Total synthesis of cryptophycins via a chemoenzymatic approach

Article abstract of DOI:10.1021/jo960972i

A highly convergent synthesis of cryptophycins in their enantiomerically-pure forms was achieved. Our strategy consists of the synthesis of the two units 3 and 4 and linking them together to form the macrocyclic ring. The upper unit 3 was prepared from 10 in four steps, and the lower unit 4 was prepared from 20 in three steps. Enantioselective biocatalytic methodology was used to prepare the requisite chiral building blocks, (R)-11 and (R)-19. The stereochemical versatility of this synthetic approach is demonstrated by the synthesis of cryptophycin A and the four diastereomers of cryptophycin C.

Full text of DOI:10.1021/jo960972i

J . Org. Chem. 1996, 61, 6893-6900
6893
Tota l Syn th esis of Cr yp top h ycin s via a Ch em oen zym a tic Ap p r oa ch
Grzegorz M. Salamonczyk, Kang Han, Zhi-wei Guo, and Charles J . Sih*
School of Pharmacy, University of Wisconsin, 425 N. Charter Street, Madison, Wisconsin 53706
Received May 28, 1996X
A highly convergent synthesis of cryptophycins in their enantiomerically-pure forms was achieved.
Our strategy consists of the synthesis of the two units 3 and 4 and linking them together to form
the macrocyclic ring. The upper unit 3 was prepared from 10 in four steps, and the lower unit 4
was prepared from 20 in three steps. Enantioselective biocatalytic methodology was used to prepare
the requisite chiral building blocks, (R)-11 and (R)-19. The stereochemical versatility of this
synthetic approach is demonstrated by the synthesis of cryptophycin A and the four diastereomers
of cryptophycin C.
In tr od u ction
The absolute stereochemistry of 1 was deduced from the
structures of cryptophycin C and D, which were obtained
from total synthesis. Since large scale propagation of
Nostoc sp. afforded 1 in very low yields (about 2 mg/L),4
we turned our attention to total synthesis for the
preparation of cryptophycins. Although Moore and co-
workers have recently completed an elegant total syn-
thesis of cryptophycin C and D,7 their synthetic route is
relatively lengthy and is not easily adaptable for the
synthesis of cryptophycins in gram quantities. The
objective of our synthetic explorations is to develop a
more concise synthesis to alleviate the scarcity of cryp-
tophycins and to prepare analogs for examining the
relationship of chemical structure to biological activity.
We herein report the total synthesis of cryptophycin A
and the four diastereomers of cryptophycin C.
A wide array of novel bioactive cyclic peptides and
depsipeptides have been isolated from Cyanobacteria.1
The cryptophycins comprise the largest class of cyano-
bacterial depsipeptides with 25 members to date.2 The
most important member of this family is cryptophycin A
(1), which was first isolated from Nostoc sp. ATCC 53789
by scientists at Merck.3 It was found to be a potent
antifungal agent, especially toward strains of Cryptococ-
cus, which is a frequent opportunistic pathogen in im-
munodeficient persons suffering from AIDS and cancer,
but unfortunately, it was too toxic to be of therapeutic
use. In 1994, Moore’s group in Hawaii described the
isolation of 1 and six additional cytotoxic analogs from a
terrestrial Nostoc sp. GSV 224.4 One of the minor
analogs was cryptophycin-24, which proved to be identi-
cal to arenastatin A (2) from an Okinawan sponge.5
Resu lts a n d Discu ssion
Retrosynthetic analysis of the cryptophycin carbon
skeleton reveals that the macrocyclic ring may be most
conveniently constructed by the assembly of the two
fragments 3 and 4, derived from the disconnection of the
bonds between O(5)-C(1′) and C(1)-N(9′). The upper
fragment 3 consists of a suitably protected derivative of
(2E,7E,5S,6R)-5-hydroxy-6-methyl-8-phenylocta-2,7-di-
enoic acid and the lower fragment 4 consists of three
subunits which are composed of (S)-2-hydroxy-4-meth-
ylvaleric acid, (R)-3-amino-2-methylpropanoic acid, and
3-chloro-O-methyl-D-tyrosine.
Cryptophycin A was discovered to inhibit tubulin
polymerization6 and was found to be effective against a
broad spectrum of solid tumors, most notably, the drug
resistant and multiple drug-resistant ones.4 Preliminary
animal data suggest that cryptophycin A may be useful
against breast, pancreatic, and colon tumors. The gross
chemical structures of cryptophycins were established
using a combination of chemical and spectral techniques.2
X Abstract published in Advance ACS Abstracts, September 15, 1996.
(1) Moore, R. E. J . Ind. Microbiol. 1996, 16, 134-143.
(2) Trimurtulu, G.; Ohtani, I.; Patterson, G. M. L.; Moore, R. E.;
Corbett, T. H.; Valeriote, F. A.; Demchik, L. J . Am. Chem. Soc. 1994,
116, 4729-4737.
(3) Schwartz, R. E.; Hirsch, C. F.; Sesin, D. F.; Flor, J . E.; Chartrain,
M.; Fromtling, R. E.; Harris, G. H.; Salvatore, M. J .; Liesch, J . M.;
Yudin, K. J . Ind. Microbiol. 1990, 5, 113-124.
(4) Golakoti, T.; Ogino, J .; Heltzel, C. E.; Husrebo, T. L.; J ensen, C.
M.; Larsen, L. K.; Patterson, G. M. L.; Moore, R. E.; Moobery, S. L.;
Corbett, T. H.; Valeriote, F. A. J . Am. Chem. Soc. 1995, 117, 12030-
12049.
(5) Kobayashi, M.; Aoki, S.; Ohyabu, N.; Kurosu, M.; Wang, W.;
Kitagawa, I. Tetrahedron Lett. 1994, 35, 7969-7972.
(6) Smith, C. D.; Zhang, X.; Mooberry, S. L.; Patterson, G. M. L.;
Moore, R. E. Cancer Res. 1994, 54, 3779-3784.
In the initial phases of our study, racemic compounds
were used to examine the feasibility and efficiency of the
important carbon-carbon bond-forming reaction for the
construction of (()-3 (Scheme 1).
S0022-3263(96)00972-3 CCC: $12.00 © 1996 American Chemical Society

6894 J . Org. Chem., Vol. 61, No. 20, 1996
Salamonczyk et al.
Sch em e 1a
a
(a) (CH3)3S+CH3OSO3-, CH2Cl2, NaOH, H2O, 50 °C, >95%; (b) MgBr2, ether, -25 °C, >95%; (c) tert-butyl 4-bromocrotonate (9),
Zn-Pb, ether-benzene 1:1, reflux 60%.
Sch em e 2
Ta ble 1. Ad d ition of 9 to (()-7
Our synthetic route begins with the commercially
available trans-4-phenyl-3-buten-2-one (5) as the starting
material. Reaction of 5 with trimethylsulfonium methyl
sulfate under phase transfer catalysis conditions led to
the unsaturated oxirane, 6 in excellent yield.8 The
epoxide 6 was then rearranged to the â,γ-unsaturated
aldehyde (()-7 by its exposure to anhydrous MgBr2
(generated in situ). The yield of this rearrangement was
quantitative and no impurities were detected by means
of NMR spectroscopy. The final and crucial step of the
synthesis was the Reformatsky reaction of the aldehyde
(()-7 with tert-butyl 4-bromocrotonate (9).9 It is well
known that the regioselectivity of addition of organome-
tallic reagents, derived from substituted allyl halides, to
carbonyl compounds is strongly dependent on solvents
and metal catalysts. Hence, the four-carbon homologa-
tion, as depicted in Scheme 2, is expected to generate
regiochemical problems arising from the behavior of the
delocalized reagents of this type. As was succinctly
pointed out by Hudlicky,10 the regioselectivity of these
additions depended heavily on the nature of the catalyst
and the solvent used. As a rule, the γ-mode of addition
increases with softness of the catalyst and with decreas-
ing polarity of the solvent.
catalyst
Zn
Zn-Cu
Zn-Cu
Zn
Zn-Cu
Zn-Cu
Zn
solvent
yield (%)
products γ/Ra
ether
ether
low (<40)
low (<40)
10:90
40:60
benzene
benzene
THF
benzene-ether low (<20)
benzene-ether low (<30)
0
0
0
cleavage of t-butyl ester
cleavage of t-butyl ester
condensation of aldehyde
∼50:50
∼40:60
Mg
Mg
ether
THF
good (∼80) 0:100
0
condensation of aldehyde
Zn-Pb wet ether
high (>80) 30:70
Zn-Pb dry ether
high (>80) 50:50
Zn-Pb dry benzene-ether high (>80) 70:30
a
The ratios of R- and γ-adducts were estimated from the NMR
signals of the allylic methylenes (∼2.4 ppm, 2H; γ-adducts) and/
or the vinylic methylenes (∼5.4 ppm, 2H; R-adducts).
tory. In this case, the rate of C-C coupling was slow
and other competitive processes such as the isomerization
of the double bond in 7, the cleavage of the tert-butyl ester
of 9, and the metal-induced coupling of the halo ester
became dominant reactions. The diastereomeric esters
(()-3 and (()-8 were easily separated by means of silica
gel column chromatography and were later used for the
synthesis of the four diastereomers of cryptophycin C.
