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Studies related to the synthesis of bisindole alkaloids of the vinblastine-Vincristine family Balsevich, J. John 1978

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STUDIES RELATED TO THE SYNTHESIS OF BISINDOLE ALKALOIDS OF THE VINBLASTINE-VINCRISTINE FAMILY  by  J . JOHN BALSEVICH B.Sc,  U n i v e r s i t y o f B r i t i s h Columbia, 1972  M . S c , U n i v e r s i t y o f B r i t i s h Columbia, 1975  A THESIS SUBMITTED IN PARTIAL FULFILMENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY  in THE FACULTY OF GRADUATE STUDIES (Dept. o f C h e m i s t r y , U n i v . o f B r i t i s h Columbia) We accept t h i s t h e s i s as conforming t o the r e q u i r e d  standard  THE UNIVERSITY OF BRITISH COLUMBIA September, 19 7 8  ©  J . John B a l s e v i c h , 1978  In p r e s e n t i n g t h i s  thesis  an advanced degree at  in p a r t i a l  f u l f i l m e n t o f the requirements f o r  the U n i v e r s i t y of B r i t i s h Columbia,  the L i b r a r y s h a l l make i t f r e e l y a v a i l a b l e I  f u r t h e r agree t h a t p e r m i s s i o n  for  r e f e r e n c e and study.  f o r e x t e n s i v e copying o f  this  representatives. thesis  It  this  thesis  i s understood that copying o r p u b l i c a t i o n  f o r f i n a n c i a l gain s h a l l  written permission.  Department of  Chemistry  The U n i v e r s i t y o f B r i t i s h 2075 Wesbrook P l a c e V a n c o u v e r , Canada V6T 1W5  Date  •6  that  s c h o l a r l y purposes may be granted by the Head of my Department or  by h i s of  for  I agree  O r t . 4. 1978  Columbia  not be allowed without my  ABSTRACT  The work done t o w a r d s t h i s t h e s i s i s o u t l i n e d i n f o u r sections.  The  indolenine  coupling'  (10).  first  Accordingly,  starting  s e c t i o n i n v o l v e d e v a l u a t i o n of the of velbanamine d e r i v a t i v e s w i t h  vindoline  (19)/  c h o s e n as  18S-carbomethoxycleavamine  m a t e r i a l , was  converted to  velbanamine c h l o r o i n d o l e n i n e  18S-carbomethoxy-3R-hydroxy-  (55) i n f i v e s t e o s .  A t t e m p t e d couu  l i n g of t h i s d e r i v a t i v e w i t h v i n d o l i n e i n methanol h y d r o g e n c h l o r i d e was  unsuccessful.  Therefore,  Consequently, i n t e r n a l  coupling.  q u a t e r n i z a t i o n of the t e r t i a r y  e s t a b l i s h e d as a c o m p e t i t i v e  side reaction.  r e s u l t s the c o n d i t i o n s of the coupling to eliminate  containing  s t u d i e s were  undertaken t o a s c e r t a i n a reason f o r the lack of  was  'chloro-  this side reaction.  carbomethoxy-3R-hydroxyvelbanamine a t t a i n e d , however, the o n l y d i m e r i c  B a s e d on  r e a c t i o n were  Coupling  nitrogen these  modified  o f v i n d o l i n e and  chloroindolenine product obtained  was was  18S-  thus 18'—  e o i v i n c a d i o l i n e (74). The s e c o n d s e c t i o n i n v o l v e d t h e s t u d y o f t h e f u n c t i o n a l i z a t i o n of 3',4'-anhydrovinblastine  (26) u s i n g  a e r i a l or t - b u t y l  hydroperoxide o x i d a t i o n .  Under the a p p r o p r i a t e c o n d i t i o n s the  n a t u r a l l y o c c u r r i n g a l k a l o i d s l e u r o s i n e (3) and C a t h a r i n e c o u l d be o b t a i n e d .  A study was  (76)  c a r r i e d out i n regards t o the  mechanism of the o x i d a t i o n w i t h the r e s u l t b e i n g i m p l i c a t i o n of a r a d i c a l mechanism. The t h i r d s e c t i o n i n v o l v e d the study of the r e a c t i o n o f 3 ' , 4 ' - a n h y d r o v i n b l a s t i n e (and r e l a t e d d e r i v a t i v e s ) w i t h potassium permanganate.  From t h i s r e a c t i o n was o b t a i n e d as the major  product 3R-hydroxyvinamidine  (8^3)—a d e r i v a t i v e o f the n a t u r a l l y  occurring a l k a l o i d vinamidine 3R-hydroxyvinamidine  was  (82).  Attempted  unsuccessful.  deoxygenation  of  Therefore, vinamidine  was prepared by potassium permanganate o x i d a t i o n of 4'-deoxyleurosidine  (93).  The f o u r t h s e c t i o n r e l a t e s t o the p r e p a r a t i o n o f v i n b l a s t i n e - v i n c r i s t i n e analogs  ( f o r the purpose of e l u c i d a t i n g  structure-activity relationships).  Thus, i o d i n e / s o d i u m b i c a r b o n -  ate o x i d a t i o n of l e u r o s i n e ( 3 ) , v i n b l a s t i n e  ( 1 ) , and  leurosidine  (JU p r o v i d e d access t o the c o r r e s p o n d i n g 19'-oxo d e r i v a t i v e s i n f a i r t o moderate y i e l d s .  O x i d a t i o n of 1 9 ' - o x o l e u r o s i n e  Jones reagent at low temperature (98).  (91) w i t h  afforded 19 ,22-dioxoleurosine 1  A l t e r n a t i v e l y 19 ,22-oxoleurosine 1  c o u l d be o b t a i n e d from  l e u r o s i n e i n b e t t e r y i e l d by f i r s t u t i l i z i n g Jones o x i d a t i o n t o a f f o r d 2 2 - o x o l e u r o s i n e and then o x i d a t i o n o f t h i s d e r i v a t i v e w i t h iodine/sodium bicarbonate.  t - B u t y l hydroperoxide o x i d a t i o n of  6 , 7-dihydro-3 , 4 ' - a n h y d r o v i n b l a s t i n e 1  (100) a f f o r d e d the n o v e l  analog 6 , 7 - d i h y d r o l e u r o s i n e (101) , Jones o x i d a t i o n of which y i e l d e d 22-oxo-6 , 7 - d i h y d r o l e u r o s i n e ( 1J02) .  - iv TABLE OF  CONTENTS Page  T i t l e page  i  Abstract Table  i i  of Contents  iv  L i s t of Figures  v  L i s t of Tables  ix  Acknowledgements  x  Introduction  1  1.1  General  1.2  S t r u c t u r e E l u c i d a t i o n of V i n b l a s t i n e , V i n c r i s t i n e , L e u r o s i n e , and L e u r o s i dine  6  S t u d i e s Aimed a t t h e S y n t h e s i s o f t h e 'Dimeric' Catharanthus A l k a l o i d s — the ' C h l o r o i n d o l e n i n e Approach  14  S t u d i e s Aimed a t t h e S y n t h e s i s o f Dimeric Catharanthus A l k a l o i d s 'Biogenetic Approach'  20  1.3  Background  1  1  1.4  the the  1.5  Vincristine Derivatives  23  1.6  Scope of C u r r e n t R e s e a r c h  25  Discussion  28  2.1  F u r t h e r E v a l u a t i o n of the C h l o r o i n d o l e nine Approach  28  2.2  Chemistry of 3',4'-Anhydrovinblastine— S y n t h e s i s o f L e u r o s i n e and C a t h a r i n e .  83  2.3  Chemistry of 3 ,4'-Anhydrovinblastine— S y n t h e s i s of Vinamidine (Catharinine) (82) and R e l a t e d D e r i v a t i v e s  93  P r e p a r a t i o n of Analogs  114  2.4  1  Vinblastine-Vincristine  Experimental  125  Bibliography  165  - v -  LIST OF FIGURES Figure  Page  1.  S t r u c t u r e o f some c l i n i c a l l y used i n g agents  alkylat-  2.  Some a n t i m e t a b o l i t e cancer drugs and t h e i r m e t a b o l i t e analogs  3  3.  S t r u c t u r e s o f some c y t o t o x i c a n t i b i o t i c s ..  4  4.  A s i g n i f i c a n t pathway f o r the f r a g m e n t a t i o n of l e u r o s i n e i n t h e mass spectrometer  9  2  5.  O u t l i n e o f Kutney and Worth's s t r u c t u r e proof of l e u r o s i n e  11  6.  P o t i e r and L a n g l o i s ' s t r u c t u r e - p r o o f o f leurosidine  13  7.  Mechanism f o r t h e f o r m a t i o n o f 18S-carbomethoxycleavamine from c a t h a r a n t h i n e  16  8.  General o u t l i n e o f the 'chloroindolenine approach  17  9.  Condensation o f the c h l o r o i n d o l e n i n e o f 4S-ethyl-18Scarbomethoxydihydrocleavamine with vindoline  19  'Biogenetic-type' coupling of catharanthineN, - o x i d e (24) w i t h v i n d o l i n e  22  11.  Some ' d i m e r i c ' catharanthus a l k a l o i d s  26  12.  P o s s i b l e mchanism f o r t h e f o r m a t i o n o f 4S,-18R-ether 37 from v i n b l a s t i n e  29  13.  "*"Hmr spectrum o f 16 ,18S-dicarbomethoxycleavamine (39)  33  14.  Comparison o f t h e u l t r a v i o l e t s p e c t r a o f 18S-carbomethoxycleavamine (A)and 1 6 , 1 8 S — dicarbomethoxycleavamine (B)  34  S p a t i a l r e l a t i o n s h i p o f the 18-hydrogen t o the N e l e c t r o n p a i r i n 18-carbomethoxycleavamine  34  "^Hmr spectrum of 16 ,18S-dicarbomethoxy— 3R,4S-epoxydihydrocleavamine (40)  36  10.  1  b  15.  <—  b  16.  - vi -  Figure 17.  Page Sequence used f o r the d e t e r m i n a t i o n o f the absolute c o n f i g u r a t i o n of 16,18S-dicarbomethoxy-3R,4S-epoxydihydrocleavamine (40) ..  39  18.  ^"Hmr  42  19.  I n f r a r e d spectrum o f formyl  20.  I n f r a r e d s p e c t r u m o f a l c o h o l 47  45  21.  Mass s p e c t r a l f r a g m e n t a t i o n p a t t e r n o f 16,18S-dicarbomethoxy-3R-hydroxyvelbanamine (51)  49  22.  "*"Hmr s p e c t r u m o f 16 , 1 8 S - d i c a r b o m e t h o x y — 3 R - h y d r o x y v e l b a n a m i n e (51)  50  23.  ^"Hmr s p e c t r u m o f 1 6 , 1 8 S - d i c a r b o m e t h o x y — 3 S , 4 R - e p o x y d i h y d r o c l e a v a m i n e (52)  51  Mass s p e c t r a l f r a g m e n t a t i o n p a t t e r n o f 16,18S-dicarbomethoxy-3S,4R-epoxydihydrocleavamine (52) ..  52  Ultraviolet i n e 55  55  24.  25. 26.  spectrum of formyl  k e t o n e 45  spectrum of the  k e t o n e 45  chloroindolen-  P r o p o s e d g e n e r a l mechanism o f ine 'dimerization  chloroindolen-  1  27.  28.  43  56  Comparison of the u l t r a v i o l e t s p e c t r a o f a t y p i c a l indole d e r i v a t i v e (A), dihydroindole d e r i v a t i v e (B) , and 'dimer' (C)  62  I n f r a r e d spectrum of 16,18S-dicarbomethoxy— 3R,4S-epoxy-19-oxodihydrocleavamine (62) ...  64  13 29. 30. 31. 32.  Cmr s p e c t r u m o f 1 6 , 1 8 S - d i c a r b o m e t h o x y - 3 R , 4 S - e p o x y - 1 9 - o x o d i h y d r o c l e a v a m i n e (62) 13 Comparison o f the Cmr c h e m i c a l s h i f t s (<0 o f h e x a h y d r o a z e p i n e (A) and c a p r o l a c t a m ( B ) . ^Hmr s p e c t r u m o f 1 6 , 1 8 S - d i c a r b o m e t h o x y - 3 R , 4 S - e p o x y - 1 9 - o x o d i h y d r o c l e a v a m i n e (62) ^Hmr s p e c t r u m o f 1 8 S - c a r b o m e t h o x y - 3 R , 4 S — e p o x y - 1 9 - o x o d i h y d r o c l e a v a m i n e (66)  65 66 67 69  -  v i i-  Figure 33.  Page Outline o f the reaction of 18S-carbomethoxy— 3R,4S-epoxy-19-oxodihydrocleavamine (66) with 1-chlorobenyotriazole 7".  70  34.  ^Hmr s p e c t r u m  o f 68  72  35.  "4lmr s p e c t r u m o f 69  73  36.  37.  1  Hmr s p e c t r u m (70) Proposed dolenine  of18 -epi-19 -oxoleurosine 1  1  75  mechanism f o r c o u p l i n g 61 w i t h v i n d o l i n e  of chloroin77  38.  "'"Hmr s p e c t r u m  of 18-epileurosine  39.  "'"Hmr s p e c t r u m  of 18'-epivincadioline  40.  ^"Hmr s p e c t r u m o f C a t h a r i n e ( s a m p l e w a s o b t a i n e d from t h e - E l i L i l l y L a b o r a t o r i e s , Indianapolis)  86  41.  ^"Hmr s p e c t r u m  87  42.  P o s s i b l e mechanism f o r t h e f o r m a t i o n o f l e u r o s i n e from 3 , 4 ' - a n h y d r o v i n b l a s t i n e  91  P o s s i b l e mechanism f o r t h e f o r m a t i o n o f Catharine from l e u r o s i n e  92  Infrared (83)  95  of synthetic  (73)  78 (74) ...  Catharine  1  43.  44.  spectrum  of  80  3R-hydroxyvinamidine  45.  ^"Hmr s p e c t r u m  o f 3R-hydroxyvinamidine  ( 8 3 ) ..  96  46.  ^Hmr  of 3R-acetoxyvinamidine  (84) ..  98  47.  "^Hmr s p e c t r u m  of triacetate  48.  ''"Hmr s p e c t r u m  of tetracetate  49.  "4lmr s p e c t r u m o f a l d e h y d e 90  103  50.  Infrared  105  51.  "*"Hmr s o e c t r u m  spectrum  spectrum  86 87  o f 1 9 - o x o l e u r o s i n e (91) .. 1  o f 1 9 ' - o x o l e u r o s i n e (91)  100 101  106  - viii -  Figure  Page  52.  ^Hmr spectrum o f vinamidine (sample was obt a i n e d from the E l i L i l l y L a b o r a t o r i e s , Indianapolis)  10 8  53.  "''Hmr spectrum  of s y n t h e t i c vinamidine  109  54.  "*"Hmr spectrum dine (95)  of  19 -oxo-4'-deoxyleurosi1  I l l  55.  P o s s i b l e mechanism f o r the formation of vinamidine from 4'-deoxyleurosidine  112  56.  P o s s i b l e mechanism f o r the formation of 3R-hydroxyvinamidine from 3 ,4'-anhydrovinblastine  113  57.  P o s s i b l e mechanism f o r the formation of 3R-hydroxyvinamidine from l e u r o s i n e  113  58.  I n f r a r e d spectrum of the product obtained from the iodine/sodium bicarbonate oxidat i o n of 4 - d e o x y l e u r o s i d i n e  116  '''Hmr spectrum of the product obtained from the iodine/sodium becarbonate o x i d a t i o n of 4'-deoxyleurosidine  117  1  1  59.  60.  61. 62. 63.  Comparison of u l t r a v i o l e t s p e c t r a of 1 9 ' — oxoleurosine (A) and 19',22-dioxoleurosine (B) . ... 1  Hmr spectrum  of 19 ,22-dioxoleurosine 1  I n f r a r e d spectrum (98) ''"Hmr spectrum 99  (98).  119 120  of 19 ,22-dioxoleurosine 1  of N -desmethyl d e r i v a t i v e f  121 122  - ix -  L I S T OF TABLES Table 1.  Page Comparison o f some fragments e x h i b i t e d i n the mass s p e c t r a of 16,18S-dicarbomethoxycleavamine (39), 16,18S-dicarbomethoxy-3R,— 4S-epoxydihydrocleavamine (40), and 16,18S— dicarbomethoxy-4S-dihydrocleavamine (41) ....  37  13 2.  Comparison of Cmr chemical s h i f t s o f l e u r o s i n e , and the epoxides 40 and 42  3.  Comparison of the chemical s h i f t of the C^ ;— hydrogen of various 16,18S-dicarbomethoxycleavamine d e r i v a t i v e s  59  Study of the a e r i a l and t - b u t y l hydroperoxide o x i d a t i o n s of 3 ' , 4 - a n h y d r o v i n b l a s t i n e , l e u r o s i n e , and d e r i v a t i v e s thereof  88  4.'  38  g  1  5.  Comparison of some c h a r a c t e r i z a t i o n data f o r 19'-oxoleurosine (91), 19'-oxovinblastine (96), and 19 '-oxoleurosidine(97) 115  -  x -  ACKNOWLEDGEMENTS  I Kutney  wish  t o e x p r e s s ray a p p r e c i a t i o n t o P r o f e s s o r J . P .  f o rh i s guidance  throughout  the course  of  this  research. I would and  staff  also  like  t o thank  who p r o v i d e d  me w i t h  the various  technicians  the necessary  spectroscopic  data. Furthermore, the of  various working  B.R. W o r t h script  post-doctoral with.  as w e l l  preparing  support.  like  t o express  f e l l o w s who  In particular,  f o rh i s help  Finally, in  I would  gratitude to  I had the pleasure  I would  like  i n the preparation  t o thank  of this  manu-  as i n c a r r y i n g o u t t h e r e s e a r c h . I would  this  like  manuscript  t o thank as w e l l  my w i f e  f o rher help  as f o rh e r g e n e r a l  -  1  -  INTRODUCTION  1.1  General Background Cancer Chemotherapy has over the past 30-40 years  evolved i n t o an e f f e c t i v e means of treatment;  and although the  somewhat l i m i t e d number of cancers t r e a t a b l e by t h i s modality has remained a major drawback, the f u r t h e r development of t h i s 1-5 area continues w i t h the  promise of g r e a t e r p o t e n t i a l .  H i s t o r i c a l l y , the e v o l u t i o n of a n t i n e o p l a s t i c chemotherapy began with a s e r i e s of independent  observations.  Thus, i n 19 39, Loeser^ reported the use of androgens i n the treatment of b r e a s t cancer, and i n 19 41, Huggins and Hodges, reported that estrogen was cancer.  By 1946,  Rhoads  7  e f f e c t i v e i n the treatment of p r o s t a t i c and many others had noted the a n t i c a n c e r 9  e f f e c t s of n i t r o g e n mustard.  In 19 48, Farber and a s s o c i a t e s ,  discovered the p o t e n t i a l of a n t i f o l i c a c i d compounds, and later,demonstrated the c l i n i c a l value of actinomycin D. i n 1955, was  the leukopenic e f f e c t of the catharanthus  noted by Beer and  10  Then,  alkaloids  Noble.  These miscellaneous r e s u l t s provided a s o l i d  founda-  t i o n , as w e l l as much impetus f o r f u r t h e r development.  Conse-  quently, the o b s e r v a t i o n that mustard g a s — b i s (2-chloroethyl)  -  2 -  s u l f i d e — h a d the a b i l i t y to produce leukopenia, l e d to the d i s covery of the l e s s t o x i c n i t r o g e n mustard.  A f u r t h e r consequence  of t h i s was the development of a s e r i e s of analgous a n t i c a n c e r  HN(CH CH CI) 2  2  H C0 SOH C(CH ) CH OS0 CH  2  2  3  Nitrogen mustard  2  2  S 2  2  2  2  3  Busulfan  0  P-N(CH CH CI)  2  — CJ N  — P  2  N Cyclophosphamide Thiotepa  Fig.  1.  S t r u c t u r e of some c l i n i c a l l y  used a l k y l a t i n g agents. 12  compounds:  the " a l k y l a t i n g agents' ( F i g . 1 ) .  Farber's  o b s e r v a t i o n , t h a t f o l i c a c i d treatment of anemic leukemic children resulted the  i n a worsened c o n d i t i o n , e v e n t u a l l y l e d to  development of m e t h o t r e x a t e — a c l i n i c a l drug which  still  13  f i n d s wide usage.  In t u r n , methotrexate has become one of 14  many a n t i m e t a b o l i t e drugs ( F i g . 2 ) .  Furthermore, the observa-  t i o n , that actinomycin D possessed i n t e r e s t i n g a n t i c a n c e r prop e r t i e s , i n i t i a t e d a search f o r other a n t i b i o t i c s which might possess s i m i l a r p o t e n t i a l .  The r e s u l t of t h i s was the i s o l a t i o n  - 3-  Drug  N ^ J l  Metabolite  / \ N  i'olic C  H  acid,  R=OH,  N  2 1  Methotrexate,  R=NH , 2  R =CH 1  3  OH  OH  O ^ N  H  H  Uracil  5-Fluorouracil  SH  NH.  "CO  N  Adenine  6-Mercaptopurine  O  H.N 2  NH  NH.  HN 1  I  2  OH Urea  Hydroxyurea  Fig.  2 .  analogs.  Some a n t i m e t a b o l i t e  cancer drugs  and t h e i r m e t a b o l i t e  R =H 1  - 4 -  9H(Me)  OC  2  -CH N Me i sarcosine L-proline i  CHfMeL CH•CO—i i NMe I  sarcosine i L-proline  '— HC  Actinomycin D  Kidamycin  Fig.  3.  Structures  of some c y t o t o x i c  antibiotics.  - 5-  and/or synthesis  of s e v e r a l new and promising compounds belonging 15-17  to the a n t i b i o t i c group  (Fig. 3).  In a s i m i l a r manner, the e a r l y work of Noble, Beer, 18 19 and  Cutts  '  cancer agents:  stimulated  the development of a new c l a s s o f a n t i -  the catharanthus a l k a l o i d s .  These workers, while  i n v e s t i g a t i n g the reputed hypoglycemic p o t e n t i a l of e x t r a c t s of Catharanthus roseus noted that i n j e c t i o n s of the e x t r a c t s r a t s produced a r a p i d onset of p e r i p h e r a l bone marrow depression.  Eventually,  into  granulocytopenia and  t h i s a c t i v i t y was l o c a l i z e d  to the a l k a l o i d f r a c t i o n s with the subsequent r e s u l t being the i s o l a t i o n of v i n b l a s t i n e  (1). Meanwhile, a t the L i l l y 20 21  t o r i e s , work c a r r i e d out by Svoboda of the r e l a t e d c y t o t o x i c (3), and l e u r o s i d i n e discovery,  '  labora-  l e d to the i s o l a t i o n  a l k a l o i d s v i n c r i s t i n e (2), l e u r o s i n e  (4). Within a decade o f the o r i g i n a l  v i n b l a s t i n e and v i n c r i s t i n e had advanced to c l i n i c a l  22 use;  and as a r e s u l t , the study of the catharanthus  alkaloids  q u i c k l y became an area of t o p i c a l i n t e r e s t t o a multitude o f researchers.  2  1, R=CH  3  (Vinblastine)  2, R=CHO ( V i n c r i s t i n e )  - 6 -  3 (Leurosine)  4 (Leurosidine)  1.2 S t r u c t u r e E l u c i d a t i o n of V i n b l a s t i n e , V i n c r i s t i n e , Leurosine, and L e u r o s i d i n e I n i t i a l l y , the chemistry of the catharanthus  alkaloids  c o n s i s t e d of work done towards the s t r u c t u r e e l u c i d a t i o n of the aforementioned  'dimeric'* a l k a l o i d s .  Thus, by means of l i t h i u m  23 aluminum hydride r e d u c t i o n , Neuss e t a l were able to convert *The term 'dimeric' a l k a l o i d s i s s t r i c t l y speaking a misnomer, however, i t has been p o p u l a r l y used i n t h i s area along with other terms such as b i s i n d o l e a l k a l o i d s to r e f e r to these compounds.  - 7 -  v i n b l a s t i n e and v i n c r i s t i n e t o the pentahydroxy  d e r i v a t i v e 5.  F u r t h e r c o r r e l a t i o n between v i n b l a s t i n e and v i n c r i s t i n e was o b t a i n e d by the development of a cleavage r e a c t i o n .  Hence,  treatment o f v i n b l a s t i n e o r v i n c r i s t i n e i n h o t c o n c e n t r a t e d hydrochloric  acid containing  stannous c h l o r i d e and m e t a l l i c t i n 23 24  l e d t o p r o d u c t s a r i s i n g from cleavage o f the C^g'-C^ bond.  '  In the case o f v i n b l a s t i n e these p r o d u c t s were velbanamine ( 6 ) ,  5  and d e a c e t y l v i n d o l i n e  ( 7 ) , w h i l e i n the case o f v i n c r i s t i n e , the  p r o d u c t s o b t a i n e d were velbanamine doline  (8).  and N ^ - d e s m e t h y l d e a c e t y l v i n -  T h e r e f o r e , the d i f f e r e n c e between v i n b l a s t i n e and  v i n c r i s t i n e was a t t r i b u t a b l e t o a d i f f e r e n c e  i n the d i h y d r o -  i n d o l e p o r t i o n o f the 'dimers'.  These r e s u l t s , coupled w i t h a 25 d e t a i l e d mass s p e c t r a l study performed by Biemann e t a l , l e d t o reasonable s t r u c t u r a l assignments. However, i t was n o t u n t i l the 26 x-ray a n a l y s i s c a r r i e d out by M o n c r i e f and Lipscomb  i n 1965 t h a t  the a b s o l u t e s t r u c t u r e of v i n c r i s t i n e (as i t s methiodide  deriv-  - 8-  ative) was c o n c l u s i v e l y  determined.  c r i s t i n e had been chemically absolute  Since v i n b l a s t i n e and v i n -  r e l a t e d , the determination of the  s t r u c t u r e of v i n c r i s t i n e meant that the absolute  of v i n b l a s t i n e was a l s o Therefore,  structure  established.  with the determination of the s t r u c t u r e s  of v i n b l a s t i n e and v i n c r i s t i n e , greater emphasis was placed on the s t r u c t u r e determination of l e u r o s i n e and l e u r o s i d i n e . Accordingly,  a comparison of s p e c t r a l data i n d i c a t e d that v i n -  b l a s t i n e and l e u r o s i n e were c l o s e l y r e l a t e d .  Reductive cleavage  of l e u r o s i n e w i t h stannous c h l o r i d e and m e t a l l i c t i n i n hot concentrated h y d r o c h l o r i c  a c i d l e d t o the i s o l a t i o n o f cleavamine  24 (9) and d e a c e t y l v i n d o l i n e  (7),  therefore  pinpointing  the d i f f e r -  ence between v i n b l a s t i n e and l e u r o s i n e to the indole p o r t i o n of  27 the  'dimers'.  High r e s o l u t i o n mass spectrometry  the molecular formula o f l e u r o s i n e as C.,H _ N.C' [  C  established —  two hydrogens  - 9 -  l e s s than that f o r v i n b l a s t i n e .  Moreover,  from the fragmenta-  t i o n p a t t e r n e x h i b i t e d i n the mass spectrum, the presence o f an oxygen f u n c t i o n a l i t y i n the p i p e r i d i n e r i n g o f the i n d o l e port i o n was i n d i c a t e d ( F i g . 