Preparative access to medicinal chemistry related chiral alcohols using carbonyl reductase technology

Article abstract of DOI:10.1016/j.tetasy.2013.09.015

Libraries of highly enantioenriched secondary alcohols in both enantiomeric forms were synthesised by enzymatic reduction of their parent ketones using selectAZyme carbonyl reductase (CRED) technology. Commercially available CREDs were able to reduce a range of substrate classes efficiently and with very high enantioselectivity. Matching substrate classes to small subsets of CREDs enabled the fast development of preparative bioreductions and the rapid generation of 100-1500 mg samples of chiral alcohols in typically >95% ee and the majority in ≥99.0% ee. The conditions for small scale synthesis were then scaled up to 0.5 kg to deliver one of the chiral alcohols, (S)-1-(4-bromophenyl)-2-chloroethanol, in 99.8% ee and 91% isolated yield.

Full text of DOI:10.1016/j.tetasy.2013.09.015

Tetrahedron: Asymmetry 24 (2013) 1369–1381
Contents lists available at ScienceDirect
Tetrahedron: Asymmetry
journal homepage: www.elsevier.com/locate/tetasy
Preparative access to medicinal chemistry related chiral alcohols
using carbonyl reductase technology
a
b
c
a
Andrew S. Rowan a, , Thomas S. Moody , Roger M. Howard , Toby J. Underwood , Iain R. Miskelly ,
⇑
Yanan He d, Bo Wang d
a Biocatalysis & Isotope Chemistry Group, Almac, 20 Seagoe Industrial Estate, Craigavon BT63 5QD, Northern Ireland, UK
b Chemical Research & Development, Pfizer Ltd, Sandwich Laboratories, Ramsgate Road, Kent, UK
c Royal Society of Chemistry, Thomas Graham House, Science Park, Milton Road, Cambridge, UK
d Biotools, Inc., 17546 Bee Line Highway, Jupiter, FL 33458, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
Libraries of highly enantioenriched secondary alcohols in both enantiomeric forms were synthesised by
enzymatic reduction of their parent ketones using selectAZyme™ carbonyl reductase (CRED) technology.
Commercially available CREDs were able to reduce a range of substrate classes efficiently and with very
high enantioselectivity. Matching substrate classes to small subsets of CREDs enabled the fast develop-
ment of preparative bioreductions and the rapid generation of 100–1500 mg samples of chiral alcohols
in typically >95% ee and the majority in P99.0% ee. The conditions for small scale synthesis were then
scaled up to 0.5 kg to deliver one of the chiral alcohols, (S)-1-(4-bromophenyl)-2-chloroethanol, in
99.8% ee and 91% isolated yield.
Received 10 June 2013
Accepted 18 September 2013
Available online 29 October 2013
Ó 2013 Elsevier Ltd. All rights reserved.
1. Introduction
O
OH OH
O
O- 0.5Ca2+
N
H
N
Chiral secondary alcohols are an important building block in
medicinal chemistry. Indeed at least 17 of the top 100 brand-name
prescription drugs by total US prescriptions in 2010 could
be derived from these structural precursors.1 Drugs such as
LipitorÒ (Atorvastatin) 1, CymbaltaÒ (Duloxetine) 2 and ZetiaÒ
(Ezetimibe) 3 have all been highly successful for their respective
pharmaceutical companies (Fig. 1).
O
N
H
S
F
1
2
OH
OH
Having an efficient and environmentally sound method of syn-
thesising these enantioselective alcohols for library protocols and
singleton synthesis is highly desirable. Often the initial medicinal
chemistry synthesis of a candidate pharmaceutical involves the
preparation of a racemate followed by separation of the enantio-
mers by chiral preparative HPLC. In many cases, this method is
not favoured for larger scale follow-up work, thus necessitating
the development of chemical asymmetric ketone reduction
processes.2
Biocatalytic processes continue to dominate the press due to
the success stories that follow their applications.3 The need for eco-
nomic, robust, scalable and reliable processes for the synthesis of
chiral APIs and intermediates has resulted in process chemists
tuning their skills at the interface of chemistry and biology and in-
deed embracing biocatalysts and biocatalytic processes in organic
N
F
O
F
3
Figure 1. A selection of drugs derived from chiral alcohol precursors.
synthesis.4 Enzymatic asymmetric reduction of prochiral ketones
using carbonyl reductase (CRED) enzymes is now well-
established tool for the efficient generation of chiral alcohols in
high enantiomeric excess with glucose or iso-propanol commonly
used as a stoichiometric reductant (Fig. 2).5,6
Although CRED technology has seen many applications in pro-
cess chemistry, scale-up and delivery of chiral product, it has seen
less uptake in medicinal chemistry labs. The research presented
herein is aimed at showing medicinal chemists the ease with
which commercially available CREDs can be used to bolster
a
⇑
Corresponding author.
E-mail address: andrew.rowan@almacgroup.com (A.S. Rowan).
0957-4166/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.tetasy.2013.09.015

1370
A. S. Rowan et al. / Tetrahedron: Asymmetry 24 (2013) 1369–1381
CREDs was screened against lead members of each substrate class
and the consistently high-performing enzymes identified as candi-
date CREDs for that class. The candidate CREDs for each substrate
class are shown in Figure 3 (selectAZyme™ CRED number).
Rapid microscale screens of the candidate CREDs against the
remaining members of each substrate class were then carried out
in order to identify those enzymes hitting the minimum perfor-
mance criteria desired for scale-up (P5% conversion to alcohol in
P95% ee). Excellent results were observed, with candidate enzyme
subsets taken from only six CREDs (A131, A161, A201, A231, A281
and A601) providing both Prelog and anti-Prelog hits for thirty-
three out of forty-one (80%) of the substrate ketones tested.
Considering that both enantiomers of each alcohol were targeted
individually, hits were found for an impressive sixty-nine of the
eighty-two target alcohols, an 84% hit rate. It was also observed
that the product was generated with exceptionally high enantio-
meric excess (P99% ee) for the majority of target alcohols (68%),
as illustrated in Figure 4.
Reduction
Recycle
Cofactor
NAD(P)H
NAD(P)
gluconate
(pH stat)
GDH
O
R1
R2
glucose
or
CRED
acetone
OH
*
R1
R2
CRED
iso-propyl alcohol
Figure 2. CRED catalysed bioreduction of prochiral ketones with cofactor recycling.
2.2. Preparative bioreductions
compound libraries with scaffolds of enantiopure chiral alcohol
building blocks, as well as providing a means to quickly generate
further amounts of these and similar alcohols when required. The
research has demonstrated that early access to enantiopure alco-
hols using CRED technology would obviate the need for chromato-
graphic or chemical resolution of racemic API, thus speeding
medicinal chemistry timelines from conception to reality.
Typically, a prochiral ketone substrate is screened against a
large number of CRED enzymes for a potential asymmetric biore-
duction and the reaction conditions for the best candidate(s) opti-
mised prior to running preparative reactions. Herein it was hoped
that the initial screening efforts could be reduced by narrowing the
number of CREDs to those most likely to provide the best (R)- and
(S)-selectivity for substrates belonging to particular structural clas-
ses. After subsequent microscale validation to confirm enzyme
activity and selectivity, these biocatalysts could be employed
directly in preparative reactions. In this manner it would be
possible to generate gram-scale libraries of enantiopure alcohols
quickly and efficiently, with the potential to up-scale without
further development.
Preparative bioreductions were carried out on thirty-five of the
ketones and gave excellent results, especially when taking into
consideration that these were one-time, un-optimised, reactions
(Table 1). Only four (6%) of the sixty-nine preparative reactions
attempted resulted in a product of less than 95.0% ee, with the
majority of the reactions (forty-six of sixty-nine, 66%) delivering
P99.0% ee and the remaining nineteen reactions (28%) generating
secondary alcohols in 95.0–99.0% ee. The yields were respectable
with an average of 68%. In cases where low yields were observed,
optimisation of the work-up procedure would likely improve the
yield considerably.
The absolute configurations of organic compounds can be deter-
mined by several methods including X-ray crystallography and the
modified Mosher’s NMR procedure.25 Herein, in cases where the
specific rotation of either enantiomer of alcohol product was not
known in the literature, the determination of enantiomeric config-
uration was conducted using vibrational circular dichroism26 (vide
infra) or, tentatively, by analogy to the selectivity observed for the
CRED employed against other members of the substrate class.
The vibrational circular dichroism (VCD) technique has played
an important role in the field of absolute structure determination,
particularly over the past decade. It is routinely used in the phar-
maceutical industry and research institutions mainly due to the
commercial availability of VCD instruments and the maturity of
the theoretical calculations used to support structure determina-
tions. Herein the absolute stereochemistries of eighteen secondary
alcohols were determined by this method, which compared the IR
and VCD spectra of enantioenriched samples with the correspond-
ing theoretically calculated spectra.27
2. Results and discussion
2.1. Carbonyl reductase screening
Structurally similar ketones, falling into seven structural clas-
ses, were selected based on their potential as precursors to
(R)- and (S)-chiral alcohol building blocks useful for target synthe-
sis and structure–activity relationship work within a medicinal
chemistry lab. A library of commercially available selectAZyme™
Set 2
(alpha-halo aryl-
ketones)
Set 3
Set 6
Set 7
Set 1
Set 4
Set 5
(fused bicyclics)
(alpha-branched
alkyl aryl-ketones)
(alpha-trifluoromethyl
aryl-ketones)
(methyl aryl-
ketones)
(beta-cyano
aryl-ketones)
(pyrrolidinone
analogues)
O
O
O
O
O
O
O
R1
X
CN
Ar
Ar
CF3
R
n
n
Ar
Ar
Me
Ar
N
R
R
R2
X = Cl or Br
A131
A161
A131
A131 (Prelog)
A131
A131
A161
A231
A281
A131
A161
A131
A161
A161
A231
A601
A161 (Anti-Prelog)
A601 (Anti-Prelog)
A161
A231
A601
A231 (Anti-Prelog)
A601
A231
A201 (Anti-Prelog)
A281 (Prelog)
Figure 3. Structural motifs of the substrate classes involved herein. High-performing carbonyl reductases associated with each class are given below in blue with their typical
selectivity given in parentheses.7

