Enzyme-mediated enantioselective hydrolysis of soluble polymer-supported carboxylates

Article abstract of DOI:10.1016/j.tet.2010.07.076

The enzyme-mediated enantioselective hydrolysis of water-soluble polymer-supported carboxylates is disclosed. The representative monomethoxy poly(ethylene glycol) (MPEG, av MW 5000)-supported substrate was synthesized by immobilization of (±)-1-phenylethanol onto the modified MPEG (MPEG/NH₂) through an carboxylate linker with a succinate spacer. For the screening of the hydrolytic enzymes, the substrate was enantioselectively hydrolyzed by lipase from Candida antarctica (Novozym 435) in a mixed solvent (hexane/buffer=9/1) at 30 °C to afford the remaining (S)-substrate and the resulting (R)-alcohol (E value>200). The products were easily separated by a simple procedure without any laborious column chromatography. The substrate was hydrolyzed with NaOH in MeOH/H₂O to afford the corresponding (S)-alcohol. We also found that the structure of the spacer between the MPEG moiety and the carboxylate linker strongly affected both the reactivity and enantioselectivity, and the substrate bearing a glutarate spacer gave the best result. Our procedure was applicable for the preparation of several optically active alcohols.

Full text of DOI:10.1016/j.tet.2010.07.076

Tetrahedron 66 (2010) 8060e8067
Contents lists available at ScienceDirect
Tetrahedron
journal homepage: www.elsevier.com/locate/tet
Enzyme-mediated enantioselective hydrolysis of soluble polymer-supported
carboxylates
Masayuki Okudomi, Kanpei Ageishi, Tomomi Yamada, Naoka Chihara, Takuya Nakagawa,
*
Katsumi Mizuochi, Kazutsugu Matsumoto
Department of Chemistry, Meisei University, Hodokubo 2-1-1, Hino, Tokyo 191-8506, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 1 July 2010
Received in revised form 28 July 2010
Accepted 28 July 2010
Available online 3 August 2010
The enzyme-mediated enantioselective hydrolysis of water-soluble polymer-supported carboxylates is
disclosed. The representative monomethoxy poly(ethylene glycol) (MPEG, av MW 5000)-supported
substrate was synthesized by immobilization of (ꢀ)-1-phenylethanol onto the modified MPEG (MPEG/
NH2) through an carboxylate linker with a succinate spacer. For the screening of the hydrolytic enzymes,
the substrate was enantioselectively hydrolyzed by lipase from Candida antarctica (Novozym 435) in
a mixed solvent (hexane/buffer¼9/1) at 30 ꢁC to afford the remaining (S)-substrate and the resulting (R)-
alcohol (E value>200). The products were easily separated by a simple procedure without any laborious
column chromatography. The substrate was hydrolyzed with NaOH in MeOH/H2O to afford the corre-
sponding (S)-alcohol. We also found that the structure of the spacer between the MPEG moiety and the
carboxylate linker strongly affected both the reactivity and enantioselectivity, and the substrate bearing
a glutarate spacer gave the best result. Our procedure was applicable for the preparation of several
optically active alcohols.
Keywords:
Carboxylates
Enantioselective hydrolysis
Enzymes
Poly(ethylene glycol)-supported substrate
Ó 2010 Elsevier Ltd. All rights reserved.
1. Introduction
molecular weight monomethoxy PEG (MPEG, av Mw 550, 750, and
5000)-supported substrate with a carbonate linker using porcine
The hydrolase-mediated kinetic resolution of racemic alcohols
and esters is one of the attractive methods for the preparation of
optically active compounds, and there have already been a great
number of studies about enzymatic hydrolysis and esterification.1
During the reaction, however, the remaining substrate and the
resulting product must be separated, and the tedious and wasteful
separation step by column chromatography is still a big problem for
developing an easy operation and a sustainable product. In order to
simplify the separation of products, an organic reaction on polymer
supports is a practical method. On the other hand, the usual organic
compounds are insoluble in water, and the solubility profile
prevents the enzymatic reaction of such compounds in some cases.
Recently, poly(ethylene glycol) (PEG) has been recognized as an
inexpensive and convenient soluble material, and the reactions
using several PEG-supported compounds in organic synthesis have
also been developed under homogeneous conditions.2e4 We
postulated that a PEG-supported strategy could be preferable for an
enzymatic reaction in water, and could also be useful for the easy
separation of the products. In this context, we have already
disclosed the enzymatic kinetic resolution of the low- and middle-
pancreas lipase (PPL, lipase Type II from Sigma), and we have
succeeded in the easy separation of the products.5,6 In particular,
the MPEG5000-bound carbonates were soluble solids and easier to
handle, and the reaction of the substrates also gave better results in
terms of both the reactivity and the enantioselectivity. Although
a carbonate bond is one of the representative linkers for polymer-
support reactions, this structure is not necessarily suitable for en-
zymatic hydrolysis, and there have been a relatively few examples
for the lipase-catalyzed kinetic resolution of carbonates. In our
previous study, many commercial enzymes scarcely hydrolyzed the
MPEG-supported carbonates.5 In addition, the coupling of alcohols
with MPEG through a carbonate linker should be constructed at
high temperature (>120 ꢁC), and the purity of the resulting com-
pounds was low in some cases. In order to extend the scope of the
application of our PEG-strategy, we then focused on the enzymatic
hydrolysis of the MPEG-supported carboxylates, which had the
usual ester bond as the linker. In this paper, we disclose the easy
preparation of new types of MPEG-supported compounds with
a carboxylate linker and the hydrolase-mediated kinetic resolution
of these substrates. We also report that the structure of an appro-
priate hydrophobic spacer between the MPEG moiety and the
carboxylate linker significantly affects both the reactivity and
enantioselectivity, and our procedure is applicable for the prepa-
ration of several optically active secondary alcohols.
* Corresponding author. Tel./fax: þ81 42 591 7360; e-mail address: mkazu@
chem.meisei-u.ac.jp (K. Matsumoto).
