Enantioselective biotransformations of racemic and meso pyrrolidine-2,5-dicarboxamides and their application in organic synthesis

Article abstract of DOI:10.1021/jo300412j

In this paper, we report the amidase-catalyzed hydrolysis of pyrrolidine-2,5-dicarboxamides and their application in organic synthesis. Catalyzed by Rhodococcus erythropolis AJ270, an amidase containing microbial whole cell catalyst, racemic trans-pyrrolidine-2,5-carboxamide was kinetically resolved into (2S,5S)-pyrrolidine-2,5-dicarboxamide and (2R,5R)-5- carbamoylpyrrolidine-2-carboxylic acid in high yields and excellent enantioselectivity. Biocatalytic desymmetrization of meso cis- pyrrolidinedicarboxamide afforded enantiomerically pure (2R,5S)-5- carbamoylpyrrolidine-2-carboxylic acid in an almost quantitative yield. In both kinetic resolution and desymmetrization, the amidase always exhibited excellent 2R-enantioselectivity, although its catalytic efficiency was influenced dramatically by the steric effect of the substituent on the nitrogen atom of pyrrolidine ring. The synthetic potential of biotransformation was demonstrated by the scalable preparation of (2R,5R)- and (2R,5S)-5-carbamoylpyrrolidine-2- carboxylic acids and their conversions to aza-nucleoside analogues and druglike pyrroline-fused diazepin-11(5H)-one compounds.

Full text of DOI:10.1021/jo300412j

Article
pubs.acs.org/joc
Enantioselective Biotransformations of Racemic and Meso
Pyrrolidine-2,5-dicarboxamides and Their Application in Organic
Synthesis
Peng Chen,† Ming Gao,† De-Xian Wang,† Liang Zhao,‡ and Mei-Xiang Wang*,‡
†Beijing National Laboratory for Molecular Sciences, CAS Key Laboratory of Molecular Recognition and Function, Institute of
Chemistry, Chinese Academy of Sciences, Beijing 100190, China
‡MOST Key Laboratory of Bioorganic Phosphorus Chemistry & Chemical Biology, Department of Chemistry, Tsinghua University,
Beijing 100084, China
S
* Supporting Information
ABSTRACT: In this paper, we report the amidase-catalyzed hydrolysis of pyrrolidine-
2,5-dicarboxamides and their application in organic synthesis. Catalyzed by
Rhodococcus erythropolis AJ270, an amidase containing microbial whole cell catalyst,
racemic trans-pyrrolidine-2,5-carboxamide was kinetically resolved into (2S,5S)-
pyrrolidine-2,5-dicarboxamide and (2R,5R)-5-carbamoylpyrrolidine-2-carboxylic acid
in high yields and excellent enantioselectivity. Biocatalytic desymmetrization of meso
cis-pyrrolidinedicarboxamide afforded enantiomerically pure (2R,5S)-5-carbamoylpyr-
rolidine-2-carboxylic acid in an almost quantitative yield. In both kinetic resolution and
desymmetrization, the amidase always exhibited excellent 2R-enantioselectivity,
although its catalytic efficiency was influenced dramatically by the steric effect of the
substituent on the nitrogen atom of pyrrolidine ring. The synthetic potential of
biotransformation was demonstrated by the scalable preparation of (2R,5R)- and (2R,5S)-5-carbamoylpyrrolidine-2-carboxylic
acids and their conversions to aza-nucleoside analogues and druglike pyrroline-fused diazepin-11(5H)-one compounds.
intermediates, remain largely unexplored and challenging.
Synthesis of (2S,5S)-2,5-pyrrolidinedicarboxylic acid, for
instance, comprises mainly multistep chemical transformations
starting from chiral materials such as S-pyroglutamate11 and S-
proline derivatives,12 meso-dimethyl 2,5-dibromoadipate,13
chiral epoxide,14 and amino acid.15 They suffer from tedious
chemical manipulations, using expensive chemical reagents and
providing low overall yields. In 1987, Achiwa16 reported
biocatalytic kinetic resolution of racemic dimethyl trans-
pyrrolidine-2,5-dicarboxylates using pig liver esterase (PLE).
The reaction gave unfortunately very low conversion and
enantioselectivity. PLE-catalyzed desymmetrization of dimethyl
cis-1-benzylpyrrolidine-2,5-carboxylate was reported.17
Although high enantioselectivity was achieved under controlled
conditions, the chemical conversion was appallingly low.
Recently, PLE-catalyzed desymmtrization of dimethyl cis-1-
(tert-butoxycarbonyl)pyrrolidine-2,5-carboxylate has been re-
ported to take place in 36 h, giving (2S,5R)-1-(tert-
butoxycarbonyl)-5-methoxycarbonyl)pyrrolidine-2-carboxylic
acid in 75% yield and >99% ee.18
INTRODUCTION
■
Chiral 2,5-disubstituted pyrrolidine derivatives are a very
important type of compound in organic chemistry. First, they
constitute the core structure of natural products1 and synthetic
compounds of pharmaceutical interest.2 Gerrardine, for
example, is a pyrrolidine alkaloid isolated from both Cassipourea
gerrardii3a and Cassipourea guianensis,3b while (2R-trans)-2-
butyl-5-heptylpyrrolidine, a potent σ receptor ligand, was found
in the culture broth of Streptomyces longispororuber.4 (2S,5S)-
Pyrrolidine-2,5-dicarboxylic acid, an interesting C2-symmetric
N-heterocyclic compound, is a marine natural product first
isolated from red alga Schizmenia dubyi in 1975.5 Second, both
trans- and cis-2,5-functionalized pyrrolidines serve as unique
building blocks in the synthesis of natural products and
bioactive compounds such as pyrrolizidines and indolizidines.6
In addition, enantiopure 2,5-substitued pyrrolidines are widely
used as chrial auxiliaries7 and ligands8 in asymmetric synthesis.
Moreover, chiral pyrrolidine2,5-dicarboxylic acid has been used
as an organocatalyst in enamine catalysis albeit the chiral
induction needs further improvement.9
Amidases [3.5.1.4] are a type of hydrolytic enzyme catalyzing
the conversion of primary amides into carboxylic acids with the
release of ammonia.19 To date, a large number of amidases and
microorganisms that contain amidases have been reported, and
Because of their importance in organic and medicinal
chemistry, chiral 2,5-disubstituted pyrrolidine derivatives have
attracted continuous attention from synthetic chemists.
Although various synthetic methods have been developed,10
general approaches to enantiomerically pure pyrrolidine
stereoisomers bearing two functional groups at the 2,5-
positions, which are versatile and invaluable synthetic
Received: February 27, 2012
Published: March 30, 2012
© 2012 American Chemical Society
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some of them have been used extensively in the kinetic
resolution of racemic amides.20 Rhodococcus erythropolis
AJ270,21 a nitrile hydratase and amidase-containing microbial
whole cell catalyst, for instance, has been shown to catalyze
enantioselective biotransformation of a large number of
structurally varied racemic amides including amino,22 hydroxyl
and alkoxy,23 alkenyl,24 alkynyl,25 allenyl,25 and azido26 bearing
amides, cyclopropanecarboxamide,27 and various heterocyclic
amides28−31 to produce highly enantiopure carboxylic acid and
amide products. Surprisingly, biocatalytic enantioselective of
dicarboxamides, including racemic, prochiral,32 and meso
ones,33 remains largely unexplored. On the basis of easy
availability of racemic and meso dicarboxamide compounds,
high catalytic efficiency, and enantioselectivity of the amidase in
Rhodococcus erythropolis AJ270, we envisioned that the amidase-
catalyzed enantioselective hydrolysis of racemic and meso
pyrrolidine-2,5-dicarboxamides33 would provide a unique
method for the synthesis of enantioenriched 2,5-functionalized
pyrrolidine stereoisomers. It is worth emphasizing that, being
different from kinetic resolution and desymmetrization of a
diester which afford only a monoester of diacid, amidase-
catalyzed reactions of diamides produce carbamoyl-substituted
acids that are more versatile and useful in organic synthesis. We
report herein the efficient and practical Rhodococcus erythropolis
AJ270 whole cell-catalyzed biotransformation of both racemic
and meso pyrrolidine-2,5-dicarboxamides.33 The synthetic
potential of the resulting enantiopure 5-carbamoylpyrrolidine-
2-carboxylic acid is also demonstrated by the synthesis of
druglike compounds.
substituent on the heterocyclic ring. For example, N-benzyl-
substituted trans-pyrrolidine-2,5-dicarboxamide substrate 1a
underwent a slower biotransformation, with 50% of the
substrate being converted within 39 h (entry 1, Table 1).
Doubling the biocatalyst loading led noticeably to the
shortening of the reaction period (entry 2, Table 1). In both
cases, enantiopure (2S,5S)-1-benzylpyrrolidine-2,5-dicarboxa-
mide 2a and (2R,5R)-methyl 1-benzyl-5-carbamoylpyrroli-
dine-2-carboxylate 3a were produced in excellent yields. In
addition to the isolation of monocarboxylate product 3a, a
small amount of enantiopure (2R,5R)-dimethyl 1-benzylpyrro-
lidine-2,5-dicarboxylate 4a was also obtained, indicating the
enzymatic activity of the amidase toward initially formed
(2R,5R)-1-benzyl-5-carbamoylpyrrolidine-2-carboxylic acid (en-
tries 1 and 2, Table 1), albeit in low catalytic efficiency.
Replacement of the benzyl substituent with an allyl group on
the pyrrolidine ring resulted in the enhancement of reaction
rate. A 7 h incubation of substrate 1b with microbial cells under
identical conditions thus gave rise to (2S,5S)-1-allylpyrrolidine-
2,5-dicarboxamide 2b, (2R,5R)-methyl 1-allyl-5-carbamoylpyr-
rolidine-2-carboxylate 3b, and (2R,5R)-dimethyl 1-allylpyrroli-
dine-2,5-dicarboxylate 4b in yields of 45%, 36%, and 10%,
respectively (entry 3, Table 1). Single enantiomers 3b and 4b
were obtained, whereas the enantiomeric excess value of 2b
decreased from >99.5% to 43.1% when the conversion of 1b
was low in a shorter reaction time (entry 4, Table 1), a
reflection of a typical kinetic resolution process. Biocatalytic
kinetic resolution of 1c, a substrate devoid of any N-substituent,
proceeded very rapidly. After interaction with biocatalyst in
10−15 min, enantiopure products 2a and 3c were obtained in
high yields. In such a short reaction period, no diacid product
4c was observed (entries 5 and 6, Table 1). It should be noted
that use of DMSO as a cosolvent improved the solubility of
substrates in buffer and therefore facilitated the biotransforma-
tion. The enantioselectivity of enzymatic reaction was, however,
not affected at all (entries 5 and 6, Table 1). Biocatalytic kinetic
resolution of 1c was also readily scalable. This was exemplified
by a 10 mmol scale reaction which produced enantiopure
(2S,5S)-pyrrolidine-2,5-dicarboxamide 2d in 47% yield and
enantioenriched (2R,5R)-5-carbamoylpyrrolidine-2-carboxylic
acid 3d in 52% with 87.1% ee. Enantiomeric purity of 3d was
improved to >99.5% ee after recrysttalization in a mixture of
water and methanol (entry 7, Table 1).