A. S. Rowan et al. / Tetrahedron: Asymmetry 24 (2013) 1369–1381
1371
Figure 4. Screening results of the best Prelog and anti-Prelog selective CREDs for each of the substrates screened. Screening was carried out with subsets of CREDs A131,
A161, A201, A231, A281 and A601, determined by substrate class (as given in Fig. 3). Prelog selectivity is denoted by positive ee values on the x-axis and anti-Prelog selectivity
by negative ee values.
2.3. 0.5 kg Scale up
4. Experimental
In order to demonstrate the scalability of the developed meth-
odology, a 0.5 kg scale reduction of ketone 15 was carried out using
CRED A161 in order to access chiral alcohol (S)-50 (Scheme 1). This
CRED was found to be able to utilise iso-propanol (IPA) for cofactor
regeneration, precluding the requirement for glucose dehydroge-
nase (GDH) and therefore the need for pH control. The reaction
profile is shown in Figure 5. The reduction was complete after
6 h and simple extraction with methyl tert-butyl ether (MTBE)
afforded (S)-50 in >99.0% ee and 91% isolated yield.
Process mass intensity (PMI) is a common green chemistry met-
ric, widely applied to enable the benchmarking of processes.28 It is
a measure of the mass of raw materials, reagents, solvents and so
on used to provide the mass of chemical synthesised, that is, kg
input/kg output. The calculation of the PMI for the bioreduction
of 15 to (S)-50 gave a value of 35, with the aqueous solvent
contributing 22 kg per kg product and organic solvent contributing
11 kg per kg product. These values compare favourably with
similar asymmetric reduction protocols.29
4.1. Chemicals and enzymes
Chemicals were purchased from Sigma Aldrich and Alfa Aesar
or, if not commercially available, synthesised using conventional
methodology. SelectAZyme™ CRED enzymes were purchased from
Almac.
4.2. Analytical methods
1H NMR spectra were recorded at 400 MHz on a Bruker AV-400
spectrometer; shifts are relative to internal TMS; J values are given
in Hz. [a]
values are given in 10ꢀ1 deg cm2 gꢀ1. Conversions and
D
enantiomeric excesses were measured by chiral stationary phase
HPLC using a Shimadzu system equipped with LC-10AS pumps, a
SIL-10AD VP auto injector and SPD-10A UV-VIS detector (see
Section 4.4 for method details).
4.3. General screening procedure
A typical screening procedure: Into a set of glass vials contain-
ing a selection of Almac CREDs (5–10 mg) were added potassium
phosphate buffer (pH 7.0, 1.7 mL), NADP or NAD (1–2 mg)
(depending on enzyme preference), glucose dehydrogenase (GDH,
3. Conclusion
Biocatalysis has proved to be a fast and efficient tool for the
generation of libraries of enantioenriched chiral alcohols with min-
imal screening and optimisation effort required to transform gram
amounts of their ketone precursors. VCD has been shown to be an
excellent tool for the determination of the absolute stereochemis-
try for small organic molecules in the solution state. Furthermore,
the successful asymmetric bioreduction of 0.5 kg of ketone 15 sup-
ports our belief that if larger quantities of the enantioenriched
alcohols presented or analogues from the same structural classes
are required in the future, the procedures described herein will
present an excellent basis for scale-up.
2 mg), a solution of glucose (75 mg) in buffer (300
lL) and finally
a solution of substrate ketone (20 mg) in DMSO (50–100
lL,
depending on solubility). For CREDs that had previously shown tol-
erance to iso-propanol, an additional reaction was run using this
cofactor recycle system, replacing GDH and glucose with iso-
propanol (300 lL). Reactions were shaken overnight at 30 °C and
then extracted with MTBE (2 mL). This was passed through a glass
pipette containing a cotton wool plug and anhydrous MgSO4
directly into an HPLC vial. The MTBE was evaporated in an oven

1372
A. S. Rowan et al. / Tetrahedron: Asymmetry 24 (2013) 1369–1381
Table 1
Results from preparative reactions
½ ꢁ
a 2D0
Substrate class
Set 1
Ketone
Alcohol product
CRED
A601
Experimental method
A
Yield (%)
88
ee (%)
99.5
Ref.
—
O
(R)-39
c
+42.5 (c 1.0, CHCl3)
(S)-39
c
A131
A
52
98.3
ꢀ38.0 (c 1.0, CHCl
3)
—
4
F
O
(R)-40b
(S)-40b
A601
A131
B
B
55
79
100
+43.5 (c 1.0, CHCl3)
ꢀ44.0 (c 1.0, CHCl3)
8
N
N
99.9
—
N
5
O
(R)-41c
(S)-41c
A601
A131
B
B
45
59
99.8
99.8
+30.3 (c 1.0, CHCl3)
ꢀ29.4 (c 1.0, CHCl3)
—
9
N
N
6
O
(R)-42c
(S)-42c
A601
A131
A
A
71
54
95.4
99.9
+65.7 (c 1.0, CHCl3)
ꢀ66.5 (c 1.0, CHCl3)
—
—
N
7
Cl
O
(R)-43d
(S)-43d
A161
A131
A
A
89
90
98.3
99.8
+29.4 (c 1.0, MeOH)
ꢀ29.5 (c 1.0, MeOH)
—
—
S
O
O
NH2
8
O
(R)-44c
(S)-44c
A161
A131
A
A
68
25
99.5
99.7
+89.0 (c 1.0, CHCl3)
ꢀ86.9 (c 1.0, CHCl3)
—
—
N
9
N
O
(R)-45c
(S)-45c
A161
A131
B then C
B
47
72
99.6
99.8
+36.4 (c 1.0, CHCl3)
ꢀ35.3 (c 1.0, CHCl3)
—
—
S
N
10
O
Set 2
(R)-46d
(S)-46d
A131
A601
B then C
B then C
54
49
98.8
96.9
ꢀ34.8 (c 1.0, CHCl3)
+33.2 (c 1.0, CHCl3)
—
—
Br
F3CO
11
O
(R)-47d
(S)-47d
A131
A161
A then B
A then B
26
24
99.6
99.2
Decomposed
Decomposed
—
—
Br
MeO
NC
12
F
O
(R)-48c
(S)-48c
A131
A601
A
B
25
42
99.9
97.6
ꢀ43.0 (c 1.0, CHCl3)
+43.5 (c 1.0, CHCl3)
—
—
Br
13
14
F
O
(R)-49a
(S)-49a
A131
A601
B
78
81
100
ꢀ38.6 (c 1.0, CHCl3)
+38.0 (c 1.0, CHCl3)
10
11
Cl
B then C
99.5
F

A. S. Rowan et al. / Tetrahedron: Asymmetry 24 (2013) 1369–1381
1373
Table 1 (continued)
½ ꢁ
a 2D0
Substrate class
Ketone
Alcohol product
CRED
Experimental method
Yield (%)
ee (%)
Ref.
O
(R)-50d
(S)-50d
A131
A161
B
B
93
91
99.6
99.9
ꢀ39.4 (c 1.0, CHCl3)
12
13
Cl
+40.2 (c 1.0, CHCl3)
15
Br
F
O
(R)-51a
(S)-51a
A131
A161
B
B
96
96
99.1
98.4
ꢀ54.6 (c 1.0, CHCl3)
+52.8 (c 1.0, CHCl3)
12
13
Cl
16
O
(R)-52a
(S)-52a
A131
A231
B
B
91
76
99.9
96.6
ꢀ60.3 (c 1.0, CHCl3)
+57.8 (c 1.0, CHCl3)
14
14
Cl
17
O
Set 3
(R)-53a
(S)-53a
A161
A131
B
A
78
76
99.8
98.3
ꢀ53.9 (c 1.0, CHCl3)
+51.7 (c 1.0, CHCl3)
15
—
N
18
O
(R)-54a
(S)-54a
A161
A131
B
A
57
44
99.1
93.5
+66.9 (c 1.0, CHCl3)
ꢀ65.1(c 1.0, CHCl3)
16
16
19
O
O
(R)-55c
(S)-55c
A161
A131
B
A
32
80
99.3
90.9
+4.0 (c 1.0, CHCl3)
ꢀ4.0 (c 1.0, CHCl3)
—
—
Br
20
O
Cl
(R)-56d
(S)-56d
A231
A281
A
A
91
62
99.0
99.3
+4.9 (c 1.0, CHCl3)
—
—
N
S
O
O
ꢀ5.7 (c 1.0, CHCl3)
21
O
(R)-57c
(S)-57c
A131
A231
A
A
78
93
95.9
96.0
ꢀ18.8 (c 1.0, CHCl3)
+17.6 (c 1.0, CHCl3)
—
—
O
22
O
(R)-58a
(S)-58a
A231
A131
A
B
70
79
96.3
99.5
ꢀ3.6 (c 1.0, CHCl3)
+2.4 (c 1.0, CHCl3)
17
18
Cl
N
23
O
(R)-59d
(S)-59d
A131
A231
B
A
99
92
99.8
98.4
+56.8 (c 1.0, CHCl3)
ꢀ55.4 (c 1.0, CHCl3)
—
—
S
O
O
24
O
(R)-60a
(S)-60a
A161
A131
B then C
B then C
93
93
97.7
99.4
+44.6 (c 1.0, CHCl3)
ꢀ46.3 (c 1.0, CHCl3)
19
20
N
Set 4
Br
25
O
(R)-61a
(S)-61a
A161
A131
B
A
92
80
99.6
100
+64.8 (c 1.0, CHCl3)
19
20
N
ꢀ61.3 (c 1.0, CHCl3)
26
(continued on next page)