0040-4020/$ e see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2010.07.076

M. Okudomi et al. / Tetrahedron 66 (2010) 8060e8067
8061
2. Results and discussion
(12), respectively (Scheme 3). The preparations of the MPEG-sup-
ported substrates were confirmed by 1H and 13C NMR and ESI-TOF
MS analyses.
2.1. Preparation of MPEG-supported compounds
Initially, we tried to prepare new MPEG-supported carboxylates
with a high purity by an easy and effective procedure. We selected
1-phenylethanol (1) as the representative target alcohol, and the
racemate (ꢀ)-1 was combined with succinic anhydride using DMAP
in CH2Cl2 to give the dicarboxylic monoester (ꢀ)-2a in 92% yield
(Scheme 1).
2.2. Screening test of enzymes
Next, we screened the enzymes having the enantioselective
hydrolytic ability of the carboxylate (ꢀ)-7a as the representative
substrate. The reaction of the screening test was performed in 0.1 M
phosphate buffer (pH 6.5) for 24 h at 30 ꢁC, and the selection of the
O
X
O
O
, DMAP
OH
Me Ph
( )-1
O
O
Ph
Me
a: X =
b: X =
CH2CH2
CH2CH2CH2
Me
HO
X
O
CH2Cl2, rt
( )-2a (92%), ( )-2b (73%)
( )-2c (82%), ( )-2d (36%)
( )-2e (59%), ( )-2f (96%)
( )-2g (94%)
c: X =
d: X =
e: X =
CH2CHCH2
Me Me
CH2CCH2
MsCl, Et3N
CH2Cl2, rt
NaN3
OMs
OH
N3
5 (94%)
Ph
Me
CH2CCH2
DMF, 95 °C
MPEG5000
4 (99%)
3
f: X =
( )-2, DMAP
O
O
DCC
CH2Cl2, rt
DIBAL-H
OTBS
CH2CHCH2
NH2
N
H
X
O
g: X =
CH2Cl2, 0 °C
6 (87%)
( )-7a (95%), ( )-7b (71%)
( )-7c (90%), ( )-7d (93%)
( )-7e (88%), ( )-7f (79%)
( )-7g (91%)
Scheme 1.
The succinate part could be utilized as the spacer between
the MPEG and carboxylate linker, and the reactions using other
carboxylic anhydrides gave the corresponding intermediates
(ꢀ)-2beg. When (ꢀ)-2a was directly coupled with MPEG5000eOH
(3, av Mw 5000), the diester (ꢀ)-8 was obtained. However, both the
two ester bonds of 8 was hydrolyzed by lipases and esterases in
many cases, and the substrate was consequently decomposed into
three components, 1, 3, and succinic acid (9) (Scheme 2).
O
O
O
R2
R1
OH
R1 R2
O
O
, DMAP
X
HO
X
O
CH2Cl2, rt
( )-13a (98%), ( )-13b (73%)
( )-14a (90%), ( )-14b (75%)
( )-15a (62%), ( )-15b (65%)
( )-10, 11, or 12
( )-13, 14 or 15
DMAP
DCC
O
O
R2
R1
6
N
H
X
O
OH
CH2Cl2, rt
Me Ph
( )-16a (98%), ( )-16b (73%)
( )-17a (90%), ( )-17b (75%)
( )-18a (62%), ( )-18b (65%)
1
+
O
Ph
Me
Hydrolase
O
OH
O
3
10, 13, 16: R1 = Et, R2 = Ph
O
+
11, 14, 17: R1 = Me, R2 = CH2Ph
12, 15, 18: R1 = Me, R2 = (CH2)2OBn
O
( )-8
HO
OH
9
O
a: X =
b: X =
CH2CH2
CH2CH2CH2
Scheme 2.
Scheme 3.
We noticed that the coupling of the MPEG part with the spacer
should be constructed by the amide bond, which could be difficult
to hydrolyze with lipases. We then tried to change the hydroxyl
group of MPEG to the amino group. The mesylate 4 derived from 3
was treated with sodium azide, and the resulting 5 was reduced by
DIBAL-H to afford the modified MPEGeNH2 (6). The coupling of 6
with compound (ꢀ)-2a using DMAP and DCC in CH2Cl2 at room
temperature gave the MPEG5000-supported carboxylic ester (ꢀ)-7a
bearing a succinate spacer. The reactions of 6 using other di-
carboxylic monoesters (ꢀ)-2beg afforded the corresponding esters
(ꢀ)-7beg with various spacers, respectively. In a similar manner,
the MPEG-supported compounds (ꢀ)-16, 17, and 18 were prepared
from the corresponding secondary alcohols, (ꢀ)-1-phenyl-1-prop-
anol (10), 1-phenyl-2-propanol (11), and 4-benzyloxy-2-butanol
enzyme was carried out on the basis of the enantioselectivity by
checking the enantiomeric excess (ee) of the product 1 by GC (CP-
Cyclodextrin-B-236-M19, Chrompack). In our previous study, al-
most every lipase, except for PPL, could not catalyze the hydrolysis
of the MPEG-supported substrates with a carbonate linker.5 In-
terestingly, all the enzymes hydrolyzed the MPEG-supported car-
boxylate (ꢀ)-7a to afford the corresponding alcohol (R)-1, although
the ees varied with the type of enzyme (Table 1).
