The Alkaloids: Chemistry and Pharmacology, Volume 44

THE ALKALOIDS Chemistry and Pharmacology VOLUME 44

This Page Intentionally Left Blank

THE ALKALOIDS Chemistry and Pharmacology Edited by Geoffrey A. Cordell College ofP/itir ni~ii:v University qf Illinois (11 Chicogo C/2iCYZ~O. IIlinois

VOLUME 44

Academic Press, Inc. A Division o j Horcoiirt Brrrce & Cotnpntz>’

San Diego New York Boston London Sydney Tokyo Toronto

This book is printed on acid-free paper. @ Copyright 0 1993 by ACADEMIC PRESS, INC. All Rights Reserved. No part of this publication may be reproduced or transmitted in any form or by any means, electronic or mechanical, including photocopy, recording, or any information storage and retrieval system, without permission in writing from the publisher.

Academic Press, Inc. 525 B Street, Suite 1900, San Diego, California 92101-4495 United Kingdom Edition published by ACADEMIC PRESS LIMITED 24-28 Oval Road, London NW 1 7DX

International Standard Serial Number: 0099-9598 International Standard Book Number:

0-12-469544-2

PRINTED IN THE UNITED STATES OF AMERICA 9 3 9 4 9 5 9 6 9 7

QW

9

8

7

6

5

4

3

2

1

CONTENTS

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

CONTRIBUTORS ...... PREFACE. . . . . . . . . . . .

vii ix

Chapter I . The Tropane Alkaloids A N D TAIUATAMMINEN MAURILOUNASMAA

I. Introduction .............................. 11. Occurrence . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . I l l . Synthesis ...................................... IV. Reactions

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

I 3 78 89 92 95 97 99 I00

Chapter 2. The Biosynthesis of Tropane Alkaloids

J. ROBINSA N D NICHOLAS J. WALTON RICHARD Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Organ Tissue Cultures for Biosynthetic Studies ............... Formation of Putrescine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . FormationofTropinone ......................... . . . . . . . . . . . . . . . . . . . Formation of Tropine and Pseudotropine . Formation of Acidic Moieties of Tropeines ........................... Formation of Tropeines ........................... Metabolism of Tropeines ......................... Degradation and Oxidation of Tropeines ............................. X. Overall Regulation of Pathway ....................... XI. Future Prospects ................................ References .......................................................

I. 11. 111. IV. V. VI. VII. VIII. IX.

I16 I19 I30 134 146 151 155

160 164 168 I80 I82

Chapter 3. Simple Indolizidine Alkaloids A N D TAKEFUMI MOMOSE HIROKITAKAHATA

I. Introduction ...................................................... 11. Indoiizidines with Alkyl and Functionalized Alkyl Appendages

III. Elaeocarpus Alkaloids

.........

.............. ._............................. V

189 190 22 I

vi

CONTENTS

IV . Slaframine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . V . Hydroxylated lndolizidines . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VI . Summary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . References .......................................................

223

228 250 250

Chapter 4 . Chemistry and Biology of Carbazole Alkaloids

D . P . CHAKRABORTY I . Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . I1 . Occurrence . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 111. Chemistry of Carbazole A loids ................................... IV . Physical Properties of Carbazole Alkaloids ........................... .... V . Biogenesis of Carbazole Alkaloids ................... VI . Biochemical and Medicinal Properties of Carbazole AIka Related Compounds . . . . . . . ....................... References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ....

CUMULATIVE INDEX OF TITLES. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . INDEX . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

258 258 258 349

351 352

360

365 373

CONTRIBUTORS

Numbers in parentheses indicate the pages on which the authors’ contributions begin

D. P. CHAKRABORTY (257), Institute of Natural Products, Calcutta 700 036, India MAURILOUNASMAA (l), Laboratory for Organic and Bioorganic Chemistry, Technical University of Helsinki, SF-02150 Espoo, Finland TAKEFUMI MOMOSE(189), Faculty of Pharmaceutical Sciences, Toyama Medical and Pharmaceutical University, Toyama 930-01, Japan RICHARDJ. ROBINS( 1 15), Agricultural and Food Research Council Institute of Food Research, Norwich Laboratory, Colney, Norwich NR4 7UA, United Kingdom (189), Faculty of Pharmaceutical Sciences, Toyama HIROKITAKAHATA Medical and Pharmaceutical University, Toyama 930-01, Japan TAIUATAMMINEN (l), Laboratory for Organic and Bioorganic Chemistry, Technical University of Helsinki, SF-02 150 Espoo, Finland NICHOLAS J. WALTON(1 IS), Agricultural and Food Research Council Institute of Food Research, Norwich Laboratory, Colney, Norwich NR4 7UA, United Kingdom

vii

This Page Intentionally Left Blank

PREFACE

The scientific accomplishments of Professor Edward Leete, who contributed so much to alkaloid chemistry and biochemistry during his brilliant and illustrious scientific career, are honored in this volume by two chapters on the tropane alkaloids, an area of alkaloid research that was especially dear to him. The format for the presentation of these two chapters is new to this series. In Chapter I , Lounasmaa and Tamminen provide a complete summary of the occurrence of the more than 200 tropane alkaloids characterized and also review the synthetic pathways that have been developed for their formation. In Chapter 2, Robins and Walton describe the tremendous progress that has been made in the level of understanding of the intricate tropane alkaloid biosynthetic pathway. As is now revealed, seminally by Leete and co-workers, the biosynthesis of the tropane alkaloids, particularly the most infamous alkaloid, cocaine, is substantially more complex than originally envisaged. As details of these processes and the enzymes involved are further studied and characterized, the intense subtlety of the pathways is becoming apparent. Takahata and Momose review the simple indolizidine alkaloids in Chapter 3. These alkaloids are widely distributed in nature and some, such as castanospermine (a potent glucosidase inhibitor), have been the subject of intense synthetic and biological interest in the recent past. Finally, in Chapter 4, the isolation, synthesis, and biological responses of the carbazole alkaloids is reviewed by Chakraborty. Ranging from the simple analogs present in Murraya species to the complex derivatives found in certain fungi, the group displays a broad spectrum of structural diversity and biological activity. Geoffrey A. Cordell University of Illinois at Chicago

This Page Intentionally Left Blank

-CHAPTER 1-

THE TROPANE ALKALOIDS MAURILOUNASMAA A N D TARJA TAMM~NEN Laboratory f o r Organic and Bioorgunic Chemistry Technical University of Helsinki SF-02150 Espoo, Finland

I . Introduction ........................ 11. Occurrence

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

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

.80

....................82

E. Lansbury Synthesis ......................

K. Jung et al. Syn L. He and Brossi

I

gongteng A ..............................

............................................. .89 ...................89

IV. Reactions .......

C. Photocyanation ..

.............................................................. .91 References .....................................................................

I. Introduction The tropane alkaloids have been reviewed on six earlier occasions in this series (1-6). Although a relatively short time has passed since the 1 THE ALKALOIDS. VOL 44 Copyright CJ 1991 by Academic Pre,,. Inc All right\ of reproduction in m y form raerved

2

MAURl LOUNASMAA A N D TARJA TAMMINEN

last review (6) in 1988, the number of known structures has increased dramatically, from 151 to 200 (i.e., 203 - 3; vide infra). To bring the record up to date, a new chapter has been prepared, following the same general lines as before (6). The focus is on areas where major changes have taken place (especially in the areas of occurrence and synthesis). Sections addressing areas in which there has been little activity are quite short (e.g., the section on spectroscopy). In the following chapter, Robins and Walton (7) review biosynthesis of the tropane alkaloids; thus, our section on biosynthesis is short, despite the significant changes that have occurred in this area. The literature is covered up to June 1992. In addition, some recent articles, appearing since the completion of the original manuscript, have been added to the reference list. The tropane alkaloids are a structurally well-defined group of natural products, and the mydriatic and anesthetic properties of several of the tropane alkaloids were exploited long before the structures were elucidated (8).Although an extensive literature exists on the pharmacological properties of tropane alkaloids, these are touched on only briefly. Readers with a deeper interest in the subject are referred to other publications (9-12) and to the references given in Section VII. The common structural element of the tropane alkaloids is the azabicyclo[3.2. lloctane skeleton, and the systematic name for tropane is 8-methyl-8-azabicyclo[3.2. ]]octane (Fig. 1). Applying the uniform numbering system presented in Fig. I , most disubstituted tropane alkaloids designated as C-3, C-6 disubstituted in the literature become C-3, C-7 disubstituted. The same principle, where applicable, is applied to the C3, C-6, C-7 trisubstituted tropane alkaloids. The C-3, C-7 notation is also used where the choice between the C-3, C-6 and C-3, C-7 notation in the literature has been arbitrary. Only in cases where the determination of absolute configuration has a solid basis, and where the structure is correctly presented by the C-3, C-6 notation also in the present numbering system, has the original C-3, C-6 notation been retained. The strict applica-

Me 7

6

5

\

4

6 FIG. 1. Ring system of the tropane alkaloids.

1.

TROPANE ALKALOIDS

3

tion of the system adopted here is certainly in several cases a simplification of the real situation and should be regarded as such. Because the systematic names in the tropane series are often long and used by very few authors, the traditional nomenclature is followed here. Trivial names are used where they exist, while for other compounds a semisystematic name based on the word “tropane” (vide infra) is adopted. In Tables II-IV, however, the nomenclature is based entirely on the semisystematic names (except for dimers and the trimer), and the trivial names are given in parentheses.

11. Occurrence The tropane alkaloids mainly occur in the plant family Solanaceae, but are found as well in the families Convolvulaceae, Erythroxylaceae, Proteaceae, and Rhizophoraceae (Tables I and 11). In addition, the presence of tropane alkaloids has occasionally been indicated in the families Brassicaceae (=Cruciferae), Euphorbiaceae, and Olacaceae (see Tables I, 111, and IV) (13-15). During the preparation of our earlier review (6), we became acutely aware of the lack of reliable information on the natural occurrence of tropane alkaloids. As a means of correcting this situation, we have collected in this chapter as much information as possible on the distribution of tropane alkaloids in plants. Thus, our main effort during the preparation of the present review was focused on the occurrence of the tropane alkaloids. The results are brought together in Tables 111 and IV. The earlier claimed natural occurrence of compounds 113, 144, and 185 (vide infra) has turned out to be erroneous. To preserve the created numbering system intact, however, and to indicate the proposed structures, compounds 113,144, and 185 are nevertheless included in Table I1 (with footnote explanations); the compounds are omitted from Tables 111 and IV. No chemotaxonomic conclusions should be drawn on the grounds of this review alone, without resort to the original papers. The plant organs from which the alkaloids have been isolated are indexed in some detail, but information concerning the geographical distribution and seasonal changes in occurrence are omitted, as well as the relative proportions of the various alkaloids. Reference to the original papers is therefore still necessary. In spite of these limitations, we believe that this compendium,

TABLE I BOTANICAL CLASSIFICATION OF PLANTS CONTAINING TROPANE ALKALOIDS Dicot yledoneae Malviflorae Euphorbiales Euphorbiaceae Phyllanthus Violiflorae Tamaricales Brassicaceae ( = Cruciferae) Cochlearia Proteiflorae Proteales Proteaceae Agaslachus Bellrndena Darlingia Knightia M yrtiflorae Rhizophorales Rhizophoraceae Bruguiera Crossostylis Pellacalyx Rutiflorae Geraniales Erythrox ylaceae Erythroxylum Sect. I1 Macrocalyx Sect. I11 Rhabdoph yllum Sect. IV Leptogramme Sect. V Heterogyne Sect. VI Archerythrox ylum Sect. IX Microphyllum Sect. X Melanocladus Sect. XI Sethia Sect. XI1 Lagynocarpus Sect. XIV Coelocarpus Sect. XVI Venelia Sect. XVIl Pachylobus Santaliflorae Geraniales Olacaceae Hcisteria

TABLE I (continued) Solaniflorae Solanales Solanaceae Anthocercoideae Anthocercideae Anthocercis Anthotroche Crenidium Cyphanthera Duboisia Grammosolen Symonanthus Cestroideae Salpiglossideae Schizanthus Solanoideae Solanineae Solaneae Cyphomandra Ph ysalinae Physalis Withania Jabroseae Latua Salpichroa Datureae Datura Sect. Brugrnansia Sect. Dutra Sect. Ceratocaulis Sect. Datura Solandreae Solandra Nicandreae Nicandra Atropoideae Atropeae Atropa Hyoscyamus Mandragora Physochlaina Przewalskia Scopolia" Convolvulaceae Calystegia Colutea Conuoluulus Erycibe Euoluulus a

Including species of Arropanthe and Anisodus.

5

6

MAURl LOUNASMAA A N D TARJA TAMMINEN

covering all known plants with tropane alkaloids, will be a valuable source of information. Reference to original articles rather than reviews has been preferred. Our original intention was to cover the literature of the nineteenth century as well. However, it became evident that referring to those papers would only have caused confusion, as in many cases both the botanical and chemical identifications were doubtful. Overviews of the older literature are included in relatively recent papers on Duboisia (16,17),Datura Sect. Brugmansia (18), Atropa (19), Withania (20), Mandragora (21,22) and Erythroxylum (23).

1 . Botanical Classification The system of Dahlgren (24,25) was used for the general botanical classification (Table I). The family Solanaceae, the principal source of tropane alkaloids, has been classified according to the chemotaxonomic system of TCtCnyi (26), but incorporating the revision of the subfamily Anthocercideae by Haegi (27). Families containing only a few “tropane genera” have not been subdivided. The main principle in listing the genera and species in Table 111 was to employ one name for one taxon. A more ambitious botanical treatment was found to be impossible. Even this seemingly trivial goal was difficult to achieve in cases where the literature covered a long time period-more than 100 years for some species of the Solanaceae. A uniform scheme was also difficult to formulate when results pertaining to the same genus flourishing in different parts of the world had to be summarized. The names used in the original papers are mentioned wherever they differ from the ones employed in this text. The nomenclature was adopted from botanical monographs when available. Fortunately Datura, perhaps the most complex genus, has been the subject of a thorough analysis by Hammer et al. (28). The section Brugmansia, not included in that monograph, has been treated according to Bristol (29,30). It was impossible to treat the species D . arborea L. and D . candida (Pers.) Saff. separately, although Bristol presents them as distinct species, as an indication of the species investigated was lacking or misleading in several cases. It is very probable that the name D . arborea has been widely used in the chemical literature as a synonym for D . candida (31). The genus Atropa has been arranged according to Heltmann (32). This approach was adopted because it covers the whole genus, even though it may not be a botanically valid revision. Scopolia is mainly treated in the manner of Weinert ( 3 3 , but even in this case some questionable species outside Weinert’s treatment (S. anomala and S . paruijloru) were included in the list.

7

1. TROPANE ALKALOIDS TABLE I1 TROPANE ALKALOID STRUCTURES Alkaloid

Structure

1. 3a -Monosubstitutedtropanes The 3a-monosubstituted tropanes consist of 49 representatives. All members (1-49)are formally derived from 3a-hydroxytropane (1)or from the not yet naturally found 3ahydroxynortropane.

3a-Hydroxytropane ( = tropine) 3a-Acetoxynortropane 3a-Acetox ytropane 3a-Propiony lox ytropane

3a-(Hydroxyacetoxy) tropane 3a-Tigloylox ynortropane

3a-Butyryloxytropane 3a-Isobutyryloxytropane ( = butropine) 3a-Isovalerylox ynortropane ( = poroidine) 10 3a-(2’-Methylbutyryloxy)nortropane ( = isoporoidine) 11 3a-Tigloylox ytropane

12 3a-Senecioylox ytropane

13 ( + )-3a-(Z’-Methylbutyryloxy)tropane ( = valtropine) 14 3a-Isovalerylox ytropane 15 3a-Benzoyloxynortropane 16 3a-(Z’-Furoyloxy)tropane 17 3a-Tigloyloxytropane N-oxide 18 3a-Benzoyloxytropane 19 3a-Phenylacetoxynortropane 20 3a-Apotropoyloxynortropane ( = aponoratropine) 21 3a-Cinnamoyloxynortropane 22 3a-Phenylacetoxytropane 23 3a-(3’-Hydroxybenzoyloxy)tropane ( = cochlearine) 24 3a-Apotropoyloxytropane ( = apoatropine) 25 3a-Cinnamoyloxytropane

R = Me, R, = H R = H, R, = acetyl R = Me, R, = acetyl R = Me, R, = propionyl R = Me, R, = hydroxyacetyl R = H, R , = tigloyl R = Me, R, = butyryl R = Me, R, = isobutyryl R = H, R, = isovaleryl R

=

H, R,

=

2-methylbutyryl

R = Me, R, = tigloyl R = Me, R, = senecioyl R = Me, R, = 2-methylbutyryl R = Me, R , = isovaleryl R = H, R, = benzoyl R = Me, R , = furoyl R = Me, 0, R, = tigloyl R = Me, R , = benzoyl R = H, R, = phenylacetyl R = H, R, = apotropoyl R = H, R, = cinnamoyl R = Me, R, = phenylacetyl R = Me, R, = 3-hydroxybenzoyl R

=

Me, R ,

R

=

Me, R, = cinnamoyl

=

apotropoyl

(continued)

8

MAURI LOUNASMAA A N D T A N A TAMMINEN

TABLE I1 (continued) Alkaloid 26 ( -)-3a-(1’,2’-Dithiolane-3’27

28 29

30 31 32 33 34 35

36 37

38 39

40

carbony1oxy)tropane ( = brugine) ( ~)-3a-Tropoyloxynortropane ( = noratropine) ( - )-3a-Tropoyloxynortropane ( = norhyoscyamine) 3a-(4‘-Methoxybenzoyloxy)tropane ( = datumetine) 3a-(3’-Hydroxyphenylacetoxy)tropane 3a-(4’-Hydroxyphenylacetoxy)tropane 3a-Vanilloyloxynortropane ( = convolidine) (?)-3a-Tropoyloxytropane (=atropine) ( - )-3a-Tropoyloxytropane ( = hyoscyamine) ( -)-3a-(2’-Hydroxy-3’phenylpropiony1oxy)tropane ( = littorine) 3a-Vanilloyloxytropane ( = phyllalbine) 3a-Veratroyloxynortropane ( = convolvine) 3a-Tropoyloxytropane N-oxide I ( = hyoscyamine N-oxide 1) 3a-Tropoyloxytropane N-oxide 2 ( = hvoscvamine N-oxide 2) . 3a-Veratroyloxytropane( = convolamine)

41 3a-Veratroylox y-N-hydroxynortropane

Structure

1,2-dithiolane-3-carbonyl

R

=

Me, R,

R

=

H, R ,

=

tropoyl

R

=

H, R,

=

tropoyl

R

=

Me, R,

=

4-methoxybenzoyl

=

R = Me, R, = 3-hydroxyphenylacetyl R = Me, R, = 4-hydroxyphenylacetyl R = H, R, = vanilloyl

R = Me, R, R = Me, R,

tropoyl tropoyl

= =

R = Me, R, = 2-hydroxy-3phen ylpropion yl R = Me, R, = vanilloyl R = H, R, = veratroyl R

=

Me, 0, R ,

R

=

Me, 0, R, = tropoyl

R = Me, R , R = OH, R,

= =

=

tropoyl

veratroyl veratroyl

( = convoline)

42 3a-Feruloyloxytropane 43 3a-Veratroyloxy-N-formylnortropane ( = confoline)

44 3a-Veratroyloxytropane N-oxide

R R

= =

Me, R , = feruloyl CHO, R , = veratroyl

R = Me, 0, R, = veratroyl

( = convolamine N-oxide)

45 30-(3’,4‘,5’-

46 47 48 49

Trirnethoxybenzoy1oxy)nortropane ( c )-3a-Veratroyloxy-Nisopropylnortropane ( = convosine) (?)-3a-Veratroyloxy-Nacetylnortropane ( = convolicine) 3a-(3’,4’,5’-Trimethoxy benzoy1oxy)tropane 3a-(3’,4‘,5‘-Trimethoxy cinnamoy1oxy)tropane

R

=

H, R,

=

3,4,5-trimethoxybenzoyl

R = i-Pr, R, = veratroyl R

=

acetyl, R, = veratroyl

R

=

Me, R,

R

= Me, R, = 3,4,5trimethox ycinnamoyl

=

3,4,5-trimethoxybenzoyl

I.

9

TROPANE ALKALOIDS

TABLE I1 (continued) Alkaloid

Structure

2. 3p-Monosubstitutedtropanes The 3P-monosubstituted tropanes comprise 3p-hydroxytropane (SO), its five naturally occurring ester derivatives (51,52,53, 55, and 56), and the nortropane derivative 3Pbenzoyloxynortropane (54).

50 51 52 53 54 55 56

3P-Hydroxytropane 3P-Acetoxytropane 3P-Tigloyloxytropane ( = tigloidine)

3P-(2’-Methylbutyryloxy)tropane 3P-Benzoyloxynortropane 3P-Benzoyloxytropane ( = tropacocaine) 3P-Cinnamoyloxytropane

R = Me, R, = H R = Me, R, = acetyl R = Me, R , = tigloyl R = Me, R, = 2-methylbutyryl R = H , R, = benzoyl R = Me, R, = benzoyl R = Me, R, = cinnamoyl

3. 3n,6/3- and 3a,7fi-disubstitutedtropanes The base compound of the large group of disubstituted tropanes (63 representatives) is 3a,7P-dihydroxytropane (57). All other compounds (58-119) are mono- or diesters of 57 or of the corresponding 3a,6P-derivative.

R

57 3a,7P-Dihydroxytropane 58 3a-Acetoxy-7P-hydroxynortropane 59 ( + )-3a-Acetoxy-7P-hydroxytropane 60 3a-Hydroxy-7P-acetoxytropane 61 3a-Hydroxy-7P-propionyloxytropane

R = Me, R, = Rz = H R = H, R , = acetyl, Rz = H R = Me, R, = acetyl, Rz = H R = Me, R , = H. Rz = acetyl R = Me, R, = H, R2 = propionyl

(continued )

10

MAURl LOUNASMAA A N D TARlA TAMMINEN

TABLE I1 (continued) Alkaloid 62 (

+ )-3a-Hydroxy-7P-

Structure

R = H, R,

=

H, R2

=

tigloyl

tigloyloxynortropane 63 64 65 66 67 68 69 70

3a-Tigloylox y-7P-h ydrox ynortropane

3a-Isobutyryloxy-7~-hydroxytropane 3a-H ydrox y -7P-isobutyrylox ytropane

3a-Hydroxy-7~-tigloyloxytropane 3a-Hydroxy-7~-angeloyloxytropane 3a-Tigloyloxy-7~-hydroxytropane 3a-Senecioyloxy-7~-hydroxytropane ( - )-3a-Hydroxy-7P-(Z'methylbutyry1oxy)tropane 3a-Isovaleryloxy-6~-hydroxytropane 3a,7P-Diacetoxytropane 3a-Benzoyloxy-7f3-hydroxynortropane 3a-Hydroxy-7~-benzoyloxytropane 3a-Benzoyloxy-7~-hydroxytropane

71 72 73 74 75 76 3a-Phen ylacetox y-7P77 78 79 80 81 82 83 84 85 86 87

88 89 90 91

R R R R R R R R

= = = = = = = =

H, R, = tigloyl, R2 = H Me, R, = isobutyryl, R2 = H Me, R, = H, R2 = isobutyryl Me, R, = H , R2 = tigloyl Me, R, = H , R2 = angeloyl Me, R, = tigloyl, R2 = H Me, R, = senecioyl, R2 = H Me, R, = H, R2 = 2-methylbutyryl

R R R R R R

= = = = = =

Me, R, = isovaleryl, R2 = H Me, R, = R, = acetyl H, R, = benzoyl, R2 = H Me, R, = R2 = H, benzoyl Me, R, = benzoyl, R2 = H H, R, = phenylacetyl, R2 = H

hydrox ynortropane 3a-Acetoxy-7~-isobutyryloxytropane R R 3a-lsobutyryloxy-7~-acetoxytropane 3a-Cinnamoyloxy-7~-hydroxynortropaneR 3a-Phenylacetoxy-7~-hydroxytropane R R 3a-Hydroxy-7P-phen ylacetoxytropane R 3a-Tigloylox y-7P-acetox ytropane 3a-Cinnamoyloxy-7P-hydroxytropane R 3a-Apotropoyloxy-7~-hydroxytropane R 3a-Tigloyloxy-7~-propionyloxytropane R R 3a-Benzoyloxy-7~-acetoxytropane ( - )-3a-Tropoyloxy-6~-hydroxytropane R [ = ( - )-anisodaminel (~)-3a-Tropoyloxy-6~-hydroxytropane R ( = 6P-hydroxyatropine) R 3a-Tropoyloxy-7f3-hydroxytropane 3a-Tigloyloxy-7~-isobutyryloxytropane R R 3a-Phenylacetox y-7P-acetox ytropane

92 3a-Acetoxy-7P-phenylacetoxytropane 93 3a.7P-Ditigloyloxytropane N94 3a-Tropoyloxy-6f3-hydroxytropane

= Me, R, = acetyl, R2 = isobutyryl = Me, R, = isobutyryl, R2 = acetyl = H, R, = cinnamoyl, R2 = H = Me, R, = phenylacetyl, R2 = H = Me, R , = H, R2.~ = phenylacetyl = Me, R, = tigloyl, R2 = acetyl = Me, R, = cinnamoyl, R2 = H = Me, R, = apotropoyl, R2 = H = Me, R, = tigloyl, Rz = propionyl = Me, R, = benzoyl, R, = acetyl = Me, R, = tropoyl, R2 = H =

Me, R,

=

tropoyl, R2 = H

Me, R , = tropoyl, R2 = H Me, R, = tigloyl, Rz = isobutyryl = Me, R, = phenylacetyl, R, = acetyl R = Me, R, = acetyl, Rz = phenylacetyl R = Me, R, = R2 = tigloyl R = Me, 0, R , = tropoyl, R2 = H = =

oxide (=6P-hydroxyhyoscyamine N oxide) 95 3a-Tigloyloxy-7~-(2'-methylbutyryloxy)R = Me, R, = tigloyl, meth ylbutyryl tropane

R2 =

2-

1.

I1

TROPANE ALKALOIDS

TABLE I1 (continued) Alkaloid 96 3a-Tigloyloxy-7~-isovaleryloxytropane R

97 3a-Cinnamoyloxy-7~-acetoxytropane R 98 3a-(3’ ,4’ ,5’-Trimethoxybenzoyloxy)-7~-R

Structure = Me, R, = tigloyl, R2 = isovaleryl = Me, R, = cinnamoyl, R2 = acetyl = H, R, = 3,4,5-trimethoxybenzoyl,

R2 = H R = Me, R, = tropoyl, R2 = acetyl R = Me, R, = 3,4,5-trimethoxybenzoyl, 100 ( + )-3a-(3’,4’,5’-Trimethoxybenzoyloxy)R2 = H 7P-hydroxytropane R = Me, R, = 4-methoxyphenylacetyl, 101 3a-(4’-Methoxyphenylacetoxy)-7PR2 = H hydroxytropane ( = physochlaine) 102 3a-(Pyrrolyl-2‘-carbonyloxy)-7~-(N”- R = Me, R, = pyrrolyl-2-carbonyl, R2 = N-methylpyrrolyl-2-carbonyl meth ylpyrrol yl-2”-carbon ylox y) tropane ( = catuabine C) R = Me, R, = methylmesaconyl, 103 3a-Methylmesaconyloxy-7PR2 = tigloyl tigloyloxytropane ( = schizanthine F) R = Me, R , = methylitaconyl, 104 3a-Methylitaconyloxy-7/3R2 = tigloyl tigloyloxytropane ( = schizanthine G ) R = Me, R , = methylmesaconyl, 105 3a-Methylmesaconyloxy-7PR2 = angeloyl angeloyloxytropane ( = schizanthine I) R = Me, R , = methylitaconyl, 106 3a-Methylitaconyloxy-7PR2 = angeloyl angeloyloxytropane ( = schizanthine H) R = Me, R, = R2 = benzoyl 107 3a,7P-Dibenzoyloxytropane 108 3a-(3‘-Ethoxycarbonylmethacryloyloxy )- R = Me, R, = 3ethox ycarbony Imethacryloyl, 7P-senecioylox ytropane R2 = senecioyl ( = schizanthine A) R = Me, R, = ethylitaconyl, 109 3a-Ethylitaconyloxy-7~R2 = angeloyl angeloyloxytropane ( = schizanthine L) R = Me, R, = ethylmesaconyl, 110 3a-Ethylmesaconylox y-7PR2 = tigloyl tigloyloxytropane ( = schizanthine K) R = Me, R, = ethylitaconyl, 111 3a-Ethylitaconyloxy-7~R2 = tigloyl tigloyloxytropane ( = schizanthine M) 1 U 3a-Tropoyloxy-7~-tigloyloxytropane R = Me, R, = tropoyl, R2 = tigloyl 113“ 3a-Acetyltropoyloxy-6~-acetoxytropaneR = Me, R, = acetyltropoyl, R2 = acetyl ( = 6P-hydroxyhyoscyamine diacetate) 114 3a-Tropoyloxy-7~-isovaleryloxytropane R = Me, R, = tropoyl, R2 = isovaleryl R = Me, R, = tropoyl, R2 = 2115 3a-Tropoyloxy-7P-(2’methylbutyryl methylbutyry1oxy)tropane R = Me, R, = R2 = cinnamoyl 116 3a,7P-Dicinnamoyloxytropane R = Me, R, = 3,4,5-trimethoxybenzoyl, 117 3a-(3’,4’,5‘-Trimethoxybenzoyloxy)-7PR2 = benzoyl benzoyloxytropane ( = catuabine B) 118 3a-(3’,4‘,5’-Trimethoxybenzoyloxy)-7P- R = Me, R, = 3,4,5-trimethoxybenzoyl, R2 = N-methylpyrrolyl-2-carbonyl (N“-methylpyrrolyl-2”-carbonyloxy) tropane ( = catuabine A) 119 3a-(3’,4’,5’-Trimethoxycinnamoyloxy)- R = Me, R, = 3,4,5trimethoxycinnamoyl, R2 = benzoyl 7P-benzoylox ytropane

h ydroxynortropane 99 3a-Tropoyloxy-7~-acetoxytropane

(continued)

12

MAURl LOUNASMAA A N D TARJA TAMMINEN

TABLE I1 (continued) Alkaloid

Structure

4. 3a,6P,7P-Trisubstituted tropanes The 3a,6P,7P-trisubstituted tropanes (120-135) are formally derived from the recently naturally found 3a,6P,7P-trihydroxytropane(120) (or from the corresponding nortropane).

120 3a,6P,7/3-Trihydroxytropane U 1 3a,7P-Dihydroxy-6P-tigloyloxytropane 122 3a-Tigloyloxy-6P,7P-dihydroxytropane ( = meteloidine)

R R R

127

128 129

130 131 132

133 134 135

= =

Me, R, = R2 = R, = H Me, R, = R, = H, R, = tigloyl Me, R, = tigloyl, R2 = R, = H

Me, R, = benzoyl, R, = R, = H Me, R, = R2 = H , R3 = benzoyl = Me, R, = cinnamoyl. Rz = R3 = H dihydroxytropane R = H, R, = phenylacetyl, 3a-Phenylacetoxy-6P.7PR, = R3 = H dihydrox ynortropane R = Me, R , = phenylacetyl, 3a-Phenylacetoxy-6P,7PR? = R, = H dihydroxytropane 3a-(2‘-Hydroxy-3’-phenylpropionyloxy)- R = Me, R, = 2-hydroxy-3phenylpripionyl, R2 = R, = H 6P,7P-dihydroxytropane ( = 6P,7Pdihydrox ylittorine) R = Me, R, = tigloyl, R, = H, 3a-Tigloyloxy-6P-hydroxy-7pR, = isovaleryl isovalerylox ytropane 3a,7P-Ditigloyloxy-6~-hydroxytropane R = Me, R, = R, = tigloyl, R2 = H 3a-(3’,4’,5’-Trimethoxybenzoyloxy)- R = Me, R, = 3,4,5-trirnethoxybenzoyl, R2 = R, = H 6P,7P-dihydroxytropane R = H, R, = 2-hydroxy-3( + )-3a-(2’-Hydroxy-3‘phenylpropionyl, R2 = H , R, = tigloyl phenylpropionyloxy)-6~-hydroxy-7Ptigloy lox y nortropane 3a-Acetoxy-6~,7~-dibenzoyloxytropaneR = Me, R, = acetyl, R2 = R, = benzoyl 3a-(3’,4’,5‘-Trimethoxycinnamoyloxy)-R = Me, R, = 3,4,5trirnethoxycinnamoyl, R2 = H, 6P-h ydroxy-7P-benzoyloxytropane R, = benzoyl 3a-(3‘,4’,5‘-Trimethoxycinnarnoyloxy)R = Me, R, = 3,4.5-trimethoxycinnarnoyl, R, = acetyl, 6~-acetoxy-7~-benzoyloxytropane R, = benzoyl

l23 3a-Benzoyloxy-6P,7P-dihydroxytropane R w 3a,6~-Dihydroxy-7P-benzoyloxytropaneR R 125 3a-Cinnamoyloxy-6p,7/3126

=

= =

1.

13

TROPANE ALKALOIDS

TABLE I1 (continued) Alkaloid

Structure

5. 3a-Substituted 6P,7P-epoxytropanes The 3a-substituted 6P.7P-epoxytropanes (136-143) are characterized by the 6P,7P-epoxy ring.

R 136 3a-Hydroxy-6@,7P-epoxytropane ( = scopine) R 137 3a-Apotropoyloxy-6P.7Pepox ynortropane (= aponorscopolamine or aponorhyoscine) 138 3a-Apotropoyloxy-6~,7~-epoxytropane R ( = aposcopolamine or apohyoscine) 139 3a-Tropoyloxy-6~,7~-epoxynortropane R ( = norscopolamine or norhyoscine) 140 ( - )-3a-Tropoyloxy-6~,7P-epoxytropaneR (=scopolamine or hyoscine) 141 (t)-3a-Tropoyloxy-6/3,7~-epoxytropaneR ( = atroscine) 142 ( - )-3a-(2’-Hydroxytropoyloxy)-6/3,7p- R epoxytropane [ = (-)-anisodine or daturamine] R 143 3a-Tropoyloxy-6P,7p-epoxytropaneNoxide ( = scopolamine N-oxide or hyoscine N-oxide)

= Me, =

H, R,

= Me, =

R, = H =

apotropoyl

R, = apotropoyl

H,R, = tropoyl

= Me,

R, = tropoyl

= Me,

R, = tropoyl

= Me,

R, = 2-hydroxytropoyl

= Me, 0, R, = tropoyl

6. 3P-Substituted 2P-carboxytropanes The 3P-substituted 2p-carboxytropanes (144-150) can be considered to be derivatives of 2P-carboxy-3P-hydroxytropane(ecgonine, 146) or of the corresponding, not yet naturally found, 2p-carboxy-3P-hydroxynortropane.

144b (-)-2P-Carboxy-3/3-formylnortropane ( =norecgonine formyl estei)

R

=

H, R, = formyl, R2 = H

(continued)

14

MAURI LOUNASMAA A N D TARJA TAMMINEN

TABLE I1 (continued) Alkaloid 145 2,3-Dehydro-2-methoxycarbonyltropane ( = A2(3)-anhydroecgoninemethyl ester or methylecgonidine) 146 ( - )-2P-Carboxy-3P-hydroxytropane ( = ecgonine) 147 ( - )-2P-Methoxycarbonyl-3Phydroxytropane ( = ecgonine methyl ester) 148 ( - )-2p-Carboxy-3P-benzoyloxytropane ( = benzoylecgonine) 149 ( - )-2p-Methoxycarbonyl-3/3benzoyloxytropane ( = cocaine) 150 (- )-2p-Methoxycarbonyl-3@cinnamo ylox ytropane ( = cinnamoylcocaine)

Structure R = Me, A2(3), R2 = Me

R = Me, R, = R2

=

H

R = Me, R, = H, R2

=

Me

R

=

Me, R, = benzoyl, R2 = H

R

=

Me, R, = benzoyl, R2

R

=

Me, R,

=

=

Me

cinnamoyl, R2 = Me

" .

7. 3a-Substituted 4a-benzvltro~anes The 3a-substituted 4a-benzyltropanes (151-155) are without functionality at C-6 and C-7.

151 3a-Acetoxy-4a-benzyltropane ( = alkaloid KD-B) 152 3a-Acetoxy-4a-hydroxybenzyltropane 153 3a-Acetoxy-4a-acetoxybenzyltropane ( = acetylknightinol) 154 3a-Benzoyloxy-4a-benzyltropane (=alkaloid KD-A) 155 3a-Benzoyloxy-4ahydroxybenzy ltropane

R, = acetyl, Rl = H R,

=

acetyl, R2 = OH

R, = acetyl, R2 = acetoxy R, = benzoyl, R2 = H R, = benzoyl, R2 = OH

8. 3a,6P-Disubstituted, 4a-benzyltropanes The 3cu-6P-disubstituted 4a-benzyltropanes (156-157) are 3a-substituted 4abenzyltropanes with an additional functionality at C-6.

15

1 . TROPANE ALKALOIDS TABLE I1 (conrinued) Alkaloid

156 3a-Acetoxy-4a-benzyl-6phydroxytropane ( = knightoline) 157 3a-Cinnamoyloxy-4a-benzyl-6phydroxytropane (=alkaloid KD-D)

Structure

R,

=

acetyl, R2

R,

=

cinnamoyl, R2 = R3

=

R3

=

H =

H

9. 3a,7p-Disubstituted 4a-benzyltropanes The 3a,7p-disubstituted 4a-benzyltropanes (158-160) are similar to those of the preceding group except that the additional functionality is at C-7 instead of C-6.

R3°Q& R2

\

158 3a-Hydroxy-4a-benzyI-7pR , = R2 = H, R3 = benzoyl benzoyloxytropane (=alkaloid KD-C) 159 3a-Cinnamoyloxy-4a-hydroxybenzyl-7p-R, = cinnamoyl, R2 = OH, R, = benzoyl benzoyloxytropane ( =alkaloid KD-E) R, = H, R2 = OH, R, = benzoyl 160 3a-Hydroxy-4a-hydroxybenzyl-7pbenzoyloxytropane ( =alkaloid KD-F) 10. 3P,6P-Disubstituted 4a-benzyltropanes The 3/3,6p-disubstituted 4a-benzyltropanes (161 and 162)are the only 4a-benzyltropanes where the C-3 substituent is p.

161 3~-Hydroxy-4a-hydroxybenzyl-6~R, acetoxytropane ( = knightalbinol) 162 3~-Benzoyloxy-4a-hydroxybenzyl-6p- R, hydroxytropane ( = knightolamine)

=

H, R2 = OH, R, = acetyl

=

benzoyl, R2

=

OH, R, = H

(continued)

16

MAURI LOUNASMAA A N D TARJA TAMMINEN

TABLE I1 (continued) Alkaloid

Structure

11. Pyranotropanes The natural products 163-166 contain a y-pyrano group attached to the 3,4-position of the tropane ring.

Me\

R1

163 Pyranotropane ( = strobiline) 164 10-Methylpyranotropane ( = bellendine) 165 1 I-Methylpyranotropane ( = isobellendine)

166 10,l I-Dimethylpyranotropane

R, = R2 = H R, = Me, R2 = H R , = H, Rz = Me R, = R2

=

Me

( = darlingine)

12. 3,4-Dihydropyranotropanes Compounds 167-168 are y-pyranotropane derivatives in which the 3,4-double bond is reduced.

MI?

\

D F H H

'O

167 1 I-Methyl-3,4-dihydropyranotropane ( = 5,ll-dihydroisobellendine) 168 lO,ll-Dimethyl-3,4dihydropyranotropane ( = 5 , l l dihydrodarlingine)

R, = H, R2 = Me R, = R2 = Me

11

1.

17

TROPANE ALKALOIDS

TABLE I1 (continued) Alkaloid

Structure

13. l0,ll-Dihydropyranotropanes The 10,l I-dihydropyranotropanes (169-174)are y-pyranotropanes in which the 10,I I double bond is reduced. It has not been possible to deduce from the data available whether the C-10 and/or C-ll substituents, when present. are Q or P .

169 10,I I-Dihydropyranotropane

Rl

=

R2

R,

=

Me,,, Rz = R,

=

H

171 10-Methyl-10,lI-dihydropyranotropane R ,

=

Me,,, R2

R,

=

H

R,

=

Rz = Meeg, R,

=

H

R,

=

R3

Ph,,

R1

=

H, R2

=

R,

=

H

( = dihydrostrobiline)

170 10-Methyl-10,ll-dihydropyranotropane ( = dihydrobellendine)

=

( = epidih ydrobellendine)

172 10,l I-Dimethyl-10.1 I dih ydropyranotropane ( = 2,3-dihydrodarlingine) 173 11-Phenyl-l0,ll-dihydropyranotropane ( = strobamine) 174 7P-Hydroxy-l I-phenyl-l0,11dihydropyranotropane ( = strobolamine)

=

H, R2 =

=

Ph,,, R,

=

OH

14. Miscellaneous tropanes The following heterogeneous group contains I3 "monomeric" compounds (175-187) not falling in any of the 13 preceding groups. Some, however, are apparent precursors for compounds mentioned earlier [e.g., chalcostrobamine (187)for strobamine (173)l.

175 3-Oxotropane

( = tropinone)

(continued )

18

MAURI LOUNASMAA A N D TARJA TAMMINEN

TABLE I1 (confinued) Alkaloid

Structure

176 Ip-Hydroxytropane (hydrochloride) ( = physoperuvine) OH

177 2P,6P-Dihydroxynortropane ( = baogongteng C)

m$ OH H

H

6

H\

FH

H

178 2P,7P-Dihydroxynortropane ( = erycibelline)

OH H

179 Ip,2a,3P-Trihydroxynortropane ( = calystegine A,) H r

Y

180 ( + )-3,4-Dehydro-4-acetyltropane ( = ferruginine)

Me

1.

19

TROPANE ALKALOIDS

TABLE I1 (continued) Alkaloid

Structure

181 Ip,2a,3/3,6P-Tetrahydroxynortropane (=calystegine B,)

mgH OHH

H

OH

a

182 lp,2a,3P,4cr-Tetrahydroxynortropane ( = calystegine B2)

q; OHH

OH

183 ( - )-2P-Hydroxy-6P-acetoxynortropane ( = baogongteng A)

184 ( + )-4a-Benzoyltropane ( = ferrugine)

(continued)

20

MAURl LOUNASMAA A N D TARJA TAMMINEN

TABLE 11 (conrinued) Alkaloid

Structure

185" (+)-2a-Benzoyloxy-3Phydrox ynortropane

186 6,7-Dehydro-3~(4'hydroxybenzoy1oxy)tropane [ = 3a-(4hydroxybenzoyloxy)trop-6-ene]

19;

7

6

0

Me

\

I

OH

I k

OH

187 ( + )-3,4-Dehydro-4-cinnarnoyl-3hydroxytropane ( = chalcostrobarnine)

0

Q I

1.

21

TROPANE ALKALOIDS

TABLE I1 (continued) Alkaloid

Structure

15. “Dmeric” and “trimeric” tropanes Fifteen “dimeric” tropane alkaloids (188-202) and one “trimeric” tropane alkaloid (203) have been found so far. 188 Schizanthine C

189 Schizanthine D

A 190 Schizanthine E

(continued)

22

MAURI LOUNASMAA A N D T A N A T A M M I N E N

TABLE I1 (continued) Alkaloid

Structure

191 a-Belladonnine

Me.

192 P-Belladonnine

Me.

II 0

193 a-Scopadonnine

23

1. TROPANE ALKALOIDS TABLE I1

(continued)

Alkaloid

Structure

194 p-Scopadonnine Me.

195 Schizanthine B Me,

C-Me

I Me

l . -

I

Me

H Me-?

-0

L"

he

1% Schizanthine X

(continued)

24

MAURI LOUNASMAA A N D TARJA TAMMINEN

TABLE I1 (continued) Alkaloid 197 7-Acetoxytropan-3-yl tropan-3’-yltruxillate

198 Subhirsine

199 Convolvidine

Structure

1.

25

TROPANE ALKALOIDS

TABLE 11 (confinued) Alkaloid

200 7-Acetoxytropan-3-yl 7’-hydroxytropan3’-yl-truxillate

HO,

0-

201 a-Truxilline

202 P-Truxilline

Structure

TABLE I1 (continued) Alkaloid

Structure

203 Grahamine

$=<”‘ a

I_

H

The indicated natural occurrence of compound 113 [Chern. Absrr. 78, 13742d (1973)l would seem to be erroneous (see Ref. 294). The indicated natural occurrence of compound 144 (6) is erroneous. The indicated natural occurrence of compound 185 (45) has turned out to be erroneous (347).

Nore. Structures of the acyl groups are as follows: 0

II

Formyl

H- C-

0

II

C-

Acetyl

Me-

Hydroxyacetyl

H&CHz-S

0

II 0

F’ropionyl

Me-

II

CHz- C0

Butyryl

Me-CH2-CH2-C-

II

TABLE 111 (continued) Me

0

I

lsobutyryl

Me-CbC-

0

Me

I

Me- CH--CHz-C-

Isovaleryl

Me

Me-

CHz-

2-Methylbutyryl

II

&A

II

0

11 C-

2

C-

I1

Tigloyl

0

Angeloyl

Senecioyl

-s

-1-

1,2-Dithiolane-3-carbonyl

O

c (3) s

3-Ethoxycarbonylmethacryloyl

Methylmesaconyl

Methylitaconyl

27


28

MAURI L O U N A S M A A A N D TARJA T A M M I N E N

TABLE I1 (continued)

Ethylmesaconyl

Ethylitaconyl

Fpi

2-Furoyl

2

c-

Benzoyl

3-H ydroxybenzoyl

"Go / \ 'd-

4-Methox ybenzoyl

MeO, Van i I I oy I

MeO.

Veratroyl

29

1. TROPANE ALKALOIDS TABLE I1 (continued) MeO. 3.45-Trimethoxybenzoyl / Me0

~ c ” - L 0

Phenylacetyl

HO

3-Hydroxyphenylacetyl

0 4-H ydroxyphenylacetyl

4-Methoxyphenylacetyl

Cinnamoyl

(continued )

30

MAURI LOUNASMAA A N D TARJA TAMMINEN

TABLE I1 (continued)

OH

I

CH2-C+C-

2-H ydroxy-3-phenylpropionyl

3

2

Apotropoyl

Tropoyl

Acetyltropoyl

II 2-H ydrox ytropoyl

OH

Pyrroly 1-2-carbonyl

QA

2

I

H

c-

0

II

31

1. TROPANE ALKALOIDS

TABLE I1 (continued) N-Methylp yrrol yl-2-carbonyl

2

I

c-

Me

Regional floras were used where possible. The oriental solanaceous genera have been classified by Baytop (34), but the classification could be used as a reference only in regard to some specific questions because species lying outside the region could not easily be incorporated in the scheme. The nomenclature of Xiao and He (35)has been adopted for Chinese plants. No botanical reference has been used for some regionally restricted genera (e.g., Solandra, Schizanthus) whose nomenclature seems to be widely accepted. The important genus Erythroxylum (Erythroxylaceae) has not been classified as a whole since Schulz (36); his division into sections is still widely used and is adopted in this work. The articles by Plowman (37-39) were taken as a guide to the cultivated Erythroxylum species, which, until that work, had been extremely poorly documented. In some cases, the only information on the species investigated was the geographical region where the sample had been collected; sometimes only the name “coca” was used. In these cases the interpretation must be considered as the personal view of the authors only. In more recent papers, only cocaine and the isomeric cinnamoylcocaines are reported for these species. Although the ecgonine derivatives reported to have been isolated earlier must now be considered artifacts, this past botanical confusion fortunately has no implications for the chemotaxonomic classification. The new flora of Australia, at the moment still incomplete, will be an invaluable source of information about the Proteaceous plants. Reviews covering all tropane alkaloid plants of this family are already available (40, 41). Some species are included in Table iII without a proper reference to the literature, and even without author names. in many cases a more precise identification might have been possible if the original articles rather than just Chemical Abstracts summaries had been available to us. The hybrids of commercially important alkaloid-producing plant species have been included in the list because they contain some interesting alkaloids not found anywhere else. Details of the hybrids could not be included, however, and the reader is referred to the original papers.

32

MAURl LOUNASMAA A N D T A N A TAMMINEN

TABLE I11 OCCURRENCE OF TROPANE ALKALOIDS I N DIFFERENT PLANTS-~ Euphorbraceae Phyllanthus P . discoides Muell. Arg.

36 3a-Vanilloyloxytropane (68?) Brassicaceae ( = Cruciferae) Cochlearia C. arctica

23 3a-(3'-Hydroxybenzoyloxy)tropane(69,*70)

Proteaceae Agastachus A . odorata R. Br.

82 3a-Tigloyloxy-7p-acetoxytropane (717) 186 6,7-Dehydro-3a-(4'-hydroxybenzoyloxy)tropane (717*) Bellendena B. montana R . Br.

59 77 78 164 165 166 167 170 171 172

+ )-3a-Acetoxy-7P-hydroxytropane(71I") 3a-Acetoxy-7~-isobutyryloxytropane (72'") 3cu-Isobutyryloxy-7p-acetoxytropane(72'") 10-Methylpyranotropane ( 7 2 , " ~73) ~ 1 I-Methylpyranotropane (72'") 10,l I-Dimethylpyranotropane (72") 1 I-Methyl-3,4-dihydropyranotropane(72") 10-Methyl-10,l I-dihydropyranotropane (72'") 10-Methyl-10, I I-dihydropyranotropane (epi) (72'") 10,l I-Dimethyl-10.1 I-dihydropyranotropane (72'") (

Darlingia D. darlingiana ( F . Muell) L. A . S . Johnson

164 10-Methylpyranotropane (747) 165 I I-Methylpyranotropane (747) 166 10.1 I-Dimethylpyranotropane (747) 168 10,l I-Dirnethyl-3,4-dihydropyranotropane (74') 180 ( + )-3,4-Dehydro-4-acetyltropane(747) D . ferncginea J . F. Bailey 155 3a-Benzoyloxy-2a-hydroxybenzyltropane (757) 166 10.1 I-Dirnethylpyranotropane(75,7 767) 180 (+)-3,4-Dehydro-4-acetyltropane(757) 184 (+)-4a-Benzoyltropane (75,7 767) Knightia K . deplunchei Vieill. e x Brongn. et Gris

151 2a-Benzyl-3a-acetoxytropane (77,"* 78) 154 3a-Benzoyloxy-4a-benzyltropane (77,"* 78) 157 3a-Cinnarnoyloxy-4a-benzyl-6~-hydroxytropane (77,"* 781 158 3a-Hydroxy-4a-benzyl-7P-benzoyloxytropane (77,"* 78) 159 3a-Cinnamoyloxy-4a-hydroxybenzyl-7~-benzoyloxytropane (7% I '*) 160 3a-Hydroxy-4a-hydroxybenzyl-7~-benzoyloxytropane (78, 79a"*)

I.

TROPANE ALKALOIDS

TABLE I11 (continued)

K . strobilina Labill. 74 3a-Hydroxy-7~-benzoyloxytropane(79’*) 83 3a-Cinnamoyloxy-7P-hydroxytropane (79’*) 152 3a-Acetoxy-4a-hydroxybenzyltropane (79):) 153 3a-Acetoxy-4a-acetoxybenzyltropane(79’*) (79’*) 156 3a-Acetoxy-4a-benzyl-6/3-hydroxytropane 161 3/3-Hydroxy-4a-hydroxybenzyl-6~-acetoxytropane (80’*) 162 3~-Benzoyloxy-4a-hydroxybenzyl-6~-hydroxytropane (801*) 163 Pyranotropane (79’,4*) 169 10,I I-Dihydropyranotropane (791,4*) 173 1 I-Phenyl-10,l I-dihydropyranotropane (80’*) 174 7P-Hydroxy-1 I-phenyl-lO,l I-dihydropyranotropane (80’*) 187 ( + )-3,4-Dehydro-4-cinnamoyl-3-hydroxytropane (80’*) Rhizophoraceae Bruguiera B . cylindrica ( L . ) BL.

( 8 / ,4 82j) 26 ( - )-3a-(1’,2’-Dithiolane-3’-carbonyloxy)tropane B . exaristnta Ding Hou 3 3a-Acetoxytropane (8j4) 4 3a-Propionyloxytropane (8j4) 7 3a-Butyryloxytropane (8j4) 8 3a-Isobutyryloxytropane (8j4) 14 3a-Isovaleryloxytropane (8j4) 18 3a-Benzoyloxytropane (8j4) 26 ( - )-3a-( 1’,2’-Dithiolane-3’-carbonyloxy)tropane (8j4) B . sexnngula (Lour.) Poir. 3 3a-Acetoxytropane (83j) 4 3a-Propionyloxytropane (8j4) 7 3a-Butyryloxytropane (8j4) 8 3a-Isobutyryloxytropane (834) 14 3a-Isovaleryloxytropane (8j4) 18 3a-Benzoyloxytropane (8j4) 26 ( - )-3a-(1‘,2‘-Dithiolane-3‘-carbonyloxy)tropane (83.4840‘) Crossostylis C. bijlora

175 3-Oxotropane ( 8 4 ’ ) C . multijlora

1 3a-Hydroxytropane (M4) 26 ( - )-3a-(1’,2’-Dithiolane-3’-carbonyloxy)tropane 42 3a-Feruloyloxytropane (M4) C . sebertii

18 3a-Benzoyloxytropane (H4) 25 3a-Cinnamoyloxytropane (M4) Pellacalyx P . axillnris Korth

21 3a-Cinnamoyloxynortropane ( 8 5 ) )

33

34

MAURl LOUNASMAA A N D TARJA TAMMINEN

TABLE Ill (continued) Erythroxylaceae Erythroxylum, Sect. I1 Macrocalyx E. macrocnemium Mart. 149 ( - )-2~-Methoxycarbonyl-3~-benzoyloxytropane (23') E. lucidum H. B. K . 149 ( - )-2~-Methoxycarbonyl-3~-benzoyloxytropane (23') Erythroxylum, Sect. 111 Rhabdophyllum E. amazonicum Peyr. 75 3-Benzoyloxy-7~-hydroxytropane(86) E. campestre 149 ( - )-2~-Methoxycarbonyl-3~-benzoyloxytropane (87') E. deciduum 149 ( - )-2~-Methoxycarbonyl-3~-benzoyloxytropane (87') E. fimbriatum Peyr. 1 3a-Hydroxytropane (68) 149 ( -)-2P-Methoxycarbonyl-3~-benzoyloxytropane (23') E. pelleterianum A. St. Hil. 149 ( - )-2~-Methoxycarbonyl-3/3-benzoyloxytropane (87') E. steyermarkii Plowman 149 ( - )-2~-Methoxycarbonyl-3~-benzoyloxytropane (23') 150 ( - )-2~-Methoxycarbonyl-3~-cinnamoyloxytropane (cis- and trans-) (23 ') Erythroxylum, Sect. IV Leptogramme E. pulchrum A. St. Hil. 149 ( -)-2/3-Methoxycarbonyl-3/3-benzoyloxytropane (23,' 87') 150 ( - )-2~-Methoxycarbonyl-3~-cinnamoyloxytropane (cis-)(23 ') E . ulei 0. E. Schulz 55 3P-Benzoyloxytropane (86) Erythroxylum, Sect. V Heterogyne E. areolatum L. 149 ( - )-2~-Methoxycarbonyl-3~-benzoyloxytropane (23,'88') E . rotundifolium Lunan 149 ( - )-2~-Methoxycarbonyl-3~-benzoyloxytropane (23') 150 ( - )-2~-Methoxycarbonyl-3/3-cinnamoyloxytropane (cis-) (23') Erythroxylum, Sect. VI Archerythroxylum E. argentinum 0 . E. Schulz 18 3a-Benzoyloxytropane (86') 58 3-Acetoxy-7~-hydroxynortropane(86") E . coca Lam. var. coca 50 3P-Hydroxytropane (892) 55 3P-Benzoyloxytropane (89') 145 2,3-Dehydro-2-methoxycarbonyltropane (903) 146 ( -)-2a-Carboxy-3P-hydroxytropane(91 ') 147 ( - )-2~-Methoxycarbonyl-3~-hydroxytropane (89.' 92') 149 ( -)-2~-MethoxycarbonyI-3/3-benzoyloxytropane (23,' 89,' 91,' 92,' 93,'.3 94,',4369.5,' 96') 150 ( - )-2~-Methoxycarbonyl-3~-cinnamoyloxytropane (cis- and trans-) (23,' 89,' 94,'.4,695,' 96')

1.

35

TROPANE ALKALOIDS

TABLE 111 (continued) (Notes: 90, 93, 96: E. coca Lam.; 92: "cusko" leaves; 94: E. coca Lam., Tingo Maria; 95: E. coca, Cuzco, Tingo Maria) E . coca var. ipadu 149 ( - )-2~-Methoxycarbonyl-3~-benzoyloxytropane (23 ') 150 ( - )-2~-Methoxycarbonyl-3~-cinnamoyloxytropane (cis-) (23') E. cumanense H.B. K. 74 3a-Hydroxy-7P-benzoyloxytropane (8S2) 76 3a-Phenylacetoxy-7P-hydroxynortropane(862*) 133 3-Acetoxy-6~,7~-dibenzoyloxytropane (86**) E. glaucum 0 . E. Schulz 74 3a-Hydroxy-7P-benzoyloxytropane (86) 149 ( - )-2~-Methoxycarbonyl-3~-benzoyloxytropane (23') E. gracilipes Peyr. 149 ( - )-2~-Methoxycarbonyl-3~-benzoyloxytropane (23') E. afi. impressum 0 . E. Schulz 149 ( - )-2~-Methoxycarbonyl-3~-benzoyloxytropane (23') E. incrassaturn 0 . E. Schulz 149 ( - )-2~-Methoxycarbonyl-3~-benzoyloxytropane (23') E . ovatum Cav. 149 ( - )-2~-Methoxycarbonyl-3~-benzoyloxytropane (88') E. mamacoca Mart. 54 3P-Benzoyloxynortropane(86*) 55 3P-Benzoyloxytropane (86) E. novogranatense (Moms)Hieron var. novogranatense 55 3P-Benzoyloxytropane (89,',* 97') 145 2,3-Dehydro-2-methoxycarbonyltropane (903) 147 ( - )-2P-Methoxycarbonyl-3P-hydroxytropane(92') 148 ( - )-2P-Carboxy-3P-benzoyloxytropane (97') 149 (-)-2~-Methoxycarbonyl-3~-benzoyloxytropane (23,' 87,' 92,' 93,'.' 96,' 97') 150 (- )-2~-Methoxycarbonyl-3~-cinnamoyloxytropane (cis-and trans-) (23,' 96,' 97') 201/ aIP-Truxilline (98') 202 (Notes: 89: E. coca L a m ; var. spruceanum Burck; 92: Java coca; 97: E. coca var. novogranatensis) E. novogranatense var. truxillense (Rusby) Machado 50 3P-Hydroxytropane (892) 55 3P-Benzoyloxytropane (89'.2) 145 2,3-Dehydro-2-methoxycarbonyltropane (99') 147 ( - )-2/3-Methoxycarbonyl-3P-hydroxytropane(89') 149 ( - )-2~-Methoxycarbonyl-3~-benzoyloxytropane (23, 89, 92, 93, 94, 95, 96, 100')

' '

' ' ' ' '

(continued)

36

MAURI LOUNASMAA A N D TARJA TAMMINEN

TABLE 111 (continued)

’

150 ( - )-2~-Methoxycarbonyl-3~-cinnamoyloxytropane (cis- and trans-) (23, 89,‘ 94,I 95,‘ 96,‘ 100I) 2011 alp-Truxilline (101 I) 202 [Notes: 94: E . coca Lam., Trujillo; 95: E. coca, Trujillo; 99, 101: cocaine side product (Peruvian coca’?); 100: Peruvian coca paste]

E. recurrens Huber 149 ( - )-2~-Methoxycarbonyl-3~-benzoyloxytropane (23’) 150 ( - )-2~-Methoxycarbonyl-3~-cinnamoyloxytropane (cis- and frans-)( 2 3 ’ ) E. shutona Macbride 149 ( - )-2P-Methoxycarbonyl-3~-benzoyloxytropane (23’) E. uacciniijolium Martius 102 3a-(Pyrrolyl-2’-carbonyloxy)-7~-(N“-methyIpy~oIyI-2”carbony1oxy)tropane (102, 1031,4) 117 3a-(3’,4’,5‘-Trimethoxybenzoyloxy)-7~-benzoyloxytropane (102, 103’,‘) 118 3a-(3’,4’,5’-Trimethoxybenzoyloxy)-7~-(N”-methylpyrrolyl-2‘’carbony1oxy)tropane (102, Erythroxylum, Sect. IX Microphyllum E. panumense Turcz. 149 ( - )-2P-Methoxycarbonyl-3p-benzoyloxytropane (87’) Erythroxylum, Sect. X Melanocladus E. zamhesiacirm N. Robson 45 3a-(3’,4‘.5’-Trimethoxybenzoyloxy)nortropane (104’*) 74 3a-Hydroxy-7p-benzoyloxytropane ( I04?) 80 3a-Phenylacetoxy-7P-hydroxytropane(104?) 98 3a-(3‘,4’,5’-Trimethoxybenzoyloxy)-7~-hydroxynortropane (104’) 100 ( + )-3a-(3’,4’,5’-Trimethoxybenzoyloxy)-7p-hydroxytropane (104?*) 119 3a-(3‘,4’,5’-Trimethoxycinnamoyloxy)-7~-benzoyloxytropane ( I 04’) 124 3a,6P-Dihydroxy-7~-benzoyloxytropane (I@*) 131 3a-(3’.4’,5’-Trimethoxybenzoyloxy)-6~,7~-dihydroxytropane (104’) 134 3a-(3’,4’.5‘-Trimethoxycinnamoyloxy)-6~-hydroxy-7~benzoyloxytropane (104*) 135 3a-(3’,4’,5’-Trimethoxycinnamoyloxy)-6~-acetoxy-7~-benzoyloxytropane (1047 Eryfhroxvlum, Sect. XI Sethia E. monogynrrm Roxb. 1 3a-Hydroxytropane(105?) 48 3a-(3’,4’,5’-Trimethoxybenzoyloxy)tropane (105,** 106l) 49 3a-(3‘,4‘,5’-Trimethoxycinnamoyloxy)tropane (105,’ 106’) 50 3P-Hydroxytropane (105?) 119 3a-(3’,4’.5‘-Trimethoxycinnamoyloxy)-7~-benzoyloxytropane (106,’ 107?*) 131 3a-(3’,4’,5’-Trimethoxybenzoyloxy)-6~,7~-dihydroxytropane (106’) 150 ( - )-2~-Methoxycarbonyl-3~-cinnamoyloxytropane (/@?I”) Eryfhroxylum. Sect. XI1 Lagynocarpus E. dekindtii (Engl.) 0.E. Schultz 1 3a-Hydroxytropane( I @ ’ ) 9 3a-Isovaleryloxynortropane( f o g ? )

1.

TROPANE ALKALOIDS

T A B L E 111 (continued) 10 3a-(2’-Methylbutyryloxy)nortropane(109’) 14 3a-Isovaleroyloxytropane (109**) 16 3a-(2’-Furoyloxy)tropane(109?) 22 3a-Phenylacetoxytropane(109?*) 50 3P-Hydroxytropane (110’) 55 3P-Benzoyloxytropane(110’) 71 3a-Isovaleryloxy-6P-hydroxytropane (109’) 146 (-)-2a-Carboxy-3P-hydroxytropane (110’) 147 ( - )-2P-Methoxycarbonyl-3P-hydroxytropane (109,’ 110’) Erythroxylum, Sect. XIV Coelocarpus E. australe F. Muell. 31 3a-(4-Hydroxyphenylacetoxy)tropane(11I?*) 62 ( + )-3a-Hydroxy-7P-tigloyloxynortropane (557) 63 (+-)-3a-Tigloyloxy-7~-hydroxynortropane (55’) 83 3a-Cinnamoyloxy-7P-hydroxytropane(111’) 122 3a-Tigloyloxy-6~,7P-dihydroxytropane (55,’ 56’) 123 (~)-3a-Benzoyloxy-6P,7~-dihydroxytropane (55,’ 111?) 125 3-Cinnamoyloxy-6P,7j3-dihydroxytropane (111’*) 132 ( + )-3a-( 2‘-Hydroxy-3’-phenylpropionyloxy)-6~-hydroxy-7~tigloyloxynortropane (5j7’) 147 ( - )-2/3-Methoxycarbonyl-3P-hydroxytropane (111’) E. ecarinaturn Burck 55 3P-Benzoyloxytropane( I 1 1 I) 73 3a-Benzoyloxy-7~-hydroxynortropane (111 ’ ) E. ellipticum R. Br. 18 3a-Benzoyloxytropane (112‘”) 49 3a-(3’.4’,5’-Trimethoxycinnamoyloxy)tropane E. cuneatum (Wall.) Kurz 55 3P-Benzoyloxytropane(111 I 4, 66 3a-Hydroxy-7P-tigloyloxytropane (1f 1 ’) 74 3a-H ydroxy-7P-benzoyloxytropane (11I I ) 107 3a,7P-Dibenzoyloxytropane( I 11 Erythroxylum, Sect. XVI Venelia E. hypericifolium Lam. 15 3a-Benzoyloxynortropane(1117,‘ 114’) 18 3a-Benzoyloxytropane(114’) 19 3-Phenylacetoxynortropane (115’*) 21 3a-Cinnamoyloxynortropane (f 1 4 ’ ) 22 3a-Phenylacetoxytropane(113,4f15’*) 25 3a-Cinnamoyloxytropane(114’) 30 3a-(3‘-Hydroxyphenylacetoxy)tropane (113.4/ 1 5 ’ * ) 49 3a-(3’,4’,5’-Trimethoxycinnamoyloxy)tropane (11j4) 56 3P-Cinnamoyloxytropane(114’*) 73 3a-Benzoyloxy-7~-hydroxynortropane(11 4 ’ ) 75 3-Benzoyloxy-7P-hydroxytropane ( 1 1 j 4 ) 76 3a-Phenylacetoxy-7P-hydroxynortropane (113,‘ 114’) 79 3-Cinnamoyloxy-7~-hydroxynortropane( 1 14’*

37

38

MAURI LOUNASMAA A N D TAIUA TAMMINEN

TABLE 111 (continued)

80 3a-Phenylacetoxy-7/3-hydroxytropane(113,4 1152*) 81 3a-Hydroxy-7P-phenylacetoxytropane ( I 14"*) 83 3a-Cinnamoyloxy-7P-hydroxytropane(114') 86 3a-Benzoyloxy-7~-acetoxytropane (11j4) 91 3a-Phenylacetoxy-7P-acetoxytropane(113,4 115'*) 92 3a-Acetoxy-7P-phenylacetoxytropane (11j4) 97 3a-CinnamoyIoxy-7~-acetoxytropane (114'*) 116 3a,7P-Dicinnamoyloxytropane(114'*) 125 3-Cinnamoyloxy-6~,7P-dihydroxytropane (114!"*) 127 3a-Phenylacetoxy-6P,7P-dihydroxytropane ( 1 13,4115**) 149 ( - )-2~-Methoxycarbonyl-3~-benzoyloxytropane (116) 197 7-Acetoxytropan-3-yl tropan-3'-yl-truxillate (114'*') 200 7-Acetoxytropan-3-yl 7'-hydroxytropan-3'-yl-truxillate(114'*?) Erythroxylum, Sect. XVII Pachylobus E. laurifolium Lamk. (complex including E. macrocarpum and E . sideroxyloides) (116) 149 ( - )-2~-Methoxycarbonyl-3~-benzoyloxytropane E . macrocarpum 0 . E. Schulz 15 3a-Benzoyloxynortropane ( I 17*1,2.4) 21 3a-CinnamoyIoxynortropane ( / / 7 ' * ' ) 50 3P-Hydroxytropane (117') 55 3P-Benzoyloxytropane (117') 73 3a-Benzoyloxy-7~-hydroxynortropane (117'*) E. sideroxyloides Lam. 15 3a-Benzoyloxynortropane (1171.2.4) 18 3a-Benzoyloxytropane (117') 55 3P-Benzoyloxytropane (117') 73 3a-Benzoyloxy-7~-hydroxynortropane (117l) 75 3a-Benzoyloxy-7P-hydroxytropane(11 7 ' * ) Olacaceae Heisteria H . olivae 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (1186) Solanaceae Anthocercoideae Anthocercideae Anthocercis A . angustifolia F. Muell 24 3a-Apotropoyloxytropane (119') 34 (-)-3a-Tropoyloxytropane (119') 52 3P-Tigloyloxytropane (119') 66 3a-Hydroxy-7P-tigloyloxytropane (1 1 9 ' ) 82 3a-Tigloyloxy-7P-acetoxytropane (119') 93 3a,7P-Ditigloyloxytropane(119') 99 3a-Tropoyloxy-7~-acetoxytropane (119') 138 3a-Apotropoyloxy-6~,7~-epoxytropane (119') 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (119')

1.

TROPANE ALKALOIDS

39

TABLE I11 (continued) A . anisantha Endl. ssp. anisantha 24 3a-Apotropoyloxytropane (1191,7) 28 ( - )-3a-Tropoyloxynortropane (119137) 34 (-)-3a-Tropoyloxytropane (1191.7) 35 ( - )-3a-(2’-Hydroxy-3’-phenylpropionyloxy)tropane( 1197)

93 130 138 140

3a,7P-Ditigloyloxytropane(1197) 3a,7P-Ditigloyloxy-6P-hydroxytropane(1197) 3a-Apotropoyloxy-6P,7P-epoxytropane(1197) ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (119’.7)

A . anisantha Endl. ssp. collina Haegi 24 3a-Apotropoyloxytropane ( I 19’) 28 ( - )-3a-Tropoyloxynortropane (119’) 34 (-)-3a-Tropoyloxytropane (119’) 140 (-)-3a-Tropoyloxy-6P,7P-epoxytropane (119’) A . fasciculara 34 ( - )-3a-Tropoyloxytropane (120’) A . genistoides Miers. 1 3a-Hydroxytropane (SO7?) 28 ( - )-3a-Tropoyloxynortropane (50,71197) 34 (-)-3a-Tropoyloxytropane (50,7 119’?.2.7) 87 ( -)-3a-Tropoyloxy-6P-hydroxytropane 51 , 2 ? 1 19*!) 93 3a,7P-Ditigloyloxytropane(119??) 122 3a-Tigloyloxy-6~,7P-dihydroxytropane (50,7 1192.7) 130 3a,7P-Ditigloyloxy-6~-hydroxytropane (119”) 137 3a-Apotropoyloxy-6~,7~-epoxynortropane (51,2?*119’) 138 3a-Apotropoyloxy-6~,7~-epoxytropane (51, 2 119’.?) 139 3a-Tropoyloxy-6~,7~-epoxynortropane (51,21191,2) 140 (-)-3a-Tropoyloxy-6~,7~-epoxytropane (50,751,2* 119’.?) (Note: 51: A . genistoides ssp. Haegi) A . gracilis Benth. 24 3a-Apotropoyloxytropane (1197!) 34 (-)-3a-Tropoyloxytropane (1197) 140 ( - )-3a-Tropoyloxy-6~,7P-epoxytropane (1197) A . ilicifolia Hook. ssp. ilicifolia 1 3a-Hydroxytropane (502?,7?) 24 3a-Apotropoyloxytropane (502,7) 27 (+)-3a-Tropoyloxynortropane (SO2) 28 ( - )-3a-Tropoyloxynortropane (507) 33 (+)-3a-Tropoyloxytropane (502) 35 ( - )-3a-(2‘-Hydroxy-3’-phenylpropionyloxy)tropane 52 3P-Tigloyloxytropane (SO?) 71 3a-Isovaleryloxy-6~-hydroxytropane (502) 87 ( - )-3a-Tropoyloxy-6~-hydroxytropane U2 3a-Tigloyloxy-6P,7P-dihydroxytropane (SO7) 138 3a-Apotropoyloxy-6~,7~-epoxytropane 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (502.7) (continued)

40

MAURl LOUNASMAA A N D T A W A TAMMINEN

TABLE 111 (continued) A . intricutu F.

28

34

Muell.

( - )-3a-Tropoyloxynortropane ( I197) ( - )-3a-Tropoyloxytropane(1197)

68 3a-Tigloyloxy-7P-hydroxytropane ( I 197) 82 3a-Tigloyloxy-7P-acetoxytropane(1197) 122 3a-Tigloyloxy-6~,7P-dihydroxytropane (1197) 140 ( - )-3a-Tropoyloxy-6~,7P-epoxytropane (1197) A . littorea Labill.

1 11 24 27 28 33 35 50 52 66 93 122 130 139 140

3a-Hydroxytropane(119,7 /2/2.7) 3a-Tigloyloxytropane(12/?’.7) 3a-Apotropoyloxytropane( 1 / 9 , 7/2/2.7) (?)-3a-Tropoyloxynortropane (I212 , 7 ) ( - )-3a-Tropoyloxynortropane (1197) (?)-3a-Tropoyloxytropane (119,7 120,’ / 2 P 7 ) ( -)-3a-(2’-Hydroxy-3’-phenylpropionyloxy)tropane(119,7 120,’ /2/?,7) 3P-Hydroxytropane ( 1 2 / 2 , 7 ) 3P-Tigloyloxytropane(119,7 121l) 3a-Hydroxy-7P-tigloyloxytropane (/217) 3a.7P-Ditigloyloxytropane( 1 / 9 7 ) 3a-Tigloyloxy-6P,7P-dihydroxytropane (119,’ 120,’ /2/2,7) 3a,7P-Ditigloyloxy-6~-hydroxytropane (119,7 12/’) 3a-Tropoyloxy-6~,7~-epoxynortropane (121 (-)-3a-Tropoyloxy-6P,7P-epoxytropane (1/9.7’/2/’,7) A . viscosa R. Br. 1 3a-Hydroxytropane (121?.7) 11 3a-Tigloyloxytropane 24 3a-Apotropoyloxytropane(/2/?.7) 27 (?)-3a-Tropoyloxynortropane (121 33 (t)-3a-Tropoyloxytropane ( I 2 1 34 ( - )-3a-Tropoyloxytropane(119,?.7120l) 35 ( - )-3a-(2’-Hydroxy-3‘-phenylpropionyloxy)tropane (/2/2.7) 50 3P-Hydroxytropane(/2/2.7) 52 3P-Tigloyloxytropane(121’) 57 3a.6P-Dihydroxytropane( / 2 / 7 ! ) 122 3a-Tigloyloxy-6~.7P-dihydroxytropane (/2/?,7) 130 3a,7P-Ditigloyloxy-6~-hydroxytropane (121’) 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (1/9,7 /2/2,7) (Note:119: A . viscosn ssp. viscosn Haegi) A . viscosa R. Br. ssp. cnudutu Haegi 1 3a-Hydroxytropane( / / 9 7 ) 11 3a-Tigloyloxytropane(1197) 24 3a-Apotropoyloxytropane( / 1 9 5 . 7 ) 28 ( - )-3a-Tropoyloxynortropane ( I197) 34 ( - )-3a-Tropoyloxytropane(1195”.7) 93 3a.i’P-Ditigloyloxytropdne ( / / 9 5 . 7 ) 122 3a-Tigloyloxy-6P,7P-dihydroxytropane (1197) 130 3a.7P-Ditigloyloxy-6P-hydroxytropane (1195,7)

1.

TROPANE ALKALOIDS

TABLE 111 (continued) 138 3a-Apotropoyloxy-6~,7~-epoxytropane ( I /97) 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (//97) Anthotroche A . myoporoides C. A . Gardn.

1 3a-Hydroxytropane (52,: / 1 9 ? ) 3 3a-Acetoxytropane (52,: // 9 ? ) 20 3a-Apotropoyloxynortropane ( 5 / , ’ *52,7 / / 9 7 ) 24 3a-Apotropoyloxytropane ( 5 / , ’ 52,7’//97) 28 (-)-3a-Tropoyloxynortropane (5/,l 52.?.7/19?.7) 34 ( - )-3a-Tropoyloxytropane ( 5 / , ’ 52,?.7//9?.7) 140 ( - )-3a-Tropoyloxy-6~.7~-epoxytropane (51 ,I 52,7 1 / 9 7 ) A . pannosa Endl. complex 1 3a-Hydroxytropane (52,?.7//92,7) 24 3a-Apotropoyloxytropane (52.7 //97) 28 ( -)-3a-Tropoyloxynortropane (52,2,71/ 9 2 . 7 ) 34 ( - )-3a-Tropoyloxytropane (52,2.71/9.?.7 / Z 7 ” ) 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (52.7 / / 9 7 ) A . walcottii F. Muell. 3 3a-Acetoxytropane (527) 20 3a-Apotropoyloxynortropane (5/,l* 52,7 //97) 24 3a-Apotropoyloxytropane (5/,l 52,7 / / 9 7 ) 28 ( - )-3a-Tropoyloxynortropane (5/,l 52,?.7//9’,7) 34 (-)-3a-Tropoyloxytropane ( 5 / , ’ 52,?,7//92.7) 140 ( - )-3a-Tropoyloxy-6~,7~-epoxytropane (51,’ 52.7’ / / 9 7 ” ) Crenidium C. spinescens Haegi

1 24 28 33 34 87 138 139 140

3a-Hydroxytropane (5U”’,7”) 3a-Apotropoyloxytropane (5U2.7) ( - )-3a-Tropoyloxynortropane (5/P7) (2)-3a-Tropoyloxytropane (5U7) ( - )-3a-Tropoyloxytropane (5U2) ( - )-3a-Tropoyloxy-6P-hydroxytropane (50?.7) 3a-Apotropoyloxy-6P,7j3-epoxytropane (5U2,7) 3a-Tropoyloxy-6~,7~-epoxynortropane (50) ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (5U?.7)

Cyphanthera C. dhicans ( A . Cunn.) Miers ssp. alhicans

3 8 11 13 24 34 64 68 71

3a-Acetoxytropane (5/,7 //9?.7) 3a-Isobutyryloxytropane (5/,7 //9?”,7) 3a-Tigloyloxytropane (5/,7 / / 9 7 ) ( + )-3a-(2’-Methylbutyryloxy)tropane(5/.7’ //9?.7”) 3a-Apotropoyloxytropane (119:) ( - )-3a-Tropoyloxytropane (5/,7 //92,7) 3a-Isobutyryloxy-7P-hydroxytropane ( 5 / , 7 */ / 9 7 ) 3a-Tigloyloxy-7p-hydroxytropane(119:) 3a-Isovaleryloxy-6P-hydroxytropane(5/,7 / / 9 7 )

41

42

MAURI LOUNASMAA AND TARJA TAMMINEN TABLE I11 (continued) 87 93 138 140

( - )-3a-Tropoyloxy-6P-hydroxytropane (1192)

3a,7P-Ditigloyloxytropane(1197) 3a-Apotropoyloxy-6~,7~-epoxytropane (1192.7) ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (51,71192.7) (Note: 51: Anthocercis albicans, coll. Haegi 1363) C . albicans A. C u m . ssp. notabilis Haegi 1 3a-Hydroxytropane (1194) 3 3a-Acetoxytropane (51,l 1 1 9 ' ) 7 3a-Butyryloxytropane (51,'* 119') 11 3a-Tigloyloxytropane (51,l 119") 24 3a-Apotropoyloxytropane (119") 34 (-)-3a-Tropoyloxytropane (1194) 64 3a-Isobutyryloxy-7P-hydroxytropane(51 I ? ) 138 3a-Apotropoyloxy-6~,7~-epoxytropane (51 ,I 119'.2,4) 140 ( -)-3a-Tropoyloxy-6P,7P-epoxytropane (51,' 119'.2.4) (Note: 51: Anthocercis albicans ssp., coll. Haegi 1379) C. anthocercidea (F. Muell.) Haegi 1 3a-Hydroxytropane (119,'.4123'.4) 24 3a-Apotropoyloxytropane (119,2.41232.4) 28 ( -)-3a-Tropoyloxynortropane ( 1 / 9 , ' 123') 34 (-)-3a-Tropoyloxytropane (119,1.2.4 123',2.4) 138 3a-Apotropoyloxy-6~,7~-epoxytropane (119,2.412324, 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (119,1.2.4123'.2.4 ) (Note: 123: Anthocercis frondosa) C . microphylla F. Muell. 34 (-)-3a-Tropoyloxytropane (1197') (1197') 138 3a-Apotropoyloxy-6~,7~-epoxytropane C . myosotidea (F. Muell.) Haegi 1 3a-Hydroxytropane (1247) 11 3a-Tigloyloxytropane (1247') 27 (~)-3a-Tropoyloxynortropane(1247) 33 (-C)-3a-Tropoyloxytropane (1247) 87 ( - )-3a-Tropoyloxy-6P-hydroxytropane (1247) 126 3a-Phenylacetoxy-6~,7P-dihydroxynortropane (1247'*) 136 3a-Hydroxy-6P,7P-epoxytropane(1247) 138 3a-Apotropoyloxy-6~,7~-epoxytropane (1247) 140 ( - )-3a-Tropoyloxy-6~,7P-epoxytropane (1247) C . odgersii (F. Muell.) Haegi 1 3a-Hydroxytropane (507') 24 3a-Apotropoyloxytropane (SO7) 28 (-)-3a-Tropoyloxynortropane (507) 34 ( - )-3a-Tropoyloxytropane (SO7) 87 ( - )-3a-Tropoyloxy-6P-hydroxytropane (SO7) 138 3a-Apotropoyloxy-6~,7~-epoxytropane (SO7) 140 ( - )-3a-Tropoyloxy-6~,7~-epoxytropane (SO7) C. odgersii ssp. occidentalis Haegi 34 ( - )-3a-Tropoyloxytropane ( 1 / 9 7 ) 87 ( - )-3a-Tropoyloxy-6~-hydroxytropane (1197)

1.

43

TROPANE ALKALOIDS

TABLE 111 (continued) 138 3a-Apotropoyloxy-6~,7~-epoxytropane ( I 19') 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane ( I 19') C . scabrella (Benth.) Miers 34 (-)-3a-Tropoyloxytropane (1 197') 71 3a-Isovaleryloxy-6~-hydroxytropane (1I 97) 138 3a-Apotropoyloxy-6~,7~-epoxytropane (1I 97) 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane ( I 19') C . tasmanica Miers. 1 3a-Hydroxytropane (XI7'.*) 33 (t)-3a-Tropoyloxytropane(507.2) 52 3P-Tigloyloxytropane (507.2) 71 3a-Isovaleryloxy-6~-hydroxytropane (%I2) U2 3a-Tigloyloxy-6j3,7P-dihydroxytropane(SO7) 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane 125') (Note: 125: Anthocercis tasmanica) Duboisia D . hopwoodii F. Muell. 34 ( -)-3a-Tropoyloxytropane (126,2.4127,'.* 128') 140 ( - )-3a-Tropoyloxy-6~,7P-epoxytropane (126,'.2.4127,2 12@) D . leichardtii F. Muell. 1 3a-Hydroxytropane (1292) 8 3a-Isobutyryloxytropane (129,'.2.4130,* 131,' 132,' 133,' 134') 13 (+)-3a-(2'-Methylbutyryloxy)tropane (129,'.2.4130,* 131,' 133,' 134') 24 3a-Apotropyloxytropane (132') 28 (-)-3a-Tropoyloxynortropane (129,1.4132') 33 (+)-3a-Tropoyloxytropane (129,2135n) 34 (-)-3a-Tropoyloxytropane(128,8 129,1.2.4131,' 136, 137,'.2,4138,' 139') 138 3a-Apotropoyloxy-6~,7~-epoxytropane (132I) 140 ( - )-3a-Tropoyloxy-6j3,7P-epoxytropane (128,8 129,1,2,4 131,' 136, 137,'.2.4 138,8 1398) 193 a-Scopadonnine (132') 194 P-Scopadonnine (132') D . myoporoides R . Br. 1 3a-Hydroxytropane (42,' 140,'.4141*) 3 3a-Acetoxytropane (140') 8 3a-Isobutyryloxytropane (42,' 140'.4) 9 3a-Isovaleryloxynortropane (16, 140,I 142'*) 10 3a-(2'-Methylbutyryloxy)nortropane (16, 142'*) 11 3a-Tigloyloxytropane (140') 13 ( + )-3a-(2'-Methylbutyryloxy)tropane(140,1,4141 143') 27 (~)-3a-Tropoyloxynortropane(140'.4) 28 ( - )-3a-Tropoyloxynortropane (42,' 144) 33 (+)-3a-Tropoyloxytropane (135,R141 ,2.4 1439) 34 (-)-3a-Tropoyloxytropane(42,' 43,' 128,R140,'.414/.?144') 52 3P-Tigloyloxytropane (16'*) 71 3a-Isovaleryloxy-6P-hydroxytropane(16,'* 136, 140,1.4141 2, 138 3a-Apotropoyloxy-6~,7~-epoxytropane (42,' 140'.4)

,'

(continued)

44

MAURl LOUNASMAA AND TARJA TAMMINEN TABLE I11 (continued) 139 3a-Tropoyloxy-6P,7P-epoxytropane(1404) 140 ( - )-3a-Tropoyloxy-6P.7P-epoxytropane(16,' 42,' 43,' 128.' 135,' 136. 140,'.4141,' 143,9 145') D.leichardtii x myoporoides 1 3a-Hydroxytropane (146') 28 ( - )-3a-Tropoyloxynortropane (146') 33 (?)-3a-Tropoyloxytropane (1359 34 ( - )-3a-Tropoyloxytropane(46.7146,' 147,' 148,' 149') 35 ( - )-3a-(2'-Hydroxy-3'-phenylpropionyloxy)tropane (148') 87 ( - )-3a-Tropoyloxy-6P-hydroxytropane (46,7 146,' 150') 89 3a-Tropoyloxy-7P-hydroxytropane(146 ') 140 ( - )-3a-Tropoyloxy-6~,7/3-epoxytropane (46,7135,' 146,' 147,' 149,' 150')

Grummosolen G. divonii (F. Muell. et R . Tate) Haegi 3 3a-Acetoxytropane (1197) 13 ( )-3a-(2'-Methylbutyryloxy)tropane(119')

+

24 34 87 138 140

3a-Apotropoyloxytropane (119') ( - )-3a-Tropoyloxytropane(119?',7) ( -)-3a-Tropoyloxy-6P-hydroxytropane(119') 3a-Apotropoyloxy-6/3,7~-epoxytropane (11Y7) ( - )-3a-Tropoyloxy-6/3,7/3-epoxytropane (11Y2,')

Symonanthus S . aromaticus (C. A. Gardner) Haegi

11 52 66 68 93

3a-Tigloyloxytropane ( / 1 9 ? ) 3P-Tigloyloxytropane ( 1 / 9 ? ) 3a-Hydroxy-7P-tigloyloxytropane (119') 3a-Tigloyloxy-7P-hydroxytropane(119',7) 3a,7P-Ditigloyloxytropane(119?) 130 3a.7/3-Ditigloyloxy-6p-hydroxytropane( I19') 138 3a-Apotropoyloxy-6~,7~-epoxytropdne (l19'.7) 140 ( - )-3a-Tropoyloxy-6P,7p-epoxytropane (1 19'.') Solanaceae Cestroideae Salpiglossideae Schizunrhus S. alpestris

1 3a-Hydroxytropane (58, 59, 60') 67 3a-Hydroxy-7/3-angeloyloxytropane (58.59. 60') 69 3a-Senecioyloxy-7P-hydroxytropane (58, 59. 607) S. grahamii

1 12 67 69 188 189

3a-Hydroxytropane (57, 58, 59, 607) 3a-Senecioyloxytropane (57) 3a-Hydroxy-7P-angeloyloxytropane(58, 59, 607) 3a-Senecioyloxy-7~-hydroxytropane (58, 59, 607) Schizanthine C (57*) Schizanthine D (57*)

1. TROPANE ALKALOIDS

45

TABLE 111 (continued) 190 196 203 S . hookerii 1 12 57

Schizanthine E (57*) Schizanthine X (IS]'*) Graharnine (1527*)

3a-Hydroxytropane (58, 59, 6@') 3a-Senecioyloxytropane (58, 59, 607) 3a,6P-Dihydroxytropane (58, 59, 6U7) 66 3a-Hydroxy-7P-tigloyloxytropane(58, 59, 60') 67 3a-Hydroxy-7~-angeloyloxytropane(58, 59, 602*') 69 3a-Senecioyloxy-7P-hydroxytropane (58, 59, 60?*7, S . littoralis 1 3a-Hydroxytropane (58, 59, 6U7) 12 3a-Senecioyloxytropane (58, 59, 607*) 57 3a,6P-Dihydroxytropane (58, 59, 6U7) 67 3a-Hydroxy-7P-angeloyloxytropane(58, 59, 607) 69 3a-Senecioyloxy-7P-hydroxytropane (58. 59, 60') S. pinnatus Ruiz et Pav. 1 3a-Hydroxytropane (153) 66 3a-Hydroxy-7P-tigloyloxytropane (153) 67 3a-Hydroxy-7~-angeloyloxytropane (153) 69 3a-Senecioyloxy-7~-hydroxytropane (153) 103 3a-( I '-Methylrnesaconyloxy)-7~-tigloyloxytropane (153') 104 301-(1'-Methylitaconyloxy)-7~-tigloyloxytropane (153*) 105 3a-( 1'-Methyrnesaconyloxy)-7~-angeloyloxytropane (153") 106 3a-( I'-Methylitaconyloxy)-7~-angeloyloxytropane (153') 108 3a-(3'-Ethoxycarbonylrnethacryloyloxy)-7~-senecioyloxytropane (15d7) 109 3a-( I '-Ethylitaconyloxy)-7~-angeloyloxytropane (153*) 110 3a-( I '-Ethylrnesaconyloxy)-7~-tigloyloxytropane (153*) 111 3a-( I '-Ethylitaconyloxyl)-7~-tigloyloxytropane (153*) 195 Schizanthine B (15d7) Solanaceae Solanoideae Solaneae Solaninae Cyphomandra C. betacea Sendt. 1 3a-Hydroxytropane (1.559 33 (*)-3a-Tropoyloxytropane (IS?') 50 3P-Hydroxytropane (1552) 52 3P-Tigloyloxytropane ( 1H 2') 175 3-Oxotropane (155*) Solanaceae Solanoideae Solaneae Physalinae

(continued)

46

MAURI LOUNASMAA A N D TARJA TAMMINEN

TABLE 111 (continued) Physalis P . alkekangi L. 1 3a-Hydroxytropane (156,*1572) 11 3a-Tigloyloxytropane (156,2157,2 1 M 2 ) 17 3a-Tigloyloxytropane N-oxide (1572*) 50 3P-Hydroxytropane (156,2.1572) 52 3P-Tigloyloxytropane (156,*1572) (Notes: 156: P . alkekangi L. var franchettii; 158: P . alkekangi L. var francheftii Hort f. bunyardii Makino) P . peruviana L. 1 3a-Hydroxytropane ( 1 5 8 ~ ) 11 3a-Tigloyloxytropane ( 1 5 8 ~ ) 5 1 3P-Acetoxytropane ( 1 5 8 ~ ) 176 Ip-Hydroxytropane (158a,159,* 160, 161) Withania W . somnifera Dunal 1 3a-Hydroxytropane (162,21632) 11 3a-Tigloyloxytropane (20,2163*) 50 3P-Hydroxytropane (162,*163*) Solanaceae Solanoideae Jabroseae Laruu L. pubiflora 33 ( *)-3a-Tropoyloxytropane (1641,3.4) 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (1641,3.4) L. uenenosa Phil. 33 (+.)-3a-Tropoyloxytropane(165) 34 ( - )-3a-Tropoyloxytropane (165) 140 ( - )-3a-Tropoyloxy-6~,7P-epoxytropane (165) Salpichroa S . organifolia (Lam.) Baillon 1 3a-Hydroxytropane (166*) 34 ( - )-3a-Tropoyloxytropane (166*’) 50 3P-Hydroxytropane (166*) Solanaceae Solanoideae Datureae Dutura, Sect. Brugmansia D . candida (Pers.) Saff. 1 3a-Hydroxytropane (167,2,430?3’) 11 3a-Tigloyloxytropane (30*) 27 (?)-3a-Tropoyloxynortropane (30,2.7168”) 28 ( -)-3a-Tropoyloxynortropane (167’.2.4) 33 (rt-)-3a-Tropoyloxytropane(30,2.7167,* 169,’ 170,1,4 171 , I 172,l 173‘) 34 (-)-3a-Tropoyloxytropane (30,7’ 167,2.4168, 174,1.2.4.5 175,*.j176.l.’ 177, 17a5)

1. TROPANE ALKALOIDS

47

TABLE 111 (continued) 35 52 93 121 122

( -)-3a-(2’-Hydroxy-3’-phenylpropionyloxy)tropane(61?)

3P-Tigloyloxytropane (61?) 3a-Tigloyloxytropane (167?) 3a,7P-Dihydroxy-6P-tigloyloxytropane ( I 79’) 3a-Tigloyloxy-6~,7P-dihydroxytropane (30,*,7167,1.2.4.’1687”) 128 3a-(2’-Hydroxy-3’-phenylpropionyloxy)-6~,7~-dihydroxytropane ( I 79’*) 130 3a,7P-Ditigloyloxy-6P-hydroxytropane (SO,? 167*) 138 3a-Apotropoyloxy-6~,7~-epoxytropane (30,71687’!) 139 3a-Tropoyloxy-6~,7~-epoxynortropane (30,2,7167,1.5168,7’!180’) 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (30,?,’ 167,‘ ~ 168,’ ~ 169,7 3 171, 173,’ 174,‘.2.4.’175,2.4.’ 176,‘.’ 177, 178,’ 180,’ 181.‘.4.5182‘) 142 ( - )-3a-(2’-Hydroxytropoyloxy-6~,7~-epoxytropane ( I 73’) (Notes: 168: D. aurea and D . candida separately; 169: Methysticodendron amesianum Schultes; 172: Brugmansia aurea and B. arborea separately; 173: D . pittieri; 175: D . arborea or B . candida; 176, 171: D . arborea; 178: Brugmansia arborea and B. candida separately; 180: Brugmansia candida; 181, 174, 177: D. urborea L.) D . chbrantha Hook. 34 ( - )-3a-Tropoyloxytropane( 1498) 140 ( - )-3a-Tropoyloxy-6~,7~-epoxytropane (149x) D . cornigera Hook. 27 (~)-3a-Tropoyloxynortropane(18,’,2.5.6 183’?) 28 (-)-3a-Tropoyloxynortropane ( I @ ) 33 (t)-3a-Tropoyloxytropane(182) 34 (-)-3a-Tropoyloxytropane (18,?183’*) 35 ( - )-3a-(2’-Hydroxy-3’-phenylpropionyloxy)tropane(61*) 52 3P-Tigloyloxytropane (612, 57 3a,7P-Dihydroxytropane (18?) 66 3a-Hydroxy-7P-tigloyloxytropane(1847*) 68 3a-Tigloyloxy-7~-hydroxytropane (1847”) 93 3a.7P-Ditigloyloxytropane (18,* 183’?) 130 3a ,7P-Ditigloyloxy-6~-hydroxytropane (18, 183 I?’!) 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane(18,1.?.3.5.6 /83’*) D.sanguinea R. and P. 1 3a-Hydroxytropane (1@) 3 3a-Acetoxytropane (185,81867*) 11 3a-Tigloyloxytropane (185,8187*) 33 (?)-3a-Tropoyloxytropane (136,1.2.4172.’ 188?) 34 ( -)-3a-Tropoyloxytropane (149,8185,8 1887) 35 ( - )-3a-(2‘-Hydroxy-3’-phenylpropionyloxy)tropane (187’.*) 63 3a-Tigloyloxy-7~-hydroxynortropane(1897*) 68 3a-Tigloyloxy-7P-hydroxytropane (1858)

‘

(continued)

48

MAURI LOUNASMAA A N D TARJA T A M M I N E N

TABLE 111 (continued) 71 72 82 93 122 129 130 138 139 140 142

3a-Isovaleryloxy-6~-hydroxytropane (136 1.3.4.’) 3a.7P-Diacetoxytropane (1XiR) 3a-Tigloyloxy-7P-acetoxytropane (1867) 3a.7P-Ditigloyloxytropane(188*) 3a-Tigloyloxy-6P,7P-dihydroxytropane (187‘.’) 3a-Tigloyloxy-6~-hydroxy-7~-isovaleryloxytropane (/87’*) 3a,7P-Ditigloyloxy-6P-hydroxytropane 3a-Apotropoyloxy-6~,7~-epoxytropane 3a-Tropoyloxy-6~,7~-epoxynortropane (18S7) ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (136.1.’.4149,x 172. 185,x 188’,’) ( - )-3a-(2’-Hydroxytropoyloxy)-6~,7~-epoxytropane (190’*) (Notes: 149: D . rosei and D. sanguinea separately; 172: Brugmansia sanguinea)

D . .suaveolens H . and B. ex Willd. 1 3a-Hydroxytropane (191’) 3 3a-Acetoxytropane (186, 1 9 / * ) 27 (+)-3a-Tropoyloxynortropane (191 7 , 28 ( - )-3a-Tropoyloxynortropane(192) 33 (+ )-3a-Tropoyloxytropane( I 91 ?.’) 34 ( - )-3a-Tropoyloxytropane(1931,4) 35 ( - )-3a-(2’-Hydroxy-3’-phenylpropionyloxy)tropane (/9/?) 68 3a-Tigloyloxy-7p-hydroxytropane (19/2.7) 95 3a-Tigloyloxy-7P-(2’-methylbutyryloxy)tropane ( I 91 ? ) 121 3a,7P-Dihydroxy-6~-tigloyloxytropane (/917*) 122 3a-Tigloyloxy-6P,7P-dihydroxytropane (191,2.7 193’,4) 130 3a,7P-Ditigloyloxy-6P-hydroxytropane (19/*.7) 138 3a-Apotropoyloxy-6~.7~-epoxytropane (1917 , 139 3a-Tropoyloxy-6~,7~-epoxynortropane ( I 91 5.7) 140 ( - )-3a-Tropoyloxy-6~.7~-epoxytropane (191,?.71931.4) Datura, Sect. Dutra D . discolor Bernh. 1 3a-H ydrox ytropane ( 194’,7) 3 3a-Acetoxytropane (194’?) 27 (+)-3a-Tropoyloxynortropane (194*”) 33 (+)-3a-Tropoyloxytropane (194’) 34 ( - )-3a-Tropoyloxytropane(185.’ 194,7 195’-3.4.’.6 ) 35 ( - )-3a-(2’-Hydroxy-3’-phenylpropionyloxy)tropane (194’) 50 3P-Hydroxytropane (1942,7) 82 3a-Tigloyloxy-7p-acetoxytropane(1947?) 93 3a,7P-Ditigloyloxytropane(1942) 122 3a-Tigloyloxy-6~,7P-dihydroxytropane (1942.7) 130 3a.7p-Ditigloyloxy-6P-hydroxytropane(194?) 138 3a-Apotropoyloxy-6P,7P-epoxytropane ( 1947) 139 3a-Tropoyloxy-6p,7P-epoxynortropane( I 942.7) 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (185, I 94,’.3.4.5.6,7 195, ‘.3.45.h I 96’.2.4)

’

1. TROPANE

ALKALOIDS

49

TABLE 111 (confinued) ~~

~

D.innoxia Mill. 1 3 4 8 11 13 24 27 28 33

34 35

50 51 52 53 57 59 66 68 70 82 85 87 89 90 93 95 96 112 114 115 120 121 122 130 136 138 139

3a-Hydroxytropane (62,’ 185,’ 197,’ 198,? 199,’ 200.8.’ 201‘.7) 3a-Acetoxytropane(185,’ 199,’ 202’) 3a-Propionyloxytropane(1Y P ) 3a-Isobutyryloxytropane(199?) 3a-Tigloyloxytropane(61,? 197,’ 199’) (+)-3a-(2’-Methylbutyryloxy)tropane(199’) 3a-Apotropoyloxytropane(199,l.’ 201,’.7 203,’ 204I.j) (t)-3a-Tropoyloxynortropane (183,’?204l.’) ( - )-3a-Tropoyloxynortropane (144,I” 197l.?) (+)-3a-Tropoyloxytropane (62,’ 17/,’.’.’ /98,1.’-’,5.h,7 204,I.j 205,7206.’”207h) ( - )3a-Tropoyloxytropane(35,5139,n 149,n 176.l.’ f83.1’ /85,’.x197.l.’ 198,l.2.7.3.6..5 199,2.4.1.5 200,8,y201,7.’ 203, 207,‘208, 209, 210,’ 2 f f 3 212,I.j ‘ 213,‘.2.4214‘) ( -)-3a-(2’-Hydroxy-3’-phenylpropionyloxy)tropane (6/’) 3P-Hydroxytropane(62,’ 197,’ 198,’ 199.’ 2IO7.’) 3p-Acetoxytropane(199’) 3P-Tigloyloxytropane(62,?199’) 3P-(2’-Methylbutyryloxy)tropane(199’) 3a,7P-Dihydroxytropane(197,’ 20/’,7) ( + )-3a-Acetoxy-7P-hydroxytropane (1W) 3a-Hydroxy-7~-tigloyloxytropane(185,x/99’.4) 3a-Tigloyloxy-7P-hydroxytropane(185,8/99’,4) ( + )-3a-Hydroxy-7p-(2’-methylbutyryloxy)tropane (199’) 3a-Tigloyloxy-7P-acetoxytropane (199’) 3a-Tigloyloxy-7P-propionyloxytropane(67,’* 199,’ 212’) (-)-3a-Tropoyloxy-6P-hydroxytropane (185,‘ 200,‘.’ 21@) 3a-Tropoyloxy-7~-hydroxytropane (214‘) 3a-Tigloyloxy-7P-isobutyryloxytropane (/99?) 3a,7P-Ditigloyloxytropane (62,’ 183,12185,n197,’ 198,’ 199.’ 200,’.’ 20/.3.7 212,? 215’*) 3a-Tigloyloxy-7P-(2‘-methylbutyryloxy)tropane(199’) 3a-Tigloyloxy-7~-isovaleryloxytropane (199:) 3a-Tropoyloxy-7P-tigloyloxytropane (I99:) 3a-Tropoyloxy-7~-isovaleryloxytropane ( I99’) 3a-Tropoyloxy-7P-(2’-methylbutyryloxy)tropane ( I99’) 3a,6P,7P-Trihydroxytropane(2008.9) 3a,7p-Dihydroxy-6P-tigloyloxytropane (2013.7) 3a-Tigloyloxy-6P,7P-dihydroxytropane (62,‘ 144,‘O 197,l.’ 198,1.2.7 199,1.4 200,8.y205,’* 208, 211,‘ 212l.’) 3a,7P-Ditigloyloxy-6P-hydroxytropane (6,’ 183.’? 185,x197,? /98.? 199.?,‘ 212’) 3a-Hydroxy-6P.7P-epoxytropane (203’) 3a-Apotropoyloxy-6p.7P-epoxytropane (197,’ fY9.’,’.’ 201 ,3.7 204I.j) 3a-Tropoyloxy-6~,7~-epoxynortropane (183,’’ 197,’.’ 200,’,y 204I.j)

50

MAURI LOUNASMAA AND TAWA TAMMINEN TABLE I11 (continued) 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (35,x62,’ 13Y,K144,‘O 14Y,x171,‘,2,3 176,1.5 /83,12 185,l.8197,1.2198

1.2.3.5.6.7

199 l.2.4.5200,s,Y201,3.7203,l.?.42~,1.3

205,7 206,” 207,6 208,

i?ll,I 212.l.’ >13,1.2.4 2/4,8216, 217’) 141 ( iz)-3a-Tropoyloxy-6P,7P-epoxytropane(622) 193 a-Scopadonnine (3963) 194 P-Scopadonnine (3963) (Notes: 61, 149: D. innoxia and D.meteloides separately; 144, 197, 205: D.meteloides D.C. ex Dun.; 21 1: D. innoxia and D. meteloides separately, diploid and haploid plants)

D . leichardtii F. Muell. ex Benth. 11 3a-Tigloyloxytropane ( 2 / 8 ! ) 34 (-)-3a-Tropoyloxytropane ( 6 1 , l 13Y,x 185,’ 21a7) 35 ( - )-3a-(2‘-Hydroxy-3’-phenylpropionyloxy)tropane (61 ’) 52 3P-Tigloyloxytropane (61,?218?) 122 3a-Tigloyloxy-6~,7P-dihydroxytropane (218?) 130 3a,7P-Ditigloyloxy-6P-hydroxytropane (218?) 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (61,? 185,’ 2187) 193 a-Scopadonnine (132l) 194 P-Scopadonnine (132’) D . leichurdtii F. Muell. ex Benth. ssp. pruinosa (Greenm.) Hammer 1 3a-Hydroxytropane (2197) 11 3a-Tigloyloxytropane (2192.7) 24 3a-Apotropoyloxytropane (2197) 27 (+)-3a-Tropoyloxynortropane (21Y7) 28 ( - )-3n-Tropoyloxynortropane (21Y2) 33 (+)-3a-Tropoyloxytropane (21Y2.7) 35 ( - )-3a-(2’-Hydroxy-3’-phenylpropionyloxy)tropane (21Y?.’) 50 3P-Hydroxytropane (219’) 52 3P-Tigloyloxytropane (2lY?.’) 93 3a,7P-Ditigloyloxytropane(219l) 122 3a-Tigloyloxy-6P,7~-dihydroxytropane (2 19 130 3a,7P-Ditigloyloxy-6P-hydroxytropane(21Y2) 138 3a-Apotropoyloxy-6~,7~-epoxytropane (2197) 139 3a-Tropoyloxy-6~,7~-epoxynortropane (21Y 7, 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (2192.7) (Note: 219: D. pruinosa Greenm.) D.metel L 1 3a-Hydroxytropane (2202) 24 3a-Apotropoyloxytropane (2215.b) 28 ( - )-3a-Tropoyloxynortropane (144IO) 29 3a-(4’-Methoxybenzoyloxy)tropane(222 I*) 33 (?)-3a-Tropoyloxytropane(175,2+4171,1,2,3 220, ’ 223,1.32247) 34 ( - )-3a-Tropoyloxytropane (35,5144,” 149,K175,‘,3,6 220,?223,’ 225,’226,1.4.5 227,3*6228’) 50 30-Hydroxytropane (2202) 93 3a,7P-Ditigloyloxytropane(2202) 122 3a-Tigloyloxy-6P.7p-dihydroxytropane(216)

1.