4). The c o n c l u s i o n  that t h i s  oxygen 13  f u n c t i o n a l i t y was a 3',4'-epoxide was corroborated  by a  C28  magnetic resonance study performed by Wenkert and a s s o c i a t e s However, the unambiguous assignment of the epoxide s t e r e o 29 chemistry was determined only r e c e n t l y by Kutney and Worth , who using the r e c e n t l y developed method of coupling v i n d o l i n e (10)  F i g . 4.  A s i g n i f i c a n t pathway f o r the fragmentation of l e u r o s i n e  i n the mass spectrometer.  - 10 -  and  catharanthine (11) to a f f o r d dimeric  compounds of the v i n -  30-33 blastine-type*,  synthesized  3R, 4S-epoxydihydrocatharanthine  (12)  and coupled t h i s compound with v i n d o l i n e to a f f o r d l e u r o -  sine  ( F i g . 5).  Since the absolute  c o n f i g u r a t i o n of the s t a r t i n g  catharanthine d e r i v a t i v e was known, the stereochemistry epoxide i n l e u r o s i n e was The followed  of the  established.  determination of the s t r u c t u r e of l e u r o s i d i n e  a s i m i l a r path.  A comparison of s p e c t r a l data between  l e u r o s i d i n e and v i n b l a s t i n e showed a c l o s e r e l a t i o n s h i p between the two a l k a l o i d s .  Reductive cleavage  (stannous c h l o r i d e and  m e t a l l i c t i n i n hot concentrated h y d r o c h l o r i c deacetylvindoline  acid)  yielded  (7) and a new i n d o l e fragment vinrosamine (13),  thus p i n p o i n t i n g the d i f f e r e n c e between the two 'dimeric' l o i d s to the i n d o l e  alka-  portion.  13 High r e s o l u t i o n mass spectrometry i n d i c a t e d l e u r o s i d i n e was isomeric w i t h v i n b l a s t i n e  that  (C^gH^gN^Og), and o r i -  g i n a l l y , based on these e a r l y r e s u l t s l e u r o s i d i n e and vinrosamine were i n c o r r e c t l y assigned as the secondary a l c o h o l s 14 and 15  *The coupling of v i n d o l i n e and catharanthine to a f f o r d 'dimers' of the v i n b l a s t i n e - t y p e c o n s t i t u t e d an important discovery and w i l l be discussed i n some d e t a i l i n s e c t i o n 1.4.  -  F i g . 5.  11  -  O u t l i n e of Kutney and Worth's s t r u c t u r e - p r o o f of  leurosine.  - I r -  respectively.  However, these s t r u c t u r e s were l a t e r  (to the t e r t i a r y a l c o h o l s  4 and  13)  corrected  on the b a s i s of f u r t h e r  14  chemical evidence as w e l l as a  C- magnetic resonance study  28 performed by Wenkert e t a l .  More r e c e n t l y , P o t i e r  and  35 Langlois  were able to confirm  unambiguous s y n t h e s i s . * anthine was  t h i s assignment by means of  Thus, v i n d o l i n e  (10)  and  dihydrocathar-  (16) were coupled to y i e l d the enamine 17.  not i s o l a t e d , but was  the 4',5'-diol  an  This enamine  reacted with osmium t e t r o x i d e to a f f o r d  18, which on treatment with sodium borohydride  yielded leurosidine  ( F i g . 6).  only have l e d to the C-4 of the other centers was  1  Since t h i s r e a c t i o n sequence could  a l c o h o l , and  since the  configuration  known, l e u r o s i d i n e s s t r u c t u r e 1  was  e s t a b l i s h e d as the C-4 epimer of v i n b l a s t i n e . *This synthesis was an a p p l i c a t i o n of the 'biogenetic approach to the synthesis of the 'dimeric' a l k a l o i d s — an approach which i s discussed i n s e c t i o n 1.4. 1  1  -13-  Leurosidine  Fig.  6.  Potier'and L a n g l o i s  1  s t r u c t u r e - p r o o f of l e u r o s i d i n e .  - 14 -  V i n b l a s t i n e , v i n c r i s t i n e , l e u r o s i n e , and  leurosidine  represented some of the f i r s t a l k a l o i d s i s o l a t e d from roseus.  Catharanthus  F u r t h e r i n v e s t i g a t i o n s i n t h i s area have continued and  to date c l o s e to one hundred i n d o l e a l k a l o i d s of v a r y i n g types have been i s o l a t e d .  Among these have been s e v e r a l new  'dimeric 36 —  1  38  a l k a l o i d s , some of which have shown c y t o t o x i c p r o p e r t i e s . 1.3 Studies Aimed at the Synthesis of the 'Dimeric' Catharanthus A l k a l o i d s — the 'Chloroindolenine Approach'. As the s t r u c t u r e s of v a r i o u s catharanthus  alkaloids  began to emerge, so too d i d i n t e r e s t i n t h e i r s y n t h e s i s .  In  p a r t i c u l a r , the s y n t h e s i s of the v i n b l a s t i n e - v i n c r i s t i n e type r a p i d l y became a dominant concern, f o r not only were these novel compounds s t r u c t u r a l l y complex and thus s y n t h e t i c a l l y c h a l l e n g i n g , but a l s o they were expensive and d i f f i c u l t to i s o l a t e possessed tremendous a n t i c a n c e r p o t e n t i a l —  a l l of which com-  bined to make them extremely a t t r a c t i v e as s y n t h e t i c Consequently, vindoline tial  and  targets.  two of the major a l k a l o i d s of C. roseus,  (10) and catharanthine (11), became important as poten-  synthons of the 'dimeric' a l k a l o i d s .  10  (Vindoline)  were r e a d i l y a v a i l a b l e  Both of these compounds  11  (Catharanthine)  (from p l a n t m a t e r i a l ) and featured de-  - 15 -  s i r a b l e f u n c t i o n a l i t y as w e l l as s u i t a b l e carbon  skeletons.  Moreover, s i n c e both of these a l k a l o i d s had r e c e n t l y been syn39-42 thesized,  any n a t u r a l products s y n t h e t i c a l l y d e r i v e d from  them could f o r m a l l y be considered as t o t a l l y s y n t h e t i c . The  f i r s t s y n t h e t i c e f f o r t s aimed at the dimeric  a l k a l o i d s were based on the ' c h l o r o i n d o l e n i n e approach'. 43 Kutney e t a l  had shown i n 1970,  that when a s o l u t i o n of  catharanthine i n hot g l a c i a l a c e t i c a c i d was  t r e a t e d with sodium  borohydride  a high y i e l d of 18S-carbomethoxycleavamine (19) c o u l d  be obtained  ( F i g . 7).  T h i s product possessed  bon s k e l e t o n of the i n d o l e p o r t i o n of the  the r e q u i r e d c a r -  'dimeric' a l k a l o i d s  and r e q u i r e d only appropriate f u n c t i o n a l i z a t i o n of the 4-double bond and subsequent condensation  3,  ( v i a the c h l o r o i n d o l e -  nine) w i t h v i n d o l i n e to provide access to a number of dimeric alkaloids  ( F i g . 8). The  f i r s t requirement,  the f u n c t i o n a l i z a t i o n of the  3,4-double bond, proved d i f f i c u l t .  This d i f f i c u l t y was  due  to  the competitive r e a c t i v i t i e s of the i n d o l e system and the b a s i c n i t r o g e n towards e l e c t r o p h i l i c reagents agents).  (e.g. many o x i d i z i n g  In general, attempted f u n c t i o n a l i z a t i o n of the  bond e i t h e r l e d to complex r e a c t i o n mixtures 44 which the double bond was  still  intact.  or to products i n  However, two  r e a c t i o n s of the 3,4-double bond were a t t a i n e d . was  hydrogenation  using a platinum c a t a l y s t .  duct obtained i n good y i e l d was 43 cleavamine  (20).  hydroboration.  double  One  selective  of these  The e x c l u s i v e pro-  4S-ethyl-18S-carbomethoxydihydro-  The other s u c c e s s f u l r e a c t i o n achieved  was  Oxidation of the intermediate alkylborane with  -16-  i  CO Me  19  F i g . 7.  (18S-Carbomethoxycleavamine)  Mechanism f o r the formation of 18S-carbomethoxy-  cleavamine from c a t h a r a n t h i n e .  - 17 -  F i g . 8.  General o u t l i n e of the 'chloroindolenine  approach'.  -  alkaline  hydrogen  derivative  21  as  peroxide  18  -  a f f o r d e d i n good  a mixture  of  C-18  yield 44  epimers.  the  Therefore,  20  least  to  a  limited  cleavamine  at  21  The a  3S-hydroxy  extent,  second  the  first  requirement,  derivative,  was  requirement  the  was  coupling of  s t u d i e d by  a number  satisfied.  vindoline of  and  research-  45-49 ers.  was  One  carried  of  out  by  the  most  Kutney  thorough 49 et  investigations  al.  These workers  4S-ethyl-18S-carbomethoxycleavamine a  solution  obtained of  a  changes  of  single  time,  products  —  acid,  showed  required product  X-ray  and  thermore,  addition  was  this  analysis  the  cleavamine the  (solvent,  temperature,  technique) products  this  was  of  extension of  to the  that  of  by  22  a t C-18 the  other  the  1  (23).  of  'natural  dimeric  the  dimer  et a l ^  Thus,  coupled  although at  the  coupled  dimers'.  c h l o r o i n d o l e n i n e method  concen-  stereo-  Clardy  were  regardless  original  completely  product  derivative  no  with  chloride  Moreover,  configuration  opposite  treating  (Fig. 9).  reaction of  hydrogen  18'-epi-4'-deoxyleurosidine  centers, 23  and  Hence,  i t t o be  vindoline  conditions  by  area  c h l o r o i n d o l e n i n e (22)  methanolic  excluding degradation  produced.  selective.  i n 1.5%  dimeric product  in reaction  tration,  were  vindoline  in this  to  a  Furnumber  —  -19-  F i g . 9.  Condensation of the c h l o r o i n d o l e n i n e of  18S-carbomethoxydihydrocleavamine with v i n d o l i n e .  4S-ethyl—  - 20 -  of other cleavamine  d e r i v a t i v e s l e d to a v a r i e t y of new  products, a l l of which possessed  the  'dimeric'  'unnatural' stereochemistry  49 at C-18 .  Therefore, as a g e n e r a l and v e r s a t i l e route to the  1  s y n t h e s i s of the  'dimeric' catharanthus  dolenine approach' as o u t l i n e d was cations of t h i s approach may  a l k a l o i d s , the 'chloroin-  a failure.  However, m o d i f i -  y e t provide some measure of success,  and f u r t h e r study towards t h i s end  continues.  1.4 Studies Aimed at the Synthesis of the 'Dimeric' Catharanthus A l k a l o i d s — t h e 'Biogenetic Approach'.* The approach' was  stereochemical outcome of the 'chloroindolenine u n f o r t u n a t e l y not p r e d i c t a b l e .  A p r i o r i considera-  t i o n s r e q u i r e d a conformational a n a l y s i s of the nine-membered r i n g of carbomethoxycleavamine —  a complex r i n g system with  p o t e n t i a l f o r conformational f l e x i b i l i t y , amenable to a n a l y s i s .  Conversely,  and thus not  readily  the p e n t a c y c l i c Iboga s k e l e t o n  of catharanthine represented a r i g i d i n f l e x i b l e moiety; and i t was  t h i s f i x e d geometry which l e d to the development -of the  'biogenetic approach* by both P o t i e r Thus, treatment  of catharanthine  30 31 32 33 ' and Kutney '  (11) with  m-chloroperbenzoic  a c i d i n methylene c h l o r i d e s o l u t i o n l e d to the formation of the N^-oxide 24.  Treatment of t h i s product with  trifluoroacetic  anhydride  (modified Polonovski r e a c t i o n ) r e s u l t e d i n the f r a g -  mentation  of the Ccj-C^g bond with concomitant  5 1  generation of a  *This approach has been termed 'biogenetic' on the b a s i s of popular b e l i e f and c i r c u m s t a n t i a l evidence, however the conc l u s i v e proof of t h i s c o n v i c t i o n has y e t to be accomplished.  -  cationic  center  appropriate blastine  C-18,  and  conditions,  good  (26)  approach  was  at  could  the  coupling.  either  concerted  seco-catharanthine quired  outcome.  trated  solution  up,  by  at  blastine  was  outcome  of  higher  i n which  more  dilute  iate  was  low  (2 5 a - » 2 5 b )  prior  the  The constituted  versatility tuting  with  was  various  syntheses  of  connection  method,  the  various  vindoline  at  31  '  with  however  the  be  their  '  53  '  52  the  were  this geometry  established  conformation lead  to  reaction  -50°C) the  of  fragmentation  in  other  hand,  that or  of  the  afforded,  a the  re-  concen-  after  work-  i n more  dilute  18'-epi-3',4'-anhydrovinof  the  r a t i o n a l i z e d by temperatures  the  stereochemical  assuming  that  reactive  intermed-  'product-like  1  in  conformation  vindoline.  in  the  the  The  'biogenetic  synthesis  several  general,  the  found  derivatives  replacement was  of  approach  not 35  as  that  into  the  substi-  in place  of  catharanthine successful.  (previously  elucidation)  disappointing  of  1  'dimeric'  investigations  i t was  leurosidine  structure  yields  out  reversal  derivatives and  the  would  40°C),  stable  In  clearly  aspidospermine  33  functionalized 29  on  launched  nucleophilic possible.  (ca  (ca  with  reaction.  had  success  reactant-like'  during  r e a l i z a t i o n of  and  1  under  3',4'-anhydrovin-  The  frozen  while  breakthrough  of  vindoline,  'reactant-like' was  higher  coupling  leurosine  models  complete  a more  alkaloids, of  ( F i g . 10).  carrying  could  successful  a major  catharanthus  The  adopt to  of  temperatures  reaction  to  (50-60%)  temperatures  solutions  able  yields  intermediate  isolated. the  of  vindoline  Accordingly, at  presence  retention  3',4'-anhydrovinblastine,  solution  in  to  -  the  Molecular  attack,  coupling  in  obtained  attributable  during  step-wise  be  21  were  mentioned  attained  (5-30%).  The  by  this  Furthermore,  -  Me  22  -  i  C0 Me 2  26  (34'-Anhydrovinblastine)  25b  F i g . 10.  'Biogenetic-type' c o u p l i n g of catharanthine-N^-oxide  with v i n d o l i n e .  (24)  - 23 -  r e a c t i o n of other catharanthine d e r i v a t i v e s such as the epoxide  27  and  the  29  and  30 r e s p e c t i v e l y . ^  lactone  2 8 afforded 4  ^  only  rearrangement products such as  As such, the u t i l i t y of the  'biogenetic  approach' i n regard to the use of f u n c t i o n a l i z e d catharanthine d e r i v a t i v e s appeared l i m i t e d . r e g i o s p e c i f i c reactions v i n b l a s t i n e has  Consequently, the development of  f o r the f u n c t i o n a l i z a t i o n of 3',4'-anhydro-  become an area of current  interest.  7  30 1.5  V i n c r i s t i n e Derivatives Although the d i s c u s s i o n  of the chemistry r e l a t i n g to  - 24  the  'dimeric'  a l k a l o i d s has  -  so f a r been concerned with v i n b l a s -  t i n e - l i k e d e r i v a t i v e s , the conversion of v i n b l a s t i n e to v i n c r i s t i n e i s an important step and has  been accomplished by  Jovanovics  57 et a l  .  Thus, treatment of a s o l u t i o n of v i n b l a s t i n e s u l f a t e i n  acetone at -50°C with chromic a c i d a f f o r d e d  vincristine.  of t h i s r e a c t i o n to l e u r o s i n e was  also successful 58 the N -formyl d e r i v a t i v e 31 i n good y i e l d .  and  Extension  provided  31  This r e g i o s p e c i f i c o x i d a t i o n development i n the chemistry of the a l k a l o i d s since i t c o n s t i t u t e d of v i n c r i s t i n e analogs. d e r i v a t i v e s had  represented a s i g n i f i c a n t  'dimeric'  catharanthus  the only method f o r the production  Thus, although various  been s u c c e s s f u l l y used i n the  aspidospermine 'biogenetic  approach' to y i e l d v i n b l a s t i n e analogs, the corresponding produc31 t i o n of v i n c r i s t i n e analogs was was  not s u c c e s s f u l .  not s u r p r i s i n g s i n c e i n the s u c c e s s f u l  aspidospermine u n i t was  coupling  This r e s u l t reactions  the  e i t h e r a n u c l e o p h i l i c methoxyaniline or  a n i l i n e system, while i n the unsuccessful  case the e l e c t r o n  don-  -  25  -  a t i n g amino group was  replaced  N ~ f o r m y l group.  r e s u l t a n t reduced r e a c t i v i t y towards  The  a  by the e l e c t r o n withdrawing  e l e c t r o p h i l i c aromatic s u b s t i t u t i o n of the N^-formyl d e r i v a t i v e thus provided  a r a t i o n a l e f o r the observed lack of r e a c t i o n .  Further dation was  due  s i g n i f i c a n c e of the r e g i o s p e c i f i c chromate o x i -  to the d i f f e r e n t spectrum of a c t i v i t i e s e x h i b i t e d  v i n b l a s t i n e and v i n c r i s t i n e .  These two  potent a n t i c a n c e r  by  agents  although s t r u c t u r a l l y very s i m i l a r were a c t i v i t y - w i s e very 59  different.  Moreover, the d i f f e r e n c e i n b i o l o g i c a l a c t i v i t y  r e s u l t i n g from the m o d i f i c a t i o n group was  further exemplified  of an N -methyl to an a  by 22-oxoleurosine  N -formyl a  (31), which  6 0  has  advanced to c l i n i c a l t r i a l s i n Hungary.  other hand, has  not been found c l i n i c a l l y  of t h i s r e a c t i o n to a v a i l a b l e s y n t h e t i c p o t e n t i a l f o r the production 1.6  of new  Leurosine on  useful.  The  extension  'dimers' shows great  anticancer  agents.  Scope of Current Research Thus, i t has been seen t h a t the chemistry of  'dimeric'  the  catharanthus a l k a l o i d s has  the  developed over the  twenty years to a p o i n t where the p o t e n t i a l synthesis  past  of a  l a r g e number of analogs and n a t u r a l products i s at hand. ever, the complete attainment of these goals  still  How-  requires  f u r t h e r development of the chemistry of the i n d o l e p o r t i o n available synthetic  'dimers'.  mentioned 'biogenetic  approach' by appropriate  of 3',4'-anhydrovinblastine of a number of n a t u r a l  Indeed, extension  of the  the of  previously  functionalizations  (26) would lead to the  'dimers', some of which are  synthesis illustrated  -  2  6  -  Vinblastine,  R ^ O H , R = C H C H , R =R =H  Leurosidine,  R-^CH^H^  Leurosine,  2  1  2  Vincadioline,  3  2  R  3  2  4  =  4  R =R =H 3  '  0  4  R  1  3  4  2  11.  Some  'dimeric'  R =R.=OH, R =CH CH  H  2  alkaloids.  3  , R =H  Vind.  Vinamidine  catharanthus  =  2  (Catharinine)  Fig.  3  R =R =R =H, R = C H C H  3  Catharine  3  2  H C0 C  Vind.  3  R =OH,  R =CH CH , ^  Deoxyvinblastine,  H C0 C  2  - 27 -  i n F i g . 11.  Furthermore,  p r e v i o u s l y developed  attainment of t h i s r e s u l t coupled with  chemistry would provide access to a number  of s t r u c t u r a l l y d i v e r s e analogs —  an important requirement f o r  the production of new drugs as w e l l as f o r the development of structure-activity  relationships.  A l t e r n a t i v e l y , another area of recent i n t e r e s t i n v o l v e s f u r t h e r e v a l u a t i o n o f the ' c h l o r o i n d o l e n i n e approach'.  The  continuing i n t e r e s t i n t h i s approach revolves around the p r e p a r a t i o n of cleavamine  d e r i v a t i v e s whose geometry would be such that 'dimer-  i z a t i o n ' with v i n d o l i n e c o u l d occur only from the oC-face to a f f o r d the  ' n a t u r a l ' stereochemistry 'dimer'.  Thus, the 4S,18R-ether 32  and the lactone 33 represent p o t e n t i a l precursors of v i n b l a s t i n e i n that t h e i r geometry should block access to t h e / 3 - f a c e l  32  33  In summary, the f u r t h e r i n v e s t i g a t i o n of the 'chloroi n d o l e n i n e approach',  the development of the chemistry o f 3',4'-  a n h y d r o v i n b l a s t i n e , and the production of v i n b l a s t i n e - v i n c r i s t i n e analogs remain as c u r r e n t t o p i c s of research.  Further discussion  of these various aspects, combined with an o u t l i n e of the work performed  i n attempting to achieve these ends, c o n s t i t u t e s the  next chapter of t h i s  thesis.  - 28 -  DISCUSSION  2.1  Further  Evaluation of the C h l o r o i n d o l e n i n e  Although previous  attempts at coupling  Approach .  chloroindole-  nines of s e v e r a l cleavamine d e r i v a t i v e s with v i n d o l i n e had s u l t e d i n the formation (at C-18')  49  of  the question  'unnatural  stereochemistry  'dimers'  of whether t h i s outcome could  reversed by u t i l i z a t i o n of other rivatives s t i l l  1  'more appropriate'  remained as a v a l i d c o n s i d e r a t i o n .  re-  be  cleavamine  de-  Indeed,  there e x i s t e d some j u s t i f i c a t i o n f o r the b e l i e f that d e r i v a t i v e s of velbanamine  (e.g. 34 and  35) might be such compounds.  The  b a s i s f o r t h i s c o n v i c t i o n was  a r e p o r t that v i n b l a s t i n e could  be cleaved  s u l f u r i c a c i d to y i e l d  i n hot aqueous 40%  deacetyl-  HO  R  34, R=H,  R]_=C0 Me 2  35, R=C0 Me, 2  vindoline  (7) and  R  R]_=H  an i n d o l e fragment whose s t r u c t u r e was  assigned  - 29 -  Vinblastine  Deacety1vindoline  (7)  H0 C 2  37  F i g . 12.  P o s s i b l e mechanism f o r the formation of  37 from v i n b l a s t i n e .  4S,18R-ether  -  as the 4S,18R-ether 37 product  30 -  ( F i g . 12). The formation of t h i s  could be explained by a sequence i n v o l v i n g p r o t o n a t i o n  of the e l e c t r o n - r i c h methoxyaniline  system of v i n b l a s t i n e  followed by i n d o l e - a s s i s t e d cleavage of the 2.8'~ 15 C  The  C  bon  ^*  r e s u l t a n t intermediate 36 could presumably be transformed  i n t o the 4S,18R-ether by i n t r a m o l e c u l a r attack of the 4S-hyd r o x y l group onto the C-18 p o s i t i o n . Since the intermediate  36 should a l s o be a v a i l a b l e  from the c h l o r o i n d o l e n i n e * 38/ i t t h e r e f o r e followed that 1 8 — carbomethoxyvelbanamine  (3_4 o r 35) should v i a i t s c h l o r o i n d o l e n i n e  37 be a precursor of the corresponding  ether 32.  Furthermore,  MeO C  37,  R=C0 Me 2  38, R=C0 H 2  the geometry of t h i s ether was such that formation of the ' n a t u r a l ' stereochemistry would be expected.  'dimer' (on coupling with v i n d o l i n e )  Therefore, coupling of 18-carbomethoxyvelban-  amine c h l o r o i n d o l i n e (37) with v i n d o l i n e should  (assuming the  *The c h l o r o i n d o l e n i n e ' d i m e r i z a t i o n ' i n v o l v e d the formation of an intermediate (analgous t o 36) according to the mechanism proposed by A.M. Treasurywala.44  - 31 -  intermediacy  of 32) a f f o r d v i n b l a s t i n e .  More g e n e r a l l y , the  synthesis of velbanamine d e r i v a t i v e s * followed by coupling of t h e i r chloroindolenines v i a b l e route  with v i n d o l i n e appeared a p o t e n t i a l l y  to v i n b l a s t i n e , v i n c r i s t i n e , v i n c a d i o l i n e , and v a r i -  ous novel analogs.  Accordingly,  e x p l o r a t i o n of t h i s route  was  undertaken. 18S-Carbomethoxycleavamine  (19), a v a i l a b l e from  43 catharanthine  i n good y i e l d  was chosen as the s t a r t i n g m a t e r i a l .  19, R=H 39, R=C0 Me 2  Thus, s u c c e s s f u l u t i l i z a t i o n of t h i s m a t e r i a l r e q u i r e d an overa l l hydration  of the 3,4-double bond.  