In the case of the reaction using PPL, which was the best enzyme
for the enantioselective hydrolysis of the MPEG-supported car-
bonates, the ee of the resulting (R)-1 was low (30% ee). These re-
sults indicated that the enzymes essentially recognize the structure

8062
M. Okudomi et al. / Tetrahedron 66 (2010) 8060e8067
Table 1
after acidification of the aqueous layer. After the chemical hydro-
lysis of the unreacted substrate 7a with NaOH aq in MeOH, the ee of
the resulting (S)-1 was also determined by GC analysis. In our
previous study, we already found that the reaction of the MPEG-
supported carbonates in a mixed solvent of buffer and hexane gave
a better result than that in a homogeneous system. We then
examined the reaction in the two-phase system. As expected,
changing the medium improved both the reactivity and enantio-
selectivity, and the conversion and E value of the reaction in
a mixed solvent containing 90% hexane were up to 0.36 and 172,
respectively (entry 2). It was found that the reaction at a lower
temperature proceeded with a higher enantioselectivity. For the
reaction at 0 ꢁC (entry 3), the E value increased to >249 and the
optically pure (R)-1 was obtained, although the conversion appar-
ently decreased to 0.19. A longer reaction time improved the
conversion and almost complete optical resolution of 1 was
accomplished, but the time taken to thoroughly react (R)-7a
needed 72 h (entry 4). As expected, increasing the amount of the
enzyme (60 mg) also improved the conversion without lowering
the enantioselectivity (entry 5). In this two-phase reaction system,
the property of a medium-molecular weight MPEG5000 allows us to
facilitate the easy work-up and separation of the products the same
as our previous studies.6 After the enzymatic reaction, the resulting
alcohol (R)-1 was already extracted in the hexane layer, and then
isolated after evaporation. On the other hand, CH2Cl2 was added to
the aqueous layer, and dried over anhydrous Na2SO4. After evapo-
ration, the residue was poured into Et2O to precipitate the mixture
of the (S)-7a and MPEG-supported carboxylic acid as a white solid,
which was collected by simple filtration.
Enantioselective hydrolysis of MPEG5000-supported carboxylic ester with hydrolytic
enzymea
O
Ph
Me
H
N
Enzyme
O
pH6.5 buffer
O
30 °C, 24 h
( )-7a
OH
Me Ph
(R)-1
O
Ph
Me
H
N
+
O
O
(S)-7a
Enzyme
ee of 1 (%)b
Lipase A (Amano)
Lipase D (Amano)
Lipase D-360 (Amano)
Lipase PS (Amano)
Newlase F (Amano)
PLE (Amano)
Lipase OF (Meito)
Lilipase (Nagase)
Esterase SNSM-87 (Nagase)
PPL (Sigma)
10
28
w0
48
11
32
42
12
w0
31
27
Lipase Type VII (Sigma)
a
-Chymotrypsin (E. Merck)
w0
96
Novozym 435 (Novozymes)
a
The reaction was performed using 5 mM of the substrate with the hydrolytic
enzyme in 0.1 M phosphate buffer (pH 6.5) for 24 h at 30 ꢁC.
b
Determined by GC analysis.
of the linker, and the carboxylate linker could be widely accepted by
hydrolases for use of a soluble polymer as the matrix. Finally, we
selected Novozym 435 from Candida antarctica (Novozymes) as the
best enzyme, and the ee of the obtained (R)-1 was 96% ee.
We speculated that a suitable spacer could increase the affinity
to the active site of the enzyme. We then investigated changing the
spacer of the substrate in order to improve the conversion and
enantioselectivity of the reaction for 24 h at 30 ꢁC (Table 3).
2.3. Enzymatic hydrolysis of MPEG-supported carboxylates
with Novozym 435
Table 3
We next tried to evaluate the determination of the enantiose-
lectivity of the reaction using Novozym 435, and these results are
shown in Table 2.
Enantioselective hydrolysis of MPEG5000-supported carboxylates 7beg with Novo-
zym 435a
OH
Me Ph
O
O
Ph
Me
O
O
Ph
Me
Novozym 435
+
N
H
X
O
N
H
X
O
hexane-buffer
Table 2
30 °C, 24 h
(R)-1
( )-7
(S)-7
Enantioselective hydrolysis of MPEG5000-supported carboxylate (ꢀ)-7a with Novo-
zym 435a
e: X =
b: X = CH2CH2CH2
OH
Me Ph
(R)-1
O
Ph
Me
O
Ph
Me
CH2CCH2
H
N
H
N
Novozym 435
Solvent
Me
CH2CHCH2
+
O
O
c: X =
O
O
f: X =
( )-7a
(S)-7a
Me Me
CH2CCH2
OTBS
CH2CHCH2
d: X =
g: X =
Entry Solvent
Temp (ꢁC) Time (h) ee of 7a/%b ee of 1/%c Conv. E value
1
2
3
Buffer
Hexane/Buffer 30
Hexane/Buffer
Hexane/Buffer
Hexane/Buffer
30
24
24
24
72
24
17
55
23
>99
95
96
98
>99
>99
>99
0.15
0.36
0.19
0.50 >1057
0.49 >747
58
172
>249
Entry
Substrate
ee of 7/%b
ee of 1/%c
Conv.
E value
0
0
0
1
2
3
4
5
6
(ꢀ)-7b
(ꢀ)-7c
(ꢀ)-7d
(ꢀ)-7e
(ꢀ)-7f
(ꢀ)-7g
>99
93
2
1
w0
w0
>99
99
89
57
3
0.50
0.48
0.02
0.02
0.05
>1057
684
18
4
5d
a
4
1
12
Unless otherwise noted, the reaction was performed using 5 mM of the sub-
strate (125 mg) with Novozym 435 (20 mg) in 0.1 M phosphate buffer (pH 6.5) or
a mixed medium (hexane/0.1 M phosphate buffer (pH 6.5)¼9/1).
84
w0
b
Determined by GC analysis after chemical hydrolysis of the unreacted substrate.
Determined by GC analysis.
Using 60 mg of Novozym 435.
a
The reaction was performed using 5 mM of the substrate with Novozym 435 in
c
a mixed medium (hexane/0.1 M phosphate buffer (pH 6.5)¼9/1) for 24 h at 30 ꢁC.
d
b
Determined by GC analysis after hydrolysis of the unreacted substrates.
