Efficient synthesis of (R)-2-chloro-1-(3-chlorophenyl)ethanol by permeabilized whole cells of candida ontarioensis

Article abstract of DOI:10.1016/S1872-2067(11)60363-X

(R)-2-Chloro-1-(3-chlorophenyl)ethanol is a key pharmaceutical intermediate in the synthesis of β₃-adrenoceptor receptor (β;₃-AR) agonists. The asymmetric reduction of 2-chloro-1-(3-chlorophenyl)ethanone to (R)-2-chloro-1-(3-chlorophenyl)ethanol catalyzed by resting cells of Candida ontarioensis was studied. At a substrate concentration of 10 g/L, the microbial cells showed excellent catalytic activity under the optimum reaction conditions, giving the product in 99.9% ee and 99.0% yield. After cetyltrimethylammonium bromide-pretreatment, the activity of permeabilized Candida ontarioensis cells was increased by more than 2-fold and the product could be produced over the significantly shortened reaction period of 24 h in 99.9% ee and 97.5%yield at a substrate concentration of 30 g/L. This work provides a practical approach for the efficient preparation of the important chiral intermediate (R)-2-chloro-1-(3-chlorophenyl) ethanol.

Full text of DOI:10.1016/S1872-2067(11)60363-X

CHINESE JOURNAL OF CATALYSIS
Volume 33, Issue 4, 2012
Online English edition of the Chinese language journal
Cite this article as: Chin. J. Catal., 2012, 33: 681–687.
ARTICLE
Efficient Synthesis of
(R)-2-Chloro-1-(3-chlorophenyl)ethanol by Permeabilized
Whole Cells of Candida ontarioensis
NI Ye*, ZHANG Beihua, SUN Zhihao
The Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122,
Jiangsu, China
Abstract: (R)-2-Chloro-1-(3-chlorophenyl)ethanol is a key pharmaceutical intermediate in the synthesis of ꢀ3-adrenoceptor receptor
(ꢀ3-AR) agonists. The asymmetric reduction of 2-chloro-1-(3-chlorophenyl)ethanone to (R)-2-chloro-1-(3-chlorophenyl)ethanol catalyzed
by resting cells of Candida ontarioensis was studied. At a substrate concentration of 10 g/L, the microbial cells showed excellent catalytic
activity under the optimum reaction conditions, giving the product in 99.9% ee and 99.0% yield. After cetyltrimethylammonium bro-
mide-pretreatment, the activity of permeabilized Candida ontarioensis cells was increased by more than 2-fold and the product could be
produced over the significantly shortened reaction period of 24 h in 99.9% ee and 97.5% yield at a substrate concentration of 30 g/L. This
work provides a practical approach for the efficient preparation of the important chiral intermediate (R)-2-chloro-1-(3-chlorophenyl)
ethanol.
Key words: (R)-2-chloro-1-(3-chlorophenyl)ethanol; asymmetric reduction; cetyltrimethylammonium bromide; permeability; Candida
ontarioensis
Optically pure chiral alcohols are valuable building blocks
for the synthesis of pharmaceuticals, agrochemicals, and liq-
uid crystals [1]. The ꢁ-halohydrins are particularly important
as synthons for the preparation of ꢀ-adrenergic drugs [2].
Several chemical and biological approaches for the synthesis of
optically active ꢁ-halohydrins have been developed [3,4] and
the asymmetric reduction of the corresponding prochiral aro-
matic ketones is regarded as the most promising methodology
[5]. Owing to its excellent stereoselectivity, mild reaction
conditions, and environmental benignancy, biocatalysis has
been reported for the production of a variety of optically pure
pharmaceutical intermediates [6–8]. In biocatalytic reactions,
whole-cell systems are often preferred to isolated enzymes as
the tedious enzyme isolation/purification processes required
are eliminated and reactions involving multiple en-
zymes/pathways as well as co-factor regeneration are allowed
[9–11].
(R)-2-Chloro-1-(3-chlorophenyl)ethanol is a versatile in-
termediate for the synthesis of ꢀ3-adrenoceptor receptor
(ꢀ3-AR) agonists, such as the anti-depressant drug SR58611
and the thermogenic anti-obesity drug BRL 37344 [12]. Sev-
eral groups have reported the biocatalytic synthesis of (R)-2-
chloro-1-(3-chlorophenyl)ethanol. Sawa et al. [13] reported the
asymmetric synthesis of (R)-2-chloro-1-(3-chlorophenyl)
ethanol using whole cells of Rhodotorula glutinis at a substrate
concentration of 3.25 g/L. Lin and co-workers [14] reported
the bioreduction of 2-chloro-1-(3-chlorophenyl) ethanone by
Saccharomyces cerevisiae, providing the product after 48 h in
99% ee and 97% yield at a substrate concentration of 28 g/L.