However, d i r e c t func-  t i o n a l i z a t i o n of the 3,4-double bond of  1 9 had p r e v i o u s l y proven  d i f f i c u l t with p a r t of the problem being  a t t r i b u t e d to the com44  p e t i t i v e r e a c t i v i t y of the i n d o l e chromophore. of t h i s problem r e q u i r e d d e a c t i v a t i o n  Elimination  ( i . e . protection)  of the  *Buchi and co-workers have r e c e n t l y reported the synthesis of 18R-carbomethoxyvelbanamine. T h e i r r e s u l t s i n regard to dimeri z a t i o n with v i n d o l i n e have not y e t appeared.  - 32 -  i n d o l e chromophore and t h e r e f o r e 18S-carbomethoxycleavamine was t r e a t e d with potassium hydride i n t e t r a h y d r o f u r a n followed by methyl chloroformate to a f f o r d a f t e r p u r i f i c a t i o n ,  16,18S—  62 63 dicarbomethoxycleavamine* of  (39) i n 78% y i e l d .  '  The i d e n t i t y  t h i s compound was v e r i f i e d by i t s s p e c t r a l p r o p e r t i e s .  Thus,  high r e s o l u t i o n mass spectrometry confirmed the molecular f o r mula as 0 2 3 ^ 2 ^ 0 ^ . spectrum  The proton magnetic resonance ("'"Hmr)  ( F i g . 13) lacked a s i g n a l f o r an N -hydrogen, while  e x h i b i t i n g the f o l l o w i n g : & 5.77 (IH, doublet, J=6 Hz, C^g-H), ^5.37 (3H,  (IH, m u l t i p l e t , C -H)-, <$ 3 . 88 (3H, s i n g l e t , -0CH ) , 3  s i n g l e t , -OCH^).  the  3 . 53  F u r t h e r supportive evidence was gained  from the u l t r a v i o l e t spectrum (Fig. of  3  a m o d i f i e d i n d o l e chromophore.  14) which was i n d i c a t i v e F i n a l l y , the assignment of  c o n f i g u r a t i o n a t C-18 as S was based upon the low f i e l d  p o s i t i o n of the C^ -hydrogen resonance (& 5.77) g  spectrum.  i n the "'"Hmr  P r e v i o u s l y i t had been noted that the C^ -hydrogen g  resonance of 18R-carbomethoxycleavamine d e r i v a t i v e s occurred at  ~<c£ 4, while that of the corresponding  d e r i v a t i v e s occurred below & 5 ( i . e .  18S-carbomethoxy 4 4 65  to lower f i e l d ) .  '  The  reason f o r the observed d i f f e r e n c e was presumed due to the s p a t i a l proximity of the 18S-hydrogen to the N of  electrons  b  unshared p a i r  ( F i g . 15).  *This compound had p r e v i o u s l y been prepared i n t h i s l a b o r a t o r y I. Itoh and A.H. R a t c l i f f e using a d i f f e r e n t procedure.  3.88  - 34 -  log I  log <f  250  300  A.  F i g . 14.  nm  300  \  nm  Comparison of the u l t r a v i o l e t spectra of 18S-carbo-  methoxycleavamine  (A) and 16,18S-dicarbomethoxycleavamine (B).  18S c o n f i g u r a t i o n  F i g . 15.  250  18?. c o n f i g u r a t i o n  S p a t i a l r e l a t i o n s h i p of the 18-hydrogen to the  Nfc e l e c t r o n p a i r i n 18-carbomethoxycleavamine.  - 35 -  With the p r o t e c t e d d e r i v a t i v e 39 a v a i l a b l e , the next requirement was oxygenation o f the 3,4-double bond.  Earlier,  i t had been observed that 39 r e a d i l y underwent a u t o x i d a t i o n * to a f f o r d 16 ,18S-dicarbomethoxy-3R, 4S-epoxydihydrocleavamine as one of many products.  (4_0)  The s t r u c t u r e of t h i s compound was de-  duced from i t s s p e c t r a l p r o p e r t i e s as w e l l as v i a a c o r r e l a t i o n to a compound of known s t r u c t u r e .  A c c o r d i n g l y , the ^Hmr spectrum  40  (Fig. 16) i n d i c a t e d that the o l e f i n i c proton was no longer present.  High r e s o l u t i o n mass spectrometry e s t a b l i s h e d the  molecular formula as C H 2 N 0 ^ . 23  g  2  Comparison o f the mass s p e c t r a l  fragmentation p a t t e r n s of the o l e f i n 39, the dihydro d e r i v a t i v e 41**,and 40 (Table 1) i n d i c a t e d the presence of an oxide funct i o n a l i t y i n the p i p e r i d e n e r i n g .  The f a c t that t h i s func-  t i o n a l i t y was a 3,4-epoxide was based on a comparison  of the  *This i n i t i a l o b s e r v a t i o n was made by I. Itoh i n t h i s laboratory. **41 was prepared by hydrogenation of 16,18S-dicarbomethoxycleavamine i n the presence of a platinum c a t a l y s t .  F i g . 16.  Hmr  spectrum of 16,18S-dicarbomethoxy-3R,4S-epoxydihydrocleavamine  (40).  - 37 -  Cmpd.  XT  39 MeO CHC m/e 208 C  1 2  H  1 8  (23%) N0  2  136 (43%)  122 (100%)  C 1  C H  H  14  N  G  1  2  N  40 Me0 CHC 2  m/e 224 (100%) C  1 2  H  1 8  N0  3  152 (15%)  138 (10%:  C H  C H  9  1 4  NO  8  1 2  NO  41 MeC> CHC 2  m/e 210 (100%) C  Table 1.  12 20 H  N O  2  138 (22%) C  9 16 H  N  124 (13%) C  8  H  1 4  N  Comparison of some fragments e x h i b i t e d i n the mass spectra  of 16,18S-dicarbomethoxycleavamine (39), 16,18S-dicarbomethcxy-3R,4Sepoxydihydrocleavamine cleavamine (41).  (40), and 16,18S-dicarbomethoxy-4S-dihydro-  - 38 -  13  28 C-magnetic resonance  s p e c t r a of 40, l e u r o s i n e (3)  Me0 C 2  and  the  C0 Me 2  41 28  epoxide  42  (Table 2) while the determination of the absolute  42  \  Compound  Carbon^x -CH2CH3 CH 2CH ^ C-2 C-3 C-4 C-5 C-7 C-19  —  Table 2. epoxides  40  Leurosine  8.9 30.0 33.6 60.6 62.7 52.7 53. 4 50.6  8.6 28.0 33.5 60.3 59.9 54.0 49.6 42.3  42* 8.1 28.6 25.4 56.0 59.8 54.6 51.3 45.5  Comparison of 13rjmr chemical s h i f t s of l e u r o s i n e and 4_0 and  42.  *The numbering system normally used i n cleavamine was a l s o a p p l i e d to the epoxide 42.  derivatives  the  - 39 -  c o n f i g u r a t i o n of the epoxide was achieved by conversion  of 40  to a d e r i v a t i v e whose c o n f i g u r a t i o n at C-4 was known ( F i g . 17). * In t h i s regard,  reduction  a f f o r d e d the d i o l 4 3 .  Fig.  17.  6 3  of 40 with l i t h i u m aluminum hydride This d i o l  (43) was a l s o obtained by  Sequence used f o r the determination of the absolute  c o n f i g u r a t i o n o f 16,18S-dicarbomethoxy-3R,4S-epoxydihydrocleavamine (40). 63 65 l i t h i u m aluminum hydride reduction  of the known t r i o l  Since the c o n f i g u r a t i o n at C-4 of the t r i o l  44.  '  44 (and hence of the  d i o l 43) had been p r e v i o u s l y determined, the c o n f i g u r a t i o n at C-3 and C-4 i n the epoxide 40 was e s t a b l i s h e d (3R,4S). * This sequence was performed by A. H. R a t c l i f f e i n t h i s  laboratory.  - 40 -  Inasmuch as the epoxide 40 represented intermediate  an  attractive  f o r the synthesis of velbanamine d e r i v a t i v e s ,  f u r t h e r study of the a e r i a l o x i d a t i o n of the o l e f i n 39 undertaken.  A u t o x i d a t i o n of 39 was  was  c a r r i e d out i n p e r o x i d e - f r e e  t e t r a h y d r o f u r a n c o n t a i n i n g a small amount of aqueous 1% acetic acid.*  Monitoring  trifluoro-  the course of the r e a c t i o n v i a t h i n  l a y e r chromatography i n d i c a t e d that a f t e r three days at ambient temperature no r e a c t i o n had occurred. f i c a n t amount of epoxide 40 had o l e f i n 39 s t i l l present.  A f t e r f i v e days, a s i g n i -  formed, however the  starting  accounted f o r the m a j o r i t y of nitrogenous  material  A f t e r e i g h t days most of the s t a r t i n g m a t e r i a l had been  consumed, however the epoxide 40 had a l s o undergone f u r t h e r reaction.  Thus, an i s o l a t i o n of the products,  i n 10% y i e l d while  the major product  40 was  (52% y i e l d ) was  obtained the  formyl-  ketone 45**. CHO  45  *These c o n d i t i o n s had o r i g i n a l l y been developed by I. Itoh and A.H. R a t c l i f f e and found to a f f o r d r e l a t i v e l y uncomplicated product mixtures. * * O r i g i n a l l y , based on the assumption that the cleavamine carbon s k e l e t o n was i n t a c t , t h i s product was i n c o r r e c t l y assigned as the lactam 46.62,6 3  - 41 -  The s t r u c t u r a l assignment of 45 was based on a n a l y s i s of s p e c t r a l p r o p e r t i e s coupled with a chemical t r a n s f o r m a t i o n . Thus, the "*"Hmr spectrum  ( F i g . 18) e x h i b i t e d the f o l l o w i n g signals*- i n  accord with the proposed s t r u c t u r e :  ^8.13 (IH, s i n g l e t ,  N -CHO), b  c$5.23 (IH, broad singlet,Cg-H) , <£4.00 (3H, broad singlet,-OCH ) , 3  ^3.66  (3H,singlet,-OCH ) , i l . 9 0  i0.79  ( 3 H , t r i p l e t , J=7 Hz, -CH CH_ ) .  3  2  (2H, quartet,J=7 Hz, -CH_ CH ) , 2  3  The i n f r a r e d  3  spectrum  (Fig. 19) e x h i b i t e d strong absorptions at 1680 and 1650 c m  - 1  which were i n agreement with the presence of an enamide functionality.  High r e s o l u t i o n mass spectrometry e s t a b l i s h e d the  molecular formula as C„,H_,N~Cv while the presence of the ketone 23  26  2  6  C  f u n c t i o n a l i t y was s u b s t a n t i a t e d by sodium borohydride r e d u c t i o n of 45 e n a b l i n g i s o l a t i o n of the a l c o h o l 47.  *The d u p l i c a t i o n of v a r i o u s resonances observed i n t h i s spectrum have a l s o been noted i n a number of a l k a l o i d s which possess an N^-formyl group.66  3.66  l  1  F i g . 18.  1  1  1  1  Hmr spectrum of f o r m y l ketone 45.  r  F i g . 19.  I n f r a r e d spectrum of formyl ketone 45.  - 44 -  CHO  47 The s t r u c t u r e 45. i t y was lactam  Accordingly/ the presence of the ketone f u n c t i o n a l -  compatible 46.  20)  only with a cleavage product  and r u l e d out  Furthermore, t h a t the a l c o h o l 47 d i s p l a y e d strong  absorptions (Fig.  above information provided s u f f i c i e n t proof f o r  i n i t s i n f r a r e d spectrum at 1669  and  1651  cm  corroborated the enamide assignment i n 47 and <~i  F i n a l l y , the a l t e r n a t i v e enamide 50 was  45 •—  while e l i m i n a t i n g s t r u c t u r e s such as the <* ,/3 -unsaturated 48 and 49.  1  ketones  r u l e d out CHO  48 due  to i n c o m p a t i b i l i t y with the observed  the resonance due was  49 "^Hmr  spectrum i n which  to the methylene hydrogens of the e t h y l group  l o c a t e d at Si.90,  i n good agreement with a methylene group 67  adjacent to a double bond  (but not to a ketone).  F i g . 20.  I n f r a r e d spectrum of a l c o h o l  47.  -  Having dation  that for  a  to  was  tested  which  had  Using  otherwise  reaction  by  In  aqueous  1%  40  to  the  aerial  starting  the  of  and  40,  acid  i t was  c a r r i e d out  in  rate.*'  as  indicated had  products the  This  after  material  sup-  the of  solvent. the after  conditions  as  well  the  olefin  of  as 39  t-butyl  appropriate  present  conjunction  25%  containing  amount  under  to  twenty-two in  of  four  converted  45  a known that  suggested  tetrahydrofuran,  that  been  the  ketoenamide  oxidation  and  of  necessary  ^°  8  autoxi-  period  be  tetrahydrofuran  found  peroxide  might  monitoring  reproducible  epoxide  this  induction  oxidation*,  and  of  reaction with  material  yield  attain  Thus  an  chromatography  trifluoroacetic  was  of  course  modification  peroxide  repeating  32%  of  to  appreciable  in peroxide-free  concentration study  of  in  yield  hydroperoxide**.  **This  an  of  I s o l a t i o n of  order  performed  by  layer  40.  the  at  undergone  majority  afforded  maximize  *The  presence  similar conditions  thin  epoxide  yield.  was  already  the  hours  occur  overall  turned  certain concentration  position  the  The  -  the  i n t e r e s t was  conditions.  reaction  hours  ascertained  reaction,  reaction  46  with  was T.  not  condi-  measured.  Hibino.  - 47 -  t i o n s 16,18S-dicarbomethoxy-3R,4S-epoxydihydrocleavamine  (40)  6 2 63 could be obtained  i n 76% y i e l d .  With the epoxide 40 now o b j e c t i v e was d i o l 51'  '  ' r e a d i l y a v a i l a b l e , the next  a c i d c a t a l y s e d h y d r o l y s i s to the trans  —a  compound possessing  d r o x y l group.  diaxial  the r e q u i r e d a x i a l C.  hy-  However, s e v e r a l attempts to e f f e c t h y d r a t i o n  of  HO  the epoxide f u n c t i o n a l i t y l e d to d i s a p p o i n t i n g r e s u l t s . general,  In  i t was  observed that at ambient temperature, except  i n concentrated  s o l u t i o n s of strong acids such as p e r c h l o r i c  a c i d , no r e a c t i o n occurred.  On heating  ( 50°C) the epoxide  40  r e a d i l y underwent r e a c t i o n i n various media y i e l d i n g i n a l l cases complex product mixtures. d i t i o n s developed  Indeed, the best r e a c t i o n con-  (aqueous 3 0% p e r c h l o r i c a c i d / t e t r a h y d r o f u r a n  at 66°C f o r 24 h or aqueous 50% t r i f l u o r o a c e t i c a c i d at 65°C f o r 4 h) a f f o r d e d the d i o l 51 i n y i e l d s of only 17-25%. The  i d e n t i t y of the d i o l 51 was  confirmed by i t s  s p e c t r a l p r o p e r t i e s as w e l l as by a chemical  transformation.  High r e s o l u t i o n mass spectrometry e s t a b l i s h e d the formula as C» ,H, N.,O , while n  c  the mass s p e c t r a l  molecular  fragmentation  -  pattern  verified  piperidine ation  ring  was b a s e d  resonance  the presence (Fig.  21).  chloride  products.  the  3S,4R-epoxide  was p r o v e n  position  From  product  t h e "*"Hmr s p e c t r u m  from)  that  (Fig.  (Fig. 22), while the  this  o f 51 w i t h  reaction  methane-  were  obtained  was a s s i g n e d a s  of i t s spectral  properties.  23) w a s s i m i l a r t o ( b u t d i f f e r e n t  o f t h e 3R,4S-epoxide  spectrometry  configur-  o f t h e C^g-hydrogen  (52% y i e l d )  52 o n t h e b a s i s  Thus,  o f t h e 18S  by t r e a t m e n t  i n pyridine.  The major  f u n c t i o n a l i t y i n the  The assignment  (e$6.54) i n t h e "'"Hmr s p e c t r u m  two  mass  of a diol  on t h e low f i e l d  3R,4R c o n f i g u r a t i o n sulfonyl  48 -  established  40  (Fig. 16).  the molecular  High  resolution  formula  as  C_oH_ N_0 fl  OH  52 while ring  53  the presence was c o n f i r m e d  o f the epoxide b y t h e mass  f u n c t i o n a l i t y i n the piperidine  spectral  fragmentation  pattern  (Fig. 24). The 3R-mesyloxy which  minor  derivative  contained  C^-H) , c(3.03 unstable  product  (29% y i e l d )  53 o n t h e b a s i s  the following  signals:  ( 3 H , s i n g l e t ,-OS0 CH ) .  and on normal  2  handling  3  was a s s i g n e d o f i t s ^Hmr  as t h e  spectrum,  ^ 4 . 6 0 ( I H , d o u b l e t , J = 2 . 5 Hz,  This  c o m p o u n d was  was p a r t i a l l y  converted  somewhat tothe  F i g . 21.  Mass s p e c t r a l fragmentation p a t t e r n of  3R-hydroxyvelbanamine (51).  16,18S-dicarbomethoxy—  3.97  3.96  Fig.  24.  Mass  spectral  3S,4R-dihydrocleavamine  fragmentation (52).  pattern of  16,18S-dicarbomethoxy—  - 53 -  3S,4R-epoxide  52*.  The f a c i l e c o n v e r s i o n o f t h e d i o l 51 t o t h e 3S ,4R-i—  epoxide  52 was  c o n c l u s i v e p r o o f o f i t s 3R,4R c o n f i g u r a t i o n , as  o n l y i n the case o f a t r a n s - d i a x i a l o r i e n t a t i o n o f the hydroxy and m e s y l o x y g r o u p s  ( i n 53) c o u l d t h e c y c l i z a t i o n have  occurred  73 so r e a d i l y i t was  now  ; and t h o u g h t h e p r e p a r a t i o n o f 51 was  inefficient,  p o s s i b l e t o t e s t t h e e f f e c t o f an a x i a l  C^-hydroxyl  g r o u p on t h e s t e r e o c h e m i c a l outcome o f t h e c h l o r o i n d o l e n i n e 'dimerization'. Consequently,  51 was  t r e a t e d w i t h a s o l u t i o n o f sodium  m e t h o x i d e i n m e t h a n o l f r o m w h i c h was quantitative yield.  o b t a i n e d the d i o l  54 i n  The i d e n t i t y o f t h i s p r o d u c t was s u b s t a n t i a t e d HO  i  C0 Me 2  54 by i t s u l t r a v i o l e t s p e c t r u m w h i c h was chromophore.  The  f a c t t h a t t h e C-18  d u r i n g t h e r e a c t i o n was  i n d i c a t i v e o f an i n d o l e p o s i t i o n was  not  c o r r o b o r a t e d by t h e c h e m i c a l  t h e "'"Hmr s p e c t r u m ) o f t h e C^g-hydrogen  epimerized shift (in  («f5.45).  * A l t h o u g h 53 was o r i g i n a l l y i s o l a t e d as a s i n g l e compound by t h i n l a y e r c h r o m a t o g r a p h y , a f t e r d e t e r m i n a t i o n o f t h e iHmr s p e c t r u m t h e sample c o n t a i n e d c l o s e t o 50% o f t h e e p o x i d e 52.  -  54  -  Treatment of 54 with 1 - c h l o r o b e n z o t r i a z o l e  in  benzene a f f o r d e d the c h l o r o i n d o l e n i n e 55 i n 57% y i e l d . * HO  i  CO  /  Me  s p e c t r a l p r o p e r t i e s of t h i s product were i n accord with assigned  structure.  phore. hydrogen  The  ^Hmr  the  Thus, high r e s o l u t i o n mass spectrometry  e s t a b l i s h e d the molecular u l t r a v i o l e t spectrum  The  formula as C^^E^^2^'  ( F i g . 25) was  w  hile  i n d i c a t i v e of the new  spectrum e x h i b i t e d a s i g n a l due  to the  (£5.79,lH,doublet of doublets,J=2,12 Hz)  and  the chromoC^g—  lacked  a s i g n a l f o r an N -hydrogen. a  With the c h l o r o i n d o l e n i n e 55 now  a v a i l a b l e , the next  step i n v o l v e d i t s coupling with v i n d o l i n e under a c i d i c c o n d i t i o n s . Treatment of 55 with v i n d o l i n e i n methanolic hydrogen c h l o r i d e however a f f o r d e d no i s o l a b l e  'dimeric' products.**  Indeed  *This compound tended to decompose during i s o l a t i o n and thus the low y i e l d of i s o l a t e d product was not t r u l y i n d i c a t i v e of the c h l o r i n a t i o n r e a c t i o n which by t h i n l a y e r chromatographic a n a l y s i s appeared to be q u a n t i t a t i v e . **This r e a c t i o n was c a r r i e d out by T. Hibino s e v e r a l times, using the various c o n d i t i o n s which p r e v i o u s l y were s u c c e s s f u l f o r the coupling of 18S-carbomethoxy-4S-dihydrocleavamine c h l o r o i n d o l e n i n e with vindoline.44  - 55 -  log  £  250  300  X F i g . 25.  350  nm  U l t r a v i o l e t spectrum of the  c h l o r o i n d o l e n i n e 55.  g r e a t e r than 90% of the v i n d o l i n e could be recovered r e a c t i o n mixture.  T h i s negative  from the  r e s u l t cast some doubt on the  o r i g i n a l premise, however due to the lack of any coupling r e a c t i o n or other t a n g i b l e consequences, no d e f i n i t i v e conclusions be made. the  1  Therefore,  dimerization  1  could  s t u d i e s were undertaken to a s c e r t a i n why  had f a i l e d and to determine whether the  presence of an a x i a l C^-hydroxyl f u n c t i o n a l i t y could change the  'normal' stereochemical  outcome o f the c h l o r o i n d o l e n i n e  coupling r e a c t i o n . Examination of the proposed general mechanism of the  - 56 -  F i g . 26.  Proposed general mechanism of  'dimerization'.  chloroindolenine  - 57 -  chloroindolenine  'dimerization  ( F i g . 26) suggested that the  1  lack of r e a c t i o n between a c h l o r o i n d o l e n i n e d e r i v a t i v e and v i n d o l i n e could be due to formation s a l t 57.  o f the quaternary ammonium  T h i s type of quaternary ammonium s a l t had p r e v i o u s l y  been prepared i n t h i s l a b o r a t o r y 49 towards v i n d o l i n e .  49 7 '  and shown to be u n r e a c t i v e  Thus, a r a p i d r a t e of i n t e r n a l  quaterniza-  t i o n r e l a t i v e to the r a t e of 'dimerization' would e x p l a i n the lack of 'dimeric'product 55 with v i n d o l i n e .  observed i n the attempted coupling of  Support f o r t h i s explanation was provided by  examination of molecular  models which showed that f o r i n t e r n a l  q u a t e r n i z a t i o n to occur the N^-lone p a i r would have to occupy an a x i a l o r i e n t a t i o n r e l a t i v e to the p i p e r i d i n e r i n g , and as the q u a t e r n i z a t i o n r e a c t i o n 'proceeded  (i-»ii) there would be a  strong s t e r i c i n t e r a c t i o n between C-18 and the C-4 a x i a l (or pseudoaxial) s u b s t i t u e n t .  I t t h e r e f o r e followed t h a t the g r e a t e r Me  the s t e r i c bulk of the C-4 s u b s t i t u e n t the l e s s f a v o r a b l e would be i n t e r n a l q u a t e r n i z a t i o n .  Accordingly,  the e f f e c t i v e  coupling  of 18S-carbomethoxy-4S-dihydrocleavamine c h l o r o i n d o l e n i n e (22) with v i n d o l i n e  (to a f f o r d the dimer 23 i n  ~ 65% y i e l d ) could be  - 58 -  a t t r i b u t e d to a slow r a t e of q u a t e r n i z a t i o n 'dimerization') provided  which i n t u r n was  due  ( r e l a t i v e to  to the s t e r i c hindrance  by the a x i a l C ^ - e t h y l group.  Conversely, the  un-  s u c c e s s f u l coupling of 18S-carbomethoxy-3R-hydroxyvelbanamine chloroindolenine increased  (55) with v i n d o l i n e could be a t t r i b u t e d to an  rate of i n t e r n a l q u a t e r n i z a t i o n  'dimerization')  ( r e l a t i v e to  r e s u l t i n g from the diminished  a f f o r d e d by an a x i a l C^-hydroxyl group. of t h i s r a t i o n a l e was  s t e r i c hindrance  Further  substantiation  f u r n i s h e d by the p r e v i o u s l y performed  coupling of 18S-carbomethoxycleavamine c h l o r o i n d o l e n i n e  (58)  with v i n d o l i n e , from which 18'-epi-3',4'-anhydrovinblastine (59) was  obtained  i n ca 10% y i e l d .  Since  the s t e r i c impedance  a f f o r d e d by the v i n y l e t h y l group* toward i n t e r n a l q u a t e r n i z a t i o n  *The s p a c i a l r e l a t i o n s h i p between C-18, the electron pair and the C-4 a x i a l (or pseudoaxial) s u b s t i t u e n t i s n i c e l y demonstrated by a comparison of the chemical s h i f t of the C]_g-hydrogen i n the Hmr s p e c t r a of various 18S-carbomethoxycleavamine d e r i v a t i v e s (Table 3). Since the chemical s h i f t of the Cl8 hydrogen depends upon i t s proximity to the electron pair, a comparison of t h i s s h i f t as a f u n c t i o n of the C o a x i a l (or pseudoaxial) s u b s t i t u e n t would r e f l e c t the 1 , 3 - d i a x i a l i n t e r a c t i o n of the Nfc, e l e c t r o n p a i r with that s u b s t i t u e n t . Indeed, Table 3 shows a p o s i t i v e c o r r e l a t i o n between the C]_g-hydrogen' s chemical s h i f t and the s t e r i c bulk a f f o r d e d by the C-4 functionality. Presumably, the g r e a t e r the s t e r i c bulk, the g r e a t e r the displacement of the N]-, e l e c t r o n p a i r away from C-18. 1  -  Table 3 . Comparison of the chemical s h i f t s of the C hydrogen of v a r i o u s derivatives.  16,18S-dicarbomethoxycleavamine  l g  —  - 60  should have been intermediate to t h a t a f f o r d e d by the a x i a l e t h y l group i n 22 and the a x i a l hydroxyl group  i n 55, the intermed-  i a t e y i e l d of 59 ( i . e . 