2428
P. Soni et al. / Tetrahedron: Asymmetry 16 (2005) 2425–2428
Chemical shifts were reported in parts per million (ppm,
d) using TMS as an internal standard. IR spectra were
recorded on a Nicolet spectrometer. Optical rotations
were measured on a Rudolph polarimeter (Rudolph
Research Autopol IV).
(760 mg). This crude reaction mixture was analyzed by
HPLC and GC–MS to determine conversion and enan-
tiomeric excess. Chromatography (silica, CH3OH 4% in
CHCl3 as eluent) of this oily mass afforded the chiral
alcohol (S)-2b (685 mg, isolated yield 85.6%) with
>99.9% enantiomeric excess determined by GC–MS on
20
D
4.2. GC–MS method
a chiral column. ½aꢃ ¼ ꢀ38.4 (c 1.0, EtOH) {lit.
24
½aꢃ ¼ ꢀ38.9 (c 1.02, EtOH)}.18 MS (ApcI): m/z = 124
D
To determine the enantiomeric excess of (S)-2b, a b-col-
umn (30 m · 0.25 mm, 0.25 lm, Sigma–Aldrich, USA)
with a chiral stationary phase of polysiloxine was used.
The temperature program used was 100 ꢁC/2 min—5 ꢁC/
min—200 ꢁC/5 min. Retention time for (R)-2b was
15.9 min and for (S)-2b was 16.1 min and was ascer-
tained on the basis of optical rotation of (S)-2b obtained
by bioreduction of 1b.
(M+1)+; 1H NMR (300 MHz, CDCl3): d = 1.46 (d,
3H), 4.91 (q, 4H), 5.39 (br s, 1H), 7.22 (dd, 1H), 7.73
(d, 1H, 7.97), 8.33 (d, 1H, 4.82), 8.41 (s, 1H); 13C
NMR (300 MHz, MeOD): d = 25.04, 67.14, 76.61,
77.03, 77.45, 123.45, 133.59, 141.92, 146.69, 147.55; IR
(neat): m = 3351.2 cmꢀ1
.
Acknowledgements
4.3. Synthesis of reference compounds (rac-alcohols)
One of the authors (P.S.) gratefully acknowledges Coun-
cil of Scientific and Industrial Research, India for pro-
viding senior research fellowship.
The racemic alcohols 2a–f were synthesized by reduction
of the corresponding ketones 1a–f with NaBH4 follow-
ing standard procedure.18 On each occasion, the product
1
was characterized by IR, H and 13C NMR, and GC–
MS. The two enantiomers exhibited distinctly different
retention times on chiral GC (for 2a–c) and chiral
HPLC (for 2d–f).
References
1. Uskokovic, M. R.; Lewis, R. L.; Partridge, J. J.;
Despreaux, C. W. J. Am. Chem. Soc. 1979, 101, 6742–
6743.
2. Kaneoya, M.; Yoshida, N.; Uchida, M. USPTO 4971909.
3. Quallich, G. J.; Woodall, T. M. Tetrahedron Lett. 1993,
34, 4145–4158.
4.4. General procedure for the microbial reduction of
heteroaryl ketones
The microorganism, C. viswanathii, was isolated by
enrichment and isolation techniques, using acetophe-
none as the sole source of carbon and energy.20 A ster-
ilized nutrient broth (100 mL) was inoculated with 5%
(v/v) of the preculture (24 h old). The mixture was incu-
bated in an orbital shaker (200 rpm) for 48 h at 30 ꢁC.
The cells were harvested by centrifugation (7000g,
20 min, 4 ꢁC) and washed thoroughly with phosphate
buffer (0.2 M, pH 7.0) and finally resuspended in the
same buffer. To the cell suspension (12 mL), an appro-
priate prochiral ketone (25 mg) in ethanol was added.
After 12 h of further incubation, an aliquot of the reac-
tion mixture was centrifuged to separate out the cells.
The supernatant as well as the cells was extracted twice
with ethyl acetate. The combined ethyl acetate extracts
were dried over Na2SO4, and evaporated under reduced
pressure. The crude product was then analyzed by
HPLC.
4. Collomb, P.; Zelewsky, A. Tetrahedron: Asymmetry 1998,
9, 3911–3917.
5. Okano, K.; Murata, K.; Ikariya, T. Tetrahedron Lett.
2000, 41, 9277–9280.
6. Brown, E.; Penfornis, A.; Bayama, J.; Touet, J. Tetrahe-
dron: Asymmetry 1991, 2, 339–342.
7. Kaneoya, M.; Naoyuki, Y.; Manabu, U. USPTO 4971909.
8. Kaminska, J.; Gornicka, I.; Sikora, M.; Gora, J. Tetra-
hedron: Asymmetry 1996, 7, 907–910.
9. Orrenius, C.; Mattson, A.; Norin, T.; Ohrner, N.; Hult, K.
Tetrahedron: Asymmetry 1994, 5, 1363–1366.
10. Garrett, M. D.; Scott, R. S.; Sheldrake, G. N. Tetrahe-
dron: Asymmetry 2002, 13, 2201–2204.
11. Nakamura, K.; Takenaka, K.; Fujii, M.; Ida, Y. Tetra-
hedron Lett. 2002, 43, 3629–3631.
12. Bradshaw, C. W.; Hummel, W.; Wong, C. H. J. Org.
Chem. 1992, 57, 1532–1536.
13. Uskokovic, M. R.; Lewis, R. L.; Partridge, J. J.;
Despreaux, D. L. J. Am. Chem. Soc. 1979, 101, 6742–6744.
14. Kawano, S.; Horikawa, N.; Yasohara, Y.; Hasegawa, J.
Biosci. Biotechnol. Biochem. 2003, 67, 809–814.
15. Takemoto, M.; Achiwa, K. Tetrahedron: Asymmetry 1995,
6, 2925–2928.
16. Soni, P.; Kaul, C. L.; Banerjee, U. C. Indian Patent Appl.
No. 1573/DEL/2004.
17. Soni, P.; Banerjee, U. C. Appl. Microbiol. Biotechnol.
2005, 67, 771–777.
18. Stampfer, W.; Edegger, K.; Kosjek, B.; Faber, K.; Kroutil,
W. Adv. Synth. Catal. 2004, 346, 57–62.
19. Soni, P.; Kamble, A. L.; Banerjee U. C. Indian Patent
Appl. No. 440/DEL/2005.
20. Kamble, A. L.; Soni, P.; Banerjee, U. C. J. Mol. Catal. B:
Enzym. 2005, 35, 1–6.
4.5. Preparative scale bioreduction
The reaction was repeated on a preparative scale by add-
ing 0.8 g of 1b (as a 100 mg/mL solution in EtOH) to
400 mL of the resting cell suspension (150 g/L) of the
yeast cells. The enzymatic reaction was allowed to pro-
ceed for 12 h (at 30 ꢁC), after which the reaction mixture
was centrifuged to separate out the cells. The superna-
tant and the cells were extracted three times using ethyl
acetate as the solvent, which was evaporated under re-
duced pressure. The organic solvent was dried over
anhydrous sodium sulfate to obtain an oily mass