51

TROPANE ALKALOIDS

TABLE I11 (continued) 130 3a,7~-Ditigloyloxy-6P-hydroxytropane (220’) 139 (?) 3a-Tropoyloxy-6~,7~-epoxynortropane (229) 140 ( - )-3a-Tropoyloxy-6~,7~-epoxytropane (35,s 144,“ 149,x I71 ,‘.I.’ 174, 175,’.2.3,5.6 220,’ 221,5.6223,‘.’ 224,’ 22.5,’ 226,‘.4.s227,’.’ 228‘) D. metel L. var. fastuosa (Bernh.) Danert 1 3a-Hydroxytropane (185,’ 220,’ 230?) 3 3a-Acetoxytropane (230’) 11 3a-Tigloyloxytropane (230’) 27 ( ?)-3a-Tropoyloxynortropane (230’) 33 (?)-3a-Tropoyloxytropane (223,’ 230’) 34 (-)-3a-Tropoyloxytropane (48,’,?139,’ 149,n 176,1,5185,’,X220.’ 223,I.j 226, 1.4.5 227’,6) 50 3P-Hydroxytropane (220,’ 230?) 52 3P-Tigloyloxytropane (61,* 230’) 72 3a,7P-Diacetoxytropane (185’) 93 3a,7P-Ditigloyloxytropane(220,*230’) 122 3a-Tigloyloxy-6P.7P-dihydroxytropane(230’) 220, 230’) 130 3a,7~-Ditigloyloxy-6P-hydroxytropane(l85, (230’) 138 3a-Apotropoyloxy-6~,7~-epoxytropane (230’) 139 3a-Tropoyloxy-6P,7P-epoxynortropane (48,I.’ 139,’ 149,R220,’ 226,’.4,’ 140 ( -)-3a-Tropoyloxy-6~,7~-epoxytropane 227.’.‘ 230,’ 231’.2.6) (Notes: 139, 176, 185: D. fastuosa; 149: D. fastuosa var. uiolacea; 223: D. fastuosa var. black) D. metel L. var. metel 33 (?)-3a-Tropoyloxytropane (2325) 34 (-)-3a-Tropoyloxytropane (176,’.5185,’232?) 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (176,’,5232,>23J5) (Notes: 174, 232: D. alba Nees; 176: D. metel L. f. alba, 233: D. alba) D. metel var. rubra 140 ( - )-3a-Tropoyloxy-6P.7~-epoxytropane (216) D . wrightii Regel 1 3a-Hydroxytropane (185’) 2 3-Acetoxynortropane (1@) 3 3a-Acetoxytropane (185’) 11 3a-Tigloyloxytropane (185’) 34 (-)-3a-Tropoyloxytropane (185.’.’211,’ 234’) 51 3P-Acetoxytropane (185‘) 59 ( )-3a-Acetoxy-7P-hydroxytropane (185’) 68 3a-Tigloyloxy-7~-hydroxytropane (185’) 93 3u,7~-Ditigloyloxytropane (185’) 122 3a-Tigloyloxy-6P,7P-dihydroxytropane (21f ’)

’ *

+

(continued)

52

MAURI LOUNASMAA A N D TARJA TAMMINEN

TABLE 111 (continued) 130 3a,7P-Ditigloyloxy-6~-hydroxytropane (185x) 140 ( - )-3a-Tropoyloxy-6P,7~-epoxytropane (/85,'.' 2 / 1 , ' 234'.') Doturci. Sect. Ceratocaulis D . ceratocaula Ort. 33 (*)-3a-Tropoyloxytropane (235 1.'.'.49 34 (-)-3a-Tropoyloxytropane (185') 70 ( + )-3a-Hydroxy-7~-(2'-methylbutyryloxy)tropane (6j7) 140 ( - )-3a-Tropoyloxy-6~,7P-epoxytropane (185,'235'.'.'.4 ') Datura, Sect. Datura D . ferox L. 1 3a-Hydroxytropane (62,'" 185x) 3 3a-Acetoxytropane ( 1858) 11 3a-Tigloyloxytropane (62,' 185') 24 3a-Apotropoyloxytropane (236'.4.6.") 27 (*)-3a-Tropoyloxynortropane (204') 28 ( - )-3a-Tropoyloxynortropane (192') 33 (*)-3a-Tropoyloxytropane (204.' 237) 34 ( -)-3a-Tropoyloxytropane (/49,* 185,'.* 238, 239l) 35 ( - )-3a-(2'-Hydroxy-3'-phenylpropionyloxy)tropane( 6 / * ) 50 3P-Hydroxytropane (62'") 68 3a-Tigloyloxy-7P-hydroxytropane ( 185') 72 3a.7j3-Diacetoxytropane (1@) 87 ( - )-3a-Tropoyloxy-6~-hydroxytropane ( /85,x2407) 93 3a.7P-Ditigloyloxytropane(62.' 185') 122 3a-Tigloyloxy-6P,7P-dihydroxytropane(62,' 238. 239,? 241 I") 130 3a,7P-Ditigloyloxy-6P-hydroxytropane(62,? 185,' 215.?* 242') 138 3a-Apotropoyloxy-6~,7~-epoxytropane (204') 140 ( - )-3a-Tropoyloxy-6P,7~-epoxytropane (62.' /49,1.8204,' 236.1.3.4.6,'1 237, 238,' 241'.'') D . y~~ercifolia H. B. K. 1 3a-Hydroxytropane (185*) 3 3a-Acetoxytropane (185') 33 (*)-3a-Tropoyloxytropane (/75'.'.6) 34 ( - )-3a-Tropoyloxytropane (149.n175.'.'.',h 185') 60 3a-Hydroxy-7P-acetoxytropane (185H) 72 3a.7P-Diacetoxytropane ( l M R ) 130 3a.7P-Ditigloyloxy-6p-hydroxytropane( 1 8 9 ) 140 ( - )-3a-Tropoyloxy-6p,7P-epoxytropane (149.x 175.'.?.'.' 185x") D . stramoniirm L. 1 3a-Hydroxytropane (61,' 70,7 /@) 3 3a-Acetoxytropane (189') 24 3a-Apotropoyloxytropane (203,1.2.4 204,l.' 2 4 3 ' ) 27 (+)-3a-Tropoyloxynortropane (204,l.' 243') 28 ( -)-3a-Tropoyloxynortropane (192,' 243') 33 ( *)-3a-Tropoyloxytropane ( I71,I,?,'204 ,I,' 243,' 244,' 245, 246'") 34 (-)-3a-Tropoyloxytropane (35,' 70,' 149,' 175.' 176,',-' 177, 185.'~x203,1~'.3.4~' 223,'.2.4.6243,' 244.' 246," 2471.'.4)

1. TROPANE ALKALOIDS

53

TABLE I11 (continued) 35 38 39 50 52 60

( - )-3a-(2'-Hydroxy-3'-phenylpropionyloxy)tropane(61')

3a-Tropoyloxytropane N-oxide 1 (248',2,3,4,5,6) 3a-Tropoyloxytropane N-oxide 2 (248'.'.3.4.".h) 3P-Hydroxytropane (61?) 3P-Tigloyloxytropane (243') 3a-Hydroxy-7P-acetoxytropane ( /85R) 68 3a-Tigloyloxy-7P-hydroxytropane (1858) 72 3a,7P-Diacetoxytropane( / 8 5 * ) 93 3a,7P-Ditigloyloxytropane(185,* 24Y2*) 122 3a-Tigloyloxy-6~,7P-dihydroxytropane (61,? 243') 130 3a,7P-Ditigloyloxy-6~-hydroxytropane (185,* 2/5?*) 136 3a-Hydroxy-6P,7/3-epoxytropane (203'.?,3.4,6) 138 3a-Apotropoyloxy-6P.7P-epoxytropane(2M3) 139 3a-Tropoyloxy-6P,7P-epoxynortropane (204'.') 140 (-)-3a-Tropoyloxy-6P,7P-epoxytropane (35.5 70.7 145,' 149,' 17/,'.'.' 176,' 185.' 203.'.2.'.4.5204,I.' 223,'.2.4.h243,' 244,3 245, 246.'" 250) 143 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane N-oxide (248'.?.7.4.5,.h) D . stramonium L. var. godronii Danert 130 3a,7P-Ditigloyloxy-6P-hydroxytropane (215?*) (Note: 215: D. tatula L. var. inermis) D. stramonium L. var. inermis (Jacq.) Lundstr. 34 (-)-3a-Tropoyloxytropane (139,' 149,' 176',3,5) 140 ( - )-3a-Tropoyloxy-6P.7P-epoxytropane (149,' 176'.') (Note: 176: D. inermis) D. stramonium L. var. strumonium 34 ( - )-3a-Tropoyloxytropane(13Y,x 149*) 140 ( - )-3a-Tropoyloxy-6~,7P-epoxytropane (1499 D. strumonium L. var tutula Torr. 34 ( - )-3a-Tropoyloxytropane(149,H176,' 234,'.? 238, 25/,'.1.7252,' 253) 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (149,' 176,' 234,' 238, 251,'.'.7 252,7 253)

D . innoxia

11 34 87 93 122

130 138 139 140 D . ferox x 1 11

(Note: 176, 251. 252, 253: D. tatula L.) x leichardtii 3a-Tigloyloxytropane(20Y2) ( - )-3a-Tropoyloxytropane(2092.7) ( - )-3a-Tropoyloxy-6P-hydroxytropane (20Y7'?) 3a,7P-Ditigloyloxytropane(20Y2) 3a-Tigloyloxy-6/3,7P-dihydroxytropane (,?OY?,') 3a,7P-Ditigloyloxy-6P-hydroxytropane (20Y2) 3a-Apotropoyloxy-6P,7P-epoxytropane(2097) 3a-Tropoyloxy-6P,7P-epoxynortropane(20Y7) ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (20Y?,7) stramonium L. 3a-Hydroxytropane(254*) 3a-Tigloyloxytropane(254?)

(continued )

54

MAURI LOUNASMAA A N D TARJA TAMMINEN

TABLE 111 (continued) 34 87 93 122 130 140

( - )-3a-Tropoyloxytropane(240,' 255,7 254?) ( - )-3a-Tropoyloxy-6~-hydroxytropane (240,' 256')

3a,7/3-Ditigloyloxytropane (254?) 3a-Tigloyloxy-6~,7p-dihydroxytropane (254?) 3a,7P-Ditigloyloxy-6P-hydroxytropane (254*) (-)-3a-Tropoyloxy-6P,7P-epoxytropane(240,l 254,l 255,7 2571,4) D . discolor x strumonium var. godronii 1 3mHydroxytropane (2587) 24 3a-Apotropoyloxytropane ( 2 S 7 ) 27 (?)-3a-Tropoyloxynortropane (2587) 34 ( -)-3a-Tropoyloxytropane (258') 50 3P-Hydroxytropane( 2 M 7 ) U 2 3a-Tigloyloxy-6p,7P-dihydroxytropane (2H7) 138 3a-Apotropoyloxy-6~,7~-epoxytropane (2H7) 139 3a-Tropoyloxy-6p,7~-epoxynortropane (2S7) 140 ( - )-3a-Tropoyloxy-6~,7~-epoxytropane (2S7) D . candidu x candida 1 3a-Hydroxytropane (185,8259n) 3 3a-Acetoxytropane (259,* 26Un) 5 3a-(Hydroxyacetoxy)tropane (26U8) 11 3a-Tigloyloxytropane( 185n) 22 3a-Phenylacetoxytropane (260') 24 3a-Apotropoyloxytropane (259,* 260') 27 (+)-3a-Tropoyloxynortropane (1687) 33 (+)-3a-Tropoyloxytropane (172') 34 ( - )-3cu-Tropoyloxytropane(168,7 185,' 26Un) 35 ( - )-3a-(2'-Hydroxy-3'-phenylpropinyloxy)tropane(26Un) 50 3P-Hydroxytropane (259,* 260') 51 3P-Acetoxytropane (260') 52 3P-Tigloyloxytropane(259,n26U8) 57 3a,7P-Dihydroxytropane (26Un) 59 ( + )-3a-Acetoxy-7p-hydroxytropane (259,* 26U8) 61 3a-Hydroxy-7~-propionyloxytropane (2598) 65 3a-Hydroxy-7P-isobutyryloxytropane(2598) 66 3a-Hydroxy-7~-tigloyloxytropane (185,8259,* 26Un) 68 3a-Tigloyloxy-7p-hydroxytropane (185,* 259,n 26@) 70 ( +)-3a-Hydroxy-7~-(2'-methylbutyryloxy)tropane(117, 26Un) 80 3a-Phenylacetoxy-7p-hydroxytropane(260') ( 185,8259,826U2) 82 3a-Tigloyloxy-7P-acetoxytropane 84 3a-Apotropoyloxy-7~-hydroxytropane (259,* 26O8) 85 3a-Tigloyloxy-7~-propionyloxytropane (26U8) (259,n260') 87 ( - )-3a-Tropoyloxy-6p-hydroxytropane 89 3a-Tropoyloxy-7p-hydroxytropane (26O8) 90 3a-Tigloyloxy-7P-isobutyryloxytropane(2608) 93 3a,7P-Ditigloyloxytropane(185,' 260') 95 3a-Tigloyloxy-7~-(2'-1nethylbutyryloxy)tropane (26Un) (26Un) 96 3a-Tigloyloxy-7~-isovaleryloxytropane

1.

TROPANE ALKALOIDS

55

TABLE I11 (continued) 122 3a-Tigloyloxy-6~,7~-dihydroxytropane (1687)

3a-Tigloyloxy-6~-hydroxy-7~-isovaleryloxytropane (260’) 3a,7P-Ditigloyloxy-6P-hydroxytropane(185,* 260’) 3a-Apotropoyloxy-6~,7~-epoxytropane (168,7259,’ 260’) 3a-Tropoyloxy-6~,7~-epoxynortropane (1687) ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (168,7 185,’ 2608) 175 3-Oxotropane (260’) (Notes: 168, 185, 260: D . aurea X candida; 172: Brugmansia aurea x candida) Solanaceae Solanoideae Solandreae Solandra S. grandij7ora Sw. 1 3a-Hydroxytropane (64’,7) 3 3a-Acetoxytropane (642,7) 11 3a-Tigloyloxytropane (642.7) 13 ( +)-3a-(2’-Methylbutyryloxy)tropane(64*,’) 27 (?)-3a-Tropoyloxynortropane (647) 33 (t)-3a-Tropoyloxytropane(647) 34 ( - )-3a-Tropoyloxytropane ( 6 4 2 ) 35 ( - )-3a-(2’-Hydroxy-3’-phenylpropionyloxy)tropane(64*) 50 3P-Hydroxytropane (64’) 52 3P-Tigloyloxytropane (642.7) 140 ( - )-3a-Tropoyloxy-6P,7p-epoxytropane (642.7) S . longij7ora Tussac 27 (?)-3a-Tropoyloxynortropane (261, 262’) 28 (-)-3a-Tropoyloxynortropane (261, 262’) 33 (?)-3a-Tropoyloxytropane (261, 262’) 34 (-)-3a-Tropoyloxytropane (261, 262’) S. guttata D. Don ex Lindley 1 3a-Hydroxytropane ( 6 4 1 , 2 , 4 ) 3 3a-Acetoxytropane (642) 11 3a-Tigloyloxytropane (64”.*) 13 ( + )-3a-(2’-Methylbutyryloxy)tropane(64”.*) 27 (?)-3a-Tropoyloxynortropane (64”.2.4) 28 ( - )-3a-Tropoyloxynortropane ( 6 4 ” ) 33 (?)-3a-Tropoyloxytropane (64’,2,4) 35 ( - )-3a-(2’-Hydroxy-3’-phenylpropionyloxy)tropane(64*’) 50 3P-Hydroxytropane (64’,2.4) 52 3P-Tigloyloxytropane (64*,4) (644) 139 (?)-3a-Tropoyloxy-6~,7~-epoxynortropane (64’.2.4) 140 ( - )-3a-Tropoyloxy-6~,7~-epoxytropane S . hartwegii N . Br. 1 3a-Hydroxytropane (642.7) 11 3a-Tigloyloxytropane (647?) 129 130 138 139 140

(continued)

56

MAURI LOUNASMAA A N D T A N A TAMMINEN

TABLE I11 (continued)

13 27 33 50 52 140

+

( )-3a-(2‘-Methylbutyryloxy)tropane(642.7’) ( *)-3a-Tropoyloxynortropane (647)

(+)-3a-Tropoyloxytropane (642,7) 3P-Hydroxytropane (642.7) 3P-Tigloyloxytropane (642) ( - )-3a-Tropoyloxy-6~,7P-epoxytropane (642.7) S . hirsuta Dun. 1 3a-Hydroxytropane (642.7) 3 30-Acetoxytropane (642.7) 11 3a-Tigloyloxytropane (642.7) 13 ( + )-3a-(2’-Methylbutyryloxy)tropane(642.7) 27 (C)-3a-Tropoyloxynortropane(642,7) 33 (~)-3a-Tropoyloxytropane(642,7) 35 ( - )-3a-(2‘-Hydroxy-3’-phenylpropionyloxy)tropane (642.7) 50 3P-Hydroxytropane (642.7) 52 3P-Tigloyloxytropane 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (642.7) S . macrocantha Dun. 1 3a-Hydroxytropane (642.7) 3 3a-Acetoxytropane (64?) 11 3a-Tigloyloxytropane (642.7’) 13 ( + )-3a-(2’-Methylbutyryloxy)tropane(642.7’) 27 (C)-3a-Tropoyloxynortropane (642.7) 33 (+)-3a-Tropoyloxytropane (642.7) 50 3P-Hydroxytropane (642.7) 52 3~-Tigloyloxytropane(642.7) 140 ( - )-3a-Tropoyloxy-6~,7P-epoxytropane (642,7) Solanaceae Solanoideae Nicandreae Nicandra N . physaloides (L.) Gaertn. 175 3-Oxotropane (263?*) Solanaceae Atropoideae At ropeae Atropa A . acuminata Royle 24 3a-Apotropoyloxytropane (32I.*) 34 ( - )-3a-Tropoyloxytropane (321.2) 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (32 A . baetica Willk. 24 3a-Apotropoyloxytropane (321,2) 34 (-)-3a-Tropoyloxytropane (321,2) 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (32’) A . belladonna L. 1 3a-Hydroxytropane (19,2203,6264,2265’.2,s) 22 3a-Phenylacetoxytropane (265 1.2.8)

1. TROPANE ALKALOIDS

57

TABLE I11 (continued) 24 3a-Apotropoyloxytropane (32,',' 203,6 204,' 264,'.?265'.2.x266,? 267.? 2682.3.4.5.lo) 21 ( ~)-3a-Tropoyloxynortropane(204) 28 ( - )-3a-Tropoyloxynortropane (265') 33 (?)-3a-Tropoyloxytropane (32,',? 171,'.*.' 204,' 244,',h264,'.?268'.2.'.J.S.h ) 34 ( - )-3a-Tropoyloxytropane (19,232,5.6139,' 149,' 174, 203,1.2.4.6 244.l.' 251,'.2.3.6,'265,'.3.x.9 266,2 267,2270") 38 3a-Tropoyloxytropane N-oxide I (248,1,2.3.4.5.6265 39 3a-Tropoyloxytropane N-oxide 2 (248,12.3.4.5.6 265 50 3P-Hydroxytropane (265*,*) 84 3a-Apotropoyloxy-7~-hydroxytropane (265?.') (265?,') 87 ( - )-3a-Tropoyloxy-6~-hydroxytropane 89 3a-Tropoyloxy-7P-hydroxytropane (2652.x) 136 3a-Hydroxy-6/3,7P-epoxytropane (203h) 138 3a-Apotropoyloxy-6~,7~-epoxytropane (265, 268'.'.'.' ) 140 (-)-3a-Tropoyloxy-6~,7P-epoxytropane (19,232,I.l 139,' 145,?149,' 171,',',' 2Oj',l.2.4.6 204,l 251,1.2.3.6.7 264 1.2 265,1.2.8 267,: 268,1.?.3.4.5.6269 2 2701 1 ) 143 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane N-oxide (248') 175 3-Oxotropane (265 I.*.*) 179 Ip,2a,3P-Trihydroxynortropane(271, 272, 273') 181 lp,2a,2P,6/3-Tetrahydroxynortropane(271, 272, 2 7 j 8 ) 182 Ip,2a,3P,4a-Tetrahydroxynortropane(271, 272, 273') 1911 Belladonnine (266.' 267.2 269,' 274) 192 (Note: 244: A . belladonna var. nigra) A. belladonna var. lutea 24 3a-Apotropoyloxytropane (32,'.?244') 33 (-t)-3a-Tropoyloxytropane(32,'.?244l.') 34 ( -)-3a-Tropoyloxytropane (244') A . caucasica Kreyer 34 ( - )-3a-Tropoyloxytropane (149') 140 ( - )-3a-Tropoyloxy-6~,7~-epoxytropane (149') A . komarovii Blin et Schal. 24 3a-Apotropoyloxytropane (32I.I) 33 (?)-3a-Tropoyloxytropane (32l.?) A . pallidi'ora Schonb.-Tern. 24 3a-Apotropoyloxytropane (32'.?) 33 (?)-3a-Tropoyloxytropane (32l.') 140 ( - )-3a-Tropoyloxy-6~,7P-epoxytropane (32' ) Hyoscyamus H . albus L. 1 3a-Hydroxytropane (146,R185,' 275,? 276,'.? 277') 3 3a-Acetoxytropane ( 1 8 j X ) 11 3a-Tigloyloxytropane (276l.') 24 3a-Apotropoyloxytropane (204.'.3275,' 2762.7) 27 (?)-3a-Tropoyloxynortropane (204',-'!) 28 ( - )-3a-Tropoylo~ynortropane(146~) (continued )

58

MAURI LOUNASMAA A N D TARJA TAMMINEN

TABLE 111 (continued)

33 34 35 50 51 52 87 89 138 139 140

(?)-3a-Tropoyloxytropane (204,',' 2762.7) ( - )-3a-Tropoyloxytropane (139,8.9146,' 149,8185,'~x214,8275,1~~277."278'~4) ( - )-3a-(2'-Hydroxy-3'-phenylpropionyloxy)tropane(276,* 277x) 3P-Hydroxytropane (185,' 277') 3P-Acetoxytropane (185') 3P-Tigloyloxytropane (2762.7) ( - )-3a-Tropoyloxy-6~-hydroxytropane (146,' 185,' 214,x277x) 3a-Tropoyloxy-7P-hydroxytropane(146,'' 214,' 277') 3a-Apotropoyloxy-6~,7~-epoxytropane (2O4,'.' 2762.7) 3a-Tropoyloxy-6~,7~-epoxynortropane (2O4'q ( - )-3a-Tropoyloxy-6~,7~-epoxytropane (139,8.9146,x149,' 185,'.' 204.l.' 214,' 275,1,2276,2.7277,' 278',4)

175 3-Oxotropane (185,' 277') H . oureus L . 1 3a-Hydroxytropane (2791,4) 24 3a-Apotropoyloxytropane (204l.') 27 (~)-3a-Tropoyloxynortropane (204'") 33 ( ?)-3a-Tropoyloxytropane (204I,') 34 (-)-3a-Tropoyloxytropane (149.x185,' 275,'.?2762.7) 50 3P-Hydroxytropane (276') 136 3a-Hydroxy-6P.7P-epoxytropane (2791,4) 138 3a-Apotropoyloxy-6~,7~-epoxytropane (204I.') 139 3a-Tropoyloxy-6~,7~-epoxynortropane (204',"!) 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (149,' 18.5,' 204,'.' 275,'.?276*.7) H . bohvmicus F. W . Schmidt 34 ( - )-3a-Tropoyloxytropane (35,' 139,s,9149,' 176,?.' 185) 140 (-)-3a-Tropoyloxy-6P,7P-epoxytropane (35,3139,x,y149,' 176,?.' 185) H . cunuriensis Ker. 34 ( - )-3a-Tropoyloxytropane (139,x,9185') 140 ( - )-3a-Tropoyloxy-6~,7P-epoxytropane (139,',' 185') H . desertorurn Tackh. 1 3a-Hydroxytropane (1858) 3 3a-Acetoxytropane (185') 34 ( - )-3a-Tropoyloxytropane (49,12.3.4.s.6 1851,x) 87 ( - )-3a-Tropoyloxy-6~-hydroxytropane (185') 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (49,1.3.4.5,6 185 I,') H . muficus L. 1 3a-Hydroxytropane (185,' 2791,4) 24 3a-Apotropoyloxytropane (204l.') 27 (?)-3a-Tropoyloxynortropane (2O4',') 33 (?)-3a-Tropoyloxytropane (2O4I.') 34 (-)-3a-Tropoyloxytropane (47,'.2.4,s.6 139,8*9149,' 185,'.' 203,',2208,'*4 2473.'+4,280',3)

50 136 139 140

3P-Hydroxytropane (185')

3a-Hydroxy-6P,7P-epoxytropane(279',4) 3a-Tropoyloxy-6~,7~-epoxynortropane (204'.') ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (47,* 139,8.9149,' 185,'.' 203,'.' 204 1.3

1. TROPANE ALKALOIDS

59

TABLE 111 (continued)

H . niger L. 1 3a-Hydroxytropane (70,7275?) 24 3a-Apotropoyloxytropane (70,7275?) 33 (?)-3a-Tropoyloxytropane ( I 71, 233) 34 (-)-3a-Tropoyloxytropane (35,370,’ 139,8,y149,n 176.’ 2/4,x231,s 233,2~f1,275,‘,~ 276,? 281,’ 282,8 283R) 38 3a-Tropoyloxytropane N-oxide I (2481,2,3.4,‘,6) 39 3a-Tropoyloxytropane N-oxide 2 (248’,’,3,4,5,6) 87 ( - )-3a-Tropoyloxy-6P-hydroxytropane(21@) 89 3a-Tropoyloxy-7P-hydroxytropane(21@) (70,7276?) 138 3a-Apotropoyloxy-6~,7~-epoxytropane (276?) 139 3a-Tropoyloxy-6~,7~-epoxynortropane (35,’ 70,’ 139,’ 145,‘ 149,K171,’,‘ 140 (-)-3a-Tropoyloxy-6~.7~-epoxytropane 176,3214,K231,’233, 241, 275,’.=276,?281.5282x) 141 (~)-3a-Tropoyloxy-6P,7P-epoxytropane (233) N-oxide (248’.?.2.4.5,.h) 143 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane 191 a-Belladonnine (707) 192 P-Belladonnine (70’) H . niger var. pallidus Rchb. 1 3a-Hydroxytropane (2032) 24 3a-Apotropoyloxytropane (203’.’.’) 34 ( -)-3a-Tropoyloxytropane (149, 203’.2.2.4.5.h) 136 3a-Hydroxy-6P,7P-epoxytropane(2031.2.’.4.5.h) 140 ( - )-3a-Tropoyloxy-6P,7~-epoxytropane (149, 203’.2.3.4.5.h) H orientalis 24 3a-Apotropoyloxytropane (2842.7) 34 (-)-3a-Tropoyloxytropane (284?,7) 140 ( - )-3a-Tropoyloxy-6~,7P-epoxytropane (284?,’) 191/ Belladonnine (2842,7) 192 H . pusillus L. 1 3a-Hydroxytropane (185,8275,’ 285,7 3317) 2 3-Acetoxynortropane (16T8) 3 3a-Acetoxytropane ( I @ ) 6 3-Tigloyloxynortropane ( l U K ) 11 3a-Tigloyloxytropane (276?) 24 3a-Apotropoyloxytropane (285,’ 3317) 27 (~)-3a-Tropoyloxynortropane(2O4’.’) 33 (?)-3a-Tropoyloxytropane (204’,’) 34 (-)-3a-Tropoyloxytropane (139,x,y176,).(‘ 185,1.K275,’,?276,?,7285.’ 3317) 35 ( - )-3a-(2’-Hydroxy-3’-phenylpropionyloxy)tropane(276’) 51 3P-Acetoxytropane ( I @ ) 138 3a-Apotropoyloxy-6~,7~-epoxytropane (204,’?,3’! 285,7 331 7 , 140 (-)-3a-Tropoyloxy-6P,7p-epoxytropane (139,K.y.1 176.’,6 204,’,’275,‘,?285,7 331’1

(continued)

60

MAURI LOUNASMAA A N D TARlA TAMMINEN

TABLE 111 (continued) H. reticularus L .

24 27 33 34 138 139 140

3a-Apotropoyloxytropane (204,’.3275’) (+)-3a-Tropoyloxynortropane (2041,3) (+)-3a-Tropoyloxytropane (171 204,’,)286”’.’) ( - )-3a-Tropoyloxytropane (275,’,’287’) 3a-Apotropoyloxy-6/3.7/3-epoxytropane (204l.’) 3a-Tropoyloxy-6/3,7~-epoxynortropane (204’,’) ( - )-3a-Tropoyloxy-6~,7P-epoxytropane (171 204,’,’ 275,‘,?286).Iu) H. senecionis Willd. 24 3a-Apotropoyloxytropane 27 (+)-3a-Tropoyloxynortropane ( 2 0 4 ’ 9 33 (r)-3a-Tropoyloxytropane (204’.)) 138 3a-Apotropoyloxy-6/3,7/3-epoxytropane (204’.)) 139 3a-Tropoyloxy-6/3,7~-epoxynortropane (2041.3) 140 ( - )-3a-Tropoyloxy-6/3,7/3-epoxytropane (204l.)) H. turcomanicus Pojark. 34 (-)-3a-Tropoyloxytropane (185’) 140 ( - )-3a-Tropoyloxy-6/3,7/3-epoxytropane (185’) Hyoscyamus x gyorffi (H. niger X H . albus allopolyploid) 34 (-)-3a-Tropoyloxytropane (139,8.9185’) 140 (-)-3a-Tropoyloxy-6/3,7P-epoxytropane (139,8.9185’) (Note: 185: Hyoscyamus X gyoerfJi) Mandragora M . cnulescens C . B. Clarke 34 ( - )-3a-Tropoyloxytropane (35.1.2.4.6 / 762.7) 87 ( - )-3a-Tropoyloxy-6/3-hydroxytropane( I 762.7) 140 ( - )-3a-Tropoyloxy-6P,7/3-epoxytropane ( I 767) M . chinghaiensis Kuang et A. M. Lu 34 ( - )-3a-Tropoyloxytropane ( 3 5 ’ 9 140 ( - )-3a-Tropoyloxy-6/3,7/3-epoxytropane (352.’0) M . officinarum L . 11 3a-Tigloyloxytropane ( 6 9 ) 24 3a-Apotropoyloxytropane (21,’! 65?) 28 ( - )-3a-Tropoyloxynortropane (21 144) 33 (?)-3a-Tropoyloxytropane (21 17/’,’) 34 (-)-3a-Tropoyloxytropane (21,?65,’288’) 38 3a-Tropoyloxytropane N-oxide 1 (248?.6.7) 39 3a-Tropoyloxytropane N-oxide 2 (248’.“’) 93 3a,7/3-Ditigloyloxytropane(65’) 136 3a-Hydroxy-6p,7/3-epoxytropane (21?’) 140 ( -)-3a-Tropoyloxy-6/3.7/3-epoxytropane (21.? 65,’ 171 289?) 1911 Belladonnine (21,*?65*?) 192 (Notes: 65: M . autumnalis Bertol. and M . uernalis Bertol. analyzed separately; 144; original isolation from M . uernulis by 0 . Hesse ( J . Pr. Chem. 4,282

,’

,’’

,’.’

1.

61

TROPANE ALKALOIDS

TABLE Ill (continued)

(1901)l under name pseudohyoscyamine; 171: M . autumnalis Bertol. ; 288: Mandragora sp., under name mandragorine) Physochlaina P . alaica E. Korot. 1 3a-Hydroxytropane (290?) 24 3a-Apotropoyloxytropane (290,’ 291) 33 (?)-3a-Tropoyloxytropane (290’) 34 (-)-3a-Tropoyloxytropane (2902) 57 3a,7P-Dihydroxytropane (290’) 87 ( - )-3a-Tropoyloxy-6P-hydroxytropane (290’) 88 (?)-3a-Tropoyloxy-6~-hydroxytropane (290,’ 292) 94 3a-Tropoyloxy-6~-hydroxytropane N-oxide (290,’ 293) 101 3a-(4’-Methoxyphenylacetoxy)-7~-hydroxytropane (290,’ 291) 138 3a-Apotropoyloxy-6~,7~-epoxytropane (290,*292) 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (290’) 191 a-Belladonnine (290’) 192 P-Belladonnine (290*) P . dubia 34 (-)-3a-Tropoyloxytropane (294’) 87 ( - )-3a-Tropoyloxy-6P-hydroxytropane(294?) P . infundubularis Kuang 33 (?)-3a-Tropoyloxytropane (295’) 34 (-)-3a-Tropoyloxytropane (35,?1767.2) 87 ( - )-3a-Tropoyloxy-6~-hydroxytropane (35,’I76,I 295’) 138 3a-Apotropoyloxy-6~,7~-epoxytropane (295*) 140 ( - )-3a-Tropoyloxy-6~,7P-epoxytropane (35,*176,’ 295’) P . orientalis (Bieb.) G. Don Fil. 34 ( - )-3a-Tropoyloxytropane (2961.2.4.’) 140 ( - )-3a-Tropoyloxy-6~,7P-epoxytropane (296’.2.4.5) P . physaloides (L.) G. Don 24 3a-Apotropoyloxytropane (35’) 34 ( - )-3a-Tropoyloxytropane (35.l.’I 76,2,7297) 87 ( - )-3a-Tropoyloxy-6~-hydroxytropane (35,’I 76,2.7297, 298’) 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (35,l.’I 767) P . praealta (Decne.) Miers 1 3a-Hydroxytropane (35*) 34 ( - )-3a-Tropoyloxytropane (35’,2.4.5.6) 87 ( - )-3a-Tropoyloxy-6~-hydroxytropane (351,4.5,6) 140 ( - )-3a-Tropoyloxy-6~,7P-epoxytropane (351.2.4.5.6) Przewalskia P . shebbearei 34 ( -)-3a-Tropoyloxytropane (I 762.4.6.7) 87 ( - )-3a-Tropoyloxy-6~-hydroxytropane (1762.4.6,7) (continued)

62

MAURl LOUNASMAA A N D TARJA TAMMINEN

TABLE 111 (continued) 140 ( - )-3a-Tropoyloxy-6P,7~-epoxytropane ( I 762.4.6.7) 142 ( - )-3a-(2’-Hydroxytropoyloxy)-6~,7~-epoxytropane ( I 76’.4.6) P . tnngutica Maxim. 1 3a-Hydroxytropane (35,’.2.3.4.5.6 44,’.2.4.s.6 299’) 24 3a-Apotropoyloxytropane (35,’.2.4.5.6 44,’.2.4.5 299*) 34 ( - )-3a-Tropoyloxytropane (35,’,2.3.4.5.6 44,1.2.3.5 176,’ 299’) 87 ( - )-3a-Tropoyloxy-6P-hydroxytropane (35,’.2.4.5.6 44 ,1.*.4.5.6 176*) 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (35,1.2.3,4.5.6 44.’,2.4,5 176,’ 299’) 142 ( - )-3a-(2’-Hydroxytropoyloxy)-6~,7~-epoxytropane (35,*44,’ 176) Scopolia S . acuiangula C . Y. Wu et C . Chun 1 3a-Hydroxytropane (353) 34 ( - )-3a-Tropoyloxytropane (35,1.3.4.5.6 76,’.’ 3009) 87 (-)-3a-Tropoyloxy-6P-hydroxytropane (35,1.3.4,5.6 176l) 140 ( - )-3a-Tropoyloxy-6~,7~-epoxytropane (35,1.3.4.5.6 176,’ 300’) 142 ( - )-3a-(2’-Hydroxytropoyloxy)-6~,7~-epoxytropane (35.’.3.4.5.h 1762,7)

S . unomala 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (301 I.’) S . carniolicu 1 3a-Hydroxytropane (302,’ 11 3a-Tigloyloxytropane (303’) 33 (?)-3a-Tropoyloxytropane (233,2244, 302,’ 304,* 305.’.?.4 306) 34 (-)-3a-Tropoyloxytropane (149,’ 233,’ 244, 303,’ 304,’ 306,’ 307’) 38 3a-Tropoyloxytropane N-oxide 1 (2481.2.4) 39 30-Tropoyloxytropane N-oxide 2 (2481,2.4) 50 3a-Hydroxytropane (303*) 136 3a-Hydroxy-6P,7P-epoxytropane (302’) 140 ( -)-3a-Tropoyloxy-6P,7P-epoxytropane(14S,’ 149,x244, 302,” 303,’ 304,’ 305,‘.2.4 306,23072) 141 (It)-3a-Tropoyloxy-6P,7P-epoxytropane (233’) 143 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane N-oxide (248’.2.4) (Note: 145, 233, 244, 304: Scopoliu atropoides) S . himalaiensis 1 3a-Hydroxytropane (308’,*) 34 ( - )-3a-Tropoyloxytropane (308,l.’ 3098) (308?‘.2) 136 3a-Hydroxy-6P,7P-epoxytropane 140 ( - )-3a-Tropoyloxy-6~,7P-epoxytropane (308, 3098) (Note: 308: Himalayan Scopolia) S . japonica Maxim. 1 3a-Hydroxytropane (.?lo2) 24 3a-Apotropoyloxytropane (310’) 28 ( - )-3a-Tropoyloxynortropane (144*) 33 (?)-3a-Tropoyloxytropane (3112, 34 (-)-3a-Tropoyloxytropane (144,’ 310,’ 311,* 312x) 140 (-)-3a-Tropoyloxy-6~,7~-epoxytropane (310,* 311 3/2,x3139) S . lurida (Link et Otto) Dunal 1 3a-Hydroxytropane (35,134,5170,2 185’) 24 3a-Apotropoyloxytropane (351,4.5)

,’

63

1. TROPANE ALKALOIDS TABLE 111 (continued) ~

~~

33 34 38 39 50 87

136 140

(?)-3a-Tropoyloxytropane (170,'.*.'.4314,7 315,'.2.3.4316,) 3178) (-)-3a-Tropoyloxytropane (35,'.4.5149,' 176,'.5 185,' 244,?.3,4 314,7 318*) 3a-Tropoyloxytropane N-oxide 1 (2481.2.4) 3a-Tropoyloxytropane N-oxide 2 (248'.2.4) 3P-Hydroxytropane (185') ( - )-3a-Tropoyloxy-6~-hydroxytropane ( I 76'.5) 3a-Hydroxy-6P,7P-epoxytropane(170*) ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (35,'.4.51 70,'.?.3.4 176,1,5 185,8301,I,* 314, 315.',2,'.4316,' 317,' 318*)

143 ( - )-3a-Tropoyloxy-6P,7P-epoxytropaneN-oxide (248'.'.4) (Notes: 149, 316: S . strarnonifolia Shestra; 185: S . strarnonifolia Sernenova; 244, 317: Anisodus luridus Link et Otto; 318: S . strarnonifolia) S . parviflora 1 3a-Hydroxytropane (3102) 24 3a-Apotropoyloxytropane (310*) 34 ( - )-3a-Tropoyloxytropane (310*) 140 ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (310*) S . sinensis Hemsl. 33 (+)-3a-Tropoyloxytropane (31Y,'.2.43201,*,4) 34 ( - )-3a-Tropoyloxytropane (35,?.3+6176*.') 140 ( - )-3a-Tropoyloxy-6~,7~-epoxytropane (35,'.3+h31Y,'.2.43201.'.4) 142 ( - )-3a-(2'-Hydroxytropoyloxy)-6~,7~-epoxytropane ( I 76'.') [Note: 35, 176: Atropanthe sinensis (Hemsl.) Pascher] S . tangutica Maxim 1 3a-Hydroxytropane (35,' 3212) 24 3a-Apotropoyloxytropane (35',?) 34 ( - )-3a-Tropoyloxytropane (35,'.*.' 176,*.5.7214,R307,1.* 321 322,7323,'324,7

,'

3257)

87 ( -)-3a-Tropoyloxy-6~-hydroxytropane (35,' I 76,'.5.7214,R3257) 89 3a-Tropoyloxy-7P-hydroxytropane(214') 140 ( -)-3a-Tropoyloxy-6~,7P-epoxytropane (35,1,?,3.4 176,*.5.7214,R307,' 321

,'

322,' 323,* 324,7 3257)

142 S . sinensis 34 140 Convovulaceae Calystegia C. sepium 179 181 182

(-)-3a-(2'-Hydroxytropoyloxy)-6~,7~-epoxytropane (35,1,*,',4176.* 3257) x tangutica (-)-3a-Tropoyloxytropane (3267) ( - )-3a-Tropoyloxy-6P,7P-epoxytropane (3267)

lp,2a,3P-Trihydroxynortropane(271, 272, 273') lp,2a,3P,6P-Tetrahydroxynortropane(271, 272, 273x) Ip,2a,3P,4a-Tetrahydroxynortropane(271, 272, 2738)

(continued )

64

MAURI LOUNASMAA A N D T A W A TAMMINEN

TABLE 111 (continued) Colutea C. orientalis

101 3a-(4’-Methoxyphenylacetoxy)-7~-hydroxytropane (327’) Conuoluulus C. aruensis

179 Ip,2a,3P-Trihydroxynortropane(271, 2728) 181 Ip,2a,3/3,6P-Tetrahydroxynortropane(271, 272n) 182 Ip,2a,3P,4a-Tetrahydroxynortropane(271, 272*) C. krauseanus Regel et Schmahl. 32 3a-Vanilloyloxynortropane (328,?*3297) 36 3a-Vanilloyloxytropane (3297) 37 3a-Veratroyloxynortropane (328,* 3297) 40 3a-Veratroyloxytropane (328,? 329’) 41 3a-Veratroyloxy-N-hydroxynortropane (3307*) 44 3a-Veratroyloxytropane N-oxide (.?.?I7*) 47 ( ~)-3a-Veratroyl-N-acetylnortropane (3297*) C. linearus 37 3a-Veratroyloxynortropane (3327) 40 3a-Veratroyloxytropane (3327) C. pseudocanrabricus Schrenck 37 3a-Veratroyloxynortropane (333,3*3 W 4 ) 40 3a-Veratroyloxytropane (334,‘ 3.V3*) 199 Convolvidine (334’*) C. subhirsurum Regel et Schmahl. 32 3a-Vanilloyloxynortropane (336,?3372) 36 3a-Vanilloyloxytropane (336,l 337?) 339,? 34@) 37 3a-Veratroyloxynortropane (336,*337,2 338,2,3.7 40 3a-Veratroyloxytropane (337,2338,2.3.73402) 41 3a-Veratroyloxy-N-hydroxynortropane(337’) (336,**337?) 43 3a-Veratroyloxy-N-formylnortropane 46 ( +)-3a-Veratroyloxy-N-isopropyhortropane(337?*) 198 Subhirsine (337,? 3 4 / * * ) 199 Convolvidine (338,3,73402) Erycibe E . elliprilimba

177 2P,6P-Dihydroxynortropane (342‘) 178 2p,7P-Dihydroxynortropane(342‘*) 183 ( - )-2P-Hydroxy-6~-acetoxynortropane(3424)

E . hainanensis 177 2/3,6P-Dihydroxynortropane (34j4) 178 2P,7P-Dihydroxynortropane (343‘) 183 ( - )-2P-Hydroxy-6P-acetoxynortropane (343‘) E. obtusifolia Benth 177 2P,6P-Dihydroxynortropane (344) 183 ( - )-2P-Hydroxy-6P-acetoxynortropane (66)

1.

TROPANE ALKALOIDS

65

TABLE 111 (continued) Euolvulus E. sericeus var. holosericeus 37 3a-Veratroyloxynortropane (345 lo) 40 3a-Veratroyloxytropane (345”) 199 Convolvidine (345’”) Key to superscripts: leaves, petioles, terminal branches root, rhizome, or root bark seeds stem or stem bark flowers, calyx, or buds fruit or pericarp aerial parts * root culture callus culture lowhole plant I’ seedlings I 2 sap ’ tentative structure * new alkaloid With one exception, all compounds whose isolation has been reported are included in the list even if the isolation was tentative. The exception is 2,6-dihydroxytropane, whose isolation from Dufura sframonium remains questionable (346). The reported isolation of tropane alkaloids from Peripenfadenia mearsii (Euphorbiaceae) has turned out to be erroneous. and we omit mention of this species (347) (see text). The parasitic species of the genera Orobanche (348) and Cusrrrfu (349) are not included either, since the reported alkaloids most probably were produced by the host plant. Names derived from 3.7-dihydroxytropane (instead of 3,6-) have been used in cases where the absolute stereochemistry is not known. Because different analytical methods were used, there is considerable variation in the degree of stereochemistry indicated in the papers. To avoid listing the same compound several times (e.g., 3-tropoyloxytropane and 3a-tropoyloxytropane), some assumptions had to be made. Thus, all compounds with the 6- and 7-substituents are described as p-derivatives. The stereochemistry of the 3substituents is more difficult to determine because several acids esterify both 3a- and 3p-hydroxytropane (acetic, tigloic, and benzoic acids). In these cases, the stereochemistry has not been indicated, although in the list of new compounds they are mentioned under the heading 3a-derivatives. ‘ The unspecified “methylbutyryl” group reported in some articles has been treated as an isovaleryl group, because ( I ) the 2-methylbutyryl derivatives had been identified separately and (2) n-valeric acid has never been found as an esterifying acid. a

’

’

2 . Chemotaxonomy of Plants Containing Tropane Alkaloids It is generally accepted that chemical evidence cannot be used as the sole criterion for taxonomic revisions. At the very least, the whole chemical spectrum of the plant must be considered, keeping in mind the wide natural variation in chemical composition. “Chemical races,” often geographically distinct, have been reported for Duboisia myoporoides (42,43) and Przewalskia tangutica, for example (44).

TABLE IV LIST OF TROPANE ALKALOIDS OF PLANTORIGIN

MW

Formula

139 CsH1,NO

Compound 175 3-Oxotropane

Plant family"

Refs

S

1,5,84,175,185,260,263,265,277

S

1,2,5,19,30,35,42,44,50,52,58-62,64,68,70,84.105, 109,l19,121,123.124,129,140,141,146,153,155157,158aJ62 163,166,167,170,185,191,I 94,197201,203,219,220,230,254,258,259,264,265,275277,279,285,290,299,302,303,308,310,321

( = tropinone)

141 C ~ H I S N O

1 3a-Hydroxytropane ( = tropine)

~

141 CBHISNO

50 3P-Hydroxytropane ( = pseudotropine)

176 1-Hydroxytropane (hydrochloride)

S, Er

1,2,5,61,62,64,89,105,110,117,121,156,157,162,

S

l63,166,185,194,197-199,20l,203,219,220,230, 258-260,265,276,277 5,158a,159-161

( = physoperuvine)

177 178 136 57 179

2P,6P-Dihydroxynortropane ( = baogongteng C) 2@,7@-Dihydroxynortropane ( = erycibelline) 3a-Hydroxy-6~,7P-epoxytropane ( = scopine) 3a,7P-Dihydroxytropane 1@,2a,3P-Trihydroxynortropane ( = calystegine

180 2 120 181

( f )-3,4-Dehydro-4-acetyltropane ( = ferruginine)

co S S, Er co, s

342,344 342,344 1,21,124,170,203,279,302 I ,18,58-60,121,197,201,260,290 271, 272

P S S co,

s

5,74.75 185 200 271,272

co,

s

271,272

co

'43)

3a-Acetoxynortropane 3a,6P,7P-Trihydroxytropane lf3,2a,3P,6P-Tetrahydroxynortropane ( = calystegine B I) 182 lp,2a,3P,4a-Tetrahydroxynortropane ( = calystegine Bz) 145 2,3-Dehydro-2-methoxycarbonyltropane ( = anhydroecgonine methyl ester or me thylecgonidine)

Er

1,90,99,1I1

3 3a-Acetoxytropane 51 3P-Acetoxytropane 146 ( - )-2a-Carboxy-3P-hydroxytropane

S Er

5,51,52,64,83,I I9,14O,185,186.I9lI194,199,202, 230,259,260 158a,185,199,260 1,2,91,110

co

66

Er P P

86 41,79 41.79

R Eu, P, S Er

5,83,199 5,72,185,259,260 1,89,92,109,110,111

S S P P P

185 260 5,41,72-74 5,41,72,74 41,72

P

41,72

P

41,72

S S, R S, R S S

185 5,51,83,115,119 1,4,5,42,51,83,119,129-134,140,199 1,16,109,140,142 1,109

S, R

( = ecgonine)

183 ( - )-2P-Hydroxy-6P-acetoxynortropane ( = baogongteng A)

58 3a-Acetoxy-7P-hydroxynortropane 163 Pyranotropane ( = strobiline) 169 10,I I-Dihydropyranotropane ( = dihydrostrobiline)

4 3a-Propion ylox ytropane 59 ( + )-3a-Acetoxy-7P-hydroxytropane 147 ( - )-2P-Methoxycarbonyl-3~-hydroxytropane ( = ecgonine methyl ester)

60 5 164 165 167

170 171 6 7 8 9 10

3a-H ydroxy-7P-acetoxytropane

3a-(H ydroxyacetoxy)tropane 10-Methylpyranotropane ( = bellendine) 2,1 I-Methylpyranotropane ( = isobellendine) 1 I-Methyl-3,4-dihydropyranotropane( = 5,11dihydroisobellendine) 10-Methyl-10,l I-dihydropyranotropane ( = dihydrobellendine) 10-Methyl- 10,l I-dihydropyranotropane ( = epidihydrobellendine) 3a-Tigloy lox y nortropane 3a-But yryloxytropane 3a-Isobutyryloxytropane ( = butropine) 3a-Isovaleryloxynortropane( = poroidine) 3a-(2'-Methylbutyryloxy)nortropane ( = isoporoidine)

(continued)

TABLE IV (continued)

MW

Formula

Compound

Plant family'

Refs.

213 CllHI9NO3 61 3a-Hydroxy-7~-propionyloxytropane 219 CI3Hl7NO? 166 10,l I-Dimethylpyranotropane ( = darlingine) 221 CI3HI9NO2 168 IO,ll-Dimethyl-3,4-dihydropyranotropane ( = 5.1 1-dihydrodarlingine) 221 C13H19N02 172 10.1 I-Dimethyl-10.1 I-dihydropyranotropane ( = dihydrodarlingine) 223 C13H2,NOz 11 3a-Tigloyloxytropane

S P P

259 5,41,72,74-76 41.74

P

41,72

S

223 CI3H2,N02

S S S. R

4,5,20,30.32,51,61,62,64,65,119,I21,124,140, 156-158,158a,163,185,187,l97,209~218,219,230, 254,276,303 5,16,32,50,6l,62,64,119,121,155-157,l99,218,219, 230,243,259.260,276 57-60 1,4,5,51,64,ll9,129-131.133,134,140,141,143,199

Er, R Er S S S S P Er Er Er S S, Er S S S

5,83,109 55 55,189 I99 51,119 259 5,75,76 lI3,l14,117 86 I09 5,157 3,459-60,111 , I 1 9,121,153,l84,185,199,2S9,260 5840,153 4,5,119,184,185,191,199,259,260 58-60,153

52 3P-Tigloyloxytropane ( = tigloidine)

12 3a-Senecioyloxytropane 13 ( + )-3a-(2'-Methylbutyryloxy)tropane ( = valtropine) 14 3cr-Isovaleryloxytropane 62 ( + )-3a-Hydroxy-7~-tigloyloxynortropane 63 3a-Tigloyloxy-7P-h ydrox ynortropane 53 3P-(2'-Methylbutyryloxy)tropane 64 3cu-Isobutyryloxy-7P-hydroxytropane 65 3a-H ydroxy-7P-isobut yrylox ytropane 184 ( + )-4a-Benzoyltropane ( = ferrugine) 15 3a-Benzoylox ynortropane 54 3P-Benzoylox ynortropane ( = nortropacocaine) 16 3a-(2'-Furoyloxy)tropane 17 3a-Tigloyloxytropane N-oxide 66 3a-Hydrox y-7P-tigloylox ytropane 67 3a-Hydroxy-7P-angeloyloxytropane 68 3a-Tigloyloxy-7P-h ydrox ytropane 69 3a-Senecioylox y-7P-hydrox ytropane

70

(

+ )-3a-Hydroxy-7&(2'-

S

5,117,199,260

S

1,3,16,51,l09,1l9,136,140,141

methylbutyry1oxy)tropane

71 3a-Isovaleryloxy-6P-hydrox ytropane ( = valeroidine)

72 18 55 19 73 121 122

3a.7P-Diacetoxytropane 3a-Benzoylox ytropane 3P-Benzoyloxytropane ( = tropacocaine)

3a-Phenylacetoxynortropane 3a-Benzoylox y-7P-hydroxynortropane

3a,7P-Dihydroxy-6P-tigloyloxytropane 3a-TigIoyloxy-6~,7~-dihydroxytropane ( = meteloidine)

20 3a-Apotropoyloxynortropane

S

185

Er, R

5,32,83,84,86,114,117 l,5,86,89,97,110,11l,Il7 115 111,114,117 5 ,I 79,182,191,201 1,4,S,30,50,55,56,61,62,119-l21,l44,167,168.187, l9l,193,194,197-200,205,208,209,21l,212,216, 218,219,230,238,239,241,243,254,258 51,52,119

Er E Er S S

S

( = aponoratropine)

21 3a-Cinnamoyloxynortropane 24 3a-Apotropoyloxytropane ( = apoatropine)

El S

22 3a-Phenylacetox ytropane Er 186 6,7-Dehydro-3a-(4'-hydroxybenzoyloxy)tropane P [ = 3a-( p-hydroxybenzoyloxy)trop-6-ene] 23 3a-(3'-Hydroxybenzoyloxy)tropane Cr

85,114,117 1,4,5,21,32,35,44,50-52,65,70,82,119,121~ 123,132, l99,201,203,204,2l9,221,236,243,244,258-260, 264-268,275,276,284,285,290,291,299,310,331 l09,113,ll5,260,265 71 69.70

( = cochlearine)

74 76 75 77 78 173

3a-H ydroxy-7P-benzoylox ytropane

3a-Phenylacetoxy-7~-hydroxynortropane 3a-Benzoylox y-7P-hydrox ytropane

3a-Acetoxy-7~-isobutyryloxytropane 3a-Isobut yrylox y-7P-acetoxytropane

I I-Phenyl-10, I I-dihydropyranotropane ( = strobamine)

P, Er Er Er P P P

79,86,l04,11I 86 113,I 14 86,I 13,I I7 5,72 5,72 41,80 I

TABLE IV (continued) MW

Formula

269 27 1 27 1 27 1 273 273 275

Compound

Plant family”

Refs.

187 ( + )-3,4-Dehydro-4-cinnamoyl-3hydroxytropane ( = chalcostrobamine) 25 3a-Cinnamo ylox ytropane 56 3P-Cinnamoy lox ytropane 137 3a-Apotropoyloxy-6~-7~-epoxynortropane ( = aponorscopolamine or aponorhyoscine) 26 ( - )-3a-(1’,2’-Dithiolane-3’carbony1oxy)tropane ( = brugine) 79 3a-Cinnamoyloxy-7~-hydroxynortropane 27 (t)-3a-Tropoyloxynortropane ( = noratropine)

P

S

I S ,18,3O,5O,64,121,124,132,14O,168,183,191,I 94,

28 ( - )-3a-Tropoyloxynortropane ( = norhyoscyamine) 29 3a-(4’-Methoxybenzoyloxy)tropane ( = datumetine) (=alkaloid KD-B) 151 2a-Benzyl-3a-acetoxytropane 30 3a-(3’-Hydroxyphenylacetoxy)tropane

S

1,4,18,21,42,50-52,64,119,123,129,132,144,146,

S

167,192,197,219,243,261,262,265 222

R Er S

84,114 114 51,119

R

4J,81-84,84a

Er

114

204.219,230,243,258.261,262

0“

275

16H21N03

275

C16H21N03

275 275 275 275 275 217 277 277 277 28 1 285

80 81 31 32

3a-Phen ylacetoxy-7P-h ydrox ytropane 3a-Hydroxy-7P-phenylacetox ytropane 3a-(4’-Hydroxyphenylacetoxy)tropane 3a-Vanilloy loxynortropane ( = convolidine) 123 3a-Benzoyloxy-6~,7~-dihydroxytropane

l24 126 82 138

3a,6~-Dihydroxy-7~-benzoyloxytropane 3a-Phenylacetoxy-6~,7~-dihydroxynortropane 3a-Tigloyloxy-7~-acetoxytropane 3a-Apotropoyloxy-6~,7~-epoxytropane ( = aposcopolamine or apohyoscine)

P

Er Er Er Er

co Er Er S

s, P S

5,41,77,78 113,115 104 1I S , I I5 260 114 111 328,329,336,337 55,111 104 124 ~

~

5,71,119,185,194,199,2S9,260 I ,4,30,42,50,51,70,119,123,124,132,l40,168,188, I9l,194,197,199,201,2O4,209,2l9,230,258-260,

265,268,276,285,290,292,295,331

285 C17H19N03

174 7P-Hydroxy-1 l-phenyl-l0,11dihydropyranotropane ( = strobolamine) 287 C17H21N03 83 3a-Cinnamoyloxy-7P-hydroxytropane 287 CI7HZINO384 3a-Apotropoyloxy-7~-hydroxytropane 289 CI7H23NO3 33 (?)-3a-Tropoloxytropane ( =atropine)

P

41.80

P, Er S S

289 CI7HZ3NO3 34

S

41,79,111,114 259,260,265 1,5,18,21,30,32,50,62,64,119-121,124,129,135, 136,141,143,155,164,165,167,169-172,173,175, 188,191,194,198,204-207,219,220,223,224,230, 232,233,235,237,243-246,26 1,262,264,268,Z76, 286,290,295,302,304-306,311,3l4-317, 319,320 1,3,4,18,19,21,30,32,35,42-44,46-52,61,64,65,70, 76,119,120,122,123,126-129,131,136-141,l44, 146-1 49,165-1 68,I 74-1 78,183,185,188,193195,197-201,203,20~-214,2 18,220,223,225-228, 231-234,238-241,243,244,246,247,251-255,258, 260-262,265-267,269,270,275-278,280-285, 287,288,290,294,296,297,299,300,303,304,306312,314,318,321-326,331 4,50,61,64,119,120,121,l48,187,191,194,2l9,260, 276,277 1,4,30,50,51,64,l19,121,140,167,168,180,183,188, 191, I 94, 197,200,204,209,219,229,230,258,276

( -)-3a-Tropoyloxytropane ( = hyoscyamine)

289 C17H23N03 35 (-)-3a-(2’-Hydroxy-3’-

S

289 C,,H19N04 139

S

289 CI6H 19N04 148

289 C17H23N03152 289 CI7Hz3NO3156

291 CI,H2,NO4 36 291 C16H21N04 37

phenylpropiony1oxy)tropane ( = littorine) 3a-Tropoyloxy-6~,7~-epoxynortropane ( = norscopolamine or norhyoscine) ( - )-2P-Carboxy-3P-benzoyloxytropane ( = benzoylecgonine) 3a-Acetoxy-4a-hydroxybenzyltropane ( = knightinol) 3a-Acetoxy-4a-benzyI-6P-hydroxytropane ( = knightoline) 3a-Vanilloyloxytropane ( = phyllalbine) 3a-Veratroyloxynortropane ( = convolvine)

Er

1,97

P

41,79

P

41,79

Eu, Co

3,68,329,336,337 1,328,329,333,334,336-340,345

co

~

(continued)

TABLE IV (continued) MW

Formula

Compound 127 3a-Phenylacetoxy-6p,7p-dihydroxytropane

85 3a-Tigloyloxy-7~-propionyloxytropane 140 ( - )-3a-Tropoyloxy-6p,7p-epoxytropane ( = scopolamine or hyoscine)

141 (~)-3a-Tropoyloxy-6P,7p-epoxytropane ( = atroscine) 149 ( - )-2P-Methoxycarbonyl-3pbenzoyloxytropane ( =cocaine) tropane 125 3a-Cinnamoyloxy-6p,7p-dihydroxy 86 3a-Benzoylox y-7P-ace tox ytropane 38 3a-Tropoyloxytropane N-oxide 1 ( = hyoscyamine N-oxide 1) 39 3a-Tropoyloxytropane N-oxide 2 ( = hyoscyamine N-oxide 2) 40 3a-Veratroyloxytropane ( = convolamine) 87 ( - )-3a-Tropoyloxy-6p-hydroxytropane [ = ( - )anisodamine] 88 (~)-3a-Tropoyloxy-6~-hydroxytropane ( = 6p-

hydroxyatropine) 89 3a-Tropoyloxy-7P-hydroxytropane 161 3/.3-Hydroxy-4a-hydroxybenzyl-6pacetoxytropane ( = knightalbinol)

Plant family" Er S 0. s

S

Refs. 113,115 5,67,199,212,260 1,4,16,18,19,21,30,32,35,42-44,46-52,61,62,64,65, 70,118,119,121,123-129,131,135-141,143-147, 149,150,167-169,171-178,180-183,185,191,193195,197-201,203-214,216-221,223-228,230-238, 240,241,243-246,250-255,257,258,260,264,265, 267-270,275-278,281,282,284-286,289,290,295, 296,299-314,318-326,331 1,62,233

Er Er Er S

111,114 113 5,248,265

S

5,248,265

co

S

1,328,329,332,334,335,337,338,340,345 5,44,46,50,51,119,124,146,150,176,185,2OO,209, 214,240,256,259,260,265,277,290,294,295,297, 298,325 290,292

S P

41,80

S

146,214,260,265,277

307

41 3a-Veratroyloxy-N-hydrox ynortropane ( = convoline)

co

330.337

309 317 317 317 319

90 3a-Tigloylox y-7p-isobutyryloxytropane

S R Er Er co

199,260 84 113,115 113 336,337

S

5,35,44,173,176,190,325

S

5.248

S S

3,18,62,65,119,167,183,185,188,194,197-201,209, 212,215,219,220,230,249,254,260 5,290,293

S

179

co

331

Er S S Er

104 5,191,199,260 199,260 I ,23,89,94-97,100,108

Er P

114 41,79

co

33 7

42 91 92 43

319 319 32 1 321 4

321

32 1 321 323 323 329 329 331 333

18H?7N04

C17H23N0S

3a-Feruloylox ytropane

3a-Phenylacetoxy-7~-acetoxytropane 3a-Acetoxy-7~-phenylacetoxytropane 3a-Veratroyloxy-N-formylnortropane ( = confoline) 142 ( - )-3a-(2’-Hydroxytropoyloxy)-6p,7Pepoxytropane [ = ( - )-anisodine or daturamine] 143 3a-Tropoyloxy-6~,7P-epoxytropane N-oxide ( = scopolamine N-oxide or hyoscine N-oxide) 93 3a,7/3-Ditigloyloxytropane N-oxide 94 3a-Tropoyloxy-6p-hydroxytropane ( = 6p-hydroxyhyoscyamine N-oxide) 128 3a-(2‘-Hydroxy-3’-phenylpropionyloxy)-6p,7/3dihydroxytropane ( = 6p,7/3dihydroxylittorine) 44 3a-Veratroyloxytropane N-oxide ( = convolamine N-oxide) 45 3a-(3‘,4’,5’-Trimethoxybenzoyloxy)nortropane 95 3a-Tigloyloxy-7~-(2‘-methylbutyryloxy)tropane

96 3a-TigIoyloxy-7~-isovaleryloxytropane 150 ( - )-2P-Methoxycarbonyl-3Pcinnamoyloxytropane ( = cinnamoylcocaine) 97 3a-Cinnamoyloxy-7~-acetoxytropane 153 3a-Acetoxy-4a-acetoxybenzyltropane ( = acetylknightinol)

46

(2)-3a-Veratroyloxy-N-isopropylnortropane ( = convosine)

(continued )

TABLE IV (continued) MW

Formula

Compound

Plant family“

Refs.

47 (~)-3a-Veratroyl-N-acetylnortropane ( = convolicine) 48 3a-(3‘.4‘,5’-Trimethoxybenzoyloxy)tropane 154 3a-Benzoyloxy-4a-benzyltropane ( = alkaloid KD-A) 130 3a,7P-Ditigloyloxy-6P-hydroxytropane

co

329

Er Er

5,105,106 5,41.77,78

S

5,18,30,62,119,121,167,183,185,188,191,194,197-

98 3a-(3’,4’,5’-Trimethoxybenzoyloxy)-7Phydroxynortropane 129 3a-Tigloyloxy-6P-hydroxy-7Pisovaleryloxytropane 99 3a-Tropoyloxy-7~-acetoxytropane 155 3a-Benzo y lox y -4a-h ydrox ybenzy ltropane 158 3a-Hydroxy-4a-benzyI-7p-benzoyloxytropane (=alkaloid KD-C) 100 ( + )-3a-(3’,4’,5’-Trimethoxybenzoyloxy)-7Phydrox ytropane 101 3a-(4’-Methoxyphenylacetoxy)-7Phydroxytropane ( = physochlaine) 102 3a-(Pyrrolyl-2’-carbonyloxy)-7P-(N”meth ylpyrrol yl-2-carbony1oxy)tropane ( = catuabine C) 49 3a-(3’,4‘,5‘-Trimethoxycinnamoyloxy)tropane 107 3a,7P-Dibenzoyloxytropane 103 3a-(1 ’-Methylmesaconyloxy)-7ptigloyloxytropane ( = schizanthine F) 104 3a-(1 ‘-Methylitaconyloxy)-7~-tigloyloxytropane ( = schizanthine G)

Er

199,209,212,215,218-220,230,242,254,260 104

S

187,260

S P P

119 41,75 5,41,77,78

Er

104

S

5,290,291,327

Er

102,103

Er Er S

4,5,105,106,112.113 111 153

S

153

106 3 4 1’-Methylitaconyloxy)-7pangeloyloxytropane ( = schizanthine H) 105 3a-( I ‘-Methylmesaconyloxy)-7Pangeloyloxytropane ( = schizanthine I)

S

I53

S

153

131 3a-(3’,4’,5’-Trimethoxybenzoyloxy)-6~,7Pdihydrox ytropane 160 3a-Hydroxy-4a-hydroxybenzyl-7Pbenzoyloxytropane ( =alkaloid KD-F) 162 3a-Benzoyloxy-4a-hydroxybenzyl-6Phydroxytropane ( = knightolamine) 157 3a-Cinnamoyloxy-4a-benzyl-6Phydroxytropane ( =alkaloid KD-D) 108 3a-(3’-EthoxycarbonyImethacryloyloxy)-7~senecioyloxytropane ( = schizanthine A) 110 3a-( 1 ‘-Ethylmesaconyloxy)-7Ptigloyloxytropane ( = schizanthine K) 109 3 4 1 ’-Ethylitaconyloxy)-7P-angeloyloxytropane ( = schizanthine L) 111 3 4 1 ‘-Ethylitaconyloxy)-7~-tigloyloxytropane ( = schizanthine M) l l 2 3a-Tropoyloxy-7P-tigloy loxytropane 132 ( + )-3a-(2’-Hydroxy-3’-phenylpropionyloxy)6~-hydroxy-7P-tigloyloxynortropane 114 3a-Tropoyloxy-7~-isovaleryloxytropane 115 3a-Tropoyloxy-7/3-(2‘methylbutyry1oxy)tropane 116 3a,7P-Dicinnamoyloxytropane 117 3a-(3‘,4’,5’-Trimethoxybenzoyloxy(-7~benzoyloxytropane (=catuabine B) 133 3a-Acetoxy-6P.7P-dibenzoyloxytropane

Er

5,104,I06

P

5 , 4 l ,78,79a

P

41,80

P

5,4l,77,78

S

154

S

153

S

I53

S

153

S

Er

199 55

S S

199 199

Er Er

114 102,103

Er

86 (continued)

TABLE IV (continued)

MW

Formula

539 C2jHj3NO3

Compound

135 191 192 193 194 195 1% 197 198 199 200

@

3a-(3',4'.5'-Trimethoxybenzoyloxy(-7~-(N"meth ylpyrrolyl-2"-carbony loxy )tropane ( = catuabine A) Schizanthine C 3a-(3',4',5'-Trimethoxycinnamoyloxy)-7~benzoyloxytropane Schizanthine D Schizanthine E 3a-Cinnamoyloxy-4a-hydroxybenzyl-7~benzoyloxytropane (=alkaloid KD-E) 3a-(3',4',5'-Trimethoxycinnamoyloxy)-6~hydroxy-7p-benzoyloxytropane 3a-(3',4',5'-Trimethoxycinnarnoyloxy)-6~acetoxy-7P- benzoyloxytropane a-Belladonnine P-Belladonnine dcopadonnine P-Scopadonnine Schizanthine B Schizanthine X 7p-Acetoxytropan-3-yI-tropan-3'-yl-truxillate Subhirsine Convolvidine 7P-Acetoxytropan-3-yI 7'P-hydroxytropan-3'-yltruxillate a-Truxilline P-Truxilline Grahamine

Plant family"

Refs.

Er

102,103

S Er

57 5,104,106,107

S S P

57 57 5,41,79a

Er

104

Er

104

S S S S

2 I ,65,70,266,267,269,274,284,2 90

s

S Er co

co Er Er Er

S

21,65,70,266,267,269,274,284,290 132,396 132,396 154 151 114 337,341 338,340,345 114

1,98.101 1,98,101 152

Key to families: Br. Brassicaceae ( = Cruciferae): Co. Convolvulaceae: Er, Erythroxylaceae: Eu. Euphorbiaceae: 0 , Olacaceae: P, Proteaceae; R, Rhizophoraceae: S . Solanaceae. . .- -

1.

TROPANE ALKALOIDS

77

Tropane alkaloids are relatively simple molecules, and it is entirely possible that the ability of plants to synthesize them has developed separately in unrelated families. Therefore, the isolation of the same alkaloid in two species may be considered as a taxonomic marker only if the alkaloid can be shown to be produced through the same biosynthetic pathway in both species. The biosynthesis of tropane alkaloids has received much attention and is rather well understood (vide infra). Variation in the spectrum of tropane alkaloids in separate plant organs as a function of growing period has been found to throw light on the site of alkaloid biosynthesis and to be informative of the accumulation of some alkaloids and the degradation of others (45-49). Despite the need for caution noted above, there are cases where chemotaxonomy based on the tropane alkaloids has been applied successfully. Romeike (13)has reviewed the occurrence of tropane alkaloids and considered their significance in plant classification. Other examples are the latest revision of the Solanaceae (26) and the new concept of the subfamily Anthocercideae (50-52). Hegnauer (53) and Evans (54) have presented a chemotaxonomic approach to the genus Erythroxylum. Links have been Erythroxylaceae and Solanaceae (55,56). At the generic level, chemotaxonomy has been used to establish the relationships among Schizanthus (57-60), Datura (61-63), Solandra (64), Mandragora ( 6 3 , and Atropa (32). In most of the studies cited, attention has been paid not only to the existence of single alkaloids, but also to the types of esterifying acids.

3. Optical Activity of Tropane Alkaloids Further information is implicit in the optical properties of the isolated alkaloids. The absolute stereochemistry is known in only a few cases, however. The optical properties of the isolated compounds are not fully reviewed here, as they are usually not mentioned in the original papers. Racemization during the isolation process causes further problems where the origin of the optical activity is tropic acid (e.g., as in hyoscyamine/ atropine). The distribution of 3a,7P-dihydroxytropane (and/or 3a,6P-dihydroxytropane) may serve as an example of the potentially informative value of absolute stereochemistry. Both optical forms have been found, either as such or esterified: the (+) form is typically found in herbaceous Datura species and the (-) form in the tree Datura, Duboisia, Anthocercis, and Erythroxylum ( f 6 , 5 f,67). In addition, optically inactive esters of 3a,7@-dihydroxytropane have been found in the Proteaceae (41,80). Because only the 3a-tropoyl ester derivatives of this alkamine (60- and

78

MAURI LOUNASMAA A N D TAFUA TA M M I N EN

7/3-hydroxyhyoscyamine) are reasonably well documented in the literature (vide infru), only they are treated separately here.

111. Synthesis

A number of new synthetic approaches to the tropane skeleton have been developed. The most important of these are described below. Readers with an interest in earlier approaches are referred to our previous review (6). A. BACKVALL SYNTHESIS OF TROPINE, PSEUDOTROPINE, SCOPINE, A N D PSEUDOSCOPINE Backvall and co-workers (350) have developed a general synthetic strategy that they have applied for the synthesis of tropine (l),pseudotropine (SO), scopine (136), and pseudoscopine (218) (Scheme 1). The common starting compound is chloroacetate 205, which is prepared from 3 3 cycloheptadienol benzyl ether (204) by palladium-catalyzed 1 ,Cacetoxychlorination. 1 . Synthesis of Tropine

Chloroacetate 205 is transformed into tosylate 206, which is catalytically [H2/RhCl(PPh,)3]reduced to compound 207. Hydrolysis of compound 207 leads to compound 208, and treatment of this with diethylazodicarboxylate (EtO,CN=NCO,Et), ZnCI,, and tributylphosphine (Bu,P), leads to compound 209. Cyclization (K,CO,), cleavage of the benzyl group (Na/NH,), and alkylation (CH,O, NaBH,) complete the synthesis of tropine (1) (Scheme 1). 2. Synthesis of Pseudotropine Chloroacetate 205 is transformed into tosylate 210, which is catalytically [H,/RhCI(PPh,),] reduced to compound 211. Hydrolysis of compound 211 to compound 212, followed by mesylation, leads to compound 213. Cyclization (K2C03),cleavage of the benzyl group (Na/NH,), and alkylation (CH,O, NaBH,) complete the synthesis of pseudotropine (50) (Scheme 1).

ooBz b,,,,6 oBz

5

L I O O O B Z-vi-viii A

R

OAC

c

204

206

iv

205

210

iV=

NHTs

i v c 2 1 9 R=Ac 220 R-H

207 R=OAc 208 R=OH 209 R=CI

211 R=Ac 212 R=H ‘ c 2 1 3 R=Ms

215 R=OH 218 R=CI

217

NHTS

NHTS

221 R=TBOMs

1

136

222 R-TBDMs

xxca R-H ‘ c 2 2 4

218

R-Ms

SCHEME1. Backvall and co-workers syntheses of tropine (l),pseudotropine (SO), scopine (136), and pseudoscopine (218). Reagents: i, Pd(OAc)?, LiCI, LiOAc, p-BQ, HOAc; ii, NaNHTs, Pd(PPh,),, CH,CN/DMSO (1 : I ) , 20°C. 12 hr; iii, H2/RhC1(PPh3)3,EtOH, 20°C 15 hr; iv, NaOH, MeOH/H20; v, Et02C-N=N-C02Et, ZnCl?, Bu,P, 20”C, 2 hr; vi, K2C03, MeOH, 20°C, 1 hr; vii, Na/NH,; viii, C H 2 0 , NaBH,; ix, NaNHTs, CH3CN/DMS0 (1: I), 80°C. 3 hr; x, MsCI/Et,N, THF; xi, DIBAL; xii, NaNHTs. DMSO, 80°C; xiii, NaNHTs, Pd(PPh,),, CH,CN; xiv, LiCI, MsCI: xv, MCPBA; xvi, NaC,,H,, THF. -78°C; xvii, MeI; xviii, H2/Pd/C, HCI,,, EtOH; xix, TBDMSCI; xx, Bu4NF, THF; xxi, H2/Pd/C, MeSO,H, MeOH.

80

MAURl LOUNASMAA A N D T A N A TAMMINEN

3. Synthesis of Scopine Chloroacetate 205 is transformed into chloroalcohol 214, which is then allowed to react with NaNHTs in the presence of Pd(PPh,),. The stereochemistry of the formed tosylate 215 at C-4 is inverted by the method of Collington and Meyers (LiCI, MsCl), with formation of chlorotosylate 216. Treatment of compound 216 with 3-chloroperoxybenzoic acid (MCPBA) affords the epoxy derivative 217. Cyclization (K,CO,), cleavage of the tosyl group (sodium naphthalide), alkylation (MeI), and cleavage of the benzyl group (H,/Pd; EtOH/HCI) complete the synthesis of scopine (136) (Scheme 1). 4. Synthesis of Pseudoscopine Chloroacetate 205 is transformed into tosylate 219, where the acetyl group is then replaced (via compound 220) by the bulky tert-butyldimethylsilyl (TBDMS) group leading to compound 221. Treatment of compound 221 with m-CPBA affords epoxy derivative 222 with the desired stereochemistry. The silyl group is then removed by tetrabutylammonium fluoride (Bu,NF). This leads to compound 223. Mesylation of 223 affords compound 224, Cyclization (K,CO,), cleavage of the tosyl group (sodium naphthalide), alkylation (MeI), and cleavage of the benzyl group (H,/Pd; MeOH/MeSO,H) complete the synthesis of pseudoscopine (218) (Scheme 1). B. BATHCATEA N D MALPASSSYNTHESIS Utilizing a similar strategy to that of Backvall et al. (vide supra), Bathgate and Malpass (351) prepared N-benzylnortrop-6-ene (225), Nbenzylnortropane (226), and nortropane (227). Cyclohepta- 1,3-diene is converted to compound 228, which by LiAIH, treatment yields compound 229. Compound 229 is transformed into the corresponding trans-chloroamine salt 230. Basification of the salt gives the free amine, which cyclizes to N-benzylnortrop-6-ene (225) (Scheme 2). To prepare the nortropanes, compound 228 is catalytically reduced to compound 231, which by LiAlH, treatment affords compound 232. Compound 232 is converted to the corresponding trans-chloroamine salt 233. Treatment of compound 233 with base gives the free amine, which cyclizes to N-benzylnortropane (226). Hydrogenolysis of compound 226 yields nortropane (227) (Scheme 2). Both the Backvall and Malpass approaches are related to the Kibayashi synthesis (6).

1. TROPANE ALKALOIDS

6

81

6.F>-) cl-

NH-COPh

NH-BZ

i

+

NH2-Bz

OH

228 iv

1

230

229

NH-COPh

;;

i

231

61

OH

() NH-BZ

225

ABZ5F) cl-

OH

El

232

233

iv

C

226

R=Bz

227

R=H

SCHEME 2. Bathgate and Malpass synthesis of N-benzylnortrop-6-ene (225), Nbenzylnortropane (226), and nortropane (227). Reagents: i, LiAIH4/Et20; ii, S0Cl2/LiCI/ CHCI,; iii, K2C03/ultrasound; iv, H2/Pd/C/MeOH; v, SOC12/CHC13;vi, pyridine.

C. DAVIESSYNTHESIS Davies et al. (352) have developed a new approach to the synthesis of tropane alkaloids. The method is based on a tandem cyclopropanation/ Cope rearrangement between metal-stabilized vinylcarbenoids and pyrroles. Decomposition of methyl 2-diazo-3-butenoate (234)by rhodium( 11) hexanoate [Rh,(OHex),] in the presence of N-{[2-(trimethylsilyI)ethoxy]carbonyl}pyrrole (235)leads to compound 236. Catalytic hydrogenation [H2/Rh,Cl(PPh,),] of compound 236 affords compound 237, which by tetrabutylammonium fluoride (Bu,NF) treatment leads to compound 238. Alkylation of compound 238 [CH,O; Na(CN)BH,] generates anhydroecgonine methyl ester (145)(Scheme 3). Similarly, decomposition of 3-diazo-4-penten-2-one (239)in the presence of N-{[2-(trimethylsilyI)ethoxy]carbonyl}pyrrole(235)leads to compound 240, catalytic hydrogenation of which affords compound 241. Bu,NF treatment of compound 241 yields compound 242,which by alkylation is transformed to (+)-ferruginine (180) (Scheme 4). Note that, owing to the absolute stereochemistry of the naturally occurring (+)-ferruginine (180)(73, we preferred to present (*)-ferruginine in Scheme 4 as the optical antipode to that given by Davies et al. (352) in the original article.

82

M A U R I LOUNASMAA A N D TAIUA T A M M I N E N

235

COOMe

237

236

234

R=COOEt

COOMs

...

COOMs

238

R=COOEt

145

SCHEME 3. Davies et ul. synthesis of anhydroecgonine methyl ester (145). Reagents: i. Rhz(OHex),; ii, Hz/RhCI(PPh,),; iii, Bu,NF; iv, C H 2 0 , Na(CN)BH,.

+

N-COOCHZCH2TMs

i

ii COMe

235

239

240 R=COOEt

...

iv

COMe

241 R=COOEt

COMe

242

COMe

180

SCHEME 4. Davies el a / . synthesis of (2)-ferruginine (180). Reagents: i, Rhz(OOct),; ii, H2/RhCl(PPh3),; iii, Bu,NF; iv, C H 2 0 , Na(CN)BH,.

D. LEETEA N D K I M SYNTHESIS

A biomimetic synthesis of tropinone (175),which consists of mercury(I1) acetate oxidation of hygrine (243) in boiling dilute acetic acid, has been described by Leete and Kim (353)(Scheme 5). E. LANSBURY SYNTHESIS Lansbury et al. (354) have shown that reaction between the dicarbanion, formed from 1,Cbisphenylsulfone (244) by n-BuLi treatment, and CY,(Y'methallyl diiodide (245)affords compound 246. Ozonolysis of compound

1.

83

TROPANE ALKALOIDS

243 175 SCHEME5. Leete and Kim biomirnetic synthesis of tropinone (175). Reagents: i, Hg(OAc),; ii, heat.

CH2

-

S02Ph

244

245

248

175

SCHEME 6. Lansbury et al. synthesis of tropinone (175). Reagents: i , n-BuLi; ii. I . O,, 2. CH,NH,.

246, followed by methylamine treatment, leads to tropinone (175) in 60-65% yield (Scheme 6). F.

MORIARTY SYNTHESIS

OF 2a-HYDROXYTROPINONE

Moriarty ef al. (355) have developed a method that permits a 2a-hydroxy group to be introduced into tropinone (175). Oxidation of tropinone (175) with iodobenzene diacetate in KOH/MeOH yields 2whydroxytropinone dimethyl acetal(247), which by hydrolysis (3 N HCI) affords 2a-hydroxytropinone (248)in 60% yield (Scheme 7 ) .

G. LALLEMAND SYNTHESIS OF 1-HYDROXYNORTROPANE SKELETON Lallemand and co-workers (356) have developed a general synthetic strategy for the 1-hydroxynortropane skeleton. The key intermediate, 2,3epoxy-4-azidocycloheptanone (250), prepared from cycloheptanone (249)by multistep synthesis, is transformed by the Staudinger reaction (PPh,, THF/H,, -50°C) in one pot to I-hydroxy-7-oxonortropane(251) (Scheme 8).

H. FURUYA SYNTHESIS Furuya and Okamoto (357)have developed a method that permits the transformation of nortropinone (252) to 2a-hydroxy-7-oxotropane (=N-

84

MAURI LOUNASMAA A N D T A W A TAMMINEN

pye- r HO

L

H

HO

ii

H

0

+Me

OMe

175

2411

247

SCHEME7. Moriarty et a / . synthesis of 2a-hydroxytropinone (248). Reagents: i. PhI(OAc),, KOHIMeOH; ii, 3 N HCI.

d

6

5

4

N3

24

250

251

SCHEME8. Lallemand and co-workers synthesis of the I-hydroxynortropane skeleton. Reagents: i, PPh,, T H F / H 2 0 , -50°C.

252

[fJq+]L

253

254

r+o > ,-

HO'

255

6

5

4

H

256

SCHEME 9. Furuya and Okamoto transformation. Reagents: i, I-BuOCI: ii. NaOMe; iii, Mel; iv, A1,O3 (basic).

methyl-2a-hydroxynortropan-7-one)(256). Treatment of nortropinone (252) with equimolar amounts of tert-butyl hypochloride and Na,CO, leads to N-chloronortropane (253), which by NaOMe/MeOH is transformed to aziridine 254. Reaction with Me1 affords the quaternary ammonium salt 255, which, after treatment with basic A1,0,, yields 2a-hydroxy-7oxotropane (256) (Scheme 9).

I . SPECKAMP SYNTHESIS Speckamp and co-workers (358) have shown that ally1 silane 257 in formic acid cyclizes to the tropane derivative 258 (Scheme 10).

1. TROPANE ALKALOIDS

G N-

C02Me

85

i

SiMe

257

2J8 R=C02Me

SCHEME 10. Speckamp and co-workers synthesis of the tropane skeleton. Reagents: i, HCOOH, 20°C.

J. XIANCel a / . SYNTHESIS OF BAOCONGTENG A (+)-BaogongtengA (183)has been synthesized by Xiang et a / . in seven steps starting from 6P-acetoxytropinone (259)(359). 6P-Acetoxytropinone (259)is brominated to the corresponding bromo derivative 260, which is transformed by Ag,CO, to the hydroxy derivative 261. Treatment of compound 261 with 1,2-ethanedithiol affords dithioketal 262, which is desulfurized with Raney nickel to compound 263. Oxidation of 263 with CrO, to compound 264, followed by NaBH, reduction to compound 265, gives the desired stereochemistry at C-2. Demethylation of compound 265 by the trichloroethyl chloroformate method (see Ref. 6) completes the total synthesis of (*)-baogongteng A (183)(Scheme 11).

K. JUNGet

a!.

SYNTHESIS OF BAOGONGTENG A

Jung et al. (360) have presented an alternative synthesis of (2)-baogongteng A (183).Reaction of the readily available N-benzyl-3-hydroxypyridinium bromide (266)with acrylonitrile gives a mixture of diastereoisomers, from which the desired exo isomer 267 is obtained after chromatographic separation. Isomer 267 is hydrogenated at room temperature to give the saturated ketone 268, which is then reduced with sodium borohydride to a mixture of diastereoisomers. The desired isomer 269 is obtained from the mixture by chromatographic separation. The secondary alcohol group is protected by conversion of 269 to the corresponding trimethylsilyl ether 270. Addition of methylmagnesium iodide to 270 produces the intermediate imine, which is treated with a solution of 15% aqueous ammonium chloride to yield the desired ketone 271 (the trimethylsilyl protecting group is also

86

MAURI LOUNASMAA A N D TARJA TAMMINEN

H,

Me-C-

Me-C-

II

0

259

EX] ti

cI1

Me-

Me-C-

II

0

II

8

260

OH

261

F$-v , E f

OH

H

1

-L Ma- C-

Me-

I1

0

262

C-

I1

0

263

264

OH N-H

4

Me-C-

Me-C-

II

0

II

0

285

103

SCHEME1 I . Xiang rt a / . synthesis of baogongteng A (183). Reagents: i. Br?; ii, Ag2C03; iii, 1,2-ethanedithiol; iv, Hz/Raney Ni; v , Cr03; vi, NaBH,; vii, 1. CICOOCH2CCI,, heat, 2. Zn, HOAc, room temperature.

Ff NC

L NC

&? 268

286

vi.

Me- C

II

0

269

270

u

183

271

1.

87

TROPANE ALKALOIDS

cleaved under these hydrolytic conditions). Baeyer-Villiger oxidation of ketone 271 affords the ester 272, from which the target compound 183 (baogongteng A) is obtained by catalytic hydrogenation (Scheme 12). L. HE A N D BROSSISYNTHESIS OF 6P-ACETOXYNORTROPANE In the synthesis of 6P-acetoxynortropane (273), He and Brossi (361) use a similar approach to that of Xiang et al. (vide supra). 6P-Acetoxytropinone (259) is reacted with 1,2-ethanedithiol in the presence of BF,.2Et,O to yield dithioketal274. Desulfurizing with Raney nickel in refluxing EtOH gives compound 275, which is demethylated with trichloroethyl chloroformate (see Ref. 6) to 6P-acetoxynortropane (273) (Scheme 13).

M. MANNSYNTHESIS OF OSCINE In the synthesis of oscine (=scopoline) (276), Mann and de Almeida Barbosa (362) react methyl pyrrole-N-carboxylate (277) with tetrabromoacetone (278) in the presence of diethylzinc (Et,Zn) in benzene to yield the dibromo adduct 279, which is debrominated to 3-0x0-8-azabicycloi3.2. I]oct-6-ene-8-carboxylate (280).Treatment of compound 280 with MCPBA affords compound 281, which is converted by diisobutylaluminum hydride (DIBAL) treatment to oscine (276) (Scheme 14). The Mann and de Almeida Barbosa synthesis of oscine (276) is an application of the Noyori method ( 6 ) .

Me-C-

II

0

1g0 259

[ e -0 I CI - O E ) ]

R=COOCH2CC13

M

rJL$(J

L o

Me-C-

II

0

e0 :

ii

Me-C-

274

-

II

0

E> 5

275

B

273

SCHEME 13. He and Brossi synthesis of 6P-acetoxynortropane (273).Reagents: i, 1,2ethanedithiol; ii, H2/Raney Ni, EtOH; iii, C1COOCH2CC13,toluene, heat; iv, Zn,HOAc, room temperature.

88

MAURI LOUNASMAA A N D TARJA TAMMINEN

277

278

280 R=COOMe

279 R=COOMe

281 R=COOMe

276

SCHEME 14. Mann and de Almeida Barbosa synthesis of oscine (276). Reagents: i, Et,Zn; ii, Zn-Cu, MeOH, NH,CI; iii, MCPBA; iv, DIBAL.

N. HARPER APPROACH

Harper and co-workers (363) have shown that the reaction between benzoylecgonine (148) and tetramethylethylenediamine(282) leads to a lipophilic ion pair 283, which can easily be converted to 2p-ethoxycarbonyI-3~-benzoyloxytropane (=benzoylecgonine ethyl ester) (284) in the absence of water (Scheme 15).

282

140

283

COOEt

284

SCHEME15. Synthesis of benzoylecgonine ethyl ester (284) by Harper and co-workers. Reagents: i, CH2C12,3 days, 20°C; ii, EtI, CH2C12.

1. TROPANE ALKALOIDS

89

IV. Reactions Since the last review in this series ( 6 ) , very few new reactions have been described in the tropane series.

A. THERMAL DEGRADATION Fowler and co-workers (364) present evidence to support the assumption that the first step in the thermal degradation of (-)-cocaine (149) consists of the formation of A3(4’-anhydroecgoninemethyl ester (=ecgonidine methyl ester) (285). Although A3‘4)-anhydroecgoninemethyl ester (285) itself could not be detected directly, its participation allowed the construction of a reasonable series of reactions leading to the formation of thermal products N-methylpyrrole (286) and methyl 3-butenoate (287) (Scheme 16). Further evidence for the intermediacy of ecgonidine methyl ester (285) in the thermal degradation of (-)-cocaine (149) is found in the generation of tropidine (288) from 3P-benzoyloxytropane (55) (=desmethoxycarbonylcocaine) at 525°C (Scheme 17).

B. DEMETHYLATION Effective photooxidation methods for N-demethylation of tropane derivatives have been presented by Santamaria et al. (365,366).N,N’-Dimethyl2,7-diazapyrenium-bis(tetrafluoroborate)(DAP2+-2BF;) sensitized photooxidation of 3a-hydroxytropane (l),3-oxotropane (173, and atropine (33) affords3a-hydroxynortropane (289), 3-oxonortropane (290),and noratropine (27), respectively, in high yield (Scheme 18). Similarly, 9,lO-dicyanoanthracene (DCA) sensitized photooxidation of 3a-hydroxytropane (l), 3-oxotropane (175), and atropine (33) yields the corresponding nor- and N-formylnortropane derivatives 289 and 291,290 and 292, and 27 and 293 in variable proportions (Scheme 19). If a salt such as LiCIO, and Mg(C104)2 is present, the formation of the nortropane derivatives 289, 290, and 27 is highly favored.

C. PHOTOCYANATION

In connection with work on the N-demethylation of tropane derivatives (vide supra), Santamaria et al. (367) developed an effective cyanation

90

MAURl LOUNASMAA A N D TARJA TAMMINEN COOMe

149

286

287

SCHEME 16. Thermal degradation of (-)-cocaine (149). Reagents: i. 550°C.

55

288

SCHEME 17. Thermal degradation of 3P-benzoyloxytropane (55). Reagents: i , 525°C.

i

1

R=H

289 R=H

35

R=tropoyl

27

i 175

R=tropoyl

IF0 290

SCHEME 18. Demethylation of 3a-hydroxytropane (l),3-oxotropane (175). and atropine (33)using N,N'-dimethyl-2,7-diazapyrenium-bis(tetrafluoroborate) (DAP2' .2BF;) sensitized photooxidation. Reagents: i, DAP2+.2BF;, CH3CN, h v / 0 2 .

91

1. TROPANE ALKALOIDS

1

R=R1=Rz=H

289

291

33

R=tropoyl: Rl=Rz=H

27

293

290

292

175

SCHEME 19. Demethylation of 3a-hydroxytropane (l),3-oxotropane (175),and atropine (33) using 9,lO-dicyanoanthracene (DCA) sensitized photooxidation. Reagents: i. DCA. hvl02.

method for tertiary amines. The (DAP2+.2BF,) sensitized photooxidation of 3a-hydroxytropane (l),3-oxotropane (175), atropine (33), and scopolamine (140) in the presence of cyanotrimethylsilane (Me,SiCN) affords the N-cyanomethyl compounds 294, 295, 296, and 297, respectively (Scheme 20).

1

R=R,=Rz=H

33

R=tropoyl; Rl=R2=H

296 R3=CHzCN

140

R=tropoyl; R1 .R2= -0-

297 R3=CHzCN

175

294 R3=CHzCN

295 R=CH2CN

SCHEME20. Photocyanation of 3a-hydroxytropane (l),3-oxotropane (175),atropine (33), and scopolamine (140).Reagents: i, Me,SiCn, DAP2+.2BF;, h v 1 0 2 .

92

MAURl LOUNASMAA A N D TARJA TAMMINEN

V. Biosynthesis An excellent review of the biosynthesis of tropane alkaloids was published in 1990 by Leete (368), and a special chapter by Robins and Walton on the same topic follows this chapter (7).Moreover, in the second edition of the authoritative book on the biosynthesis of secondary metabolites, Herbert brings the description of the biosynthesis of tropane alkaloids up to date (369). In view of this, we note here only the essential changes required in the biosynthetic schemes published in our earlier chapter in this treatise ( 6 ) . In addition, we comment on a few results appearing after the publication of Leete (368). Contrary to what was thought earlier (e.g., Refs. 370-373), Leete (368) now proposes that 6-N-methylornithine (298) is not an intermediate in the formation of N-methylputrescine (299). To explain the integrity of the C 2 and C-5 carbons found in feeding experiments with certain Datura species [ D . innoxia, D. metel, and D. stramonium (374-376)], Leete further suggests (368) that ornithine (300) is decarboxylated to afford a “bound form” of putrescine (301), which is then methylated to yield first a “bound form” of N-methylputrescine (302) and then N-methylputrescine (299) (an asymmetrical intermediate). Oxidation of 299 leads to 4-methylaminobutanal (303), which then cyclizes to the N-methyl-A’-pyrrolinium salt (304) (Scheme 21). In Duboisia leichhardtii, Hyoscyamus albus, Nicandra physaloides, and Erythroxylum coca, a symmetrical intermediate is needed to explain equal labeling of the C-1 and C-5 bridgehead carbons found in feeding experiments (377-384). Leete proposes (368) that, in these cases, the above-mentioned “bound form” of putrescine (301) is first transformed to putrescine (305) and then methylated to N-methylputrescine (299) (this time racemic). The N-methyl-A’-pyrrolinium salt (304) is subsequently formed via 4-methyl-aminobutanal (303) (Scheme 21). In further steps, two units of acetyl-coenzyme A condense with the N-methyl-A’-pyrrolinium salt (304) to form the intermediate 305, which then leads, via the normal route (305 + 306 -+ 307 + 308), first to tropinone (175) and tropine (1) and then to hyoscyamine (34) and similar compounds (see Ref. 6 ) (Scheme 22). Leete (368) also proposed a new biosynthetic scheme for cocaine (1491, and suggests that the N-methyl-A’-pyrrolinium salt (304) (racemic) reacts with one unit of acetyl-coenzyme A to give the coenzyme A thioester of 1 -methylpyrrolidine-2-acetic acid (309). Reaction of 309 with a second acetyl-coenzyme A unit yields thioester 310, which is first oxidized to the iminium ion 311 and then transformed by a Mannich reaction to the

1. TROPANE ALKALOIDS

300

305

301

298

93

299

303

t

I

302

304

SCHEME21. Biosynthetic formation of N-methyl-A'-pyrrolinium salt (304).

304

305

307

308

306

175

F%IHE N H 0 II

--c

O-C-CHPh

I

CH20H

1

34

SCHEME22. Biosynthetic formation of tropinone (175). tropine (I),and hyoscyamine (34)

from N-methyl-A'-pyrrolinium salt (304).

94

MAURI LOUNASMAA AND TARJA TA M M I N EN

X

-

-

ENM -FO

304

COSCoA

-

309

310

COSCOA

COSCoA

-

F:M&O

p

31 1

&

COOMe

-

O

COOMe

147

p

$

z

O

-

313

312

E X H-

-

EW-M&O

COOMe

p

$

p

-

P

h

148

SCHEME 23. Biosynthetic formation of cocaine (149) from N-methyl-A'-pyrrolinium salt (3041.

tropinone intermediate 312. Transesterification to 2-methoxycarbonyl-3tropinone (313) followed by reduction leads to the corresponding alcohol, methylecgonine (147). Benzoylation of 147 completes the biosynthetic formation of cocaine (149) (Scheme 23). It was recently shown (385-389) that scopolamine (140), a well-known metabolite of hyoscyamine (34), is not formed via 6,7-dehydrohyoscyamine (314) as previously thought (6). Rather, the 6p-hydroxy group 0

II

Q- C-

CHPh I &$OH

314

of the initially formed 60-hydroxyhyoscyamine (=anisodamine) (87) attacks directly the C-7 position of 87, giving rise to the epoxy ring (Scheme 24).

1.

II

I CH~OH

95

TROPANE ALKALOIDS

p x H -QTXH

34

(t*CHPh II I

(tC-CHPh I CH~OH

CH~OH

87

140

SCHEME 24. Biosynthetic formation of scopolamine (140) from hyoscyamine (34).

VI. Spectroscopy We discuss here only new structure elucidations by 'H-NMR and I3CNMR spectroscopy and mass spectrometry, which are the three most important spectroscopic methods for the tropane alkaloids. Since our last review in 1988 ( 6 ) , relatively few new spectroscopic data for the tropane alkaloids have been published (vide infra). A. 'H-NMR

AND

13C-NMR SPECTROSCOPY

Anisodamine (87)and anisodine (142) have been studied by 'H- and I3C-NMR spectroscopy using two-dimensional and double-resonance techniques (390). From the results, it was surprisingly concluded that the N-methyl group in anisodamine (=6P-hydroxyhyoscyamine) (87)is equatorial. Earlier 13C-NMR measurements (391) had suggested that the N-methyl group in anisodamine 87 is axially oriented. The N-methyl group stereochemistry in tropane alkaloid salts has been studied by 'H and I3C NMR with application of the slow exchange limit (SEL) technique (392).At equilibrium, most tropane alkaloid salts showed a preponderance for the equatorial position (equatorial/axial ratio, -7/ 1 in D 2 0 , -1811 in CD2C12).Scopolamine hydrobromide expressed a strong solvent dependence in its equatorial/axial ratio (1118 in D,O; 18/1 in CD,C12). Complete 'H-NMR (Table V) and 13C-NMR (Fig. 2 ) data have been published for oscine (=scopoline or 3a,6a-oxido-7P-hydroxytropane) (276)(393). Complete and unequivocal 'H-NMR (Table V ) and "C-NMR (Fig. 2 ) spectral assignments have been reported for scopolamine (140) (free base in CDCI, solution). Each atom of the piperidine ring could be differentiated, and some earlier assignments were corrected. In contrast to the earlier results and general presumptions, it was shown that the N methyl group is equatorially disposed relative to the piperidine ring (394).