Compound (()-3, eluted from the column as the less polar
isomer, was transformed into cryptophycin C2,7a (26) and
30 (Scheme 8), thereby establishing that the stereochem-
istry of (()-3 is anti as shown in Scheme 1. However, as
these compounds (()-3 and (()-8 are highly susceptible
to dehydration on silica gel to give the corresponding
triene derivative (14 in Scheme 4), it was necessary to
include triethylamine in the eluting solvent to avoid this
problem.
Therefore, a series of experiments were conducted with
a view to defining the most suitable conditions favoring
the yield of the desired γ-addition product. Indeed, after
much experimentation we found that the best yield of
the γ-addition products, 3 and 8 (60%), was attained
using “dry” Zn-Pb couple in a gently boiling mixture of
ether-benzene (1:1) (Table 1). However, the use of Zn-
Pb catalyst in neat benzene was found to be unsatisfac-
Having established the applicability of the Refor-
matsky reaction, we then turned our attention to the
synthesis of the upper fragment 3 in its enantiomerically
pure form using a chemoenzymatic approach. We envis-
aged that the racemic ester (()-11, which was prepared
by the alkylation of methyl trans-styrylacetate (10) with
dimethyl sulfate and LDA11 in high yield, could be
conveniently resolved using a lipase-catalyzed enanti-
(7) (a) Barrow, R. A.; Hemscheidt, T.; Liang, J .; Paik, S.; Moore, R.
E.; Tius, M. A. J . Am. Chem. Soc. 1995, 117, 2479-2490. (b) See also
Kobayashi, M.; Kurosu, M.; Wang, W.; Kitagawa, I. Chem. Pharm. Bull.
1994, 42, 2394-2396, for the synthesis of arenastatin A.
(8) (a) Corey, E. J .; Chaykovsky, M. J . Am. Chem. Soc. 1965, 87,
1353-1364. (b) Merz, A.; Ma¨rkl, G. Angew. Chem., Int. Ed. Engl. 1973,
12, 845-846.
(9) Pinza, M.; Pifferi, G. J . Pharm. Sci. 1978, 67, 120-121.
(10) Rice, L. E.; Boston, M. C.; Finklea, H. O.; Suder, B. J .; Frazier,
J . O.; Hudlicky, T. J . Org. Chem. 1984, 49, 1845-1848.

Total Synthesis of Cryptophycins
J . Org. Chem., Vol. 61, No. 20, 1996 6895
Sch em e 3a
a
(a) (CH3O)2SO2, LDA, THF, -60 to 5 °C, 92%; (b) 2-propanol-treated Candida rugosa lipase, 0.2 M phosphate buffer, rt, 48%; (c)
DIBALH, ether, -70 °C, 95%; (d) tert-butyl 4-bromocrotonate, Zn-Pb.
Sch em e 4a
a
(a) 2,4-Dinitrobenzoic acid, Ph3P, DEAD, THF; (b) K2CO3, methanol, (93%).
oselective hydrolysis. The requisite aldehyde (R)-7 for
the Reformatsky reaction could be easily obtained by the
reduction of (R)-11.
Zn-Pb couple as catalyst under the same reaction
conditions as described for the homologation of the
racemate. For no apparent reason, in this case the yield
of the diastereomeric mixture, (+)-3 and (+)-8, was found
to be only 40% in contrast to the 60% that was repeatedly
obtained using (()-7, prepared by an alternate route.
Since the homologation of 9 to (R)-7 was not stereose-
lective and two diastereomers, (+)-3 and (+)-8, were
obtained in almost equal amounts, we decided to trans-
form the syn-isomer 8 into the desired anti-isomer 3 by
means of the Mitsunobu reaction.17 After much experi-
mentation, we found that the best yield of (+)-3 was
obtained with 2,4-dinitrobenzoic acid in the presence of
triphenylphosphine and diethyl azodicarboxylate. Under
these conditions the diester 13 was obtained in 59% yield
accompanied by a slight quantity of the triene 14. When
4-nitrobenzoic or phenoxyacetic acid was used instead,
the predominant product formed was 14. The activated
ester 13 was conveniently hydrolyzed with K2CO3 in
methanol to furnish the desired alcohol, (+)-3.
Since Candida lipases have been widely used for the
kinetic resolution of R-substituted carboxylic esters,12 we
examined three commercially available preparations for
their suitability in catalyzing this transformation. Un-
fortunately, the lipases of Candida cylindracea OF-360
(Meito Sangyo), immobilized Candida antartica (Novo
Nordisk), and Candida rugosa (CRL) (Sigma) all dis-
played poor enantioselectivity toward this substrate with
E13 values ranging from 1.5 to 5.0. However, it is well
known that the enantioselectivity of CRL could often be
enhanced by organic solvent treatment.14 Kazlauskas15
reported the conversion of CRL to a high enantioselec-
tivity form by a simple 2-propanol treatment. Recent
X-ray structures identified two conformational forms of
CRL-open and -closed.16 It was suggested that the
2-propanol treatment increased the activity and enanti-
oselectivity by converting the closed form of CRL to the
open form. Indeed, we found that the 2-propanol-treated
CRL catalyzed the hydrolysis of (()-11 with high degree
of enantioselectivity with an enantiomeric ratio, E of
>100, to yield (S)-12 in 45% yield and (R)-11 in 48% yield.
The enantiomeric purity of both the acid and ester was
estimated to be greater than 96% ee from the analysis
Having completed the synthesis of the upper fragment
3, we then focused our attention on the synthesis of the
lower fragment 4, which is composed of three subunits.
A chemoenzymatic approach was chosen for the synthesis
of the â-amino acid subunit (R)-19. Reaction of methyl
1
of their respective H NMR spectra in the presence of a
chiral shift reagent. In general, CRL has shown a
consistent (S)-stereochemical preference for the hydroly-
sis of arylpropionic ester derivatives.12 Moreover, the
empirical rule, advanced by Kazlauskas,15 for the 2-pro-
panol-treated CRL also predicts that this enzyme pref-
erentially cleaves the (S)-ester. Consequently, the re-
maining ester (R)-11 was reduced with DIBALH at -70
°C to give the aldehyde (R)-7 in 95% yield, which was
stable for about 6 days at -20 °C under argon. The
fourth and last step of the synthesis of 3 entails the
Reformatsky reaction of (R)-7 with the halo ester, 9, using
(11) (a) Rathke, M. W.; Lindert, A. J . Am. Chem. Soc. 1971, 93,
2318-2310. (b) Cregge, R. J .; Herrmann, J . L.; Lee, C. S.; Richman, J .
E.; Schlessinger, R. H. Tetrahedron Lett. 1973, 26, 2425-2428.
(12) Sih, C. J .; Wu, S. H. Top. Stereochem. 1989, 19, 63-125.
(13) Chen, C. S.; Fujimoto, Y.; Girdaukas, G.; Sih, C. J . J . Am. Chem.
Soc. 1982, 104, 7294-7299.
(14) Wu, S. H.; Guo, Z. W.; Sih, C. J . J . Am. Chem. Soc. 1990, 112,
1990-1995.