RESULTS AND DISCUSSION
■
We initiated our study by examining the biocatalytic kinetic
resolution of racemic trans-pyrrolidine-2,5-dicarboxamides 1a
and 1b. To facilitate the isolation of products, acids and diacids
were converted into their methyl esters 3 and 4, respectively,
using CH2N2. In the case of the reaction of 1c, the substrate
contains no substituent on pyrrolidine nitrogen (R = H), and
the acid product was transformed into its corresponding benzyl
ester using benzyl bromide as an alkylation reagent. It should
be pointed out that, under the basic reaction conditions,
benzylation also occurred on pyrrolidine nitrogen to afford the
final products 2a and 3c (R = R′ = Bn) (Scheme 1). As
summarized in Table 1, under very mild conditions such as in
neutral aqueous potassium phosphate buffer at 30 °C,
Rhodococcus erythropolis AJ270 whole cell catalyst was able to
catalyze the hydrolysis of the testing racemic trans-pyrrolidine-
2,5-dicarboxamides in a highly enantioselective manner. The
reaction velocity, however, was governed dramatically by the
All resulting dicarboxamides 2 and 5-carbomylpyrrolidine-2-
carboxylates 3 were easily hydrolyzed chemically under acidic
conditions, allowing the preparation of both antipodes of
pyrrolidine-2,5-dicarboxylic acids. Demonstrated in Scheme 2,
for instance, is the chemical transformation of (2S,5S)-2a and
(2R,5R)-3c into (2S,5S)-4a and (2R,5R)-4a, respectively.
Products were isolated in high yields albeit partial racemization
was observed in both cases.
Scheme 1. Biocatalytic Kinetic Resolution of Racemic trans-
Pyrrolidine-2,5-dicarboxamides
In order to synthesize other diastereomers of 5-carbamoyl-
pyrrolidine-2-carboxylic acid derivatives and to explore the
enzymatic activity of the amidase in Rhodococcus erythropolis
AJ270 in catalyzing desymmetrization reactions, hydrolysis of
cis-pyrrolidine-2,5-dicarboxamides was studied (Scheme 3). In
comparison to racemic trans-1-benzylpyrrolidine-2,5-dicarbox-
amide 1a, the meso substrate cis-1-benzylpyrrolidine-2,5-
dicarboxamide 5a underwent a very sluggish biocatalytic
hydrolysis. It took 4 days to produce (2R,5S)-methyl 1-
benzyl-5-carbamoylpyrrolidine-2-carboxylate 6a in 15% yield,
with a large amount of starting material (75%) being intact
(entry 1, Table 2). A slow reaction rate was also observed in the
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a
Table 1. Enantioselective Biotransformations of Racemic Dicarboxamides 1
b
c
b
c
b
c
entry
1
R
time
39 h
2 (yield, %) (ee, %)
3 (R′) (yield, %) (ee %)
4 (R′) (yield, %) (ee, %)
1
2
3
4
5
1a
1a
1b
1b
1c
1c
1c
Bn
Bn
allyl
allyl
H
2a (49) (>99.5)
2a (49) (>99.5)
2b (45) (>99.5)
2b (61) (43.1)
2a (45) (>99.5)
2a (45) (>99.5)
2d (47) (>99.5)
3a (Me) (39) (>99.5)
3a (Me) (32) (>99.5)
3b (Me) (36) (96.8)
3b (Me) (27) (>99.5)
3c (Bn) (36) (93.5)
3c (Bn) (44) (93.0)
4a (Me) (6) (>99.5)
4a (Me) (12) (>99.5)
4b (Me) (10) (93.3)
4b (Me) (8) (>99.5)
d
15 h
7 h
3.5 h
e
10 min
15 min
1 h
4c (N.O.)
f
e
6
g
H
4c (N.O.)
h
e
7
H
3d (H) (52) (87.1) (>99.5%)
4d (N.O.)
a
Substrate (1 mmol) was incubated with Rhodococcus erythropolis AJ270 cells (2 g wet weight) in potassium phosphate buffer (0.1 M, pH 7.0, 50
b
c
d
mL) at 30 °C. DMSO (2.5 mL) was used as a cosolvent. Isolated yield. Determined by chiral HPLC analysis. Rhodococcus erythropolis AJ270 cells
(4 g wet weight) were used. N.O. = not observed. 1c (3 mmol) was used in the absence of DMSO. 1c (10 mmol) was used. After
e
f
g
h
recrystallization.
Scheme 2. Chemical Hydrolysis of 2a and 3c
5c was effected within 1 h, and (2R,5S)-benzyl 1-benzyl-5-
carbamoylpyrrolidine-2-carboxylate 6c was isolated in 96% with
94.8% ee (entry 3, Table 2). In the absence of DMSO as a
cosolvent, enantiopure 6c (>99.5% ee) was produced in 90%
yield (entry 4, Table 2). The amidase in Rhodococcus
erythropolis AJ270 was robust, and it was able to tolerate high
concentrations of substrate 5c and product 6d. In a scaled-up
biocatalytic reaction, for example, 20 g of meso dicarboxamide
5c was transformed almost quantitatively into a single
enantiomeric product (2R,5S)-5-carbamoylpyrrolidine-2-car-
boxylic acid 6d (R = R′ H) within 12 h33 (entry 5, Table 2).
The outcomes of biocatalytic hydrolysis of racemic trans- and
meso cis-pyrrolidine-2,5-dicarboxamide compounds showed
clearly that the amidase within Rhodococcus erythropolis AJ270
displayed higher enzymatic activity against trans-configured
dicarboxamides 1 than the cis-isomers 5. This is most likely
attributable to the steric effect of the substrates, as two cis-
positioned amido groups in 5 may mutually prevent the
interaction from reaching the active site of the amidase, whereas
trans-orientation of two amido groups in 1 does not pose steric
encumbrance to the substrate−enzyme interaction. The
sensitivity of the amidase toward the steric feature of the
substrates is in agreement to previous observations.20a−c It has
been hypothesized that the amidase in Rhodococcus erythropolis
AJ270 may comprise a deep-buried and size-limited active
site.27d−f
The structure of all products was established on the basis of
spectroscopic data and microanalysis. The absolute config-
uration of biocatalytic kinetic resolution products was assigned
on the basis of the comparison of the optical rotation of
(2R,5R)-dimethyl 1-benzylpyrrolidine-2,5-dicarboxylate 4a with
that of authentic sample reported in literature.16 Since 4a was
derived from 3a, compound 3a therefore has the same
stereochemistry as 4a. The absolute configuration of bio-
catalytic desymmetrization products 6 was determined by X-ray
diffraction analysis of the single-crystal structure of the salt of
(2R,5S)-5-carbamoylpyrrolidine-2-carboxylic acid 6c with HCl,
which was cultivated conveniently from crystallization by means
of diffusion of diethyl ether into a mixture of 6c in aqueous HCl
and ethanol.33 It is interesting to address that the amidase
within Rhodococcus erythropolis AJ270 shows 2R-enentioselec-
tivity against both trans- and cis-pyrrolidine-2,5-dicarboxamides.
Functionalized enantiopure pyrrolidine derivatives produced
from biotransformation of trans- and cis-pyrrolidine-2,5-
dicarboxamides are conceivably invaluable chiral building
blocks for the synthesis of natural products and druglike
compounds. Starting from (2R,5S)-5-carbamoylpyrrolidine-2-
carboxylic acid 6d, we33 have previously synthesized a pair of
Scheme 3. Biocatalytic Desymmetrization of Meso
Dicarboxamides 5
Table 2. Biocatalytic Desymmetrization of Meso
a
Dicarboxamides 5
b
c
entry
5
R
time (h)
6 (R′) (yield, %) (ee %)
d
1
5a
5b
5c
5c
5c
Bn
allyl
H
96
96
1
6a (Me) (15) (>99.5)
6b (Me) (60) (95.2)
e
2
f
3
6c (Bn) (96) (94.8)
g
f
4
g,h
H
1
6c (Bn) (90) (>99.5)
5
H
12
6d (H) (94) (>99.5)
a
Substrate 5 (1 mmol) was incubated with Rhodococcus erythropolis
AJ270 cells (2 g wet weight) in potassium phosphate buffer (0.1 M,
pH 7.0, 50 mL) at 30 °C. DMSO (2.5 mL) was used as a cosolvent.
b
c
d
Isolated yield. Determined by chiral HPLC analysis. Starting
e
material 5a (75%) was recovered. Starting material 5b (25%) was
f
recovered. Benzylation also occurred on heterocyclic nitrogen.
g
h
DMSO was not used. Substrate (20 g) in 250 mL of buffer was used.
hydrolysis of N-allyl-substituted cis-pyrrolidine-2,5-dicarboxa-
mide 5b, which afforded 60% of (2R,5S)-methyl 1-allyl-5-
carbamoylpyrrolidine-2-carboxylate 6b along with the recovery
of reactant 5b in 25% (entry 2, Table 2). In both cases, the
amidase exhibited excellent enantioselectivity as enantiomeric
excess values of 6a and 6b reached >99.5% and 95.2%,
respectively (entries 1 and 2, Table 2). To our delight, amidase-
catalyzed desymmetrization of cis-pyrrolidine-2,5-dicarboxa-
mide 5c proceeded efficiently.33 Quantitative conversion of
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enantiomers of aza-sugar containing nucleoside analogues in
which the anomeric carbon is substituted by a tetrazole moiety,
compounds of pharmaceutical interest.34 Now having enan-
tioenriched (2R,5R)-5-carbamoylpyrrolidine-2-carboxylic acid
3d in hand, we attempted the synthesis of compound 10, a
diastereomer of tetrazole-bearing aza-nucleoside analogues. As
depicted in Scheme 4, N-protection with CbzCl followed by
sugar containing nucleoside analogues, compounds of signifi-
cance in the study of anticancer and antivirus agents.33,34 As
illustrated in Scheme 6, compound 11 underwent exhaustive
Scheme 6. Synthesis of Both Antipodes of Aza-nucleotide
Analogues 17 and 22
Scheme 4. Synthesis of Aza-nucleoside Diastereomer 10
esterification via acyl chloride led to the intermediate 7 in 87%.
