2-0-trichlo- I>-, 121 roacctyl-P-u-, 249 Glucosyltrimethglsmi~io~~iiim halide dc-, tctra-O-acetyl-I,-, 216 rivat>ives. See u n d e r respecfive h m -, 2,3,4,6-tetra-o-acetyl-o-, 215 nionirim halide. DI anomer, 209, 211, 219, 222, 247 Glucuronic acid, I>-, 76, 314 p anomer, 209,211,212,210,221,248 in beech wood, 301 rearrangement of, 234 incorporation of, into mucopolgsac-, 2,3,5,6-tetra-O-acetyl-u-, 247 charides, 367 supposed, 222 2,6-lactone, 66 -, 2,3,4,6-tetra-0-benzoyl-a-~-, 248 -, 2,5-di-O-acetyl-l-hromo-I -deoxy-a-, 2,3,4,6-tet,ra-O-mesyl-a-u-, 248 u-, 6,3-lact,onc, 252 -, 2,3,4,6-tetra-O-iiiethyl-a-u-, 221, 248 -, 2,5 - di- 0 - acetyl-1- chloro -1-deoxy-, 2,3,4,6~t,etra-O-propiorlyl-a-1,-,248 a - D - ,6,3-lactone, 247 -, 2,3,4,6-tetra-O-Sulfo-a-o-, 248 methyl ester, 247 -, 2,3,4,6-tct.rn-O-tosylol-~-,215, 243 -, 4-O-methyl-~-,64, 72, 91, 286, 303, -, 2,3,4-tri-O-acetyl-au-u-, 228, 247 313, 325 -, 3,4,6-t,ri-o-acet3-1-p-~-, 217, 242, 248 Glutamine, I,-, in synthesis of 2-nmino-2methanolysis of, 236 deoxy-D-glucose, 368 reaction with ammonia, 243 Glycals, 205 -, 2,3,4-tri -0-acetyl-6-chloro-6-deoxy-aaddition t o , of halogen acids and of D - , 247 halogens, 228 -, 3,4,6-tri-O-acetyl-2-chloro-2-deoxyGlyceraldehyde. See Glycerose. WD-, 235, 247 Glyceritol, 66 -, 2,3,4-tri-O-acetyl-6-deoxy-a-~-, 248 -, 1,3-O-benzylidenc-, isomers of, 30 -, 3,4,6-tri-0-acetyl-2-dcoxy-o-, 228 -, 2,3-S-benzylidene-2,3-dideoxy-2,3-, 3,4,6-tri-0-acetyl-2-0-tosgl-a-~-, 247 dithio-DL-, 28 -, 3,4,6-tri-O-acety1-2-O-t richloroace- Glyceronic acid, D - , 117 tyl-p-D-, 217, 238, 248 Glycerose, 72, 174,204 methanolgsis of, 235, 236 -, 3-phenyl-, 174 Glucosyl fluoride, 2,3,4,6-tctra-O-acedimeric, 204 t,J’l-a-D-, 246 Glycine, 360 ethyl ester, 185 p anomer, 221,246 -, 2,3,4,6-tetra-O-henzoyl-cu-1,-, 246 -, N-o-glucosyl-, barium salt,, 149 -, 2,3,4-tri-0-acet~.vl-6-chloro-6-deoxy- ethyl ester, 140 hydrolysis of, 105 a-~246 , Glucosyl halides, tetrn-0-acetyl-D-, 232 sodium salt, 149 Glucosyl iodide, 2 , 3 , 4 , 6 4 e tra-0-acetyl- -, N-D-glUCoSylg~laIlyl-,149 W D - , 256 -, N-D-glUCOSylgUaIlylglycyl-, 149 -, 2,3,4,6-tet,ia-O-benzoyl-a-D-, 256 Glycitols, 1-amino-1-deoxy-, 200 -, 2,3,4-tri-O-acet~yl-6-deoxy-6-iodo-ay-, anhydro-, formation of, 245 D - , 256 -, 1-arylamino-1-deoxy-, 191

414

SUBJECT INDEX

Glycofuranosides, from thioacetals, 23 methyl 3,6-anhydro-, formation of,

-,

22 Glycofuranosyl halides, poly-0-acetyl-,

preparation of, 97 structure of, 102 N-substituted, hydrogenation of, 103,

200

reaction with hydrocyanic acid, 201 the term, 97 Glycogens, animal, fractionation by tritylation of, 116 colunin chromatography, 79 uses of, 124 structure of, 86 -, N-aryl-, hydrolysis of, 105 Glycolaldehyde, 72,73 oxidation of, 119 Glycolic acid, 2,3,4,6-tetra-O-acetyl-p-, Karyldeoxy-, hydrolysis of, 105 D-glucoside, ethyl ester, 222 Glycols, cis-l,3-, cuprammonium rom- -, N-0-carboxyphenyl-, 98 -, N-phenyl-D-, separation of anomeric plexes of, 23 tetraacetates, 111 Glycopyranoses, 1,4-anhydro-, 21 Glycopyranosides, methyl 3,6-anhydro , -, N-p-sulfophenyl-D-, 99 -, Ni-p-tolyl-, 98 rearrangement of, 22 Glycosans, conversion t o poly-0-acyl- Glycosyl bromides, poly-0-acetyl-, staglycosyl halides, 227 bility of, 233 -, 1,2-cis-poly-O-acetyl-, 220 Glycoseen-l,2, 229 -, poly-0-acyl-, preparation of, 213 formation of, 243 Glycosides, alkyl and aryl, 229 reaction with amines, 239 -, poly-0-benzoyl-, stability of, 233 cardiac, 209 Clycosyl chlorides, poly-0-acetyl-, 82, conversion t o glycosyl halides, 222 233 flavonoid, 93 formation of, 240 1,Z-transisomers, 220 -, poly-0-acyl-, preparation of, 212 hydroxyanthraquinorie, 209 infrared absorption spectra of snomers, Glycosyl fluorides, 233.See fable of properties on page 246. 358 -, poly-0-acetyl-, stability of, 233 methyl, hydrolysis of, 322 naturally occurring P-D-, 240 -, poly-0-acyl-, 212,221 in seeds, 94 preparation of, 210 -, 1-deoxy-1-thio-, reductive desulfuri - Clycosyl halides, 01 and 6 anomers of, 209 aation of, 245 of the ‘‘0-series,” 210,219 -, glycolic ester p-D-, 2,3,4,6-tetraacepreparation of, 2@9 tate, 222 simultaneous formation and acetylation of, 214 -, p-phenylazophenyl poly-0-acetyl-, 82 “N-Glycosides.” See Glycosylamines. the term, 207 Glycosylamines, 95, 106, 243. See also -, 2-deoxy-, preparation of, 228,229 tables of properties on pages 126 to 164. -, poly-0-acetyl-, of the “p-series,” 234 acetylation of, 110 chemical properties of, 233 as antioxidants, 125 optical rotations of, 231 benzoylation of, 115 optical rotatory properties of, 230 decomposition w i t h benzaldehyde, 105 physical properties of, 233 formation of, 243 reduction with lithium aluminum hyhydrolysis of, 104 dride, 245 methylation of, 109 solvolytic reactivity of, 235 stable forms of, 232 mutarotation of, 104,183 nomenclature of, 96 -, poly-0-acyl-, configurations of, 231 physical properties of, 101 from glycosans, 227 in plastics, 125 from ortho esters, 227

234

415

SUBJECT INDEX

preparation of, from glycosides, 222 with hydrogen halide in acetic acid,

sapote, 7!) Slerculia setigera, 71

213 with hydrogen halide in ether, 212 with liquid hydrogen halide, 210 with phosphorus halides, 215 with titanium tetrahalide, 218 reaction of, with nucleophilic reagents,

234 with primary, secondary, and tertiary amines, 243 reactions of, 239 stable anomers of, 210 structure of, 229 unstable anomers of, 210 -, poly-0-benzoyl-, reactions of, 238 Glycosyl iodides, 209 -, poly-0-acetyl-, stability of, 233 -, poly-0-acyl-, 219 Glyoxylic acid, 312,314 Grapes, 93 Grass, couch, rhizomes of, 91,92 esparto, 90 leafy cocksfoot, fructan of, 91,92 rye, fructan of, 91,92 Grignard reagents, for scission of anhydro sugars, 53 Guanidine, amino-, 194 -, N-D-frUCtOSyl-, 130 -, N-D-glUCOSyl-, 149 -, N-D-mannosyl-, 156 Guanosine, synthesis of, 244 Guaran, acetate, 86 Gulitol, 1,5-anhydro-~-,246 Gulopyranose, 1,6-anhydro-b-~-,50 Gulose, D-, 66 Guloside, methyl 2,3-anhydro-4,6-0benzylidene-D-, 52 a anomer, 51,93 p anomer, 51 -, methyl 2,6- dideoxy - 6 - iodo - 3 - 0methyl-4-0-tosyl-or-~-,92 p anomer, 92 Gum, Acacia pycnanlha, 71 apple, golden, 71 cherry, 71,90 Karaya, 90 lemon, 71 myrrh, 72 peach, 71

H Halogens, radii of atoms of, 231 Hardwoods, alpha-cellulose of, 328 hemicelluloses of, 302,313 isolation of carbohydrates from, 317 mixed, sulfite pulp from, 325 pulping of, 324 sulfate pulps from, 327 Heartwood, of oak, 301,302 Hemicellulose-A, of beech wood, 91 Hemicellulose-A, -B,and -C, of black spruce, 296 of slash pine, 296 Hemicelluloses, of American white oak,

300 of aspen wood, 286 of beech wood, 300 of black spruce, 64,313 of conifers, 313 effects of, on pulps, 310,311 of hardwoods, 302,313 of jute fiber, 304 of mesquite wood, 286 nitrate, from Western hemlock wood,

330 of oak, 301 of oak wood, 313 preparation of, from holocelluloses, 318 of Scots pine, 64 of slash pine, 313 of softwoods, 302 of spruce, 290 of white pine, 313 wood, alkaline degradation of, 326 uronic acids of, 313 Hemlock (the t r e e ) , alpha-cellulose of,

328 Western, 292,319,320,330 Heparin, acetylation of, 365 N-acetyl content of, 353 acid hydrolysis of, 348 action on, of acid, 346 of alkali, 346 of enzymes, 345,365 of nitrous acid, 357 adsorption-front analysis of, 344 antibacterial action of, 36@

