Enantioselective hydrolysis of various substituted styrene oxides with Aspergillus Niger CGMCC 0496

Article abstract of DOI:10.1039/b312469j

Enantioselective biohydrolysis of various substituted styrene oxides using whole fungus cells of Aspergillus niger CGMCC 0496 are described. The results show not only para- but also some ortho- substituted styrene oxides can achieve high enantioselectivity during the hydrolysis.

Full text of DOI:10.1039/b312469j

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Enantioselective hydrolysis of various substituted styrene oxides
with Aspergillus Niger CGMCC 0496†
Hao Jin, Zu-yi Li* and Xiao-Wei Dong
State Key Laboratory of Bioorganic & Natural Products Chemistry,
Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, 354 Fenglin Lu,
Shanghai 200032, China. E-mail: lizy@mail.sioc.ac.cn; Fax: 86-21-64166128;
Tel: 86-21-64163300-1406
Received 7th October 2003, Accepted 17th November 2003
First published as an Advance Article on the web 12th January 2004
Enantioselective biohydrolysis of various substituted styrene oxides using whole fungus cells of Aspergillus niger
CGMCC 0496 are described. The results show not only para- but also some ortho- substituted styrene oxides can
achieve high enantioselectivity during the hydrolysis.
hydrolysis of the epoxide by selecting appropriate substrates for
Introduction
the study of the enantioselectivity of this strain. During the
research, the substituted styrene oxides, especially nitrostyrene
oxides, were found to be stable and resistant to a non-enzymatic
spontaneous degradation. In order to get reliable results on the
enantioselectivity and specificity of the strain, p-nitrostyrene
oxides were used as the model substrates.
Temperature is an important factor in a biocatalytic system.
It has been reported that the temperature greatly affected not
only the performance of the reaction but also the enantio-
selectivity of the kinetic resolution.11 As a result, in this case,
the most favorable temperature for the biocatalytic hydrolysis
reaction was determined first. (Table 1)
From Table 1 it clearly appears that the best temperature for
the whole cell biocatalytic system was 25 ЊC. After 109 min,
(S)-epoxide was recovered in 42% yield with 94% ee and the
corresponding (R)-diol was obtained in 45% yield with 95% ee.
When the reaction was carried out at 30 ЊC and the product diol
reached 93% ee, the residue epoxide was only recovered with
78% ee. When the reaction was carried out at 20 ЊC, no observ-
able hydrolysis could be found even with a prolonged reaction
time.
The stereochemical pathways for the biohydrolysis of 1,2-
epoxides using epoxide hydrolases are complex.12 Two enantio-
mers of diols may be found when hydrolyzing epoxides using
a single enantiomer. When the enzyme attacks the terminal
carbon of the epoxide, the configuration of the product can be
retained. Otherwise, the enzyme attacks the secondary carbon
of the epoxide, via a S2 mechanism and the configuration of
the product is reversed.
Enantiomerically pure forms of substituted phenylethanediols
and substituted styrene oxides are important intermediates for
the synthesis of various biologically active molecules. Recently,
a number of chemical and biochemical methods have been
developed to obtain these optically pure building blocks.1 One
of the most useful bioapproaches is enantioselective hydrolysis
of racemic epoxides by epoxide hydrolases.
Epoxide hydrolases (EHs) are very important enzymes which
have been detected in many organisms such as mammals,2
plants,3 insects,4 and microorganisms.5 Because of the wide-
spread distribution in metabolism of various xenobiotics,
mammalian epoxide hydrolases have been extensively studied
during the past two decades.6 However, due to their low avail-
ability, the potential of mammalian enzymes as chiral catalysts
is seriously limited in the light of the preparative scale.
Recently detailed searches for EHs from microbial sources
have been undertaken by several groups. A lot of microbial
origin EHs have been identified,7 and some show extremely
good enantioselectivity when hydrolyzing para-substituted
styrene oxides.8 However, there have been few results on the
substrates of ortho-9 and meta-substituted styrene oxides. More
recently, we screened EHs among several strains of Aspergillus
niger which were isolated from soil. Several of them were found
to have extremely highly active and enantioselective EHs,10
especially A. niger CGMCC 0496.9
In this paper, we investigated the biocatalytic hydrolysis of
various substituted styrene oxides using whole cells of A. niger
CGMCC 0496.
In order to obtain detailed information about the bio-
process, (S)-p-nitrostyrene oxide (>99% ee) and (R)-p-nitro-
styrene oxide (>98% ee) were selected as microbial substrates
for hydrolysis. (S)-p-Nitrostyrene oxide was obtained by bio-
hydrolysis of its racemic form. (R)-p-Nitrostyrene oxide was
prepared from its corresponding (R)-diol (Scheme 1). The
(R)-diol was also obtained by hydrolysis and could easily be
recrystallized from CHCl3 leading to enantiopure material.
Results and discussion
As we know, most epoxides are unstable in acidic and basic
conditions. Hence it was important to minimize the chemical
† Electronic supplementary information (ESI) available: experimental
details. See http://www.rsc.org/suppdata/ob/b3/b312469j/
Table 1 Biohydrolysis of p-nitrostyrene oxide by A. niger CGMCC 0496a under different temperatures
Reaction
time/min
Yield of
epoxide (%)
Ee of the epoxide (%)
Ee of the diol (%)
Temperature/ЊC
(abs. config.b)
Yield of diol (%)
(abs. config.b)
30
25
20
83
109
120
45
42
78 (S)
94 (S)
39
45
93 (R)
95 (R)
Biohydrolysis was not detected by TLC
a The reactions were performed with 10 g cells : 200 mg substrate. b The absolute configurations were confirmed by comparing the optical rotations
reported in the literature.8
O r g . B i o m o l . C h e m . , 2 0 0 4 , 2, 4 0 8 – 4 1 4
T h i s j o u r n a l i s © T h e R o y a l S o c i e t y o f C h e m i s t r y 2 0 0 4
408

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Table 2 Biohydrolysis of (S)- and (R)-p-nitrostyrene oxide with A. niger CGMCC 0496a
Substrate
Reaction time/min
Yield of diol (%)
Ee of diol (%) (abs. config.b)
(S)-1
(R)-1
720
32
8
>95
89(S)
97(R)
a The reactions were carried out at 25 ЊC with 10 g cells : 0.1 g substrate. b The absolute configurations were established by comparisons with the
elution order of the standard sample, (S)-p-nitrostyrene oxide, on a Chiralpak AD column.
Table 3 Different rate constants for the production of the (S)- and (R)-diols from the corresponding chiral epoxides by A. niger CGMCC 0496
Substrate
Rate constants of production (S)-diol k/minϪ1
Rate constants of production (R)-diol k/minϪ1
(R)-1
(S)-1
>1.4 × 10Ϫ3
1.09 × 10Ϫ4
>9.22 × 10Ϫ2
6.4 × 10Ϫ5
Table 4 Biohydrolysis of various substituted nitrostyrene oxides by A. niger CGMCC 0496
Substitution of
nitro group
Reaction
time/min
Yield of
Ee of epoxide
(%) (abs. conf.)
Yield of
diolc
Ee of diol
Entry
epoxidec (%)
(%) (abs. conf.)d
1
2
3
4
5
6
7
p-a
p-b
m-a
m-b
o-a
o-b
o-f
109
23
140
39
170
68
68
42
47
45
40
52
34
42
94(S)d
97(S)
55(S)e
57(S)
58(S)e
98(S)
84(S)
45
46
48
33
42
38
—
95(R)
93(R)
66(R)
79(R)
96(R)
>99(R)
—
a 200 mg substrate with 10 g cells. b 100 mg substrate with 20 g cells. c Isolated yield. d By comparing the optical rotation reported in the literature.8,13
e The absolute configuration was obtained by checking the optical rotation of the epoxides which were prepared from corresponding optical diols
with a chemical correlation method.14 f Same conditions as the entry 6, except adding 7 mg of (R)-diol.9
Scheme 1 Preparation of (R)-p-nitrostyrene oxide from (R)-(4-nitro-
phenyl)-1,2-ethanediol.