Recombinant E. coli cells and cell-extracts from Nocardia
globerula and Rhodotorula glutinis have also been reported for
the asymmetric production of (R)-2-chloro-1-(3-chlorophenyl)
ethanol in over 99% ee. However, the inclusion of expensive
additives such as NADP+/NAD+ and glucose dehydrogenase
Received 2 Novebmer 2011. Accepted 6 January 2012.
*Corresponding author. Tel/Fax: +86-510-85329265; E-mail: yni@jiangnan.edu.cn
This work was supported by the National Basic Research and Development Program of China (973 Program, 2011CB710800), New Century Excellent Talents in
University (NCET-11-0658), Natural Science Foundation of Jiangsu Province (BK2011150), the Program of Introducing Talents of Discipline to Universities
(111-2-06), and a Project Funded by the Priority Academic Program Development of Jiangsu Higher Education Institutions.
Copyright © 2012, Dalian Institute of Chemical Physics, Chinese Academy of Sciences. Published by Elsevier BV. All rights reserved.
DOI: 10.1016/S1872-2067(11)60363-X

NI Ye et al. / Chinese Journal of Catalysis, 2012, 33: 681–687
The reaction system consisted of 1 g of wet cells (collected
O
OH
X
X
from 30 ml of culture broth), glucose (5%, w/v), the appropri-
ate amount of 2-chloro-1-(3-chlorophenyl)ethanone, and po-
tassium phosphate buffer (0.2 mol/L, pH 6.5), in a final vol-
ume of 10 ml. The reaction was conducted in a 50-ml Erlen-
meyer flask with stopper and shaken at 30 °C and 180 r/min
for 72 h. The mixture was extracted with an equal volume of
ethyl acetate and the organic layer was concentrated under
vacuum and diluted with mobile phase for subsequent HPLC
analysis.
Candida ontarioensis
NADPH
NADP+
R
ꢀ
1 6
R
Gluconolactone
Glucose
O
O
O
Cl
COOEt
OC2H5
7
8
R = H, m-Cl, p-Br, or m-NH2, p-NO2; X = H, Cl, or Br.
Scheme 1. Bioreduction of compounds 1–8 with Candida ontarioensis
whole cells.
1.4 Activity assay
was required for regeneration of the cofactor [15–18].
In this study, Candida ontarioensis was selected from a
screening panel of 90 strains in our laboratory collection be-
cause of its high carbonyl reductase activity towards various
halo-acetophenone substrates and 2-chloro-1-(3-chlorophenyl)
ethanone was used as a model substrate to investigate the
asymmetric reduction catalyzed by whole-cells of Candida
ontarioensis (Scheme 1). Using cetyltrimethylammonium
bromide (CTAB)-permeabilized cells, the reaction time was
significantly reduced from 72 to 24 h and the product was
obtained in 99.9% ee and 97.5% yield at a 30 g/L substrate
concentration.
The carbonyl reductase activity of Candida ontarioensis
cells was measured according to the reaction conditions men-
tioned above by analyzing the reaction during the first 30 min
to determine the initial velocity. One unit of the enzyme activ-
ity (U) is defined as the amount of enzyme required for gen-
eration of 1 ꢂmol (R)-2-chloro-1-(3-chlorophenyl)ethanol per
minute.
The enantiomeric excess (ee) and analytic yield of
(R)-2-chloro-1-(3-chlorophenyl)ethanol were determined on
an Agilent 1100 HPLC (California, USA) equipped with a 5
ꢂm Chiralcel OB-H (0.46 mm × 250 mm, Tokyo, Japan)
column using an UV detector at 220 nm. The HPLC was per-
formed using n-hexane/isopropanol (9/1, HPLC grade, Xingke
Chemical Reagent Co., Ltd, China) as the mobile phase at a
flow rate of 1.0 ml/min and the column temperature was 30
°C.