96

MAURI LOUNASMAA A N D TARJA TAMMINEN

TABLE V ’H NMR DATAOF SCOPOLAMINE (140) ANDOSCINE ( = SCOPOLINE) (276)“

H- 1 H-2m H-2eq H-3 H-4m H-4eq

H-5 H-6 H-7 H-2’ H-3’a H-3’b 2 X 0-H 2 x m-H 1 X p-H CH3-N

140

276

2.94 1.99 1.31 4.98 2.08 1.55 3.08 3.37 2.70 3.72 3.77 4.30 7.21 7.29 7.18 2.42

3.67 2.37 1.24 4.25 1.89 1.43 2.97 3.89 4.50 2.58

’ Data were recorded in CDCI,. 56.4 57.9

31.0

CH~OH 64.0

276

140

FIG.2. I3C-NMR data of scopolamine (140) and oscine (276).

The structure of the “trimeric” tropane alkaloid grahamine (203) was mainly solved by mass spectrometry (vide infra) and two-dimensional NMR spectroscopy (152). Full I3C-NMR spectral assignments of grahamine 203 are given (Fig. 3) (152). Finally, some ’H-NMR data have been published for a-belladonnine (191), p-belladonnine (192), a-scopadonnine (193), and p-scopadonnine (194) (395,396).

B. MASS SPECTROMETRY Electron-impact (EI) mass spectrometry (often combined with gas chromatography techniques) has been abundantly used for the determination

1. TROPANE ALKALOIDS

0

97

H

of tropane structures of various sources (e.g., (397,398).The mass spectral fragmentation of the tropane alkaloids was described in detail in our earlier review (6). Fast atom bombardment (FAB) mass spectrometry, combined with two-dimensional NMR spectroscopy (uide supra) was used in the structure elucidation of the “trimeric” tropane alkaloid grahamine (203) (152). The results showed that the molecular weight of grahamine is 871, which is the highest value found among the tropane alkaloids thus far.

VII. Pharmacology An extremely large number of pharmacological reports on the tropane alkaloids have appeared. The section “Pharmacology” in Chemical Abstracts [Vols. 106-1 17 (1987-1992)] lists about 500 articles. As it would go far beyond the scope of this chapter to deal in an adequate way with

98

MAURI LOUNASMAA A N D TARJA TAMMINEN

these articles, interested readers are referred to Chemical Abstracts. Many of the same articles are mentioned in Natural Product Reports (9). Only some of the papers (vide infra) are discussed here. The ideal conformations of Atropa belladonna alkaloids were shown by molecular mechanics MMPM calculations to be quite different from those found in the crystalline state. An intramolecular hydrogen bond was present between the hydroxy group and the carbonyl oxygen. A model involving four recognition sites between the muscarinic acetylcholine receptor and the anticholinergic drug-receptor interaction in mydriasis has been suggested. Calculated ideal conformations were used as the pharmacologically active ones (399). The effects of several analogs of atropine (33) have been studied in the specific binding of 3H-labeled quinuclidinyl benzilate to muscarinic receptors in rat brain (400). Acute carbofuran intoxication can be prevented and antagonized by atropine (401). The calming effect of atropine (33) on heart rate fluctuations, especially at high altitudes, has been documented (402,403).Evidently, intramuscularly administered atropine also prevents the effects of physostigmine on human vision (404). On the negative side, atropine has been found to cause abnormal gait sequence in locomotion (405,406)and to affect the Randt Memory Test (407). Study has been made of the effect of intravenous atropine on gastric emptying (408). Pretreatment of asthmatics with atropine sulfate affects adenosineinduced bronchoconstriction in humans (409).The effects of atropine on glucose metabolism have been detailed (410),and a recent review summarized the effects of atropine (33) on performance and physiology (411). A review article has also been published on the neurochemical and physiological effects of cocaine (149), as related to behavioral and learning mechanisms (412). The mechanism of cocaine-induced teratogenesis has been shown to involve inhibition of the uptake of norepinephrine and placental vasoconstriction (413). In mice, cocaine affects the heart rate and locomotor activities (414), as well as the immune system (415). The neuropharmacological aspects of cocaine ingestion have been studied (416,417),and the pharmacokinetics and biotransformations of cocaine (149) in man have been reviewed (418). Light has recently been thrown on the interaction of cocaine with central serotonergic neuronal systems (419-421). Four earlier unreported metabolites (ecgonidine, norecgonidine methyl ester, norecgonine methyl ester, and m-hydroxybenzoylecgonine) have been found in urine (422).Considerable attention has been focused on the clinical pharmacology and toxicity of cocaine (149) (423). Several groups have examined the cocaine receptors on dopamine transporters (e.g., 424,425)and the structure-activity relationships of cocaine analogs (e.g., 4 2 6 4 2 9 ) . Both topics have recently been reviewed (430).

1.

TROPANE ALKALOIDS

99

In a study (431) of whether prenatal cocaine exposure alters the later reward efficacy of cocaine in rats, evidence was found to suggest that offspring so exposed are less likely to develop a preference for stimuli associated with cocaine. In contrast to this, another group (432) has reported that cocaine in utero increases the response to cocaine in adult offspring. An annotated bibliography on cocaine is also available (433). Mechanisms have been proposed for scopolamine (140) in its role as an amnestic (434) and inducant of dementia (435,436).Evidently, scopolamine has dissociative effects on the working memory (437-439) and on the performance and learning of motor skills (440,441).The effects on memory, language, visuospatial praxis, and psychomotor speed have also been reinvestigated (442). It has been shown that scopolamine affects the cognitive processes involved in selective object exploration (443). The behavioral effects of scopolamine have been investigated (444-447). The cognitive effects of scopolamine are quantitatively more pronounced in elderly than in young subjects (448). I n uiuo metabolism of scopolamine (140) has been studied in rats, mice, guinea pigs, and rabbits. (449). In an elucidation of the structures of eight urinary metabolites by mass spectrometry and NMR spectroscopy, it was shown that scopolamine metabolism is highly species specific.

VIII. Perspectives Thus far 200 tropane alkaloids have been isolated from different plant sources, and there is no reason to doubt that the search for new tropane alkaloids will continue with equal intensity during the coming years. Despite the new syntheses developed, most of the medicinally important tropane alkaloids are still obtained more economically by extraction from plant material. Tremendous efforts have recently been made to develop economically feasible methods for the production of tropane alkaloids, especially medicinally important ones, by applying cell culture techniques. Only a fraction of the extensive literature concerning the production of tropane alkaloids in cell cultures is mentioned below. Readers with a deeper interest in the subject are referred to recent review articles (450-456) and references therein. The main efforts toward the industrial preparation of tropane alkaloids by cell culture methods have been concentrated on hyoscyamine (34) and scopolamine (140), and to a lesser extent on anisodamine (87)and aniso-

100

MAURI LOUNASMAA AND TARJA TAMMINEN

dine (142) (4.57462). Unfortunately, the alkaloid content in “normally” cultured cells has so far generally been lower than that in intact plants (463). However, root cultures of some species (464-473), especially when Agrobacteriurn-mediated transformation systems (214,259,277,283,474484) are used, may hold some promise for tropane alkaloid production. Finally, it can be expected that methods based on gene transfer systems will be commercially competitive in the production of medicinally important tropane alkaloids in the future.

REFERENCES I . H. L. Holmes, in “The Alkaloids” (R. H . F. Manske and H . L. Holmes, eds.), Vol. 1, p. 271. Academic Press, New York, 1950. 2. G. Fodor, in “The Alkaloids” (R. H. F. Manske, ed.), Vol. 6, p. 145. Academic Press, New York, 1960. 3. G. Fodor, in “The Alkaloids” (R. H. F. Manske and H. L. Holmes, eds.), Vol. 9, p. 269. Academic Press, New York, 1967. 4. G. Fodor, in “The Alkaloids” (R.H. F. Manske and H. L. Holmes, eds.), Vol. 13, p. 351. Academic Press, New York, 1971. 5. R. L. Clarke, in “The Alkaloids” (R. H . F. Manske and H. L. Holmes, eds.), Vol. 16, p. 83. Academic Press, New York, 1977. 6. M. Lounasmaa, in “The Alkaloids” (A. Brossi, ed.), Vol. 33, p. 1. Academic Press, New York, 1988. 7. R. J. Robins and N . J. Walton, this volume, Chap. 2. 8. See, for example, G. A. Swan, “An Introduction to the Alkaloids.” p. 50. Blackwell, Oxford, 1967. 9. G. Fodor and R. Dharanipragada, Nut. Prod. Rep. 5 , 67 (1988); G. Fodor and R. Dharanipragada, Nar. Prod. Rep. 7,539 (1990); G. Fodor and R. Dharanipragada, Nut. Prod. Rep. 8, 603 (1991); G. Fodor and R. Dharanipragada, Nut. Prod. Rep. 10, 199 ( 1993). 10. L. N. Bereznegovskaya, “Physiology and Biochemistry of Tropane Alkaloids,” Izd. Tomsk. Gos. Uniu., Tomsk. ( U . S . S . R . )(1974); Chem. Ahsrr. 82, 121987f (1975). 11. R. E. Schultes and A. Hofmann, “The Botany and Chemistry of Hallucinogens,” 2nd Ed., Charles C. Thomas Publ., Springfield, Illinois, 1980. 12. Various authors, J. Ethnopharmacol. 3, I 1 1 (1981). 13. A. Romeike, Bot. Nor. 131, 85 (1978). 14. W. C. Evans, in “The Biology and Taxonomy of the Solanaceae” ( J . G. Hawkes, R. N . Lester, and A. D. Skelding, eds.), Linnean Society Symposium Series, No. 7, p. 241. Academic Press, London, 1979. 15. R. Hegnauer, “Chemotaxonomie der Pflanzen,” Vols. 1-10, Birkhauser, Basle and Stu t tgart , 1962- 1992. 16. G. Barger, W. F. Martin, and W. Mitchell, J. Chem. SOC., 1820 (1937). 17. P. Bachmann, K. Steffen, and F.-C. Czygan, 2. Phytother. 11, 30 (1990). 18. W. C. Evans and M. Pe Than, J. Phurm. Pharmacol. 14, 147 (1962). 19. H. King and L. L. Ware, J. Chem. SOC., 331 (1941).

1. TROPANE ALKALOIDS

101

J. D. Leary, K. L. Khanna, A. E. Schwarting, and J. M. Bobbitt, Lloydia 26,44 (1963). H. Staub, Helu. Chim. Acta 45, 2297 (1962). M. I. Berry and P. Jackson, Planta Med. 30, 281 (1976). T . Plowman and L. Rivier, Ann. Bot. (London) 51, 641 (1983). R. Dahlgren, Bot. J . Linn. Soc. 80, 91 (1980). R. Dahlgren, Rev. Latinoam. Quim. 1 (Suppl.), 10 (1989). P. TCtenyi, Ann. Mo. Bot. Card. 74, 600 (1987). L. Haegi, Telopea 2, 173 (1981). K. Hammer, A. Romeike, and C. Tittel, KulturpJlanzr 31, 13 (1983). M. L. Bristol, Bot. Mus. Leap., Haru. Uniu.22, 165 (1966). M. L. Bristol, Bot. Mus. Leap., Hnru. Uniu. 21, 229 (1966). M. L. Bristol, W. C. Evans, and J. F. Lampard, Lloydia 32, 123 (1969). H. Heltmann, Herba Hung. 18, 101 (1979); Chem. Absfr. 93, 200948~(1980). E. Weinert, Feddes Repertorium 82, 617 (1972). A. Baytop, in “Flora of Turkey and the East Aegean Islands” (P. H . Davis, ed.), Vol. 6, p. 437. University Press, Edinburgh, Scotland, 1978. 35. P. Xiao and L. He, J . Ethnopharmacol. 8, 1 (1983). 36. 0. E. Schulz, in “Das Pflanzenreich” (A. Engler, ed.), Vol. 4, Sect. 134, Erythroxylaceae, p. 1. Wilhelm Engelmann, Leipzig, 1907. 37. T. Plowman, J. Psychedelic Drugs 11, 103 (1973). 38. T. Plowman, Bot. J. Linn. SOC. 84, 329 (1982). 39. B. A. Bohm, F. R. Ganders, and T. Plowman, Syst. Bot. 7, 121 (1982). 40. I. R. C. Bick and H.-M. Leow, J. Indian Chem. SOC. 55, I103 (1978). 41. I. R. C. Bick, J. W. Gillard, H-M. Leow, M. Lounasmaa, J. Pusset, and T. SCvenet, Planta Med. 41, 379 (1981). 42. W. Gritsanapan and W. J. Griffin, Phytochemistry 30, 2667 (1991). 43. L. Cosson, J. C. Vaillant, and E. Dequeant, Phytochemistry 15, 818 (1976). 44. P. Xiao and L . He, Planta Med. 45, 112 (1982). 45. S. R. Johns, J. A. Lamberton, and A. A. Sioumis, Aust. J. Chem. 24, 2399 (1971). 46. 0. Luanratana and W. J. Griffin, J. Nut. Prod. 43, 552 (1980). 47. M. 0. A. El Sheikh, G. M. El Hassan, A.-R. El Tayeb Abdel Hafeez, A. A. Abdalla, and M. D. Antoun, Planta Med. 45, I16 (1982). 48. S. Gupta, V. S. Prabhakar, and C. L. Madan, Planta Med. 23, 370 (1973). 49. N. N. Sabri, S. El-Masry, and S. M. Khafagy, Planta Med. 23, 4 (1973). 50. Y. M. A. El-Imam and W. C. Evans, Planta Med. 50, 86 (1984). 51. W. C. Evans and K. P. A. Ramsey, Phytochemistry 20, 497 (1981). 52. W. C. Evans and K. P. A. Ramsey, J. Pharm. Pharrnacol. 33 (Suppl.), 14P (1981). 53. R. Hegnauer, J. Ethnopharmacol. 3, 279 (1981). 54. W. C. Evans, J . Ethnopharmacol. 3, 265 (1981). 55. W. J. Griffin, Aust. J . Chem. 31, 1161 (1978). 56. S. R. Johns and J. A. Lamberton, Aust. J. Chem. 20, 1301 (1967). 57. A. San-Martin, C. Labbe, 0. Munoz, M. Castillo, M. Reina, G. de la Fuente, and A. Gonzales, Phytochemistry 26, 819 (1987). 58. V. Gambaro, C. Labbe, and M. Castillo, Phytochemisfry 22, 1838 (1983). 59. V. Gambaro, C. Labbe, and M. Castillo, Bol. Soc. Chi/. Quim. 27, 296 (1982). 60. A. San-Martin, J. Rovirosa, V. Gambaro, and M. Castillo, Phytochemisfry 19, 2007 (1980). 61. W. C. Evans, A. Ghani, and V. A. Woolley, Phyfochemistry 11, 2527 (1972). 62. W. C. Evans and M. Wellendorf, J. Chem. Soc., 1406 (1959). 63. P. J. Beresford and J. G. Woolley, Phytochemistry 13, 2511 (1974).

20. 21. 22. 23. 24. 25. 26. 27. 28. 29. 30. 31. 32. 33. 34.

102

MAURI LOUNASMAA A N D TARJA TAMMINEN

64. W. C. Evans, A. Ghani, and V. A. Woolley, Phytochemistry 11, 470 (1972). 65. B. P. Jackson and M. I. Berry, Phytochernistry 12, 1165 (1973). 66. T. Yao, Z. Chen, D. Yi, and G. Xu, Yaoxue Xuebao 16, 582 (1981); Chem. Abstr. 96, 48972c (1982). 67. P. J. Beresford and J. G. Woolley, Phytochemistry 13, 1249 (1974). 68. J. Parello, P. Longevialle, W. Vetter, and J. A. McCloskey, Bull. Soc. Chim. Fr.. 2787 (1963). 69. V. I. Muraveva. A. G. Mamedova, and A. 1. Bankovskii, Zh. Obshch. Khim. 33,1690 (1963); Chem. Abstr. 59, 13114g (1963). 70. T. F. Platonova and A . D. Kuzovkov, Med. Prom. SSSR 17, 19 (1963). 71. I. R. C. Bick, J. W. Gillard, H.-M. Leow, and N. W. Preston, Ailst. J. Chem. 32, 2071 (1979). 72. I . R. C. Bick. J. W. Gillard, and H.-M. Leow, Aust. J. Chem. 32, 1827 (1979). 73. W. D. S. Motherwell, N. W. Isaacs, 0. Kennard, I. R. C. Bick, J. B. Bremner, and J. W. Gillard, J. Chem. Soc. D , 133 (1971). 74. I. R. C. Bick, J. W. Gillard, and H.-M. Leow. Aust. J. Chrm. 32, 2523 (1979). 75. I. R. C. Bick, J. W. Gillard, and H.-M. Leow, Aust. J. Chem. 32, 2537 (1979). 76. 1. R. C. Bick, J. W. Gillard, and M. Woodruff, Chem. Itid. (London), 794 (1975). 77. C. Kan-Fan and M. Lounasmaa, Acta Chem. Scand. 27, 1039 (1973). 78. M. Lounasmaa, P. M. Wovkulich, and E. Wenkert. J. Org. Chem. 40, 3694 (1975). 79. M. Lounasmaa, J. Pusset, and T. Sevenet, Phytochemistry 19, 949 (1980). 79a. M. Lounasmaa, Planta Med. 27, 83 (1975). 80. M. Lounasmaa, J. Pusset, and T. Sevenet, Phytochemistry 19, 953 (1980). 81. A. Kato, Phytochemistry 14, 1458 (1975). 82. A. Kato and J. Takahashi, Phytochemistry 15, 220 (1976). 83. J. W. Loder and G. B. Russell, Aust. J. Chem. 22, 1271 (1969). 84. D. H. Gnecco Medina, M. Pusset, J. Pusset, and H. P. Husson, J . Nut. Prod. 46, 398 ( 1983). 84a. J. W. Loder and G. B. Russell, Tetrahedron Lett., 6327 (1966). 85. D. Arbain, R. D. Wiryani, and M. V. Sargent, Aust. J. Chem. 44, 1013 (1991). 86. Y. M. A. El-Imam, W. C. Evans, and T. Plowman, Phytochemistry 24, 2285 (1985). 87. G. H. Aynilian, J. A. Duke, W. A. Gentner, and N. R. Farnsworth, J. Pharm. Sci. 63, 1938 (1974). 88. C. Hartwich, Arch. Pharm. (Weinheim, Ger.) 241, 617 (1903). 89. R. Hegnauer and L. H. Fikenscher, Pharm. Acta Helu. 35, 43 (1960). 90. J. R. Matchett and J. Levine, J . Am. Chem. Soc. 63, 2444 (1941). 91. W. Lossen, Liebigs Ann. Chem. 133, 351 (1865). 92. A. W. K. de Jong, Recl. Trau. Chim. Puys-BUS59, 687 (1940). 93. B. Holmstedt, E . Jaatmaa, K. Leander, and T. Plowman, Phyfocheniistry 16, 1753 (1977). 94. M. Youssefi, R. G. Cooks, and J. McLaughlin, J . Am. Chem. Soc. 101, 3400 (1979). 95. C. E. Turner, C. Y. Ma, and M. A. Elsohly, J. Ethnophurrnucol. 3, 293 (1981). 96. L . Rivier, J. Ethnopharmacol. 3, 313 (1981). 97. G. Espinel Ovalle and I. Guzman Parra, Reu. Colomb. Cienc. Quim-Farm.1,95 (1971); Chem. Abstr. 76, 89994s (1972). 98. 0. Hesse, Ber. Dtsch. Chem. Gas. 22, 665 (1889). 99. C. Liebermann, Ber. Dtsch. Chem. Ges. 40, 3602 (1907). 100. J. Moore, J. Assoc. Ofl.Anal. Chem. 56, 1199 (1973). 101. C. Liebermann, Ber. Dtsch. Chem. Ges. 21, 2342 (1888). 102. E. Graf and W. Lude, Arch. Pharm. (Weinheim, Ger.) 310, 1005 (1977).

1.

TROPANE ALKALOIDS

103

103. E. Graf and W. Lude, Arch. Pharm. (Weinheim. G e r . ) 311, 139 (1978). 104. Y. M. A. El-Iman, W. C. Evans, R. J. Grout, and K. P. A. Ramsey, Phytochemistry 26, 2385 (1987). 105. J. T. H. Agar, W. C. Evans, and P. G. Treagust, J. Pharm. Pharmacol. 26 (Suppl.), l l l P (1974). 106. J . T. H. Agar and W. C. J. Evans, J. Chem. Soc., Perkin Trans. 1. 1550 (1976). 107. J. T. H. Agar and W. C. J. Evans, J . Pharm. Phurmacol. 27 (Suppl.), 85P (1975). 108. R. N. Chopra and N. N. Ghosh, Arch. Pharm. (Weinheim, G e r . ) 276, 340 (1938). 109. M. A. I. Al-Yahya, W. C. Evans, and R. J . Grout, J. Chem. S O C . , Perkin Trans. I , 2130 (1979). 110. M.T. Campos-Neves and A. S. Campos-Neves, Garcia de Orta 14,97 (1966); Chem. Abstr. 69, 93608s (1968). 11 1. Y. M. A. El-Imam, W. C. Evans, and R. J. Grout, Phytochemistry 27, 2181 (1988). 112. S. R. Johns, J. A. Lamberton, and A. A. Sioumis, Aust. J . Chem. 23, 421 (1970). 113. M. S. Al-Said, W. C. Evans, and R. J. Grout, Phytochemistry 28, 671 (1989). 114. M. S. Al-Said, W. C. Evans, and R. J. Grout, Phytochernistry 28, 3211 (1989). 115. M. S. Al-Said, W. C. Evans, and R. J. Grout, J . Chem. Soc., Perkin Trans. I , 957 ( I 986). 116. J. Bosser and R. Pernet, Nut. Malgache 9, 195 (1957);Chem. Abstr. 54,25068f (1960). 117. M. S. Al-Said, W. C. Evans, and R. J. Grout, Phytochemistry 25, 851 (1986). 118. V. G . Cairo, J. De Budowski, F. Delle Monache, and G. B. Marini-Bettolo, Atti Accad. Naz. Lincei, Cl. Sci. Fis., Mat. Nut., Rend. 62, 363 (1977); Chem. Abstr. 89, 87163q ( 1978). 119. W. C. Evans and K. P. A. Rarnsey, Phytochemistry 22, 2219 (1983). 120. J. R. Cannon, K. R. Joshi, G. V. Meehan, and J . R. Williams, A u s f . J. Chem. 22, 221 ( 1969). 121. W. C. Evans and P. G. Treagust, Phytochemistry U ,2505 (1973). 122. J. B. Bremner and J. R. Cannon, Aust. J. Chem. 21, 1369 (1968). 123. W. C. Evans and K. P. A. Ramsey, J. Pharm. Pharmacol. 31 (Suppl.), 9P (1979). 124. Y. M. A. El-Imam, W. C. Evans, and L. Haegi, Int. J. Crude Drug Res. 28, 237 (1990). 125. 1. R. C. Bick, J. B. Bremner, J. W. Gillard, and K. N . Winzenberg, Aust. J. Chem. 27, 2515 (1974). 126. G. S. Kennedy, Phytochemistry 10, 1335 (1971). 127. 0. Luanratana and W. J. Griffin, Phytochemistry 21, 449 (1982). 128. T. Endo and Y. Yamada, Phytochernistry 24, 1233 (1985). 129. W. J. Griffin, Aust. J . Pharm. 46, S128 (1965). 130. W. Deckers and J. Maier, Chem. Ber. 86, 1423 (1953). 131. E. 1. Rosenblum and W. S. Taylor, J. Pharm. Phurmacol. 6, 410 (1954). 132. K. Kagei, M. Ikeda, T. Sato, Y. Ogata, T . Kunii, S. Toyoshima, and S. Matsuura, Yakugaku Zasshi 100, 216 (1980); Chem. Abstr. 92, 194445s (1980). 133. K. Kagei, M. Ikeda, T. Sato, Y. Ogata, S. Toyoshima, and S. Matsuura, Yakugaku Zasshi 100, 574 (1980); Chem. Abstr. 93, 146285~(1980). 134. Eisai Co., Ltd., Jpn. Kokai Tokkyo Koho J P 81,121,494 (1981); Chem. Abstr. 96, I16542 (1982). 135. Y. Kitamura, Y. Sugimoto, T. Samejima, K. Hayashida, and H. Miura, Chem. Pharm. Bull. 39, 1263 (1991). 136. R. E. A. Drey and G. E. Foster, J. Pharm. Pharmacol. 5 , 839 (1953). 137. K. Kagei, S. Hemmi, H. Shirai, and S. Toyoshima, Shoyakugaku Zasshi 32,228 (1978); Chem. Abstr. 91, 16688~(1979).

104

MAURI LOUNASMAA A N D TARJA TAMMINEN

138. Y. Yamada and T. Endo, Plant Cell Rep. 3, 186 (1984). 139. T. Hashimoto, Y. Yukimune, and Y. Yamada, J. Plant Physiol. 124, 61 (1986). 140. J. F. Coulsen and W. J. Griffin, Planta Med. 15, 459 (1967). 141. J. F. Coulsen and W. J. Griffin, Planta Med. 16, 174 (1968). 142. G. Barger, W. F. Martin, and W. Mitchell, J. Chem. Soc., 1685 (1938). 143. K. J. Sipply and H. Friedrich, Planta Med. (Suppl.), 186 (1975). 144. F. H. Carr and W. C. Reynolds, J. Chem. Soc. 101, 946 (1912). 145. E. Schmidt, Arch. Pharm. (Weinheim, Ger.) 230, 207 (1892). 146. K. Ishimaru and K. Shimomura, Phytochemistry 28, 3507 (1989). 147. T. Ikenaga, M. Abe, A. Itakura, and H. Ohashi, Planta Med. 35, 51 (1979). 148. W. J. Griffin and G. D. Lin, Naturwissenschaften 76, 582 (1989). 149. E. Knopp, A. Strauss, and W. Wehrli, Plant Cell Rep. 7, 590 (1988). 150. W. J. Griffin, Naturwissenschaften 62, 97 (1975). 151. 0. Munoz, R. Hartmann, and E . Breitmaier, J. Nut. Prod. 54, 1094 (1991). 152. R. Hartmann, A. San-Martin, 0. Munoz, and E. Breitmaier, Angew. Chem., Int. Ed. Engl. 29, 385 (1990). 153. G. de la Fuente, M. Reina, 0. Munoz, A. San Martin, and J . P. Girault, Heterocycles 27, 1887 (1988). 154. H. Ripperger, Phytochemistry 18, 171 (1979). 155. W. C. Evans, A. Ghani, and V. A. Woolley, J. Chem. Soc., Perkin Trans. I , 2017 ( 1972). 156. K. Basey and J. G. Woolley, Phytochemistry 12, 2557 (1973). 157. H. Yamaguchi, A. Numata, and K. Hokimoto, YakugakuZasshi94,11 I5 (1974);Chem. Abstr. 82, 73255q (1975). 158. H. Yamaguchi and K. Nishimoto, Chem. Pharm. Bull. 13, 217 (1965). 158a. C. Kubwabo, B. Rollmann, and B. Tilquin, Planta Med. 59, 161 (1993). 159. A. B. Ray, M. Sahai, and P. D. Sethi, Chem. Znd. (London), 454 (1976). 160. A. B. Ray, Y. Oshima, H . Hikino, and C. Kabuto, Heterocycles 19, 1233 (1982). See also M. Sahai and A. B. Ray, J. Org. Chem. 45, 3265 (1980), and A. R. Pinder, J . Org. Chem. 47, 3607 (1982). 161. A. T. McPhail and A. R. Pinder, Tetrahedron 40, 1661 (1984). 162. K. L. Khanna, A. E. Schwarting, A. Rother, and J. M. Bobbitt, Lloydia 24, 179 (1961). 163. A. E . Schwarting, J. M. Bobbitt, A. Rother, C. K. Atal, K. L. Khanna, J. D. Leary, and W. G. Walter, Lloydia 26, 258 (1963). 164. T. Plowman, L . 0. Gyllenhaal, and J. E . Lindgren, Bot. Mus. LeaJI., H a w . Ilniv. 23, 61 (1971). 165. K. Bodendorf and H. Kummer, Pharm. Zentralhalle 101, 620 (1962); Chem. Abstr. 58, 12367f (1963). 166. W. C. Evans, A . Ghani, and V. A. Woolley, Phytochemistry 11, 469 (1972). 167. W. J. Griffin, Planta Med. 14, 468 (1966). 168. S. W. El-Dabbas and W. C. Evans, Planta Med. 44, 184 (1982). 169. I. J. Pachter and A. F. Hopkinson, J. A m . Pharm. Assoc. 49,621 (1960). 170. R. Zielinska-Sowicka and M. Szymanska, Akad. Wiss. Berlin, KI. Chem., Geol. Biol., 551 (1966); Chem. Abstr. 66, 83046f (1967). 171. T. Baytop and N. Giiner, Istanbul Univ. Eczacilik Fak. Mecm. 19, 47 (1983); Chem. Abstr. 101, 147881a (1984). 172. L. A. de Garcia, P. H. Rodriguez, and M. Martinez, Rev. Mex. Cienc. Farm. 16, 11 (1985); Chem. Abstr. 103, 138638r (1985). 173. R. Ptleger, Mitt. Dtsch. Pharm. Ges. 34,182(1964); Suppl. to [Arch.Pharm. (Weinheim, Ger.) 297, 182 (1964)l.

1. TROPANE ALKALOIDS

105

174. E. Schmidt, Arch. Pharm. (Weinheim, Ger.) 243, 303 (1905). 175. A. Kircher, Arch. Pharm. (Weinheim, Ger.) 243, 309 (1905). 176. P.-K. Hsiao, K.-C. Hsia, and L.-Y. Ho, Chih Wu Hsueh Pao (Zhiwu Xuebao) [Acta Bot. Sin. (Enel. Trans/.)]15, 187 (1973). 177. N. Mollov, H . Fernandez de Cordova, and V. L. Sanchez, Reu. Cubana Farm. 10, 139 (1976); Chem. Abstr. 87, 2396d (1977). 178. 0. E. Roses, C. M. Lopez, and J. C. Garcia Fernandez, Acta Farm. Bonaerense 6, 167 (1987); Chem. Abstr. 109, 107760~(1988). 179. W. J. Griffin, Aust. J. Chem. 29, 2329 (1976). 180. 0. E. Roses, V. E. Gambaro, and R. Rofi, Acta Farm. Eonaerense 7 , 8 5 (1988);Chem. Abstr. 110, 209372g (1989). 181. C. S. Shah and A. N. Saoji, Planta Med. 14, 465 (1966). 182. W. J. Griffin, Econ. Bot. 30,361 (1976). 183. W. C. Evans and W. J. Griffin, J. Pharm. Pharmucol 16, 337 (1964). 184. W. C. Evans and W. J. Griffin, J . Chem. Soc., 4348 (1963). 185. A. J. Parr, J. Payne, J. Eagles, B. T. Chapman, R. J. Robins, and M. J. C. Rhodes, Phytochemistry 29, 2545 (1990). 186. W. C. Evans and V. A. Major, J. Chem. Soc. C, 1621 (1966). 187. W. C. Evans and V. A. Major, J. Chem. Soc. C, 2775 (1968). 188. W. C. Evans, V. A. Major, and M. Pe Than, Planta Med. 13, 353 (1965). 189. W. C. Evans and V. A. Woolley, Phytochemistry 17, 171 (1978). 190. C. F. Moorhoff, Planta Med. 28, 106 (1975). 191. W. C. Evans and J. F. Lampard, Phytochemistry 11, 3293 (1972). 192. A. Romeike, Naturwissenschaften 49, 426 (1962). 193. S. I. Balbaa, A. H . Saber, M. S. Karawya, and G. A. El-Hossary, J. Pharm. Sci. U.A.R. 10, 125 (1969); Chem. Abstr. 73, 127727e (1970). 194. W. C. Evans and A. 0. Somanabandhu, Phytochemistry 13, 304 (1974). 195. A. H. Saber, S. 1. Balbaa, G. A. El Hossary, and M. S. Karawya, Lbydia 33, 401 (1970). 196. P. H. Kalemkiarian and 0. H. Miller, J. A m . Pharm. Assoc. 46, 393 (1957). 197. W. C. Evans and J. G. Woolley, J. Chem. Soc., 4936 (1965). 198. C. H. Shah and P. N. Khanna, J. Pharm. Pharmacoll7, I I5 (1965). 199. L. Witte, K. Mueller, and H.-A. Arfmann, Planla Med. 53, 192 (1987). 200. G. Petri, in “Cell Culture and Somatic Cell Genetics of Plants” (F. Constabel and I. K. Vasil, eds.), Vol. 5 , p. 263. Academic Press, San Diego, 1988. 201. S. F. Aripova and S. N . Yunusov, Khim. Prir. Soedin., 36(1989) [Chem. Natl. Compd. (Engl. Trans/.)25, 30 (1989)l. 202. A. Romeike and 0. Aurich, Phytochemistry 7, 1547 (1968). 203. A. Romeike, Pharmazie 8, 729 (1953). 204. J. M. Pelt, C. Younos, and J . C. Hayon, Ann. Pharm. Fr. 25, 101 (1967). 205. F. L. Pyman and W. C. Reynolds, J. Chem. Soc. 93, 2077 (1908). 206. F. J. Al-Muhtadi, M. M. A. Hassan, and 0. A. Aziz, Bu/l. Fac. Sci., Riyad Univ.5, 1 (1973); Chem. Abstr. 81, 101848k (1974). 207. F. G. ACMuhtadi, Bull. Fac. Sci., Riyad Uniu. 7, 113 (1975);Chem. Abstr. 85, 106689b (1976). 208. W. C. Evans and M. W. Partridge, Nature (London) 171, 656 (1953). 209. W. C. Evans, N. A. Stevenson, and R. F. Timoney, Planta Med. 17, 120 (1969). 210. N. A. Trofimova, I. F. Gusev, and N . V. Doshchinskaya, Rastit. Resur. 9, 168 (1973); Chem. Abstr. 79, 15797~(1973). 211. E. Mechler and H . W. Kohlenbach, Planta Med. 33, 350 (1978).

106

MAURl LOUNASMAA A N D TARJA TAMMINEN

212. F. Pagani, Boll. Chim. Farm. 121, 230 (1982). 213. K.-H. Plank and K. G. Wagner, Z. Naturforsch. C: Biosci. 41C, 391 (1986). 214. K. Shimomura, M. Sauerwein, and K. Ishimaru, Phytochemistry 30, 2275 (1991). 215. W. C. Evans and M. Wellendorf, J. Chem. SOC., 1991 (1958). 216. G. Verzar-Petri and S. Sarkany, Acta Pharm. Hung. 31, 22 (1961); Chern. Abstr. 55, 9788a (1961). 217. 1. Chandrasekharan, V. A. Amalraj, H. A. Khan, and A. Ghanim, Trans. Indian Soc. Desert Technol. Uniu. Cent. Desert Stud. 9, 23 (1984); Chem. Abstr. 102, 1637358 (1985). 218. W. C. Evans and N. A. Stevenson, J. Pharm. Pharmacol. 14, 107T (19621. 219. W. C. Evans and P. G. Treagust, Phytochemistry 12, 2077 (1973). 220. C. S. Shah and P. N. Khanna. Lloydia 28, 71 (1965). 221. F. Bi, Z. Kapadia, W. Qureshi, and Y. Bader, Pak. J . Sci. Ind. Res. 25, 66 (1982); Chem. Abstr. 98, 68887y (1983). 222. S. Siddiqui, N. Sultana, S. S. Ahmed, and S. I. Haider, J. N o t . Prod. 49, 511 (1986). 223. A. E. Andrews, J . Chem. Soc. 99, 1871 (1911). 224. A. Somanabandhu and N . Suntorncharoenon, Varasarn Paesachasarthara (Warasan Phesatchasat.) 7, 53 (1980); Chem. Abstr. 95, 39126g (1981). 225. E. Schmidt, Arch. Pharm. (Weinheim, G e r . )248, 641 (1910). 226. C. S. Shah and P. N. Khanna, Ind. J. Pharm. 25, 370 (1963). 227. C. S. Shah and P. N . Khanna, Ind. J . Pharm. 26, 140 (1964). 228. P. Tantivatana, R. Bavovada, and V. Jirawongse, J. Natl. Res. Counc. Thailand 10, 77 (1978); Chem. Abstr. 91, 305206d (1979). 229. E. Gmelin, Arch. Pharm. (Weinheim, Ger.) 279, I12 (1941). 230. K. Anwar and A. Ghani, Bangladesh Pharm. J . 2, 25 (1973); Chem. Absrr. 80,57429~ (1974). 231. Y. Yamada and T. Hashimoto, Plant Cell Rep. 1, 101 (1982). 232. 0. Hesse, Liebigs Ann. Chem. 303, 149 (1898). 233. 0. Hesse, Liebigs Ann. Chem. 309, 75 (1899). 234. V. Spurna, M. Sovova, E. Jirmanova, and A. Sustackova, Ptunta Med. 41,366 (1981). 235. R. Zielinska Sowicka and K. Szepczynska, Diss. Pharm. Pharmacol. 20, 539 (1968); Chem. Abstr. 70, 1 7 5 4 2 ~(1969). 236. A. H. Saber, S. I. Balbaa, and A. Youssef, Pak. J. Sci. Ind. Res. 4,254 (1961);Chem. Abstr. 57, 14184h (1962). 237. N . C. Saraiva de Siqueira, G. A. Brasil e Silva de Assis, B. Santana, M. Schmitt, A. C. Ballve, and L . Bauer, Trib. Farm. 47, 71 (1979); Chem. Abstr. 92, 211826m (1980). 238. A. Romeike, Flora 143, 67 (1956). 239. A. Romeike and R. Zimmermann, Naturwissenschafien 45, 187 (1958). 240. A. Romeike. Naturwissenschafren 49, 281 (1962). 241. W. C. Evans and M. W. Partridge, J . Pharm. Pharmacol. 1, 593 (1949). 242. A. Romeike, Naturwissenschaften 46, 492 (1959). 243. 1. Lalezari, Pazhoohandeh 16, 118 (1977); Chem. Abstr. 88, 71501s (1978). 244. W. Schutte, Arch. Pharm. (Weinheim, G e r . ) 229,492 (1891). 245. M. C. R. Belda and A. B. Rocha, R e v . Far. Farm. Odontol. Araraquara 6, 11 (1972); Chem. Abstr. 79, 75839v (1973). 246. Y. D. Sadykov, M. Khodzhimatov, K. Khafizov, and Y. M. N. Begovatov, Dokl. Akad. Nauk Tadzh. S S R 21, 34 (1978); Chem. Abstr. 90, 51420~(1979). 247. W. R. Dunstan and H. Brown, J . Chem. SOC. 79,71 (1901). 248. J. D. Phillipson and S. S. Handa, Phytochemistry 14,999 (1975).

1.

TROPANE ALKALOIDS

107

W. C. Evans and M. W. Partridge, J. Chem. Soc., 1102 (1957). W. C. Evans and M. W. Partridge, J. Pharm. Pharmacol. 21, 126 (1948). W. C. Evans and M. W. Partridge, J. Pharm. Pharmacol. 5, 772 (1953). L. Cosson, P. Chouard, and R. Paris, Lloydia 29, 19 (1966). L. Cosson, Plant. Med. Phytother. 2, 269 (1968); Chem. Abstr. 70, 75136~(1969). 1. Lubis, Ann. Bogor. 4, 163 (1967); Chem. Abstr. 69, 8 4 1 8 2 ~(1968). A. Romeike, Kulturpjanze 14, 129 (1966). A. Romeike, Naturwissenschafren 47, 64 (1960). A. Romeike, Planta Med. 6, 426 (1958). M. Al-Yahya and W. C. Evans, J . Pharm. Pharmacol. 27 (Suppl.), 87P (1975). P. Christen, M. F. Roberts, J. D. Phillipson, and W. C. Evans, Plant Cell Rep. 9, 101 ( 1990). 260. R. J. Robins, A. J. Parr, J. Payne, N. J. Walton, and M. J. C. Rhodes, Planta 181, 414 (1990). 261. 3. M. Petrie, Proc. Linn. Soc. N . S . W . 32, 789 (1907). 262. J. M. Petne, Proc. Linn. SOC. N . S . W . 41, 815 (1916). 263. A. Romeike, Naturwissenschaften 53, 82 (1966). 264. J. Wilms, E. Roeder, and H . Kating, Planta Med. 31, 249 (1977). 265. T. Hartmann, L. Witte, F. Oprach, and G. Toppel, Planta Med. 52, 390 (1986). 266. 0. Hesse, Liebigs Ann. Chem. 261, 87 (1891). 267. E. Steinegger and G. Phokas, Pharm. Acta Helv. 31, 330 (1956). 268. G. Clair, D. Drapier-Laprade, and R.-R. Paris, C. R. Hebd. Seances Acad. Sci., Ser. D 283, 503 (1976). 269. G. Phokas and E. Steinegger, Pharmazie 11,652 (1956). 270. P. Reinouts van Haga, Nature (London) 173, 692 (1954). 271. D. Tepfer, A. Goldmann. N . Pamboukdjian, M. Maille. A. Lepingle, D. Chevalier, J. Dtnarie, and C. Rosenberg, J . Bacteriol. 170, 1153 (1988). 272. A. Goldmann, M.-L. Milat, P.-L. Ducrot, J.-Y. Lallemand, M. Maille, A. Lepingle, I. Charpin, and D. Tepfer, Phytochemistry 29, 2125 (1990). 273. P.-H. Ducrot and J. Y. Lallemand, Tetrahedron Lett. 31, 3879 (1990). 274. W. Kussner, Arch. Pharm. (Weinheim, Ger.) 276, 617 (1938). 275. A. Baytop and M. Tanker, Istanbul Uniu. Tip Fak. Mecm. 25,259 (1962);Chem. Abstr. 61, 6044e (1964). 276. A. Ghani, W. C. Evans, and V. A. Woolley, Bangladesh Pharm. J . 1, 12 (1972). 277. M. Sauerwein, K. Ishimaru, and K. Shimomura, Phytochemistry 30, 2977 (1991). 278. U. Mahmood, Y. N. Shukla, and R. S. Thakur, Phytochemistry 24, 1618 (1985). 279. A. Saint-Firmin and R. R. Paris, C . R. Acad. Sci. Paris, Ser. D 267, 1448 (1968). 280. W. R. Dunstan and H. Brown, J . Chem. SOC., Trans. 75, 72 (1899). 281. G. K. Dhoot and G. G. Henshaw, Ann. Bot. (London) 41, 943 (1977). 282. T. Hashimoto and Y. Yamada, Planta Med. 47, 195 (1983). 283. M. Jaziri, M. Legros, J. Homes, and M. Vanhaelen. Phytochemistry 27, 419 (1988). 284. N. 1. Telezhko, Akrual. Vopr. Farm. 2, 45 (1974); Chem. Abstr. 84, 102349~(1976). 285. S. F. Aripova, Khim. Prir. Soedin., 274 (1985) [Chem. Nut. Compd. (Engl. Transl.) 21, 261 (1985)l. 286. H. Golgolab, Pazhoohandeh 23, 35 (1979); Chem. Abstr. 92, 143255~(1980). 287. R. A. Konowalowa and 0. J. Magidson, Arch. Pharm. (Weinheim, Ger.) 266, 449 (1928). 288. F. B. Ahrens, Ber. Dtsch. Chem. Ges. 22, 2159 (1889). 289. H. Thorns and M. Wentzel, Ber. Dtsch. Chem. Ges. 34, 1023 (1901). 290. R. T. Mirzamatov and K. L. Lutfullin. Khim. Prir. Soedin., 128 (1985) [Chem. Nut. 249. 250. 251. 252. 253. 254. 255. 256. 257. 258. 259.

108

291. 292. 293. 294. 295. 296. 297. 298. 299. 300. 301. 302. 303. 304. 305. 306. 307. 308. 309.

310. 311. 312. 313. 314. 315. 316. 317. 318.

MAURl LOUNASMAA A N D TARJA TAMMINEN

Compd. (Engl. Transl.) 21, 129 (19831. See also R. T. Mirzamatov, K. L. Lutfullin, V. M. Malikov, and S. Y. Yunusov, Khim. Prir. Soedin., 680 (1973) [Chem. Nar. Compd. (Engl. Transl.) 9, 655 (197311. R. T. Mirzamatov, K. L . Lutfullin, V. M. Malikov, and S. Y. Yunusov, Khim. Prir. Soedin., 415 (1974) [Chem. Nut. Compd. (Engl. Transl.) 10, 426 (1974)l. R. T. Mirzamatov, K. L . Lutfullin, V. M. Malikov, and S. Y. Yunusov, Khim. Prir. Soedin., 416 (1974) [Chem. Nut. Compd. (Engl. Trans.) 10, 427 (1974)l. R. T. Mirzamatov, K. L. Lutfullin, V. M. Malikov, and S. Y. Yunusov, Khim. Prir. Soedin., 540 (1974) [Chem. Nut. Compd. (Engl. Transl.) 10, 558 (1974)l. R. T . Mirzamatov, V. M. Malikov, K. L . Lutfullin, and S. Y. Yunusov, Khim. Prir. Soedin., 493 (1972) [Chem. Nar. Compd. (Engl. Transl.) 8, 486 (1972). Z. Chen, X. Chang, G. Qin, B. Wang, and R. Xu, Zhongcaoyao 12, 1 (1981): Chem. Abstr. 96, 11552q (1982). S . M. Aslanov, Rasrit. Resur. 19, 65 (1983): Chem. Abstr. 98, 104344e (1983). Y.-L. Liu and F.-Z. Xie, Yao Hsueh Hsueh Pao 14, 497 (1979); Chem. Abstr. 92, 72723k (1980). L . Chen and D. Si, Yaoxue Xuebao 19, 869 (1984); Chem. Ahstr. 102, 757332 (1985). X. Yang and J. Yang, Zhongcaoyao 12,534,553 (1981); Chem. Absrr. 97,3550t (1982). K. Cheng and C. Liang, Plant Tissue Culr., [Proc. Symp.], 469. (1981); Chem. Abstr. 98, 122833t (1983). M. Szymanska, Herba Pol. 32, 161 (1986); Chem. Abstr. 109, 70422g (1988). 1. Bendik, 0. Bauerova, S. Bauer, J. Mokry, and J. Tomko, Chem. Zuesti 12, 181 (1958); Chem. Abstr. 52, 12324e (1958). S. W. Zito and J. D. Leary, J . Pharm. Sci. 55, 1150 (1966). C. Siebert, Arch. Pharm. (Weinheim, Ger.) 228, 139 (1890). R. Nikolic, M. Gorunovic, and P. Lukic, Acra Pharm. Jugosl. 26, 257 (1976); Chem. Abstr. 86, 13807~(1977). 0. T. Sluka and G. Y. Genig, Farm. Zh. (Kiev), 85 (1978): Chem. Absrr. 90,69147d ( 1979). G. M. Ulicheva, Rastir. Resur. 7, 18 (1971); Chern. Absrr. 74, 108126n (1972). E. S. Gritsaeva, Tr. I [Peruogo] Mosk. Med. Insr. 61, 329 (1968); Chem. Abstr. 71, 98981j (1969). L. N . Bereznegovskaya and A. V. Smorodin, Usp. Izuch. Lek. Rust. Sib., Marer. Mezhuuz. Nauchn. Konf., 50-1 (1973); [(L. N. Bereznegovskaya, ed.), Tomsk. Univ., Tomsk, USSR, 19731; Chem. Abstr. 81, 166562g (1974). M. Tabata, H. Yamamoto, N . Hiraoka, A. Oka, K. Kawashima, and M. Konoshima, Shoyakugaku Zasshi 23, 83 (1969); Chem. Abstr. 73, 73844v (1970). E. Schmidt and H . Henschke, Arch. Pharm. (Weinheim, Cer.) 226, 185 (1888). Y. Mano, S. Nabeshima, C. Matsui, and H. Ohkawa, Agric. Biol. Chem. 50, 2715 (1986). M. Konosima, M. Tabata, N. Hiraoka, and H. Miyake, Shoyakugaku Zasshi 21, 108 (1967); Chem. Abstr. 69, 65150t (1968). C. Siebert, Arch. Pharm. (Weinheim, Ger.) 228, 145 (1890). M. Szymanska, Acra Pol. Pharm. (Engl. Transl.) 23, 319 (1966); Chem. Absrr. 66, 17097~(1967). M. Gorunovic, N. Prum, and J. Raynaud, Plant. Med. Phytother. 4,286 (1970);Chem. Abstr. 74, 108125m (1971). V. Jovanovic, D. Grubisic, Z. Giba, N. Menkovic, and M. Ristic, Planta Med. 57 (Suppl. 2), A102 (1991). S . A. Minina and L . P. Mashkova, Rastit. Resur. 12, 546 (1976): Chem. Abstr. 86, 13948r (1977).

1.

TROPANE ALKALOIDS

109

319. R. Zielinska-Sowicka and J. Gudej, Ann. Acad. Med. Lodz. 12, 411 (1971); Chem. Abstr. 78, 156651j (1973). 320. M. Szymanska, Pol. J . Pharmacol. Pharm. 25, 201 (1973); Chem. Abstr. 79, 102854e (1973). 321. I. A. Barene and S. A. Minina, Khim. Prir. Soedin., 379 (1971) [Chem. Nut. Compd. (Engl. Trans/.) 7, 360 (1971)l. 322. B. A. Samoryadov and S. A. Minina, Khim. Prir. Soedin., 209 (1971) [Chem. Narl. Compd. (Engl. Transl.) 7, 205 (1971)l. 323. S. A. Minina and I. Barene, Biol. Akt. Veshchesrva Flory Fauny Dal’n. Vosr. Tikhogo Okeana 22-3. From Ref. Zh., Biol. Khim. 1972, Abstr. No. 7F1052 1971; Chem. Abstr. 77,111461k (1972). 324. S. A. Minina, T. V. Astakhova, and N. V. Nazarova, Rasrit. Resur. 11, 493 (1975); Chem. Abstr. 84, 5648Ij (1976). 325. S. A. Minina, T. V. Astakhova, and D. A. Fesenko, Khim. Prir. Soedin., 712 (1977) [Chem. Nut. Compd. (Engl. Trans/.) 13, 598 (1977)l. 326. I. B. Sandina, Rastir. Resur. 8, 524 (1972); Chem. Abstr. 78, 82095g (1973). 327. M.G. Abutalybov, S. M. Aslanov, and E. N. Novruzov, U.S.S.R. SU 639888. From Otkrytiya, Izobret., Prom. Obraztsy, Tovarnye Znaki 55, 90 (1978); Chem. Absrr. 90, 100395j (1979). 328. S. F. Aripova, V. M. Malikov, and S. Y. Yunusov, Khim. Prir. Soedin., 290 (1977) [Chem. Nut. Compd. (Engl. Transl.) 13, 255 (1977)l. 329. S. F. Aripova and S. Y. Yunusov, Khim. Prir. Soedin., 527 (1979) [Chem. Nut. Compd. (Engl. Trans/.) 15, 457 (1979)l. 330. S. F. Aripova, E. G. Sharova, U. A. Abdullaev, and S. Y. Yunusov, Khim. Prir. Soedin., 749 (1983) [Chem. Nut. Compd. (Engl. Trans/.) 19, 712 (1983)l. 331. S. F. Aripova, Khim. Prir. Soedin., 275 (1985) [Chem. Nut. Compd. (Engl. Trans/.) 21, 261 (1985)l. 332. I. Israilov, K. A. Abduazimov, and S. Y. Yunusov, Dokl. Akad. Nauk Uzbek. S.S.R. 22, 18 (1965); Chem. Abstr. 63, 7346e (1965). 333. A. Orechoff and R. A. Konowalowa, Arch. Pharm. (Weinheim, Ger.) 271, 145 (1933). 334. A. Orechoff and R. A. Konowalowa, Ber. Dtsch. Chem. Ges. 68, 814 (1935). 335. A. Orechoff and R. A. Konowalowa, Ber. Dtsch. Chem. Ges. 67, 1153 (1934). 336. E. G. Sharova, S. F. Aripova, and S. Y. Yunusov, Khim. Prir. Soedin., 672 (1980); Chem. Absrr. 94, 117787~(1981). 337. S. F. Aripova and S. Y. Yunusov, Khim. Prir. Soedin., 618 (1986) [Chem. Nut. Compd. (Engl. Trans/.) 22, 581 (1986)l. 338. S. Y. Yunusov, T. T. Shakirov, and N. V. Plekhanova, Dokl. Akad. Nauk Uzbek. SSR, 17 (1958); Chem. Abstr. 53, 18391g (1959). 339. S. F. Aripova, V. M. Malikov, and S. Y. Yunusov, Khim. Prir. Soedin., 401 (1972) [Chem. Nut. Compd. (Engl. Trans/.) 8, 403 (1972)l. 340. S. F. Aripovaand S. Y. Yunusov, Khim. Prir. Soedin., 657 (1986)[Chem. Nut/. Compd. (Engl. Trans/.) 22, 622 (1986)l. 341. S. F. Aripova, E. G. Sharova, and S. Y. Yunusov, Khim. Prir. Soedin., 640 (1982) [Chem. Nut. Compd. (Engl. Trans/.) 18, 606 (1982)l. 342. Y. Lu, T. Yao, and Z. Chen, Yaoxue Xuebao 21,829 (1986); Chem. Abstr. 106,153028~ ( 1987). 343. P. Wang, T. Yao, and Z. Chen, Zhiwu Xuebao 31, 616 (1989); Chem. Absfr. 113, 3769911( 1990). 344. Z. Chen, P. Xu, and T. Yao, Zhongcaoyao 17,386 (1986); Chem. Absfr. 106, 153016s (1987).

110

MAURI LOUNASMAA A N D TARJA TAMMINEN

345. L. C. Fonseca and G. A. Salive, Rev. Colomb. Cienc. Quim.-Farm.2,27 (1972); Chem. Abstr. 79, 1 5 7 9 8 ~(1973). 346. E. G. Sharova, S. F. Aripova, and 0. A. Abdilalimov, Khim. Prir. Soedin., 126 (1977) [Chem. Nut. Compd. (Engl. Trans/.) 13, 117 (1977)l. 347. J. A. Lamberton, Y. A. G. P. Gunawardana, and I. R. C. Bick, J. Nut. Prod. 46, 235 (1983). See also D. J . Collins, C. C. J. Culvenor, J . A. Lamberton, J . W. Loder, and J. R. Price, “Plants for Medicines,” p. 93. CSIRO Publications, Melbourne, 1990. 348. S. I. Balbaa, A. Y. Zaky, M. Y. Haggag, and M. El-Azizi, Bull. Fac. Pharm., Cairo Uniu. 14, 231 (1976); Chem. Abstr. 87, 164210r (1977). 349. F.-C. Czygan, B. Wessinger, and K. Warmuth, Biochem. Physiol. Pfanz. 183, 495 ( 1988). 350. H. E. Schink, H. Pettersson, and J.-E. Backvall, J . Org. Chem. 56, 2769 (1991). See also J. E . Backvall, Z. D. Renko, and S. E. Bystrom, Tetrahedron Lett. 28, 4199 (1987). 351. A . Bathgate and J. R. Malpass, Tetrahedron Lett. 28, 5937 (1987). See also A. Naylor (nee Bathgate), N. Howarth, and J . R. Malpass, Tetrahedron 49, 451 (1993). 352. H. M. L. Davies, E. Saikali, and W. B. Young, J. Org. Chem. 56, 5696 (1991). See also H. M. L. Davies, W. B. Young, and H. D. Smith, Tetrahedron Lett. 30, 4653 (1989). and H. M. L. Davies and N . J. S. Huby, Tetrahedron Lett. 33, 6935 (1992). 353. E. Leete and S. H. Kim, 1. Chern. Soc., Chem. Comnzun., 1899 (1989). 354. P. T. Lansbury, C. J . Spagnuolo, B. Zhi, and E. L. Grimm, Tetrahedron Lett. 31, 3965 (1990). 355. R. M. Moriarty, 0. Prakash, P. R. Vavilikolanu, R. K. Vaid. and W. A. Freeman, J. Org. Chem. 54, 4008 (1989). 356. P.-H. Ducrot, J . Beauhaire, and J . Y. Lallemand, Tetrahedron Lett. 31, 3883 (1990). See also 0. Duclos, M. Mondange, A. DurCault, and J . C. Depezay. Tetrahedron Lett. 33,8061 (1992); F.-D. Boyer, P.-H. Ducrot, V. Henryon, J. Soulie, and J.-Y. Lallemand, Synlerr., 357 (1992); F.-D. Boyer and J . - Y . Lallemand, Synlett., 969 (1992). 357. S. Furuya and T. Okamoto, Heterocycles 27, 2609 (1988). 358. P. M. Esch, H. Hiemstra, W. J. Klaver, and W . N . Speckamp, Heterocycles 26, 75 (1987). 359. Z. Xiang, J . E. Zhou, Z. N. Chen, L. P. Wang, H. N. Wang, T. R. Yao. J. X. Xie, G. Y. Xu, and D. N . Yi, Yaoxue Xuebuo 24, 105 (1989); Chem. Abstr. 112, 198846~ ( 1990). 360. M. E. Jung, L.-M. Zeng, T.-S. Peng, H.-Y. Zeng, Y. Le, and J . - G . Su, J. Org. Chern. 57, 3528 (1992). 361. X. S. He and A. Brossi, J. Heterocycl. Chem. 28, 1741 (1991). See also H. J . Yang and X. J. Xie, Yaoxue Xuebao 26, 948 (1991); Chem. Abstr. 117, 131412t (1992). 362. J . Mann and L.-C. de Almeida Barbosa, J. Chem. Soc., Perkin Trans. 1 , 787 (1992). 363. M. R. Brzezinski, C. D. Christian, M.-F. Lin, R. A. Dean, W. F. Bosron, and E. T. Harper, Synth. Commun. 22, 1027 (1992). 364. N. J . Sisti, F. W. Fowler, and J . S. Fowler, Tetrahedron Lett. 30, 5977 (1987). See also M. Novak, J. Novak, and C. A. Salemink, Tetrahedron Lett. 32, 4405 (1991). 365. J . Santamaria, R. Ouchabane, and J. Rigaudy, Tetrahedron Lett. 30, 2927 (1989). 366. J. Santamaria, R. Ouchabane, and J. Rigaudy, Tetrahedron Lett. 30, 3977 (1989). 367. J. Santamaria, M. T. Kaddachi, and J . Rigaudy, Tetrahedron Lett. 31, 4735 (1990). 368. E. Leete, Planta Med. 56, 339 (1990). and references therein. 369. R. B. Herbert, “The Biosynthesis of Secondary Metabolites,” 2nd Ed., p. 133. Chapman & Hall, London, 1989. 370. A. Ahmed and E. Leete, Phytochemisrry 9, 2345 (1970).

1.

TROPANE ALKALOIDS

111

371. F. E. Baralle and E. G. Gros, Chem. Commun., 721 (1969). 372. E. Leete, Planta Med. 36, 97 (1979). 373. H. W. Liebisch and H. R. Schiitte, in “Biochemistry of Alkaloids” (K. Mothes, H. R. Schiitte, and M. Luckner, eds.), p. 108. VEB Deutscher Verlagder Wissenschaften, Berlin, 1985. 374. E. Leete, J. A m . Chem. Soc. 84, 55 (1962). 375. E. Leete, Tetrahedron Lett., 1619 (1964). 376. H. W. Liebisch, H . Radwan, I. Schoffinius, and H . R. Schutte, Z . Naturforsch. B: Anorg. Chem. Org. Chem. Biochem. Biophys. Biol. 20B, 1183 (1965). 377. E. Leete, T. Endo, and Y. Yamada, Phytochemistry 29, 1847 (1990). 378. T. Hashimotq, Y. Yamada, and E. Leete, J . A m . Chem. Soc. 111, 1141 (1989). See also, U. Sankawa, H . Noguchi, T. Hashimoto, and Y. Yamada. Chem. Pharm. Bull. 38, 2066 (1990). 379. E. Leete, J. A m . Chem. Sac. 104, 1403 (1982). 380. E. Leete, Phytochemistry 24, 953 (1985). 381. E. Leete and S. H. Kim, J. A m . Chem. Soc. 110, 2976 (1988). 382. E. Leete, Heterocycles 28, 481 (1989). 383. E. Leete, J. A. Bjorklund. M. M. Couladis, and S . H. Kim, J. Am. Chem. Soc. 113, 9286 (1991). 384. E. Leete, S. H. Kim, and J . Rana. Phytochemistry 27, 401 (1988). See also E. Leete. J. A. Bjorklund, and S . H. Kim, Phytochemistry 27, 2553 (1988). and R. B. Herbert, N u t . Prod. Rep. 7, 105 (1990). 385. T. Hashimoto and Y. Yamada, Agric. B i d . Chem. 53, 863 (1989). 386. Y. Yamada and T. Hashimoto, Proc. J p n . Acad. Ser. B 65, 156 (1989). 387. K. Cheng, H. Fang, W. Zhu, C. Mang, D. Yang. and L. Li. Planta Med. 55, 391 (1989). 388. Y. Yamada, T. Hashimoto, T. Endo, Y. Yukimune, J . Kohno, N. Hamaguchi, and B. Drager, in “Secondary Products from Plant Tissue Culture” (B. V. Charlwood and M. J. C. Rhodes, eds.), p. 227. Oxford Univ. Press (Clarendon), Oxford, 1990. 389. T. Hashimoto, J. Matsuda, S . Okabe, Y. Amano, D. J . Yun, A . Hayashi, and Y. Yamada, in “Progress in Plant Cellular and Molecular Biology” (H. J. J. Nijkamp. L. H. W. van der Plas, and J. van Aartrijk, eds.). p. 775. Kluwer Academic Publ., Dordrecht, The Netherlands, 1990. 390. G. Wang, H. Li. L. Shen, R. Quian, and F. Huang, Bopuxite Zuchi 4, 247 (1987); Chem. Abstr. 109, 38009s (1988). 391. M. R. Yagudaev and S. F. Aripova, Khim. Prir. Soedin., 80 (1986) [Chem. N o t . Compd. (Engl. Transl.) 22, 74 (1986)l; Chem. Ahstr. 105, 24479a (1986). 392. R. Glaser, Q.-J. Peng, and A. S. Perlin, J . Org. Chem. 53,2172 (1988). See also R. Glaser, J.-P. Charland, and A. Michel, J . Chem. Soc. Perkin Trans. 2 , 1875 (1989). 393. P. Scheiber and K. Nador, Magn. Res. Chem. 27, 1097 (1989). 394. C. Sarazin, G. Goethals, J.-P. Seguin, and J.-N. Barbotin, Magn. Res. Chem. 29, 291 ( 1991). 395. S. F. Aripovaand S . Y. Yunusov, Khim. Prir. Soedin., 447 (1991) [Chrm. Nat. Compd. (Engl. Transl.) 27, 389 (1991)l. 396. S . F. Aripova and B. Tashkhodzhaev, Khim. Prir. Soedin., 532 (1991) [Chern. Nat. Compd. (Engl. Transl.) 27, 464 (1991)l. 397. M. G. Ikonomou, A. T . Blades, and P. Kegarle, Anal. Chem. 62, 957 (1990). 398. S. P. Jindal and T. Lutz, J . Pharm. Sci. 78, 1009 (1989). 399. W.-X. Hu, L.-H. Yun, and S . 3 . Li, Chin. Chem. Lett. 3, 271 (1992); Chem. Abstr. 117, 171767~(1992).

I12

MAURI LOUNASMAA A N D TARJA TAMMINEN

400. X.-Y.Niu and Z.-H. Ren, Yaoxue Xuebao 19, 326 (1984); Chem. Abstr. 103, 47831 (1985). 401. R. C. Gupta and W. L . Kadel, J. Toxicol. Enuiron. Health 28, 1 1 1 (1989). 402. E. A. Koller, S. Drechsel, T. Hess, P. Macherel, and U. Boutellier, Eur. J. Appl. Physiol., Occup. Physiol. 57, 163 (1988). 403. F., Weise, K . Baltrusch, and F. Heydenreich, J. Aufon. Neru. Syst. 26, 223 (1989). 404. C. D. Kay and J . D. Morrison, J. Exp. Physiol. 73, 511 (1988). 405. S. M. Pellis, V. C. Pellis, R. M. Chesire, N. Rowland, and P. Teitelbaum, Proc. Natl. Acad. Sci. U . S . A . 84, 8750 (1987). 406. L. Molinengo, A. Fundaro, and M. Orsetti, Pharmacol. Biochem. Behau. 32, 1075 (1989). 407. L . Walters, P. Bartel, De K. Sommers, and P. Becker, Methods Find. Exp. Clin. Pharmacol. 10, 419 (1988). 408. M. U. Rashid and D. N. Bateman, Br. J. Clin. Pharmacol. 30, 25 (1990). 409. M. Kung and L. Diamond, Pulm. Pharmacol. 3, 17 (1990). 410. M. Durkot, 0. Martinez, V. Pease, R. Francesconi, and R. Hubbard, Auiat. Space Enuiron. Med. 61, 424 (1990). 41 I . D. M . Penetar, Drug. Deu. Res. 20, 117 (1990). 412. S. Castellani and E. H. Ellinwood, Psychopharmacology (Amsferdam)2,442 (1985). 413. M. P. Mahalik, R. F. Gautieri, and D. E. Marin, Jr., Res. Commun. Subst. Abuse 5, 279 (1984). 414. J. A. Ruth, E. A. Ullman, and A. C. Collins, Pharmacol. Biochem. Behau. 29, 157 ( 1988). 415. D. W. Ou, M. L. Shen, and D. Ying, Clin. Immunol. Immunopafhol. 52, 305 (1988). 416. I. K. Ho, Life Sci. 42, 2063 (1988). 417. F. H . Gaven, J . Clin. Psychiatry 49 (Suppl.), I I , (1988). 418. T. Inaba. Can. J. Physiol. Pharmacol. 67, 1154 (1989). 419. J. M. Lakoski and K. A. Cunningham, NIDA Res. Monogr. 88, 78 (1988). 420. K. A. Cunningham and J. M. Lakoski, Neuropsychopharmacology 3,41 (1990). 421. E. A. Loh and D. C. S. Roberts, Psychopharmacology (Berlin) 101, 262 (1990). 422. Y. Zhang and R. L. Foltz, J. Anal. Toxicol. 14, 201 (1990). 423. E.g., W. J. Crumb, Jr., and C. W. Clarksom, Biophys. J. 57, 589 (1990). 424. M. C. Ritz, R. J. Lamb, S. R. Goldberg, and M. J. Kuhar, Science 237, 1219 (1987). 425. J. Bergman, 9. K. Madras, S. E. Johnson, and R. D. Spealman, J. Pharmacol. Exp. Ther. 251, 150 (1989). 426. M. C. Ritz, E. J. Cone, and M. J. Cone, Life Sci. 46,635 (1990). 427. F. 1. Carroll, Y. Gao, M. A. Rahman, P. Abraham, K. Parham, A. Lewin, J. W. Boja, and M. J. Kuhar, J. Med. Chem. 34,2719 (1991). 428. P. Abraham, J . B. Pitner, A. H. Lewin, J. W. Boja, M. J. Kuhar, and F. I. Carroll, J . Med. Chem. 35, 141 (1992). 429. A. H. Lewin, Y. Gao, P. Abraham, J. W. Boja, M. J. Kuhar, and F. I. Carroll, J . Med. Chem. 35, 135 (1992). 430. F. I. Carroll, A. H. Lewin, J . W. Boja, and M. J. Kuhar, J . Med. Chem. 35, 969 (1992). 431. C. J. Heyser, G. A. Goodwin, C. A. Moody, and L. P. Spear, Pharmacol. Biochem. Behau. 42, 169 (1992). 432. J. Pens, M. Coleman-Hardee, and W. J. Millard, Pharmacol. Biochem. Behau. 42, 509 (1992). 433. C. E. Turner, 9. S. Urbanek, G. M. Wall, and C. W. Walker, “Cocaine, an Annotated Bibliography.” Univ. Press of Mississippi, Jackson, Mississippi, 1988.

1 . TROPANE ALKALOIDS

113

434. 1. Izquierdo, Trends Pharmacol. Sci. 10, 175 (1989). 435. G. C. Preston, C. Brezell, C. Ward, P. Brooks, M. Traub, and S. M. Stahl, J. Psychopharmacol. (Oxford)2, 67 (1988). 436. C. Brazell, G. C. Preston, C. Ward, C. R. Lines, and M. Traub, J. Psychopharmacol. (Oxford) 3, 76 (1989). 437. J . M . Rusted and D. M. Warburton, Psychopharmacology (Berlin) 96, 145 (1988). 438. J. M. Rusted, Psychopharmacdogy (Berlin) 96, 487 (1988). 439. A. Santi and S. Bridson, Pharmacol. Biochem. Behau. 39, 935 (1991). 440. C. D. Frith, M. A. McGinty, I. Gergel, and T. J. Crow, Psychopharmacology (Berlin) 98, 120 (1989). 441. D. K. Rush and K. Streit, Psychopharmacology (Berlin) 106, 375 (1992). 442. C. Flicker, M. Serby, and H. Ferris, Psychopharmacology (Berlin) 100, 243 (1990). 443. M. C. Buhot, M. Soffle, and B. Poucet, Psychobiology 17, 409 (1989). 444. M. Soffle, M. Bronchart, and Y. Lamberty, J. Psychopharmacol. (Oxford) 3, 142 (1989). 445. R. F. Genovese, T . F. Elsmore, and J. M. Witkin, Pharmacol. Biochem. Behau. 37, 117 (1990). 446. C. H. Vanderwolf, C. T. Dickson, and G. B. Baker, Pharmacol. Biochem. Behau. 35, 847 (1990). 447. M. W.Parsons and P. E. Gold, Behau. Neural Biol. 57, 90 (1992). 448. C. Flicker, S. H. Steven, and M. Serby, Psychopharmacology (Berlin)107,437 (1992). 449. S. Wada, T. Yoshimitsu, N . Koga, H. Yamada, K. Oguri, and H. Yoshimura,Xenobiorica 21, 1289 (1991). 450. Y. Kitamura, in “Biotechnology in Agriculture and Forestry” (Y. P. S. Bajaj, ed.), Vol. 4, p. 419. Springer-Verlag, Berlin and Heidelberg, 1988. 451. G.-Z. Zheng, in “Biotechnology in Agriculture and Forestry” (Y. P. S. Bajaj, ed.), Vol. 7, p. 23. Springer-Verlag, Berlin and Heidelberg, 1989. 452. G. Petri and Y. P. S. Bajaj, in “Biotechnology in Agriculture and Forestry” (Y. P. S. Bajaj, ed.), Vol. 7, p. 135. Springer-Verlag, Berlin and Heidelberg, 1989. See also G. Petri, in “Cell Culture and Somatic Cell Genetics of Plants” (F. Constabel and I. K. Vasil, eds.), Vol. 5, p. 263. Academic Press, New York, 1988. 453. A. Straws, in “Biotechnology in Agriculture and Forestry” (Y. P. S. Bajaj, ed.), Vol. 7, p. 286. Springer-Verlag. Berlin and Heidelberg, 1989. 454. Y. P. S. Bajaj and L. K. Simola, in “Biotechnology in Agriculture and Forestry” (Y. P. S. Bajaj, ed.), Vol. 15, p. 1. Springer-Verlag, Berlin and Heidelberg, 1991. 455. Y. P. S. Bajaj, in “Biotechnology in Agriculture and Forestry” (Y. P. S. Bajaj, ed.), Vol. 9, p. 1. Springer-Verlag, Berlin and Heidelberg, 1989. 456. R. Verpoorte, R . van der Heijden, W. M. van Gulik, and H. J. G. ten Hoopen. in “The Alkaloids” (A. Brossi, ed.), Vol. 40, p. 52. Academic Press, San Diego. See also M. W. Fowler, R. C. Cresswell, and A. M. Stafford, in “Bioactive Compounds from Plants” (D. J. Chadwick and J. Marsh, eds.), p. 157. Wiley, Chichester, 1990, and W. Gritsanapan and W. J. Griffin, Phyrochemistry 31, 3069 (1992). 457. Y. Yamada and T . Endo, Plant Cell Rep. 3, 186 (1984). 458. T. Hashimoto and Y. Yamada, Agric. Biol. Chem. 51, 2769 (1987). 459. L. K. Simola, R. Parviainen, A. Martinsen. A. Huhtikangas, R. Jokela. and M. Lounasmaa, Acra Chem. Scand. 43, 702 (1989). 460. L. K. Simola, R. Parviainen, A. Martinsen, A. Huhtikangas, R. Jokela, and M. Lounasrnaa. Phyrochemisrry 29, 35 I7 (1990). 461. K. Yoshimatsu, T . Hatano, M. Katayama, S. Muramo, H. Kamada, and K. Shimomura. Phytochemisrry 29, 3525 (1990).

114 462. 463. 464. 465. 466. 467. 468. 469.

470. 471. 472. 473. 474.

475. 476. 477. 478. 479. 480. 481. 482. 483. 484.

MAURI LOUNASMAA AND TARJA TAMMINEN Y. Kitamura, A. Taura, Y. Kajiya, and H. Miura, J . Plant Physiol. 140, 141 (1992). T. Hashimoto and Y. Yamada, Planfa Med. 47, 195 (1983). and references therein. T. Hashimoto, Y. Yukumune, and Y. Yamada, J. Plant Physiol. U4, 61 (1986). Y. Mano, S. Nabeshima, C. Matsui, and H . Ohkawa, Agric. B i d . Chem. 50, 2715 ( 1986). J. Payne, J. D. Hamill, R. J. Robins, and M. J. C. Rhodes, Planta Med. 53,474 (1987). T. Hashimoto, Y. Yukumune, and Y. Yamada, Planta 178, 123 (1989). T. Hashimoto, Y. Yukumune, and Y. Yamada, Planta 178, 131 (1989). Y. Yamada and T. Hashimoto, in “Progress in Plant Cellular and Molecular Biology” (H. J . J. Nijkamp, L. H. W. van der Plas, and J. van Aartrijk, eds.), p. 547. KIuwer Academic Publ., Dordrecht, The Netherlands, 1990. E. Knopp, A. Straws, and W. Wehrli, Plant Cell Rep. 7, 590 (1988). Y. Kitamura, M. Sato, and H. Miura, Phytochemistry 31, 1191 (1992). J. M. Furze, M. J. C. Rhodes, A. J. Parr, R. J. Robins, I. M. Withehead, and D. R. Threlfall, Plant Cell Rep. 10, 11 1 (1991). Y. Kitamura, Y. Sugimoto, T. Samejima, K. Hayashida, and H. Miura, Chem. Pharm. Bull. 39, 1263 (1991). M. J. C. Rhodes, R. J. Robins, J. D. Hamill, A. J. Parr, M. G. Hilton, and N. J. Walton, in “Secondary Products from Plant Tissue Culture” (B. V. Charlwood and M. J. C. Rhodes, eds.), p. 201. Oxford Univ. Press (Clarendon), Oxford, 1990. R. J. Robins, A. J. Parr, J. Payne, N . J. Walton, and M. J. C. Rhodes, Planfa 181, 414 (1990). N. J. Walton, R. J. Robins, and A. C. J. Peerless, Planta 182, 136 (1990). R. J. Robins, A. J. Pam, E. G. Bent, and M. J. C. Rhodes, Planta 183, 185 (1991). R. J. Robins, A. J . Pam, and N. J. Walton, Planta 183, 196 (1991). R. J. Robins, E. G. Bent, and M. J. C. Rhodes, Planta 185, 385 (1991). R. J. Robins, N. J . Walton, J. D. Hamill, A. J. Parr, and M. J. C. Rhodes, Planta Med. 57 (Suppl. I), S27 (1991). P. Christen, M. F. Roberts, J . D. Phillipson, and W. C. Evans, Plant Cell Rep. 8, 75 ( 1989). H. Kamada, N. Okamura, M. Satake, H. Harada, and K. Shimomura, Plant Cell Rep. 5, 239 (1986). M. Sauerwein and K. Shimomura, Phytochemistry 30, 3277 (1991). P. Christen, T. Aoki, and K. Shimomura, Plant Cell Rep. 11, 597 (1992).

-CHAPTER 2-

THE BIOSYNTHESIS OF TROPANE ALKALOIDS RICHARDJ . ROBINSA N D NICHOLAS J . WALTON Agricultural and Food Research Council Institute of Food Research Norwich Laboratory Colney, Norwich NR4 7 U A , England

I. Introduction

.......

11. Organ Tissue Cultures for Biosynthetic Studies .....................................

111.

IV.

V. VI. VII.

VIII.

IX. X. XI.

A. Hyoscyamus Species ................... B. Datura stramoniurn ..................... C. Duboisia Species ......................................................................... Formation of Putrescine A. Incorporation of Lab ............................................... B. Effect of Enzyme Inhibitors .......................................................... C. Metabolite Tunneling ................................................................... Formation of Tropinone .................................................................... A. Conversion of Putrescine to N-Methylpyrrolinium B. Conversion of N-Methylpyrrolinium to Tropinone . Formation of Tropine and Pseudotropine ................. A. Tropinone Reductase I ........ .......................................... B. Tropinone Reductase I1 ....... .......................................... Formation of Acidic Moieties of Tropeines ........................................... A. Formation of Phenyllactic and Tropic Acids ... B. Formation of Other Esterifying Acids ............................................. Formation of Tropeines ........................ A. Esters of Aliphatic Acids .............................................................. B. Hyoscyamine and Other Aromatic Esters ........................................ Metabolism of Tropeines ...... A. Conversion of Hyoscyamin ........................................ B. Further Esterifying Reactions ........................................................ Degradation and Oxidation of Tropeines .... Overall Regulation of Pathway ........................................................... A. Regulation at Biochemical Level .................................................... B. Regulation at Whole-Plant Level ................... Future Prospects .............................................................................. References ......................................................................................

119

129 130 132 132 133 134

146 147 151 154 156 158 160 160 164 164 168 169 177 180 182

DEDICATION: EDWARD LEETE While preparing this chapter, we were saddened to learn of the death of Edward Leete. It is our wish to dedicate this account of tropane alkaloid biosynthesis to him in recognition of the breadth and significance of the contributions he made over many years, both in his

I I5

THE ALKALOIDS. VOL. 44 Copyrtgh! 0 1993 by Academic Press, Inc. All rights of reproduction in any form reserved.

116

RICHARD J . ROBINS A N D NICHOLAS J . WALTON

personal work and by stimulation and critical appraisal of the work of many others, including our own. For his insight, vigor, and infectious enthusiasm, he will be greatly missed.

I. Introduction

It is our intention in this chapter to present as complete a biochemical picture of the pathway of tropane alkaloid synthesis (Scheme 1) as is presently feasible. Thus, we focus on the activities and properties, in uiuo and in uitro, of the enzymes thought to be involved in this process. Inevitably, the picture is incomplete, some parts of the pathway having been well described, while others remain more or less speculative. Our aim is to show the mechanisms by which established intermediates are converted one to another in living tissues. It is not our intention to cover in detail the evidence obtained by chemical labeling studies on which the intermediacy of the metabolites in the pathway has been based. These studies have been comprehensively reviewed ( I , 2 ) ;such data will be analyzed in this chapter only when they are of value in assessing specific biochemical evidence. Neither do we wish to cover biomimetic chemical investigations, these having been effectively covered in previous volumes in this treatise (3,4). As discussed by Lounasmaa and Tamminen in Chapter 1 of this volume, the tropane alkaloids occur widely within a subsection of the plant family Solanaceae (3,being most well known from species of Darura, Atropa, Hyoscyamus, Scopolia, and Duboisia. Tropeines (tropane esters) also occur in the families Erythroxylaceae, Convolvulaceae, Proteaceae, and Rhizophoraceae ( 4 ) ,but they remain relatively little investigated from all but the foremost of these sources. The tropane moiety forms the alkamine base of hyoscyamine, scopolamine, cocaine, and related alkaloids. The use of tropane alkaloids in medicine has a long history ( 6 ) , and at times they have been applied as an effective means of disposing of unwanted rivals. The particular importance of the major aromatic tropeines hyoscyamine and hyoscine (Scheme 2) as acetylcholine antagonists (7)has stimulated considerable interest in determining their biosynthetic pathway ( I ,2). It is only since the 1980s, however, that any serious inroads into understanding the biochemistry of their formation have been made. This advance has been considerably aided by the use of root tissue cultures (see Section 11) of tropane-producing species, notably Hyoscyamus, Datura, Atropa, and Duboisia. Root tissue cultures have provided an excellent experimental system in which the feeding of precursors may

2.

BIOSYNTHESIS OF TROPANE ALKALOIDS

ORNlTHlNE

ARGlNlNE

4 J.

\

AGMATINE

\

\

4 -

POLYAMINES

117

N-CARBAMYLPUTRESCINE

PUTRESCINE

1 \

HYDROXYCINNAMOYLPUTRESCINES

N-METHYLPUTRESCINE

PHENYLALANINE

N-METHY LPYRROLlNlUM

PHENYLPYRUVIC ACID

HYGRINE

PHENYLLACTICACID

TROPINONE

TROPIC ACID

HYOSCYAMINE

CUSCOHYGRINE

PSEUDOTROPINE

3-ACETYLTROPINE

I

w

3-ACETYLPSEUDOTROPINE 3-TIGLYLPSEUDOTROPINE

CHYDROXYHYOSCYAMINE

SCOPOLAMINE

SCHEME 1 . Schematic pathway for the fbrmation of tropanes from the amino acids arginine or ornithine and phenylalanine.

118

RICHARD 5 . ROBINS A N D NICHOLAS J . WALTON

Hyoscyamine

Hyoscine

Littorine Phenylacetyluopine SCHEME 2. Structures of the aromatic tropane esters.

be performed simply and from which enzyme extracts may readily be prepared. Much of the information detailed in this chapter results from the use of this tool. The characteristics of these cultures are described below. Several enzymes involved in hyoscine formation have now been fully or substantially purified. In one example, namely, L-hyoscyamine 6P-hydroxylase (see Section VIII,A), a gene coding for the enzyme has been isolated and is being used to improve the ability of plants to make hyoscine (see Section XI). Thus, the phase has now been reached in which the powerful techniques of molecular biology are beginning to facilitate the metabolic engineering of this pathway. A biochemical approach to the pathway can enable a distinction to be made between alternative biosynthetic routes. Thus, it has been shown that the hypothetical intermediacy of 5-N-methylornithine (8)is improbable, because the necessary enzymatic activities for the synthesis and further metabolism of this compound are apparently absent (see Section IV,A). An integrated approach using enzymatic analyses combined with

2.

BIOSYNTHESIS OF TROPANE ALKALOIDS

119

labeling studies is proving most effective in clarifying such problems. An important area of uncertainty, yet to be resolved, relates to the intermediacy of hygrine, as discussed in Section IV,B. Little is known of the pathway by which the phenylalanine-derived moiety is made. A major challenge still facing biochemists is to define the reaction or sequence of reactions by which tropine is esterified with a tropic acid equivalent to form hyoscyamine. This chapter aims to detail those areas of tropane alkaloid metabolism in which progress has been made and to highlight those parts of the pathway for which significant information is still lacking.

11. Organ Tissue Cultures for Biosynthetic Studies

Callus and suspension cultures of tropane alkaloid-producing species were established some time ago, but they were never found to accumulate any alkaloids (Table I). Only following the “rediscovery” of root organ cultures have in uitro systems been available that are active in biosynthesizing and accumulating tropane alkaloids. The ability to culture excised roots was shown many years ago (9). Success may require very carefully controlled culture conditions, particularly in relation to the balance of phytohormones in the growth medium. This approach is rather limited in the range of species that can be cultured successfully and found little application in the secondary product field. Nevertheless, it was shown in the late 1950s that excised roots of tobacco species could make alkaloids (lo),while Mitra (11) provided comparable evidence for atropine production in excised roots of Atropa belladonna. At the same time, the value of isolated roots for feeding experiments was recognized (12,13). Owing in part to the difficulty of sustaining root cultures, it was not for another 10 years, however, that the full potential of these systems came to be exploited. Their potential use for product formation was patented by Merck in 1964 ( I # ) , but this patent has now expired. In 1983, Yamada and colleagues described the production of hyoscine in redifferentiated roots of Hyoscyarnus niger (15). Since then, a number of additional species have been cultured in this way (Table I). An improved approach, pioneered simultaneously in a number of laboratories, was the introduction of root cultures made by transforming the appropriate plant species with the soil bacterium Agrobacteriurn vhizogenes (16). The transformation process involves the insertion of a small piece of DNA, carried in the bacterium on a plasmid, into the genome of the plant. This transferred DNA (T-DNA) carries a number of genes that

TABLE I PRODUCTION OF TROPANE ALKALOIDS B Y TISSUE CULTURES Alkaloid ('3 dry weight: highest reported value is given)h Type of culture"

Hyoscine

Hyoscyamine

Anisodus fanguticus

S

nd

nd

Atropa belladonna

C S

nd nd

<0.01

R R R

0.02 0.26 0.21

0.27 0.50 0.28

R TR TR TR TR

0.08 nd 0.02 0.02 0.09

TR TR

0.01

0.55 0.20 0.20 0.37 0.95 0.37 0.12

R TR TR

nd nd 0.30

Svecies

Atropa caucasica Calystegia sepum Datura candida x D . aurea

Other

Hyoscyarnine + 6p-hydroxyhyoscyarnine (10% conversion to product). 6p-Hydroxyhyoscyamine hyoscine (4% conversion to product)

-

nd

0.05 <0.01

0.47

Biotrdnsformation

3-Acetyltropine ( 1 0 . 0I ). 3-tiglyltropine (<0.01)

Refs.' i

2 3 2

Hygrine (0.07). cuscohygrine (0.14). 6p-hydroxyhyoscyamine (0.04). 1 1 other bases Cuscohygrine (0.17) Calystegin (0. I )

4 3

Cuscohygrine (0.28)

Cuscohygrine (0.2) Cuscohygrine (0.3) Apoatropine (0. IS). apohyoscine (0.17). 32 other bases

5 5 10

TR

0.57

0.11

5.11

Tropine (0.45).

3-acetoxy-6-hydroxytropine(0.14),

6-hydroxyhyoscyamine (0.16). apohyoscine (0.1 I ) . I 1 other bases Dafura chloranrha Datirra fasfuosa

Darura fastuosa var. uiolacea Datura ferox

Dafura innoxia

TR

10.01

0.24

TR

0.11 0.01

0.12 0.56

TR

0.07

0.13

TR TR

<0.01 0.04

0.23 0.85

C

TR TR

nd 0.05 0.02 0.04 0.30

nd 0.38 0.22 0.17 I .OO

TR

0.11

0.49

R

R TR

7

2 12

3.6-Ditiglyl-7-hydroxytropine(0.02). 5 other bases

7 7 12 Hygrine (0.07). tropine (0.07). 3.6-diacetyltropine (0.06). 3-tiglyl-6-hydroxytropine(0.03). (0.06). 3 .6-ditiglyl-7-hydroxytropine 6-hydroxyhyoscyamine (0.04). 4 other bases Tropine

66-Hydroxyhyoscyamine (0.04) Cuscohygrine. 3-tigloyloxytropine. 3-hydroxy-6-tigloyloxytropine 6-hydroxyhyoscyamine. 6-tigloyloxyhyo5cyamine. various other bases Hygrine (0.04). tropine (0.02). 3-acetyltropine (0.03).

-

3-acetyltropine

13 2 7 14 IS

12

TABLE I (continued) Alkaloid (% dry weight; highest reported value is given)b Species

Type of culture"

Hyoscine

Hyoscyamine

Other cuscohygrine (0.09). 3-tiglyl-6-hydroxytropine (0.02). 3-hydroxy-6-tiglyltropine (0.04). 6-hydroxyhyoscyamine (0.03). 3 other bases

-

N

N

Datura leichhardtii Datura metel Datura meteloides Datura qitercifolia

Datura rose; Datura sanguinea

Datura srramonirrm

R

nd

TR TR

<0.01

0.06

0.08

TR TR

<0.01 10.01

0.26 0.42

TR TR TR

<0.01 <0.01

0.17

0.17 0.13 0.27

C

nd 0.01 nd

nd 0.31 1.05

TR TR

0.19

Biotransformation

Refs.'

12 16 7

0.31

7 7 12 Tropine (0.02). 3-acetyltropine (0.05). 3.6-diacetyltropine (0.07). 3 other bases

Tropine (0.05). 3-tiglyltropine (0.03), 6 other bases

Hygrine (0.02). tropine (0.05). 3,6-diacetyltropine (0.021, 3.6-ditiglyl-7-hydroxytropine (0.02). 5 other bases

7 7 12

2 7 12

Darura srramonircm var. inermis

R TR TR

nd 10.01 <0.01

0.22 0.60

Darura stramonium var. taticla Darura stramonium var. stramonium Datura wrighrii

TR

0.01

0.40

7

R TR

nd <0.01

0.42 0. I9

2 7

TR

0.02

0.82

R

0.07

0.13

C Sh R R R

nd nd 0.37 1.16 0.11

nd nd 0.16 0.53 0.08

TR R

1.80 0.56

0.39

Du boisia hopwoodii Duboisia leichhardtii

0.20

0.40

Tropine (0.05). 3-acetyltropine (0.05). 3,6-diacetyltropine (0.04). apoatropine (0.08)

--

Tropinone tropine

acetyltropine. acetyltropine

Hygrine (0.27). tropine (0.07). acetyltropine ( I .26). 3-acetyl-6-hydroxytropine(0.14), 3-tiglyltropine (0.14). cuscohygrine (0.07), 3-tiglyl-6-hydroxytropine(0.14). 3.6-ditiglyl-7-hydroxytropine(0.10). 4 other bases Nicotine (0.85)

12

12

17

Hyoscyamine --* hyoscine Nicotine (0.28) Nicotine (0.47) Nicotine (0.07). apoatropine

2

7

18 18 18

17 19 20 16 (continued)

TABLE I (confinued) ~

Alkaloid (% dry weight; highest reported value is given)* Species

Duboisia mvoporoides

-

Type of culture”

Hyoscine

Hyoscyamine

C C S R

Trace nd nd 0.04

Trace nd nd 0.05

R R

0.20 0.02

0.05 0.01

TR R TR

0.15 0.18 0.25

1.42 0.30 0.21

TR C

0.11 10.01 0. I5 0.58

0.02 0.03 1.12 1.15

TR

0.40

2.00

TR TR

0.05 0.14

0.52 1.36

TR

0.46

0.54

P t 4

Dirboisia myporoides x D . leichhardrii Dirboisia hybrid Hvoscvamirs albirs

R R

Other

Biotransformation

Valtropine (tr) Tropine Tropine Nicotine (0. lo), nornicotine (0.03) Nicotine (0.15) Nicotine (0.04). anabasine (0.01) 6P-Hydroxyhyoscyamine (0.08) 6@-Hydroxyhyoscyarnine(0.12)

6P-Hydroxyhyoscyarnine (0.40). 76-hydroxyhyoscyarnine (0.23) Cuscohygrine (0.2). littorine (0.9). several other bases

--* --*

acetyltropine acetyltropine

Refs.‘

21 22 22 23 17 19

20 14 14 7 16 16.24 14

25

7 26

Littorine (0.25). b~-hydroxyhyoscyamine(0.28) 6P-Hydroxyhyoscyarnine (0.17). 7~-hydroxyhyoscyamine(0.01)

14 12

TR

0.34

0.73

Norhygrine ( 0 . 0 6 ) . hygrine (0.06). tropinone (0.10). tropine (0.04). pseudotropine (0.06). 3 other bases

Hyoscyamus aureus Hyoscyamus bohemicus

TR

<0.01

0.66

7

C

Hyoscyumus canariensis Hyoscyamus desertorum Hyoscyamus gyorffii

C

10.01 0.20 0.04 <0.01 0.14 0. I6


16 16 7 16 16 12

R TR

10.01 0.33 Present

0.01 0.33 Present

C C

<0.01 <0.01

<0.01 <0.01 1.81 0.12 0.20 0.05 0.90 0.58

Hyoscyamus muticus

R TR R TR C

R R R TR TR TR Hyoscyamus niger

C

s s

R

0.23 <0.01

0.02 10.01 <0.01

nd 0.22

0.02 <0.01 0.05 0.03

Hygrine (0.03). 4 other bases

Cuscohygrine. littorine. several other bases

Tropine (0.03). 5 other bases

16 16 25

16 27 28 16 27 7 28 12 2 2 29 16

(continued)

TABLE I (continued) Alkaloid (% dry weight; highest reported value is given)" Species

Hyoscyamus niger var. pallidus Hyoscyamus pusillus Nicandra physaloides Scopolia carniolica Scopolia japonica Scopolia parvifolia Scopolia stramonifolia

Type of culture"

Hyoscine

Hyoscyamine

R

0.50

0.46

TR

1.2s

0.08

TR TR

0.13 0.03

0.32

C R TR

10.01 0.08 nd

<0.01

S TR TR

nd <0.01 0.50

0.01 0.1s I .30

C

10.01

R TR TR

nd 0.03

Other

Biotransformation

6p-Hydroxyhyoscyamine (0.181, 7j3-hydroxyhyoscyamine (10.01 ) 66-Hydroxyhyoscyamine (0.25). 7p-hydroxyhyoscyamine (
14 14

7 7

0.05

0.20 nd

16 16 30

Hygrine (0.06). 7 other bases Unidentified M , 343

-

<0.01

0.08 0.02 0.05

Refs.'

Tropic acid hyoscine, hyosc yamine Tropine (0.02). 4 other bases

31 7 32 33 33 7 12

Scopolia fannutica

R TR

<0.01

0.02

0.01 0.05

66-Hydroxyhyoscyamine (<0.01) 6~-Hydroxyhyoscyamine(0.01

14 14

Type of culture: C, callus; S , suspension; Sh, shoot; R, root; TR. transformed root. nd, Not detected; tr, trace. Key to references: (I) C. Ke-Di, Z. Wei-Hua, L. Xin-Lan, M. Chao, S . Ming-Zai, and Y. Dan-Hua, Planta Med. 53, 211 (1987); (2) Y. Yamada and T. Hashimoto, Planf Cell R e p . 1,101 (1982); (3) T. Hartmann, L. Witte, F. Oprach, and G. Toppel, Planta Med., 390 (1986): (4) J. Lang, R. Hamilton, H. Pedersen, and C.-K. Chin, in “Plant Cell Biotechnology” (M. S. S. Pais. F. Mavituna. and J. M. Novais. eds.). Vol. H18. p. 245. Springer-Verlag, Berlin, Heidelberg. and New York, 1988: (5) G. Jung and D. Tepfer, Plant Sci. 50, 145 (1987); (6) B. Drager, personal communication (1991); (7) E. Knopp, A. Strauss, and W. Wehrli, Plant Cell R e p . 7 , 590 (1988); (8) H . Kamada, N. Okamura. M. Satuke, H. Harada, and K. Shimomura. Plant Cell R e p . 5 , 239 (1986); (9) J. M. Sharp and P. M. Doran, J. Biotechnol. 16, 171 (1990): (10) R. J. Robins, A. J. Parr. J. Payne, N. J. Walton. dnd M. J. C. Rhodes, Planta 181,414 (1990); (11) P. Christen, M. F. Roberts, J. D. Phillipson. and W. C. Evans, Plant Cell R e p . 9, 101 (1990); (12) A. J. Parr, J. Payne. J. Eagles, B. T. Chapman, R. J. Robins, and M. J. C. Rhodes, Phytochemistry 29,2545 (1990); (13) N. Hiraoka, M. Tabata, and M. Konoshima. Phytochemistry 12,795 (1973); (14) K. Shimomura. M. Sauerwein, and K. Ishimaru, Phytochemistry 30, 2275 (1991); (15) I. Ionkova, L. Witte, and A. W. Alfermann, Planta Med. 55, 229 (1989); (16) T. Hashimoto, Y. Yukimune. and Y. Yamada, J. Plant Physiol. W, 61 (1986); (17) T. Endo and Y. Yamada. Phytochemistr?; 24, 1233 (1985);(18) Y. Yamada and T. Endo, Plant Cell Rep. 3, 186 (1984); (19) Y. Kitamura, Y. Sugimoto, T. Samejima. K. Hayashida, and H. Miura. Chem. Pharm. Bull. 39, 1263 (1991);(20) Y. Mano, H. Ohkawa, and Y. Yamada, Plant Sci. 59, 191 (1989): (21) K.-J. Sipply and H. Friedrich, Planta Med. 28, 186 (1975);(22) Y. Kitamura. H. Miura, and M. Sugii, Phytochemistry 25,2541 (1986); (23) Y. Kitamura, H. Miura, and M. Sugii, Planta Med. 51,489 (1985);(24) T. Hashimoto, Y. Yukimune, and Y. Yamada, Planta 178, 131 (1989); N . I(25) K. Doerk, L. Witte, and A. W. Alfermann. Z. Naturforsch. Cc Biosci. 46C, 519 (1991); (26) M. Sauerwein and K. Shimomura, Phyrochemisfry 30, 3277 (1991); (27) K.-M. Oksman-Caldentey, H. Vuorela, A. Strausse, and R. Hiltunen, PIanta Med. 53, 349 (1987); (28) K.-M. Oksman-Caldentey, 0. Parkkinen, E. Joki, and R. Hiltunen, Planta Med. 55, 682 (1989); (29) T. Hashimoto and Y. Yamada, Agric. Biol. Chem. 51, 2769 (1987);(30) A. J. Parr, Plant Cell R e p . 11, 270 (1992); (31) H . J. Scholten, S. Batterman, and J. F. Visser. Planta Med. 55, 230 (1989); (32) Y. Mano, S . Nabeshima, C. Matsui, and H. Ohkawa, Agric. Biol. Chem. 50, 2715 (1986); (33) M. Tabata, H. Yamamoto, N . Hiraoka, and M. Konoshima, Phytochemistry 11, 949 (1972). a

-

128

RICHARD J . ROBINS A N D NICHOLAS J . WALTON

stimulate the formation of roots from the point of infection. These roots may be excised and propagated aseptically in uitro using simple, defined media. The primary advantage of this approach is that it is more easily applied to a wider range of species and provides a culture requiring less stringent cultivation conditions. It is also possible to include further foreign DNA in the transformation event, enabling other genes to be carried into the plant (see Section XI). As most of the biochemical understanding of tropane alkaloid biosynthesis has been obtained from studies of a limited number of cultures, their basic properties are described here, in order that the reader may understand the context in which the information was derived. A. Hyoscyamus SPECIES Following a survey of alkaloid production in root and callus cultures of seven species of Hyoscyamus, Hashimoto er al. (17) identified root cultures of H . albus as most effective for the production of hyoscyamine and cultures of H . niger and H . gyorfii as hyoscine rich. Alkaloid levels in 8-day-old differentiated cultures of all species were 50- to 100-fold greater than in 28-day-old callus. In some cases, the root content of hyoscyamine in H . albus exceeded 1.2% dry weight. Cultures of both H . albus and H . niger grew rapidly, increasing 6-fold in about 10 days. The alkaloid production profiles, however, were rather different. Hyoscyamus albus cultures had a maximum hyoscyamine content in stationary phase roots, whereas H . niger roots showed a rather constant, low hyoscyamine level and a similarly constant, but higher, hyoscine content. The root cultures were initiated by excising roots from seedlings and culturing on medium, in which they grew in the absence of exogenous phytohormones. Growth of both cultures was stimulated by adding indole-3-acetic acid (IAA) analogs, notably 3-indolebutyric acid (IBA), to the medium, but the more rapidly growing cultures accumulated lower levels of total alkaloids. Root cultures of both H . albus and H . niger, growing in a basal medium without phytohormones, were found to provide good material for biosynthetic studies. An examination in H . albus of the enzymes ornithine decarboxylase (ODC), arginine decarboxylase (ADC), and putrescine N-methyltransferase (PMT) showed that all were present at readily determinable levels and at sufficiently high activities to account for the amounts of alkaloids observed (18). The enzymes are not equally active at all stages of culture, but rather show a distinct peak of activity at the period of maximum growth and minimum hyoscyamine content. The role of these enzymes is discussed later (see Sections 111, IV).

2.

BIOSYNTHESIS OF TROPANE ALKALOIDS

129

B. Datura strarnonium Datura stramonium is a major source of hyoscyamine and hyoscine for pharmaceutical use in Europe and the United States (7). Seed samples from a wide variety of sources were germinated, and the derived plants were examined for their ability to accumulate these alkaloids (19). Transformed root cultures were generated from a range of plants (20) and compared to cultures of several other Datura species (Table I). The highest biosynthetic capacity was found in D . stramonium, and one line, D15/5, showing good growth characteristics, was selected for biochemical studies (21). The hyoscyamine content of D . stramonium D15/5 cultures shows a relationship to growth very similar to that seen in H . albus. The two cultures grow at comparable rates, but D . stramonium is subcultured at a lower level (0.1-0.2 versus 0.5 g fresh weight) and therefore shows a more extended culture period. The total alkaloid content closely follows growth, and the sum of hyoscyamine plus apoatropine constitutes at least 80% of the alkaloids present at all ages of culture. In contrast to H . niger and some other Datura cultures (22), hyoscine is only found at trace levels. Rather, apoatropine forms the major product of hyoscyamine metabolism, being the principal alkaloid present a few days following subculture. Most of the identified intermediates in the pathway have also been demonstrated at various levels in these roots (21). Because the cultures are transformed, phytohormones are not required to stimulate root growth. Indeed, in contrast to the cultures of Hyoscyamus species, adding a-naphthaleneacetic acid (NAA) plus kinetin causes a rapid loss of alkaloid formation and aprogressive loss of rootiness, leading, after prolonged exposure, to a nonproductive suspension culture (23). Such experiments have been valuable in providing information on the importance of root morphology in regulating the overall activity of the pathway (see Section X,A). A further similarity between the D . stramonium and Hyoscyamus spp. cultures is, however, seen in the enzyme activity profiles. Again, a maximum in activity is seen around the time of most rapid growth, and the activity of PMT is found to decrease at a lower rate than the activities of other enzymes.

C . Duboisia SPECIES Several types of cultures have been derived from the Duboisia species D. myoporoides and D . leichhardtii. The genus Duboisia is particularly interesting in biosynthetic terms as in whole plants it forms both the pyridine alkaloid nicotine and hyoscine: some racial variation apparently

130

RICHARD J . ROBINS A N D NICHOLAS J . WALTON

occurs in the ratio to which these types accumulate (24; see Section X,B). Callus (25,26), shoot (26,27), root (28,29), and transformed root (30,311 cultures of one or both species have been used in experiments examining tropane alkaloid formation (Table I). Neither alkaloid occurs at detectable levels in fully established callus cultures of either species (25,26),although some biotransformation of tropine has been reported (25).The callus may be induced to form shoots by phytohormone treatment (26,27),and these are capable of biotransforming both tropine (25) and hyoscyamine (26). When, however, root organogenesis is induced, cultures fully competent in alkaloid biosynthesis are generated (28,29).The alkaloid spectra of root cultures of both D. rnyoporoides and D . feichhardtii differ significantly from those of the aerial parts of whole plants, having a higher pyridine to tropane ratio and a lower hyoscine to hyoscyamine ratio (26).Considering that the conversion of hyoscyamine to hyoscine occurs readily in shoot cultures (26), it is reasonable to conclude that, at least in these species, significant conversion in whole plants occurs in the aerial parts (see Section XB). Transformed root cultures, generated from leaf or root tissue inoculated with Agrobacterium rhizogenes, also produce alkaloids of both classes (30,31). Duboisia rnyoporoides transformed roots appear less efficient at converting hyoscyamine to hyoscine than the comparable untransformed culture, but they grow much better (31). As with D. stvarnonium, exogenous phytohormones tend to differentiate roots and suppress alkaloid formation. A clone derived from a D. leichhardtii transformed culture that is particularly rich in hyoscine has been identified (32).