Scheme 2 Biohydrolysis process of substituted styrene oxide with EHs
The biohydrolysis of (S)- and (R)-p-nitrostyrene oxide
were carried out respectively under the same conditions
(Table 2).
in A. niger strain.
In order to obtain further reaction data on the effect of the
substitution pattern, various nitrostyrene oxides were investi-
gated at 25 ЊC (Table 4).
No hydrolysis of (S)-p-nitrostyrene oxide could be detected
by TLC after 60 min. Even after an additional 660 min,
the yield of the product diol only reached 8.0% with the
(S)-configuration in 89% ee. This was not the result of chemical
hydrolysis because only a yield of 1.2% of the diol was found by
HPLC in a blank test after 720 min in the same buffer. When
(R)-p-nitrostyrene oxide was tested, the rate of the reaction was
much faster than that of the (S)-form. No residue epoxide
could be detected by TLC after 32 min. The (R)-configuration
product was obtained with a 97% ee.
Through the methods of Michaelis–Menten we checked the
rate constants of the forming (S)- and (R)-diols from the corre-
sponding chiral epoxides (Table 3). The results showed that the
stereochemical pathway of the biohydrolysis of the EHs is more
complex.
From the results in Table 3, we found the following profiles of
the bioprocess: (1) The rate constants for the production of
(R)-diols from the corresponding (R)-epoxides were approx-
imately 1.44 × 103 times faster than the one which formed
from (S)-epoxide. (2) The rate constants for the production
of (S)-diols from the corresponding (R)-epoxides were
approximately 12 times than that formed from (S)-epoxide. It
was suggested that the EHs in A. niger favorably attack the
(R)-configuration enantiomer, especially at the first carbon
atom of the 1,2-epoxide. (Scheme 2)
Excellent results were found when the strain hydrolyzed p- and
o-substituted nitrostyrene oxides. The high ratio of cells to
substrate reduced the time of the complete resolutions. When
o-nitrostyrene oxide was used as the substrate, a high ratio of cells
to substrate was required for complete resolution. We thought it
probably due to the product inhibition. In order to get further
evidence to support our surmise, a carefully designed reaction
was carried out with the same conditions as entry 6 except adding
7 mg of (R)-diol to the substrate epoxide. The residue epoxide
was recovered in 42% yield and 84% ee when the reaction was
stopped within 68 min. This confirmed our hypothesis.9
After investigating the substitute site specificities of the
strain, a range of substituted styrene oxides were investigated in
order to broaden the applicability of this fungus (Scheme 3 and
Table 5).
The strain was found to have high affinity for a wide range of
substituted styrene oxides. The enantioselectivity of the reac-
tions strongly depended on the position of the substituent
group. Remarkable results were obtained in most cases when
hydrolyzing para- and ortho-substituted styrene oxides. Product
inhibitions were also found during the hydrolysis of other
ortho-substituted substrates, such as ( )-7, when using the low
ratio of cells to substrate. To our surprise, unexpectedly low
O r g . B i o m o l . C h e m . , 2 0 0 4 , 2, 4 0 8 – 4 1 4
409

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Table 5 Different rate constants for the production of (S)- and (R)- diols from the corresponding chiral epoxides by A. niger CGMCC 0496
Residue epoxide
Product of diol
Substratea
Entry
Time/min
Entryb
Yield (%)
Ee (%)
Entryc
Yield (%)
Ee (%)
( )-1
109
140
68
32
60
24
45
44
40
38
65
52
43
41
215
126
(S)-1
(S)-2
(S)-3
(S)-4
(S)-5
(S)-6
(S)-7
(S)-7
(S)-8
(S)-9
(S)-10
(S)-11
(S)-12
(S)-13
(S)-14
(S)-15
42
45
34
24
32
47
17
22
30
43
22
25
45
29
15
34
94
55
98
41
95
28
47
94
(R)-16
(R)-17
(R)-18
(R)-19
(R)-20
(R)-21
(R)-22
(R)-22
(R)-23
(R)-24
(R)-25
(R)-26
(R)-27
(R)-28
(R)-29
(R)-30
45
48
38
33
46
40
37
30
43
43
46
39
43
41
36
31
95
66
>99
( )-2
( )-3*
( )-4
74
85
74
84
84
86
70
80
72
63
70
52
82
( )-5
( )-6
( )-7
( )-7*
( )-8
>99
( )-9
35
>99
97
70
63
75
98
( )-10
( )-11
( )-12
( )-13
( )-14
( )-15*
a The reactions were carried out at 25 ЊC with 200 mg substrate and 10 g cells except *, which were carried out with 100 mg substrate and 20 g cells.
b The absolute configurations of (S)-4, (S)-5, (S)-6, (S)-7, (S)-8, (S)-12, (S)-14 were established by comparisons with the optical rotation reported in
the literatures.8,15,16 The absolute configurations of (S)-11, (S)-13 were defined by analogy with the common spectroscopic behavior of (S)-para-
substituted styrene oxide. The absolute configurations of (S)-9, and (S)-10 were obtained by checking the optical rotations of the epoxides which
were prepared from the corresponding optical diols with a chemical correlation method.14 The absolute configuration of (S)-15 was deduced by
common behavior of A. niger hydrolyzing substituted styrene oxide. c The absolute configurations of (R)-19, (R)-20, (R)-21, (R)-22, (R)-23, (R)-25,
(R)-27, (R)-29 were established by comparison of the optical rotations reported in the literature.8,13,17 The absolute configurations of (R)-24, (R)-26,
(R)-28, (R)-30 were deduced by common behavior of A. niger hydrolyzing substituted styrene oxide.
Institute of Microbiology, the Chinese Academy of Sciences
(Beijing, China) under no. CGMCC 0496. All melting points of
the compounds are uncorrected. IR spectra were recorded on a
Shimadzu IR-440 spectrometer. EI mass spectra (MS) were run
on a HP-5989A mass spectrometer. 1H NMR spectra were
recorded on a Bruker AMX-300 spectrometer with tetramethyl-
silane as the internal standard. Chemical shifts are reported in
ppm and J are in Hz. Optical rotations were measured on a
Perkin-Elmer 241 polarimeter. HPLC was carried out using
Chiralcel OD or OJ, and Chiralpak AD or AS (0.46 cm ꢀ ×
25 cm) detected at UV 254 nm. GC was carried out using a
Rt-βDEXcstcolumn (Resteck) or Chiraldex G-PN column
Scheme 3
(Astec); TLC was carried out using HSG F254 silica gel plates
and silica gel (200–400 mesh) was used for chromatography.
enantioselectivities were found in the hydrolysis of ( )-4,
( )-12, ( )-13. The fair results of the reaction of ( )-4 were
2
General procedure for the culture of microorganisms
probably due to the small size of the fluorine atom, which
makes the enantioselectivity of the reaction much more like the
case of (ϩ)-14. On the other hand, the poor results of ( )-12,
( )-13 may be attributable to the electron donating properties
of the methyl and ethyl groups which decrease the essential
stability of the epoxides and increase the non-enzymatic,
concomitant hydrolysis during the attempted resolution.
The results reported in Table 5 also showed that more time
was needed for complete resolution when there was no substi-
tuted group in the aromatic ring [case ( )-14], and the substi-
tuent group in the benzene ring accelerated the rate of the
biohydrolysis.
