R. S. Topgi et al. / Bioorg. Med. Chem. 7 (1999) 2221±2229
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Deacylation of ethyl 3-(N-phenylacetyl)-5-trimethylsilyl-
4-pentynoate, 4, using Enzyme Penicillin G amidohy-
drolase. Phosphate buer (500 mL, 0.01 M prepared
from 0.1 M phosphate buer, pH 7.4) was added to
racemic ethyl 3-(N-phenylacetyl)-5-trimethylsilyl-4-pen-
tynoate, (4), (5 g, 15 mmol). The pH was 7.4. Immobi-
lized Penicillin G amidohydrolase (2 g, 345 units/g) was
added. The reaction mixture was stirred at room tem-
perature. During the reaction the pH of the reaction
medium was maintained around 7.4 using dilute
KHCO3 solution (1:1, saturated KHCO3:water) with an
auto titrator. After 24 h stirring, the reaction mixture
was extracted with ethyl acetate (3Â75 mL). The com-
bined organic extract was extracted with dilute hydro-
chloric acid (pH 2, 3Â75 mL). The hydrochloric acid
extracts were combined, and preserved. The organic
extract was washed with dilute potassium bicarbonate
(pH 8, 2Â25 mL), dried over sodium sulfate and eva-
porated to dryness under reduced pressure to obtain
(2.4 g, 96%) unreacted (S)-amide, 6a, as a white crys-
talline compound. M.p.: 74±76ꢁC. IR (MIR): 3243,
3059, 2963, 2174, 1742, 1641, 1550, 1495, 1312, 1275,
1250, 1152, 1041, 964, 898, 845, 648 cm 1; Sp. Rot.
(c=2 cm, methanol): [a]=d= 71.9ꢁ 1H NMR (400
MHz, CDCl3): d 0.12 (s, 9H, TMS); 1.18±1.24 (t, 3H,
CH3), 2.54±2.72 (m, 2H, CH2), 3.58 (m, 2H, CH2), 4.07±
4.15 (m, 2H, CH2), 5.08±5.15 (m, H, C-H), 6.21±6.28 (d,
broad, H, N-H), 7.26±7.39 (m, 5H, aromatic); 13C
NMR (500 MHz, CDCl3): d 0.25, 14.15, 38.54, 39.90,
43.59, 60.75, 88.07, 102.79, 127.30, 128.89, 129.31,
134.46, 169.65, 170.10; anal.: C18H25NO3Si requires C:
65.22, H: 7.60, N: 4.23. Found C: 64.87, H: 7.80, N:
4.25%.
units/g, Boehringer±Mannheim) was added. The reac-
tion mixture was stirred at room temperature. During
the reaction the pH of the reaction medium was main-
tained around 7.5 using dilute KHCO3 solution (1:1,
saturated KHCO3:water) with an auto titrator. After 24
h stirring, the reaction mixture was extracted with ethyl
acetate (3Â75 mL). The organic extracts were combined
and extracted with dilute hydrochloric acid (pH 2.0,
3Â75 mL). The hydrochloric acid extracts were com-
bined and preserved. The organic extract was washed
with dilute potassium bicarbonate (pH 8, 2Â25 mL),
dried over sodium sulfate and evaporated to dryness
under reduced pressure to obtain unreacted (S) amide,
(6b), (0.92 g, 95%) as a white crystalline solid. IR (MIR):
3279, 3205, 1733, 1641, 1530, 1413, 1351, 1317, 1180,
1114, 1073, 1030, 954, 763, 699 cm 1; Sp. Rot. (c=2 cm,
methanol): [a]d= 41.4ꢁ 1H NMR (400 MHz, CDCl3): d
1.20±1.25 (t, 3H, CH3), 2.22±2.25 (d, H, acetylenic),
2.58±2.75 (m, 2H, CH2), 3.8 (s, 2H, CH2), 4.08±4.17 (q,
2H, CH2), 5.05±5.13 (m, H, C-H), 6.26±6.36 (d broad, H,
N-H), 7.22±7.39 (m, 5H, aromatic); 13C NMR (500
MHz, CDCl3): d 14.10, 37.68, 39.42, 43.56, 60.89, 71.36,
81.34, 127, 37, 128.96, 129.33, 134.35, 169.83, 170.11; MS
m/z: 168, 190, 198, 200, 214, 221, 234, 242, 246, 261
(M+1, 100); anal.: C15H17NO3 requires C: 69.48, H: 6.61,
N: 5.40. Found C: 69.30, H: 6.52, N: 5.34%.
