LETTER
Markovnikov Addition of Azoles to Vinyl Esters in Organic Solvents
2435
Table 1 D-Aminoacylase-Catalyzed Markovnikov Addition of
In conclusion, we have developed a facile biotransforma-
tion path to perform Markovnikov additions between
azoles and vinyl esters by D-aminoacylase catalysis. Sol-
vents play an essential role in the enzymatic addition re-
action. The result of addition was affected by the structure
of substrate and vinyl ester. By this novel strategy, a num-
ber of pharmacologically active azole derivatives were
successfully synthesized in moderate to excellent yields.
Azoles to Vinyl Estersa
N
N
N
N
N
H
N
N
H
H
1a
1c
1b
Entry
Azole
1a
Vinyl Ester Product Time (h) Yieldb (%)
1
2
3
4
5
6
7
8
9
2a
2b
2c
2d
2e
2f
3a
3b
3c
3d
3e
3f
84
96
85
82
75
72
91
26
93
90
76
1a
D-Aminoacylase-Catalyzed Markovnikov Addition of Azoles to
Vinyl Esters; General Procedure
1a
96
A suspension of substrate (0.6 mmol) and D-aminoacylase (100 mg)
from Escherichia coli (EC 3.5.1.81, purchased from Amano En-
zyme Inc.) in hexane (2 mL) was incubated at 50 °C at 200 r.p.m.
(orbitally shaken) for 5 min. Then, vinyl ester (8 equiv) was added
in order to initiate the reaction. After the time indicated (Table 1),
the reaction was terminated by filtering off the enzyme. The crude
product was purified by chromatography on silica gel (petroleum
ether–EtOAc).
1a
96
1a
96
1a
192
84
1b
1b
1c
2a
2e
2a
3g
3h
3i
96
References
94
a Substrate (0.6 mmol), vinyl ester (8 equiv), D-aminoacylase (100
mg), hexane (2 mL), 50 °C.
(1) (a) Rogers, R. S. Chem. Eng. News 1999, 77 (29, July 19),
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3194. (c) Faber, K. Biotransformations in Organic
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b Isolated yields.
were run with covalently inhibited D-aminoacylase by
adding 50 mM of the noncompetitive inhibitor ZnCl2,11
the inhibited enzyme did not show any acylase activity to
catalyze the hydrolysis of N-acetyl-D-methionine, and the
specific activity for the Markovnikov addition was that of
the non-enzymatic reaction. These results suggest that the
tertiary structure and the specific active site of D-amino-
acylase are responsible for the Markovnikov addition
reaction.
The generally accepted acylase mechanism of D-amino-
acylase usually involves the polarization of a carbonyl
group by the binding of a zinc ion followed by proton
transfer from water to a leaving group mediated by Asp.12
Herein, we propose a tentative mechanism for the enzy-
matic Markovnikov addition. The tightly bound zinc ion
first interacts with the carbonyl group of the vinyl ester
and draws electron density away from the carbon; owing
to the electron-withdrawing effect of the carboxyl group,
the a-carbon of the vinyl group carries a partial positive
charge. When the substrate enters the active site, the Asp
functions as a general base, removing the N-proton, while
the nucleophile simultaneously adds to the C-b position.
The resulting negative charge at C-b could be stabilized
by the zinc ion. Finally, the Asp, now functioning as a
general acid, would deliver the proton to C-b to complete
the reaction. Preliminary experiments confirmed that D-
aminoacylase was not able to catalyze any reaction be-
tween azoles and vinyl ethers. This indicated that the car-
boxyl group of vinyl ester played an extremely significant
role. Further study of the catalytic mechanism is in
progress in our laboratory.
(6) Branebby, C.; Carlqvist, P.; Magnusson, A.; Hult, K.;
Brinck, T.; Berglund, P. J. Am. Chem. Soc. 2003, 125, 874.
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Matsoyan, S. G. Zh. Org. Khim. 1988, 24, 1339.
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A. Chem. Heterocycl. Compd. (Engl. Transl.) 2002, 38, 981.
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Commun. 2005, 2348.
Synlett 2005, No. 16, 2433–2436 © Thieme Stuttgart · New York