1 Experimental
1.1 Screening and identification of the target strain
Three strains were screened from the 90 strains in our
laboratory using 10 g/L 2-chloro-1-(3-chlorophenyl)ethanone
(Porton Co., Ltd, China) as the substrate and Candida sp. BC
2A showed the highest carbonyl reductase activity. The 18S
rRNA gene sequence of Candida sp. BC 2A was determined
(GenBank number: JN820129), identifying the strain as Can-
dida ontarioensis.
2 Results and discussion
2.1 Cell growth and carbonyl reductase production by
Candida ontarioensis
The time course of cell growth and carbonyl reductase
production by Candida ontarioensis was investigated. As
shown in Fig. 1, the cell growth continued throughout the
entire fermentation process and the carbonyl reductase activ-
ity of whole-cells increased rapidly over the first 20 h. A
maximum activity of 25.4 U/L (0.74 U/gWCW) and a cell mass
of 34.5 gWCW/L (WCW = wet cell weight) was reached at the
20 h mark. A decrease in enzymatic activity was evident after
20 h because of the degradation of reductase by protease at the
end of the fermentation process. Thus, Candida ontarioensis
cells were collected after 20 h of the fermentation process for
further bioreduction studies.
1.2 Cultivation conditions of Candida ontarioensis
The fermentation medium was composed of 1% (w/v) glu-
cose, 2% (w/v) corn steep liquor power, and 0.005% (w/v)
CaCl2 and was adjusted to pH 6.5. The microorganism was
cultured at 30 °C for 20 h with 180 r/min shaking. The cells
were collected by centrifugation at 2350 ꢀ g for 5 min and
washed twice with 0.85% saline.
The cell pellet was treated with 4 g/L CTAB at 4 °C for 20
min to improve permeability. The CTAB-treated and untreated
Candida ontarioensis cells were used as biocatalysts in the
bioconversion of 2-chloro-1-(3-chlorophenyl)ethanone.
2.2 Optimization of bioreduction conditions
1.3 Asymmetric bioreduction of
2-chloro-1-(3-chlorophenyl)ethanone in aqueous medium
Major parameters in the asymmetric reduction of 2-chloro-
1-(3-chlorophenyl)ethanone were investigated, including
buffer pH, temperature, cell concentration, and cosubstrate (for

NI Ye et al. / Chinese Journal of Catalysis, 2012, 33: 681–687
100
80
60
40
20
27
24
21
18
15
12
9
40
35
30
25
20
15
100
80
60
40
20
0
Cell concentration (g/L)
50
100
150
200
250
ee
ee
6
Activity of whole cells
3
Cell mass
0
14
16
18
20
22
24
Fermentation time (h)
0
10
20
30
40
50
60
70
Reaction time (h)
Fig. 1. Time courses of carbonyl reductase activity and cell growth of
Candida ontarioensis. Fermentation conditions: 50 ml fermentation me-
dium in 250 ml shake flask, 2% inoculum, 30 °C, and 180 r/min shaking.
Fig. 3. Effects of cell concentration on product ee and yield in the bio-
reduction. Reaction conditions: 5% (w/v) glucose, 10 g/L substrate, po-
tassium phosphate buffer (0.2 mol/L, pH 6.5) and different cell concen-
trations in a final volume of 10 ml, 30 °C, 72 h.
cofactor recycling).
the bioreduction reaction.
2.2.1 Effect of pH and temperature
2.2.2 Effect of cell concentration
To evaluate the effect of pH on the carbonyl reductase ac-
tivity, the reaction was carried out in buffers at various pH
values, including sodium acetate buffer (pH 4.5–6) and potas-
sium phosphate buffer (pH 6–8). Figure 2(a) shows that the
buffer pH did not affect the ee of the product and that the
product yield was greater than 90% in the pH range of 5–8. The
asymmetric reduction was sensitive to acidic environments and
the product yield was observed to decrease from 94.3% to
72.9% when the pH was reduced from 5.0 to 4.5. Therefore, pH
6.5 was chosen as the optimum pH for this study.
Reaction temperature could also affect the activity and sta-
bility of the enzyme. An increase in the temperature could
make the activation energy for the reaction easier to reach and
the mass transfer of substrate/product could also be improved.
The effect of reaction temperature was investigated (Fig. 2(b))
and the results indicated that the product yield was maintained
at 98%–99% between a temperature range of 30–45 °C.