1374
A. S. Rowan et al. / Tetrahedron: Asymmetry 24 (2013) 1369–1381
Table 1 (continued)
½ ꢁ
a 2D0
Substrate class
Ketone
Alcohol product
CRED
Experimental method
Yield (%)
ee (%)
Ref.
O
(R)-62c
(S)-62c
A161
A131
A
C
54
44
99.8
99.6
+11.5 (c 0.2, CHCl3)
ꢀ10.6 (c 0.2, CHCl3)
—
—
N
N
27
O
Set 5
(R)-63c
(S)-63c
A201
A131
A
B
39
31
94.8
99.2
ꢀ44.5 (c 1.0, CHCl3)
+44.4 (c 1.0, CHCl3)
—
—
N
N
N
28
O
O
(R)-64c
(S)-64c
A161
A131
A
B
56
56
95.6
99.9
ꢀ29.3 (c 1.0, CHCl3)
+34.1 (c 1.0, CHCl3)
—
—
N
N
N
29
30
(R)-65
(S)-65c
No hit
A131
—
B
—
—
—
—
—
N
N
40
99.9
+68.5 (c 1.0, CHCl3)
O
Set 6
(R)-66c
(S)-66c
A231
A131
A
A
60
58
99.4
99.0
ꢀ36.3 (c 1.0, CHCl3)
+32.1 (c 1.0, CHCl3)
—
—
Br
N
31
O
(R)-67c
(S)-67c
A601
A131
A
B
52
54
99.7
99.9
ꢀ36.2 (c 1.0, CHCl3)
+32.7 (c 1.0, CHCl3)
—
—
32
O
(R)-68c
(S)-68c
A161
A131
B
B
93
94
99.7
98.7
+52.2 (c 1.0, CHCl3)
ꢀ52.0 (c 1.0, CHCl3)
—
—
N
33
O
(R)-69c
(S)-69c
A161
A131
A
B
58
76
99.5
97.7
+30.6 (c 1.0, CHCl3)
ꢀ31.6 (c 1.0, CHCl3)
—
—
Cl
34
O
F
Set 7
(R)-70c
(S)-70c
A131
A231
C
A
71
89
99.9
99.9
ꢀ14.8 (c 1.0, CHCl3)
+12.9 (c 1.0, CHCl3)
—
—
CF3
N
Me
35
O
O
(R)-71a,c
(S)-71a,c
A131
A161
B
B
68
65
99.8
98.7
ꢀ28.7 (c 1.0, CHCl3)
+26.1 (c 1.0, CHCl3)
21
22
CF3
CF3
Cl
Cl
36
(R)-72a
(S)-72a
A131
A161
B
C
89
81
95.3
99.6
ꢀ24.5 (c 1.0, CHCl3)
+23.1 (c 1.0, CHCl3)
23
—
37

A. S. Rowan et al. / Tetrahedron: Asymmetry 24 (2013) 1369–1381
1375
Table 1 (continued)
½ ꢁ
a 2D0
Substrate class
Ketone
Alcohol product
CRED
Experimental method
Yield (%)
ee (%)
Ref.
O
(R)-73a
(S)-73a
A131
A161
C
C
90
78
99.7
93.4
ꢀ34.6 (c 1.0, MeOH)
+24.5 (c 1.0, MeOH)
24
—
CF3
HO2C
38
Assigned by comparison to literature ½a D20
ꢁ
.
a
Assigned by comparison to ½a D20
ꢁ
of the commercially available compound.
b
c
Assigned (or assignment corroborated) by VCD analysis.
Assigned by comparison to selectivities achieved for other members of this compound set.
d
OH
O
shaken at room temperature overnight and the reaction progress
was checked by 1H NMR. If not complete, additional CRED (100–
200 mg) and NADP or NAD (10 mg) were added and shaking was
continued. This was repeated until the reaction reached comple-
tion. If the reaction appeared to stall or proceed very slowly, exper-
imental method C was used. Method C: Almac CRED (200 mg) and
NADP or NAD (10 mg) were measured into a 250 mL round-
bottomed flask then dissolved in 0.1 M potassium phosphate buf-
fer (pH 7, ca. 50 mL). IPA (7 mL) was added, followed by a solution
of ketone (900–1700 mg) in DMSO (2.5–5 mL, depending on solu-
bility). This was stirred at 35 °C under 500 mbar reduced pressure
to aid removal of acetone formed by IPA oxidation. Standard
work-up procedure: The pH of the reaction mixture was adjusted
if necessary (basic or acidic depending on estimated pKa of alcohol
product) with either 1 M NaOH or 1 M HCl. The reaction mixture
was then extracted with MTBE (3 ꢂ 100 mL). The organic solvent
was washed with brine (ca. 100 mL), dried over anhydrous magne-
sium sulfate, filtered and concentrated in vacuo to afford the
desired alcohol. If recovery of the alcohol was low, the aqueous
phase was re-extracted with EtOAc (3 ꢂ 100 mL), washed with
brine (ca. 100 mL), dried (MgSO4), filtered and concentrated in
vacuo, then combined with the alcohol recovered from the MTBE
extraction. If the reaction did not go to >95% completion, or the
purity of the isolated alcohol was <95% by NMR, flashmaster silica
purification was carried out to afford a clean alcohol. Analysis of
the enantiomeric excess was carried out by chiral HPLC.
CRED A161
NAD
Cl
Cl
IPA
Br
Br
0.1 M KH2PO4 (pH 7)
15
50
(S)-
Scheme 1. Enzymatic reduction of ketone 15 by CRED A161 to afford chiral alcohol
(S)-50.
Figure 5. Reaction profile for the CRED A161-catalysed asymmetric reduction of
para-bromophenacyl chloride 15.
4.4.1. 1-(3-Fluoro-4-methylphenyl)ethanol 39
Analysis of the enantiomeric excess was performed on a Chiral-
cel OB-H column with eluent 98:2 hexane/IPA, flow rate
1 mL minꢀ1, detection 254 nm and run time 22 min; retention
times for (S)-39 and (R)-39 were 14.3 and 17.4 min, respectively.
and the residue analysed by chiral HPLC after re-dissolving in
mobile phase.
(S)-39. Pale yellow oil (523 mg, 52%). ½a D20
¼ ꢀ38:0 (c 1.0, CHCl3);
ꢁ
4.4. Small scale preparative scale-ups
1H NMR (CDCl3, 400 MHz): d 7.06 (1H, m, Ar-H), 6.94 (2H, m,
2 ꢂ Ar-H), 4.76 (1H, m, CH-OH), 2.17 (3H, s, Ar-CH3), 1.38 (3H, d,
J = 6.5 Hz, CH-CH3); ee = 98.3%. (R)-39. Pale yellow oil (892 mg,
Preparative experimental method A: Almac CRED (200 mg),
NADP or NAD (10 mg), GDH (20 mg) and glucose (1.5 equiv) were
measured into a 100 mL round bottomed flask then dissolved in
0.1 M potassium phosphate buffer (pH 7, ca. 50 mL) and stirred
at 30 °C. Next, a solution of ketone (900–1700 mg) in DMSO
(2.5–5 mL, depending on solubility) was added to the reaction
and this was allowed to stir overnight under pH-stat control (pH
7.0, adjusted with 1 M NaOH solution). The following day, reaction
progress was checked by 1H NMR. If not complete, additional CRED
(100–200 mg), NADP or NAD (10 mg) and GDH (10 mg) were
added and stirring was continued. This was repeated until the reac-
tion reached completion or had stalled and would not go any fur-
ther. Method B: Almac CRED (200 mg) and NADP or NAD (10 mg)
were charged into a 100 mL round bottomed flask and dissolved
in 0.1 M potassium phosphate buffer (pH 7, ca. 50 mL). Next, IPA
88%). ½a 2D0
ꢁ
¼ þ42:5 (c 1.0, CHCl3); 1H NMR (CDCl3, 400 MHz): d
7.06 (1H, m, Ar-H), 6.94 (2H, m, 2 ꢂ Ar-H), 4.77 (1H, m, CH-OH),
2.18 (3H, s, Ar-CH3), 1.39 (3H, d, J = 6.4 Hz, CH-CH3); ee = 99.5%.
4.4.2. 1-(4-(1H-1,2,4-Triazol-1-yl)phenyl)ethanol 40
Analysis of the enantiomeric excess was performed on a Chiral-
cel OJ column with eluent 6:4 hexane/IPA + 0.1% DEA, flow rate
1 mL minꢀ1, detection 254 nm and run time 10 min; retention
times for (S)-40 and (R)-40 were 7.0 and 8.7 min, respectively.
(S)-40. White solid (916 mg, 79%). ½a D20
ꢁ
¼ ꢀ44:0 (c 1.0, CHCl3); 1H
NMR (CDCl3, 400 MHz): d 8.54 (1H, s, triazole-H), 8.10 (1H, s, tria-
zole-H), 7.66 (2H, m, 2 ꢂ Ar-H), 7.52 (2H, m, 2 ꢂ Ar-H), 4.99 (1H, m,
CH-OH), 1.53 (3H, d, J = 6.5 Hz, CH3); ee = 99.9%. (R)-40. White solid
(640 mg, 55%). ½a D20
ꢁ
¼ þ43:5 (c 1.0, CHCl3); 1H NMR (CDCl3,
(7 mL) was added, followed by
a solution of ketone (900–
400 MHz): d 8.54 (1H, s, triazole-H), 8.10 (1H, s, triazole-H), 7.66
1700 mg) in DMSO (2.5–5 mL, depending on solubility). This was