(15) Colton, I. J .; Ahmed, S. N.; Kazlauskas, R. J . J . Org. Chem.
1995, 60, 212-217.
(16) Grochulski, P.; Li, Y.; Schrag, J . D.; Cygler, M. Protein Sci.
1994, 3, 82-91.
(17) (a) Dodge, J . A.; Nissen, J . S.; Presnell, M. Org. Synth. 1995,
73, 110-115. (b) Hughes, D. L. Org. React. 1992, 42, 335-656.

6896 J . Org. Chem., Vol. 61, No. 20, 1996
Salamonczyk et al.
Sch em e 5a
Reaction of 4 with (()-8 (syn) using the same reaction
conditions furnished the analogs 28 and 29 in similar
yields whereas compounds 26 and 30 were obtained when
(()-3 (anti) was used instead for the condensation
(Scheme 8). To confirm the structural assignments of
28 and 29, we conducted the macrocyclization of (-)-8
(derived from (S)-(+)-11), with the lower fragment 4. The
chromatographic behavior and spectral properties of this
cyclized product were identical to 29, thus validating our
structural assignments of 28 and 29 as shown.
a
(a) Benzylamine, methanol, (90%); (b) H2, Pd/C, AcOH,
methanol; (c) di-tert-butyl dicarbonate, dioxane, H2O, (90% from
16); (d) immobilized Candida antarctica lipase (31%).
Con clu sion
We have developed a highly convergent synthesis of
the cryptophycins in their enantiomerically-pure forms.
Our strategy entails the construction of the two frag-
ments 3 and 4 and joining them together to form a
macrocyclic ring. The upper fragment 3 was synthesized
from 10 in four steps, and the lower fragment 4 was
prepared from 20 in three steps. The requisite chiral
building blocks (R)-11 and (R)-19 were prepared using
enantioselective biocatalytic methodology. The stereo-
chemical flexibility of this synthetic route is illustrated
by the synthesis of cryptophycin A and the four diaster-
eomers of cryptophycin C, in a relatively few steps.
methacrylate, 15, with benzylamine at 70 °C for 4 days
afforded the Michael addition product 16, in 90% yield.
Catalytic hydrogenation followed by amino group protec-
tion furnished (()-18 in 90% yield. A variety of lipases
and proteases were examined for their abilities to cata-
lyze the kinetic resolution of (()-18, but most of them
exhibited low enantioselectivity with E values less than
5. The only exception was the immobilized lipase of
Candida antartica which showed moderate enantioselctiv-
ity with a E value of around 10. Therefore, it was
necessary to deploy a multistep kinetic resolution13,18
procedure to secure (R)-19 of high optical purity. Thus,
(()-18 was first exposed to enantioselective hydrolysis
in aqueous medium catalyzed by the immobilized Can-
dida lipase to yield (R)-19 of moderate optical purity (48%
ee), which in turn was subjected to enzymatic esterifi-
cation with methanol in CCl4 to yield the ester, (R)-(-)-
18, in 89% ee. This enriched ester was again incubated
with the same lipase to furnish the acid (R)-19, with an
optical purity of 97% ee. The overall yield for the three-
step procedure was 31% (Scheme 5).
Exp er im en ta l Section
NMR spectra were obtained in CDCl3 on a Bruker AM-300
instrument operating at 300 MHz for 1H and 75 MHz for 13C.
Coupling constants are reported in hertz (Hz). EI and FAB
mass spectra and high-resolution mass measurements were
performed on a Kratos MS-80RFA DS55/DS90 mass spectrom-
eter, and IR spectra were recorded on a Perkin Elmer 599 B
infrared spectrophotometer. Optical rotations were measured
on a Perkin Elmer 241 polarimeter at 23 °C in chloroform
unless noted otherwise. Thin layer chromatography (TLC) was
performed on Merck precoated silica gel 60 F254 plates. Flash
chromatography was carried out on Baker 40 µm silica gel.
Anhydrous solvents were obtained using standard procedures.
(3E)-1,2-Ep oxy-2-m eth yl-4-p h en ylbu ten e (6). To a solu-
tion of (3E)-4-phenylbuten-2-one (5) (0.1 mol, 14.6 g) in CH2Cl2
(100 mL) were added trimethylsulfonium methyl sulfate (0.12
mol, 22.6 g), 50% aqueous NaOH (50 mL), and benzyltriethyl-
ammonium chloride (500 mg). The reaction mixture was
stirred at 50 °C for 12 h. Ether (200 mL) and water (200 mL)
were added, organic layer was separated, washed with water
(2 × 100 mL), dried (MgSO4), and evaporated under reduced
pressure to give product 6 (15.4 g, 96%) as an orange oil. NMR
The diester 20, prepared following a known procedure,7a
was converted into the trifluoroacetate 21 by treatment
with TFA. The 3-chloro-O-methyl-D-tyrosine was pre-
pared by the monochlorination of N-acetyl-O-methyl-D-
tyrosine with sulfuryl chloride in acetic acid.19 After
protection of the amino group, the t-Boc derivative 22
was obtained in 71% overall yield. The coupling reaction
of 21 with 22 was accomplished in 86% yield in tetrahy-
drofuran with a small excess of DCC, 1-hydroxybenzot-
riazole (HOBT), and diisopropylethylamine (DIEA) at 23
°C. Allyl ester cleavage was effected in THF containing
dry morpholine and a catalytic amount of tetrakis(tri-
phenylphosphine)palladium to furnish 4 in 90% yield7a
(Scheme 6).
The final stages of synthesis entails the assembly of
fragments 3 and 4 (Scheme 7). This was achieved by
using DCC/ DMAP in dichloromethane to give the adduct
24 in 74% yield. After deprotection with TFA, macro-
lactamization of 25 with pentafluorophenyl diphenylphos-
phinate (FDPP) produced cryptophycin C, 26 (71% yield),
whose physical constants were found to be identical to
reported values.7a Epoxidation of 26 with m-CPBA2,7
gave cryptophycin A, 1, and the corresponding R-epoxide
27 in a ratio of 2:1 (78% yield), which were separated by
HPLC (tR of 1 ) 5.65 min; tR of 27 ) 6.61 min). The
analytical data of cryptophycin A, obtained from this total
synthesis, were found to be in full agreement with those
reported for cryptophycin A isolated from natural
source.2,7a
1
δ H: 1.58 (s, 3H), 2.87 (AB, 5.2, 2H), 6.00 (d, 16.2, 1H), 6.65
(d, 16.2, 1H), 7.09-7.21 (m, 5H) ppm; δ 13C: 19.81, 55.84,
56.21, 126.4 (2C), 127.9, 128.7 (2C), 130.7, 131.7, 136.4 ppm.
(3E)-2-Meth yl-4-p h en ylbu ten a l [(()-7]. Magnesium (300
mg) was covered with a solution of 1,2-dibromoethane (17.4
mmol, 1.5 mL) in 10 mL of anhyd ether under an argon
atmosphere. After the reaction ceased, the heterogeneous
mixture was cooled to -25 °C, and the solution of epoxide 6
(0.42 mol, 6.7 g) in ether (20 mL) was added. The reaction
mixture was stirred at that temp for 0.5 h, quenched with 5
mL of 3 M NH4Cl, and diluted with ether (50 mL). The
ethereal layer was washed with 0.5 M citric acid, dried
(Na2SO4), and concentrated to give aldehyde (()-7 (6.5 g, 97%)
as yellow oil. No impurities in (()-7 were detected by NMR.
This compound is stable in the freezer for few days. NMR δ
1H: 1.32 (d, 7.0, 3H), 3.25 (dq, 7.0, 7.5, 1H), 6.15 (dd, 7.5, 16.0,
1H), 6.51 (d, 16.0, 1H), 7.19-7.42 (m, 5H), 9.64 (s, 1H) ppm;
δ
13C: 13.53, 50.25, 125.7, 126.3 (2C), 127.8, 128.6 (2C), 133.4,
137.1, 201.3 ppm.
ter t-Bu tyl 4-Br om ocr oton a te (9). Crude 4-bromocrotonic
acid9 (17.2 g obtained from 0.116 mol of crotonic acid) was
suspended in 25 mL of ether containing 1.8 g of sulfuric acid.