The strain of A. niger CGMCC 0496 was maintained on agar
medium which contained K2HPO4 (1.0 g LϪ1), KCl (0.5 g LϪ1),
MgSO4ؒ7H2O (0.5 g LϪ1), FeSO4ؒ7H2O (0.0 g LϪ1), fructose
(10 g LϪ1), corn steep liquor (15 g LϪ1), and agar (3 g LϪ1) pH
6.8–7.0. The cells were grown on medium19 of K2HPO4 (1.0 g
L
Ϫ1), KCl (0.5 g LϪ1), MgSO4ؒ7H2O (0.5 g LϪ1), FeSO4ؒ7H2O
(0.01 g LϪ1), fructose (10 g LϪ1) and corn steep liquor (15 g LϪ1
)
in tap water. For large-scale cultures, a two-stage process was
used. 100 mL of medium (in a 500 mL flask) was first inocula-
ted and then cultured at 30 ЊC on a reciprocating shaker set at
100 cpm for 20 h. In the second stage, 1000 mL of medium (in a
5 L flask) was inoculated under the same conditions with 5% of
the first stage culture. The second stage lasted for 52 h. The cells
were harvested by centrifugation (HITACHI, CR20B2) at 7000
rpm and 5 ЊC for 30 min, and were then washed twice with 100
mL of 0.1 mol LϪ1 (pH 8.0) phosphate buffer. Fresh wet
mycelium was used for further biotransformation studies.
Conclusion
In summary, the A. niger CGMCC 0496 strain in asymmetric
hydrolysis of various substituted styrene oxides is a useful
methodology for preparing chiral styrene oxides and its corre-
sponding diols, especially for some para- and ortho-substituted
styrene oxides.
3
General procedure for the biotransformation of racemic
epoxide [( )-1]-[( )-15] with A. niger CGMCC 0496
Experimental
Wet mycelium 10 g or 20 g was suspended in 100 mL of sodium
phosphate buffer 0.1 M (pH = 8.0) and maintained at 25 ЊC. A
portion of 100 or 200 mg of the epoxide as a solution in DMF
(3 mL) was added to the cells suspension, and the medium
was stirred by a mechanical stirrer at 600 rpm. The course of
1
General
The strain of A. niger used in this work is registered at the
China General Microbiological Culture Collection Center,
O r g . B i o m o l . C h e m . , 2 0 0 4 , 2, 4 0 8 – 4 1 4
410

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bioconversion was monitored by the residual epoxide and the
formation of the corresponding diol by TLC. When an
appropriate degree of conversion was reached, the reaction was
stopped by adding 50 mL of ethyl acetate. The mycelium was
filtered off. The fungal cake was washed twice in 20 mL ethyl
acetate. The aqueous phase was saturated with sodium chloride,
and then extracted three times with ethyl acetate. The combined
organic layers were dried over Na2SO4 and the solvent was
removed under reduced pressure. Flash chromatography
(petroleum ether–EtOAc, gradient from 8 : 1 to 1 : 1) of the
residue gave the epoxides and corresponding diols. The yields,
the enantiomeric purity and the absolute configuration of the
reside epoxides and corresponding diols are summarized in
Tables 1–4.
excess was measured by capillary GC analysis using Rt-
βDEXcst(Resteck).
3.5 (S )-(؉)-2-(4-Chlorophenyl)-oxirane (S )-5. Following
the general procedure (10 g cells to 200 mg substrate), epoxide
(S)-5 was obtained in the yield as indicated in Table 5. Light oil;
95% ee; [α]2D7 ϩ18.7 (c 1.8, CHCl3), {lit.8 [α]D ϩ19.3 (c 1.16,
20
CHCl3) for >98% ee, (S)}; MS m/z (rel. intensity %): 156, 154
(M ϩ 1, 2, 8), 155, 153 (Mϩ, 3, 7), 138(3), 125(40), 119(39),
1
91(29), 89(100), 63(34), 50(17); H NMR (300 MHz, CDCl3,
TMS): δ 2.74 (dd, 1H, J1 = 5.4 Hz, J2 = 2.4 Hz), 3.13 (dd, 1H,
J1 = 5.4 Hz, J2 = 4.2 Hz), 3.82 (dd, 1H, J1 = 4.2 Hz, J2 = 2.4 Hz),
7.10–7.21 (m, 2H), 7.26–7.35 (m, 2H); IR (film): νmax 3054,
2992, 2920, 1602, 1496, 1478, 1417, 1381, 1199, 1090, 1015, 987,
879, 831, 769 cmϪ1. The enantiomeric excess was determined by
HPLC analysis using Chiralcel OJ column (eluent at V = 0.8
mL minϪ1, hexane : 2-propanol = 9 : 1).
3.1 (S )-(؉)-2-(4-Nitrophenyl) oxirane (S)-1. Following the
general procedure (10 g cells to 200 mg substrate), epoxide
(S)-1 was obtained in the yield as indicated in Table 5. Light
yellow solid; mp 73–75 ЊC; 94% ee; [α]2D5 ϩ35.0 (c 1.20, CHCl3),
{Lit.8 [α]2D0 ϩ37.6 (c 1.99, CHCl3) for >98% ee, (S)}; MS m/z
(rel. intensity %): 166([M ϩ 1]ϩ, 2), 165(Mϩ, 3), 164(6), 148(47),
118(68), 91(30), 89(100), 77(12), 65(21), 63(47), 51(21); 1H
NMR (300 MHz, CDCl3, TMS): δ 2.78 (dd, 1H, J1 = 5.5 Hz,
J2 = 2.5 Hz), 3.23 (dd, 1H, J1 = 5.5 Hz, J2 = 4.1 Hz), 3.96 (dd,
1H, J1 = 4.1 Hz, J2 = 2.5 Hz), 8.22, 7.46 (AB, 4H, J = 8.7 Hz); IR
(KBr): νmax 1610, 1530, 1410, 1350, 855 cmϪ1. The enantiomeric
excess was determined by HPLC analysis using a Chiralpak AD
column (eluent at V = 0.8 mL minϪ1, hexane : 2-propanol =
9 : 1).
3.6 (S )-(؉)-2-(3-Chlorophenyl) oxirane (S )-6. Following
the general procedure (10 g cells to 200 mg substrate), epoxide
(S)-6 was obtained in the yield as indicated in Table 5. Light oil;
28% ee; [α]2D7 ϩ3.6 (c 1.0, CHCl3), {lit.15 [α]2D2 Ϫ11.15 (c 1.56,
CHCl3) for >99% ee, (R)}; MS m/z (rel. intensity %): 157,
155(M ϩ 1, 0.8, 1.4), 156, 154(Mϩ, 10, 24), 141(15), 139(14),
125(50), 111(11), 91(56), 89(100); 1H NMR (300 MHz, CDCl3,
TMS): δ 2.79 (dd, 1H, J1 = 5.3 Hz, J2 = 2.4 Hz), 3.17 (dd, 1H,
J1 = J2 = 4.8 Hz), 3.86 (dd, 1H, J1 = J2 = 2.8 Hz), 7.16–7.62 (m,
4H); IR (film): νmax 3059, 2993, 1602, 1575, 1481, 1435, 1386,
1079, 999, 880, 823, 692 cmϪ1. Anal. calcd. For C8H7ClO: C,
62.15; H, 4.56; Cl, 22.93. Found: C, 62.11; H, 4.58; Cl, 23.35.
The enantiomeric excess was measured by capillary GC analysis
using Chiraldex G-PN column (Astec).