High-yielding dehydration of amide 7 produced nitrile 8, which
underwent 1,3-dipolar cycloaddition with azide to afford
tetrazole product 9 effieciently. Treatment of 9 with LiAlH4
led to simultaneous reduction of the ester and N-Cbz groups,
furnishing the desired product 10 in a good yield (Scheme 4).
To explore the synthetic potential of enantiopure 6d,
available in tens of grams from biocatalytic desymmetrization
of cis-pyrrolidine-2,5-carboxamide 5c (entry 5, Table 2), we
conducted the synthesis of the druglike pyrrole-fused benzo-
[1,4]diazepinone derivative 14 (Scheme 5). Biotransformation
reduction with LiAlH4 to afford amino alcohol 15 in 79% yield.
Reductive amination with 2-phenylacetaldehyde, which gave
16, and consecutive Pictet−Spengler reaction led conveniently
to the formation of 17. Its enantiomer 22 was then synthesized
through the intermediate 19 which was resulted from the high-
yielding ester hydrolysis of 7 and subsequent amide formation
upon the treatment with 1,2,3,4-tetrahydroisoquinoline
(THIQ) in the presence of chlorodimethoxytriazine
(CDMT) and N-methylmorpholine (NMM).37 Conversion of
primary amide 19 into methyl ester 21 via nitrile 20 was
effected under mild conditions, and the final reduction of both
ester and amide functionalities with LiAlH4 afforded the desired
product 22 (Scheme 6).
Scheme 5. Synthesis of Bicyclic Ring Products 14
CONCLUSION
■
In summary, we have showed that the amidase-catalyzed
hydrolysis of pyrrolidine-2,5-dicarboxamides provides unique
and practical synthetic routes to highly enantiopure function-
alized pyrrolidine derivatives under very mild conditions. In the
presence of Rhodococcus erythropolis AJ270, an amidase
containing microbial whole cell catalyst, kinetic resolution of
racemic trans-pyrrolidine-2,5-dicarboxamides give (2S,5S)-
pyrrolidine-2,5-dicarboxamides and (2R,5R)-5-carbamoylpyrro-
lidine-2-carboxylic acids or (2R,5R)-pyrrolidine-2,5-dicarboxylic
acids in excellent yields, while desymmetrization of meso cis-
pyrrolidine-2,5-dicarboxamides produced nearly quantitatively
(2R,5S)-5-carbamoylpyrrolidine-2-carboxylic acids. In both
reactions, although the amidase exhibited the substrate- or N-
substituent-dependent catalytic efficiency, it always displayed
excellent 2R-enantioselectivity, giving products of >99.5% ee.
The resulting chiral pyrrololidine derivatives, which are not
readily available by other synthetic methods, are versatile
building blocks in organic synthesis. Their synthetic applica-
tions have been demonstrated by the construction of tetrazole-
substituted pyrrolidine and pyrroline-fused diazepin-11(5H)-
one compounds. We have also shown the practical
enantiodivergent preparation of a pair of enantiomers of aza-
product 6d was first converted into 11 in excellent yield simply
following the same procedure for the transformation of 3d to 7.
The N-Cbz group was removed easily by catalytic hydro-
genolysis and subsequent N-benzylation with o-bromobenzyl
bromide in the presence of Ag2O afforded intermediate 12 in
72% yield. Selective reduction of the ester group was achieved
using a combination of NaBH4 and LiCl35 in a mixture of
ethanol and THF (1:1), yielding compound 13 in an excellent
yield. A CuI-catalyzed intramolecular cross-coupling reaction36
resulted in the effective construction of (3R,11aS)-3-(hydrix-
ymethyl)-2,3,10,11a-tetrahydro-1H-benzo[e]pyrrolo[1,2-a]-
[1,4]diazepin-11(5H)-one 14 in good yield.
Considering the cis configuration of 2,5-functional groups
and their facile interconversions, we also envisioned the concise
and enantiodivergent construction of both antipodes of aza-
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The reaction, monitored by TLC or HPLC, was quenched after a
specified period of time (see Tables 1 and 2) by removing the biomass
through a Celite pad filtration, and the filtrate was freeze-dried. For
biotransformation of 1a−c, the residue was mixed with methanol (2
mL), and a freshly prepared solution of CH2N2 in diethyl ether (10
mL) was added while the solution was kept cool with an ice−water
bath. The mixture was stirred overnight. Water (15 mL) was added,
and the resulting mixture was extracted with ethyl acetate (8 × 50
mL). The combined organic layer was dried with anhydrous Na2SO4.
After removal of solvent, the residue was chromatographed using a
silica gel column eluted with a mixture of petroleum ether and ethyl
acetate (12:1) to give diester product 4, a mixture of petroleum ether
and ethyl acetate (1: 2) to give monoester product 3, and then a
mixture of ethyl acetate and methanol (5:1) to give dicarboxamide
product 2. The aqueous phase from extraction was freeze-dried again,
and the residue was chromatographed using a silica gel column eluted
with a mixture of ethyl acetate and methanol (5:1) to give pure
dicarboxamide product 2. For biotransformation of 5a or 5b, the
residue was mixed with methanol (2 mL), and a freshly prepared
solution of CH2N2 in diethyl ether (10 mL) was added while the
mixture was kept cool with an ice−water bath. The mixture was stirred
overnight. Water (10 mL) was added, and the resulting mixture was
extracted with ethyl acetate (3 × 10 mL) and dried with anhydrous
Na2SO4. After removal of solvent, the residue was chromatographed
using a silica gel column eluted with ethyl acetate to give product 6a or
6b. For the biotransformation of 5c, the residue was mixed with DMF
(2 mL), BnBr (1 mL), and K2CO3 (1.38 g, 10 mmol). After the
mixture was stirred for 24 h, water (10 mL) was added, and the
resulting mixture was extracted with ethyl acetate (3 × 100 mL). The
combined organic layer was washed with brine (3 × 2 mL) and dried
with anhydrous Na2SO4. After removal of solvent, the residue was
chromatographed using a silica gel column with ethyl acetate as eluent
to give pure product 6c.
Procedure for Large-Scale Biocatalytic Kinetic Resolution of
1c. Rhodococcus erythropolis AJ270 cells21,23a (2 g wet weight) were
suspended in aqueous potassium phosphate buffer (0.1 M, pH 7.0, 50
mL) in an Erlenmeyer flask (150 mL) with a screw cap and activated
at 30 °C for 0.5 h with orbital shaking. Dicarboxamide 1c (1.57 g, 10
mmol) was added in one portion to the flask, and the resulting mixture
was incubated at 30 °C using an orbital shaker (200 rpm). The
reaction, monitored by HPLC, was quenched after 1 h by removing
the biomass by filtration through a Celite pad. After removal of
solvent, the residue was loaded to an ion-exchange column (Dowex,
50WX8 200−400 mesh) and eluted with pure water until pH was
around 7.0. Elution with ammonia solution (1%) gave a mixture of 2d
and 3d. The mixture was chromatographed on a C-18 reverse phase
column eluted with water to afford enantioenriched products 2d (740
mg, 47%) and 3d (826 mg, 52%).
sugar containing nucleoside analogs from a common
intermediate (2R,5S)-5-carbamoylpyrrolidine-2-carboxylic acid.
Further exploration of the application of amidase and its
catalytic mechanism is being actively pursued in this laboratory
and the results will be reported in due course.
EXPERIMENTAL SECTION
■
Starting pyrrolidine-2,5-dicarboxamides 1 and 5 were synthesized from
chemical hydrolysis of pyrrolidine-2,5-dicarbonitriles using concen-
trated H2SO4.38 Dicarbonitriles in turn were prepared from the simple
Strecker reaction using succinaldehyde bis-sodium bisulfite and amines
according to a method reported in the literature.39
( )-trans-1-Benzylpyrrolidine-2,5-dicarboxamide (1a): 1.8 g,
1
74%; mp 218−219 °C; IR (KBr) ν 3414, 1670, 1628 cm−1; H NMR
(300 MHz/DMSO-d6) δ 7.32−7.21 (m, 5 H), 7.17 (br, s, 2 H), 6.88
(br, s, 2 H), 3.81 (d, J = 13.5 Hz, 1 H), 3.69 (d, J = 13.5 Hz, 1 H),
3.56−3.54 (m, 2 H), 2.23−2.09 (m, 2 H), 1.71−1.66 (m, 2 H); 13C
NMR (75 MHz/DMSO-d6) δ 176.4, 139.6, 129.0, 128.6, 127.3, 64.5,
53.5, 29.4; MS (ESI) m/z 248.2 (M + 1, 100), 270.2 (M + 23, 92).
Anal. Calcd for C13H17N3O2: C, 63.14; H, 6.93; N, 16.99. Found: C,
62.94; H, 6.92; N, 17.05.
( )-trans-1-Allylpyrrolidine-2,5-dicarboxamide (1b): 1.1 g,
57%; mp 223−224 °C; IR (KBr) ν 3413, 3378, 3200, 1660, 1633,
1459, 1415 cm−1; 1H NMR (300 MHz/DMSO-d6) δ 7.23 (br, s, 2 H),
6.87 (br, s, 2 H), 5.97−5.84 (m, 1 H), 5.15−5.01 (m, 2 H), 3.55−3.53
(m, 2 H), 3.28 (dd, J = 13.6, 7.2 Hz, 1 H), 3.15 (dd, J = 13.5, 5.7 Hz, 1
H), 2.21−2.07 (m, 2 H), 1.69−1.59 (m, 2 H); 13C NMR (75 MHz/
DMSO-d6) δ 176.0, 136.4, 116.5, 64.3, 52.5, 28.9; MS (ESI) m/z 198.1
(M + 1, 100), 220.2 (M + 23, 17). Anal. Calcd for C9H15N3O2: C,
54.81; H, 7.67; N, 21.30. Found: C, 54.75; H, 7.71; N, 21.36.