416

SUBJECT INDEX

anticoagulant activity of, 336,343,353, 359, 360 ash content of, 352 bacteriostatic activity of, 360 biological activity of, 359 commercial, electrophoresis of, 353 fractionation of, 344 complex of, with protein, 347 with Toluidine ]Hue, 339, 344 composition of, 339 countercurrent distribution of, 344 degradation of, 357 N- (2,4-dinitrophenyl) derivat ive, 355 discovery of, 336 distribution of, in hotly tissues, 344 distril)ution of sulfate groups in, 363 electrometric titration of, 352, 353 electrophoretic study of, 344 enzymic inactivation of, 365 enzymic sulfation of, 363 extraction of, 348 filter-paper electrophoresis of, 345 fractionation of, Nith brucine, 352 from beef liver, 335 from beef lung, 33s from dog liver, 337 n-glucuronic acid of, 351 helical structure of, 366 hexosaminc content of, 351 hexuronic acid content of, 351 histological demonstration of, 339 homogeneity of, 344 inactivation of, by acetic acid, 366 by dilute acid, 365 by enzymes, 365 by nitrous acid, 352, 365 infrared absorption spectrum of, 358 as inhibitor of pancreatic ribonuclease, 360 interaction of, p i t h proteins, 335 isolation of, 337 in mammalian tissues, 330 medical uses for, 360 metachromatic activity of, 344 molecular shape of, 364 molecular weight of, 363 mutagenic action of, 360 nitrogen content of, 352 nitroso derivative of, 354 occurrence of, 337,339, 340

octylamine-protein complex of, 348 oxidation of, in presence of ascorbic acid, 359 paper chromatography of, 343 partial acid hydrolysis of, 352, 355 partial formula of, 349, 350 polyelectrolyte character of, 364 potency of, from various sources, 347, 348 potentiometric titration of, 353 properties of, 337, 338 provisional international standard for, 361 purification of, 338, 339, 355 purity of, criteria of, 343 regeneration of, from heparin-octyl:tmiiie-protein complexes, 345 from heparin-protein complexes, 345 rctention of acetic acid by, 353 Roche, 314 simultaneous acetylation and desulfation of, 356 simultaneous hydrolysis and oxidation of, 351 structure of, 348, 360 sulfaniic acid groupings of, 354 sulfate bridges in, 364 “N-sulfate” group of, 354, 363 sulfation of, 364 sulfur content of, 352 supposed species-specificity of, 347 surgical uses for, 360 the term, 336 total, acid hydrolysis of, 351 ultraviolet absorption spectrum of, 343 Vittum, 344 p-Heparin, 367 $-Heparin, 349, 350,352 acid hydrolysis of, 350 action of nitrous arid on, 358 derivatives of, 360 -, N-isonicotinyl-, 360 -, N-nicotinvl-, 360 -, N-trifluoromethylphenyl-, 360 Heparinnse, 345 Heparinic acid, 336, 353 ammonium salt, 340,341 barium acid salt, 340, 341, 347, 352 analysis of, 31

RUBSECT INDEX

417

barium-sulfur ratio of, 352 CJf y C a S t , , 36s ~~CXOkill&SC, coni1)osit ion of, 351 Ilesonic u i d , di-O-iiict.Ii~leiic-, 42 iiitrogeri-sulfur ratio of, 352 Hesosans, 326 slruct~ii~.e of, 357 of cellulose prqi:ir:~ti~~iis, 321, :;25 bariiun salt, 343, 348, 3% Hcose-:i.niiiioni:i, I06 honwgcneous, 345 Ilesositle, methyl 2,6-tlideosy-3,4-cli-0benzitline salt,, 33‘3,355 1,osyI-01- I ) -.ryfo-, !U brucine s:tlt, 338, 339 p anomer, 92 tlecamethylenediamine salt, 343 Hexulose, 1 ,6-dideosy-l -(p-tnIui~litin)dodecylamine salt, 343 L-orrrbino-, 205 isoperitylamine salt, 343 -, r.ibo-. See T’sic,ose. n-pent>ylaminesalt, 343 Hexiironic acid, O-metli~.l-,313 piperidine salt, 343 of hardwoods, 302 prot,ein complex of, 336 of oak heniicelluloses, 301 sodium salt, 336, 346 of softwoods, 302 analysis of, 340 Ilist,idine, N , N ’ - d i - ( n - g l i ~ c o s ~ ~ l g i i a n ~ ~ l ) Heparosinsulfuric acid, 356 L-, 136 -, N-acetyl-, 356 Holocellulose, 288, 313, 318, 331 Heptonic acid, ~ - g l ~ c e r . o - ~ ) - g uamide, lo-, of birch wood, 303, 314 77 of black spruce, 296,319 Heptose, r~-glycero-o-qulo-,66, 77 carbohydrate content of, 319 Heptosyl hromide, 2,3,4, 6,7-pentma-0- chlorite, 332 acetJ.l-P-D-gl!/cer.o-I,-ynlacto-, 253 from aspen wood, 308, 309, 310 -, 2 , 3 , 4 , 6 , 7 - p e n t ~ ~ - O - n c e t g l - ~ - n - g Z ~ ~ r e ~ oprocedure for, 317, 318 w y u l o - , 253 of slash pine, 318 -, 2,3,4,6,7-pent,a-O-acet~yI-ol-~-glyce~o- delignified, 312 D-ido-, 253 of Douglas fir, 319 -, 2,3,4,6,7-penta-O-acet,yl-ol-~-gl~~cer.o- hydrolysis of, 320 ~ - t n l o -253 , isolation of, 317 _ _, 2,3,4,6-tetra-O-ncet~.l-i-~~eoxy-01-~~of loblolly pine, 319 glycero-L-gulncto-, 251 maple, analysis of, 316 Heptosyl chloride, 2,3,4,6,7-penta-Ofrom non-woody plant material, 318 Ltcetyl-ol-D-glycCrO-D-g~ZO-,248 of overcup oak, 319 -, 2,3,4,6,7-penta-O-acetyl-p-D-gl?/ce,.oof slash pine, 290, 295, 296, 319 D - g U b , 248 of softwoods, 325 -, 2,3,4,6-tetra-U-acet~~1-7-deosy-~-gly- of Southern red oak, 319 cero-L-galacto-,2-17 of spruce, 314 Heptulosan, ~ - i d o - 73 , of sugar maple, 320 Heptulose, D-gluco-, 66 the term, 316 D-~ZU.TWLO-,66 of Western hemlock, 319 -, I-deosy-I-diazo-keto-L-guluc~o-, penof Western red cedar, 319 taacetate, 87 from wood, 312 -, 1-deoxy-I-diazo-keto-L-nlanno-, pen- Horses, polysaccharide from intestines taacetate, 87 of, 90 Hexaric acids, 2,4:3,5-diacetaIs of, 40 Huckleberry, leaves, 93 -, 2,4:3,5-di-O-methylene-,7 Hudson’s rilles, of isorotation, 112 Hexitols, 1-amino-1-deosy-, deaminafor naming anomers, 230 tion of, 23 Humin, formation of, 193 -, 1,4-anhydro-,23 Hpaluronic acid, 346,349,366 -, 1-arylamino-1-deoxy-, 200 biosynthesis of, 367

418

SUBJECT INDEX

microbial, 368 oxidation of, in presence of ascorbic acid, 359 11) dantoic acid, N-o-glucosyl-, 119 ethyl ester, tetraacetate, 1.1-9 potassiiim salt, 149 -, N-(o-glucosy1)thio-, ethyl ester, 150 ethanolate, 150 tetraacetate, 150 potassium salt, 150 Hydant oin, N - o -gluc osyl-, 140 -, N-(1~-glucosyl)-2-thio-,150 Hydrasine, action on o-friictose, 187 -, (2,4-dinitrophenyl)-, 305 -, (p-nitropheny1)-, 195, 305 -, phenyl-, 305 Hydrindan. See Indan, hesahydro-. 1-Hydrindanone. See 1-Indanone, hexahydro-. Hydrobenzoin. See 1,2-Ethanediol, 1,2diphenyl-. Hydrogen bromide, liquid, 211, 228 Hydrogen chloride, liynid, 211, 222, 224 Hydrogen cyanide, 102 reaction with X-suhstituted glyrosylamines, 201 Hydrogen fluoride, anhydrous, 225 liquid, 212 Hydrogen iodide, 219 Hydrogen-ion concentration, effect on glycosylamines, 106 Hydrophilicity, of wood pulps, 315 Hydroxylamine, hydrochloride, 306

, 2,1:3,5-di-0-metIiy.Icnc

- 1,6-di-Otosyl-lr-, 12 -, 1 , 3 :2,1:5,6-tri-O-l,eiixyliclene-~-, 39 Idopj-ranow, 1,6 anhydro-p-r-, 50 Idose, 6-deouy-l , 2:3,5-di-O-isopropylidene-6-nitro-1-, 36 Idoside, methyl 4,6-O-benzylidene-a-o-, 43 4 anomer, 44 Idosylamine, 6 - d e o ~ g - h ' - p h e n y l - ~ -3-, methyl ether, 151 Indan, hexahydro-, 20 1-Indnnone, hexahpdro-, 20 Indophenol, 2,6-dirhlorophenol-, 187, 188, 201, 202 use in column chromatography, 65 Infrared absorption spectra, 100, 102, 103, 186 Inositol, rnyo-, 66, 87 Interferometer, use in column chromatography, 58 Iiiulin, 91, 92 D-glucose content of, 91 sulfated, 361 Invertase, of yeast, 64 Ion-exchange resins, for column chromatography, 75 Iris, galactomannan of seed, 72, 90 Iris ochroleuca, seeds, 289 Iris sibirica, seeds, 289 Isoglucosamine, 173 -, N-p-tOlyl-D-, 173, 199 Isoguanosine, synthesis of, 244 Isomaltose, 86, 87 I p - , acetate, 86 isolation of, b y column chromatogIdaric acid, 2,4:3,5-di-O-meth~lene-r,-, 40 raphy, 63 epimerieation of, 41 p-, octaacetate, 86 Iditol, L-, 66 Isoquercitrin, 94 2,4:3,5-diacetals, 39 Isorhamnosylamine. See Glucosylamine, -, 2,5-diamino-l,4:3,6-dianhydro-Z, 56-deoxy-. dideoxy-L-, 48 Isosucrose, octaacetate, 88 -, 1,4:3,6-dianhydro-~-,46, 49, 67 Ivory nut, mannan of, 289,290 -, 1,4:3,6-dianhydro -5 -chloro-5-deoxy 2-O-mesyl-~-,48 J -, 1,4:3,6-dianhydro-2,5-di-O-tosyl-~-, Juniperis commzinis, 286 47, 49 -, 1,4:3,6-dianhydro-2,5-O-methylene- Jute, 324 fiber, hemicelluloses of, 304 L - , supposed, 47 -

.