The mixture was cooled to -40 °C, and isobutylene (about 30
mL) was added. The resulting reaction was stirred at 23 °C
(18) Guo, Z. W. J . Org. Chem. 1993, 58, 5748-5752.
(19) Allen, G. R.; Baker, B. R.; Dornbush, A. C.; J oseph, J . P.;
Kissman, H. M.; Weiss, M. J . J . Med. Pharm. Chem. 1960, 2, 391-
412.

Total Synthesis of Cryptophycins
J . Org. Chem., Vol. 61, No. 20, 1996 6897
Sch em e 6a
a
(a) TFA; (b) 1-hydroxybenzotriazole, DCC, THF, DIEA, (86%); (c) Pd(PPh3)4, morpholine (90%).
Sch em e 7a
a
(a) DCC, DMAP, CH2Cl2 (74%); (b) TFA; (c) FDPP, DMF (71% from 24); (d) m-CPBA, CH2Cl2 (78%).
for 3 h and diluted with 50 mL of hexane containing 2 mL of
triethylamine. After washing (2 × 20 mL of 1 M Na2CO3),
drying (MgSO4), and concentration, the crude product was
passed through a short pad of silica gel (hexane-ether 50:1)
to give 16.1 g (overall yield 64%) of 9 as an oil. Rf ) 0.50
silica gel (CH2Cl2-acetone 40:1). The ratio of regioisomers was
1
measured by H NMR.
ter t-Bu tyl (2E,7E)-5-Hyd r oxy-6-m eth yl-8-p h en ylocta -
2,7-d ien oa te [(()-3] a n d [(()-8]. “Dry” zinc-lead couple was
used as a catalyst. The mixture of anhyd ether-benzene 1:1
(5 mL) was used as a solvent, and the above procedure was
followed exactly until the concentration step. The residue was
chromatographed on silica gel using CH2Cl2-hexane-Et3N-
acetone 200:60:3:2 as an eluent. The following compounds
were obtained: R-isomers (75 mg, 25%) not separated (Rf )
0.64-0.66, CH2Cl2-acetone 20:1) colorless oil; (()-3: (80 mg,
27%) colorless oil, Rf ) 0.52 (CH2Cl2-acetone 20:1); HRMS
m/ z (M - 1) calcd (for C19H26O3 - 1): 301.1804, found
301.1810; NMR δ 1H: 1.11 (d, 6.9, 3H), 1.49 (s, 9H), 1.77 (brd,
3.6, 1H), 2.27-2.51 (m, 3H), 3.65 (m, 1H), 5.84 (dt, 15.6, 1.3,
1H), 6.13 (dd, 15.9, 8.6, 1H), 6.48 (d, 15.9, 1H), 6.91 (dt, 15.6,
7.0, 1H), 7.10-7.40 (m, 5H) ppm; δ 13C: 17.51, 28.85 (3C),
37.90, 43.93, 74.58, 80.95, 126.2, 126.9 (2C), 128.1, 129.3 (2C),
131.7, 132.6, 137.7, 144.7, 166.5 ppm; (()-8: (100 mg, 33%)
colorless oil, Rf )0.45; HRMS m/ z calcd (for C19H26O3):
1
(hexane-ether 10:1); NMR δ H: 1.49 (s, 9H), 4.00 (dd, 7.4,
1.0, 2H), 5.95 (dt, 15.3, 1.0, 1H), 6.90 (dt, 15.3, 7.4, 1H) ppm;
δ
13C: 28.61 (3C), 29.89, 81.44, 127.0, 141.1, 165.2 ppm.
P r ep a r a tion of Zin c-Meta l Cou p les. Zinc (500 mg, 20
mesh) was covered with either 3 M copper sulfate or 3 M lead
acetate (3 mL). The mixture was stirred vigorously for 5 min.
The plated couple was then rinsed with methanol and with
solvent used in the reaction (for “wet” conditions it was used
immediately) and dried at ∼120 °C (0.2 mmHg) for 4 h (for
“dry” conditions).
Ad d ition s to (3E)-2-Meth yl-4-p h en ylbu ten a l [(()-7].
The appropriate metal catalyst (∼4 fold excess by weight) was
covered with 4 mL of solvent of choice, under argon, and a
crystal of iodine was added. A mixture of aldehyde (()-7 (1
mmol) and halo ester 9 (1.2 mmol) was dissolved in the same
solvent, and a few drops of this mixture were added to the
stirred suspension of catalyst. The reaction was then heated
to the gentle reflux while the rest of reagents were added (30
min), and reflux was continued for the next 30 min. After
cooling, the reaction was diluted with ether (20 mL), the
catalyst and other precipitates were filtered off, and after
concentration the residue was passed through short pad of
1
302.1837, found 302.1831; NMR δ H: 1.15 (d, 6.8, 3H), 1.48
(s, 9H), 1.82 (brd, 3.9, 1H), 2.29 (ddd, 6.8, 7.2, 8.0, 1H), 2.39-
2.51 (m, 2H), 3.67 (m, 1H), 5.82 (d, 15.6, 1H), 6.13 (dd, 15.9,
8.0, 1H), 6.46 (d, 16.0, 1H), 6.89 (dt, 15.6, 7.2, 1H), 7.12-7.38
(m, 5H) ppm; δ 13C: 16.04, 28.80 (3C), 37.96, 43.72, 74.62,
80.94, 126.3, 126.8 (2C), 128.0, 129.2 (2C), 131.6, 132.6, 137.9,
144.9, 166.4 ppm.

6898 J . Org. Chem., Vol. 61, No. 20, 1996
Salamonczyk et al.
Sch em e 8
(()-Meth yl 2-Meth yl-tr a n s-styr yla ceta te [(()-11]. To a
stirred solution of diisopropylamine (0.9 mL, 6.4 mmol) in 4
mL of anhyd THF under argon was added at -4 °C the solution
of butyllithium (2.7 mL, 2.5 M) in hexanes. After 5 min the
mixture was cooled to -60 °C and a solution of ester 10 (1.06
g, 6 mmol) in 2 mL of ether was added. The reaction mixture
was stirred at that temperature for 10 min, and then dimethyl
sulfate (11 mmol, 1 mL) was added. The temperature of the
reaction was quickly increased to 5 °C, and the resulting
mixture was stirred for 15 min. The reaction was quenched
with 1 M citric acid (10 mL) and diluted with 50 mL of hexane.
The organic layer was washed with water, dried (MgSO4), and
concentrated. The residue was chromatographed over a short
column of silica gel using hexane-ether 40:1 as an eluent to
give (()-11 (1.05 g, 92%) as a colorless liquid. Rf ) 0.35
trated to give (R)-(-)-11 (360 mg, 48%, ee > 96%). [R]D -52.5°
(c ) 5.2); HRMS m/ z calcd (for C12H14O2): 190.0994, found
190.0991; NMR data was identical to that of (()-11.
(2S)-Meth yl 2-Meth yl-tr a n s-styr yla ceta te [(S)-(+)-11].
The acid (S)-(+)-12 (300 mg) was stirred with 5 mL of standard
diazomethane solution in ether at 25 °C for 5 min. Concentra-
tion gave 323 mg of (S)-(+)-11 (100%); [R]D +52° (c ) 2.2); ee
> 96%.
Sta n d a r d P r oced u r e for Deter m in a tion of Op tica l
P u r ity for Com p ou n d s. To a solution of 1.6 µmol of the
appropriate compound in CDCl3 was added 18 mg of tris[3-
[(heptafluoropropyl)hydroxymethylene]-(+)-camphorato]-
1
europium(III). The H NMR spectra (useful spectral param-
eters: O1 ) 3900, SW ) 3800, NS ) 160, LB ) 0, GB ) 0)
were measured both for scalemic (+)-3, (+)-8, (R)-(-)-11, (S)-
(+)-11, and corresponding racemic compounds.