( )-trans-Pyrrolidine-2,5-dicarboxamide (1c): 279 mg, 89%;
mp 189−190 °C; IR (KBr) ν 3401, 3317, 3171, 1703, 1628 cm−1; 1H
NMR (300 M Hz/D2O) δ 3.86−3.82 (m, 2 H), 2.21−2.10 (m, 2 H),
1.85−1.72 (m, 2 H); 13C NMR (75 MHz/D2O) δ 180.0, 60.3, 30.4;
MS (ESI) m/z 113.19 (M − 44, 100), 158.3 (M + 1, 20). Anal. Calcd
for C6H11N3O2: 158.0930 (M + 1)+. Found: 158.0925.
cis-1-Benzylpyrrolidine-2,5-dicarboxamide (5a): 12.1 g, 84%;
mp 156−157 °C; IR (KBr) ν 3384, 3174, 1693, 1663 cm−1; 1H NMR
(300 M Hz/DMSO-d6) δ 7.82 (br, s, 2 H), 7.33−7.21 (m, 5 H), 7.03
(br, s, 2 H), 3.73 (s, 2 H), 3.24−3.21 (m, 2 H), 2.03−1.91 (m, 2 H),
1.75−1.62 (m, 2 H); 13C NMR (75 MHz/DMSO-d6) δ 176.1, 137.2,
129.4, 128.0, 127.1, 66.6, 57.1, 29.9; MS(ESI) m/z 248.2 (M + 1, 22),
270.2 (M + 23, 100), 286.2 (M + 39, 21.2). Anal. Calcd for
C13H17N3O2: C, 63.14; H, 6.93; N, 16.99. Found: C, 63.44; H, 6.97;
N, 17.11.
cis-1-Allylpyrrolidine-2,5-dicarboxamide (5b): 1.2 g, 61%; mp
148−149 °C; IR (KBr) ν 3367, 3166, 1697, 1667 cm−1; 1H NMR (300
M Hz/DMSO-d6) δ 7.78 (br, s, 2 H), 7.07 (m, 2 H), 5.98−5.84 (m, 1
H), 5.18−5.06 (m, 2 H), 3.18−3.13 (m, 4 H), 2.09−1.96 (m, 2 H),
1.72−1.61 (m, 2 H); 13C NMR (75 MHz/DMSO-d6) δ 176.2, 134.7,
117.9, 66.9, 56.9, 29.9; MS (ESI) m/z 198.2 (M + 1, 22), 220.2 (M +
23, 100). Anal. Calcd for C9H15N3O2, C, 54.81; H, 7.67; N, 21.30.
Found: C, 54.73; H, 7.76; N, 20.92.
Procedure for Large-Scale Biocatalytic Desymmetrization of
5c. Rhodococcus erythropolis AJ270 cells21,23a (2 g wet weight) were
suspended in aqueous potassium phosphate buffer (0.1 M, pH 7.0, 250
mL) in an Erlenmeyer flask (500 mL) with a screw cap and activated
at 30 °C for 0.5 h with orbital shaking. Dicarboxamide 5c (20 g) was
added in one portion to the flask, and the resulting mixture was
incubated at 30 °C using an orbital shaker (200 rpm). The reaction,
monitored by HPLC, was quenched after 12 h by removing the
biomass by filtration through a Celite pad. The filtrate was
concentrated to 100 mL using a rotary evaporator under reduced
pressure, and product 6d (8.0 g) precipitated from solution. The
mother liquid was subjected to an ion-exchange column and eluted
with aqueous ammonia solution (1%) to give another portion of
product 6d (11.0 g).
cis-Pyrrolidine-2,5-dicarboxamide (5c). 280 mg, 91%; mp 220−
1
221 °C; IR(KBr) ν 3356, 3209, 1643 cm−1; H NMR (300 M Hz/
DMSO-d6) δ 8.09 (br, s, 1 H), 7.77 (br, s, 1 H), 4.21 (t, 2 H, J = 5.8
Hz), 2.39−2.26 (m, 2 H), 1.93−1.78 (m, 2 H); 13C NMR (75 MHz/
DMSO-d6) δ 169.3, 59.3, 29.6; MS (ESI) m/z 158.1 (M + 1, 100),
180.1 (M + 23, 95). Anal. Calcd for [M + H] C6H12N3O2: 158.09295
[M + H]. Found: 158.09246.
General Procedure for the Biotransformation of Pyrrolidine-
2,5-dicarboxamides. To an Erlenmeyer flask (150 mL) with a screw
cap were added Rhodococcus erythropolis AJ270 cells21,23a (2 g wet
weight) and potassium phosphate buffer (0.1 M, pH 7.0, 50 mL), and
the resting cells were activated at 30 °C for 0.5 h with orbital shaking.
Racemic trans-pyrrolidine-2,5-dicarboxamides 1 or meso cis-pyrroli-
dine-2,5-dicarboxamides 5, which may be dissolved in DMSO (2.5
mL) (see Tables 1 and 2), were added in one portion to the flask, and
the mixture was incubated at 30 °C using an orbital shaker (200 rpm).
All biotransformation products were fully characterized, and
characterization data are as follows.
(2S,5S)-1-Benzylpyrrolidine-2,5-dicarboxamide (2a): 121 mg,
49%; mp 235 °C; [α]25 −48.0 (c 0.5, CH3OH); ee >99.5% (HPLC
D
with ADH). Spectroscopic data identical to those of racemic
compound 1a were obtained.
(2S,5S)-1-Allylpyrrolidine-2,5-dicarboxamide (2b): 88.7 mg,
45%; mp 220−221 °C; [α]25 −58.8 (c 0.5, CH3OH); ee >99.5%
D
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Article
(HPLC with ADH). Spectroscopic data identical to those of racemic
1b were obtained.
129.0, 128.2, 127.2, 63.4, 54.1, 51.5, 28.4; MS (ESI) m/z 278.2 (M + 1,
100), 300.2 (M + 23, 8.0). Anal. Calcd for C15H19NO4: 277.1314.
Found: 277.1318.
(2S,5S)-Pyrrolidine-2,5-dicarboxamide (2d): 738 mg, 47%; mp
170 °C; [α]25 −111.3 (c 0.75, H2O). Spectroscopic data identical to
(2R,5R)-Dimethyl 1-allylpyrrolidine-2,5-dicarboxylate (4b):
D
those of 1c were obtained. To determine the enantiomeric excess (ee)
value and the absolute configuration of product 2d, a transformation of
2d into 2a was conducted. Thus, a mixture of 2d (75 mg), K2CO3 (0.7
g), and benzyl bromide (0.5 mL) in dry DMF (1 mL) was stirred at
ambient temperature for 24 h. The mixture was submitted to a short
silica column and eluted with ethyl acetate to remove side product and
then with a mixture of ethyl acetate and methanol (20:1−10:1) to gave
2a with ee >99.5% (HPLC analysis with a Daicel ADH column).
(2R,5R)-Methyl 1-benzyl-5-carbamoylpyrrolidine-2-carboxy-
23 mg, 10%; oil; [α]25 = +72.0 (c 1.5, CHCl3); ee 93.3% (HPLC
D
1
with OJH); IR ν (KBr) 1738 cm−1; H NMR (300 MHz/CDCl3) δ
5.96−5.82 (m, 1 H), 5.18−5.05 (m, 2 H), 3.89−3.85 (m, 2 H), 3.70
(s, 6 H), 3.45−3.33 (m, 2 H), 2.36−2.28 (m, 2 H), 1.94−1.89 (m, 2
H); 13C NMR (75 MHz/CDCl3) δ 174.7, 135.3, 117.7, 63.6, 53.5,
51.6, 28.4; MS (EI) m/z 227 (M+, 3), 168 (100). Anal. Calcd for
C11H17NO4: 227.1158. Found: 227.1161.
(2R,5S)-Methyl 1-benzyl-5-carbamoylpyrrolidine-2-carboxy-
late (6a): 39 mg, 15%; oil; [α]25 +10.7 (c 3.0, C6H6); ee >99.5%
D
late (3a): 84 mg, 32%; mp 140−141 °C; [α]25 +121.5 (c 4.0,
1
(HPLC with ADH); IR (KBr) ν 3415, 1736, 1678 cm−1; H NMR
D
CHCl3); ee >99.5% (HPLC with ADH); IR (KBr) ν 3446, 1734,
1685, 1658 cm−1; 1H NMR (300 MHz/CDCl3) δ 7.36−7.21 (m, 5 H),
6.91 (br, s, 1 H), 5.50 (br, s, 1 H), 3.92 (d, 1 H, J = 13.1 Hz), 3.84 (d,
1 H, J = 13.1 Hz), 3.80−3.72 (m, 2 H), 3.69 (s, 3 H), 2.58−2.51 (m, 1
H), 2.13−2.03 (m, 1 H), 2.00−1.85 (m, 2 H); 13C NMR (75 MHz/
CDCl3) δ 177.7, 173.5, 138.0, 128.74, 128.66, 127.6, 65.9, 62.9, 54.3,
51.4, 29.4, 28.6; MS (ESI) m/z 263.2 (M + 1, 100%), 285.2 (M + 23,
64.3), 301.2 (M + 39, 4.9). Anal. Calcd for C14H18N2O3: C, 64.10; H,
6.92; N, 10.68. Found: C, 63.85; H, 6.90; N, 10.94.
(300 MHz/CDCl3) δ 7.97 (br, s, 1 H), 7.33−7.25 (m, 5 H), 5.57 (br,
s, 1 H), 3.89 (d, J = 13.2 Hz, 1 H), 3.82 (d, J = 13.2 Hz, 1 H), 3.63−
3.51 (m, 2 H), 3.58 (m, 3 H), 2.20−1.85 (m, 4 H); 13C NMR (75
MHz/CDCl3) δ 177.9, 175.7, 137.2, 129.3, 128.5, 127.6, 67.7, 66.2,
59.0, 52.0, 30.7, 30.4; MS (ESI) m/z 263.2 (M + 1, 60), 285.2 (M +
23, 100). Anal. Calcd for: C14H18N2O3, C, 64.10; H, 6.92; N, 10.68;
Found: C, 64.25; H, 6.87; N, 10.92.