10%) was reasonable. Another d e r i v a t i v e a v a i l a b l e to f u r t h e r evaluate the 'standard c h l o r o i n d o l e n i n e d i m e r i z a t i o n ' was the epoxide 60 which was a v a i l a b l e from 40 i n 50% y i e l d * by the a c t i o n of  4 0 , R=C0 Me 2  *****  6 0 , R=H  sodium methoxide i n methanol.  In accord with the proposed  s t r u c t u r e , the u l t r a v i o l e t spectrum was i n d i c a t i v e o f the i n d o l e chromophore.  High r e s o l u t i o n mass spectrometry confirmed, the  *This compound was found to be s e n s i t i v e to a c i d i c and b a s i c media and the low y i e l d of i s o l a t e d product obtained l i k e l y r e f l e c t s the occurrence of decomposition during the r e a c t i o n and subsequent p u r i f i c a t i o n by chromatography on alumina.  - 61 -  molecular formula as 2 1 2 6 2 ° 3 ' ^^--'C  18S  H  N  w  e t l l e  assignment of the  c o n f i g u r a t i o n was based upon the low f i e l d p o s i t i o n of the  C - h y d r o g e n resonance i n the "'"Hmr spectrum ig  (^5.19,doublet o f  doublets,J=10,5 Hz). Treatment of 60 with 1 - c h l o r o b e n z o t r i a z o l e the c h l o r o i n d o l e n i n e  61 i n 67% y i e l d .  afforded  High r e s o l u t i o n mass  i  C0 Me 2  61  spectrometry confirmed the molecular formula as C ^ H ,.N 0 C1, 2  2  while the u l t r a v i o l e t spectrum was analgous to that  2  3  obtained  f o r 55. Treatment of 61 with a s o l u t i o n of v i n d o l i n e i n methanolic hydrogen c h l o r i d e a f f o r d e d four  'dimeric'  products  i n a combined y i e l d of ca 17%. Although s t r u c t u r e s were not assigned  to these products the f a c t t h a t they were 'dimers'  was e s t a b l i s h e d by t h e i r u l t r a v i o l e t s p e c t r a which were i n d i c a t i v e o f the s u p e r p o s i t i o n of an i n d o l e and dihydroindole chromophore ( F i g . 27). The i s o l a t i o n of s e v e r a l dimers was l i k e l y due to the i n s t a b i l i t y of the epoxide f u n c t i o n a l i t y i n the s t r o n g l y a c i d i c r e a c t i o n c o n d i t i o n s employed.  Of p a r t i c u -  l a r i n t e r e s t however was the observed o v e r a l l y i e l d of 'dimeric'  -  62  -  log £  250  300  250  X  )\ nm  F i g . 27.  300  nm  Comparison of the u l t r a v i o l e t spectra of a t y p i c a l  derivative  (A), d i h y d r o i n d o l e  m a t e r i a l which was  d e r i v a t i v e (B), and  s i m i l a r to that obtained  r e a c t i o n of the o l e f i n 59.  Since  i n the  'dimer'  indole  (C).  'dimerization'  the s t e r i c requirements f o r the  i n t e r n a l q u a t e r n i z a t i o n were s i m i l a r i n both cases the observed r e s u l t s were c o n s i s t e n t . To t h i s p o i n t the important r o l e of i n t e r n a l quatern i z a t i o n as a competing r e a c t i o n i n the c h l o r o i n d o l e n i n e ' d i merization'  had been s t r o n g l y implied but not proven.  t h i s proof was  undertaken.  Oxidation  Thus,  of the 3R,4S-epoxide 40  77 with e i t h e r chromium t r i o x i d e i n p y r i d i n e or mercuric acetate 78 i n aqueous dioxane a f f o r d e d the lactam epoxide 62. This de-  - 63 -  r i v a t i v e could not undergo i n t e r n a l q u a t e r n i z a t i o n as the e l e c t r o n p a i r was now i n v o l v e d i n resonance s t a b i l i z a t i o n o f the adjacent carbonyl group.  Me0 C 2  C0 Me 2  62 The s t r u c t u r a l assignment of 62 was based on a n a l y s i s o f s p e c t r a l data.  Thus, the presence o f the amide f u n c t i o n a l i t y  was corroborated by a strong absorption a t 164 2 cm ^ i n the i n f r a r e d spectrum  ( F i g . 28). High r e s o l u t i o n mass spectrometry  e s t a b l i s h e d the molecular formula as C»_.H_^N-0,.  Furthermore,  2 3 26 2 6  that the lactam obtained was indeed the 19-oxo d e r i v a t i v e 62 and not the 5- or 7-oxo d e r i v a t i v e s 63 or 64 was deduced from the  13  Cmr and  1 Hmr s p e c t r a .  63  The  13  Cmr spectrum  64  ( F i g . 29) e x h i b i t e d  F i g . 28.  I n f r a r e d spectrum of 16,18S-dicarbomethoxy-3R,4S-epoxy-19-oxodihydro-  cleavamine (62).  61.4 i  59.3  171.9  lU F i g . 29.  / O ' w  V^rv.  TMS  13 Cmr spectrum of 16,18S-dicarbomethoxy-3R,4S-epoxy-19-oxodihydrocleavamine  (62).  -  signals  66 -  f o r C-3 a n d C-4 a t J61.4 a n d £59.3  Comparison  o f these  signals  with  40 (£60.6 a n d £62.7) i n d i c a t e d field  b y 0.8  ppm w h i l e  Since  a c a r b o n y l group  those  that  C-4 h a d b e e n normally  respectively.  of the starting  epoxide  C-3 h a d b e e n s h i f t e d downshifted  causes  upfield  b y 3.4  a substantial  ppm.  downfield  79 shift  o f the adjacent  observed ture basis  63.  position  carbon  o f t h e C-4  F i n a l l y , structure  o f t h e "'"Hmr s p e c t r u m .  resonance signal  ( c f F i g . 30), t h e  was i n c o m p a t i b l e w i t h  6 4 was d e e m e d u n a c c e p t a b l e Thus,  a 7-oxo d e r i v a t i v e  A  struc-  on t h e  w o u l d be  B  13 Fxg.  30.  Comparison  hexahydroazepine  expected as  to exhibit  was o b s e r v e d  spectrum  8 0  ( F i g . 31)  exhibiting  Cmr c h e m i c a l  (A) a n d c a p r o l a c t a m  7 9  shifts  (S) o f  (B).  a n AB q u a r t e t d u e t o t h e C-8 h y d r o g e n s  f o r 65  "'"Hmr s p e c t r u m — F i g . while  of the  81  (<$3.94, AB q u a r t e t ) .  l a c k e d such 3 2--of  resonances  a signal  Indeed,the  (more e a s i l y  the N -descarbomethoxy a  i n accord with  seen  such  1  Hmr  i n the  derivative  structure  66)  62 U4.32,  - 68 -  b  OMe 65  lH,multiplet,C -H*; 2  J 4 .1, 1 H , d o u b l e t ,J-6  With the i d e n t i t y s t e p was r e g e n e r a t i o n 62 w i t h  Hz,C -H). 1 8  of the lactam  6 2 secured  o f t h e i n d o l e chromophore.  sodium m e t h o x i d e i n m e t h a n o l a f f o r d e d  the next  Treatment of  66 i n 95% y i e l d .  Me0 C 2  66  The  p r e s e n c e o f t h e i n d o l e chromophore was s u b s t a n t i a t e d by  the  ultraviolet  spectrum .  Further  corroboration  66 was a v a i l a b l e from t h e "^Hmr s p e c t r u m the  f o l l o w i n g s i g n a l s : <4 8.67  of t r i p l e t s , J = 1 3 , 3  ( F i g . 32) w h i c h  (lH,bs,N -H) ; & 4.33 a.  f o r structure exhibited  (IH,doublet  —  H z , C ~ H ) , & 4.05 (IH,dd,J=10.5,1.5 2  Hz,C -H). l g  *The low f i e l d p o s i t i o n o f t h e C^-hydrogen r e s o n a n c e was a s c r i b e d t o t h e f a c t t h a t i t was e c l i p s e d w i t h t h e a d j a c e n t c a r b o n y l group (at C-19) and thus d e s h i e l d e d s t r o n g l y by t h e a n i s o t r o p i c e f f e c t of the carbonyl. Some d e s h i e l d i n g c o u l d a l s o be a t t r i b u t e d t o t h e s t e r i c a l l y crowded e n v i r o n m e n t o f t h e C 2 h y d r o g e n . _  6  3.71  - 70 -  F i n a l l y , the 18S c o n f i g u r a t i o n was assigned on the b a s i s of previous r e s u l t s where no e p i m e r i z a t i o n of t h i s center had been observed. Having 66 a v a i l a b l e , the next step attempted was c h l o r i n a t i o n of the i n d o l e .  Treatment of 6 6 with  1-chloro-  b e n z o t r i a z o l e i n benzene however d i d not y i e l d the expected c h l o r o i n d o l e n i n e 67, i n s t e a d two i n d o l i c products were i s o l a t e d (Fig. 33).  The major product  (40% y i e l d ) was assigned as the  CI  Fig.  33.  O u t l i n e of the r e a c t i o n of 18S-carbomethoxy-3R,4S—  epoxy-19-oxodihydrocleavamine  (66) with 1 - c h l o r o b e n y o t r i a z o l e .  - 71 -  1 8 R - b e n z o t r i a z o l e adduct 68. I n a c c o r d w i t h t h i s s t r u c t u r e the "'"Hmr spectrum  ( F i g . 34) l a c k e d a C^g-hydrogen resonance w h i l e  e x h i b i t i n g s i g n a l s c o r r e s p o n d i n g t o f o u r new aromatic p r o t o n s . The c h e m i c a l s h i f t s o f these p r o t o n s i n d i c a t e d t h a t they were m a g n e t i c a l l y d i f f e r e n t from each o t h e r and thus l e d t o t h e assignment o f the l i n k a g e between C-18 and the 1 - p o s i t i o n o f benzotriazole.  The t e n t a t i v e assignment o f the s t e r e o c h e m i s t r y  as 18R was based on the f a c t t h a t i n p r e v i o u s ' c o u p l i n g s ' the nucleophile  ( i . e . v i n d o l i n e ) approached  from the '/3-face . 1  F i n a l l y , h i g h r e s o l u t i o n mass spectrometry confirmed t h e molecul a r formula as C H N 0 . 2 7  2 7  5  4  The minor product  (27%) was a s s i g n e d as the 18R-hy-  droxy adduct 69. High r e s o l u t i o n mass spectrometry e s t a b l i s h e d the m o l e c u l a r formula as 2 i 2 4 2 " 4 *-'--'- ^he ^"Hmr spectrum C  H  N  <  )  w 1  e  ( F i g . 35) l a c k e d a C ^ - h y d r o g e n resonance  i n d i c a t i n g the hy-  droxy 1 group a t t h a t p o s i t i o n . Although t h e c h l o r o i n d o l e n i n e 6_7 was n o t o b t a i n e d , the adducts 6 8 and 6 9 were as s u i t a b l e f o r t h e r e q u i r e d purpose ( i . e . c o u p l i n g w i t h v i n d o l i n e ) , as under a c i d i c c o n d i t i o n s t h e 44  e q u i l i b r i u m i i i ^ = s . i v would be a t t a i n e d .  Therefore, a solu-  t i o n o f 68 i n methylene c h l o r i d e was t r e a t e d w i t h v i n d o l i n e and a s m a l l amount o f t r i f l u o r o a c e t i c a c i d .  From t h i s r e a c t i o n  a s i n g l e product was o b t a i n e d (89% y i e l d ) which on the b a s i s of i t s s p e c t r a l p r o p e r t i e s was a s s i g n e d as 18'-epi-19'-oxoleuros i n e (70).  High r e s o l u t i o n mass spectrometry e s t a b l i s h e d the  m o l e c u l a r formula as 4 6 5 4 4 ° ] _ o ' C  H  N  T l l e  ^  H m r  spectrum  ( F i g . 36)  3.83  i F i g . 34.  1  r  i  Hmr spectrum of 68.  1  1  1  r  - 74 -  contained  singlets at i6.87  (C,.-H) and 5.9 8 (C,_-H) i n accord with  a 15-substituted v i n d o l i n e d e r i v a t i v e . was assigned  the 'unnatural'  Finally, this  stereochemistry  on the b a s i s of i t s c i r c u l a r dichroism  'dimer'  at C-18' ( i . e . 18R)  (cd) curve which e x h i b i t e d 82  strong absorptions  at 208 nm  P r e v i o u s l y the cd curves  (A£=+12.4)  and 222 nm  (A£=-41.3).  of various n a t u r a l and 'unnatural  Thus, the 'natural dimers'  ( i . e . 18'S c o n f i g u r a t i o n )  e x h i b i t e d a strong negative absorption at ca 208 nm and a strong p o s i t i v e one at ca 222 nm while the 'unnatural dimers' 18'R c o n f i g u r a t i o n ) e x h i b i t e d the reverse.  '  dimers'  (at C-18') had been c o r r e l a t e d and shown to be of i n t e r p r e t i v e value.  8  (i.e.  2 . 0 6  T  F i g . 36.  Hmr  1  1  r  spectrum of 18'-epi-19'-oxoleurosine (70).  - 76 -  The e x c e l l e n t y i e l d of dimer obtained i n the c o u p l i n g r e a c t i o n of the lactam 68 with v i n d o l i n e versus the poor  yield  obtained i n the r e a c t i o n of the amine 61 e s t a b l i s h e d the s i g n i - • f i c a n c e of i n t e r n a l q u a t e r n i z a t i o n as a side r e a c t i o n during attempted d i m e r i z a t i o n s . 1  1  Having e s t a b l i s h e d q u a t e r n i z a t i o n as a s e r i o u s drawback, a more e a s i l y reversed method of b l o c k i n g t h i s path sought.  One  p o s s i b i l i t y was  was  formation of the N^-iminium s p e c i e s ,  which upon r e d u c t i o n with sodium borohydride would regenerate the parent amine.  T h e r e f o r e , 18S-carbomethoxy-3R,4S-epoxy-  dihydrocleavamine c h l o r o i n d o l i n e  (61) was  N^-oxide 71 i n the presence of v i n d o l i n e  converted to i t s (by the a c t i o n of  chloroperbenzoic acid) and t h i s mixture was  t r e a t e d with  f l u o r o a c e t i c a c i d / t r i f l u o r o a c e t i c anhydride  (presumably  generating the iminium species 72*)  ( F i g . 37).  m—  tri-  Quenching of the  r e a c t i o n with sodium borohydride a f f o r d e d a f t e r p u r i f i c a t i o n 18 -epileurosine 1  was  (73) i n 40% y i e l d .  The i d e n t i t y of t h i s product  i n accord with i t s s p e c t r a l p r o p e r t i e s .  High r e s o l u t i o n mass  spectrometry confirmed the molecular formula as C^gH^gN^Og, while the cd curve e s t a b l i s h e d the c o n f i g u r a t i o n at C-18' F i n a l l y , the "^Hmr  spectrum  ( F i g . 38) was  as R.  s i m i l a r to those of 44  various  'unnatural dimers' obtained e a r l i e r .  *The formation of the iminium d e r i v a t i v e i s an a p p l i c a t i o n of the Polonovski reaction.51 On the b a s i s of geometric and e l e c t r o n i c c o n s i d e r a t i o n s i t was f e l t that the e l i m i n a t i o n process to form the iminium ion would occur r a t h e r than any C-C bond fragmentation process.  - 77 -  trifluoroacetic acid trifluoroacetic anhydride  72  Vindoline  Fig.  37.  Proposed mechanism f o r coupling of  61 with v i n d o l i n e .  chloroindolenine  - 79 -  Therefore, chloroindolenine  having  modified  the c o n d i t i o n s of the  ' d i m e r i z a t i o n ' so as t o m i n i m i z e  internal  q u a t e r n i z a t i o n as w e l l as t h e d e c o m p o s i t i o n o f p r o d u c t s , t h e n e x t a i m was t h e c o u p l i n g o f 1 8 S - c a r b o m e t h o x y - 3 R - h y d r o x y v e l banamine c h l o r o i n d o l e n i n e  (55) w i t h v i n d o l i n e .  To t h i s e n d ,  55 was t r e a t e d w i t h v i n d o l i n e u s i n g t h e m o d i f i e d tions.  I n v e s t i g a t i o n of the product mixture  'dimeric' product  reaction condi-  afforded a single  (19% y i e l d ) w h i c h on t h e b a s i s o f i t s s p e c t r a l  p r o p e r t i e s was i d e n t i f i e d as 1 8 ' - e p i v i n c a d i o l i n e ( 7 4 ) . H i g h  HO  74 r e s o l u t i o n mass s p e c t r o m e t r y as -4g 5g 4 - ]_o' <  H  N  <  )  w n i  l  e  t  n  e  c  &  e s t a b l i s h e d the molecular  formula  c u r v e a f f i r m e d t h e 18R a s s i g n m e n t .  F u r t h e r m o r e , t h e ^Hmr s p e c t r u m  ( F i g . 39) was s i m i l a r t o t h o s e 44  of p r e v i o u s l y prepared  'unnatural  dimers'.  I n summary, i t was shown t h a t i n t e r n a l q u a t e r n i z a t i o n c o u l d be a s e r i o u s s i d e r e a c t i o n . ' d i m e r i z a t i o n ' was m o d i f i e d reaction.  Using  Hence, t h e c h l o r o i n d o l e n i n e  so as t o m i n i m i z e t h i s  the modified  conditions,  3R-hydroxyvelbanamine c h l o r o i n d o l e n i n e  18S-carbomethoxy—  (55) c o u l d be  with v i n d o l i n e to a f f o r d only the 'unnatural' 'dimer'  ( i . e .74).  undesirable  coupled  stereochemistry  F i g . 39.  Hmr  spectrum of 1 8 - e p i v i n c a d i o l i n e 1  (74).  -  In c o n c l u s i o n , indolenine of the was  -  81  the i n i t i a l premise, that the  of 18-carbomethoxyvelbanamine would be  4S,18R-ether 32, was  underwent i n t e r n a l q u a t e r n i z a t i o n . coupling  s s e s s i n g an a x i a l C^-hydroxyl group  which was  reported  product obtained  po-  velbanamine  Furthermore, the s t r u c t u r e  as being the  37,  indolic  from the a c i d c a t a l y z e d cleavage of v i n b l a s t i n e  here.  agents, tended to contravene the  In view of these r e s u l t s i t was  more p l a u s i b l e s t r u c t u r e would be t h i s proposal  readily  of d e r i v a t i v e s  ( i . e . various  i n the l i t e r a t u r e  i n the absence of reducing s u l t s obtained  at C-18  Thus, the argument f o r the  (with v i n d o l i n e )  d e r i v a t i v e s ) became f a l l a c i o u s .  Rather i t  an a x i a l C41-hydroxy1  f u n c t i o n a l i t y as w e l l as a c a t i o n i c center  and  precursor  found to be i n c o r r e c t .  seen that d e r i v a t i v e s possessing  preparation  a  chloro-  must be  although the s y n t h e s i s  considered and  f e l t that a  the ammonium s a l t 75, purely  speculative.  'dimerization'  re-  however  Finally,  (with v i n d o l i n e )  of  cleavamine d e r i v a t i v e s whose geometry precludes r e a c t i o n with nucleophiles  from the  '/9-face'  (e.g.  32 and  33)  still  represents  - 82 -  a v a l i d g o a l , the d i f f i c u l t y encountered i n i n t r o d u c i n g an a x i a l C^-hydroxyl group i n t o 18S-carbomethoxycleavamine  (20)  f o r c e d abandonment o f the approach i n favor o f other more promising routes, such as the f u n c t i o n a l i z a t i o n of 3',4 -anhydro1  vinblastine  (26)  (Sec. 2.2,  2.3)..  - 83 -  2.2  Chemistry  of  3',4 -Anhydrovinblastine— 1  S y n t h e s i s o f L e u r o s i n e and C a t h a r i n e . Although approach'  f u r t h e r e v a l u a t i o n of the ' c h l o r o i n d o l e n i n e  d i d n o t l e a d t o t h e a n t i c i p a t e d outcome,  t h a t was d e v e l o p e d cleavamine,  the chemistry  ( f o r the e l a b o r a t i o n o f 18S-carbomethoxy-  19) d i d p r o v i d e a f o u n d a t i o n f o r t h e i n i t i a l  aimed a t f u n c t i o n a l i z a t i o n o f 3 ' , 4 ' - a n h y d r o v i n b l a s t i n e  19 Accordingly, tection  (26).  26  the f i r s t m o d i f i c a t i o n of 26 attempted, was  of the i n d o l e chromophore.  and methyl chloroformate  pro-  However, despite treatment of  26 with v a r y i n g amounts of potassium hydride  tures  studies  i n tetrahydrofuran  (2-5  equivalents)  at v a r i o u s  tempera-  (0-50°C), only s t a r t i n g m a t e r i a l or decomposition products  could be i s o l a t e d . *  Indeed, no evidence f o r the formation  N i-carbomethoxy d e r i v a t i v e s could be obtained. a  r e a c t i o n at the N i p o s i t i o n a  of the  'dimer' was  reduced a c c e s s i b i l i t y of t h i s p o s i t i o n hindrance a f f o r d e d by the C-18'  *This study was  lack of  presumed due  to  as a r e s u l t of s t e r i c  substituents  c a r r i e d out by T.  The  of  Hibino.  (i.e. vindoline  and  - 84 -  the carbomethoxy group). cated  that the C-18'  Examination of molecular models i n d i -  s u b s t i t u e n t s would a l s o a f f o r d s t e r i c  pedance to approach of reagents at the  ft-position  chromophore  reasoned that  ( i . e . C-9),  and  thus i t was  of the  t i o n of the i n d o l e chromophore might not be necessary. epoxidation  of 3 , 4 ' - a n h y d r o v i n b l a s t i n e using the  indole  protecTherefore,  previously  1  developed c o n d i t i o n s *  im-  (tetrahydrofuran,  aqueous 1%  trifluoro-  a c e t i c a c i d , t - b u t y l hydroperoxide) was  performed.  From t h i s  62 reaction leurosine  (3) was  obtained  i n 51% y i e l d .  i d e n t i t y of the s y n t h e t i c l e u r o s i n e was son with an authentic  Further  63 '  The  e s t a b l i s h e d v i a compari-  sample.  examination of the t - b u t y l hydroperoxide r e a c t i o n i n -  d i c a t e d that under prolonged r e a c t i o n times l e u r o s i n e would undergo f u r t h e r r e a c t i o n * * to y i e l d another n a t u r a l l y o c c u r r i n g  *This r e a c t i o n was  i n i t i a l l y performed by T.  Hibino.  * * O r i g i n a l l y , the s t r u c t u r e of the o v e r o x i d a t i o n product of l e u r o s i n e was i n c o r r e c t l y assigned as the lactam 77.63/64  - 85 -  catharanthus  (76).  Catharine  a l k a l o i d ,  The  of  s t r u c t u r e  85 C a t h a r i n e  h a d  b e e n  e l u c i d a t e d  an x - r a y  b y  a n a l y s i s  ,  w h i l e  19 CHO  76 the  i d e n t i t y  t a b l i s h e d and  by  of  77  t h e s y n t h e t i c  c o m p a r i s o n  w i t h  m a t e r i a l an  as  a u t h e n t i c  C a t h a r i n e sample  w a se s -  (cf.  Fig.  40  41) . The p r o d u c t i o n o f two n a t u r a l p r o d u c t s  coupled  t h e u n u s u a l s t r u c t u r e o f one o f them prompted f u r t h e r tion of their a radical fore,  formation.  and/or a c i d  Under t h e o r i g i n a l  c a t a l y s e d mechanism  a number o f r e a c t i o n s were  with  examina-  c o n d i t i o n s employed  was p o s s i b l e .  There-  c a r r i e d out (Table 4) i n order  t o d i s t i n g u i s h between t h e p o s s i b i l i t i e s . A e r i a l o r t - b u t y l hydroperoxide tetrahydrofuran rise  c o n t a i n i n g aqueous  to Catharine  respectively reaction  (Rx #i #2, #12} t  via thin  the p r e c u r s o r  ( c a 30% y i e l d )  layer  1% t r i f l u o r o a c e t i c  a f t e r e l e v e n and f i v e  M o n i t o r i n g t h e course  chromatography  of Catharine.  Accordingly, a e r i a l  Alternatively,  v i n b l a s t i n e or leurosine with  acid  gave  days  of the  i m p l i c a t e d l e u r o s i n e as  hydroperoxide oxidation of leurosine also (Rx #10 and #11).  o x i d a t i o n o f 26 i n  or t-butyl  afforded Catharine  t h e r e a c t i o n o f 3',4'-anhydro-  t - b u t y l hydroperoxide  c o u l d be  F i g . 40. Hmr spectrum of C a t h a r i n e (sample was o b t a i n e d from the E l i L i l l y L a b o r a t o r i e s , Indeanapolis).  - 88 -  Table 4.  Study of the a e r i a l and t - b u t y l hydroperoxide o x i d a t i o n s  of 3 4 - a n h y d r o v i n b l a s t i n e , 1  leurosine,  and d e r i v a t i v e s  thereof.  Rx# 1.  t-BuOOH  3',4 -anhydrovinblastine 1  THF,1%TFA(aq) 22 h 3' , 4 - a n h y d r o v i n b l a s t i n e 1  l e u r o s i n e (+++) C a t h a r i n e (±) 78 o r 79 (±)  t-BuOOH THF,1%TFA(aq) 5 days  l e u r o s i n e (+) C a t h a r i n e (++] 78  o r  79 (+)  ~ o t h e r s  3.  t-BuOOH  3',4 -anhydrovinblastine 1  THF-H 0 22 h 2  (+)  l e u r o s i n e  (+)  C a t h a r i n e  (+)  78  o r  79 (+)"  " o t h e r (+++)  t-BuOOH  3 ,4'-anhydrovinblastine 1  THF 22 h  t-BuOOH  3 ' , 4 ' - a n h y d r o v i n b l a s t i n e  THF,1%TFA(aq) MeOH 22 h 3 ,4'-anhydrovinblastine  t-BuOOH  1  THF,1%TFA(aq) rad. i n h i b . * rx c a r r i e d out i n dark  3 ' , 4'-anhydrovinblastine N i-oxide b  3',4'-anhydrovinblastine  t-BuOOH THF,1%TFA(aq) 22 h t-BuOOH THF,5 %TFA (aq) 22 h  S.M. (±) l e u r o s i n e (±) Catharine (++) 78 or 79 (±) Others (++) S.M. (++) l e u r o s i n e (++] 78 or 79 (±)  S.M. (+++) l e u r o s i n e (+) 78 or 79(+)  no rx  no rx  *The r a d i c a l i n h i b i t o r used was 3-t-butyl-4-hydroxy-5-methylphenyl s u l f i d e .  - 89 -  Table 4 con't Rx# 9.  leurosine  10.  leurosine  THF,5 %/TFA (aq) 22 h  11. leurosine 12. or 3 , 4 - a n h y d r o v i n b l a s t i n e 1  13.  t-BuOOH  t-BuOOH  leurosine  Air THF,1%TFA(aq) 11 days  leurosine  15.  leurosine  16.  leurosine-N, ,-oxide (PleurosineT  17.  leurosine  =2*  t-BuCOH cn ci 2  14.  