1
(hexane-ether 10:1); NMR δ H: 1.41 (d, 7.4, 3H), 3.36 (dq,
7.5, 1H), 3.72 (s, 3H), 6.32 (dd, 7.6, 15.9, 1H), 6.53 (d, 15.9,
1H), 7.14-7.40 (m, 5H) ppm; δ 13C: 17.45, 43.17, 51.95, 126.4
(2C), 127.6, 128.1 (2C), 128.6, 131.5, 136.9, 174.9 ppm.
(2R,3E)-2-Meth yl-4-p h en ylbu ten a l [(R)-(-)-7]. The so-
lution of ester (R)-(-)-11 (300 mg, 1.57 mmol) in 3 mL of anhyd
ether was cooled to -75 °C, and 1.75 mL of DIBALH (1.1 equiv,
1 M solution in hexane) was added carefully. The reaction
mixture was stirred at -75 °C to -70 °C for 3 h and then
quenched with 2 mL of 1 M NH4Cl. Ether (30 mL) was added
and then the organic phase was washed successively with 1M
citric acid (2 × 10 mL), water (15 mL), dried (Na2SO4), and
concentrated to give (R)-(-)-7 (245 mg, 95%) as a colorless oil.
[R]D -118° (c ) 2.3); NMR data were identical to that of (()-
7.
ter t-Bu tyl (2E,7E,5S,6R)-5-Hyd r oxy-6-m eth yl-8-p h en yl-
octa -2,7-d ien oa te [(+)-3] a n d ter t-Bu tyl (2E,7E,5R,6R)-5-
Hydr oxy-6-m eth yl-8-ph en ylocta-2,7-dien oate [(+)-8]. Zinc
(500 mg, 20 mesh) was covered with 3 M lead acetate (3 mL).
The mixture was stirred vigorously for 5 min. The plated
couple was then rinsed successively with methanol (2 × 4 mL)
and ether (2 × 4 mL) and dried at ∼120 °C (0.2 mmHg) for 4
h. The catalyst was then covered with 5 mL of anhyd ether-
benzene 1:1, under argon, and a crystal of iodine was added.
A mixture of aldehyde (R)-(-)-7 (224 mg, 1.4 mmol) and halo
ester 9 (1.2 equiv, 370 mg, 1.7 mmol) were dissolved in the
same solvent (1 mL), and a few drops of this mixture were
added to the stirred suspension of catalyst. The reaction was
then heated to the gentle reflux while the rest of reagents was
added (30 min), and reflux was continued for the next 30 min.
After cooling, the reaction was diluted with ether (20 mL), the
catalyst and other precipitates were filtered off, and after
concentration of the residue was chromatographed on silica
gel using CH2Cl2-hexane-Et3N-acetone 200:60:3:2 as an
eluent. Compound (+)-3 eluted as less polar (78 mg, 18%, ee
> 96%); [R]D +69° (c ) 2.3); the other analytical data were
2-P r op a n ol Tr ea tm en t of C. r u gosa Lip a se. Crude C.
rugosa lipase (10 g, Sigma) was dissolved in MES buffer (50
mL, 50 mM, pH 6.0, 4 °C, methanesulfonic acid was used to
adjust pH) by stirring at 4 °C for 30 min. 2-propanol (50 mL)
was added dropwise over 1 h at 4 °C. The cloudy solution was
stirred at 4 °C for 46 h. The precipitate was removed by
centrifuging at 2000g for 35 min at 2 °C. The supernatant
was dialyzed against deionized distilled water (3 × 2 L) for 24
h at 4 °C. This lipase solution (120 mL) can be stored at 4 °C
with 0.02% wt/vol sodium azide.
Hyd r olysis of (()-Meth yl 2-Meth yl-tr a n s-styr yla ceta te
[(()-11] w ith 2-P r op a n ol Tr ea ted C. r u gosa Lip a se. To
a solution of 4 mL of CRL in 10 mL of phosphate buffer (0.2M,
pH 7.0) was added 750 mg of ester (()-11. The cloudy mixture
was stirred at 25 °C for 120 h and then the pH of the reaction
was adjusted to 11 by the addition of 2 M Na2CO3. Extraction
with hexane-ether 5:1 (3 × 15 mL), drying (MgSO4), and
concentration of organic layer yielded ester (R)-(-)-11 (373 mg,
ee 93%). The water layer was diluted with 25 mL of 2 M citric
acid, extracted with ether (3 × 15 mL), dried (MgSO4), and
concentrated to give acid (S)-(+)-12 (310 mg, 45%). [R]D +37.6°
(c ) 2.1); mp 80-81 °C; NMR δ 1H: 1.40 (d, 7.1, 3H), 3.34 (dq,
7.0, 7.8, 1H), 6.27 (dd, 7.8, 15.9, 1H), 6.51 (d, 15.9, 1H), 7.16-
7.45 (m, 5H) ppm; δ 13C: 17.24, 40.17, 126.4 (2C), 127.7, 127.9
(2C), 128.6, 131.7, 136.7, 181.3 ppm. The ester (R)-(-)-11 (373
mg, ee 93%) was stirred with 2 mL of CRL solution and 4 mL
of phosphate buffer (0.2 M, pH 7.5) at 37 °C for 16 h. The pH
of this reaction mixture was adjusted to 11, extracted with
hexane-ether 5:1 (3 × 15 mL), dried (MgSO4), and concen-

Total Synthesis of Cryptophycins
J . Org. Chem., Vol. 61, No. 20, 1996 6899
identical to that of (()-3. Compound (+)-8 - eluted as more
polar (91 mg, 22%, ee > 96%); [R]D +50° (c ) 3.1); the other
analytical data were identical to that of (()-8.
In ver sion of ter t-Bu t yl (2E,7E,5R,6R)-5-Hyd r oxy-6-
m eth yl-8-p h en ylocta -2,7-d ien oa te, (+)-8, to ter t-Bu tyl
(2E,7E,5S,6R)-5-H yd r oxy-6-m et h yl-8-p h en yloct a -2,7-d i-
en oa te, (+)-3.
dissolved in 10 mL of acetonitrile) in phosphate buffer (0.2M,
pH 7. 95 mL) was stirred with immobilized Candida antartica
(Novo Nordisk) lipase (1.5 g) at 25 °C for 42 h. The reaction
was acidified to pH 3 with 5% KHSO4, and EtOAc (40 mL)
was added. The enzyme was filtered off, and the two layers
were separated. The aqueous phase was extracted with EtOAc
(2 × 10 mL). The combined organic solutions were dried
(MgSO4) and concentrated, and the residue was passed
through short plug of silica gel (hexane-EtOAc-HOAc 60:40:
1) to give the acid (R)-(-)-19 (5.44 g, 48% ee) and the ester
(S)-(+)-18 (3.83 g, 74% ee). The acid (R)-(-)-19 (5.16 g, 48%
ee) was then esterified with MeOH (10 mL) in the presence of
immobilized C. antarctica lipase (1.29 g) in CCl4 (200 mL) at
50 °C for 29 h. The enzyme was removed, and the filtrate was
concentrated. Separation of the products on silica gel (as
above) gave ester (R)-(-)-18 (3.63 g, 89% ee) and acid (R)-(-
)-19 (1.66 g, 34% ee). The (-)-ester (3.6 g, 89% ee, dissolved
in 3.4 mL of acetonitrile) was hydrolyzed again with im-
mobilized C. antarctica lipase (0.7 g) in phosphate buffer (0.2
M, pH 8, 34 mL) at 25 °C for 48 h. The reaction was worked
up, and the products were separated as described before to
give acid (R)-(-)-19 as colorless crystals (2.863 g, 97% ee). [R]D
-17.9° (c ) 2.3, MeOH), lit.7a [R]D -18.4° (c ) 2.0, MeOH).
The overall yield of (R)-(-)-19 was 31%. Other physical data
are identical with those reported in the literature.7a
(a ) ter t-Bu tyl (2E,7E,5S,6R)-5-(2,4-Din itr oben zoyloxy)-
6-m eth yl-8-p h en ylocta -2,7-d ien oa te (13). To a solution of
alcohol (+)-8 (80 mg, 0.265 mmol), triphenylphosphine (3.9
equiv, 263 mg, 1 mmol), and 2,4-dinitrobenzoic acid (4 equiv,
233 mg, 1.1 mmol) in 3 mL of anhyd THF was added diethyl
azodicarboxylate (3.9 equiv, 160 mL, 1 mmol) at 0 °C under
argon. The reaction mixture was stirred at 40 °C for 50 h and
then was diluted with 30 mL of ether, washed with 0.5 M
solution of NaHCO3 (10 mL), dried (MgSO4), and concentrated.