Determined by GC analysis.
c
In a typical experiment, 125 mg of (ꢀ)-7a (sub. concn, 5 mM)
and 20 mg of Novozym 435 were added to a medium (5 mL) in
a test tube, and the mixture was stirred for 24 h at a constant
temperature. The reaction was first carried out in 0.1 M phosphate
buffer (pH 6.5) at 30 ꢁC (entry 1). Although the hydrolysis of (ꢀ)-7a
proceeded, the reactivity was low (conv.¼0.15) and the enantiose-
lectivity was moderate (E value¼58).7 In this case, the resulting
(R)-1 should be extracted from the mixture with AcOEt, and the
remaining substrate (S)-7a was obtained by extraction with CH2Cl2
Beyond our expectation, changing the spacer significantly
affected not only the conversion, but also the enantioselectivity.
Surprisingly, the substrate (ꢀ)-7b bearing the glutarate spacer was
smoothly hydrolyzed, and the conversion was up to 0.50 (entry 1).
In addition, the reaction also proceeded with an excellent enan-
tioselectivity (E value>1057) to afford both almost optically pure
enantiomers. The reaction was also performed using 3.0 g of the

M. Okudomi et al. / Tetrahedron 66 (2010) 8060e8067
8063
substrate (ꢀ)-7b in a recovery flask, and we finally obtained (R)-1
substrate 7a. Interestingly, the hydrolysis of the acetate of the al-
cohol 12 with Novozym 435 in phosphate buffer for 24 h at 30 ꢁC
smoothly proceeded (conv.¼0.52), but the E value was only 46.
(>99% ee, [
a]
20 þ34.5 (c 1.77, MeOH); lit.8 [
a
]
20 þ45 (c 5.15, MeOH))
D
D
in 25% and (S)-1 (94% ee) in 24% isolated yields (conv.¼0.49, E
value>712). Interestingly, the reaction of compound (ꢀ)-7c bearing
the 3-methylglutarate spacer gave almost the same result as the
case of 7b (entry 2, conv.¼0.48, E value¼684). However, the in-
troduction of a dimethyl group to the glutarate spacer drastically
decreased both the reactivity and enantioselectivity, and the con-
version and E value of the reaction using 7d bearing the 3,3-
dimethylglutarate spacer were only 0.02 and 18, respectively (entry
3). As expected, the compounds 7e, 7f, and 7g bearing the bulky
spacer (3,3-tetramethyleneglutarate, phthalate, and 3-(tert-butyl-
dimethylsilyloxy)glutarate spacers, respectively) were scarcely
hydrolyzed, and the enantioselectivities were quite low in all cases
(entries 4e6, respectively).
Next, in order to apply the concept of this reaction for the kinetic
resolution of the other secondary alcohols, we examined the en-
zymatic hydrolysis of several MPEG-supported carboxylates bear-
ing the succinate or glutarate spacer. These results are shown in
Table 4. While the enantioselective hydrolysis of the 1-phenyl-1-
propanol derivative (ꢀ)-16a bearing the succinate spacer hardly
proceeded (entry 1), changing the spacer to the glutarate ((ꢀ)-16b)
significantly improved both the reactivity and enantioselectivity
(entry 2; conv.¼0.53, E value¼51). In the case of the 1-phenyl-2-
propanol derivative (ꢀ)-17b bearing the glutarate spacer (entry 4),
the enzyme also catalyzed the hydrolysis of the compound with an
excellent enantioselectivity to afford the highly optically active (R)-
11 (>99% ee) and (S)-17b (95% ee) (conv. 0.49, E value>747). On the
other hand, the reaction of the carboxylate (ꢀ)-17a, which had the
succinate spacer, slowly proceeded with a moderate enantiose-
lectivity (entry 3; conv. 0.09, E value¼54). These results indicate
that the enzyme apparently prefers the R-enantiomer of the sub-
strates having not the succinate, but the glutarate spacer in all
cases. It was noteworthy that both reactions of the 4-benzyloxy-2-
butanol derivatives (ꢀ)-18a (entry 5) and 18b (entry 6) proceeded
to afford the optically active compound (R)-12 in both cases,
although the conversions were low (conv.¼0.16 and 0.18,
respectively). The excellent enantioselectivities (E value>239 and
>249, respectively) were substantially the same as that in the
reaction of 17b. Although the reactions of 18a and 18b were not
carried out under the optimized conditions, we supposed that
prolonging the reaction time and increasing the amount of the
enzyme could improve the conversions in a manner similar to the
3. Conclusions
In summary, we have demonstrated the enzyme-mediated
kinetic resolution of soluble polymer MPEG5000-supported car-
boxylates to afford optically active secondary alcohols 1, 10, 11,
and 12. We also have disclosed that the reactivity and enantio-
selectivity can be controlled using a suitable hydrophobic spacer
between the MPEG moiety and the carboxylate linker. In our
method, the separation and isolation of the reaction products
were achieved by a simple precipitation technique without using
time- and solvent-consuming column chromatography. Because
the use of the PEG matrix could change the physical properties
of the insoluble organic compounds to solubilize them in an
aqueous medium, the PEG-strategy could pave the way for
a novel enzymatic system.
4. Experimental
4.1. General
1H (500 MHz or 300 MHz) and 13C (125 MHz or 75 MHz) NMR
spectra were recorded on a JEOL a-500 with tetramethylsilane as
the internal standard. IR spectra were recorded with a Shimadzu IR
Prestige-21 spectrometer. ESI-TOF mass spectra were measured in
MeOH/H2O solution including AcONa with a JEOL JMS-T100. All
enzymatic reactions were performed in a SANYO incubator MIR-
253. Kieselgel 60 F254 Art.5715 (E. Merck) was used for analytical
thin-layer chromatography (TLC), and preparative TLC was per-
formed on Kieselgel 60 F254 Art. 5744 (E. Merck). The optical rota-
tions were measured with a Jasco DIP-1000 polarimeter. HPLC data
were obtained on a Shimadzu LC-10ADvp, SPD-10Avp, and sic 480II
date station (System Instruments Inc.). GLC data were obtained on
GL Sciences GC 353B and sic 480II. MPEG5000eOH (3) was pur-
chased from Fluka, and all other chemicals and enzymes were also
obtained from commercial sources.
The yields of the MPEG-supported compounds were based on
the weights of the starting materials. The purities were determined
by 1H NMR analysis, and the terminal methyl group and/or the PEG-
methylenes were used as the reference.