111. Formation of Putrescine

It has been convincingly established by radio- and stable-isotope labeling that the tropane moiety of tropeines derives its pyrrolidine ring from either ornithine or arginine (Scheme 3). The details of how putrescine (or N-methylputrescine; see Section IV,A) is derived are, however, less well defined. When either [14C]arginineor [14C]ornithineis fed to D. stramoniurn root cultures, the I4Cfrom both is readily incorporated into hyoscyamine (33). Similarly, label from these amino acids is incorporated by H. albus into hyoscyamine (34). The direct incorporation of both amino acids by decarboxylation to the respective amines is quite possible in these cultures, since the required enzymes, ADC and ODC, are present. The product of ADC, agmatine, is apparently rapidly converted to putrescine

2.

131

BIOSYNTHESIS OF TROPANE ALKALOIDS

H I CH,CH2CH,C--COOH I I NH NH2 I C=NH I NH2

Arginine

Omithine

1 Putrescine

N-Carbamylputrescine

Agmatim

N-Methylputrescine SCHEME 3. Formation of putrescine and N-rnethylputrescine from ornithine or arginine.

via N-carbamylputrescine (35).Biosynthetically, ornithine is derived from arginine (36),and in most plants, including Datum ( 3 3 ,the enzyme arginase that cleaves the guanidino group from arginine is present. Arginase activities are quite substantial in D . stramonium and A . belladonna root cultures (33), though the activity is barely detectable in root cultures of H . albus (18). Thus, it is possible that arginine incorporation might, in some cases, proceed via ornithine, the agmatine pathway not being involved. Two lines of evidence, however, indicate that arginine is directly metabolized to hyoscyamine.

132

RICHARD J . ROBINS A N D NICHOLAS J . WALTON

A. INCORPORATION OF LABELED PRECURSORS [U-14C]Agmatineis readily prepared from [U-14C]arginineby enzymatic decarboxylation. When labeled agmatine was fed to D.stramonium root cultures, an excellent incorporation (8.7%) was obtained, better than with either arginine or ornithine under the same conditions (33). Agmatine could not have been incorporated via ornithine but, rather, must have been converted directly to putrescine via N-carbamylputrescine. When [2,3-3H,]arginine was fed to H. albus cultures, it was rapidly absorbed and metabolized to form free amines and alkaloids (34). Negligible amounts could be detected in the ornithine pool. Thus, again, it can be concluded that arginine is decarboxylated and incorporated directly. Corroborative evidence has been obtained in Nicotiana tabacum callus cultures, where label from [U-I4C]arginine was much more efficiently incorporated into nicotine than was label from [U-14C]ornithine(38). Unfortunately, no attempt was made to determine the levels in agmatine in the experiments.

B. EFFECTOF ENZYME INHIBITORS The second line of evidence comes from experiments employing inhibitors of ODC and ADC, namely, DL-a-difluoromethylornithine (DFMO) and DL-a-difluoromethylarginine (DFMA), respectively. These active sitedirected, catalytically activated inhibitors are highly specific and irreversibly inhibit the target enzyme (39). The growth of D. stramonium root cultures is unaffected by the presence of DFMO or DFMA at up to 5 mM exogenous concentration. Roots grown with DFMO for 18 days contain normal levels of hyoscyamine (33),even though the ODC activity, determined at 10 days, had been completely inhibited (40). Growing roots with DFMA similarly eliminates ADC activity (40), but, in contrast, a much smaller alkaloid accumulation is subsequently observed (33). Lntermediates in the pathway are also found at much lower levels than in control tissue. The situation in H. afbus is, however, different (34). Only DFMO was tested in this culture, but an inhibition of incorporation was seen. These experiments, however, followed a different protocol, the cultures being fed DFMO (5 m M ) for only 24 hr and then ~ ~ - [ 5 - ' ~ C ] o r n i t h ifor ne 7 hr. Under these conditions, DFMO appeared to have decreased the incorporation of radiolabel into hyoscyamine and scopolamine, but the amount of activity in alkaloids in both control and treated tissues was extremely low. These inhibitors have been effectively applied in other cultures and have provided supporting evidence for a role of ADC in alkaloid formation.

2.

BIOSYNTHESIS OF TROPANE ALKALOIDS

133

In N. tabacum callus cultures, DFMA was somewhat more effective at inhibiting nicotine formation than was DFMO, although the diminution was not great, and, even at 1 mM, growth was also inhibited (38). The influence of these inhibitors has also been examined in other alkaloidproducing genera. Similar, and particularly convincing, results were obtained for pyrrolizidine alkaloid formation in Senecio vulgaris root cultures (41). These were shown to contain very low levels of ODC, and DFMO was found to have little influence on the culture. DFMA, in contrast, severely suppressed alkaloid accumulation. C. METABOLITE TUNNELING Both ornithine and arginine may apparently be directly incorporated into tropeines via the common intermediate putrescine. This symmetrical diamine is involved in other metabolic events in cells (see Section X), and it may be argued that the first true metabolite of the tropane pathway is, in fact, N-methylputrescine (see Section IV). An alternative is that a molecule of arginine or ornithine becomes committed to a specific biosynthetic pathway at the stage of decarboxylation (see Section X,A). The first evidence that this may occur in Datura was obtained when [2-I4C1ornithine was fed to intact D. stramonium plants (42,43):the label was only incorporated into the C-1 position of tropine (42,43).This indicated that incorporation had not involved a free, symmetrical intermediate such as putrescine. Similar data were produced for D. metel (44) and D. innoxia (45). In contrast, ~~-[5-'~C]ornithine fed to H. albus (46) or Duboisia leichhardtii (47)root cultures yielded symmetrically labeled tropane rings. Leete initially interpreted the data from Datura by suggesting that 4N-methylornithine might be an intermediate, with the methylation prior to decarboxylation ensuring that the C-2 and C-5 of ornithine remained metabolically distinct (8). This attractive hypothesis has since been rejected on biosynthetic grounds (see Section IV,A). Instead, the less experimentally testable proposal has been made (2) that in Darura (though not in H. albus) putrescine remains bound to the decarboxylase prior to methylation. ODC and PMT are suggested to be closely associated, and their sequential reactions to occur in a concerted manner. Because ODC and PMT are distinct and separable activities in uitro, there is no enzymological evidence for this proposal. Nevertheless, physiological data from feeding experiments indicate that some degree of concerted metabolism may occur. When 25 nmol of DL[5-I4C]ornithine (58 kBq) was fed to D. stramonium roots in 50 ml of medium, an incorporation of 4.5% was obtained (33). Feeding the same level of radiolabel in the presence of 50 pmol of unlabeled putrescine only diminished incorporation from ornithine to 2.7%, less than 50% at a

I34

RICHARD J . ROBINS A N D NICHOLAS J . WALTON

dilution of label of 2000 : 1. Identical experiments with [U-'4C]arginine produced a decrease from 3.7 to 1.6%. At the same time, however, labeled putrescine fed at high concentration was readily and efficiently incorporated. Thus, it may be speculated that to some extent an enzyme-bound intermediate may exist, there being limited exchange of putrescine with the free pool. Rather more convincing indications have been obtained that cadaverine may be enzyme bound during anabasine formation from lysine in Nicotiana hesperis root cultures (48),again supporting previous labeling experiments. The mechanism by which this phenomenon occurs will be difficult to establish. Metabolite tunneling might involve, as suggested, the direct transfer of the product of one enzyme to the active site of the next enzyme in a sequence of reactions. Alternatively, it could be caused by compartmentation of the reaction sequence within the cell, the boundary of the compartment having poor permeability to the metabolites in question. In each case, dilution of the pools of intermediates by molecules exogenous to the system will be decreased, leading to the apparent directing of molecules to a particular product without dilution, as observed in the tropane and tobacco alkaloid pathways.

IV. Formation of Tropinone A. CONVERSION OF PUTRESCINE TO N-METHYLPYRROLINIUM Although 5-N-methylornithine, when fed to D.stramonium plants, has been found to be incorporated into hyoscyamine and scopolamine (8), this does not, of course, prove that metabolism via this compound occurs in uiuo. Indeed, when Hedges and Herbert (49) isolated labeled 5-Nmethylornithine from Atropa belladonna plants fed with labeled ornithine, no label at all could be detected in the alkaloids. More recently, it has been demonstrated that ornithine 5-N-methyltransferase and 5-Nmethylornithine decarboxylase, the enzymes required for the conversion of ornithine to N-methylputrescine via this route (Scheme 4), are undetectable in root cultures of Hyoscyamus albus (34),D . stramonium, and A . belladonna (33). On the basis of these findings, it can be concluded that essentially all of the carbon framework destined for tropane alkaloid biosynthesis from ornithine passes through putrescine (Scheme 5 ) , in common with that arising from arginine via agmatine, as already described (Section 111).

2.

-

BIOSYNTHESIS OF TROPANE ALKALOIDS

H

I CHZCHZCH~F-COOH NH2 I NH2

CH2CH2CH2CH2 I I

NH2

Omithine I I I I I

135

-

NH2

Putrescine

4---

--

'I

SAM

5-N-Methylomithine

-----&

I

Pyrroline

[SAM]

--_

I I I

; ?? 'I

N-Methylpumscine

N-Methylpyrrolinium

SCHEME 4. Alternative routes by which the N-methyl group could be introduced. SAM, S-Adenosylrnethionine.

This conclusion is fully supported by the presence in roots and root cultures active in tropane alkaloid biosynthesis of appreciable levels of the enzyme putrescine N-methyltransferase (PMT; EC 2.1.1.53), and, conversely, by the absence of the enzyme from aerial and other tissues incapable of nicotine or tropane alkaloid biosynthesis (Section X,A). In

-

Putrescine

CH,CH,CH,CH, I I NH NH2 I CH3

-

CH,CH,CH,CH I II NH 0 I CH3

4-methy laminobutanal

N-Methylputrescine

c I -

I-J y

x-

CH3

II

0 Tropinone

Hygrine SCHEME5. Formation of tropinone from putrescine.

N-Methy lpyrrolinium

136

RICHARD J. ROBINS A N D NICHOLAS J . WALTON

general, under the conditions of assay, the activity of PMT is comparable to, o r slightly greater than, that determined for either ODC or ADC. This enzyme, first described in roots of N. tabacum (50), appears to be solely responsible for N-methylation in this pathway. The enzyme was purified 30-fold using ammonium sulfate fractionation and gel chromatography, and some kinetic and other properties were determined. The pH optimum was found to be between 8 and 9, the molecular weight was about 60,000 D (estimated by gel filtration), and the apparent K, values for putrescine and S-adenosylmethionine were 0.40 and 0.1 1 mM, respectively. The enzyme was found to be particularly sensitive to sulfhydry1 reagents, requiring 2-mercaptoethanol to prevent a rapid loss of activity and being inhibited by the sulfhydryl-binding reagents AgNO, and 4-chloromercuribenzoate. Specificity for putrescine as the methylaccepting substrate appeared to be virtually absolute, with no activity being observed with cadaverine, A'-pyrroline, o r ornithine and only slight methylation being detectable with N-methylputrescine. Since 1971, only one further paper reporting the properties of PMT has appeared (51). Using a new high-performance liquid chromatography (HPLC) assay, the authors quoted apparent K, values of 0.88 m M (putrescine) and 0.15 m M (S-adenosylmethionine)for the enzyme extracted from a root organ culture of D. stramonium. The absence of further information from more highly purified preparations appears to be related to stability problems. Feth et al. (51) reported that 35% of the activity was lost after 3 hr at 4"C, although the enzyme could be stabilized to some extent by freezing in the presence of 12.5% (w/v) glucose. In a procedure involving precipitation by polyethylene glycol, ion-exchange chromatography, and affinity chromatography on a column of S-adenosylhomocysteinylaminohexyl-Sepharose 4B, using 12.5% (w/v) glucose and 10 m M 2mercaptoethanol throughout, a reliable purification of between 200- and 500-fold has been achieved (52). The properties of this more purified preparation are now being investigated. The extent to which PMT reacts with unbound putrescine in vivo may be variable between genera o r species (see Section III,C), though there is no doubt that exogenous putrescine is actively incorporated into hyoscyamine by transformed root cultures. Walton et al. (33) observed a 7.4% incorporation into hyoscyamine of millimolar levels of [ 1,4-14C]putrescine fed to transformed root cultures of D. stramonium. This metabolism occurred without substantial dilution of the label, as the specific incorporation (ratio of the specific radioactivity of hyoscyamine to that of the putrescine supplied) was 75%. Experimental evidence suggests that the activity of PMT, as the enzyme apparently responsible for committing the metabolism of putrescine to the production of alkaloids, is closely

2.

BIOSYNTHESIS OF TROPANE ALKALOIDS

137

regulated in Nicotiana and Datura roots, though, as indicated below (see Section X,A), other factors, as yet poorly understood, may contribute to the maintenance of the overall substrate specificity and organized metabolic flow. The second step in the conversion of putrescine to N-methylpyrrolinium is catalyzed by a diamine oxidase, N-methylputrescine oxidase (MPO; EC 1.4.3.6). The properties of the enzyme from root cultures of Hyoscyamus niger have been studied in a preparation purified about 40-fold using ammonium sulfate fractionation followed by chromatography on DEAE-Sephadex and hydroxyapatite (53).The apparent K , values for Nmethylputrescine, putrescine, and cadaverine were found to be 0.33,2.85, and 6.25 mM, respectively, indicating a substantially higher affinity for N-methylputrescine than for either of the other two substrates; consistent with this, the enzyme was inhibited by N-methylated amines somewhat more strongly than by the corresponding primary amines. The V,, values determined with N-methylputrescine and putrescine were similar, whereas that with cadaverine was only about 20-25% as high, leading to specificity constants (V,,,/apparent K,) for the three substrates of 100, 1 1 , and 1%, respectively. In contrast, corresponding specificity constants of 21, 82, and 100% were observed by the same authors for diamine oxidase from the epicotyls of pea (Pisum satiuum). Rather different results were obtained using extracts of transformed roots of Nicotiana tabacum var. SC58 (54, where lower apparent K, values for all three substrates, especially Nmethylputrescine and putrescine, were obtained. Furthermore, the highest V,, was with cadaverine, consistent with the production of the cadaverine-derived alkaloid, anabasine, observed when transformed root cultures of Nicotiana species were fed this diamine (55). A summary of kinetic parameters obtained for MPO from H. albus and N. tabacum is given in Table 11. The causes of the differences between the values observed are unclear, but they might relate to the potential for anabasine production in Nicotiana. There is good spectroscopic and electrochemical evidence that, like bovine amine oxidase and a number of bacterial oxidoreductases, diamine oxidase from Pisum satiuum contains copper and the tightly bound pyrroloquinoline quinone cofactor (56). Although absolute proof of the presence of this cofactor has not been obtained for MPO from solanaceous genera, it has been strongly indicated by the behavior of phenylhydrazine as an active site-directed inhibitor of the N. tabacum enzyme (57). Inhibition by phenylhydrazine was irreversible but was partially prevented by the substrate, N-methylputrescine. It was possible to label the enzyme irreversibly with ['4C]phenylhydrazine, and analysis of the labeled enzyme by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE)

TABLE I1 KINETICPROPERTIES OF N-METHYLPUTRESCINE OXIDASES FROM Hyoscyamus albus A N D Nicotiana tabacum

Species

Source

Purification (-fold)

Hyoscyamus albus Nicotiana tabacum var. SC58 Nicotiana tabacum cv. Bright Yellow Nicotiana tabacum cv. Sarnsun

Root culture Transformed root culture Root

41.8 Gel-filtered extract 152

Root

Gel-filtered extract

a

Apparent K , ( m M )

V,,, (relative)a

N-Meputb

Put

Cad

N-Meput

Put

Cad

Refs.'

100

100

91 139

22 290

0.33 0.08

2.85 0.35

6.25 2.15

2

100

ndd

nd

0.45

nd

nd

3

100

nd

nd

1.9

nd

nd

4

1

V,, values are relative, in each case, to that observed with N-methylputrescine, set at 100%.

* N-Meput, N-Methylputrescine; Put, putrescine; Cad, cadaverine.

Key to references: (1) T. Hashimoto, A. Mitani, and Y. Yamada, Plant Physiol. 93, 216 (1990); (2) N. J . Walton and W. R. McLauchlan, Phyfochemistry 29, 1455 (1990); (3) S. Mizusaki, Y. Tanabe, M. Noguchi, and E. Tamaki, Phytochemisfry 11,2757 (1972); (4) F. Feth, V . Wray, and K. G. Wagner, Phyfochernisrry 24, 1653 (1985). nd. Not determined.

2.

BIOSYNTHESIS OF TROPANE ALKALOIDS

139

enabled a subunit molecular weight of around 70,000 D to be deduced. This is fairly close to the values for the subunit molecular weight of a number of other plant diamine oxidases. A somewhat lower value for the molecular weight of this enzyme has been obtained by other workers (58). In this case, SDS-PAGE has indicated a subunit molecular weight of 53,000 D, and comparison with gel filtration data suggests that the native enzyme is a dimer, in common with other diamine oxidases. The causes of the differing molecular weight estimates for the Nicotiana enzyme are uncertain. Polyclonal antibodies raised against the purified protein have been found to recognize a single protein band of similar molecular weight in unpurified extracts of transformed root cultures of several solanaceous species (Atropa belladonna, Saracha edulis, and Datura stramonium) and also several nonsolanaceous ones (Emilia coccinea, Cynoglossum australe, Securinega suffruticosa, and Beta vulgaris), but there is little or no cross-reactivity with partially purified preparations of diamine oxidase from etiolated seedlings of Pisum sativum (58). The greater affinity of MPO for N-methylputrescine, relative to putrescine, promotes the preferential oxidation of the former. Competition experiments in extracts of transformed roots of N. tabacum confirmed that, at both 1 and 0.1 mM concentrations, provision of an equal concentration of either putrescine or cadaverine did not inhibit the oxidation of N methylputrescine (54). Conversely, the oxidation of these unsubstituted diamines was diminished by N-methylputrescine. In root cultures of D . stramonium, the concentration of soluble, free (in contrast to conjugated) putrescine was found to be comparable with that of free N-methylputrescine (21), and a kinetic adaptation of the enzyme may be essential to prevent the formation of 4-aminobutanal from putrescine. As pointed out by Hashimoto et al. ( 5 3 , direct oxidation of putrescine may be involved in the formation of a small number of nortropine esters in which the Nbridge is unmethylated (59). In the formation of the pyrrolizidine alkaloids by members of the Asteraceae and Boraginaceae, putrescine oxidation appears to be catalyzed by a diamine oxidase (60). Furthermore, oxidation of putrescine is clearly an essential aspect of the metabolism of cell lines of Nicotiana cultures selected for growth on putrescine as a sole carbon and nitrogen source (61). To what extent the regulation of putrescine oxidation is the result of further subcellular factors, however, is not clear (see Section X,A). 4-Methylaminobutanal, originally established as an intermediate in the biosynthesis of nicotine, is in equilibrium with the N-methylpyrrolinium ion (62). It is at this point that the pathways of biosynthesis of nicotine and the tropane alkaloids diverge. Little is known of the enzymology of nicotine formation from N-methylpyrrolinium and nicotinic acid, though

140

RICHARD J. ROBINS A N D NICHOLAS J . WALTON

one paper (63)reports a “nicotine synthase” activity of 80 fkat/mg protein in extracts of Nicotiana gfutinosa and N. tabacum. The activity was O2 dependent, was stimulated by divalent metal ions, and gave rise specifically to the (S) isomer of nicotine.

B. CONVERSION OF N-METHYLPYRROLINIUM TO TROPINONE Whereas the enzymology of the conversion of ornithine and arginine to N-methylpyrrolinium has now been described in outline, even if the enzymes have not been thoroughly purified and characterized, the subsequent reactions leading from N-methylpyrrolinium to tropinone (Scheme 5 ) are not well understood. No enzymes have yet been demonstrated, and even the mechanism of the process remains in some doubt. Schematically, the reaction sequence may be depicted as shown in Scheme 6. N-Methylpyrrolinium is proposed to condense with an aceto-

H3CC-OH II 0

-

HS-CoA

H3CC-S-CoA II 0 Acetyl-CoA

-

HS-CoA

H3C\

/cEr21

C II 0

C-S-COA I1 0

Acetoacetyl-CoA

\ 0

Tropinone

Hygrine

SCHEME 6. Conversion of N-rnethylpyrroliniurn to tropinone.

0

2.

BIOSYNTHESIS OF TROPANE ALKALOIDS

141

acetyl unit, with the release of CO,, to form hygrine. This is then oxidized to 5-acetonyl-1-methyl-A'-pyrrolinium,which, by a Mannich reaction, condenses to form tropinone. Several questions arise in relation to this scheme. These concern the source of the acetonyl unit, its manner of condensation with N-methylpyrrolinium, and the role of (I?)- and ( S ) hygrines as precursors of tropane alkaloid formation. Acetoacetate was found to be a good precursor of hyoscyamine in Datura plants (64) and of hygrine in Nicandra physaloides (65). The latter study established, using sodium [4-14C]acetoacetatewick-fed to N. physaloides plants and chemical degradation of the labeled hygrine subsequently formed, that condensation with N-methylpyrrolinium occurred at the C-2 atom of acetoacetate. Over 95% of the label recovered in the isolated hygrine was at the C-3' position. Condensation, however, is a facile nonenzymatic process. Acetoacetate, acetonedicarboxylate, and, to a much lesser extent, acetoacetyl-CoA produced hygrine when incubated with Nmethylpyrrolinium in uitro (66). The pH optimum of the reaction with acetoacetate was 9, but the reaction rate was approximately half-maximal at pH 5. Over a 5-hr period at 30°C and pH 7, with 1 mM acetoacetate and approximately 2.4 mM N-methylpyrrolinium in a reaction volume of 1 ml, the rate of hygrine formation was about 70 nmol/hr. The rate was not influenced by the addition of about 1 mg protein in cell-free preparations from cultures of Duboisia leichhardtii or Hyoscyamus albus. In a further experiment, hygrine formation was observed from 10 mM acetoacetate fed alone to a Nicotiana root culture (66). Synthesis apparently occurred using N-methylpyrrolinium generated endogenously. Substantial levels of N-methylpyrrolinium (- 100 nmol/g fresh weight), similar to the levels of free putrescine, have been measured in transformed root cultures of Nicotiana rustica (67). Over a 24-hr period, 4.3 pmol of hygrine per gram dry weight of tissue was produced. (A much smaller amount, 0.35 pmollg dry weight, was produced from acetonedicarboxylate.) Using an approximate general estimate of 50 mg/g dry weight for the protein content of Nicotiana roots, the mean rate of formation of hygrine from acetoacetate was therefore about 1 pkat/mg of protein. These values compare with enzyme activities of MPO (responsible for the generation of Nmethylpyrrolinium) of about 70 pkat/mg protein in 10-day-oldtransformed root cultures of N. tabacurn (54) and 10-50 pkat/mg protein in 10-dayold transformed root cultures of D. strarnoniurn (21,40). The substantial nonenzymatic rates of hygrine formation imply that it will be very difficult to determine accurately the rate of any simultaneous enzyme-catalyzed reaction, and so far this has not been achieved. Indeed, it might be argued that hygrine formation in uiuo need not involve an enzyme-catalyzed process. There is a precedent for the occurrence of

142

RICHARD J . ROBINS A N D NICHOLAS J . WALTON

nonenzymatic steps in the middle of a biosynthetic pathway in the formation of the Rauwolfia alkaloids (68). As a result of experiments on the biosynthesis of cocaine, it has become necessary to question the assumption that acetoacetate (or its coenzyme A thioester) is an obligate precursor of hyoscyamine and scopolamine. diethylaceLeete and Kim (69) fed 4 4 1-'3C,'4C,'SN](methylarnino)butanal tal to Erythroxylum coca plants and analyzed the I3C-NMR spectrum of the labeled cocaine produced. Comparison with natural cocaine revealed an enhancement of the signal resulting from C-5 and an upfield satellite of this signal resulting from I3Cadjacent to lSN;no satellites were observed for the signal due to C-1. Carbon-1 of the labeled diethylacetal substrate (assumed to be hydrolyzed to 4-methylaminobutanal in the plant tissue) was apparently metabolized to C-5 of cocaine, and not to C-1 as predicted by the generally established metabolic route involving the attack of the methylene carbon of the acetoacetate at C-2 of N-methylpyrrolinium (cf. Scheme 6). The newly proposed biosynthetic route of Leete and Kim (69) is shown in Scheme 7. It is suggested that N-methylpyrrolinium reacts with malonylCoA to yield the CoA thioester of 1-methylpyrrolidine-2-acetic acid; this then reacts with a further molecule of malonyl-CoA to give the CoA thioester of 1-methylpyrrolidine-2-acetoaceticacid. Oxidation of the pyrrolidine ring and subsequent Mannich condensation afford the bicyclic tropane ring system. Some further circumstantial evidence for this scheme

Ecgonine methyl ester

SCHEME 7. Suggested route for the formation of ecgonine methyl ester in cocaine biosynthesis

2.

BIOSYNTHESIS OF TROPANE ALKALOIDS

143

has been the isolation of 1-[2-'4C]methylpyrrolidine-2-aceticacid from E . coca plants fed N-[2-'4C]methylpyrrolinium; the labeled product was isolated from an aqueous alkaline extract to which the unlabeled compound had been added as a trap. No conclusive experiments of this kind have yet been reported in species producing hyoscyamine and scopolamine, and the performance of such studies must therefore be a priority (see below). Prompted by these proposals (69) for the biosynthesis of cocaine, the mode of synthesis of the lysine-derived alkaloid N-methylpelletierine, a higher homolog of hygrine, has been examined using 13C-NMR spectroscopy (70). However, these studies have vindicated the accepted scheme of biosynthesis for this alkaloid, and its reduction product N methylallosedridine, in Sedum sarmentosum by proving the incorporation of an intact acetoacetate unit, with condensation of the methylene carbon of an acetoacetate unit with C-2 of the piperideine ring. Separate experiments were performed using cuttings fed sodium [ 1,2-I3C,]acetate and In particular, using labeled acetoacesodium [ 1,2,3,4-'3C4]acetoacetate. tate, a two-bond long-range coupling was observed between the C-methyl carbon and the methylene carbon of the side chain of N-methylpelletierine, indicating incorporation without cleavage to labeled acetate. NMR spectroscopy has also been used in an attempt to establish, from the I3C-I3C coupling, the mode of acetate incorporation into C-2 and C-4 of hyoscyamine by untransformed root cultures of H . albus fed [ 1,213Czlacetate.The labeled acetate molecules were incorporated intact into the pairs C-2 and C-3 and C-3 and C-4 at a 1 : 1 ratio. From this evidence, it was again concluded that both isomers of hygrine were apparently utilized in hyoscyamine biosynthesis (72). Very similar results have recently been obtained from feeding [ 1,2-I3Cz]acetateto transformed root cultures of D . stramonium (72). No conclusion could be drawn regarding the mode of acetate incorporation into hyoscyamine and, in particular, whether the malonyl-CoA route (69) is involved in the biosynthesis of hyoscyamine. Evidence on the role of the different isomers of hygrine as precursors of tropane alkaloids has been conflicting. McGaw and Woolley (73) found that (R)-hygrine was incorporated into hyoscyamine much more effectively than the (S) isomer by D . innoxia plants. On the other hand, the same authors found that both (R)- and (S)-hygrines were precursors of tropane alkaloids, with approximately equal efficiency, in H . niger, A . belladonna, and Physalis alkekengi (74). The enantiomers of hygrine were isolated as their tartrate salts and fed to the roots of intact plants; these salts are stable, although the optically active free base readily racemizes. However, the intermediacy of hygrine has been questioned (64)and recent

144

RICHARD J . ROBINS A N D NICHOLAS J . WALTON

experiments have cast further doubt on the role of free hygrine in this pathway. When [2',3'-I3C,]hygrine was fed to transformed root cultures of D. stramonium over a 16-day period, relative specific incorporations of 20 and 37% were observed in cuscohygrine and in an unidentified condensation product of N-methylpyrrolinium and hygrine, respectively (75). Although the relative specific incorporation into tropinone was about 8%, there was virtually no incorporation into tropine (0.03%) and no detectable incorporation whatever into hyoscyamine. Similarly, no incorporation into hyoscyamine was detected in whole plants of D. innoxia or root cultures of H. niger (76). This result appears at variance with the earlier observations of McGaw and Woolley (73,74). It would appear that hygrine, used in the biosynthesis of cuscohygrine and condensation products with N-methylpyrrolinium, may not gain full access to the hyoscyamine biosynthetic pathway. Further experiments along these lines are clearly necessary to elucidate the conditions required for the incorporation of exogenous hygrine into hyoscyamine, as well as the relationship of this incorporation to the synthesis of cuscohygrine and other, more immediate, products of hygrine metabolism. Advances in understanding the enzymology involved will assist in establishing the details of the metabolic pathway. Although no enzyme capable of converting hygrine to tropinone has yet been demonstrated, conditions have been established for a biomimetic reaction (77). Hygrine was boiled for 18 hr with excess mercury(I1) acetate in 2% acetic acid; the reaction mixture was cooled and decanted and then treated briefly with sodium cyanoborohydride, which was essential to release the reaction products from complexation with mercury( 11) acetate. The mixture contained unreacted hygrine (50-60%), together with tropinone (10-15%) and its isomer 2,l'-dehydrohygrine (20-30%). The proposed reaction mechanism is shown in Scheme 8. Following the formation of a quaternary ammonium compound with mercury(I1) acetate, loss of a proton from C-5 leads to the formation of the 5-acetonyl-l-methyl-A'pyrrolinium salt which, by a spontaneous Mannich reaction, leads directly to the production of tropinone. By contrast, loss of a proton from C-2 yields the 2-acetonyl-l-methyl-A'-pyrroliniumsalt, which cannot cyclize but which then rearranges, with loss of a proton from C-l', to give the vinylogous amide 2,l '-dehydrohygrine. This compound has been detected in root extracts of the solanaceous shrub Vussobia breuijlora (78), and it may, perhaps, occur more widely; on account of its weak basicity, however, it will readily extract into CH,CI, from acidic solutions and may have been missed in conventional alkaloid extractions. Huhtikangas and co-workers have made the interesting suggestion that the oxidation of hygrine to tropinone via dehydrohygrine might involve

2.

BIOSYNTHESIS OF TROPANE ALKALOIDS

145

Loss of H ai C-5

Loss of H at C-2

2,1'-Dehydrohygrine

SCHEME8. Biomimetic conversion of hygrine to tropinone or 2, I '-dehydrohygrine.

an autocatalytic loop via a specific tropinone-water adduct (79,80). This theory is based on considerations of molecular electrostatic potential and the solvation of tropinone in water. It will be difficult to find direct experimental support for this theory, though it may well be consistent with some of the apparent anomalies of hygrine metabolism referred to above. Although it is not possible to be clear about the mechanism of conversion of N-methylpyrrolinium to tropinone, recent experiments have at least clarified the important questions which now need to be asked. The proposal of Leete and Kim (69) regarding the mechanism of cocaine biosynthesis is currently being examined in relation to hyoscyamine and scopolamine biosynthesis in a range of genera. Although hygrine may be formed by a rapid nonenzymatic reaction between N-methylpyrrolinium and acetoacetate, the physiological significance of this remains unclear. First, it is not certain that acetoacetate is an obligate precursor of hyoscyamine, even though earlier work (64) demonstrated clearly that radioactive acetoacetate was incorporated into this alkaloid. Second, evidence that hygrine (65) and N-methylpelletierine (70) are formed from acetoacetate without degradation to acetate does not imply that tropinone and hyoscyamine are necessarily formed by the same mechanism, especially in different genera. Third, earlier uncertainty regarding the role of the ( R )and ( S ) enantiomers of hygrine as precursors of hyoscyamine has deepened, so that it is necessary to question whether free hygrine is an intermediate in tropinone and hyoscyamine formation. Taken together, present evidence suggests the possibility that the conversion of N-methylpyrrolinium to tropinone is a concerted process and that hygrine may be a shunt metabolite with variable access to the pathway. This could, of course, explain the inability so far to demonstrate any enzymology for the conversion.

I46

RICHARD J. ROBINS A N D NICHOLAS J . WALTON

V. Formation of Tropine and Pseudotropine Surprisingly, it is only recently that labeling data have been provided showing that tropinone is incorporated into hyoscyamine (81).Tropinone is reduced stereospecifically to either tropine or pseudotropine by the enzymes tropinone reductase I and I1 (TR I and 11), respectively. These activities have been identified, separated, and purified from several sources (Table 111). Initially, it was not clear whether only one protein existed, the tropine-forming activity being lost during purification. Recent work from Drager and co-workers, using dye-affinity matrix binding and hydrophobic interaction chromatography has, however, clarified this situation and demonstrated that there are two separable proteins (82). The ratio of TR I to TR I1 varies considerably with the species examined (83). TR I is particularly dominant in D. stramonium, where up to a 20-fold higher level has been determined. It has, however, proved to be less stable than TR 11, and has therefore been more difficult to purify.

I A. TROPINONE REDUCTASE The presence of tropine-forming tropinone reductase in sterile excised roots of D. stramonium was first described by Koelen and Gross (84). They reported that the enzyme reaction was reversible, that tropine was the exclusive product of reduction, and that only NADPH was active as the reductant. All these conclusions have, however, subsequently been questioned. Drager et al. found no activity with NADH in extracts of root cultures of H. niger, D. stramonium, or A . belladonna (85), but, in contrast, Couladis el al. (86) report that TR I in extracts of D . innoxia roots is active with NADH at 40-60% of the NADPH-utilizing rate. The assay was performed using [N-rnethyl-'4C]tropinoneand differed from the methods used by other workers, who have followed the oxidation of NAD(P)H spectrophotometrically and/or quantitated the alkaloidal products by gas chromatography (GC). The levels of activity are extremely low compared to those determined by other workers, however (see Table III), and the cause of this activity remains to be clarified. TR I activities from both H. niger (83) and D . stramonium (87) have been purified to near homogeneity. TR I from D. stramonium shows a sharp pH optimum at pH 6.0. The activity from both sources will reduce several other substrates (Table 111), but, in contrast to TR I1 (see below), none of the substrates tested is more efficiently reduced than tropinone.

2.

BIOSYNTHESIS OF TROPANE ALKALOIDS

147

I1 B. TROPINONE REDUCTASE It has been shown that TR I1 is present in D . stramonium. TR I1 was first isolated from H. niger root cultures, a 760-fold purification being achieved in a three-step procedure (85).The activities from D . stramonium (87),H . niger (85), and A . belladonna (88)show some variation in properties. All three show maximal activity around pH 5.8 to 6.0, but, whereas the D . stramonium enzyme shows remarkably little pH dependence, the activities from the other cultures are strongly inhibited by acidic conditions yet retain about 80% maximal activity at pH 8.5. These activities again use NADPH and only oxidize NADH at 5% or less of the rate with NADPH. They are completely free of TR I activity. The preparations show activity with a number of tropinone analogs, but, in contrast to TR I, are more active with N-methyl-4-piperidone and N-propyl-4-piperidone than with tropine. The affinity for these nonnatural substrates is, however, 2- and 8-fold lower, respectively (Table 111). These activities appear to be irreversible, with no activity being detected with NADP+ and either tropine or pseudotropine (Table 111). These findings confirm at the biochemical level the conclusion reached by Leete (89) on the basis of the retention of 3H in hyoscyamine, hyoscine, and meteloidine when D . meteloides was fed [N-methyl-'4C,3P-3H]tropine. In contrast, Koelen and Gross reported a higher activity for the oxidation of tropine with NADP+ than for the reduction of tropinone (84).They also found their extract to oxidize aromatic alcohols (benzyl, vanillyl, and coniferyl) at significant rates (10-40%), and thus it might be that their oxidation of tropine is artifactual, involving a NADP+-utilizing, nonspecific alcohol dehydrogenase. In H. niger (85) and D. innoxia (90) no interconversion of tropine and pseudotropine could be measured. It appears, therefore, that the TR I1 enzyme has a distinct role in producing the known tropane alkaloids of the 3p configuration. Recently, it has been proposed (88)that pseudotropine is the precursor of the polyhydroxynortropanes known as the calystegins (91). Certainly, the root cultures that produce these compounds have high TR I1 activity (90),but a direct demonstration of this pathway has yet to be provided. The effect that varying the TR I to TR I1 ratio may have on the alkaloid composition of the tissue is discussed later (see Section X). In biochemical studies, it is always valuable to have available specific enzyme inhibitors. Unfortunately, Drager et al. (85) did not identify any inhibitors among the alternative substrates exhibited. Recently, during studies of the ability of D . stramonium root cultures to metabolize tropane analogs (92), an inhibitor of TR I was discovered. 8-Thiabicyclo[3.2.1]octan-3-one (TBON), the sulfur analog of nortropinone, is metabolized to the alcohol 8-thiabicyclo[3.2.l]octan-3-ol(TBOL) and its 3-0-acetyl ester

TABLE 111 PROPERTIES OF TROPINONE REDUCTASESFROM VARIOUSSOURCES Activity

Species

e

m P

Atropa belladonna

Tissue

Type

Transformed TR I1 root culture

Level Purification ( p k a t h g protein) (-fold) 94

17.472 0 54.687

pH optimum 5.9(broad to alkaline)

12,405

-

0

-

Substrate Tropinone Tropinone N-Methyl-4-piperidone Tetrah ydrothiopyran-4-one 3-Quinuclidinone

Substrate

Cofactor

Km

Km

(mM)

Cofactor NADPH NADH NADPH NADPH NADPH

Inhibitor K0.5

(mM)

Inhibitor

0.021

None identified

-

(mM)

Refs.O 1

-

-

-

Datura innoxia

Excised root

Datura Root stramonium culture

TR 1 TR Ib

None

TR II*

None

Mixed

None

Dehydrogenase

6.8

0 Tropinone 0.125 6.4 (broad) Tropinone 0.063 Tropinone 0.88 Tropinone 0.242 6.4 (broad) Tropinone 0.067 Tropinone 1.21 Tropinone 0 Tropinone 0 2-Carbomethox y-3-tropinone Tropine 147 9.5 0 Tropine Pseudotropine 0 64.6 Scopine 17.6 Nortropine 55.9 Benzyl alcohol 16.2 Vanillyl alcohol 27.9 Coniferyl alcohol

NADPH NADPH NADH NADPH NADPH NADH NADPH NADP NADPH NADP NAD NADP NADP NADP NADP NADP NADP

2

+

NADH

+

NADH

3

Mixed Transformed TR I root culture

80.1

13‘

TR I1

Hyoscyamus niger

Root culture

TR I1

7611

0 10,490 0 1678 2308 8708 7868 3150 2130 0 I 07 3770 2343 0 0 17.540 950 40.342 52,620 13.506 5437 4385 1754 0

Tropinone Tropinone 6.0 (narrow) Tropinone Tropinone 6-Hydroxytropinone N-Methyl-4-piperidinone 6.8 -

-

Tetrahydrothiopyran-4-one

-

3-Quinuclidinone TBON~ 6.0 (broad) Tropinone Tropinone 6-H ydrox ytropinone N-Methyl-4-piperidinone -

Tetrahydrothiopyran-4-one

-

3-Quinuclidinone TBON~ Tropinone Tropinone N-Methyl-4-piperidone N-Propyl-4-piperidone 4-Pipendone 4-Methylcyclohexanone Tetrahydrothiopyran-4-one Nortropinone Eight other analogs

-

5.8 -

-

-

-

0.83

-

1.39

-

-

0.035 0.22

-

-

0.035

-

0.057 0.299 -

-

-

NADPH NADH NADPH NADH NADPH NADPH NADPH NADPH NADPH NADPH NADH NADPH NADPH NADPH NADPH NADPH NADPH NADH NADPH NADPH NADPH NADPH NADPH NADPH NADPH

None identified P-TBOLd a

-

~

TBONd

3

~

0.021 0.081 ~ 0.25

4 ~

d5

6

None identified

None identified

7

* Key to references (1) B. Drager and A. Schaal, in “Phytochemistry and Agriculture” (T. A. van Beek, ed.), p. 52. Wageningen Agric. Univ. (UNIPUB), Wageningen, 1992; (2) M. M. Couladis, J. B. Friesen, M. E. Landgrebe, and E. Leete, Phyrochemisrry 30, 801 (1991); (3) K. J. Koelen and G. G. Gross, PIanta Med. 44, 227 (1982); (4) A. Portsteffen, B. Drager, and A. Nahrstedt, Phytochemistry 31, 1135 (1992); ( 5 ) A. Portsteffen, B. Drager, and A. Nahrstedt, in “Phytochemistry and Agriculture” (T. A. van Beek, ed.), p. 63. Wageningen Agric. Univ. (UNIPUB), 1992; (6) B. Drager, A. Portsteffen, A. Schaal, P. H. McCabe, A. C. J. Peerless, and R. J. Robins, PIanfa 188, 581 (1992); (7) B. Drager, T. Hashimoto, and Y. Yamada, Agric. B i d . Chem. 52, 2663 (1988). TR 1 and TR I1 were not separated in these assays: levels were estimated from products ratios. Versus mixed activity in ammonium sulfate precipitate. For structure. see Scheme 9.

150

RICHARD J . ROBINS A N D NICHOLAS J . WALTON

(Scheme 9). A number of sulfoxides are also generated, presumably by nonenzymatic oxidations. TBON markedly diminished the accumulation of hyoscyamine in cultures to which it was fed (92). Using purified TR I and TR I1 from D . stramonium root cultures, it was shown that TR I1 is insensitive to TBON (93). TR I, in contrast, is able to metabolize TBON at a rate of about 30% that with tropinone and has a much lower K , of 0.035 mM. As the affinity for TBON is thus much greater than that for tropinone ( K , 1.1 mM), TBON acts as an effective inhibitor, reducing the rate of tropinone reduction by 70% at 0.5 mM. P-TBOL 0.02 m M ) and, to a lesser extent, a-TBOL 0.08 m M ) have also been found to be extremely effective inhibitors of TR I (93). These analogs may well prove useful in attempting to unravel the roles of TR I and TR I1 at this important branch point in the pathway. Causative evidence that pseudotropine leads to calystegins has been obtained from TBON-fed A . belladonna root cultures. In the presence of TBON at 4.2 mM, hyoscyamine accumulation is depressed 50%, whereas calystegin production simultaneously doubles (90).

8-~iabicyclo[3.2.110ctan-3-one (TBON)

QH

OH

8-Thiabicyclo[3.2.1]octan-3a-Ol (a-TBOL)

8-Thiabicyclo[3.2.l]octm-3~-ol (P-TBOL)

QH

0,C-CH3

d' 3-0-Acetyl a-TBOL

SCHEME 9. Structures of 8-thiabicyclo[3.2.Iloctan-3-one and metabolic products derived therefrom.

2.

BIOSYNTHESIS OF TROPANE ALKALOIDS

15 1

VI. Formation of Acidic Moieties of Tropeines

A. FORMATION OF PHENYLLACTIC A N D TROPIC ACIDS The incorporation of phenylalanine into hyoscyamine was demonstrated some time ago in D . stramonium plants by 14C-labelingstudies. Hydrolysis of the alkaloid confirmed the label to be entirely in the acidic moiety of the ester (94,95).More recent studies in a number of species, using both whole plants and root cultures, have confirmed this origin (2). Subsequent analyses using chirally labeled phenylalanine (96-98) or I3C-labeledintermediates (98-101) have shown that the carboxyl group undergoes a C-2’ to C-3’ shift during tropic acid formation. Cinnamic acid, the product of the irreversible action on phenylalanine of the well-described enzyme phenylalanine ammonia-lyase, was not incorporated (100,102,103), even though this is the initial step in the formation of many phenylalaninederived secondary products (68). It is probable, therefore, that the first step in phenylalanine metabolism in tropane alkaloid formation involves a transamination reaction, converting phenylalanine to phenylpyruvic acid. The subsequent reduction of this unstable keto acid to phenyllactic acid followed by a C-l’-C-2’ to C-1’- C-3’ shift will form tropic acid (see Scheme 10). The activation of phenyllactic and tropic acids, perhaps as the CoA thioesters (see Section VII,B), will enable their direct combination with tropine to form littorine and hyoscyamine, respectively. Support for this putative pathway has been obtained by feeding D . stramonium plants (RS)-[l4C]pheny1alaninealone or in combination with unlabeled phenylpyruvic or (RS)-phenyllactic acid (104). The latter, but, surprisingly, not the former, strongly inhibited [14C]phenylalanineincorporation into tropate esters of tropine. In contrast, unlabeled phenylalanine did not affect (RS)-[’4C]phenyllacticacid incorporation. Thus, it appeared likely that the rearrangement occurred at the level of phenyllactic rather than phenylpyruvic acid. The readiness with which these two compounds might be interconverted, however, makes it hard to establish the sequence of events by chemical labeling alone. A transamination reaction converting phenylalanine to phenylpyruvate at pH 8.0 using 2-oxoglutarate as amino group acceptor has been reported for extracts of D . stramonium suspension cultures and whole Datura plants (105).The activity extracted from plants was much higher than that from cultures. The nature of the products has not been demonstrated, however (106). If this reaction can be confirmed, it is possible that it

152

RICHARD J . ROBINS A N D NICHOLAS J . WALTON

~

~

OH

~

I

H Phenylalanine

1

Phenylppvic acid

H

O

O * i - * $ - S d A CHzOH Tropoyl-CoA

- c_‘>-

Phenylladc acid

OH CH,--F-*~--S-CoA I

H

O

F‘henyllactoyl-CoA

1 (-t

#H3

H CH*-y-*$-O I OH 0

Hyoscyamine

Littorine

SCHEME 10. Proposed pathway for the incorporation of phenylalanine into hyoscyamine and littorine.

represents a specific activity of tropane alkaloid metabolism, as in plants the pathway to phenylalanine proceeds via prephenate and arogenate, not phenylpyruvate as it can in bacteria (107). Obtaining further information about the enzyme has remained difficult, however, and it is only recently that a phenylalanine aminotransferase from a tropane alkaloid-producing species has again been reported, this time from transformed roots of H. albus (108). No further details are currently available. Aminotransferases showing activity with phenylalanine have been reported from plants of other genera (109), though their relevance to phenylalanine metabolism is unclear (see Refs. 107 and 110). To date, no enzyme responsible for the reduction of phenylpyruvate to phenyllactate has been described. This is somewhat surprising in view of the ease with which NADPH-linked oxidoreductases can be assayed; however, attempts to measure this reductase in H. albus root cultures have so far proved unsuccessful (111). Phenylpyruvic acid is readily incorporated into tropane alkaloids (112). It has been shown in root cultures of D.stramonium that [3’-*H2]phenylpyruvic acid is incorporated into both hyoscyamine and littorine with the predicted retentions of one and two 2H atoms/molecule, respectively.

,

2.

BIOSYNTHESIS OF TROPANE ALKALOIDS

153

Incorporation was determined by G U M S on a DB-17 column, which fully resolves these isomeric alkaloids (78). Similarly, the incorporation of phenyllactic acid into littorine and hyoscyamine has been clearly established (103). Woolley and colleagues acid was converted to showed that ( R S ) - [l’,3’-’3C,,3‘-’4C]phenyllactic hyoscyamine by whole plants of Datura (98,100).More recently, we have shown that ( R S ) - [1’ ,3’-’3C2]phenyllacticacid was incorporated into hyoscyamine in transformed root cultures of D.stramonium with a specific incorporation of about 60% (113). Similarly, this compound was incorporated into littorine, hyoscyamine, 6-hydroxyhyoscyamine, hyoscine, apoatropine, and apohyoscine by root cultures of a Datura hybrid (101) at levels of up to 65% specific incorporation. In both experiments, coupling in the tropate moiety between the 1’- and 3’-I3C nuclei of phenyllactic acid was shown by NMR, indicating that the predicted shift in carbon bonding had occurred. Only littorine showed no coupling, as expected. In parallel experiments in which ’H-labeled (RS)-phenyllactic acids were fed to D. stramonium and Anthocercis littorea root cultures, it was demonstrated that (RS)-[3-2H2]-,(RS)-[2-,H]-, and (RS)-[2-2H,3-2H2]phenyllactic acids all gave rise to ’H in littorine and hyoscyamine at the level predicted if this molecule were incorporated directly or following a C- 1’-C-2’ to C1’-C-3’ bond rearrangement (78). In Datura, the retention of the 2-% in hyoscyamine (23%) and littorine (18%) was similar, suggesting that phenyllactic acid was not metabolized via phenylpyruvic acid. Some loss of the 2-’H might be expected, however, as the phenyllactate dehydrogenase reaction is likely to be reversible. Thus, incorporation from the C2‘ might be lower than from the C-3’, but this has yet to be confirmed by ’H NMR. The analysis is further complicated by the possible loss of some 2-’H in hyoscyamine by racemization. Furthermore, we have found that both (R)-[l’,3‘-’3C2]-and ( S ) - [I ’,3’-’3C2]phenyllacticacids are incorporated into hyoscyamine by D . stramonium root cultures at equivalent levels (113). This may indicate that racemization of the phenyllactate takes place in uiuo during the feeding period. These apparently conflicting results need to be resolved. In cultures of D . strarnonium, it has been found that the (R) isomers of P-phenyllactic acid, mandelic acid, and 2-phenylpropionic acid are less toxic than the (S) isomers (78). This may indicate a greater ability to metabolize the (R) isoforms, since the toxicity of these compounds is usually a general effect of acidity. Interestingly, feeding either (RS) or (R)-phenyllactic acid increased the littorine level, but not the hyoscyamine content, of these cultures (78,113).The isolation of the enzymes concerned along with study of their properties in uitro will enable a clearer picture to be established of the exact pathway.

154

RICHARD J . ROBINS A N D NICHOLAS J . WALTON

Despite these uncertainties, however, it seems reasonable to argue that this pathway involves the metabolism of phenylalanine to phenylpyruvate and, subsequently, phenyllactate. In contrast, the route by which phenyllactic acid is metabolized to hyoscyamine is much less clear-cut. That the C-2’ of phenylalanine forms the carboxymethyl group of hyoscyamine was elegantly established by Leete and Louden (95). Exactly when, and how, the carbon group migration occurs is undescribed, as is the mechanism of ester formation (see Section VI1,B). What has been established is that the carbon shift does not involve the oxidation of the C-2’-C-3’ bond. The 3-2Hand 2-2Hlabels are both retained during the rearrangement, indicating that double bond formation has not occurred (78,97).The intermediacy of an epoxide has not, however, been ruled out. Recent experiments examining the incorporation of label from phenylpyruvic, phenyllactic, and tropic acids into tropoyl-tropanes have caused the putative role of free tropic acid in hyoscyamine biosynthesis to be questioned (see Section X,A). A considerable effort in enzymology is clearly required to establish the metabolic events occurring in the conversion of phenyllactic acid to hyoscyamine. It may be, for example, that the rearrangement in the side chain takes place after the activation of phenyllactic acid, possibly as the CoA thioester (Scheme 10). Not unexpectedly, the benzoic acid component of cocaine is derived from phenylalanine (124,115). Although, for example, the properties of phenylalanine ammonia-lyase (PAL), responsible for the conversion of phenylalanine to trans-cinnamic acid during cocaine biosynthesis in E. coca, are likely to be similar to those of PAL characterized from other plant sources (116), there is no specific evidence on this point. B. FORMATION OF OTHERESTERIFYING ACIDS Besides tropic and phenyllactic acids, a wide range of other acids are found esterified with tropane bases (4). These range from such common and simple compounds as formic and acetic acids to highly unusual molecules, such as the sulfur-containing 1,2-dithiolane-3-carboxylicacid, the esterifying acid in brugine, an alkaloid of the Rhizophoraceae (117). The structures of the esterifying acids found in the tropane alkaloids have been thoroughly reviewed previously (4) in this treatise (see also Chapter 1, this volume) and need not be repeated here. Limited labeling evidence is available on the origin and likely biosynthetic pathway of these acids. Tiglic acid, the acidic component of tigloidine and meteloidine, has been shown to be derived from isoleucine, probably via 2-methylbutanoic acid (Scheme 11) (2,118-120) and not via trans-2,3-dimethylacrylic (angelic) acid, as is apparently the route for

2.

Angelic acid

BIOSYNTHESIS OF TROPANE ALKALOIDS

Tiglic acid

155

2-Hydroxy-2-methylbutanoic acid

SCHEME 1 1 . Pathway of tiglic acid formation from isoleucine.

heliosupine formation in Cynogfossum(121). N o specific biochemical characterization at the cell-free level has been reported, however, and, where a particular reaction or sequence of reactions is not shared with a wellcharacterized pathway, no information at all is available directly. In the case of tiglic acid formation, this reflects the lack of detailed characterization of the catabolic pathway for isoleucine in plants, though this pathway has been well studied in animals and bacteria (122). It is likely that the final stage of esterifying acid addition involves the formation of CoA thioesters, since, in the few cases where evidence is available, the esterifying reactions use the CoA esters as substrates, as discussed below (see Section VII).

VII. Formation of Tropeines The reactions described in Sections V and VI result in the biosynthesis of the two intermediates that constitute the alkamine and acidic moieties which are brought together as an ester in the tropeines. The reactions by which the ester bonds are formed have remained essentially uncharacterized until recently. A number of reported properties of esterases appear more related to tropane ester hydrolysis (see Section IX). It has now been demonstrated, however, that a number of esters can be formed in uitro by acyltransferase reactions involving the transfer of the acidic group

156

RICHARD J. ROBINS A N D NICHOLAS J . WALTON

from the relevant coenzyme A thioester to tropine or pseudotropine. The reactions by which the CoA thioesters are formed have yet to be described. ACIDS A. ESTERSOF ALIPHATIC Although in D. stramonium root cultures (see Section II,B) the dominant alkaloid is hyoscyamine, the roots accumulate substantial quantities of acetyltropine in response to feeding with tropine or tropinone (21). Callus cultures of D. innoxia also accumulate acetyltropine when fed tropine (123, whereas suspension cultures of Duboisia myoporoides produce both acetyltropine and isobutyryltropine (25). Similarly, some enhanced tiglyltropine formation occurs when tiglic acid is fed to a Datura hybrid (22). The ability of these cultures to make high levels of acetyltropine led us to investigate the acetyl-CoA: tropine acyltransferase activity present in D. strumonium root cultures in the anticipation that, by understanding the properties of the enzyme, it might be possible to define the conditions required to assay hyoscyamine biosynthesis successfully. A preliminary analysis of extracts of 10-day-old roots indicated that enzymes able to esterify tropine and pseudotropine with acetyl-CoA were present (124). Surprisingly, considering that the a series of tropane alkaloids strongly dominates in this species, the activity with pseudotropine was 5- to 10fold higher than that with tropine. The acetylation of tropine and pseudotropine by extracts of D. stramonium roots shows maximal activity at pH 8.5-9.5 and only about 20% of this level at pH 7. This may indicate that tropine needs to be in the neutral state to act as substrate for the enzymes. The reaction transfers an acetyl unit to the C-3 hydroxyl of the substrate, and the nature of the products has been confirmed by cochromatography of the products derived by incubating ['4C]tropine or ['4C]pseudotropine with unlabeled acetyl-CoA with the products formed by incubating [ I-'4C]acetyl-CoA with tropine or pseudotropine (124). On G U M S , the products of tropine or pseudotropine acetylation co-elute with authentic synthetic standards and show the same fragmentation characteristics. Preliminary studies of the two activities show there to be two separable enzymes present (124). It is possible to obtain a fraction from anionexchange chromatography containing only the acetylpseudotropineforming activity. The tropine-acetylating activity has not, however, been obtained free of any pseudotropine-utilizing enzyme. In crude extracts it shows apparent K , values of 0.22 and 0.24 mMfor acetyl-CoA and tropine, respectively. The observed acetylation of tropine in suspension cultures of D. innoxia (123) has also been further examined. Unpublished studies (125) of this

2.

BIOSYNTHESIS OF TROPANE ALKALOIDS

157

system showed an activity to be present that acetylated tropine with acetyl-CoA and which peaked at the maximum growth rate of the culture. The product was confirmed by crystallization to constant specific activity to be 3-acetyltropine. In contrast to the D. sframonium enzyme, activity appeared to be highest at pH 7.5. Whether this represents a different enzyme activity from that found in D. sframonium roots has yet to be resolved. A further activity that forms P-tigloxyltropane (tigloidine) from tiglylCoA and pseudotropine has also recently been found in root cultures of D. stramonium ( I 26). Again, although tigloxyltropane extracted from roots of this species is predominantly (-80 : I ) the 3a stereoisomer, the activity that forms the 3p isomer is present at high levels in the extracts. Only a trace of 3cu-forming activity can be detected. The enzyme has been purified about 100-fold by sequential ammonium sulfate precipitation, hydrophobic interaction chromatography, anion-exchange chromatography, and chromatofocusing. The activity that forms 3-acetylpseudotropine copurifies with the 3-tiglylpseudotropine-formingenzyme to about the same level of purification. At present, it appears that a single enzyme may be responsible for both activities, based on the copurification and kinetic experiments that indicate competitive kinetics between acetyl-CoA and tiglyl-CoA. The enzyme shows a K , of 0.30 mM for acetyl-CoA, similar to that of the tropine-acetylating enzyme. The K, of 1.1 mM for tiglylCoA is rather higher, but the V,,, is very much greater. The similarities of the K , values for pseudotropine (0.34 mM with tiglyl-CoA and 0.36 mM with acetyl-CoA) supports the conclusion that this is a single activity. The enzyme shows a high degree of specificity for the alkaloidal substrate, being active only with pseudotropine (100%) and 4-hydroxy-lmethylpiperidine (14%). In this respect it is dissimilar to TR I and TR 11, with the latter enzymes showing a broader substrate specificity. With the acyl-CoA thioester, however, a broad range of substrates was utilized, although all at rates of less than 10% that of tiglyl-CoA. The range of acyl groups accepted could indicate that the wide range of aliphatic esters found (4) is derived from the activities of a small number of broad-specificity acyltransferases. The differences in the apparent affinities of the 3a-forming and 3pforming enzymes for the CoA thioesters and 3-hydroxyltropane bases may also be an important factor in determining the alkaloidal spectrum formed in uivo. In D. stramonium, TR I activity is 5- to 10-fold higher than that of TR I1 (82,87),and the tropine to pseudotropine ratio is typically about 30 : 1. Thus, even though the total potential enzyme activity as determined in uitro is much lower for 3cu-acetylation than for 3p-acetylation, the availability of the substrate, coupled with the slightly greater affinity of

I58

RICHARD J . ROBINS A N D NICHOLAS J . WA LTO N

the 3a-acetylating enzyme for acetyl-CoA or of the 3P-enzyme for both its substrates, establishes conditions in which the 3a-acetyl is the favored product. A similar situation may arise with 3-tigloxyltropane formation, but this analysis awaits the description of the tiglyl-CoA:tropine acyltransferase enzyme.

B. HYOSCYAMINE A N D OTHERAROMATIC ESTERS The formation of the tropoyl ester of tropine, hyoscyamine, is, in many species, quantitatively the dominant metabolic reaction involving tropine esterification in v i m . Investigations in the early to mid 1960s claimed to measure ester formation, but these results have subsequently failed to be substantiated. Extracts of whole leaves of wild D. srramonium plants and of 2- to 3week-old cultured seedlings were used by Kaczkowski to prepare enzyme extracts (127). Following ammonium sulfate fractionation and DEAEcellulose chromatography, he reported the separation of a tropine esterase (see Section IX) from a synthesizing activity. The 75-100% saturated ammonium sulfate cut was described as having an “irregular but considerable” activity. When the desalted extract was incubated at pH 6 (0.2 M phosphate buffer) with 6 p M tropine and 6 p M tropic acid, 1 mg/1.7 ml ATP, and traces of Mg2+and Mn2+,up to 3% of the substrate was reported to have been converted to hyoscyamine. The reaction was optimal at pH 6, 25”C, and with 6 p M substrates, higher levels causing a loss of activity. The report concluded that “atropine synthase” had only been found because of the successful separation of this enzyme from atropine esterase, which was reported present at a 7-fold higher level. There appear, however, to be a number of problems associated with the findings of Kaczkowski, and later workers have had difficulty reproducing the results. The extracts were made from the leaves, yet synthesis is known to occur in roots. The “irregular” nature of the occurrence of hyoscyamine might indicate contamination with alkaloid from the tissue, which we know can be carried through a number of purification stages tightly bound (nonspecifically) to protein. Yet the rates Kaczkowski obtained tend to preclude this possibility. Another reason to question these reports, however, is the improbability that tropine and tropic acid, even in the presence of MgATP, are combined directly. Evidence obtained subsequently from numerous other biological ester-forming systems shows that the acid is normally activated, frequently as the CoA thioester. Jindra and co-workers recognized a number of these deficiencies and reinvestigated this work. They incubated the expressed sap of D.srrumonium roots with tropine and tropic acid, but without effect (128). The addition of CoA and ATP to the reaction mixture did not stimulate biosyn-

2.

159

BIOSYNTHESIS OF TROPANE ALKALOIDS

thesis (129). Subsequently, however, in a brief communication, success was claimed using extracts made from 27- or 31-day-old D. stramonium suspension cultures (105). Expressed juice was concentrated 10-fold by lyophilization and incubated with 30 mM tropic acid, 30 mM tropine, 5 mg/ml ATP, and 0.15 mg/ml CoA at pH 7. After 6 to 48 hr, samples were analyzed and found to contain hyoscyamine by cochromatography. The amount present appeared to increase with time, and no product was found in boiled extracts or when reactions were incubated at pH 5.3. Details of these experiments have not been published (106), and neither has the report, made simultaneously, that the suspension cultures would convert added tropine and tropic acid to hyoscyamine. In view of the findings of later workers, both these claims need to be substantiated by further analysis. The potential role of tropoyl-CoA in hyoscyamine formation led to the development of a synthetic route for this putative intermediate in which tropic acid is first activated as the N-hydroxysuccinimide ester (yield 50%) and tropoyl-CoA made by transesterification with free CoA (yield -35%) (130).Incubations made with this substrate, however, have proved ineffective (83,131,132). Nevertheless, it is likely that a CoA thioester is involved. The role of thioesters in the acyl transfer of aromatic groups is well established for the phenylpropanoid pathway (68). In recent work, an enzyme activity in transformed root cultures of D. stramonium has been identified that acylates pseudotropine with phenylacetate from phenylacetyl-CoA and with benzoate from benzoyl-CoA (132). Furthermore, when labeled benzoic acid was activated as the S-(2acetamidoethyl)ester, incorporation rates into cocaine were greatly improved (133). Interestingly, N-methylecgonine was not benzoylated by the extract from D. stramonium, indicating that a quite separate enzyme, able to accept the presence of the 4-carboxyl group, is probably present in Erythroxylon species. An intriguing possible alternative route was investigated by McGaw and Woolley (134). They argued that esterification might occur prior to cyclization if hygrine were reduced first to hygroline (Scheme 12). The

@ - @ - @ H3C,N,6 C II 0

Hygrine

N = H3C\yC

H I ‘OH

Hygroline

N : H3C\C/C

H

-

\

/,c - CH3

C - CH3

H’ ‘ W C - 7 II 0 CH3

Tiglylhygroline

It

0

\ CH3

3-Tigloyloxytopane

SCHEME 12. Proposed hygroline/tiglylhygnne pathway.

160

RICHARD J . ROBINS AND NICHOLAS J . WALTON

hypothesis was tested by feeding intact D . meteloides plants with [1',2'''C2]tigloyl hygrine and found not to be valid, as the 1 ' : 2' ratio of I4C was not maintained in the tigloxytropane esters, as shown by degradation.

VIII. Metabolism of Tropeines A. CONVERSION

OF

HYOSCYAMINE TO HYOSCINE

In many whole plants, and some root cultures, the major product that accumulates is not hyoscyamine, but the 6,7-P-epoxide, hyoscine. The conversion of hyoscyamine to hyoscine is the most thoroughly investigated part of the pathway at the biochemical level, with a series of studies since the late 1980s having resulted in the purification of the pertinent activities, the clear demonstration of the mechanism of the reaction, and the cloning of a gene. This work demonstrates effectively how biochemical studies can clarify uncertainties that cannot be resolved by chemical labeling studies alone. Prior to this biochemical analysis, the route by which the conversion occurred was not clear. Feeding experiments with 6,7-dehydrohyoscyamine in a Daturaferox scion showed that this compound was efficiently incorporated into hyoscine, but not hyoscyamine (135). When [N-methyl14C,6P,7P-3H2]tropine was fed to whole plants of D . innoxiu and D . metebides, both 3Hatoms were lost (136), supporting the proposed intermediacy of the 6,7-dehydro compound. Two alternative pathways were proposed (see Scheme 13). One pathway suggested that hyoscyamine is first dehydrated and then hydrated to 6-hydroxyhyoscyamine, this subsequently being oxidized to form the 6,7-epoxide (137);the other pathway hypothesized that 6-hydroxyhyoscyamine is formed directly from hyoscyamine followed by dehydration to 6,7-dehydrohyoscyamine and subsequent conversion to hyoscine. A flaw in both schemes was the complete absence of the putative dehydro intermediate in alkaloid extracts made from hyoscine-forming species. In any event, neither scheme proved correct, although it is the case that the first event is the hydroxylation of hyoscyamine to form the 60hydroxy derivative. The correct sequence of reactions can be assigned on the basis of the enzymology. In 1986, Hashimoto and Yamada (138) described an enzyme (hyoscyamine 6P-hydroxylase, H6H) in extracts of root cultures of Hyoscyamus niger that would carry out this reaction. The enzyme shows an absolute requirement for molecular oxygen, Fez+,2-

2.

BIOSYNTHESIS OF TROPANE ALKALOIDS

161

6,7-~ehydmhyoscyamine

SCHEME 13. Alternative routes for the conversion of hyoscyamine to hyoscine.

oxoglutarate, and hyoscyamine. I n uitro,the activity is also stimulated 3to 6-fold by ascorbic acid, but it is not clear whether this effect is due to its acting to maintain the iron in its Fe(1I) oxidation state or is due to some other effect on the enzyme reaction mechanism. The reaction shows a I : 1 stoichiometry between the production of 6P-hydroxyhyoscyamine, as determined by GC, and the decarboxylation of 2-oxoglutarate to succinate, as determined by the loss of 14C02from 2-[l-'4C]oxoglutarate. The reaction product was confirmed to be (-)-6P-hydroxyhyoscyamine by G U M S , NMR, melting point, and ORD. The presence of this activity correlated well with the ability of cultures to make hyoscine de nouo. It shows variable activity with the age of the cultures, showing a peak of activity about 7 days after switching the cultures to IBA-free medium (139). The H6H in Datura similarly shows a peak during the growth of the cultures (22). Following a purification of about 300-fold (17% yield) by sequential hydrophobic interaction, ion-exchange, and hydroxyapatite chromatographies, the properties of the enzyme were described in detail (139). The enzyme appears to be a monomer with a molecular weight of about 41,000. The activity is maximal at about pH 7.8 and shows a moderately high affinity for both its organic substrates, with a K , of 35 p M f o r hyoscyamine and a K , of 43 p M for 2-oxoglutarate. Although the specificity for 2oxoglutarate was very high, hyoscyamine could be replaced by a number

162

RICHARD J . ROBINS A N D NICHOLAS J . WALTON

of other tropanes, some of which were inhibitors of hyoscyamine hydroxylation (Table IV). The enzyme showed good activity with a number of nor compounds, and there appeared little requirement for the carboxymethyl group of the tropate moiety. The nonnatural analog homatropine was readily hydroxylated. In all cases, the enzyme showed complete regiospecificity, the reaction placing a hydroxyl only at the C-6 position. It also showed an absolute requirement for the ester to be 3a and to have the d configuration. With the exception of Mg2+and Ca*+,the enzyme appeared to be completely inhibited (>95% at 0.4 m M ) by all other divalent metal ions tested. Surprisingly, however, in view of the putative role of ascorbate, Fe3+inhibits activity only by about 30%. Like other known 2TABLE IV SUBSTRATE SPECIFICITY OF HYOSCYAM~NE 6P-HYDROXYLASE FOR ALKALOIDAL SUBSTRATES" Relative activity Substrate

(9%)

Product

I-Hyosc yamine Tropine Acetyltropine lsobut yry ltropine Apoatropine I-Norh yosc yamine Noratropine-N-acetic acid Noratropine-N-butyric acid Atropine methyl bromide Phenylacet yltropine 2-Phenylacrylylpseudotropine Phenylalanyltropine tert-Cinnamoy ltropine

100

( - )-6P-Hydroxyhyoscyamine

p-H ydroxyatropine I-Homatropine 2-H ydrox y-3-phen ylpropion y ltropine 3-H ydrox y-3-phenylpropionyltropine 6.7-Dehydrohyoscyamine

0 1

I5 45 81 17 0 1

81

3 8 39 26 81 15 56 119

-

nd nd 6-H ydrox yapoatropine 6-H ydrox ynorh yosc yamine nd

-

nd 6-H ydrox yphen ylacet yltropine nd nd terr-Cinnamoyl-6h ydroxytropine nd 6-H ydrox yhomatropine 2-H ydroxy-3-phenylpropion yl6-h ydrox ytropine 3-H ydrox y-3-phenylpropionyl6-h ydrox ytropine Scopolamine

Inhibition (%Y ndd nd nd nd 29 nd 19 nd nd 12 nd nd nd nd 8 nd

nd 32

" Data from T. Hashimoto and Y. Yamada, Eur. J . Biochem. 164, 277 (1987): Y . Yamada and T. Hashimoto, Proc. J p n . Acad. 65B, 156 (1989). Activities (relative to that observed with I-hyoscyamine) were measured in the presence of each alkaloid at 0. I mM. Percentage inhibition of I-hyoscyamine hydroxylation with an inhibitor concentration of 0.2 m M . nd, Not determined.

2.

BIOSYNTHESIS OF TROPANE ALKALOIDS

163

oxoglutarate/Fe2+-dependentdioxygenases, H6H is totally inhibited (>92% at 0.1 mM) by superoxide scavengers such as nitro blue tetrazolium. The availability of purified hyoscyamine 6P-hydroxylase enabled studies to be performed that clarified the aberrant metabolism of 6,7-dehydrohyoscyamine previously observed. In uitro, this compound proved to be an excellent substrate for H6H, being converted to hyoscine. Thus, it was still conceivable that 6,7-dehydrohyoscyamine was an intermediate in viuo. This possibility was eliminated, however, by examining the metabolic fate of I8O in [6-'80]hydroxyhyoscyamine (140). The labeled intermediate was prepared by incubating I8O2with an extract of H6H from H . niger. An incorporation of 85% was obtained based on the isotope ratios determined by derivatized G U M S analysis. The metabolism was tested in shoot cultures of Duboisia myoporoides, themselves unable to make hyoscine but previously demonstrated to convert hyoscyamine efficiently to 6-hydroxyhyoscyamine and hyoscine (26). The hyoscine extracted from the shoots after 6 days of feeding showed an isotopic ratio of 84.6%, indicating that complete retention of the 6-0 occurs during epoxidation. This finding fully supports that of Leete and Lucast (136). Furthermore, when [7P-*H]6P-hydroxyhyoscyamine was fed to D . myoporoides shoot cultures, all the 2H was lost on conversion of the substrate to hyoscine (141).

What has proved much more difficult to establish, however, has been the nature of the epoxidase enzyme. Experimentally, it has proved impossible to separate the hyoscyamine-hydroxylating activity from the 6-hydroxyhyoscyamine epoxidase activity (139,142). The latter enzyme shows a similar absolute requirement for 2-oxoglutarate and Fe", stimulation by ascorbate (about 3-fold), and sensitivity to divalent metal ions. During molecular exclusion chromatography, the activities were found to coelute at 41,000 D, and the two activities eluted in the same fractions from both hydrophobic interaction and ion-exchange columns. In the course of purification, however, the epoxidase activity was substantially lost. In a crude extract, the ratio of the specific activities was 2.3 : 5 . 3 in favor of the hydroxylase, but after purification a ratio 3.3 : 255 was observed (139). Thus, the epoxidase appears to be much less stable than the hydroxylase. Nevertheless, an indication that the two activities may be associated with a single protein is that both hyoscyamine 6P-hydroxylation and 6,7dehydrohyoscyamine epoxidation were found to be inhibited in parallel by a monoclonal antibody raised to purified H6H protein (143,143~).Furthermore, when the H6H clone (see Section XI) is expressed in transgenic, hyoscyamine-rich A . belladonu plants (83,1449,the major product accumulated is hyoscine, not hyoscyamine, as found in control plants. This could

164

RICHARD J. ROBINS A N D NICHOLAS J. W A L T O N

result from an abundance of epoxidase activity in the plants and a deficiency of only the hydroxylase. However, when the clone was expressed in Nicotiana tabacum, which cannot normally make these products, hyoscine accumulated in response t o feeding hyoscyamine or 6-hydroxyhyoscyamine (83), indicating that epoxidase activity was present. Thus, it appears that both activities are indeed the property of a single protein.

B. FURTHER ESTERIFYING REACTIONS The introduction of a 6p-hydroxyl into tropane alkaloids occurs in many instances other than hyoscyamine (4). Because hyoscyamine 6phydroxylase does not act on tropine as a substrate (139), it is apparent that other enzymes must be present that catalyze the formation of 6pand 7p-hydroxyltropanols and esters. Although no enzymology is available for the formation of these compounds, some interesting indications of possible activities are available. By examining the results from D. innoxia and D. meteloides plants (145-147) challenged with 3c-w-tigloxyltropane labeled with I4C in both the alkamine and acidic moieties and with I4Clabeled tiglic acid, and from the findings of competitive feeding experiments, it proved possible to propose an ordered metabolic sequence to the di- and trioxygenated tropanes and, in particular, to deduce which reactions were unlikely to take place. It is also conceivable that the further oxygenation actually occurs prior to reduction of the 3-one, since 6P-hydroxytropan-3-one is a substrate for both TR I and TR I1 (87). When extracts of D. stramonium root cultures were incubated with 6phydroxytropan-3-one and tiglyl-CoA, no 6/3-tigloxyltropan-3-onecould be detected, however (132). Unraveling these complex hydroxylations and tigloylations will prove extremely challenging.

IX. Degradation and Oxidation of Tropeines

In contrast to the increasing information on the biosynthesis of the tropane alkaloids, their degradation remains a comparatively neglected field. To obtain a clear picture in whole plants, it is necessary to distinguish between degradation and translocation. If the total alkaloid content of a plant is not determined, it cannot be concluded that a decrease in the alkaloid content of a particular organ does not simply reflect the movement

2.

BIOSYNTHESIS OF TROPANE ALKALOIDS

165

of alkaloid to other sites within the plant (see Section X). Convincing evidence of the occurrence of tropane alkaloid degradation has, however, been obtained both in studies using isolated organs and, perhaps more satisfactorily, in experiments using labeled alkaloids wick-fed to whole plants. Neumann and Tschope (148) reported evidence for the degradation of alkaloids in plants of a number of solanaceous species, including Atropa belladonna, Withania somnifera, Cyphornandra betacea, and Nicotiana glauca. There appeared to be substantial degradation of atropine fed to isolated leaves of Withania and Cyphomandra, in both the light and the dark. When [ l’-I4C]atropine was fed to isolated leaves of Cyphomandra betacea, almost 45% of the label was recovered after 24 hr in neutral organic substances. It was therefore suggested that, following ester hydrolysis, an early step in atropine degradation might be the decarboxylation of the resulting tropic acid. Hamon and Youngken (149) later investigated the degradation of [3H]atropine in mature D. innoxia plants. They found that almost 60% degradation of the alkaloid occurred during the first day and that after 20 days only 1.5% of the label remained in the alkaloid fraction. Because this degradation could also be shown in maturing plants, the total alkaloid contents of which were increasing, it was concluded that synthesis and degradation might occur simultaneously. Alkaloid degradation has not yet been studied directly in transformed root cultures. Circumstantial evidence for its occurrence has come from alkaloid determinations in D. strarnoniurn cultures that have been induced to differentiate by being subcultured in media containing NAA and kinetin (23; see also Section X,A). A rapid decline in alkaloid content occurs which, in apparent contrast to the situation in cultures of Nicotiana rustica (150), cannot be ascribed entirely to dilution of the alkaloid present at subculture. It is also noticeable that the ratio of alkaloids changes; in particular, there is a much greater loss of hyoscyamine than of 3-acetyltropine (see Section X,A). Data from these experiments are summarized in Table V. It is probable that the initial step in tropane-alkaloid degradation is ester hydrolysis. Cleavage of atropine to tropine and tropic acid was studied by Jindra and co-workers (128,151). Esterase activity catalyzing this reaction was found in the sap prepared from roots of D. strarnonium plants; the highest activity was obtained from plants approximately 24 weeks old, at a time of low alkaloid accumulation. No activity was found in the root sap of 9-week-old plants or in the sap from stems or leaves. The pH optimum was between 5.0 and 5.8, and the reaction was inhibited by physostigmine. The extract would also hydrolyze homatropine and nova-

TABLE V CHANGESI N ALKALOIDCONTENTOF Datitra stramoniitm ROOT CULTURESI N RESPONSETO MEDIUMCONTAINING PLANTGROWTHREGULATORS~ ~

Treatment*

B5O

NK5

a

Time from subcultureC (days)

HYg

cus

Tro

Tri

ACT

DaT

PaT

Apo

Hyo

Total

0 8 14 5 8 14

0 39 0 0 0

38 66 0 0 1

15 18 5 0 0 4

110

65 370 21 1 194 98 8

20 87 131 7 4 2

-

280 29 229 9 6 2

850 800 1490 117 56 13

1380 1393 2546 340 166 29

Alkaloid content (nmolig FW)d

30 203 8 2 0

19 49 0 0 0

Data from Robins et a[. (23).

* B50 medium contained Gamborg’s B5 salts with the addition of 3% sucrose: NK5 contained, additionally, 2.0 mgiliter a-naphthaleneacetic acid and 0.2 mgi liter kinetin. The initial subculture was made with roots that were 14 days old. Hyg, Hygrine; Cus, cuscohygrine; Tro. tropinone: Tri. tropine: ACT.3-acetyltropine: DaT. 3.6-diacetyltropine: PaT, 3-phenylacetyltropine; Apo, apohyoscyamine; Hyo, hyoscyamine. The total includes some further minor alkaloids.

2.

BIOSYNTHESIS OF TROPANE ALKALOIDS

I67

tropine, although cocaine was not attacked. Esterase activity and its possible relationship with hyoscyamine synthesis were also studied by Kaczkowski (127; see Section VI1,B). Cosson and Paris (152) estimated hyoscyamine and scopolamine esterase activity in extracts made from the roots and aerial parts of Datura tatula plants at various stages of development. They found that root extracts generally gave a higher level of activity, whereas in extracts of aerial parts the greatest activity could be demonstrated prior to flowering; during fruiting, the activity disappeared from the aerial organs but persisted in the roots. The activity of atropine esterase in intact roots and cultured roots of various solanaceous tropane alkaloid-producing species has recently been thoroughly documented by Kitamura and colleagues (153). Appreciable levels of activity were found in roots of plants of Darura farufa,Hyoscyamus niger, Atropa belladonna, Scopolia japonica, and Duboisia leichhardtii. The highest activities were found in roots taken from fruiting plants of D.tatula and H . niger; roots taken from growing plants of these species in full leaf, or from flowering plants, gave activities of around 15% of these values. Roots from plants of A . belladonna, whether growing, flowering, or fruiting, gave broadly similar values intermediate between these limits. In no case were measurable activities found in the leaves or stems of plants. Furthermore, in contrast to the activities found in normal plant roots, no activity was detected in cultured roots of these species, despite the presence of substantial accumulations of alkaloids. It appears that the presence of stems and leaves is essential for the development of atropine esterase activity, suggesting that the function of the enzyme may be the hydrolysis of alkaloid transported from the aerial parts back to the roots. In support of this, atropine esterase in regenerated plantlets of Duboisia myoporoides was initially very low in relation to that of seedlings and appeared to be correlated with a very low alkaloid concentration in the leaves of the plantlets (154). The picture that emerges, therefore, is one in which the roots are likely to be the chief sites of both alkaloid synthesis and degradation, with alkaloids being transported to and from the aerial parts as required in response to developmental or other factors (see Section X). At present, however, the metabolic fate of the basic and acidic fragments of hydrolysis remains largely uninvestigated. It would appear that, to some extent, they are reutilized for alkaloid biosynthesis. When littorine containing I4Clabel in the acidic moiety and 3Hlabel in the alkamine moiety was fed to Datura innoxia plants, both isotopes became incorporated into hyoscyamine (102). The initial ratio of 3H to I4C was not maintained,

168

RICHARD J. ROBINS A N D NICHOLAS J . W A L T O N

however, indicating deesterification of the littorine followed by the differential utilization of the two resulting fragments. A number of tropane alkaloid-producing genera form small quantities of nor-type alkaloids in which the nitrogen atom is unmethylated (59). In principle, these compounds could be produced directly from putrescine, via pyrroline, without methylation (see Section IV,A). Alternatively, however, they might be produced by demethylation of the methylated alkaloids (e.g., norhyoscyamine from hyoscyamine), by analogy with the formation of nornicotine from nicotine, shown to occur in leaf extracts of Nicotiana species (155,156). Evidence on this possibility is presently lacking. Either way, however, these compounds can hardly be regarded as degradation products, and, furthermore, it is doubtful whether they are any more susceptible to extensive degradation than their methylated counterparts. A quite general reaction of alkaloids is N-oxidation. Phillipson and Handa (157) isolated the two isomeric N-oxides of hyoscyamine from species of Atropa, Datura, Hyoscyamus, Scopolia, and Mandragora and one N-oxide of hyoscine from species of the first four genera. Substantial amounts of the hyoscyamine N-oxides were present in A. belladonna (between 5 and 40% of the amount of tertiary base), with particularly large quantities being present in the seeds of mature fruits. In view of the insolubility of the N-oxides in diethyl ether, it is possible for their occurrence to be overlooked in routine alkaloid extraction procedures. Nothing is known, however, of the further metabolism of these compounds, or whether they represent end products more or less stable than the tertiary bases from which they are derived. Interestingly, in Senecio vulgaris root cultures, the major stable end product accumulated is senecionine Noxide (158). Furthermore, it appears that it is in this form that the alkaloid is translocated in the plant (41).

X. Overall Regulation of Pathway Alkaloid biosynthesis is regulated, indeed made possible, by organizational features at the cellular and tissue levels that ensure that an ordered sequence of reactions occurs and that appropriate relationships are maintained with other metabolic processes. In contrast, at the level of the whole organism, alkaloid production is affected and regulated by a wide range of nutritional and environmental factors. The mechanisms relating such influences at the whole-plant level with their consequences at the biochemical level are the province of both physiologists and molecular

2.

BIOSYNTHESIS OF TROPANE ALKALOIDS

169

biologists and pose problems that are particularly difficult to identify and to solve. AT BIOCHEMICAL LEVEL A. REGULATION

With the exception of hyoscyamine 6P-hydroxylase (and its associated activities), which is known in roots to be present specifically in the pericycle (143; see Section VIII,A), no information is yet available on the tissue or subcellular localization of enzymes of tropane alkaloid biosynthesis. Progress on both of these questions needs to be made before a full understanding of the organization of the pathway can be achieved. As a result of recent investigations, however, insights into the biochemical regulation of the pathway are beginning to emerge. Simple feeding experiments with metabolites of the pathway supplied to transformed root cultures of Datura species have indicated likely limiting steps in hyoscyamine formation. Feeding individually putrescine, agmatine, Nmethylputrescine, tropinone, or tropine to root cultures of D . stramonium was, in each case, largely ineffective in increasing hyoscyamine formation (Table VI) (21). On the other hand, substantial increases in the production of hygrine, cuscohygrine, tropine, and 3-acetyltropine were found after feeding the amine precursors, as well as increases in tropine and 3acetyltropine after supplying tropinone; tropine was also metabolized to 3-acetyltropine (Table VI). Broadly consistent data from cultures of the D. candida X D . aurea hybrid have also been obtained (22). Of special interest is the observation that exogenous tropinone was entirely converted to tropine and 3-acetyltropine virtually without stimulating the production of hyoscyamine. Even when fed at 5 mM, tropinone is totally reduced within 48 hr. This suggests that the reduction of tropinone is rapid in relation to hyoscyamine formation and unlikely to be of regulatory significance. This conclusion is further supported by the presence of substantial tropinone reductase activities in transformed root cultures of both D. stramonium (82,87) and the D . candidu X D . aurea hybrid (22; see Section V). It may also be inferred from Table V1 that the formation of 3-acetyltropine occurs in response to a surfeit of tropine. Its seemingly adventitious formation may thus bear little direct relationship to the mechanism of hyoscyamine formation (see Section VII). This interpretation is also consistent with the effect of feeding TBON, which is metabolized to the 3-O-acetyl derivative of the corresponding alcohol, TBOL (see Scheme 9) (92). From these results it might be deduced that the supply of tropic acid is likely to be an important limitation to hyoscyamine production. This has been tested in a number of systems by feeding free tropic acid

TABLE VI EFFECTS OF FEEDING TROPINE A N D METABOLIC PRECURSORS ON INTRACELLULAR ALKALOID LEVELS I N TRANSFORMED ROOT CULTURES OF Datura stramoniuma

Alkaloid content (nmol/g FW)c Experiment I

Compound suppliedb None Putrescine Agmatine

2

None N-Methylputrescine Tropinone Tropine

a

Concentration (mM)

Growth (g FW)

Hyg

Cus

Tro

Tri

ACT

DaT

Apo

Hyo

1 .O 5.0 1.o 5.0 1 .O 5.0 1 .O 5.0 1.o 5.0

6.7 7.4 7.1 7.8 6.8 9.5 8.2 9.0 9.2 9.5 10.1 12.2

7 58 98 28 89 7.8 98.5 88.5 6.6 2.8 7.6 10.4

9 6 278 83 344 10.6 306.7 237.6 5.6 14.2 7.1 20.7

ndd nd nd nd nd 4.5 9.8 7.1 5.9 5.9 2.8 4.3

150 520 775 357 504 65 46 1 290 213 1469 592 1322

90 289 419 34 1 49 1 82 33 1 235 254 1425 460 78 1

nd nd nd nd nd 66 I07 74 65 90 65 91

278 443 157 265 128 218 137 106 260 181 162 154

1205 1314 1271 1472 670 1734 1721 1906 1698 2019 2034 2039

Data from Robins ef a / . ( 2 1 ) . Additions of compounds were made to roots at subculture, and alkaloids were determined 21 days later. Hyg, Hygrine; Cus. cuscohygrine; Tro, tropinone; Tri, tropine. ACT. 3-acetyltropine; DaT. 3,6-diacetyltropine; Apo, apohyoscyamine; Hyo, hyoscyamine. nd, Not determined.

2.

BIOSYNTHESIS OF TROPANE ALKALOIDS

171

(21,22,123,159,160). The effect has been variable, but, in the main, no

enhanced tropoyltropane levels were obtained. This could simply indicate that a further step is limiting, for example, the activation of tropic acid. In some instances tropic acid was detrimental to hyoscyamine production. For example, when fed at 1-2 mM to root cultures of D. stramonium, exogenous (RS)-tropic acid caused a 60% reduction in hyoscyamine accumulation. In contrast, 1-2 mM (RS)-phenyllactic acid (while also failing to stimulate hyoscyamine production) did not have the same inhibitory effect (21,f13). (RS)-Phenyllactic acid did, however, enhance the formation of littorine, causing a change in the littorine to hyoscyamine ratio from 0.07: 1 to 0.26: 1 when fed at 1.5 mM (113). The implication of these experiments, namely, that free tropic acid is acting indirectly on the formation of hyoscyamine, is strengthened by recent feeding experiments. Root cultures of Duboisia leichhardtii incorporated ~ ~ - [ l - ' ~ C ] t r o acid p i c into hyoscyamine very poorly (<1% in 3 weeks) compared to L-[l-'4C]phenylalanine (-50%) (161). Using the Daturu hybrid culture, the influence of feeding (RS )-tropic acid simultaneously with ( R S ) - [1',3'-'3C2]phenyllactic acid was examined (101,113). In cultures supplied with these acids at 1 : 1 or 1 : 3 molar ratios, the total amounts of littorine, hyoscyamine, and hyoscine formed were decreased. In particular, littorine formation was suppressed, there being a 90% lower yield of this alkaloid at 1.5 mM (RS)-tropic acid, whereas hyoscyamine was diminished only about 50%. In addition, the percentage incorporation of [ I ',3'-13C2]phenyllacticacid into hyoscyamine and its derivatives was very much higher than might be expected if free DL-tropic acid were competing with tropoyl moieties derived from phenyllactic acid for incorporation. Particularly significant was the observation that free tropic acid did not decrease the specific incorporation of the 13C label into hyoscyamine and hyoscine, suggesting that the free tropic acid had not prevented the [ '3C]phenyllactic acid from being converted to the tropoyl moiety of hyoscyamine. Furthermore, the specific incorporation of the label into littorine was enhanced about 5-fold, indicating that the presence of the free tropic acid had encouraged the I3Clabel to be metabolized to littorine. Thus, it appears that exogenous tropic acid may act, specifically or otherwise, as an inhibitor of the formation of phenyllactic and tropic acids from phenylalanine, while being itself only poorly incorporated. In addition, it appears that there may be some level of interaction between the reactions forming different tropane esters. With the D.candidu x D. aurea hybrid, it was found that, of a range of esterifying acids supplied (including tropic acid), none stimulated the formation of hyoscyamine (22). Indeed, the levels of both hyoscyamine and scopolamine were diminished in virtually every instance. Both tiglic acid and,

172

RICHARD J . ROBINS A N D NICHOLAS J . WALTON

unexpectedly, DL-P-phenyllactic acid caused some increase in the levels of 3-tigloyloxytropanes, whereas DL-mandelic acid, while not itself esterified, nevertheless reduced the formation of both 3-tigloyloxy- and 3tropoyloxytropanes (including hyoscyamine and hyoscine) and led to an appreciable increase in the level of 3-acetyltropine. This stimulation of 3-acetyltropine formation is further evidence that it is an adventitious metabolite (see above). In general, these results support the theory that the physiological limitation in the supply of esterifying moieties is not the provision of the acids themselves, but rather of their CoA thioesters (see Sections VI and VII). A second, conceptually simple approach to the identification of regulatory reactions is the measurement and comparison of enzyme activities. A thorough survey of the enzyme activities involved in nicotine biosynthesis and their implications for the control of the nicotine biosynthetic pathway has been made by Wagner and co-workers (162,163). Data on the enzyme activities in suspension cultures, callus cultures, and (plant) roots of Nicotiana tabacum cv. “Samsun” strongly suggest a regulatory role for PMT because, unlike ODC and MPO, the enzyme was virtually absent from suspension cultures, which did not produce nicotine (162), and, furthermore, showed a good correlation with nicotine production when callus cultures were induced to form nicotine by transfer to an “induction medium” containing reduced levels of the plant growth regulators NAA and kinetin (163). On the other hand, there was an inverse correlation between PMT activity (in the roots) and the aerial nicotine content when comparing the low-nicotine-producing cultivar N . tabacum, cv. “Sarnsun” with the high-nicotine producer N . tabacum cv. “Bursa.” In this case, there was instead a correlation between nicotine content and ODC activity. In a variety of plants and root cultures (Nicotiana, Hyoscyamus, Atropa, and Datura), the activities of ODC, ADC, and PMT are fairly similar and within the range 1-40 pkat/mg protein. Under the conditions of assay, the activity of PMT very often exceeds that of ODC or ADC (18,21,33,162,164). The activity of MPO falls within the range 170 pkat/mg protein (21,54,162,164). Interestingly, appreciably greater activities of ODC, PMT, and MPO, both in roots and in root cultures, have been measured in N . tabacum than in D . stramonium (162). Rhodes et al. (150) compared the synthetic capacities of ODC, ADC, and PMT with the observed flux into alkaloids in a transformed root culture of D . stramonium (Table VII). In all cases, the maximum flux capacity of the enzymes is greater than the rate of alkaloid formation. On the other hand, this flux capacity is below 7-fold for each enzyme during the phase of maximal accumulation of alkaloid. Because the flux capacities of the enzymes are measured at nonphysiological, near-saturating levels

TABLE VII ESTIMATES OF SYNTHETIC CAPACITIES OF INDIVIDUAL ENZYMES OF TROPANE ALKALOID BIOSYNTHESIS A N D FLUXINTO ALKALOIDS IN TRANSFORMED ROOTCULTURE OF Datura stramonium" Total alkaloid accumulation over the time interval (pmol/flask)

Time interval (days)

Total synthetic capacity over time interval of biosynthetic enzymes (pmol/flask)bJ ODC + ADC

(A)

ODC

ADC

(9)

Excess B over A (-fold)

1.4 11.2 8.8 1.4

3.0 14.8 22.2 12.5

2.5 14.7 23.5 12.6

5.5 29.5 45.7 25.1

3.9 2.6 5.2 18

PMT (C)

Excess C over A (-fold)

MPO (D)

Excess D over A (-fold)