S.M. (++) catharine (+) 79 ( + )  THF,1%TFA(aq) 22 h  1  2  t-BuOOH THF,1%TFA(aq) 22h, rad i n h * & r x i n dark 1%TFA(aq) THF t-BuOOH 44h, rad i n h * , * * rx i n dark t-BuOOH THF , 1%TFA (aq) 22 h Air CH C1 22 h 2  no rx  l e u r o s i n e (+) catharine (++) others (+) others (+) catharine (+++) (48% y i e l d ) S.M. (+++) catharine (±) 79 (±)  79  (+++)  no rx  no rx 2  *The r a d i c a l i n h i b i t o r used was 3-t-butyl-4-hydroxy-5-methylphenyl s u l f i d e . **The d i f f e r e n c e between t h i s r e a c t i o n and r e a c t i o n #14 (aside from the length of time) was i n the amount of r a d i c a l i n h i b i t o r used ( i . e . greater i n t h i s case).  - 90 -  significantly on  addition  inhibited,  of a radical  The  role  the absence  mixtures sine.  complex  (Rx #4  Finally,  a n d #4),  Catharine  a n d #13).  i n methylene  Although  and c o n t a i n e d  hand  l e u r o s i n e i n 48%  peroxide  (Rx #3  Indeed  yield  changed,  i n t h e o x i d a t i o n o f 3',4'—  acid  not clear.  of acid  On t h e o t h e r  or water from  were  was  o f the reaction  (Rx #6,#14,#15).  inhibitor  o f aqueous  anhydrovinblastine in  o r the course  l e u r o s i n e was  the resultant  formed  product  a low p r o p o r t i o n o f l e u r o formation  Catharine  by the a c t i o n  d i d not require c o u l d be  acid  obtained  of t-butyl  hydro-  chloride.  the intermediacy  of the N ,-oxides b  78* a n d  86  79  was, u n d e r  the reaction  c o n d i t i o n s employed,  78 Therefore, in (Rx  78  a n d 79 w e r e  their  a n d #16). intermediacy In  treated with  Since was  no r e a c t i o n  t-butyl 1%  hydroperoxide  trifluoroacetic  had o c c u r r e d  after  22  acid hours  ruled out.  summary, 3 , 4 - a n h y d r o v i n b l a s t i n e 1  1  *3 ' , 4 — A n h y d r o v i n b l a s t i n e — N j - ) < — o x i d e p r e p a r e d b y G.H. B o k e l m a n . 6 2,63 1  possibility.  79  t e t r a h y d r o f u r a n c o n t a i n i n g aqueous #7  a  c o u l d be o x i d i z e d  (7,9) h a d p r e v i o u s l y b e e n  - 91 -  to l e u r o s i n e i n the presence of peroxides presumably v i a a r a d i c a l mechanism.  Aqueous a c i d was necessary f o r a s y n t h e t i -  c a l l y u s e f u l conversion side r e a c t i o n s .  p o s s i b l y due to i n h i b i t i o n of unwanted  Catharine  was formed from l e u r o s i n e v i a a r a d -  i c a l mechanism with n e i t h e r a c i d nor water being intermediacy  were  a c h i e v e d  the appropriate  the f i r s t syntheses of l e u r o s i n e a n d from  conditions.  3  1  , 4 ' - a n h y d r o v i n b l a s t i n e  catharine  F i g . 42.  could be put f o r t h  1  u t i l i z i n g  of l e u r o s i n e  ( F i g . 42 and 43).  P o s s i b l e mechanism f o r the formation  3',4 -anhydrovinblastine.  by  Furthermore, based on the o x i d a t i o n  s t u d i e s , p o s s i b l e mechanisms f o r the formation and  The  of the N, .-oxides 7 8 and 79 was r u l e d out. b — ~~  In c o n c l u s i o n , C a t h a r i n e  required.  of l e u r o s i n e from  - 92  F i g . 43. leu rosine-  -  P o s s i b l e mechanism f o r the f o r m a t i o n  of c a t h a r i n e  from  - 93 -  2.3  Chemistry of 3 ' , 4 - A n h y d r o v i n b l a s t i n e — S y n t h e s i s 1  of Vinamidine*  (Catharinine) (82) and Related D e r i v a t i v e s .  F u r t h e r i n v e s t i g a t i o n s i n t o the f u n c t i o n a l i z a t i o n of 3 ,4 -anhydrovinblastine 1  1  (and l e u r o s i n e ) l e d to the d i s c o v e r y of  the potassium permanganate o x i d a t i o n as a p r o p i t i o u s  method.  Thus, from the r e a c t i o n of 3',4'-anhydrovinblastine with a s o l u t i o n of potassium permanganate i n acetone was obtained two 8T main products.  The minor product  (10%) was assigned as the 5'  *Vinamidine o r i g i n a l l y was i n c o r r e c t l y assigned as the C]_gi-C2i cleavage d e r i v a t i v e 80.3 6 Subsequently i t was found to be i d e n t i c a l with the a l k a l o i d c a t h a r i n i n e 8 8 — compound whose a  19  CHO  s t r u c t u r e was assigned as the C 4 1 - C 5 1 , cleavage d e r i v a t i v e 82 on the b a s i s of an x-ray a n a l y s i s of a degradation product.  - 94 -  lactam 81 based on a comparison with an a u t h e n t i c sample.* The major product  (42%) was i d e n t i f i e d  3R-hydroxyvinamidine  (ex post facto) as  ( 8 3 ) — a novel d e r i v a t i v e o f the n a t u r a l l y 88  o c c u r r i n g a l k a l o i d vinamidine  (82). The i d e n t i t y of t h i s  product was e s t a b l i s h e d v i a s e v e r a l chemical transformations and e x h i b i t e d s p e c t r a l p r o p e r t i e s i n agreement with the a s s i g n ment.  High r e s o l u t i o n mass spectrometry  confirmed the molecular  formula as C^gH^gN^O^^, while the presence of the  t-formyl  group was i n d i c a t e d by the strong bond a t 16 60 cm''" i n the -  i n f r a r e d spectrum  ( F i g . 44). Further evidence f o r the  i  —  formyl group was provided by the s i g n a l at 6 7.32 ( I H , s i n g l e t , N^i-CHO) i n the ^Hmr spectrum  ( F i g . 45).  The presence of a secondary  a l c o h o l was proven by  a c e t y l a t i o n of 83 using a c e t i c a n h y d r i d e / p y r i d i n e .  From t h i s  r e a c t i o n was obtained the ketoacetate 84 i n 65% y i e l d .  In accord  CHO  Me0 C 2  Vind. 84  * P r e v i o u s l y , t h i s compound had been prepared by G.H. Bokelman i n t h i s l a b o r a t o r y from the r e a c t i o n of 3 ,4'-anhydrovinblastine-N]-, i -oxide with one e q u i v a l e n t of osmium t e t r o x i d e . The s t r u c t u r a l assignment was based on a n a l y s i s of s p e c t r a l p r o p e r t i e s and should be considered t e n t a t i v e . 6 2,6 3 1  F i g . 44.  I n f r a r e d spectrum of 3R-hydroxyvinamidine (83).  - 97 -  with t h i s s t r u c t u r e , high r e s o l u t i o n mass spectrometry gave the molecular formula as 4 8 5g 4°]_2' C  H  N  w  n  i  i  e  t  n  ^Hmr  e  spectrum  (Fig. 46) s u b s t a n t i a t e d the presence of the new secondary t a t e with s i g n a l s a t & 4.80  (IH,broad sing l e t /  ace-  , -H) and cf 2.10  (3H,singlet,-OCOCH ). 3  The presence of a ketone  f u n c t i o n a l i t y was proven by  sodium borohydride r e d u c t i o n of 83 from which was obtained the triol  85 i n 6 6 % y i e l d .  A c c o r d i n g l y , high r e s o l u t i o n mass spec-  trometry gave the molecular formula as 46 5g 4°]_i' while the C  i n f r a r e d spectrum  H  N  confirmed the presence o f s e v e r a l a l c o h o l  f u n c t i o n a l i t i e s with a moderately  strong bond a t 3585 cm  i  F i g . 46.  Hmr  1  1  r  spectrum of 3R-acetoxyvinamidine  (8A-) .  - 99 -  A c e t y l a t i o n of the t r i o l The major product Confirming  85 y i e l d e d two products.  (25% y i e l d ) was assigned as the t r i a c e t a t e 86.  t h i s assignment, high r e s o l u t i o n mass  gave the molecular formula as 5 o 6 2 4 ° 1 3 ' C  (Fig. 47) r e v e a l e d the presence with s i g n a l s at<£4.89  H  N  w n :  *-  l e  t  spectrometry h  e  "*"  of two new secondary  Hmr  spectrum  acetates  ( l H , m u l t i p l e t , C , -H) , & 4 . 41 (IH, t r i p l e t , 4  J=6 Hz ,C ,-H) , & 2. 08 (3H, singlet,-OCOCH ) , and «£ 1.97 (3H, 3  3  singlet,_OCOCH ).  The minor product  3  as the t e t r a a c e t a t e 87.  (24% y i e l d ) was assigned  Thus, high r e s o l u t i o n mass  e s t a b l i s h e d the molecular  formula as C - H , . N . O . w h i l e the c  52  ^Hmr spectrum  spectrometry  64 4 14  ( F i g . 48) e x h i b i t e d s i g n a l s corresponding  to four  acetates at £ 2.08 (6H,singlet,2X-OCOCH ) and £ 1.98 (6H,singlet, 3  2X-OCOCH ). 3  O x i d a t i o n of 83 with c u p r i c acetate i n hot methanol a f f o r d e d the o<-diketone 88.  Since t h i s o x i d a t i o n i s a s p e c i f i c  MeCLC 2  ... . Vind.  88 89 method f o r the conversion of c<-ketols to o<.-diketones presence  of t h i s f u n c t i o n a l i t y  , the  (oC-ketol) i n 83 was e s t a b l i s h e d ,  In r e l a t i o n to c h a r a c t e r i z a t i o n o f the <X-diketone 88, high r e s o l u t i o n mass spectrometry C  4 6 5 4 4 ° 1 1 ' '-- H  N  wh:  1  e  t  h  e  gave the molecular  formula as  i n f r a r e d spectrum provided f u r t h e r e v i -  - 102 -  dence with a strong a b s o r p t i o n at 1713 cm  i n agreement with  the presence o f the diketone group. E l i m i n a t i o n o f the a l t e r n a t e cleavage s t r u c t u r e '89 as a p o s s i b i l i t y  f o r the potassium  permanganate product was  CHO  Me0 G 2  Vind.  89  accomplished d i o l 85. of  by p e r i o d i c a c i d o x i d a t i o n of the k e t o l 83 or the  The e x c l u s i v e nitrogenous product obtained from  these r e a c t i o n s was the aldehyde  90.  The ^"Hmr  both  spectrum  90  (Fig.  49) of 90 e x h i b i t e d one methyl  t r i p l e t at £ 0.74  r  as w e l l  as s i n g l e t s at &7.61 and &9.20 corresponding to the N^-formyl and a l d e h y d i c hydrogens r e s p e c t i v e l y .  High r e s o l u t i o n mass  trometry e s t a b l i s h e d the molecular formula as 4 3 5 o 4 ° i o ' C  confirming the l o s s of a three carbon  fragment..  H  N  t  spech  u  s  F i g . 49.  Hmr  spectrum of aldehyde 90.  - 104 -  F i n a l l y , the stereochemistry a t C-3 was assigned 1  as R on the b a s i s of the r e a c t i o n of l e u r o s i n e with permanganate.  potassium  From t h i s r e a c t i o n were obtained two products.  The major product  (27% y i e l d ) was found t o be i d e n t i c a l with 83.  Since the absolute c o n f i g u r a t i o n o f l e u r o s i n e was known, the assignment of the absolute c o n f i g u r a t i o n of 83 was based on the assumption that the stereochemical i n t e g r i t y of C-3' was maintained. The minor product oxoleurosine  (19% y i e l d ) was assigned as 1 9 ' —  (91) on the evidence of i t s s p e c t r a l p r o p e r t i e s .  High r e s o l u t i o n mass spectrometry as C.,H .N . 0 , w h i l e 46 54 4 10 r  a f f i r m e d the molecular  the i n f r a r e d spectrum ^  a strong absorption a t 1644 cm  formula  ( F i g . 50) contained ^  i n accord with the presence o f  1  13 an amide c a r b o n y l . 61.6  and  The  Cmr spectrum e x h i b i t e d s i g n a l s a t  59.8 corresponding  to C-3  1  and C-4'.  The lack o f any  s i g n i f i c a n t change i n the p o s i t i o n of these s i g n a l s r e l a t i v e to the analgous s i g n a l s i n l e u r o s i n e (£60.3 and ^59.9) corroborated the l9'-oxo spectrum  (rather the 5'-oxo) assignment.  F i n a l l y , the "*"Hmr  ( F i g . 51) contained a s i g n a l a t £ 4 . 7 6  which was a t t r i b u t a b l e to the C •-hydrogen. 2  (IH,multiplet)  F i g . 51.  Hmr  spectrum of 19'-oxoleurosine (91).  - 107 -  Having of  vinamidine  3R-hydroxvinamidine a v a i l a b l e , a s y n t h e s i s  (82) was  sought.  Since various methods f o r de-  oxygenation of a l c o h o l s and t h e i r d e r i v a t i v e s were a v a i l a b l e , t h i s route was  initially  investigated.  However,  attempted  formation of the thiobenzoate 9 2 — a d e r i v a t i v e which normally CHO  OCII Me0 C  Vind.  2  92 90 should be e a s i l y deoxygenated of  decomposition  — r e s u l t e d only i n the formation  products, while the use of the acetate 84 as a  s u b s t r a t e f o r deoxygenation r e a c t i o n s (such as z i n c metal i n hot 91 92 acetic acid or l i t h i u m dimethyl cuprate i n t e t r a h y d r o f u r a n ) a f f o r d e d s t a r t i n g m a t e r i a l (84) and/or a l c o h o l 83.  Therefore,  an a l t e r n a t i v e means of s y n t h e s i z i n g vinamidine, v i a extension of  the potassium permanganate r e a c t i o n was  ment of 4 - d e o x y l e u r o s i d i n e 1  (93)  undertaken.  ( a v a i l a b l e from 3',4'-anhydro31 33  v i n b l a s t i n e by hydrogenation over a platinum c a t a l y s t vinamidine i n 22% y i e l d . was  '  ) afforded  The i d e n t i t y of the s y n t h e t i c m a t e r i a l  based on a comparison with an a u t h e n t i c sample  and 53).  Treat-  (cf F i g . 52  Moreover, the o p t i c a l r o t a t i o n of the sodium borohydride  r e d u c t i o n product 94 was 88 ature value.  found to be i n agreement with the  liter-  F i g . 53.  Hmr  spectrum of s y n t h e t i c vinamidine.  - no  -  A l s o o b t a i n e d from t h i s r e a c t i o n was t h e lactam 95 i n 11% y i e l d .  I n agreement w i t h t h i s s t r u c t u r e h i g h r e s o l u t i o n  95 mass spectrometry gave t h e m o l e c u l a r formula as C^gH^gN^Og w h i l e the "''Hmr spectrum  ( F i g . 54) e x h i b i t e d a s i g n a l a t £ 4.84 ( I H ,  m u l t i p l e t ) a t t r i b u t a b l e t o the  ,-hydrogen.  F i n a l l y , the i n f r a -  red spectrum possessed an a b s o r p t i o n a t 1640 cm  1  corresponding  t o the presence of the amide f u n c t i o n a l i t y . In summary, potassium permanganate o x i d a t i o n o f v a r i 93 ous a v a i l a b l e 'dimers  1  gave r i s e t o 'normal  1  lactam products  (81,91 and 95) as w e l l as t o 'abnormal'  C^,-C^, cleavage  products  (vinamidine and 3R-hydroxyvinamidine).  The mechanism f o r f o r -  mation o f these cleavage p r o d u c t s remains ambiguous, however, p o s s i b i l i t i e s have been suggested i n F i g . 55, 56 and 57.  3.82  -  Fig.  55.  from  4'-deoxyleurosidine.  112  -  P o s s i b l e mechanism f o r t h e f o r m a t i o n o f  vinamidine  -  113  -  OH  F i g . 56.  P o s s i b l e mechanism f o r the formation of 3R-hydroxyvina-  midine from  3',4 -anhydrovinblastine. 1  3.:— Hy droxyvin am i d ine F i g . 57.  P o s s i b l e mechanism f o r the formation of 3R-hydroxyvina-  midine from l e u r o s i n e .  - 114 -  2.4  P r e p a r a t i o n o f V i n b l a s t i n e - V i n c r i s t i n e Analogs. A s i d e from the chemistry  synthesis of n a t u r a l products,  developed i n r e l a t i o n t o the  a number of r e a c t i o n s were c a r r i e d  out i n order t o o b t a i n n o v e l analogs o f v i n b l a s t i n e and v i n c r i s t i n e f o r the purpose o f e l u c i d a t i n g s t r u c t u r e - a c t i v i t y r e lationships.  I n g e n e r a l , the p r o d u c t i o n  good o v e r a l l c o n v e r s i o n  o f analogs r e q u i r e d a  of r e a d i l y a v a i l a b l e materials.  Pre-  v i o u s l y , i t was seen t h a t potassium permanganate o x i d a t i o n access t o s e v e r a l i n t e r e s t i n g lactams.  afforded  The low y i e l d s o f lactams  produced by t h i s method prompted use o f o t h e r c o n d i t i o n s t o achieve improved r e s u l t s .  Accordingly,  i t was determined t h a t  o x i d a t i o n w i t h i o d i n e , i n aqueous t e t r a h y d r o f u r a n  containing  94  sodium b i c a r b o n a t e c o n s t i t u t e d such an improvement. Thus, l e u r o s i n e c o u l d be c o n v e r t e d t o 1 9 - o x o l e u r o s i n e i n 56% y i e l d . 1  63,64  Extension  o f t h i s r e a c t i o n t o v i n b l a s t i n e and l e u r o s i d i n e *  a f f o r d e d the c o r r e s p o n d i n g lactams 96 and 97 i n y i e l d s o f 33% and 62% r e s p e c t i v e l y .  The s p e c t r a l p r o p e r t i e s a s s o c i a t e d  with  V i n b l a s t i n e and l e u r o s i d i n e were s u p p l i e d by the E l i L i l l y laboratories, Indianapolis.  - 115 -  the lactams were i n accord with the s t r u c t u r a l assignments are p a r t l y summarized i n Table 5. . C=0 of lactam i n i . r . soectrum  Compound 91  16 44 cm  96  1640  cm"  1  97  1644  cm"  1  1  and  ,  Cp'-H i n iHmr spectrum  Molecular formula (from high r e s . ms.)  4. 76  C  46 54 4°10  4.70  C  46 56 4°10  4. 73  C  46 56 4°10  H  H  H  N  N  N  Table 5.  Comparison of some c h a r a c t e r i z a t i o n data f o r 19'-oxo-  leurosine  (91), 1 9 - o x o v i n b l a s t i n e (96), and 1 9 - o x o l e u r o s i d i n e 1  Attempted  1  extension of the iodine/sodium bicarbonate  o x i d a t i o n to 4 - d e o x y l e u r o s i d i n e was  not s u c c e s s f u l .  1  of the a n t i c i p a t e d product  Instead  (19'-oxo-4'-deoxyleurosidine)  d e r i v a t i v e of unknown s t r u c t u r e was was  (97)  obtained.  a  new  This derivative  not a lactam as evidenced by i t s i n f r a r e d spectrum  ( F i g . 5 8).  High r e s o l u t i o n mass spectrometry gave the molecular formula as C^gH^N^Og while the ''"Hmr spectrum n a l at c/8.78 ( I H , s i n g l e t ) which was  ( F i g . 59) possessed a s i g -  s p e c i f i c to t h i s  'dimer'.  The s t r u c t u r e of t h i s product remains mysterious, however e f f o r t s aimed at i t s e l u c i d a t i o n continue. Having a v a i l a b l e v a r i o u s analogs of v i n b l a s t i n e , a t t e n t i o n was  turned to the p r e p a r a t i o n of analogs of v i n c r i s t i n e .  Treatment reagent  95*  of 19'-oxoleurosine i n acetone at -7 8°C with l e d to the i s o l a t i o n of two products.  96 97 '  Jones The major  *This procedure represents an adaptation of the chromic a c i d o x i d a t i o n reported by Jovancvics et- 'al- -for the conversion of v i n b l a s t i n e to v i n c r i s t i n e . ^ T h i s adaptation was o r i g i n a l l y developed by T. Honda and P.H. L i a o i n t h i s l a b o r a t o r y . 5  F i g . 58. I n f r a r e d spectrum of the product obtained from the iodine/sodium bicarbonate o x i d a t i o n of 4'-deoxyleurosidine.  3.64  F i g . 59. Hmr spectrum of the product obtained from the i o d i n e / sodium bicarbonate o x i d a t i o n of 4'-deoxyleurosidine.  - 118 -  product  (29% y i e l d ) was a s s i g n e d as t h e N ~ f o r m y l  d e r i v a t i v e 98.  a  Accordingly, the u l t r a v i o l e t  spectrum  of this  derivative  ( F i g . 60)  4.5H 4.5-  4.CH 4.0 H  300  250  \  I  250  nm  300  A  F i g . 60. C o m p a r i s o n o f u l t r a v i o l e t and 1 9 , 2 2 - d i o x o l e u r o s i n e ( B ) .  nm  spectra of 19 -oxoleurosine 1  (A)  1  e s t a b l i s h e d the presence t i o n mass s p e c t r o m e t r y  o f t h e new c h r o m o p h o r e .  C  ( F i g . 61) l a c k e d a s i g n a l  to  Finally,  group.  resolu-  gave t h e m o l e c u l a r f o r m u l a as 4 g 5 2 4 ~ i i '  w h i l e t h e ^Hmr s p e c t r u m an N - m e t h y l  High H  N  <  )  attributable  t h e i n f r a r e d spectrum  ( F i g . 62)  e x h i b i t e d a s t r o n g a b s o r p t i o n a t 16 7 8 cm ^ i n a c c o r d w i t h t h e presence  o f the N - f o r m y l group. 3.  The  minor product  N ~ d e s m e t h y l d e r i v a t i v e 99. a  (17% y i e l d ) was a s s i g n e d a s t h e A c c o r d i n g l y , h i g h r e s o l u t i o n mass  s p e c t r o m e t r y v e r i f i e d t h e m o l e c u l a r f o r m u l a as 4 5 5 2 4 ° ] _ o ' C  the  Hmr s p e c t r u m  H  N  wn;  ( F i g . 6 3) l a c k e d a s i g n a l f o r an N - m e t h y l  "-  xe  group,  - 11-9 -  91, R=Me 98, R=CHO 99, R=H  A l t e r n a t i v e l y , a more e f f i c i e n t r o u t e f o r the p r e p a r a t i o n o f 19 ,22-dioxoleurosine 1  (98) was achieved by f i r s t c o n v e r t i n g  leurosine t o 22-oxoleurosine*  (37) (75% y i e l d ) .  The i d e n t i t y o f t h i s product was c o r r o b o r a t e d by the u l t r a v i o l e t spectrum which was i n d i c a t i v e o f the new chromophore as w e l l as by h i g h r e s o l u t i o n mass spectrometry which confirmed  *Jones reagent was again used t o e f f e c t t h i s t r a n s f o r m a t i o n r a t h e r than the c o n d i t i o n s g i v e n by J o v a n o v i c s e t a l "  F i g . 62.  I n f r a r e d spectrum  of 19 ,22-dioxoleurosine 1  (98).  3.81  - 123 -  2 2  CHO  C0 Me 2  37  the.molecular formula as C, H .N .0, . 46 54 4 10 C  C  n  O x i d a t i o n of 37 with  i o d i n e i n aqueous t e t r a h y d r o f u r a n c o n t a i n i n g sodium bicarbonate y i e l d e d 19 ,22-dioxoleurosine 1  (67% y i e l d ) i d e n t i c a l with the  m a t e r i a l obtained p r e v i o u s l y . Finally, blastine  u t i l i z a t i o n of  6,7-dihydro-3 ,4 -anhydrovin1  1  (100) as a substrate gave r i s e to two more novel  analogs  Thus, e p o x i d a t i o n of 100 using the p r e v i o u s l y developed c o n d i t i o n ( t - b u t y l hydroperoxide,  t e t r a h y d r o f u r a n , aqueous 1% t r i f l u o r o -  a c e t i c acid) a f f o r d e d 6,7-dihydroleurosine High r e s o l u t i o n mass spectrometry  a f f i r m e d the molecular  as C gH^gN O , while the ^"Hmr spectrum 4  resonances  4  g  due to v i n y l  hydrogens.  (101) i n 41% y i e l d .  lacked any  formula  -  -  124  Oxidation of 101 with Jones reagent as before a f f o r d e d 22-oxo-6,7-dihydroleurosine  (102) i n 72% y i e l d .  In accord with  t h i s s t r u c t u r e , high r e s o l u t i o n mass spectrometry gave the molec u l a r formula as 4 6 5 g 4 ° i o ' C  H  N  wn;  "-  le  s t a n t i a t e d the presence of the new  t  n  e  u l t r a v i o l e t spectrum  sub-  chromophore.  F i n a l l y , the p r o d u c t i o n of the various d e r i v a t i v e s o u t l i n e d represented a p o r t i o n of an ongoing program. biological results in relation  Preliminary  to the a n t i c a n c e r p r o p e r t i e s of  s e v e r a l analogs have been obtained, however, the e l a b o r a t i o n of t h i s data was  not p o s s i b l e at t h i s  time.  - 125 -  EXPERIMENTAL  M e l t i n g p o i n t s (mp) were determined and are u n c o r r e c t e d .  U l t r a v i o l e t (uv) s p e c t r a were recorded  on a Cary 15 spectrophotometer i n ethanol solution.  on a K o f l e r b l o c k  and were o b t a i n e d w i t h samples  The a b s o r p t i o n maxima are r e p o r t e d i n  nanometers (nm) w i t h l o g <£ v a l u e s i n parentheses.  Infrared ( i r )  s p e c t r a were r e c o r d e d on a P e r k i n Elmer model 457 s p e c t r o p h o t o meter and were o b t a i n e d w i t h samples i n c h l o r o f o r m s o l u t i o n . The a b s o r p t i o n maxima are r e p o r t e d i n wavenumbers  ( c m ) , and - 1  are c a l i b r a t e d w i t h r e s p e c t t o the a b s o r p t i o n band of p o l y s t y r e n e a t 1601 cm . 1  P r o t o n magnetic resonance (''"Hmr) s p e c t r a were  r e c o r d e d on a V a r i a n HA-10 0, XL-100, NTCFT-100, o r Bruker-270 MHz spectrometer.  S p e c t r a were o b t a i n e d w i t h samples i n deutero-  c h l o r o f o r m s o l u t i o n a t ambient temperature..  Chemical  s h i f t values  are g i v e n i n the £ (ppm) s c a l e r e l a t i v e to t e t r a m e t h y l s i l a n e which was used as the i n t e r n a l s t a n d a r d .  The i n t e g r a t e d peak  a r e a s , s i g n a l m u l t i p l i c i t i e s , and proton assignments are g i v e n i n parentheses.  Low r e s o l u t i o n mass s p e c t r a (ms) were  determined  on e i t h e r an AEI-MS-902 o r an a t l a s CH-4B spectrometer.  