(2R,5S)-Methyl 1-allyl-5-carbamoylpyrrolidine-2-carboxy-
late (6b): 127 mg, 60%; oil; [α]25 +6.0 (c 4, CH2Cl2); ee 95.2%
D
1
(HPLC with ADH); IR (KBr) ν 3415, 1738, 1681 cm−1; H NMR
(2R,5R)-Methyl 1-allyl-5-carbamoylpyrrolidine-2-carboxy-
late (3b): 57 mg, 27%; mp 104−105 °C; [α]25 = +78.0 (c1.0,
(400 MHz/CDCl3) δ 7.99 (br, s, 1 H), 5.87−5.78 (m, 1 H), 5.45 (br,
s, 1 H), 5.20−5.12 (m, 2 H), 3.73 (s, 3 H), 3.57 (t, 1 H, J = 7.6 Hz),
3.45−3.42 (m, 1 H), 3.32 (d, 2 H, J = 6.8 Hz), 2.20−2.12 (m, 2 H),
2.07−2.01 (m, 1 H), 1.92−1.86 (m, 1 H); 13C NMR (75 MHz/
CDCl3) δ 178.1, 175.8, 133.8, 118.7, 67.0, 65.7, 57.2, 52.1, 30.7, 30.4;
MS (ESI) m/z 213.2 (M + 1, 43), 235.2 (M + 23, 100). Anal. Calcd
for C10H16N2O3: C, 56.59; H, 7.60; N, 13.20. Found: C, 56.62; H,
7.68; N, 13.16.
D
CHCl3); ee >99.5% (HPLC with ADH); IR(KBr) ν 3377, 3190, 1728,
1
1654 cm−1; H NMR (300 MHz/CDCl3) δ 6.91 (br, s, 1 H), 5.89−
5.78 (br, s, 1 H, m, 1 H), 5.20−5.10 (m, 2 H), 3.93 (d, J = 7.5 Hz, 1
H), 3.69 (s, 3 H), 3.61 (dd, J = 10.8, 3.3 Hz, 1 H), 3.36−3.34 (m, 2
H), 2.54−2.47 (m, 1 H), 2.16−2.09 (m, 1 H), 1.96−1.87 (m, 2 H);
13C NMR (75 MHz/CDCl3) δ 178.2, 173.5, 134.8, 118.0, 65.4, 63.3,
53.1, 51.4, 29.3, 28.6; MS (ESI) m/z 213.2 (M + 1, 100), 235.2 (M +
23, 6.5). Anal. Calcd for C10H16N2O3: C, 56.59; H, 7.60; N, 13.20.
Found: C, 56.57; H, 7.61; N, 12.94.
(2R,5S)-Benzyl 1-benzyl-5-carbamoylpyrrolidine-2-carboxy-
late (6c): 304 mg, 90%; mp 58−59 °C; [α]25D +19.2 (c 1, CH2Cl2); ee
>99.5% (HPLC analysis with a Diacel ADH column); IR (KBr) ν
(2R,5R)-Benzyl 1-benzyl-5-carbamoylpyrrolidine-2-carboxy-
1
3422, 3369, 1740, 1663 cm−1; H NMR (300 MHz/CDCl3) δ 7.89
late (3c): 446 mg, 44%; mp 104−105 °C; [α]25 = +156.0 (c 1.0,
D
(br, s, 1 H), 7.37−7.22 (m, 10 H), 5.90 (br, s, 1 H), 4.98 (s, 2 H), 3.88
(d, J = 13.2 Hz, 1 H), 3.79 (d, J = 13.2 Hz, 1 H), 3.63 (t, 1 H, J = 7.4
Hz), 3.54−3.50 (m, 1 H), 2.19−1.88 (m, 4 H); 13C NMR (75 MHz/
CDCl3) δ 177.1, 174.1, 136.4, 134.7, 128.5, 127.8, 127.6, 127.57,
127.3, 126.8, 67.1, 65.9, 65.5, 58.2, 29.8, 29.5; MS (ESI) m/z 339 (M +
1, 86), 361 (M + 23, 100). Anal. Calcd for C20H22N2O3: C, 70.99; H,
6.55; N, 8.28. Found: C, 70.65; H, 6.71; N, 8.53. Slow vapor diffusion
of diethyl ether into a mixture of 6c (15.7 mg), concentrated
hydrochloric acid (15 μL) and ethanol (0.5 mL) at 5 °C for 2 days led
to the formation of an X-ray-quality single crystal of 6c·HCl.33
(2R,5S)-5-Carbamoylpyrrolidine-2-carboxylic acid (6d): 19.0
CHCl3); ee 93.0% (HPLC with ADH); IR (KBr) ν 3396, 3172, 1726,
1
1657, 1161 cm−1; H NMR (300 MHz/CDCl3) δ 7.41−7.10 (m, 10
H), 6.96 (br, s, 1 H), 5.49 (br, s, 1H), 5.18 (d, J = 12.0 Hz, 1 H), 5.08
(d, J = 12.0 Hz, 1 H), 3.91−3.72 (m, 4 H), 2.61−2.47 (m, 1 H), 2.15−
1.85 (m, 3 H); 13C NMR (75 MHz/CDCl3) δ 177.9, 172.9, 137.9,
135.6, 128.8, 128.6, 128.5, 127.5, 66.3, 65.8, 62.9, 54.1, 29.4, 28.6; MS
(ESI) m/z 339.1 (M + 1, 100), 361.2 (M + 23, 82). Anal. Calcd for
C20H22N2O3: C, 70.99; H, 6.55; N, 8.28. Found: C, 70.60; H, 6.60; N,
8.37.
(2R,5R)-5-Carbamoylpyrrolidine-2-carboxylic acid (3d): 822
mg, 52%; mp 251 °C (sublimation); [α]25 = +95.3 (c 1.0, H2O); IR
D
g, 94%; mp 233−234 °C; [α]25 −9.6 (c 1, H2O); IR (KBr) ν 3308,
(KBr) ν 3396, 3340, 3129, 3050, 1692, 1689, 1657, 1613, 1405 cm−1;
1H NMR (300 MHz/D2O) δ 4.43 (t, J = 7.3 Hz, 1 H), 4.20 (t, J = 7.3
Hz, 1 H), 2.41−2.28 (m, 2 H), 2.13−1.98 (m, 2 H); 13C NMR (75
MHz/D2O) δ 173.8, 171.4, 61.9, 60.1, 29.6, 28.8; MS (ESI) m/z 339.1
(M + 1, 100), 361.2 (M + 23, 82). Anal. Calcd for C6H11N2O3:
159.0769. Found: 159.0760. To determine the ee value and the
absolute configuration of product 3d, transformation of 3d into 3c was
conducted. Thus, a mixture of 3d (75 mg), K2CO3 (0.7 g), and benzyl
bromide (0.5 mL) in dry DMF (1 mL) was stirred at ambient
temperature for 24 h. Water (10 mL) was then added, and the mixture
was extracted with ethyl acetate (3 × 10 mL). The combined organic
layer was washed with brine (3 × 10 mL) and dried with anhydrous
Na2SO4. After removal of organic solvent, the residue was chromato-
graphed on a silica gel column eluted with ethyl acetate to give 3c with
ee 87.1% (HPLC analysis with a Daicel ADH column). Enantiomeric
purity of 3d was improved to >99.5% ee after recrysttalization in a
mixture of water and methanol
D
3153, 2361, 1696, 1642, 1576 cm−1; 1H NMR (300 MHz/D2O) δ 4.34
(t, J = 7.2 Hz, 1 H), 4.11 (t, J = 6.3 Hz, 1 H), 2.39−2.25 (m, 2 H),
2.06−1.91 (m, 2 H); 13C NMR (75 MHz/D2O) δ 173.8, 171.4, 61.8,
60.31, 29.8, 29.2; MS (ESI) m/z 159 (M + 1, 7), 181 (M + 23, 100).
Anal. Calcd for C6H10N2O3: C, 45.57; H, 6.37; N, 17.71. Found: C,
45.54; H, 6.53; N, 17.78. To determine the enantiomeric excess (ee)
value and the absolute configuration of product 6d, transformation of
6d into 6c was conducted. Thus, a mixture of 6d (158 mg), K2CO3
(1.38 g), and benzyl bromide (1 mL) in dry DMF (2 mL) was stirred
at ambient temperature for 24 h. Water (10 mL) was then added, and
the mixture was extracted with ethyl acetate (3 × 10 mL). The
combined organic layer was washed with brine (3 × 10 mL) and dried
with anhydrous Na2SO4. After removal of organic solvent, the residue
was chromatographed on a silica gel column eluted with ethyl acetate
to give 6c [ee >99.5% (HPLC analysis with a Daicel ADH column)].
General Procedure for Chemical Hydrolysis of 2a and 3c. A
solution of (2S,5S)-2a (50 mg, 0.2 mmol, ee >99.5%, HPLC analysis
with a Daicel ADH column) or (2R,5R)-3c (67.6 mg, 0.2 mmol, ee
88.5%, HPLC analysis with a Daicel ADH column) in hydrochloric
acid (6 N, 8 mL) was refluxed for 12 or 8 h. The solvent was removed
under vacuum, and water in the residue was further removed by
azotropic distillation with methanol (3 × 3 mL) using a rotary
(2R,5R)-Dimethyl 1-benzylpyrrolidine-2,5-dicarboxylate
(4a): 33 mg, 12%; oil; [α]25 = +58.7 (c 1.5, CHCl3); ee >99.5%
D
1
(HPLC with ODH); IR (KBr) ν 1737 cm−1; H NMR (300 MHz/
CDCl3) δ 7.31−7.21 (m, 5 H), 3.97 (d, J = 12.9 Hz, 1 H), 3.86−3.82
(m, 2 H), 3.79 (d, J = 13.1 Hz, 1 H), 3.64 (s, 6 H), 2.38−2.24 (m, 2
H), 1.99−1.88 (m, 2 H); 13C NMR (75 MHz/CDCl3) δ 174.7, 138.5,
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evaporator. The resulting residue was mixed with methanol (2 mL),
and then CH2N2 (5 mL) was added dropwise at ambient
temperature.After the mixture was stirred overnight, water (2 mL)
was added, and the mixture was extracted with ethyl aceate (3 × 10
mL). The combined organic layer was dried with anhydrous sodium
sulfate. After removal of solvent, the residue was purified through a
silica gel column eluted with a mixture of petroleum ether and ethyl
acetate (6:1) to give pure (2S,5S)-4a (50 mg, 90%) with 89.2% ee
(HPLC analysis with a Daicel ODH column) or (2R,5R)-4a (55 mg,
99%) with 85.9% ee (HPLC analysis with a Daicel ODH column).
(2S,5S)-Dimethyl 1-benzylpyrrolidine-2,5-dicarboxylate (4a):
Synthesis of 10. Under argon protection, a mixture of 9 (397 mg,
1.2 mmol) and LiAlH4 (456 mg, 12 mmol) in dry THF (10 mL) was
refluxed for 24 h. While the mixture was kept at −20 °C, NaOH
aqueous solution (2 N, 1 mL) was slowly injected through a syringe.