9.2 75 12.3 90

6.6 6.7 14.0 64

6.6 44 69 54

4.7 3.9 7.8 38

~

~~~

0-7 7-14 14-22 22-3 I ~

~~

From Rhodes ef a / . (150). The activities of ODC and ADC were determined by release of 14C02from ~-[l-'~C]ornithine (2 m M ) and ~-[U-'~Clarginine (0.5 m M ) , respectively; the activity of PMT was determined by transfer of a C-1 unit from [merhv/-"C]SAM (0.1 m M ) to putrescine (4 m M ) and that of MPO by conversion of N-[mefhyl'Hlmethylputrescine ( 5 m M ) to N-[mefhyl-'H]methyIpyrrolinium. ODC, Ornithine decarboxylase; ADC, arginine decarboxylase; PMT, putrescine N-methyltransferase; MPO, methylputrescine oxidase.

174

RICHARD J . ROBINS A N D NICHOLAS J. WALTON

of substrate, the levels of available enzyme activity might limit the rate of alkaloid biosynthesis. This is particularly so in the case of ODC and ADC, especially in view of the role of these enzymes in providing putrescine for the biosynthesis of polyamines. The likelihood of a significant overall limitation to alkaloid biosynthesis at the stage of putrescine formation has been further borne out by the results of Hamill et a f . (67). These authors inserted into the genome of Nicotiana rustica transformed root cultures a gene for ornithine decarboxylase cloned from Saccharomyces cerevisiae and driven by the powerful enhanced cauliflower mosaic virus 35 S promoter. Expression of the gene and elevated ODC activity were shown to be correlated with modest increases in putrescine and nicotine production. Whereas the mean nicotine content of (genetically manipulated) control lines 14 days after subculture was 2.28 0.22 pmol/g fresh weight, that of six culture lines containing the s. cerevisiae ODC gene was 4.04 +- 0.48 pmol/g fresh weight. Detailed analysis of one of these lines showed that ODC activity was elevated, relative to that present in a control line, from around 20 to 50 pkat/mg protein 7 days after subculture and, more strikingly, from about 1 to 30 pkat/mg protein 24 days after subculture. Not unexpectedly, perhaps, exogenous metabolites have been found to affect extractable enzyme activities, although the full significance of these observations remains unclear. Putrescine, agmatine, or spermidine, supplied at 1 , 1 , and 0.5 mM, respectively, to transformed root cultures of D. stramonium decreased by around half the ADC activity extracted 10 days later (21). The enzyme activity was in each case determined after rigorous gel filtration of the extract to remove small molecules, since these amines act as inhibitors in vitro (21). Agmatine alone had essentially no effect on extractable PMT activity but, interestingly, was found to prevent the diminution of activity caused by the addition to the culture medium of the ADC inhibitor DFMA (40). When provided in the medium at 2 mM concentration, it completely offset the 75% reduction in extractable activity caused by 5 mM DFMA. Besides providing further circumstantial support for the importance of ADC in tropane alkaloid biosynthesis (see Section III), this finding suggests the possibility that agmatine acts, directly or indirectly, as an inducer of PMT activity, with its concentration in the tissue presumably being diminished by DFMA treatment. Further work is necessary to characterize these interactions, however, especially since (i) DFMO was also found to have some effect on extractable PMT activity and (ii) the effects of other amines in conjunction with DFMA and DFMO have not yet been reported. Finally, 1 mM tropine caused a modest (-20%) decrease in the extractable activity of both ADC and PMT; this effect became more marked at concentrations of 2.5 and 5 mM,

*

2.

BIOSYNTHESIS OF TROPANE ALKALOIDS

175

when a reduction in the extractable activities of ODC and MPO was also apparent (21). It is unclear whether these observations are of physiological significance. The profiles of the levels of some enzyme activities and of metabolic intermediates throughout the culture cycle have been described in detail for cultures of H. afbus (18,34) and Datura (21,22). It is difficult to assign metabolic significance to these data, especially since the tissue and intracellular compartmentation of the enzymes and metabolites remains unknown. Noteworthy, however, is the high level of conjugated putrescine, varying between 1 and 3 pmollg fresh weight throughout the culture cycle in transformed roots of D. stramonium and being very much higher than the level of unconjugated putrescine (21). By contrast, N-methylputrescine is chiefly found in its unconjugated form and at levels comparable with those of free putrescine. Whether conjugated amines are metabolically active in alkaloid biosynthesis is an important question which remains to be investigated. It is intriguing, however, to see that conjugated putrescines and hyoscyamine show inverse accumulation profiles in D. stramonium (165). Useful insights into the overall control of the pathway have been obtained from studies in which metabolite and enzyme levels have been measured in transformed root cultures caused to differentiate into callus tissue by plant growth regulators. Adventious root cultures respond quite differently to exogenous plant growth regulators (17,166,167). From experiments with a number of transformed root cultures, including those of H. muticus (168) and N . rustica (169,170),it has been shown that maintenance of the root phenotype is essential to continued alkaloid production. The subculture of transformed roots of D. stramonium into medium containing NAA (2 mglliter) and kinetin (0.2 mglliter) causes alkaloid production to cease rapidly. After 5 days, the alkaloid content of the tissue is only about 25% of that present initially in roots of the same age subcultured into normal medium (23). Interestingly, the content of 3-acetyltropine appears to diminish less rapidly than that of other alkaloids, a further indication that the metabolism of this compound is not tightly controlled (see above; Table V). Accompanying the decline in alkaloid content are changes in the activities of certain enzymes, relative to those measured in roots subcultured normally. Thus, ADC, ODC, and MPO do not show the normal transient rise in activity observed 4-5 days after subculture. More significantly, the activity of PMT falls rapidly to only 10% of its normal value 2 days after subculture, with a substantial diminution being apparent even within 1 day. The rapid loss of PMT activity, occurring well before the start of any visible morphological changes, has also been observed in cultures of

176

RICHARD J . ROBINS A N D NICHOLAS J . WALTON

N. rustica (150,170). The particular sensitivity of PMT to the transfer to dedifferentiation-inducing medium has been taken to indicate that the level of this enzyme is especially tightly controlled and may represent a crucial point of regulation of the tropane biosynthetic pathway. This is in agreement with the conclusions reached by Wagner and colleagues from analyses of callus and suspension cultures of N. tabacum (162, 163). On the other hand, the changes in ODC and ADC activity, although less striking, appear equally reproducible; furthermore, from a very low level of activity 2 days after subculture into dedifferentiation-inducing medium, the activity of PMT is later partially restored, without an accompanying restoration of alkaloid production. The theory that PMT is crucial in the regulation of tropane alkaloid biosynthesis from arginine and ornithine is fully consistent with its apparent function as the branch-point enzyme, committing putrescine to alkaloid biosynthesis rather than to polyamine formation. An alternative view is that it is the compartmentation of putrescine which represents the true branch point, a view supported indirectly by the inability of tropane alkaloid-producing plants to form substantial quantities of nonmethylated, nor-type alkaloids even though they undoubtedly possess the enzymatic capacity to do so (see Section IV). In a pathway for which there is good, if indirect, evidence for metabolic tunneling and a concerted sequence of reactions (see Section III,C), however, the distinction between these alternatives is a fine one and difficult, if not impossible, to substantiate experimentally. To summarize, it is now possible to reach certain preliminary conclusions regarding the regulation of tropane alkaloid formation at the biochemical level. (1) Enzyme activity measurements, and also genetic manipulation studies, in Nicotiana suggest limitations to metabolic flux at the stage of arginine and ornithine decarboxylation (see also Section X,B). (2) From its position at a metabolic branch point and from correlations of enzyme activity with overall alkaloid biosynthetic capacity, PMT is implicated in the overall regulation of the pathway, even though its activity may not be flux limiting in the steady state. (3) Agmatine may play a role in the induction of PMT. (4)Tropine esterification is a potential, if not an actual, limitation to tropeine production. Esterification (except for acetylation) may be partially limited by the provision of esterifying acids in their activated form. Over the next few years, it is expected that these tentative mechanisms will be further assessed and characterized. There is also the need to extend investigations to a much wider range of species and also beyond the transformed root cultures used for many of these studies.

2.

BIOSYNTHESIS OF TROPANE ALKALOIDS

I77

B. REGULATION AT WHOLE-PLANT LEVEL

There is a considerable literature concerned with the whole-plant aspects of alkaloid biosynthesis. Much older work, which is still relevant, was comprehensively reviewed by James (171), Mothes (172), Sokolov (173), and Fluck (174), and only reference to selected aspects and to recent developments is made here. In general, tropane alkaloid biosynthesis occurs largely, though not exclusively, in the roots. Translocation then occurs via the xylem to the aerial parts, where limited further metabolism may take place, for example, the conversion of hyoscyamine to hyoscine in many Daturu species. The root origins and subsequent movements of alkaloids in a number of species were deduced in part from grafting experiments, frequently involving stocks and scions from plants of different solanaceous genera producing dissimilar alkaloids, for instance, Datura, Nicotiana, and Solanurn (171,172). It is clear, however, that even where synthesis is predominately limited to the roots, some synthesis may potentially occur de nouo in aerial parts. For example, scions of A. belladonna grown on foreign stocks nevertheless contain concentrations of mydriatic alkaloids, and detached leaves of A . belladonna, kept in darkness, were found to show a rise in alkaloid content after 5 days which was correlated with a decline in protein nitrogen (171). The ontogeny of tropane alkaloid accumulation has been documented in several cases. For example, Guillon (175) studied the appearance and disappearance of alkaloids in seedlings of D. srrarnoniurn, noting that in the young root they appeared in the cortex from the sixth day after germination and within 1 month appeared in the pith and the periderm. In the leaves, alkaloids appeared first in the epidermis and subsequently in the mesophyll and the parenchyma of the vascular bundles. At a later stage, particularly in Hyoscyarnus, a correlation has been demonstrated between the onset of flowering and a reduction in alkaloid biosynthesis (172,174). In addition, the alkaloid composition may vary substantially with the developmental stage of the plant. Thus, the principal alkaloid in germinating seeds and seedlings of A. belladonna was found to be cuscohygrine, with hyoscyamine and hyoscine appearing only at a later stage (176,177). Tropane alkaloid formation may be appreciably stimulated in response to mechanical damage. This is consistent with the probable role of alkaloids as toxic protectants against insects and herbivores. In plants of Atropa acuminata, alkaloid levels have been determined following varying degrees of damage inflicted either mechanically or by the feeding of mol-

178

RICHARD J . ROBINS A N D NICHOLAS J . WALTON

luscs or moth larvae (178,179). When up to 5.3% of the tissue of an individual, attached leaf had been removed, very little change in alkaloid content was observed; however, when 9% had been removed, the alkaloid content of the remainder of the leaf was approximately doubled. The consequences of mechanical damage and insect damage were essentially identical (179). This was a short-range effect, however, and there was little or no increase in alkaloid levels in the undamaged leaves and seemingly no reproducible change in the stems or roots (178,179). Similar results were obtained in N . tabacum, where simulated herbivory increased the concentration of nicotine by up to 550% of the control values after 11 days (180). It is likely that more drastic treatments can markedly increase alkaloid biosynthetic capacity. The PMT activity in roots of N . tabacum was increased 10-fold following decapitation of the plants, and smaller increases were observed in the activities of ODC and MPO (164). Similar effects on PMT were obtained by Saunders and Bush (180, though not by Feth and co-workers (51). These experiments need to be repeated in tropane alkaloid-producing plants and extended to other enzymes. There is abundant evidence that less extreme environmental influences also substantially affect the alkaloid content. Alkaloid levels are responsive to temperature, light, water deficit, and nutrient status. Low temperatures, especially frost, can markedly reduce the alkaloid content in a wide range of plants. The effects of low temperatures on Datura and other species were reviewed by Sokolov (173). For example, D. stramonium and D . ceratocaula plants, in which growth was halted at the second or third leaf stage by a cold spell in June, were found to lose alkaloids completely from their leaves and stems, even though the temperatures fell only from 15-18°C to 8- 10°C. These comparatively early physiological observations need to be followed up by more detailed investigations of enzymological changes. The effects of light and, in particular, of day length are complex and closely associated with ontogeny. Of particular interest, perhaps, are the observations of Zolonitskaya (for review, see Ref. 173) that alkaloid composition may be manipulated by day length. Thus, by growing the shortday plant D. innoxia, which normally produces principally hyoscine, under long-day conditions, it was possible to achieve an enhanced production of hyoscyamine (45-50% of the total alkaloid), whereas, conversely, plants of D . stramonium, a long-day plant, produced elevated levels of hyoscine when grown under short-day conditions. It is possible that day length is one factor which regulates the alkaloid composition of plants native to particular localities.

2.

BIOSYNTHESIS OF TROPANE ALKALOIDS

179

The existence of particular races of plants with discrete alkaloid compositions and geographical distributions is perhaps most conspicuous among tropane alkaloid producers in the case of Duboisia (171). In north Queensland, the principal alkaloid of Duboisia myoporoides is scopolamine. Further south, the pyridine alkaloids nicotine and anabasine predominate (124).

In common with other alkaloid-producing plants (182),tropane alkaloid producers appear generally to accumulate increased concentrations of alkaloid in response to diminished water availability. Thus, in H . muticus, the alkaloid content of (dried) flowering plants grown on sandy soil irrigated sparingly was 1.3%, in contrast to a content of 0.8% for plants grown on well-irrigated clay soil (183). Similar behavior has been reported for D. innoxia (173). Such effects need not simply reflect a reduction in vegetative growth associated with increasing water stress, however. For N. tabacum, nicotine production could be increased by 85% in an unirrigated crop compared with a freely irrigated one, whereas the yield of cured leaves was diminished by only 15% (184). Clearly, some more specific effect on alkaloid biosynthesis is implied. Whether the stimulation of alkaloid production by water stress is related in any interpretable manner to the accumulation of free amino acids (especially proline, glutamine, and asparagine) known to occur in many plants under these conditions is uncertain (182). On the other hand, it has long been known that tropane alkaloids are accumulated to higher concentrations in plants which receive nitrogen fertilization than in plants which are unfertilized or which are deficient in nitrogen (171,173,174).In general, fertilization with ammonium appears to have a more direct effect on tropane alkaloid biosynthesis than that with nitrate, and ammonium can continue to increase the alkaloid concentration even when growth is inhibited. An apparently more direct relationship between nutrition and tropane alkaloid formation can be demonstrated in the case of potassium deficiency. In many plants, potassium stress has been shown to be associated with increased levels of amines, including putrescine and agmatine (185). In potassium-deficient barley, for example, increases of 2- to 4-fold in the activities of two enzymes of putrescine formation, ADC and N-carbamylputrescine amidohydrolase, have been shown to occur (185). In A . acuminata plants, potassium deficiency has been shown to cause increases of around 30% in the concentrations of alkaloids in roots, stems, and leaves, which could be attributed only partially to decreases in biomass; these changes were accompanied by approximately 2- to 3-fold increases in the measured activities of ODC and ADC and in the levels of putrescine (186). These findings are further evidence suggesting a flux

180

RICHARD J . ROBINS A N D NICHOLAS J . WALTON

limitation to alkaloid biosynthesis at the stage of ornithine or arginine decarboxylation (see Section X,A). Given the much clearer biochemical characterization achieved since the earlier physiological and agronomic work was performed, along with increasing insights into the regulation of tropane alkaloid biosynthesis, it is now reasonable to expect a new wave of progress in understanding the biochemical basis of nutritional and environmental effects on tropane alkaloid production.

XI. Future Prospects Writing in the first volume of The Alkaloids, published in 1950, W. 0. James (171) observed that “difficulties are involved in securing access of the substance fed to the site of synthesis, in choosing a tissue suitable for handling and at the same time capable of alkaloid synthesis, in providing appropriate conditions, the optima being at present largely unknown, and in maintaining a reasonable degree of freedom from interfering microorganisms.” Although the difficulties to which James referred have been appreciably lessened by the advent of root cultures able to be maintained indefinitely in uitro (see Section II), some seemingly intractable biochemical problems nevertheless remain. For example, although it is true that the early speculations of Cromwell (187) on the role of ornithine, arginine, and putrescine in hyoscyamine biosynthesis have now been vindicated, and the enzymes responsible at least partially characterized, the assertion of James that “the biosynthesis of the tropic acid half of the molecule has not yet received attention” (171) would, in a strictly biochemical sense, require disappointingly little amendment more than 40 years later. Perhaps the most powerful new approach being brought to bear on this pathway is the use of genetic manipulation to alter the levels of enzymes. To date, the only metabolic sequence in tropane alkaloid biosynthesis successfully modified in this way is the conversion of hyoscyamine to scopolamine. The gene for hyoscyamine 6/3-hydroxylase, cloned from H. niger root cultures (188), has been expressed in both A. belladonna and N . tabacum under the control of the powerful cauliflower mosaic virus 35 S promoter (83,144).In the former species, which normally accumulates hyoscyamine, over 99% of the leaf and stem alkaloids of transgenic plants was found to be hyoscine (83).Transgenic N. tabacum, when fed hyoscya-

2.

BIOSYNTHESIS OF TROPANE ALKALOIDS

181

mine or 6P-hydroxyhyoscyamine, also accumulated hyoscine, a biotransformation not found in untransformed plants. From the latter result, it can be concluded with considerable confidence that both steps in the conversion of hyoscyamine to hyoscine are encoded by the same gene and are, therefore, the property of a single enzyme. This particular genetic manipulation has given conclusive results partly because it was apparent at the outset that the hyoscyamine 6P-hydroxylase enzyme (though not necessarily the gene) required for the conversion was deficient or absent in the untransformed plants. Clearly, it is an advantage to be able to establish conclusively that a specific enzyme limits a metabolic process. In many pathways, however, the limitation of metabolic flux is the result of partial limitation by each of several enzymes, and it is not possible to predict with any certainty that genetic manipulation will bring about the desired result, even if the expression of the transferred gene is achieved (189). In these circumstances, conversely, genetic manipulation might permit insights into metabolic regulation which would be difficult to achieve by biochemical or physiological means. Furthermore, both gene and antibody probes can provide valuable information concerning the tissue and cellular specificity of gene expression and enzyme localization. Because such information is itself highly desirable in the formulation of strategies to “engineer” a metabolic pathway, genetic manipulation might best develop as an iterative process in which the biochemical information gained from its practice contributes to successive, increasingly refined cycles of gene transfer and expression. The application of these concepts to tropane alkaloid metabolism is as yet untried, though some initial work has been reported using Nicotiana, in which nicotine production was enhanced by the introduction of an ornithine decarboxylase gene from yeast (67; see Section X,A). In the future, it may therefore be possible to increase the production of hyoscyamine and hyoscine beyond normal levels and, perhaps, to generate transgenic plants in which useful or valuable minor components, for example, tigloidine (tigloylpseudotropine), are produced in increased quantities. Conversely, genetic manipulation approaches are available in principle for “shutting off” the production of undesirable minor components, provided that genes associated with the unwanted metabolic sequences can be isolated (190). Finally, gene isolation might ultimately lead to the production of microorganisms capable of synthesizing useful tropane alkaloids. That tropane-type skeletons, for example, anatoxin-a of the cyanobacterium Anabaena Jos-aquae (191), are already made in such organisms reveals the possibility that it may not be necessary to transfer the entire pathway from the plant.

182

RICHARD J . ROBINS AND NICHOLAS J . WALTON

Acknowledgments

We are grateful to the following individuals for providing us with information prior to publication: A. W. Alfermann, P. Bachmann, K. Doerk, B. Drager, E. Leete, J. B. Harborne, W. J. Griffin, T. Hashimoto, M. B. Khan, Y. Kitamura, H. Noguchi. U. Sankawa, M. Sauenvein, E . J . Staba, J . G. Woolley, and Y. Yamada. We thank our colleagues Mike Rhodes and Adrian Parr at the AFRC Institute of Food Research, who have been most helpful in discussing various points of tropane metabolism with us, and Birgit Drager for discussions and for reading and criticizing the manuscript.

REFERENCES

1. E. Leete, Planta Med. 36, 97 (1979). 2. E. Leete, Planfa Med. 56, 339 (1990). 3. R. L . Clarke, in “The Alkaloids” (R. H. F. Manske and H. L. Holmes, eds.), Vol. 16, p. 83. Academic Press, New York, 1990. 4. M. Lounasmaa, in “The Alkaloids” (A. Brossi, ed.), Vol. 33. p. I . Academic Press, New York, 1988. 5. W. C. Evans, Linn. SOC. Symp. Ser. 7, 241 (1979). 6. W. C. Evans, Pharm. J. 244, 651 (1990). 7. W. C. Evans, “Trease and Evans’ Pharmacognosy,” 13th Ed., p. 832. Bailliere Tindall, London, 1989. 8. A. Ahmad and E. Leete, Phytochemistry 9, 2345 (1970). 9. D. N. Butcher and H. E. Street, Bot. Rev. 30, 513 (1964). 10. M. L. Solt, Plant Physiol. 32, 480 (1957). 11. G. C. Mitra, Indian J. Exp. Biol. 10, 217 (1972). 12. H.-W. Liebisch, W. Maier, and H.-R. Schiitte, Tetrahedron Lett. 34, 4079 (1966). 13. A. von Romeike, in “Biochemie und Physiologie der Alkaloide” (K. Mothes, K. Schreiber, and H.-R. Schiitte, eds.), p. 223. Akademie-Verlag, Berlin, 1972. 14. H. Metz, Chem. Ind. 2nd Int. Ferment. Symp., 552 (1964). 15. T. Hashimoto and Y. Yamada, Planta Med. 47, 195 (1983). 16. J. D. Hamill, A. J. Parr, M. J. C. Rhodes, R. I. Robins, and N. J. Walton, B i d Technology 5, 800 (1987). 17. T . Hashimoto, Y. Yukimune, and Y. Yamada, J . Plant Physiol. 124, 61 (1986). 18. T. Hashimoto, Y. Yukimune, and Y. Yamada, Plantu 178, 123 (1989). 19. A. J. Parr, J . D. Hamill, J. Payne, M. J. C. Rhodes, R. J. Robins, and N . J. Walton, in “Manipulating Secondary Metabolism in Culture” (R. J. Robins and M. J . C. Rhodes, eds.), p. 101. Cambridge Univ. Press, Cambridge, 1988. 20. A. J. Parr, J. Payne, J. Eagles, B. T. Chapman, R. J. Robins, and M. J. C. Rhodes, Phytochemistry 29, 2545 (1990). 21. R. J. Robins, A. J. Parr, E. G. Bent, and M. J. C. Rhodes, Plnntu 183, 185 (1991). 22. R. J. Robins, A. J. Parr, J. Payne, N . J. Walton, and M. J. C. Rhodes, Planta 181, 414 (1990). 23. R. J. Robins, E. G. Bent, and M. J. C. Rhodes, Planta 185, 385 (1991). 24. W. Gritsanapan and W. J. Griffin, Phytochemistry 30, 2667 (1991).

2. 25. 26. 27. 28. 29.

BIOSYNTHESIS OF TR O P A N E ALKALOIDS

I83

Y. Kitamura, H. Miura, and M. Sugii, Phytochemistry 25, 2541 (1986). Y. Yamada and T. Endo, Plant Cell Rep. 3, 186 (1984). Y. Kitamura, H. Miura, and M. Sugii, Chem. Pharm. Bull. 33, 5445 (1985). T. Endo and Y. Yamada, Phytochemistry 24, 1233 (1985). Y. Kitamura, Y. Sugimoto, T. Samejima, K. Hayashida. and H. Miura. Chem. Pharrn.

Bull. 39, 1263 (1991). 30. P. R. Bachmann and F.-C. Cygan, Pharm. Weekhl. Sci. Ed. 9, 219 (1987). 31. H. Deno. H. Yamagata, T. Emoto, T. Yoshioka, Y. Yamada. and Y. Fujita, J. Plant Physiol. 131, 315 (1987). 32. Y. Mano, H . Ohkawa, and Y. Yamada, Plant Sci. 59, 191 (1989). 33. N. J. Walton, R. J. Robins, and A. C. J. Peerless, Plantu 182, 136 (1990). 34. T. Hashimoto, Y. Yukimune. and Y. Yamada, Plantu 178, 131 (1989). 35. T. A. Smith, Prog. Phytochem. 4, 27 (1977). 36. P. D. Shargool, J. C. Jain. and G. M. McKay, Phytochemistry 27, 1571 (1988). 37. W.C. Fuller and M. R. Gibson. J. A m . Pharm. As.soc. Sci. Ed. 41, 263 (1952). 38. A. F. Tiburcio and A. W. Galston, Phytochemistry 25, 107 (1986). 39. P. Bey, C. Danzin, and M. Jung, in “Inhibition of Polyamine Metabolism” (P. P. McCann, A. E . Pegg, and A. Sjoerdsma. eds.), p. I . Academic Press. San Diego. 1987. 40. R. J. Robins, A. J. Parr, and N. J . Walton. Planta 183, 196 (1991). 41. T. Hartmann, A. Ehmke, U. Eilert, K. von Borstel. and C. Theuring. Plantu 177, 98 (1989). 42. E. Leete, J . A m . Chem. Soc. 84, 55 (1962). 43. E. Leete, Tetruhedron Lett. 24, 1619 (1964). 44. H.-W. Liebisch and H.-R. Schiitte, 2. PJanzenphysiol. 57,434 (1967). 45. E. Leete and A. J. McDonell, J. Am. Chem. Soc. 103, 658 (1981). 46. T. Hashimoto, Y. Yamada, and E. Leete, J . A m . Chem. Soc. 111, 1141 (1989). 47. E. Leete, T. Endo, and Y. Yamada, Phytochemistry 29, 1847 (1990). 48. N. I . Walton and N. J. Belshaw, Plant Cell Rep. 7, I15 (1988). 49. S. H. Hedges and R. B. Herbert, Phytochemistry 20, 2064 (1981). 50. S. Mizusaki, Y. Tanabe. M. Noguchi. and E. Tamaki, Plurit Cell Physiol. 12, 633 ( I97 1). 51. F. Feth, H.-A. Arfmann, V. Wray, and K. G. Wagner. Phvtochemistry 24,921 (1985). 52. N. J. Walton, A. C. J. Peerless, H. D. Boswell, and R. J. Robins, unpublished results (1992). 53. T. Hashimoto, A. Mitani, and Y. Yamada. Plant Physiol. 93, 216 (1990). 54. N. J. Walton and W. R. McLauchlan, Phytochemistry 29, 1455 (1990). 55. N. J. Walton, R. J. Robins, and M. J. C. Rhodes, Plant Sci. 54, 125 (1988). 56. L. Glatz, J. Kovar, L . Macholan, and P. Pec. Biochem. J . 242, 603 (1987). 57. H. M. Davies, D. J. Hawkins, and L. A. Smith, Phytochemistry 28, 1573 (1989). 58. W. R. McLauchlan, personal communication (1992). 59. A. von Romeike, Bot. Nor. 131, 85 (1978). 60. D. J. Robins, Chem. Soc. Rev. 18, 375 (1989). 61. H. E. Flores and P. Filner, in “Primary and Secondary Metabolism of Plant Cell Cultures” (K.-H. Neumann, W. Barz. and E. Reinhard, eds.), p. 174. Springer-Verlag. Berlin, Heidelberg, and New York, 1985. 62. S. Mizusaki, T. Kisaki, and E. Tamaki, Plant Physiol. 43, 93 (1968). 63. J. B. Friesen and E . Leete, Tetrahedron Lett., 6295 (1990). 64. H.-W. Liebisch, K. Peisker, H . Radwan, and H.-R. Schiitte, Z . Pflanzenphysiol. 67, l(1972). 65. B. .A McGaw and J. G. Woolley, Tetrahedron Lett. 34, 3135 (1979).

184

RICHARD J . ROBINS A N D NICHOLAS J . WALTON

66. T. Endo, N. Hamaguchi, T. Hashimoto, and Y. Yamada, FEBS Lett. 234, 86 (1988). 67. J. D. Hamill, R. J. Robins, A. J. Parr, D. M. Evans, J. M. Furze, and M. J. C. Rhodes, Plant Mol. Biol. 15, 27 (1990). 68. M. J. C. Rhodes and R. J. Robins, in “The Biochemistry of Plants” (D. D. Davies, ed.), Vol. 13, p. 65. Academic Press, San Diego, 1987. 69. E. Leete and S. H. Kim, J . A m . Chem. Soc. 110, 2976 (1988). 70. T. Hemscheidt and I. D. Spenser, J. A m . Chem. Soc. 112, 6360 (1990). 71. U. Sankawa, H . Noguchi, T. Hashimoto, and Y. Yamada, Chem. Pkarm. Bull. 38, 2066 (1990). 72. U. Sankawa, H. Noguchi, and R. J. Robins, unpublished results (1991). 73. B. A. McGaw and J. G. Woolley Phytochemistry 17, 257 (1978). 74. B. A. McGaw and J. G. Woolley, Phytochemistry 18, 189 (1979). 75. R. J. Robins, E. Leete, and A. J. Parr, unpublished results (1992). 76. E. Leete, personal communication (1991). 77. E. Leete and S. H. Kim, J . Chem. Soc., Chem. Commun., 1899 (1989). 78. P. R. Bachmann, personal communication (1992). 79. A. Huhtikangas, A. Martinsen, A. Yliniemela, H. Konschin, and L. Laaksonen, Acta Pharm. Fenn. 99, 7 (1990). 80. L. K. Simola, A. Martinsen, A . Huhtikangas, R. Jokela, and M. Lounasmaa, Acta Chem. Scand. 43, 702 (1989). 81. M. E . Landgrebe and E. Leete, Phytochemistry 29, 2521 (1990). 82. A. Portsteffen, B. Drager, and A. Nahrstedt, Phytochemistry 31, 1135 (1992). 83. Y. Yamada and T. Hashimoto, personal communication (1992). 84. K. J. Koelen and G. G. Gross, Planta Med. 44, 227 (1982). 85. B. Drager, T. Hashimoto and Y. Yamada, Agric. B i d . Chem. 52, 2663 (1988). 86. M. M. Couladis, J. B. Friesen, M. E. Landgrebe, and E. Leete, Phytochemistry 30, 801 (1991). 87. A . Portsteffen, B. Drager, and A. Nahrstedt, in “Phytochemistry and Agriculture” (T. A. van Beek, ed.), p. 63. Wageningen Agric. Univ. (UNIPUB), Wageningen, The Netherlands, 1992. 88. B. Drager and A. Schaal, in “Phytochemistry and Agriculture” (T. A. van Beek, ed.), p. 52. Wageningen Agric. Univ. (UNIPUB), Wageningen, The Netherlands, 1992. 89. E. Leete, Phytochemistry 11, 1713 (1972). 90. B. Drager, personal communication (1991). 91. A. Goldmann, M.-L. Milat, P.-H. Dudrot, J.-Y. Lallemand, M. Maille, A. Lepingle, I. Charpin, and D. Tepfer, Phytochemistry 29, 2125 (1990). 92. A. J. Pam, N. J. Walton, S. Bensalem, P. H. McCabe, and W. Routledge, Phytochemisfry 30, 2607 (1991). 93. 9. Drager, A. Portsteffen, A. Schaal, P. H. McCabe, A. C. J. Peerless, and R. J. Robins, Planta 188, 581 (1992). 94. E. Leete, J. A m . Chem. Soc. 82, 612 (1960). 95. E. Leete and M. L. Louden, Chem. Ind. (London), 1405 (1961). 96. E. Leete, J . A m . Chem. Soc. 106, 7271 (1984). 97. E. Leete, Can. J. Chem. 65, 226 (1987). 98. M. Ansarin, Ph.D. Thesis, Leicester Polytechnic, Leicester, England (1980). 99. E. Leete, N. Kowanko, and R. A. Newmark, J. A m . Chem. SOC.97, 6826 (1975). 100. M. Ansarin, S. C. Woodland, and J. G. Woolley, IUPAC Congress, Manchester, Poster Abstracts 5P.30 (1985). 101. R. J . Robins, J. Eagles, 1. Colquhoun, J. G. Woolley, and M. Ansarin, in “Phytochemistry and Agriculture” (T. A. van Beek, ed.), p. 66. Wageningen Agric. Univ. (UNIPUB), Wageningen, The Netherlands, 1992.

2. BIOSYNTHESIS

OF TROPANE ALKALOIDS

I85

102. E. Leete and E. P. Kirven, Phytochemistry 13, 1501 (1974). 103. W. C. Evans, J. G. Woolley, and V. A. Woolley, in “Biochemie und Physiologie der Alkaloide” (K. Mothes, K. Schreiber, and H.-R. Schutte, eds.), p. 227. AkademieVerlag, Berlin, 1972. 104. M. Ansarin and J. G. Woolley, J. Pharm. Pharmacol. 30, 83P (1978). 105. A. Jindra and E. J. Staba, Phytochemistry 7, 79 (1968). 106. E. J. Staba, personal communication (1991). 107. R. A. Jensen, Recent Adu. Phytochem. 20, 57 (1986). 108. K. Doerk, I. Ionkova, L. Witte, and A. W. Alfermann, in “Abstracts IAF’TC Hamburg, Germany 18-19 September 1991” (W. 0. Abel, H. Lorz, W. Preil, and 0. Schieder, eds.), p. 45. 109. C. V. Givan, in “The Biochemistry of Plants” (B. J. Miflin, ed.), Vol. 5, p. 329. Academic Press, New York and London, 1980. 110. M.Mazelis, in “The BiochemistryofPlants” (B. J. Miflin, ed.), Vol. 5 , p. 542. Academic Press, New York and London, 1980. 111. A. W. Alfermann, personal communication (1991). 112. H.-W. Liebisch, G. C. Bhasvar, and H. J. Schaller, in “Biochemie und Physiologie der Alkaloide” (K. Mothes, K. Schreiber, and H.-R. Schutte, eds.), p. 233. AkademieVerlag, Berlin, 1972. 113. R. J . Robins and J. G. Woolley, unpublished results (1991). 114. D. Gross and H.-R. Schiitte, Arch. Pharm. (Weinheim, Ger.), 2961 (1963). 115. E. Leete, Phytochemistry 22, 699 (1983). 116. K. R. Hansen and E. A. Havir, in “The Biochemistry of Plants” (E. E. Conn, ed.), Vol. 7, p. 577. Academic Press, New York and London, 1981. 117. J. W. Loder and G. B. Russell, Aust. J. Chem. 22, 1271 (1969). 118. J. G. Woolley, in “Biochemie und Physiologie der Alkaloide” (K. Mothes, K. Schreiber, and H.-R. Schutte, eds.), p 531. Akademie-Verlag, Berlin, 1966. 119. K. Basey and J. G. Woolley, Phytochemistry 12, 2197 (1973). 120. K. Basey and J. G. Woolley, Phytochemistry 12, 2883 (1973). 121. D. H. G. Crout, J. Chem. Soc., C , 1233 (1967). 122. V. W. Rodwell, in “Metabolic Pathways,” 3rd Ed., Vol. 3, p. 191. Academic Press, New York and London, 1969. 123. N. Hiraoka, M. Tabata, and M. Konoshima, Phyrvchemistry 12, 795 (1973). 124. R. J. Robins, P. Bachmann, T. Robinson, M. J. C. Rhodes, and Y. Yamada. FEES Lett. 292, 293 (1991). 125. H. Mizukami, personal communication (1990). 126. S. Rabot and R. J. Robins, in “Phytochemistry and Agriculture” (T. A. van Beek, ed.), p. 67. Wageningen Agric. Univ. (UNIPUB), Wageningen, The Netherlands, 1992. 127. J. Kaczkowski, Bull. Acad. Pol. Sci. Ser. Sci. Biol. 12, 375 (1964). 128. A. Jindra, A. Cihak, and P. Kovacs, Collect. Czech. Chem. Commun. 29, 1059 (1964). 129. A. Jindra, D. Sofrova, and S. LCbova, Collect. Czech. Chem. Commun. 27, 2467 (1962). 130. G. G. Gross and K. J. Koelen, Z. Naturforsch. 35C,363 (1980). 131. G. G. Gross, personal communication (1991). 132. R. J. Robins and S. Rabot, unpublished results (1992). 133. E. Leete, J. A. Bjorklund, and S. H. Kim, Phytochemistry 27, 2553 (1988). 134. B. A. McGaw and J. G. Woolley, Phytochernistry 21, 2653 (1982). 135. G. Fodor, A. Romeike, G. Janzso, and I. Koczor, Tetrahedron Lett. 7, 19 (1959). 136. E. Leete and D. H. Lucast, Tetrahedron Lett. 38, 3401 (1976). 137. G. R. Waller and E. K. Nowacki, “Alkaloid Biology and Metabolism in Plants” p. 71. Plenum, New York and London, 1978.

186

RICHARD J . ROBINS A N D NICHOLAS J . WALTON

T. Hashimoto and Y. Yamada, Plant Physiol. 81, 619 (1986). T. Hashimoto and Y. Yamada, Eur. J. Biochem. 164, 277 (1987). T. Hashimoto, J. Kohno. and Y. Yamada. Plant Physiol. 84, 144 (1987). T. Hashimoto and Y. Yamada, Agric. Biol. Chem. 53, 863 (1987). T. Hashimoto, J. Kohno, and Y. Yamada, Phytochemistry 28, 1077 (1989). T. Hashimoto, A. Hayashi, Y. Amano, J . Kohno. H. Iwanari, S. Usuda, and Y. Yamada, J . B i d . Chem. 266, 4648 (1991). 143a. Y. Yamada, S. Okabe, and T. Hashimoto, Proc. Jpn. Acad. 66B, 73 (1990). 144. Y. Yamada, T. Hashimoto, and F. Sato, Proc. IAPME: Tucson, 5 (1991). 145. K. Basey and J. G. Woolley, Phytochemistry 14, 2201 (1975). 146. P. J. Beresford and J. G. Woolley, Phytochemisrry 14, 2205 (1975). 147. P. J. Beresford and J. G. Woolley, Phytochemistry 14, 2209 (1975). 148. D. von Neumann and K. H. Tschope, Nora (Jena) Abst. A . 156, 521 (1966). 149. N. W. Hamon and H . W. Youngken, Lloydia 34, 199 (1971). 150. M. J. C. Rhodes, R. J. Robins, E. L. H. Aird, J. Payne, A. J. Parr, and N. J. Walton, in “Primary and Secondary Metabolism of Plant Cell Cultures 11” (W. G. W. Kurz, ed.), p. 58. Springer-Verlag. Berlin, Heidelberg, and New York, 1989. 151. A. Jindra and A. Cihak, Symp. II Alkaloidtagungc Halle. 201 (1960). 152. L. Cosson and R.-R. Paris, C. R . Acad. Sci. Paris. Ser. D 265, 202 (1967). 153. Y. Kitamura, M. Sato, and H. Miura. Phytochemistry 31, I191 (1992). 154. Y. Kitamura, H . Miura, and M. Sugii, J. Plant Physiol. 133, 316 (1988). 155. B. C. Bose, H . N. De, and S. Mohammed, Indian J . Med. Res. 44, 91 (1956). 156. H. B. Schroter, Abh. Dtsch. Akad. Wiss. Berlin, K l . Chem. Geol. Biol. 3, 157 (1966). 157. J. D. Phillipson and S. S. Handa, Phytochemistry 14, 999 (1975). 158. T. Hartmann and G. Toppel, Phytochemistry 26, 1639 (1987). 159. T. Hashimoto and Y. Yamada, Agric. B i d . Chem. 51, 2769 (1987). 160. M. Konoshima, M. Tabata, N. Hiraoka. and H. Miyake. Shoymkugaku Zasshi 21, 108 (1967). 161. Y. Kitamura, A. Taurd, Y. Kajiya, and H . Miura. J. Plant Physiol. 140, 141 (1992). 162. R . Wagner, F. Feth. and K. G. Wagner, Phvsiol. Plant. 68, 667 (1986). 163. F. Feth, R. Wagner, and K. G. Wagner, Planta 168, 402 (1986). 164. S. Mizusaki, Y. Tanabe, M. Noguchi, and E. Tamaki, Plant Cell Physiol. 14, 103 (1973). 165. A. J. Parr, R. J. Robins, J. D. Hamill, and M. J. C. Rhodes, in “Polyamines and Ethylene: Biochemistry, Physiology and Interactions” ( H . E. Flores, R. N . Arteca, and J. C. Shannon, eds.), p. 299. American Society of Plant Physiologists, Rockville, Maryland, 1990. 166. K. Shimomura, M. Sauerwein, and K. Ishimaru, Phytochemistry 30, 2275 (1991). 167. M. Sauerwein, M. Wink, and K. Shimomura, in “Phytochemistry and Agriculture” (T. A. van Beek, ed.), p. 69. Wageningen Agric. Univ. (UNIPUB), Wageningen, The Netherlands, 1992. 168. H. E . Flores, in “Symposium, American Chemistry Society, Application of Biotechnology to Agricultural Chemistry” (H. Lebaron, R. 0. Mumma, R. C. Honeycutt, and J. H. Dussing, eds.), p. 66. American Chemical Society, Washington, D.C., 1987. 169. E. L. H. Aird, J. D. Hamill, R. J. Robins, and M. J. C. Rhodes, in “Manipulating Secondary Metabolism in Culture” (R. J. Robins and M. J. C. Rhodes, eds.), p. 137. Cambridge Univ. Press, Cambridge, 1988. 170. E. L. H. Aird, Ph.D. Thesis, University of East Anglia, Norwich, U.K. (1988). 171. W. 0. James, in “The Alkaloids” (R. H. F. Manske and H. L. Holmes, eds.), Vol. 1, p. 15. Academic Press, New York, 1950. 138. 139. 140. 141. 142. 143.

2.

BIOSYNTHESIS OF TROPANE ALKALOIDS

187

172. K. Mothes, in “The Alkaloids” (R. H . F. Manske, ed.), Vol. 6. p. I . Academic Press, New York, 1960. 173. V. S. Sokolov, S y m p . Soc. Exp. Biol. 13, 230 (1959). 174. H. Fluck, in “Chemical Plant Taxonomy” (T. Swain ed.), p. 167. Academic Press, New York and London, 1963. 175. A. Guillon, C. R . Acad. Sci. Fr. 230, 1604 (1950). 176. P. Reinouts van Haga, Nature (London) 173,692 (1954). 177. P. Reinouts van Haga, Nature (London) 174, 833 (1954). 178. B. Khan and J. B. Harborne, Chemoecology 1,77 (1991). 179. B. Khan and J. B. Harborne, Biochem. Syst. Ecol. 19, 529 (1991). 180. I. T. Baldwin, Oecologia 77, 378 (1988). 181. J. W. Saunders and L. P. Bush, Plant Physiol. 64, 236 (1979). 182. J. Gershenzon, Recent Adu. Phytochem. 18, 273 (1984). 183. Z. F. Ahmed and 1. R. Fahmy, J . A m . Phurm. Assoc. S c i . E d . 38, 484 (1949). 184. c. H. M. von Bavel, AgrOn. J . 45, 61 I (1953). 185. T. A. Smith, Recent Adu. Phytochem. 18, 7 (1984). 186. B. Khan and J. B. Harborne. Phytochemistrv 30, 3559 (1991). 187. B. T. Cromwell, Biochem. J. 37, 717 (1944). 188. J. Matsuda, S. Okabe, T . Hashimoto, and Y. Yamada, J . B i d . Chrm. 266,9640 (1991). 189. G. Stephanopoulos and J . J . Vallino, Science 252, 1675 (1991). 190. R. Offringa, P. J. M. van den Elzen. and J. J . Hookyaas. Trunsgenic Res. 1, I14 (1992). 191. J. R. Gallon, K. N . Chit. and E. G. Brown, P/zytochemi.stry 29, I107 (1990).

This Page Intentionally Left Blank

-CHAPTER 3-

SIMPLE INDOLIZIDINE ALKALOIDS HIROKITAKAHATA A N D TAKEFUMI MOMOSE Faculty of Pharmac~e~~ticul Sciences Toyama Medical and Pharmuceutical Uniurrsit) Toyama 930-01, Japan

. 189 s ................... 190

I. Introduction ............................................................................... 11. Indolizidines with Alkyl and Functionalized

111.

IV. V.

VI.

A. Indolizidine Alkaloids from Ants ......... B. Indolizidine Alkaloids from Amphibians .......................................................... Elaeocarpus Alkaloids ..... Slaframine.............................................. Hydroxylated Indolizidines ....................... A. I-Hydroxyindolizidines and 1,2-Dihydroxyindolizidines........................ B. Swainsonine .................................................................................. C. Castanospermine ............................................................................ Summary ....................................... References .........................................................................................

221 .228 233 239 250 250

I. Introduction Indolizidine alkaloids possessing the l-azabicyclo[4.3.0]nonaneskeleton are widely distributed in nature and comprise a wide range of structural and stereochemical features. The chemistry of indolizidine alkaloids was previously reviewed by Howard and Michael in Volume 28 of this treatise (1). The synthetic work on these alkaloids published during 1979-1985 was reviewed by Govindachari in 1987 (2). Alternative syntheses of indolizidines, including pyrrolizidines, have been treated in an article encompassing the period 1976 to mid-1987 by Nishimura (3 ). Since 1985, a continuous series of excellent reviews by Grundon and Michael has appeared in Natural Products Reports ( 4 4 , covering all aspects of the literature on indolizidine natural products. In addition, specialized reviews on “Amphibian Alkaloids: Chemistry, Pharmacology, and Biology” by Daly and Spande ( 9 ) and “The Chemistry and Biochemistry of Simple Indolizidine and Related Polyhydroxy Alkaloids” by Elbein and Molyneux (10) have appeared. In the 1980s, methodology for the asymmetric syntheI89 THE ALKALOIDS. VOL 44 Copyright 6 1997 hy Academic P r e s l n c All n g h h of reproduction in m y form re\erved

I90

HIROKI TAKAHATA A N D T A K E F U M I MOMOSE

sis of these alkaloids has been extensively developed. Accordingly, this chapter focuses heavily on the chiral synthesis, which has mostly been accomplished in the period from 1986 to date (1992), and on the isolation and structures of new alkaloids.

11. Indolizidines with Alkyl and Functionalized Alkyl Appendages

A. INDOLIZIDINE ALKALOIDS FROM ANTS The chemistry of alkaloids from ants and other insects has been reviewed by Numata and Ibuka in Volume 31 of this treatise (If). Five 3,5-dialkylindolizidines, namely, monomorine I (1) (f2), monomorine VI (2) (f.?), cis(3,5),cis(3H,8aH)-3-ethyl-5-methylindolizidine (3) (Id), cis(3,5),cis(3H,8aH)-3-hexyl-5-methylindolizidine (4) (fd), and tvans(3,5),trans(3H,8aH)-3-butyl-5-(4-pentenyl)indolizidine( 5 ) ( 1 3 , have been isolated from ant venoms (Fig. I). The first asymmetric synthesis of (-)-monomorine I (the enantiomer of 1) was achieved by Royer and Husson in 1985 via the chiral 2-cyano-6oxazolopiperidine synthon 6, prepared from (-)-phenylglycinol, glutaraldehyde, and potassium cyanide (16).The synthesis of an enantiomer of the (3S,5R,9R) absolute configuration established the absolute configuration of natural (+)-monomorine I (1) as (3R,5S,9S). Alkylation of 6 with the iodo ketal 7 and subsequent removal of the cyano group afforded the oxazolopiperidine 10 having the (2s)configuration. The stereospeci-

& / A + & @

H

\

1

4

3

2

5

FIG. 1. 3,5-Dialkylindolizidines isolated from ant venom.

3.

191

SIMPLE INDOLIZIDINE ALKALOIDS

"'

b,c

: P i

9

ficity implied an elimination-addition mechanism wherein hydride ion attacked the preferred iminium conformer 9 from the axial direction under stereoelectronic control. Compound 10 was alkylated with methylmagnesium iodide to give a 4 : 1 mixture of the cis-alcohol 11and its trans isomer. Hydrogenation of 11 under acidic conditions afforded the enantiomer of 1 via the iminiurn intermediate 12 (Scheme 1). In 1988, the first asymmetric synthesis of natural (+)-monomorine I (1) was accomplished by Yamazaki and Kibayashi (17) via 1,2-asymrnetric induction based on the highly diastereoselective addition of hydride to an acyclic a,P-dialkoxy ketone (13) ( l a ) , which was available from diethyl L-tartrate (Scheme 2). Reduction of ketone 13 with zinc borohydride (19) gave the syn,anti-alcohol 14 with a diastereoselectivity greater than 99% under a-chelation control. Compound 14 was converted to the aldehyde 15 by Mitsunobu inversion of the hydroxyl with phthalimide followed by debenzylation and Swern oxidation. After a sequence of homologation-oxidation steps, diastereoselective reduction of the resulting ketone 16 with L-Selectride (lithium tri-sec-butylborohydride) provided the alcohol 17 with high syn selectivity (syn:anti = 98:2), possibly via the Felkin-Anh model. Replacement of the phthaloyl group by N benzyloxycarbonyl followed by mesylation and base-induced cyclization afforded the (2S,SR)-pyrrolidine 18, which was deoxygenated to give the 3-pyrroline 19 via an oxirane. Wacker oxidation of the terminal olefin led

I 1

192

HIROKI TAKAHATA A N D TAKEFUMI MOMOSE

-

a_ Brio+ OMOM HO

b

0

MOMO

&onB

hi

MOM0

C0,EI

13

f,e

mo+ MOMO

NPMh

14

0

MOMO

15

no

OMOM

OMOM

MOMO

NPMh

16

NPMh

17

0

Cbz

Cbr

18

19

-

d Cbz

20

21

SCHEME2. Reagents: a, Ref. 18; b, Zn(BHJ2; c, phthalimide, Ph3P, DEAD; d, H2, Pd/ C; e, (COCI),, Et,N, DMSO; f, CH2=CH(CH2),MgBr; g, L-Selectride; h, NH2NH2;i, CbzCI, aq. Na2C03;j, MsCI, Et3N; k, ‘BuOK; I, conc. HCI; m, imidazole, triiodoimidazole, Ph,P; n, Ph3P, Zn; 0 , 02,PdC12, DMF-H20.

to the conversion of 19 to the ketone 20, which on hydrogenation provided exclusively (+)-monomorine I (1) via the iminium intermediate 21. An alternative enantioselective total synthesis of 1 was achieved by Ito and Kibayashi (20) (Scheme 3). The key stage in the synthesis is the asymmetric 1,3-dipolar cycloaddition of the prochiral nitrone 22 with a chiral (S)-allylic ether (23), a dipolarophile available from diethyl L-tartrate in eight steps. The rrans(3,5)-isoxazolidine 24 resulted as a major product stereoselectively via a transition state with the alkoxy group (BnO) inside, and the relative and absolute stereochemistry were suitable for the synthesis of 1 according to the stereoelectronic concept of Huck er al. (21). The major adduct 24 was transformed into the iodide 25 by a conventional procedure, and the coupling of 25 was effected with a mixed higher order organocuprate, derived from 3-butenylmagnesium bromide and lithium 2thienylcyanocuprate, leading to 26. Reductive N - 0 bond cleavage followed by benzyloxycarbonylation gave 27. After Wacker oxidation of 27, the resultant ketone 28 underwent simultaneous reductive cyclization and

I

3.

SIMPLE INDOLIZIDINE ALKALOIDS

24

25

26

27 OH

29

I93

28

OH

30

31

SCHEME 3. Reagents: a, toluene; b, LiAIH,; c, TsCI, DMAP, 'Pr,NEt; d, NaI; e , CH2=CH(CH2)2MgBr, (2-thienyl)Cu(CN)Li; f, Zn, AcOH-H20-THF; g, CbzCI, aq. NazC03; h, 02.PdCl2, CuCI2, DMF-H20; i, H2. 10% Pd/C; j, H2, 10% Pd/C, 10% HCI-MeOH; k, BnBr, Na2C03;1, MsCI, Et3N; m, H2, 10% Pd/C; n, Et3N; 0 , H2. 10% Pd/ C, Et,N.

debenzylation to yield the cis(2,6)-piperidine 29, which was subsequently converted to 30 by selective N-benzylation. Di-0-mesylation of 30 followed by hydrogenolysis and cyclization afforded the indolizidine 31, which was finally converted to 1 by nucleophilic displacement followed by reductive deiodination. Momose et al. reported the total synthesis of 1 via asymmetric aketonic cleavage of 8-azabicyclo[3.2.l]octan-8-one(32)(Scheme 4) (22). Asymmetric deprotonation of a cT-symmetric ketone (32)using a chiral lithium amide (33)(23), followed by trimethylsilylation, gave the enol silyl ether 34. Ozonolysis of 34 and subsequent reduction with sodium borohydride followed by esterification afforded the cis-2,5-disubstituted pyrrolidine 35. Modification of the appendages in several steps provided the iodide 36, which was transformed by Grignard cross-coupling and Wittig elongation reactions into the diolefin 37. As previously described in Scheme 2, Wacker oxidation of the terminal olefin in 37, followed by hydrogenation, led to 1 via 21. A short and enantiogenic synthesis of 1from L-alanine was achieved by Jefford et al. (24) (Scheme 5). Condensation of 2,Sdimethoxytetra-

194

HIROKI TAKAHATA A N D TAKEFUMI MOMOSE

Me

33

36

37

SCHEME 4. Reagents: a, 33, TMSCI-HMPA; b, O,, NaBH4; c, CH2N,; d, MOMCI, 'Pr,NEt; e , Super-Hydride; f, TsCI, pyridine; g, NaI; h, allylrnagnesiurn chloride, CuI; i, conc. HCI; j, (COCl),, DMSO, Et,N; k, CH,CH,CH=PPh,; I, 02,PdCI,, CuCI; rn, HI, 5% PdIC.

hydrofuran with L-alanine afforded the 1-pyrrolylacetic acid 38. The mixed anhydride 39 prepared in situ from 38 underwent an intramolecular electron transfer through a six-membered transition state (40) by aluminum chloride catalysis to give 42 via the dipolar intermediate 41. Chain elongation of 42 using the Arndt-Eistert procedure afforded the homologous acid 43, which was converted to the a-diazo ketone 44. Rhodium(11) acetate-catalyzed decomposition of 44 then afforded the bicyclic ketone

.

38

44

45

46

41

SCHEME 5. Reagents: a, 2,5-dirnethoxytetrahydrofuran,NaOAc; b, PrCOCI, N-rnethylrnorpholine; c, AICIj; d, isobutyl chloroformate, N-methylmorpholine; e , CH2N2;f, AgOAc; g, Rhz(OAc),; h, Hz. RO2, HCI.

3.

SIMPLE INDOLIZIDINE ALKALOIDS

40

51

49

195

50

52

Cbr

53

SCHEME 6. Reagents: a, Ref. 26; b, "Pr2CuLi; c, Me2S04;d, Meldrum's acid, N i ( a ~ a c ) ~ ; e, MeONa; f, H2, Raney Ni, HCI; g, LiAIH,; h, CbzCl; i, PCC; j , CH,COCH=PPh,; k, Hz,P t 0 2 ; I, Hz, Pd/C.

45, which was stereoselectively reduced on hydrogenation to afford 1 along with indolizidinols 46 and 47.

In addition, Lhommet and co-workers achieved the total synthesis of 1from (S)-pyroglutamic acid via the iminium intermediate 21, in the final stage, as precedented in Schemes 2 and 4 (Scheme 6) (25). Coupling of the tosylate 48 (26), derived from (S)-pyroglutamic acid, with lithium dipropylcuprate afforded 5-butylpyrrolidinone (49), which was transformed into the p-enamino ester 50. Stereoselective hydrogenation of 50 gave a mixture (96 :4) of the cis- and trans-2,5-disubstituted pyrrolidines 51. After transformation of the major cis isomer 51 into the aldehyde 52, carbon chain elongation of 52 by the Wittig reaction gave the enone 53, which, on hydrogenation, provided 1 exclusively. Recently, a short total synthesis of 1via the enantiomer of 12 using a cis-2,6-disubstituted piperidine 55, available from L-alanine, has been performed by Takahata et al. (27) (Scheme 7). Intramolecular amidomercuration (28) of N-alkenylurethane 54 (29), derived from L-alanine, followed by oxidative demercuration (30) gave a mixture (5.5 : 1) of cis(2,6)- and trans(2,6)-piperidino alcohols 55 and 56. Swern oxidation of 55 followed by modification at the ring appendage in the resulting aldehyde 57 using the Horner-Emmons reaction afforded the a$-unsaturated ketone 58, which, on hydrogenation, provided 1 along with indolizidine 195B (59), which has been isolated from the frog Dendrobates as a minor product. Furthermore, the synthesis of the indolizidines 3 and 4 isolated

I96

HIROKI TAKAHATA A N D T A K E F U M I MOMOSE

fic"o - 4 + - rg f

[ent-l2j

-

(+)-I

+

cbr

58

57

59

h

57

SCHEME7. Reagents: a, Ref. 29; b, Hg(OCOCF,),; c , NaBr, NaHCO,; d, 0 2 NaBH4; , e, (COCI),, DMSO, Et3N;f, C4H9COCH2PO(OMe)2, NaH; g, H,, Pd(OH),; h, C2H5COCH2. PO(OMe),, NaH for 60 and C6H13COPO(OMe)2, NaH for 61.

from venom of the ant Solenopsis (Diplorhoptrum) (14) has been achieved via 60 and 61, respectively, derived from the aldehyde 57 in a similar way. One new indolizidine alkaloid, compound 5 (Fig. I), has been isolated from the New Zealand ant Monomorium smithi, together with its monocyclic analog (15). The relative configuration [trans(3,5),trans(3H,SaH)] was deduced by gas chromatographic and mass spectrometric (GUMS) comparison with synthetic samples of the four diastereomers of 3-butyl5(4-penteny1)indolizidine. The stereoselective synthesis of 5 based on hydrogenation of a 2,5-disubstituted pyrrole allowed unambiguous assignment of the relative stereochemistry (Scheme 8).The 2-pyrrolyl keto acid 62 obtained from 5-butyl-2-pyrrolylmagnesium bromide and succinic acid was condensed with hydrazine to form the tetrahydropyridazinone 63. Wolff-Kishner reduction of 63 with potassium hydroxide followed by hydrogenation of the pyrrole ring and cyclization of the resulting pyrrolidine provided the indolizidinone 64. Stereoselective introduction of the pentenyl group at C-5 in 64 was carried out using the cyanoamine method of Stevens and Lee (31) to give 5 , the I3C-NMR chemical shifts of which were very close to those reported for indolizidine 223 (65).

B. INDOLIZIDINE ALKALOIDS FROM AMPHIBIANS 1 . Structure Daly and Spande published a major review on the chemistry, pharmacology, and biology of amphibian alkaloids in 1986 (9). An additional

3.

5

65

n 62

197

SIMPLE INDOLIZIDINE ALKALOIDS

63

64

SCHEME8. Reagents: a, NH2NH2;b, KOH; c , H2, PtO,, AcOH; d, EtOH; e DIBAL; g, HC104; h, KCN; i, 4-pentenylmagnesium bromide.

important review by Daly ef al. deals with the classification of alkaloids isolated from the skin of neotropical poison frogs (Dendrobatidae), and it includes a general survey of toxic or noxious substances in amphibians (32). Accordingly, this chapter introduces up-to-date information. Four simple 3,5-disubstituted indolizidines, namely, indolizidines 223AB (65) (33,34), 239AB (66) (34,351, 239CD (67) (34,35), and 195B (59) (35) (Fig. 2), have been isolated in quantities sufficient for measurement of spectroscopic and physical parameters. All four indolizidines have been shown to be trans(3,5),trans(3H,8aH)-disubstituted indolizidines. One enantiomer of indolizidine 223AB (65), namely, (3R,5R,8aR)-3-butyl-5-propylindolizidine, has been synthesized by Royer and Husson (36) and shown to be levorotatory, indicating that the absolute stereochemistry of indolizidine 223AB (65) is (3R,5R,8aR). Both indolizidines 239AB (66)and 239CD (67) are levorotatory, suggesting that their absolute stereochemistry is the same as that of indolizidine 223AB (65). Recently, (-)-indolizidine 239AB (66) and (-)-indolizidine 239CD (67) have been synthesized by Machinga and Kibayashi (37),and their absolute configurations have been shown to be (3R,5S,8aR) and (3R,5R,8aR), respectively. On the other hand, natural indolizidine 195B (59) is dextrorotatory, and the absolute stereochemistry is tentatively proposed to be (3S,5S,8aS). Indeed, the absolute configuration has been confirmed by the chiral synthesis of (+)-indolizidine 195B (59) (37,38). A number of simple congeneric indolizidine alkaloids, the 5-substituted 8-methylindolizidines, whose mass spectra all show a base peak at mlz 138, have been detected in extracts of skins of members of the family Dendrobatidae of neotropical arrow poison frogs. The base peak arises as the result of loss of the C-5 side chain from the molecular ion. These 5-substituted 8-methylindolizidines, which constitute a new subclass of

198

65 223A0

Hl RO K I T A K A H A T A A N D T A K E F U M I MOMOSE

66 239AB

59 1958

67 239CD

FIG.2. 3.5-Disubstituted indolizidenes.

dendrobatid indolizidine alkaloids, have been found, mainly by G U M S analysis, to be present in certain dendrobatid frogs. Seven indolizidines, namely, indolizidines 203A (68) (39),205A, (69) (40),207A (70)(41), 2338 (71)(39), 235B (72)(do), 235B' (73)(41), and 251B (74)(39), have been isolated in quantities sufficient to permit further characterization (Fig. 3). 'H- and "C-NMR and Fourier transform infrared (FTIR) spectra analysis showed that each compound possesses an equatorial 8-methyl group and an equatorial 5-substituent. The proposed structures for indolizidines 205A (691, 207A (70),235B (72),and 235B' (73)were confirmed by Holmes and co-workers through racemic synthesis (42). In addition, the chiral synthesis of indolizidines 205A (69), 207A (70),and 235B (72)has been achieved by Shishido and Kibayashi (43). All of the synthetic (-)-indolizidines 205A (69), 207A (70),and 235B (72)possess the (5R,8R,8aS)stereochemistry and are levorotatory. The signs of the specific rotations of natural indolizidines 205A (69) and 207A (70)were found to be identical, thereby proving the absolute stereochemistry of natural 205A (69) and 207A (70)to be (5R,8R,8aS). On the other hand, the specific rotation of natural 235B (721,[aID+ 1 I .3", was different

70 207A R= 71 233D R=

72 2358 R= 73 2358' R=

\

-

/

-

\

74 2510 R=

OH

FIG.3. 5-Disubstituted 8-methylindoizidines.

3.

SIMPLE INDOLIZIDINE ALKALOIDS

199

in both magnitude and sign from the value for the synthetic 235B (72) prepared by the two groups ([a], -85.4" or -73.4'). At present, except for natural 235B (72),the known natural indolizidines 203A (68),205A (69), 207A (70),233D (71),and 235B' (73)are all levorotatory. The discrepancy between the optical rotation of the synthetic 235B (72)and that of natural 235B (72)may be the result of contamination of a dextrorotatory impurity in the natural sample. Thus, natural 235B (72)should be levorotatory. The 5,s-disubstituted indolizidines act as noncompetitive blockers at nicotinic receptor channels (44,45). In the 20 years since the initial isolation of pumiliotoxins A [indolizidines 307A' (75)and 307A" (76)]and B (323A) (77)(Fig. 4), Daly has discovered more than 20 members of this family of dendrobatid alkaloids ( 9 ) . The exact constitution of these structurally unusual alkaloids remained elusive until 1980 when an X-ray analysis of the crystalline hydrochloride salt of pumiliotoxin 25 1 D (78)revealed both the structure and absolute configuration of this simple toxin (46). From this information, together with NMR and mass spectral data, it was deduced that the pumiliotoxin A family of dendrobatid alkaloids possessed, as their defining structural element, the (Z)-alkylideneindolizidine ring system. The pumiliotoxin A alkaloids differ from one another primarily in the nature of the side chain attached to C12 of the alkylidene moiety. In the case of the most widely studied toxin, pumiliotoxin B, the stereochemistry of the allylic diol moiety of the side chain [E-alkene and (15R, 16R)] was deduced initially in model systems, and then rigorously confirmed by total synthesis. Another group of more highly functionalized indolizidines, the allopumiliotoxin A alkaloids, bears an additional hydroxyl group on the indolizidine backbone and contains the most complex members of this alkaloid family known to date. The representative allopumiliotoxins 267A (79),323B' (SO), 323B" (Sl), 339A (82),and 339B (83), the latter two of which differ from pumiliotoxin B solely by incorporation of the C-7 alcohol functionality, are shown in Fig. 4. Recently, total syntheses of the allopumiliotoxin A alkaloids 267A (79) (47), 339A (82)(49,and 339B (83) (47) have been performed and their stereostructures rigorously defined. Since the earlier review ( I ) , several new (allo)pumiliotoxin A alkaloids have been isolated and their structures determined. Two N-oxides, allopumiliotoxin 267A (84) and pumiliotoxin 323A (85) (Fig. 5 ) , were isolated from Dendrobates species (41). An alkaloid similar to pumiliotoxin B (323A) was detected from an Australian myobatrachid frog, Pseudophyne coriacea (49).The structure of the compound was assigned as a diastereomer at C-15,C-16 of pumiliotoxin B from the difference in chemical shifts for the signals of the side chain. The presence of a naturally occurring 16s) erythro diastereomer of pumiliotoxin B (86),specifically the (15R,

200

HIROKI TAKAHATA A N D TAKEFUMI MOMOSE

75 A (307A) R=

OH

U

I

76 A (307A) R= OH

77 B (323A) R= &#

OH

78 251D R=CH&H&H,

79 267A R=CH2CH&H3, R'=OH, R2=H

80 3238' R=

82 339A R=

4OH

R'=OH, R ~ = H

&L#R ~ = H R1=OH,

OH

83 3398 R=

OH

R'=H, R*=OH

FIG.4. Representative pumiliotoxin A alkaloids.

isomer, was confirmed by comparison with synthetic derivatives. In addition, from the genus Pseudophryne, pumiliotoxins 267C (87), 277 (88) (perhaps formed by degradation of pumiliotoxin B), and 325B (89) were isolated in trace quantities (Fig. 5) (50). Further skin extracts from Dendrobates purnilio have afforded a trace amount of alkaloids of the pumiliotoxin A class [209F (90), 225F (91), and 307F (92)] and allopumiliotoxin subclass [225E (93), 309D (94), and 325A'/325A (95)], shown in Fig. 6 (40,51). The pumiliotoxin A alkaloids have strong potentials to affect cardiac activity (52-57). The compounds that contain two hydroxyls in the side chain, of which pumiliotoxin B is the most potent, cause marked increases

3.

20 I

SIMPLE INDOLIZIDINE ALKALOIDS

84 N-oxide of 267A

85 N-oxide of 323A

&Ly

Lq;

OH

86 erylhro-323A

87 267C

88 277

89 3258

FIG.5. New alkaloids of the pumiliotoxin A class.

in both the rate and force of contraction of isolated guinea pig atria. Simple natural toxins, such as pumiliotoxin 251D (78) and synthetic congeners having either nonoxygenated side chains or only protected alcohol functionalities, are mild cardiodepressants. The stimulatory activity, which is likely mediated by calcium mobilization, has recently been shown to be initiated by the binding of the toxins to voltage-dependent sodium channels. This event triggers sodium ion influx and phosphatidylinositol breakdown.

90 209F R= CH3

93 225E R=CH3

91 225F R=CH20H

94 309D R=

92 307F R= 0

95 325KI325A" R= OH

FIG. 6. New alkaloids of the pumiliotoxin A class and allopumiliotoxin subclass from Dendrobates pumilio.

202

HIROKI TAKAHATA A N D TAKEFUMI MOMOSE

2. Synthesis The asymmetric synthesis of one of the most simply substituted indolizidines, 5-n-propylindolizidine or indolizidine 167B (96 and 97) ( 9 ) , has been accomplished by three groups. Because this alkaloid occurs in only trace amounts, the absolute configuration has been inferred by analogy to the structurally related indolizidine 223AB (65), whose absolute stereochemistry is known. Polniaszek and Belmont reported the first enantioselective synthesis of indolizidine 167B (96, 97) via the key intermediate amino nitrile 98 prepared from ( S ) - (-)-a-phenethylamine in seven steps (Scheme 9) (58).The amino nitrile 98 served as a pivotal precursor for both diastereomers of alkaloid 167B. Thus, deprotonation of 98 with lithium diisopropylamide (LDA) followed by alkylation with propyl bromide afforded the a-propylated amino nitrile 99. The alkylation reaction produced only one of the two possible diastereomeric products. The orientation of the propyl group in 99 was assigned as equatorial by analogy to the results obtained in alkylation reactions of structurally related cyclic a-amino nitriles (59). Reduction of the amino nitrile 99 with sodium borohydride produced the (SR,9R)-indolizidine 167B (96). Alternatively, reaction of 98 with propylmagnesium bromide afforded the (SS,9R)-indolizidine 167B (97). In a similar way, the indolizidine alkaloids 209D (100 and 101) were prepared from the amino nitrile 98 (Scheme 9). The high stereoselectivity associated with both the hydride reduction of the amino nitriles 99 and 102 and the Grignard addition to the amino nitrile 98 is a result of stereoelectronically controlled nucleophilic addition to iminium ion 103 ac-

H a-phenethylamine

@2 CN

98

Jc

($)b 6 CN

99, R= n-propyl 102, R= n-hexyl

R

96, R= n-propyl 100, R= n-hexyl

0 R

97, R= n-propyl 101, R= n-hexyl

SCHEME 9. Reagents: a, LDA, RBr; b, NaBH,; c , RMgBr.

3.

203

SIMPLE INDOLIZIDINE ALKALOIDS

stereoelectronic control

Nu

CN

H 103

98, R= H 99, R= n-propyl 102, R= n-hexyl

R

Nu

SCHEME 10.

cording to the insightful stereoelectronic principles developed by Stevens (60) (Scheme 10). Jefford er al. synthesized indolizidine 167B from D-norvaline according to the method described for the synthesis of monomorine I (1) (24) (see Scheme 5 ) . The 1-pyrrolylacetic acid 104, prepared by condensation of D-norvaline with 2,5-dimethoxytetrahydrofuran,was converted to the adiazo ketone 105 by reaction of the mixed anhydride, obtained by the action of isobutyl chloroformate, with diazomethane (Scheme 11). Treatment of 105 with silver acetate brought about a Wolff rearrangement, giving the homologous acid 106. Repetition of the mixed anhydride-diazomethane procedure on 106 afforded the corresponding a-diazo ketone 107, and decomposition of 107 with a catalytic amount of rhodium( 11) acetate provided the didehydroindolizidinone 108. The submission of 108 to hydrogen under 15 atm of pressure over Adams’ catalyst under acidic conditions gave the desired (5R,9R)-indolizidine 167B (96). Lhommet and co-workers also synthesized (5R,9R)-indolizidine 167B (96) via diastereoselective reduction of the intermediate 103 (Scheme lo), enantiospecifically prepared from (S)-pyroglutamic acid (61). The first total synthesis of ( + )-indolizidine 195B (59) was accomplished by Yamazaki and Kibayashi (38) starting from the known ketone 13, readily available from L-tartaric acid (62), as illustrated in Scheme 12. Hydride reduction of ketone 13 with zinc borohydride effected a high anti selectivity of over 99: 1, yielding the anti-alcohol 14 via the achelation model, as shown in Fig. 7A. In marked contrast, reduction with L-Selectride displayed an excellent syn selectivity (leading to the

204

HIROKI TAKAHATA A N D TAKEFUMI MOMOSE

-

"

a

D-norvaline

0L:H

b*c

HOOC Pr*Na 104

-

105

e

r

106

f&

-

b,c

COOH

d

-

PriCNa

Pr

107

96

108

SCHEME1 1 . Reagents: a, 2,5-dimethoxytetrahydrofuran,NaOAc; b, isobutyl chloroformate, N-methylmorpholine; c , CH2N2;d, AgOAc; e , R ~ * ( O A C f, ) ~H2/Pt02/HCI. ;

-

-

OMOM

a

diethyl L-tartrate

B n O -

MOMO

0

OMOM + B

MOMO

OMOM

d,e

B n O NPhth

OHC* MOMO

f

OMOM

0

e

w

OH OMOM

OMOM

NPhth

112 b

- /

113

/

on

MOMO

NPhth

111

on

g.h

O

MOW

109

* * ** 110

- /

n

14

-

OMOM

C

on

MOMO

13

-

OMOM

B n O -

nq

OMOM

115

114

+

/

HO

OMOM

i-k

-H>

116

Cbz 117

Cbz 120

(+)-59

(+)-121

SCHEME12. Reagents: a, Ref. 62; b, Zn(BH,) or L-Selectride; c , phthalimide, Ph3P, DEAD; d, H2, 10% Pd/C; e, (COC1)2,DMSO, Et3N; f, CH2=CH(CH2)3MgBr;g, NH2NH2; h, CbzCI, Na2C03;i, MeS02CI, Et3N;j, 'BuOK; k, HCI; I, 2,4,5-triiodoimidazole,imidazole, Ph3P, Zn; m, 02,PdCI2, CuCI2, DMF-H20.

3.

205

SIMPLE INDOLIZIDINE ALKALOIDS si attack

0-

I

A a-chelation model

si attack

I

B P-chelation model

\

OMOM

C Felkin-Anh model

FIG.I .

syn-alcohol 109) of 92: 8, possibly via the P-chelation model (Fig. 7B) and/or the Felkin-Anh model (Fig. 7C). Mitsunobu reaction of the synalcohol 109 with phthalimide afforded 110 with stereoinversion, which was converted to aldehyde 115 by debenzylation followed by Swern oxidation. Grignard reaction of 115 proceeded in a nonstereoselective manner, yielding a mixture of diastereomeric alcohols 112, which were transformed into the ketone 114 via the sequence involving removal of the phthaloyl group, benzyloxycarbonylation, and Swern oxidation. The same diastereoselective behavior toward borohydride reagents as for 107 was encountered in the reaction of 114 by following the predicted pattern. Thus, as shown in Scheme 12, anti,syn,anti- and syn,syn,anti-alcohols 115 and 116 were obtained by employing zinc borohydride and L-Selectride, respectively, with excellent facial selectivity. The syn,syn,anti-amino alcohol 116 was subjected to mesylation, basepromoted cyclization, and removal of the methoxymethyl (MOM) group to provide the pyrrolidine 117. The trans-diol 117 was converted to the Z-olefin 119 via epoxide 118 by treatment with triiodoimidazole, triphenylphosphine, and zinc. Wacker oxidation was applied for site-selective oxidation of the terminal olefin, affording the methyl ketone 120, which, on hydrogenation over palladium on carbon, provided ( + bindolizidine 195B [( +)-591 along with its C-5 epimer (+)-121 in an 86 : 14 ratio. In addition, (-)-indolizidine 195B [(-)-591 was synthesized from 14 in a similar manner by Kibayashi. Recently, total syntheses of both enantiomers of all four 3S-disubstituted indolizidines [indolizidines 195B (59), 223AB (65), 239AB (66), and 239CD (67)] have been reported (37), starting from 3,4-dideoxy-~-threohexitol (125) as a single and common chiral synthon. The retrosynthesis for a general approach to the series of (-) enantiomers is illustrated in Scheme 13. This approach suggests a 2-fold disconnection providing the (R,R)-2,5-dialkylated pyrrolidine 123, having a C2 symmetrical structure, as an advanced intermediate, which might be prepared from the C2 symmetrical (R,R)-diepoxide building block 124. This synthetic strategy is

206

HIROKI TAKAHATA A N D T A K E F U M I MOMOSE

(-)-59, R=Me, X=H (-)-65, R= n-Pr, X=H (-)-66. R= -(CH2)30H, X=H (-)-67, R= n-Pr, X=OH

125

(R.4-124

(+)-59,R=Me, X=H (+)-65, R= n-Pr, X=H (+)-66,R= -(CH2)3OH. X=H (+)-67, R= n-Pr, X=OH

-

n .A0

(S,S)-124

SCHEME 13. Retrosynthesis of both enantiomers of indolizidines 195B (59), 223AB (65), 239AB (a), and 239CD (67).

adaptable also for the synthesis of the series of ( + ) enantiomers using the (S,S)-diepoxide 124. Both enantiomers of the diepoxides, (R,R)-124 and (S,S)-124, are available by utilizing 125, which is readily prepared from D-mannitol. The selective protection of 125 with pivaloyl chloride afforded the 1,6bispivalate 126, which was converted to (R,R)-124by mesylation followed by treatment with base. The attempt to prepare (S,S)-124 aimed at the regioselective introduction of bromine at C-1 and C-6 in order to obtain an efficient precursor for epoxidation. After bisacetalization of 125, exposure of the resulting bisacetal 127 to N-bromosuccinimide (NBS) led to regioselective ring opening (63),furnishing the I ,bdibromide 128, which was converted under basic conditions to (S,S)-124 (Scheme 14). First, (-)-indolizidine 239CD [( -)-671 was synthesized from (R,R)-l24 (37). The diepoxide (R,R)-124 was converted by a copper( I)-mediated coupling reaction to the diol 129, which subsequently was transformed by a known method (64) into 130. Benzoylation followed by desilylation of 130 yielded the diol 131, which was converted by the Sharpless procedure (65) to the cyclic sulfate ( +)-132. Subsequent nucleophilic ring opening of (+)-132with LiN, proceeded in a nonregioselective manner, providing an inseparable 1 : 1 mixture of isomers (133a,b), which was then converted to a 1 : 1 mixture of the corresponding mesylates 134a,b. Hydro-

3.

D-mannitol

-

207

SIMPLE INDOLIZIDINE ALKALOIDS

OH

HoO -H

-

Ph 04-H

d

OH 125

0"-O P h y

127

la OH l B u C O , W OCOBu' OH 126

QCOPh

:& Br Br OCOPh 120

wo

0

( R ,R)-124

(S,S)-124

SCHEME14. Reagents: a, 'BuCOCI, Py; b, MsCI, EtlN; c, NaOH; d, PhCHO, TsOH; e, NBS.

genation of the azide function afforded the trans-pyrrolidine 135 as a single product via intramolecular cyclization. N-Protection of 135, followed by alkaline hydrolysis, afforded the pyrrolidine alcohol 136. Oxidation of 136 with pyridinium dichromate (PDC), followed by homologation of the resulting aldehyde 137 via reaction with the Grignard reagent n-PrMgBr and oxidation of the resulting alcohol, furnished the ketone 138. Hydrogenation of 138 provided the cyclization product 139, which underwent further hydrogenolysis under acidic conditions, thereby affording (-)indolizidine 239CD [(-)-671 as a single isomer (Scheme 15). An identical strategy was applied for the synthesis of the ( + ) enantiomer [(+)-671 starting from the (S,S)-diepoxide 124 via the cyclic sulfate ( 9 - 1 3 2 (Scheme 15). Having established a route to both enantiomers of indolizidine 239CD (67) in a stereo-defined manner via the trans-2,5-dialkylated pyrrolidine intermediate, the synthesis of the other w-hydroxy congener indolizidine 239AB [(-)-661 was achieved by a methodology very similar to that used for the synthesis of indolizidine 239CD (67). The pyrrolidine 136, prepared from the epoxide (R,R)-124, was converted via several steps, as illustrated in Scheme 16, to the pyrrolidino ketone (-)-143, which on catalytic hydro-

208

HIROKI TAKAHATA A N D TAKEFUMI MOMOSE

a

(R.R)-124

OTBDMS

OH

e

n OBn

On

b

OH

e

n

~

B

on

n OCOPh

OH

OTBDMS

129

e

130

131

(+)-132

133b R=H 134b R-MS

k.1

c

137

135, R= H, X= PhCO 136, R= Cbz, X=H

A L L O Bi n Cbz

OR

N-

138

(-)-132

(+)-67

SCHEME 15. Reagents: a, BnO(CH2)3MgBr, Cul; b, Ref. 64;

c, PhCOCI, DMAP; d, Bu4NF; e, SoC12, Et3N; f, RuC13, NaIO,; g, LiN3; h, aq. H2S04; i, MsCI, Et3N;j, H2, P d C k, CbzCI, K2C03;I, aq. KOH; rn, PDC, CH2CI2;n, "PrMgBr; 0 , H,, Pd/C, HCI.

genation gave indolizidine 239AB [(-)-661 and its C-5 epimer 144 in an 88 : 12 ratio. In an analogous manner, the aldehyde (-)-142 was readily converted to (-)-indolizidines 195B [(-)-591 and 223AB [(-)-651. Thus, (-)-142 was subjected to a Grignard reaction using MeMgBr and n-PrMgBr to afford the alcohols (2R,5R)-145 and (2R,5R)-146, respectively, which were converted to the corresponding ketones (-)-147 and (-)-148. Finally, hydrogenation of ketones (-)-147 and (-)-148 produced the unnatural (-) enantiomer [(-)-591 of indolizidine 195B along with its C-5 epimer (-)-121 in

3.

209

SIMPLE INDOLlZlDINE ALKALOIDS

-,...aH f,e

Cbz

(-)-142

0

-\.-

O0n

g.c

Cbz

(-)-143

(-)-66

+ 144, C-5 epirner

SCHEME 16. Reagents: a, TsCI, DMAP; b, LiAlH,; c, H2. PdlC, HCI; d, CbzCI, aq. K$03; e , PDC, CH2CI2;f, B I I O ( C H , ) ~ M ~ B g,~H2. ; Pd/C.

a ratio of 89 : 1 1 and (-)-indolizidine 223AB [(-)-651along with its C-5 epimer (149)in an 86 : 14 ratio, respectively (Scheme 17). Preparation of the ( + ) enantiomers of indolizidines 195B [( + )-591, 223AB [( +)-651,and 239AB [( +)-MIwas achieved using a similar strategy starting with the (S,S)-diepoxide 124. Thus, the (S,S)-diepoxide 124 was converted to truns-pyrrolidine 150 according to the procedure described for ( +)- and ( -)-67(Scheme 15). N-Protection of 150,followed by alkaline hydrolysis, produced pyrrolidino alcohol ( +)-141,which was oxidized to the aldehyde (+)-142.Following the same procedure as described for the (-) enantiomers of indolizidines 195B, 223AB, and 239AB (Schemes 16 and 17), (+)-142was subjected to homologation of the alkyl side chain, followed by cyclization of the resulting ketones ( + )-143,( + )-147,and (+)-148,to yield the natural ( + ) enantiomer of indolizidine 195B [( + 1591 and the unnatural ( + ) enantiomers 223AB [( +)-651and 239AB [( +)661,respectively (Scheme 18). A short synthesis of indolizidine 195B [( +)-591was performed by using the intramolecular amidomercuration of an N-alkenyl urethane (151)available from L-norleucine (27). The known 151 (66) underwent stereoselective, intramolecular amidomercuration followed by reductive oxygenation to provide the truns-pyrrolidino alcohol 152, Swern oxidation of 152,followed by Wittig homologation of the resulting aldehyde, afforded the olefin 153,which was converted, by hydrogenation followed by benzyloxycarbonylation, to the same intermediate [( + )-1411as described above (Scheme 18). Swern oxidation of (+)-141,followed by a Wittig reaction, provided the olefin 154. Wacker oxidation of 154,followed by hydrogenation of the resulting ketone (+)-147,gave the natural indolizidine

210

HIROKI TAKAHATA A N D TAKEFUMI MOMOSE

-

-

a

(-)-I42

b

- \ * ' ' m R Cbz

Cbz

(2559-145, R= Me

(-)-147, R= Me

(2R.5R)-146, R= n-Pr

(-)-I@, R= n-Pr

%

%

v-

(-)a

(-)-65

+

+

(-)-121, C-5 epimer

149. C-5 epimer

SCHEME 17. Reagents: a, MeMgBr or "PrMgBr;b, PDC, CH2C12;c, H2, Pd/C.

Cbz

H

(+)-141

150

-

0-

as in Scheme 17 d

'

'

*

>

A

cbz (+)-I42

H

c

7

\ 2l7

(+)-59

as in Scheme 16

./'

p

HO

\A

(+)-66

SCHEME 18. Reagents: a, Ref. 64; b, CbzCI, aq. K2C03;c, aq. NaOH; d, (COC1)2,DMSO, Et,N.

3.

21 1

SIMPLE INDOLIZIDINE ALKALOIDS

195B [(+)-591 along with its C-5 epimer (+)-Elin a ratio of 84: 16 (Scheme 19). During the preparation of this chapter a novel synthesis of (-)-65 using an enantioselective RuC1,/2,2'-bis(diphenylphosphino)- 1,l '-binaphthyl (B1NAP)-mediated reduction (67) of p-keto esters was published by Taber el al. (68). Holmes and co-workers reported the first synthesis of 5-substituted 8methylindolizidines using a highly stereoselective, intramolecular nitrone cycloaddition of the (Z)-N-alkenylnitrone 155 as a key step (42). Intramolecular dipolar cycloaddition of 155, prepared by condensing the hydroxylamine 156 with acetoxybutanol, afforded the oxazolidine 157 as the only isolated product. Subsequent conversion of 157 to the corresponding mesylate resulted in spontaneous intramolecular cyclization to yield a fivemembered ring. Reductive cleavage of the N - 0 bond resulted in the construction of an indolizidine skeleton (158) with complete stereocontrol at the centers C-5, C-8, and C-8a (Scheme 20). The key axialhydroxymethyl compound 158 was epimerized via the aldehyde to the equatorial alcohol 159, which was subsequently reduced to give indolizidine 205A [(?)-691. Lindlar reduction of (?)-69 then afforded indolizidine 207A [(?)-701. An asymmetric synthesis of the indolizidine 209B [(-)-la] was achieved by using a homochiral N-alkenylhydroxylamine 161 derived from (S)-5-(hydroxymethyl)-2-pyrrolidinone(162) by a chainextension sequence (Scheme 20). Thus, the nitrone precursor 163, obtained from 161, was converted via 164, in a strategy similar to that described for the synthesis of the racemic form, to (-)-209B [(-)-la].

VvQ..-OB" Cbz 153

_f,g,h

d,i 4..,,ApOH Cbz

__

(+)-141

d..*,- 4.%2 f

(+)-59+ (+)-I21

Cbz

154

Cbz

(+)-I47

SCHEME19. Reagents: a, Ref. 66; b, Hg(OAc),; c, 02,NaBH4, DMF; d, (C0C1)2, DMSO, Et3N; e , Ph3P*(CH2)30BnBr-, "BuLi; f, H2, Pd(OH)2;g, H2. W/C, HCI; h, CbzCI, NaOH; i, Ph3P+MeI-,"BuLi; j, 02,PdCI2, CuC1, DMF-H20.

212

HIROKI TAKAHATA A N D TAKEFUMI MOMOSE

c-0

-

/-Ya

&VO d

-/-

158 158

c c

.." OH

159

(*)-69

(+)-70

12 steps

O-OHN H 162

NHOH

L - L

n-CsH11

" - C , H l l y A c O U

161

b

0163

(-)-160 164

165. R= (CH2)3CCCH2CH,

(4-72, R = (Z)-(CH&CH=CHCH&H3

166. R= (CH,)&H=CH,

(+)-73,R = (CH2)5CH=CHZ

SCHEME 20. Reagents: a, CHzC12; b, toluene; c , K2C0,; d, MsCI, Et,N; e , Zn, AcOH-HzO; f, (COCI)z, DMSO, Et3N; g, K,CO,; h, NaBH4; i, LiEt,BH; j, Hz,Lindlar catalyst.

In addition, the total synthesis of racemic indolizidines 235B [(2)-721 and 235B' [( ?)-73]was performed by the intramolecular thermal cycloaddition of the (Z)-alkenylnitrones 165 and 166, respectively (Scheme 20) (69). Marazano and co-workers reported a short synthesis of a 5-substituted 8-methylindolizidine, alkaloid (+)-209B [( +)-1601, using a new chiral 1,4-

3.

NO2

170 169

168

d.e

213

SIMPLE INDOLlZlDlNE ALKALOIDS

-

n Rl

M N

e R2

,#.-Me

+ R

N

O M e "R2

OH

OH 172

171

167

173

+ R'"' N

"R2

OH

R'= n-C5Hll

-

0

R'

R2=

do]

(+)-160

174

SCHEME 21. Reagents: a, (R)-phenylglycinol; b, Na2S20,, K2C0,; c, alumina; d, nCSHIlMgBr; e, R2MgBr; f, H2, H + .

dihydropyridine (167) (70). Treatment of Zincke's salt 168 with (R)phenylglycinol afforded pyridinium salt 169 having a chiral carbon atom linked directly to the nitrogen of the pyridine ring (71). Reduction of 169 with Na,S,O, provided the unstable 1,Cdihydropyridine 167, which was isomerized into a mixture of oxazolidines 170 and 171. Grignard reaction of the oxazolidine mixture, followed by homologation using a second Grignard reaction, provided the piperidines 172, 173, and 174 in a ratio of 5 : 3:2. Hydrogenation of the major isomer 172 in an acidic medium directly furnished indolizidine (+)-209B [( +)-1601(Scheme 21). The enantioselective total synthesis of indolizidine alkaloids (-)-205A [(-)-691 and (-)-235B [(-)-731 was achieved via the common intermediate a-aminonitrile 175 by Polniaszek and Belmont (72). Absolute stereochemical control over the C-8 and C-8a stereocenters was achieved by a stereoselective reaction between the chiral acyliminium ion generated from the sulfonyl lactam 176 and crotylmagnesium chloride, resulting in formation of a mixture of two crotyl lactams 177 and 178 in the ratio 7 : 3, which was hydroborated to afford a mixture of the primary alcohols 179a,b. Swern oxidation of the major isomer 179a, followed by Wittig homologation and subsequent acetalization, provided the dimethyl acetal 180. Reduction of 180 to the corresponding pyrrolidine, and subsequent hydrogenolytic removal of the chirality-directing group, afforded the amino acetal 181, which was converted to the key intermediate 175. Alkylation of the amino nitrile 175, followed by reduction with sodium borohydride of the iminium derived from the resulting a-amino nitrile 181 according to the procedure described previously (Schemes 9 and lo), produced indolizidines 205A [(-)-691 and 235B [(-)-721 (Scheme 22). The total synthesis of (-)-indolizidines 205A, 207A, 209A, and 235B via a common chiral oxazino-lactam (184), itself prepared from the N -

214

HIROKI TAKAHATA A N D TAKEFUMI MOMOSE

176

180

A

177

178

181

179a

175

A

182. R= (CH2)jCCTMS

(-)-69.R= (CH2)jCCH

183, R= (Z)-(CH&CH=CHCH,CH3

(-)-72. R= (Z)-(CHZ)jCH=CHCH&H,

SCHEME22. Reagents: a, rrans-crotylmagnesiurn chloride, ZnBr,; b, (Sia)2BH;c, H,Ol; d , separation; e, (COCI),, Et3N,DMSO; f, Ph3P=CHOMe; g. camphorsulfonic acid, MeOH; h, LiAIH,; i, 10% Pd/C, NH4+HCO?-, j, KCN, HCI; k , LDA, RX; I, KF; m, NaBH4.

acylnitroso intermediate 185 by an asymmetric intramolecular Diels-Alder reaction, was reported by Shishido and Kibayashi (43,73).The precursor hydroxamic acid 186 was prepared from the known (R)-4-methyl-5hexenoic acid (187) (74) in seven steps. Oxidation of 186 generated the N-acylnitroso 185, which spontaneously underwent intramolecular [4 t 21 cycloaddition to afford a 1.8 : 1.0 mixture of rrans- and cis-bicyclic oxazinolactams (184 and 188). Subsequent introduction of the side chain (the C-5 moieties of indolizidines 205A, 207A, 209A, and 235B) was accomplished with full stereochemical control using the dihydro-oxazinolactam 189. Grignard reaction of 189 followed by NaBH, reduction under acidic conditions (through stereoelectronically controlled addition of hydride ion to the transient iminium ion 190) afforded the single products 191, 192, and 193, depending on the Grignard reagent used. Reductive cleavage of the N - 0 bond in 191 followed by cyclodehydration of the resulting piperidine 195 using PPh,/CBr,/Et,N provided the indolizidine 198. Removal of the trimethylsilyl group from 195 furnished indolizidine 205A [( -)-69]. Similarly, compounds 192 and 194 were converted to indolizidines 207A [(-)-701 {and to 209B (-)-1601) and 235B [(-)-721, respectively, as shown in Scheme 23. Overman and Bell reported earlier the enantioselective synthesis of pumiliotoxin 25 1 D (79) by use of an iminium-vinylsilane cyclization (46),

3.

Me

7steps

HO&

0

-

a

[ 03Me]

NHOH 187

184

215

SIMPLE INDOLIZIDINE ALKALOIDS

186

188

.$%.$y f

185

L

189

J

190

.\>.q#MLoH H

Q

191, R= (CHZ)3CCSi(CH3)3

195, R= ( C H ~ ) ~ C C S I ( C H ~ ) ~

192, R= (CH,),CH=CHp

196, R= (CH,),CH=CHp

193, R= ( C H Z ) ~ C C C H ~ C H ~

197, R= (Z)-(CH2)3CH=CHCHpCH3

[194,

R= (Z)-(CH2)3CH=CHCH2CH3

[I

198. R= (CH,),CCSI(CH,)3

h

(-)-69,R= (CH2)3CCH (-)-70,R= (CH2)3CH=CH2 (-)-160, R= (CH2)4CH3 (-)-72, R= (Z)-(CH2)3CH=CHCHzCH3

SCHEME 23. Reagents: a, “Pr4NI04;b, H2. Pd/C; c , RMgBr; d, NaBH4, AcOH; e , Hz, Pd, BaSO,, quinoline; f, Zn, AcOH; g, Ph3P, CBr4, Et3N; h. KOH.

and a full account together with the total synthesis of pumiliotoxin B (323A) (78) has since been published (75).The same approach was applied to a convergent synthesis of purniliotoxin A (307A’) (75) (76). The fully elaborated side chain segment of purniliotoxin A, (-)-silylalkyne 199, was -)-2-methyl- l-pentene-3-01(200) in eight steps. Alkyne prepared from (S)-( 199 was converted to the silylvinyl alanate and coupled with the (S)proline-derived epoxide 201 to give the bicyclic carbarnate 202. Hydrolysis of 202, followed by addition of formalin, afforded the key intermediate 203, which was cyclized via irniniurn-vinylsilane cyclization to provide the indolizidine 204. Debenzylation of 204 afforded purniliotoxin A (307A’) [( +)-751 (Scheme 24). Furthermore, Overrnan and Sharp reported an alternate cyclization strategy in which the Z stereochemistry of the alkylidene side chain resulted from the antarafacial stereochemistry of a reductive iminiurn ionalkyne cyclization (77). Coupling of the epoxide 201 with the alkyne 205, followed by hydrolysis of the carbamoyl moiety, yielded the important

216

HIROKI TAKAHATA A N D TAKEFUMI MOMOSE

201

f

25

c

CH3

CHI

CH3

c C:; H

203

SCHEME24. Reagents: a, BuiAIH; b, MeLi; c, 201; d, KOH; e, formalin; f, PPTS, pyridine; g, Li, NH,.

intermediate 206. The iodide-promoted iminium ion cyclization of 206, by treatment with paraformaldehyde, in aqueous acidic media containing a large excess of iodide ion, provided the alkylidene indolizidine 208 via 207, which was converted by deiodination and debenzylation to ( + )pumiliotoxin A (307A') [( +)-751 (Scheme 25). Two groups have succeeded in the total synthesis of allopumiliotoxin 339B [( +)-831. Overman and Goldstein reported a short, highly stereocontrolled synthesis of ( + )-83, together with that of allopumiliotoxin 267A [( + )-791, from the indolizidino ketone 209, assembled from L-proline (47).

205

201

206

e.f,g 207

(+)-75.R= X= H

SCHEME 25. Reagents: a , 205, "BuLi;b, Et2AICI, 201; c , Ba(OH)*;camphorsulfonic acid, (CH,O),,, NaI; e, "BuLi; f, MeOH; g, Li, NH3.

3.

SIMPLE INDOLIZIDINE ALKALOIDS

217

The N-tert-butoxycarbonyl (N-Boc) acetylpyrrolidine 210 was deprotected to afford the labile 2-acetylpyrrolidine salt 211, which was directly treated with excess I-lithio- I-methoxyallene to provide the allenylpyrrolidine 212 as a single diastereomer (diasteroselectivity >97 : 3) resulting from the chelation control operating between the lithium cation and the free amine. Cyclization of the allene 212 with p-toluenesulfonic acid, followed by hydrolysis of the resulting bicyclic enol ether 213, gave the 7-indolizidinone 209. Aldol condensation of 209 with aldehyde 214 followed by dehydration afforded the enone 215. Stereoselective reduction of 215, followed by silylation of the resulting allylic alcohol, provided the indolizidine 216. Debenzylation of 216 followed by Swern oxidation furnished the aldehyde 217, the coupling of which with the known ylide 218 provided the a’-silyloxylated (E)-enone219. Threo-selective reduction of 219 with LiAIH, was accompanied by desilylation to afford (+)-allopumiliotoxin 339B [( +)-831 (Scheme 26). Trost and Scanlan reported a novel synthesis of ( +)-allopumiliotoxin 339 B [( + )-831 using an innovative, Pd(0)-catalyzed 6-endo-trigonal mode of cyclization of the vinyl epoxide 224 (78). Thus, the treatment of the Lproline-derived ketone 211 with the allyltitanium reagent 221, followed by epoxidation, afforded 220, which was diastereofacially cyclized with Pd(0) to provide the indolizidine 223 as a single isomer. Hydroxyl groupdirected epoxidation of 223 gave the vinyl epoxide 224, which, under Pd(0)-induced condensation with the ally1 sulfone 225, provided the glycol 226 possessing the side chain profile of the target alkaloid. Desulfonylation of 226 and subsequent threo-selective reduction, accompanied by concomitant desilylation, gave ( + )-allopumilitoxin 339B [( +)-831 (Scheme 27). The first total synthesis of ( + )-allopumiliotoxin 339A [( + )-82], an indolizidinediol containing a @-oriented C-7 hydroxyl, was achieved via the pivotal, nucleophile-promoted iminium ion-alkyne cyclization step by Overman et ul. (48). Alkyne 227, provided with the full side chain profile of the pumiliotoxin A alkaloids (77, 82, and 83), was prepared from (R)2-methyl-4-pentenol (228) in eight steps. Addition of the lithio derivative of 227 to the a-benzyloxyaldehyde 229 provided the alcohols 230 and 231 with 4 : I selectivity. Treatment of 230 with silver triflate afforded the pyrrolidino-oxazine 232, the precursor for the key cyclization step. Iodinepromoted cyclization of 232 provided the alkylidene-indolizidine 233, with no other stereoisomer. Deiodination of 233, followed by debenzylation afforded ( + )-allopumiliotoxin 339A [( +)-82] (Scheme 28). Both the Overman and Trost groups have used various strategies based on (S)-proline for the synthesis of the indolizidine skeleton. Gallagher has reported the enantioselective synthesis of pumiliotoxin 25 ID (78) employing an allene-based electrophile-mediated cyclization and the rapid

218

HIROKI TAKAHATA A N D TAKEFUMI MOMOSE

210

21 1

21 2

e-g

d

3

213

209

21 5

CH3 21 4

-

I

h,i

"OTDMS

"OTDMS

216

21 7 218

21 9

(+)-a3

(+)-79

SCHEME 26. Reagents: a , CF3C02H;b, 1-lithio-1-methoxyallene; c, TsOH; d, HCI; e, Ph3CLi;f, 214; g, (CF3C0)2,DBU; h, NaBH4, CeC13;i, TBDMSCI, n-BuLi; j, Na, NH3; k, (COCI),, DMSO, EtjN; I, 218; m, LiAIH4.

elaboration of the indolizidine skeleton by the correct choice of an electrophilic trigger (79). Pd( 11)-mediatedcyclization of the enantiomerically pure allenic arnine 234 provided the 2-substituted pyrrolidines 235a,b with a negligible level of diastereoselectivity (1 : 1). Reduction of 235a to the corresponding allylic alcohol, followed by Claisen rearrangement, effected homologation to the ester 236. Hydrolysis of 236 and reaction of the resulting carboxylate salt with acetic anhydride led directly to the bicyclic lactam 237 as a single enantiomer. Hydration (hydroxymercuratiod reduction) of 237 provided the tertiary alcohol 238 with a high level of stereoselectivity (10: 1) in favor of the expected axial alcohol. The introduc-

3.

219

SIMPLE INDOLIZIDINE ALKALOIDS

+ a __c

SCzH,

-

222 21 1

220

('PrOkTi -SC,H,

221

-

-

C

223

224

0 225

226

(+)-83

SCHEME27. Reagents: a, 221, (CF3CO),0; b, (CH3),0BF4, NaOH; c , catalytic (dba)3Pdz-CHC13,(RO),P, H,O; d, CF3C03H;e , 225, (dba)3Pd2-CHC13,dppf, H,O; f. Na(Hg), NaHPO,; g, LiAIH4.

CN

b

230, R'= OH, R2= H 231, R'=

232

H,R ~ OH = CHI OH CH3,,.& OH

d.e

r

233, R= En, X=l (+)-82.R= X=H

SCHEME28. Reagents: a, 227, "BuLi; b, AgOS02CF3;c, TsOH, NaI, (CH,O),, acetone-H20; d, "BuLi, MeOH; e , Li, NH3.

220

HIROKI TAKAHATA A N D TAKEFWMI MOMOSE

tion of the (Z)-alkylidene unit at C-6 was performed by an aldol condensation of 238 with (R)-2-methylhexanal followed by syn elimination and anti elimination of the resulting alcohols 239a and 239bc,respectively. Finally, 1,2-reduction of the unsaturated lactam moiety of the (Z)-alkylidene indolizidine (Z)-240 thus obtained was achieved by using LiAICI, to afford pumiliotoxin 251D [( +)-781 (Scheme 29). Honda et al. reported a 16-step synthesis of the key intermediate 237 from the product (R)-241 of the Katsuki-Sharpless kinetic resolution of the racemic 2-furylmethanol 241, derived from 2-lithiofuran and 5-trimethylsilyl-4-pentynal (80).