SUBJECT INDEX

K Karaya gum, 90 Kestose, 64 Ketohexopyranosyl halides, poly-0-acetyl-, stable forms of, 233 Ketoses, condensation with aliphatic amines, 100 dehydrogenation of, by hydrazines, 197 differentiation from aldoses, 197 reagents for detection of, 193, 194 Seliwanoff test for, 201 -, 1-amino-I-deoxy-, formation of, 169, 170 nomenclature of N-substituted, 173 optical rotations of, 186, 203 -, 1-arylamino-1-deoxy-, hydrogenation of, 191 for preparation of phenylosazones, 197 -, 1 - deoxy-1-(N-methylanilino)- 3 phenyl-, 174 semicarbazone, 175 Ketosylamines, N-substituted, 99 Knoevenagel reaction, 179 Koenigs-Knorr reac'tion, 209, 231, 240 improvements in conditions for, 241 Kojic acid, synthesis of, 229 Konjac mannan, 289 Kraft process, for pulping, 326,329

L Lactaldehyde, DL-, 73 Lactic acid, 184 in pulping waste liquors, 326 Lactitol, 66, 77 Lactobacillus gayonii, 306 Lactobacillus nrannitopoeus, 306 Lactose, 66, 76, 80, 83 action of ammonia on, 73 azoate, a-,80 p nnomer, 80 monohydrate, 77 octancetate, 215, 216 I,actosylnmine, N-acetyl-, dihydrate, 151 heptaacetate, monohydrate, 151 -, N-4-biphenyl-, 151 -, N-p-bromoplienyl-a-, heptaacetate, 1.51

0 anomer, 151 -, N-carbamoyl-, 151

, N

419

- (4-carboxy-3-hydroxy.phenyl)-, phenyl ester, tetrahydrate, 151 sodium salt, 151 -, N,N-dimethyl , heptaacetate, 151 -, N-I-dodecyl-, 151 -, N-p-ethoxyphenyl-, 151 -, N-phenyl-a-, heptaacetate, 151 @ anomer, 151 -, N-p-sulfamylphenyl-, 151 trihydrate, 151 -, N-p-tolyl-a-, heptaacetnte, 151 P anomer, 151 Lactosyl hromide, hepta-0-acetyl-a-, 254 I,actosyl chloride, hepta-O-acet~l-,215, 216 a anomer, 249 I~actosylfluoride, 246 heptaace t at e, 246 1,actosyl iodide, hepta-0-acetyl-a-, 256 Lactosyl isothiocyanate, hepta-0-acetyl-, 151 Lactulose, 73 Laminaribiose, a-. See Glucopyranose, 3-0-(p-o-glucop~ranosgl) -1)Larch, e-galactan of, 72 Lead tetraacetate, oxidation I)y. See Oxidation. I>ecithin, 360 Lemonflavin. See Oak, bark. Lemon gum, 71 Leucine, 360 Leuconostoc mesentei oitbs, 362 Levans, bacterial, 89 Levoglucosan. See Glucopyranose, 1,6anhydro-p-o-. Levulinic acid, 305 lichenin, fully methyhted, 82 triacetate, 223 J,ignin, 284, 316 association with xylan, 310 dissolution of, 326 i n hemirelluloscs, 287 of maple holort.llnlose, 316 of maple wood, 316 sodium salt, 325 -, chloro-, 316, 317 h p e m i a , alimentnrp, 360 Lithium aluminum hgdride. See Aluminum lithium hydride. -

420

SUBJECT INDEX

Lobry de Brngn and Alhcrda van Ekenstein transformation, 184, 195 LoIZ'IL~Z perenne, 91 Lucerne, galnctornannan of, 72 galactomannan of seeds, 90 Lysinc, N2,N6-di-(D-glucosy1) - DL- , 136 -, N , Nl-di- (D-glucosylguanyl)- D L - , 136 -, N-D-glUcosyl-, 149 Lyxitol, 1-amino-1-deoxy-n-, 173, 205 Lysonic acid, D-, amide, 77 -, 2,3, 4-tjri-O-methyl-~-,lactone, 266 phcnylhydrazidc, 266 -, 2,3,5-tri -0-methyl-o-, 266 lactone, 266 phcnylhydrazide, 266 Lyxosc, D-, 66 -, 2,3,4-tri-O-methy1-1-,266 -, 2,3,5-tri-O-mcthyl-~-, 266 Lyxosidc, methyl 2,3-anhgdro-5-0methyl-a-o-, 266 -, methyl 2,3,4-tri-O-nicthyl-~-,266 Lyxosglaminc, D - , 152 -, N-(4,5-dimcthyl-2-nitrophcnyl) -D-, 152 2,3,4-triacctate, 152 -, N-(4,5-dimethyl-2-1iitrophenyl)-~-, 152 2 , 3 ,4-triacctate, 152 -, N - (3,4-dimethylphcnyl) - D- , 152 -, N-p-nitrophcnyl-I)-,152 -, iv-phcnyl-D-, comples with sodium sulfate, 124 I ~ g ~ o s ybromide, l 2,3,4-tri-O-acctyl-aD - , 253 1,ysosyl halides, tri-O-:tcet,yl-i)-, 232

M nragllcsol, 7s, 70, SO, s 1 , m , SG, 8 7 , N for coliimn chroiiiat,ogr:tl,liy, 93 hlaillard reactiou, 172, 103 Malic acid, levo-, 66 Rfaloriic acid, 177 ethyl e s k r , 177, 201 hlultitol, acetate, 8G Maltotlextrins, prep~r;~!ion of, 63 Malt.ohept.aose,prep:irat.ion of, 03 Rlsltose, 66, 76, 83, 87 action of ammonia on, 7:; P - , aeoatc, 80 degradation of acetate, 224

heptaacetate, 213 monohydratc, 77 g-, octaacctatc, 83,88, 213 -, hexa-O-aoetyl-l,2-0-(1-chlorocth~lidene)-, 213, 227 Maltosylaminc, 153 heptaacctatc, 153 -, N-(l-carbomethoxy- 3 -hydrosyphcny1)-, 153 -, N - (4-cnrboxy-3-hydroxyphenyl)-, 153 -, N-(o-carbosyphcny1)-, 98, 153 -, N-(l-dodccyl)-, 153 hydrolysis of, 105 -, N-l-hcryl-, 153 -, N-1-octadecyl-, 153 -, N-phcnyl-, 153 hcptaacctate, 153 -, N-p-sulfamylphenyl-, 153 -, N-p-tolyl-, hcptaacctatc, 153 Maltosyl bromide, hepta-0-acetyl-, 214, 222 01 anomcr, 254 from potato starch, 223 Maltosgl chloride, hcpta-0-acctyl-a-, 213, 249 -, hesa-O-acet~l-2-O-tI.ichloro~~cctyl-p~, 217, 249 Ma1tosyl fluoride, hept:t-O-acetyl-, 216 Pvlaltosyl halides, poly-0-acctgl-, 232 Maltotetraosc, isolation of, by column chromatography, 63 Rf a1tot riosc, 86 isolat,ion of, by column chromatography, 63 hfaltulosc, 73 Malviri chloride, 2440 hlandelic acid, D L - , ethyl ester, 240 Mannans, 284, 319, 320 of alpha-ocllnlose, 328 of birch, 326 compound wit,h cellulose, 296 content of, in coniferous woods, 296 detcrrninatior~of, 290, 294 optical rotntion of'. See tuble on prc~lc 290.

oxidation wit,li lcail t.ptr:i:t of pine wood, 326 properties of, 205 st,ructurc of, 288 of sulfitc pulps, 323, 325

SUBJECT IXDEX

42 1

the term, in wood chemistry, 284 Mannopyranoside, methyl WD-, 67 of unbleached, sulfate pulps, 327 B anomer, 16, 322 of woods, 285,290 -, methyl di-0-benzylidrne-oI-u-, 35 -xylaii combination, of wood pulp, 329 -, methyl 6 - o - t o s y l - a - ~ -22, illarman A , 288,289,290 Maniiopyranosyl iodide, O - ( B - ~ - g l u c o of ivory nut, 290 pyranosyl) - o ~ - D - , heptaacetate, 256 Mannan B, 288, 289, 290 Mannosaminic w i d , penta-0-acetyl-NMannaric acid, 2,4:3,5-di O-methylenemethyl-L-, 87 D - , epimerization of, 41 Mannose, 318, 322 Mannitol, D-, 64, 66, 76, 77,87 D-, 64, 66, 77, 173 effect of hot hydrochloric acid on, 23 determination of, 293 hesaacetate, 88 I)-, 1-dincetamide derivative, 154 -, 1-amino-1-deoxy-I)-, 205 I ) - , 1-dibenzamide derivative, 124, 154 -, 1,4-anhydro-~-,23,66 pentabenzoate, 154 -, 1,5-anhydro-1)-,23, 66, 245 disaccharide of, 296 di-0-benzylidene acetal, 35 n-, monobenzamide derivative, 124 6-tosylate, 23 phenylhydrazone, 291, 292 -, 6-deoxy-l , 3:2,5-di-O-methylene-r-, D - , phenylhydrazone, 102 44 D - , reaction of, with dihenzylamine, -, 1-deoxy-1-p-toluidino-o-, 199 200 -, 1,4:3,6-dianhydro-1-, 46, 66 with morpholine, 200 -, 1,5:3,6-dianhydro-~-,67 with p-phenetidine, 98 -, 1,4:3,6-dianhydro-2,5-dideoxy-2,5- with piperidine, 200 imino-u-, 48 with p-toluidine, 200 -, 1,4 :3,6-dianhydroS,5-di-O-t,osyl-r)-, -, 2,5-anhydro-o-, 358 47 -, 2 , 3 :5,6-di-O-isopropylidene-u-,32, reaction with ammonia, 48 34, 221 -, 1 , 4 :3,6-dianhydro-2,5-O-methylene--, 6-O-(~~-~-galactopyranosyl)-p-1,-, 73 n-, 47 -, O-glucosyl-, 296 -, 1,B-di-O-benzoyl-o-,41 -, 1,2,3,4,6-penta-O-benzoyl-~-, action 2,5-0-methylene acetal, 35 of ammonia on CY and p anomers, 124 -, 1 ,6-di-O-benzoyl-3,4-O-benzylidene--, 2,3,4,6-tetra-O-benzoyl-~-, 115 D-,35 -, tetra-0-methyl-o-, 288 -, 2,3,4-tri-0-acetyl-I,6-anhydro-o-, 228 2,5-O-methylene acetal, 35 -, 1,6-dichloro-l,6-dideosy-~-, 2,3,4,5- Mannoside, methyl 2,3-anhydro-4,6-0benzylidene-a-D-, 51, 52 di-0-ethylidene acetal, 41 isomeric di-0-isopropylidene acetals of, -, methyl 2,3-anhydro-4,6-di-O-methyI6-D-, 52 41 Mannosylamine, D-, N-substituted, 184 -, 2,j:3,5-di-O-methylene-u-, 6, 40 -, N-benzoyl-D-, 154 1,6-ditosylate, 43 tetraacetate, 154 -, 2,5-o-methylene-o-,45 -, N-n-butyl-D) 154 -, mono-0-benzylidene-mono-0-meth- -, N-p-carbethoxyphenyl-D-, 154 ylene-D-, 41 -, N-o-carbomethoxyphenyl-o-,154 -, 1,3:2,5:4,6-tri-O-ethylidene-~-, 44, -, N - (4-carboxy -3-hydroxyphenyl)-D-, 45 154 -, 1 , 3 :2,5 :4,6-tri-O-methylene-, 44 phenyl ester, 154 Mannonic acid, D-, 66 sodium salt, 154 from sulfite waste liquors, 322 -, N-ni-carboxyphenyl-D-, 154 51 Mannopyranose, 1,6-anhydro-p-~,-, -, N-a-carboxyphenyl-n-, 98, 154