1376
A. S. Rowan et al. / Tetrahedron: Asymmetry 24 (2013) 1369–1381
(2H, m, 2 ꢂ Ar-H), 7.53 (2H, m, 2 ꢂ Ar-H), 4.99 (1H, m, CH-OH),
1.53 (3H, d, J = 6.4 Hz, CH3); ee = 100%.
(1H, s, thiazole-H), 5.22 (1H, m, CH-OH), 2.44 (3H, s, thiazole-
CH3), 1.56 (3H, d, J = 6.4 Hz, CH-CH3); ee = 99.6%.
4.4.3. 1-(Pyrazin-2-yl)ethanol 41
4.4.8. 2-Bromo-1-(4-(trifluoromethoxy)phenyl)ethanol 46
Analysis of the enantiomeric excess was performed on a Chir-
alpak AS-H column with eluent 9:1 hexane/IPA + 0.1% DEA, flow
rate 1 mL minꢀ1, detection 254 nm and run time 7 min; retention
times for (S)-41 and (R)-41 were 5.6 and 5.0 min, respectively.
Analysis of the enantiomeric excess was performed on a Chir-
alpak IA column with eluent 9:1 hexane/IPA, flow rate 1 mL minꢀ1
,
detection 254 nm and run time 20 min; retention times for (S)-46
and (R)-46 were 6.5 and 6.0 min, respectively. (S)-46. Pale yellow
(S)-41. Pale yellow oil (656 mg, 59%). ½a D20
ꢁ
¼ ꢀ29:4 (c 1.0, CHCl3);
oil (652 mg, 49%). ½a D20
ꢁ
¼ þ33:2 (c 1.0, CHCl3); 1H NMR (CDCl3,
1H NMR (CDCl3, 400 MHz): d 8.68 (1H, s, Ar-H), 8.52 (2H, m,
2 ꢂ Ar-H), 5.00 (1H, m, CH-OH), 1.57 (3H, d, J = 6.6 Hz, CH3);
400 MHz): d 7.43 (2H, m, 2 ꢂ Ar-H), 7.24 (2H, m, 2 ꢂ Ar-H), 4.95
(1H, m, CH-OH), 3.64 (1H, dd, J = 10.5, 3.4 Hz, CHAHB), 3.52 (1H,
dd, J = 10.5, 8.9 Hz, CHAHB); ee = 96.9%. (R)-46. Pale yellow oil
ee = 99.8%. (R)-41. Pale yellow oil (502 mg, 45%). ½a D20
¼ þ30:3 (c
ꢁ
1.0, CHCl3); 1H NMR (CDCl3, 400 MHz): d 8.68 (1H, s, Ar-H), 8.52
(2H, m, 2 ꢂ Ar-H), 5.00 (1H, m, CH-OH), 1.57 (3H, d, J = 6.6 Hz,
CH3); ee = 99.8%.
(707 mg, 54%). ½a D20
ꢁ
¼ ꢀ34:8 (c 1.0, CHCl3); 1H NMR (CDCl3,
400 MHz): d 7.43 (2H, m, 2 ꢂ Ar-H), 7.24 (2H, m, 2 ꢂ Ar-H), 4.95
(1H, m, CH-OH), 3.64 (1H, dd, J = 10.5, 3.3 Hz, CHAHB), 3.52 (1H,
dd, J = 10.5, 8.9 Hz, CHAHB); ee = 98.8%.
4.4.4. 1-(3-Chloropyridin-4-yl)ethanol 42
Analysis of the enantiomeric excess was performed on a Chir-
alpak IA column with eluent 95:5 hexane/IPA + 0.1% DEA, flow rate
1 mL minꢀ1, detection 254 nm and run time 14 min; retention
times for (S)-42 and (R)-42 were 11.9 and 10.3 min, respectively.
4.4.9. 2-Bromo-1-(3-fluoro-4-methoxyphenyl)ethanol 47
Analysis of the enantiomeric excess was performed on a Chir-
alpak AD-H column with eluent 95:5 hexane/IPA, flow rate
1 mL minꢀ1, detection 254 nm and run time 25 min; retention
times for (S)-47 and (R)-47 were 18.2 and 21.1 min, respectively.
Optical rotation measurements were not made due to product deg-
radation. (S)-47. Pale yellow oil (360 mg, 24%). 1H NMR (CDCl3,
400 MHz): d 7.12 (2H, m, 2 ꢂ Ar-H), 6.95 (1H, t, J = 8.4 Hz, Ar-H),
4.87 (1H, m, CH-OH), 3.89 (3H, s, OCH3), 3.60 (1H, dd, J = 10.4,
3.4 Hz, CHAHB), 3.50 (1H, dd, J = 10.5, 8.8 Hz, CHAHB); ee = 99.2%.
(R)-47. Colourless oil (340 mg, 26%). 1H NMR (CDCl3, 400 MHz): d
7.12 (2H, m, 2 ꢂ Ar-H), 6.95 (1H, t, J = 8.5 Hz, Ar-H), 4.87 (1H, m,
CH-OH), 3.89 (3H, s, OCH3), 3.60 (1H, dd, J = 10.4, 3.4 Hz, CHAHB),
3.50 (1H, dd, J = 10.5, 8.8 Hz, CHAHB); ee = 99.6%.
(S)-42. Yellow oil (926 mg, 54%). ½a D20
ꢁ
¼ ꢀ66:5 (c 1.0, CHCl3); 1H
NMR (CDCl3, 400 MHz): d 8.27 (2H, m, 2 ꢂ Ar-H), 7.50 (1H, d,
J = 5.0 Hz, Ar-H), 5.11 (1H, m, CH-OH), 1.38 (3H, d, J = 6.5 Hz,
CH3); ee = 99.9%. (R)-42. Yellow oil (800 mg, 71%). ½a D20
¼ þ65:7
ꢁ
(c 1.0, CHCl3); 1H NMR (CDCl3, 400 MHz): d 8.47 (2H, m, 2 ꢂ
Ar-H), 7.56 (1H, d, J = 5.0 Hz, Ar-H), 5.23 (1H, m, CH-OH), 1.50
(3H, d, J = 6.5 Hz, CH3); ee = 95.4%.
4.4.5. 3-(1-Hydroxyethyl)benzenesulfonamide 43
Analysis of the enantiomeric excess was performed on a Chir-
alpak AD-H column with eluent 93:7 hexane/EtOH, flow rate
1 mL minꢀ1, detection 210 nm and run time 110 min; retention
times for (S)-43 and (R)-43 were 91.5 and 83.6 min, respectively.
4.4.10. 3-(2-Bromo-1-hydroxyethyl)benzonitrile 48
Analysis of the enantiomeric excess was performed on a Chir-
alpak AS-H column with eluent 8:2 hexane/IPA, flow rate
1 mL minꢀ1, detection 254 nm and run time 8 min; retention times
for (S)-48 and (R)-48 were 4.5 and 5.3 min, respectively. (S)-48.
(S)-43. Yellow oil (1275 mg, 90%). ½a D20
ꢁ
¼ ꢀ29:5 (c 1.0, MeOH); 1H
NMR (DMSO-d6, 400 MHz): d 7.91 (1H, s, Ar-H), 7.74 (1H, m,
Ar-H), 7.61–7.54 (2H, m, 2 ꢂ Ar-H), 4.86 (1H, m, CH-OH), 1.40
(3H, d, J = 6.4 Hz, CH3); ee = 99.8%. (R)-43. Yellow oil (1182 mg,
Colourless oil (609 mg, 42%). ½a D20
ꢁ
¼ þ43:5 (c 1.0, CHCl3); 1H NMR
89%). ½a 2D0
ꢁ
¼ þ29:4 (c 1.0, MeOH); 1H NMR (DMSO-d6, 400 MHz):
(CDCl3, 400 MHz): d 7.73 (1H, s, Ar-H), 7.63 (2H, m, 2 ꢂ Ar-H),
7.50 (1H, m, Ar-H), 4.98 (1H, m, CH-OH), 3.65 (1H, dd, J = 10.6,
3.5 Hz, CHAHB), 3.51 (1H, dd, J = 10.6, 8.4 Hz, CHAHB); ee = 97.6%.
d 7.85 (1H, s, Ar-H), 7.68 (1H, m, Ar-H), 7.55–7.48 (2H, m, 2 ꢂ
Ar-H), 4.81 (1H, m, CH-OH), 1.35 (3H, d, J = 6.4 Hz, CH3); ee = 98.3%.
(R)-48. Pale yellow oil (253 mg, 25%). ½a D20
¼ ꢀ43:0 (c 1.0, CHCl3);
ꢁ
4.4.6. 1-(3-Methylpyrazin-2-yl)ethanol 44
1H NMR (CDCl3, 400 MHz): d 7.73 (1H, s, Ar-H), 7.63 (2H, m,
2 ꢂ Ar-H), 7.51 (1H, m, Ar-H), 4.98 (1H, m, CH-OH), 3.65 (1H, dd,
J = 10.6, 3.5 Hz, CHAHB), 3.51 (1H, dd, J = 10.6, 8.5 Hz, CHAHB);
ee = 99.9%.
Analysis of the enantiomeric excess was performed on a Chir-
alpak AD-H column with eluent 9:1 hexane/IPA + 0.1% DEA, flow
rate 1 mL minꢀ1, detection 254 nm and run time 11 min; retention
times for (S)-44 and (R)-44 were 8.0 and 7.3 min, respectively.
(S)-44. Pale yellow oil (349 mg, 25%). ½a D20
ꢁ
¼ ꢀ86:9 (c 1.0, CHCl3);
4.4.11. 2-Chloro-1-(2,4-difluorophenyl)ethanol 49
1H NMR (CDCl3, 400 MHz): d 8.41 (1H, d, J = 2.6 Hz, Ar-H), 8.37
(1H, d, J = 2.4 Hz, Ar-H), 5.03 (1H, m, CH-OH), 2.58 (3H, s, Ar-
CH3), 1.45 (3H, d, J = 6.4 Hz, CH-CH3); ee = 99.7%. (R)-44. Brown
Analysis of the enantiomeric excess was performed on a Chir-
alpak AD-H column with eluent 95:5 hexane/IPA, flow rate
1 mL minꢀ1, detection 254 nm and run time 15 min; retention
times for (S)-49 and (R)-49 were 10.2 and 9.0 min, respectively.
oil (907 mg, 68%). ½a D20
ꢁ
¼ þ89:0 (c 1.0, CHCl3); 1H NMR (CDCl3,
400 MHz): d 8.41 (1H, d, J = 2.5 Hz, Ar-H), 8.37 (1H, d, J = 2.4 Hz,
Ar-H), 5.04 (1H, m, CH-OH), 2.58 (3H, s, Ar-CH3), 1.45 (3H, d,
J = 6.4 Hz, CH-CH3); ee = 99.5%.
(S)-49. Colourless oil (1140 mg, 81%). ½a D20
¼ þ38:0 (c 1.0, CHCl3);
ꢁ
1H NMR (CDCl3, 400 MHz): d 7.53 (1H, m, Ar-H), 6.92 (1H, m,
Ar-H), 6.81 (1H, m, Ar-H), 5.18 (1H, m, CH-OH), 3.81 (1H, dd,
J = 11.3, 3.3 Hz, CHAHB), 3.62 (1H, dd, J = 11.2, 8.4 Hz, CHAHB);
4.4.7. 1-(4-Methylthiazol-5-yl)ethanol 45
ee = 99.5. (R)-49. Pale yellow oil (1070 mg, 78%). ½a D20
¼ ꢀ38:6 (c
ꢁ
Analysis of the enantiomeric excess was performed on a Chir-
alpak AS-H column with eluent 98:2 hexane/IPA + 0.1% DEA, flow
rate 1 mL minꢀ1, detection 254 nm and run time 24 min; retention
times for (S)-45 and (R)-45 were 21.4 and 18.4 min, respectively.
1.0, CHCl3); 1H NMR (CDCl3, 400 MHz): d 7.53 (1H, m, Ar-H), 6.94
(1H, m, Ar-H), 6.81 (1H, m, Ar-H), 5.18 (1H, m, CH-OH), 3.81 (1H,
dd, J = 11.2, 3.3 Hz, CHAHB), 3.62 (1H, dd, J = 11.1, 8.3 Hz, CHAHB);
ee = 100.
(S)-45. Pale yellow oil (762 mg, 72%). ½a D20
¼ ꢀ35:3 (c 1.0, CHCl3);
ꢁ
1H NMR (CDCl3, 400 MHz): d 8.59 (1H, s, thiazole-H), 5.21 (1H,
m, CH-OH), 2.41 (3H, s, thiazole-CH3), 1.56 (3H, d, J = 6.4 Hz,
CH-CH3); ee = 99.8%. (R)-45. Pale yellow oil (505 mg, 47%).
4.4.12. 1-(4-Bromophenyl)-2-chloroethanol 50
Analysis of the enantiomeric excess was performed on a Chiral-
cel OJ column with eluent 9:1 hexane/IPA, flow rate 1 mL minꢀ1
,
½
a 2D0
ꢁ
¼ þ36:4 (c 1.0, CHCl3); 1H NMR (CDCl3, 400 MHz): d 8.61
detection 254 nm and run time 16 min; retention times for