The residue was purified by silica gel chromatography to give
13 (78 mg, 59%) as a yellow oil. Rf ) 0.19 (hexane-ether 4:1);
[R]D +59° (c ) 3.1); HRMS m/ z calcd (for C26H28N2O8
-
C4H9): 439.1141 found, 439.1120; NMR δ 1H: 1.19 (d, 7.0, 3H),
1.48 (s, 9H), 2.57-2.84 (m, 3H), 5.29 (q, 6.1, 1H), 5.84 (d, 15.6,
1H), 6.06 (dd, 15.9, 8.7, 1H), 6.43 (d, 15.9, 1H), 6.81 (dt, 15.6,
7.5, 1H), 7.17-7.38 (m, 5H), 7.75 (d, 8.4, 1H), 8.42 (dd, 8.4,
2.2, 1H), 8.78 (d, 2.2, 1H) ppm; δ 13C: 17.58, 28.12 (3C), 34.27,
42.96, 79.86, 81.51, 119.6, 126.2 (2C), 126.7, 127.4, 127.8, 128.7
(2C), 129.6, 131.2, 132.1, 133.1, 133.6, 141.3, 148.9 (2C), 163.8,
165.3 ppm.
Allyl (2S,2′R)-2-[(3′-Am in o-2′-m eth ylp r op a n oyl)oxy]-4-
m eth ylp en ta n oa te (21). The solution of allyl (2S,2′R)-2-[[3′-
[(ter t-bu t oxyca r bon yl)a m in o]-2′-m et h ylpr opa n oyl]oxy]-4-
methylpentanoate7a (20) (300 mg, 0.84 mmol) in 3 mL of
CH2Cl2 was treated with TFA (7 mL) at 0 °C then stirred at
rt for 2 h. The resulting mixture was concentrated and dried
in vacuo. The crude product 21 (312 mg, TFA salt) was used
directly in the next reaction.
(b ) ter t-Bu t yl (2E,7E,5S,6R)-5-H yd r oxy-6-m et h yl-8-
p h en ylocta -2,7-d ien oa te, (+)-3. To a solution of diester 13
(70 mg, 0.141 mmol) in 4 mL of methanol was added 100 mg
of K2CO3. The reaction mixture was stirred for 0.5 h, diluted
with ether (30 mL), washed with 10 mL of water, dried
(MgSO4), and then concentrated. The residual oil was passed
through short pad of silica with CH2Cl2-hexane-Et3N-
acetone 200:60:3:3 to give compound (+)-3 (39 mg, 92%)
possessing analytical data ([R]D +70° (c ) 2.2)) as previously
described.
Meth yl 3-(Ben zyla m in o)-2-m eth ylp r op a n oa te (16). A
mixture of 70 mL (0.67 mol) of methyl methacrylate (15),
benzylamine (60 mL, 0.55 mol), and methanol (50 mL) was
stirred at 70 °C for 4 days. After evaporation of the volatiles,
the crude product was purified by flash chromatography
(CH2Cl2-hexane-MeOH, 7:2:1) to give compound 16 (53g,
90%) as a colorless oil. NMR δ 1H: 1.17(d, 6.9, 3H), 1.63 (brs,
NH), 2.6 (m, 2H), 2.88 (dd, 10.1, 3.7, 1H), 3.68 (s, 3H), 3.78 (s,
2H), 7.20-7.35 (m, 5H) ppm; δ 13C: 15.33, 40.12, 51.64, 52.13,
53.71, 126.9, 128.0 (2C), 128.4 (2C), 140.3, 176.3 ppm; EIMS
m/ z (rel intensity): 207 (M+, 4), 120 (84), 106 (71), 91 (100);
HRMS calcd (for C12H17NO2) 207.1259, found: 207.1255.
Meth yl 3-Am in o-2-m eth ylp r op a n oa te (17). To a solu-
tion of ester 16 (2.0 g, 9.66 mmol) and acetic acid (1 mL) in 35
mL of ethanol was added Pd/C (10%, 0.3 g). The resulting
mixture was stirred under hydrogen at 25 °C for 16 h. The
catalyst was filtered off, and the filtrate was concentrated to
give 17 as an acetate (2.3 g). This crude compound was used
directly for next step without purification.
Met h yl 3-(ter t-Bu t oxyca r b on yla m in o)-2-m et h ylp r o-
p a n oa te (18). To a stirred mixture of compound 17 (2.3 g,
12.9 mmol), dioxane (7.5 mL), and H2O (7.5 mL) at 0 °C was
added Et3N (3.2 mL) followed by di-tert-butyl dicarbonate (3.0
g, 13.6 mmol). The reaction was stirred at 0 °C for 1 h and
then at 25 °C for 7 h. The mixture was concentrated, and the
residue was extracted with EtOAc (3 × 20 mL). The combined
EtOAc solution was washed with brine (30 mL), dried (MgSO4),
and concentrated. The crude product was purified by silica
gel column chromatography (hexane-EtOAc 3:2) to give
compound 18 as a colorless oil (1.9 g, 90% overall). NMR δ
1H: 1.17 (d, 7.2, 3H), 1.44 (s, 9H), 2.67 (m, 1H), 3.26 (m, 2H),
3.70 (s, 3H), 4.95 (m, 1H) ppm; δ 13C: 14.73, 28.32 (3C), 39.91,
42.91, 51.84, 79.33, 155.91, 176.2 ppm; EIMS m/ z: 218 (M+
+ 1, 4), 202 (1), 161 (54), 144 (55), 130 (42), 112 (60), 101 (48),
88 (80), 57 (100); HRMS calcd (for C10H19NO4 + H) 218.1392,
found 218.1389.
Com p ou n d 23. To a stirred solution of crude ester 21 (312
mg, 0.84 mmol), N-(tert-butoxycarbonyl)-3-(3-chloro-4-meth-
oxyphenyl)-D-alanine (22) (277 mg, 0.84 mmol), 1-hydroxy-
benzotriazole (129 mg, 0.84 mmol), and diisopropylethylamine
(350 µL) in anhyd THF (10 mL) at 0 °C was added DCC (190
mg, 1.1 equiv, 0.92 mmol). The mixture was stirred at 0 °C
for 30 min and then at 23 °C for 16 h. All the precipitate was
filtered off, and the filtrate was concentrated. The residue was
dissolved in EtOAc (150 mL) and washed with 5% KHSO4 (20
mL) and 5% NaHCO3 (20 mL), dried (MgSO4), and concen-
trated. Silica gel column chromatographic purification (hex-
ane-acetone 7:3) gave ester 23 as a colorless gum (410 mg,
86%). [R]D -34.2° (c ) 2.44); NMR δ 1H: 0.93 (d, 6.3, 3H),
0.96 (d, 6.3, 3H), 1.18 (d, 7.0, 3H), 1.39 (s, 9H), 1.66-1.83 (m,
3H), 2.76 (m, 1H), 2.97 (m, 2H), 3.16 (ddd, 13.9, 9.5, 4.8, 1H),
3.69 (m, 1H), 3.86 (s, 3H), 4.33 (m, 1H), 4.65 (brd, 5.2, 2H),
5.11 (brd, 9.1, 1H), 5.19 (m, NH), 5.28 (d, 10.4, 1H), 5.36 (d,
17.2, 1H), 5.90 (m, 1H), 6.83 (d, 8.4, 1H), 6.86 (m, NH), 7.03
(d, 8.4, 1H), 7.19 (brs, 1H) ppm; δ 13C: 14.72, 21.55, 23.03,
24.83, 28.32 (3C), 37.80, 39.51, 40.40, 41.83, 55.74, 56.11,
66.43, 70.73, 79.70, 112.2, 119.2, 122.1, 128.6, 130.0, 131.0,
131.2, 153.8, 155.1, 171.1 (2C), 173.8 ppm; MS m/ z: 569 (M+
+ 1, 52), 515 (13), 513 (33), 497 (20), 495 (42), 453 (82), 451
(100), 417 (50), 341 (56), 313 (65), 252 (53), 195 (85), 184 (74),
155 (63), 141 (83); HRMS calcd (for C28H4135ClN2O8 + H):
569.2630, found: 569.2640.