422

-, N-p-carboxyphenyl-D-, 154

SUBJECT INDEX

-, 4-0-(~-~-glucopyranosyl)-~-1t-, hepN- (3,4-dimethyl-2-nitrophenyl) -D-, taacetate, 24'3 154 -, 2,3,4,6-tetra-O-acetyl-c-~-, 220, 248 -, 2,3,4,6-tetra-O-benzoyl-a-~-, 248 tetraacetate, 154 -, N-(4,5-dimethyl-2-nitrophenyl)-u-, -, 3,4 ,6-tri-0-acetJy1-o-,249 154 2-trichloroacetate, 249 Mannosyl fluoride, 3,6-di-O-acety1-4-0tetraacetate, 154 (2,3,4,6 - tetra - 0 - acetyl - 0 - D 6-trityl ether, triacetate, 154 glUC0Syi)-a-0-, 212, 246 -, N- (3,4-dimethylphenyl)- D - , 154 -, N-p-ethoxyphenyl-1)-, 154 -, 4-O-(p-~-glucopyranosyl)-a-~-, heptaacetate, 246 -, N-1-hexyl-o-, 155 246 -, N-l-(2-hydroxynaphthyl)-N-phenyl- -, 2,3,4,6-tetra-O-acetyl-c~-~-, Mannosyl halides, poly -0-srety 1-1-0ethyl-D-, 155 (D-glUCOSyl)-D-,232 -, N-(4-methyl-2-nitrophenyl) -D-, 155 -, tetra-0-acetyl-D-,232 tetraacetate, 155 Mannosyl iodide, 2,3,4,6-tetra-O-ace-, N-2-naphthyl-~-,155 tyl-, 256 -, N-nL-nitrophenyl-0-D-, 155 -, 2,3,4,6-tetra-O-benzoyl-, 256 -, N-o-nitrophenyl-B-D-, 155 Maple, holocellulose, analysis of, 316 tetraacetate, 155 holocellulose of sugar-, 320 -, N -p-nitrophenyl-0-o-, 155 wood, 316 dihydrate, 155 analysis of, 316 tetraacetate, 155 Melanoidins, 193 -, N-1-pentyl-u-, 155 Melezitose, 66, 77,80 -, N-phenyl-o-, 102, 155 azoate, 80 complex with sodium sulfate, 124 Melibiitol, 66 2,3-dimethyl ether, 155 Melibiose, 66, 218, 242 2,3,4,6-te tmme thy1 ether, 155, 156 action of ammonia on, 73 2,3,6-trimethyl ether, 155 0-, azoate, 80 2,4,6-trimethyl ether, 155 octaacetate, 218 3,4,6-trimethyl ether, 155 8-anomer, 88 -, N-4-phenylazophenylene-~-, 155 synthetic, 242 -, N-p-sulfamylphenyl D-, 156 synthesis of, 242 monohydrate, 156 Melibiosyl bromide, hepta-0-acetyl-a-, -, N-p-sulfophenyl-o-, 156 254 -, N-P-tolyl-D-, 102, 156, 199 Melibiosyl chloride, hepta-0-acetyl-a-, tetraacetate, 156 249 tetrabenzoate, 115 Melibiosyl fluoride, hepta-0-acetyl-, 246 2,3,4,6-tetrabenzoate, 156 Mannosyl bromide, 4-o-(@-D-glUCOpy- Melibiosyl halides, poly-0-acetyl-, 232 Melibiosyl iodide, hepta-0-acetyl-a-, 256 ranosyl)-a-o-, heptaacetate, 254 -, 2,3,4,6-tetra-O-acetyl-a-~-, 220, 241, Melibiulose, 73 Metasaccharinic acid, a-o-galacto-, 66 245, 253 Methane, diphenyl-, 177 methanolysis of, 236 Methylamine, 354 reaction with secondary amines, 239 Methylation, of glycosylamines, 109 solvolysis of, 236, 237 Methylene Blue, 188,201, 202 -, 2,3,4,6-tetra-O-benzoyl-a-~-, 253 Micrococcus pyogenes, 360 Mannosyl chloride, 2,3 :5,6-di-O-isopro- Milk, nitrogenous tetrasaccharide from pyiidene-D-, 221 human, 64 a anomer, 248 Mill, Lamp&, 308 -,

423

SUBJECT INDEX

Mit,osis, 360 Molasses, beet, 68 cane, blackstrap, 68, 87 fermentation residue of, 87 cane, yeast-fermentation residue of, 68 Molisch reagent, 89 Monosaccharides, separation from disaccharides, 57 Morpholine, 177, 200 Mucilage, flasseed, 72 slippery elm, 72 MucopolysBccharides, biosynthesis of, 367 classification of, 352 enzymic desulfation of, 356 infrared absorption spectra of, 358 Mucoproteins, linkages in, 172 Mycobacterium tuberculosis, polysaccharides of, 79 Myrrh, gum, 72

N Napthalene, ris-decahydro-, 18 trans isomer, 18 Neighboring-group effect, 235, 237 Neokestose, 64 Neolactosyl chloride, hepta-0-acetyl-, 216 a anomer, 249 Neurospora crassa, 368 Nicotinamide, glycosyl , 245 -, o-ribofuranosyl-, 245 Nigeran, 91 Norcamphane, 21 Nortropane, 21 Noslac, polysaccharide of, 72 Novocaine. See Procaine. Kucleic acid, deoxyribo-, 360 ribo-, 360 Nucleosides, purine and pyrimidine, permanganate oxidation of, 119 synthesis of, 243

0 Oak, 324 Ameriran white, 300 bark of, 93 hemicellulose-A and -B, from heartwood of, 301 from sapwood of, 301

hemicelluloses of, 301 overcup, 319 Southern red, 319 wood, hemicelluloses of, 313 Obituary, of Edmund George Vincent Percival, xiii Octadecylamine, reaction with o-fructose, 100 Octulose, D-glycero-D-g?do-, 87 keto form, heptaacetate, 87 -, 1-deoxy-1-diazo-keto-D-glycero-D-galacto-, hexaacetate, 87 Oligosaccharides, 208 acetates of, 295 a-D-linked, synthesis of, 241, 242 synthesis of, 240 of D-xylose, 300 Optical rotations, of poly-0-acetylglycosy1 halides, 231 relation to structure, of sugars, 230 Optical superposition, Van’t Hoff’s principle of, 230 Orcinol reagent, 76 Osazones, phenyl-, formation of, 197 mechanism of formation of, 197 preparation of, 172 7-Oxabicyclo[4.1.O]heptane, 21 Osalic acid, 314 Oxidation, with alkaline hypoidite, 312 with bromine, 312 with bromine-sulfuric acid, 351 with chlorous acid, 312 with chromium trioxide in pyridine, 43 with cupric salts, 187 187 with 2,6-dichlorophenolindophenol, with 0-dinitrobenzene, 187 with hydrazine(l87 with lead tetraacetate, 118,294 with nitric acid, 286 with nitrogen dioxide, 312 waith nitrogen tetroxide, 314 with oxygen in the presence of ascorbic acid, 359 with periodate, 116, 190, 196, 200, 312, 356, 357 with selenious acid, 187

P Panitol, dodecaacetate, 86 Panose, 86

424

SUBJECT INDEX

acetate, 86 -, 1-deoxy-1-piperidino-D-lhreo-, 203 isolation of, by column chromatog5-trityl ether, 196,203 raphy, 63 hydrochloride, 203 Paper, beater additive for, 289 -, 1 - deoxy - 1 - (p-toluidino)-L-erythl.o-, determination of pentosan in, 304 205 Paper birch, wood, 318 -, l-deosy-l-(p-toluidino)-D-threo-, 205 xylan of, 299 Peonin chloride, 240 Paper chromatography, 76,90,106, 287, Percival, Edmund George Vincent, obit295,307,318,322,326,327 uary of, xiii formation of glycosylamines during, Periodate oxidation. See Oxidation, with 121 periodate. of heparin, 343 Perseitol, 1,3:5,7-di-O-benzylidene-o-, quantitative, 292 30 of wood hydrolyzates, 285 -, 1,3:5,7-di-O-henzylidene-~-, 30 Papermaking, 315 Perseulose, D-,66 Pager pulp. See Pulps. L-, 66 Paraformaldehyde, 72. See also s-Tri- pH. See Hydrogen-ion conceptration oxane. Phenethylamine, 185 Paraldehyde, 15 p-Phenetidine, reaction of, with D-frucPeach, gum, 71 tose, 98 Pears, xylan from, 300 with D-galactose, 171 Peas, enzyme of, 73 with D-glucose, 97,170, 171,175 Pectic acid, 91 with D-mannose, 98 from wood, 313 with L-sorbose, 98 Pelargonin chloride, 240 Phenol, relative affinity for carbon, 59 Pentaerythritol, 67 -, p-phenylazo-, 82 Pentalene, octahydro-, 22 o-Phenylenediamine, 96, 198 cis form, 17 -, N , AT’-di- (tetra-0-acetyl-D-glucosyl)trans form, 17 4,5-dimethyl-, 112 2,4-Pentanedione, 177,201 I’hlorizin, 67 Pentitols, 1-arylamino-1-deoxy-, 200 Phloroglucinaldehyde. See BenzaldePentosans, 326 hyde, 2,4,6-trihydroxy-. alkali-insoluble, of wood, 308 Phloroglucinol, 305 of maple holocellulose, 316 Phosphoglucomutase, 368 of maple wood, 316 Phosphorus pentabromide, 228 removal of, from unbleached sulfite Phosphorus pentachloride, 215,221 pulps, 325 Photometer, Zeiss Schnell-, 291 resistant, of wood, 308 Picea ezcelsa,286,314 of wood pulps, 324 3-Picoline, 239 Pentulose, D-threo-, 73 Pine, alpha-cellulose of, 328 oxidation of, with cupric salts, 187 hemicelluloses of, 302 with 2,6-dichlorophenolindophenol, loblolly, 319,320 187 pectic material from, 313 with o-dinitrobenzene, 187 mannan of, 290 -, 1-alkylamino-l-deoxy-o-th,eo-,203 pulp from, 327 -, 5-deoxy-~-fhreo-, 73 slash, 319,320 -, 1 - deouy-l-(3,4-dimethylanilino)-~chlorite holocellulose of, 318 erythro-,171,205 -, 1 - deoxy-l-(3,4-dimethylanilino)-~- hemicelluloses of, 313 sulfate pulp from, 331 erythro-, 205 hydrogenation of, 192 sulfite cooking of, 307,322

SUBJECT INDEX

srilfit,e p u l p from, 310, 328 whit,e, hemicellulose of, 313 wood, glucan and niannari of, 326, 327 s y l a n of, 327 Pinus caribaea, 318 Pinus silvestris, 286 Piperidine, 200, 239 acetat,e, 179, 195 action of, on penta-O-ncet,yl-o-glucopyranoses, 112 on 2,3,4,6-t,etra-O-:icet,yI-D-gliicosr, 112 on 3,4,6-tri - 0 - acet,yl - D - glucosgl chloride, 112 -, N-o-arabinosyl-, 126 -, N-~-arabinosyl-,128 -, N-o-galactosyl-, 134 mutarotation of, 105, 194 -, i\i-D,-glUCOSyl-,97, 140 2-carbanilate, 150 decomposition of, 101 hydrochloride, 149 hydrogenation of, 104 2-methyl ether, 150 3-methyl ether, 150 reaction with hydrogen cyanide, 102 supposed mutarotation of, 104 2,3,4,6-tetraacetate, 113, 149 tetramethyl ether, 150 2-p-toluenesulfonate, 150 3,4,6-triacetate, 112, 149 2-carbanilate, 149 2-methyl ether, 149 2-p-nitrobenzoate, 149 2-p-toluenesulfonate, 149 -, N-D-mannosyl-, 156 mutarotation of, 105 -, A’%-xylosgl-, hydrochloride, 164, 196 Plasma. See Blood plasma. Plastic materials, use of glycosylnmines in, 125 Polygalitol. See Glucitol, 1,5-anhydroD-.