4.2. Preparation of dicarboxylic monoesters
Table 4
Enantioselective hydrolysis of MPEG5000-supported carboxylates 16e18 with No-
vozym 435a,b
4.2.1. 3-((1-Phenylethoxy)carbonyl)propanoic acid ((ꢀ)-2a). Under
an argon atmosphere, succinic anhydride (489.5 mg, 4.892 mmol),
and DMAP (598.7 mg, 4.901 mmol) were added to a solution of
1-phenyletanol ((ꢀ)-1, 299.1 mg, 2.448 mmol) in CH2Cl2 (10 mL),
and the solution was stirred for 2 h at room temperature. After the
product was extracted with satd NaHCO3 aq, the separated water
layer was acidified by 2 M HCl. The products were extracted with
AcOEt, washed with brine, and dried over Na2SO4. After evapora-
tion in vacuo, the residue was purified by column chromatography
on silica gel (hexane/AcOEt¼3/1) to give the (ꢀ)-2a as a colorless oil
in 97% yield (527.9 mg); IR (KBr) 2982, 2934, 1736, 1713, 1495, 1450,
R2
R1
O O
X
R2
R1
OH
O
O
Novozym 435
+
R1 R2
N
H
X
O
N
H
O
hexane-buffer
30 °C, 24 h
( )-16, 17 and 18
(S)-16, 17 and 18 (R)-10, 11 and 12
10, 16: R1 = Et, R2 = Ph
a: X =
b: X =
CH2CH2
CH2CH2CH2
11, 17: R1 = Me, R2 = CH2Ph
12, 18: R1 = Me, R2 = (CH2)2OBn
Entry
Substrate
ee of substrate/%c
ee of product/%d Conv.
E value
1
2
3
4
5
6
(ꢀ)-16a
(ꢀ)-16b
(ꢀ)-17a
(ꢀ)-17b
(ꢀ)-18a
(ꢀ)-18b
w0
97
10
95
19
23
68
85
94
>99
>99
>99
w0
0.53
0.09
0.49
0.16
0.19
<5
51
54
>747
>239
>249
1375, 1285, 1207, 1169, 1063, 959, 762, 700 cmꢂ1
;
1H NMR
(500 MHz, CDCl3)
d
1.53 (d, J¼6.5 Hz, 3H), 2.56e2.74 (m, 4H), 5.89
(q, J¼6.5 Hz, 1H), 7.21e7.41 (m, 5H); 13C NMR (125 MHz, CDCl3)
d
22.1, 28.9, 29.1, 72.9, 126.0, 127.9, 128.5, 141.3, 171.4, 178.2; HRMS
a
m/z (ESI) 245.0788 (calcd for C12H14O4Na: 245.0790, MþNaþ).
The other compounds (ꢀ)-2, 13, 14, and 15 were synthesized by
the same procedure. In the cases of (ꢀ)-2e, 2f, 2g, 13a, 13b, 14b, 15a,
and 15b, the products were directly extracted with AcOEt without
the extraction step using satd NaHCO3.
The reaction was performed using 5 mM of the substrate with Novozym 435 in
a mixed medium (hexane/0.1 M phosphate buffer (pH 6.5)¼9/1) for 24 h at 30 ꢁC.
b
The absolute configurations of the compounds 10e12 were determined by GC
analysis comparing the retention times to those of the authentic samples.
c
Determined by GC analysis after hydrolysis of the unreacted substrates.
Determined by GC analysis.
d

8064
M. Okudomi et al. / Tetrahedron 66 (2010) 8060e8067
4.2.2. 4-((1-Phenylethoxy)carbonyl)butanoic acid ((ꢀ)-2b). Yield
73% from (ꢀ)-1 with glutaric anhydride; IR (KBr) 2980, 2936, 1732,
1709, 1495, 1452, 1375, 1287, 1246, 1207, 1155, 1063, 935, 762,
170.15, 176.63 and 176.68; HRMS m/z (ESI) 389.1756 (calcd for
C
19H30O5SiNa: 389.1760, MþNaþ).
700 cmꢂ1
;
1H NMR (500 MHz, CDCl3)
d
1.53 (d, J¼6.5 Hz, 3H), 1.95
(tt, J1¼7.5 Hz, J2¼7.5 Hz, 2H), 2.35e2.48 (m, 4H), 5.89 (q, J¼6.5 Hz,
1H), 7.21e7.41 (m, 5H); 13C NMR (125 MHz, CDCl3)
19.8, 22.2, 32.9,
4.2.8. 3-((1-Phenylpropyloxy)carbonyl)propanoic acid ((ꢀ)-13a). Yield
98% from (ꢀ)-10 with succinic anhydride; IR (KBr) 2970, 2936, 2880,
1738, 1713, 1495, 1454, 1381, 1258, 1169, 1084, 964, 843, 758,
d
33.4, 72.5, 126.0, 127.9, 128.5, 141.5, 172.1, 179.0; HRMS m/z (ESI)
259.0953 (calcd for C13H16O4Na: 259.0946, MþNaþ).
700 cmꢂ1
;
1H NMR (500 MHz, CDCl3)
d
0.87 (t, J¼7.5 Hz, 3H),
1.75e1.98 (m, 2H), 2.55e2.74 (m, 4H), 5.68 (t, J¼7.0 Hz, 1H), 7.16e7.40
(m, 5H); 13C NMR (125 MHz, CDCl3)
9.8, 28.9, 29.1, 29.2, 77.9, 126.4,
d
4.2.3. 3-Methyl-4-((1-phenylethoxy)carbonyl)butanoic acid ((ꢀ)-2c).
Yield 82% from (ꢀ)-1 with 3-methylglutaric anhydride; IR (KBr)
2974, 2878, 1732, 1709, 1495, 1456, 1375, 1287, 1207, 1065, 974,
127.8, 128.3, 140.2, 171.5, 178.2; HRMS m/z (ESI) 259.0946 (calcd for
C13H16O4Na: 259.0946, MþNaþ).