A. S. Rowan et al. / Tetrahedron: Asymmetry 24 (2013) 1369–1381
1377
(S)-50 and (R)-50 were 13.6 and 12.3 min, respectively. (S)-50.
4.4.17. 5-Bromo-1,2,3,4-tetrahydronaphthalen-1-ol 55
White solid (1105 mg, 91%). ½a D20
ꢁ
¼ þ40:2 (c 1.0, CHCl3); 1H NMR
Analysis of the enantiomeric excess was performed on a Chir-
alpak IB column with eluent 95:5 hexane/IPA, flow rate
1 mL minꢀ1, detection 254 nm and run time 10 min; retention
times for (S)-55 and (R)-55 were 7.6 and 8.3 min, respectively.
(CDCl3, 400 MHz): d 7.51 (2H, m, 2 ꢂ Ar-H), 7.27 (2H, m, 2 ꢂ
Ar-H), 4.88 (1H, m, CH-OH), 3.72 (1H, dd, J = 11.3, 3.5 Hz, CHAHB),
3.60 (1H, dd, J = 11.2, 8.6 Hz, CHAHB); ee = 99.9%. (R)-50. White
solid (1131 mg, 93%). ½a D20
ꢁ
¼ ꢀ39:4 (c 1.0, CHCl3); 1H NMR (CDCl3,
(S)-55. White solid (806 mg, 80%). ½a D20
ꢁ
¼ ꢀ4:0 (c 1.0, CHCl3); 1H
400 MHz): d 7.51 (2H, m, 2 ꢂ Ar-H), 7.27 (2H, m, 2 ꢂ Ar-H), 4.87
(1H, m, CH-OH), 3.72 (1H, dd, J = 11.2, 3.4 Hz, CHAHB), 3.60 (1H,
dd, J = 11.2, 8.6 Hz, CHAHB); ee = 99.6%.
NMR (CDCl3, 400 MHz): d 7.48 (1H, m, Ar-H), 7.41 (1H, m, Ar-H),
7.08 (1H, t, J = 7.8 Hz, Ar-H), 4.77 (1H, m, CH-OH), 2.85 (1H, m,
CHAHB), 2.67 (1H, m, CHAHB), 2.04-1.79 (4H, m, 2 ꢂ CH2);
ee = 90.9%. (R)-55. White solid (397 mg, 32%). ½a D20
¼ þ4:0 (c 1.0,
ꢁ
4.4.13. 2-Chloro-1-(4-fluorophenyl)ethanol 51
CHCl3); 1H NMR (CDCl3, 400 MHz): d 7.48 (1H, m, Ar-H), 7.42
(1H, d, J = 7.7 Hz, Ar-H), 7.09 (1H, t, J = 7.8 Hz, Ar-H), 4.78 (1H, m,
CH-OH), 2.84 (1H, m, CHAHB), 2.68 (1H, m, CHAHB), 2.04–1.80
(4H, m, 2 ꢂ CH2); ee = 99.3%.
Analysis of the enantiomeric excess was performed on a Chiral-
cel OB-H column with eluent 95:5 hexane/IPA, flow rate
1 mL minꢀ1, detection 210 nm and run time 17 min; retention
times for (S)-51 and (R)-51 were 13.7 and 12.0 min, respectively.
(S)-51. Dark yellow oil (1175 mg, 96%). ½a D20
ꢁ
¼ þ52:8 (c 1.0, CHCl3);
4.4.18. 6-Chloro-1-(methylsulfonyl)-1,2,3,4-tetrahydroquinolin-
4-ol 56
Analysis of the enantiomeric excess was performed on a Chir-
alpak AD-H column with eluent 9:1 hexane/IPA, flow rate
1 mL minꢀ1, detection 254 nm and run time 27 min; retention
times for (S)-56 and (R)-56 were 19.1 and 16.8 min, respectively.
1H NMR (CDCl3, 400 MHz): d 7.37 (2H, m, 2 ꢂ Ar-H), 7.07 (2H, m,
2 ꢂ Ar-H), 4.89 (1H, m, CH-OH), 3.72 (1H, dd, J = 11.3, 3.5 Hz,
CHAHB), 3.62 (1H, dd, J = 11.2, 8.7 Hz, CHAHB); ee = 98.4%. (R)-51.
Dark yellow oil (1185 mg, 96%). ½a D20
ꢁ
¼ ꢀ54:6 (c 1.0, CHCl3); 1H
NMR (CDCl3, 400 MHz): d 7.37 (2H, m, 2 ꢂ Ar-H), 7.07 (2H, m,
2 ꢂ Ar-H), 4.89 (1H, m, CH-OH), 3.72 (1H, dd, J = 11.2, 3.5 Hz,
CHAHB), 3.62 (1H, dd, J = 11.2, 8.7 Hz, CHAHB); ee = 99.1%.
(S)-56. White solid (766 mg, 62%). ½a D20
ꢁ
¼ ꢀ5:7 (c 1.0, CHCl3); 1H
NMR (CDCl3, 400 MHz): d 7.73 (1H, d, J = 9.0 Hz, Ar-H), 7.40 (1H,
m, Ar-H), 7.26 (1H, m, Ar-H), 4.80 (1H, m, CH-OH), 4.02 (1H, m,
CHAHB), 3.77 (1H, m, CHAHB), 2.92 (3H, s, CH3), 2.10 (2H, m, CH2);
4.4.14. 2-Chloro-1-phenylethanol 52
Analysis of the enantiomeric excess was performed on a Chir-
alpak AS-H column with eluent 95:5 hexane/IPA, flow rate
1 mL minꢀ1, detection 254 nm and run time 13 min; retention
times for (S)-52 and (R)-52 were 10.3 and 11.2 min, respectively.
ee = 99.3%. (R)-56. White solid (1257 mg, 91%). ½a D20
¼ þ4:9 (c 1.0,
ꢁ
CHCl3); 1H NMR (CDCl3, 400 MHz): d 7.73 (1H, d, J = 8.9 Hz,
Ar-H), 7.40 (1H, m, Ar-H), 7.26 (1H, m, Ar-H), 4.79 (1H, m,
CH-OH), 4.02 (1H, m, CHAHB), 3.77 (1H, m, CHAHB), 2.92 (3H,
s, CH3), 2.10 (2H, m, CH2); ee = 99.0%.
(S)-52. Pale yellow oil (925 mg, 76%). ½a D20
¼ þ57:8 (c 1.0, CHCl3);
ꢁ
1H NMR (CDCl3, 400 MHz): d 7.40–7.30 (5H, m, 5 ꢂ Ar-H), 4.91
(1H, m, CH-OH), 3.75 (1H, dd, J = 11.2, 3.4 Hz, CHAHB), 3.65 (1H,
dd, J = 11.2, 8.8 Hz, CHAHB); ee = 96.6%. (R)-52. Pale yellow oil
4.4.19. Isochroman-4-ol 57
Analysis of the enantiomeric excess was performed on a Chir-
alpak IA column with eluent 95:5 hexane/IPA, flow rate
1 mL minꢀ1, detection 254 nm and run time 18 min; retention
times for (S)-57 and (R)-57 were 14.0 and 15.1 min, respectively.
(1111 mg, 91%). ½a D20
ꢁ
¼ ꢀ60:3 (c 1.0, CHCl3); 1H NMR (CDCl3,
400 MHz): d 7.41-7.30 (5H, m, 5 ꢂ Ar-H), 4.91 (1H, m, CH-OH),
3.75 (1H, dd, J = 11.2, 3.4 Hz, CHAHB), 3.65 (1H, dd, J = 11.2,
8.8 Hz, CHAHB); ee = 99.9%.
(S)-57. Yellow oil (1023 mg, 93%). ½a D20
ꢁ
¼ þ17:6 (c 1.0, CHCl3); 1H
NMR (CDCl3, 400 MHz) d 7.45 (1H, m, Ar-H), 7.28 (2H, m, 2 ꢂ
Ar-H), 6.99 (1H, m, Ar-H), 4.75–4.63 (2H, m, CH2), 4.54 (1H, m,
CH-OH), 4.10 (1H, dd, J = 11.9, 2.7 Hz, CHAHB), 3.87 (1H, dd,
J = 11.8, 2.7 Hz, CHAHB); ee = 96.0%. (R)-57. Yellow oil (994 mg,
4.4.15. 5,6,7,8-Tetrahydroisoquinolin-5-ol 53
Analysis of the enantiomeric excess was performed on a Chir-
alpak AD-H column with eluent 95:5 hexane/IPA + 0.1% DEA, flow
rate 1 mL minꢀ1, detection 254 nm and run time 28 min; retention
times for (S)-53 and (R)-53 were 21.9 and 23.9 min, respectively.
78%). ½a 2D0
ꢁ
¼ ꢀ18:8 (c 1.0, CHCl3); 1H NMR (CDCl3, 400 MHz) d
7.45 (1H, m, Ar-H), 7.28 (2H, m, 2 ꢂ Ar-H), 7.00 (1H, m, Ar-H), 4.81–
4.65 (2H, m, CH2), 4.55 (1H, m, CH-OH), 4.11 (1H, dd, J =
12.0, 2.8 Hz, CHAHB), 3.87 (1H, dd, J = 11.9, 2.7 Hz, CHAHB);
ee = 95.9%.
(S)-53. Pale orange solid (951 mg, 76%). ½a D20
¼ þ51:7 (c 1.0,
ꢁ
CHCl3); 1H NMR (CDCl3, 400 MHz): d 8.39 (1H, d, J = 5.1, Ar-H),
8.34 (1H, s, Ar-H), 7.39 (1H, d, J = 5.0 Hz, Ar-H), 4.74 (1H, m, CH-
OH), 2.76 (2H, m, CH2), 2.15–1.76 (4H, m, 2 ꢂ CH2); ee = 98.3%.
(R)-53. Pale orange solid (921 mg, 78%). ½a D20
ꢁ
¼ ꢀ53:9 (c 1.0,
4.4.20. 2-Chloro-6,7-dihydro-5H-cyclopenta[b]pyridin-7-ol 58
Analysis of the enantiomeric excess was performed on a Chiral-
cel OB-H column with eluent 3:2 hexane/IPA + 0.1% DEA, flow rate
1 mL minꢀ1, detection 254 nm and run time 9 min; retention times
for (S)-58 and (R)-58 were 6.4 and 5.0 min, respectively. (S)-58.
CHCl3); 1H NMR (CDCl3, 400 MHz): d 8.38 (1H, d, J = 5.1, Ar-H),
8.33 (1H, s, Ar-H), 7.39 (1H, d, J = 5.1 Hz, Ar-H), 4.74 (1H, m, CH-
OH), 2.76 (2H, m, CH2), 2.12 (1H, m, CHAHB), 2.00 (1H, m, CHAHB),
1.81 (2H, m, CH2); ee = 99.8%.
Pale yellow oil (991 mg, 79%). ½a D20
ꢁ
¼ þ2:4 (c 1.0, CHCl3); 1H NMR
4.4.16. Chroman-4-ol 54
(CDCl3, 400 MHz) d 7.53 (1H, d, J = 8.0 Hz, Ar-H), 7.19 (1H, d,
J = 8.0 Hz, Ar-H), 5.20 (1H, m, CH-OH), 3.02 (1H, m, CHAHB), 2.81
(1H, m, CHAHB), 2.57 (1H, m, CHCHD), 2.08 (1H, m, CHCHD);
Analysis of the enantiomeric excess was performed on a Chir-
alpak AS-H column with eluent 85:15 hexane/IPA, flow rate
1 mL minꢀ1, detection 254 nm and run time 8 min; retention times
for (S)-54 and (R)-54 were 3.6 and 4.3 min, respectively. (S)-54.
ee = 99.5%. (R)-58. Pale yellow oil (948 mg, 70%). ½a D20
¼ ꢀ3:6 (c
ꢁ
1.0, CHCl3); 1H NMR (CDCl3, 400 MHz) d 7.53 (1H, d, J = 8.0 Hz,
Ar-H), 7.18 (1H, d, J = 8.0 Hz, Ar-H), 5.20 (1H, m, CH-OH), 3.02
(1H, m, CHAHB), 2.81 (1H, m, CHAHB), 2.57 (1H, m, CHCHD), 2.09
(1H, m, CHCHD); ee = 96.3%.
White solid (594 mg, 44%). ½a D20
ꢁ
¼ ꢀ65:1 (c 1.0, CHCl3); 1H NMR
(CDCl3, 400 MHz): d 7.32 (1H, m, Ar-H), 7.21 (1H, m, Ar-H), 6.93
(1H, m, Ar-H), 6.85 (1H, m, Ar-H), 4.80 (1H, m, CH-OH), 4.28 (2H,
m, CH2), 2.13 (1H, m, CHAHB), 2.04 (1H, m, CHAHB); ee = 93.5%.
(R)-54. White solid (758 mg, 57%). ½a D20
ꢁ
¼ þ66:9 (c 1.0, CHCl3); 1H
4.4.21. 3-Hydroxy-2,3-dihydrobenzo[b]thiophene 1,1-dioxide 59
Analysis of the enantiomeric excess was performed on a
Chiralpak AS-H column with eluent 3:2 hexane/IPA, flow rate
1 mL minꢀ1, detection 254 nm and run time 18 min; retention
NMR (CDCl3, 400 MHz): d 7.31 (1H, m, Ar-H), 7.21 (1H, m, Ar-H),
6.93 (1H, m, Ar-H), 6.85 (1H, m, Ar-H), 4.80 (1H, m, CH-OH), 4.28
(2H, m, CH2), 2.13 (1H, m, CHAHB), 2.04 (1H, m, CHAHB); ee = 99.1%.