Poly-D-glucoses. See Glucans. Polyose, the term, 284 of wood, 311 Polysaccharides, associated with wood cellulose, 283 from horse intestines, 90 noncellulosic, of wood, 328 from sheep rumen, 90

425

from sprnce wood, 318 sulfated, reaction with Tolrridine Blue, 344 synthetic, 361 test for, 365 sulfatiori of, 361 triacetates of, linked 3 --f ~ - c Y - D - 224 , linked 4 -+ LwD-,224 linked 6 --f I-WD-, 224 Phlyuronides, of pulp, 313 Porphyrn uinhi/iraZis, mannan o f , 289, 290 Primeverasyl chloride, hexa-0-acetyl-a-, 249 Procaine, N-o-ghicosyl-, monohydrochloride, 140 monohydrate, 140 1,2-Propanediol, 67 2-Propanone, 1,3-dihydroxy-, oxidation of, with 2,6-dichlorophenolindophenol, 187 with o-dinitrobenzene, 187 -, hydroxy-, 195 -, phenyl-, 177 Propionic acid, 3-mercapto-, 177 Propylene glycol. See 1,2-Propanediol. Proteins, complexes with heparin, 338 Prothrombin, 359 Psicofuranose, 1,2:3,4-di-O-isopropylidene-D-, 73 Psicose, D - , 66, 73 phenylosazone, 73 Pulping, alkaline, 307 of hardwoods, 324 procedures for, 321 soda process, 321 sulfate process, 321 sulfite process, 321 Pulps, acetylation grade of, 295, 328 alkaline, 325 from hardwoods, 308 alpha-cellulose content of, 298 from aspen wood, 308, 309 bleached kraft, 309 bleached, soda, 308, 309 bleached, sulfate, 308, 309 bleached, sulfite, 308, 309 chlorite holocellulose, 308, 310 kraft-cooked holocellulose, 309 overhleached sulfate, 309

426

SUBJECT INDEX

resistant xylnn of, 309 xylan of, 309 bleached coniferous, 330 carboxyl content of, 298 for chemical conversion, 311 determination of pentosan in, 304 difficultly and easily accessible material of, 331 dissolving, 323 folding resistance of, 311 hardwood, 292 high-alpha, 324, 329 hydration capacity of, 311 mannan content of, 298 of mixed hardwoods, 325 for papermaking, 310 pentosan content of, 298, 324 pine sulfite, 290 rag, free from hemicellulose, 311 softwood, 296, 325 sulfate, alpha-cellulose content of, 327 carbohydrate composition of, 326 comparison with sulfite pulps, 327 glucan content of, 327 from hardwoods, 310, 327 prehydrolyzed, 327 , from softwoods, 327,328 sulfite, 310 alpha-cellulose content of, 327 comparison with sulfate pulps, 327 glucan content of, 327 hydrolysis of, 323 pentosans of, 324 from softwoods, 308, 310, 328 sulfite cooking of, 332 tear resistance of, 311 technical, uronic acid of, 313 tensile strength of, 311 unbleached sulfate, 292 carbohydrate composition of, 327 unbleached sulfite, removal of pentosans from, 325 uronic acid anhydride of, 328 white birch, 325 wood, 284, 321 alkaline extraction of, 308 carboxpl content of, 315 carboxyl groups of, 315 carboxylic acids in, 312 effect of beating, 308

effect of drying, 308, 309 effect of grinding, 308 gamma-cellulose of, 329 hydrolyzates of, 292 hydrophilicity of, 315 mannan content of, 298 mannan plus xylan content of, 329 pentosan in, 303 rate of beating of, 315 technical, 321 for viscose rayon, 311 xylan content of, 298 Pyran, 2,3-dichlorotetrahydro-,235 -, tetrahydro-, 16 Pyranose ring, conformations of, 232 Pyridine, 239 -, N-~-ribosyl-3-carbamoyl-,160 -, N-(2,3,4,6-tetra-O-acetyl-~-glucosyl) -3-carbethoxy-l , 2-dihydro-, 148 -, N - (2,3,4,6-tetra-O-acetyl-o-glucosyl)-l,2-dihydro-, 148 Pyridinium bromide, N-~-glucosyl-3carbamoyl-, 148 -, N-~-glucosyl-3-carbnmopl-l ,2 (or 1,6)-dihydro-, 148 2’, 3’, 4’, 6’-tetraacetate, 148 -, N-~-ribosyl-3-carbamoyl-,160 N’-acetyl derivative, 160 - , N-(tetra-0-acetyl-D-glucosyl)-, 150 -, N-(2,3,4,6-tetra-O-acetyl-u-glucosyl)-3-cyano-l,2-dihydro-, 148 Pyrrole, N-D-glucosyl-, 150 Pyruvaldehyde, 174, 195 bis-p-nitrophenylhydrazone,195 m-nitrobenzoylosazone, 174 p-nitrophenylosazone, 198 -, 3-phengl-, m-ni t rohenzoylosazone, 174

Q Quercetin, 94 -, glycosyl-, 93 Quercetrin, 94 Quercitol, 66 Quercitrin, 93 Quercus robur ,286 Quinoxaline, 2-(u-arabino-tetrshydroxybutyl)-, 198,200 Quinoxalines, preparation of, 172

SUBJECT INDEX

R Rafinose, 66,75 azoate, 80 hendecaacetate, 88 pentahydrate, 77 in raw beet sugars, 74 separation of, by column chromatography, 57 Rayon, cellulose acetate, 329 textile, 323 viscose, 331 Redox indicators, 188 Reductic acid, 188 Reductones, 101, 184, 188 formation of, 194 six-carbon, 193 the term, 188 Resorcinol, 193 Rhamnitol, 174 D - , 66 L - , 66 -, 1-amino-1-deoxy-L-, 205 Rhamnoglucosides, 94 Rhamnonic acid, 3,4-di-O-metliyl-~-, 267 amide, 267 lactone, 267 -, 2-O-methyl-~-,amide, 266 l,.l-lactone, 266 -, 4-O-methyl-~-,lactone, 267 -, 5-O-methyl-~-,lactone, 267 -, 2,3,4-tri-O-methyl-~-,lactone, 268 phenyIhydrazide, 268 -, 2,3,5-tri-O-methyl-~-,phenylhydrazide, 268 Rhamnopyranoside, methyl 2-0-methylL-, 266 I~harnnopyranosyluniine, 2,3-di-0methyl-N-phenyl-~.,267 Hhamnose, L - , 64, 67, 174 of aspen Nood, 287 henzoylhydrazone, 287 methyl ethers of. See the tables on pages 266 lo 268.

monohydrate, 77 -, 2,3-di-O-methyl-r-, 267 1,5-dit)enzyl ether, 267 -, 2,4-di -@methyl -I,-, 267 - , 3,4-di-O-methyl-r , 267 1,2-(methyl orthoacetate), 267

427

-, 2-O-methyl-1~-,266 -, 3-O-methyl-~-,266 phenylosazone, 266 hydrate, 266 -, 4-O-methyl-~-,266 phenylhydrazone, 266 phenylosazone, 266 triacetate, 266 -, 5-O-methyl-~-,267 2,3-0-methylene acetal, 267 phenylhydrazone, 267 phenylosazone, 267 triacetate, 267 -, 1,2,3,4-tetra-O-acetyl-~-, 218 -, 2,3,4-tri-O-methyl-~-,267 Rhamnoside, methyl 5-0-benzoyl-2,3di-0-methyl-L-, 267 -, methyl 2,3-di-O-methyl-a-~-,267 -, methyl 4-O-methyl-a-~-,267 p anomer, 267 -, methyl 5-O-methyl-a-~-,267 -, methyl 2,3,4-tri-O-methyl-a-~-, 268 @ anomer, 268 Rhamnosylamine, L-, with ethanol of crystallization, 157 with methanol of crystallization, 157 -, N-n-butyl-L-, 157 -, N-p-carbethoxyphenyl-L-,157 -, N-o-carboxyphenyl-L-, 98, 157 --, N-p-carboxyphenyl-L-, 157 -, N,N-dimethyl-L-, isopropylidene acetal, 157 -, N-(4,5-dimethyl-2-nitrophenyl) -L-, triacetate, forms I and 11, 157 -, N-ethyl-L-, 157 -, N-1-heptyl-L-, 157 -, N-1-hexyl-L-, 157 -, N-l-(2-hydroxynaphtliyl)-N-phenylmethyl-L-, 157 -, N-methyl-L-, 157 -, N-(4-methyl-2-nitrophenyl)-~-, 157 triacetate, 157 -, N-m-nitrophenyl-L-, 157 -, N-o-nitrophenyl-I,-, 157 triilcet:rte, 157 -, N-p-nitrophenyl-L-, 157 triacetate, 157 -, N-I-pentyl-I-, 157 -, K-pheny-L-, 168 2,3-dimethyl ether, 158