3.2 (S )-(؉)-2-(3-Nitrophenyl)-oxirane (S )-2. Following the
general procedure (10 g cells to 200 mg substrate); epoxide
(S)-2 was obtained in the yield as indicated in Table 5. Yellow
oil; 57% ee; [α]1D8 ϩ2.5 (c 2.8, CHCl3); MS m/z (rel. intensity %):
165(Mϩ, 18), 150(32), 136(68), 120(25), 105(17), 90(100),
3.7 (S )-(؉)-2-(2-Chlorophenyl) oxirane (S )-7. Following
the general procedure (20 g cells to 100 mg substrate), epoxide
(S)-7 was obtained in the yield as indicated in Table 5. Oil; 94%
ee; [α]2D3 ϩ48.3 (c 1.2, CHCl3) {lit.16 [α]2D5 ϩ32.2 (c 1.19, CHCl3)
for 99% ee, (S)}; MS m/z (rel. intensity %): 156, 154(Mϩ, 4, 16),
155, 153(Mϩ Ϫ 1, 10, 23), 134(9), 124(28), 119(75), 91(33),
1
77(22), 74(12), 65(52), 63(59); H NMR (300 MHz, CDCl3,
TMS): δ 2.80 (dd, 1H, J1 = 4.8 Hz, J2 = 2.4 Hz), 3.22 (dd, 1H,
J1 = 4.2 Hz, J2 = 0.9 Hz), 3.97 (dd, 1H, J1 = 3.9 Hz, J2 = 1.2 Hz),
7.40–7.75 (m, 2H), 8.02–8.24 (m, 2H); IR (film): νmax 3113,
2995, 1517, 1343, 1301, 1042, 983, 887, 789, 741 cmϪ1. The
enantiomeric excess was determined by HPLC analysis using a
Chiralpak AD column (eluent at V = 0.8 mL minϪ1, hexane :
2-propanol = 9 : 1).
1
89(100), 63(17); H NMR (300 MHz, CDCl3, TMS): δ 2.65
(dd, 1H, J1 = 5.9 Hz, J2 = 3.6 Hz), 3.19 (dd, 1H, J1 = 5.3 Hz,
J2 = 4.2 Hz), 4.21 (dd, 1H, J1 = J2 = 3.3 Hz), 7.10–7.50 (m, 4H);
IR (film): νmax 3061, 2993, 1699, 1593, 1482, 1442, 1383, 1249,
1121, 1053, 1035, 880, 755 cmϪ1. The enantiomeric excess was
measured by capillary GC analysis using Chiraldex G-PN
column (Astec).
3.3 (S )-(؊)-2-(2-Nitrophenyl)-oxirane (S )-3. Following the
general procedure (20 g cells to 100 mg substrate), epoxide
(S)-3 was obtained in the yield as indicated in Table 5. Light
yellow solid; mp 51–52 ЊC; 98% ee; [α]1D9.5 Ϫ107.2 (c = 1.7;
CHCl3); MS m/z (rel. intensity %): 165(Mϩ, 0.3), 149(2),
3.8 (S )-(؉)-2-(4-Bromophenyl) oxirane (S )-8. Following
the general procedure (20 g cells to 100 mg substrate); epoxide
(S)-8 was obtained in the yield as indicated in Table 5. Oil;
1
135(21), 105(10), 104(10), 91(79), 89(21), 79(71), 77(100); H
>99% ee; [α]2D3 ϩ13.9 (c 1.2, CHCl3) {lit.8 [α]D ϩ13.6 (c 1.46,
20
NMR (300 MHz, CDCl3, TMS): δ 2.67 (dd, 1H, J1 = 5.4 Hz,
J2 = 2.4 Hz), 3.30 (dd, 1H, J1 = 5.4 Hz, J2 = 4.2 Hz), 4.48 (dd,
1H, J1 = 4.5 Hz, J2 = 3.0 Hz), 7.40–7.55 (m, 1H), 7.58–7.75 (m,
2H), 8.15 (dd, 1H, J1 = 8.1 Hz, J2 = 1.2 Hz); IR (KBr): νmax 3150,
2997, 1532, 1353, 1254, 899, 859, 809, 737, 684 cmϪ1. The
enantiomeric excess was determined by HPLC analysis using
Chiralcel OD column (eluent at V = 0.8 mL minϪ1, hexane :
2-propanol = 9 : 1).
CHCl3) for >98% ee, (S)}; MS: m/z (rel. intensity %): 200,
198(Mϩ, 4), 199, 197(M Ϫ 1, 3), 169(14), 119(41), 89(100),
63(36); 1H NMR (300 MHz, CDCl3, TMS): δ 2.73 (dd, 1H, J1 =
5.4 Hz, J2 = 2.7 Hz), 3.13 (dd, 1H, J1 = 5.1 Hz, J2 = 3.6 Hz), 3.81
(dd, 1H, J1 = 3.9 Hz, J2 = 2.4 Hz), 7.05–7.18 (m, 2H), 7.40–7.48
(m, 2H); IR (film): νmax 3051, 2991, 2919, 1595, 1490, 1415,
1378, 1101, 1073, 1011, 987, 878, 828 cmϪ1. The enantiomeric
excess was determined by HPLC analysis using a Chiralcel
OJ column (eluent at V = 0.8 mL minϪ1, hexane : 2-propanol =
100 : 1).
3.4 (S )-(؉)-2-(4-Fluorophenyl)-oxirane (S )-4. Following
the general procedure (10 g cells to 200 mg substrate), epoxide
(S)-4 was obtained in the yield as indicated in Table 5. Oil; 41%
ee; [α]2D7 ϩ7.0 (c 1.9, CHCl3), {lit.8 [α]2D0 ϩ15.6 (c 0.97, CHCl3) for
98% ee, (S)}; MS m/z (rel. intensity %): 139 (M ϩ 1, 0.92),
138(Mϩ, 13), 110(13), 109(100), 107(11), 83(22), 81(4); 1H
NMR (300 MHz, CDCl3, TMS): δ 2.75 (dd, 1H, J1 = 5.4 Hz,
J2 = 2.5 Hz), 3.13 (dd, 1H, J1 = 5.3 Hz, J2 = 4.1 Hz), 3.84 (dd,
1H, J1 = 3.9 Hz, J2 = 2.7 Hz), 6.90–7.12 (m, 2H), 7.14–7.35 (m,
2H); IR (film): νmax 3055, 2995, 2926, 1608, 1514, 1479, 1383,
1222, 1157, 1015, 988, 882, 836, 820 cmϪ1. The enantiomeric
3.9 (S )-(؉)-2-(3-Bromophenyl) oxirane (S )-9. Following
the general procedure (10 g cells 200 mg substrate), epoxide
(S)-9 was obtained in the yield as indicated in Table 5. Oil; 35%
ee; [α]2D7 ϩ4.0 (c 1.1, CHCl3); MS m/z (rel. intensity %): 200,
198(Mϩ, 23, 23), 199, 197(Mϩ Ϫ 1, 32, 32), 169(27), 141(16),
1
119(67), 91(60), 89(100); H NMR (300 MHz, CDCl3, TMS):
δ 2.77 (dd, 1H, J1 = 5.4 Hz, J2 = 2.5 Hz), 3.15 (dd, 1H, J1 =
5.4 Hz, J2 = 4.1 Hz), 3.81 (dd, 1H, J1 = 4.0 Hz, J2 = 2.5 Hz),
O r g . B i o m o l . C h e m . , 2 0 0 4 , 2, 4 0 8 – 4 1 4
411

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7.10–7.50 (m, 4H); IR (film): νmax 3057, 2992, 1600, 1571, 1478,
1431, 1385, 1369, 1201, 1070, 997, 877, 786, 691 cmϪ1. The
enantiomeric excess was determined by HPLC analysis using a
Chiralpak AS column (eluent at V = 0.8 mL minϪ1, hexane : 2-
propanol = 9 : 1).
cmϪ1. The enantiomeric excess was measured by capillary GC
analysis using Chiraldex G-PN column (Astec).