Therefore, 30 °C was chosen as the optimum temperature for
The amount of cell mass in the biocatalytic reaction system
directly affects the product yield of the bioconversion. The
reaction time course at different cell concentrations was also
investigated (Fig. 3). As shown in Fig. 3, the yield of product
increased rapidly over the first 24 h for all of the cell concen-
trations tested. A yield of 100% was reached at cell concentra-
tions of 200 and 250 g/L after 36 h of reaction. Thus, a cell
concentration of 200 g/L was used as the optimum cell con-
centration.
2.2.3 Effect of different cosubstrates
Cofactor regeneration is crucial for the continuous progress
of asymmetric reduction, because of the limited amount of
NAD(P)H in microbial cells [19,20]. Thus, a cosubstrate
should be added to the reaction system for the cofactor to be
100
100
(a)
95
90
85
80
75
70
99
98
97
96
95
94
(b)
Yield
ee
Yield
ee
4.0 4.5 5.0 5.5 6.0 6.5 7.0 7.5 8.0 8.5
pH
25
30
35
40
45
50
Temperature (oC)
Fig. 2. Effects of pH (a) and temperature (b) on product ee and yield in the bioreduction. (a) Reaction conditions: 1 g wet cells, 5% (w/v) glucose, 10 g/L
substrate, various buffers (0.2 mol/L, pH 4.5ꢁ8.0 sodium acetate buffer and potassium phosphate buffer) in a final volume of 10 ml, 30 °C, 72 h. (b) Reac-
tion conditions: the same as above except different temperatures and potassium phosphate buffer (0.2 mol/L, pH 6.5).

NI Ye et al. / Chinese Journal of Catalysis, 2012, 33: 681–687
m-aminoacetophenone, and p-nitroacetophenone. The corre-
100
80
60
40
20
0
sponding alcohols reached over 99.9% ee and 99.0% yield in
all cases. However, Candida ontarioensis whole-cells showed
no activity for ethyl 4-chloroacetoacetate (COBE) and ethyl
2-oxo-4-phenylbutyrate (OPBE). The results suggest that the
carbonyl reductase activity of Candida ontarioensis has high
substrate specificity and enantioselectivity towards various
halo-, amino-, and nitro-substituted acetophenones, but not for
keto ester substrates.
2.4 Reaction acceleration by improved membrane
permeability
Cosubstrate
Whole-cell catalyzed reactions often exhibit much slower
reaction rate than that of isolated enzymes due to the mass
transfer limitations imposed by the cellular membrane. Our
previous studies demonstrated that the reaction rate of
whole-cell biocatalysis could be significantly accelerated by
modulating membrane permeability [21,22]. Various chemical
and physical strategies including detergents, solvents, chela-
tors, and freeze-thaw treatments have been employed to ad-
dress the permeability issue. CTAB is a cationic surfactant and
has long been utilized as an effective permeabilizer to enhance
the permeability of microbial cell membranes [23]. Low
product yields and prolonged reaction times were often re-
ported in whole-cell asymmetric reductions involving unnatu-
ral and hydrophobic substrates. Yu and co-workers [24] re-
ported that the reactions catalyzed by alcohol dehydrogenase
Fig. 4. Effect of various cosubstrates on the bioreduction reaction.
Reaction conditions: 2 g wet cells, 5% (w/v) cosubstrates, 10 g/L sub-
strate, potassium phosphate buffer (0.2 mol/L, pH 6.5) in a final volume
of 10 ml, 30 °C, 72 h.
recycled by glucose dehydrogenase or alcohol dehydrogenase
mediated oxidation of the cosubstrate. Sugars and alcohols,
including glucose, sucrose, ethanol, and isopropyl alcohol, are
commonly used as cosubstrates. In this study, several cosub-
strates were investigated (Fig. 4). In comparison with the con-
trol (no cosubstrate), the highest yield was observed in groups
supplemented with glucose or xylose. In contrast, the use su-
crose, glycerol, and isopropanol resulted in no obvious positive
effect and negative effects were actually observed. The dif-
ferent cosubstrates did not affect the ee of product and the
optical purity achieved in all cases was 99.9%. Glucose was
therefore selected as cosubstrate due to its low cost.
and
glucose-6-phosphate
dehydrogenase
from
CTAB-permeabilized fresh brewer's yeast were increased by
525- and 7.9-fold, respectively. Cortez and co-workers [25]
compared various permeabilization methods including CTAB,
Triton X-100 and freezing-thawing and found that
CTAB-pretreatment could effectively improve the accessibility
of glucose-6-phosphate dehydrogenase in Candida guillier-
monedii. In this study, the effect of CTAB-treatment on the
bioreduction reaction was investigated in detail.