428

SUBJECT INDEX

2,4-dimethyl ether, 158, 267 I) , phosphates, chromatography of, 76 2-methyl ether, 266 D-, separation from o-arabinose, 124 2,3,4-trimethyl ether, 158, 268 DL-, 72 -, N-4-phenylazophenylene-L-, 158 DL-, p-tolylsulfonylhydrazone,72 -, N-n-propyl-L-, 158 -, 3,5-di-O-benzoyl-~-,1,2-0rthobenzoate, 239 -, N - p sulfamylphenyl-L-, 158 -, N-p-sulfophenyl-L-, 158 -, 2,3-di-O-methyl-o-, 265 -, N-p-tolyl-L-, 158 -, 3,5-di-O-methyl-o-, phenylosazone, Rhamnosyl bromide, 2,3,4-tri-O-acetyl265 a - ~218, , 230, 253 -, 1,2-0-isopropylidene 3,5-di-O-tosyl-, 2,3,4-tri-O-benzoyl-a-~-, 253 I ) - , 35 Rhamnosyl chloride, 2,3,4-tri-O-acetyl- -, 5-O-methyl-o-, 265 W L - , 248 p-bromophenylosazone, 265 -, 2,3,4-tri-O-benzoj.l-a-~-, 248 -, 1,2,3,4-tetra-O-acetyl-o-, 112 Rhamnosyl halides, tri-0-acetyl-L-, 232 -, 1,2,3,5-tetra 0-acetyl-D-, 211, 212 Rhamnosyl iodide, 2,3,4-tri-O-henzoyl- -, 2,3,4,5-tetra-O-acetyI-aldehydo-D-, a-L-,256 100 Rhaninus jranguln, 94 -, 1 , 2 , 3 , 4 tetra-0-benzoyl-p-D-, 220 Ribitol, 66 --, 2,3,5-tri-O-benzoyl-13-~-, 239 -, 1-amino-1-deoxy-, 173 -, 2,3,4-tri-O-rnethyl-o-, 265 -, 1-amino-1-deoxy-D-, 206 -, 2,3,5-tri-O-methyl-~-,265 -, 1-amino-1-deoxy-L-, 205 Rihoside, methyl 2,3-anhpdro-4-0-, 1-deoxy-1- (3,4-dimethylanilino) -D-, methj l - p - ~ . , 265 171 -, methyl 2,3,4-tri-O-methj~l-o-, 265 -, l-deoxy-l-(3,4-dimethylanilino)-~-,-, methyl 2,3,5-tri 0-methyl-D-. 265 Rihosylamine, D-, 159 192 Riboflavine, preparation of, 124, 171 -, N - a r y h - , 98 Ribofuranose, 2,3-0-isopropylidene-~-,-, 2-deoxy-N-phenyl-1-, 160 34 - , 2-deoxy-N-phenyl-~-,161 derivatives of, 32 -, 2-deoxy-N-p-tolyl-~-,160 tosylation of, 34 -, 2,3-di-O-acetyl-N-(2-amino-4,5-diRibonic acid, D-, amide, 77 methylphenyl)-5-O-trityl-~-, 159 -, 2,3,4-tri-O-methyl-o-, 265 -, N-(4,5-dimethyl-2-nitrophenyl) -D , 159 lactone, 265 -, 2,3,5-tri-O-methyI-~-, 265 triacetate, 159 tritylation of, 116 lactone, 265 5-trityl ether, 159 phenylhydrazide, 265 2,3-diacetate, 159 Ribonuclease, pancreatic, 360 Ilihopyranosylamine, N-p-carl)oxy- -, N-(3,4-dimethylphenyl)-o-, 150 p heny 1-a-o-, 159 isomers A and B, 159 -, N-o-chlorophenyl-a-D-, 159 -, N-ethyl N-2-nitrophenyl-~-,159 -, N-(4,5-diethyl-2-nitrophenyl)-~-, 159 -, N - (4-ethyl-2-nitrophenyl)-I)-, 159 -, N-(3-hydroxy-4-methylphenyl)-~-, -, N-o-nitrophenyl-I)-, 159 isomers A arid B, 159 159 -, N -4-methoxyphenyl - a - ~ -159 , -, N-phenyl-D , 109 -, N-l-Ilaphthyl-ol-I>-,159 acetvlation of, 111 -, N-2-naphthyl-n-, 359 attempted methylation of, 110 comple\ with sodium sulfate, 124 -, N-phenyl-i)-, acet:itc, 101 infrared absorption spectrum, 102 Rihose, D - , 66 isomer A, 159, 160 D - , condensation with amines, 124

SUBJECT INDEX

isomer B, hemihydrate, 160 isomeric forms, 98 triacetate, 160 2,3,5-trimethyl ether, 160,265 tritylation of, 116 -, N-p-tOlyl-D-, 160 hemihydrate, 160 with 2 ethanol of crystallization, 160 -, 2,3,4-tri-O-acetyl-N- (4,5-dimethyl2-thioformamidophenyl)-~-,159 Ribosyl bromide, 2,3,4-tri-O-acetyI-pD-, 253 -, 2,3,4-tri-O-acetyl-~-,253 -, 2,3,5-tri-O-acety1-11-,210, 253 LY anomer, 243, 244, 245 -, 2,3,4-tri-O-benzoyl-a-~-, 237,253 p anomer, 220, 237, 253 -, 2,3,5-tri-O-benzoyl-~-,239 Ribosyl chloride, 3,4-di-O-acetyl-2-deOXY-D-, 245, 247 -, 2,3,4-tri-O-acetyl-p-o-, 248 -, 2,3,5-tri-O-acetyl-~-,212 a anomer, 248 -, 2,3,4-tri-O-bensoyl-a-~-, 220,237,248 p anomer, 220, 237, 248 Ribosyl halides, poly-0-acyl-, 208 Robinobiosyl chloride, hem-0-acetyla-,249 Rutin, 94 Itutinosyl chloride, hem-0-acetyl-a-, 249 S

Saccharic acid. See Glucaric acid, D - . Saccharin, tetra-O-acetyl-I)-glucosyl-,137 Saccharinic acid, 184 in pulping waste liquors, 326 Sacchnrorrryces ba!lanus, 294 Salep, mannan of, 288,290 Salicin, 67 Salicylic acid, 4-amino-, 125 Saponins, 209 of wood, 70 Sapote, gum, 79 Sapwood, of oak, 301, 302 Scots pine, hemicelluloses of, 64 Sedoheptulosan, 76 Sedohept.ulose,73,76 Selenions acid, act,ion on D-fructose, 187 Seliwanoff reagent, 193

429

Seliwanoff test, for ketose, 201 Serine, acyl derivatives, periodate oxidation of, 117 -, N-D-glucOSJ’l-DL-,150 Sheep, polysaccharide from riimen of, 90 Shock, anaphylactic, 337 Silene EF, 77, 78, 83, 88 Silica, f o r column chromatography, 89 Silicic acid, 80, 81, 86, 94 for column chromatography, 93 Silicon tetrachloride, 219 Silver chloride, activated, 210, 219, 220 Silver fluoride, 221 Slash pine, alpha-cellulose of, 295, 296 holocellulose of, 290 mannan of, 290, 296 Slippery elm, mucilage, 72 Soda process, for pulping wood, 321, 325 Sodium nitroprusside, 195 Sodium sulfate, complexes with glycosylamines, 124 Softwoods, hemicelluloses of, 302 holocellulose of, 325 sulfate pulps from, 327, 328 sulfite pulps from, 325, 328 Sophorose, 87 Sorbitol. See Glucitol, D Sorbose, L-, 66, 77, 175 bimolecular dianhydrides of, 63 reaction of, with aliphatic amines, 100 with p-phenetidine, 98 -, 6-deoxy-~-,73 Sorbosylamine, N-p-ethosyphenyl-I,-, 161 -, N-phcnyl-L-, 124 Sorbosyl chloride, 1,3,4,5 tetra-0)-acetyla - ~248 , Spruce, alkaline pulping of, 326 alpha-cellulose of, 328 black, 319, 320 ’ hemicelluloses o f , 64, 313 holocellulose of, 296 mannan in, 296 pectic materiel from, 313 hemicellulose of, 290 holocellulose of, 314 mannan of, 290 pulp from, 327 sulfite cooking of, 307,322

430

SUBJECT INDEX

sulfite pulp from, 310,323,325,331 wood, 318 Stachyose, 66,75 preparation of, 63 structure of, 91 Stannic chloride, 219 Starch, as adsorbent in column chromatography, 74,75 banana, 89 corn, fractionation of, 70 degradation of, 223 fractionation of, by column chromatography, 79 highly polymerized, 346 hydrolysis products from, 86 separation of, by column chromatography, 63 oxidation of, in the presence of ascorbic acid, 359 potato, 223 fractionation of, 70 rice, 89,223 structure of, 224 -, 6-deoxy-6-iodo-2,3-di-O-tosyl-, 224 -, tri-0-methyl-, 224 -, tri -0-tosyl-, 224 Sterculia setigera, gum, 71 Steric hindrance, 237 Straw, of wheat, 90 Styracitol. See Mannitol, 1,5-anhydroD-.

“Substance X,” from hemicelluloses, 287 Succinamide, N - (tetra-0-acetyl-D-gluco~ y l ) - ,150 Succinic acid, 66 Succinimide, N-o-glucosyl-, 150 dihydrate, 150 Sucrose, 64,66, 76, 77, 80 azoate, 80 octaacetate, 88 chemical synthesis of, 87,241 synthesis of, 241 x-ray study of, 13 Sugar acids, column chromatography of, 65 Sugar beets, aralmn of, 90 Sugars, alkyl orthoesters of, reaction of, with hydrogen halides, 227 with tit nriium tetrachloride, 227 aiihgdro derivatives of, 208

arylhydraxones of, 99 azoates. See table of sugar azoates suitable f o r chromatography, on page 80.

complexes with borate ion, 75 deoxyseleno derivatives, 208 deoxythio derivatives, 208 diazo derivatives, 87 effect of boric acid on electric conductivity of, 231 invert, 69 methyl ethers, characterization of, 124 mutarotation of, 106, 107 orthoesters of, 208 partially methylated, azoates of, 89 p-phenylazobenzoates, 79 raw beet, raffinose in, 74 reducing, reaction with ammonia, 106 relative affinity of, for carbon, 59 Sulfamic acid, 354 -, cyclohexyl-. See Cyclohexanesulfamic acid. Sulfapyridine, condensation with sugars,

125

-,

N-D-galactosyl-, 134 -, N-D-glUCOSyl-, 150 Sulfate cook, of sulfite pulp, 325 Sulfate process, for pulping wood, 321,

326 sulfate liquor for, 326 Sulfation, of polysaccharides, 361 Sulfite cooking, removal of carbohydrates during, 332 Sulfite process, for pulping wood, 321,

322 Sulfite pulp, mnnnan content of, 325

T Tagntose, D-, 71, 73 -, 1 -deoxy-1-(p-to1uidino)-u-, 205 Taka-diastase, 64 action on oak hemicelluloses, 301, 302 Talitol, u - , 66 -, 1 , B : 2,4:5,G-tri-O-l)eiizylidciir-i~-,38 -, 1 , 3 :2,4:5,6-tri-0-mrt hylene-I)-, 38 Talopyranose, 1,G:2,3-dianhydro-p-~-, 51

Talose, 1 ,G:2,3-cliatilri~dro-P-u-,52 Tnlose, 1,6:3,4-dianhydro-p-~-,51, 52

SUBJECT INDEX

43 1

Taloside, methyl 2,3-unhydro-4,6-0itcetylcellobiosyl bromide, 113 bcneylidene-D-, 52 on hepta-0-acetyllactosyl bromide, 01 anomer, 51, 93 113 p anomer, 51 quaternary ammonium salts from, 114 Talosyl bromide, 2,3,4,6-tetra-O-acetyl- Triose, phosphate, 73 D-, 253 Triose-reduct one, 188 Tartaric acid, 314 s-Trioxane, 15 dextro-, 66 Triphenylmethyl ethers. See under reTextiles, fibers for, 323 spective tri tyl ethers. Theophylline, silver salt, 228, 229, 245 s-Trithiane, 15 -, D-arahinofuranosyl-, 244 -, 2,4,6-trimcthyl-, 15, 31 -, (2-deoxy-o-glucopyranosy~)-,229 Y‘rilicum repens L., 91 -, (2-deoxy-~-ribopyranosyl)-,229 Triulose, 1 deoxy-1-(N-met hylnnilino) -, 7-(3,4-di-O-acetyl-2-deoxy-or,p-~3-phenyl-, 204 ribosy1)-, 245 hydrochloride, 204 -, D-xylofuranosyl-, 244 semicarbaxone, 204 Thiapyran, 2,3-dichloro-tetrahydro-, 235 Tryptophan, 360 Threitol, 1 , 3:2,4-di -0-methylene-w , 19 Turanose, 66 Threonine, 360 Turanosyl bromide, hepta-0-acetyl-p-, Thrombin, 359 254 Thrombokinase, 359 Turanosyl chloride, hepta-0-acetyl-p-, 249 Thrombosis, 359 Thymine, nucleosides of, 243 Turanosyl iodide, hepta-0-acetpl-, 256 Tires, cord for, 323 Tyrosine, 360 Titanium tetrabromide, 218, 228 Titanium tetrachloride, 218,222, 227,228 U Titanium trichloride, 188 0-Toluidine, reaction with o-glucose, 176 U h u s julva. See Slippery elm. Ultraviolet absorption spectra, 186 p-Toluidine, 99, 200 action of, on 2,3,4,6-tetra-o-acetyl-n-Unibilicaria pustulata, 72 sugars of, 64 glucose, 113 See Arabitol, 3-0-(p-~-galacUmbilicin. on 2,3,4,6-tetra-0-acetyl-01-~-glucotopyranosy1)-D-. syl bromide, 113 Urea, N,N’-di-(L-arabinosyl)-, 128 condensation with aldose methyl hexabenzoate, 128 ethers, 110 -, N , N’-di-(D-gIucosyl) -, 136 reaction with D-glucose, 97, 170, 175 octaacetate, 136 ultraviolet absorption spectrum of, octabcnzoate, 136 186 -, N , N’-di- (D-mannosyl) -,154 -, N-formyl-, 190 -, N,N’-di- (D-xylosyl) -, 162 -, N-methyl-, 196 Uridine, synthesis of, 243 Toluidine Blue, 339, 344, 365 Uronic acids, associated with cellulose, Tosylation-iodination, of starch, 224 284 Transglycosidation, 218, 219 column chromatography of, 94 Transglycosylation, of aldosylamines, 99 determination of, 286 Trehalose, 66, 76 in cellulose and wood, 314, 315 01,01-, 64 2-furaldehyde from, 304 azoate, 80 of holocellulose, 319 B,B-, 87 moieties of, in wood, 284, 286 azoate, 80 monomethyl ether, 306 Trimethylamine, action of, on hept,a-0-