3.15 (S )-(؉)-2-((2-Trifluoromethyl)phenyl)oxirane (S )-15.
Following the general procedure (10 g cells to 200 mg sub-
strate), epoxide (S)-15 was obtained in the yield as indicated in
Table 5. Oil; 98% ee; [α]2D1 ϩ35.4 (c 0.2, CHCl3) MS m/z (rel.
intensity %): 188(Mϩ, 16), 187(51), 174(33), 173(75), 159(39),
157(30), 138(27), 119(100), 108(25), 95(11); 1H NMR (300
MHz, CDCl3, TMS): δ 2.63 (dd, 1H, J1 = 6.0 Hz, J2 = 2.7 Hz),
3.18 (dd, 1H, J1 = 5.1 Hz, J2 = 4.5 Hz), 3.83 (dd, 1H, J1 = J2 = 1.8
Hz), 7.20–7.85 (m, 4H); IR (film): νmax 1655, 1606, 1458, 1316,
1168, 1121, 1060, 1035, 887, 769 cmϪ1. The enantiomeric excess
was measured by capillary GC analysis using Chiraldex-G-
PN column (Astec).
3.10 (S )-(؉)-2-(2-Bromophenyl) oxirane (S )-10. Following
the general procedure (20 g cells to 100 mg substrate), epoxide
(S)-10 was obtained in the yield as indicated in Table 5. Oil;
>99% ee; [α]1D8 ϩ68.7 (c 1.1, CHCl3); MS m/z (rel. intensity %):
200, 198(Mϩ, 17, 18), 199, 197(Mϩ Ϫ 1, 16, 15), 185(1), 171(9),
169(10), 141(1), 120(7), 119(84), 91(63), 90(41), 89(100); 1H
NMR (300 MHz, CD3COCD3, TMS): δ 2.64 (dd, 1H, J1 =
5.9 Hz, J2 = 2.6 Hz), 3.18 (dd, 1H, J1 = 5.9 Hz, J2 = 4.1 Hz), 4.15
(dd, 1H, J1 = 4.1 Hz, J2 = 2.6 Hz), 7.08–7.38 (m, 3H), 7.54 (dd,
1H, J1 = 7.9 Hz, J2 = 1.1 Hz); IR (film): νmax 3055, 2991, 2916,
1569, 1472, 1440, 1381, 1248, 1045, 1026, 879, 753 cmϪ1. The
enantiomeric excess was measured by capillary GC analysis
using Chiraldex G-PN column (Astec).
3.16 (R)-(؊)-1-(4-Nitrophenyl)-1,2-ethanediol (R)-16. Fol-
lowing the general procedure (10 g cells to 200 mg substrate),
diol (R)-16 was obtained in the yield as indicated in Table 5.
Yellow solid; mp 89–90 ЊC; 94% ee; [α]2D5 Ϫ19.5 (c 1.0, EtOH)
{lit.13 [α]2D0 Ϫ20.0 (c 1.15, MeOH) for 96% ee, (R)}; MS m/z (rel.
intensity %): 184([M ϩ 1]ϩ, 100), 182 (2) 166 (17), 152 (63), 136
(26), 122 (16), 106 (43), 94 (35), 91 (11), 78 (44); 1H NMR (300
MHz, CD3COCD3): δ 3.45 (br s, 2H), 3.58 (dd, 1H, J = 11.1,
6.7 Hz), 3.66 (dd, 1H, J = 11.1, 4.9 Hz), 4.85 (dd, 1H, J = 6.7,
4.9 Hz), 8.18, 7.69 (AB, 4H, J = 8.7 Hz); IR (KBr): νmax 3250,
1600, 1510, 1410, 1350, 1100, 1060, 850, 800, 720 cmϪ1. The
enantiomeric excess was determined by HPLC analysis using
Chiralcel OD column (eluent at V = 0.8 mL minϪ1, hexane :
2-propanol = 9 : 1).
3.11 (S )-(؉)-2-(4-Iodophenyl)oxirane (S )-11. Following the
general procedure (20 g cells to 100 mg substrate), epoxide
(S)-11 was obtained in the yield as indicated in Table 5. Yellow
oil; 97% ee; [α]2D6 ϩ25.1 (c 1.0, CHCl3); MS m/z (rel. intensity %):
247(M ϩ 1, 10), 246(Mϩ, 53), 245(36), 233(19), 230(10),
1
217(53), 127(6), 119(75), 91(69), 90(54), 89(75); H NMR (300
MHz, CDCl3, TMS): δ 2.74 (dd, 1H, J1 = 5.4 Hz, J2 = 2.6 Hz),
3.13 (dd, 1H, J1 = 5.3 Hz, J2 = 2.6 Hz), 2.80 (dd, 1H, J1 = 3.8 Hz,
J2 = 2.5 Hz), 6.90–7.05 (m, 2H), 7.60–7.78 (m, 2H); IR (film):
νmax 3053, 2990, 1589, 1474, 1413, 1377, 1056, 1006, 876, 825,
792cmϪ1. The enantiomeric excess was determined by HPLC
analysis using a Chiralpak AS column (eluent at V = 0.8 mL
minϪ1, hexane : 2-propanol = 9 : 1).
3.17 (R)-(؊)-1-(3-Nitrophenyl)-1,2-ethanediol (R)-17. Fol-
lowing the general procedure (10 g cells to 200 mg substrate),
diol (R)-17 was obtained in the yield as indicated in Table 5.
Yellow solid; mp 76–77 ЊC; 79% ee, [α]2D5 Ϫ13.9 (c 1.6, EtOH)
3.12 (S )-(؉)-2-(4-Methylphenyl)oxirane (S )-12. Following
the general procedure (10 g cells to 200 mg substrate), epoxide
(S)-12 was obtained in the yield as indicated in Table 5. Oil;
70% ee; [α]1D6 ϩ19.5 (c 1.2, CHCl3); MS: m/z (rel. intensity %):
235(M ϩ 1, 3), 134(Mϩ, 2), 121(100), 91(42), 77(24), 65(12),
20
{lit.13 [α]D ϩ26 (c 1.5, MeOH) for 92% ee, (S)}; MS m/z
(rel. intensity %): 184([M ϩ 1]ϩ, 5.1), 166 (70), 152 (100), 136
(34.4), 105 (60), 91 (16), 77 (52); 1H NMR (300 MHz,
CD3COCD3): δ 3.40 (br s, 2H), 3.70–3.50 (m, 2H), 4.87 (dd,
1H, J1 = J2 = 5.7 Hz), 7.61 (t, 1H, J1 = J2 = 8.2 Hz), 7.83 (d,
1H, J = 7.5 Hz), 8.10 (d, 1H, J = 8.2 Hz), 8.28 (s, 1H); IR
1
51(6); H NMR (300 MHz, CDCl3, TMS): δ 2.34 (s, 3H), 2.80
(dd, 1H, J1 = 5.4 Hz, J2 = 2.7 Hz), 3.13 (dd, 1H, J1 = 5.7 Hz, J2 =
4.2 Hz), 3.83 (dd, 1H, J1 = 3.9 Hz, J2 = 2.4 Hz), 7.10–7.20 (m,
4H); IR (film): νmax 3051, 2988, 2922, 1519, 1477, 1386, 1131,
1255, 1199, 1109, 986, 881, 818 cmϪ1. The enantiomeric excess
was measured by capillary GC analysis using Rt-βDEXcst
column (Resteck).
(KBr): νmax 3300, 1520, 1340, 1070, 1030, 860, 800, 720 cmϪ1
.