1378
A. S. Rowan et al. / Tetrahedron: Asymmetry 24 (2013) 1369–1381
times for (S)-59 and (R)-59 were 10.8 and 6.7 min, respectively.
times for (S)-64 and (R)-64 were 19.3 and 21.9 min, respectively.
(S)-59. Pale yellow solid (1110 mg, 92%). ½a D20
ꢁ
¼ ꢀ55:4 (c 1.0,
(S)-64. Yellow oil (622 mg, 56%). ½a D20
ꢁ
¼ þ34:1 (c 1.0, CHCl3); 1H
CHCl3); 1H NMR (CDCl3, 400 MHz) d 7.76–7.64 (3H, m, 3 ꢂ Ar-H),
7.59 (1H, m, Ar-H), 5.52 (1H, br, CH-OH), 3.81 (1H, dd, J = 13.5,
6.7 Hz, CHAHB), 3.47 (1H, dd, J = 13.5, 4.3 Hz, CHAHB); ee = 98.4%.
NMR (CDCl3, 400 MHz) d 8.15 (1H, m, Ar-H), 7.44 (1H, m, Ar-H),
6.54 (1H, m, Ar-H), 6.37 (1H, d, J = 8.5 Hz, Ar-H), 4.61 (1H, m, CH-
OH), 3.67–3.50 (4H, m, 2 ꢂ CH2), 2.21–2.04 (2H, m, CH2);
(R)-59. Pale yellow solid (1120 mg, 99%). ½a D20
ꢁ
¼ þ56:8 (c 1.0,
ee = 99.9%. (R)-64. Yellow oil (573 mg, 56%). ½a D20
¼ ꢀ29:3 (c 1.0,
ꢁ
CHCl3); 1H NMR (CDCl3, 400 MHz) d 7.77–7.65 (3H, m, 3 ꢂ Ar-H),
7.60 (1H, m, Ar-H), 5.53 (1H, br, CH-OH), 3.82 (1H, dd, J = 13.5,
6.8 Hz, CHAHB), 3.48 (1H, dd, J = 13.5, 4.3 Hz, CHAHB); ee = 99.8%.
CHCl3); 1H NMR (CDCl3, 400 MHz) d 8.15 (1H, m, Ar-H), 7.44 (1H,
m, Ar-H), 6.54 (1H, m, Ar-H), 6.37 (1H, d, J = 8.5 Hz, Ar-H), 4.61
(1H, m, CH-OH), 3.67–3.50 (4H, m, 2 ꢂ CH2), 2.19–2.06 (2H, m,
CH2); ee = 95.6%.
4.4.22. 3-(4-Bromophenyl)-3-hydroxypropanenitrile 60
Analysis of the enantiomeric excess was performed on a Chir-
alpak AD-H column with eluent 95:5 hexane/IPA, flow rate
1 mL minꢀ1, detection 254 nm and run time 25 min; retention
times for (S)-60 and (R)-60 were 18.9 and 21.1 min, respectively.
4.4.27. 1-(Pyrimidin-2-yl)pyrrolidin-3-ol 65
Analysis of the enantiomeric excess was performed on a Chir-
alpak AD-H column with eluent 95:5 hexane/IPA + 0.1% DEA, flow
rate 1 mL minꢀ1, detection 254 nm and run time 28 min; retention
times for (S)-65 and (R)-65 were 23.3 and 21.0 min, respectively.
(S)-60. Yellow oil (1245 mg, 93%). ½a D20
ꢁ
¼ ꢀ46:3 (c 1.0, CHCl3); 1H
NMR (DMSO-d6, 400 MHz) d 7.56 (2H, m, 2 ꢂ Ar-H), 7.38 (2H, m,
2 ꢂ Ar-H), 4.90 (1H, m, CH-OH), 2.90 (1H, dd, J = 16.7, 5.0 Hz,
CHAHB), 2.82 (1H, dd, J = 16.7, 6.4 Hz, CHAHB); ee = 99.4%. (R)-60.
(S)-65. Off-white solid (421 mg, 40%). ½a D20
¼ þ68:5 (c 1.0, CHCl3);
ꢁ
1H NMR (CDCl3, 400 MHz) d 8.32 (2H, m, 2 ꢂ Ar-H), 6.49 (1H, t,
J = 4.8 Hz, Ar-H), 4.62 (1H, br, CH-OH), 3.79–3.63 (4H, m,
2 ꢂ CH2), 2.20–2.04 (2H, m, CH2); ee = 99.9%.
Yellow oil (1136 mg, 93%). ½a D20
ꢁ
¼ þ44:6 (c 1.0, CHCl3); 1H NMR
(DMSO-d6, 400 MHz) d 7.56 (2H, m, 2 ꢂ Ar-H), 7.38 (2H, m,
2 ꢂ Ar-H), 4.90 (1H, m, CH-OH), 2.90 (1H, dd, J = 16.7, 5.0 Hz,
CHAHB), 2.82 (1H, dd, J = 16.7, 6.4 Hz, CHAHB); ee = 97.7%.
4.4.28. (4-Bromophenyl)(cyclopropyl)methanol 66
Analysis of the enantiomeric excess was performed on a Chir-
alpak AD-H column with eluent 95:5 hexane/IPA, flow rate
1 mL minꢀ1, detection 254 nm and run time 14 min; retention
times for (S)-66 and (R)-66 were 11.2 and 10.0 min, respectively.
4.4.23. 3-Hydroxy-3-phenylpropanenitrile 61
Analysis of the enantiomeric excess was performed on a Chiralcel
OJ column with eluent 7:3 hexane/IPA, flow rate 1 mL minꢀ1, detec-
tion 254 nm and run time 12 min; retention times for (S)-61 and
(R)-61 were 8.2 and 9.2 min, respectively. (S)-61. Brown oil
(S)-66. Pale yellow oil (616 mg, 58%). ½a D20
¼ þ32:1 (c 1.0, CHCl3);
ꢁ
1H NMR (CDCl3, 400 MHz) d 7.50 (2H, m, 2 ꢂ Ar-H), 7.33 (2H, m,
2 ꢂ Ar-H), 4.00 (1H, d, J = 8.2 Hz, CH-OH), 1.19 (1H, m, CH), 0.71–
0.60 (4H, m, 2 ꢂ CH2); ee = 99.0%. (R)-66. Pale yellow oil (611 mg,
(903 mg, 80%). ½a D20
ꢁ
¼ ꢀ61:3 (c 1.0, CHCl3); 1H NMR (CDCl3,
400 MHz) d 7.42–7.32 (5H, m, 5 ꢂ Ar-H), 5.03 (1H, m, CH-OH), 2.75
(2H, m, CH2); ee = 100%. (R)-61. Brown oil (1160 mg, 92%).
60%). ½a 2D0
ꢁ
¼ ꢀ36:3 (c 1.0, CHCl3); 1H NMR (CDCl3, 400 MHz) d
7.50 (2H, m, 2 ꢂ Ar-H), 7.33 (2H, m, 2 ꢂ Ar-H), 4.00 (1H, m, CH-
OH), 1.19 (1H, m, CH), 0.70–0.61 (4H, m, 2 ꢂ CH2); ee = 99.4%.
½
a 2D0
ꢁ
¼ þ64:8 (c 1.0, CHCl3); 1H NMR (CDCl3, 400 MHz) d 7.43–7.32
(5H, m, 5 ꢂ Ar-H), 5.04 (1H, m, CH-OH), 2.77 (2H, m, CH2); ee = 99.6%.
4.4.29. Cyclopropyl(pyridin-4-yl)methanol 67
4.4.24. 3-Hydroxy-3-(pyridin-4-yl)propanenitrile 62
Analysis of the enantiomeric excess was performed on a Chir-
alpak AS-H column with eluent 9:1 hexane/IPA + 0.1% DEA, flow
rate 1 mL minꢀ1, detection 254 nm and run time 16 min; retention
times for (S)-67 and (R)-67 were 11.4 and 8.0 min, respectively.
Analysis of the enantiomeric excess was performed on a Chir-
alpak AS-H column with eluent 4:1 hexane/IPA + 0.1% DEA, flow
rate 1 mL minꢀ1, detection 254 nm and run time 12 min; retention
times for (S)-62 and (R)-62 were 10.2 and 8.3 min, respectively.
(S)-67. Brown oil (564 mg, 54%). ½a D20
ꢁ
¼ þ32:7 (c 1.0, CHCl3); 1H
(S)-62. Orange solid (486 mg, 44%). ½a D20
ꢁ
¼ ꢀ10:6 (c 0.2, CHCl3); 1H
NMR (CDCl3, 400 MHz) d 8.55 (2H, m, 2 ꢂ Ar-H), 7.36 (2H, m,
2 ꢂ Ar-H), 4.02 (1H, d, J = 8.4 Hz, CH-OH), 1.17 (1H, m, CH), 0.70-
0.43 (4H, m, 2 ꢂ CH2); ee = 99.9%. (R)-67. Brown oil (536 mg, 52%).
NMR (DMSO-d6, 400 MHz) d 8.56 (2H, m, 2 ꢂ Ar-H), 7.42 (2H, m,
2 ꢂ Ar-H), 4.94 (1H, m, CH-OH), 3.02–2.83 (2H, m, CH2); ee = 99.6%.
(R)-62. Orange solid (620 mg, 54%). ½a D20
ꢁ
¼ þ11:5 (c 0.2, CHCl3); 1H
½
a 2D0
ꢁ
¼ þ36:2 (c 1.0, CHCl3); 1H NMR (CDCl3, 400 MHz) d 8.56 (2H,
NMR (DMSO-d6, 400 MHz) d 8.56 (2H, m, 2 ꢂ Ar-H), 7.42 (2H, m,
2 ꢂ Ar-H), 4.94 (1H, m, CH-OH), 3.02–2.83 (2H, m, CH2); ee = 99.8%.
m, 2 ꢂ Ar-H), 7.36 (2H, m, 2 ꢂ Ar-H), 4.02 (1H, d, J = 8.4 Hz, CH-
OH), 1.16 (1H, m, CH), 0.70–0.43 (4H, m, 2 ꢂ CH2); ee = 99.7%.
4.4.25. 1-(Pyrimidin-2-yl)piperidin-3-ol 63
4.4.30. 2-Methyl-1-(pyridin-3-yl)propan-1-ol 68
Analysis of the enantiomeric excess was performed on a Chiral-
cel OJ column with eluent 4:1 hexane/IPA + 0.1% DEA, flow rate
1 mL minꢀ1, detection 254 nm and run time 18 min; retention
times for (S)-63 and (R)-63 were 7.5 and 6.7 min, respectively.
Analysis of the enantiomeric excess was performed on a Chiral-
cel OD-H column with eluent 9:1 hexane/IPA + 0.1% DEA, flow rate
1 mL minꢀ1, detection 254 nm and run time 15 min; retention
times for (S)-68 and (R)-68 were 10.7 and 12.2 min, respectively.
(S)-63. Colourless oil (341 mg, 31%). ½a D20
ꢁ
¼ þ44:4 (c 1.0, CHCl3);
(S)-68. Colourless oil (1046 mg, 94%). ½a D20
¼ ꢀ52:0 (c 1.0, CHCl3);
ꢁ
1H NMR (CDCl3, 400 MHz) d 8.29 (2H, m, 2 ꢂ Ar-H), 6.46 (1H, t,
J = 4.7 Hz, Ar-H), 4.11 (1H, m, N-CHAHB), 3.95–3.83 (2H, m, N-CHCHD,
CH-OH), 3.66–3.58 (2H, m, N-CHAHB, N-CHCHD), 1.98–1.50 (4H, m,
2 ꢂ CH2); ee = 99.2%. (R)-63. Colourless oil (369 mg, 39%).
1H NMR (CDCl3, 400 MHz) d 8.50 (2H, m, 2 ꢂ Ar-H), 7.68 (1H, m,
Ar-H), 7.27 (1H, m, Ar-H), 4.45 (1H, d, J = 6.5 Hz, CH-OH), 1.98
(1H, m, CH(CH3)2), 0.99 (3H, d, J = 6.7 Hz, CH3), 0.83 (3H, d,
J = 6.8 Hz, CH3); ee = 98.7%. (R)-68. Colourless oil (1018 mg, 93%).
½
a 2D0
ꢁ
¼ ꢀ44:5 (c 1.0, CHCl3); 1H NMR (CDCl3, 400 MHz) d 8.29 (2H,
½
a 2D0
ꢁ
¼ ꢀ52:0 (c 1.0, CHCl3); 1H NMR (CDCl3, 400 MHz) d 8.47
m, 2 ꢂ Ar-H), 6.46 (1H, t, J = 4.7 Hz, Ar-H), 4.12 (1H, dd, J = 13.1,
3.4 Hz, N-CHAHB), 3.96–3.83 (2H, m, N-CHCHD, CH-OH), 3.66–3.57
(2H, m, N-CHAHB, N-CHCHD), 1.98–1.50 (4H, m, 2 ꢂ CH2); ee = 94.8%.
(2H, m, 2 ꢂ Ar-H), 7.68 (1H, m, Ar-H), 7.27 (1H, m, Ar-H), 4.44
(1H, d, J = 6.6 Hz, CH-OH), 1.97 (1H, m, CH(CH3)2), 0.99 (3H, m,
CH3), 0.83 (3H, m, CH3); ee = 99.7%.
4.4.26. 1-(Pyridin-2-yl)pyrrolidin-3-ol 64
4.4.31. 1-(3-Chlorophenyl)-2-methylpropan-1-ol 69
Analysis of the enantiomeric excess was performed on a Chir-
alpak AD-H column with eluent 95:5 hexane/IPA + 0.1% DEA, flow
rate 1 mL minꢀ1, detection 254 nm and run time 30 min; retention
Analysis of the enantiomeric excess was performed on a
Chiralpak IB column with eluent 95:5 hexane/IPA, flow rate
1 mL minꢀ1, detection 254 nm and run time 8 min; retention times