2.3 Substrate specificity
The asymmetric synthesis of several optically active alco-
hols by Candida ontarioensis was also tested (Table 1). Can-
dida ontarioensis whole-cells showed high reaction activity
and enantioselectivity towards various halo-acetophenones
including 2-chloroacetophenone, 2-bromoacetophenone,
2-chloro-1-(3-chlorophenyl)ethanone, p-bromoacetophenone,
2.4.1 Effect of CTAB pretreatment time
Herein, the effect of CTAB-pretreatment time on the initial
reaction rate was studied (Fig. 5(a)). The initial reaction rate
reached a maximum when Candida ontarioensis cells were
pretreated with 2% CTAB for 20 min at 4 °C and the activity of
whole-cells reached 74.8 U/L, representing a greater than
2-fold increase relative to the cell without CTAB-treatment.
Hence the favorable CTAB pretreatment time was determined
to be 20 min at 4 °C.
Table 1 Bioreduction of compounds 1–8 by Candida ontarioensis
whole cells
Configu- Yield
Substrate
R
X ee (%)
ration
R
R
(%)
99.0
99.0
2-Chloroacetophenone
2-Bromoacetophenone
2-Chloro-1-(3-chlorophe
nyl)ethanone
1
2
H
H
Cl > 99.9
Br > 99.9
3
m-Cl Cl > 99.9
R
99.0
p-Bromoacetophenone
m-Aminoacetophenone
p-Nitroacetophenone
COBE
4
5
6
7
8
p-Br
m-NH2
p-NO2
—
H
H
H
—
—
> 99.9
> 99.9
> 99.9
—
R
R
S
—
—
99.0
99.0
99.0
—
2.4.2 Effect of CTAB pretreatment concentration
The effect of CTAB concentration on the asymmetric re-
duction was also examined. As shown in Fig. 5(b), the greatest
yield was achieved at a CTAB concentration of 4 g/L. Further
OPBE
—
—
—
Reaction conditions: 2 g wet cells, 10 g/L substrate, potassium phosphate
buffer (0.2 mol/L, pH 6.5) in a final volume of 10 ml, 30 °C, 72 h.

NI Ye et al. / Chinese Journal of Catalysis, 2012, 33: 681–687
80
70
60
50
40
30
20
100
(a)
(b)
98
96
94
92
90
88
86
84
Yield
ee
0
10
20
30
40
50
0
2
4
6
8
10
12
Pretreatment time of CTAB (min)
Concentration of CTAB (g/L)
Fig. 5. Effects of CTAB pretreatment time (a) and concentration (b) on the activity of Candida ontarioensis whole-cells and the product ee and yield. (a)
Reaction conditions: 2 g wet cells, 2% CTAB, 30 g/L substrate, and potassium phosphate buffer (0.2 mol/L, pH 6.5) in a final volume of 10 ml, 30 °C, 30
min. (b) Reaction conditions: 2 g wet cells, 30 g/L substrate, and potassium phosphate buffer (0.2 mol/L, pH 6.5) in a final volume of 10 ml, 30 °C, 72 h
increases in the CTAB concentration, however, led to a slight
decline in the yield and also compromised the enantioselectiv-
ity. An appreciable decrease in product ee at over 10 g/L of
CTAB was noted. It was speculated that CTAB might interact
with carbonyl reductase and interfere with its enzymatic ac-
tivity and enantioselectivity. Also, the increased membrane
permeability could promote the formation of the alcohol
product of the opposite configuration catalyzed by other re-
ductases, leading to a decreased ee value in the product.
Therefore, a CTAB concentration of 4 g/L was used for per-
meabilization treatment.
2.6 Isolation and identification of the bioreduction
product
The reaction system consisted of 24 g wet cells, 1200 mg
substrate, and 0.2 mol/L potassium phosphate buffer (pH 6.5)
in a final volume of 120 ml. The process was carried out at 180
r/min and 30 °C over 24 h. The mixture was extracted with
ethyl acetate and the organic layer was concentrated under
vacuum. The crude product was separated by silica gel chro-
matography using ethyl acetate/petroleum ether (1:6) as elu-
ent. The optical purity and yield of (R)-2-chloro-
1-(3-chlorophenyl)ethanol were determined by HPLC analy-
sis. After purification, 800 mg of the product with > 99.9% ee
and 99% purity was obtained. The yields of biotransformation
and purification processes were calculated to be 99% and
66.7%, respectively.