432

SUBJECT INDEX

in wood hemicelluloses, 313 of wood pulps, 327 Uronic anhydrides, of alpha-cellulosc, 320 of holocelluloses, 319 in pulps, 328 of woods, 285

V Vaccznium n2 yrtzlliis, 93 Vrgetahle ivory, mannan of, 288 Viscose, process for, 311 rayon, 284 effect of mannan and sylan on tensile strength of, 297 laundering resistance of, 311 wood pulps for, 311 Viscosity, intrinsic, of cellulose acetate, 297 Viscosity ratio, the term, 297 Vitamin B,? , 244

W Walden inversion, 230, 234 of glycosyl halides, 210 Western hemlock, holocellulosc from, 312 pectic acid from, 313 Wheat, 324 Wohl degradation, 123 Wood, anatomy of, 321 ash, 286 aspen, 286, 303 hydrolysis of, 287 birch, 286 hlack spruce, 285 carbohydrate composition of, 285, 326 cedar, 286 cellulose from, 284, 328 carbohydrates in, 287 preparation of, 316 characterization of, 317 commercial cellulose from, 321 coniferous, 285, 327 isolation of Carbohydrates from, 317 mannan content of, 296 deciduous, 286 delignification of, 316 determination of mannan in, 290 Douglas fir, 285

extraction with a l l d i , 318 cvtractivcs of, 284 hemicelluloses of, 313 hydrolyzates of, 292 isolation of carbohydrates from, 317 loblolly pine, 285 maple, 316 analysis of, 316 mesquite, 286 nitrated cellulosic material from, 330 oak, 286 paper birch, 318 pectic material in, 312, 313 pine, 286 polysaccharides of, 283, 328 pulps. See Pulps, wood. saponins of, 90 Southern red oak, 285 spruce, 286, 318 sulfate cook of, 307 sulfite cook of, 307 summative analysis of, 304 Western hemlock, 285 nitration of, 330 Wrstcrn red cedar, 285

X Xanthic acid, glycosyl esters, reductive desulfurization of, 245 Xanthorhnmnin, 94 X-rays, for determining crystal strncturc of a-o-glucopyranose, 231 Xylan, 284, 296, 299, 319, 320, 324 acetate, 86 acetylation of, 311 acidic, 310, 313 from birch, 303 of alpha-cellulose, 328 association of, u i t h cellulose, in wood, 307,309 with lignin, 310 from barley straw, 299 of birch, 327 from corn cobs, 300 crystalline, properties of, 299 determination of, 303 diacetate, 311 effect of different alkalis on, 308 from esparto grass, 90, 299 hemicellulosic. 311

SUBJECT INDEX

433

neutral type of, 310 -, 3-O-methyl-~-,lactone, 258 -, 2,3,4-tri-O-rnethyl-o-, lactone, 260 from paper birch, 299 phenylhydrazide, 260 partial hydrolysis of, 63 -, 2,3,5-tri-O-methyl-o-,amide, 260 from pears, 300 lactone, 260 of pine, 327 phenylhydrazide, 260 properties of, in wood cellulose, 307 resistant, 311 Xylopentaose, D-, 63 of aspenwood pulp, 308, 309 Xylopyranoside, methyl B-D-, 322 isolation of, 309 -, methyl 2-0-methyl-p-o-, 257 cf wood, 308 3,4-diacetate, 257 structure of, 299, 300 3,4-ditosylate, 257 sulfated, 361 Xylopyranosylamine, 2,3-di-O-methyl of sulfite pulps, 323 N-phenyl-D-, 258 -, 2-O-methyl-N-phenyl-~-, 257 the term, in wood chemistry, 284 -, 3-O-methyl-N-phenyl-~-D-,258 of unbleached sulfate pulps, 32i of \+heat straw, 90 Xplose, 318, 322 D-, 64, 67, 77, 173, 314 of woods, 285,299,300 determination of, 293, 306 3,4-Xylidine, condensation with uarahinose, 171 D-, fermentation of, 306 Xylitol, 66 D-, 2-furaldehyde from, 305 -, 1,3-anhydro-2,4-0-methylene-~~-, 20 W D - , tetraazoate, 80 -, l-deoxy-2,4-0-methylene-~-, 30 D - , 5-trityl ether, 173 -, l-deoxy-2,4-0-methylene-3,5-di-O- DL-, 72 tOSyl-D-, 30 -, 3,5-anhydro-1,2-0-isopropylidene-, 2,4:3,5-di-0-benzylidene-l-0-tosylu-, 20 DL-, 43 -, di-0-benzylidene-D-, dimethyl acetal, -, 2,4: 3,5-di-O-methylene-l-O-tosyl306 D L - , 43 -, di-0-benzylidene-L-, dimethyl acetal, Xylobiose, D-, 63, 300 306 X ylof uranose , 3-O-met h yl -D -, 257 -, di-0-(p-isopropylbenzy1idene)-D-,di1,2-O-isopropylidene acetal, 257 methyl acetal, 306 5-tosylate, 257 -, di-0-(p-isopropylbenzy1idene)-L-,diXyloheptaose, D-, 63 methyl acetal, 306 Xylohexaose, D - , 63, 300 -, 1,2: 3,5-di-O-isopropylidene-ol-o-, 32, Xylonic acid, compound with cadmium 36 bromide, 306 -, 2,3-di-O-methyl-a-~-,91, 258 D - , from sulfite waste liquors, 322 -, 2,4-di-O-methyl-p-n-, 259 -, 2,3-di-O-methyl-~-,amide, 259 -, 2,5-di-O-methyl-~-,259 p-bromophenylhydrazide, 259 -, 3,4-di-O-methyl-o-, 259 lactone, 259 -, 3,5-di-O-methyl-~-,260 phenylhydrazide, 259 p-bromophenylosazone, 260 -, 2,4-di-O-methyl-o-, amide, 259 1,2-O-isopropylidene acetal, 260 lactone, 259 -, hexa-0-acet yl - d d e hydo -D -, 86 phenylhydrazide, 259 -, 1,2-0-isopropylidene-3-0-methyl-5-, 3,4-di-o-methyl-o-,lactone, 259 0-tosyl-D-, 257 phenylhydraxide, 260 -, 1,2-O-isopropylidene-5-0-methyl-3-, 3,5-di-O-methyl-1-, lactone, 260 0-tosyl-D-, 258 phenylhydrazide, 260 -, 2-0-methyl-p-~-,257 -, 2-O-methyl-n-, amide, 257 triacetate, 257 -, S - o - m e t h y l - ~ ~258 -, lactone, 257

434

SUBJECT INDEX

p-hromophcnylosazone, 258 -, N-o-nitrophenyl-u-, 162 phenylosazone, 258 triacetate, 162 -, 4-O-methyl-~-,258 -, N-p-nitrophenyl-a-u-, 162 phenylosaxone, 258 p anomer, 162 -, 5-O-methpl-~-,258 -, N-phenyl-D-, 124, 163 p-bromophenylosazone, 258 2,3-dimethyl ether, 163 -, 2,3,4-tri-O-benzoyl-~-,115 2,4-dimethyl ether, 163, 259 -, 2,3,4-tri-O-methyl-o-, 260 3,4-dimethyl ether, 163 -, 2,3,5-tri - O - m e t h y - ~ -260 , 2-methyl ether, 163 Xyloside, methyl 2,3-di-O-methyl-a-~-, 3-methyl ether, 163 258 triacetate, 163 p anomer, 258 2,3,4-trimethyl ether, 163,260 4-tosylate, 258 -, N-p-sulfacetamidophenyl-u-, 164 -, methyl 2,4-di-O-methyl-p-u-, 259 -, N-p-sulfamylphenyl-n-, 164 3-tosylate, 259 -, N-p-sulfophenyl-o-, 164 -, methyl 3,4-di-O-methyl-p-o-, 259 -, N-P-tolyl-D-, 164 2-tosylate, 259 tribenxoate, 115, 164 -, methyl 2,5-di-0-methyl-3-0-t osyl-a- Xylosyl bromide, 2,3,4-t ri-0-acctyl-aD-, 259 D- , 237, 253 0 anomer, 259 methanolysis of, 236 -, methyl 4-O-rnethyl-B-o-, 258 reaction with secondary amines, 239 -, methyl 5-0-methyl-3-0-tosyl-a-~-, -, 2,3,4-tri-O-acetyl-~-, 253 258 -, 2,3,4-tri-O-benxoyl-n-, 253 p anomer, 258 01 anomer, 237 -, methyl 2,3,4-tri-O-metliyl-a-~-, 260 -, 2,3,4-tri-U-l,enzoyI-~-, 253 p anomer, 260 Sylosyl chloride, 3,4-di-O-benxoyl-2-, methyl 2,3,5-tri-O-methgl-u-,260 ch~oro-2-deoxy-a-o-, 248 Xylosylamine, D-, 162 -, 2,3,4-t ri-0-acetyl-a-D-, 248 triacetate, 162 p anomer, 248 -, N-o-aminophenyl-o-, 162 Xylosyl fluoride, 2,3,4-tri-U-acetyl-atriacetate, 162 D-, 246 -, N-n-butyl-D-, 162 Xylosyl halides, tri-0-acety-D-, 232 -, N-p-carboethoxyphenyl-u-, 162 Xylosyltrimethylammonium bromide de-, N-o-carbomethoxyphenyl-D-, 162 rivative. See under Ammonium -, N-o-carboxyphenyl-D-, 162 bromide. -, N-p-carboxyphenyl-D-, 162 Xylotetraosc, D-, 63 -, N-(4-chloro-2-nitrophenyl)-~-,162 Sylotriose, D - , 63 triacetate, forms I and 11, 162 Xylulose, u-. See Pentulose, D-threo-. -, 2-deoxy-N-phenyl-~-,163, 164