The enantiomeric excess was determined by HPLC analysis
using Chiralcel OD column (eluent at V = 0.8 mL minϪ1
hexane : 2-propanol = 9 : 1).
,
3.18 (R)-(؉)-1-(2-Nitrophenyl)-1,2-ethanediol (R)-18. Fol-
lowing the general procedure (10 g cells to 200 mg substrate),
diol (R)-18 was obtained in the yield as indicated in Table 5.
Light yellow solid; mp 108–109 ЊC; >99% ee, [α]1D9.5 ϩ53.4
(c = 1.1; EtOH) {lit.13 [α]2D0 ϩ9 (c 0.8, MeOH) for 47% ee, (R)};
MS m/z (rel. intensity %): 184([M ϩ 1]ϩ, 1.1), 166 (21), 152 (88),
3.13 (S )-(؉)-2-(4-Ethylphenyl)oxirane (S )-13. Following
the general procedure (10 g cells to 200 mg substrate), epoxide
(S)-13 was obtained in the yield as indicated in Table 5. Oil;
63% ee; [α]1D6 ϩ18.4 (c 1.4, CHCl3); MS m/z (rel. intensity %):
149(M ϩ 1, 4), 148(Mϩ, 14), 131(5), 119(62), 117(11), 115(5),
104(9), 91(21), 77(7); 1H NMR (300 MHz, CDCl3, TMS): δ 1.23
(t, 3H, J = 7.6 Hz), 5.30 (q, 2H, J = 7.6 Hz), 2.80 (dd, 1H, J1 =
5.5 Hz, J2 = 2.6 Hz), 3.13 (dd, 1H, J1 = J2 = 4.7 Hz), 3.83 (dd,
1H, J1 = 4.0 Hz, J2 = 2.5 Hz) 7.02–7.25 (m, 4H); IR (film): νmax
3057, 2967, 2874, 1908, 1617, 1519, 1384, 1255, 1128, 987, 881,
834 cmϪ1. The enantiomeric excess was determined by HPLC
analysis using a Chiralpak AS column (eluent at V = 0.8 mL
minϪ1, hexane : 2-propanol = 9 : 1).
1
135 (52), 104 (100), 91 (54), 79 (69), 77 (88); H NMR (300
MHz, CD3COCD3): δ 3.45 (br s, 2H), 3.60 (dd, 1H, J1 = 11.1Hz,
J2 = 7.0 Hz), 3.74 (dd, 1H, J1 = 11.1 Hz, J2 = 4.0 Hz), 5.28 (dd,
1H, J1 = 7.0 Hz, J2 = 4.0 Hz), 7.53 (dd, 1H, J1 = J2 = 7.8 Hz),
7.73 (dd, 1H, J1 = J2 = 7.8 Hz), 7.91–7.86 (m, 2H); IR (KBr):
νmax 3250, 1610, 1530, 1065, 825, 795, 750, 700 cmϪ1. The
enantiomeric excess was determined by HPLC analysis using
Chiralcel OJ column (eluent at V = 0.8 mL minϪ1, hexane :
2-propanol = 9 : 1).
3.14 (S )-(؊)-2-Phenyloxirane (S )-14. Following the gener-
al procedure (10 g cells to 200 mg substrate), epoxide (S)-14
was obtained in the yield as indicated in Table 5. Oil; 75% ee;
MS m/z (rel. intensity %): 122(M ϩ 2, 15), 121(M ϩ 1, 43),
3.19 (R)-(؊)-1-(4-Fluorophenyl)-1,2-ethanediol (R)-19. Fol-
lowing the general procedure (20 g cells to 100 mg substrate),
diol (R)-19 was obtained in the yield as indicated in Table 5.
White solid; mp 55–56 ЊC; 74% ee; [α]2D4 Ϫ23.5 (c 1.5, EtOH)
{lit.8 [α]2D0 Ϫ49 (c 1.07, CHCl3) for 81% ee, (R)}; MS m/z (rel.
intensity %): 156(Mϩ, 2), 140(3), 139(47), 126(6), 125(100),
123(15), 121(5), 119(4), 109(9), 107(4), 95(26), 91(3), 77(38),
1
120(Mϩ, 20), 105(100), 91(68), 77(88); H NMR (300 MHz,
CDCl3, TMS): δ 2.67 (dd, 1H, J1 = 5.7 Hz, J2 = 2.7 Hz), 3.15
(dd, 1H, J1 = 5.4 Hz, J2 = 3.9 Hz), 3.86 (dd, 1H, J1 = 3.9 Hz, J2 =
2.7 Hz) 7.15–7.45 (m, 4H); IR (film): νmax 3040, 2991, 2913,
1608, 1497, 1477, 1453, 1390, 1254, 1202, 985, 877, 759, 699
1
70(4); H NMR (300 MHz, CD3COCD3, TMS): δ 3.29(s, 2H),
O r g . B i o m o l . C h e m . , 2 0 0 4 , 2, 4 0 8 – 4 1 4
412

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3.45–3.78 (m, 2H), 4.70–4.83 (m, 1H), 7.02–7.20 (m, 2H), 7.22–
7.54 (m, 2H); IR (KBr): νmax 3207, 2934, 2878, 1608, 1512, 1470,
1245, 1234, 1105, 1086, 1033, 891, 831 cmϪ1. The enantiomeric
excess was determined by HPLC analysis using a Chiralpak AS
column (eluent at V = 0.8 mL minϪ1, hexane : 2-propanol =
9 : 1).
diol (R)-24 was obtained in the yield as indicated in Table 5. Oil;
70% ee; [α]2D3 Ϫ8.5 (c 1.2, EtOH); MS: m/z (rel. intensity %): 218,
216(Mϩ, 8), 191(22), 187(70), 185(74), 157(34), 117(7), 105(12),
1
81(21), 77(100); H NMR (300 MHz, CDCl3, TMS): δ 3.50–
2.80(br s, 2H), 3.58–3.85 (m, 2H), 4.74 (dd, 1H, J1 = 8.2Hz, J2 =
3.3 Hz), 7.12–7.52 (m, 3H), 7.55 (s,1H); IR (film): νmax 3260,
2918, 1656, 1594, 1569, 1415, 1193, 1111, 1068, 1057, 1022, 995,
906, 779, 695 cmϪ1. The enantiomeric excess was determined by
HPLC analysis using Chiralcel OD column (eluent at V = 0.8
mL minϪ1, hexane : 2-propanol = 95 : 5).
3.20 (R)-(؉)-1-(4-Chlorophenyl)-1,2-ethanediol
(R)-20.
Following the general procedure (10 g cells to 200 mg sub-
strate), diol (R)-20 was obtained in the yield as indicated in
Table 5. White solid; mp 83–84 ЊC; 95% ee; [α]2D7 ϩ18.7 (c 1.8,
CHCl3) {lit.13 [α]2D0 ϩ26 (c 2.1, EtOH) for 83% ee, (R)}; MS: m/z
(rel. intensity %): 172(Mϩ, 2), 155(11), 141(86), 113(26), 89(8),
77(100), 51(21); 1H NMR (300 MHz, CD3COCD3, TMS):
δ 3.13 (br s, 2H), 3.52 (dd, 1H, J1 = 10.9 Hz, J2 = 7.4 Hz), 3.60
(dd, 1H, J1 = 11.0Hz, J2 = 4.6 Hz), 4.71 (dd, 1H, J1 = 11.9 Hz,
J2 = 4.5 Hz), 7.30–7.40 (m, 2H), 7.40–7.50 (m, 2H); IR (KBr):
νmax 3310, 2977, 2939, 2879, 1596, 1490, 1457, 1341, 1110, 1085,
1031, 1015, 890, 823 cmϪ1. The enantiomeric excess was deter-
mined by HPLC analysis using Chiralcel OD column (eluent at
V = 0.8 mL minϪ1, hexane : 2-propanol = 95 : 5).