Com p ou n d 4. A solution of allyl ester 22 (727 mg, 1.28
mmol), Pd(PPh3)4 (149 mg, 0.1 equiv), and morpholine (1.1 mL,
10 equiv) in anhyd THF (16 mL) was stirred at 23 °C for 1 h.
The solvent was removed, and the residue was dissolved in
EtOAc (150 mL), washed with 5% KHSO4 (30 mL), dried
(MgSO4), and concentrated. Silica gel column chromatographic
purification (hexane-acetone-HOAc 120:80:1) afforded acid
4 as a white solid (608 mg, 90%). [R]D -20.9° (c ) 1.46); NMR
δ 1H: 0.93 (d, 6.1, 3H), 0.97 (d, 6.1, 3H), 1.17 (d, 6.5, 3H), 1.38
(s, 9H), 1.77 (m, 3H), 2.78 (m, 1H), 2.85-3.06 (m, 2H), 3.21
(m, 1H), 3.63 (m, 1H), 3.86 (s, 3H), 4.25 (dd, 14.7, 7.8, 1H),
4.88 (brs, OH), 5.13 (m, 1H), 5.31 (m, NH), 6.83 (brd, 8.3, 1H),
6.91 (m, NH), 7.02 (brd, 8.3, 1H), 7.19 (brs, 1H) ppm; δ 13C
(CDCl3 + trace MeOH): 14.52, 21.55, 23.03, 24.83, 28.20 (3C),
37.84, 39.45, 40.14, 41.83, 55.79, 56.11, 70.93, 80.32, 112.1,
122.1, 128.6, 129.8, 131.0, 153.8, 155.6, 171.8, 173.3, 174.0
ppm.
(2R )-3-(t er t -B u t o x y c a r b o n y la m in o )-2-m e t h y lp r o -
p a n oic Acid [(R)-(-)-19]. A suspension of ester 18 (10 g,

6900 J . Org. Chem., Vol. 61, No. 20, 1996
Salamonczyk et al.
Com p ou n d 24. To a stirred solution of acid 4 (129 mg,
0.244 mmol), alcohol 3 (70 mg, 0.232 mmol) and DMAP (7.1
mg, 0.06 mmol) in CH2Cl2 (3 mL) at 0 °C was added DCC (53
mg, 0.255 mmol). The resulting mixture was stirred at 0 °C
for 1 h and then at 23 °C for 16 h. The precipitate was filtered
off, and the filtrate was concentrated. The residue was
dissolved in EtOAc (50 mL), washed with brine-5% KHSO4
(pH ∼ 3, 10 mL) and 5% NaHCO3 (10 mL), dried (MgSO4),
and concentrated. Chromatography on silica gel (hexane-
EtOAc 3:2) gave the pure product 24 as a white solid (140 mg,
74%). [R]D -12.9° (c ) 4.3); NMR δ 1H: 0.79 (d, 6.4, 3H), 0.84
(d, 6.4, 3H), 1.13 (d, 6.9, 3H), 1.16 (d, 7.3, 3H), 1.35 (s, 9H),
1.48 (s, 9H), 1.51-1.71 (m, 3H), 2.38-2.71 (m, 4H), 2.87 (m,
1H), 3.16 (m, 1H), 3.23 (dd, 14.0, 5.1, 1H), 3.71 (m, 1H), 3.84
(s, 3H), 4.38 (m, 1H), 4.98 (dd, 9.7, 3.7, 1H), 5.06 (m, 1H), 5.74
(m, NH), 5.83 (d, 15.6, 1H), 6.01 (dd, 15.8, 8.7, 1H), 6.42 (d,
15.8, 1H), 6.84 [2H: (d, 8.4, 1H), (m, 1H)], 7.02 (m, NH), 7.11
(dd, 8.4, 2.1 1H), 7.17-7.36 (m, 6H) ppm; δ 13C: 14.60, 17.04,
21.33, 22.91, 24.74, 28.23 (3C), 28.33 (3C), 35.13, 37.65, 39.63,
40.92, 41.12, 42.24, 56.14 (2C), 70.74, 76.61, 79.50, 80.53,
112.1, 122.1, 126.2 (2C), 126.4, 127.5, 128.6 (3C), 130.0, 130.8,
131.2, 131.8, 136.9, 141.7, 153.7, 155.6, 165.8, 171.0, 171.6,
173.6 ppm; FAB-MS m/ z: 813 (M+ + 1), 713, 657, 412, 228;
FAB-HRMS calcd (for C44H6135ClN2O8 + H): 813.4093, found:
813.3093.
overall yield starting from (()-8] by means of chromatography
on silica gel with CH2Cl2-acetone 4:1 afforded 28 (less polar,
Rf ) 0.24, 23 mg) and 29 (more polar, Rf ) 0.18, 27 mg) as
white solids. (5R)-cryptophycin C (28): [R]D +59.9° (c ) 1.8);
NMR δ 1H: 0.90 (brd, 4.8, 3H), 0.94 (brd, 4.8, 3H), 1.14 (d,
6.7, 3H), 1.24 (d, 7.4, 3H), 1.69 (m, 2H), 2.39 (m, 1H), 2.70 (m,
3H), 3.06 (dd, 14.4, 7.4, 1H), 3.15 (dd, 14.4, 5.2, 1H), 3.41 (m,
1H), 3.56 (m, 1H), 3.86 (s, 3H), 4.83 (m, 2H), 5.15 (m, 1H),
5.75 (d, 15.7, 1H), 5.92 (brd, 8.0, 1H), 6.00 (dd, 15.9, 8.6, 1H),
6.47 (m, 1H), 6.51 (d, 15.9, 1H), 6.86 (d, 8.4, 1H), 6.95 (m, NH),
7.08 (dd, 8.4, 2.0, 1H), 7.21 (d, 2.0, 1H), 7.21-7.36 (m, 5H)
ppm; δ 13C: 14.42, 17.31, 21.31, 23.14, 24.72, 34.94, 35.81,
39.10 (2C), 40.32, 40.56, 53.90, 56.24, 71.02, 78.63, 112.4,
122.4, 126.2 (2C), 126.6, 127.7, 128.3, 128.7 (2C), 129.6, 129.7,
131.0, 132.2, 136.7, 139.0, 154.0, 165.8, 170.8 (2C), 174.4 ppm;
MS m/ z: 640 (M+ with 37Cl, 4), 638 (M+ with 35Cl, 6), 414 (16),
412 (45), 368 (26), 280 (6), 227 (50), 195 (26), 91 (65); HRMS
calcd (for C
35H4335ClN2O7): 638.2759, found: 638.2759. (6S)-
cryptophycin C (29): [R]D +2.6° (c ) 2.3); NMR δ 1H: 0.86 (d,
6.2, 3H), 0.88 (d, 6.2, 3H), 1.15 (d, 6.8, 3H), 1.23 (d, 7.2, 1H),
1.46 (m, 1H), 1.72 (m, 2H), 2.40 (m, 1H), 2.56 (m, 2H), 2.71
(m, 1H), 3.03 (dd, 14.5, 7.3, 1H), 3.14 (dd, 14.5, 5.6, 1H), 3.30
(m, 1H), 3.49 (brddd, 13.5, 4.3, 4.3, 1H), 3.86 (s, 3H), 4.83 (m,
1H), 4.88 (dd, 9.6, 3.8, 1H), 5.05 (brdd, 9.2, 6.3, 1H), 5.77 (m,
2H), 6.06 (dd, 15.9, 7.6, 1H), 6.43 (d, 15.9 1H), 6.69 (ddd, 15.3,
9.8, 5.4, 1H), 6.83 (d, 8.4 1H), 6.97 (brdd, 5.5, 5.5, 1H), 7.08
(dd, 8.4, 2.0, 1H), 7.22 (d, 2.0, 1H), 7.22-7.35 (m, 5H) ppm; δ
13C: 14.13, 15.75, 21.50, 22.91, 24.62, 35.13, 36.54, 38.32,
39.72, 41.10, 41.56, 53.73, 56.14, 71.51, 77.34, 112.2, 122.3,
125.0, 126.2 (2C), 127.6, 128.4, 128.6 (2C), 129.9, 130.5, 131.0,
131.4, 136.8, 141.7, 153.9, 165.5, 170.8, 171.1, 175.6 ppm; MS
m/ z: 640 (M+ with 37Cl, 6), 638 (M+ with 35Cl, 15), 414 (34),
412 (91), 368 (28), 280 (16), 227 (100), 195 (47), 91 (56); HRMS
calcd (for C35H4335ClN2O7): 638.2759, found: 638.2743.