The acidic aqueous phase containing the amine was
basi®ed using dilute potassium bicarbonate solution to
attain pH ꢀ7.8 and extracted with ethyl acetate (3Â75
mL). The ethyl acetate extracts were combined, dried
over sodium sulfate, and evaporated to dryness under
reduced pressure to obtain (R)-amine, (7b), (0.49 g,
92%). The small quantity of (R)-amine was derivatized
using 1-naphthoyl chloride. 1-Naphthoyl (R)-amide is a
white crystalline compound. IR (MIR): 3288, 1734,
1636, 1522, 1430, 1376, 1288, 1187, 102ꢁ5, 783 cm 1; Sp.
Rot. (c=2 cm, methanol): [a]d= 5.6 1H NMR (400
MHz, CDCl3): d 1.26±1.32 (t, 3H, CH3), 2.37 (d, H,
acetylenic), 2.84±2.97 (m, 2H, CH2), 4.17±4.26 (m, 2H,
CH2), 5.40±5.47 (m, H, C-H), 6.92±6.98 (d, broad, H,
N-H), 7.44±8.36 (m, 7H, aromatic); 13C NMR (500
MHz, CDCl3): d 14.16, 38.04, 39.53, 61.06, 71.69,
124.65, 125.26, 126.45, 127, 22, 128.31, 131.05, 133.29,
133.67, 168.25, 170.44: Anal.: C18H17NO3 requires C:
73.20, H: 5.80, N: 4.74. Found C: 69.97, H: 5.73, N:
4.84%.
The acidic aqueous phase containing the amine was
basi®ed using dilute potassium bicarbonate solution to
attain pH ꢀ7.8 and extracted with ethyl acetate (3Â75
mL). The ethyl acetate extracts were combined, dried
over sodium sulfate and evaporated to dryness under
reduced pressure to obtain (1.45 g, 91%) (R)-amine, 7a.
The small quantity of (R)-amine was derivatized using
1-naphthoyl chloride. 1-Naphthoyl (R)-amide is a white
crystalline compound. IR (MIR): 3273, 2958, 2175,
1734, 1642, 1513, 1371, 1293, 1248, 1093, 1026, 839,
778, 759, 699 cm 1; Sp. Rot. (c=2 cm, methanol):
[a]d=+43.3ꢁ 1H NMR (400 MHz, CDCl3): d 0.19 (s,
9H, TMS), 1.26±1.33 (t, 3H, CH3), 2.80±2.95 (m, 2H,
CH2), 4.16±4.28 (m, 2H, CH2), 5.40±5.48 (m, H, C-H),
6.81±6.88 (d broad, H, N-H), 7.44±8.38 (m, 7H, aro-
matic); 13C NMR (500 MHz, CDCl3): d 0.22, 14.23,
39.06, 40.07, 60.91, 88.51, 102.81, 124.69, 125.22,
125.32, 126.43, 127, 15, 128.30, 130.16, 130.93, 133.56,
133.70, 168.15, 170.37; MS m/z: 190, 204, 224, 268, 282,
296, 340, 368 (M+1, 100). Anal.: C21H25NO3Si requires
C: 68.63, H: 6.86, N: 3.81. Found C: 68.38, H: 6.82, N:
3.71%.
Acylation of ethyl 3-amino-5-(trimethylsilyl)-4-pentynoate,
3, using Enzyme Penicillin G amidohydrolase. Crude
free amine, 3, (racemic, 500 mg) was added drop by
drop to dilute HCl acid (50 mL, pH 1.9) with stirring.
After 10 min, the insoluble material was extracted using
t-butyl methyl ether (25 mL). The layers were separated,
and the pH of the aqueous phase was adjusted to 6
using ammonium hydroxide (2.5 N). Phenylacetic acid
(2 equiv) was added and the mixture was stirred for
about 10 min. The pH was adjusted to 6 and the mix-
ture was stirred for about 10 min. Soluble enzyme (250
units, Boehringer±Mannheim) was added, and the mix-
ture was stirred at room temperature for 42 h. The pH
of the reaction mixture was adjusted to 4 and stirring
was continued for 10 min. The product was collected by
Deacylation of ethyl 3-(N-phenylacetyl)-4-pentynoate, 5,
using Enzyme Penicillin G amidohydrolase. Phosphate
buer (250 mL, 0.01 M prepared from 0.1 M phosphate
buer, pH 7.4) was added to racemic ethyl 3-(N-phenyl-
acetyl)-4-pentynoate (1.94 g, 7.5 mmol). The pH was
7.57. Immobilized Penicillin G amidohydrolase (2 g, 345