935, 762, 700 cmꢂ1
;
1H NMR (500 MHz, CDCl3)
d
1.02 (d,
4.2.9. 4-((1-Phenylpropyloxy)carbonyl)butanoic acid ((ꢀ)-13b). Yield
73% from (ꢀ)-10 with glutaric anhydride; IR (KBr) 2970, 2938, 2878,
J¼6.5 Hz, 3H), 1.536 (d, J¼6.5 Hz) and 1.539 (d, J¼6.5 Hz) (3H),
2.20e2.34 (m, 2H), 2.36e2.54 (m, 3H), 5.90 (q, J¼6.5 Hz, 1H),
1734, 1709, 1495, 1452, 1383, 1246, 1204, 1084, 968, 758, 700 cmꢂ1
;
7.20e7.43 (m, 5H); 13C NMR (125 MHz, CDCl3)
d
19.66 and 19.71,
1H NMR (500 MHz, CDCl3)
1.87e2.00 (m, 4H), 2.34e2.49 (m, 4H), 5.67 (t, J¼7.0 Hz, 1H),
7.22e7.39 (m, 5H); 13C NMR (125 MHz, CDCl3)
9.9, 19.8, 29.3, 32.9,
d
0.88, (t, J¼7.5 Hz, 3H),1.76e1.86 (m, 1H),
22.18 and 22.20, 27.2, 40.42 and 40.47, 41.0, 72.4, 126.1, 127.9,
128.5, 141.5, 171.6, 178.4; HRMS m/z (ESI) 273.1096 (calcd for
C14H18O4Na: 273.1103, MþNaþ).
d
33.4, 77.5, 126.5, 127.8, 128.4, 140.4, 172.2, 179.1; HRMS m/z (ESI)
273.1104 (calcd for C14H18O4Na: 273.1103, MþNaþ).
4.2.4. 3,3-Dimethyl-4-((1-phenylethoxy)carbonyl)butanoic
acid
((ꢀ)-2d). Yield 36% from (ꢀ)-1 with 3,3-dimethylglutaric anhy-
4.2.10. 3-((1-Methyl-2-phenylethoxy)carbonyl)propanoic
acid
dride; IR (KBr) 2963, 2876, 1734, 1707, 1495, 1450, 1371, 1287,
((ꢀ)-14a). Yield 90% from (ꢀ)-11 with succinic anhydride; IR (KBr)
1234, 1152, 1063, 964, 760, 698 cmꢂ1
;
1H NMR (500 MHz, CDCl3)
2980, 2932, 1730, 1713, 1497, 1454, 1377, 1233, 1173, 1049, 962, 845,
d
1.11 (s, 3H), 1.12 (s, 3H), 1.54 (d, J¼6.5 Hz, 3H), 2.42 (d, J¼14.5 Hz,
748, 702 cmꢂ1
;
1H NMR (500 MHz, CDCl3)
d
1.22 (d, J¼6.5 Hz, 3H),
1H), 2.44 (d, J¼14.5 Hz, 1H), 2.46 (d, J¼14.0 Hz, 1H), 2.49 (d,
2.49e2.69 (m, 4H), 2.77 (dd, J1¼6.5 Hz, J2¼13.5 Hz, 1H), 2.91 (dd,
J¼14.0 Hz, 1H), 5.90 (q, J¼6.5 Hz, 1H), 7.21e7.42 (m, 5H); 13C NMR
J1¼6.5 Hz, J2¼13.5 Hz, 1H), 5.14 (qt, J1¼6.0 Hz, J2¼6.5 Hz, 1H),
(125 MHz, CDCl3)
d
22.2, 27.72, 27.74, 32.6, 45.0, 45.2, 72.4, 126.1,
7.15e7.34 (m, 5H); 13C NMR (125 MHz, CDCl3)
d 19.3, 28.9, 29.1, 42.1,
127.9, 128.4, 141.4, 171.4, 177.5; HRMS m/z (ESI) 287.1257 (calcd
72.0, 126.5, 128.3, 129.4, 137.4, 171.5, 178.1; HRMS m/z (ESI) 259.0947
for C15H20O4Na: 287.1259, MþNaþ).
(calcd for C13H16O4Na: 259.0946, MþNaþ).
4.2.5. 2-(1-(((1-Phenylethoxy)carbonyl)methyl)cyclopentyl)acetic
acid ((ꢀ)-2e). Yield 59% from (ꢀ)-1 with 3,3-tetramethyleneglu-
taric anhydride; IR (KBr) 2955, 2872, 1732, 1705, 1495, 1454, 1371,
4.2.11. 4-((1-Methyl-2-phenylethoxy)carbonyl)butanoic acid ((ꢀ)-14b).
Yield 75% from (ꢀ)-11 with glutaric anhydride; IR (KBr) 2978, 2934,
1728, 1713, 1497, 1454, 1379, 1248, 1155, 1057, 943, 822, 746, 700 cmꢂ1
;
1285, 1209, 1161, 1062, 953, 762, 698 cmꢂ1
;
1H NMR (500 MHz,
1H NMR (300 MHz, CDCl3)
d
1.23 (d, J¼6.0 Hz, 3H), 1.80e1.94 (m, 4H),
CDCl3)
d
1.53 (d, J¼6.5 Hz, 3H),1.55e1.69 (m, 8H), 2.54 (d, J¼15.5 Hz,
2.25e2.39 (m, 2H), 2.77 (dd, J1¼6.0 Hz, J2¼13.5 Hz, 1H), 2.90 (dd,
J1¼7.0 Hz, J2¼13.5 Hz, 1H), 5.14 (qt, J1¼6.0 Hz, J2¼6.0 Hz, 1H), 7.14e7.33
1H), 2.55 (d, J¼15.5 Hz, 1H), 2.57 (d, J¼14.5 Hz, 1H), 2.60 (d,
J¼15.0 Hz, 1H), 5.89 (q, J¼6.5 Hz, 1H), 7.20e7.41 (m, 5H); 13C NMR
(m, 5H); 13C NMR (125 MHz, CDCl3)
d 19.5, 19.7, 32.8, 33.4, 42.2, 71.6,
(125 MHz, CDCl3)
d
22.2, 23.9, 37.98, 38.02, 42.1, 42.3, 43.0, 72.4,
126.5, 128.3, 129.3, 137.5, 172.3, 178.9; HRMS m/z (ESI) 273.1091 (calcd
126.1, 127.8, 128.4, 141.5, 171.8, 177.7; HRMS m/z (ESI) 313.1411
for C14H18O4Na: 273.1103, MþNaþ).