High  r e s o l u t i o n mass s p e c t r a were measured on an AEI-MS-902  instru-  ment.  C i r c u l a r d i c h r o i s m (cd) curves were recorded .on a Jasco  - 126 -  J-20  s p e c t r o p o l a r i m e t e r and were o b t a i n e d w i t h samples i n  methanol s o l u t i o n .  The  curves were c a l i b r a t e d w i t h 10-camphor-  s u l f o n i c a c i d and were o b t a i n e d a t 25.0±0.1°C.  The wavelengths  of a b s o r p t i o n maxima are r e p o r t e d i n nanometers w i t h A £ v a l u e s 13 C-magnetic resonance ( 13 Cmr) s p e c t r a were i n parentheses. o b t a i n e d on a V a r i a n CFT-20 spectrometer decouple mode.  Samples were i n d e u t e r o c h l o r o f o r m s o l u t i o n  were measured a t ambient temperature. are g i v e n i n the was  used i n the p r o t o n  Chemical  and  s h i f t values  s c a l e r e l a t i v e to t e t r a m e t h y l s i l a n e which  used as the i n t e r n a l s t a n d a r d .  g i v e n i n parentheses.  The carbon assignments are  M i c r o a n a l y s e s were c a r r i e d out by  Mr. P. Borda of the M i c r o a n a l y t i c a l L a b o r a t o r y , U n i v e r s i t y o f B r i t i s h Columbia. T h i n l a y e r chromatography ( t i c ) u t i l i z e d Merck gel  G (acc. t o S t a h l ) c o n t a i n i n g 2-3%  fluorescent indicator.  For p r e p a r a t i v e l a y e r chromatography ( p i c ) , p l a t e s (20x20 of one mm  t h i c k n e s s were used.  silica  V i s u a l i z a t i o n was  cm)  e f f e c t e d by  v i e w i n g under u l t r a v i o l e t l i g h t and/or by c o l o r r e a c t i o n w i t h e e r i e s u l f a t e spray reagent. Merck s i l i c a g e l 60  Column chromatography u t i l i z e d  (70-230 mesh) o r Merck aluminum o x i d e  90  (neutral) . P o r t i o n s o f the f o l l o w i n g work have been p u b l i s h e d p r i o r to  the completion of t h i s t h e s i s .  c o n t i n u i t y and convenience  However, f o r the sake of  these experiments  have been i n c l u d e d .  -  127  16,18S-Dicarbomethoxycleavamine  (502 a  mg,  A  solution  1.5  mmole)  nitrogen  hydride  was  f o r 10  treated  stirred  for a  stirring and  the  ether-benzene)  1 h.  43  (19)  (50 m l )  under  a suspension of potassium KH,  1.7  mmole).  After  temperature, the r e s u l t a n t  chloroformate Acetic  (29 0 mg,  acid  The  on  3.1  (0.5 ml)  solvent  chromatographed  to afford  7 8 % ) , mp  with  22.5%  f o r 5 min.  was  tetrahydrofuran  treated  a t ambient  methyl  continued  ( 4 6 1 mg,  was  further  residue  18S-carbomethoxycleavamine  o i l (300 mg,  min  with  (39)  i n anhydrous  atmosphere  i n mineral  stirring  of  -  was  mmole)  and  was,added  removed  alumina  mixture  in  and vacuo  (Act.. I l l , p e t .  16,18S-dicarbomethoxycleavamine*  124-125°C  uvA  (acetone-ether);  :  (39)  294  (3.66),  ITLclX  283  sh  1725;  (3.80),  268  iHmr  5.77  8.08  ( I H , d,  - O C H ) , 3.53 396  (-C0 CH ), 2  3  J=6  s,  C  (4.11),  ~ H ) , 7.23  1 4  (3H,  t , J=7  136,  124;  0.98  3  (base p e a k ) ,  (-C0 CH ), 2  141.1  3  1 Q  ),  124.3  (C  1 2  ) , 122.8  118.5  (C  1 3  ),  115.8  (C  1 4  ),  ( C ) , 51.8  (-OCH ) , 38.6  (-CH CH ),  12.7  3  54.5 (C  3  determination  (-CH CH ).  calcd.  2  (3H, m,  -OCH_ ),  (C  2  (4.37);  ( I H , m,  129.9  5  228  C-^-H) , 5.37  (M+) , 273 152.0  262  ( I H , m,  Hz,  (3H,  3  ms: m/e  (4.09),  C  (C  l g  ),  ) , 35.9  N  Cmr  137.7  (C  53.3  :  -  2 0 4 8  2  : 1 ?  ),  3  173.7 136.0  & -OCH ),  7  3  (C- ), 2  molecular  '"  s,  -CH CH_ ) ;  3  (Cg) , 34.8  3 9 6  (3H,  ( C ) , 119.5 (C  resolution  3 28 2°4 H  2  Hz,  1 3  :  C ^ - C ^ - H ' s) ,  , 3.88  3  ( C ^ ) , 122.3  High  3  for  l g  (C^),  C ~H)  i r V  found:  (C  1 5  (C ), g  52.5  27.6 weight 396 . 2 0 7 0 .  * T h i s compound had p r e v i o u s l y b e e n p r e p a r e d i n t h i s laboratory b y I . I t o h a n d A.H. R a t c l i f f e u t i l i z i n g a d i f f e r e n t p r o c e d u r e . The c h a r a c t e r i z a t i o n d a t a f o r t h i s compound, w i t h t h e e x c e p t i o n o f t h e 1 3 c m r s p e c t r u m , was c o m p i l e d b y t h e s e w o r k e r s .  ),  - 128 -  Anal.calcd. for 3 2 8 2 ° 4 C  H  N  :  C  6  9  2  -  6  7  ;  H  7  - '* 12  N  7  -  0 7  ?  found:  C 69.86; H 7.10; N 6.87.  A u t o x i d a t i o n of 16,18S-dicarbomethoxycleavamine (39)  A s o l u t i o n of 16,18S-dicarbomethoxycleavamine (400 mg, 1 mmole) i n p e r o x i d e - f r e e t e t r a h y d r o f u r a n t a i n i n g aqueous  1% t r i f l u o r o a c e t i c a c i d  (10 ml) con-  (1 ml) was s t i r r e d a t  ambient temperature i n the presence of a i r f o r a p e r i o d of 8 days.  After drying  Chromatography  (K^CO^), the s o l v e n t was removed i n vacuo.  o f the residue on alumina (Act. I l l ,  benzene)  4 0 * (40 mg, 10%) and the ketoenamide 4 5  a f f o r d e d the epoxide (230 mg, 54%).  16,18S-dicarbomethoxy-3R,4S-epoxydihydrocleavamine mp 131-132°C  (40),  (ether); uv A , : 294 (3.66), 283 sh (3.76), 268 m  v  UIGLX  (4.08), 262 (4.09), 227 (4.32);  ir *  1113. X  :  1728; Hmr <£: 8.1 X  (IH, m, C -H) , 7.3 (3H, m, C^-C-^-H ' s) , 5.84 (IH, d, J=6 Hz, 14  C -H) , 3.91 (3H, x, -OCH ) , 3.57 (3H, s, -OCH_ ), 1.00 (3H, t , 18  3  3  J=7.5 Hz, -GH CH_ ); ms: m/e 412 (M ) , 224 (base peak), 152, 138; +  2  13  3  C m r S: 173.4 (-C0 CH ), 152.0 (-C0 CH ), 137.1 ( C ) , 136.0 2  3  2  1 ?  (C ),  129.8 ( C ) ,  (C ),  115.9 ( C ) , 62.7 ( C ) , 60.6 ( C ) , 53.4 ( C & -OCH ),  1 5  1 3  1 Q  124.5 ( C ) ,  3  ± 2  1 4  122.9 (C.^), 119.5 ( C ) , 118.2 g  4  3  ?  52.7  ( C ) , 51.9 (-OCH ), 50.6 ( C ) ,  30.0  (-CH CH ) , 26.3 (C-^ , 8.9 (-CH CH ) .  5  3  2  3  39.5 ( C ) ,  i g  l g  2  3  c u l a r weight determination c a l c d . f o r C ^ H O  3  33.6 ( C & C ) , g  2  High r e s o l u t i o n moleO O  N T 0 : 412.1997; c  *This compound was o r i g i n a l l y i s o l a t e d as a decomposition product of 3 9 by I . Itoh and A.H. R a t c l i f f e . The c h a r a c t e r i z a t i o n data l i s t e d here was compiled by these workers.  - 129 -  f o u n d : 412.2027. N  Anal.calcd.  for  C  3 28 2°5 H  2  N  :  C  6  6  -  9  7  ;  H  -  6  8 4  ?  6.79; f o u n d : C 66.81; H 6.87; N 6.71. Ketoenamide  281  s h (3.78),  45  265 (4.05),  (as a foam); uv *  : 293 (3. 67), max ' 227 (4.38); i r V :  259 (4.06),  rricLX  1732,  1680, 1650; Hmr <£: 8.13 (IH, s, N -CHO), 8.10 (1H, m, 1  b  C -H),  7.3 (3H, m, C ^ - C ^ - H ' s ) ,  l 4  (3H,  5.23 (IH, b s , Cg-H), 4.00  b s , -OCH_ ) , 3.66 (3H, s, -OCH_ ) , 1.90 (2H, q, J=7 Hz, 3  3  -CH_ CH ) , 0.79 (3H, t , J=7 HZ, -CH CH_ ); ms: m/e 426 ( M , base +  2  3  peak),  2  394, 228; C m r  2  (C  1 3  (C  ),  ),  1 2  1 5  1 1  ),  3  1 Q  122.7 (C^), 119.9 ( C ) , 118.9 g  1 4  12.3  (-CH CH ).  3  44.3 ( C ) ,  3  2  2  2 3  H  l g  2 6  2  3  ),  30.3 (Cg),  High r e s o l u t i o n molecular  3  for C  40.0 ( C  2 6  N O : 2  g  6  ?  24.5 (_CH CH ), 2  weight  426.1791; f o u n d : 426.1795.  C H N 0 . l i C H O H : C 63. 80; 2 3  2  116.0 ( C ) , 53.6 (-OCH ) , 52.5 (-OCH ) , 48.7 (C ) ,  (C ),  for  1 7  123.3 ( C  44.9  calcd.  b  ( C ) , 132.3 ( C ) , 128.8 ( C ) , 126.9 (Cg) ,  3  125.1  188.8 (N CHO), 163.0 ( - c p C H ) , 160.7  1 3  (-C0 CH ) , .135.9  3  3  3  determination Anal.calcd.  H 6.33; N 6.33; f o u n d : C 64.08;  H 6.11; N 6.31.  Oxidation 1  o f 16,18S-dicarbomethoxycleavamine  (39)  in  preoxidized'*tetrahydrofuran A s o l u t i o n o f (39) (900 mg, 2.27 -mole) i n ' p r e oxidized  1  tetrahydrofuran  fluoroacetic  acid  (20 ml) c o n t a i n i n g aqueous 1% t r i -  (2 ml) was s t i r r e d  a t ambient t e m p e r a t u r e f o r  * ' P r e o x i d i z e d ' t e t r a h y d r o f u r a n r e f e r s t o t e t r a h y d r o f u r a n which had undergone a e r i a l o x i d a t i o n and c o n t a i n e d an u n s p e c i f i e d amount o f p e r o x i d e s .  - 130  20 hr.  The  r e a c t i o n mixture  was  s o l u t i o n of sodium bicarbonate e t h y l acetate  (2x40 ml).  The  washed with water (3x15 ml) (Na SC> ) the s o l v e n t was 2  4  -  poured i n t o a s a t u r a t e d (20 ml)  and e x t r a c t e d with  combined organic p o r t i o n  and b r i n e (1x20 ml).  (Act. I l l ,  epoxide  and the ketoenamide  26%).  (306 mg,  A f t e r drying  removed i n vacuo and the residue  chromatographed on alumina (40)  was  33%)  benzene) to a f f o r d (45)  the  (250  These compounds were i d e n t i c a l to the products  was  mg,  obtained  above.  O x i d a t i o n of 16,18S-dicarbomethoxycleavamine t-butyl  (39) using  hydroperoxide A s o l u t i o n of 39  (1.76  g, 4.4 mmole) i n f r e s h l y  d i s t i l l e d tetrahydrofuran  (50 ml)  fluoroacetic acid  and t-buty hydroperoxide  (10 ml)  c o n t a i n i n g aqueous 1%  s t i r r e d at ambient temperature f o r 21 h. was  The  tri-  (9 ml)  was  r e a c t i o n mixture  poured i n t o a saturated s o l u t i o n of sodium bicarbonate  (40 ml)  and e x t r a c t e d with e t h y l acetate  organic p o r t i o n was (1x20 ml), water  washed with 5% sodium hydroxide  (1x20 ml), and b r i n e  (Na S0 ) the s o l v e n t was 2  4  Chromatography on alumina epoxide 40 (1.22  (2x30 ml).  g, 67%)  (1x20 ml).  The  combined  solution  A f t e r drying  removed i n vacuo to give a viscous o i l . (Act. I l l ,  benzene) a f f o r d e d the  i d e n t i c a l with that obtained  before.  - 131 -  16,18S-Dicarbomethoxy-4S-dihydrocleavamine  (41 )  To a s o l u t i o n o f 16,18S-dicarbomethoxy cleavamine (39) (15 mg, 0.038 mmole) i n 95% ethanol Adam's c a t a l y s t  (2 mg, 0.009 mmole).  (2 ml) a t 5°C was added  The r e a c t i o n mixture was  hydrogenated a t atmospheric pressure f o r 1 h, a t which p o i n t the  s o l v e n t was removed i n vacuo and the residue was chromato-  graphed on s i l i c a g e l t o y i e l d dihydrocleavamine uv * ir ^  m a x  16,18S-dicarbomethoxy-4S—  (41) (14 mg, 93%) as a c o l o r l e s s  film;  : 2 9 3 (3.72), 283 (3.80), 262 (4.08), 226 (4.34); : 2962, 1732; Hmr i : 8.17 (IH, m, C^-H) , 7.6-7.2 1  m a x  (3H, m, C - C - H ' s ) , 5.66 (IH, m, C^-H) , 3.96 (3H, s, -OCH_) , 11  14  3  3.64 (3H, s, -OCH ) , 0.90 (3H, t , J=7 Hz, -CH CH ); ms: m/e 398 3  (M ), +  2  3  396, 210 (base peak), 114. High r e s o l u t i o n molecular  weight determination c a l c d . f o r C ^ H ^ N ^ :  398. 2206; found:  398.2231.  Hydroxenamide 47 A s o l u t i o n of the ketoenamide 45 (10 mg, 0.023 mmole) i n 95% ethanol mg, 0.1 mmole).  (2 ml) was t r e a t e d with sodium borohydride (4 The r e a c t i o n mixture was s t i r r e d a t ambient  temperature f o r 20 min, taken up i n water with methylene c h l o r i d e  (3x5 ml).  (10 ml) and e x t r a c t e d  A f t e r drying  (Na„S0.), the  - 132 -  s o l v e n t was removed i n vacuo and the residue chromatographed on s i l i c a g e l (ether) to a f f o r d the hydroxyenamide as a f i l m , uv A  : 293  47 (6 mg,  (3.50), 281 sh (3.68), 262  60%)  (4.03),  ITISX  224  (4. 32); i r t>  : 3530, 3420 , 1730 , 1669 , 1651;  ^mr  IU3.X  8.05  (IH, m, C - H ) , 14  C - C - H ' s ) , 5.48 11  13  (M ), 315, 201, 126 +  (IH, s, N^CHO), 7.6-7.2 (3H, m,  (IH, bs, C -H) , 4.00 5  (3H, s, -OCH_) , 0.98 3  8.02  (3H, s, -OCH_ ), 3.66 3  (3H, t , J=7.5 Hz, -CH CH_ ) ; ms: m/e 2  (base peak).  428  High r e s o l u t i o n molecular  weight determination c a l c d . f o r 2 3 2 8 2 ° 6 C  3  H  N  :  428.1947; found:  428.1928.  16,18S-Dicarbomethoxy-3R-hydroxyvelbonamine A.  Using p e r c h l o r i c  (51)  acid  A s o l u t i o n of the epoxide 40 (25 mg, tetrahydrofuran  0.6 mmole) i n  (2 ml) c o n t a i n i n g aqueous 25% p e r c h l o r i c  acid  (0.25 ml) was heated at 66°C under a n i t r o g e n atmosphere f o r 2 4 h.  The r e a c t i o n mixture was cooled to ambient temperature,  c a u t i o u s l y added to c o l d concentrated ammonium hydroxide and e x t r a c t e d with methylene c h l o r i d e  (3x8 ml).  The combined  organic p o r t i o n was washed with 5% sodium bicarbonate (1x8 ml) and d r i e d  (Na„SO.).  (10 ml),  solution  Removal of the s o l v e n t i n vacuo  - 133 -  a f f o r d e d a brown foam which was  shown by t i c to c o n s i s t of a  r a t h e r complex mixture of products. (Act. I l l , 17%), mp  Chromatography on  ether) a f f o r d e d the t r a n s - d i o l  153-157°C (ether); uv A  : 294  51 (R  alumina  0.35) (4.5  f  (3.66), 282  sh  mg,  (3.76),  in 9.x  263  (4.09), 227  (4.37); i r  insx  : 3600, 3440, 1728;  (IH, m, C -H) , 7.5-7.2 (3H, m,  ^ - C ^ - H ' s ) , 6.54  14  J=8.5 Hz, C -H) , 3.97 lg  C -H) , 3.59  3  (3H, s, -OCH_) , 0.97  3  ms: m/e  (3H, S, -OCH_) , 3.63  (M ) , 315, +  242  Hmr  c£:  8.14  (IH, d,  (IH, d, J-2  Hz,  (3H, t , J=7.5 Hz, -CH CH_ ) ;  3  430  1  2  (base peak),170, 158,  156.  3  High  r e s o l u t i o n molecular weight determination c a l c d . f o r  C„-,H- N_0-: 23 30 2 6 ri  430.2104; found:. 430.2112. B.  Using t r i f l u o r o a c e t i c  acid  A s o l u t i o n of the epoxide  40 (108 mg,  0.262 mmole)  i n aqueous 52% t r i f l u o r o a c e t i c a c i d  (10 ml) was  heated at  6 5°C under a n i t r o g e n atmosphere f o r 4 h. was  cooled to ambient temperature,  concentrated ammonium hydroxide methylene c h l o r i d e was  dried  (3x15 ml).  The r e a c t i o n  mixture  c a u t i o u s l y added to c o l d  (50 ml), and e x t r a c t e d with The combined organic p o r t i o n  (K^CC^) and the s o l v e n t was  removed i n vacuo•  Chromatography of the r e s i d u e on s i l i c a g e l (ether) a f f o r d e d the t r a n s - d i o l 51  (28 mg,  25%)  i d e n t i c a l with the product  obtained above. 16,18S-Dicarbomethoxy-3S,4R-epoxydihydrocleavamine A s o l u t i o n of the t r a n s - d i o l in pyridine  (2 ml)  51  (13 mg,  (52) 0.03  c o n t a i n i n g methanesulfonyl c h l o r i d e  mmole) (3 drops)  - 134 -  was s t i r r e d  a t ambient temperature under a n i t r o g e n  for  The r e a c t i o n mixture was poured i n t o 5% sodium  2.5 h.  atmosphere  bicarbonate s o l u t i o n and e x t r a c t e d with methylene c h l o r i d e (3x12 ml) .  The combined organic p o r t i o n was d r i e d  s o l v e n t was removed i n vacuo.  (K CC> ) and the 2  Chromatography  on s i l i c a g e l (ether) a f f o r d e d two products. (R  0.8) was the 3S,4R-epoxide  f  product  3  of the residue The major product  52 (6.5 mg, 52%) while the minor  (R^ 0.6) was the unstable mesylate 53 (4.5 mg, 29%)  which during normal handling was p a r t i a l l y ^Hmr) t o the 3S,4R epoxide 5 2 .  converted  (ca 50% by  The i n s t a b i l i t y of the mesylate  5 3 r e s u l t e d i n i t s incomplete c h a r a c t e r i z a t i o n . 16,18S-Dicarbomethoxy-3S,4R-epoxydihydrocleavamine ( 5 2 ) ; uv>  : 296 (3.67)  227  (4.29); i r D  (3H, m,  C 1 1  284 sh (3. 76),  266 (4.03), 262 (4.03),  : 1730; Hmr <£: 8.2 (IH, m, C..-H), 7.5-7.2 1  - C - H ' s ) , 6.24 (IH, d, J=7.5 Hz, C^-H) , 3.96 1 3  (3H, s, -OCH ) , 3.63 (3H, s, -OCH_) , 0.97 (3H, t , J=7.5 Hz, 3  3  -CH CH ) ; ms: m/e 412 (M ), 383 (base peak), 353, 224, 152, +  2  3  140, 110. High r e s o l u t i o n molecular weight determination c a l c d . f o r C H g N 0 : 412.1998; found: 412.2020. 2 3  2  2  5  Mesylate 53; Hmr 1  (3H, C - C - H * s ) , 1 1  8.18 (IH, m, C ~ H ) , 7.6-7.2 14  6.37 (IH, d, J=8.5 Hz, C ~ H ) , 4.60 (IH, d,  1 3  18  J=2.5 Hz, C -H), 3.97 (3H, s, - O C H 3 ) , 3.65 (3H, s, - O C H 3 ) , 3  3.03  (3H, s, -SOCH ), 0.98 (3H, t , J=7.5 Hz, -CH CH_ ) . 3  2  3  - 135 -  18S-Carbomethoxy-3R-hydroxyvelbanamine A 1.5%  54  s o l u t i o n of sodium methoxide i n methanol  (3 ml)  under a n i t r o g e n atmosphere was t r e a t e d with a s o l u t i o n of 16,18S-dicarbomethoxy-3R-hydroxyvelbanamine mmole) i n anhydrous methanol stirred  (0.5 ml).  (51) (29 mg,  The r e a c t i o n - mixture was  at ambient temperature f o r 4 h.  Acetic acid  was added and the s o l v e n t was removed i n vacuo. taken up i n methylene c h l o r i d e pension was  0.067  (ca 10 drops)  The residue was  (5 ml) and the r e s u l t a n t sus-  f i l t e r e d through a short plug of alumina (Act. I l l ,  methylene c h l o r i d e ) .  Removal of the s o l v e n t i n vacuo a f f o r d e d  18S-carbomethoxy-3R-hydroxyvelbanamine  (54) (25 mg,  a white foam; uv ?\  (3. 79),  : 292  (3.76), 283  100%)  as  277 sh (3.75),  ITtctX  224  (4.39); i r V  : 3605, 3453, 1720; Hmr 1  ITldX  NftH) , 7.60-7.06 (4H, m, C-^-C^-H ' s) , 5.45 C -H) , 3.69  (3H, S, -OCH_) , 3.50  lg  3  158. C  3  N  :  3 7 2  -  2 0 4 9  ?  & 2 Hz,  (M ) , 170 +  (base peak),  found: 372.2049.  18S-Carbomethoxy-3R-hydroxyvelbanamine A s o l u t i o n of (54)  (IH, dd, J=12  High r e s o l u t i o n molecular weight determination c a l c d . f o r  21 28 2°4 H  (IH, bs,  (IH, d, J=2 Hz, C^-H) , 0.99  (3H, t , J=7 Hz, -CH CH ) ; ms'.m/e 372 2  <4: 8.03  (24 mg,  c h l o r o i n d o l e n i n e (55)  18S-carbomethoxy-3R-hydroxyvelbanamine  0.065 mmole) i n benzene  (3 ml) at 0°C under a  n i t r o g e n atmosphere was t r e a t e d with 1 - c h l o r o b e n z o t r i a z o l e (11 mg,  0.072 mmole).  The r e s u l t a n t s o l u t i o n was  stirred for  15 min a t which p o i n t i t was concentrated i n vacuo.  Chroma-  tography of the residue on s i l i c a g e l (ether) a f f o r d e d the  - 136  chloroindolenine  55  (15 mg,  57%)  -  as a yellow f i l m ; uv X : • max (4.19); i r D : 3602, 1730, z  307  (3.28), 288-268 (3.43), 223  rricix  1571; J=12  "''Hmr cf: 7.7-7.1 (4H, m, & 2 Hz,  J=7.5 Hz,  C -H) , 3.65 18  (3H,  -CH CH ) ; ms: m/e 2  peak), 170.  C ^ - C ^ - H ' s ) , 5.79  408,  3  S, -OCH_) , 0.96 3  406  (M ) , 343, +  (IH, (3H, 311,  dd, t,  209  (base  High r e s o l u t i o n molecular weight determination  calcd. for C  2 1  H  2 7  N 0 2  Attempted coupling  3 5 4  C 1 : 406.1660; found: 406.1655.  of 18S-carbomethoxy-3R-hydroxyvelbanamine (54)  with v i n d o l i n e * A s o l u t i o n of 18S-carbomethoxy-3R-hydroxvelbanamine (54)  (17 mg,  0.046 mmole) i n methylene c h l o r i d e  0°C under a n i t r o g e n  atmosphere was  t i o n of t - b u t y l h y p o c h l o r i t e 0.046 mmole).  The  (1.5 ml)  t r e a t e d with a 0.05  s o l v e n t was  M solu-  i n carbon t e t r a c h l o r i d e (0.92  r e a c t i o n mixture was  removed i n vacuo and  with a s o l u t i o n of v i n d o l i n e  (20.8  methanolic hydrogen c h l o r i d e  (3 ml).  and  formation.  the residue was  mg,  ml,  s t i r r e d f o r 10 min  checked by t i c to insure complete c h l o r o i n d o l e n i n e The  at  treated  0.046 mmole) i n 3% The  r e a c t i o n mixture  was  s t i r r e d under a n i t r o g e n atmosphere at ambient temperature f o r 4 days at which p o i n t i t was ammonium hydroxide (3x8 ml).  The  (8 mg,  e x t r a c t e d with methylene c h l o r i d e p o r t i o n was  removed i n vacuo.  on s i l i c a g e l  vindoline  and  combined organic  the solvent was due  (10 ml)  poured i n t o c o l d concentrated  dried  (Na S0 ) 2  4  Chromatography of the  and  resi-  (10% methanol/ethyl acetate) gave back 38%)  and  deacetylvindoline  (10 mg,  Also present were s e v e r a l minor more p o l a r products  *This r e a c t i o n was c a r r i e d ^ o u t by T. Hibino i n t h i s and i s i n c l u d e d here f o r the sake of completeness.  53%). (total  laboratory,  - 137 -  18S-Carbomethoxv-3R,4S-epoxycleavamine  (60)  To a 2% s o l u t i o n of sodium methoxide i n methanol (25 ml) under a n i t r o g e n atmosphere was added 16,18S-dicarbomethoxy-3R,4S-epoxydihydrocleavamine  (40) (500 mg, 0.824 mmole).  The r e a c t i o n mixture was s t i r r e d a t ambient temperature f o r a p e r i o d o f 2.5 h at which p o i n t i t was made n e u t r a l by the addition of acetic acid.  The r e s u l t a n t s o l u t i o n was concen-  t r a t e d i n vacuo and the residue was d i s s o l v e d i n e t h y l acetate . (30 ml) and washed with 5% sodium bicarbonate s o l u t i o n and b r i n e  (1x10 m l ) .  A f t e r drying  (1x10 ml)  ( N a S 0 ) , the s o l v e n t was 2  4  removed i n vacuo to y i e l d a gum which was chromatographed on alumina  (Act. I l l , benzene/ethyl acetate) to give 18S-car-  bomethoxy-3R,4S-epoxydihydrocleavamine a white foam; uv A 224  (4.58); i r V  (60) (146 mg, 50%) as  : 292 (3.99), 283 (4.00), 276 sh (3.94),  max : 3458, 1721; Hmr J : 8.62 (IH, bs, NH), max —  7.56-7.08 (4H, m, C  1  1()  - C - H ' s) , 5.19 (IH, dd, J=10 & 5 Hz, 14  C 1Q  H) , 3.71 (3H, s, OCH ) , 1.10 (3H, t , J=7.5 Hz,. - C H J C H J ) ; ms: 3  m/e 354 (M , base peak), 325, 152, 138. +  High r e s o l u t i o n molecu-  l a r weight determination c a l c d . f o r 2 1 2 6 2 ^ 3 354.1944;. found: 354.1939. A n a l . c a l c d . f o r C-. H_ ,,N-0, • h CH-.CO-C-H,.: C  H  21  N  26 2 3  :  3  2 2 5  C 69.35; H 7.54; N 7.04; found: C 69.13; H 7.45; N 7.15.  18S-Carbomethoxy-3R,4S-epoxydihydrocleavamine  c h l o r o i n d o l e n i n e (61)  A s o l u t i o n of the epoxide 60 (48 mg, 0.136 mmole) i n benzene  (2 ml) a t 0°C under a n i t r o g e n atmosphere was t r e a t e d with  a s o l u t i o n of 1 - c h l o r o b e n z o t r i a z o l e  (24 mg, 0.156 mmole) i n  - 138 -  benzene  (1.5 m l ) .  The r e a c t i o n mixture was s t i r r e d f o r a f u r -  ther 10 min at which p o i n t i t was concentrated i n vacuo and the residue  chromatographed on s i l i c a g e l (benzene/10% e t h y l ace-  tate) to a f f o r d the c h l o r o i n d o l e n i n e uv A  max  61 , mp 149-150°C  : 303 (3.38), 288-272 (3.45), 225 (4.32); i r >  (ether) ; : max  1730; Hmr £: 7.6 (IH, m, C ~H) , 7.4 (3H, m, C ^ - C ^ - H ' s) , 4.48 1  14  (IH, d, J = l l Hz, C -H) , 3.70 (3H, s, -OCH_) , 1.11 (3H, t , ig  3  J=7.5 Hz, -CH CH ); ms: m/e 390, 388 (M , base peak), 353, 152. +  2  3  High r e s o l u t i o n molecular weight determination c a l c d . f o r C  21 25 2°3 H  N  3 5 c l :  3 8 8  -  1 5 5 4  ''  Attempted d i m e r i z a t i o n cleavamine  (60)  found: 388.1567.  o f 18S-carbomethoxy-3R,4S-epoxydihydro-  with v i n d o l i n e i n methanolic hydrogen c h l o r i d e  A s o l u t i o n o f the epoxide 6 0 (90 mg, 0.25 mmole) and t r i e t h y l a m i n e  (0.05 ml) i n methylene c h l o r i d e  0°C under a n i t r o g e n  atmosphere was t r e a t e d with a 0.05 M s o l u -  t i o n of t - b u t y l h y p o c h l o r i t e 0.27 mmole).  (9 ml) a t  i n carbon t e t r a c h l o r i d e (5.4 ml,  The r e a c t i o n mixture was s t i r r e d at 0°C f o r a  f u r t h e r 15 min and checked by t i c to ensure complete formation of the c h l o r o i n d o l e n i n e  61.  The solvent was removed i n vacuo  and the residue was t r e a t e d with a s o l u t i o n of v i n d o l i n e (10) (112 mg, 0.25 mmole) i n 1.8% methanolic hydrogen c h l o r i d e (10 ml).  