Com p ou n d 25. To a solution of compound 24 (57 mg, 0.07
mmol) in 1 mL of CH2Cl2 at 0 °C was slowly added TFA (4
mL), and resulting solution was stirred at 23 °C for 2 h. The
reaction mixture was concentrated and dried in vacuo. The
crude product (56 mg) was used directly in the next reaction.
Cr yp top h ycin C (26). To a solution of compound 25 (56
mg, 0.07 mmol) in anhyd DMF (9.5 mL) at rt were added FDPP
(41 mg, 1.5 equiv, 0.1 mmol) and DIEA (37 µL). The reaction
mixture was stirred under argon at 23 °C for 16 h, diluted
with 20 mL of EtOAc, and then washed successively with 5
mL portions of 5% KHSO4, 5% NaHCO3, and water. The
organic layer was dried (MgSO4) and concentrated. The
residue was subjected to chromatography on silica gel (CH2Cl2-
acetone 4:1) to give the desired product as a white solid (32
mg, 71%). [R]D +36.4° (c ) 2.1), {lit.7a [R]D +28.8° (c ) 0.65)}.
Other analytical data were in full agreement with literature.7a
Cr yp top h ycin A (1) a n d (7S,8S)-Cr yp top h ycin A (27).
A solution of cryptophycin C (26) (50 mg, 78.3 µmol) in 10 mL
of CH2Cl2 was stirred at 23 °C for 45 h, while m-CPBA (52
mg, 300 µmol) was added portionwise. The mixture was then
diluted with 50 mL of EtOAc, washed successively with 10 mL
portions of 5% NaHCO3 and water, dried (MgSO4), and
concentrated to give colorless residue, which was passed
through a short pad of silica gel with hexane-acetone 3:2 to
give 1 and 27 as a mixture (40.7 mg, 78%). Epoxides 1 and
27 were separated by HPLC (Waters 4 µm C18 cartridge,
internal diameter 8 mm, eluent (isocratic) MeOH-H2O 65:35,
flow 3 mL/min) to give cryptophycin A (1) (26 mg): tR)5.65
min; [R]D +32.5° (c ) 2.0, MeOH), {lit.2 [R]D +33.8° (c ) 1.8,
MeOH)]}, with NMR and MS data in agreement with lit.,2 and
(7S,8S)-cryptophycin A (27) (13 mg). 27: tR ) 6.61 min; [R]D
+23.6° (c ) 1.1, MeOH); NMR δ 1H: 0.89 (d, 6.4, 3H), 0.91 (d,
6.4, 3H), 1.05 (d, 7.0, 3H), 1.25 (d, 7.2, 3H), 1.51 (m, 1H), 1.74
(m, 3H), 2.50-2.81 (m, 3H), 2.91 (brd, 7.4, 1H), 3.04 (dd, 14.4,
7.2, 1H), 3.16 (dd, 14.4, 5.4, 1H), 3.32 (m, 1H), 3.51 (brddd,
13.5, 4.5, 4.5, 1H), 3.60 (brs, 1H), 3.88 (s, 3H), 4.82 (brdd, 13.7,
7.2, 1H), 4.92 (m, 1H), 5.14 (brdd, 10.0, 4.0, 1H), 5.73-5.87
(m, 2H), 6.71 (ddd, 15.3, 9.6, 5.3, 1H), 6.85 (d, 8.4, 1H), 7.00
(m, NH), 7.09 (brd, 8.4, 1H), 7.24 (m, 3H), 7.34 (m, 3H) ppm;
(5R,6S)-Cr yp top h ycin C (30). Compound 30 was pre-
pared starting from alcohol (()-3 (60 mg, 0.2 mmol) and acid
4 (111 mg, 0.21 mmol). The synthetic procedures for com-
pounds 24-26 were followed. Separation of the mixture of
products 26 and 30 [52% overall yield starting from (()-3] by
means of column chromatography on silica gel with CH2Cl2-
acetone 4:1 afforded 26 (less polar, Rf ) 0.27, 35 mg) and 30
(more polar, Rf ) 0.19, 31 mg) as white solids (compound 26
possessed exactly the same analytical data as given above).
(5R, 6S)-cryptophycin C (30): [R]D +11.3° (c ) 2.2); NMR δ
1H: 0.81 (brs, 6H), 1.16 (d, 6.7, 3H), 1.24 (d, 8.4, 3H), 1.59 (m,
3H), 2.42 (m, 1H), 2.68 (m, 3H), 3.06 (dd, 14.4, 7.4, 1H), 3.15
(dd, 14.4, 5.1, 1H), 3.40 (m, 1H), 3.54 (m, 1H), 3.87 (s, 3H),
4.86 (m, 2H), 5.08 (m, 1H), 5.78 (d, 15.9 1H), 5.97 (m, NH),
6.01 (dd, 15.9, 8.3, 1H), 6.43 (d, 15.9, 1Hz), 6.46 (m, 1H), 6.86
(d, 8.5, 1H), 6.93 (m, NH), 7.08 (brd, 8.5, 1H), 7.22 (d, 2.0, 1H),
7.23-7.36 (m, 5H) ppm; δ 13C: 14.41, 16.62, 21.20, 22.92,
24.65, 34.57, 35.81, 39.10, 39.20, 40.33 (2C), 54.05, 56.23,
71.02, 78.24, 112.4, 122.5, 126.2 (2C), 126.6, 127.6, 128.3, 128.6
(2C), 129.6, 130.0, 130.9, 131.9, 136.8, 139.0, 154.2, 165.9,
170.7, 170.8, 174.4 ppm; MS m/ z: 640 (M+ with 37Cl, 10), 638
(M+ with 35Cl, 21), 414 (33), 412 (100), 368 (26), 280 (13), 227
(64), 195 (31), 91 (53); HRMS calcd (for C35H4335ClN2O7):
638.2759, found: 638.2772.
Ack n ow led gm en t . This investigation was sup-
ported by National Institutes of Health grant GM331449.
δ
13C: 13.50, 14.11, 21.41, 23.14, 24.70, 35.15, 36.73, 38.33,
39.32, 41.06, 41.21, 53.73, 56.23, 56.43, 63.21, 71.50, 77.34,
112.2, 122.5, 125.2, 125.4 (2C), 128.3, 128.4, 128.6 (2C), 129.8,
131.0, 137.1, 141.4, 154.0, 165.5, 170.8, 171.0, 175.7 ppm; MS
m/ z: 656 (M+ with 37Cl, 10), 654 (M+ with 35Cl, 27), 412 (18),
Su p p or tin g In for m a tion Ava ila ble: NMR (1H, 13C, 1H-
1H COSY) spectra of compounds 1, (+)-3, 4, (-)-7, (+)-8, 23,
24, 26-30; MS and IR spectra of compounds 1, 26-30 (39
pages). This material is contained in libraries on microfiche,
immediately follows this article in the microfilm version of the
journal, and can be ordered from the ACS; see any current
masthead page for ordering information.
280 (18), 227 (47), 195 (58), 91 (100); HRMS calcd (for C35H43
-
35ClN2O8): 654.2708, found: 654.2715.
(5R)-Cr yp t op h ycin C (28) a n d (6S)-Cr yp t op h ycin C
(29). Compounds 28 and 29 were prepared starting from
alcohol (()-8 (49 mg, 0.162 mmol) and acid 4 (90 mg, 0.168
mmol). The synthetic procedures for compounds 24-26 were
followed. Separation of the final products 28 and 29 [49%
J O960972I