d,e

b,c PhACH3

235a

234

236

235b, epimer at C-2

on

OH

OH

f.g '0 1.1.

237 I

1

239a,b,c

(0-240

16steps

li

TMS

(R)-241

EH,

(+)-78

EH3

(2)-240

SCHEME 29. Reagents: a, 1% PdCl,, CO, CH30H, CuCI,; b, DIBALH; c, (EtO),CCH,, H + ;d, NaOH; e, (CH,CO),O; f, Hg(02CCH3)2;g, NaBH,, NaOH; h, LDA, (R)-2-methylhexarial; i, DCC, Cu(I)Cl;j, CH3S02C1, pyridine, KOH ( E : Z = 2.6: 1); k, LiAIH,-AICI,; I, Ref. (80).

3.

SIMPLE INDOLIZIDINE ALKALOIDS

22 1

111. Elueoculpus Alkaloids

Following the discovery (81) of the Elaeocarpus alkaloids, several total syntheses concerning racemic elaeokanines were reported (82). However, the absolute configuration of these alkaloids remained unknown. The first asymmetric total synthesis of elaeokanines A (242) and B (243) via an optically active a-sulfinylketimine (244) was achieved by Hua et af.,which permitted the absolute stereochemistry of these alkaloids to be established (83). (+)-(R)-Sulfinylketimine 246, prepared by reaction of the anion of pyrroline 245 with (-)- 1-menthyl p-toluenesulfinate, was alkylated with 1,3-diiodopropane to provide the key bicyclic intermediate 244. Reduction of 244 with NaBH, gave the four separable diastereomeric indolizidines 247a,b and 248a,b in the ratio 4 : 4 : 1 : 1 . The stereochemistry at C-8a of the sulfoxides was determined by desulfurization of each isomer, which gave either a (-)-indolizidine or a ( )-indolizidine. a-Hydroxybutylation of the major isomer 247a or 24713 furnished a mixture of the alcohols 249a and 24913 in a 2 : 1 ratio, whereas the similar reaction of each minor isomer 248a,b provided a 2 : 1 mixture of 250a,b. Single-crystal X-ray analysis of 249a established the relative stereochemistry. Dehydrosulfinylation of 249a or 249b furnished the natural enantiomer (-)-elaeokanine B [( -)2431, which was oxidized to afford the unnatural antipode (-)-elaeokanine A [(-)-2421. Under the same reaction conditions, dehydrosulfinylation of 250a,b provided (+)-251, which was oxidized to afford the natural enantiomer (+)-elaeokanine A [( +)-2421 (Scheme 30). Comins and Hong reported the asymmetric total synthesis of elaeokanine C (2521, together with that of elaeokanine A (242), using a chiral dihydropyridone intermediate (84). Reaction of the chiral l-acylpyridinium salt 253, prepared from 4-menthoxy-3-(triisopropylsilyl)pyridine(85) and the chloroformate of (-)-8-(4-phenoxyphenyl)menthol,with the Grignard reagent 254 gave the alcohol 255 in 92% diastereomeric excess, which was converted to the chloride 256 by treatment with triphenylphosphine and N-chlorosuccinimide (NCS). On removal of the chiral auxiliary with sodium methoxide, concomitant cyclization occurred to afford the enone 257. Carbamylation of 257 followed by desilylation yielded the amide 258. Stereoselective reduction of the enaminone moiety of 258 using catalytic hydrogenation over PtO, gave a mixture of alcohols 259a,b in a 95 :5 ratio. Treatment of 259a with cerium chloride and n-propylmagnesium chloride gave elaeokanine C [( +)-2521. Because the optical rotation for the isolated natural (-)-elaeokanine C is levorotatory, it was demonstrated that the absolute configuration is (7R,8S,8aS). Elaeokanine C was converted to

+

222

HIROKI T A K A H A T A A N D T A K E F U M I MOMOSE

U 245

246

d

247a,b

244

G!J;+< 249a

+

e l (-)-243

(-)-242

249b

248a,b

250a

(+)-251

(+)-242

250b

SCHEME 30. Reagents: a, LDA, (-)-(S)-I-menthylp-toluenesulfinate;b, LDA, 1,3-diiodoPropane; c, NaBH,; d, LDA, butyraldehyde; e, toluene; f, pyridinium chlorochromate.

+ )-elaeokanine A [( + )-2421 by treatment with sodium hydroxide (Scheme 31). The asymmetric synthesis of elaeokanines A and C based on a novel approach involving diastereoselective addition of an enolic nucleophile to a tricyclo-iminium species, and subsequent removal of the stereo template by a retro-Diels-Alder reaction, was achieved by Koizumi and co-workers (86). Oxidation of the maleimide 260, prepared by the addition of 10mercapto-isoborneol to N-tert-butyldimethylsilyl (N-TBDMS) maleimide followed by chlorination and dehydrochlorination, afforded the sulfinylmaleimide 259 as a single diastereomer. Diels-Alder reaction of 259 with cyclopentadiene produced the em-sulfinyl adduct 262 with greater than 99% diastereomeric excess (87). Desilylation of 262 and subsequent N alkylation gave the acetal 263. Regioselective reduction of 263 followed by desulfinylation with samarium( 11) diiodide afforded the y-hydroxylactam 264, which was transformed into 265. Addition of 2-(trimethylsilyloxy)pent-I-ene to 265 in the presence of BF,-Et,O provided the lactam 266 as a single diastereomer possessing a p-oriented five-carbon chain at

(

3.

258

SIMPLE INDOLIZIDINE ALKALOIDS

259a, R'= OH, R2=H

223

(+)-252

259b. A'= H. R2= OH

SCHEME 31. Reagents: a. 254; b, H,O+ ;c , Ph,P, NCS; d, NaOMe; e, LDA, dirnethylcarbamoyl chloride; f, (COOH),; g, HI, PtO,; h, PrMgCI, CeCI,; i. NaOH.

C-5. Acid-catalyzed aldol reaction of 266 afforded the tetracyclic lactam 267, which was converted to the dehydroindolizidinone 268a,b, separable diastereomers at C-8a, in a 3 : 1 ratio by flash vacuum pyrolysis (retroDiels-Alder). Hydrogenation of the major isomer 268a and subsequent protection of the ketone produced the lactam 269. Reduction of 269 followed by acid hydrolysis gave elaeokanine C [( +)-2521, which was transformed into elaeokanine A [( + 1-2421 according to the same procedure as described in Scheme 31 (Scheme 3 2 ) .

IV. Slaframine Forage materials contaminated with fungus Rhizocronia legitminicola (88) are responsible for a disease in ruminants known as black patch (89). The most obvious symptom associated with ingestion of contaminated feed is excessive salivation, which is thought to be caused by the alkaloid slaframine (270). Slaframine has previously been mentioned on several occasions in this treatise. The chapter by Howards and Michael offered a comprehensive review, covering the isolation, structure, biosynthesis, synthesis, and biological activity of this alkaloid ( I ) . Thereafter, further biological studies on slaframine included a potentially important use of the alkaloid in deliberately altering ruminal fermentation to improve the efficiency of feed utilization, lactation, and growth in sheep and cattle, and the ability of slaframine to stimulate circulating concentrations of a

224

HIROKI TA K A H A T A AND T A K E F U M I MOMOSE

259

260

c,d

-

262. R= TBDMS 263, R=

I

h

'

264, R= H c265,

266

R=Me

268a,b

qo]

267

269

(+)-252

SCHEME32. Reagents: a, 3-chloroperbenzoic acid; b, cyclopentadiene, ZnCI,; NaH; e , NaBH,; f, Sm(II)12;g, pyridinium c, silica gel; d, 2-(2-bromoethyl)-l,3-dioxolane, p-toluenesulfonate; h. 2-(trimethylsilyloxy)pent-I-ene, BF,-Et,O; i. conc. HCI; j , flash vacuum pyrolysis; k, H,, Pt on alumina: I , (EtO),CH, TsOH; m, LiAlH.,; n, 10% H2S04: 0, NaOH.

growth hormone in broiler chicks was also investigated (10). Biosynthetic studies by Harris et d.have been actively pursued (90). Recently, the asymmetric synthesis of (-)-slaframine has been achieved by three groups. Pearson et ul. reported the first synthesis of (-)-270 via reductive cyclization of an epoxy azide as a key step (Scheme 3 3 ) (91). N-Benzylation of L-glutamic acid, followed by protection, provided the N benzyloxycarbonyl derivative 271. Diborane reduction of both carboxyls afforded the diol 272, which was selectively silylated to produce 273, presumably because of the different steric and electronic environments of the two hydroxyl groups. Conversion of 273 to the azide 274 was accomplished with the Mitsunobu reaction. Deprotection of 274 and oxidation of the resultant hydroxyl at C-4 gave the azido aldehyde 275, which was converted to the Z-alkene 276 by a highly stereoselective Wittig olefination. Epoxidation of 276 with m-chloroperbenzoic acid (MCPBA) was nonselective, producing a 1 : 1 ratio of the diastereomeric epoxides 277 and 278. Tosylation of 277 afforded the azide 279, and selective reduction of 279 to an amine in the presence of the two benzyl protecting

3.

S I M P LE I N D O L I Z I D I N E ALKALOIDS

'

27 1

272, X=CHpOH, Y= OH

225

276

273, X=CH20TBDMS, Y= OH

d'e

c

274. X=CH,OTBDMS, Y= N3

275. X=CHO, N3

278 R= H i

283 R=Ts '

c

285 R,= En, R2= Cbz, R3= Ac 286 R,= H, R2= H, R= , A

SCHEME 33. Reagents: a, BH,-THF; b, TBDMSCI. pyridine. catalytic DMAP: c. Ph3P, DEAD. (PhO),P(O)N,; d , "BudNF; e . (COCI)?. DMSO, NEt3; f. Ph3P+ (CH2),0HBr-, KN(SiMe,),. TMSCI. 275; g. HCI; h, MCPBA; i. TsCI, pyridine. catalytic DMAP; j , 10% Pd/C ( 5 wt %), H?; k, K2C03;I, Ac,O, pyridine: m, 10% Pd/C (100 wt %, 10 mol 7% Pd). H,;n, Ac,O, pyridine.

groups was accomplished by hydrogenolysis. The resultant amine was not isolated, but was directly heated in the presence of a base. Intramolecular epoxide opening and subsequent alkylation of the nitrogen by the tosylate afforded the indolizidine 280. Acetylation of the secondary alcohol in 280

226

HIROKI TAKAHATA A N D TAKEFUMI MOMOSE

gave 281, which was deprotected to afford (-)-slaframine [( -)-2701. The signs of the specific rotations of both synthetic N-acetylslaframine 282 [a];' -11.2' (EtOH) and natural 282 [ a ] $ -15.9" (EtOH) are the same, indicating that the absolute stereochemistry of natural ( 4 - 2 7 0 is (lS,6S,8aS). A similar sequence afforded (-)-I ,8a-diepislaframine 286 in good yield from the epoxy tosylate 283 (Scheme 33). Cha and co-workers reported an enantioselective synthesis of ( 3 - 2 7 0 through an intramolecular azide [2 + 31 dipolar cycloaddition reaction (Scheme 34) (92). The chiral phosphonium salt 287 was prepared from the readily available aldehyde 288 (obtained from L-aspartic acid in eight steps) (93). Diisobutylaluminum hydride (DIBAL) reduction of the known and readily available lactone 290 (94) and subsequent Wittig reaction with the ylide derived from 287 afforded the adduct 291. Subsequent tosylation followed by DIBAL reduction yielded the alcohol 292. Selective N-tosylation of 292 followed by azidation with sodium azide via an intramolecular azide [2 + 31 cycloaddition produced the imine 293. Bicyclic ring closure was achieved uneventfully by selective 0-mesylation and subsequent NaBH, reduction to produce the protected slaframine 294. Finally, a series of straightforward deprotection steps and 0-acetylation afforded (-)slaframine [( -)-2701. The synthesis of (-)-270 was executed in 11 steps via radical cyclization from resolved (S)-3-hydroxy-4-pentenamide (295) by Knapp and Gibson

OA N A

a-c

kN, PMB

PMP

PMP

R

c288

g-i

- Hbo \

2pso\ e.f

OTs

290

291

OH

292

R= CHo

L289

R= CH2Br 287 R= CHZ'PPh3Bi

293

PMP= pmethoxyphenyl PM0= pmethoxybenzyl

294

(-)-270

SCHEME34. Reagents: a, NaBH,; b, TsCI, catalytic DMAP; c, LiBr, NaHCO,; d, PPh,; e, KN(SiMe,h; f, 290, DIBAL; g, N-tosyl-N-methylpyrlidine perchlorate; h, NaN,; i, toluene; j, MsCI, Et3N; k, K2C03;I, ceric ammonium nitrate; m, "Bu,NF; n, Na, NH,; 0,HCI, AcOH.

3.

SIMPLE INDOLIZIDINE ALKALOIDS

296

295

300a a-isomer R= THP

2917

302

?A'

227

299

303

300b p-isomer R= THP 301a a-isomer R= H 301b D-isomer R=H

WN3 - PN"' 0

AcO

ti

304

AcO

(-)-270

35. Reagents: a, TMSOTf, Et3N: b, 12; c, aq. Na2SOl: d. PhSeSePh, NaBH4; e, TBDMSCI, irnidazole: f, NaH, 298: g, "Bu4NF;h, AczO, pyridine: i, (Me$i),SiH, AlBN: j, AcOH; k, MsCI, 'PrzEtN; I, NaN3; rn, BH3-SMe2: n, TMDEA; 0,H2 5% Pd/C. SCHEME

(95) (Scheme 35). Stereoselective iodolactamization of 295 provided the

iodolactam 296 (96). Replacement of the iodo in 296 with the phenylseleno group and subsequent protection of the hydroxyl in 296 afforded 297. N Alkylation of 297 with 3-iodo-2-(2-tetrahydropyranyloxy)propene (298) and subsequent replacement of the silyl group and installation of the acetyl group gave the required substrate 299. Formation of an indolizinone from 299 by radical-initiated cyclization using tris(trimethylsi1yl)silane proceeded to yield two pairs of diastereomers 300a,b in the ratio 7 : I owing to the tetrahydropyranyl (THP) group. Hydrolysis of the 6a-hydroxyl diastereomer (300a) provided a single alcohol (301a). Conversion of 301a to its methanesulfonate 302 followed by SN2 displacement of the mesylate with azide gave the azidoindolizidinone 303. Reduction of the lactam with a borane-dimethyl sulfide complex and liberation of the free amine with tetramethylenediamine afforded the indolizidine 304. Finally, azidoslaframine (304) was converted to (-)-slaframine [( -)-2701 by catalytic hydrogenation.

228

H l R O K I T A K A H A T A A N D T A K E F U M I MOMOSE

H OH

305

H

OH

9H

306

OH

307

308

31 0

309

FIG.8.

V. Hydroxylated Indolizidines The alkaloids to be considered in this section are the l-hydroxyindolizidines 305 and 306, indolizidinediols 307 and 308, swainsonine (309), castanospermine (310), and their analogs (Fig. 8). A. ~-HYDROXYINDOLIZIDINES A N D I ,2-DIHYDROXYINDOLlZlDlNES

(IS,8aS)-l-Hydroxyindolizidine(305) and (IR,8aS)-l-hydroxyindolizidine (306) were recognized as key precursors for the biosynthesis of the toxic indolizidine alkaloids slaframine (270) and swainsonine (309), respectively, in the fungus Rhizoctonici lc~griminic~olrr. Investigation of their elegant biosynthesis has been performed by Harris et al. (90), and the current view of their biosynthesis in R . iegrrtninicola is summarized in Scheme 36. It was shown that the alkaloids are formed from L-lysine via L-pipecolic acid, 1 -oxoindolizidine 310, and 1 -hydroxyindolizidines 305 and 306. Harris and Harris prepared the four diastereomers of l-hydroxyindolizidine (305,306,312, and 313) in high optical purity (>%%I by NaBH, reduction of the ( + )- and (-)-3-bromocamphor-8-sulfonic acid (BCS) salts of I-oxoindolizidine [( ?)-311], followed by separation of the resulting diastereomeric alcohols with ion-exchange chromatography (Scheme 37) (97). Enantiocontrolled synthesis of 305 has been achieved through a condensation of (3R,2S)-hydroxyprolinal (314) with a three-carbon synthon by Sibi and Christensen (98). Baker's yeast reduction of the protected 3ketoproline ester 315 afforded 3-hydroxyproline ester 316. Interchange of the benzyloxycarbonyl with the tert-butoxycarbonyl protecting group, followed by protection of the hydroxyl group and conversion of the ester to the aldehyde using DIBAL reduction, provided the aldehyde 314. Wittig

3.

SIMPLE INDOLIZIDINE ALKALOIDS

L-lysine

-eH L-pipecolic acid

305

u

229

OH

306

310

H

307

OAC

270

309

SCHEME36. Biosynthesis of slaframine and swainsonine

reaction of 314 with the phosphorane 317 gave a mixture of cis- and trcinsolefins 318. Hydrogenation of 318 followed by mesylation furnished the mesylate 319, which was treated with 3M HCl to provide the desired (IS,8aS)-1-hydroxyindolizidine (305) (Scheme 3 8 ) . An asymmetric synthesis of 1-hydroxyindolizidines 305 and 306 was performed by a short reaction sequence involving intramolecular amidomercuration as shown in Scheme 39 (99). The kinetic resolution and asymmetric epoxidation of racemic N-benzyloxycarbonyl-3-hydroxy-4-

7 (f)-311

(+)-BCS salt

\

306

(-)-BCS salt

313

SCHEME37. Reagents: a, ( + ) - B C S ; b, (-)-BCS; c, NaBH,; d, Dowex 50.

230

HIROKI TAKAHATA A N D TAKEFUMI MOMOSE

,.,OH

-2-

,OTBDMS

b-e

q..cmE,

f Q'CHO

b

Ph3P'(CH*),OHCI-

Cbz

Cbz

Boc

31 5

31 6

31 4 317

p

s

g,h

(3 Boc

BOC

OH

OMS

31 8

i

-

305

31 9

SCHEME38. Reagents: a, baker's yeast; b, H,, Pd; c , BoczO; d, TBDMSCI, base; e , DIBAL; f, 317, LiHMDS; g, H2, Pd; h, MsCI, Et,N; i , 3 M HCI.

pentenylamine (320), capitalizing on the Katsuki-Sharpless reagent, provided three kinds of products, (S)-320, the epoxy alcohol 321, and the pyrrolidine 322 (100). Stereoselective intramolecular amidomercuration of (S)-320 followed by radical Michael addition to methyl acrylate afforded the cis-2,3-disubstituted pyrrolidine 323 without isolation of the trans isomer. Exposure of 323 to hydrogenation gave the indolizidinone 324, which on reduction with LiAlH, provided 305. Conversion of 324 to 306

c-c Y"

Cbz

(+)-320

a

/

+c

KY+--$ YH I

Cbz

(S)-320

Cbz

321

OH

Cbz

322

If ! T

OCOPh

p : ( y J 0

325

kj OH

306

SCHEME39. Reagents: a, D-(-)-diisopropyl tartrate, 'BuOOH, Ti(O'Pr),; b, Hg(O COCFh; c , CHz=CHCOOMe, NaBH(OMe),; d , H,, Pd(OH),; e, LiAIH,; f, Ph3P,diethy1 azodicarboxylate, PhCOOH.

,

3.

23 1

SIMPLE INDOLIZIDINE ALKALOIDS

was achieved by inversion of the hydroxyl group in 324 employing the Mitsunobu reaction followed by reduction. Indolizidinediol307 has been isolated from R. legurninicola and Astuagalus lentignosus (101). In addition, 307 has been demonstrated to be a biosynthetic precursor of swainsonine (309) as shown in Scheme 36 (90). The absolute configuration of (1S,2R,8aS) was confirmed by efficient reutilization of the compound in the further biosynthesis of swainsonine. Lentiginosine (308), isolated from A . lentignosus, inhibited the a-glycosidase amyloglucosidase (102). The structure was confirmed by spectroscopic data. On the basis of biosynthetic considerations, the absolute configuration is assumed to be (IS,2S,8aS). Heitz and Overman reported the enantiodivergent synthesis of both enantiomers of 1,2-indolizidinedioI (307) from the common intermediate 326, which was prepared in three steps from commercially available Disoascorbic acid (Scheme 40) (103).The enantiodifferentiation stems from cyclization at either the carbon site represented by the carbonyl or the one

X c-e

three steps

D-isoascorbic acid

326

327

328

330

329

(-)-307

0 x 0

TMS

332

(+)-307

331

SCHEME 40. Reagents: a, MsCI, Et3N;b, NaH; c, Lawesson reagent; d, Et,0BF4, 2.6di-rerr-butylpyridine; e , LiBEt3H; f , Cu(OS0,CF3),; g, H2. PdlC; h, 2M HCI; i, S03-py, DMSO; j, Ac,O, DMAP; k, BF,-OEt2; I, LiAIH4.

232

H l R O K l T A K A H A T A A N D T A K E F U M I MOMOSE

bearing the hydroxymethyl in the amide 326. The lactam 327, a potential precursor to an iminium ion intermediate, was obtained from 326 by conversion via mesylation followed by cyclization. The conversion of 327 to the 2-ethylthiopyrrolidine 328 was performed by a sequence involving thioamidation, methylation, and reduction. The iminium ion-vinylsilane cyclization was accomplished by treatment of 328 with copper triflate to provide the tetrahydroindolizidine 329, which was transformed by reduction followed by deprotection of the resulting acetonide 330 into the desired (-)-307, [a];' -39.4' (0.58, CHCI,). On the other hand, the conversion of 326 to the acetate 331 was carried out by oxidation of the hydroxymethyl followed by acetylation of the resulting hydroxy lactam. The acetoxy lactam 331, on treatment with boron trifluoride. underwent cyclization on the less hindered convex face to give 332, which was converted by the standard transformation to the unnatural ( 1-307 [a];4 +40.2" (0.88, CHCI,). An enantiocontrolled synthesis of the acetonide form (330) of (-1-307 was achieved starting with the pyrrolidino ester 323, described previously (Scheme 39), by Takahata et al. (104). Reduction of 323 followed by selective monotritylation provided the 3-hydroxypyrrolidine 333. Elimination of the hydroxyl group in 333 afforded the 3-pyrroline 334, which was dihydroxylated from the opposite side to the ring appendage at C-2 to furnish the diol335 as a single diastereomer. Subsequent acid hydrolysis. acetonide formation, mesylation, and cyclization afforded the desired 330 (Scheme 41).

+

,+OH Q..,,-Co0Me Cbz

323

335

ab

+OH

1Q.Cbz "

f l M O T r

333

336

Cbz

334

330

SCHEME 41. Reagents: a, DIBAL; b, 'HCI, Et,N; c , NaH, CS2, Mel; d , 170°C; e, catalytic OsO,, N-methylmorpholine oxide; f, HCI; g, (CH3)2C(OCH3)2. TsOH; h, MsCI. pyridine; i , H2, Pd(OH),; j. aq. K2C0,.

3.

233

SIMPLE I N D O L I Z I D I N E A L K A L O I D S

B . SWAINSONINE From the fungus Rhizoctonici legrrrninic~olciwas isolated the toxic indolizidine alkaloid swainsonine (309) (105). Swainsonine has also been shown to be present in locoweed (Astrrrgcrlus lentiginosus)(106) and Sirwinsoniu canescens (107), as well as in the fungus Metarrhizium unisopliue (108). The pronounced a-mannosidase inhibitory (109) and immunoregulative properties (110) of swainsonine have stimulated considerable chemical, biosynthetic, and pharmacological interest (111 1, and its total synthesis by several groups, including five enantioselective syntheses ( 112-1 161, has already been described ( I ) . Most of the syntheses are based on a route from carbohydrates. Hashimoto and co-workers carried out a short, enantiospecific synthesis of (-) swainsonine [(-)-3091 from D-mannose using. a5 a key step, the double cyclization of the epoxy amino ester337 (Scheme 42) ( 1 17).Conversion of the oxime 338, readily accessible from D-mannose, to the protected amine 339 was followed by selective hydrolysis to form a diol, which led to the epoxide 340. Oxidation of 340 to an aldehyde and Wittig reaction of the latter furnished the a$-unsaturated ester 341. The required 337 was obtained by reduction of 341 with sodium borohydride in the presence of trifluoroethanol as a proton source. Release of the amino group and subsequent reflux in ethanol resulted in double cyclization of 337 to the lactam 338, which was converted to (-)-309. Fleet and co-workers devised a new design in which the mannosederived azidoepoxide 343 played a pivotal role (Scheme 43) (118). The

D-mannose

NHCbz

338

341

340

339

337

NHCbz

338

(-)-309

SCHEME 42. Reagents: a , LiAIH,: b, CbzCl; c. MsCI, pyridine; d, TsOH; e, Amberlite IRA 400 resin; f, Collin’s reagent; g. Ph3P=CHC02Et; h, NaBH4. CFICH20H; i. H2. Pdi C; j , EtOH; k, LiAIH,; I, 6M HCI.

234

HIROKI TAKAHATA A N D TAKEFUMI MOMOSE

344

346

345

338

SCHEME43. Reagents: a, NaN,: b, camphorsulfonic acid: c, Ba(OH),; d, (CF3S02),0; e , LiCH2CO2But;f, H2, Pd/C: g, NaOMe: h, BH3-Me2S; i, CF3C02H-H20:j, ion-exchange chromatography.

dimesylate 344, readily available on a large scale from mannose, underwent selective displacement of the primary mesylate by azide anion to give the azido mesylate, which, on partial hydrolysis, afforded the diol 345. Base-mediated epoxidation of 345, followed by two-carbon extension at the terminal hydroxyl, provided the azido epoxide 343. Hydrogenation of 343 gave the pyrrolidino ester 346 by intramolecular S,2 attack of the resulting amine on the oxirane. Subsequent heating with sodium methoxide provided the same &lactam (338) as described above (Scheme 42). Preparation of the highly valuable intermediate 347 developed by Fleet and co-workers for the synthesis of (-)-swainsonine was achieved by a 15-step sequence starting from D-mannose (Scheme 44) (119). The application of selective tosylation, acetonization, and silylation to benzyl-a-Dmannoside (348) gave the tosylate 349. Nucleophilic displacement at the tosyloxymethyl in 349 by reaction with allylmagnesium chloride followed by desilylation and Swern oxidation, provided the olefinic ketone 350. Oxidation of the olefinic terminus of 350, followed by esterification of the resulting carboxyl group, furnished the ester 351. NaBH,-mediated reduction of 351 by attack of hydride from the a side and subsequent trifluoromethanesulfonation gave the triflate 352, which, on treatment with sodium azide, was transformed into the azide 353. Hydrogenation of 353 followed by lactamization gave the bicyclic lactam 354, which was converted with LiAlH, to the pivotal intermediate 347 for the synthesis of (-)-swainsonine [(-)-3091. A practical, enantioselective synthesis of (-1-309 was achieved in seven steps from 2,3-O-isopropylidene-~-erythrose (355) (120). The key step involves the construction of the bicyclic imine 358 via intramolecular 1,3dipolar cycloaddition in the olefinic azide 356 to give the triazoline 357 (Scheme 45). The requisite azide 356 was prepared in three steps (i.e., Wittig reaction of 355, tosylation, and azidation). Heating of 357 led to

3.

SIMPLE INDOLIZIDINE ALKALOIDS

352

351

354

235

353

ref. 112 (-)-309 347

SCHEME 44. Reagents: a, TsCI, pyridine; b, CH,C(OCH3)2, catalytic camphorsulfonic acid (CSA); c, TMSCI, Et3N: d, allylmagnesium bromide; e, "Bu4NF; f, (COC1)2,DMSO, &N; g, NaI04, RuOZ-H~O;h, CH2N2; i, NaBH4: j, (CF3S02)20,pyridine; k, NaN,; I, H2, Pd; rn, toluene; n, LIAIH,.

361

362

SCHEME 45. Reagents: a, Br-Ph3P+(CH2),C02Et,KN(TMS),; b, TsCI, Et3N; c, NaN,; d, K2CO3; e, toluene: f , BH3; g, H202-NaOH; h, 6N HCI.

236

HlROKl TAKAHATA A N D TAKEFUMI MOMOSE

I ,3-dipolar cycloaddition to yield the cyclic imine 358 via the triazoline intermediate 357. Imino ester 358 was then hydrolyzed to afford the acid 359, which was heated to provide the enamide 361 via the spirocyclic lactone 360. A highly diastereoselective hydroboration of 361, accompanied by reduction of the lactam carbonyl, produced the swainsonine acetonide 362, which, on hydrolysis, was converted to (- 1-309. A similar route to (-)-309 was achieved by Pearson and Lin (121). Wittig reaction of 355 followed by Mitsunobu reaction of the resulting alcohol afforded the olefinic azide 363. Intramolecular 1,3-dipolar cycloaddition of 363 resulted in the formation of the bicyclic iminium ion 364, which, on treatment with ?err-butylamine, was converted to the enamine 365. Enamine 365, without isolation, was hydroborated to afford swainsonine acetonide 362, along with a small amount of the indolizidine diol 366 (Scheme 46). A novel approach to swainsonine (309) from the hydroxy lactam 367 via intramolecular cyclization to form an enantiomerically pure cyclic acyliminium ion intermediate was achieved by Miller and Chamberlin (122). The requisite lactam 367 was prepared in four steps from D-(-)lyxose. Mesylation of 367 was followed by spontaneous formation of the acyliminium ion intermediate and subsequent cyclization to yield the indolizinone 368 as a single diastereomer. The ketone dithioacetal368 was converted to the a-bromo ester by treatment with NBS, and subsequent treatment with 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU) gave the unsaturated ester 369. Treatment of 369 with Meerwein’s reagent, followed by reduction with NaBH,CN, did not terminate at the vinylogous ester 370, but instead resulted directly in the formation of a single, fully saturated indolizidine (371). The C-8 ketone 372 was prepared by a-hydroxylation

363

362

365 S C H E M E 46.

364

366

Reagents: a, Br-Ph3Pt (CH2)dCI. KN(TMS)?;b. (PhO)?P(O)N1,Ph3P. DEAD;

c, PhH; d, ‘BuNH2; e, BH3-THF; f. NaOAc. H 2 0 2 ;g, 6N HCI: h, IRA-400 ion-exchange

chromatography .

3.

237

SIMPLE INDOLIZIDINE ALKALOIDS

of ester 371 followed by LiAIH, reduction to the diol and oxidative cleavage with NaIO,. Reduction of 372 with Na/NH, gave the desired equatorial alcohol 373, which was transformed by removal of the cyclohexylidene ketal moiety into (-)-309 (Scheme 47). A noncarbohydrate route to (-)-309 by Ikota and Hanaki (124) was achieved by way of face-selective osmylation of the optically active butenolide 374 ( 1 2 3 , available from (R)-glutamic acid (Scheme 48). Cisdihydroxylation of 374 followed by protection of the diol gave the isopropylidene derivative 375, which was converted by successive reduction, mesylation, and azidation to the azido mesylate 376. Pyrrolidine construction to 377 was effected by reduction accompanied by concomitant cyclization via intramolecular S,2 displacement. Introduction of both a threecarbon unit and the fourth stereocenter (C-8) to give the target molecule was achieved by oxidation of the dibenzylated pyrrolidine 378 followed by diastereoselective allylation. Swern oxidation of 378, followed by allylation of the resulting aldehyde with Grignard reagent, furnished the desired diastereomer 379 and its epimer 380 in a ratio of 3 : I . Successive hydroboration-oxidation of 379, mesylation, and debenzylation afforded the desired ( 4 - 3 0 9 . The same operation for 380 provided 8-episwainsonine

QM -1ao g-] d,e

f-h

0

369

-

0

371

370

i

(-)-309

373 372 SCHEME 47. Reagents: a, MsCI, Et,N; b, NBS; c, DBU: d , Et,OtBF,-: e. NCHBH,; f, LDA, 0,; g, LiAIH,; h, NaIO,; i, Na/NH3:j , 6 M HCI.

238

HlROKl TAKAHATA A N D T A K E F U M I MOMOSE

-

a,b

Q..,,,OTr

0

374

0x0

8 0 x 0

ref. 123

(RJ-glutamicacid

-

..,,,OTr

0

0x0 ..,,,OTr

375

BnO

OBn

BnO

OBn

..,,,OTr

Bn

H

378

377

V

379 ref. 125

V : / O H

I

4

0-1

(-)-309

0

D-ribonolactone

SCHEME 48. Reagents: a, catalytic OsO,, 4-methylmorpholine N-oxide; b, 2,2-dimethoxypropane, TsOH; c, LiAIH,; d, MsCI, pyridine; e , NaN,; f, H2, Pd black; g, BnBr, K2C03; h, conc. HCI; i, MOMCI, N,N-diethylaniline; j , 10% HCI; k , BnBr, NaH; 1, 10% HCI; m, (COCI)2, DMSO, Et3N; n, allylmagnesium bromide; 0,BH3; p, H 2 0 2 ,NaOH; q, MsCI, Et3N; r, H2. Pd/C, HCI.

(381). The key intermediate 378 was also obtained from D-ribonolactone by the same group (125). Hart and co-workers developed a synthesis of (-)-309 from the tartarimide 382 via radical-initiated cyclization as a crucial step (Scheme 49). (126). Mitsunobu reaction of 382, accessible from D-tartaric acid, with acetylenic alcohol provided the tartarimide 383. Reduction of 383 followed by sulfenylation of the resulting carbinol lactam gave the radical precursor 384, which on cyclization afforded a mixture of geometric isomers of indolizidines (385). Ozonolysis of 385 followed by stereoselective reduction of the resulting ketone furnished the C-8 hydroxyl (386). Inversion of the stereochemistry of C-1 of 386 was achieved by displacement of the triflate by the acetate ion to give 387. Removal of the lactam carbonyl was carried out by desulfurization via the thiolactam. Subsequent deacylation provided (-)-309. Finally, a number of syntheses of stereoisomers of swainsonine (309) such as 1-epi- (388) (1277, 8-epi- (381) (123,128),8a-epi- (389) (128,129), 8,ga-diepi- (390) (128,130), 2,ga-diepi- (391) (130), 1 $-diepi- (392) (1277,

3.

SIMPLE INDOLIZIDINE ALKALOIDS

385

386

239

387

SCHEME 49. Reagents: a, Ph3P, DEAD, PhCCCH2CH2CH20H;b, NaBH,; c, "Bu3P, PhSSPh; d, "BulSnH; e, AIBN; f, 03;g, Me& h, NaBH4; i, Me3CCOCI (PivCI), DMAP, pyridine; j, NH3, MeOH; k, Tf20; I, KOAc, 18-crown-6; m, Ac,O, Et,N, DMAP; n, ( p MeOC6H4PS2)2; 0,Raney Ni; p, MeNH2.

and 2,8-diepiswainsonine (393) (115), as shown in Fig. 9, have been developed.

C . CASTANOSPERMINE Castanospermine (310), isolated from seeds of the Australian legume Castanospermum australe (131) and the dried pods of Alexa leiopetala (132), is a potent, competitive, and reversible inhibitor of several glucosi-

381

390

388

39 1

389

392

393

FIG.9. Stereoisomers of swainsonine.

240

H l R O K l T A K A H A T A A N D T A K E F U M I MOMOSE

H

HoO

HO"'

OH a

HO&

H2

H

Hoo

d

HO

HO

31 0

OH

394

395

FIG. 10. Naturally occurring stereoisomer5 of castanospermine.

dases (133). It has potential for treatment of diabetes (1341, obesity (13% cancer (136),and viral infections (1377, including human immunodeficiency virus-1 (HIV-1) (138). Stereoisomers of castanospermine, 6-epicastanospermine (394) (139) and 6,7-diepicastanospermine(395) (140),also have been isolated from Castunospermum uustrule (Fig. lo), and both inhibited amyloglucosidase. It is likely that other stereoisomers and analogs of these compounds will also inhibit glucosidase and potentially result in providing materials possessing interesting and useful biological properties. In the previous review ( I ) , only one synthesis of 310 was cited (141). To date, a number of syntheses of 310 have been achieved. Recently, an excellent review by Burgess and Henderson of synthetic approaches to stereoisomers and analogs of castanospermine has appeared (142).Most syntheses of 310 inevitably utilize carbohydrates as starting materials because of the sugar-like structure. This section reviews syntheses of the 1,6,7,8-tetrahydroxyindolizidines310, 394, and 395 isolated from natural sources (Fig. 10). Other analogs are not discussed. The absolute stereochemistry was confirmed during the first total synthesis of 310 by Bernotas and Ganem to be (IS,6S,7R,gR,SaR) (141). The protected D-glucopyranose 396 was transformed via amide 397 into the epoxide 398. Cleavage of the amide group in 398 with NaBH, followed by spontaneous cyclization gave the desired piperidine 399 and the azepane 400. Swern oxidation of 399 provided the aldehyde 401, which was immediately treated with tert-butyl lithioacetate to afford I : 1 mixture of the diastereomers 402 and 403. The less polar epimer 402 was transformed by hydrogenolysis and acid treatment into the lactam 404. Reduction of which with DIBAL gave (+)-castanospermine [( +)3101. The same reaction sequence transformed the more polar epimer 403 into 1-epicastanospermine (405) (Scheme 50). Hashimoto and co-workers reported a synthesis of ( + )-310 involving double cyclization of the epoxy amino ester 406, utilizing a strategy similar to that adopted in the indolizidine ring formation of swainsonine (Scheme 42) (143). The D-mannose-derived diol 407 was transformed in four steps into the aldehyde 408, which was epimerized by treatment with K,CO,

3.

397

396

399

+

24 1

SIMPLE INDOLIZIDINE ALKALOIDS

401

390

402

404

+

400

405 403

SCHEME 50. Reagents: a, BnNH,: b, LiAIH,; c , trifluoroacetylation; d. TBDMSCI, imidazole; e . mesylation: f. "Bu4NF;g, MeONa: h. NaBH4: i , (COCI)?. DMSO, Et,N: j, terfbutyl lithioacetate: k, hydrogenolysis; I , trifluoroacetic acid; rn,DIBAL.

in MeOH to give aldehyde 409, which possesses the requisite stereochemistry. Aldehyde 409 was converted in seven steps to the epoxy alcohol 410, which was then oxidized to the aldehyde 411. Without isolation. aldol reaction of 411 with rm-butyl lithioacetate gave a mixture of diastereomers 412 in a ratio of 3 : 2. The transformation of 412 via protection of the hydroxyl group and subsequent hydrogenolysis to the pivotal amine 406 was followed by the double cyclization reaction to provide a mixture of indolizidinones 413 and 414. After separation, reduction of 413 with borane followed by treatment with acid afforded ( + )-310. Similarly, epimer 414 was converted to 405 (Scheme 51). Ganem and co-workers devised a stereoselective synthesis of ( )-310 via Sakurai allylation with highly selective chelation control of the Dglucopyranose-derived aldehyde 401 (Scheme 52) (144).Sakurai condensation of the aldehyde 401, prepared according to the method described earlier (see Scheme 50), with allyltrimethylsilane in the presence of TiCI, produced only diastereomer 415 as a result of the excellent stereocontrol derived from selective chelate formation between TiCl, and the a-amino carbonyl in 401. Ozonolysis of 415, followed by reduction, gave the diol 416. Selective monomesylation of 416 followed by hydrogenolysis afforded (+)-310. Good stereocontrol also operated in the Sakurai allylation of the

+

242

HIROKI TAKAHATA A N D TAKEFUMI MOMOSE

407

408

409

41 2

41 1

410

KI*

TBDMSO

-

OH

q,r

,,.OH Z

H

OH

n

414

405

SCHEME 51. Reagents: a, BzCI, pyridine; b, TBDMSCI, imidazole; c, 1 M NaOH; d, DMSO, DCC, TFA, pyridine; e, K2C03;f, H2NOH-HCI, NaHCO,; g, LiAIH4; h, CbzCI; i, TsOH;J, "Bu4NF; k, MsCI, pyridine; I, MeONa; m, Cr03-pyridine;n, tert-butyl lithoacetate; 0 , H,, 10% PdIC; p, methoxyethanol; q; BH3-THF; r, 6 M HCl.

D-mannose-derived aldehyde 417. The product 418 was then converted to 6-epicastanospermine (394) by the same manipulation. An efficient synthesis of ( + )-310 was achieved starting from glucuronolactone (419) by Anzeveno et al. (145) (Scheme 53). The starting material

41 7

418

394

SCHEME 52. Reagents: a, allytrimethylsilane, TiCI,; b, 0,; c, NaBH,; d, MsCI, Et3N; e, H,, PdIC.

3.

SIMPLE INDOLIZIDINE ALKALOIDS

243

421

+

423

SCHEME53. Reagents: a, LDA, EtOAc; b, H,, PtO2; c, HC02H;d, Dowex 1-X2 (OH-) resin; e , LiAIH,; f, CF3C02H;g, H2, 5% Pt/C.

419, containing four of the five chiral centers in (+)-310, was converted in four steps to the hemiketal 420. Catalytic hydrogenation of 420 over PtO, via the favored conformer of the keto ester 421 gave a 7 : 2 mixture of 422 and 423. The desired alcohol 422 was transformed by successive deprotection, cyclization, and reduction into the pyrrolidine 424. Deprotection of 424 followed by hydrogenation gave ( + )-310. Carrying the epimer 423 through the same series of reactions gave 405. Miller and Chamberlin (122) reported a synthesis of ( +)-310 based on the intramolecular cyclization of an enantiomerically pure polyhydroxylated acyliminium ion adopted in the synthesis of swainsonine (309), as described earlier (Scheme 47). Preparation of the requisite hydroxy lactam 425 began with the known D-gluconolactone 426, which was treated with I ,3-dithiane, followed by lead tetraacetate oxidation, to afford the lactam 425. Mesylation of 425 was followed by spontaneous formation of the acyliminium ion intermediate and cyclization to give the epimers 427 and 428 in a ratio of 1 : 1. After separation, oxidation of 427 with singlet oxygen produced an unstable ketone, which was reduced selectively by L-Selectride to the indolizidinone 429. Reduction of 429 followed by hydrogenolysis gave (+)-310. The other epimer 428 was also transformed via indolizidinone 430 into 1 $a-diepicastanospermine 431 (Scheme 54). Gerspacher and Rapoport (146) devised a methodology to prepare ( + ) -310 and 6-epicastanospermine (394) based on stereoselective reduction

244

HIROKI T A K A H A T A A N D T A K E F U M I M O M O S E

427

429

+ q ~

~

6

d,e '

Brio$ fa

BnO"'

BnO'"

0

428 C.

0

0

430

431

SCHEME 54. Reagents: a, 2-(3-aminopropylidene)-l.3-dithiane;b, Pb(OAc),, AcOH; MsCI, Et,N; d, '02; e , L-Selectride; f, BH,-DMS: g, H2. 10% PdlC. HCI.

of cyclic ketone 432 (Scheme 55). The manno azide 433, obtained from D-gluco-&lactone, has four stereocenters arranged in the same manner as required for C-6, C-7, C-8, and C-8a in 394. The azide 433 was transformed in four steps into a-amino aldehyde 434, protected by a phenylfluorenyl (Pf)group, which allowed the introduction of a C? unit by an organometallic reagent. The product 435 was then cyclized in three steps to the fivemembered ring ketone 432. Reduction of 432 with NaBH, gave the alcohol 436 as a single epimer. Selective tosylation of 436 followed by removal of the Pf group, effected cyclization to give the indolizidine 437, which was then deprotected to provide the tetraol394. Synthesis of ( + b310 was accomplished by inversion of the C-6 hydroxyl in the alcohol 432. Selective acylation of 432 followed by formation of the triflate gave the pyrrolidinone 438, which was treated with acetate anion to give the inverted acetate 439. Reduction of 439 then gave a single alcohol (440).After deacetylation of 440, the resulting trio1 was converted to ( + )-310 by a series of reactions analogous to the synthesis of 394. The first synthesis of the unnatural enantiomer of castanospermine [(-I -3101 was achieved by two successive SN2-typecyclization reactions from D-xylose derivative 441 (Scheme 56) (147). Grignard reaction of the known pentose ether 441 with chelation control, followed by protection, gave

3. SIMPLE INDOLlZlDlNE ALKALOIDS

433

4

0

0

OH

434

r f n

436

432

435

j,k

437

439

I,m

-

HO

245

432

Pf

554

430

440

SCHEME 55. Reagents: a, H,, PdIC; b, PfBr, Et,N, Pb(N03),; c, DIBAL; d. NCS, Me2S; e, vinylmagnesium bromide; f, HBr; g, NaHCO?, Na2C03;h, NaBH4;i, N-methyltosylimidazolium trillate, N-methylimdazole; j , CF,COOH; k, Dowex 50W; I, Ac@, pyridine: m, (CF,S02)2,pyridine: n, Bu4NOAc; 0,AczO, pyridine, DMAP; p. K?COi.

the di-MOM derivatives 442 and 443 in the ratio 87 : 13. After separation, the major isomer 442 was subjected to ozonolysis to provide the aldehyde 444, which was followed by Hiyama-Nozaki allylation (148) under Felkin-Anh control to furnish a diastereomeric mixture of 445 and 446 in the ratio 85 : 15. The major product 445 was transformed into the mesylate 447 in seven steps. Hydrazinolysis of 447 induced cyclization to the pyrrolidine 448. After deprotection of 448, the resulting pyrroiidine alcohol was cyclized by means of a tetravalent phosphonium species in the form of the Appel reagent (Ph,P, CCI,, Et,N) (149) to afford 449, which was converted by debenzylation to (- )-castanospermine [( - 1-3101. Fleet e t a / . devised a total synthesis of 6-epicastanospermine 394 starting from L-gulonolactone (150). Three syntheses of castanospermine 310 have been performed by use of noncarbohydrate starting materials. Vogel and co-workers (151) devel-

246

HIROKI TAKAHATA A N D TAKEFUMI MOMOSE

OBn OBn

441

d

442

OBn OBn OH

OBn OBn OH

BnO

BnO

OMOM

445

k

443

444

OBn OBn OBn

BnO

OMOM

447

446

OBn OBn

0on H

'BuPhpSiOCH, ti

440

OMS

BnO

gBn

n

HO

449

(-)-310

SCHEME 56. Reagents: a, vinylmagnesium bromide; b, MOMCI, EtN('Pr):; c , 0, PPH3; d, ally1 bromide, CrC13, LiAIH,; e , NaH, BnBr; f , LiAIH,; g. phthalimide, PPh3, DEAD; h. HCI; i, MsCI, pyridine; j, 'BuPhzSiC1, imidazole; k, NzH,-H20; I , "Bu,NF; m, PPh,, CCI4, Et,N; n, H2, 10% pd/C, HCI.

oped a highly stereoselective synthesis of (+)-310 starting with (-) -( IS,4S)-7-oxabicyclo[2.2. IIhept-5-en-2-one [( -)-4501, "a naked sugar" (Scheme 57). Bromination of the benzyl acetal451 obtained from (-)-450 occurred on the less-hindered, convex face, and this was followed by stereoselective I ,3-migration of the endo-benzyloxy group of the acetal to afford the bromo ketone 452. Baeyer-Villiger oxidation of 452 followed by ring cleavage provided a 4 : 1 mixture of the methyl furanosides 453 and 454. The minor furanoside 454 was re-equilibrated to give 453. Furanoside 453 was transformed in three steps into the fused pyrrolidine 455 by an intramolecular S,2 aminocyclization, and it was then converted to the phosphonoacetamide 456 in three steps including the Arbuzov reaction. Intramolecular Horner-Emmons condensation of 456 followed by acetylation gave the indolizinone 457. Conversion of 457 to the epoxide 458 was followed by regioselective opening of the epoxide with H,O and acetylation to give the triacetate 459. Reduction of 459 followed by debenzylation provided (+)-310 (Scheme 57). A chemoenzymaatic avenue to (+)-310 utilizing the proline ester 460 as a chiral building block was developed by Sih and co-workers (152) (Scheme 58). The requisite chiron 460 was prepared by two procedures. First, an enzyme-catalyzed reduction of p-ketoester 461 gave 460 in over 99% enantiomeric excess. Alternatively, 460 was obtained via an enantioselective hydrolysis of the acetate (+)-462 using lipases [Pseudomonas

0

(+)-310

459

SCHEME 57. Reagents: a. BnOTMS, TMSOTf: b, Br,: c , NaHCO!: d. MCPBA, NaHCO,: e , SOCI?, MeOH; f , DIBAL; g. MsCI, pyridine; h, NH,; i , CICHzCOCl, pyridine; j, AczO, H2SOd: k , P(OEt),: I , K2C03: m, Ac?O, DMAP; n, Brz. AgOAc, AcOH. AQO: 0, 2-(ferf-butylimino)-2-[(diethylamino)imino]I ,3-dimethylperhydro- I .3.2-diazaphosphonne, polystyrene; p. 2-(t~r~-butylimino)-2-[(diethylamino)imino]-l,3-dimethylperhydro1,3,2-diazaphosphorine, polystyrene, H 2 0 ; q. A c 2 0 , DMAP: r. BH,-DMS: s, H2. PdlC.

~ . .

460

(k)- 461

460

-

0

c-e

OTBDMS

(+)-462

(2R,35)-462

(2S,3R)-460

-

h

464

463

465

TMSO

+

+

HO

467

469

468

Ik

Ik

(+)-310 HO

395

SCHEME 58. Reagents: a, Dipoduscus sp., Vogel's medium; b, Cundidu cylindrucea or Pseudomonus sp. (AK); c , TBDMSCI, imidazole: d, CF3C02H;e, methyl acrylate, Et,N; f, Na, TMSCI: g, DBU; h, TMSCI, LiN(TMS),; i, BH3-DMS: j , Me,NO; k , "BudNF.

248

HIROKI TAKAHATA A N D TAKEFUMI MOMOSE

sp. (AK) and Candida cylindrucea]. Conversion of 460 to the diester 463 in three steps included Michael addition. Intramolecular acyloin condensation of 463,followed by treatment with DBU, gave a mixture of indolizidines 464 and 465. Either 464 or 465 was converted to the enol silyl ether 466,which, on hydroboration/oxidation, gave a mixture of three products, 467,468,and 469).Desilylation of 468 and 469 afforded ( + 1-310and 6,7diepicastanospermine (393,respectively. A stereoselective synthesis of ( )-310 via a noncarbohydrate-based approach utilizing the chiral allylic alcohol 470, available from L-tartaric acid, was achieved by h a and Kibayashi (153) (Scheme 59). KatsukiSharpless oxidation of 470, followed by regiospecific cleavage of the resultingoxirane with Et,AINBn,, gave the 1,3-glycol471. The amino alcohol 471 was transformed in four steps into the aldehyde 472,and aldol condensation of 472 with ethyl lithioacetate provided an inseparable 89 : 1 I mixture of the p-and a-hydroxy esters 473 and 474 under p-chelation control. The diastereomeric mixture of 473 and 474 was converted to a separable

+

-

L-tartaric cid

TBDMSO

TBDMSO

470

471

,

/

-

T B D M S O ~ ~ C H O

9

Y-:

+

TBDMSO&-co2E:

MOM

472

-

Y

TBDMSO&C02Et

e.h

OTBDMS MOM

+

OTBDMS

TBDMSO

-

6H

474

473

TBDMSO

0.

j.k

MOM

475

477

470

SCHEME 59. Reagents: a, 'BuOOH. Ti('OPr)(4), diethyl L-tartrate; b, Et,AINBn,; CH3COC1, Et3N; d, MOMCI, Pr2NEt; e , LiAIH,; f, DMSO, (COCI),, Et3N; g, AcOEt. LiN(TMS),; h, TBDMSCI, imidazole; i, AcOH, Ph3P. DEAD; j, "Bu4NF;k. TsCI, pyridine; 1, H2, PD(OH),; m, Et3N; n, HCI. C,

3.

nos

SIMPLE I NDOLI ZI DI NE ALKALOIDS

249

no

31 0

ent-310 Ho,.& Ho

HO'"

OH

HO"'

405

H

Hc

O OH

~

HofY5

HO"'

HO

ent-394

395

394

ent-479

479

H O , , . g HO"'

431

480

481

482

no 483

FIG. I I . 1.6.7.8-Tetrahydroxyindolizidines synthesized to date,

mixture of the p-alcohol 475 and the a-alcohol 476. After conversion of the 0-alcohol 475 to 476 by Mitsunobu inversion, the a-alcohol 476 was transformed in two steps into the tosylate 477. Hydrogenolysis of 477 gave the protected indolizidine 478, which was deprotected by treatment with HCI-MeOH to provide ( + )-310. Three isolated products, 310, 394, and 395, plus a further nonisolated ten compounds, namely, ent-310 (147), I-epi-405 (14/,143,145,/47,~53), L-6-epi-ent-394( I N ) , L-1 ,6-diepi-ent-479, (/50), 1,6-diepi-479(150), 1 $adiepi-431 (122),8-epi-480 (144) 1,6,8-triepi-481(154),1,7,8-triepi-482(/54), 483 (/54),of the possible 32 stereoisoand 1,6,7,8-tetraepicastanospermine mers of 1,6,7,8-tetrahydroxyindoIizidine have been obtained to date (Fig. 1 I).

250

HIROKI TAKAHATA A N D TAKEFUMI MOMOSE

VI. Summary Investigation of synthetic routes to the simple indolizidine alkaloids has been active. With increased interest in the pharmacological properties of indolizidines, it is expected that further methodologies will be developed.

REFERENCES

1. A. S. Howards and J. P. Michael, in “The Alkaloids” (A. Brossi. ed.), Vol. 28, p.

183. Academic Press, New York, 1986. 2. S. Rajeswari, S. Chandrasekharan, and T. R. Govindachari. Heterocycles 25, 659 (1987). 3. Y. Nishimura, in “Studies in Natural Product Chemistry” (Atta-ur-Rahman, ed.). Vol. 1, p. 227. Elsevier, Amsterdam, 1988. 4. M. F. Grundon, N u t . Prod. Rep. 2, 235 (1985). 5. M. F. Grundon, Nut. Prod. R e p . 4, 415 (1987). 6. M. F. Grundon. Nut. Prod. Rep. 6 , 523 (1989). 7. J. P. Michael, Nut. Prod. Rep. 7, 485 (1990). 8. J. P. Michael, Nut. Prod. Rep. 8, 553 (1991). 9. J. W. Daly and T. F. Spande, in “Alkaloids: Chemical and Biological Perspectives” (S. W. Pelletier, ed.), Vol. 4. p. 1. Wiley. New York, 1986. 10. A. D. Elbein and R . J. Molyneux, in “Alkaloids: Chemical and Biological Perspectives” (S. W. Pelletier, ed.), Vol. 5, p. 1. Wiley, New York, 1987. 11. A. Numataand T. Ibuka, in “The Alkaloids” (A. Brossi, ed.). Vol. 31. p. 193. Academic Press, New York, 1987. 12. F. J. Ritter, L. E. M. Rotgans, E. Talman. P. E. J. Verwiel. and F. Stein, Experientia 29, 530 (1973). 13. F. J. Ritter, I . E. M. Bruggeman-Rotgans. E. Verkuil. and C. J . Persoons. in “Proceedings of the Symposium on Pheromones and Defensive Secretions in Social Insects” (C. Noirot, P. E. Howse, and G. Le Masne, eds.). p. 99. University of Dijon Press, Dijon, France, 1975. 14. T. H. Jones, R. J. Hight. M. S. Blum. and H. M. Fales, J . Chem. Ecol. 10, 1233 (1984). 15. T. H. Jones, A. Laddago, A. W. Don, and M. S . Blum, J . Nut. Prod. 53, 375 (1990). 16. J. Royer and H.-P. Husson, J . Org. Chem. 50, 670 (1985). 17. N. Yamazaki and C. Kibayashi, Tetralzedron Lett. 29, 5767 (1988). 18. H. Iida, N. Yamazaki, and C. Kibayashi, J . Org. Chem. 51, 3769 (1986). 19. T. Nakata, T . Tanaka, and T . Oishi, Tetrahedron Lett. 24, 2653 (1983). 20. M. Ito and C. Kibayashi, Tetrahedron 47, 9329 (1991). 21. K. N. Houk, S . R. Moses, Y.-D. Wu, N. G. Rondan, V. Jager, R. Schohe, and F. R. Fronczek, J . A m . Chem. Soc. 106, 3880 (1984); K. N. Houk, H.-Y. Duh. Y.-D. Wu, and S . R. Moses. J . Am. Chem. Soc. 108, 2754 (1986). 22. T. Momose, N. Toyooka, S. Seki, and Y. Hirai, Chem. Pharm. Bull. 38, 2072 (1990). 23. R. Shirai, M. Tanaka, and K. Koga, J . A m . Chem. Soc. 108, 543 (1986). 24. C. W. Jefford, Q. Tang, and A. Zaslona, J . A m . Chem. Soc. 113, 3513 (1991).

3.

SIMPLE INDOLIZIDINE ALKALOIDS

25 1

25. C. Saliou, A. Fleurant, J. P. Celerier, and G. Lhommet, Tetruhedron Lett. 32, 3365 (1991). 26. A. L. Smith, S. F. Williams, and A . B. Holmes, J. Am. Chem. Soc. 110, 8696 (1988). 27. H . Takahata, H. Bandoh, and T. Momose, 34th Symposium on The Chemistry of Natural Products, p. 663. Tokyo, 1992. 28. K. E. Harding and T. H. Marman, J. Org. Chem. 49, 2838 (1984). 29. H. Takahata, H. Bandoh, M. Hanayama, and T. Momose. Tetruhedron: Asymtnetrv 3, 607 (1992). 30. C. L. Hill and G. M. Whitesides, J . A m . Chetn. Soc. 96, 870 (1974). 31. R. V. Stevens and A. W. M. Lee, J. Chem. Soc., Chrm. Cotnmim., 103 (1982). 32. J . W. Daly, C. W. Myers, and N. Whittaker, Toxicon 25, 1023 (1987). 33. J . W. Daly, G. B. Brown, M. Mensah-Dwuman. and C. W. Myers, Toxicon 16, 163 (1978); T. F. Spande. J. W. Daly. D. J . Hart, Y.-M. Tsai. and T. L. Macdonard, Experientia 37, 163 (1981). 34. J. W. Daly, T. F. Spande, N. Whittaker, R. J. Highet, D. Feigl, N. Nishimori. T. Tokuyama, and C. W. Myers, J. N a f . Prod. 49, 265 (1986). 35. T. Tokuyama, N . Nishimori, I. L. Karle, M. W. Edwards, and J . W. Daly. Tetrahedron 42, 3453 (1986). 36. J. Royer and H.-P. Husson, Tetrahedron Lett. 26, 1515 (1985). 37. N. Machinaga and C. Kibayashi, J . Org. Chetn. 57, 5178 (1992). 38. N. Yamazaki and C. Kibayashi, J. Am. Chem. Soc. 111, 1396 (1989). 39. T. Tokuyama, T. Tsujita, A. Shimada, H. M. Garrdffo, T. F. Spande, and J. W. Daly, Tetrahedron 47, 5401 (1991). 40. T. Tokuyama. N . Nishimori, A. Shimada. M. W. Edwards, and J. W. Daly, Tetrahedron 43, 643 (1987). 41. M. W. Edwards, J. W. Daly, and C. W. Myers, J. Nor. Prod. 51, 1188 (1988). 42. A. L. Smith, S. F. Williams, A . B. Holmes, L. R. Hughes. Z. Lidert. and C . Swithenbank, J. A m . Chem. SOC.110,8696 (1988): A. H. Holmes, A. L. Smith. S. F. Williams. L. R. Hughes, Z. Lidert, and C. Swithenbank, J . Org. Chetn. 56, 1393 (1991). 43. Y. Shishido and C. Kibayashi. J. Org. Chem. 57, 2876 (1992). 44. J. W. Daly, Y. Nishizawa, M. W. Edwards, and J. A. Waters. Nrrrroc~hem.Res. 16, 489 (1991). 45. J. W. Daly, Y. Nishizawa, W. L. Padgett, T. Tokuyama. A . L. Smith, A. B. Holmes. C. Kibayashi, and R. Aronstam, Nertrochetn. Res. 16, 1213 (1991). 46. L. E. Overman and K. L. Bell, J. Am. Chem. Soc. 103, 1851 (1981). 47. L. E. Overman and S. W. Goldstein, J . A m . Chem. Soc. 106, 5360 (1984); S. W. Goldstein, L. E. Overman, and M. H. Rabinowitz, J . Org. Chetn. 57, I179 (1992). 48. L. E. Overman, L. A. Robinson, and J. Zablocki, J. Am. Chem. Soc. 114, 368 (1992). 49. H. M. Garraffo, M. W. Edwards, T. F. Spande, J . W. Daly. L. E. Overman, C. Severini, and V. Erspamer, Tetruhedron 44,6795 (1988). 50. J . W. Daly, H. M. Garrafo, L. K. Pannell, T. F. Spande, C. Severini. and V . Erspamer, J . N a t . Prod. 53, 401 (1990). 51. T. Tokuyama, T. Tsujita, H. M. Garraffo, T. F. Spande. and J. W. Daly, Tetruhedron 47, 5415 (1991). 52. M. Mensah-Dwumah and J. W. Daly, Toxicon 16, 189 (1978). 53. E. X. Albuquerque, J. E . Warnick, M. A. Maleque, M. C. Kaufman, R. Tamburini, Y. Nimit, and J . W. Daly, Mol. Pharmacol. 19, 411 (1981). 54. J. W. Daly, E. McNeal, F. Gusovsky, F. Ito, and L. E. Overman, J. Med. Chetn. 31, 477 (1988).

252

HlROKl TAKAHATA A N D TAKEFUMI MOMOSE

55. F. Gusovsky, D. P. Rossignol. E. T. McNeal. and J. W. Daly. Proc.. Ntrtl. Actrd. Sc,i. U . S . A . 85, 1272 (1988). 56. K. S. Rao. J . E. Warnick, J. W. Daly. and E. X . Albuquerque. J . Phtrrnicrcol. Exp. Ther. 243, 775 (1987). 57. J. W. Daly, F. Gusovsky, E. T. McNeal, S. Secunda, M. Bell. C . R. Creveling. Y. Nishizawa. L. E. Overman, M. J . Sharp, and D. P. Rossignol. Biochc,nr. Phorintrcnl. 40, 315 (1990). 58. R. P. Polniaszek and S. E. Belmont. J . Org. Chern. 55, 4688 (1990). 59. M. Bonin, R. Besselievre, D. S. Grierson. and H.-P. Husson. Tc.trcrhedro)i Leti. 24, 1493 (1983). 60. R. V. Stevens, Acc. Cliern. R r s . 17, 289 (1984). 61. A. Fleurdnt, J. P. Celerier. and G. Lhommet. Tc~trcrhedron:Asyninic,try 3, 695 (1992). 62. H. Iida. N. Yamazaki, and C. Kibayashi, J . Org. Clreni. 51, 1069 (1986). 63. S. Hanessian, Curhohydr. Res. 2, 86 (1966). 64. N. Machinaga and C. Kibayashi, J . O r g . Chcwi. 56, 1386 (1991). 65. Y. Gao and K . B. Sharpless, J . A m . Cheni. Soc.. 110, 7538 (1988). 66. H. Takahata, H. Takehara. N . Ohkubo. and T. Momose, Tr~trrilic,dron: A.synznze/ry 1, 561 (1990). 67. D. F. Taber. and L. Silverberg. Tetrrrlirdron Lett. 32, 4227 (1991): D. F. Taber. L. Silverberg, and E. D. Robinson. J . A m . C/ic,/n.Soc. 113, 6639 (1991). 68. D. F. Taber, P. B. Deker. and L. Silverberg. J. Org. Clien?. 57, 5990 (1992). 69. I . Collins, M. E. Fox. A. B. Holmes. S. F. Williams, R . Baker, I . J. Forbes. and M. Thompson. J . Ckrnz. Soc.. Perkin Trrrns. 1. 175 (1991). 70. D. Gnecco, C. Marazano, and B. C. Das, J . Chern. Soc.. Chrrn. Cornmiin. 625 (1991). 71. M. Mehmandoust. C. Marazano. R. Singh, B. Gillet. M. Cesario. J . L. Fourrey. and B. C. Das, Tetruhedron Lctt. 29, 4423 (1988). 72. R. P. Polniaszek and S. E. Belmont, J . Org. Clrem. 56, 4868 (1991). 73. Y. Shishido and C. Kibayashi. J . Cliern. Soc., Chem. Corn,nrin.. 1237 (1991). 74. J. Plesek, Chern. Listy. 50, 1854 (1956). 75. L. E. Overman and K. L. Bell. and F. Ito. J . Ani. Chetn. So(,. 106, 4192 (1984). 76. L. E. Overman and N.-H. Lin, J . Org. Chc,ni. 50. 3669 (1985). 77. L. E. Overman and M. J . Sharp, Tetrtihedroii Lett. 29, 901 (1988). 78. B. M. Trost and T. S. Scanlan. J . A m . Chc~m.Soc. 111, 4988 (1989). 79. D. N. A. Fox, D. Lathbury. M. F. Mahon. K. C . Molloy. and T. Gallagher. J . Am. Chcni. Soc. 113, 2652 (1991). 80. T. Honda. M. Hoshi, and M. Tsubuki. Hrt~~rocvc.les 34, 1515 (1992). 81. N. K. Hart. S. R. Johns, and J. A. Larnbei-ton, Arist. J . Chc,tn. 25, 817 (1972): S. R. Johns and J. A. Lamberton. in “The Alkaloids” (R. H. F. Manske. ed.). Vol. 14, p. 325. Academic Press, New York. 1973. 82. C. Fann, M. C. Malone. and L . E. Overman. J . Am. Clrern. Soc.. 109, 6097 (1987): G . W. Gribble. F. L. Switzer. and R. M . Sol. J . Org. Clieni. 53, 3164 (1988); D. L. Comins and Y. C. Myoung. J . O r g . Chert?.55, 292 (1990);D. F. Taber. R. S. Hoerner, and M. D. Hagan, J . Org. Chern. 56, 1287 (IY91). 83. D. H. Hua, N. Bharathi. P. D. Robinson. and A. Tsujirnoto. J . O ~ KClion. . 55, 2128 ( 1990). 84. D. L. Comins and H. Hong. J . A m . Clicwr. SOC. 113, 6672 (1991). 85. D. L. Comins, R. R. Goehring, S. P. Josepf. and S. O’Connor. J . O r g . Chcni. 55,374 ( 1990). 86. Y. Arai, T . Kontani. and T . Koizurni, Ti,/rcihedron: A s y n i n z ~ t r y3, 535 (1992). 87. Y. Arai. M. Matsui. and T. Koizurni. J . Org. C/rc,m. 56, 1983 (1991).

3.

SIMPLE lNDOLIZlDINE ALKALOIDS

253

88. D. P. Rainey, E. B. Smalley. M. H. Crump, and F. M. Strong, Ntrfrrre (London)205, 203 (1965);S. D. Aust and H. P. Broquist. Ntrfrrre (London) 205, 204 (1965). 89. H. P. Broquist. Annrc. Reu. Nrtfr. 5 , 391 (1985). 90. C. M. Harris, M. J. Schneider. F. S. Ungemach. J . E. Hill. and T. M. Harris. J . Am. Chem. Soc. 110, 940 (1988). 91. W. H. Pearson and S. C. Bergmeier, J . Org. Clicm. 56, 1976 (1991):W. H. Pearson. S. C. Bergmeier, and J . P. Williams, J . Org. Chew. 57, 3977 (1992). 92. J.-R. Choi, S. Han. and J. K. Cha, Tetrahedron Lett. 32, 6469 (1991). 93. G . J. McGarvey. J. M. Williams. R. N . Hiner, Y. Matsubara. and T. Oh, J . Am. Chem. Soc. 108,4943 (1986). 94. D. B. Collum. J. H. MacDonard. and W. C. Still, J . Am. Cheru. Soc. 102,21 17 (1980). 95. S. Knapp and F. S. Gibson, J . O r g . Clieni. 57, 4802 (1992). 96. S. Knapp and F. S. Gibson. Org. Synt/i. 70, 1 0 1 (1991). 97. C . M. Harris and T . M. Harris, Terrrrhedron L e f t . 28, 2559 (1987). 98. M. P. Sibi and J . M. Christensen. Tetra/iedro/i Lett. 31, 5689 (1990). 99. H. Takahata, Y. Banba, and T. Momose, Tefruhedron:Asynirnetry 1, 763 (1990). 100. H. Takahata, Y. Banba, M. Tajima, and T. Momose, J . Org. Chem. 56, 240 (1991). 101. T. M. Harris, C. M. Harris, J. E. Hill. F. S. Ungemach, H. P. Broquist. and B. M. Wickwrite, J . O r g . Chem. 52, 3094 (1987). 102. 1. Pastuszak. R. J. Molyneux, L. F. James. and A. D. Elbein. Bioc,/iemistry 29, 1886 ( I 990). 103. M.-P. Heitz and L. E. Overman, J . Org. Chem. 54, 2591 (1989). 104. H . Takahata, Y. Banba, and T. Momose. Tetrrrhedront Asvmmetry 3, 999 (1992). 105. M. J. Schneider. F. S. Ungemach, H. P. Broquist, and T. M. Harris, Tetrtr/iedro,r 39, 29 (1983):F. P. Guengerich. S. J. DiMari, and H . P. Broquist. J . Am. Chem. Soc. 95, 2055 (1973). 106. R. J. Molyneux and L. F. James. S(,ic,nce 216, 190 (1982);D. Davis. P. Schwarz, T. Hernandez, M. Mitchell, B. Warnock. and A. D. Elbein. Pltrrit Pliysiol. 76,972(1984). 107. S. M. Colegate. P. R. Dorling. and C. R. Huxtable. Airsr. J . Chc,m. 33, 435 (1980). 108. M. Hino, 0.Nakayama, Y. Tsurumi. K. Adachi. T . Shibata. H. Terano, M . Kohsaka. H. Aoki, and H. Hamanaka. J . Antihior. 38, 926 (1985). 109. L.F0ddy.J. Feeney.andR. C. Hughes.Bio(,/ie)?i.J.233,697(1986):R. W . McLawhon. E. Berry-Kravis, and G. Dawson. Biochem. Biophvs. R r s . Comnirrn. 134, 1387 (1987); D. R. P. Tulsiani and 0. Touster, J . Biol. Clicwi. 262, 6506 (1987):J. E. Tropea. R. T. Swank, and H. L. Segal. J . B i d . Chem. 263,4309 (1988):A. L. Vlasova. N. A. Ushakova, and M. E. Preobrazhenskaya. Biokhimiyrr ( M o s c w i . ) 56, 1479 (1991):J . F. Haeuw, G. Strecker, J. M. Wieruszeski. J. Montreuil. and J. C. Michalski. Errr. 1. Biochem. 202, 1257 (1991); R. De Gasperi, S. Al Daher, 9. G. Winchester, and C. D. Warren, Biochem. J . 286, 55 (1992). 110. M. J. Humphries, K. Matsumoto, S. L. White, and K. Olden. Proc. Nntl. Accrd. S c i . U.S.A. 83, 1752 (1986):J. W. Dennis, Cancer Res. 46,5131 (1986):D. A. Granato and J. R. Neeser, M o l . Immrrnol. 24,849(1987):S. L.White, K. Schwitze. M. J. Humphries. and K. Olden, Biochern. Biopliys. Res. Commrrn. 150, 615 (1988); M. J . Humphries, K. Matsumoto, S. L. White, R. J. Molyneux, and K. Olden, Cancer Res. 48, 1410 (1988); K. Olden, P. Breton. K. Grzegorzewski, Y. Yasuda, B. L. Bause. 0. A. Oredipe, S . A. Newton, and S. L. White, Pknrmacol. Ther. 50, 285 (1991):G. T. Tan. J. F. Miller, A. D . Kinghorn, S. H. Hughes, and J . M. Pezzuto. Biochem. Biophys. Res. Comrnun. 185, 370 (1992). I l l . M. J. Humphries and K. Olden. Phnrmacol. Ther. 44, 85 (1989); K. Grzegorzewski. S. A. Newton, S. K . Akiyama. S. Sharrow, K. Olden. and S . L . White. Ccincw

254

HIROKI TAKAHATA A N D TAKEFUMI MOMOSE

Commun. 1, 373 (1989); P. Breton, A. Asseffa, K. Grzegorzewski, S . K. Akiyama, S. L. White, J. K. Cha, and K . Olden, Cancer Commun. 2, 333 (1990); P. S. Wright, D. E. Cross-Doersen, K . K . Schroeder, T. L. Bowlin, P. P. McCann. and A. J . Bitonti, Biochem. Pharmacol. 41, 1855 (1991); S. L. White, T . Nagai, S. K. Akiyama. E. J. Reeves, K . Grzegorzewski. and K. Olden, Cancer Commim. 3, 83 (1991); A. Myc, P. DeAngelis, P. Lassota. M. R. Melamed, and Z. Darzynkiewicz, Clin. Exp. Immunol. 84, 406 (1991). 112. G. W. J . Fleet, M. J . Cough, and P. W. Smith, Tetrahedron Lett. 25, 1853 (1984). 113. M. H. Ali, L. Hough. and A. C. Richardson, J . Cheni. Soc., Chem. Cornmun. 447 (1984); M. H. Ah, L. Hough. and A. C . Richardson, Carbohydr. Res. 136,225 (1985). 114. T. Suami, K. Tadano, and Y. limura. Chem. L e t t . 513 (1984); T . Suami, K. Tadano, and Y. limura. Carbohydr. Res. 135, 67 (1985). 115. N. Yasuda, H. Tsutsumi, and T . Takaya, Chem. Lett., 1201 (1984). 116. C . E. Adams, F. J. Walker, and K. B. Sharpless. J . Org. Chrm. 50, 3948 (1985). 117. H. Setoi, H. Takeno, and M. Hashimoto, J . Org. Chem. 50, 3948 (1985). 118. N. M. Carpenter, G . W. J . Fleet, I. C. di Bello, B. Winchester, L. E. Fellows, and R. J. Nash, Tetrahrdron Lett. 30, 7261 (1989). 119. F. B. Gonzalez, A. L . Barba, and M. R. Espina. B d l . Chern. Soc. J p n . 65, 567 (1992). 120. R. B. Bennett 111, J.-R. Choi, W. D. Montgomery, and J. K . Cha, J . A m . Chem. Soc. 111, 250 (1989). 121. W. H. Pearson and K.-C. Lin. Tetrahedron L e t t . 31, 7571 (1990). 122. S. A. Miller and A. R. Chamberlin, J . A m . Chem. Soc. 112, 8100 (1990). 123. K . Tomioka, T. Ishiguro, and K. Koga, Tetruhedron Lett. 21, 2973 (1980). 124. N. lkota and A. Hanaki, C h m . Phurm. Bull. 35,2140 (1987); N. Ikota and A. Hanaki, Chem. Phurm. Bull. 38, 2712 (1990). 125. N. lkota and A. Hanaki, Chem.Phurm. Bull. 36, 1143 (1988). 126. J . M. Dener, D. J. Hart, and S. Ramesh. J . Org. Chem. 53, 6022 (1988). 127. N. lkota and A. Hanaki. Heterocycles 26, 2369 (1987). 128. Y. G . Kim and J. K. Kim, Tetruhedron Lett. 30, 5721 (1989). 129. K. Tadano, Y. Hotta, M. Morita. T. Suami. B. Winchester, and I . Cenci di Bello, Chem. Lett., 2105 (1986);K. Tadano. Y. Hotta, M. Morita, T. Suami. B. Winchester, and I . Cenci di Bello, Bull. Chrm. Soc. Jpn. 60, 3666 (1987). 130. K . Tadano. M. Morita. Y. Hotta, S. Ogawa. B. Winchester, and I. Cenci di Bello, J . Org. Chem. 53, 5209 (1988). 131. L. D. Hohenschutz. E. A. Bell, P. J. Jewess, D.P. Leworthy. R. J. Pryce, E. Arnold, and J. Clardy, Phytochemistry 20, 81 I (1981). 132. R. J . Nash. L. E. Fellows, J. V. Dring. C. H. Stirton, D. Carter, M. P. Hegarty, and E. A. Bell, Phytochemistry 27, 1403 (1988). 133. R. Saul, J . P. Chambers, R. J . Molyneux, and A . D. Elbein, Arch. Biochem. Biophys. 221, 593 (1983); Y. T. Pan. H. Hori, R. Saul, B. A. Sanford. R. J . Molyneux, and A. D. Elbein, Biochemistry 22, 3975 (1983); H . Hori, Y. T. Pan, R. J. Molyneux, and A. D. Elbein, Arch. Biochem. Biophys. 228, 525 (1984); R. Saul, J. J . Gindoni, R. J. Molyneux, and A. D. Elbein. Proc. Natl. Acad. Sci. U . S . A . 82, 93 (1985); V. W. Sasak, J . M. Ordovas, A. D. Elbein, and R. W . Berninger, Biochem. J . 232,759 (1985); T. Szumilo, G . P. Kaushal. and A. D. Elbein, Arch. Biochem. Biophys. 247,261 (1986); B . C . Chambel. R. J. Molyneux. and K. C . Jones, J . Chem. Ecol. 13, 1759 (1987); B. G. Winchester, I. Cenci di Bello, A. C . Richardson, R. J . Nash. L. E . Fellows, N. G. Ramsden, and G . W. J . Fleet, Biochem. J . 269, 227 (1990); T. G. Cooper, C . H. Yeung, D. Nashan, F. Jockehoevel, and E . Nieschlag, Int. J . Androl. 13,297

3.

SIMPLE INDOLIZIDINE ALKALOIDS

255

(1990); A. D. Elbein, FASEB J . 5, 3055 (1991); M. A. Speaman, B. C . Ballon. J . M. Gerrard, A. H . Greenberg, and J. A. Wright, Cancer L e u . 60, 185 (1991);H. Takahashi and P. G. Parsons, J . Invesi. Dermaiol. 98, 481 (1992); P. M. Grochowicz. K. M. Bowen, A. D. Hibberd, D. A. Clark, W. B. Cowden, and D. 0. Willenborg, Transpluni Proc. 24, 295 (1992). 134. B. L. Rhinehart. K. M. Robinson, A. J. Payne, M. E. Wheatley, J . L. Fisher, P. S . Liu, and W. Cheng, Life Sci. 41,2325 (1987);G . Trunan, M. Rousset, and A. Zweibaum, FEBS Lett. 195, 28 (1986). 135. E. Truscheit, W. Frommer, B. Junge, L. Muller, D. D. Schmidt, and W. Wingender, Angew. Chem., h i . Ed. Engl. 20, 744 (1981). 136. M. J . Hamprie, K. Matsumoto. S. White, and K. Olden. Crrrzcer Res. 46,5215 (1986); J . W. Dennis, Cancer Res. 46, 5131 (1986); J . W . Dennis. S. Laferte. C . Waghorne, M. L. Breitman. and R. S. Kerbel, Science 236, 582 (1987): P. B. Ahrens and H. Ankel, J . B i d . Chem. 262, 7575 (1987); G. K. Ostrancer. N . K . Scribner, and L. R. Rohrschneider, Cancer Res. 48, 1091 (1988). 137. P. S. Sunkara, T . L. Bowlin, P. S. Liu, and A. Sjoerdsma. Biochem. Biophys. R e s . Commun. 148, 206 (1987); E. J. Nicols. R. Manger. S. Hakornori, A. Herscovics, and L. R. Rohrschneider. Mol. CellBiol. 5,3467 (1985);M. G . Hollingshead, L. Westbrook, B. J. Toyer, and L. B. Allen, Aniivirul Chem. 2 , 119 (1991): M. G. Hollingshead, G. Melinda. L. Westbrook, M. J . Ross. J. Bailey, K. J. Qualls. and L. B. Allen. Antiviral Res. 18, 267 (1992). 138. B. D. Walker, M. Kwalski, W. C . Goh. K. Kozarsky. M. Krieger, C. Rosen, L. Rohrschneider, W. A. Haseltine. and J . Sodroski. Proc. N a f l . Acad. Sci. U . S . A . 84, 8120 (1987); R . Dagani, Chem. Eng. News, 25 (1987); R. A. Gruters, J . J. Neejes. M. Tersmette. R. E. Y. De Goede, A. Tulp. H. G . Huisman. F. Miedema. and H. L. Plogh, Nature (London) 330, 74 (1987); G. W. J . Fleet, A. Karpas, R. A. Qwek. L. E. Fellows, A. S. Tyms. S. Petursson, S. K . Narngoong, N . G. Ramsden. P. W. Smith, J . C. Son, F . Wildon. D. R. Witty, G . S. Jacob. and T. W. Rademacher. FEBS Lett. 237, 128 (1988); A. Karpas, G. W. J. Fleet, R. A. Dwek, S. Petursson, S. K. Namgoong, N . G . Ramsden, G. Jacob. and T. W. Rademacher, Proc. Nriil. Accid. Sci. U . S . A . 85, 9229 (1988); A. Karpas. G. W. J . Fleet, R. A. Dwek. S . Petrsson. S . K. Namgoong. N. G. Ramsden. G . Jacob, and T. W. Rademacher, Proc. N a f l . Acud. Sci. U . S . A . 86, (1989); R. M. Ruprecht, S . Mullaney, J. Andersen, and R. Bronson, J . Acquired Imrnruze Dejic. Syndr. 2, 149 (1989);P. S. Sunkara, D. L. Taylor, M. S. Kang, T. L. Bowlin, P. S. Liu, A. S. Tyms, and A. Sjoerdsma. Loncei, 1206 (1989): D. C. Montefiori, W. E. Robinson, and W. M. Mitchell. Proc. Nail. Acad. Sci. U . S . A . 85, 9248 (1988); A. S. Tyms. E. M. Berrie. T. A. Ryder. R. J . Nash, M. P. Hegarty, D. L. Taylor. M. A. Mobbeley, J. M. Davis. E. A. Bell, D. J. Jeffries, D. Taylor-Robinson, and L. E. Fellows, Lancet, 1025 (1987);H. C. Holmes, N . Mahmood, A. Karpas. J. Petrik, D. Kinchington, T. O’Connor, D. J . Jeffries, J. Desmyter, and E. De Clercq. Antiviral Chem. 2, 287 (1991); D. Schols. R. Pauwels. M. Witvrouw. J. Desmyter. and E. De Clercq, Aniiviral Chem. Chemoiher. 3, 23 (1992). 139. R. J . Molyneux, J. N . Roitman. G. Dunnheirn, T. Szumilo. and A. D. Elbein, Arch. Biochem. Biophys. 251, 450 (1986). 140. R. J . Molyneux, Y. T . Pan, J. E. Tropea, M. Benson, G. P. Kaushal, and A. D. Elbein, Biochemisiry 30, 9981 (1991). 141. R. C. Bernotas and B. Ganem. Teirahedron Leii. 25, 165 (1984). 142. K. Burgess and I. Henderson, Tetrahedron 48, 4045 (1992). 143. H. Setoi, H. Takeno, and M. Hashimoto, Teirahedron Lett. 26, 4617 (1985).

256

HIROKI TAKAHATA AND TAKEFUMI MOMOSE

144. H. Hamana. N . Ikota, and B. Ganem, J. Org. Chem. 52, 5494 (1987). 145. P. B. Anzeveno, P. T. Angell, L. J. Creemer, and M. R. Whalon, Tetrahedron Lett. 31, 4321 (1990). 146. M. Gerspacher and H. Rapoport, J. Org. Chern. 56, 3700 (1991). 147. J. Mulzer, H. Dehmlow, J. Buscmann, and P. Luger, J. Org. Chem. 57, 3194 (1992). 148. Y . Okude, S. Hirano, T. Hiyama, and H. Nozaki, J. Am. Chern. Soc. 99, 3179 (1977). 149. R. Appel and H.-D. Wihler, Chem. Bey. 109, 3446 (1976). 150. G. W. J. Fleet. N. G. Ramsden, R. J. Molyneux, and J. S. Jacob, Tetrahc&on Lett. 29, 3603 (1988); G. W. J. Fleet, N. G. Ramsden. R. J . Nash, L. E. Fellows, G. S. Jacob, R. J. Molyneux, I. Cenci di Bello, and B. Winchester, Carbohvdr. Res. 205, 269 (1990). 151. J.-L. Reymond, A. A. Pinkerton, and P. Vogel, J . Org. Chem. 56, 2128 (1991). 152. R. Bhide, R. Mortezaei, A. Scilimati, and C. J. Sih, Tetrahedron Lett. 31,4827 (1990). 153. H. h a and C. Kibayashi, Tetrahedron Lett. 32,4147 (1991); H. Ina and C. Kibayashi. J . Org. Chem. 58, 52 (1993). 154. K. Burgess, D. A. Chaplin, and I Henderson, J. Org. Chem. 57, I103 (1992).

-CHAPTER 4-

CHEMISTRY AND BIOLOGY OF CARBAZOLE ALKALOIDS D. P. CHAKRABORTY of' Nirtririil Prodrrcts Cdcrrttn 700 036. Itidia

Institute

1. Introduction ................................................. 11. Occurrence ............................................................................... 111. Chemistry of Carbazole Alkaloids .................................................

................ 258 Tricyclic Alkaloids from Higher Plants Tricyclic Alkaloids from Other Source 290 Synthesis of Tricyclic Alkaloids ........................................................ ........................ 297 Tetracyclic Alkaloids from Higher Plants 306 Tetracyclic Alkaloids from Streptomyces ............................................ F. Synthesis and Transformation of Tetracyclic Alkaloids ..... G. Hexacyclic Alkaloids from Higher Plants ........................ H. Penta- and Hexacyclic Alkaloids from Aspergillus.. .......... .............................. 317 1. Hexa- and Octacyclic Indolocarbazoles J. Synthesis of Hexacyclic Bases ............ .............................. 331 K. Biscarbazole Alkaloids .................................................................... 332 ................ 347 L. Synthesis of Biscarbazoles IV. Physical Properties of Carbazole Alkaloids .............................................. 349 A. Ultraviolet Abso B. Infrared Spectra C. NMR Spectra ... D. Mass Spectra ................................................................................. 350 ............... 350 E. X-Ray Crystallography ... V. Biogenesis of Carbazole Alk VI. Biochemical and Medicinal Properties of Carbazole Alkaloids and Related Compounds .................................. .... 352 A. Antimicrobial Properties .................................................................. 352 B. Antitumor and Tumor-Promoting Activity. ............ C. Antiviral Activity ............................................. D. Cardiovascular-Modulating Activity ..................... ................................. 356 E. Central Nervous System Activity F. Anti-inflammatory Properties .......................................... G. Modulation of Enzyme Activity, Metabolism, and Allergic R ........................ 359 H. Miscellaneous Effects References ......................................................................................... 360 A. B. C. D. E.

257 T H E A L K A L O I D S . VOL. 44 Copyright h IY93 hy Aciidemic Pre\\. Inc. All rightc of reproduction in a n y form reberved

258

D. P. CHAKRABORTY

I. Introduction Graebe and Glaser (1) discovered carbazole (1) from abiological coal tar. The first carbazole from a biological source, the alkaloid murrayanine (2), was isolated by Chakraborty et al. in 1962 (2) from Murraya koenigii Spreng. The structure and antibiotic properties of murrayanine (2) were published in 1965 (3,4).Since then several reviews on carbazole alkaloids have appeared (5-13). This chapter reviews work published after the review in this treatise by Husson in 1985 ( 9 ) .

11. Occurrence

Carbazole alkaloids were first isolated from the taxonomically related genera Murrayu, Glycosmis, and Clausena of the family Rutaceae (subtribe Clausanae, subfamily Aurantodoae). The genera Micromelum (Rutaceae) and Ekebergia (Meliaceae) have also been reported to elaborate carbazole alkaloids (13). Murraya euchrestijolia, obtained from Taiwan, has been found to be the richest source of carbazole alkaloids, providing a variety of novel structures. Some bioactive carbazole alkaloids have been reported from other sources (actinomycetes, blue-green algae) and from mammalian systems. From the aspect of structural considerations, tricyclic to octacyclic alkaloids have been reported. The occurrence of alkaloids reviewed after Husson ( 9 ) is summarized in Table I .

111. Chemistry of Carbazole Alkaloids

A. TRICYCLIC ALKALOIDS FROM HIGHER PLANTS 1. Carbazole Carbazole (1, CI2H9N,mp 225°C) was reported from Glycosmis pentaphylla. It was identified based on IR, UV, and mass spectral data as well as from direct comparison with a pure sample (14).

4.

CHEMISTRY AND BIOLOGY OF CARBAZOLE ALKALOIDS

259

TABLE I OCCURRENCE OF CARBAZOLE ALKALOIDS Compound (formula no.)

Source (Ref.)

Tricyclic alkaloids from higher plants Carbazole (1)

Glycosmis pentaphylla Retz. (DC) (14) Murraya euchrestifolia Murrayafoline-A (3) Hyata (15) Murraya koenigii Spreng. (16) 2-Hydroxy-3-methylcarbazole (4) M . koenigii (16) 2-Methoxy-3-methylcarbazole (7) M . euchrestifolia (17) Murrayastine (8) M . euchrestifolia (17) Murrayaline-A (9) M . koenigii (18) Mukoline (17) M . koenigii (18) Mukolidine (20) M . koenigii (19) Koenoline (27) M . euchrestifolia (20) 3-Formylcarbazole (29) M . euchrestifolia ( 2 l c ) 3-Formyl-7-hydroxycarbazole (29) M . euchrestifolia (20) N-Methoxy-3-formylcarbazole (30) N-Methoxy-3-hydroxymethylcarbazole (35A) M . euchrestifolia (21c) Clausena lansium (21a) 3-Formyl-6-methoxycarbazole (36) M . koenigii (23) Mukonal (37) Murraya siamensis (24) 0-Methylmukonal (38) M . siamensis (24) 7-Methoxy-0-methylmukonal (39) Clausena harmandiana Pierre (25) 7-Methoxymukonal (40) C. lansium (21a) 6-Methoxymurrayanine (41) Clausena anisata (26) 0-Demethylmurrayanine (42) Glycosmis pentaphylla (27) Glycozolidal (43) M . euchrestifolia (28) Murrayaline-B (45) M . euchrestifolia (28) Murrayaline-C (46) C. lansium (21a) Carbazole-3-methylcarboxylate (47) C . lansium (21a) Carbazole-3-carboxylic acid (48) C . lansium (21a) 6-Methoxycarbazole-3-methylcarboxylate

(49) Murrayafoline-B (50) Isomurrayafoline-B (52) Clausenapin (53) Glycomaurrol (55) Euchrestine-A (57) Euchrestine-B (58) Euchrestine-C (59) Euchrestine-D (60) Euchrestine-E (62) Murrayanol(63) Eustifoline-C (64) Ekebergenine (66) Murrayaline-D (67) 7-Methoxyheptaphylline (68A)

M . euchrestifolia (22) M . euchrestifolia (30) Clausena heptaphylla ( I I ) Glycosmis mauritiana (32) M . euchrestifolia (33) M . euchrestifolia (33) M . euchrestifolia (33) M . euchrestifolia (33) M . euchrestifolia (34) M . koenigii (35) M . euchrestifolia (36) Ekebergia senegalensis (37) M . euchrestifolia (34) Clausena harmandiana (25) (continued)

260

D. P. CHAKRABORTY

TABLE I (continued) Compound (formula no.) Carbazoloquinones Murrayaquinone-A (69) Murrayaquinone-B (51) Murrayaquinone-C (87) Murrayaquinone-D (88) Tricyclic alkaloids from other sources Hyellazole (89) 6-Chlorohyellazole (90) I-Methylcarbazole (103) I-Acetylcarbazole (104) Carbazomycin-C (105) Carbazomycin-D (106) Carbazomycinal (107) Carbazomycin-E 6-Methoxycarbazomycinal (109) Carbazomycin-F Carbazomycin-G (110) Carbazomycin-H (111) Carazostatin (112) 3-Chlorocarbazole (120) Tetracyclic alkaloids Furanocarbazoles Eustifoline-D (177) Furostifoline (178) Pyranocarbazoles Dihydroxygirinimbine (179) Pyrayafoline-A (183) Pyrayafoline-B (186) Pyrayafoline-C (190) Pyrayafoline-D (191) Pyrayafoline-E (193) Mukonicine (194) Glycomaurin (eustifoline-A) (195) Eustifoline-B (195A) 7-Methoxymurrayacine (199) Isomahanine (201) Heptazolicine (201A) Pyranocarbazoloquinones Pyrayaquinone-A (202) Pyrayaquinone-B (203) Pyrayaquinone-C (208) Tetracyclic alkaloids from Strepiomyces Kinamycin-A (216)

Source (Ref.)

M. M. M. M.

euchrestifolia euchresiifolia euchresiijolia euchresiifolia

(29) (29) (29) (29)

Hyella cuespiiosa Born et Flah (40) H . caespitosa (40) Tedaniu ignis (42) Tedania ignis (42) Sireptoueriicillium ehimense H-1051-NY 105 (43) S. ehimense (43) s. sp. (44) S . ehimense (43) s. sp. (44) S. ehimense (43) S. ehimense (45) S. ehimense (45) Strepiomyces chromofuscus DC 118 (46) Bovine urine (48)

M . euchresiijolia (36) M . ruchresiifolia (36) M . euchresiijolia (60) M . euchresiijolia (61) M . euchresrifolia (33) M . euchresiijolia (33) M . euchrestijolia (33) M . euchresiijolia (28) M . koenigii (62) Glycosmis mauriiiana (32) M . euchresiifolia (36) Murruya siamensis (24) M . kornigii (35) Clausena hepiaphylla ( 6 ) M . euchresiijolia (62) M . euchresiijolia (62) M . euchrestijolia (20) Sirepiomyces murayamaensis (64,65a,h )

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

261

TABLE I (continued) Compound (formula no.) Kinamycin-B (217) Kinamycin-C (209) Kinamycin-D (218) Prekinamycin (219) Ketoanhydrokinamycin (220) Kinamycin-E (221) Kinamycin-F (deacetylkinamycin-C) (212) Hexacyclic alkaloids from higher plants ( + )-Murrayazoline (241) Murrayazolinol (245) Penta- and hexacyclic alkaloids from Aspergillus Tubingensin-A (246) Tubingensin-B (247) Alfavazole (249) Hexa- and octacyclic Indolocarbazoles Arcyriaflavin-B (252) Arcyriaflavin-C (253) Protein kinase C inhibitor K-252c (262) Protein kinase C inhibitor K-252d (263) Rebeccamycin (254) AT 2433-A1 (272) AT 2433-A2 (273) AT 2433-B 1 (274) AT 2433-82 (275) Staurosporine (251) Tan-1030A (293) Tan-999 (296) Protein kinase C inhibitor K-252a (297) Protein kinase C inhibitor K-252b (298) UCN-OI (298A) Biscarbazoles Indole dimer (305) Bismurrayafoline-A (306) Chrestifoline-D (307B) Bismurrayafoline-B (308) Bismurrayafolinol (309) Oxydimurrayafoline (311) Murrafoline-F (312) Murrastifoline-A (313) Murrastifoline-B (314) Chrestifoline-A (315)

Source (Ref.) S. murayamaensis S.murayamaensis S.murayamaensis S.murayamaensis S.murayamaensis S . murayamaensis S.mrrrayamaensis

(64,65a,b) (64,65a,b) (64,65a,b) (66.67) (67) (67) (67)

M. euchrestifolia (29) M . koenigii (79) Aspergillus tubingensis (80) A . tubingensis (81) Aspergillus f l a w s (82) Arcyria denudata (85) A . denudata (85) Nocardiopsis sp. K-290 (91,92) Nocardiopsis sp. (91,921 Nocardia aerocolonigenes (86-88) Actinomadura melliaura sp. nov. (SCC 1655) (93,94) A . melliaura sp. (93,94) A . melliaura s p. (93,94) A . melliaura sp. (93,94) Streptomyces staurosporsus Anaya, Takahashi, and Omura sp. nov. (83,84) Streptomyces sp. C-71799 (98,99) Nocardiopsis dassonvillei (98,99) Nocardiopsis sp. K-252 and K-290 ( I 00,92) Nocardiopsis sp. (91,92) Streptomyces sp. ( l o l a ) Murraya gleni (102) Murraya euchrestifolia (15) M . euchrestifolia ( 2 l c ) M. euchrestifolia (15) M . euchrestifolia (30) M. euchrestifolia (30) M. euchrestifolia (20) M . euchrestifolia (103) M . euchrestifolia (103) M . euchrestifolia (103)

(continued)

262

D. P. CHAKRABORTY

TABLE I (confinued) Compound (formula no.) Bismurrayafoline-C (316) Bismurrayafoline-D (317) Murrafoline-B (319) Murrafoline-D (320) Murrafoline-E (321) Murrastifoline-D (322) Murrastifoline-E (323) Chrestifoline-B (324) Murrastifoline-C (325) Chrestifoline-C (326) Murrafoline-C (327) Murranimbine (328) Bis-7-hydroxygirinimbine A (328A) Bis-7-hydroxygirinimbine B (328B) Murrafoline (329)

Source (Ref.) euchresfifolia (28) euchrestifolia (28) euchresfifolia (104) euchresfifoh (104) euchrestifoh (20) euchrestifoliu (103) M . euchrestifolia (105) M . euchresfifolia (103) M . euchrestifoliu (103) M . euchrestifoh (103) M . euchrestifoliu (104) M . euchrestifolia (106) M . euchrestifolia ( 1 0 6 ~ ) M . euchrestifolia ( 1 0 6 ~ ) M.euchrestifoh (107) M. M. M. M. M. M.

2. Murruyufoline-A From an ethanolic extract of the root bark of Mirrruyu euchrestfoliu Hyata, murrayafoline-A [3, C,,H,,NO (M+21I ) , mp 52-57"CI was obtained as colorless plates (15). The 1-methoxycarbazole skeleton in 3 was readily detected from the characteristic UV spectrum [A, 225, 243, 251 (sh), 283 (sh), 292, 330, 344 nm, with log E 4.47, 4.58, 4.44, 3.83, 4.01, 3.53, 3.491 and was supported by IR data (v,,, 1640, 1610, 1590, 1505 cm-'). The 'H-NMR data for 3 indicated that ring A was unsubstituted. The signals for an aromatic methoxy group (6 3.76) and an aromatic methyl (6 2.42, s), together with the signals for H-4 (6 7.33) and H-2 (6 6.44), showed that the methoxy group was at C-1 and the methyl at C-3. The structural assignments were substantiated by nuclear Overhauser effects (NOE) in which the enhancement of signals for H-4 and H-2 were observed arising from irradiation of the methyl at C-3 and the methoxy at C-1,respectively. From these data, murrayafoline-A was formulated as l-methoxy-3-methylcarbazole, which was previously known as a synthetic intermediate for murrayanine (6).

4.

CHEMISTRY AND BIOLOGY OF CARBAZOLE ALKALOIDS

263

3 . 2-Hydroxy-3-methylcarbazole

The roots of M . koenigii furnished (16) a compound with the formula C,,H,,NO (M+ 197) and a melting point of 245°C (4). From the IR (urnax3520, 3400, 1635, 1600 cm-') and U V (A, 235, 254, 258, 304, 322 nm with log E 4.65,4.25,4.26,4.19, 3.66) spectral data, the isolate was considered to be a phenolic carbazole. The 'H-NMR data showed that it has a phenolic group (6 8.1), a methyl substituent (6 2.37), and an unsubstituted ring A; signals were observed for H-4 (6 7.68) and H-1 (6 7.0). The characteristic U V spectrum of acetate 5 for 3-methylcarbazole (6) and its isolation from 4 by zinc dust distillation confirmed the position of the methyl group at C-3. From these data, compound 4 was assigned the structure 2-hydroxy-3-methylcarbazole, which was confirmed by 13CNMR data and direct comparison with a synthetic specimen (6).

4. 2-Methoxy-3-methylcarbazole

From a petroleum ether extract of the seeds of Murraya koenigii, an isolate with the formula C,,HI3NO (7, mp 245°C) was obtained (16). The IR (v,,, 3425, 1640, 1600, 1708, 820, 750 cm-') and UV (A, 235, 255, 300, 328 nm with log E 4.35, 3.8, 3.9, 3.30) spectra showed it to be a carbazole derivative. 'H-NMR data showed that ring A was unsubstituted, and signals for an aromatic methoxy group (6 3.77), and an aromatic Cmethyl (6 2.35) as well as for H-4 (6 7.5) and H-1 (6 6.95) were observed. On zinc dust distillation, the compound furnished 3-methylcarbazole ( 6 ) , and on demethylation it furnished 2-hydroxy-3-methylcarbazole (4). Consequently, the isolate was formulated as 2-methoxy-3-methylcabazole (7) (6).

5 . Murrayastine From the bark of Murraya euchrestifolia, murrayastine [8, C,,H,,NO, 224, 247, 255, 298, 332) and (M' 271.1191)] was isolated (17). UV (A, IR data showed it to be a carbazole alkaloid, and the 'H-NMR spectrum showed an aryl methyl signal (6 2.47), aryl methoxy groups (6 3.95, 3.96,

264

D. P. CHAKRABORTY

and 4.0), and meta-coupled H-4 (6 7.32) and H-2 (6 6.65) signals. The enhancements of the H-4 and H-2 resonances on irradiation of the aryl methyl group showed that the methyl group was at C-3. The ortho-coupled H-5 signal (6 7.56, d, J = 8 Hz) showed that C-5 and C-6 were unsubstituted. A 1,7,8-trimethoxy-3-rnethylcarbazolestructure for 8 is consistent with the physical data and has been confirmed by synthesis.

OCH3

CH3O

CH~O

H

OCH;

CHO

6. Murrayalinr-A

From the bark of M. ei~hrrstifolim, murrayaline-A [9, C,,HISNO3 ( M + 269.1002), mp 248.5"CI was obtained as pale yellow prisms (17). The U V and IR data showed the isolate to be a carbazole derivative with a , ~ cm-I). The 'H-NMR spectrum hydrogen-bonded formyl group ( v , , , ~1640 showed signals for an aryl methyl (6 2.35), two aromatic methoxy groups (6 3.90, 3.99, H-4 (6 7.95, s) and H-1 protons (6 6.91, s), and orthocoupled H-5 (6 8.02, d , J = 9 Hz) and H-6 protons (6 6.77, d, J = 9 Hz). In NOE experiments irradiation of the methyl caused enhancement of the H-4 signal, whereas irradiation of the methoxy signals resulted in enhancement of the H-1 and H-6 signals, indicating that the methoxy groups were located at C-2 and C-7. From these data, murrayaline was carbazole (9), which has formulated a 2,7-dimethoxy-3-methyl-8-formyl been confirmed by synthesis. Synthesis of Murrayastine and Murrayaline-A. Alkaloids 8 and 9 were synthesized using the diphenylamine route (17). l-Bromo-2-methoxy-4methylbenzene (lo), on reaction with 5,6-dimethoxyaniline acetate (11) in pyridine in the presence of Cu and K,CO,, followed by hydrolysis, provided the diphenylamine derivative 12 required for the synthesis of murrayastine. On the other hand, l-bromo-3-methoxy-4-methylbenzene (13) on reaction with the dimethyl acetal of 6-formyl-5-methoxyaniline acetate (14) furnished the diphenylamine derivative 15. On hydrolysis, compound 15 furnished the diphenylamine aldehyde 16 required for murrayaline synthesis. Compounds 12 and 16, on cyclization with palladium acetate in N,N-dimethylformamide (DMF), furnished murrayastine (8) and murrayaline-A (9), respectively.

4.

CHEMISTRY AND BIOLOGY OF CARBAZOLE ALKALOIDS

265

7 . Mukoline Mukoline [17, C,,HI,NO2 ( M + 227), mp 115-12o"C], isolated from the roots of Murraya koenigii ( I @ , indicated the presence of a I-methoxycarbazole chromophore with an additional alcoholic hydroxyl group, based 221, 242, 252, 255, 280, 290, 320 nm with log E 4.60, 4.85, on UV ,A( 4.65,4.0, 3.4, 3.6, 3.0) and IR (v,,, 3440, 3240, 1610 cm-') data and color reactions. In the 'H-NMR spectrum, a benzylic methylene (6 4 . 7 3 , a hydroxyl group (6 4 . 9 , and an aromatic methoxy group (6 3.9), besides the signals for H-5, H-4, and four other protons of the carbazole nucleus, were discernible. The isolate afforded the N-methyl derivative 18 and an 0-acetate 19 with a U V spectrum similar to that of 17, showing that the hydroxyl group was not phenolic. On oxidation with active MnO,, 17 furnished aldehyde 20 (mp 152-155"C), the UV spectrum of which was similar to that of 3-formylcarbazole. On decarbonylation 20 furnished 1-methoxycarbazole (21). From the nonidentity of 20 with murrayanine (2) and other data, mukoline was formulated as l-methoxy-6hydroxymethylcarbazole (17).

8. Mukolidine Mukolidine [20, C,,H,,N02 (M+ 225), mp 152-55"C], isolated from M . koenigii (18), was shown by IR and UV spectroscopy to be a 3formylcarbazole derivative. From the 'H-NMR data, the presence of an aldehydic proton (6 10.8) and an aromatic methoxy group (6 4.25), in addition to the aromatic proton signals, including those of H-4 and H-5, was readily discernible. The physical data for mukolidine and the identity of its borohydride reduction product with mukoline, as well as its identity

266

D. P. CHAKRABORTY

Q +H3c'0\ -

HOHC

(23) N=NCI +

0

NHN (2.4)

(22) H

3

c

I

i

+

w

0

Dehydrogenation methylation 4

( Diazomethane)

H (25)

OCH~

(26)

'

O

I D D Q oxdn.

(20)

Sodium borohydride

reduction -

(17)

with the oxidation product of mukoline, showed it to be l-methoxy-6formylcarbazole (20). The structures of both 17 and 20 have been confirmed by synthesis as follows. 2-Hydroxymethylenecyclohexanone (22) on condensation with diazonium chloride (23,) obtained from p-toluidine under JappKlingemann conditions, furnished the hydrazone 24, which on indolization furnished the ketotetrahydrocarbazole 25. Dehydrogenation (Pd/C) and subsequent methylation with diazomethane furnished l-methoxy-6methylcarbazole (26). On oxidation with 2,3-dichloro-5,6-dicyano-l,4benzoquinone (DDQ), 26 furnished mukolidine (20), which on subsequent reduction with sodium borohydride, yielded mukoline (17). 9. Koenoline

Koenoline (27, C,,H,,N02, mp 213"C), isolated from the root bark of Murraya kocnigii (19), displayed a UV spectrum (A, 225, 241, 251, 258, 279, 289, 323, 335 with log E 3.58, 3.63, 4.04, 3.86, 4.36, 4.22, 4.71, 4.56)

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

267

f&-JqCH20R

H

OCHJ

(27)R = H ;( 2 8 ) R = COCH3

that indicated the presence of a 1-methoxycarbazole chromophore. The IR spectrum (v,,, 3445, 3235 cm-I) showed the presence of NH and OH groups, and the isolate formed a monoacetate (28, mp 110°C). In the 'HNMR spectrum, the presence of an aromatic methoxy (6 4.01), a benzylic methylene (6 4 . 3 , H-4 (6 7.66, lH, br s), a H-2 proton (6 6.95, lH, br s), s), and an unsubstituted ring A were readily discernible. Consequently, (27) the formulation of koenoline as 1-methoxy-3-hydroxymethylcarbazole was rational and was confirmed from I3C-NMR data and partial synthesis from murrayanine (2) by sodium borohydride reduction (6). w

c

H

2

O

H

Sodium borohydride reduction

"

OCH3

(2)

( 27)

10. 3-Formylcarbazole From Murraya euchrestifolia, 3-formylcarbazole [29, C13H,N0 (M+ 195.0683)] was obtained as a colorless oil (20). It was identified from its characteristic UV spectrum, which was also supported by IR and

H (29) R = H (29A) R = OH

I

O C H ~ (30) R =CH0;(35A)

R = CH2OH

'H-NMR data. 3-Formylcarbazole reported from Clausena fansiurn (21a) melted at 158-159°C like synthetic 29 (mp 153-154°C) (21b). 11. 3-Formyl-7-hydroxycarbazole

From Murraya eucharestifolia, 3-formyl-7-hydroxycarbazole [29A, C,,H,NO, (M+ 21 l)] was obtained as a colorless powder. The structure was determined from physical (UV, IR, 'H NMR) data ( 2 1 ~ ) .

268

D. P. CHAKRABORTY

12. N-Methoxy-3-formylcarbazole

From M . euchrrstifolia, the oil N-methoxy-3-formylcarbazole [30, C14HIINOZ (M+ 225.04)] was isolated (20). UV (A,, 236, 272, 288, 320 nm) and IR (Y,,, 1690 cm-I) spectral data showed it to be a 3-formylcarbazole derivative. The 'H-NMR spectrum indicated signals for a 3formylcarbazole except that the signal for N-H was replaced by a methoxy group (6 4.27). These data led to formulation of the isolate as N-methoxy3-formylcarbazole, which has been confirmed by synthesis as described below (22). Treatment of 4a,9a-cis-l,2,3,4,4a,9a-hexahydroxycarbazole (31) with methyl chloroformate in methylene chloride and trimethylamine produced the corresponding 9-methoxycarbonyl compound 32 (mp 68-69°C). The 6-iodo derivative 33 was obtained by treatment of 32 with iodine and sodium periodate, and reaction of 33 with 30% aqueous HIOz and sodium tungstate, followed by methylation with diazomethane, furnished 1,2,3,4tetrahydro-6-iodo-9-methoxycarbazole (34),from which the 6-formyl compound 35 was obtained by treatment with butyllithium in tetrahydrofuran (THF) and quenching in dry DMF. The formyl derivative, on dehydroge(30). nation with DDQ in benzene, afforded 9-methoxy-3-formylcarbazole

Methyl c h l o r o f o r m a t e in CHZCIZ+N(CH~)~

I

H (31)

COOCH3 (32)

A Ikali n e hydrolysis, sod. t u n g s t a t e 0xdn.I and m e t h y l a t i o n

I

OCH3

(3L)

I

C 0OC H - j

(33)

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

269

13. N-Methoxy-3-hydroxymethylcarbazole From M . euchresrifolia (21c), N-methoxy-3-hydroxymethylcarbazole [35A, CI4Hl3NO2 (M+ 227.0895)] was isolated as a colorless oil. From the UV, IR, 'H-NMR, and I3C-NMR data it was considered to be N-methoxy3-hydroxymethylcarbazole. The proposed structure was further supported by conversion of 35A to 3-hydroxymethylcarbazole by catalytic hydrogenation and by NaBH, reduction of 30 to yield 35A.

14. 3-Formyl-6-methoxycarbazole 3-Formyl-6-methoxycarbazole [36, C I 4 H'NOz I ( M + 225), mp Clausena lunsium (21a), showed a UV spectrum 135-136"C], isolated from (A, 229, 245, 281, 297, 334 nm with log E 4.38, 4.17, 4.35, 4.47, 3.80) similar to that of 3-formylcarbazoles. The structure of 36 was established from 'H-NMR data, which were similar to those for glycozoline with the aryl methyl signal being replaced by a signal for a formyl group.

(36)

15. Mukonal From the stem bark of M . koenigii (23),mukonal [37, C,,H,NO, (M+ 211), mp 238"CI was isolated. From the UV (A,, 234, 247, 278, 297, 342 nm with log F 4.42, 4.21, 4.54, 4.58, 4.06) and 1R (v,,, 1640 cm-') data, and its color reactions, the isolate was considered to be a 3-formylcarbazole derivative with a chelated hydroxyl group. The 'H-NMR data showed the presence of an unsubstituted ring A, an H-4 singlet (6 8.4) deshielded by the formyl at C-3, a chelated hydroxyl (6 11.76), and an aldehydic proton at C-3 (6 10.16). From these data, a 2hydroxy-3-formylcarbazole formulation of mukonal (37) was advanced, which was supported by the "C-NMR spectrum and the identity of the isolate with an authentic sample of 3-formyl-2-hydroxycarbazole ( 6 ) . 16. O-Merhylmukonal

O-Methylmukonal [38, C,,HI,N02 (M' 225), mp 189-189.5"CI was obtained from the roots of Murruya siamensis (24). The U V and IR

270

D. P. CHAKRABORTY