A. S. Rowan et al. / Tetrahedron: Asymmetry 24 (2013) 1369–1381
1379
Figure 6. Optimised geometries, relative energies and Boltzmann populations of the three calculated conformers of the (S)-configuration.
for (S)-69 and (R)-69 were 6.0 and 6.6 min, respectively. (S)-69. Col-
rate 1 mL minꢀ1, detection 254 nm and run time 18 min; retention
ourless oil (812 mg, 76%). ½a D20
ꢁ
¼ ꢀ31:6 (c 1.0, CHCl3); 1H NMR
times for (S)-73 and (R)-73 were 10.0 and 14.0 min, respectively.
(CDCl3, 400 MHz) d 7.33–7.16 (4H, m, 4 ꢂ Ar-H), 4.37 (1H, m, CH-
OH), 1.94 (1H, m, CH(CH3)2), 1.85 (1H, d, J = 3.4 Hz, CH-OH), 0.98
(3H, d, J = 6.7 Hz, CH3), 0.83 (3H, d, J = 6.8 Hz, CH3); ee = 97.7%. (R)-
(S)-73. White solid (823 mg, 78%). ½a D20
¼ þ24:5 (c 1.0, MeOH);
ꢁ
1H NMR (DMSO-d6, 400 MHz) d 7.98 (2H, m, 2 ꢂ Ar-H), 7.62 (2H,
m, 2 ꢂ Ar-H), 5.28 (1H, m, CH-OH); ee = 93.4%. (R)-73. White solid
69. Colourless oil (673 mg, 58%). ½a D20
ꢁ
¼ þ30:6 (c 1.0, CHCl3); 1H
(965 mg, 90%). ½a D20
ꢁ
¼ ꢀ34:6 (c 1.0, MeOH); 1H NMR (DMSO-d6,
NMR (CDCl3, 400 MHz) d 7.33–7.17 (4H, m, 4 ꢂ Ar-H), 4.37 (1H, m,
CH-OH), 1.94 (1H, m, CH(CH3)2), 1.85 (1H, d, J = 3.4 Hz, CH-OH),
0.98 (3H, d, J = 6.7 Hz, CH3), 0.83 (3H, d, J = 6.8 Hz, CH3); ee = 99.5%.
400 MHz) d 7.98 (2H, m, 2 ꢂ Ar-H), 7.62 (2H, m, 2 ꢂ Ar-H), 5.29
(1H, m, CH-OH); ee = 99.7%.
4.5. Large scale bioreduction
4.4.32. 2,2,2-Trifluoro-1-(6-fluoro-1-methyl-1H-indol-3-yl)ethanol
70
Analysis of the enantiomeric excess was performed on a Chir-
alpak AD-H column with eluent 9:1 hexane/IPA, flow rate
1 mL minꢀ1, detection 254 nm and run time 17 min; retention
times for (S)-70 and (R)-70 were 10.9 and 13.3 min, respectively.
A161 cell free extract powder (5 g) was dissolved in 0.1 M phos-
phate buffer (pH 7, 10 L) and stirred at 30 °C. Next, para-bromoph-
enacyl chloride 15 (500 g, 2.14 mol) was dissolved in DMSO
(200 mL) and added to the stirring solution, followed by NAD
(1 g) and isopropyl alcohol (1 L). Reaction progress was monitored
(S)-70. White solid (911 mg, 89%). ½a D20
ꢁ
¼ þ12:9 (c 1.0, CHCl3); 1H
NMR (CDCl3, 400 MHz) d 7.64 (1H, m, Ar-H), 7.20 (1H, s, Ar-H),
7.01 (1H, m, Ar-H), 6.94 (1H, m, Ar-H), 5.30 (1H, m, CH-OH), 3.75
(3H, s, CH3); ee = 99.9%. (R)-70. White solid (777 mg, 71%).
½
a 2D0
ꢁ
¼ ꢀ14:8 (c 1.0, CHCl3); 1H NMR (CDCl3, 400 MHz) d 7.64
(1H, m, Ar-H), 7.20 (1H, s, Ar-H), 7.01 (1H, m, Ar-H), 6.95 (1H, m,
Ar-H), 5.31 (1H, m, CH-OH), 3.76 (3H, s, CH3); ee = 99.9%.
4.4.33. 1-(4-Chlorophenyl)-2,2,2-trifluoroethanol 71
Analysis of the enantiomeric excess was performed on a Chir-
alpak IB column with eluent 9:1 hexane/IPA, flow rate 1 mL minꢀ1
,
detection 254 nm and run time 7 min; retention times for (S)-71
and (R)-71 were 5.4 and 4.8 min, respectively. (S)-71. Pale yellow
oil (716 mg, 65%). ½a D20
ꢁ
¼ þ26:1 (c 1.0, CHCl3); 1H NMR (CDCl3,
400 MHz) d 7.44–7.37 (4H, m, 4 ꢂ Ar-H), 5.02 (1H, m, CH-OH);
ee = 98.7%. (R)-71. Pale yellow oil (758 mg, 68%). ½a D20
¼ ꢀ28:7 (c
ꢁ
1.0, CHCl3); 1H NMR (CDCl3, 400 MHz) d 7.44–7.37 (4H, m,
4 ꢂ Ar-H), 5.02 (1H, m, CH-OH); ee = 99.8%.
4.4.34. 1-(3-Chlorophenyl)-2,2,2-trifluoroethanol 72
Analysis of the enantiomeric excess was performed on a Chir-
alpak IB column with
a Phenomenex Sphereclone 5l silica
(150 ꢂ 4.6 mm) pre-column with eluent 95:5 hexane/IPA, flow rate
1 mL minꢀ1, detection 254 nm and run time 15 min; retention
times for (S)-72 and (R)-72 were 10.9 and 10.0 min, respectively.
(S)-72. Pale yellow oil (853 mg, 81%). ½a D20
¼ þ23:1 (c 1.0, CHCl3);
ꢁ
1H NMR (CDCl3, 400 MHz) d 7.50 (1H, s, Ar-H), 7.41–7.32 (3H, m,
3 ꢂ Ar-H), 5.02 (1H, m, CH-OH); ee = 99.6%. (R)-72. Colourless oil
(1006 mg, 89%). ½a D20
ꢁ
¼ ꢀ24:5 (c 1.0, CHCl3); 1H NMR (CDCl3,
400 MHz) d 7.50 (1H, s, Ar-H), 7.41–7.32 (3H, m, 3 ꢂ Ar-H), 5.02
(1H, m, CH-OH); ee = 95.3%.
4.4.35. 4-(2,2,2-Trifluoro-1-hydroxyethyl)benzoic acid 73
Analysis of the enantiomeric excess was performed on a Chir-
alpak AD-H column with eluent 9:1 hexane/IPA + 0.05% TFA, flow
Figure 7. The IR and VCD spectra of each of the three lowest energy-conformers
compared with the observed spectra.

