552
N.W.Fadnavis et al./ Tetrahedron: Asymmetry 15 (2004) 549–553
2
D
5
corresponding to (R)-configuration with ½aꢀ ¼ )17.1
residual dark yellow oil was extracted with diisopropyl
ether at room temperature. The combined extracts on
removal of solvent gave a pale yellow viscous oil (129 g,
92%). CAUTION: It was important to maintain the
temperatures below 40 ꢁC to avoid formation of poly-
meric materials during solvent evaporation. The crude
material so obtained was sufficiently pure for use in the
enzymatic resolution. However, a small portion was
purified by column chromatography using hexane–ethyl
acetate (98:2) as eluent (r.f. 0.64) for analytical purpose.
IR (neat): 3484, 3061, 3029, 2933, 2361, 1753, 1654,
1579, 1495, 1451, 1372, 1293, 1216, 1122, 1071, 1021,
16
25
D
(
c 2.0, CHCl
3
);lit ½aꢀ ¼ )67.7 (c 1.03, CHCl
3
for the
corresponding methyl ester). Although the specific
rotation of the methyl and ethyl esters cannot be com-
pared directly, the negative sign of rotation for 7
obtained from 2 suggests the (R)-configuration of 2.
3. Conclusion
In conclusion, present work describes the preparation of
enatiomerically pure cyanohydrin esters 2 and 3 using
lipases of opposing stereoselectivities immobilized in
gelatin matrix. Conditions for analysis by chiral HPLC
have been established to determine the enantiomeric
purity of the products without derivatization. Although
preparation of hydroxyester 7 and its antipode via
enantiospecific reduction of the corresponding a-keto
ꢁ
1
1
968, 922, 837, 750, 694, 601, 559, 505 cm . H NMR
(CDCl , 200 MHz): d 2.2 (s, 3H), 5.99 (d, 1H
J ¼ 6:8 Hz), 6.16 (dd, 1H, J ¼ 6:4 Hz, J ¼ 15:9 Hz),
6.95 (d, 1H, J ¼ 14:7 Hz), 7.25–7.45 (m, 5H). C NMR
(CDCl , 200 MHz): d 41.2, 76.12, 94.88, 98.17, 107.12
3
1
2
13
3
(2C), 108.55, 109.15 (2C), 114.22, 117.52, 148.25. Anal.
15;20
acid has been well demonstrated,
the necessity of
calcd for C H NO : C, 71.63;H, 5.51;N, 6.96;found
2
12
11
growing the appropriate cultures in an organic chemistÕs
laboratory is a handicap. Use of commercially available
lipases solves these problems. The recycling of the
lipases immobilized in gelatine matrix for several recy-
cles has already been demonstrated and thus our
procedure can be used for large scale preparations.
Although our first experiments with conversion of
enantiomerically pure 2–7 without racemization have
not been completely successful, efforts are being made to
develop an appropriate methodology.
C, 71.21;H, 5.48;N, 6.52.
4.2. Enzymatic preparation of (R)-2-acetoxy-4-phenyl-
(E)-but-3-enenitrile 2
12
(RS)-1 (20.1 g, 0.1 mol) in diisopropyl ether (200 mL)
and 1-butanol (14.8 g, 0.2 mol) were stirred magnetically
with Amano Ps gelozyme (20 g, 3.6 g lipase) at room
temperature in a round-bottom flask. The progress of
the reaction was monitored by chiral HPLC. The reac-
tion was continued till all the (R)-ester had reacted (6 h,
50% conversion). The reaction mixture was then de-
canted, the filtrate was stirred with 1 M imidazole
solution (50 mL, pH 8.5) for 20 min to decompose the
cyanohydrin to cinnamaldehyde. The reaction mixture
was then stirred with 30% sodium bisulfite solution for
30 min. After separating the bisulfite adduct, the organic
layer was washed with water, phosphate buffer (0.1 M,
pH 8.2) and brine. After evaporation of the solvent
under high vacuum at room temperature, the residue
obtained (9.2 g, 92% of theoretical) consisted of >95%
(R)-2 (HPLC analysis). A small portion was purified by
column chromatography using hexane–ethyl acetate
4. Experimental
Lipase from C.rugosa , porcine pancreas and M.miehei
were obtained from Sigma, USA. Lipase Ps was
obtained from Amano Pharmaceutical Corporation
Ltd. Japan. All other reagents were A.R. grade obtained
from SD Fine Chem, India. HPLC analyses were carried
out on Hewlett Packard HP1090 unit with diode array
detector and HP Chem station. The enzymes were
12;14
immobilized as described earlier.
(
A loading of 18%
w/w) was obtained in the gelatin matrix for both the
enzymes. Almond meal was prepared from freshly
ground almonds, defatted with cold hexane and used as
2
5
(98:2) as eluent (r.f. 0.64). ½aꢀ ¼ )22.0 (c 2.0, chloro-
D
22
described in literature.
form), ee >99%.
4.1. Preparation of (RS)-2-acetoxy-4-phenyl-(E)-but-3-
enenitrile 1
4.3. Enzymatic preparation of (S)-2-acetoxy-4-phenyl-
(E)-but-3-enenitrile 3
Sodium cyanide (49 g, 1 mol) and tetrabutyl ammonium
chloride (5.55 g, 20 mmol) were dissolved in water
(RS)-1 (2.01 g, 0.01 mol) in hexane (100 mL) and
1-butanol (1.48 g, 0.02 mol) were stirred magnetically
with C.rugosa gelozyme (10 g, 1.8 g lipase) at room
temperature in a round-bottom flask. The progress of
the reaction was monitored by chiral HPLC. After every
8 h, the reaction mixture was decanted from the immo-
bilized enzyme and vigorously stirred with sodium
bisulfite solution (30%, 50 mL) for 10 min. The bisulfite
adduct was filtered off, the organic layer was washed
with sodium phosphate buffer (3 · 20 mL, 0.1 M, pH
8.2), dried over anhydrous magnesium sulphate and
again subjected to enzymatic reaction after addition of
1 mL 1-butanol to compensate for its loss during aque-
(
600 mL) and the solution was cooled in ice with stirring.
A solution of cinnamaldehyde (92.5 g 0.7 mol) in
dichloroethane (500 mL) was added and the reaction
mixture was stirred for 4 h at 4–5 ꢁC. Acetic anhydride
(
71 g, 0.7 mol) in dichloroethane (200 mL) was added
dropwise with stirring over 2 h. The reaction mixture
was stirred further for 4 h in cold and then allowed to
reach room temperature. The layers were then separated
and the organic layer was washed with sodium carbon-
ate, water and brine;dried over anhydrous magnesium
sulfate and the solvent was removed under vacuum. The
Fadnavis, Nitin W.