The resulting mixture was filtrated through a Celite pad and washed
thoroughly with a mixture of CH2Cl2 and MeOH (20:1). After
removal of solvent, the residue was mixed with hydrochloric acid (1
N) with pH value of the mixture being adjusted to around 2.0. The
resulting mixture was subjected to an ion-exchange column (Dowex,
50WX8 100−200 mesh) and then eluted with pure water. When the
pH of the fraction was around 7, the column was eluted with aqueous
ammonia solution (1%). After removal of solvent, product 10 was
[α]25 = −40.0 (c 1.5, CHCl3); ee >89.2% (HPLC with ODH).
D
Identical spectroscopic data as that of (2R,5R)-4a were obtained.
Synthesis of 7. To a mixture of 3d (1.19 g, 7.5 mmol) in ethanol
(20 mL) and saturated NaHCO3 aqueous solution (20 mL) was added
dropwise CbzCl (3 mL) at room temperature with stirring. The
resulting mixture was allowed to stire overnight. The reaction was
quenched by addition of hydrochloric acid (2 N) until the pH was
adjusted to around 7.0. Under reduced pressure, the solvent was
completely removed. The residue was mixed with methanol (50 mL)
and SOCl2 (4 mL) at ambient temperature. After the misture was
stirred for another 4 h at ambient temperature, saturated NaHCO3
aqueous solution (100 mL) was added. The mixture was extracted
with ethyl acetate (3 × 50 mL). The combined organic layer was
washed with brine (3 × 10 mL) and dried with anhydrous Na2SO4.
After removal of organic solvent, the residue was chromatographed on
a silica gel column eluted with ethyl acetate to give product 7 (2.0 g,
obtained. 10 (182 mg, 83%): mp 194−195 °C; [α]25 +16.0 (c 1.0,
D
H2O); IR (KBr) ν 3412, 3155, 1575, 1461, 1408, 1089 cm−1; 1H NMR
(600 MHz/D2O) δ 5.00 (t, J = 12.6 Hz, 1 H), 3.87−3.86 (m, 2 H),
3.58−3.55 (m, 1 H), 2.54−2.34 (m, 3 H), 2.41 (s, 3 H), 2.06−2.00
(m, 2 H); 13C NMR (75 MHz/D2O) δ 158.0, 67.0, 60.7, 59.3, 35.7,
27.4, 25.5; MS (ESI) m/z 184.5 (M + 1, 100). Anal. Calcd for
C7H14N5O1: 184.1198. Found: 184.1194.
Synthesis of 11. To a stirred mixture of 6d (4.0 g, 25 mmol) in
ethanol (20 mL) and saturated aqueous NaHCO3 solution (20 mL)
was added dropwise CbzCl (6 mL) at room temperature. The
resulting mixture was allowed to stir overnight at room temperature.
After concentration by removing ethanol using a rotary evaporator
under reduced pressure, the resulting aqueous solution (about 15 mL)
was neutralized with hydrochloric acid (2 N) to pH 7.0. The solvent
was removed under vacuum, and water in the residue was further
removed by azotropic distillation with methanol (3 × 30 mL) using a
rotary evaporator. The resulting residue was mixed with methanol
(100 mL) and then SOCl2 (4 mL) at room temperature. After being
stirred for another 4 h, the reaction mixture was poured slowly into a
saturated aqueous solution of NaHCO3 (200 mL), and the resultant
mixture was extracted with ethyl acetate (3 × 100 mL). The combined
organic layer was washed with brine (3 × 20 mL) and dried with
anhydrous sodium sulfate. The solvent was removed and the residue
was chromatographed on a silca gel column eluted with ethyl acetate
to give pure product 11 (6.96 g, 91%): mp 104−105 °C; [α]25D +46.6
(c 0.6, CHCl3); IR (KBr) ν 3435, 3401, 1734, 1700, 1683, 1415, 1356,
87%): oil; [α]25 +30 (c 1, CHCl3); IR (KBr) ν 3421, 1743, 1698,
D
1
1683 cm−1; H NMR (400 MHz/DMSO-d6, 405 K) δ 7.34−7.28 (m,
5 H), 6.76 (br, s, 2 H), 5.04 (t, J = 14.0 Hz, 2 H), 4.43 (dd, J = 8.8, 1.7
Hz, 1 H), 4.33−4.30 (m, 1 H), 3.58 (s, 3 H), 2.32−2.13 (m, 2 H),
1.92−1.86 (m, 2 H); 13C NMR (75 MHz/CDCl3) δ 175.2, 174.5,
172.9, 172.7, 1584.9, 154.7, 136.2, 136.0, 128.0, 127.9, 127.7, 127.5,
67.4, 60.6, 60.1, 59.7, 52.3, 52.1, 29.5, 29.3, 28.0, 27.6; MS (ESI) m/z
329.4 (M + 23, 100). Anal. Calcd for C15H19N2O5: 307.1294. Found:
307.1291.
Synthesis of 8. To a solution of 7 (1.6 g, 5.2 mmol) in dry DMF
(10 mL) was added SOCl2 (1.1 mL) at 0 °C. After being stirred for
another 1 h at room temperature, the reaction was quenched by the
addition of saturated NaHCO3 aqueous solution (10 mL). The
mixture was extracted with ethyl acetate (3 × 30 mL). The combined
organic layer was washed with brine (3 × 10 mL) and dried with
anhydrous Na2SO4. After removal of organic solvent, the residue was
chromatographed on a silica gel column eluted with a mixture of ethyl
acetate and petroleum ether (1:2) to give compound 8 (1.3 g, 90%):
1
1213, 1117, 1004 cm−1; H NMR (300 MHz/DMSO-d6, 375 K) δ
7.35−7.25 (m, 5 H), 6.97 (br, s, 2 H), 5.09 (d, J = 12.6 Hz, 1 H), 5.05
(d, J = 12.6 Hz, 1 H), 4.42 (t, J = 7.2 Hz, 1 H), 4.18 (dd, 1 H, J = 3.6,
8.3 Hz), 3.66 (s, 3 H), 2.31−2.17 (m, 2 H), 1.98−1.87 (m, 2 H); 13C
NMR (75 MHz/DMSO-d6) δ 175.0, 174.8, 174.3, 174.0, 154.4, 154.1,
136.9, 136.7, 128.8, 128.4, 128.3, 127.7, 127.6, 67.2, 67.1, 62.5, 62.1,
60.5, 60.1, 53.1, 53.0, 30.6, 29.7,29.6, 28.8; MS (ESI) m/z 307.35 (M +
1, 20), 329.29 (M + 23, 100). Anal.Calcd for C15H18N2O5: C, 58.82;
H, 5.92; N, 9.15. Found: C, 58.87; H, 5.99; N, 9.10.
oil; [α]25 +31.3 (c 1.3, CHCl3); IR (KBr) ν 1747, 1717, 1407, 1351
D
1
cm−1; H NMR (300 MHz/DMSO-d6) δ 7.41−7.27 (m, 5 H), 5.20−
4.94 (m, 3 H), 4.55−4.46 (m, 1 H), 3.65 (s, 1.2 H), 3.56 (s, 1.7 H),
2.44−2.31 (m, 2 H), 2.26−2.08 (m, 2 H); 13C NMR (75 MHz/
CDCl3) δ 171.8, 153.5, 135.6, 135.57, 128.59, 128.51, 128.3, 128.0,
118.5, 118.3, 68.3, 68.0, 59.0, 58.8, 52.6, 52.4, 48.1, 47.5, 29.9, 29.7,
28.9, 28.6; MS (ESI) m/z 311.5 (M + 23, 100). Anal. Calcd for
C15H17N2O4: 289.1188. Found: 289.1188.
Synthesis of 12. To a flask charged with Pd/C catalyst (10%, 300
mg) was added a solution of 11 (3.06 g, 10 mmol) in methanol (50
mL). The mixture was allowed to stir at room temperature for 4 h
under hydrogen atmosphere with a H2 balloon. After removal of
catalyst by filtration, the filtrate was concentrated to 20 mL. Ag2O
(2.32 g, 10 mmol) and 1-bromo-2-(bromomethyl)benzene (2.5 g, 10
mmol) were added, and the resulting mixture was stirred at room
temperature for 2.5 h. Ethyl acetate (10 mL) was added, and the
reaction mixture was filtrated to remove solid residue. The filtrate was
concentrated under vacuum, and the residue was chromatographed on
a silica gel column eluted with ethyl acetate to afford pure 12 (2.46 g,
72%): mp 110−111 °C; [α]25D −8.0 (c 2.0, CHCl3); IR (KBr) ν 3419,
Synthesis of 9. A mixture of 8 (576 mg, 2 mmol), NaN3 (260
mg,4 mmol), and ZnBr2 (225 mg,1 mmol) in water (6 mL) and 2-
propanol (3 mL) was refluxed for 4 h. After the mixture was cooled to
room temperature, hydrochloric acid (1 N, 10 mL) was added. The
mixture was extracted with ethyl acetate (3 × 20 mL). The combined
organic layer was washed with brine (3 × 4 mL) and dried with
anhydrous Na2SO4. After removal of organic solvent, the residue was
chromatographed on a silica gel column eluted with a mixture of ethyl
acetate and methanol (1:2) to give compound 9 (596 mg, 90%): mp
1
1737, 1678 cm−1; H NMR (300 MHz/CDCl3) δ 7.91 (br, s, 1 H),
7.47 −7.05 (m, 5 H), 5.80 (br, 1 H), 3.94 (d, J = 13.5 Hz, 1 H), 3.91
(d, J = 13.5 Hz, 1 H), 3.60−3.47 (m, 2 H), 3.49 (s, 3 H), 2.20−1.97
(m, 3 H), 1.90−1.77 (m, 1 H); 13C NMR (75 MHz/CDCl3) 177.7,
175.3, 136.5, 133.0, 131.8, 129.4, 127.5, 125.0, 67.8, 66.6, 58.8, 52.0,
30.7, 30.4. MS (ESI) m/z 341.3, 343.1 (M + 1, 20), 363.2, 365.2 (M +
23, 100). Anal. Calcd for C14H17BrN2O3: C, 49.28; H, 5.02; N, 8.21.
Found: C, 49.32; H, 5.20, N, 8.07.