data suggested the presence of a 3-formylcarbazole skeleton. From the 'H-NMR data, an unsubstituted ring A, an aromatic methoxy group (6 4.50), an aldehydic proton (6 10.45), deshielded singlets for H-4 and H-5(6 8.50 and 7.12), and a singlet for H-1 were detected. The 3-formyl2-methoxycarbazole formulation for 0-methylmukonal(38) was consistent with the above data and was supported by I3C-NMR data. 17. 7-Methoxy-0-methylmukonal 7-Methoxy-0-methylmukonal [39, C,,H,,NO, (M+ 255.0875), mp 219-220"Cl was obtained from Murruya siumensis (24).The UV and IR data suggested the presence of a 3-formylcarbazole system, and the 'HNMR data for 39 were similar to those for 38 with the exception of an additional methoxy group (6 3.85). The signals for H-5(6 7.97, d , J = 8.3) and H-6 (6 6.83, dd, J = 8.3,2.2) showed that H-5 was ortho-coupled, whereas H-6 was both ortho- and meta-coupled, suggesting the location of an additional methoxy group at C-7. Consequently, formulation 39 was consistent with the data and was subsequently confirmed by 13C-NMR data. 18. 7-Methoxymukonal 7-Methoxymukonal [40, CI,H,,NO, (M+ 241)] was isolated from the root bark of Clausena harmandiana (25). U V (A, 222, 238, 244, 254, 288, 302, 340 nm with log E 4.27, 4.30, 4.30, 4.20, 4.33, 4.54, 3.8) and IR (vmax 3350, 1619 cm-') data for the isolate showed the presence of a 3formylcarbazole skeleton with a hydroxyl group chelated to the formyl group. The 'H-NMR data were very similar t o those of 39 except that the methoxy group was replaced by a hydroxyl(6 1 I .56) chelated to the formyl group (6 9.88) at C-3. From these data, formulation of the alkaloid as 2hydroxy-3-formyl-7-methoxycarbazole(40) was advanced and later confirmed by I3C-NMR data. 19. 6-Methoxymurrayanine 6-Methoxymurrayanine [41, CI5Hl3NO3 (M+ 255), mp 23 1-233"C], isolated from Clausena lansium (21a), showed UV data characteristic for a 3-formylcarbazole derivative (A,, 239, 251, 287, 294, 335, 349 nm with log E 4.35, 4.20, 4.40, 4.36, 3.85, 3.30), which was also supported from IR data. The 'H-NMR data showed the alkaloid to possess an additional methoxy group as well as signals for murrayanine. From the 'H-NMR data for H-5, the methoxy group was placed at C-6. This conclusion was also supported by I3C-NMR data.

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

271

20. O-Demethylmurrayanine

O-Demethylmurrayanine [42, C,,H,NO, (M+2 1 I), mp 237-239"Cl was isolated from the root bark of Clausena anisata (26). From the IR and UV spectra (A, 226, 244, 255, 278, 291, 336, 346 nm with log E 4.40, 4.50, 4.39, 4.59, 4.45, 4.22, and 4.22) and color reactions, the isolate was considered to be a phenolic 3-formylcarbazole derivative. 'H-NMR spectroscopy showed that ring A was unsubstituted and that it had metacoupled signals for H-4 and H-2, as well as an aldehydic proton at 9.89. The hydroxyl group was located at the 1 position, and the l-hydroxy-3formylcarbazole structure for 42 was further supported by the 13C-NMR spectrum. 21. Glycozolidal

Glycozolidal [43, C,,H13N0, (M+ 255), mp 185"C], isolated from the roots of Glycosmis pentaphylfa (27), appeared to be a 3-formylcarbazole derivative, judging from the UV ,,A( 235, 245, 303, 340 nm with log E 4.4, 4.2, 4.2, 4.06) and IR (vmax 3500, 1675 cm-I) data. The 'H-NMR spectrum showed the presence of a formyl group (6 9.9), two aromatic methoxyl groups, a singlet for the H-4 proton (deshielded because of the formyl at C-3), and a meta-coupled H-5 proton (6 7.6), besides the signals for three other aromatic protons (H-1, H-7, and H-8). Thus, structure 43 was advanced for glycozolidal, which was confirmed by synthesis from glycozolidine (44)by DDQ oxidation.

22. Murrayaline-B Murrayaline-B [45, C15H,,N03(M+ 255), mp 240-247"C], isolated from 223, 259 (sh), Murraya euchrestifolia (28),showed a UV spectrum [A,

272

D . P. CHAKRABORTY

303, 380 nm] similar to that of murrayaline-A (9). The 'H-NMR spectrum of 45 showed the presence of an aryl methyl (6 2.34), a methoxy group (6 4.02), a C-8 aldehydic proton (6 10.58), H-4 (6 7.79) and H-5 protons (6 8.16), and two deuterium-exchangeable protons at 6 8.42 and 10.79. In NOE experiments, irradiation of the aromatic methoxy (6 4.02) and aryl methyl signals (6 2.34) showed enhancement in the H-6 (6 6.89) and H-4 (6 7.73) signals, demonstrating that the aromatic methoxy was at C-7 and the aromatic methyl at C-3. On methylation with CHJ, murrayaline-B (45)furnished murrayaline-A (9).

-

(9)

OH

23. Murrayaline-C

Murrayaline-C (46, C,,H,,NO,), obtained as a pale yellow powder from M . euchrestifolia (28),had a UV spectrum similar to that of murrayalineA (9). The 'H-NMR spectrum showed signals similar to those for murrayaline-B, except that it had an additional aldehydic proton signal (6 9.95) chelated to a hydroxyl group (6 11.41) replacing the methyl at C-3. Thus, the structure of murrayaline-C was assigned as 46.

24. Carbazole-3-methylcurboxylate

Carbazole-3-methylcarboxylate [47, C,,H, ,NO2 (M 225), mp 168-170°C], isolated from Clausena lansiirm ( 2 1 ~ 1showed , a U V spectrum [A, 241, 251, 273 (sh), 287, 320, 332, 346 nm with log E 4.57, 4.50,4.57, 4.68 (sh), 3.82 (sh), 3.73 (sh), 3.301 typical of a carbazole derivative. The 'H-NMR spectrum showed a signal for a COOCH, group and other signals for a carbazole derivative. From the 'H-NMR data alkaloid 47 was considered to be carbazole-3-methylcarboxylate. +

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

273

25. Carbazole-3-carboxylic Acid Carbazole-3-carboxylic acid (48, C,,H,NO,, mp 270-272"C), also isolated from Clausena lansium (21a),showed absorption maxima in the UV at 230, 237, and 269 nm (log E 4.18, 4.7, 4.22). From the 'H-NMR data and the identity of its methyl ester with 47, the compound was formulated as carbazole-3-carboxylic acid (48). 26. 6-Methoxycarbazole-3-methylcarboxylate

6-Methoxycarbazole-3-methylcarboxylate [49, C,,H,,NO, (M 255), mp 147-14SoC], isolated from Clairsena Iunsium (21a).showed a UV spectrum characteristic for a carbazole derivative. 'H-NMR and IR data revealed a methoxycarbonyl function. 'H-NMR spectroscopy showed a signal for an aromatic methoxy group (6 3.91) and other signals for the protons of a carbazole nucleus, and compound 49 was considered to be a 6-methoxy3-methylcarboxylate. +

27. Murrayajkline-B Murrayafoline-B [50, C,,H,,NO, (Mi 295)] was obtained as a colorless syrup from Murraya euchrestifolia (29). The UV data of 50 (A,, 234,254, 304, 338 nm) and its shift in alkali showed it to be a phenolic carbazole, which was consistent with the IR data (v,,, 3600, 3475, 1620, 1595 cm-I). The 'H-NMR spectrum of 50 showed the presence of an aromatic methoxy group (6 3.90), an aromatic methyl group (6 2.42), and a dimethylallyl side chain (6 1.88, 1.72, 3.90, 5.31). In addition, it showed two broad

274

D. P. CHAKRABORTY

singlets arising from the H-4 and H-2 protons and ortho-coupled H-5 and H-6 proton signals. Photooxidation of murrayafoline-B (50) gave murrayaquinone-B (51)(29), and thus murrayafoline-B was represented by structure 50.

28. Isomurrayafoline-B Isomurrayafoline-B [52, CI9H,,NO, (M+ 295.1567), mp 158-161"C] was obtained from M. euchrestifoliu (30).The IR and UV spectra ,X,(, 213, 237, 264, 310, 330 (sh)] showed it to be a carbazole derivative. The 'HNMR spectrum showed the presence of an aromatic methoxy (6 3.90), an aromatic methyl (6 2.38), a hydroxyl function (6 4.74), and signals for H-5 (6 7.70, d, J = 8 Hz) and H-4 (6 7.67, s). Enhancement of the H-4 and H-6 signals by irradiation of the aromatic methyl and aromatic methoxy signals, respectively, showed that the methyl was at C-3 and the methoxy group at C-6. From these data, structure 52 was assigned as isomurrayafoline-B, an isomer of 50.

29. Clausenapin

Clausenapin [53, C,,H,,NO ( M + 279), mp IOI"C] was obtained from Clausena heptaphylla (31). The UV spectrum (A, 224, 240, 250, 290, 320,335 nm with log E 4.55,4.50,4.59,4.29,5.39,5.50) showed the isolate to have a I-methoxycarbazole chromophore, which was also supported by the IR data. In the 'H-NMR spectrum, signals for an aromatic methyl (6 2.28), a dimethylallyl side chain (6 1.75, 1.85, 3.62, and 5.31, t), an unsubstituted ring A, and a H-4 singlet were observed. Carbazole (1)and 3-methylcarbazole (6) were obtained by zinc dust distillation of 53, and

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

275

compound 54 was obtained by cyclization with HBrIacetic acid. From these chemical studies, structure 53 was assigned to clausenapin, which was previously reported as the Huang-Minlon reduction product of indizoline (6). 30. Glycomaurrol

Glycomaurrol [55, C,,H,,NO (M+ 265.1466), mp 149-15o"C], isolated from the stem bark of Glycosmis mauritiana (32),was shown by U V [A, 230, 243 (sh), 255 (sh), 268 (sh), 289, 298, 327 nm with log E 4.15, 4.08, 3.97,3.89,3.79,3.88,3.05]and IR spectroscopy to be a phenolic carbazole. 'H-NMR data showed the presence of a dimethylallyl side chain [6 I .76, 1.95 (3H, s each), 3.99 (d, J = 6.5 Hz), 5.40 (d, J = 6.5 Hz)], a phenolic hydroxyl (6 4.68), and an aromatic methyl group (6 2.52), in addition to the aromatic proton signals similar to those of glaucomaurrol. From these data, glycomaurrol was considered to be 3-methyl-5-(3',3'-dimethylallyl)6-hydroxycarbazole ( 5 9 , which was confirmed by conversion to dihydroglycomaurin (56).

31. Euchrestine-A

Euchrestine-A [57, C,,H,,NO, (M+ 281.1414)] was obtained as a colorless oil from M. euchrestifolia (33). The UV spectrum of 57 showed the presence of a 2,7-oxygenated carbazole chromophore. The 'H-NMR spectrum showed the presence of an aromatic methyl (6 2.3% a dimethylally1 side chain [6 1.78, 1.89 (3H, s), 3.59 ( 2 H , d), 5.36 (t, J = 6.7 Hzl, unsubstituted H-4 and H-1 protons, and ortho-coupled H-5 and H-6 protons. By comparison of the 'H-NMR data with those of isomurrayafolineB, structure 57 was assigned to euchrestine-A.

32. Euchrestine-B Euchrestine-B [58, C,,H2,N0, (M+ 363.2173)], a pale yellow oil from M. euchrestifolia (33), showed a UV spectrum, like 57, characteristic of

276

D. P . CHAKRABORTY

(57) (58) (59) (60)

R1

R2

H

H

R3 DMA

R.4

Euchrcstinc-A Euchrestine-B Euchrestine-C Euchrcstinc-D

H

H

H

Gcranyl Gc rany I

H Gcranyl H

CH3 H H

Gcranyl H H 0 x 0 Gcranyl

CH3 H

H

H

(61) O-Dimethyl Euchrcstinc -D (62) Euchrestinc-E

CH3

H

Abbr. :

DMA=

0 x 0 Gcranyl =

+ ; Gcranyl=

/

/

.s

a 2,7-dioxygenated carbazole. From the 'H-NMR data, the presence of H-4, H-1, and ortho-coupled H-5 and H-6 protons were discernible. The compound showed signals for an aromatic methoxy (6 3.90), an aromatic methyl (6 2.38), and the protons of a geranyl side chain [6 I .57, 1.62, 1.88 (3 methyl, s), 2.06 (4H, m), 3.62 (2N, d, J = 6.7 Hz), 5.32 (lH, t , J = 6.7 Hz), 5.07 (lH, m)]. In NOE experiments, irradiation of the methoxyl group led to enhancement of the H-6 resonance, proving the methoxy group to be at C-7. From these data, euchrestine-B was assigned structure 58. 33. Euchrestine-C Euchrestine-C [59, C2,H,,N02 (M+ 349)] was obtained as a brown powder from M. euchrestifolia (33).Like euchrestine-A and -B, it had a UV spectrum characteristic for a 2,7-dioxygenated carbazole system. The mass spectral peak at mlz 266 (M+ - 83) and the 'H-NMR signal for HI (6 6.81), together with the similarity to euchrestine-B of signals for other aromatic protons and an aromatic methyl, but without the methoxyl signals, allowed placement of the geranyl chain at C-8. Consequently, structure 59 was consistent with the physical data for euchrestine-C. 34. Euchrestine-D (M+ 349.201 l)] was obtained as a pale Euchrestine-D [60, C23H27N02 yellow oil from M . euchrestiJofia (33).The U V data were similar to those

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

277

for other euchrestines, suggesting the presence of a 2,7-dioxygenated carbazole system. The 'H-NMR data showed signals for the H-5 (6 7.74, dd, J = 8.4, 1 Hz), H-6 (6 6.88, dd, J = 8.4, 2.4 Hz), H-8 (6 6.82, d, J = 2.4 Hz), and H-4 protons (6 7.56), as well as signals for a geranyl side chain. The 0-dimethyl ether 61 also showed similar signals for the aromatic protons and geranyl chain, besides signals for two aromatic methoxy groups. From these data, structure 60 was assigned to euchrestine-D, which was substantiated by NOE data.

35. Euchrestine-E Euchrestine-E 162, C,,H,,NO, (M+ 365.1993)] was obtained as a pale brown oily racemate from M . e u c h r e s t i f o h (34). 'H-NMR spectroscopy of 62 showed signals for an aryl methyl (6 2.38), an H-4 proton (6 7.651, and an H-5 (6 7.66) doublet coupled with H-6 (6 6.73, d , J = 8.4 Hz). 'H-'H correlation spectroscopy (COSY) data showed the presence of three directly coupled protons (6 2.84, 3.10, and 3.99) arising from a benzylic methylene and adjacent methine bearing an oxirane ring, suggesting the presence of a geranyloxy chain. Another methylene proton signal (6 2.17) coupling with an olefinic proton (6 5.07) and other oxymethine protons overlapping with signals of ally1 methyls and methyls attached to oxygen were observed. The 'H-NMR data of 60 and 62 were similar, except for some signals in the geranyloxy chain of 62, with the mass spectral peak at mlz 226 (M+ - 139) being due to loss of a fragment from the geranyloxy chain. These data showed structure 62 to be consistent for euchrestine-E; however, the stereochemistry of 62 was not assigned. 36. Murruyanol

Murrayanol [63, C,,H,,NO, (M+ 363.2196), mp 161"C], isolated from M . koenigii ( 3 3 , showed UV [A,, 208,236,262,296,316,328; log E 4.66, 4.69, 4.88, 4.25 (sh), 4. I 1 (sh), 4.631 and IR spectra typical for a phenolic carbazole derivative. 'H-NMR spectroscopy showed signals for a geranyl side chain [6 1.57, 1.63, 1.88 (Me groups), 3.62 (2H,J = 6.7 Hz), 5.02-5.12 ( I H , s) 2.06-2.09 (4H, m), 5.33 ( l H , m)], as well as H-5 (6 7.74) H-4 (6 7.72, l H , d, J = 8.6 Hz), H-8 (6 6.80), and H-3 protons (6 6.83, l H , J = 1.6 Hz), an aromatic methyl at C-6 (6 2.38), and an aromatic methoxy at C-7 (6 3.91). These data established the compound to be l-geranyl-2-

OH

278

D . P. CHAKRABORTY

hydroxy-6-methyl-7-methoxycarbazole(63). The structure of murrayanol (63) has also been supported by mass spectral (13) and I3C-NMR data. 37. Eustifoline-C

Eustifoline-C [64, C,,H2,N0 (M+ 333)] was obtained as a brown powder from M. euchrestifolia (36). The UV and IR data suggested the presence of a 3- or 6-oxygenated carbazole skeleton. 'H-NMR data showed the presence of an aromatic methyl group (6 2.51), a hydroxyl function (6 4.901, and a geranyl side chain [6 1.56, 1.63, 1.94 (3H, s each), 2.09 (4H, m), 4.0 (2H, d, J = 6.4 Hz), 5.05 (IH, m), and 5.41 ( I H , J = 6.4 Hz)] which was also supported by the mass spectral peak at mlz 210 (13). In NOE experiments, irradiation of the benzylic methylene of the geranyl chain caused enhancement of the H-4 signal, (6 7.90) whereas irradiation of the methoxy group of the 0-methyl derivative (65) of eustifoline-C enhanced the H-7 signal (6 7.08). These data indicated that the hydroxyl group in 64 was at C-6 and the geranyl chain at C-5; thus, eustifoline-C is represented as structure 64. Geranyl I

\

38. Ekebergenine

Ekebergenine [66, Cl9HI9N0,(M+ 293), mp 230-23loC] was obtained from Ekebergia senegalensis (Meliaceae) (37). The 'H-NMR spectrum of the N-methyl derivative 66A (C,,H,,NO,, mp 155- 157°C) showed signals for an aldehyde (6 10.37), one aromatic methoxy (6 3.99), and an N-methyl (6 4.13) group, as well as signals for a dimethylallyl side chain (6 1.70, 1.89, 4.19, 5.28, t). In addition, it showed signals for H-5 (6 8.10), H-6 (6

Y ( 6 6 ) R = H ;(66A) R = CH3

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE A L K A L O I D S

279

7.70), H-7 (6 7 . 9 , H-8 (6 7.4), and H-2 protons (6 7.4). It lacked signals for H-4, H-3, and H-1, thereby allowing assignments of the substitutions in ring C. 13C-NMR data for 66A, including heteronuclear correlation spectra, confirmed the structures of N-methylekebergenine (66A) and ekebergenine (66). 39. Murrayaline-D

Murrayaline-D [67, C,,H,,NO, ( M + 377.1984)] was obtained as a brown oil from M . euchrestifofia (34). In the 'H-NMR spectrum, signals for a methoxy (6 3.93, an aldehydic function at C-3 (6 9.98), ortho-coupled H5 and H-6 protons (6 7.88, 6.95, J = 8.8 Hz each), singlets for H-1 and H-4 protons (6 7.40, 8.08), and signals for a geranyl side chain [a 1.56, 1.60, 1.91 (3H each), 2.10 (4H, m), 3.68 (2H), 5.08 (IH), and 5.36 ( l H , t)] were readily discernible. The mass spectral peak at mlz 254 represented by the ionic species 68 (13) also confirmed the geranyl side chain in 67. From NOE experiments, the E configuration of the double bond of the side chain was suggested. Enhancement of the proton signal at C-6 (6 6.95) showed that the methoxy group was at C-7. These data led to the formulation of murrayaline-D as 67.

CHJO

40. 7-Methoxyheptaphylline

7-Methoxyheptaphylline [68A, C,,H,,NO, (M+ 30911, isolated from Cfausena harmandiana (25), showed evidence (IR, UV, and 'H-NMR spectra) for the presence of CHO, OH, NH, and OMe functions, as well as a dimethylallyl side chain on a carbazole skeleton. By comparison of the 'H-NMR data of 68A with those of 40, the DMA chain was placed at C-1. From the 13C-NMR data for C-8 (6 95.57), the methoxy group was located at C-7. and the alkaloid was formulated as structure 68A.

280

D. P. CHAKRABORTY

41. Murrayaquinone-A

Several carbazoloquinones have been discovered in Murraya euchrestifolia. Murrayaquinone-A [69, Cl,H,,NO2 (M+ 271), mp 246-247OC1, the first carbazoloquinone alkaloid, was isolated from M . erichrestifolia (29). IR (vmaX3200, 1650, 1595 cm-I) and UV ,A[ 225, 258, 293 (sh), 395 nm with log E 4.63, 4.51, 3.85, 3.931 data showed it to be a carbazole-1,4quinone. This was substantiated by I3C-NMR data (6 180.4 and 183.4). 'H-NMR spectroscopy showed the presence of an unsubstituted ring A, an ally1 methyl signal at 6 2.19, and a vinylic proton signal at 6 6.31. From these and biogenetic considerations, the methyl was placed at C-3, showing murrayaquinone-A to be 1,4-quinonoid-3-methylcarbazole (69). Compound 69 has also been obtained by oxidation with Fermi's salt, as well as by photochemical oxidation, of 1-hydroxy-3-methylcarbazole(70). Palladium-assisted cyclization of analogs of diphenylamine derivatives has been utilized in the synthesis of murrayaquinone-A (37a). 0

H

O

42. Murrayaquinone-B Murrayaquinone B [51, C,,HI,N03 (M+ 309), mp 221-223OC1, also obtained from M . euchrestifolia (29), showed IR (v,,, 3280, 1655, 1640,

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

281

210 (sh), 232, 264, 310 (sh), 400 nm; log E 1610 cm-') and U V [A,, 4.28, 4.58,4.44, 3.21, 3.661 data characteristic for a 1,4-~arbazoloquinone fragment, like murrayaquinone-A. This was supported by 13C-NMRdata for the carbonyl functions (6 179.8, 183.7). The characteristic ally1 methyl signal (6 2.13) and the vinylic proton (6 6.42) were readily discernible from the 'H-NMR data. In addition, signals for ovtho-coupled H-5 and H-6 protons (6 7.98, 7.02, d , J = 9 Hz), one methoxy group (6 3.91), and a dimethylallyl side chain [6 1.74, 1.85, 3.6, 5.23(t)] were observed, which

(51)CH3O

WCH O C H ~

282

D. P. CHAKRABORTY

were also supported by 13C-NMR data. From the results of NOE experiments, the methoxy group was placed at C-7 and the dimethylallyl side chain at C-8. Thus, the structure of murrayaquinone-B was advanced as 51, which was confirmed by synthesis (38). The azidocinnamate 73, obtained by condensation of 4-(1 , I-dimethylal1yloxy)benzaldehyde (74) with methyl azidoacetal(75), by heating in tolu(76). The methoxy indole ene, furnished 6-hydroxy-7-dimethylallylindole 77 obtained by methylation of 76 with CH,I, on Claisen condensation with 4-methylbutyrolactone, gave lactone 78. On heating in aqueous dioxane, 78 gave alcohol 79 (mp 79-80°C), which, on pyridinium chlorochromate oxidation, gave the aldehyde 79A. Through cyclization at room temperature with boron trifluoride etherate, 79A furnished 1,7-dimethoxy-3methyl-8-(3’-methylbut-2-enyl)-9H-carbazole (80), which, by photooxidation, furnished murrayaquinone-B (51). Synthesis of murrayaquinone-B has been achieved by Ramesh and Kapil via murrayafoline-B by several routes (39), one of which is mentioned here. 7-Hydroxy-3-methyl- 1-oxo- 1,2,3,4-tetrahydrocarbazole(81) on acetylation furnished a 7-acetoxy derivative (82), which, on aromatization with Pd/C in diphenyl ether, yielded l-acetoxy-3-methyl-7hydroxycarbazole (83) as one of the products. Condensation with 2methyl-3-buten-2-01 (84) in presence of boron trifluoride etherate gave l-acetoxy-7-hydroxy-3-methyl-8-(3’-methyl-but-2-enyl)carbazole (85). On methylation of 85, the 7-methoxy derivative 86 was obtained which,

HO

R H

O

(81) R=OH (02) R=OCOCH3

OCOCH3 (83)

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

283

on hydrolysis with sodium acetate in methanol, gave murrayafoline-B (50). Oxidation of 86 with pyridinium chlorochromate (PCC) furnished murrayaquinone-B (51). 43. Murrayaquinone-C Murrayaquinone-C [87, C,,H,, NO, (M+ 377), mp 158-159"C], obtained from M. euchrestifolia (29), showed the presence of a 1,4-benzoquinone system in the UV (A, 233, 267, 460 nm) and I3C-NMR spectra (6 183.6 and 179.7), like other carbazoloquinones. The IH-NMR spectrum was similar to that of murrayaquinone-B (51)except that the signals for the dimethylallyl side chain were replaced by signals for a geranyl side chain [6 1.56, 1.61, 1.85 (3 methyl signals), 2.05 (4H, s), 3.58 (2H, d, J = 7 Hz), 5.03 ( I H , m), and 5.26 (IH, t, J = 7 Hz)], which was substantiated by the mass spectral peak at mlz 254. The positions of the methoxy group at C-7 and the geranyl group at C-8 were substantiated by the results of NOE experiments. Thus, murrayaquinone-C was formulated as 87. 0

(87) R1 = OCH3;Rz = Geranyl ( 8 8 ) R1 = OH ;R2 = Geranyl

44. Murrayaquinone-D Murrayaquinone-D [88, C,,H,,NO, (M+ 363), mp 164- 168"C], obtained (29)along with murrayaquinone-C, showed IR, UV, and I3C-NMR characteristics very similar to those of 87. The 'H-NMR spectrum of 88 was very similar to that of 87, with the exception that one aromatic methoxy signal was replaced by a phenolic hydroxyl group, showing the isolate to be a demethylated murrayaquinone-C, which was confirmed by methylation of murrayaquinone-D to murrayaquinone C with diazomethane. Therefore, murrayaquinone-D was formulated as 88. B. TRICYCLIC ALKALOIDS FROM OTHERSOURCES 1 . Hyellazole

The first carbazole alkaloid from a marine source (40), namely, hyellazole [89, C,,H,,NO (M+ 287), rnp 133-134"C], isolated from the bluegreen alga Hyella caespitosa, showed IR (v,,, 3490 cm-') and UV (A,,,

284

D . P. CHAKEUBORTY

226, 232, 250, 260, 292, 304, 338, 352 nm with log E 4.53, 4.55, 4.53, 4.13, 4.09, 4.26, 3.65, 3.69) spectra characteristics of a carbazole derivative. 'H-NMR spectroscopy showed the presence of an unsubstituted ring A and signals for an aromatic methoxy (6 3.79), an aromatic methyl (6 2.14), an H-4 proton (6 7.70), and five aromatic protons (6 7.6-7.35). These data (89), suggested hyellazole to be l-phenyl-2-methyl-3-methoxycarbazole which has been supported by I3C-NMR spectroscopy.

2. 6-Chlorohyellazole The 6-chloro derivative of hyellazole 190, C2,,HlhCIN0(M+ 312), mp 163-164"CI was isolated from the same source along with hyellazole (40). It displayed a 'H-NMR spectrum very similar to that of 89 except for the absence of the H-6 signal, indicating the chloro substituent to be at C-6. The proposed structures of hyellazole (89) and its 6-chloro derivative (90) were confirmed by X-ray crystallography of 6-chlorohyellazole and by several syntheses. Synthesis of Hyellazole and Chlorohyelluzole. The syntheses of 89 and 90 by Kano et al. (41) involved the formation of a 2-vinylindole derivative as the starting material. Subsequently, formation of 2,3-divinylindole and thermal ring closure of the 2,3-divinyl system led to the syntheses of the natural products as described below. N-Benzoylsulfonylindole (91) or its 5-chloro derivative (91A), on reacting with propiophenone, furnished alcohols 92 and 93. These compounds, on hydrolysis, furnished the indoles 94 and 95. In the case of hyellazole, the intermediate formyl derivative 96 was obtained by Vilsmier-Haack reaction, whereas in case of chlorohyellazole the intermediate formyl derivative was obtained via oxalyl chloride derivative 97 and the keto ester 98. Compound 98, on hydrolysis, furnished the ketoacid 99, which, by decarboxylation, afforded the formyl derivative 100. The formyl derivatives 96 and 100, on Wittig reaction, furnished the divinylindoles 101and 102, which were converted to alkaloids 89 and 90 by thermal cyclization.

4.

285

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

R q = R y & - J Propiono\

7

I 7O2 Ph

I

02

Ph (92) R = H

Ph (91) R = H

(93) R = C I

(91A) R = C I

J

Hydrolysis

-

~CcgJ-JCy

/

R N

(94) R = H

/

\

N

Ph

I

(95) R = C I

i

Vilsmier-Haack

R

4 0 2 C 'QJ-$y?

a

(96) R = H (100) R - CI

Ph

(97)

Ph

(98) R = E t (99) R = H

Wittiq

RQq.p I c

H

~

c

H

~

~

~

m

Ph

0

~C"3 y

s

~

\

(101) R = H (102) R = C I

/

(89) R = H (90) R = C I

3 . 1-Methylcarbazole

From the marine sponge Tedania ignis (42) was isolated I-methylcarbazole [103, C,,H,,N (M+ 181.0811), mp 120°C]. The 'H-NMR spectrum of 103 included aromatic protons typical of a carbazole system, except that H-1 was replaced by a signal for an aromatic methyl group (6 2.56).

~

~

0

c

D. P. CHAKRABORTY

286

(103) R = CH3 (lob) R = COCH3

4. I-Acetylcarbazole

Also from the marine sponge Tedania ignis was isolated l-acetylcarbazole (104, C,,H,,NO). The isolate showed in the IR spectrum the presence of an amine and an acetyl function (v,,, 3444, 1671 cm-'1. The 'H-NMR spectrum showed signals for a carbazole system and lacked the signal for a H-1 proton, showing instead a signal for an acetyl group (6 2.8, 3H, s). The physical data indicated the compound to be I-acetylcarbazole (104), which was supported by I3C-NMR data.

5. Carbazomycin-C The structural determinations of eight antibiotic alkaloids isolated from Streptoverticillium ehminse and a Streptomyces sp. by Japanese groups primarily rests on detailed work on carbazomycin-B. The chemistry of carbazomycin-A and -B has been reviewed by Husson ( 9 ) .The chemistry of the six other alkaloids (13) is reviewed here. Carbazomycin-C [105, C,,H,,N03 ( M + 291.1 19), mp 198-198.5"C], obtained from Streptoverticillium ehimense (43), was considered to be a carbazole derivative from UV data [A, 227, 248, 260 (sh), 287 (sh), 295, 341, 354 nm with E 24,900, 24,000, 12,550, 7600, 12,200, 3000,42001. The 'H-NMR spectrum showed the presence of two aryl methyl (6 2.33, 2.36), two aromatic methoxy (6 3.74, 3.84), and one hydroxyl group (6 8.06), as well as signals for meta-coupled H-5 (6 7.7), H-7, and H-8 protons (6 R2 R 3 q $ : 3

(105) Carbazomycin C ; R1 = C H 3 , R2= OH9 R 3 = OCH3 (106) Carbazomycin D; R1 = C H 3 9 R2 = R 3 = OCH3

(107) Carbazomycinal ; R1 = CHO Y R2 = OH * R 3 = H (108) 0-Methyl carbazomycinal;Ri = C H O , R 2 = OCH3, R 3 = H (109) 6-Methoxy carbazomycinal; R1 = CHO * R 2 = OH * R3 = OCH3

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

287

6.91 and 7.3 dd). These data suggested a formulation of I ,2-dimethyl-3,6dimethoxy-4-hydroxycarbazole for carbazomycin-C (105). This has been supported by I3C-NMR data in which the methoxy-induced shifts of C-6, C-5, C-7, C-8, and C-8a were reported at 6 154 (s), 106.2 (d), 113.9 (i), 1 11.5, and 135.6. The structure was confirmed by X-ray crystallographic studies. 6. Carbazomycin-D

Carbazomycin-D [106, C,,H,,NO, (M+ 285.1374), mp 129.5-130"Cl (43) showed IR and UV data similar to those of carbazomycin-C. Compared to the 'H-NMR data of carbazomycin-C, it had an additional aromatic methoxy group, which suggested it to be the O-methyl ether of carbazomycin-C. This was confirmed by methylation of carbazomycinC with dimethyl sulfate and alkali to yield carbazomycin-D. Thus, carbazomycin-D was formulated as 106.

7. Carbazomycinal Carbazomycinal [107, CI5Hl5NO3 (M 255.0875), mp 224"C], isolated from a member of the genus Strrptouerticillii~m(44),was considered to be a I-formylcarbazole derivative from IR (vmax1660 cm-l) and UV (A,, 214, 227, 263, 295, 320, 372 nm with F 24,400, 23,200, 12,200, 16,300, 5000,8400) data. From the 'H-NMR data, ring A was considered unsubstituted, and I3C-NMR data confirmed the presence of a carbonyl group. NOE enhancement after irradiation of the methyl group showed proximity to the formyl group, suggesting the methyl to be at C-2. In the IH-NMR spectrum of O-methylcarbazomycinal (lOS), the signal of the additional O-methyl group was deshielded (6 3.8) owing to ring A, which suggested that the hydroxyl group of carbozomycinal was at C-4. Thus, carbazomycinal was formulated as 1 -formyl-2-methyl-3-methoxy-4-hydroxycarbazole (107). +

8. 6-Methoxycarbazomycinul

Along with carbazomycinal was isolated an antibiotic substance [109, C,,H,,NO, (M+ 285.1017), mp 221"CI (44).Like carbazomycinal, the UV (A,, 215, 227, 245, 268, 310, 382 with E 27,000, 24,500, 13,000, 13,000, 17,000,8500) and IR data for the isolate showed it to be a I-formylcarbazole derivative. The IH-NMR spectrum, as conipared to carbazomycinal, showed an additional methoxy group that was placed at the 6 position in view of the meta-coupled signals of €3-5 (6 7.72, d , J = 2.4 Hz) and orthocoupled H-8. In a long-range selective proton decoupling experiment (LSPD), irradiation of H-3 caused collapse of both C-4b and C-6, whereas collapse of the signals for C-7 and C-8a was observed on irradiation of

288

D. P. CHAKRABORTY

H-5 and collapse of C-5 and C-8a on irradiation of H-7. The alkaloid was therefore formulated as 6-methoxycarbazomycinal (109) which was supported also by I3C-NMR data. Nakamura and co-workers isolated both carbazomycinal and its 6-methoxy derivative and named them carbazomycin-E and carbazomycin-F (43). 9 . Carbazomycin-G

Carbazomycin-G [110, CI5H,,NO, (M 257), mp 241-243"C], isolated from Streptoverticillium ehimense (4.9, showed UV and IR data suggesting a carbazole chromophore. The 'H-NMR data showed signals for two aromatic methyl (6 1.60, 2.01), and one aromatic methoxy (6 3.7), and H5 [6 8.05 (deshielded by C=O at 4)], H-6, H-7, and H-8 protons (6 7.21-7.50). The I3C-NMR spectrum showed signals for a tertiary methyl (6 27.9), a quarternary carbon (6 67.3) carrying a hydroxyl, a vinylic methyl (6 10.10), and a methoxyl group (6 59.2). The carbonyl signal at 6 177.5 could be reconciled with a 2-methoxydienone formulation. From these data carbazomycin-G could be represented by structure 110, which has been confirmed by X-ray crystallographic analysis. +

(110) Carbazomycin G ; R = H ( 1 1 1 ) Carbazomycin H ; R = OCH3

10. Carbazomycin-H

Along with carbazomycin-G was isolated carbazomycin-H [111, C,,H,,N04 (M+ 287), mp 228-230°C](45). The similarity of the IR and UV spectra with those of carbazomycin-G suggested a similar chromophoric system. The 'H-NMR spectrum showed an additional methoxy group (6 3.84) as compared with carbazomycin G. The H-5 signal was meta coupled (6 7.66, d , J = 2.4 Hz) and showed a diamagnetic shift of 0.57 ppm as compared with that of 110, which could be due to the position of the methoxy group at C-6. The methoxy-induced shifts of the aromatic protons of ring A were in conformity with the assignment of the methoxy at C-6. Thus, carbazomycin-H (111)was identified as 6-methoxycarbazomycin-G. 1 1 . Carazostatin Carazostatin [ l U , C2,H25N0 (M+ 295), mp 149-152"C], isolated as a pale yellowish powder from Streptomyces chromofuscus DC 118 (461,

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE A L K A L O I D S

289

displayed IR (v,,, 3460, 3360 cm-I) and U V (A,,,, 218, 235, 254, 266,303, 342; E 153,000, 141,000, 76,000, 60,000, 83,000, 20,000) data that showed it to be a phenolic carbazole. The 'H-NMR spectrum showed signals for an aromatic methyl (6 2.38) and H-4 (6 7.3), H-5 (6 7.94, dd, J = 8.0, 1.5 Hz), H-6 (6 7.17, ddd, J = 8.0, 8.0, 1.5 Hz), H-7 (6 7.37, IH, ddd, J = 8.0, 8.0, 1.5 Hz), and H-8 protons (6 7.42, dd, J = 8.0, 1.5 Hz), demonstrating that ring A and C-4 were unsubstituted. Dimethylcarazostatin (113) showed signals for an aromatic methoxy (6 4.05) and an N methyl (6 3.9) group. The signals for a normal heptyl chain were readily ascertained (6 2.87, 1.65, 1.46-1.30, 8 H , m, 0.91, 3H and J = 7.0 Hz for the C-methyl). Like many carbazoles of microbial origin, the aromatic

H

(120)

290

D. P. CHAKRABORTY

methyl was placed at C-2 and the hydroxyl at C-3. Thus, carazostatin could be formulated as l-heptyl-2-methyl-3-hydroxycarbazole(112), which has also been confirmed by I3C-NMR data and synthesis (47). Diels-Alder reaction of 1-heptylpyrano [3,4-b]indole-3-one (116) [prepared from indolylacetic acid (114) with octanoic anhydride (115)l with 3-trimethylsilylpropynoate (117) afforded carbazole (118,) which, on reduction, gave carazostatin (112). 1,2-Dialkyl-3-trimethylsilylcarbazole (119) also furnished carazostatin (112) when it was mercurodesilylated followed by hydroboration and oxidation in the same reaction sequence used in case of hyellazole (89) and carbazomycin-B (127). 12. 3-Chlorocarbazole

One alkaloid, 3-chlorocarbazole 1120 ( M + 201)], was obtained from bovine urine (48). From IR, UV, and 'H-NMR data (6 7.18-7.50, 7H, m and 6 7.90-8.15,3H, m), it was considered to be 3-chlorocarbazole, which was confirmed by direct comparison with a synthetic specimen.

C. SYNTHESIS OF TRICYCLIC ALKALOIDS The tricyclic carbazole alkaloids have attracted the attention of synthetic organic chemists because of their structural novelty and biological activity. As a result, various new strategies have been developed to synthesize these alkaloids. Transition metal-catalyzed metal diene complexes have been utilized in the total synthesis of carbazomycins, koenoline, murrayanine, and mukonine. For the synthesis of carbazomycin-A and -B, iron tricarbonyl hexadiene complex (121) has been utilized in the electrophilic aromatic substitution (122, 123) at room of 2,3-dimethyl-4-methoxy-5-hydroxy/methoxyaniline temperature (49). The 5-methoxy derivative provided the substituted product 124 on standing for 3 days, whereas the substituted product 125 was obtained after 2 hr of refluxing. Acetyl derivative 126, on MnO, oxidation, gave O-acetylcarbazomycin-B (127A). Deacetylation of 127A gave carbazomycin-B (127). Selective oxidation of 124 furnished the iron-complexed product 128, demetallation of which with methylamine oxide gave 3demethylcarbazomycin A (129). Methylation of 129 furnished carbazomycin-A (130). Optimized iron-mediated amine cyclization (50) has been utilized for broad and general access to 1-oxygenated carbazole alkaloids, resulting in the synthesis of mukonine (131), murrayanine (2), koenoline (27). The iron complexed cation 121, when reacted with arylamines 132 and 135, furnished the iron-complexed intermediates 133 and 136, which, on cyclization under different experimental conditions, furnished the alkaloids

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

1

?CH3

OH

H (130)

(132)

(133)

CH3

291

292

D. P. CHAKRABORTY

mukonine (131) and murrayanine (2). Reduction of murrayanine (2) furnished koenoline (27).

N H2

N 02 (134)

(135) CH3CN

I

; 25OC

Mn02

Toluene

Lewis acid-catalyzed aliphatic diazo coupling has been utilized in the synthesis of glycozoline (141) by Chakraborty and Roy (13). Iodinecatalyzed thermal cyclization of anthranilic acid (142) has been found to yield carbazole (1) as one of the products (51).

C

H

$

m

c

H

3

W-K. reduction -dc h y drogc nat ion

H

H

O

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

293

3-Methylcarbazole (6), glycozoline (141), and glycozolidine (44) have been synthesized from appropriate diphenylamines using degassed Raney nickel in the presence of p-cymene in a sealed tube (13). R 2 0 , N n ‘ 1 H 3

Degassed Raney n i c k e i

H

H

Synthesis of murrayafoline-A was reported by Martin and Moody (52). Under Claisen condensation conditions, reaction of indole-2-methylcarboxylate (144) with 4-methylbutyrolactone (145) yielded the product 146, from which the alcohol 147 was obtained by hydrolysis and decarboxylation. Oxidation of 147 with PCC gave the aldehyde 148, which on cyclization with boron trifluoride-methanol gave murrayafoline-A (3).

CO 0 C H3

H (ILL)

(1L 5 ) H O (146) Hydrolysis and Decar b o x y l a t i o n

294

D. P. CHAKRABORTY

2-Hydroxy-3-methylcarbazole (4). an important biogenetic intermediate, has been synthesized by Bergman and Carlson (53)via alkylation of 2-methylindole (149) with aldehyde 150. Reaction of 2-methylindole (149) with a,@-unsaturatedketone 152 in the presence of Pd/C and a molecular sieve afforded a better yield of 2-methylcarbazole (153), a key intermediate in the pathway to most of the carbazole alkaloids of fungi and bacteria (54). 2-Hydroxy-3-methylcarbazole (4) was also obtained by biomimetic hydroxylation (Fe2+,EDTA, and oxygen) of 3-methylcarbazole (6) (55). CHO

I

C H 3C H-CO 0 C2H 5

CH3

--

(150)

H (149)

H (151)

+

1

9 R1

Cat 10% Pd, HOAc r x L8 h r

R = CH3 R1 = R2 = (1521

CH3 (153)

H

Biomimetic

(6)

Hydroxylation-

(L)

Diels-Alder reaction for the synthesis of carbazomycin and hyellazole using indole-2,3-quinodimethaneanalogs pyrano [3,4-h]indole-3-one (154) and 155 has been utilized by Moody and Shah (56). Diels-Alder reaction of 154 with trimethyl silylpropynoate (117) gave carbazole 156, which, on reduction with LiAIH,, yielded the 2-methylcarbazole derivative 158. Mercurodesilylation of 158 gave 160, which by hydroboration and oxidation gave phenol 162. On methylation, the phenol gave 4-deoxycarbazomycin-B (164). Starting with 155, following the reaction sequence 155 + 157 + 161 + 163, hyellazole (89) was obtained. Treatment of 4-deoxycarbazomycin B (164) with N-bromosuccinimide, following protection of the NH group with a tert-butoxycarbonyl group (165), afforded the 4-bromo derivative 166. Subsequent treatment of 166, with l-butyllithium in THF at -78"C, followed by reaction with aryllithium and trimethylborate and alkaline hydrogen peroxide treatment, provided the 4-hydroxy derivative 61. After removal of the N-butoxycarbonyl group carbazomycin-B (127) was obtained. Pindur and Pfeuffer (57) synthesized 4-demethoxycarbazomycin-A(164) and its isomer using [4 21 cycloaddition reactions of appropriate die-

+

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

(156) R = C H 3 (157) R = Ph

(154) R = C H 3 (155) R - P h

W

H

q

1 cAH3

O

c

4

H R (160) R = CH3 (161) R = P h

(158) R = CH3 (159) R = P h

1

Hydro bo ra t i o n oxidation

woH c H3

\

>

\

H R (162) R = CH3 (163) R = P h

W

t-BUO

C

A.

H CH3 3

cH3

= t-BUO

(165)

(167)

A.

bH3

295

296

D. P. CHAKRABORTY

nophiles to a 3-vinylindole. Bergman and Pelcman (54) synthesized 3demethoxyhyellazole (168) starting from 2-vinylindole through several steps using cycloaddition reactions. A new synthesis of 3-demethoxycarbazomycin-B (169) has been reported by Bergman et al. in which intramolecular acyl indole cyclization has been utilized (58). A convenient synthesis of hyellazole has been effected by benzannelation of 2-methoxyindolin-3-one (170) by Sakamato and co-workers (59) Wittig reaction of 170 with phosphonium compound 171 gave 172, which yielded 3-buta- 1,3-dienylindole (173) after reaction with trimethylsilylindole in the presence of hexamethyldisilazane (HMDS). On heating, 173 yielded 174 via an electrocyclic reaction. Ready removal of methanol from 174 gave 3-hydroxycarbazole derivative 175 and the silyl ether 176. The

OCH3 Ac

(170)

(171)

i

c

(175) R = H ( 1 7 6 ) R = Si(CH3)3

(89) R = H (176A) R = A c

4.

CHEMISTRY AND BIOLOGY OF CARBAZOLE A L K A L O I D S

297

latter, on treatment with tetra-l-butylammonium flouride (TBAF), afforded 175, from which hyellazole (89)was obtained after methylation and deacetylation. ALKALOIDS FROM HIGHER PLANTS D. TETRACYCLIC

Pyranocarbazoles of the girinimbine and mahanimbine groups constitute the largest classes of the tetracyclic alkaloids from higher plants. Furanocarbazoles and benzo[b]carbazoles of the kinamycin group constitute further additions to these groups. 1 . Eustifoline-D

Eustifoline-D [177,C,,H,,NO ( M + 221.08411, obtained as a colorless oil from M . euchrestifolia (361, represents a new variant of the tetracyclic carbazole alkaloids with a furan ring system. The IR (vmaX3470 cm-I) and UV (A, 206, 224, 250, 268, 298, 310, 340, 354 nm) data, together with the 'H-NMR signals for the a-and p-furan protons (6 7.81,7.32, H1 each, d, J = 2.6 Hz), suggested a furanocarbazole system. On irradiation of H4 (6 7.97) enhancement of the p-furan proton was observed, showing that the furan was fused with the carbazole system at the 5,6 positions. These findings, together with the 'H-NMR signals for an aromatic methyl and other protons of carbazole ring system, led to the formulation of eustifoline-D at 177.

2. Furostifoline Furostifoline [178,CI5H,,NO( M + 221.0840)], obtained as an oil (36) from the same source as 177 was shown to be a 2-oxygenated carbazole derivative based on IR and UV data. The 'H-NMR spectrum of 178 showed two doublets (6 7.73, 7.00, 1H each, J = 2.0 Hz) attributable to the aand p-furan protons. The 'H-NMR data showed the absence of substitution in ring A (6 8.06, 7.25, 7.37, 7.49), one aromatic methyl, and a singlet for H-4, indicating the compound to have structure 178. This was also supported from NOE experiments when enhancement of H-4 was observed on irradiation of the methyl at C-3.

298

D. P. CHAKRABORTY

3. Dihydroxygirinimbine Dihydroxygirinimbine (179, CI8H,,NO, (M+ 297.1365), mp 189-190"C, [aID-40" (MeOH) } was obtained from the root bark of M . euchrestifolia

(60). A 3-methylcarbazole skeleton with a hydroxyl group was readily 215, 238, discernible from the IR (v,,, 3450 cm-') and UV spectra (A,, 254, 259, 303, 332 nm). The 'H-NMR data for 179 showed an absence of substitution in ring A and signals for H-4 (6 7.72, s) and the aryl methyl group at C-3 (6 2.28), showing that the C-2 was substituted. Doublets at 6 3.79 and 4.9 ( J = 8 H z each), which shifted in the diacetate 180 (mp 159-161°C) to 6 5.8 and 6.22, respectively, indicated the presence of a methine group. These data could be reconciled with a 3-methyl-2',2'dimethyldihydroxypyranocarbazolesystem in 179. Girinimbine (1811, on oxidation with chloroperbenzoic acid, furnished dihydroxygirinimbine and its cis isomer, suggesting that alkaloid 179 is trans-dihydroxygirinimbine.

(179)R = H (trans) (180 1 R = C O C H 3 (182 1 R = H ( c i s )

4 . Pyrayafoline-A

Pyrayafoline-A (183), C,,H,,NO,) was obtained the root bark of M. euchrestifolia ( 1 7 ) . The IR and UV [A,, 222, 239, 286 (sh), 295, 334 nml data suggested it to be a carbazole derivative with a 2',2'-dimethyl-A3' -pyran (DMP) system, which was substantiated by a high-intensity mass spectral peak at mlz 278 (M+ - 15) (6). The 'H-NMR spectrum showed signals for an aromatic C-methyl (6 2.33), an aromatic methoxy (6 3.89), and a DMP( system (6 1.47, 6H and two one-proton doublets at 6 5.67 and 6.58, J = 10 Hz), as well as signals for H-4 (6 7.62, s), H-5 (6 7.63, J = 8 Hz), and H-6 protons (6 7.77 br s). From these data, pyrayafolineA was formulated as 183, which was confirmed by synthesis as follows (17). The diphenylamine derivative 184, obtained by condensing l-bromo3-methoxy-4-methylbenzene (13) with 2,3-(2' ,2'-dimethyl-A3'-pyrano)anilinoacetate (185), on cyclization with Pd(Ac), in DMF furnished pyrayafoline-A (183) (17).

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

299

5 . Pyrayafoline-B Pyrayafoline-B [186, C,,H,,NO, (M+ 279.1266)], obtained from the stem 228,252,285, bark of M. euchrestifolia (33),showed a UV spectrum [A, 296 (sh), 329, 353 nm] similar to that of carbazole alkaloids with a DMP system, and this was substantiated by a high-intensity peak at mlz 264 (M' - 15) in the mass spectrum. The 'H-NMR spectrum showed an aromatic methyl group (6 2.37), H-4 and H-5 protons (6 7.62 and 7.49), resonances for a DMP system [6 1.46, two doublets at 6.48, 5.58 (IH, J = 6 Hz each)], and aromatic signals at 6 6.74 and 6.71. The isolate afforded a monomethyl derivative on treatment with diazomethane, suggesting the presence of a phenolic group. NOE experiments on the methyl ether involving irradiation of the methoxy group and the aromatic methyl group produced enhancement of the signals for H-1 and for H-4, showing that the methyl was at C-3 and the methoxy at C-2. From these data,

H (186) R - H (189) R = C H 3

Ac

( 1 88)

t

300

D. P. CHAKRABORTY

pyrayafoline-B was formulated as structure 186 which was confirmed by synthesis. 7-Acetylaminochromene (187), on reaction with 4-bromo2-methoxytoluene (lo), after refluxing in presence of copper pyridine and K,CO, for 43 hr, furnished the diphenylamine derivative 188. Hydrolysis and subsequent cyclization of 188 with Pd(OAc), in DMF furnished 0methylpyrayafoline-B (189). 6 . Pyrayufoline-C

Pyrayafoline-C [190, C,,H,,NO, (M+ 279.1258)] was obtained along with 186 (33). IR and UV data and the characteristic high-intensity mass spectral peak for the carbazolopyrillium ion at mlz 264 (M+ - 15) (6) showed it to be a phenolic carbazole derivative with a DMP system. This conclusion was also substantiated from the 'H-NMR spectrum, in which signals for a DMP system and H-4 (6 7.64), H-5 (6 7.63, d, J = 8.4 Hz), and H-l protons (6 6.79) appeared. On treatment with diazomethane it gave an 0methyl ether identical with synthetic pyrayafoline-A, showing pyrayafoline-C to be 190.

7 . Pyrayufoline-D Pyrayafoline-D (191, C,,H,,NO,, [aIDOO} was obtained from the same source as 186 and 190 (33) in the form of pale brown powder. The UV [A, 222, 238, 268 (sh), 296, 331, 342 nm] and IR (v,,, 3600, 3450, 3380 cm-') data and the high-intensity peak at mlz 265 showed it to be a phenolic pyranocarbazole derivative, which was also supported by 'HNMR data. Compared with pyrayafoline-C (190), the isolate showed an additional CH,CH,CH=C(CH,), side chain [a 1.75 (2H, m), 2. I5 (2H), 5.70 ( I H , t), 1.57 (3H, s), and 1.65 (3H, s)] in the pyran fragment. On methylation, it afforded a monomethyl ether. In NOE experiments with 0-methylpyrayafoline-D (192), the H-4 and H- 1 signals showed enhancement on irradiation of the aromatic methyl and methoxy groups, respectively. From these data, structure 191 was advanced for pyrayafoline-D.

4.

CHEMISTRY A N D BIOLOGY O F CARBAZOLE ALKALOIDS

301

8 . Pyrayufoline-E Pyrayafoline-E (193, C,,H,,NO,), obtained as a pale brown oil from M . euchrestifolia (28), displayed U V , IR, and 'H-NMR spectra similar to those of pyrafoline-B, suggesting the presence of a phenolic pyranocarbazole system. The mass spectrum showed the characteristic high-intensity peak for a carbazolopyrillium ion (6) at rnlz 264 (M+ - 83), proving the presence of the carbazolopyrillium ion, and a C, unit comprising CH,CH,CH=C(CH,),. 'H-NMR data also supported the presence of this six-carbon fragment [6 5.10 ( l H , t, J = 7.3 Hz), 2.13 (2H, m), 1.70 (2H), 1.65 and 1.57 (3H each)]. From the physical data the structure 193 has been proposed for pyrayafoline-E.

H2C

I

c H*C H=C'

CH3 'CH3

9. Mukonicine

Mukonicine [194, C20H,,N0, (M+ 323), mp 231-233"CI was obtained 226, 240, 300, 342 nm; log E from Murraya koenigii ( 6 1 ) . The U V (A, 4.70, 4.69, 4.59, 4.26) and IR data were close to those of koenimbine, suggesting the presence of a pyranocarbazole system, which was also supported by the characteristic high-intensity peak for the carbazolopyrillium ion at mlz 308 (6) (M+ - 15) as well as 'H-NMR signals for the DMP system [6 I .44 (6H, s), 5.65, 6.5 ( I H each, d, J = 10 Hz)]. It had a singlet for H-4 at 6 7.5, and H-5 appeared at 6 7.4 because of the methoxy at C6. Signals for the aromatic C-methyl at C-3 and two aromatic methoxy groups at 6 3.9 ( 6 H , s) were observed. Zinc dust distillation furnished 3methylcarbazole, and chromic acid oxidation furnished acetone arising from the DMP system. From these data, structure 194 has been suggested for mukonicine.

302

D. P. CHAKRABORTY

10. Glycomaurin

Glycomaurin (195, CIBH,,NO, mp 195-196"C), isolated in 1989 from Glycosmis mauritiana (32), showed a UV spectrum [A,, 215, 231, 262, 270 (sh), 284 (sh), 297 (sh), 312, 322, 378 nm with log E 4.18, 4.23, 3.90, 3.87,3.65,3.57,3.82,3.85,3.23]characteristic of a pyranocarbazole derivative. The base peak in the mass spectrum at m / z 248 showed the presence of a DMP system (6).Signals for H-4 (8 7.92) and for a DMP system were apparent in the 'H-NMR spectrum [8 1.46 (6H, s), 5.82 and 7.28 (2d, J = 9 Hz each)], and the isolate gave a dihydro derivative (56) on hydrogenation. From the 'H-NMR data, the structure of glycomaurin was advanced as 195, which was confirmed by synthesis of 195 along with the linear isomer 198. 6-Hydroxy-3-methylcarbazole (196) and 3-chloro-3methylbutyne (197), in the presence of the phase-transfer catalyst tetrabutylammonium bromide, furnished glycomaurin (195) and isoglycomaurin (198). Eustifoline-A (36) and 195 are identical. F?.

,

(197) H

H

(196)

11. Eustifoline-B

Eustifoline-B [195A, C,,H,,NO (M+ 33 I)] (36) showed a high-intensity mass fragment at mlz 248, showing it to be a pyranocarbazole, like 195, with an additional dimethylallyl unit. The structure of 195A was deduced from UV, IR, and 'H-NMR data as being similar to that of glycomaurin, except that one of the methyls in the pyran ring was extended by a dimethylallyl unit as in mahanimbine. 12. 7-Methoxymurrayacine 7-Methoxymurrayacine [199, CI9H,,O3(M+ 307.1208), mp 21 1-213"CI, 306, 354 nm; isolated from Murraya siamensis (24, showed UV (A,,

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

303

(201)

log E 4.58, 4.19) and IR (Y,,,,, 3460, 1660, 1628, 1603, 1156 cm-I) data characteristic of a carbazole alkaloid. The IH-NMR spectrum showed signals for H-4 [6 8.23 (deshielded by an adjacent formyl group)], H-5 (6 7.97, d, J = 8.4 Hz), H-6 (6 6.83, dd, J = 8.4, 2.2 Hz), and H-8 protons (6 6.97, l H , d, J = 2.2 Hz), as well as signals for a DMP system [6 1.54 (6H, s), 6.90, 5.90 (1H each, J = 9.8 Hz)], an aldehyde proton (6 10.45, s), and an aromatic methoxy (6 3.85). The formyl group was placed at C3, and the DMP group was considered to be fused at the 2 : 1 position; the methoxy was placed at C-7 based on the coupling of H-8. A carbazolopyrillium ion at rnlz 292 (m+ - 15) was also observed. Thus, structure 199 was determined for 7-methoxymurrayacine, which was confirmed by I3C-NMR spectroscopy. 13. Isomahanine

Isomahanine 1200, C2,H2,N02 (M+ 347.1885), mp 184"C], isolated from M. koenigii (35),showed UV [A, 220, 238, 286, 294, 324, 338 nm; log E 4.51, 4.48 (sh), 4.29, 5.52, 3.68, 3.741 and IR spectra suggesting it to be a phenolic carbazole like mahanine (6). IH-NMR data showed signals for a DMP system in which a methyl has been extended by a dimethylallyl fragment, like that in mahanimbine. The mass fragment at mlz 264 (M+ - C,H,,) supports this. The resonance H-5 (6 7.56) was a singlet, and the H-4 and H-3 signals were ortho coupled, showing that the methyl was at C-6 and the phenolic group at C-7. The pyran unit was fused, as in mahanimbine at the 2 : 1 position. 14. Heptazolicine

Heptazolicine [201,C,,H,,N03 (M+ 295), mp 285"C], obtained from Clausena heptaphylla, was shown to be a phenolic carbazole with an

304

D. P. CHAKRABORTY

aldehyde function, judging from the UV and IR spectra. From 'H-NMR (201A), data it was shown to be a 3-formyl-2,2-dimethylpyranocarbazole and it was identified with cycloheptazoline by direct comparison ( 6 ) .

15. Pyrayaquinone-A Pyrayaquinone-A [202, C,,H,,NO, (M+ 293.1053), mp 22"C], a pyranocarbazoloquinone obtained from the stem bark of M . euchrestifolia (62), showed UV ,,A( 220 (sh), 252, 308 (sh), 460 nm] and IR (A,, 1660, 1640, 1010 cm-') spectra typical of a carbazoloquinone system. The 'H-NMR data (6 1.48, 6H, s, 5.78 and 6.4, 1H each, J = 10 Hz) and the mass spectral peak at mlz 278 (M+ - 15) showed the presence of a DMP system. The doublet at 6 6.46 and a 3 H signal at 6 2.16 for a vinylic methyl showed the presence of a substituted quinonoid system. The singlets for H-5 (6 7.79) and H-8 (6 6.83) demonstrated that the DMP system was fused to ring A of the carbazole system, with oxygen substitution at C-7. From these data, structure 202 was advanced for pyrayaquinone-A. 16. Pyrayaquinone-B

Pyrayaquinone-B [203, C,,H,,NO, (M+ 293.10531, mp 244"CI was isolated from M . euchrestifolia (62) along with pyrayaquinone-A (202). The UV and IR data were consistent with the presence of a 1,4-~arbazoloquinone system. A DMP system fused to the carbazoloquinone nucleus was indicated from the 'H-NMR data (6 1.48, 6H, s, 5.70 and 6.60, 1H each, J = 10 Hz) and the characteristic high-intensity mass spectral peak for the carbazolopyrilliurn ion (M+ 278). The signals for 1H at 6 6.44 and 3H for a methyl group at 6 2.14 with long-range coupling were reconcilied with the presence of a vinylic methyl at C-3 and no substitution at C-2. The ortho-coupled doublets for H-5 at 6 7.94 and H-6 at 6 6.87 (J = 9 Hz) showed that the DMP system was fused to ring A at C-7 and C-8, with oxygen at C-7. Therefore, pyrayaquinone-B is the angular isomer of pyrayaquinone-A. For structural confirmation, pyrayaquinone-A and -B have been synthesized using the method for the synthesis of murrayaquinone A (63). 7Amino- (204) and 5-Amino-2,2-dimethyl-2H-chromene (205) were condensed with methylbenzoquinone (71) to afford 2-(2,2-dimethyl-2ffchrornen-7-y1amino)- and 2-(2,2-dimethyl-2H-chromen-5-ylarnino)-5methyl- 1 ,Cbenzoquinones (206 and 207), respectively. Subsequent treatment of benzoquinones 206 and 207 with Pd(OAc), in acetic acid furnished pyrayaquinone-A (202) and pyrayaquinone-B (203).

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

305

(20L)

17. Pyrayaquinone-C Pyrayaquinone-C [208, C,,H,,NO, (Mi361.1661), mp 222"CI was isolated, like pyrayaquinone-A (202) and pyrayaquinone-B (203), from M. euchrestifolia (20a-c). The UV and IR data were consistent with the presence of a 1,4-carbazoloquinonoid system, which was also indicated from the 'H-NMR signals for an aryl methyl at C-3 (6 2.15, d , J = 1.7 Hz) and the olefinic proton doublet at 6 6.45 ( l H , J = 1.7 Hz). The quinonoid carbazolopyrillium ion at mlz 278 (M+ - 83), the 'H-NMR signals (6 5.68, 6.63, l H , J = 10 Hz), the singlet for a methyl at 6 1.44, and other data suggested that one of the methyls of the DMP system was

306

D. P.

CHAKRABORTY

appended by a dimethylallyl unit like those in mahanimbine and related alkaloids. The ortho-coupled 'H-NMR signals for H-5and H-6 (6 7.94 and 6.80) suggested that the pyran ring was fused angularly as in pyrayaquinone-B. Consequently, pyrayaquinone-C was represented by 208. U

E. TETRACYCLIC ALKALOIDSFROM Streptomyces Eight antibiotics of the kinamycin type isolated from Streptomyces murayamaensis form an interesting group of antibiotic carbazole alkaloids (13). The detailed structural work on kinamycin-C forms the basis for structural determinations of the other alkaloids (64,65a,b). 1. Kinamycin-C Kinamycin-C (209, C,,H,,O,,N, (M+ 496), mp 150-153"C, [aID- 24" (CHCI,)}, isolated from S. murayamaensis, was considered to be a pheno-

W

C

H

O

/

/COCH3

\

OH

AN

CH AH

w

0 c

' I

?H H

I

0 '

\

OH

3

?

CN

I

bH

'CH3

4.

CHEMISTRY AND BIOLOGY OF CARBAZOLE ALKALOIDS

307

lic naphthaquinone derivative from UV spectral data and its red shift in alkali. 'H-NMR data of kinamycin-C showed the signals of one tertiary methyl (6 1.3, s), two alcoholic acetoxy groups (6 2.0-2.3), a hydroxyl group (6 2.57), a chelated phenolic group (6 12.0), two protons on vicinal carbons carrying acetoxy groups (6 5.4, 6.2), and three aromatic protons at 6 7.83, 7.5, and 7.6. The isolate yielded a diacetate (210,&,H,O,,N) and an 0-methyl derivative (211).The IR data of derivatives 210 and 211 showed that the phenolic hydroxyl group in 209 was peri to the quinonoid carbonyls. The presence of three acetyl groups in kinamycin was also mp 137-1 36°C). confirmed from its deacetylated product (212,C18H,407N2, From the 'H-NMR data of 212, the presence of an 8-hydroxynaphthaquinone fragment, two vicinal hydroxy groups, a tertiary methyl, and nitrile or isonitrile functions was ascertained. On sodium periodate oxidation, deacetylkinamycin furnished an aldehyde (213,C,,H ,,,06N2), which confirmed the presence of an alcoholic hydroxyl at C-1 and a tertiary methyl at C-2 in 212. By means of preparation of the isopropylidene derivative 214 of deacetylkinamycin-C (212)with acetone, the position of the tertiary hydroxyl at C-2 was ascertained. X-Ray crystallographic analysis of the bromobenzoate of kinamycin-C showed its structure to be 215. Hence, the structure of kinamycin-C was ascertained as 209. The presence of an N-cyano group in kinamycin-C was confirmed by hydrolysis of deacetylkinamycin-C 212 when ammonia was liberated; X-ray crystallography did not unambiguously ascertain the presence of a nitrile or isonitrile function in kinamycin-C.

2 . Kinamyein-A Kinamycin-A (216,C24H20010N2 (M+ 496), mp 139-142°C (dec.), [a]?$ -60" (CHCI,)} displayed UV, IR, and 'H-NMR data broadly similar to those of kinamycin-C, suggesting a structural similarity. Thus, kinamycinA has a hydroxyl group at C-4. The monoacetates of kinamycin-A and

.*- R2

CH3 OH

CN

6R1

308

D. P. CHAKRABORTY

kinamycin-C were identical, showing the isolate to be 4-deacetylkinamycin C. Consequently, the structure of kinamycin-A was advanced as 216. 3. Kinamycin-B

Kinamycin B (217, C,,H,,O,N, (M+ 412), mp 190-193°C (dec.), [a15 -48" (CHCI,)} had physical properties (IR, UV, and 'H-NMR) similar to those of kinamycin-A, showing the two carbonyl functions to be hydrogen bonded. Tetraacetylkinamycin-B was identical with the diacetate of kinamycin-C. From these and other data, the structure of kinamycin-B was advanced as 217. 4 . Kinamycin-D

Kinamycin-D (218, C,,H,,O,N, (M+ 452), mp 170-175°C (dec.), [a]% -37" (CHCl,)} yielded IR, UV, and IH-NMR data similar to those of kinamycin-A, showing it to have a hydroxyl at the 4 position. On acetylation kinamycin-D furnished deacetylkinamycin-C, proving its structure to be 218.

5 . Prekinamycin Prekinamycin [219, C,,H,,N,O,, mp 300°C (dec.)], obtained from Strep254, 288.4, 342, tomyces murayamaensis (66,67), showed UV (A,,

(219)

n

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

309

574 nm; E 5500, 21,700, 5770, 37,700) and IR data characteristic of a carbazole system with a hydroxyl group chelated to a quinonoid function and is thus related to the kinamycins. In the 'H-NMR spectrum, the signals for two meta-coupled H-1 and H-3 protons (6 6.60, 6.69, 1H each, d , J = 1.5 Hz), three aromatic protons at H-5', H-6', and H-7', and an aromatic methyl group (6 2.29) were readily discernible. Two proton signals at 6 11.60 and 12.32 disappeared in the diacetate {C,,H,,N,O, (M+ 402.239), mp >300"C (dec.)]. In the I3C-NMR spectrum of the diacetate, signals for two quinonoid carbonyls (6 174.4 and 192.45), two ester carbonyls (6 170.28 and 170.64), and a cyanamide function (6 83.71) were detected. These data, together with I3C-NMRdata of a sample obtained biosynthetically through feeding experiments, supported the structure of prekinamycin as 219. 6. Ketoanhydrokinamycin

Ketoanhydrokinamycin [220, CI8HIONZO6 (M+ + H + 351), mp 300°C (dec.)] was isolated by Seaton and Gould (67). The U V and IR spectra were similar to those of kinamycins. The 'H-NMR data of 220 showed the presence of two hydroxyls (6 5.94 and 12.06), one of which was a hydrogen-bonded phenolic hydroxyl (6 12.06), but no acetate signals as in the other kinamycins. I3C-NMR data indicated the presence of 1,4quinonoid system (6 183.67, 180.85) and a conjugated carbonyl function (6 188.62). The coupling (2 Hz) of H-3 and H-4 (6 3.89 and 5.34) showed the regiochemistry of the hydrogens. The signal at 6 5.34 showed coupling to a hydroxyl at 6 5.94. The long-range heteronuclear shift correlation (HETCOR) spectrum showed the coupling of the methyl hydrogens at 6 1.53 and the carbonyl at 6 188.62, suggesting the presence of the carbonyl at C-1 of ring C. In NOE experiments, enhancement of resonances at 6 3.89 and 5.34 by irradiation of the methyl group confirmed the positions of H-3 and H-4 in ring C. These data permitted the formulation of ketoanhydrokinamycin as 220.

7. Kinamycin-E Kinamycin-E [221, C,oH,,N,O, (M+ 412.0906), mp >200"C (dec.)] was obtained from a culture extract of S. murayamaensis (67). From the U V (A, 255.6, 277, 295, 408 nm) and IR spectra, it was considered to have a kinamycin-C chromophore, and from the 'H-NMR spectrum it was inferred that the isolate had a kinamycin-D type acetylation pattern. Treatment of kinamycin-D with methanolic potassium carbonate gave a small amount of kinamycin-E, showing the latter to have structure 221.

310

D . P. CHAKRABORTY

8. Deacetylkinamycin-C. Kinamycin-F (212), also isolated from S . murayamaensis (67), was found to be identical with deacetylkinamycin-C based on physical data. Compound 212 was obtained by hydrolysis of kinamycin-D (218).

F. SYNTHESIS AND TRANSFORMATION OF TETRACYCLIC ALKALOIDS 1 . Synthesis

The novelty of the structures of the antibiotics of the kinamycin group has attracted the attention of synthetic organic chemists, and several strategies have been developed to build up the tetracyclic system or proposed intermediates in the biogenesis of kinamycin. Palladium-catalyzed

CONH-t- BU (222)

(224 1

(223)

2 steps

[Me301 BFL OCH3 OH

0 (226) NH2OH in MeOH+ Demethylation BBr3 in C H C l 2 a t -70' C

OH OH

0 (225)

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

311

coupling of 2-bromojuglone (222)with the stannane 223 furnished the intermediate 224, which was transformed in two steps to 225. Treatment of 225 with [(Me,O)+]BF,- in T H F at 23°C gave 226, which, on treatment with NH,OH in methanol and subsequent demethylation, furnished the naphthaquinonoid quinolone 227 considered to be an intermediate in the biosynthesis of prekinamycin (219)(69). In another approach to the synthesis of prekinamycin (68),the entire skeletal unit of prekinamycin was obtained except for the cyano group of the carbazole fragment. Bromonaphthaquinone (228),on annelation with cyclic N-benzylaminoketone (229),furnished in moderate yield (36-50%) tetrahydrobenzo[b]carbazole-l,6, I I-trione (230),which, in refluxing dioxane in the presence of DDQ, gave the benzo[b]carbazole (231),having a carbon framework identical to that prekinamycin, except for the cyano

(231

(230)

substitution at the carbazole nitrogen. A simple approach to the benzo[b]carbazoloquinone skeleton (232) of the kinamycins has also been worked out through the oxidative cyclization of 2-benzoylindole containing a benzylic alcohol in the orrho position (233)with tetra-n-butylammonium perruthenate (TBAP) (70).

& z & N

\

O (232)

H

N

/

O (233)

H

312

D . P. CHAKRABORTY

The synthesis of norgirinimbine (235)and its linear isomer (236)provides an illustration of the thermal insertion of a C, unit into a carbazole skeleton. 2-Hydroxycarbazole-3-carboxylic acid (234), when heated at 145°C in the presence of SbCl,, provided, after workup in three steps, norgirinirnbine and its linear isomer as the end products (13,71).

+ H (236)

Pate1 has reported (72-74) some interesting synthetic studies in relation to analogs of girinimbine and mahanimbine, which have been reviewed previously (13). Synthesis of normahanimbine (237) and regioisomer 238 from 2-hydroxycarbazole (239), under conditions of citral condensation (13), show that the reaction is regioselective, not regiospecific (75).

.

~itral

Py r c f l u x

OH

2. Transformations a. Cyclomer Formation in Muhanimbine. Some results on the formation of cyclomers of mahanimbine have been enumerated previously (6), and further results are described here. Mahanimbine (240) has been thermally transformed into murrayazoline (241) and rnurrayazolidine (242). On prolonged heating only murrayazoline is obtained (i3,76). When passed

4.

-a

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

313

qcH2 (2.40)

\

C H 2 CH=c 'CH3 /CH3

(2.41)

3

through a column of Dowex 50-X8 resin (H form), mahanimbine furnishes racemic murrayazolidine (77). +

b. Transformation of Optical Properties. In isooctane solution, mahanimbine (240) and normahanimbine (237) undergo racemization at 90°C when kept in the dark (77). On sitting in ethanolic solution at room temperature (37"C), mahanimbine undergoes racemization and subsequent optical inversion (+45" to -24.8") when kept for 6 days. The reaction is catalyzed by hydrochloric acid. In chloroform solution the compound does not undergo inversion. The racemization of mahanimbine and its congener, as well as the inversion of 240, could be explained as arising from enone-chromene transformation (13).

c. Photochemical Transformation of Girinimbine and Mahanimbine. Studies on the photochemical transformation of the tetracyclic

(2.43)

H (2.4.4)

314

D.

P. CHAKRABORTY

carbazoles girinimbine (181) and mahanimbine (240) have been reported to result in the expansion of the pyran ring (78). The transformation has been perceived to proceed through an 0-quinolide intermediate and subsequent 1,4-shift and cyclization. The nitrogen lone pair probably participates in the ring expansion, as N-sulfonyl derivatives of the alkaloids do not undergo the transformation.

G. HEXACYCLIC ALKALOIDS

FROM

HIGHER PLANTS

The penta- and hexacyclic carbazole alkaloids of higher plants are derived from a tricyclic carbazole unit and a monoterpene unit. Thus, murrayazolidine, murrayazoline, and related compounds are derived from mahanimbine. 1 . Murrayzoline

(+)-Murrayazoline (241, C,,H2,N0 (M+ 3 3 3 , mp 276-278"C, [ale +2.25" (CHCI,)} was isolated from Murruya euchrestfolia (29).The U V , IR, and IH-NMR spectra showed it to be identical with murrayazoline, except for the optical activity. Thus, compound 241 has been considered to be an optical antipode of murrayazoline. 2. Murrayazolinol Murrayazolinol(245, C,,H,,NO, (M+ 347), mp 290"C, [ a ] ,0" (CHCI,)}, isolated from Murruya koenigii (79), showed UV data (A,,, 248, 365, 305 nm with log ~4.85,4.32,4.40)indicating the presence of a 2-oxygenated carbazole system. The 'H-NMR data were similar to those of murrayazoline, except for an additional hydroxyl group and a carbinyl hydrogen signal at 6 3.8. By comparison of the mass spectral data and the benzylic proton signals of murrayazolinine (245A) (6 3.65) (6) and isomurrayazoline (6 3.08) (13) with those of murrayazoline (6 3.83) and consideration of the spectral pattern of the carbinyl hydrogen signal at 6 3.80, the hydroxyl group was placed on a carbon adjacent to the oxygen linked to the carbon atom of the pyran ring, and structure 245 was advanced for murrayazolinol.

4. H.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS PENTA- A N D

315

HEXACYCLIC ALKALOIDS FROM Aspergillus

1 . Tubingensin-A

From the sclerotia of Aspergillus tubingenesis was isolated tubingensinA (246, C,,H,,NO (M+ 401.2698), mp 95-98"C, [aID13.6" (CHCI,)} (80). The UV spectrum suggested acarbazole chromophore (A, 218,239,262, 302, 326, 341 nm with E 14,900, 18,200, 6930, 6780, 480, 480), and 'HNMR data indicated the presence of an isolated four-spin system, showing that there was no substitution in ring A . The signals for H-1 (6 7.92) and H-4 (6 7.11) showed that C-2 and C-3 were substituted, which was also supported through insensitive nuclei enhanced by polarization transfer (INEPT) correlation data. The attachment of C-10 to an sp2 center was inferred from the downfield shifts of two of the C-10 protons (6 2.99,2.88, br dd). The six-carbon chain attached to C-13 was evident from the 'HNMR data and was also supported from the mass spectral peak at mlz 318 (M+ - 83, base peak). From the detailed INEPT studies, NMR correlation studies, and biogenetic considerations, structure 246 was proposed.

-4 ..

H

H

The stereochemistry of tubingensin-A (246)follows from NOE spectroscopy (NOESY) correlation studies. The equatorial disposition of H-14 is in accord with a trans-diaxial coupling between H-14 and the neighboring proton. The gauche relationship between C-18 and C-19 is inferred from correlation studies, and the cis orientation of H-17 and C-1 1 is consistent with the cross-peak correlating the axial proton at C- 10 with a signal centred at 6 1.74 (H-17). The methyl groups have also been considered to be cis, and the substituents at C-12 and C-17 and their relative disposi-

3 16

D. P. CHAKRABORTY

tions with respect to C-1 1 and C-14 were deduced from additional correlations.

2 . Tubingensin-B Tubingensin-B (247, C,,H,,NO (Mi 401.2733), mp 152- 154°C [ a ]+ 6.7" (CHCI,)} was isolated along with tubingensin-A (81). It displayed U V data (A, 218,237, 260,299, 325,338 nm with E 17,200,25,500, 10,100, 10,100, 2200, 6700) suggestive of a carbazole derivative. The base peak in the mass spectrum at mlz 218 (M+ - 183) represented the tetracyclic ring with a 9H carbazole system arising from loss of a C,2H2,0unit from the molecule. IH-NMR data showed that ring A was unsubstituted, and the broad singlets for H-4 (6 8.05) and H-1 (6 7.32) showed that the C-2 and C-3 positions were substituted. The spin systems were also established by homonuclear decoupling and 'H-IH COSY experiments. Selective INEPT and HETCOR experiments were used for precise assignments of I3C-NMR signals. From these data, especially those obtained from selective INEPT experiments and from various other correlations, structure 247 was assigned to tubingensin-B. From the similarity of the W-NMR data of nominine (248) and tubingensin-A (246), together with the results of NOESY experiments, the relative stereochemistry of tubingensin-B was deduced. In consideration of the spatial requirements of the rings, the isopropyl group at C-10 was considered to be cis to C-14. 3 . Aflavazole

Aflavazole [249, C,,H,,NO, (M+ 417.2694), mp 156-16o"C], isolated 219, 243, from the sclerotia of Aspergillus j a m s (821, showed UV (A,, 263, 297, 327, 341 nm, with E 15,300, 17,300, 7500, 7600, 1300, 1600) and IR data for a carbazole derivative with hydroxyl groups. In consideration of its molecular formula, the degrees of unsaturation, the nature of the oxygen functions, and the presence of 12 aromatic carbazole carbons, as evidenced from the I3C-NMR spectrum, it was suggested that aflavazole is a hexacyclic base.

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

317

The evidence for a 2,3,4-trisubstituted carbazole skeleton came from 'HNMR decoupling experiments and COSY data, together with the results of INEPT experiments. By analogy with the physical data of the aflavanine skeleton (250),the skeletal system for rings D and E and the methyl at C-2 were assigned. The direct linkage of C-13 with the aromatic ring was deduced from the correlation data, and the downfield shift of H-13 (8 4.18 ppm), was attributed to the deshielding effect of the aromatic ring. The aromatic methyl signal was correlated with three aromatic carbon signals (8 126.60, 135.13, 110.25 for C-3, C-2, C-l), confirming its position meta to C-4 and ortho to C-1. From IH-NMR and 13C-NMR data and INEPT and NOESY correlations, structure 249 was assigned to alfavazole. The relative stereochemistry was assigned by analogy with the stereochemistry of aflavanine (250), which has also been supported by NOESY data. I. HEXA-A N D OCTACYCLIC INDOLOCARBAZOLES

Staurosporine (251),the first member of the novel bioactive indolocarbazole* alkaloids, was isolated by Omura e f a/. in 1977 (83,84) from a member of the genus Streptomyces. In 1980 arcyriaflavins (252,253) containing the indolocarbazole skeleton were isolated from Arcyria deni4data (85). They were considered primarily indolic pigments. Biosynthetic work (86) on the rebeccamycin (e.g., 254) (86-88) and staurosporine (251)(89) led to the understanding that these alkaloids, like the carbazomycins of Streptouerticilfium ehimense (13), originate from tryptophan, though the nitrogen of the phthalimide unit does not come from tryptophan. The indolocarbazole skeleton present in these alkaloids has been classified with carbazoles (21b);hence, this group of bioactive indolocarbazoles is included here.

I . Arcyriajlauin-B and -C Arcyriaflavin-B (252, C,,H 13N303,mp 35OOC) and arcyriaflavin-C (253, C,,H,,N,O,) were isolated along with the indolic pigments arcyriarubin B (255)and arcyriarubin C (256)from the fruiting bodies of the slime mold Arcyria denudata (85). The structures of the indolocarbazoles 252 and 253 were based on structural studies of the associated indolic pigments 255 and 256. The maleimide/phthalimide moiety in the compounds was detected from the IR bands at around 1750 and 1705 cm-I, and the I3C* In consideration of the Chemical Abstracts conventions for the tricyclic carbazole system, the numbering of carbon atoms in the aglycone fragments of the indolocarbazoles has been modified from that reported by the respective authors.

318

D. P. CHAKRABORTY O+N

H

(252) R = H ( 2 5 3 ) R = OH

R

HO

NMR signals at 6 175.3 and 129.2 of the indolic pigments acryriarubin-B (255) and -C (256) supported such assignments. The absence of the 2,2' signals present in the 'H-NMR spectra of 255 and 256 from those of acryriaflavin-B and -C showed that the linkage of the indolic fragments in 252 and 253 was through the 2- and 2'- positions in the indolocarbazole. The UV spectra of arcyriaflavin-B and -C were similar to those of staurosporine, showing the presence of the indolocarbazole chromophore. The signals of H-5 and H-13 (6 8.93) in the 'H-NMR spectra of the compounds showed that the maleimide fragment was fused to the carbazole skeleton at the 3,4 positions. These structures have been confirmed by transforma-

H ~ S O ~ ( 2 5 2 ) ( R = H)

( 2 5 3 ) ( R = OH) H (255) R - H ;

H (256) R = O H

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

319

tion of arcyriarubin-B and -C to arcyriaflavin-B and -C, respectively, with H,SO,. Synthesis of arcryiaflavin-B (252) was accomplished by Hughes and Raphael as follows (90). The phosphonium bromide 257 was converted to 258 with nitrosocinnamaldehyde to yield a mixture of stereoisomeric dienes, which was converted to pure E,E-diene by room temperature treatment with iodine in toluene. Heating with neat maleimide gave a Diels-Alder adduct (259), which, on DDQ treatment, afforded substituted terphenyl 260. Deoxygenation with triphenyl phosphonium in refluxing collidine for 40 hr yielded the double nitrene insertion product 261. On demethylation with pyridine hydrochloride arcyriaflavin-B (252) was obtained.

2. Protein Kinase Inhibitor K-252c

Alkaloid K-252c [262, C,oH,3N30(M+ 31 1.1079), mp >300°Cl, a protein kinase inhibitor, was isolated from Nocardiopsis sp. K-290 (91,92). The

320

D. P. CHAKRABORTY

UV spectrum [A, 230, 238, 246, 257, 287, 320, 331, 341, 358 nm, with E 37,000, 34,000, 28,000 (sh), 29,000 (sh), 86,000, 16,000, 20,000, 16,000, 1 1001showed the presence of an indolocarbazole chromophore like staurosporine. The 'H-NMR spectrum displayed the NH protons at 6 1 1.46 and 1 I .38, a secondary amide (6 8.49), and signals similar to those K-252a (K252a and K-252b are discussed below), which showed that the indole nitrogen atoms were unsubstituted. K-252c was also obtained from K252a by drastic hydrolysis, showing it to be the aglycone of K-252a. Consequently, structure 262 was assigned to K-252c.

(262) R = H (263 ) R =

HO

OH

3. Protein Kinase Inhibitor K-252d Alkaloid K-252d (263, C,,H,,N,O,, mp 240-245°C (dec.), [a],, + 35" (CH,OH)} (91,92)displayed physical properties similar to those of K-252c and K-252a. It produced an 'H-NMR signal for one NH (6 11.68) and a secondary amide signal (6 8.54), showing that, unlike K-252c, one of the NH nitrogens was substituted, suggesting the incorporation of a sugar moiety at the carbazole nitrogen. From the high-resolution electron-impact (EI) mass spectrum, the composition of the sugar moiety was found to be C&,404, and on hydrolysis of K-252d rhamnose was isolated. From detailed considerations of the 'H- and ' T - N M R data of the compound, the structure of K-252d (263) was found to be 9-N-(a-~-rhamnopyranose)K-252c, where the rhamnopyranosyl moiety has a 'C4 configuration. 4 . Rebeccamycin

+

Rebeccamycin (254, C,,H,,N,O, (M+ 1, 570, mp 326-330°C (dec.), [aID 131" (THF)}, obtained from (93)Nocardia aerocofonigenes(87,88),

+

4.

CHEMISTRY A N D BIOLOGY O F CARBAZOLE A L K A L O I D S

321

showed the presence of hydroxyl and cyclic imide functions (v,,, 3418, 3355, 1752 cm-I). The IV data showed maxima at 238 and 3 14 nm, besides shoulders at 256,293,362, 390 nm. The IH-NMR data showed signals for an amide N H (6 11.37) and an indole N H (6 10.30), in addition to signals for aromatic protons and sugar fragments. In rebeccamycin the aromatic protons H-5 and H-13 are both deshielded owing to the anisotropic deshielding effect of the phthalimide function, unlike staurosporine where the lactam function deshields only the ortho H-5 proton. Rebeccamycin U

& \

CI

N

N H

(266)

c1

322

D . P. CHAKRABORTY

afforded a tetraacetate which showed a UV spectrum similar to that of staurosporine. In consideration of the 'H-NMR signals of the glycosidic protons and the proton signals of the aglycone moiety, a 4-0-methyl glycosidic linkage with the aglycone was indicated. The chemical shift of H-I' in both rebeccamycin and its acetate showed that the sugar residue was either a C- or N-glycoside. The structure of rebeccamycin was deduced from X-ray crystallographic studies and its absolute configuration from synthesis (87).7-Chloroindole (264),on treatment at room temperature with MeMgI and I-benzyloxymethyl-2,3-dibromomaleimide(265) in benzene containing small amount of hexamethylphosphoramide (HMPA), furnished the desired 2 : I adduct 266 and a 1 : I by-product. The 2 : 1 adduct was photocyclized to 267 in the presence of iodine. In a one-pot method, cyclization and glycosidation were effected by reacting 264 and 265 in the presence of Ag,O in benzene, probably via thermal cyclization to 267 of the intermediate triene system 266. Subsequent glycosidation with 1-bromo-2,3,6-tri-0-acetyl-4-O-methylglucose (268) in refluxing benzene gave 269. The benzyloxymethyl group was removed by hydrogenation and the acetyl groups by ammonolysis, by which rebeccamycin (254) was obtained in 95% yield. Because the sugar moiety was prepared from D-glucose, the absolute configuration was determined to be the same as D-glucose. A shorter route to the synthesis of the aglycone and thus to a rebeccamycin was found. 7,7-Dichlorobisindigo (270) on Wolff-Kishner reduction and acetylation furnished (271) monoacetyl bisindole which, on heating with N-benzoyloxymethylmaleimide in a sealed tube at IO5"C for 8 days afforded the aglycone. The reaction probably involves an initial Diels-Alder reaction followed by loss of acetic acid and subsequent dehydration.

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

323

5 . Antitumor Alkaloids AT 2433-Al, AT 2433-A2, AT 2433-B1, and AT 2433-B2 Antitumor compounds AT 2433-A1 [272), C3,H,,N4O9C1 (M+ 678)], AT 2433-A2 [273, C33H3,N40&1 (M+ 664)], AT 2433-B1[274, C34H36NdO9 (M' 64411, and AT 2433-B2 [275, C,,H3,N40, (Mi 630)] were obtained as yellow solids from Actinomaduru melliaura sp. nov. (SCC 1655) (91). Because AT 2433-A1 was obtained in large quantity, structural determinations of compounds 272-275 were based primarily on the reaction profile and degradation sequence of AT 2433-A1 (94).

(272 1 (273) (274) (275)

AT AT AT AT

2433-A1 2133-A2 2L33-81 2L33-82

R = CI 3 R'= H3C R = C I , R'=H R = H , R'=CH3 R = H 9 R'=H

AT 2433-A1 showed a U V spectrum (A,, 200, 235, 283, 316, 395 nm, with log E 30,781, 39,934, 34,307, 45,562, 3865) similar to that of rebeccamycin, suggesting the presence of an indolocarbazole chromophore. The IR spectrum showed the presence of hydroxyl and cyclic imide functions (v,,, 3120, 3365, 1750, 1962 cm-I). Unlike rebeccamycin, the imide NH signal (6 3.25) in the 'H-NMR spectrum of AT 2431-A1 was replaced by an N-CH, signal, indicating methyl substitution at the imide NH. Like rebeccamycin, both H-5 and H-13 were anisotropically deshielded (6 9.27 and 9.18) owing to the imide carbonyl functionalities. The elemental composition of the compound was determined from the high-resolution flamedetection (FD) mass spectrum, in which isotopic clusters for molecular ions indicated the presence of one chlorine atom. Important information was also obtained from the EI mass spectrum of the volatile tetracetate. The presence of a 4-methoxyglucopyranoside fragment was ascertained from the 'H-and 13C-NMRdata of A T 2433-A). The 'H-NMR sjgnals for

324

D.

P. CHAKRABORTY

the C-4' rnethoxy (6 3.68) and the C-I' anomeric rnethine protons (6 6.91) as well as the 13C absorptions of C-I' or C-5' and 4'-OCH, were very close to those of rebeccarnycin. The downfield shift of the C-6' absorption to 6 66.00 pprn as compared with rebeccarnycin suggested further substitution. The downfield shift of the anomeric proton of AT 2433-A1 as cornpared with the dechloro congeners showed the proximity of H-I' to an electronegative chlorine. Further, from the "C-NMR signal of the anomeric carbon, a N-C glycosidic bond was located at N-9. The triplet arising from the 6'-OH proton in rebeccamycin was absent in AT 2433A l , showing the linkage of the aminosugar at that position. Studies on the methanolic hydrolysis products of the carbazole provided information on the structure of the arninosugar in AT 2433-A1. The 'HNMR spectrum at I 10°C* of the 3,4-di-4-brornobenzoate of methyl 2,4dideoxy-4-N-rnethylaminopyranoside provided information regarding the a configuration at C-1" and an equatorial substitution of the pyranoside at C-3". The CD spectrum of the derivative showed that the chilarity of 3,4-dibromobenzoate is 2 - ~ - t h r e o . The CD spectrum of AT 2433-A1 was superirnposable with that of rebeccamycin, showing identical chilarity for the two compounds and thereby confirming the absolute structure of AT 2433-A I (272). Structures of AT 2433-A2 (273), AT 2433-Bl (274), and AT 2433-B2 (275) have been deduced from the 'H-NMR spectra and other physical data. In the case of AT 2433-A2, the aglycone part was identical with that of AT 2433-A I . In the case of AT-2433-B1 (274) and AT-2433-B2 (275), the aglycone parts were found to be dechlorinated congeners, resulting in shifts in the proton and carbon spectra of the respective compounds. The arninosugar fragments of 272 and 274 were identical, whereas those of 273 and 275 were N-demethylated. The chilarity of the antitumor compounds is identical to that of rebeccamycin. 6. Stuurosporinr

Staurosporine (251, CZRH26N403 (M+ 466), rnp 270°C (dec.), [ a ] + 35.0" (MeOH)} was isolated from Streptomyces stuurosporeus Anaya, Takahashi, and Ornura sp. nov. (83).The U V (h,,,243, 263 (sh), 292, 322 (sh), 335, 356, 372 nrn] and IR (vmax 3200, 3500 crn-I) spectra showed the presence of hydroxyl and arnine groups, and an aromatic system. The structure and stereochemistry of the isolate were established by X-ray crystallographic analysis of the methanol solvate (84).Complete and unam-

* Owing to a rotational barrier about the amide bond, a highly complex spectrum is observed below 1 I O T .

4. CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

325

biguous assignments of the IH- and 13C-NMRspectra of 251 have been accomplished (942). There have been several attempts to synthesize staurosporine aglycone by various methods, as detailed below. The staurosporine aglycone was prepared from tryptamine and p-indoloacetic acid in two consecutive cyclization reactions (95). The starting amide 276 was prepared from tryptamine and the appropriate acid chloride. DDQ oxidation of the amide in a mixture of water and THF yielded the diketo compounds 277, which on selective reduction with NaBH, gave rise to the hydroxyketone 278. On acetylation in the presence of 4-dimethylaminopyridine (DMAP), the cyclized pentaacetate 279 was obtained. Reduction of 279 with TiCI, in aqueous acetone afforded the lactam 280, which on deacetylation with NaHCO, in aqueous methanol and subsequent irradiation furnished the aglycone of staurosporine. In another method (96),dibromomaleimide was N-benzylated to afford 280A, which on treatment with indolylmagnesium bromide gave 280B. Oxidation with p-toluene sulfonic acid/DDQ provided a skeleton related to the staurosporine alkaloids (280C). Magnus and Sear (97) reported the synthesis of the aglycone of staurosporinone using indole-2,3-quinodimethanemethodology. Tryptamine was converted to its phthalimide derivative 281, and the indole nitrogen was protected by treatment with p-methoxybenzene sulfonyl chloride with NaH and DMF to give the sulfonyl derivative 282. Subsequent formylation with 2,2-dichloromethyl methyl ether-titanium tetrachloride at 35°C gave 283, which, on condensation with 2-aminostyrene, afforded the imine 284. The 2,3-quinodimethane intermediate generated in the usual way yielded the pentacyclic carbazole 285, along with other reaction products.Dehydrogenation of 285 (DDQ-toluene reflux) gave the indolocarbazole 286. The phthalimide protecting group was selectively removed by treatment of 286 with hydrazine hydrate in THF at 20°C to afford 287. Treatment

NaBHL redn.

&Cm I

\

-

1

\

I

1

N

N H

N

N Ac

i-,s’Bn H

Ac

H (278)

(279) I

Ticl3

H

& \

Ac N

H

NaHC03

Ac

% +*

(280)

/

\

‘h

D D8, 7

Ph H/A TsO H

4. CHEMISTRY

A N D BIOLOGY OF CARBAZOLE ALKALOIDS

mR,

327

f NPht

R

(284)

of 287 with phosgene in dichloromethane followed by TiCl, (0°C) gave the hexacyclic carbazole 288. The p-MeOC,H,SO, group was selectively removed using LiNH,-THF to yield 289. With KOH/glyme, 289 gave 290, thereby offering the potential to attach a carbohydrate substrate regiospecifically.

= H

In the course of the synthesis of arcyriaflavin-B, Hughes and Raphael (90) also reported an approach to the synthesis of staurosporine starting with 1,4-(dinitrophenyl)butadiene and N-benzylmaleimide. The same series of reactions as used in the case of arcyriaflavin provided the

328

D. P. CHAKRABORTY

(2911

(292)

N-benzylmaleimide derivative 291, which on Clemenson reduction gave the lactam 292, which might be employed as an intermediate in the synthesis of staurosporine. 7. Tan-1030A

Tan-l030A (293, C2,H2,N4O4 [(M + H)+ 4671, mp 290-295°C (dec.)}, obtained from Streptomyces sp. C-71799, was isolated by Tanida et al. (98). The UV (A,, 233, 244, 263 (sh), 275 (sh), 289, 359 (sh), 333, 352, 369 nm, with E 29,400,28,000,31,300,4200,71,000,13,400,17,700, 12,100, 13,4001 and IR spectra (v,,, 3430, 1680 cm-') showed that it was an indolocarbazole derivative with NH, OH, and amide functions, like staurosporine. IH-NMR and I3C-NMR data were close to those of staurosporine and supported the presence of an amide group, nineteen s p 2 carbons, one quaternary carbon, two methines, two methylenes, one methyl, and one

(295)

R = NHCOCHj9R1 = R 2 = H

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

329

methoxy. On hydrogenation, Tan- 1030A gave an amine (294, C,,H2,N403 ( M + 452), [aID +90.4" (CHCI,)}, which was acetylated to the N-acetyl ( M + 474)]. The complete structure of this derivative 295 [C,,H,,N,O, bioactive substance was finally ascertained by NMR (99). 'H-'H COSY provided information as to the partial structures, which were further explained and clarified using IH-13C COSY and correlation spectroscopy via long-range couplings (COLOC). For COLOC experiments, the optimization was obtained using refocused INEPT experiments. From the results of COLOC experiments, further expansion of the partial structure was obtained that substantiated the proposal that it had a y-lactam, a 1,2-disubstituted benzene, and another 1,2-disubstituted benzene fragment with two quaternary carbons. Tan- 1030A (293) had a fragment containing methoxymethine carbons as NMR spectroscopy showed the methoxy carbon (6 58.29) coupled with a methine (H-4', 6 4.73) and vice versa. From the long-range coupling data, the presence of an oxime function was detected. The oxime carbon was correlated with the methine protons H-4' and H-l', which showed that the CHCH, group and the methoxymethine group were connected through the oxime group. The data were in agreement with the fact that the CHCH, group was part of a six-membered ring moiety carrying the oxime function. The downfield shifts of the methine (C-l', 6 82.17) and quaternary carbons (C-5', 6 98.16) show that the two carbons are bonded to heteroatoms (N, N, 0). From the data obtained in COLOC experiments with J = 12.5 Hz, it was shown how ten of the thirteen quaternary carbons were connected. Two of the remaining three carbons (6 114.98, 114.02, and 128.03) were observed in the refocused INEPT experiments when the delay was set at 60 msec. From the long-range coupling experiments, two quaternary carbons (C-4a, 6 114.98, and C-3, 6 114.02) were found to be coupled to H-5 and H-13, respectively. From these data, the two quaternary carbons C-14a and C-9a were considered to be bonded to two nitrogen atoms. For the placement of the lactam function, the two disubstituted benzenoid fragments, and the six-membered cyclic fragments carrying the oxime function, NOE experiments were helpful. On irradiation of the methylene protons, enhancement of the amide NH proton (H-16) and H-13 were observed. In addition, irradiation of the methyl (H-6', 6 2.47) and the methine (H-l', 6 7.04) groups led to enhancement of H-10 and H-8 signals, which supported a structure in which the lactam function, the disubstituted benzene ring, the hexacyclic sugar fragment, and another aromatic ring were connected in order. On irradiation of the oxime proton (6 10.45), enhancement of the signals for the methoxy group (6 3.3) and the methyl proton (H-6', 6 2.47) were observed, from which the stereochemistry of

330

D. P. CHAKRABORTY

the oxime group was ascertained. These data led to the formulation of Tan-1030A as 293. 8. Tan-999

Tan-999 (296, C,,H,,N,O, (M+ 496), mp 221°C (dec.), [aID+42" (DMF)} was isolated along with Tan-1030A from Nocardiopsis dassonuillei (98,99). Like Tan-1030A and staurosporine, it showed IR and UV data suggesting the presence of an indolocarbazole chromophore with a lactam functionality. The 'H-NMR spectrum of Tan-999 was similar to that of staurosporine except for the presence of an additional methoxy group, resulting in the presence of a 1,2,4-trisubstituted benzene ring instead of a 1,2disubstituted benzene ring. From the results of 'H-lH COSY and 'H-I3C COSY, refocused INEPT and COLOC experiments were carried out to determine the position of the methoxy substitutent. In the COLOC spectrum (J = 4.2 Hz), a quaternary carbon was correlated with the methoxy protons (6 3.95) and the aromatic proton H-13. A NOESY experiment showed a cross-peak between the methoxy protons and two aromatic protons (H-12 and H-10). NOE enhancement was also observed between the aromatic proton H-10 and the methyl protons H-6'. These data showed that the aromatic methoxy group was located at C-l 1. The NOESY spectrum also showed the correlation of H-14 and H-13 with the signals of H-l'and H-8. From these data, structure 296 was advanced for Tan-999 (99). 9. Protein Kinase C Inhibitors K-252a and K-2526