2.5 Time courses of bioreduction catalyzed by
CTAB-pretreated and untreated cells
To evaluate the potential of CTAB-pretreated cells as
whole-cell biocatalysts in the biosynthesis of (R)-2-chloro-
1-(3-chlorophenyl)ethanol, an asymmetric reduction catalyzed
by CTAB-pretreated and untreated Candida ontarioensis cells
was conducted (Fig. 6). During the first 10 h, the reaction rate
of CTAB-pretreated cells was 2-fold greater than that of the
cells without CTAB-treatment. The reaction catalyzed by
CTAB-pretreated cells was almost finished after 24 h (97.5%
yield), whereas a yield of only 86.6% was attained by un-
treated cells over a prolonged reaction period of 72 h. It was
speculated that the reaction progress was hindered in the later
stage of bioconversion by high levels of intracellular product,
because of mass transfer resistance across the untreated-cell
membrane. The optical purity of product was maintained at
99.9% ee throughout the bioconversion process. Conse-
quently, the CTAB-pretreatment resulted in a greatly reduced
reaction time at an elevated substrate concentration of 30 g/L
and was therefore feasible for the bioreduction catalyzed by
whole- cells of Candida ontarioensis. Compared with the
bioreduction of 2-chloro-1-(3-chlorophenyl)ethanone by Sac-
charomyces cerevisiae [14] on a 1-L reaction scale, the reac-
tion time was shortened considerably in this study.
The product was identified by 1H (400 MHz) and 13C (400
MHz) NMR using CDCl3 as solvent, and the 1H and 13C NMR
spectra are as follows. 1H NMR (CDCl3, 400 MHz): ꢀ 2.70 (s,
1H, OH), 3.60 (dd, 1H, CH2Cl), 3.72 (dd, 1H, CH2Cl), 4.87
100
80
60
40
Yield
ee
20
0
0
10
20
30
40
50
60
70
Reaction time (h)
Fig. 6. Time courses of bioreduction catalyzed by CTAB-pretreated
(solid) and untreated (hollow) Candida ontarioensis cells. Reaction con-
ditions: 200 g wet cells, 30 g/L substrate, potassium phosphate buffer (0.2
mol/L, pH 6.5) in a final volume of 1 L, 30 °C, 72 h.

NI Ye et al. / Chinese Journal of Catalysis, 2012, 33: 681–687
(dd, 4H, aromatic H), 7.26–7.32 (m, 1H, CH); 13C NMR
(CDCl3, 100 MHz): ꢀ 51, 73, 124, 126, 129, 130, 135, 142.
2
3
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3 Conclusions
4
5
The asymmetric reduction of 2-chloro-1-(3-chlorophenyl)
ethanone to its corresponding chiral alcohol by Candida on-
tarioensis whole-cells was investigated. In the aqueous system,
the microbial cells showed excellent catalytic ability under the
optimum reaction conditions (30 °C, pH 6.5, 10 g/L substrate
concentration), giving the product in 99.9% ee and 99.0%
yield. A substrate specificity study indicated that the carbonyl
reductase from Candida ontarioensis shows high substrate
specificity and enantioselectivity for various halo-, amino, and
nitro-substituted acetophenones, but not for keto ester sub-
strates. CTAB-treatment was applied to enhance the cellular
membrane permeability and to accelerate the reaction. Using
cells pretreated with 4 g/L CTAB for 20 min, the reaction rate
was increased by 2-fold over the first 10 h of the biotransfor-
mation and a product yield of 97.5% was achieved in 24 h,
which was 48 h earlier than that without CTAB-treatment
(86.6% yield at 72 h). The results demonstrate the importance
of good mass transfer across the cell membrane for the
whole-cell biocatalysis and that the efficiency of biocatalytic
reactions can be greatly improved by increasing membrane
permeability. This study provides a feasible and green ap-
proach for the efficient production of (R)-2-chloro-1-(3-
chlorophenyl)ethanol. To further enhance the enzymatic activ-
ity and efficacy of the biocatalytic reaction, asymmetric
preparation of (R)-2-chloro-1-(3-chlorophenyl) ethanol using
recombinant strains overexpressing carbonyl reductase is un-
derway.
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