1380
A. S. Rowan et al. / Tetrahedron: Asymmetry 24 (2013) 1369–1381
by HPLC analysis. After 6 h, the reaction was complete. The reac-
tion mixture was extracted with MTBE (2 ꢂ 2 L), and the organic
solvent was dried (MgSO4, 300 g), filtered and concentrated in
vacuo to afford compound (S)-50 (458 g, 1.94 mol, 91% yield) as a
pale orange oil. Analysis of the ee was performed as described in
converted to Lorentzian bands with 6 cmꢀ1 half-width for compari-
son to experiment. Three of the nine conformers had energies of less
than 2 kcal/mol difference to that of the lowest-energy conformer
(Fig. 6). By comparing the observed VCD and IR spectra of the sam-
ples with those of the Boltzmann averaged VCD spectra of (S)- and
(R)-configuration (Fig. 8; Boltzmann averaged from the predicted
VCD spectra of the individual conformers shown in Fig. 7), the abso-
lute configuration of chiral alcohol 39 was assigned as (R).
section 4.4.12; ee of (S)-50 = 99.8%; ½a D20
¼ þ40:2 (c 1.0, CHCl3);
ꢁ
1H NMR (CDCl3, 400 MHz): d 7.53 (2H, m, 2 ꢂ Ar-H), 7.30 (2H, m,
2 ꢂ Ar-H), 4.89 (1H, m, CH-OH), 3.73 (1H, dd, J = 11.2, 3.7 Hz,
CHAHB), 3.63 (1H, dd, J = 11.2, 8.4 Hz, CHAHB).
Acknowledgements
4.6. Exampleoftheprocedurefortheabsolutestructuredetermination
by VCD
The authors wish to thank the UK Technology Strategy Board for
its generous financial support of this project. Meredith Lloyd-Jones
(BioBridge Ltd) and the Chemistry Innovation and Biosciences
Knowledge Transfer Networks are also thanked for co-ordinating
the project and guidance in setting up this collaboration, respec-
tively. Furthermore, Olivier Drap, Neal Sach and Andrew Cronin
(Pfizer) are thanked for support and helpful discussions. This study
was part financed by the European Regional Development Fund
under the European Sustainable Competitiveness Program for
Northern Ireland.
VCD measurement: Chiral alcohol 39 from the reduction of
ketone 4 by CRED A601 was dissolved in CDCl3 (17.9 mg/0.2 mL)
and placed in a 100 lm path-length cell with BaF2 windows. Next,
the IR and VCD spectra were recorded on a ChiralIRTM FT-VCD spec-
trometer with Dual PEM (BioTools, Inc.), with 4 cmꢀ1 resolution
and 10 h collection. The spectrum of CDCl3 was measured for 9 h
for VCD baseline correction and also used for subtraction of solvent
for IR. The PEMs were optimised at 1400 cmꢀ1
.
Structure modelling and calculations of IR and VCD spectra: The
(S)-configuration of chiral alcohol 39 was built with Hyperchem8
(Hypercube, Gainesville, FL). A conformational search was carried
out with Hyperchem8 at the molecular mechanics level and resulted
in nine low energy conformers. Geometry optimisation, frequency
and IR and VCD intensity calculations were then carried out at the
DFT level with GAUSSIAN 09 (Gaussian Inc., Wallingford, CT) with the
B3LYP hybrid functional and the 6-31G(d) basis set. The calculated
frequencies were scaled by 0.97 and the IR and VCD intensities were
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