Structures of the hexacyclic protein kinase C inhibitors K-252c (262) and K-252d (263) are discussed above. The octacyclic alkaloids K-252a and K-252b are treated here (92,100). Alkaloid K-252a (297, C,,H21N205(M+ 467), mp 262-273"C, [a]-23" (CHCI,)} was isolated from Nocardiopsis sp. K-252 and K-290. The UV and IR spectra of the compound showed it to be an indolocarbazole with NH, OH, ester, and amide functions. 'H-NMR and I3C-NMRdata further showed that it possessed one secondary amide group, one methoxycarbony1 group, one tertiary hydroxyl, eighteen sp2 carbons, one quaternary carbon, one methine, two methylenes and one methyl. The presence of the following structural moieties was inferred from different decoupling experiments: two 1,2-disubstituted benzenes, a y-lactam, and a sugar moiety. The y-lactam contained two quaternary carbons (6 119.5, 132.9). In addition, it had two other quaternary carbons (6 114.6, 115.8). One quaternary carbon (6 123.9) was connected to the sugar moiety through a heteroatom. From NOE experiments of the methylene protons, it was apparent that the methylene is at the C-15 position, confirming the pres-

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

331

( 2 9 7 ) R = CH3 (298) R = H

ence of a staurosporine-type aglycone. The 2-deoxyfuranoside structure of the sugar moiety was derived from COLOC experiments. Further observations of long-range coupling and NOE experiments showed that C-I' is bonded to N-9 and C-4' to N-14. From these data, the structure of protein kinase inhibitor K-252a was advanced as 297. The 3' configuration and the structure of 297 was confirmed by X-ray crystallographic studies. The structure is identical with the antibiotic SF-2370 (101). Alkaloid K-252b (298, C,,H,,N,O, (M+ 453), mp 262-266°C (dec.), [aID +97" (DMF)} was shown to have similar UV and 1R spectra. The 'HNMR and I3C-NMR data of K-252b were also similar to those of K-252a, except for the absence of the methyl group resonance of the carbomethoxy group. From this evidence it was concluded that K-252a is the methyl ester of K-252b. This was confirmed by hydrolysis of K-252a to give K252b and the methylation of K-252b with diazomethane to yield K-252a. Hence, K252b could be represented by structure 298. 10. UCN-01

UCN-O1(298A), a protein kinase inhibitor, was isolated from Streptomyces sp. (IOla).The IR and UV spectral data of 298A showed it to contain

an indolocarbazole chromophore, like staurosporine, and its 13C-NMR spectrum showed the presence of one amide group, eighteen sp2 carbons, eight aromatic protons, one quaternary carbon, four methines, one methoxy group, one N-methyl group, and one methyl group. From the 'HNMR spectrum the presence of a hydroxyl group at C-14 was detected. Consideration of the 'H-NMR and l3C-NMR data led to structure 298A being assigned to UCN-01 .

J. SYNTHESIS OF HEXACYCLIC BASES In the course of the synthesis of analogs of mahanimbine, Pate1 reported (72) the formation of hexacyclic bases 301 and 302. Photocitral-A (303),

332

D. P. CHAKRABORTY Citral condensation*

R2

OH ( 2 9 9 ) R1 = H 3 R 2 = CH3

obtained by irradiation of citral in sunlight or by reflux in pyridine, reacted with 2-hydroxy-3-methylcarbazole (4) to yield bicyclomahanimbine (304) (13).

H (L1

(303)

1

I

Pyridine reflux

% 3

H

(30L)

K. BISCARBAZOLE ALKALOIDS Biscarbazole alkaloids are enumerated here according to the constituent fragments, beginning with an alkaloid derived from a tricyclic precursor and then progressing to biscarbazoles formed from two tricyclic fragments, one tricyclic and one tetracyclic fragment, two tetracyclic bases, and finally dimers made from tetra- and pentacyclic fragments. 1. Indole Dimer

Kumar et al. (102) isolated from the roots of Murraya gleni an indole dimer [305, C,,H,,N,O, (M+ 414)l. It showed U V (A, 222, 271, 284 nm) and IR (v,,, 3400, 1730 cm-') data for an indole with an ester function. The mass spectral peaks at mlz 231 and 184 showed it to be an indole dimer comprising C,,H,,N02 and C,,H,,N units. The 'H-NMR signals of 305 at 6 7.98 and 6.83 (1H each) showed that one of the indole N H and one H-2 of an indole moiety were substituted. From the 'H-NMR and

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

333

'H-IH COSY data, the presence of a carbomethoxy group (6 3.33) and two sets of CH2CH,CCH3 groups, one carrying a carbomethoxy group and the other a -NHCHCH cyclized to the indole skeleton of the nonoxygenated fragment, was ascertained. From the coupling of the CHCH group, the stereochemistry of the substituent on the tetrahydrocarbazole was considered to be trans diequatorial. Consequently, structure 305 was derived, which was also substantiated by I3C-NMR and other physical data.

2. Bismurrayafoline-A Bismurrayafoline-A [306, C2,H2,N202 (M+ 4201, mp 176- 177OC1, obtained fromMurraya euchrestfolia (15),had a UV spectrum [A, 228,244, 253 (sh), 284 (sh), 293 nm; log ~4.81,4.95,4.84,4.15,4.80]characteristic of carbazole derivative. The IH-NMR spectrum showed signals for H-5 and H-5' protons (6 7.72, 7.85, d , J = 7 Hz each), H-4 and H-4' protons (6 7.32, 7.36, br s), a complex pattern of eight aromatic protons, an aryl methyl (6 2.46, s), two aromatic methoxy groups (6 3.71 and 3.82), and a benzylic methylene at 6 5.83. The high-intensity peak at mlz 210 (M2+) suggested the isolate to be a dimeric compound. The signal for a twoproton singlet at 6 5.83 and the mass spectral peak at m/z 210 suggested that the benzylic methylene was bonded to the nitrogen atom. On treatment with sodium in liquid ammonia, 306 afforded 307 (mlz 210 and 181), showing the loss of a methoxy group. An N-methyl derivative (307A), obtained on methylation of 307, showed a high-intensity mass spectral

bCH3

(306)

334

D. P. CHAKRABORTY

peak at mlz 224 instead of at mlz 210 and 181, supporting structure 307 for the liquid ammonia reduction product. On hydrogenolysis, 306 afforded murrayafoline-A. Thus, bismurrayafoline-A was formulated as 306.

3. Chrestifoline-D Chrestifoline-D [307B, C,,H,,N,O, (Mi 434)] was isolated as a colorless oil from M. euchrestifolia (21c). From physical data and partial synthesis from bismurrayafoline-A by DDQ oxidation, compound 307B was considered to be the 3-formyl analog of bismurrayafoline-A (307). 4 . Bisrnurrayafoline-B

Bismurrayafoline-B [308, C,,H,,N,O, (M + 588; M2+294)] was obtained from M. euchrestifolia (15). From IR (v,,, 3550, 3150, 1615 cm-') and UV [A, 225 (sh), 265 (sh), 285 (sh), 312, 333 nm, bathochromic shift on addition of alkali] data, it appeared to be a phenolic carbazole. The nineteen carbon signals in the I3C-NMR spectrum, the molecular ion peak at rnlz 588, and the molecular di-ion peak at rnlz 294 showed it to be a dimer comprised of two symmetrical monomer units. The 'H-NMR spectrum of 308 was similar to that of murrayafoline-B except that it lacked the singlet at H-2. From NOE experiments, in which enhancements of the proton signals at H-4 and H-4' were observed on irradiation of the aryl methyl

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

335

protons, the linkage of the monomers was assigned at the 2,2' position. From these data, structure 308 was assigned to bismurrayafoline-B.

5 . Bismurrayafolinol Bismurrayafolinol [309, C,,H,,N,O, (M 436.1784), colorless oil], obtained from M. euchrestifolia (30), showed UV (A,, 225, 244, 252, 281, 292, 329, 340 nm) and IR spectra suggesting it to be a carbazole alkaloid with a hydroxyl function. From the base peak in the mass spectrum at mlz 210 (Mz+), it was considered to be a dimer. From the 'H-NMR spectrum, the presence of two aromatic methoxy groups, two orthocoupled H-5and H-5'protons (6 7.92 and 8.07, d , J = 8 Hz each), a singlet for H-4 (6 7.74), H-2 and H-2' protons (6 6.78, 7.06), two benzylic methylenes, one attached to the nitrogen (6 6.01) and the other to oxygen (6 4.86), and additional signals for seven protons (6 7.10-7.50) was determined. On comparison of the data with those for bismurrayafoline-A, structure 309 was advanced for bismurrayafolinol, which has been confirmed by the synthesis of its acetate as follows. Murrayanine (2), on reduction with sodium borohydride, gave 3-hydroxymethyl-1-methoxycarbazole, which, on treatment with acetic anhydride, furnished bismurrayafolinol acetate (310). +

(309) R-H ( 3 1 0 ) R-OC.CH3

6 . Oxydimurrayafoline

Oxydimurrayafoline [311, C,,H,,N,03 (M 436.1809)l was obtained as a colorless oil from M. euchrestifolia (30). The UV spectrum (A,, 226, 242, 253, 260, 280, 291, 324, 331 nm) and the base peak at mlz 21 1 in the mass spectrum suggested the presence of a dimeric carbazole skeleton comprising of two murrayafoline A units. 'H-NMR data showed the presence of two NH protons (6 8.27), two aromatic methoxy groups (6 4.01), and two benzylic oxymethylene groups. From NOE experiments, it was found that there was enhancement of the H-2 signal on irradiation of the aromatic methoxy group (6 4.61) and enhancement of the H-4 and H-2 +

336

D. P. CHAKRABORTY

signals on irradiation of the benzylic methylene. Thus, oxydimurrayafoline was formulated as 311.

7. Murrafoline-F Murrafoline F [3U,C2,H,,N202 (M+ 420.181)] was obtained as a colorless oil from M . euchrestifolia (20). It had a characteristic U V spectrum for a carbazole skeleton, and 'H-NMR data showed the presence of two rnethoxy groups (6 4.14, 3.93) and an aromatic methyl (6 2.38). The isolate showed two four-spin proton signals and one three-spin proton resonance, suggesting the presence of two unsubstituted ring A units and a trisubstituted ring C. The signal at 6 4.43 (2H) and a carbon signal at 6 32.2 in the 'H- and 13C-NMR spectra, respectively, could be attributed to a rnethylene group. Enhancement of the H-4 signal on irradiation of a methyl signal and the enhancement of H-4 and H-4' caused by irradiation of the methylene signal showed that H-4, H-4', and H-2' were unsubstituted. 'H-'H COSY data showed the long-range coupling of the proton signal at 6 7.74 with the aryl methyl. Lack of enhancement of any proton signals following irradiation of either methoxy group indicated that one of the rnethoxy groups was at the 1 position and the other was bonded to nitrogen. The location of the second methoxy group bonded to nitrogen was also supported by the intense mass spectral fragment at mlz 390 and the presence of 12 sp2 carbons as doublets in the I3C-NMR spectrum. Thus, murrafoline-F was formulated as structure 312.

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

337

8 . Murrastifoline-A and -B

Murrastifoline-A [313, C2,H,,N20, (M 420)] and murrastifoline-B [314, C2,H2,N202(M+ 406)], two biscarbazoles comprised of two tricyclic units, were obtained from M . euchrestifolia (103),along with murrastifoline-D and -E, which have tricyclic and tetracyclic units. All of these alkaloids have U V spectra characteristic of a carbazole chromophore. ‘H-’H COSY spectra of the compounds showed that all had common signals for 0methoxy (6 3.57-4.11) and aromatic methyl groups (6 2.52-2.56). Two singlets arising from H-4 (6 7.56-7.63) and another signal for the H-2 proton (6 6.85) were observed in the ’H-NMR spectrum of 313 and 314. The signals for a four-spin proton system indicated the absence of substitution in ring A of one of the constituent carbazole fragments of the bisalkaloids. The position of the methoxy group at C-1 and the methyl group at C-3 of this unit was ascertained from NOE studies. This was further corroborated by the mass spectral fragment at r n l z 210 or 21 1, which could be assigned to the murrayafoline-A structural unit (3), common to all the murrastifoline bisalkaloids. +

’”3-J-y \

In murrastifoline-A, besides the ‘H-NMR signals for the common structural units, signals for a C-methyl (6 2.48), an aromatic methoxy (6 4.13), and H-4 (6 7.54), H-l’, and H-2 protons were observed. It also showed

338

D. P. CHAKRABORTY

signals for H-5 (6 8.10, J = 1.2 Hz), H-6 (6 7.40, dd, J = 1.2, 7.9 Hz), and H-8 protons (6 7.67, d , J = 7.9 Hz). Because H-5 was meta coupled, the linkage of the second unit was at C-6-C-6'. From this evidence, the structure 313 was assigned to murrastifoline-A. In murrastifoline-B (3141, the 'H-NMR data for the second unit showed the presence of an additional four-spin system, demonstrating the absence of substitution in ring A'. The signals for H-4 (6 7.61) and H-2 (6 7.04) were meta coupled (J = 1.5 Hz). Irradiation of the 0-methoxy signal at 6 4.03 showed enhancement of the H-2' signal (6 7.04). Thus, the two units were linked through C-3' of the second unit, and the structure was assigned as 314. 9. Chrestifoline-A

Chrestifoline-A [315, C,,H,,N,O, (M 420)], along with chrestifoline-B and -C (uide infra), was obtained from M. euchrestifolia as an oil (103). It showed a UV spectrum characteristic for a carbazole chromophore. The chrestifolines are built on a common l-methoxy-3-(substituted methy1ene)carbazole unit, as revealed by 'H-NMR spectroscopy. On irradiation of the benzylic methylene, enhancement of the signals of H-4 and H-2 were observed, and the mass spectral fragment at mlz 210 also supports the structure. From the mass spectral data of chrestifoline-A (M+ 420), it was inferred that it had two C,, units. Besides the 'H-NMR signals for the common unit, it had one aromatic methoxy group (6 4.02), an aryl methyl group (6 2.47), an aromatic proton signal at 6 6.94 and a N H proton at 6 10.28. In NOE experiments, irradiation of the aromatic methoxy caused enhancement of the H-2' signal, whereas irradiation of the methyl signal (6 2.07) caused enhancement of the H-2' signal and the benzylic methylene signal. Irradiation of the benzylic methylene signal caused enhancement of the aromatic methyl and the H-5' signal. From these data, structure 315 has been proposed for chrestifoline A. +

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

339

10. Bismurrayafoline-C and -D Bismurrayafoline-C (316, C,&,N,O,), obtained as a pale yellow oil from M . euchrestifolia (28),showed a UV spectrum characteristic for I oxygenated carbazoles [A, 226,235,260 (sh), 285 (sh), 31 1,335 nm (sh)]. The 'H-NMR signals were similar to those of bismurrayafoline-B (308), but the signal for a dimethylallyl group 308 was replaced by a geranyl chain [6 1.42, 1.47, 1.60 (6Heach) 1.67 (4H, m), 1.75 (4H, m), 3.42 (4H, m), 6.16 (t, 7 . 9 , 4.85 (t, 7.9)] and that for a methoxy by a hydroxyl. The proposed structure was supported by the results of NOE experiments. Irradiation of the aryl methyl signals at 6 2.48 showed enhancement of H-4 and H-4', and irradiation of the benzylic methylene of the side chain showed enhancement of the ally1 methyl (6 1.42) and the N-H. Dimethyl ether derivative 317 showed enhancement of the doublet at 6 6.85 (H-6 and H-6'), confirming the location of the oxygenated substituents at C7 and C-7'. Irradiation of the second methoxy group at 6 3.4 caused enhancement of the N H proton signal at 6 7.85 and the aromatic methyl signal on the other unit, suggesting the location of the second methoxy at C-I. On irradiation of the aromatic methyl at 6 2.52, area increases of the singlets at 6 7.85 (H-4 and H-4') and in the methoxy signal of the other carbazole unit (6 3.4) were observed. These data led to the formulation of bismurrayafoline-C as 316. Bismurrayafoline-D [317, C,,H,,N,O, (M' 724), mp 198-20O"Cl was isolated as a colorless prism (28) along with bismurrayafoline-C. The UV, IR, and 'H-NMR data were very close to those of bismurrayafoline-C. The 'H-NMR spectrum showed the presence of an additional methoxy group (6 3.89). The geranyl side chain was readily discernible from the mass spectral fragmentation at mlz 293 (M*+ - C,H,) and at mlz 239 (M2+ - C,H,). On treatment with diazomethane, the isolate gave an 0tetramethyl ether of bismurrayafoline-C (318). NOE experiments on bismurrayafoline-D (317), after irradiation of the methoxy group (6 3.8),

340

D. P. CHAKRABORTY

indicated an enhancement of the signals of H-6 and H-6', demonstrating the methoxy groups to be at C-7 and C-7' and the hydroxyls at C-1 and C- I '. Thus, bismurrayafoline-D was formulated as 317, the dimethyl ether of bismurrayafoline-C (316). 1 I . Murrafoline-B Murrafoline-B [319, C,,H,,N,O, (M + 474.32), mp 234-237"C], obtained from M. euchrestifolia (104), was considered to be a dimeric carbazole alkaloid from the UV spectrum (A,, 208, 226, 240, 292, 304, 330 nm) and its molecular di-ion peak (M2+ 237). From the 'H-NMR spectrum the presence of one aryl methoxy group (6 3.87), two aryl methyl groups (6 2 4 8 and 2.49), and a dimethyldihydro pyran unit (6 2.30, 2.38,4.69, 1.46, 1.56) was evident. From decoupling experiments, it was inferred that the carbazole had one unsubstituted ring A and another ring A with substitution at C-8. From NOE experiments enhancement of the signals at H-2', H-4', and H-4 were observed on irradiation of the aromatic methyl group, showing that these positions were unsubstituted, whereas irradiation of the methoxy group (6 3.87) led to enhancement of the signal at H-2', indicating the methoxy to be at C-1'. These results, and the mass spectral peaks at m/z 263 and 21 1, indicate that the base is built on a dihydrogirinimbine skeleton and a murrayafoline-A skeleton. Thus, the structure of murrafoline-B was advanced as 319 and confirmed by synthesis, namely, treatment of a mixture of murrayafoline-A and girinimbine with Nafion 117 in refluxing aqueous methanol.

(319 1

(320)

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

341

12. Murrafoline-D Murrafoline-D (320),isolated from M . euchrestifolia (104, was identified from IR, 'H-NMR, and mass spectral data. It was found to be have TLC behavior identical to a compound obtained during the Nafion 117catalyzed reaction of murrayafoline-A with girinimbine (104). 13. Murrafoline-E

Murrafoline-E [321,C,,H,,N,O, (M+ 420.183 I)]. obtained as a colorless oil (20), showed UV [A,, 228 (sh), 238, 255 (sh), 263 (sh), 287, 320, 340, 352 nm] and IR spectral data for a carbazole derivative. From the mass spectral fragment at mlz 262 and 21 I , it was considered to be a biscarbazole alkaloid with girinimbine and murrayafoline-A units. From the 'H-NMR spectrum, the presence of an aryl methyl (6 2.51). an aryl methoxy (6 3.94), a DMP system [6 1.44 (6H, s), 5.79, 6.92 ( I H , d, J = 10 Hz each)], and a nitrogen-bonded benzylic methylene group (6 6.01) was readily discernible. From 'H-'H COSY experiments, two four-spin systems showing the presence of unsubstituted rings A and A' were ascertained. It appeared from the 'H-'H COSY data and the results of NOE experiments that H-4 and H-2 in the murrayafoline fragment of 321 were unsubstituted. Consequently, the structure of murrafoline-E was assigned as 321. 14. Murrustifoline-D

Murrastifoline-D (322,C,,H,,N,O, (M+ 440.2227), [aID0" (CHCI,)} was obtained from M . euc.'zrestifofiu(103) as a colorless oil. Like its congeners murrastifoline-A and -B, it contained a tricyclic murrayafoline-A unit in which the second tetracyclic unit was bonded through nitrogen. From the mass spectral fragments at mlz 280 (base peak) and at mlz 21 I , the two constituent units were suggested. Oxygen-linked gerninal dimethyl group signals were observed at 6 1.55 and 1.42 along with a double doublet (J = 5.9, 9.2 Hz) at 6 4.38 which was coupled to a hydroxyl proton at 6 4.48 (d, J = 5.9 Hz). The signal at 6 7.2 ( I H , d , J = 9.2 Hz) was as-

dH3 q

\

"

/

(322)

R=OH

(323)

R=H

3

342

D. P. CHAKRABORTY

signed to H-4' as a benzylic carbon directly bonded to the nitrogen of the murrayafoline-A unit. The trans relationship between C-3" and C-4" was ascertained from the coupling constants and the absence of an NOE effect between the methine protons. The 'H-NMR data showed the absence of substitution in rings A and A' of the two units, and the signals at 6 7.87 and 7.56 were attributed to the H-4 and H-4' protons, showing substitution in ring C. From these data, structure 322 was assigned to murrastifoline-D. 15. Murrastifoline-E

Murrastifoline-E (323, C,,H3,,N,02 (M + 474.2280), [aID -5.7" (CHCI,)} was obtained from the same source as its congeners (105). It had a carbazole-like UV spectrum and showed mass spectral fragments at mlz 21 1 and 264 (base peak) arising from the two units. The 'H- and 13C-NMR spectra showed signals for an NH, an aryl methoxy group (6, 4.11, aC 56.89), and two aryl methyl groups (6, 2.56, 2.42, aC 17.56, 22.11). The 'H-NMR data indicated the presence of H-4 and H-4' resonances, and, from proton-proton decoupling experiments, two pairs of four-spin systems were detectable, showing an absence of substitution in rings A and A'. The presence of a dihydropyran (DMPH) ring was detectable from the 'H-NMR and I3C-NMR data. The 'H-NMR signal at 6 7.42 and the appearance of eleven sp2 carbon signals as doublets in the I3C-NMR spectrum showed that the two carbazole units were linked through the benzylic carbon at C-4" and the nitrogen atom of the murrayafoline A unit. From these data, and the results of NOE experiments, structure 323 was assigned to murrastifoline-E, which is evidently deoxygenated murrastifoline-D (322). 16. Chrestifoline-B

Chrestifoline-B [324, C,2H28N202 (M+ 472)] was obtained from M . euchrestifolia (103) as an oil showing a characteristic UV spectrum for carbazoles. The mass spectral fragments at mlz 262 and 248, together with a base peak at mlz 210, suggested the presence of a murrayafolineA unit and a girinimbine unit in the binary system. The presence of a DMP system was readily discernible from the 'H-NMR data [6 1.36 (6H, s), doublets at 6 6.91, 5.53 ( I H , d, J = 9.9 Hz each)]. In a series of NOE experiments, enhancement of the H-4' signal on irradiation of the aryl methyl signal (6 2.27) and enhancements of the signals for H-4" (6 6.91), H-2 (6 6.90), H-8' (6 7.39), and H-4 (6 7.38) were observed on irradiation of the benzylic methylene. These data permitted the formulation of chrestifoline-B as 324.

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

343

17. Murrastifoline-C Murrastifoline-C (325, C,,H,,H,O, (M+ 540.2773), [aID0" (CHCI,)}, obtained from M . euchrestifolia (103)as a colorless oil, showed, like the other murrastifolines, spectral evidence (UV and IH NMR) for the presence of a 1-methoxy-3-methylcarbazolesystem. Mass spectral fragments at mlz 330 (base peak) and 211 suggested a murrayafoline-A unit and a mahanimbine unit in the binary system. The detection of two sets of four-spin systems in 325 along with results of 'H-'H COSY experiments suggested that rings A and A' were unsubstituted. The 'H-NMR data showed characteristic signals for a DMP system in which one of the methyl groups was extended by an isoprene unit. This inference was also supported from the mass spectrum, in which a fragment at mlz 457 (M+ - 83) was observed. IH-'H COSY exchange and NOESY correlation data showed a broad singlet at 6 7.36 (deshielded H-4') and a signal at 6 6.13 for the benzylic methylene linked to the nitrogen of the murrayafoline-A unit. Thus, murrastifoline-C was formulated as 325.

18. Chrestifoline-C

Chrestifoline-C [326, C,,H,,N,O, (M+ 540)], obtained as a colorless oil from M . euchrestifolia (lo.?),showed a characteristic U V spectrum for carbazoles. Like other chrestifolines, the IH-NMR data and the

344

D. P. C H A K R A B O R T Y

mass spectral fragment at rnlz 210 of chrestifoline-C showed it to contain a l-methoxy-3-(substituted methy1ene)carbazole nucleus. In 'H'H COSY experiments it showed two sets of four aromatic protons, like murrastifoline-C. The 'H-NMR data showed the presence of a DMP system in which one of the methyls was extended by an isoprene unit, like that in mahanimbine. The mass spectral fragment at r n l z 457 (M+ - 83) and the peak at rnlz 248, corresponding to a carbazolopyrillium ion arising from a mahanimbine unit, substantiated the above findings. In NOE experiments, irradiation of the aryl methyl signal (6 2.30) caused enhancement of the H-4' signal, whereas irradiation of the benzylic methylene (6 5.80) caused enhancement of the signals at H - 4 (6 6.95), H-8' (6 7.38), H-4 (6 7.381, and H-2 (6 6.90). From these data the structure 326 has been advanced for chrestifoline-C (103). 19. Murrafoline-C

Murrafoline-C [327, C3,H,,N,0, (M+ 526.261 l)] was obtained as a colorless oil from M . euchrestifolia (104),and the IR and U V data suggested the presence of a carbazole skeleton. The 'H-NMR data showed the presence of four oxygen-linked tertiary methyls (6 1.38, 1.41, 1.42, 1 S2). ABX type signals (6 2.27, 2.31, and 4.63) and AB type signals (6 5.56 and 6.07) represented 2',2'-dimethyldihydropyran and 2',2'-dimethylpyran systems in the molecule. The structure for murrafoline-C has been advanced from comparative assessment of the 'H-NMR data of 327 and rnurrafoline-B, which suggested that a girinimbine unit is linked with a dihydrogirinimbine unit through C-9' and C-8. Consequently, structure 327 is consistent for murrafoline-C and is also supported by the mass spectral fragmentation pattern.

CH3

(327)

20. Murran irn bin e

Murranimbine [328, C3&3&0~ (M' 528.2585)], obtained (106) as a pale racemic oil from M . euchrestifolia, displayed UV and IR data characteristic of a carbazole unit. The characteristic mass spectral fragment at mlz 262.13 showed it to be a bis alkaloid. 'H-NMR data indicated the

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

345

presence of two aryl methyls (6 2.30, 2.32), four methyls attached to carbons linked with oxygen (6 0.60, 1.54, 1.44, 1.60), and one D,Oexchangeable NH proton (6 10.55). From the ‘H-’H COSY experiments two sets of four-spin aromatic systems were readily detected, which inTwo deshielded signals at 6 7.65 and 8.00 having a cluded H-5 and H-5’. long-range coupling with the aryl methyls were discernible. The results of NOE experiments after irradiation of the aryl methyls confirmed the presence of H-4 and H-4’. ‘H-’H COSY experiments showed the relationship of the signals at 6 6.12 (H-12’, J = 2.9 Hz), 6 2.74 (H-11, J = 2.9, 5.1 Hz), 6 4.07 (H-12), 6 2.21 (H-11, J = 5.16, 12.8 Hz), and 6 2.08 (HI I ) overlapped by a solvent signal. These data, and the results of COSY experiments, showed that the alkaloid contained two girinimbine units in it. Analysis of the homonuclear broad band decoupling (HMBC) spectrum (J = 8 Hz) of murranimbine and the cross-peaks of three-bond correlation related to the connectivity of two units was undertaken. The proton (H12’, 6, 6.12) of the benzylic methylene attached to a nitrogen was related to another benzylic methine ((2-12, 6, 32.4) on the upper unit and to the oxygenated carbon (C-lo’, SC80.3l)in the lower unit, which is also related to the benzylic methine proton (H-12, 6, 4.07) in the upper unit. The methine proton (H-l I ’ , 6,2.74) coupled with both of the benzylic protons (H-12’, 6, 6.12; H-12, 6, 4.07), had a three-bond relationship with an aromatic carbon at C-1 (6, 103.74), and was related to H-11 (6, 2.21) in the upper unit. The methylene carbon at C-11 (6,37.10) was related to the geminal methyl protons at 6,1.44 and 1.60. The three-bond relationships in the HMBC spectrum and other relationships were taken into consideration in assigning the structure of murranimbine. NOE experiments provided information regarding the stereochemistry of the molecule. Enhancements of the signals for H-ll’, H-12, and the N-H signals were observed on irradiation of H-12’. On the other hand, irradiation of H-l 1’ caused enhancements of the signals of H-12’ and H-12. Enhancements of H-l I’ and the methyl signal at 6 1.44 were noticed on irradiation of H-12. Consequently, murranimbine was formulated as 328.

346

D. P. CHAKRABORTY

21. Bis-7-hydroxygirinimbine-Aand -B

7-Hydroxygirinimbine-A [328A, C3&3+04 (M + 556)] and 7hydroxygirinimbine-B (328B,C,,H,,N,04), isolated as pale yellow oils from M . euchrestifolia (106a), had U V spectra very similar to that of girinimbine. The IH-NMR spectrum of 328A showed an aryl methyl singlet, two quaternary methyl singlets, two pairs of AB-type doublets (J = 8.55, 9.6 Hz), and an aromatic proton singlet. The 'H-NMR data, together with ',C-NMR data, suggested the presence of a 7,8-disubstituted girinimbine as the structural unit. Compound 328A gave an 0-methyl derivative on methylation. In differential NOE experiments, enhancement of the H-4 and H-6 signals was observed, indicating methyl and methoxy groups to be at C-3 and C-7, respectively. The linkage of the two girinimbine units was at C-8, C-8'. This was also supported from the HMBC spectrum of the 0-methyl derivative of 328A. The structure of 328B was also determined from U V , IH-NMR, and I3C-NMR data. The compound gave an 0-methyl derivative on methylation, and NOE experiments on the derivative indicated the linkage of the

HO

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

347

two girinimbine units to be at C-8 and C-6'. The possibility of the coupling of the two monomers during isolation has not been ruled out. 22. Murrafoline Murrafoline (329, C,,H,,N,O, (M+ 594), [aID0" (CHCI,)}, the first bis alkaloid to be reported, was isolated from the root bark of Murraya euchrestifolia (107). The IR and UV data for 329 [A,,, 218, 243, 260 (sh), 307, 332 nm, with log E 4.66,4.80,4.66,4.45, 3.911 revealed it to be a carbazole derivative. The 'H-NMR spectrum showed the presence of two aryl methyl groups (6 2.28, 2.37), a vinyl methyl (6 1.54), three tertiary methyls on a carbon linked to oxygen, a triplet arising from a benzylic group (6 4.58), a multiplet (6 3.24), and overlapping unresolved signals for thirteen protons in the aromatic region [S 6.70-7.90, two of which disappeared on deuteration (NH)]. The molecular di-ion peak at rnlz 297 and the ion peak at rnl z 594 showed it to be a bis alkaloid. The complete structure and relative stereochemistry were obtained from single-crystal X-ray analysis as 329.

L.

S Y N T H E S I S OF

BISCARBAZOLES

Biscarbazole formation in connection with studies of carbazole alkaloids is discussed here. Various other methods of dimerization, including electrochemical methods, have been reviewed by Joule ( 1 0 7 ~ ) . Under Udenfriend reaction conditions, 3-methylcarbazole (6) furnished a dimeric compound (330) in poor yield, along with other hydroxycarbazoles (55). A methanolic solution of murrafoline-A and girinimbine, when

348

D. P. CHAKRABORTY

refluxed in methanol in presence of Nafion 117, yielded a mixture of biscarbazoles, namely, murrafoline D and two other alkaloids (331 and 332) (104). When girinimbine and derivatives 333 and 334 were treated for a short time with boron trifluoride etherate, dimeric carbazoles were obtained (337-339). The reaction has been mechanistically represented as shown (108).

4.

CHEMISTRY A N D BIOLOGY O F CARBAZOLE ALKALOIDS

349

IV. Physical Properties of Carbazole Alkaloids Applications of physical methods for the structural determination of carbazole alkaloids have been previously reviewed (6,13). Here we report some of the salient features and new findings. A. ULTRAVIOLETABSORPTION SPECTRA Extensive application of ultraviolet absorption spectroscopy to carbazole alkaloids has been made in detecting the carbazole and indolocarbazole chromophores in the respective alkaloids. The positions of the formyl, methoxy, and methyl groups continue to be deduced from UV data (16,13,33) and are confirmed subsequently by other data or by synthesis. UV absorption data for a large number of alkaloids have been detailed in previous reviews (6, 13).

B. INFRARED SPECTRA The characteristic IR bands for carbazoles and methylsubstituted carbazoles as reported by Richards (109) were utilized by Chakraborty in the structural determination of degradation products (6). Recently, the IR bands near 750 cm-' have been found to be characteristic for the unsubstituted ring A of carbazomycins. The IR bands at low frequency (1630-1645 cm-I) have been considered characteristic for a formyl group at C-3 chelated to the hydroxyl at C-2 in several alkaloids (13). C. NMR SPECTRA 1. ' H - N M R Spectra The presence of a carbazole skeleton in the first carbazole alkaloid murrayanine was detected by 'H-NMR spectroscopy (6). Discussions on 'H-NMR spectra of the alkaloids have been presented with data for various compounds in previous reviews (6,13). The low-field H-5 and H-4 signals (6 7.17-7.50) and their shift caused by neighboring protons have been extensively utilized for the detection of substitution in rings A or C. The presence of an isolated four-spin system (6 7.06-7.90) has been considered indicative of the presence of an unsubstituted ring A. The characteristic signals for a 3,3-dimethylallyl (DMA) side chain have been utilized in their detection. The olefinic doublets at around 6 6.2 and 5.4 (J = 10 Hz each) and the signal for a gem-dimethyl group have been utilized for the detection

350

D. P. CHAKRABORTY

a 2,2-dimethyl-A3-pyran (DMP) system fused to the carbazole skeleton. In compounds of the mahanimbine type, where one of the methyl signals has been extended by a DMA unit, consequent changes in the spectrum have been observed. NOE experiments have been extensively used for confirming the assignments of proton signals. In the case of some complex compounds the more informative INEPT method has been utilized. 2. 13C-NMR Spectra Diagnostic chemical shifts for C-1 to C-8 were first utilized in the case of carbazomycins-A and -B (10,13). A short discussion on the shift of the carbon signals as compared with those of benzene signals has been presented by Chakraborty and Roy (13). Carbon-1 in 2-oxygenated carbazoles experiences shift effects owing to both oxygen and nitrogen atoms (13). Similar shifts of C-8 have also been found in the case of 7-oxygenated compounds (25). Thus, in the case of several 2- and 7-oxygenated compounds (e.g., 38 and 39) both C-1 and C-8 signals appear at around 6 96.0. Modern two-dimensional NMR techniques like COSY, HETCOR, COLOC, and other techniques have been utilized for the confirmation of structures of complex alkaloids. The structural determination of Tan1030A and Tan-999 are interesting in this respect (98). I3C-NMR data of several carbazole alkaloids have appeared in previous reviews (10,13). D. MASS SPECTRA Mass spectral investigations have been extensively utilized for the structure determination of carbazole aklaloids. Various modern mass spectral methods have been employed in arriving at a correct mass for a molecule. Apart from this, mass spectrometry studies have provided interesting information on various structural aspects. For example, the carbazolopyrillium ion found in mass spectra of pyranocarbazoles like girinimbine and mahanimbine has given adequate information for the presence of this tetracyclic system. When the pyran ring is hydrogenated, a different characteristic spectrum is obtained. Molecular di-ion peaks have also been utilized for detecting some bisalkaloids. Brief discussions on this subject have appeared in previous reviews (6,13). E. X-RAYCRYSTALLOGRAPHY X-Ray crystallographic methods have been utilized in the structural determination of various alkaloids, the first being that of murrayazoline. The structures of murrafoline (329) and staurosporine (251) have primarily been elucidated by X-ray crystallographic methods, whereas for

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

351

carbazomycin-B (1271, -C (105), -G (1101, chlorohyellazole (90), K-252a (2971, and kinamycin-C (209) X-ray methods have confirmed the structures deduced by other methods.

V. Biogenesis of Carbazole Alkaloids Previous reviews have discussed the biogenesis of 3-methylcarbazole, the origin of the functional variants of aromatic methyl group in carbazole alkaloids, the hydroxylation pattern of the compounds, and the biosynthesis of carbazomycins and kinamycins (5,6,10,13). 3-Methylcarbazole (6) has been considered by Kapil ( 5 ) to be the key precursor compound in higher plants. Chakraborty (12) suggested that 2-methylcarbazole (153) could be the key compound in the pathway operating in fungi and bacteria and also suggested that both 3- and 2-methylcarbazoles arise from 2- and 3-prenylated indoles. Since the report on the structure of echinulin in 1943 (110), there had been wider interest in studies on the prenylation of indoles (111,112). The primary electrophilic attack of the prenyl chain occurs at the 3 position of the indole skeleton, which may subsequently migrate to other positions. Kapil (3,as well as Erdtman (112a), conceived that 3-prenylated indole gives rise to 3-methylcarbazole via prenylated 2-indole after rearrangement. Isolation from Murraya gfeni of the dimeric indole 305 by Kumar et a f . (102), which contains a 2-methyltetrahydrocarbazole with another unit of 3-prenylated indole containing a carbomethoxy group, provides the first circumstantial evidence for the formation of 2-methylcarbazole from a 3prenylated indole, although paniculidin-A (340) was isolated from Murraya panicufata in 1985 (113). Thus, evidence supporting the formation of 2methylcarbazole from prenylated indole is strengthened. The occurrence of tubingensin-A, -B, and aflavazole in members of the genus Aspergillus containing polyprenylated indoles also contributes to the idea that 3- or 2-methylcarbazole or their terpenoid analogs arise from prenylated or polyprenylated indoles.

H (340)

352

D.

P.

CHAKRABORTY

Biosynthetic experiments (1 14) on carbazomycin-B have shown that C2 and C-1 with their methyl substituents arise from a pyruvate unit via acetyl-coenzyme A. Thus, in the biosynthesis of carbazomycins both the tryptophan and pyruvate pathways participate. Insight into the formation of indolocarbazoles has been derived from studies on the biosynthesis of staurosporine (251) and rebeccamycin (254). Biosynthetic studies of rebeccamycin (86) have shown that rebeccamycin arises from two molecules of tryptophan, one of glucose (341), and one of methionine (342). Evidence has also been provided that the a-amino group of tryptophan does not contribute to the phthalimide fragment, suggesting that no symmetrical indole intermediate is involved in the biosynthesis of the alkaloid. Studies on the biosynthesis of staurosporine by Meksuriyen and Cordell (89) have shown that the aglycone moiety (343) of staurosporine is derived from two units of tryptophan with the carbon skeleton intact. Further experimental results are necessary to establish the nature of the intermediate in the biotransformation of tryptophan to staurosporine.

VI. Biochemical and Medicinal Properties of Carbazole Alkaloids and Related Compounds Earlier in this treatise, Kapil(5) mentioned some of the biological properties of carbazole alkaloids. Since then, wider interest in this area has developed (6,8,10,13). Some important properties and some new reports are summarized below. A. ANTIMICROBIAL PROPERTIES Antimicrobial properties of carbazole alkaloids received attention beginning with the report of the activity of murrayanine (2) and other alkaloids of M. koenigii (4). The antibiotic properties of carbazomycin-B (127) and

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

353

related compounds attracted wider attention. Carbazomycin-B was active against the fungi Glomerella cingulata N 13 and Elsinoe fawcettii at a minimum inhibitory concentration (MIC) of 3 pg/ml, whereas against Trychophyton asteroides 429 and Trychophyton mentagrophytes 833 it was active at a concentration of 12.5 pg/ml. Demethylated glycozoline (197) was active against Trychophyton sp. at aconcentration of 10 pglml. Phenolic compounds were more active than their O-methyl derivatives (13). Tetracyclic kinamycins have been found to be active against a large number of bacteria and fungi. The activities of the kinamycins have been found to decrease with the larger number of acetoxy groups. Thus, the relative order of antimicrobial properties are kinamycin-B, -D, -A, and -C (217,218, 216, 209). The MIC values of kinamycin-B are 0.012 pg/ml against Bacillus subtilis PCI-219, Staphylococcus aureus, Bacillus anthracis, and Staphylococcus albus, and 0.9 to 0.19 pg/ml against Vihrio coma. Deacetylkinamycin-D derivatives show substantial activity against Mycobacterium ATCC 607 and gram-negative Escherichia coli NIHJ, Klebsiella pneumoniae, and Shigella sonnei (65b). The antimicrobial properties of the indolocarbazoles have received widespread attention. The activity of staurosporine (251)against various bacteria and fungi has been examined (83). It has been reported to be active against Candida albicans, Cundida pseudotropicalis, Saccharomyces sake, Aspergillus brevipes, Trichophyton rubum,Sclerotinia cinerea, and Piricularia oryzae at MIC values of 6.25, 3.13, 3.13, 3.13, 6.25, 0.78, and 0.78 pg/ml, respectively. Alkaloids AT 2433-Al, -A2, -B1, and -B2 were found to be active against gram-positive Micrococcus luteus, Bacillus subtilis, Staphylococcus aureus, Streptococcus jaecalis, and Streptococcus faecium. AT 2433-B 1 was also active against the gramnegative bacterium Escherichia coli SS 1431 (91,100).Alkaloid SF 2370

(216 (217)

R1 = R2= R 3 = COCH3; R b = H R ~ = R J = R L = H ; R ~ = COCH3

2 1 8 R 1 = R3 = C O C H3 ;R2 = RL = H ( 2 0 9 ) R ~ = R ~ = R L = C O C H ~R;2 = H (

354

D. P. CHAKRABORTY

(identical with K-252a) has been examined with several microbes and was found to be active against Micrococcus luteus, Micrococcus frcluus FDA 16, and Corynebacteriurn bouis 1810 at a concentration of 6.25 pg/ml (202). B. ANTITUMOR A N D TUMOR-PROMOTING ACTIVITY Kinamycin C (209) has weak antitumor activity against Ehrlich ascites carcinoma and sarcoma 180 (66). In the brine shrimp assay, 7methoxymukonal (40) was more active than 7-methoxyheptaphylline (@A), whereas against the NT-29 cell lines, 7-methoxyheptaphylline was more active than 7-methoxymukonal (25). Antibiotics AT 2433-A 1 and AT 2433-B 1 (272,274) were found to be effective in prolonging the lifespans of mice transplanted with the leukemia P338 tumor (92,200).Rebeccamycin (254) has been found to have antitumor activity (86a). Growth inhibition of mammary carcinoma by compound 344 has been observed at 1 Fg/ml (225). Staurosporine is cytotoxic against NB- 1 cells; it induces elongation . also shows in uitro of neurites and cell enlargement ( 2 2 5 ~ )Staurosporine antineoplastic activities against cancer cells (2256)and a strong cytotoxic M effect on the growth of HeLa S3 cells, with an IC,, value of 4 x (I I5c).

mcoo c2 H 5

I

I

N-Methylcarbazole (345) is a carcinogen and mutagenic constituent of tobacco smoke. This alkaloid is converted by primary cultures of rat heptatocytes to N-hydroxymethylcarbazole (346)and carbazole (1).The cytotoxicity of N-hydroxymethylcarbazole was greater than that of the

4. CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

355

parent N-methylcarbazole. Hydroxylation of N-methylcarbazole is considered a toxicant step, whereas its dealkylation to carbazole is likely to be a detoxification step (116). 3-Hydroxymethyl-I-methoxycarbazole (koenoline-27) has been shown to be a cytotoxic agent (19). l,lO-Bis(6methyl-5H-benzo[b]carbazol-1 I-y1)decane (346A) has a potential bifunctional DNA intercalating property (117). C. ANTIVIRAL ACTIVITY Several carbazoles have been examined for antiviral activity. In uitro activity against herpes simplex virus type- 1 by tubingensin-A (246) and tubingensin-B (247) were reported. Carprofen (347) increased interferon (IFN) produced by 10-carboxymethyl-9-acridenone (CMA, suboptimal concentration) in murine cell cultures. The CMA-induced IFN production was increased 500-fold in pure bone marrow-derived macrophage cultures. The activity has been shown to be related to cyclooxygenase (118). Some bis-basic ethers of carbazoles are antiviral. When tested against Encephalomyocardis virus infection, several 9-ethyl-substituted bis-basic carbazoles with the general formula 348 have been shown to be active (13).

“‘m /COOH

\

N

“CH3

H

(347 1

R

2

m

R

t.’

R (348) R=H,Me,Et R1 = COOR, C O N M e 2

3 R2 = Et

N H S 0 , G

H

(349)

’

/ Bu / h e x y l

OH

(2L5A)

D. CARDIOVASCULAR-MODULATING ACTIVITY Murrayaquinone A (69) has been found to produce a triphasic inotropic response (119). This triphasic pattern of inotropism was unaffected by reserpine, metoprolol, or cimetidine treatment and is not mediated through a receptor mechanism but, rather, through a mechanism involving ATP

356

D. P. CHAKRABORTY

production. In rabbits, the sudden death produced by arachidonic acid was antagonized by pretreatment with Bay U 3405 (349), showing it to be a thromboxane A2 antagonist with antithrombolic activity. Murrayazolinine (245A) caused sudden lowering of blood pressure in experimental subjects. The action of (245A) is probably not mediated through muscarine H, or H, receptors (120). Staurosporine has been found to have hypotensive activity (83). K-252b and K252c seriously affect the functions of platelets, mast cells, and several other cells and tissues (91).

E. CENTRAL NERVOUS SYSTEM ACTIVITY Behavioral changes in animals have been recorded following administration of some tetrahydrocarbazoles having alkylamino substitution in the aromatic ring (121). Compounds 350 and 352 induce an abnormal behavioral pattern, whereas 351 does not. Some tetrahydrocarbazoles of general formula 353 induce inhibition of gastric secretion (13).

(350) R = C H 2 P h (351) R = H

(352)

(353) R = Me/ E t / C H M q / C H 2 C M e 3 / C M e

R1 = N M e 2 / N E t 2 / p y r i m i d y n y I / 1-piper I d y n y l

Some carbazole derivatives are useful in the treatment of psychotic disorders, anxiety, pain, and gastrointestinal dysfunction (122). Some neuroleptic agents like cycloindole (354) and flucindole (355) have found use in therapy; whereas cycloindole has antidepressant activity, flucindole shows antipsychotic activity (123). Tetrahydrocarbazoles of the general formula 356 are useful central nervous system agents. They have also antiparkinson activity (124).

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

R

R

357

=H

(35L

Cycloindole

(355

Flucindole R = F

-f3zo-zN3R1 \

N

/

H

(356) R = Alkyl, Alkoxy, Alkylthio, S u l f o n y l s u l f one , O H , F , C I , B r , C F 3 , Cyano, Methylene dioxy Z = CH2 /H2-CH2

R1= Ph

Rincazole (357),a novel neuroleptic and antipyretic agent (125,126),has evoked a great interest. The effects induced by phencyclidine, a 6-opioid antagonist, are counteracted by 357. It indirectly affects dopamine neurons, displaying selectivity for A 10 dopamine cells. It increases the concentration of neurotensin in the caudate nucleus. A constituent of bovine urine, 3-chlorocarbazole (UO),has been shown to have diazepam-like activity (48).

F. ANTI-INFLAMMATORY PROPERTIES Anti-inflammatory properties of several carbazole derivatives have attracted widespread attention. 6-Chloro- I ,2,3,4-tetrahydrocarbazolehas been found to be useful against treatment of gout (10). I-Ethyl-8-N-propyl1,2,3,4-tetrahydrocarbazole1-acetic acid (358) is an anti-inflammatory

358

D.

P. CHAKRABORTY

agent (10). Other complex tetrahydrocarbazoles of general structure 359 and 360 have also been found to have anti-inflammatory properties (13). Carprofen (347) has received significant attention as a substitute for nonsteroidal and nonalkaloidal anti-inflammatory substances. Its activity is comparable to indomethacin, with less toxic side effects (production of gastric ulcer and blockade diarrhea) (127). In uitro cellular effects of carprofen were found to be greater than those of ibuprofen and almost comparable to those of hydrocortisone (128).The anti-inflammatory activity of carprofen is probably dependent on inhibition of some neutrophil macrophage function. It stimulates acid secretion without affecting basal acid secretion (129). This enhancement of secretagogue-stimulated acid secretion was dependent extracellular calcium. It has been suggested that the compound acts at postreceptor site between adenylate cyclase and a protein pump. The drug probably increases calcium efflux through the plasma membrane and decreases the endogenous prostaglandin E, content.

G. MODULATION OF ENZYME ACTIVITY, METABOLISM, AND ALLERGIC REACTIONS 3-Chlorocarbazole (120) is a potent inhibitor of rat liver monoamine oxidase (13). The inhibition of lipid peroxidation induced by free radicals generated in the presence of Fe2+and ascorbic acid by carazostatin (112)

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

359

is higher than that with the brain protective agent lunarizine, which has afree radical-scavenging activity. As an antioxidant, it is better than butylated hydroxytoluene. Carazostatin may be helpful for alleviation of tissue damage due to the action of superoxide radicals and subsequent peroxidative disintegration of cell membranes (46).Carbazomycin-B and -C (127, 105) were found to inhibit lipooxygenase activity, which could be related to free radical-scavenging effects (130). Alkaloids K-252a, -b, -c, -d (91), and UCN-01 ( 1 0 1 ~have ) been found to be inhibitors of protein kinase C. The indolocarbazole aglycone moiety has been considered to be required for such inhibitory activity. The protein kinase C inhibitory activity of Tan-999, Tan- 1030A, and staurosporine also support this contention. Unlike K-252a, K-252b appeared to lose the calmodulin selectivity; not only Ca2+/calmodulin-dependent activity but also the basal activity of Ca2+/calmodulin-dependent phosphodiesterase were affected (91). Macrophage-activating activity is associated with the antitumor and antimicrobial activity through augmentation of phagocytic activity. Tan-999, Tan-l030A, and staurosporine were found to be macrophageactivating substances. They augment phagocytic activity of murine macrophage cells. The activity of p-glucouronidase, a lysosomal enzyme, is a marker for macrophages related to bacterial infection. Tan-999 increased the activity of this enzyme at 5 pg/ml. Tan-1030A had substantial activity, but staurosporine had little activity on this enzyme. The phagocytosisdependent respiratory burst in mouse peritoneal macrophages was found to increase with Tan-999 (0.01-1 pg/ml) and Tan-1030A (1-10 pg/ml). It has been suggested that the specific inhibitors of protein kinase-C are expected to act as macrophage activators that could provide protection against tumor development and microbial infection through phagocytic activity (83,98,99). Oxarbazole (361) shows antiallergic activity (123).

H. MISCELLANEOUS EFFECTS Trypanocidal effects have been shown by substituted 1,2,3-tetrahydro carbazoles (131). Some carbazole derivatives have larvacidal and insecticidal activity (132). Inhibitory effects (80,81) on the crop pest Heliofhis zea

360

D. P. CHAKRABORTY

have been shown by tubingensin-A and -B. Carbazole-N-carboxamide (362)has been found to have growth inhibitory activity (133). SF-2370 has been found to have effects against rice plant diseases and the greenhouse blights caused by Rhizocotonia solani, Xanthomonas campestris pv oryzae, and Piricularia oryzae (101).

REFERENCES 1. C. Graebe and C. Glazer, Chem. Eer. 5 , 12 (1872).

D. P. Chakraborty. B. K. Barman, and P. K. Bose, Sci. Cult. 30,445 (1964). D. P. Chakraborty, B. K. Barman, and P. K. Bose, Tetrahedron 21, 681 (1965). K. C. Das, D. P. Chakraborty, and P. K. Bose, Experientia 21, 340 (1965). R. S . Kapil, in “The Alkaloids” (R. H. F. Manske. ed.), Vol. 13, p. 273. Academic Press, New York and London, 1971. 6. D. P. Chakraborty, Forschr. Chem. Org. Nuturst. 34, 299 (1977). 7. D. P. Chakraborty, Planta Med. 39, 97 (1980). 8. D. P. Chakraborty, Trans. Bose R r s . Inst. (Calcutta) 47, 49 (1984). 9. H . Husson in “The Alkaloids” (A. Brossi, ed.), Vol. 26, p. 1. Academic Press, New York and London. 1985. 10. P. Bhattacharyaand D. P. Chakraborty, Fortschr. Chem. Org. Nufurst.52, 159 (1987). 1 1 . I . Mester, in “The Chemistry and Chemical Taxonomyofthe Rutales” (P. G. Waterman and M. F. Grundon, eds.), p. 31. Academic Press, London, 1983. 12. D. P. Chakraborty, J. Indian Chem. S O C . 66, 843 (1989). 13. D. P. Chakraborty and S. Roy, Forfschr. Chem. Org. Nuti~rst.57, 71 (1991). 14. B. K . Choudhury, A. Mustafa, M. Garba, and P. Bhattacharyya. Phytochemistry 26, 2138 (1988). 15. H. Furukawa, T. S. Wu, and T. Ohta, Chem. Pharm. Bull. 33,4202 (1983). 16. P. Bhattacharyya and B. K. Chowdhury, Indian J. Chem. 24B,452 (1985). 17. H . Furukawa, C. Ito, M. Yogo, and T. S. Wu, Chem. Pharm. Bull. 33, 2672 (1985). 18. S . Roy, L. Bhattacharyya. and D. P. Chakraborty, J. Indian Chem. SOC.59, 1369 (1982). 19. N. Fiebig, J . M. Pezzuto, D. Soejarto, and A. D. Kinghorn, Phyfochemistry 24, 3041 (1985). 20. C. Ito, T. Wu, and H. Furukawa, Chem. Pharm. Bull. 16, 2372 (1988). 21a. W. S. Li, J. D. McChesney, and F. G. El-Feraly, Phytochemistry 30, 343 (1991). 21b. R. Livingstone In “Organic Chemistry“ (E. H. Rodd, ed.), p. 486. Elsevier, Arnsterdam, 1973. 21c. C. Ito, N. Okahana. T. S. Wu, M. Wang, J . Lai, C . S. Khon, and H . Furukawa. Chem. Pharm. Bull. 40, 230 (1992). 22. K . Toshiya and S. Masonori, Heterocycles 31, 1605 (1990). 23. P. Bhattacharyya and A. Chakraborty, Phytochumistry 23, 471 (1984). 24. N. Ruangrungsi, J. Ariyaprayoon, G. L. Lang, and H. G. Organ, J. Nut. Prod. 53, 946 (1990). 25. C. Chaichantipyuth, S. Pummangure, K. Naowseran. D. Thanyavuthi, J. Anderson, and J . L . McLeughlin, J. N u t . Prod. 51, 1285 (1988). 2. 3. 4. 5.

4. CHEMISTRY

A N D BIOLOGY OF CARBAZOLE ALKALOIDS

361

26. B. T. Ngadjui, J . P. Ayafor, B. L. Sondengam, and J . D. Connolly, Phvtochemistry 28, 1517 (1989). 27. P. Bhattacharyya and B. K. Chowdhury. J . Nut. Prod. 48, 465 (1985). 28. C. Ito, M. Nakagawa, T. S. Wu, and H. Furukawa. Chem. Phurm. Birll. 39, 2525 (1991). 29. H . Furukawa, T. S. Wu, T. Ohta, and C. S . Kuoh, Chrm. Phurm. Bull. 33,4132 (1985). 30. C. Ito, T. S. Wu, and H. Furukawa, Chem. Pharm. Bull. 15, 450 (1987). 31. P. Bhattacharyya and B. K. Chowdhury, Chem. Ind., Lond. 301 (1984). 32. V. Kumar, J . Reisch, and A. Wickramasinghe, Aust. J . Chem. 42, 1375 (1989). 33. C. Ito, M. Nakagawa, T. S . Wu, and H. Furukawa, Chem. Phurm. Bull. 39, 1668 ( 1991). 34. C. Ito, M. Nakagawa, T. S . Wu, and H . Furukawa, Chem. Pharm. Bull. 39, 2525 (1991). 35. J. Reisch, 0. Goz. A. Wickramsinghe, H. M. T. Baudra Herath, and G . Henkel, Phytochemistry 31, 2877 (1992). 36. C. It0 and H. Furukawa, Chem. Pharm. Bull. 38, 1548 (1990). 37. D. Lontsi, J. P. Ayafor, D. L . Sondengam, J. D. Conolly, and D. S . Rycroft. Trtrahedron Lett. 26, 4249 (1985). 37a. M. Yogo, C. Ito, and H. Furukawa, Chem. Pharm. Bull. 39, 328 (1991). 38. T. Martin and C. J . Moody, J . Chem. Soc., Chem. Commun., 1391. (1985). 39. K. Ramesh and R. S . Kapil, Indian J . Chem. 25B, 462 (1986). 40. J . H. Cardellina 11, M. C. Kirkup, R. E. Moore, J. S. Mynderse. and K . Seff, Tetrahrdron Lett., 4915 (1979). 41. S. Kano, E. Sugino, S. Shibuya, and S . Hibino, J . Org. Chem. 46, 3856 (1981). 42. R. L. Dillman and J. H. Cardellina 11, J . Nut. Prod. 51, 1056 (1991). 43. T. Naid, T. Kitahara, M. Kaneda, and S . Nakarnura, J . Antihior. 40, 157 (1984). 44. S . Kondo, M . Katayama. and S . Marumo, J . Antihiot. 39, 727 (1986). 45. M. Kaneda, T. Naid, T. Kitahara, and S . Nakamura. J . Antihiot. 41, 602 (1988). 46. S . Kato, H. Kawai, T. Kawasaki, Y . Toda, T. Urata, and Y . Hayakawa, J . Antihiot. 42, 1879 (1989). 47. P. M. Jackson, J . Moody, and R. J. Mortimer, J . Chem. S o c . , Perkin Trans. I , 2941 (1991). 48. K. Luk, L. Stern, M. Weigel, R. A . O’Brien, and N. Spirt, J . Nut. Prod. 46, 852 (1983). 49. H. J . Knolker and M. Bauermeister, J . Chem. Soc., Chem. Commun.,1468 (1989). 50. H. J . Knolker and M. Bauermeister, J . Chem. Soc., Chem. Commun., 664 (1990). 51. P. Bhattacharyya, M. Sarkar, A. K. Biswas, and D. P. Chakraborty. J . Indian Chem. Soc. 6, 328 (1979). 52. T. Martin and C. J. Moody, Tetrahedron Lett. 26, 5841 (1985). 53. J . Bergman and R. Carlson, Tetrahedron Lett.. 4051 (1978). 54. J . Bergman and B. Pelcman, Tetrahedron 44, 5215 (1988). 5 5 . S . Roy, R. Guha, S. Ghosh. and D. P. Chakraborty, Indian J . Chem. 21B, 617 (1982). 56. C. J . Moody and P. Shah, J . Chem. Soc., Perkin Trans. I , 2463 (1989). 57. U. Pindur and L . Pfeuffer, Heterocycles 26, 325 (1987). 58. J . Bergman, L . Venemalm, and A. Gogoll, Tetrahedron 46, 6067 (1990). 59. T. Kawasaki, Y. Nonaka, and M. Sakamoto. J . Chem. Soc., Chem. Commun., 43 ( 1989). 60. H. Furukawa, T. S . Wu, and C. Kuoh, Heterocycles 23, 1391 (1985). 61. M. Mukherjee, S . Mukherjee, A. K. Shaw. and S . N. Ganguly. Phytochemistry 22, 2328 (1983).

362

D. P. CHAKRABORTY

62. H. C. Furukawa, C. Ito, M. Yogo, T. S. Wu, and C. Kuoh, Chem. Pharm. Bull. 33, 1320 (1985). 63. M. Yogo, C. Ito, and H . Furukawa, Chem. Pharm. Bull. 39, 328 (1991). 64. C. Ito, T. Matsuya, S. Omura, M. Otani, A. Nakagawa, Y. Iwai, M. Ohtani, and T. Hata, J . Antibiot. 23, 315 (1970). 65a. S. Omura, A. Nakagawa, H. Yamada, T. Hata, A. Furusaki, and T. Watanabe, Chem. Pharm. Bull. 19, 2428 (1971). 65b. S. Omura, A. Nakagawa, H. Yamada, T. Hata, A. Furusaki, and T. Watanabe, Chem. Pharm. Bull. 21, 931 (1973). 66. M. C. Cone, P. J. Seaton, K. A. Halley, and S. J. Gould, J. Antibiot. 42, 179 (1989). 67. P. J. Seaton and S. J. Gould, J. Antibiot. 42, 189 (1989). 68. P. J. O’Sullivan, R. Moreno, and W. S. Murphy, Tetrahedron Lett. 33, 535 (1992). 69. N. Tamayo, A. M. Echavarren. and M. C. Paredes, J. Org. Chem. 56,6488 (1991). 70. W. P. Griffith, S. V. Ley, G. P. Whitcornb, and A. D. White, J . Chem. Soc., Chem. Commun., 1625 (1987). 71. D. P. Chakraborty, S. Roy, and A. K. Dutta, J. Indian Chem. SOC. 64, 215 (1987). 72. B. P. J. Patel, Indian J. Chem. 21B, 612 (1982). 73. B. P. J. Patel, Synth. Commun. 11, 823 (1981). 74. B. P. J . Patel, Indian J. Chem. 21B, 20 (1982). 75. V. Kane, A. R. Martin, and J. A. Peters, Heterocycles 16, 1445 (1981). 76. N. S. Narashirnhan and S. L. Kelkar, Indian J . Chem. 14B, 430 (1976). 77. M. Banderanayake, M. J. Begeley, B. 0. Brown, D. G. Clarke, L. Crombie. and D. A. Whiting, J. Chem. SOC. Parkin Trans. I , 998 (1974). 78. A. Chakrabarti and D. P. Chakraborty, Tetrahedron Lett. 29, 6625 (1988). 79. L. Bhattacharyya, S. Roy, S. Chatterjee, and D. P. Chakraborty, J . Indian Chem. Soc. 66, 140 (1989). 80. M. R. Te Paske, J. B. Gloer, D. T. Wicklow, and P. F. Dowd, J. Org. Chem. 54,4743 (1989). 81. M. R. Te Paske, J. B. Gloer, D. T. Wicklow, and P. F. Dowd, Tetrahedron Lett. 30, 5965 (1989). 82. M. R. Te Paske, J. B. Gloer, D. T. Wicklow, and P. F. Dowd, J. Org. Chem. 55,5299 ( 1990). 83. S. Omura, Y. Iwai, A. Hirano, A. Nakagawa, J. Awaya, H. Tsuchiya, Y. Takahasi, and R. Masuma, J. Antibiot. 30, 275 (1977). 84. A. Furusaki, N. Hashibe, T. Matsumoto, A. Hirano, Y. Iwai, and S. Omura, J. Chem. Soc., Chem. Commun., 800 (1978). 85. W. Steglich, R. Steffan, L. Kopanski, and G. Echardt, Angew. Chem. Int. End. Engl. 19, 459 (1980). 86. C. J. Pearce, T. W. Doyle, S. Forenza, K. S. Lem, and D. R. Schroeder, J . Nut. Prod. 51, 937 (1988). 86a. J. A. Bush, B. H. Long, J. J. Catino, and W. T. Bradner. J . Antibiot. 40,668 (1987). 87. D. E. Nettleton, T. W. Doyle, and B. Krishnan, Tetrahedron Lett. 26, 401 1 (1985). 88. T. Kaneko. H. Wong, K. T . Okamoto. and J. Clardy, Tetrahedron Letr. 26, 4015 ( 1985). 89. D. Meksuriyen and G. A. Cordell, J. Nut. Prod. 51, 893 (1988). 90. I. Hughes and R. A. Raphael, Tetrahedron Lett. 24, 1441 (1983). 91. S. Nakanishi, Y. Matsuda, K . Iwahasi, and H. Kase, J . Anribiot. 39, 1066 (1986). 92. T. Yasuzawa, T. Iida, M. Yoshida, N. Nirayama, M. Takashi, K. Shirahata, and H. Sano, J. Antibiot. 39, 1072 (1986).

4.

CHEMISTRY A N D BIOLOGY OF CARBAZOLE ALKALOIDS

363

93. J. A. Matson, C. Claridge, J. A. Bush, J. Titus, W. T. Bradner, T. W. Doyle, A. C. Horan, and M. Patel, J. Antibiot. 42, 154 (1989). 94. J. Golik, T. W. Doyle, B. Krishnan, G. Dubay, and J. A. Matson, J . Antibiot. 42, 178 (1989). 94a. D. Meksuriyen and G. A. Cordell, J. Nut. Prod. 51, 884 (1988). 95. B. Sarstedt and E. Winterfeldt, Heferocycles 20, 469 (1983). 96. S. M. Weintraub, R. S. Garigipati, and J. A. Goiuor, Heterocycles 21, 309 (1984). 97. P. D. Magnus and N. L. Sear, Tetrahedron 40, 2795 (1984). 98. S. Tanida, M. Takizawa, T. Takahasi, S. Tsubotani, and S. Harda, J . Antibior. 42, 1619 (1989). 99. S. Tsubotani, S. Tanida, and S. Harda, Tefrahedron 47, 3536 (1991). 100. H. Kase, K. Iwahasi, and Y. Matsuda, J. Antibior. 39, 1059 (1986). 101. M. Sezaki, T. Sasaki, T. Nakazawa, U. Takeda, M. Iwata, T. Watanabe, M. Koyama, F. Kai, T. Shomura, and N. Kojima, J. Antibiot. 38, 1437 (1985). 101a. A. I. Takahasi, E. Kobayashi, K. Asano, M. Yoshida, and H. Nakano, J . Antibiof. 40, 1782 (1987). 102. V. Kumar, D. B. M. Wickramartne, and U. Jacobson, Terrahedron Lett. 31, 5217 (1990). 103. C. Ito, T. S. Wu, and H. Furukawa, Chem. Pharm. Bull. 38, 1143 (1990). 104. H. Furukawa, T. S. Wu, and C. Kuoh, Chem. Pharm. Bull. 33, 2611 (1985). 105. C. Ito and H. Furukawa, Chem. Pharm. Bull. 38, 1548 (1990). 106. C. Ito and H. Furukawa, Chem. Pharm. Bull. 39, 1355 (1991). 106a. T. S. Wu, M. Wang, J. Lai, C. Ito, and H. Furukawa, Phytochemisrry 30, 1052 (1991). 107. A. T . McPhail, T. W. Wu, T. Ohta, and H. Furukawa, Tetrahedron Lett., 5377 (1983). 107a. J. Joule, “Advances in Heterocyclic Chemistry” (A. R. Katritzky, ed.), Vol. 35, p. 83. Academic Press, New York, 1984. 108. A. Chakrabarti and D. P. Chakraborty, Tetrahedron 45, 7007 (1989). 109. R. E. Richards, J. Chem. Soc., 978 (1947). 110. T. Hino and M. Nakagawa, in “The Alkaloids” (A. Brossi, ed.), Vol. 34, p. I . Academic Press, San Diego, 1988. I 1 I. G. Casnati, A. Francion, A. Gnareschi, and A. Pochim, Tetrahedron Lett. 2485 (1969). 112. A. H. Jackson and A. E. Smith, Tefrahedron 21, 989 (1965). 112a. H. Erdtman in “Perspectives in Phytochemistry” ( J . B. Harborne and T. Swain, eds.), p. 107. Academic Press, New York, 1969. 113. J. Kinoshita, S. Tataya, and U. Sankawa, Chem. Pharm. Bull. 33, 1770 (1985). 114. M. Kaneda, T . Kitahra, K. Yamasaki, and S. Nakamura. J. Anribiot. 43, 1623 (1990). 115. L. M. Rice and K. B. Scott, J. Med. Chem. 13, 308 (1970). 115a. H. Morioka, M. Ishihara, H . Shibai, and T. Suzuki, Agric. B i d . Chem. 49, 1959 (1985). 115b. H. Morioka, H. Shibai, Y. Yokogawa, M. Ishihara, T. Kida, and T. Suzuki. J p n . Kokai Tokkyo K o h o , J P 60, 185, 1719; Chem. Abstr. 104, 18649~(1986). 115c. T. Tamaki, H. Nomoto, I. Takahasi, Y. Kato, M. Morimoto, and T. Tomita, Biochem. Biophys. Res. Commun. 135, 397 (1986). 116. W. Yang, T. Jiang, P. J. Davis, and D. Acosta. Toxicology 68, 217 (1991). 117. U . Pindar and H. Erfanian-Ardoust, Chem. Rev. 89, 1681 (1989). 118. E. Storch, H. Kirchner, K. Hueller, M. G. Maritonitti, and D. Gernsa, Chem. Abstr. 109, 3531 1 (1980). 119. K. Takeya, M. Itoigawa. and H. Furukawa, Eur. J. Pharmacol. 169, 137 (1989). 120. A. R. Mitra, Ph.D. Thesis, Calcutta University (1974).

364

D. P. CHAKRABORTY

121. W. A. Sexton, in “Chemical Constitution and Biological Activity,” 3rd Ed. p. 306. F. N. Spoon, Ltd.. London, 1973. 122. Glaxo Group, Eur. Pat. Appl. 219 193/1956: Chem. Abstr. 107, 176032d (1987). 123. D. Lednicer and L. A. Mitscher, in “The Organic Chemistry of Drug Synthesis.” Vol. 3, p. 168. Wiley, New York, 1984. 124. H. H. Huasberg and H. Bettcher, Ger. Pat. DE 330094; Chem. Abstr. 101, 210979f ( 1984). 125. R. M. Ferris, H. L. White, F. L. M. Tang, A. Russel, and M. Harfenist, Drug. Deu. Res. 9, 171 (1987); and previous references [Chem. Abstr. 106, 27732 (1987)l. 126. B. Levant, G. Bissette, F. Widerloev, and C. B. Nerneroff, Regul. Pept. 32, 193 (1991); Chem. Abstr. 114, 1359554 (1991). 127. M. T. Maski, S. Yushiro, S. Tsutomu, and N. Keiju, Nippon Yakurigaku Zasshi 73, 757 (1977); Chem. Abstr. 88, 69038c (1978). 128. A. Tursi, M. P. Loria, G. Specchia, and D. Cassaccima, Eur. J. Rheumatol. InJlammation 5 , 488 (1982); Chem. Abstr. 98, 46621e (1983). 129. R. A. Levin. J. Nandi, and R. L. King, Gastroenterology 101, 765 (1991). 130. D. J. Hook, J. J. Yacobucci, S. O’Connor. M. Lee, Ed. Kers, B. Krishnai, J. Matson, and G. Hesler, J. Antibiof. 43, 1347 (1990). 131. J. C. Pecca and S. M. Albonico. J. Med. Chem. 13, 327 (1970). 132. B. P. Das, I n t . Pest Control 31, 144 (1989). 133. T. Karmakar, M. Mukherjee. and D. P. Chakraborty, Curr. Sci. 55, 828 (1986).

CUMULATIVE INDEX OF TITLES

Aconitum alkaloids, 4, 275 (19S4), 7,473 (1960), 34,95 (1988) C I 9diterpenes, 12, 2 (1970) Cz0 diterpenes, 12, 136 (1970) Acridine alkaloids, 2, 353 (1952) Acridone alkaloids, experimental antitumor activity of acronycine, 21, 1 ( 1983) N-Acyliminium ions as intermediates in alkaloid synthesis, 32, 27 1 ( 1988) Ajmaline-Sarpagine aklaloids, 8,789 (1965), 11,41 (1968) Alkaloid production, plant biotechnology of 40, 1 (1991) Alkaloid structures spectral methods, study, 24, 287 (1985) unknown structure, 5, 301 (1955), 7,509 (1960), 10,545 (1967), 12, 4.55 (1970), 13,397 (1971), 14, 507 (1973), 15,263 (1975), 16, 511 ( 1977) X-ray diffraction, 22, 5 1 (1983) Alkaloids forensic chemistry of, 32, 1 (1988) histochemistry of, 39, I (1990) in the plant, 1, 15 (1950), 6, 1 (1960) Alkaloids from Amphibians, 21, 139 (1983), 43, 185 (1993) Ants and insects, 31, 193 (1987) Chinese Traditional Medicinal Plants, 32, 241 (1988) Mammals, 21, 329 (1983),43, I19 (1993) Marine organisms, 24,25 (1989, 41,41 (1992) Mushrooms, 40, 189 (1991) Plants of Thailand, 41, 1 (1992) Allelochemical properties o r the raison d'Ctre of alkaloids, 43, 1 (1993) A110 congeners, and tropolonic Colchicum alkaloids, 41, 125 (1992) Alstonia alkaloids, 8, 159 (1965), 12,207 (1970),14, 157 (1973) Amaryllidaceae alkaloids, 2,33 1 (1952), 6, 289 (1 9601, 11, 307 ( 1968), 15,83 (1975), 30, 251 (1987) Amphibian alkaloids, 21, 139 (1983). 43, 185 (1983) Analgesic alkaloids, 5, 1 ( 1 9 s ) 36s

366

CUMULATIVE INDEX OF TITLES

Anesthetics, local, 5, 21 1 (1955) Anthranilic acid derived alkaloids, 17, 105 (1979), 32, 341 (1988), 39, 63 ( 1990) Antifungal alkaloids, 42, 117 (1992) Antimalarial alkaloids, 5, 141 (1955) Antitumor alkaloids, 25, 1 (1985) Apocynaceae alkaloids, steroids, 9, 305 (1967) Aporphine alkaloids, 4, 119 (1954), 9, I (1967), 24, 153 (1985) Arisrolochia alkaloids, 31, 29 (1987) Arisrotelia alkaloids, 24, 113 (1985) Aspergillus alkaloids, 29, 185 (1986) Aspidosperma alkaloids, 8, 336 (1965), 11, 205 (1968), 17, 199 (1979) Azafluoranthene alkaloids, 23, 301 (1984) Bases simple, 3, 313 (1953),8, 1 (1965) simple indole, 10, 491 (1967) simple isoquinoline, 4, 7 (1954), 21, 255 (1983) Benzodiazepine alkaloids, 39, 63 (1990) Benzophenanthridine alkaloids, 26, 185 (1985) Benzylisoquinoline alkaloids, 4, 29 (I954), 10,402 (1967) Betalains, 39, I (1990) Biosynthesis, isoquinoline alkaloids, 4, I (1954) Bisbenzylisoquinoline alkaloids, 4, 199 (1954), 7,429 (19601, 9, 133 (1967), 13, 303 (1971), 16, 249 (1977), 30, 1 (1987) synthesis, 16, 319 (1977) Bisindole alkaloids, 20, 1 (1981) Bisindole alkaloids of Catharanthus, C-20’ position as a functional hot spot in, 37, 133 (1990) isolation, structure elucidation and biosynthesis, 37, 1 (1990) medicinal chemistry of, 37, 145 (1990) pharmacology of, 37,205 (1990) synthesis of, 37,77 (1990) therapeutic use of, 37,229 (1990) Buxus alkaloids, steroids, 9, 305 (l967), 14, 1 (1973), 32,79 (1988) Cactus alkaloids, 4, 23 (1954) Calabar bean alkaloids, 8, 27 (1965),10,383 (1967), 13,213 (1971), 36, 225 (1989) Calabash curare alkaloids, 8,515 (1965), 11, 189 (1968) Calycanthaceae alkaloids, 8,581 (1965) Camptothecine, 21, 101 (1983) Cancentrine alkaloids, 14,407 (1973)

CUMULATIVE INDEX OF TITLES

367

Cannabis satiua alkaloids, 34, 77 (1989) Canthin-6-one alkaloids, 36, 135 (1989) Capsicum alkaloids, 23, 227 (1984) Carbazole alkaloids, 13,273 (1971), 26, 1 (1985) chemistry and biology of, 44,257 (1993) Carboline alkaloids, 8,47 (1969, 26, I (1985) P-Carboline congeners and Ipecac alkaloids, 22, 1 (1983) Cardioactive alkaloids, 5,79 (1955) Celastraceae alkaloids, 16, 2 15 (1977) Cephalotaxus alkaloids, 23, 157 (1984) Cevane group of Veratrum alkaloids, 41, 177 (1992) Chemotaxonorny of Papaveraceae and Fumaridaceae, 29, 1 (1986) Chinese medicinal plants, alkaloids from, 32, 241 (1988) Chromone alkaloids, 31, 67 (1987) Cinchona alkaloids, 3, 1 (1953), 14, 181 (1973), 34, 332 (1989) Colchicine, 2, 261 (1952), 6, 247 (1960), 11,407 (1968), 23, 1 (1984) Colchicum alkaloids and allo congeners, 41, 125 (1992) Configuration and conformation, elucidation by X-ray diffraction, 22, 5 1 (1983) Corynantheine, yohimbine, and related alkaloids, 27, 131 (1986) Cularine alkaloids, 4, 249 (1954), 10, 463 (1967),29, 287 (1986) Curare-like effects, 5, 259 (1955) Cyclic Tautomers of Tryptamine and Tryptophan, 34, 1 (1989) Cyclopeptide alkaloids, 15, 165 (1975) Daphniphyllum alkaloids, 15,41 (1975),29, 265 (1986) Delphinium alkaloids, 4,275 (1954), 7,473 (1960) Clo-diterpenes, 12,2 (1970) Czo-diterpenes, 12, 136 (1970) Dibenzazonine alkaloids, 35, 177 (1989) Dibenzopyrrocoline alkaloids, 31, 101 ( 1987) Diplorrhyncus alkaloids, 8, 336 (1965) Diterpenoid alkaloids Aconitum, 7,473 (1960), l 2 , 2 (1970), 12, 136 (1970),34,95 (1989) Delphinium, 7,473 (1960), 12, 2 (1970), 12, 136 (1970) Garrya, 7,473 (1960), 12,2 (1960), 12, 136 (1970) chemistry, 18,99 (1981), 42, 151 (1992) general introduction, 12,xv (1970) structure, 17, 1 (1970) synthesis, 17, 1 (1979) Eburnamine-vincamine alkaloids, 8, 250 (1965), 11, 125 (1968), 20, 297 (1981), 42, l(1992)

368

CUMULATIVE INDEX OF TITLES

Efaeocarpus alkaloids, 6, 325 (1960) Ellipticine and related alkaloids, 39, 239 (1990) Enamide cyclizations in alkaloid synthesis, 22, 189 (1983) Enzymatic transformation of alkaloids, microbial and in uito, 18, 323 (1981) Ephedra alkaloids, 3, 339 (1953) Ergot alkaloids, 8,726 (1965),15, 1 (I975),39,329 (1990) Erythrina alkaloids, 2,499 (1952), 7, 201 (1960), 9,483 (1967), 18, 1 (1981) Erythrophfeurn alkaloids, 4, 265 (1954), 10, 287 (1967) Eupomutia alkaloids, 24, I (1985)

Forensic chemistry, alkaloids, 12, 5 I4 (1970) by chromatographic methods, 32, 1 (1988) Gafbulirnirna alkaloids, 9, 529 (1967), 13, 227 (1971) Gardneria alkaloids, 36, 1 (1989) Garrya alkaloids, 7,473 (l960), 12, 2 (1970), 12, 136 (1970) Geissosperrnum alkaloids, 8, 679 (1965) Gelsemiurn alkaloids, 8,93 (1963, 33, 84 (1988) Glycosides, monoterpene alkaloids, 17,545 (1979) Guatteria alkaloids, 35, 1 (1989) Haplophyton cimicidurn alkaloids, 8, 673 (1965) Hasubanan alkaloids, 16,393 (1977), 33, 307 (1988) Histochemistry of alkaloids, 39, 165 (1990) Hofarrhenagroup, steroid alkaloids, 7, 3 19 (1960) Hunteria alkaloids, 8, 250 (1965) lhoga alkaloids, 8,203 (1965),11,79 (1968) Imidazole alkaloids, 3, 201 (1953), 22, 281 (1983) Indole alkaloids, 2, 369 (1952), 7, 1 (1960), 26, I (1985) distribution in plants, 11, I (1968) simple, 10, 491 (1967), 26, 1 (1985) Reissert synthesis of, 31, 1 (1987) Indolizidine alkaloids, 28, 183 (1986). 44, 189 (1993) 2,2'-Indolylquinuclidine alkaloids, chemistry, 8, 238 (196% 11, 73 ( 1968) Ipecac alkaloids, 3, 363 (1953), 7,419 (1960), 13, 189 (1971), 22, 1 (1983) Isolation of alkaloids, 1, I (1950) Isoquinoline alkaloids, 7, 423 (1960) biosynthesis, 4, 1 (1954) "C-NMR spectra, 18,217 (1981) simple isoquinoline alkaloids, 4 , 7 (1954), 21, 255 (1983) Reissert synthesis of, 31, 1 (1987)

CUMULATIVE INDEX OF TITLES

369

Isoquinolinequinones, from Actinomycetes and sponges, 21, 55 (1983) Khat (Catha edulis) alkaloids, 39, 139 (1990) Kopsia alkaloids, 8, 336 (1965) Lead tetraacetate oxidation in alkaloid synthesis, 36, 70 (1989) Local anesthetics, 5, 21 I (1955) Localization in the plant, 1, 15 (1950), 6, 1 (1960) Lupine alkaloids, 3, 119 (1953), 7, 253 (1960), 9, 175 (1967), 31, 16 ( 1987) Lycopodium alkaloids, 5,265 (1955),7,505 (1960), 10, 306 (l967), 14, 347 (1973), 26,241 (1985) Lythraceae alkaloids, 18, 263 (1981), 35, 155 (1989) Mammalian alkaloids, 21, 329 (l983), 43, 119 (1993) Marine alkaloids, 24, 25 (1983, 41,41 (1992) Maytansinoids, 23,71 (1984) Melanins, 36, 254 (1989) Melodinus alkaloids, 11, 205 (1968) Mesembrine alkaloids, 9,467 (1967) Metabolic transformation of alkaloids, 27, 323 (1986) Microbial and in uifro enzymatic transformation of alkaloids, 18, 323 (1981) Mifrugyna alkaloids, 8, 59 (1965), 10, 521 (1967), 14, 123 (1973) Monoterpene alkaloids, 16, 431 (1977) glycosides, 17, 545 (1979) Morphine alkaloids, 2, 1 (part 1, 1952), 2, 161 (part 2, 1952), 6, 219 (1960), 13, 1 (1971) Muscarine alkaloids, 23, 327 (1984) Mushrooms, alkaloids from, 40, 190 (1991) Mydriatic alkaloids, 5, 243 (1955) a-Naphthophenanthridine alkaloids, 4, 253 (1954), 10,485 (1967) Naphthylisoquinoline alkaloids, 29, 141 (1986) Narcotics, 5 , 1 (1955) Nuphar alkaloids, 9,441 (1967), 16, 181 (1977), 35,215 (1989) Ochrosia alkaloids, 8, 336 (1963, 11, 205 (1968) Ourouparia alkaloids, 8, 59 (1965), 10, 521 (1967) Oxazole alkaloids, 35, 259 (1989) Oxaporphine alkaloids, 14, 225 (1973) Oxindole alkaloids, 14, 83 (1973)

Papaveraceae alkaloids, 19,467 (1967), 12, 333 (l970), 17,385 (1979) pharmacology, 15,207 (1975) toxicology, 15,207 (1975)

370

CUMULATIVE INDEX OF TITLES

Pauridiantha alkaloids, 30, 223 (1987) Pavine and isopavine alkaloids, 31, 3 17 (1987) Pentaceras alkaloids, 8,250 (1965) Peptide alkaloids, 26, 299 (1985) Phenanthrene alkaloids, 39,99 (1990) Phenanthroindolizidine alkaloids, 19, 193 (1981) Phenanthroquinolizidine alkaloids, 19, 193 (1981) P-Phenethylamines, 3, 313 (1953), 35,77 (1989) Phenethylisoquinoline alkaloids, 14, 265 (19731, 36, 172 (1989) Phthalideisoquinoline alkaloids, 4, 167 (1954), 7, 433 (19601, 9, 117 (1967), 24, 253 (1985) Picralima alkaloids, 8, 119 (1965), 10, 501 (19671, 14, 157 (1973) Piperidine alkaloids, 26,89 (1985) Plant biotechnology, for alkaloid production, 40, 1 (1991) Plant systematics, 16, 1 (1977) Pleiocarpa alkaloids, 8, 336 (19651, 11, 205 (1968) Polyamine alkaloids, 22, 85 (1983) Pressor alkaloids, 5 , 229 (1955) Protoberberine alkaloids, 4, 77 (1954),9, 41 (1967), 28,95 (1986) transformation reactions of, 33, 141 (1988) Protopine alkaloids, 4, 147 (1954), 34, 181 (1989) Pseudocinchoma alkaloids, 8,694 (1965) Purine alkaloids, 38, 226 (1 990) Pyridine alkaloids, 1, 165 (1950), 6, 123 (1960), 11,459 (1968), 26, 89 ( 1985) Pyrrolidine alkaloids, 1,91 (1950), 6, 31 (1960), 27,270 (1986) Pyrrolizidine alkaloids, 1, 107 (1950), 6, 35 (1960), 12, 246 (1970), 26, 327 (1985)

Quinazolidine alkaloids, see Indolizidine Alkaloids Quinazoline alkaloids, 3, 101 (1953), 7, 247 (1960), 29, 99 (1986) Quinazolinocarbolines, 8,55 (1965),21,29 (1983) Quinoline alkaloids related to anthranilic acid, 3, 65 (1953), 7,229 (1960), 17, 105 (1979), 32,341 (1988) Quinolizidine alkaloids, 28, 183 (1985) Rauwolfia alkaloids, 8, 287 (1965) Reissert synthesis of isoquinoline and indole alkaloids, 31, 1 (1987) Reserpine, chemistry, 8, 287 (1965) Respiratory stimulants, 5 , 109 (1955) Rhoeadine alkaloids, 28, 1 (1986)

CUMULATIVE INDEX OF TITLES

Sulumandru group, steroids, 9, 427 (1967) Sceletium alkaloids, 19, 1 (1981) Secoisoquinoline alkaloids, 33, 23 1 (1988) Securinega alkaloids, 14,425 (1973) Senecio alkaloids, see Pyrrolizidine alkaloids Simple indole alkaloids, 10,491 (1967) Simple indolizidine alkaloids, 28, 183 ( I 986), 44, 189 (1993) Sinomenine, 2,219 (1952) Solanum alkaloids chemistry, 3, 247 (1953) steroids, 7,343 (1960), 10, 1 (1967), 19,81 (1981) Sources of alkaloids, 1, 1 (1950) Spectral methods, alkaloid structures, 24,287 (1985) Spermidine and related polyamine alkaloids, 22,85 (1983) Spermine and related polyamine alkaloids, 22, 85 (1983) Spirobenzylisoquinolinealkaloids, 13, 165 (1971),38, 157 (1990) Sponges, isoquinolinequinone alkaloids from, 21, 55 (1983) Stemona alkaloids, 9,545 (1967) Steroid alkaloids Apocynaceae, 9,305 (1967),32,79 (1988) Buxus group, 9, 305 (1967), 14, 1 (1973), 32,79 (1988) Holarrhena group, 7,319 (1960) Salamandra group, 9,427 (1967) Solanum group, 7,343 (1960), 10, 1 (1967), 19,81 (1981) Veratrum group, 7 , 363 (1960), 10, 193 (1967), 14, 1 (1973), 41, 177 ( 1992) Stimulants respiratory, 5, 109 (1955) uterine, 5, 163 (1955) Structure elucidation, by X-ray diffraction, 22, 51 (1983) Strychnos alkaloids, 1,375 (part 1, 1950), 2,513 (part 2, 1952), 6, 179 (1960), 8,515, 592 (1965), 11, 189 (1968), 34,211 (1989), 36, 1 ( 1989) Sulfur-containingalkaloids, 26, 53 (1983, 42, 249 (1992) Synthesis of alkaloids, Enamide cyclizations for, 22, I89 (1983) Lead tetraacetate oxidation in, 36,70 (1989) Tabernaemontana alkaloids, 27, 1 (1983) Taxus alkaloids, 10, 597 (1967), 39, 195 (1990) Thailand, alkaloids from the plants of, 41, 1 (1992) Toxicology, Papaveraceae alkaloids, 15, 207 (1975)

37 I

372

CUMULATIVE I N D E X OF TITLES

Transformation of alkaloids, enzymatic microbial and in uitro, 18, 323 (1981) Tropane alkaloids biosynthesis of, 44, 115 (1993) chemistry, 1,271 (1950), 6, 145 (1960), 9,269(1967), 13, 351 (1971), 16,83 (1977), 33,2 (1988), 44, I (1993) Tropoloisoquinoline alkaloids, 23, 301 (1984) Tropolonic Colchicurn alkaloids, 23, 1 (1984), 41, 125 (1992) Tylophora alkaloids, 9, 517 (1967) Uterine stimulants, 5 , 163 (1955) Veratrum alkaloids cevane group of, 41, 177 (1992) chemistry, 3, 247 (1952) steroids, 7, 363 (1960), 10, 193 (1967), 14, 1 (1973) Vinca alkaloids, 8,272 (1965), 11,99 (1968), 20,297 (1981) Voacanga alkaloids, 8, 203 (1963, 11,79 (1968) X-ray diffraction of alkaloids, 22, 51 (1983) Yohimbe alkaloids, 8, 694 (1965), 11, 145 (1968), 27, 131 (1986)

INDEX

Acetoacetate, as precursor of hyoscyamine and hygrine. 141-142 ( + )-6P-Acetoxynortropane, He and Brossi synthesis, 87 I -Acetylcarbazole, 286 Aflavazole, 316-317 Agmatine, 174 L-Alanine, in monomorine I synthesis, 193-195 Aliphatic acids, esters, biosynthesis, 156-158 Allopumiliotoxin A alkaloids, 199-201 Allopumiliotoxins, total synthesis, 216-219 Amine precursors, feeding studies, 169- 170 Amphibians, indolizidine alkaloids from. 196-220 Anhydroecgonine methyl ester, Davies synthesis. 81-82 Anthocercoideae, tropane alkaloids in, 38-44 Anti-inflammatory properties, carbazole alkaloids. 357-358 Antimicrobial properties, carbazole alkaloids, 353-354 Antitumor alkaloids, 323-324, 354 Antiviral activity, carbazole alkaloids, 355 Ants, indolizidine alkaloids from, 190-196 Arcyriaflavin-B, 3 17-319 Arcyriaflavin-C, 317-319 Arginine decarboxylase, activity changes, 176 Aromatic tropane esters, structures, 118 Atropine demethylation, 89, 91 photocyanation, 91 Atropine esterase, activity, 167 Atropoideae, tropane alkaloids in, 56-63 8-Azabicyclo[3.2. Iloctan-8-one, asymmetric a-ketonic cleavage, I 93- 194 373

Backvall synthesis, tropine, pseudotropine. scopine, and pseudoscopine, 78-79 Baogongteng A Jung et d.synthesis, 85-87 Xiang c/ a / . synthesis, 84, 86 Bathgate and Malpass syntheses, tropane alkaloids, 80-81 Belladonnines, structure, 22 Benzoylecgonine ethyl ester, synthesis, 88 3P-Benzoyloxytropane, thermal degradation, 89-90 N-Benzylnortrop-6-ene. Bathgate and Malpass synthesis, 80-81 Benzyltropanes disubstituted, 14-15 substituted, 14 Biogenesis, carbazole alkaloids, 35 1-352 Biscarbazole alkaloids, 332-348 synthesis. 347-348 Bis-7-hydroxygirinimbine-A,346-347 Bis-7-hydroxygirinimbine-B, 346-347 Bismurrayafoline-A, 333-334 Bismurrayafoline-B, 334-335 Bismurrayafoline-C, 339-340 Bismurrayafoline-D, 339-340 Bismurrayafolinol, 335 Brassicaceae, tropane alkaloids in, 32

Carazostatin, 288-290 Carbazole, 258 Carbazole alkaloids, 257-360; see also Biscarbazole alkaloids anti-inflammatory properties, 357-358 antimicrobial properties. 353-354 antitumor activity, 354-355 antiviral activity, 355 biogenesis, 351-352 cardiovascular-modulating activity, 355-356

374

INDEX

central nervous system activity, 356-357 hexacyclic alkaloids, 3 14 hexacyclic base synthesis, 331-332 hexa- and octacyclic indolocarbazoles, 317-331 infrared spectra, 349 mass spectra, 350 modulation of enzyme activity, metabolism, and allergic reactions, 358-359 NMR spectra, 349-350 occurrence, 258-262 penta- and hexacyclic alkaloids, 315-317 tetracyclic alkaloids, 297-3 14 from higher plants, 297-306 optical properties transformation, 3 I3 from Streptomyces, 306-310 synthesis, 310-312 transformations, 312-314 tricyclic alkaloids from higher plants, 258-283 from non-plant sources, 283-290 synthesis, 290-297 tumor-promoting activity, 354-355 ultraviolet absorption spectra, 349 X-ray crystallography, 350-35 I Carbazole-3-carboxylic acid, 273 Carbazole-3-methylcarboxylate,272-273 Carbazomycin, 294-295 Carbazomycin-A, 290-29 I Carbazomycin-B, 290-291 Carbazomycin-C, 286 Carbazomycin-D, 287 Carbazomycin-G, 288 Carbazomycin-H, 288 Carbazomycinal, 287 Carbofuran, acute intoxication, 98 Carboxytropanes, substituted, 13-14 Cardiovascular-modulating activity, carbazole alkaloids, 355-356 Carprofen anti-inflammatory activity, 358 antiviral activity, 355 Castanospermine, 239-249 stereoisomers, 240 synthesis based on intramolecular cyclization, 243-244 based on stereoselective reduction of cyclic ketone, 243-245

chemoenzymatic avenue, 246-248 double cyclization of epoxy amino ester, 240-242 from glucuronolactone, 242-243 noncarbohydrate starting materials, 245-247 stereoselective, 241-242, 248-249 total, 240-241 unnatural enantiomer, synthesis, 244-246 Central nervous system, carbazole alkaloid activity, 356-357 Cestroideae, tropane alkaloids in, 44-45 3-Chlorocarbazole, 290 6-Chlorohyellazole, 284-285 Chrestifoline-A, 338 Chrestifoline-B, 342 Chrestifoline-C, 343-344 Chrestifoline-D, 334 Clausenapin, 274-275 Cocaine biosynthesis, 92, 94, 142 pharmacology, 98-99 thermal degradation, 90

Datum strnrnonium alkaloid degradation, 165-166 tropane alkaloids, biosynthesis and flux, 129, 172-174 Datureae, tropane alkaloids in, 46-55 Davies synthesis, tropane alkaloids, 81-82 Deacetylkinamycin-C, 3 10 Degradation thermal, tropane alkaloids, 89-90 tropeines, 164-168 2,I'-Dehydrohygrine. biomimetic conversion of hygrine to, 144-145 6.7-Dehydrohyoscyamine. 163 4-Demethoxycarbazomycin-A, synthesis, 294-296 Demethylation, tropane alkaloids, 89-9 1 0-Demethylmurrayanine, 271 DL-a-Difluoromethylarginine, effect on putrescine biosynthesis, 132-133 DL-a-Difluoromethylornithine, effect on putrescine biosynthesis, 132-133 Dihydropyranotropanes, 16-17 Dihydroxygirinimbine, 298

INDEX

I ,2-Dihydroxyindolizidines,228-232 Dihydroxynortropanes, 18 Dihydroxytropanes, distribution, 77-78 Drrboisiu, tropane alkaloid biosynthesis, 129-1 30

Ecgonine methyl ester, formation, 142 Ekebergenine, 278-279 Elncwcurpris alkaloids, 221-224 Elaeokanines. total synthesis, 221-224 Epoxytropanes, substituted, 13 Erythroxylaceae, tropane alkaloids in, 34-38 Euchrestine-A. 275 Euc hrestine-B, 275-276 Euchrestine-C, 276 Euchrestine-D. 276-277 Euchres t ine- E. 277 Euphorbiaceae, tropane alkaloids in, 32 Eustifoline-B, 302 Eustifoline-C. 278 Eustifoline-D, 297

bFerruginine. Davies synthesis. 81-82 3-Formylcarbazole, 267 3-Formyl-7-hydroxycarbazole. 267 3-Formyl-6-methoxycarbazole, 269 Furostifoline, 297 Furuya synthesis, tropane alkaloids, 83-85

(2

Girinimbine, photochemical transformation, 3 13-3 14 Glycomaurin, 302 Glycomaurrol, 275 Glycozolidal, 271 Glycozolidine. 293 Glycozoline, synthesis, 292-293 Grahamine. 13C-NMR spectroscopy, 96-97

Harper approach. tropane alkaloids, 88 He and Brossi synthesis. ( + )-6P-acetoxynortropane, 87

375

Heptazolicine, 303-304 6-Hydroxyhyoscyamine epoxidase, 163 I-Hydroxyindolizidines. 228-232 Hydroxyltropanols, 164 2-Hydroxy-3-methylcarbazole, 263. 294 I -Hydroxynortropopane skeleton, Lallemand synthesis, 83-84 3a-H ydrox ytropane demethylation. 89-90 photocyanation, 91 2a-Hydroxytropinone. Moriarty synthesis, 83-84 Hyellazole, 283-285 synthesis, 294-295 Hygrine acetoacetate as precursor. 141-142 biomimetic conversion to tropinone or 2,1 ‘-dehydrohygrine, 144-145 isomers. as precursors of tropane alkaloids, 143- I44 Hyoscine hyoscyamine conversion to, 160-164 production by tissue cultures, 120-127 structure, I18 Hyoscyamine acetoacetate as precursor, 141-142 biosynthesis, 92-93, 158-159 tropic acid effect, 169. 171 conversion to hyoscine, 160-164 production by tissue cultures. 120-127 structure, I18 Hyoscyamine 6P-hydroxylase. SUbStrdte specificity. 161- I62 H v o s c y m r r s nlbrrs, N-methylputrescine oxidase, kinetic properties, 137-138 H.voscymrrs species, tropane alkaloid biosynthesis, 128

lndole carbazole dimer. 332-333 Indolizidine alkaloids. 189-250 from amphibians, 196-220 structure, 196-20 1 synthesis, 202-220 from ants, 190-196 diepoxides, enantiomer synthesis, 206-208 3,5-disubstituted, 197-198 enantiomer synthesis, 205-206

376 Elaeocarpus alkaloids, 221-224 ( - )-enantiomer synthesis, 206-209 ( + )-enantiomer synthesis, 209-21 I hydroxylated, 228-249 ( + )-Indolizidine 195B, total synthesis, 203-205 I ,2-lndolizidinediol, 231-232 Indolocarbazoles, hexa- and octacyclic, 317-331 Infrared spectra, carbazole alkaloids, 349 Isomahanine, 303 Isomurrayafoline-B, 274 fmns(3,5)-lsoxazolidine, 192-193

Jabroseae, tropane alkaloids in, 46 Jung et a / . synthesis, baogongteng A, 85-87

Ke toanh ydrokinamycin, 309 Kinamycin-A, 307-308 Kinamycin-B, 308 Kinamycin-C, 306-307 antitumor activity, 354 Kinamycin-D, 308 Kinamycin-E, 309 Koenoline, 266-267 synthesis, 290, 292

Lallemand synthesis, I-hydroxynortropopane skeleton, 83-84 Lansbury synthesis, tropane alkaloids, 82-83 Leete and Kim synthesis, tropane alkaloids. 82-83

Mahanimbine cyclomer formation in, 312-313 photochemical transformation, 3 13-314 Mann synthesis, oscine, 87-88 Mass spectrometry carbazole alkaloids, 350 tropane alkaloids, 96-97

INDEX

Metabolite tunneling, in putrescine biosynthesis, 133- 134 6-Methoxycarbazole-3-rnethylcarboxylate, 273 6-Methoxycarbazomycinal, 287-288 7-Methoxyheptaphylline, 279-280

N-Methoxy-3-hydroxymethylcarbazole, 269

2-Methoxy-3-methylcarbazole,263 7-Methoxy-O-methylmukonal, 270 7-Methoxymukonal, 270 7-Methoxymurrayacine, 302-303 6-Methoxymurrayanine, 270-271 1-Methylcarbazole, 285-286 3-Methylcarbazole, 293 N-Methylcarbazole, tumor-promoting activity. 354-355 8-Methylindolizidines. 5-substituted, 197-198 synthesis, 21 1-213 0-Methylmukonal, 269-270 N-Methylpelletierine. synthesis, 143 N-Methylputrescine, 175 biosynthesis, 130- 13 I N-Methylputrescine oxidases, kinetic properties, 137-138 N-Methylpyrroliniurn conversion of putrescine to, 134-140 conversion to tropinone, 140- 145 Monomorine I asymmetric a-ketonic cleavage of 8-azabicyclo[3.2.1Iloctan-8-one. 193- 194 synthesis from L-alanine, 193-195 asymmetric, 190-192 enantioselective total, 192-193 from (S)-pyroglutamic acid, 195 Moriarty synthesis, 2a-hydroxytropinone. 83-84 Mukolidine, 265-266 Mukoline, 265-266 Mukonal, 269 Mukonicine, 301 Mukonine, synthesis, 290-291 Murrafoline, 347 Murrafoline-B, 340 Murrafoline-C, 344 Murrafoline-D, 341 Murrafoline-E, 341 Murrafoline-F, 336

INDEX

Murranimbine, 344-345 Murrastifoline-A, 337-338 Murrastifoline-B, 337-338 Murrastifoline-C. 343 Murrastifoline-D. 34 1-342 Murrastifoline-E, 342 Murrayafoline-A, 262, 293 Murrayafoline-B, 273-274 Murrayaline-A, 264-265 Murrayaline-B, 271-272 Murrayaline-C. 272 Murrayaline-D. 279 Murrayanol, 277-278 Murrayaquinone-A, 280 Murrayaquinone-B, 280-283 Murrayaquinone-C, 283 Murrayaquinone-D, 283 Murrayastine. 263-264 Murrayazoline, 3 14 Murrayazolinol, 3 14

Nicandreae, tropane alkaloids in, 56 Nicoriunu rabricum, N-methylputrescine oxidase, kinetic properties, 137-138 Nicotine, biosynthesis, 172 NMR. see Nuclear magnetic resonance Norgirinimbine, synthesis, 312 Nuclear magnetic resonance I 'C carbazole alkaloids, 350 tropane alkaloids, 95-97 'H carbazole alkaloids, 349-350 tropane alkaloids, 95-96

Olacaeae, tropane alkaloids in, 38 Optical activity, tropane alkaloids, 77-78 Organ tissue cultures, tropane alkaloid biosynthesis, 119-120 Dcitirrci sfrumonium. 129 Dtrhoisici, 129-130 Hyosc-yumirs. 128 Ornithine decarboxylase. activity changes. I76 Oscine Mann synthesis. 87-88 NMR spectroscopy, 95-96

377

N-Oxidation, tropeines, 168 3-Oxotropane demethylation, 89. 91 photocyanation, 91 structure, 17 Oxydimurrayafoline, 335-336

Pharmacology. tropane alkaloids. 97-99 Phenylalanine, metabolism, 151-152 Phenyllactic acid, biosynthesis. 151-154 Phenylpyruvic acid, 152, 154 Photocyanation, tropane alkaloids, 89. 91 Piperidine, cis-2,6-disubstituted, 195-196 Plants, containing tropane alkaloids, 4-5 Prekinamycin, 308-309, 31 1 Proteaceae, tropane alkaloids in, 32-33 Protein kinase C inhibitors K-252a, 330-331 K-252b. 330-331 K-252~.319-320 K-252d, 320 Pseudoscopine, Backvall synthesis, 79-80 Pseudotropine acetylation, 156 Backvall synthesis, 78-79 biosynthesis, 146-150 Pumiliotoxins A, 199-201 synthesis, 215-216 Pumiliotoxins D, enantioselective synthesis. 217-218, 220 Putrescine biosynthesis, 130- I34 enzyme inhibitor effect, 132-133 labeled precursor incorporation. 132 metabolite tunneling, 133-134 conversion to N-methylpyrrolinium. 134-140 formation. 174 Putrescine N-methyltransferase biosynthesis regulatory role, 172 induction, 174 Pyranotropanes, structure, I6 Pyrayafoline-A, 298-299 Pyrayafoline-B, 299-300 Pyrayafoline-C, 300 Pyrayafoline-D, 300 Pyrayafoline-E. 301 Pyrayaquinone-A, 304 Pyrayaquinone-B. 304-305

378

INDEX

Pyrayaquinone-C, 305-306 acid, in monomorine I synthesis, 195

( S)-Pyroglutamic

Rebeccamycin, 320-322 Rhizophoraceae, tropane alkaloids in, 33 Rincazole, 357

Schizanthines, 21, 23 Scopadonnines, 22-23 Scopine, Backvall synthesis, 79-80 Scopolamine biosynthesis, 94-95 NMR spectroscopy, 95-96 pharmacology, 99 Sikabaceae, tropane alkaloid degradation, I65 Slaframine. 223-227 asymmetric synthesis, 224-226 enantioselective synthesis, 226 synthesis via radical cyclization, 226-227 Solandreae, tropane alkaloids in, 55-56 Solaneae, tropane alkaloids in. 45-46 Speckamp synthesis, tropane alkaloids, 84-85 Staurosporine, 324-328 Swainsonine, 233-239 noncarbohydrate route, 237-238 stereoisomers, 238-239 synthesis enantioselective, 234-236 enantiospecific, 233 from hydroxy lactam, 236-237 from D-mannose, 234-235 from tartarimide, 238-239

Tan-999, 330 Tan-I030A, 328-330 I ,6,7,8-Tetrahydroxyindolizidines, 249 Tetrahydroxynortropanes, 19 8-Thiabicyclo[3.2. Iloctan-3-one, 149- 150 Tiglic acid, biosynthesis, 154-155 Tigloidine, synthesis, 157 /3-Tigloxyltropane, synthesis, 157

3-Tigloylox ytropane formation. I7 1- 172 synthesis, 159-160 Trihydroxynortropanes, 18 Tropane alkaloids, 1-100 acyl group structures, 26-31 in Anthocercoideae, 38-44 in Atropoidea, 56-63 biosynthesis, 92-95 day length effect, 178 formation of tropanes from amino acids, 116-117 organ tissue cultures, 119-130 pathway regulation biochemical level, 169-176 whole-plant level, 177-180 temperature effects, 178 water stress effect, 179 in Brassicaceae, 32 in Cestroideae, 44-45 chemotaxonomy of plants, 65-77 in Concoculaceae. 63-65 in Datureae, 46-55 demethylation. 89-91 disubstituted, structures, 9-12, 14-15 in Erythroxylaceae, 34-38 in Euphorbiaceae, 32 industrial preparation, 99- 100 in Jabroseae. 46 mass spectrometry, 96-97 medicinal use. 116 3cu-monosubstituted, structures, 7-8 3P-monosubstituted, structures, 9 in Nicandreae, 56 NMR spectra, 95-97 numbering system, 2 in Olacaceae, 38 ontogeny of accumulation, 177 optical activity, 77-78 pharmacology, 97-00 photocyanation. 89. 91 plant origin, 66-76 plants containing, 4-6 potassium stress effect, 179 in Proteaceae, 32-33 in Rhizophoraceae, 33 ring system, 2 in Solandreae, 55-56 in Solaninae, 45-46 in Solanoideae, 45-46

INDEX

structures, 6-31 synthesis, 78-88 thermal degradation, 89-90 trisubstituted, structures, 12 Tropeines biosynthesis, 155-160 acidic moieties, 151-155 esters of aliphalic acids, 156-158 hyoscyamine and aromatic esters, 158-160 degradation and oxidation, 164-168 metabolism, 160-164 Tropic acid biosvnthesis. 15 1- 154 effect on hyoscyamine production, 169, 171 Tropine acetylation, 156-157 Backvall synthesis, 78-79 biosynthesis, 92-93, 146-150 feeding studies, 169-170 Tropinone biosynthesis, 92-93, 134-145 N-methylpyrrolinium conversion to tropinone, 140-145

3 79

putrescine conversion to N-methylpyrrolinium, 134- 140 Lansbury synthesis, 82-83 Tropinone reductase I , 146-148 Tropinone reductase 11, 147-150 Truxillines, 25 Tubingensin-A, 3 15-316 Tubingensin-B, 316 Tumors, promotion by carbazole alkaloids. 354-355

UCN-01, 331 Ultraviolet absorption spectra, carbazole alkaloids. 349

Xiang et a / . synthesis, boagongteng A. 84, 86 X-ray crystallography, carbazole alkaloids. 350-35 I

This Page Intentionally Left Blank