Synthesis of 13. To a solution of 12 (342 mg, 1 mmol) in a
mixture of THF and ethanol (10 mL, 1:1) were added NaBH4 (80 mg,
2.1 mmol) and LiCl (80 mg, 1.9 mmol). After the mixture was stirred
136−137 °C; [α]25 +28.0 (c 1.0, MeOH); IR (KBr) ν 1746, 1713,
D
1
1412, 1353, 1202 cm−1; H NMR (400 MHz/DMSO-d6, 405 K) δ
7.23−7.13 (m, 5 H), 5.42−5.40 (m, 1 H), 4.92 (s, 2 H), 4.48 (d, J =
6.7 Hz, 1 H), 3.60 (s, 3 H), 2.42−2.20 (m, 2 H), 1.95−1.81 (m, 2 H);
13C NMR (75 MHz/DMSO-d6) δ 172.6, 172.3, 153.6, 153.2, 136.6,
136.4, 128.3, 128.1, 127.8, 127.3, 126.5, 66.3, 65.8, 59.3, 58.8, 53.3,
53.0, 52.0, 31.2, 30.0, 28.4, 27.1; MS (ESI) m/z 330.5 (M − 1, 100).
Anal. Calcd for C15H18N5O4: 332.1359. Found: 332.1355.
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at room temperature for 12 h, the reaction was quenched by adding a
saturated aqueous NaHCO3 solution (10 mL). The mixture was
extracted with ethyl acetate (3 × 10 mL), and the combined organic
layer was dried with anhydrous sodium sulfate. The solvent was
removed using a rotavapor and the residue was chromatographed on a
silica gel column eluted with ethyl acetate to yield pure 13 (287 mg,
92%): mp 114−115 °C; [α]25D +14.2 (c 1.5, CHCl3); IR(KBr) ν 3405,
3318, 1669 cm−1; 1H NMR (300 MHz/DMSO-d6) δ 7.58−7.17 (m 5
H), 6.95 (br, s, 1 H), 4.67 (t, 1 H, J = 5.1 Hz), 3.95 (d, J = 13.5 Hz, 1
H), 3.78 (d, J = 13.5 Hz, 1 H), 3.41−3.37 (m, 2 H), 3.21−3.17 (m, 1
H), 2.94 (br, s, 1 H), 2.05−1.96 (m, 1 H), 1.86−1.58 (m, 3 H).13C
NMR (75 MHz/DMSO-d6) δ 176.6, 137.8, 132.5, 132.0, 129.1, 127.5,
124.0, 67.2, 66.8, 62.7, 57.9, 29.2, 27.4; MS (ESI) m/z 313.3, 315.2 (M
+ 1, 50), 335.2, 337.2 (M + 23, 100). Anal. Calcd for C13H17BrN2O2:
C, 49.85; H, 5.47; N, 8.94. Found: C, 49.79; H, 5.57; N, 8.58.
Synthesis of 14. Under argon protection, a mixture of 13 (157
mg, 0.5 mmol), CuI (38.2 mg, 0.2 mmol), N,N-dimethylglycine
hydrochloric acid (DMGC) (56 mg, 0.4 mmol), and Cs2CO3 (326 mg,
1 mmol) in dry 1,4-dioxane (15 mL) was refluxed for 12 h. Water (10
mL) was added, and the resulting mixture was extracted with ethyl
acetate (3 × 10 mL). The combined organic layer was dried with
anhydrous sodium sulfate. The solvent was removed using a rotavapor,
and the residue was chromatographed on a silica gel column eluted
with ethyl acetate to produce 14 (101 mg, 87%): oil; [α]25D +22 (c 1.0,
aqueous NaHCO3 to pH 8.0 and extracted with dichloromethane (3 ×
10 mL). The combined organic layer was dried with anhydrous
sodium sulfate. After removal of solvent, the residue was chromato-
graphed on a C-18 reversed-phase column eluted with a mixture of
methanol and water (1:1) to yield product 17 (100 mg, 78%): oil;
[α]25 −50.7 (c 1.5, CHCl3); IR (KBr) ν 3424, 2943, 1641, 1457,
D
1
1036 cm−1; H NMR (300 MHz/DCl) δ 6.95−6.80 (m, 4 H), 4.31
(dd, J = 15.0, 7.8 Hz, 1 H), 4.11 (dd, J = 14.9, 5.4 Hz, 1 H), 3.79−3.73
(m, 1 H), 3.67−3.34 (m, 6 H), 3.24−3.13 (m, 1 H), 3.02−2.88 (m, 1
H), 2.81−2.68 (m, 1 H), 2.76 (s, 1.5 H), 2.74 (s, 1.5 H), 2.35−2.23
(m, 1 H), 2.02−1.90 (m, 1 H), 1.80−1.58 (m, 2 H); 13C NMR (75
MHz/D2O) δ133.6, 133.2, 128.7, 126.8, 126.7, 126.1, 68.7, 65.4, 61.9,
59.4, 55.3, 50.5, 39.7, 28.9, 27.1, 25.9; MS (ESI) m/z 261.4 (M + 1,
100). Anal. Calcd for C16H25N2O: 261.1967. Found: 261.1961.
Synthesis of 18. A mixture of 11 (306 mg, 1 mmol), LiOH (80
mg, 2 mmol), THF (4 mL), and water (4 mL) was stirred at room
temperature for 2 h. The mixture was acidified with hydrochloric acid
(10%) to pH 2.0. THF was removed using a rotavapor, and white
solids precipitated from solution. After the mixture was cooled in an
ice−water bath, filtration and washing with cold water afforded pure
product 18 (277 mg, 95%): mp 198−199 °C; IR (KBr) ν 3374, 1712
cm−1; 1H NMR (300 MHz/DMSO-d6) δ 13.55 (br, s, 1 H), 7.88 (br,
s, 0.5 H), 7.86 (br, s, 0.5 H), 7.60 (br, s, 0.5 H), 7.58 (br, s, 0.5 H),
7.39−7.33 (m, 5 H), 5.15−5.03 (m, 2 H), 4.41−4.24 (m, 2 H), 2.36−
2.21 (m, 2 H), 1.92−1.85 (m, 2 H); 13C NMR (75 MHz/DMSO-d6) δ
175.3, 175.1, 174.9, 174.7, 153.6, 153.5, 136.41, 136.36, 128.3, 127.8,
127.1, 127.0, 66.51, 66.46, 61.5, 61.0, 60.5, 60.0, 30.2, 29.3, 28.2; MS
(ESI) m/z 293.3 (M + 1, 10), 315.3 (M + 23, 100). Anal. Calcd for
C14H16N2O5: C, 57.53; H, 5.52; N, 9.58. Found: C, 57.75; H, 5.51; N,
9.58.
1
CHCl3); IR (KBr) ν 3417, 1455, 1424 cm−1; H NMR (300 MHz/
DMSO-d6) δ 9.73 (s, 0.5 H), 7.18−6.96 (m, 4 H), 4.63 (t, 0.3 H, J =
5.1 Hz), 4.28 (d, J = 15.3 Hz, 1 H), 3.92 (d, J = 15.3 Hz, 1 H), 3.53−
3.48 (m, 1 H), 3.39−3.28 (m 2 H), 2.77−2.72 (m, 1 H), 2.23−2.14
(m, 1 H), 1.81−1.76 (m, 2 H), 1.51−1.43 (m, 1 H). 13C NMR (75
MHz/DMSO-d6) δ 172.3, 136.9, 136.7, 129.3, 127.73, 127.68, 127.1,
123.0, 122.9, 120.8, 120.7, 67.0, 66.9, 65.8, 65.7, 63.8, 63.7, 56.9, 56.8,
27.3, 24.1; MS (EI) m/z 232 (M+). Anal. Calcd for C13H16N2O2:
232.1212. Found: 232.1214.
Synthesis of 19. To a suspension of 18 (1.46 g, 5 mmol) in
dichloromethane (10 mL) were added 2-chloro-4,6-dimethoxytriazine
(CDMT) (1.05 g, 6 mmol) and N-methylmorpholine (NMM) (1.52
g, 15 mmol). After the mixture was stirred at room temperature for 1
h, tetrahydroisoquinoline (732 mg, 5.5 mmol) was added, and the
resulting mixture was stirred overnight. The reaction was quenched by
adding hydrochloric acid (1 N, 10 mL), and was extracted with
dichloromethane (3 × 10 mL). The combined organic phase was dried
with anhydrous sodium sulfate. After removal of the solvent, the
residue was chromatographed on a silica gel column eluted with ethyl
acetate to give product 19 (1.83 g, 90%) as a glassy solid: mp 77−78
°C; [α]25D +22.6 (c 0.26, CHCl3); IR (KBr) ν 3421, 1579, 1446, 1410,
Synthesis of 15. A mixture of 11 (1.53 g, 5 mmol) and LiAlH4
(1.9 g, 50 mmol) in dry THF (50 mL) was refluxed for 24 h. After the
mixture was cooled to room temperature, aqueous NaOH (2 N) was
added, and the resulting mixture was filtrated through a Celite pad.
The filter cake was washed thoroughly with a mixture of ethyl acetate
and methanol (20:1). After removal of solvent, the residue was loaded
to an ion-exchange column (Dowex, 50WX8 100−200 mesh) and
eluted with pure water until the pH was around 7.0. Elution with
ammonia solution (1−5%) gave product 15 (568 mg, 79%): oil;
[α]25D −18 (c 1.0, CHCl3); IR (KBr) ν 3403, 2952, 2875, 1637, 1569
cm−1; 1H NMR (300 MHz/CDCl3) δ 3.64−3.59 (m, 1 H), 3.42−3.37
(m, 1 H), 2.77−2.64 (m, 2 H), 2.58−2.42 (m, 2 H), 2.30 (s, 3 H),
1.82−1.69 (m, 6 H), 1.59−1.50 (m, 1 H); 13C NMR (75 MHz/
CDCl3) δ 68.7, 67.5, 61.7, 44.5, 39.3, 27.2, 26.1; MS (ESI) m/z 145.1
(M + 1, 100). Anal. Calcd for C7H17N2O: 145.1341. Found: 145.1336.