(calcd for C17H22O4Na: 313.1416, MþNaþ).
4.2.12. 3-((1-Methyl-4-(benzyloxy)propyloxy)carbonyl)propanoic
acid ((ꢀ)-15a). Yield 62% from (ꢀ)-12 with succinic anhydride; IR
(KBr) 2978, 2932, 2870, 1732, 1713, 1497, 1454, 1377, 1258, 1171,
4.2.6. 2-((1-Phenylethoxy)carbonyl)benzoic acid ((ꢀ)-2f). Yield 96%
from (ꢀ)-1 with phthalic anhydride; IR (KBr) 2976, 2833, 2683,
2581, 1724, 1694, 1601, 1493, 1449, 1418, 1315, 1288, 1258, 1125,
1099, 1028, 961, 739, 698 cmꢂ1; 1H NMR (500 MHz, CDCl3)
d 1.24 (d,
1063, 932, 795, 743, 698 cmꢂ1
;
1H NMR (500 MHz, CDCl3)
d
1.66
J¼6.5 Hz, 3H), 1.76e1.95 (m, 2H), 2.46e2.68 (m, 4H), 3.43e3.55 (m,
(d, J¼6.5 Hz, 3H), 6.14 (q, J¼6.5 Hz,1H), 7.26 (tt, J1¼1.5 Hz, J2¼7.5 Hz,
1H), 7.34 (tt, J1¼1.5 Hz, J2¼7.5 Hz, 2H), 7.41 (td, J1¼1.5 Hz, J2¼8.0 Hz,
2H), 7.56 (td, J1¼1.5 Hz, J2¼7.5 Hz, 1H), 7.59 (td, J1¼1.5 Hz, J2¼7.5 Hz,
1H), 7.69 (dd, J1¼2.0 Hz, J2¼7.0 Hz, 1H), 7.88 (dd, J1¼2.0 Hz,
2H), 4.47 (d, J¼12.0 Hz, 1H), 4.48 (d, J¼12.0 Hz, 1H), 5.05e5.16 (m,
1H), 7.22e7.39 (m, 5H); 13C NMR (75 MHz, CDCl3)
d 20.1, 28.9, 29.1,
35.9, 66.4, 69.1, 73.0, 127.6, 127.7, 128.3, 138.2, 171.6, 178.1; HRMS
m/z (ESI) 303.1181 (calcd for C14H18O4Na: 303.1203, MþNaþ).
J2¼7.0 Hz, 1H); 13C NMR (125 MHz, CDCl3)
d 21.7, 74.2, 126.2, 128.0,
128.5, 128.8, 129.7, 130.0, 130.8, 132.1, 133.4, 141.0, 167.2, 172.4;
HRMS m/z (ESI) 293.0779 (calcd for C16H14O4Na: 293.0790,
MþNaþ).
4.2.13. 4-((1-Methyl-4-(benzyloxy)propyloxy)carbonyl)butanoic acid
((ꢀ)-15b). Yield 65% from (ꢀ)-12 with glutaric anhydride; IR
(KBr) 2976, 2936, 2870, 1732, 1709, 1497, 1452, 1377, 1248, 1206,
1146, 1099, 1028, 941, 739, 698 cmꢂ1 1H NMR (500 MHz, CDCl3)
;
4.2.7. 3-(tert-Butyldimethylsilyloxy)-4-((1-phenylethoxy)carbonyl)
butanoic acid ((ꢀ)-2g). Yield 94% from (ꢀ)-1 with 3-(tert-butyl-
dimethylsilyloxy)glutaric anhydride; IR (KBr) 2930, 2857, 1736,
1713, 1495, 1462, 1377, 1256, 1204, 1099, 1063, 972, 837, 779,
d
1.24 (d, J¼6.0 Hz, 3H), 1.71e2.02 (m, 4H), 2.19e2.51 (m, 4H),
3.39e3.58 (m, 2H), 4.47 (d, J¼12.0 Hz, 1H), 4.48 (d, J¼12.0 Hz, 1H),
5.01e5.17 (m, 1H), 7.15e7.47 (m, 5H); 13C NMR (75 MHz, CDCl3)
d
19.8, 20.2, 32.9, 33.4, 35.9, 66.4, 68.7, 73.0, 127.6, 127.7, 128.3,
698 cmꢂ1; 1H NMR (500 MHz, CDCl3)
d
0.01 (s), 0.059 (s) and 0.065
138.2, 172.4, 178.8; HRMS m/z (ESI) 317.1337 (calcd for
(s) (6H), 0.81 (s) and 0.84 (s, 9H), 1.54 (d, J¼6.5 Hz, 3H), 2.50e2.66
(m, 4H), 4.50e4.58 (m, 1H), 5.883 (q, J¼6.5 Hz) and 5.887 (q,
J¼6.5 Hz) (1H), 7.25e7.38 (m, 5H); 13C NMR (125 MHz, CDCl3)
C14H18O4Na: 317.1359, MþNaþ).