3.25 (R)-(؊)-1-(2-Bromophenyl)--1,2-ethanediol
(R)-25.
Following the general procedure (10 g diol to 200 mg substrate),
diol (R)-25 was obtained in the yield as indicated in Table 5.
White solid; mp 118–119 ЊC; 80% ee; [α]1D8 Ϫ24.8 (c 1.3, EtOH)
{lit.17 [α]2D0 Ϫ7.5 (c 0.99, CHCl3) for (R)}; MS m/z (rel. intensity
%): 219,217(M ϩ 1, 0.3, 0.2), 218,216(Mϩ, 2, 2), 201(2), 199(2),
188(7), 187(72), 185(76), 159(15), 157(19), 155(3), 137(3),
119(3), 107(2), 105(11), 91(5), 89(6), 78(44), 77(100); 1H NMR
(300 MHz, CD3COCD3, TMS): δ 3.29 (s, 2H), 3.42 (dd, 1H, J1 =
11.2 Hz, J2 = 7.9 Hz), 3.74 (dd, 1H, J1 = 11.2 Hz, J2 = 3.1 Hz),
5.07 (dd, 1H, J1 = 7.9 Hz, J2 = 3.1 Hz), 7.20 (ddd, 1H, J1 = J2 =
7.7 Hz, J3 = 1.8 Hz), 7.39 (ddd, 1H, J1 = J2 = 7.4 Hz, J3 =
1.2 Hz), 7.54 (ddd, 1H, J1 = J2 = 8.0 Hz, J3 = 1.2 Hz), 7.66 (dd,
1H, J1 = 7.8 Hz, J2 = 1.6 Hz); IR (KBr): νmax 3276, 2923, 1589,
1568, 1467, 1431, 1363, 1193, 1127, 1093, 1069, 1023, 953, 898,
836, 756 cmϪ1. Anal. calcd. for C8H7BrO: C, 44.27; H, 4.18; Br,
36.81. Found: C, 44.32; H, 4.27; Br, 36.90%. The enantiomeric
excess was determined by HPLC analysis using Chiralcel OD
column (eluent at V = 0.8 mL minϪ1, hexane : 2-propanol =
9 : 1).
3.21 (R)-(؊)-1-(3-Chlorophenyl)-1,2-ethanediol
(R)-21.
Following the general procedure (20 g cells to 100 mg sub-
strate), diol (R)-21 was obtained in the yield as indicated in
Table 5. Oil; 74% ee; [α]2D4 Ϫ15.8 (c 1.1, EtOH) {lit.13 [α]2D0 ϩ17
(c 1.4, EtOH) for 55% ee, (S)}; MS m/z (rel. intensity %): 175,
173(M ϩ 1, 0.38, 1.09), 174, 172(Mϩ, 3.65, 11.65), 143(25),
141(80), 139(4), 125(2), 115(10), 113(33), 105(4), 89(4), 77(100);
1H NMR (300 MHz, CD3COCD3, TMS): δ 2.92 (s, 2H), 3.59
(dd, 1H, J1 = 11.2 Hz, J2 = 8.1 Hz), 3.73 (dd, 1H, J1 = 11.2 Hz,
J2 = 2.6 Hz), 4.76 (dd, 1H, J1 = 7.8 Hz, J2 = 2.8 Hz) 7.01–7.28
(m, 3H), 7.35 (s, 1H); IR (film): νmax 3369, 2926, 2878, 1599,
1575, 1479, 1431, 1197, 1102, 1077, 1029, 786, 693 cmϪ1. Anal.
calcd. for C8H9ClO2: C, 55.67; H, 5.26; Cl, 20.54. Found: C,
55.69; H, 5.22; Cl, 20.53%. The enantiomeric excess was deter-
mined by HPLC analysis using Chiralcel OD column (eluent at
V = 0.8 mL minϪ1, hexane : 2-propanol = 9 : 1).
3.26 (R)-(؊)-1-(4-Iodophenyl)-1,2-ethanediol (R)-26. Fol-
lowing the general procedure (10 g cells to 200 mg substrate),
diol (R)-26 was obtained in the yield as indicated in Table 5.
White solid; mp 120–121 ЊC; 72% ee; [α]2D7 Ϫ17.7 (c 1.1, EtOH);
MS m/z (rel. intensity): 265(M ϩ 1, 3), 264(Mϩ, 30), 234(13),
233(100), 205(3), 141(11), 127(3), 107(25), 105(21), 79(25),
78(99); 1H NMR (300 MHz, CD3COCD3, TMS): δ 3.25 (s, 2H),
3.52 (dd, 1H, J1 = 11.0 Hz, J2 = 7.6 Hz), 3.61 (dd, 1H, J1 =
11.2 Hz, J2 = 4.2 Hz), 4.70 (dd, 1H, J1 = 7.6 Hz, J2 = 4.3 Hz),
7.20, 7.66 (AB, 4H, J = 8.1 Hz); IR (KBr): νmax 3369, 2923,
1586, 1484, 1396, 1092, 1066, 1034, 1023, 1006, 895, 832, 821,
523 cmϪ1. The enantiomeric excess was determined by HPLC
analysis using a Chiralpak AS column (eluent at V = 0.8 mL
minϪ1, hexane : 2-propanol = 9 : 1).
3.22 (R)-(؊)-1-(2-Chlorophenyl)-1,2-ethanediol
(R)-22.
Following the general procedure (20 g cells to 100 mg sub-
strate), diol (R)-22 was obtained in the yield as indicated in
Table 5. White solid; mp 99–100 ЊC; 84% ee; [α]2D3 Ϫ50.4 (c 1.7,
EtOH) {lit.13 [α]2D5 Ϫ50 (c 2.0, EtOH) for 66% ee, (R)}; MS m/z
(rel. intensity %): 172(Mϩ, 1.8), 143(38), 141(100), 113(24),
77(70);1 H NMR (300 MHz, CD3COCD3, TMS): δ 3.39 (dd,
1H, J1 = 11.2 Hz, J2 = 7.8 Hz), 3.46 (br s, 2H), 3.69 (dd, 1H, J1 =
11.2 Hz, J2 = 3.0 Hz), 5.12 (dd, 1H, J1 = 7.8 Hz, J2 = 3.0 Hz),
7.61–7.29 (m, 4H); IR (KBr): νmax 3300, 1635, 1440, 1070, 1040,
760 cmϪ1. The enantiomeric excess was determined by HPLC
analysis using Chiralcel OD column (eluent at V = 0.8 mL
minϪ1, hexane : 2-propanol = 8 : 2).
3.27 (R)-(؊)-1-(4-Methylphenyl)-1,2-ethanediol
(R)-27.
Following the general procedure (10 g cells to 200 mg sub-
strate), diol (R)-27 was obtained in the yield as indicated in
Table 5. White solid; mp 68–69 ЊC; 63% ee; [α]2D7 Ϫ48.1 (c 1.1,
EtOH) {lit.8 [α]2D5 Ϫ44.7 (c 0.75, CHCl3) for 66% ee, (R)}; MS
m/z (rel. intensity %): 152(Mϩ, 2.4), 135(11), 121(100), 105(5),
93(45), 77(25), 65(6); 1H NMR (300 MHz, CD3COCD3, TMS):
δ 2.32 (s, 3H), 3.18 (br s, 2H), 3.50 (dd, 1H, J1 = 11.1 Hz, J2 =
8.1 Hz), 3.60 (dd, 1H, J1 = 11.1 Hz, J2 = 4.2 Hz), 4.68 (dd, 1H,
J1 = 7.8 Hz, J2 = 4.2 Hz), 7.14, 7.28 (AB, 4H, J = 7.8 Hz);
IR (KBr): νmax 3264, 2921, 2863, 1515, 1347, 1097, 1070, 1033,
900, 849, 819 cmϪ1. The enantiomeric excess was determined
by HPLC analysis using a Chiralpak AS column (eluent at
V = 0.8 mL minϪ1, hexane : 2-propanol = 95 : 5).