Synthesis of 16. To a solution of 15 (90 mg, 0.625 mmol) in
dichloromethane (4 mL) were added 2-phenylacetaldehyde (72 mg,
0.6 mmol) and NaBH3CN (110 mg, 1.9 mmol), and the resulting
mixture was stirred at room temperature for 24 h. The organic phase
was washed with aqueous HCl (2 N) (3 × 5 mL), and product was
transferred into aqueous solution. The combined aqueous solution was
basified with NaHCO3 to pH around 8.0, and was extracted with ethyl
acetate (3 × 10 mL). After removal of solvent, the residue was
subjected to a silica gel column coated with C18H38 using a mixture of
methanol and water (4:6) as an eluent. Product 16 (84 mg, 54%) was
obtained. 16: oil; [α]25D −24.6 (c 1.4, CHCl3); IR (KBr) ν 3420, 1628,
1
1106, 1057, 1034 cm−1; H NMR (300 MHz/DMSO-d6, 363 K) δ
8.51 (br, s, 1 H), 7.32−7.17 (m, 9 H), 6.71 (br, s, 1 H), 5.09−4.91 (m,
3 H), 4.67 (br, s, 2 H), 4.20−4.16 (m, 1 H), 3.73−3.69 (m, 2 H), 2.80
(br, s, 2 H), 2.39−2.17 (m, 2 H), 2.01−1.76 (m, 2 H); 13C NMR (75
MHz/CDCl3) δ 175.62, 175.16, 172.03, 171.87, 171.83, 154.44,
153.80, 136.06, 135.84, 135.61, 134.75, 134.67, 133.71, 133.62, 132.68,
132.49, 131.59, 131.45, 128.96, 128.51, 128.38, 128.32, 128.30, 128.18,
128.13, 128.08, 127.95, 127.79, 127.75, 127.72, 127.3, 126.9, 126.8,
126.6, 126.0, 125.96, 67.9, 67.7, 67.6, 62.2, 61.98, 58.9, 58.7, 58.2, 47.1,
47.0, 45.0, 44.8, 43.3, 43.1, 40.8, 30.8, 30.7, 30.1, 29.4, 29.1, 28.7, 28.6,
28.4, 28.2; MS (ESI) m/z 408.5 (M + 1, 60), 430.5 (M + 23, 100).
Anal. Calcd for C23H26N3O4: 408.1923. Found: 408.1909.
Synthesis of 20. To a well-stirred solution of 19 (1.63 g, 4 mmol)
in dry DMF at 0 °C was added dropwise SOCl2 (1 mL). The reaction
mixture was allowed to stire at room temperature for another 1 h. The
reaction was quenched by adding water (20 mL) followed by
extraction with ethyl acetate (3 × 20 mL). The organic layers were
combined and washed with saturated aqueous NaHCO3 (3 × 4 mL).
After drying with anhydrous sodium sulfate and removal of solvent, the
residue was chromatographed on a silica gel column eluted with a
mixture of petroleum ether and ethyl acetate (1:1) to yield 20 (1.21 g,
78%): mp 175−176 °C; [α]25D −16.0 (c 1, CHCl3); IR (KBr) ν 1712,
1
1605 cm−1; H NMR (300 MHz/CDCl3) δ 7.32−7.19 (m, 5 H),
3.63−3.58 (m, 1 H), 3.39−3.35 (m, 1 H), 2.93−2.71 (m, 4 H), 2.64−
2.54 (m, 3 H), 2.35−2.32 (m, 1 H), 2.28 (s, 3 H), 1.88−1.72 (m, 3 H),
1.59−1.50 (m, 1 H); 13C NMR (75 MHz/CDCl3) δ 139.9, 128.7,
128.5, 126.2, 67.5, 66.6, 61.5, 53.2, 51.7, 39.7, 36.2; MS (ESI) m/z
249.5 (M + 1, 100). Anal. Calcd for C15H25N2O: 249.1967. Found:
249.1962.
1
1644 cm−1; H NMR (300 MHz/DMSO-d6, 375 K) δ 7.26−7.15 (m,
9 H), 5.07 (br, s, 2 H), 4.92 (dd, J = 7.5, 5.7 Hz, 1 H), 4.84 (dd, J =
7.4, 5.1 Hz, 1 H), 4.70 (d, J = 16.2 Hz, 1 H), 4.60 (d, J = 16.2 Hz, 1
H), 3.72−3.63 (m, 2 H), 2.80 (br, s, 2 H), 2.45−2.18 (m, 3 H), 1.98−
1.93 (m, 1 H); 13C NMR (75 MHz/CDCl3) δ 169.53, 169.48, 153.4,
153.3, 135.7, 135.6, 135.5, 135.0, 134.9, 133.9, 133.7, 133.2, 133.1,
Synthesis of 17. A mixture of 16 (120 mg, 0.48 mmol),
formaldehyde (35%, 1.2 mL), concentrated hydrochloric acid (2.4
mL), and chloroform (2.4 mL) was refluxed for 24 h. After being
cooled to room temperature, the mixture was basified with saturated
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131.9, 131.8, 129.1, 129.05, 128.6, 128.4, 128.3, 128.25, 128.17,
128.12, 127.97, 127.1, 126.8, 126.7, 126.5, 126.0, 125.9, 118.1, 117.7,
68.2, 67.9, 57.8, 57.5, 57.2, 48.2, 47.6, 47.2, 47.1, 44.7, 44.65, 43.3,
43.0, 40.4, 30.6, 30.5, 29.8, 29.6, 29.5, 29.2, 28.7, 28.6, 28.4, 28.3; MS
(ESI) m/z 390.4 (M + 1, 50), 412.5 (M + 23, 100). Anal. Calcd for
C23H23N3O3: C, 70.93; H, 5.95; N, 10.79. Found: C, 70.68; H, 5.90;
N, 10.78.
Keohane, C. A.; Milligan, J. A.; Rosenbach, M. J.; Shei, G.-J.; Mandala,
S. M. Bioorg. Med. Chem. Lett. 2006, 16, 2905.
(3) (a) Wright, W. G.; Warren, F. L. J. Chem. Soc. C 1967, 284.
(b) Kato, A.; Okada, M.; Hashimoto, Y. J. Nat. Prod. 1984, 47, 706.
(4) Kumagai, K.; Shono, K.; Nakayama, H.; Ohno, Y.; Saji, I. J.
Antibiot. 2000, 53, 467.
(5) Impellizzeri, G.; Mangiafico, S.; Oriente, G.; Piattelli, M.; Sciuto,
S.; fattorusso, E.; Magno, S.; Santacroce, C.; Sica, D. Phytochemistry
1975, 14, 1549.
Synthesis of 21. To a solution of 20 (155 mg, 0.4 mmol) in a
mixture of dichloromethane (5 mL) and methanol (10 mL) at −10 °C
was bubbled into dry HCl gas for 1 h, and the reaction mixture was
kept at −20 °C overnight. The mixture was added dropwise into
hydrochloric acid (6 N, 10 mL) at 0 °C, and then was neutralized with
saturated aqueous NaHCO3 solution to pH 7.0. After extraction with
ethyl acetate (3 × 20 mL), drying with anhydrous sodium sulfate, and
removal of solvent under vacuum, the residue was chromatographed
on a silica gel column eluted with a mixture of petroleum ether and
(6) For a recent example, see: Gartner, M.; Weihofen, R.; Helmchen,
̈
G. Chem.Eur. J. 2011, 17, 7605.
(7) (a) Whitesell, J. K. Chem. Rev. 1989, 89, 1581. (b) Meth-Cohn,
O.; Chen, Y. Tetrahderon Lett. 1999, 40, 6069. (c) Kim, B. H.; Lee, H.
B.; Hwang, J. K.; Kim, Y. G. Tetrahedron: Asymmetry 2005, 16, 1215.
(8) (a) Sweet, J. A.; Cavallari, J. M.; Price, W. A.; Ziller, J. W.;
McGrath, D. V. Tetrahedron: Asymmetry 1997, 16, 1215. (b) Chen, H.;
Sweet, J. A.; Lam, K.-C.; Rheingold, A. L.; McGrath, D. V.
Tetrahedron: Asymmetry 2009, 20, 1672.
ethyl acetate (1:2) to furnish product 21 (86 mg, 51%): oil; [α]25
D
1
−34.0 (c 1, CHCl3); IR (KBr) ν 1635, 1019 cm−1; H NMR (300
MHz/DMSO-d6, 375 K) δ 7.34−7.13 (m, 9 H), 5.01 (s, 2 H), 4.89−
4.84 (m, 1 H), 4.65 (s, 2 H), 4.46 (t, J = 6.72 Hz, 1 H), 3.73−3.57 (m,
2 H), 3.66 (s, 3 H), 2.95 (br, s, 1 H), 2.80−2.78 (m, 2 H), 2.30−2.12
(m, 3 H), 1.94−1.86 (m, 1 H); 13C NMR (75 MHz/MeOD) δ 172.1,
171.8, 171.0, 170.8, 170.6, 154.8, 154.7, 136.3, 136.0, 135.8, 134.8,
134.6, 134.4, 134.2, 132.9, 132.8, 132.7, 132.6, 128.4, 128.3, 128.2,
128.1, 128.0, 127.9, 127.7, 127.6, 127.5, 127.45, 127.3, 126.6, 126.3,
126.2, 126.1, 125.84, 125.79, 67.4, 67.1, 60.2, 59.9, 58.7, 58.3, 57.9,
51.3, 51.2, 44.4, 44.3, 43.1, 42.8, 40.5, 29.3. 29.0, 28.9, 28.8, 28.6, 28.5,
28.2, 27.8, 27.75; MS (ESI) m/z 423.5 (M + 1, 25), 445.5 (M + 23,
100). Anal. Calcd for C24H27N2O5: 423.1920. Found: 423.1910.
Synthesis of 22. A mixture of 22 (42 mg, 0.1 mmol) and LiAlH4
(60 mg, 1.58 mmol) in dry THF (4 mL) was refluxed for 24 h. After
the mixture was cooled to room temperature, aqueous NaOH solution
(2 N, 5 mL) was added, and the mixture was extracted with ethyl
acetate (3 × 10 mL). The combined organic layer was dried with
anhydrous sodium sulfate. After removal of solvent, the residue was
chromatographed on a C-18 reversed-phase column eluted with a
mixture of methanol and water (1:1) to afford product 22 (21 mg,
81%): [α]25D +54.7 (c 1.5, CHCl3). Compound 22 gives spectroscopic
data identical to those for 17.
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̈
ASSOCIATED CONTENT
■
S
* Supporting Information
1
Chiral HPLC analysis and H and 13C spectra of all products.
(18) Chen
Asymmetry 2008, 19, 1333.
̂ ̀
evert, B.; Jacques, F.; Giguere, P.; Dasser, M. Tetrahedron:
This material is available free of charge via the Internet at
http://pubs.acs.org.
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AUTHOR INFORMATION
Corresponding Author
*E-mail: wangmx@mail.tsinghua.edu.cn.
Notes
■
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The authors declare no competing financial interest.
ACKNOWLEDGMENTS
■
We thank the Ministry of Science and Technology
(2009CB724704), the National Science Foundation of China
(20820102034), the Chinese Academy of Sciences, and
Tsinghua University for financial support.
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