The r e a c t i o n mixture was s t i r r e d a t ambient temper-  ature under a n i t r o g e n  atmosphere f o r 48 h at which p o i n t i t  was poured i n t o c o l d ammonium hydroxide s o l u t i o n e x t r a c t e d with methylene c h l o r i d e i c p o r t i o n was d r i e d  (3x15 ml).  (30 ml) and  The combined organ-  (Na2S0 ) and the s o l v e n t was removed 4  -  i n vacuo.  The r e s i d u e was  -  139  chromatographed on s i l i c a g e l to  a f f o r d four d i m e r i c products  ( t o t a l ~18  mg).  were a s c e r t a i n e d by t h e i r cd curves to be chemistry  These products  'unnatural' s t e r e o -  'dimers', however due to the low y i e l d s no  c h a r a c t e r i z a t i o n was  further  obtained.  P r e p a r a t i o n of 16,18S-dicarbomethoxy-19-oxo-3R,4S-epoxydihydrocleavamine A.  (62) Using S a r r e t t reagent To f r e s h l y prepared S a r r e t t reagent  (75 mg CrO^ i n  2 ml p y r i d i n e ) a t 0°C under a n i t r o g e n atmosphere was  added a  s o l u t i o n of 16,18S-dicarbomethoxy-3R,4S-epoxydihydrocleavamine 40 (71 mg,  0.172  mixture was  mmole) i n methylene c h l o r i d e  sodium b i s u l f i t e s o l u t i o n  chloride  (10 ml)  and  reaction stirred  taken up i n 10%  and t r e a t e d with a c e t i c a c i d  The r e s u l t a n t mixture was  e x t r a c t e d with methylene  (2x15 ml) and the combined organic p o r t i o n was  (Na S0 ). 2  The  allowed to warm to ambient temperature  f o r a f u r t h e r 17 h at which p o i n t i t was  (0.5 ml).  (1 ml).  dried  Removal of the s o l v e n t i n vacuo followed by chroma-  4  tography of the r e s i d u e on alumina a f f o r d e d the lactam epoxide 62 (methanol); uv X  : 293  (Act. I l l ,  (50 mg,  (3.58), 282  e t h y l acetate)  68%), mp  202-204°C  (3.65), 262  (4.01),  IUclX  227  (4.28); i r v  7.5  (3H, m,  3.55 426 152.0  : 1732 , 1642 ; Hmr  C - C - H s ) , 4.32 ,  1 1  1 3  (3H, S, -OCH ), 1.00 +  394 2  3  C -H) , 3.93  C..-H),  (3H, s, -OCH_) ,  2  2  (base peak);  (-C0 CH ) , 136.0  (IH, m,  (IH, m,  3  (3H, t , J=7.5 Hz, -CH CH ); ms: m/e  3  (M ),  <£: 8.25  1  13  Cmr  (C ), 1 5  £: 135.5  171.9 (C ), 1 ?  3  (-C0 CH ), 167.7 2  129.1  3  (C ), 1 Q  (C ), 1 9  125.2  - 140 -  (C ), 1 2  123.4 ( C ) , 119.5 ( C ) , 118.4 ( C ) , 116.1 ( C ) , 61.4 n  g  13  1 4  (C ) , 59.3 (C ) , 53.6 (-OCH ) , 52.4 (-OCH ), 47.7 (Cg) , 47.1 (C 3  38.6  4  3  3  (C ) , 31.1 ( C ) , 30.6 (C ) , 24.3 (-CH CH ), 22.1 ( C ^ , 2  1 8  9.4 (-CH CH ). 2  3  g  2  3  High r e s o l u t i o n molecular weight determination  c a l c d . f o r C~-,H .N-C>: 426. 1791; found: 426.1785. 2.6 26 2 6 0/  Anal.calcd.  f o r C H gN O .35CH OH: C 63. 80; H 6.37; N 5.88; found: C 64.01; 23  2  2  g  3  H 6.30; N 5.91. B .  Using mercuric acetate A s o l u t i o n o f the epoxide  p-dioxane  40 (25 mg, 0.061 mmole) i n  (2 ml) was t r e a t e d with a s o l u t i o n o f the disodium  s a l t of ethylenediaminetetraacetic acid and mercuric acetate acid  (2 ml).  (65 mg, 0.232 mmole)  (55 mg, 0.173 mmole) i n 1% aqueous  acetic  The r e a c t i o n mixture was s t i r r e d a t ambient temp-  e r a t u r e f o r 19 h, f i l t e r e d , and d i l u t e d with e t h y l acetate (10 ml). acid brine  The o r g a n i c p o r t i o n was washed with 5% h y d r o c h l o r i c  (1x5 ml), 5% sodium bicarbonate s o l u t i o n (1x5 ml).  A f t e r drying  (1x5 ml), and  ( N a S 0 ) , the s o l v e n t was removed 2  4  i n vacuo and the residue was chromatograohed on s i l i c a g e l (benzene/10% e t h y l acetate) t o a f f o r d the lactam epoxide 62 (13 mg, 50%) i d e n t i c a l t o t h a t obtained above.  18S-Carbomethoxy-19-oxo-3R,4S-epoxy-dihydrocleavamine (66) To a 1% s o l u t i o n of sodium methoxide i n methanol (4 ml, 0.741 mmole) a t ambient temperature under a n i t r o g e n atmosphere was added the lactam epoxide 62 (45 mg, 0.106 mmole) The r e s u l t a n t suspension was s t i r r e d f o r 3 h a t which p o i n t  - 141 -  a l l the s o l i d had d i s s o l v e d . with a c e t i c a c i d  The r e a c t i o n mixture was t r e a t e d  (5 drops) and the s o l v e n t was removed i n vacuo.  The residue was t r i t u r a t e d with methylene c h l o r i d e  (2 ml)  and the r e s u l t a n t suspension was f i l t e r e d through c e l i t e .  The  f i l t r a t e was concentrated i n vacuo and chromatographed on s i l i c a gel  (ether) t o a f f o r d 18S-carbomethoxy-19-oxo-3R,4S-epoxy-  dihydrocleavamine 292  (66) (37 mg, 95%) as a white foam; uv A  : max : 3452,  (3.96), 283 (3.99), 276 (3.92), 224 (4.57); i r $ III 3.X  1728,  1648; -"-Hmr i : 8.67 (IH, bs, NH) , 7.56 (IH, m, C^-H) , 7.2  (3H, m, C - C - H ' s ) , 4.33 (IH, doublet of t r i p l e t s , J=3 & 13 1 1  1 3  Hz, C -H), 4.05 (IH, dd, J=10.5 & 1.5 Hz, C^-H) , 3.71 (3H, s, 2  -OCH ) , 1.15 (3H, t , J=7.5 Hz, -CH CH_ ) ; ms: m/e 368 (M ) , 228 +  3  2  3  (base peak), 215, 183, 170, 168, 167. High r e s o l u t i o n molecular weight determination c a l c d . f o r 2 i 2 4 2 ° 4 368.1736. C  H  N  :  368.1736;  f  D u n c  j:  Reaction of 18S-carbomethoxy-19-oxo-3R,4S-epoxydihydrocleavamine (66)  with 1 - c h l o r o b e n z o t r i a z o l e A s o l u t i o n of the lactam epoxide 66 (32 mg, 0.087 mmole)  i n benzene  (2 ml) a t ambient temperature  under a n i t r o g e n a t -  mosphere was t r e a t e d with 1 - c h l o r o b e n z o t r i a z o l e (14.7 mg, 0.096 mmole).  The r e a c t i o n mixture was s t i r r e d  f o r 10 min a t  which £>oint i t was chromatographed on s i l i c a g e l (ether) to a f f o r d two products. was  The major and l e s s p o l a r (Rf 0.8) product  the b e n z o t r i a z o l e d e r i v a t i v e  minor product 27%).  68 (17 mg, 40%), while the  (R 0.7) was the hydroxy d e r i v a t i v e f  Both compounds were obtained as c o l o r l e s s  69 (9 mg, films.  - 142 -  18-Benzotriazole d e r i v a t i v e 68; uv A  : 291 (3.87),  ITtciX  283  (3.94), 277 sh (3.90), 258 (3.86), 221 (4.45); i r  max 3442, 1732 , 1649; """Hmr «T 9.39 (IH, bs, NH) , 8.03 (IH, dd, :  J=8  & 2 Hz, 1 b e n z o t r i a z o l e H) , 7.6-7.0 (6H, m, n " " c  c  4~ ' H  1  s  &  2 b e n z o t r i a z o l e H's), 6.34 (IH, dd, J=8 & 2 Hz), 4.65 (IH, m, C ~H), 3.83 (3H, s, -OCH ), 1.07 (3H, t , J=7.5 Hz, -CH CH ); 2  3  2  ms: m/e 485 ( M , base peak), 426 , 367, 228, 154 , 119 . +  3  High  r e s o l u t i o n molecular weight determination c a l c d . f o r C^H^NgO^: 485.2063; found: 485.2061. 18-hydroxv d e r i v a t i v e 69, uv A  : 292 (3.76),  max (3. 81), 276 sh (3. 78), 223 (4 . 48); ir S)  283  : 3586, 3515, 3467,  IIlclX  3442 , 1734 , 1641; Hmr J: 8.43 (IH, bs, NH) , 7.56 (IH, m, C -H) , 1  14  7.25  (3H, m,  J=13  & 3 Hz, C -H) , 3.94 (3H, s, -OCH_ ), 2.02 (2H, q, J=7.5 Hz,  C 1 1  ^C  ; L 3  - H ' s) , 4.40 (IH, doublet of t r i p l e t s ,  2  3  -CH CH ) ; ms: m/e 384 (M , base peak), 325, 228, 215, 182, 164, +  2  3  115. High r e s o l u t i o n molecular weight determination c a l c d . f o r C 1 2 4 2 ° 4 384.1685; found: 384.1689. H  N  :  2  18'-epi-19'-oxoleurosine 70 A. s o l u t i o n of v i n d o l i n e benzotriazole derivative chloride  (12 mg, 0.026 mmole) and the  68 (12 mg, 0.025 mmole) i n methylene  (2 ml) under a n i t r o g e n atmosphere a t ambient temper-  ature was t r e a t e d with t r i f l u o r o a c e t i c a c i d  (5 drops).  The  r e a c t i o n mixture was s t i r r e d f o r 1 h, d i l u t e d with methylene chloride  (20 ml), washed with 5% sodium bicarbonate s o l u t i o n  (5 ml), and d r i e d  (K C0 ). 2  3  Removal of the s o l v e n t i n vacuo  followed by chromatography o f the residue on s i l i c a g e l ( e t h y l  - 143 -  acetate/10% methylene c h l o r i d e , 2X) a f f o r d e d oxoleurosine uv ^  18'-epi-19' —  (70) (18 mg, 89%) as a white amorphous s o l i d ;  : 309 (3.94), 295 (4.07), 284 (4.04), 261 (4.08), ill 3.X  218  (4.73); i r T)  : 3468, 1740, 1639, 1618; Hmr Si 1  bs, NH) , 7.5-7.0 (4H, m, C  u  , -C  1 4  9.16 (IH,  , -H' s) , 6.87 (IH, s, C ~H) , 14  5.98 (IH, s, C - H ) , 5.93 (IH, dd, J=10.5 & 4 Hz, C -H), 5.33 ?  ?  (IH, s, C.-H), 5.31 (IH, d, J=10.5 Hz, C -E), 4 6  4.71 (IH, m,  r  C ,-H), 3.86 (3H, s, -OCH_) , 3.82 (3H, s, -OCH_) , 3.75 (3H, s, 2  3  3  -OCH ) , 3.69 (Iff, s, C -H) , 2.62 (3H, s, N-CH_ ) , 2.06 (3H, s, 3  2  3  -OCOCH ), 1.02 (3H, t , J=7 Hz, -CH CH_ ) , 0.59 3  2  (3H, t , 0.65,  3  -CH CH ) ; ms: m/e 822 (M , base peak), 762, 663, 662, 661, 555, +  2  135;  3  cd A max : 208 (+12.4), 222 (-41.3).  High r e s o l u t i o n molecu-  l a r weight determination c a l c d f o r C. <-H N .0 : 822. 3829; 46 54 4 10 found: 822.3856.  18' - E p i l e u r o s i n e (73) A s o l u t i o n of v i n d o l i n e and the c h l o r o i n d o l e n i n e chloride  (10) (20 mg, 0.044 mmole)  61 (18 mg, 0.046 mmole) i n methylene  (1 ml) a t ambient  temperature under a n i t r o g e n  phere was t r e a t e d with m-chloroperbenzoic a c i d mmole).  (8 mg, 0.0 46  The r e a c t i o n mixture was s t i r r e d f o r 5 min, t r e a t e d  with a 1:1 mixture  (by volume) of t r i f l u o r o a c e t i c  f l u o r o a c e t i c anhydride min.  atmos-  The r e s u l t a n t  (6 drops) and s t i r r e d  s o l u t i o n was c a u t i o u s l y  s i o n of sodium borohydride (5 ml) at 0°C.  acid/tri-  f o r a f u r t h e r 15 added to a suspen-  (100 mg, 2.63 mmole) i n methanol  This mixture was d i l u t e d with water  (10 ml) and  - 144 -  e x t r a c t e d with methylene c h l o r i d e organic  p o r t i o n was d r i e d  i n vacuo.  (3x8 ml).  (Na S0 ) 2  The combined  and the solvent was  4  removed  Chromatography of the residue on s i l i c a g e l ( e t h y l  acetate/10% methanol) a f f o r d e d  18'-epileurosine  as a white amorphous s o l i d ; uv A  : 307  (15 mg,  (3.77), 290  40%)  (3.97),  IT13.X  283  (3. 98),  262  (4.04), 217  (4. 63);  ir  : 3463 , 1739,  1616;  in 3.x  W  9.10  7.01  (IH, bs, NH) , 6.9-7.5 (4H,m, C  (IH, s , C - H ) , 1 4  4 Hz, C -H), 5.38  6.00  3.84  (3H, s, -OCH_) , 3.77  3.68  (IH, s, C -H) , 2.61  0.97  (3H, t , J=7 Hz, -CH CH ) , 0.61  ms m/e cd  822, 808  max : 207  3  3  2  (IH, dd, J=10 &  (3H, s, -OCH_) , 3.75 (3H, s, NCH_) , 2.06  2  3  g  3  (3H, s, -OCOCH_ ) , 3  (3H, t , J=7 Hz, -CH CH_ ) ; 2  +  (-41.4).  C -H),  (3H, s, -OCH_ ),  (M ), 610, 510, 282, 135, 121, 106  (+21.8), 222  ,-H s) , 1  1 4  (IH, d, J=10 Hz,  4  3  , -C  (IH, s, C^-H) , 5.91  (IH, s, C ~H), 5.30  7  n  3  (base peak);  High r e s o l u t i o n molecular r  weight determination c a l c d . f o r C gH,-gN 0g: 808.4046; found: 4  4  808.4036.  1 8 ' - e p i v i n c a d i o l i n e (74) A s o l u t i o n of v i n d o l i n e and  the c h l o r o i n d o l e n i n e  (10) (18 mg,  55 (24 mg,  c h l o r i d e at 0°C under a nitrogen m-chloroperbenzoic a c i d (9 mg,  0.039 mmole)  0.059 mmole) i n methylene  atmosphere was t r e a t e d with  0.052 mmole).  The r e a c t i o n  mixture was s t i r r e d at 0°C f o r 30 min, t r e a t e d with a 1:1 mixture  (by volume) of t r i f l u o r o a c e t i c a c i d / t r i f l u o r o a c e t i c  anhydride  (8 drops),  and s t i r r e d f o r a f u r t h e r 35 min at 0°C.  The r e s u l t a n t s o l u t i o n was c a u t i o u s l y added to a suspension of  - 145 -  sodium borohydride This s o l u t i o n was  (130 mg,  dried  mmole) i n methanol at 0°C.  d i l u t e d with water  methylene c h l o r i d e was  3.42  (3x8 ml).  (10 ml)  The combined organic p o r t i o n  (Na S0 ) and the s o l v e n t was 2  residue was  and e x t r a c t e d with  removed i n vacuo.  4  The  chromatographed on s i l i c a g e l (76% e t h y l a c e t a t e /  12% methylene c h l o r i d e / 1 2 % methanol) to y i e l d 1 8 ' - e p i v i n c a d i o l i n e (9.5 mg,  19%)  as a white amorphous s o l i d ; uv A  : 30 8  (3.81),  (4.58), 209  (4.66);  in 9.x  292  (3.97), 283  ir V  (3.95), 252  : 3438, 1737,  1616;  (4.10), 222 1  Hmr  (4H, m, C , - C , - H s ) , 6.98  (IH, bs, NH) , 7.5-7.0  (IH, s, C -H),6.01 (1H, s, C-^-H) ,  ,  11  <$: 9.00  sh  14  14  5.88  (IH, dd, J=10.5 & 4 Hz, C -H), 5.34  (IH,  d, J=10.5 Hz, C -H), 3.87  ?  c  (IH, s, C ~H),  (3H, s, -OCH.), 3.77  o  -OCH_) , 3.76 NCH ), 2.09 3  (IH, s, Cj-H), 2.64  (3H, s, -OCOCH^), 0.93  0.66  (3H, t , J=7 Hz,  169,  168,  135,  121,  (3H, s,  — j  (3H, s, -OCHj), 3.68  3  5.30  4  (3H, t , J=7 Hz,  -CH CH ); ms: m/e 2  115,  3  107  826  (3H, .s,  -CH CH ), 2  (M ) , 824, +  3  344,  170,  (base peak); cd A  : 209 (+18.0), max 223 (-42.0). High r e s o l u t i o n molecular weight determination c a l c d . f o r C . ,H N .0,.: 826.4152; found: 826.4129. 46 58 4 10 co  3',4'-Anhydrovinblastine  (26)  3',4'-Anhydrovinblastine  (26) was  prepared using a 31  s l i g h t l y modified procedure  to those p r e v i o u s l y p u b l i s h e d .  A s o l u t i o n of catharanthine i n methylene c h l o r i d e  (30 ml)  (11)  (1.35 mg,  33 '  4 mmole)  a t 0°C under a n i t r o g e n atmosphere  was  t r e a t e d with m-chloroperbenzoic  The  r e a c t i o n mixture was  stirred  acid  (0.73 g, 4.2  a t 0°C f o r 5 min  and  mmole). checked  - 146 -  by t i c to ensure complete N-oxide formation.  V i n d o l i n e (10)  (1.2 g, 2.6 mmole) was added and the r e s u l t a n t s o l u t i o n was cooled to -50°C.  T r i f l u o r o a c e t i c anhydride (2.4 ml, 12 mmole)  was added and the r e a c t i o n mixture was s t i r r e d f o r a f u r t h e r 3.5 h a t -40°C.  The r e s u l t a n t s o l u t i o n was c a u t i o u s l y added to  a suspension of sodium borohydride (4 g, 0.105 mmole) i n methanol  (100 ml) a t 0°C. This s o l u t i o n was d i l u t e d withwater  (200 ml) and e x t r a c t e d with methylene c h l o r i d e combined organic p o r t i o n was d r i e d removed  i n vacuo.  (3x60 ml).  The  (Na S0 ) and the s o l v e n t was 2  4  C r y s t a l l i z a t i o n of the residue from methanol  (ca 25 ml) a f f o r d e d 3 , 4 ' - a n h y d r o v i n b l a s t i n e (26) (1.55 g, 69% 1  based on v i n d o l i n e , 48% based on c a t h a r a n t h i n e ) .  Leurosine ( 3 ) Leurosine blastine  ( 3 ) was prepared from 3',4 -anhydrovin1  (26) v i a the method of T. Hibino  utilizing  t-butyl  hydroperoxide i n t e t r a h y d r o f u r a n c o n t a i n i n g 1% aqueous fluoroacetic acid.  tri-  However, the i s o l a t i o n procedure used was  d i f f e r e n t from t h a t p u b l i s h e d .  Thus, upon completion o f the  r e a c t i o n as i n d i c a t e d by t i c (usually 9 to 22 h ) the r e a c t i o n mixture was t r e a t e d with methanol v i n b l a s t i n e used) and d r i e d  (ca 10 ml/g of 3 ,4 -anhydro-  (K^CO^).  1  1  The s o l v e n t was removed  i n vacuo and the residue was c r y s t a l l i z e d from methanol t o afford leurosine  (3).  - 147  Catharine A.  (76) A s o l u t i o n of l e u r o s i n e (3)  in tetrahydrofuran acid The  (0.2 ml)  was  (2 ml)  (3x5 ml).  The  the s o l v e n t was  (5 ml)  and e x t r a c t e d with methylene c h l o r i d e  removed i n vacuo.  (4 mg)  and  catharine  had mp  213-215°C (acetone) 85  authentic sample*;  trifluoroacetic  d i l u t e d with a s a t u r a t e d s o l u t i o n of  combined organic p o r t i o n was  (lit.  0.037 mmole)  c o n t a i n i n g aqueous 1%  on s i l i c a g e l ( e t h y l acetate/12%  (ethanol)  (30 mg,  s t i r r e d i n the presence of a i r f o r 11 days.  r e a c t i o n mixture was  sodium bicarbonate  The  -  (4 mg,  d r i e d (Na2SO^) and  Chromatography of the  residue  methanol) a f f o r d e d l e u r o s i n e  15%). (lit.  The  s y n t h e t i c catharine  213-215°C); mp  162-166°C  171-175 C); undepressed on admixture with oC = -49°  (C, 0.7,  CHC1 ) ( l i t .  an  -51°).  8 5  3  ''"Hmr, uv, i r , and mass s p e c t r a were superimposable with  those of authentic m a t e r i a l . B.  Oxidation of 3 , 4 - a n h y d r o v i n b l a s t i n e 1  a f f o r d e d c a t h a r i n e i n 34% C.  1  (26)  yield.  A s o l u t i o n of l e u r o s i n e (3)  (45 mg,  0.56  mmole) i n  methylene c h l o r i d e c o n t a i n i n g t - b u t y l hydroperoxide was  as above  s t i r r e d at ambient temperature f o r 24 h.  The  (0.06  solvent  ml) was  removed i n vacuo and the residue chromatographed on s i l i c a g e l (methylene c h l o r i d e / 5 % methanol) to a f f o r d catharine  *The a u t h e n t i c catharine was Indianapolis.  provided by the E. L i l l y  (22 mg,  Co.,  48%).  - 148 -  Comparative leurosine  o x i d a t i o n s of 3',4'-anhydrovinblastine ( 2 6 ) , ( 3 ) , and d e r i v a t i v e s thereof u t i l i z i n g  t-butyl  hydroperoxide Reactions were a l l c a r r i e d out u t i l i z i n g  10 mg  of  substrate d i s s o l v e d i n 0.5 ml t e t r a h y d r o f u r a n c o n t a i n i n g 0.05 ml of t - b u t y l hydroperoxide. f u r t h e r reagents used. t i o n s was  To these s o l u t i o n s were added any  The product composition of the reac-  ascertained v i a t i c .  A u t h e n t i c samples  of  3',4'-anhydrovinblastine ( 2 6 ) , l e u r o s i n e ( 3 ) , c a t h a r i n e ( 7 6 ) , 86 63 pleurosine ( 7 9 ) , and 3 ,4'-anhydrovinblastine-N^,-oxide 1  (78)  were used f o r comparison  used f o r the chromatographic  purposes.  The s o l v e n t systems  a n a l y s i s were e t h y l acetate/20%  methanol and methylene c h l o r i d e / 6 % methanol.  Visualization  achieved by spraying with e e r i e s u l f a t e spray reagent heating a t 10 0°C f o r 1 h.  was  and  The r e l a t i v e amounts of products  formed were estimated from the v i s u a l i z e d chromatograms.  Rx . Conditions Substrate  Time  26  22 h  26  5 days  26  22 h  26  22 h  26  22 h  Products  Additive  0.05 ml 1% t r i fluoroacetic acid II  0.05  ml water  0.05 ml 1% t r i fluoroacetic acid 0.1 ml methanol  76  (%)  26  78 or 79  -  5  -  5  30  5  60  Others  5  95  40  25  5  30  -  30  10  5  10  45  —  50  45  5  —  - 149 -  cont' d Rx. Conditions Substrate  Time  Additive  Products 76  3  26  (%) 78 or 79  -  25  -  -  -  -  -  100  0.05 ml 1% t r i fluoroacetic acid  20  65  15  22 h  0.05 ml 1% t r i fluoroacetic acid 5 mg r a d i c a l i n hibitor*  5  90  5  3  44 h  0.05 ml 1% t r i fluoroacetic acid 10 mg r a d i c a l i n hibitor*  100  79  22 h  0.05 ml 1% t r i fluoroacetic acid  100  26  22 h  0.05 ml 1% t r i fluoroacetic acid 5 mg r a d i c a l i n hibitor*  78  22 h  0.05 ml 1% t r i fluoroacetic acid  26  22 h  0.05 ml 5% t r i fluoroacetic acid  3  22 h  3  22 h  3  II  * 3-t-Butyl-4-hydroxy-5-methylphenyl  sulfide  70  Others  5  -  100  -  100  -  -  - 150 -  Reaction of 3',4'-anhydrovinblastine  (26),with potassium  permanganate A s o l u t i o n of 3 4 ' - a n h y d r o v i n b l a s t i n e 0.316 mmole) i n methylene c h l o r i d e  (26) (250 mg,  (2 ml) and acetone  (5 ml)  was t r e a t e d at 0°C with a s o l u t i o n of potassium permanganate (105 mg, 0.665 mmole) i n acetone  (5 ml).  The r e a c t i o n mixture  was s t i r r e d at 0°C f o r 5 min and the s o l v e n t was removed i n vacuo.  The r e s i d u e was t r i t u r a t e d with methylene c h l o r i d e  and f i l t e r e d through s i l i c a g e l (ethyl acetate/25%  (5 ml)  methanol).  Removal of the s o l v e n t i n vacuo f o l l o w e d by chromatography of the residue on s i l i c a g e l (ethyl acetate/15% methanol) a f f o r d e d the k e t o l 83 (111 mg, 42%) as the major product 19'-Oxo-3',4 -anhydrovinblastine 1  to  an a u t h e n t i c sample  minor product  (R  (R 0.4). f  (81) (25 mg, 9.8%) i d e n t i c a l  ( t i c , ms, ^Hmr) was obtained as the  0.75).  f  K e t o l 8 3 , mp 198-202°C (ethanol); uv 294 1734,  max (4.08), 284 (4.14), 268 (4.19), 212 (4.72); i r 1660, 1612; Hmr  IuclX  : 3475,  : 7.88 (IH, bs, NH), 7.51 (IH, m,  1  C , - H ) , 7.32 (IH, s, NCHO) , 7.14 (3H, m, C 14  (IH,  : 310 (3.80),  1 1  , -C ,-H's) , 6.71 13  s, C - H ) , 6.00 (IH, s , C - H ) , 5.86 (IH, dd, J=10, 4 Hz, 14  1 7  C -H), 5.49 (IH, s, C -H), 5.30 (IH, d, J=10 Hz, C -H), 3.97 7  (IH,  4  g  bs, C ,-H), 3.79 (3H, s, -OCH_) , 3.73 (3H, s, -OCH) , 3  3  3.51 (3H, s, ' -OCH ) , 2.69 (3H, s, -NCH_) , 2.12 (3H, s, -OCOCH_ ), 3  3  3  0.79 (3H, t , J=7 Hz, -CH CH_ ) , 0.70 (3H, t , J=7 Hz, -CH CH_ ); 2  3  ms m/e 840 (M ), 781, 680, 573, 135 (base peak). +  2  High  3  resolu-  t i o n molecular weight determination c a l c d . f o r C . ,H_,N .0,,: ^ 46 56 4 11 840.3945; found: 840.3966.  - 151 -  Reaction of l e u r o s i n e  (3) with potassium  A s o l u t i o n of l e u r o s i n e i n acetone  permanganate  (3) (105 mg,  (1 ml) and methylene c h l o r i d e  (0.5 ml) was  with a s o l u t i o n of potassium permanganate i n acetone  (4 ml).  0.13 mmole)  (40 mg,  treated  0.25 mmole)  The r e a c t i o n mixture was s t i r r e d a t ambient  temperature  f o r 3 min and the s o l v e n t was removed i n vacuo.  The residue  was t r e a t e d as above to a f f o r d the k e t o l  83 (30 mg,  27%) i d e n t i c a l with that obtained above, and 19'-oxoleurosine (91)  (20 mg,  19%) as a c o l o r l e s s  19'-Oxoleurosine (4.00), 284  (4.05), 262  film.  (91), uv  (4.13), 214  : 309sh (3.74), 294 max (4.66); i r  : 3470, 1738, max  1644; •'"Hmr  : 8.06  (IH, bs, NH) , 7.57  (3H, m, C , - C , - H s ) , 6.65 ,  1 1  5.90  1 3  (IH, m, C , - H ) 1 4  (IH, s, C - H ) , 6.19  (IH, d, J-10.5 Hz, C -H), 4.76  3  CH ) , 0.84 3  282, 135  (3H, s, -OCH_ ), 2.76  3  (3H, t , J=7.5 Hz,  3  (3H, t , J=7 Hz, -CH CH_ ) ; ms m/e 2  (base peak);  13  Cmr  (-CH £H ), 8.5  3  : 163.0  (C ,),  8.9  weight  determination c a l c d . f o r 4 6 5 4 4 ° i o  2  3  (-CH CH ). 2  H  N  :  -CH  2  (M ) , 763, +  (Cj,),  High r e s o l u t i o n  3  C  822 61.6  1 9  (C ,), 4  (3H,  3  (3H, s, -OCOCH ), 1.01  5.33  (3H, s,  2  (3H, s, -OCH_ ), 3.63  s, NCH ), 2.12  4  (IH, m, C ,-H), 3.85  6  3  1?  (IH, s, C ~H),  7  -OCH ) , 3.83  (IH, s, C - H ) ,  14  (IH, dd, J=10.5, 3.5 Hz,C ~H), 5.51  7.18  f  59.8  molecular  822.3839;  found:  822.3806. Ketoacetate 84 A s o l u t i o n of the k e t o l 83 (22 mg, pyridine  (2 ml) a t ambient temperature  phere was t r e a t e d with a c e t i c anhydride  0.026 mmole) i n  under a n i t r o g e n (4 drops).  The  atmosreaction  - 152 -  mixture was s t i r r e d f o r 30 h.  Methanol  (1 ml) and toluene  (10 ml) were added and the s o l v e n t was removed i n vacuo. Chromatography o f the residue on s i l i c a g e l (ethyl  acetate/10%  methanol.) a f f o r d e d the ketoacetate 84 (15 mg, 65%) as a white amorphous s o l i d , uv  : 310 (3.81), 294 (4.10), 285 (4.17), n 3.x (4.21), 213 (4.73); i r : 3470, 1738, 1660, 1612;  270  m  Iu3X  •"•Hmr  : 7.90 (IH, bs, NH) , 7.51 (IH, m, C , - H ) , 7.32 (IH, s, 14  NCHO), 7.12 (3H, m,  , -C ,-H's) , 6.64 (IH, s, C^-H) , 6.00 13  (IH, s, C - H ) , 5.86 (IH, dd, J=10, 4 Hz, C -H) , 5.48 (IH, s, 1 ?  ?  C -H), 5.31 (IH, d, J=10 Hz, C -H), 4.80 (IH, bs, C ,-H), 3.79 4  g  3  (3H, s, -OCH ), 3.76 (3H, s, -OCH_) , 3.72 (IH, s, C ~H) , 3.62 3  3  2  (3H, s, -OCH_) , 2.69 (3H, s, NCH_ ), 2.14 (3H, s, -OCOCH_ ) , 2.10 3  3  3  (3H, s, -OCOCH ) , 0.77 (3H, t , J=7 Hz, -CH CH_ ), 0.70 (3H, t , 3  2  3  J=7 Hz, -CH CH ); ms m/e 882 (M ), 822, 762, 720, 613, 555, +  2  354,  3  181, 169, 135, 131, 119 (base peak).  High  resolution  molecular weight determination c a l c d . f o r 4 8 5 8 4 ° 1 2 C  found:  H  N  :  8  8  2  •  4  0  5  0  ;  882.4046.  D i o l 85 A s o l u t i o n o f the k e t o l 95% ethanol 0.16  mmole).  8 3 (50 mg, 0.06 mmole) i n  (3 ml) was t r e a t e d with sodium borohydride  The r e a c t i o n mixture was s t i r r e d at ambient temper-  ature f o r 30 min and t r e a t e d with acetone was  ( 6 mg,  ( 1 ml).  The s o l v e n t  removed i n vacuo and the residue was t r i t u r a t e d with methylene  chloride  (20 ml) and f i l t e r e d through c e l i t e .  Removal of the  s o l v e n t i n vacuo followed by chromatography of the residue on  - 153 -  s i l i c a g e l (methylene c h l o r i d e / 5 % methanol) a f f o r d e d the d i o l 85 (33 mg, 66%) as a c o l o r l e s s f i l m , uv A  : 310 (3.65),  ITlcLX  295  (4.00), 284 (4.06), 267 (4.11), 212 (4.65); i r V : 3585, msx  3472, 1738,1660, 1613; Hmr &: 7.98 (IH, s, NH), 7.53 (IH, m, 1  C , - H ) , 7.45 (IH, s, NCHO), 7.15 (3H, m, 14  6.76  , -C  , -H s) , 1  1 3  (IH, s, C - H ) , 6.12 (IH, s, C-^-H), 5.90 (IH, dd, J=10, 14  4 Hz, C -H), 5.46 (IH, s, C ~H), 5.35 (IH, d, J-10 Hz, C -H), 7  4  g  3.82  (3H, s, -OCH ) , 3.80 (3H, s, -OCH_) , 3.76 (IH, s, C -H) ,  3.55  (3H, s, -OCH ),  0.84  (3H, t , J=7.5 Hz, -CH CH ) , 0.79 (3H, t , J=7 Hz, -CH CH_ ) ;  3  3  3  2  2.71 (3H, s, NCH_ ), 2.10 (3H, s, -OCOCH_ ), 3  2  3  3  2  ms: m/e 842 (M ) , 783, 681, 574, 516, 135 (base peak).  3  High  +  r e s o l u t i o n molecular weight determination c a l c d . f o r C gH^gN 0^ 4  842.4102; found:  4  842.4060.  Acetylation of diol85 A s o l u t i o n o f the d i o l pyridine  8 5 (32 mg, 0.38 mmole) i n  (2 ml) a t ambient temperature under a n i t r o g e n atmos-  phere was t r e a t e d with a c e t i c anhydride  (0.1 ml).  The r e a c t i o n  mixture was s t i r r e d f o r 22 h a t which p o i n t methanol (0.5 ml) and toluene  (10 ml) were added.  The s o l v e n t was removed i n  vacuo and the residue was chromatographed on s i l i c a g e l (methylene c h l o r i d e / 5 % methanol) t o y i e l d the t e t r a a c e t a t e 8 7 (9 mg, 24%, R  f  0.4) and the t r i a c e t a t e 86 (9 mg, 26%, R  compounds were obtained as c o l o r l e s s T e t r a a c e t a t e 8 7, uv A  284  f  0.35).  Both  films.  310 (3.73), 294 (4.06), max (4.11), 267 (4.15), 212 (4.70); i r v : 3468, 1732, 1666, :  IT13.X  1618; Hmr <£: 8.06 (IH, bs, NH) , 7.50 (H, m, C, ,-H), 7.36 1  4  - 154 -  (IH, s, NCHO), 7.14 (3H, m, 6.13  , - C , -H' s) , 6.53 (IH, s, C ~H) , 13  14  ( I H , s, C -H) , 5.89 (IH, dd, J=10, 5 Hz, C -H), 5.52 17  (IH,  ?  s, C -H), 5.30 (IH, d, J=10 Hz, C -H), 4.90 (IH, m, C ,-H), 4  g  4.54  4  (IH, t , J=6 Hz, C ,-H), 4.03 (IH, s, Cj-H), 4.82 (3H, s, 3  -OCH ), 4.77 (3H, s, -OCH_) , 4.59 (3H, s, -OCH ) , 2.85 (3H, s, 3  3  3  NCH ), 2.08 (6H, s, 2X-OCOCH_ ), 1.98 (6H, s, 2X-OCOCH_ ), 0.75 3  3  3  (3H, t , J=7 Hz, -CH-CH ), 0.48 (3H, t , J=7 Hz, -CH.CH-); ms: m/e 968 ( M ) , 765, 659, 600, 135 (base peak).  High  +  resolu-  t i o n molecular weight determination c a l c d . f o r ^52^64^4^14 968.4419; found:  :  968.4395.  T r i a c e t a t e 86, uv A  : 309 (3.70), 294 (4.03),  max 284 (4.08), 269 (4.10), 212 (4.67); i r 0  : 3480, 1732, 1664, IHclX  1617; W  7.95 (IH, bs, NH) , 7.54 (IH, m, C ^ . - H ) , 7.38  (IH, s, NCHO), 7.15 (3H, m, C , - C , -H' s) , 6.54 (IH, s, C ~H) , n  6.17  1 3  14  (IH, s, C -H) , 5.88 (IH, dd, J=10, 4 Hz, C -H) , 5.55. 17  ?  (IH, s, C -H), 5.30 (IH, d, J=10 Hz, C -H), 4.89 (IH, m, C ,-H), 4  4.41  g  4  (IH, t , J=6 Hz, C ,-H), 3.84 (3H, s, -OCH ), 3.82 (3H, s, 3  3  -OCH ) , 3.75 (IH, s, C -H) , 3.58 (3H, s, -OCH_) , 2.73 (3H, s, 3  2  3  NCH ) , 2.12 (3H, s, -OCOCH ) , 2.08 (3H, s, -OCOCH_ ), 1.97 3  3  3  (3H, s, -OCOCH ), 0.76 (6H, t , J=7 Hz, 2X-CH CH_ ); 3  (M ), +  2  867, 765, 659, 600, 135 (base peak).  3  High  ms: m/e 926  resolution  molecular weight determination c a l c d . f o r C H g N 0 ^ : 926.4313; 5 Q  2  4  3  found: 926.4331. Diketone 8 8 A s o l u t i o n of the k e t o l 83 (30 mg, 0.036 mmole) and c u p r i c acetate  (monohydrate) (20 mg, 0.11 mmole) i n methanol  - 155 -  (3 ml) was  heated at r e f l u x f o r 25 min.  removed i n vacuo and the r e s i d u e was chloride  (2 ml)  The  s o l v e n t was  t r i t u r a t e d with methylene  and f i l t e r e d through c e l i t e .  Removal of the  s o l v e n t i n vacuo followed by chromatography of the residue on s i l i c a g e l (methylene tone  88 (16 mg,  293  53%)  (4.03), 277  c h l o r i d e / 5 % methanol) a f f o r d e d the dikeas a p a l e yellow f i l m ; uv ? *  (4.11), 260  ( 4.20), 207  max  : 308  (4.70); i r v  (3. 70), : 3450,  max 1739,  1713,  (IH,  m,  1662,  14  n  4  3  (3H, s, -OCOCH_ ), 1.04 3  (3H, t , J=7  3  (base peak), 570,  , -H' s) , 6.77  2  135.  4 Hz, C -H), ?  Hz, C -H), 3.82 g  (3H, s,  (3H, t , J=7.5 Hz,  Hz, -CH CH_ ); ms: m/e 3  838  7.52  (IH, s,  (3H, s, -OCH.^) , 2.74  3  3  CH_) , 0.79  (IH, d, J=10  (3H, bs, -OCH_) , 3.62  s, NCH ), 2.12  1 3  NCHO),  (IH, dd, J=10,  1 ?  (IH, s, C -H), 5.34  -OCH ) .3.74  678  (3H, m, C , - C  (IH, s, C - H ) , 5.91  14  (2H, bs, NH,  1  C , - H ) , 7.18  C - H ) , 6.10 5.51  1615; Hmr<^: 7.94  (3H, -CH  2  (M ) , 779 , +  High r e s o l u t i o n molecular weight  determination c a l c d . f o r C . ,H_ .N.O., , : 838. 3789; found: 838. 3770. 46 54 4 11 P e r i o d i c a c i d cleavage of the d i o l 8 5 A s o l u t i o n of the d i o l 8 5 (25 mg, tetrahydrof.uran atmosphere was mg,  (2 ml)  0.030 mmole) i n  at ambient temperature  t r e a t e d with a s o l u t i o n of p e r i o d i c a c i d  0.066 mmole) i n t e t r a h y d r o f u r a n (0.5 ml).  mixture was  s t i r r e d f o r 2 min  Chromatography of the  The  (8.5  reaction  and concentrated i n vacuo.  r e s i d u e on s i l i c a g e l (methylene  5% methanol/0.1% ammonium hydroxide) aldehyde  under a n i t r o g e n  a f f o r d e d the  chloride/  N^-formyl-  9 0 (15 mg, 65%) as a c o l o r l e s s f i l m , uv h : 310 ^ max (3.67), 294 (4.00), 284 (4.08), 268 (4.15), 213 (4.66);  - 156 -  i  =  r  7.96  3464, 1732, 1661,  1612; "'•Hmr J : 9.20 (IH, s, -CHO),  (IH, bs, NH), 7.61 (IH, s, NCHO), 7.56 (IH, m, C , - H ) , 7.18 14  (3H, m, C  1 1 1  -C  , - H s ) , 6.64 (IH, s, C - H ) , 6.11 (IH, s, C-^-H) , ,  1 3  14  5.88  (IH, dd, J=10,4 Hz, C -H), 5.49 (IH, s, C ~H), 5.29 (IH, d,  J=10  Hz, Cg-H) , 3.80 (6H, s, 2X-OCH_ ) , 3.62 (3H, s, -OCH_) ,  3.50  (IH, s, C -H) , 2.71 (3H, s, NCH_ ), 2.11 (3H, s, -OCOCH ),  0.74  (3H, t , J=7 Hz, -CH CH_ ); ms'-m/e 782 (M ) , 723, 621 , 514,  ?  4  3  2  3  3  3  +  2  135  (base peak).  calcd. for C  4 3  H  5 0  3  High r e s o l u t i o n molecular weight determination N O 4  1 Q  : 782.3515; found: 782.3484.  P e r i o d i c a c i d cleavage o f the k e t o l 83 A s o l u t i o n of the k e t o l tetrahydrofuran  (2 ml) a t ambient  83 (40 mg, 0.048 mmole) i n temperature under a n i t r o g e n  atmosphere was t r e a t e d with a s o l u t i o n o f p e r i o d i c a c i d 0.117 was  mmole) i n t e t r a h y d r o f u r a n (1 ml).  (15 mg,  The r e a c t i o n mixture  s t i r r e d f o r 4 h and the s o l v e n t was removed i n vacuo.  Chromatography of the residue on s i l i c a g e l (methylene  chloride/  5% methanol/0.1% ammonium hydroxide) a f f o r d e d the N^-formylaldehyde 90 (20 mg, 54%) i d e n t i c a l with that obtained above.  Reaction o f 4'-deoxyleurosidine (93) with potassium permanganate A s o l u t i o n o f 4 - d e o x y l e u r o s i d i n e (93) (260 mg, 1  0.33  mmole) i n acetone  ambient  (5 ml) and methylene  (2 ml)  at  temperature under a n i t r o g e n atmosphere was t r e a t e d  with a s o l u t i o n of potassium permanganate acetone  chloride  (3 ml).  (15 8 mg, 1 mmole) i n  The r e a c t i o n mixture was s t i r r e d f o r 20 min  157 -  and the s o l v e n t was removed i n vacuo. ated with methylene c h l o r i d e  The r e s i d u e was  (10 ml) and f i l t e r e d  tritur-  through  s i l i c a g e l ( e t h y l acetate/25% methylene c h l o r i d e / 1 5 % methanol). Removal of the s o l v e n t i n vacuo followed by chromatography of the r e s i d u e on s i l i c a g e l (ethyl acetate/13% methanol) a f f o r d e d vinamidine (95)  (82) (60 mg,  (30 mg,  11%).  22%) and  The s y n t h e t i c  88 (C=l.l, CHC1 ) ( l i t 3  o -33 ).  19'-oxo-4 -deoxyleurosidine 1  vinamidine 1  The  had  oC= -35°  Hmr and mass s p e c t r a as  w e l l as the t i c p r o p e r t i e s of the s y n t h e t i c m a t e r i a l were i n accord with those e x h i b i t e d by an a u t h e n t i c sample.* 19'-Oxo-4 -deoxyleurosidine  (95), uv X : 311 m3.x  1  294  (4.03), 284  (4.08), 263  1736, 1640, 1615; Hmr  <£: 8.05  1  C , - H ) , 7.17 14  (IH,  (3H, m, C' , - C 11  s, C - H ) , 5.88 1?  C -H), 5.31  , -H' s) , 6.65  (3H, s, NCH_ ) , 2.12 3  -CH CH ), 0.84  (IH, m,  (IH, s, C^-H) , 6.16 (IH, s,  ?  g  3  690, 646  (IH, bs, NH) , 7.58  : 3476,  (IH, dd, J=10.5, 4 Hz, C -H), 5.51  (6H, s, 2X-OCH ), 3.78  3  1 3  (4.72); i r  (IH, d, J=10 Hz, C -H), 4.84  4  2  (4. 13), 212  (IH, m, C ,-H), 2  (IH, s, C ~H) , 3.61 2  (3H, s, -OC0CH_ ),  0.92  3  (3.92),  3.82  (3H, s, -OCH_) , 2.74 3  (3H, t , J=7 Hz,  (3H, t , J=7 Hz, -CH CH_ ); ms:m/e 808 ( M ) , 749, +  2  3  (base peak), 589, 540, 135.  High r e s o l u t i o n molecular  weight determination c a l c d . f o r C. ,H .N ,O : 808.4079; 46 56 4 9 808.4046. r/  /  n  found:  *The a u t h e n t i c sample was provided by the E. L i l l y Co., Indianapolis.  - 158 -  C a t h a r i n i n o l (94) C a t h a r i n i n o l was prepared from the s y n t h e t i c  (vinamidine)  88  catharimne had  v i a the l i t e r a t u r e procedure.  oC^-78  0  (C=0.42, CHC1 ) 3  This material  ( l i t -80°).  19 '-Oxoleurosine(91) A suspension o f l e u r o s i n e (3; (70 mg, 0.087 mmole) i n aqueous 1.8% sodium bicarbonate s o l u t i o n  (3.5 ml) was t r e a t e d  with a s o l u t i o n of i o d i n e (45 mg, 0.35 mmole) i n t e t r a h y d r o furan  (3 ml).  temperature solution ml).  The r e a c t i o n mixture was s t i r r e d at ambient  f o r 10 min, d i l u t e d with 5% sodium bicarbonate  (15 ml), and e x t r a c t e d with methylene c h l o r i d e (3x15  The combined organic p o r t i o n was washed with 10% sodium  b i s u l f i t e solution  (1x15 ml) and b r i n e (1x15 ml).  ( N a S 0 ) , the s o l v e n t was removed 2  4  After drying  i n vacuo and the residue  chromatographed on s i l i c a g e l ( e t h y l acetate/12% methylene c h l o r i d e / 1 2 % methanol) to a f f o r d 19 -oxoleurosine 1  (91) (40 mg,  56%) i d e n t i c a l with the m a t e r i a l prepared p r e v i o u s l y .  19'-Oxoleurosidine (96) Treatment of l e u r o s i d i n e oxoleurosidine  (4) as above a f f o r d e d 1 9 ' —  (96) (62%) as an amorphous s o l i d , uv A : 309 max  (3.74), 294 (4.00), 285 (4.04), 262 (4.11), 214 (4.67); ir ^  : 3477, 1738, 1644, 1615; Hmr i : 8.07 (IH, bs, NE), 1  m a x  7.51  (IH, m, C , - H ) , 7.16 (3H, m, C ^ , - C 14  1 3  ,-H's) , 6.63 (IH,  s, C - H ) , 6.16 (IH, s, C - H ) , 5.88 (IH, dd, J=10, 4 Hz, C -H), 14  17  ?  - 159 -  5.50  (IH, s, C -H), 5.32 (IH, d, J=10 Hz, C -H), 4.73 4  g  (IH,  m, C ,-H), 3.82 (6H, s, 2X-OCH_ ) , 3.60 (3H, s, -OCH_ ), 2.74 2  3  3  (3H, s, NCH_ ), 2.11 (3H, s, -OCOCH_ ), 0.96 (3H, t , J=7.5 Hz, 3  3  -CH CH_ ) , 0.83 (3H, t , J=7 Hz, -CH CH_ ) ; ms: m/e 824 (M ) , 765, +  2  3  2  665, 282, 135 (base peak).  3  High r e s o l u t i o n molecular weight  determination c a l c d . f o r C^H^N^O., : 824. 3996; found: n  824. 3953  19'-Oxovinblastine (97) Treatment of v i n b l a s t i n e oxovinblastine  (1) as above a f f o r d e d 1 9 ' —  (97) (33%) as an amorphous s o l i d , uv ?\ : 311 max  (3.74), 294 (3.99), 285 (4.02), 259 (4.14), 213 (4.69); ir  \l  : 3475, 1740, 1640; Hmr i : 8.06 (IH, bs, NH) , 7.54 1  IUclX  —•  (IH, m, C , - H ) , 7.18 (3H, m,  , - C ,-H' s) , 6.68 (IH, S, C -  14  13  1 4  6.18 (IH, s, C - H ) , 5.91 (IH, dd, J=10.5, 3.5 Hz, C -H), 1 ?  5.54  ?  (IH, s, C -H), 5.34 (IH, d, J=10.5 Hz, C -H), 4.70 4  (IH,  g  m, C ,-H), 3.82 (3H, s, -OCH_) , 3.80 (3H, s, -OCH_) , 3.6 0 2  3  3  (3H, s, -OCH ) , 2.75 (3H, s, NCH_) , 2.12 (3H, s, -OCOCH_ ), 3  3  3  0.87 (6H, b t , J=7.5 Hz, 2X-CH CH_ ) ; ms: m/e 824 (M ) , 765, +  2  665, 135 (base peak).  3  High r e s o l u t i o n molecular weight d e t e r -  mination c a l c d . f o r C..H N.O. . : 824. 3996; found: 46 56 4 10 £  cc  824. 3971.  - 160 -  Reaction of 4'-deoxyleurosidine (93) with iodine/sodium bicarbonate. Treatment  of 4'-deoxyleurosidine (93) (40 mg,  mmole) as above a f f o r d e d as the major product  (28 mg,  0.05 69%)  a compound possessing the f o l l o w i n g c h a r a c t e r i s t i c s , i r ^ 3472, 1735, 1616; W  i:  (IH, m, C , - H ) , 7.16 14  6.13  (IH, s ) , 8.10  (3H, m,  (IH, s, C - H ) , 5.88 1?  (IH, s, C -H), 5.32  8.78  (IH, bs, NH)  , - C , -H' s) , 6.52  (3H, s, -OCH ), 3.64  2.10  (3H, s, -OCOCH ), 0.89  (3H, s, -OCH  14  (3H, s, -OCH ),  ), 2.72  3  (3H, s, NCH ), 3  (3H, t , J=7.5Hz, -CH CH ),  3  2  (3H, t , J=7.5Hz, -CH CH ); ms: m/e 3  647, 448, 282, 135 (base peak).  3  Oxidation of 19'-oxoleurosine  +  High r e s o l u t i o n molecular  (100 mg, 0.122 mmole)  (6 ml) c o n t a i n i n g a c e t i c anhydride  (1.5 ml) at -78°C  under a n i t r o g e n atmosphere was t r e a t e d with Jones 3  found:  (91) with Jones reagent  A s o l u t i o n of 19'-oxoleurosine  (0.4 ml, 1.06 mmole C r 0 ) .  0.79  808 ( M ) , 806, 779, 749,  weight determination c a l c d . f o r C . ,-H_^N .0. : 808.4046; ^ 46 56 4 9 808.4048.  i n acetone  7.53  f  ?  3.81  2  x  (IH, dd, J=4.5,10 Hz, C -H), 5.46 g  3  a  (IH, s, C ~H) ,  13  (IH, d, J=10 Hz, C -H), 3.83  4  : m  reagent  The r e a c t i o n mixture was s t i r r e d a t  -78°C f o r 35 min, t r e a t e d with ammonium hydroxide  solution  - 161 -  (10 ml), and allowed t o warm t o 0 C.  The r e s u l t a n t  solution  was d i l u t e d with 1% sodium b i s u l f i t e s o l u t i o n  (15 ml) and  e x t r a c t e d with methylene c h l o r i d e  The combined  organic p o r t i o n was d r i e d i n vacuo.  (3x15 ml).  (Na S0 ) and the s o l v e n t was removed 2  4  Chromatography of the residue on s i l i c a g e l (methy-  lene c h l o r i d e / 5 % methanol) a f f o r d e d s t a r t i n g m a t e r i a l (15 mg, Rp 0.7), N -desmethyl-19'-oxoleurosine R  f  0.6) and the N ~ f o r m y l A  derivative  (98) (17 mg, 17%, 99 (30 mg, 29%, R 0.5). f  The products were obtained as amorphous N -Desmethyl-19*-oxoleurosine, A  solids. uv ?\  : 314 sh (3.75),  ITlclX  294 (4.07), 286 (4.11), 279 (4.09), 252 (4.10), 224 sh (4.58), : 3476, 1735, 1645, 1620; W  212 (4.70); i r 1  8.02  max (IH, bs, N ,-H), 7.56 (1H-, m, C ^ . - H ) , 7.17 (3H, m, C-^.-C^,a  H's),  6.66 (IH, s, C -H) , 6.30 (IH, s, C^-H) , 5.89 (IH, dd, 14  10, 3.5 Hz, C -H), 5.57 (IH, s, C ~HJ, 5.34 (IH, d, J=10 Hz, C,-H), 4.78 (2H, m, C,,-H, N -H), 4.19 (IH, bs, C „ - H ) , 3.81 (6H, b 2 a — A ?  4  s, 2X-OCH ) , 3.62 (3H, s, -OCH_ ), 2.14 (3H, s, -OCOCH_ ) , 1.02 3  3  3  (3H, t , J=7.5 Hz, -CH CH ) , 0.84 (3H, t , J=7 Hz, -CH CH_ ) ; 2  3  2  3  ms: m/e 808 (M') , 806, 749, 648, 296, 207, 174 (base peak).  High  r e s o l u t i o n molecular weight determination c a l c d . f o r 4 5 5 2 4 ° i o C  808.3593; found:  N  808.3618.  N -Formvl 288  H  d e r i v a t i v e 99, uv A  (4.23), 280 (4.17),. 258  • 296 (4.27),  (4.24), 217 (4.78); i r i  : 3478,  ITlclX  1740, 1678, 1645, 1600; Hmr c£: 8.06 (IH, bs, NH) , 7.54 (IH, 1  m, C , - H ) , 7.21 (3H, m, C , - C , -H s) , 7.16 (IH, s, C ~H) , 1  14  6.93  n  1 3  14  (IH, bs, C - H ) , 5.97 (IH, dd, J=10, 4 Hz, C -H), 5.47 1?  ?  - 162 -  (IH, d, J=10 Hz, Cg-H) , 5.28 (IH, s, C ~H) , 4.74 (2H, m, C ,-H, 4  2  C -H) , 3.95 (3H, s, -OCH_) , 3.78 (3H, s, -OCH > , 3.68 (3H, 2  3  3  s, -OCH_) , 2.10 (3H, s, -OCH_) , 1.02 (3H, t , J=7 Hz, -CH CH_ ) , 3  3  2  3  0.87  (3H, t , J=7 Hz, -CH CH_ ) ; ms: m/e 836 (M ) , 834, 820, 777,  761,  677, 661, 636, 121 (base peak).  +  2  3  High r e s o l u t i o n molecu-  l a r weight determination c a l c d . f o r C.^H N.O,,: 836.3620; ro  4o  found:  OZ  4 JL±  836.3686.  22-Oxoleurosine (37) O x i d a t i o n of l e u r o s i n e oxoleurosine  (3) as above a f f o r d e d 2 2 —  (75%), mp 211-216°C (methanol), uv A  : 296  IuclX  (4.22), 289 sh (4.18), 262 sh (4.15), 253 (4.20), 219 (4.68), : 3470, 1738, 1688; Hmr S: 8.80-8.20 (IH, m, NCHO),  ir ) 8.01  1  (IH, bs, NH) , 7.53 (IH, m, C  C ,-H s)  - H ) , 7.18 (3H, m, C ^ , -  6.88 (2H, bs, C-^-H, C-^-H), 5.94 (IH, dd, J=10,  ,  1 3  1 4 I  /  4 Hz, C -H), 5.44 (IH, d, J=10 Hz, C -H), 5.26 (IH, s, C ~H), ?  g  4  4.66  (IH, m, C -H) , 3.92 (3H, s, -OCH_ ), 3.75 (3H, bs, -OCH_) ,  3.71  (3H, s, -OCH ),  2  3  3  Hz, -CH CH ), 2  3  3  2.09 (3H, s, -OCOCH ), 1.62 (2H, q, J=7.5 3  0.97 (3H, t , J=7.5 Hz, -CH CH_ ), 2  3  0.84 (3H, t ,  J=7 Hz, -CH CH_ ) ; ms: m/e 822 (M ) , 820, 683 , 588, 121 (base +  2  3  peak), 106. High r e s o l u t i o n molecular weight determination c a l c d . f o r C.,H .N.O, : 822.3839; found: 822.3806. 46 54 4 10 r  n  O x i d a t i o n of 22-oxoleurosine 22-Oxoleurosine  (37) with i o d i n e  (37) was o x i d i z e d with i o d i n e i n  t e t r a h y d r o f u r a n as d e s c r i b e d p r e v i o u s l y i n the p r e p a r a t i o n o f 19'-oxoleurosine  (91) from l e u r o s i n e  ( 3 ) . 19',22-Dioxo-  -  leurosine  -  163  (98), i d e n t i c a l with the product r e s u l t i n g  Jones o x i d a t i o n of 19'-oxoleurosine, was  6,7-Dihydroleurosine  6,7-dihydro-3',4'-anhydrovinblastine  0.076 mmole) i n t e t r a h y d r o f u r a n (3 ml) c o n t a i n i n g  1% aqueous t r i f l u o r o a c e t i c a c i d peroxide The  (0.3 ml) was  sodium bicarbonate The  (0.3 ml)  and t - b u t y l  s t i r r e d at ambient temperature  r e a c t i o n mixture was  (3x6 ml) .  (67%).  (101)  A s o l u t i o n of (100) (60 mg,  obtained  from  f o r 9 hr.  d i l u t e d with a s a t u r a t e d s o l u t i o n of  (10 ml)  and e x t r a c t e d with methylene c h l o r i d e  combined o r g a n i c p o r t i o n was  the s o l v e n t was  hydro-  removed i n vacuo.  dried  (Na^O^) and  Chromatography of the  due on s i l i c a g e l ( e t h y l acetate/12% methanol) a f f o r d e d dihydroleurosine  (101) (25 mg,  41%)  as a g l a s s , u v A  resi6,7—  : 310 max  (3.75), 294 i  r  ^L = ~  :  (4.00), 284  3478, 1738,  (4.03), 260  1612;  1  Hmr  (4.12), 213  <£: 8.03  (4.68);  (IH, bs, NH) ,  III 3.X  (IH, m,  7.54  —  C , - H ) , 7.18 14  (3H, m,  C ,-C ,-H's), ±1  6.57  13  (IH, s,  C - H ) , 6.13  (IH, s, C - H ) , 5.64  (IH, s, C ~H), 3.84  -OCH_ ), 3.82  (3H, s, -OCH_ ), 3.73  (IH, s, C ~H) , 3.6 2 (3H, s,  -OCH ), 2.68  (3H, s, NCH_) , 2.12  (3H, s, -OCOCH_ ) , 0.98  1 4  3  3  17  3  3  t, J=7.5 Hz, -CH CH ) , 0.82 2  3  4  (3H, s,  2  (3H, t , J=7  3  Hz,  (3H,  -CH CH_ ) ; ms: m/e 2  3  810 (M ), 808, 671, 453, 106 (base peak). High r e s o l u t i o n molecular weight determination c a l c d . f o r C . ,-H,_N .0„ : 810.4203; ' 46 58 4 9 found: 810.4203. +  n  22-OXO-6,7-dihydroleurosine  (102)  Jones o x i d a t i o n of 6,7-dihydroleurosine  (101) as  - 164 -  described p r e v i o u s l y f o r 19'-oxoleurosine 22-OXO-6,7-dihydroleurosine  (91), a f f o r d e d  (102) (72%), mp 201-205°C  (methanol); uv A  : 296 (4.22), 287 sh (4.18), 252 (4.20),  218 (4.68); i r i>  : 3478, 1738, 1687; Hmr $: 8.00 (IH, bs, 1  NH), 6.82 (IH, s, C - H ) , 5.40 (IH, s, C ~H), 4.67 (IH, bs, 1 7  4  C -H) , 3.92 (3H, s, -OCH_ ), 3.81 (3H, s, -OCH^), 3.66 (3H, s 2  3  -OCH_ ), 2.08 (3H, s, -OCOCH_ ) , 0.99 (6H, t , J=7 Hz, 2X-CH CH 3  3  ms: m/e 824 (M ), 686, 658, 138, 124 (base peak), 106. +  r e s o l u t i o n molecular weight determination c a l c d . f o r C .H . N.O. 46 56 4 10 /l<  1  c  824. 3995; found: 824.3955.  2  High  -165-  BIBLIOGRAPHY  1.  E.S. 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T h e s y n t h e s i s o f l e u r o s i n e and 3 ' - h y d r o x y v i n b l a s t i n e . J.P. Kutney, J . B a l s e v i c h , and G.H. Bokelman, H e t e r o c y c l e s , 4, 1377 (1976). S t u d i e s on t h e s y n t h e s i s o f b i s i n d o l e a l k a l o i d s . V I . Novel l a c t a m d e r i v a t i v e s in the v i n b l a s t i n e area. J.P. Kutney, J . B a l s e v i c h , G.H. Bokelman, T. H i b i n o , T. Honda, I . I t o h , a n d A.H.  R a t c l i f f e , Can. J . Chem., 56, 6 2 ( 1 9 7 8 ) .  T o t a l s y n t h e s i s o f i n d o l e and  dihydroindole a l k a l o i d s . XII. Selective f u n c t i o n a l i z a t i o n o f various b i s indoles.  An e f f i c i e n t s y n t h e s i s o f l e u r o s i n e and r e l a t e d b i s i n d o l e d e r i v a t i v e s .  J.P. K u t n e y , J . B a l s e v i c h , T. Honda, P.-H. L i a o , H.P.M. T h i e l l i e r , and B.R. Worth, H e t e r o c y c l e s , 9_, 201 ( 1 9 7 8 ) . S t u d i e s on t h e s y n t h e s i s o f b i s i n d o l e alkaloids. XII. Derivatives o f vincristine. J.P. Kutney, J . B a l s e v i c h , T. Honda, P.-H. L i a o , H.P.M. T h i e l l i e r , and B.R. Worth, Can. J . Chem., 56, i n p r e s s ( 1 9 7 8 ) . D e r i v a t i v e s o f v i n b l a s t i n e and v i n c r i s t i n e : change o f f u n c t i o n a l i t y i n t h e v i n d o l i n e u n i t . J.P. K u t n e y , J . B a l s e v i c h , and B.R. Worth, H e t e r o c y c l e s , 9, 4 9 3 ( 1 9 7 8 ) . S t u d i e s on t h e s y n t h e s i s o f b i s i n d o l e a l k a l o i d s . X I I I . A s y n t h e s i s o f C a t h a r i n e . J.P. Kutney, J . B a l s e v i c h , and B.R. Worth, H e t e r o c y c l e s , i n p r e s s ( 1 9 7 8 ) . on t h e s y n t h e s i s o f b i s i n d o l e a l k a l o i d s . XV. A s y n t h e s i s o f v i n a m i d i n e (catharinine).  Studies  J.P. Kutney, J . B a l s e v i c h , R. C a r r u t h e r s , A. Markus, M.J. M c G r a t h , R.N. Young, and B.R. Worth, B i o o r g . Chem., i n p r e s s ( 1 9 7 8 ) . The c h e m i s t r y o f t h u j o n e . 1. S y n t h e s i s o f i n s e c t j u v e n i l e hormone a n a l o g s v i a W i t t i g c o u p l i n g .  

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