3.23 (R)-(؊)-1-(4-Bromophenyl)--1,2-ethanediol
(R)-23.
Following the general procedure (10 g cells to 200 mg sub-
strate), diol (R)-23 was obtained in the yield as indicated in
Table 5. White solid; mp 104–105 ЊC; 86% ee; [α]2D3 Ϫ41.4 (c 1.2,
EtOH) {lit.8 [α]2D0 Ϫ37.2 (c 1.03, CHCl3) for 79% ee, (R)}; MS
m/z (rel. intensity %): 218,216(Mϩ, 3,3), 187(71), 185(83),
159(16), 157(19), 77(100), 51(18); 1H NMR (300 MHz,
CD3COCD3, TMS) δ 3.26 (br s, 2H), 3.53 (dd, 1H, J1 = 11.4 Hz,
J2 = 7.5 Hz), 3.61 (dd, 1H, J1 = 11.1 Hz, J2 = 4.8 Hz), 4.70 (dd,
1H, J1 = 7.5 Hz, J2 = 4.8 Hz), 7.20–7.38 (m, 2H), 7.40–7.60 (m,
2H); IR (KBr): νmax 3051, 2991, 2919, 1595, 1490, 1073, 1011,
828 cmϪ1. The enantiomeric excess was determined by HPLC
analysis using a Chiralpak AS column (eluent at V = 0.8 mL
minϪ1, hexane : 2-propanol = 8 : 2).
3.28 (R)-(؊)-1-(4-Ethylphenyl)-1,2-ethanediol (R)-28. Fol-
lowing the general procedure (10 g cells to 200 mg substrate),
diol (R)-28 was obtained in the yield as indicated in Table 5.
White solid; mp 64–65 ЊC; 70% ee; [α]2D3 Ϫ23.8 (c 1.2, EtOH);
MS m/z (rel. intensity %): 166(Mϩ, 0.16), 165(2), 155(1),
149(100), 136(3), 135(51), 133(7), 131(64), 120(4), 105(6),
91(20), 79(39); 1H NMR (300 MHz, CD3COCD3, TMS): δ 1.23
(t, 3H, J = 5.2 Hz), 2.65 (q, 2H, J = 7.6 Hz) 2.74–3.15 (br s, 2H),
3.24 (R)-(؊)-1-(3-Bromophenyl)-1,2-ethanediol (R)-24. Fol-
lowing the general procedure (10 g cells to 200 mg substrate),
O r g . B i o m o l . C h e m . , 2 0 0 4 , 2, 4 0 8 – 4 1 4
413

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3.60–3.78 (m, 2H), 4.78 (dd, 1H, J1 = 8.0 Hz, J2 = 3.5 Hz), 7.20,
7.26 (AB, 4H, J = 8.1 Hz); IR (KBr): νmax 3259, 2959, 2921,
2867, 1513, 1456, 1358, 1097, 1072, 1050, 1031, 900, 849, 831
cmϪ1. The enantiomeric excess was determined by HPLC analy-
References
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sis using a Chiralpak AS column (eluent at V = 0.8 mL minϪ1
hexane : 2-propanol = 9 : 1).
,
3.29 (R)-1-(؊)-Phenyl-1,2-ethanediol (R)-29. Following the
general procedure (10 g cells to 200 mg substrate), diol (R)-29
was obtained in the yield as indicated in Table 5. white solid; mp
66–67 ЊC; 52% ee; MS: m/z (rel. intensity %): 138(Mϩ, 3),
1
121(11), 107(100), 79(85), 77(65), 63(4), 51(14); H NMR (300
MHz, CDCl3, TMS) δ 2.92 (br s, 2H), 3.56–3.81 (m, 2H), 4.82
(dd, 1H, J1 = 8.1 Hz, J2 = 3.3 Hz), 7.26–7.50 (m, 5H); IR (KBr):
νmax 3203, 3061, 3030, 2934, 1469, 1448, 1343, 1228, 1101, 1089,
1054, 887, 833, 760, 748, 699 cmϪ1. The enantiomeric excess
was determined by HPLC analysis using Chiralcel OD column
(eluent at V = 0.8 mL minϪ1, hexane : 2-propanol = 95 : 5).
7 (a) M. Mischitz, W. Kroutil, U. Wandel and K. Faber, Tetrahedron:
Asymmetry, 1995, 6, 1261; (b) P. Moussou, A. Archelas and
R. Furstoss, J. Mol. Catal. B: Enzym., 1998, 5, 447.
8 S. Pedragosa-Moreau, C. Morisseau, J. Zylber, A. Archelas and
R. Furstoss, J. Org. Chem., 1996, 61, 7402.
9 H. Jin and Z-Y. Li, Biosci. Biotechnol. Biochem., 2002, 66,
1123.
10 H. Jin and Z-Y. Li, Chinese J. Chem., 2001, 3, 272.
11 T. Sakai, T. Kishimoto, Y. Tanaka, T. Ema and M. Utaka,
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12 A. Steinreibern, S. F. Mayer, R. Saf and K. Faber, Tetrahedron:
Asymmetry, 2001, 12, 1519.
3.30 (R)-(؊)-1-((2-Trifluoromethyl)phenyl)-1,2-ethanediol
(R)-30. Following the general procedure (20 g cells to 100 mg
substrate), diol (R)-30 was obtained in the yield as indicated in
Table 5. White solid; mp 66–67 ЊC; 82% ee; [α]2D1 Ϫ38.4 (c 0.5,
EtOH); MS: m/z (rel. intensity %): 206(Mϩ, 4), 175(44),
173(25), 155(100), 145(18), 127(55); 1H NMR (300 MHz,
CD3COCD3, TMS): δ): δ 3.18(br s, 2H), 3.48 (dd, 1H, J1 = 11.4
Hz, J2 = 8.1 Hz), 3.54 (dd, 1H, J1 = 11.1 Hz, J2 = 3.0 Hz), 5.10
(dd, 1H, J1 = 8.1Hz, J2 = 1.5 Hz), 7.40–7.56 (m, 1H), 7.58–7.78
(m, 2H), 7.82–7.98 (m, 1H); IR (KBr): νmax 3323, 2939, 2877,
13 Z-L. Wei, G-Q. Lin and Z-Y. Li, Bioorg. Med. Chem., 2000, 8,
1129.
1610, 1586, 1456, 1316, 1164, 1110, 1025, 770, 755, 667 cmϪ1
.
The enantiomeric excess was determined by HPLC analysis
14 Z-L. Wei, Z-Y. Li and G-Q. Lin, Tetrahedron, 1998, 54,
using Chiralcel OD column (eluent at V = 0.8 mL minϪ1
hexane : 2-propanol = 9 : 1).
,
13059.
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4, 2468.
18 C. Morisseau, G. Venturi, P. Moussou and J. Baratti, Biotechnol.
Tech., 1998, 12, 805.
Acknowledgements
The authors are grateful to the National Natural Science
Foundation of China (No. 200032020, No. 29770020) and
Chinese Academy of Sciences for financial support.
O r g . B i o m o l . C h e m . , 2 0 0 4 , 2, 4 0 8 – 4 1 4
414