, N-(4,5-dimethyl-2-nitrophenyl)-~-,

-

162 triacetate, 162 -, N - o - (1-ethoxyethylideneamin0)phenyl-D-, triacetate, 162 -, N-1-hesyl-D-, 162 -, N - (4-methyl-2-nitrophenyl) -D-, 162 triacetate, forms I and 11, 162

Y Yarns, rayon, 297 viscose, strengt,h of, 311 Teasts, 306 crystalline hexokinase of, 368 mannan of, 289, 290

ADVANCES IN CARBOHYDRATE CHEMISTRY Volume 1 C . S . HUDSON. The Fischer Cyanohydrin Synthesis and the Configurations of Higher-Carbon Sugars and Alcohols . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . NELSONK . RICHTMYER, The A1t)rose Group of Substances . . . . . . . . . . . . . . . . . . . . EUGENE PAC.SU.Carbohydrat.e Orthoesters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ALBERTI, . RAYMOND. Thio- and Seleno-Sugars. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ROBERTC . ELDERFIELD, The Carbohydrate Components of the Cardiac Glycosides . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . C . JELLEFF CARRand JOHN C . KRANTZ, JR., Metabolism of t.he Sugar Alcohols and Their Derivatives . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . R . STUARTTIPSON, The Chemistry of the Nucleic Acids . . . . . . . . . . . . . . . . . . . . . . THOMAS JOHN SCHOCH, The Fractionation of Starch . . . . . . . . . . . . . . . . . . . . . . . . . . . ROYL . WHISTLER,Preparation and Properties of Starch Esters . . . . . . . . . . . . . . . CHARLESR . FORDYCE, Cellulose Esters of Organic Acids . . . . . . . . . . . . . . . . . . . . . ERNESTANDERSONA N D LILA SANDS,A Discussion of Methods of Value in Research on Plant Polyuronides . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

1 37 77 129

147 175 193 247 279 309 329

Volume 2 C . S . HUDSON, Melezitose and Turanose . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . STANLEY PEAT, The Chemistry of Anhydro Sugars . . . . . . . . . . . . . . . . . . . . . . . . F . SMITH,Analogs of Ascorbic Acid . . . . . . . . . . . . . . . . . . . . . . . . . . R . LESPIEAU,Synthesis of Hexitols and Pentitols from Unsaturated Polyhydric Alcohols . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . HARRY J . DEUEL,JR. and MARGARET G . MOREHOUSE, The 1nterrelat)ionof Carbohydrate and F a t Metabolism . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . M . STACEY, The Chemistry of Mucopolysaccharides and Mucoproteins . . . . . . . . . TAYLOR H . EVANSand HAROLD HIBBERT,Bacterial Polysaccharides . . . . . . . . . . E . L . HIRSTand J . K . N . JONES, T h e Chemistry of Pectic Mat.erials. . . . . . . . . . EMMAJ . MCDONALD, The Polyfructosans and Difructose Anhydrides . . . . . . . . . . JOSEPH F . HASKINS,Cellulose Ethers of Industrial Significance . . . . . . . . . . . . . . .

1

37

107 119 161 203 235 253 279

Volume 3 C . S. HUDSON, Hist.orical Aspects of Emil Fischer’s Fundamental Conventions for Writing Stereo-Formulas in a Plane . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . E . G . V . PERCIVAL, The Structure and Reactivity of the Hydrazone and Osazone Derivatives of the Sugars . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . HEWITTG . FLETCHER, J R . , The Chemistry and Configuration of the Cyclitols., BURCKHARDT HELFERICH, Trityl Ethers of Carbohydrates. . . . . . . . . . . . . . . . . . . . . LOUIS SATTLER,Glutose and the Unfermentable Reducing Substances in Cane Molasses . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . JOHNW . GREEN,The Halogen Oxidation of Simple Carbohydrates, Excluding the Action of Periodic Acid . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 435

1 23 45 79 113 129

43G

ADVANCES IN CARBOHYDRATE CHEMISTRY

JACK ConirToN. The Molecular Coiist.it.ut.ionof Cellulose .

. . . . . . . . . . . . . . .

SAMUEL OURIN. 1sot.opic Tracers in the Study of C:tabohydrate htet.abolism . . KARL~ Z Y R B A C K . Products of the Enzymic Degradation of Starch and Glycogen . S. rAcm and P . . KENT.The Polysaccharides of MycoDacteriirni tuberculosis. It . 17 . 1,E M I E U X :tnd M . L . WOLFROM. The Chemistry of Streptomycin . . . . . .

w

185 229 251 311 337

Volume 4

IRVING LEVI A N D CLIFFORD B . PURVSS. The Structure and Configuration of Sucrose (alpha-D-GlucoIJyrnnosyl Deta-D- Fructofuranoside) . . . . . . . . . . . . . . 1 H . G . BRAYand M . STACEY. Blood Group Polys.iccharides . . . . . . . . . . . . . . . . . 37 C . S. HUDSON. Apiose and the Glycosides of t.he Parsley Plant . . . . . . . . . . . 57 CARLNEUBERG, Diochemical Reductions a t t.he Expense of Sugars . . . . . . . . . . 75 VPNABCIODEIJLOKEU, Thc Acylated Nit.riles of Alclonic Acids and Their Degra119 dation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ELWINE . HARRIS.Wood Saccharificat.ion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 153 J . BBESEKEN. The Use of Boric Acid for the Determination of the Configuration of Carbohydrat.es . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 189 ROLLANDLOHMAR and R . M . GOEPP.JR.,The Hexit.ols and Some of Their Derivatives . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 211 J . K . N . JONES and F . S%cI.rH.Plant. Gums and Mucilages . . . . . . . . . . . . . . . 243 I. . F . WIGGINS, The Utilization of Sucrose . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 203 Volume 6 HEWITTG . FLETCHER. J R . ,and NELSON K . RICHTMYER. Applications in t.he Carbohydrate Field of Reductive Desu1furizat)ion by Raney Nickel . . . . . . . . . 1 W . Z. HASSII)and M . DOUDOROFF. Enzymatic Synthesis of Sucrose and Other 29 Disaccharides . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ALFREDGOTTSCHALK. Principles Underlying Enzyme Specificity in the Domain of Carbohydrates . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 49 Z . I . KERTESZ and R . J . MCCULLOCH. Enzymes Acting on Pectic Subst.ances. . . . 79 R . F . NICKERSON. T h e Relative Crystallinity of Celluloses . . . . . . . . . . . . . . . . . . . 103 G . R . DEANand J . B . GOTTFmED. The Commercial 1’roduct)ion of Crystalline 127 Dextrose . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . E . J . BOURNE and STANLEY PEAT,The Methyl Ethers of n-Glucose . . . . . . . . 145 L . F . WIGGINS,Anhydrides of the Pentitols and Hexitols . . . . . . . . . . . . . . . . . . 191 MARYL . CALDWELL and MILI)RED ADAMS, Action of Certain Alpha Amylases . . . 229 ROYI, . WHISTLER. Svlan . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 269 Volume 6 E . L . HIRST, Obituary of Walter Norman Haworth . . . . . . . . . . . . . . . . . . . . . . . . . . D . J . BELL,The Met.hyl Ethers of D-(falactose . . . . . . . . . . . . . . . . . . . . . . . . . W . L . EVANS. D . D . REYNOLDS arid E . A . TALLEY, The Synthesis of Oligosaccharides . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . F . H . NEWTH.The Formation of Furan Compounds from Hexoses . . . . . . . . . . . . . RICHARD E . REEVES,Cuprammonium-Glycoside Complexes . . . . . . . . . . . . . . . . . ROGERW . JEANLOZ and HEWITTG . FLETCHER. J R . , The Chemistry of Ribose . . . NELSONK . RICHTMYER. The 2-(Aldo-polyli~~droxyalkyl)benzimidazoles ....... ELLIOTTP. BARRETT, Trends in the Development. of Granular Adsorbents for Sugar Refining., . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

1

11

27 83 107 135 175 205

A DV.kNCES I N CAIEBOHYDIEATE CRISMISTRY

437

ROBERTELLSWORTH MILLERand SIDNEY M . CAN.roR. Aconit.ic Acid. a Byproduct. in the Manufacture of Sugar . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 231 WrrmAM A . RONNER. FriedelLCrafts and Grignard Processes in the Carbohydrat.e Series . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 251 JOHN C . SOWDEN. T h e Xitromethane and 2-Nitroethanol Syntheses . . . . . . . . . . 291

Volume 7

R . -4. LAIDLAW and I<;. G . V . I’ERCIVAL, T h e Methyl Ethers of the Aldopentoses rid of Rharnnose and Fucose . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

1

R . J . DIMLER, 1,6-Anhydrohexofuranoses, a New Class of Ilexosans . . . . . . . . . . . 37 C . P . BARRYand .JOHN HONEYMAN, Fructose arid I t s Derivat.ives . . . . . . . . . . . . 53 99 J . v . I
Volume 8

JAMES M . SUGIHARA, Relative Reactivit ies of Hydroxyl Groups of Carliohydrates W . G . OVERENI) and M . STACEY, The Chemistry of the 2-Desoxysugars . . . . . . . . It . STUART TIPSON, Sulfonic Esters of Carbohydrates . . . . . . . . . . . . . . . . . . . . . . . . . G . 0. ASPINALL,The Met.hyl Et.hers of u.ilZannose . . . . . . . . . . . . . . . . . . . . . . . . . . . C . 1, . MEHLTRETTER, The Chemical Synthesis of D-Glucuronic Acid . . . . . . . . . . . H . G . BRAY,1)-Glucuronic: Acid in Met.abolisrn . . . . . . . . . . . . . . . . . . . . . . . EI)WARD J . HEHRE,The Suhstit.uted-Sucrose St.ructure of Melezit.ose . . . . . . . . w . w . H I N K L E Y and hf . L . WOLFROM, Cornposit.ion of Cane Juice and &.lie Filial Molasses

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T . AM^^^^, S e a w e d Polysaccharidcs

1 45 107 217 231 251 277 291 315

Volume 9

It.. U . T.EmE:Ux, Some Implications in Carbohydrate Chemistry of Theories Relating t o t.he Mechanisms of Replacement Itcactions . . . . . . . . . . . . . . . . . . 1 59 CLINTON E . BALLOU. Alkali-Sensitive Glgcosides . . . . . . . . . . . . . . . . . . . . . . . . ~ ‘ I A RGRACE Y HIAIR.The 2-Hydroxyglycals . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . (37 G . 0 . ASI.INAI.I..The Met.hyl Ethers of Hexuronic Acids . . . . . . . . . . . . . . . . . . . . . 131 D E x r m FRENCH. The Raffiriose Family of Oligosaccharides . . . . . . . . . . . . . . . . . . RORERTS. TEAGUE, The Conjugates of I)-Glucuroiiic Acid of Animal Origin . . TT and VIc1’oR R . DYITZ.Color and Turbidity of Sugar Products . J . V . I ~ A R A B I Nant1 O S MARJORIE H I X D E R TCarbox-ymet.hylrcllulose . . . . . . . . . . GEORGE N KOWKAB ~ N Y Paper . Chromatographv of Carhohvdrates and Related Compounds

149 185 247 285

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E. G. V. PERCIVAL

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