4.3. Preparation of MPEG5000-supported compounds
d
ꢂ5.05 and ꢂ5.03, ꢂ4.92 and ꢂ4.91, 17.8 and 17.9, 22.2 and 22.3,
25.59 and 25.62, 42.0 and 42.1, 42.4 and 42.5, 65.98 and 66.03,
72.7, 126.07 and 126.12, 127.9, 128.5, 141.38 and 141.41, 170.10 and
4.3.1. Compound (ꢀ)-7a. Under an argon atmosphere, methane-
sulfonyl chloride (0.35 mL, 4.51 mmol) and triethylamine (1.50 mL,

M. Okudomi et al. / Tetrahedron 66 (2010) 8060e8067
8065
10.8 mmol) were added to a solution of 3 (3.00 g, 0.60 mmol) in
CH2Cl2 (13 mL), and the solution was stirred overnight at room tem-
perature. Afterevaporationinvacuo, theresiduewaspouredintoEt2O
to precipitate a white solid. After the solid was washed with 2-
propanol, evaporation and precipitation gave the mesylate 4 as
a white solid in 99% yield (3.03 g; purity, ca. >99%); IR (KBr) 2886,
40.50, 40.55, 40.9, 58.9, 63.2, 68.9, 70.4 (PEG), 70.6, 71.8, 72.1, 125.9,
127.7, 128.3, 141.45, 141.47, 171.4, 172.1; ESI-TOF MS m/z 2468 Da
(2466 Da calcd for [CH3(OCH2CH2)105C14H18NO3$2Na]2þ).
4.3.4. Compound (ꢀ)-7d. Yield 93% (purity, ca. 98%) from 6 with
(ꢀ)-2d; IR (KBr) 2886, 1732, 1653, 1466, 1342, 1281, 1242, 1109, 962,
1636, 1466, 1360, 1342, 1281, 1242, 1113, 962, 843 cmꢂ1
;
1H NMR
843 cmꢂ1; 1H NMR (500 MHz, CDCl3)
d 1.05 (s, 3H), 1.09 (s, 3H), 1.55
(500 MHz, CDCl3)
d
3.09 (s, 3H) 3.38 (s, 3H), 3.46e3.82 (m, PEG-
(d, J¼6.5 Hz, 3H), 2.21 (s, 2H), 2.39 (d, J¼13.5 Hz, 1H), 2.42 (d,
J¼13.5 Hz, 1H), 3.38 (s, 3H), 3.39e3.44 (m, 2H), 3.47e3.81 (m, PEG-
methylenes), 5.89 (q, J¼6.5 Hz, 1H), 6.77 (br s, 1H), 7.15e7.41 (m,
methylenes), 4.39 (t, J¼4.5 Hz, 2H); 13C NMR (125 MHz, CDCl3)
d 37.6,
58.9, 68.9, 69.2, 70.4 (PEG), 70.5, 70.6, 71.8.
Under an argon atmosphere, sodium azide (256 mg, 3.94 mmol)
was added to a solution of 4 (4.00 g, 0.788 mmol) in toluene
(10.5 mL), and the solution was stirred overnight at 95 ꢁC. The
mixture was evaporated in vacuo, and the residue was filtrated
through a Celite pad. After evaporation, the residue was poured into
Et2O to precipitate the compound 5 as a white solid in 95% yield
(3.75 g; purity, ca. >99%); IR (KBr) 2886, 2100,1636,1468,1342,1281,
5H); 13C NMR (125 MHz, CDCl3)
d 22.1, 28.26, 28.27, 33.4, 38.8, 45.0,
47.2, 58.9, 69.8, 70.0, 70.4 (PEG), 70.6, 71.8, 72.3, 126.0, 127.8, 128.4,
141.3, 171.0, 171.9; ESI-TOF MS m/z 2475 Da (2473 Da calcd for
[CH3(OCH2CH2)105C15H20NO3$2Na]2þ).
4.3.5. Compound (ꢀ)-7e. Yield 88% (purity, ca. 83%) from 6 with
(ꢀ)-2d; IR (KBr) 2886, 1732, 1653, 1466, 1342, 1281, 1242, 1113, 964,
1242, 1115, 963, 843 cmꢂ1; 1H NMR (500 MHz, CDCl3)
d
3.38 (s, 3H),
843 cmꢂ1 1H NMR (500 MHz, CDCl3)
; d 1.40e1.73 (m, 8H), 1.55 (d,
3.40 (t, J¼5.0 Hz, 2H), 3.46e3.82 (m, PEG-methylenes); 13C NMR
(125 MHz, CDCl3) 50.5, 58.9, 69.9, 70.4 (PEG), 70.55, 70.62, 71.8.
Under an argon atmosphere, DIBAL-H (7.1 mL, 1.0 M solution in
toluene) was added to a solution of 5 (2.70 g, 0.537 mmol) in CH2Cl2
(22 mL) at 0 ꢁC, and the solution was warmed to room temperature.
After stirring overnight, the reaction was quenched with water, and
the suspension was filtrated through a Celite pad. After evapora-
tion, the residue was poured into Et2O to precipitate the compound
6 as a white solid in 87% yield (2.33 g; purity, ca. 97%); IR (KBr)
J¼6.5 Hz, 3H), 2.29 (s, 2H), 2.48 (d, J¼14.0 Hz, 1H), 2.49 (d,
J¼14.0 Hz, 1H), 3.38 (s, 3H), 3.45e3.83 (m, PEG-methylenes), 5.89
(q, J¼6.5 Hz, 1H), 6.78 (br s, 1H), 7.21e7.38 (m, 5H); 13C NMR
(125 MHz, CDCl3)
d 22.1, 23.7, 37.92, 37.98, 38.8, 42.2, 44.1, 58.9,
69.8, 70.0, 70.4 (PEG), 70.6, 71.8, 72.4, 126.0, 127.8, 128.4, 141.4,
171.5, 172.3; ESI-TOF MS m/z 2487 Da (2486 Da calcd for
[CH3(OCH2CH2)105C17H22NO3$2Na]2þ).
4.3.6. Compound (ꢀ)-7f. Yield 79% (purity, ca. 48%) from 6 with
(ꢀ)-2f; IR (KBr) 2886, 1715, 1653, 1466, 1342, 1281, 1242, 1113, 964,
3464, 2887, 1651, 1468, 1342, 1281, 1242, 1113, 964, 843 cmꢂ1 1H
;
NMR (500 MHz, CDCl3)
d
2.88 (t, J¼5.0 Hz, 2H), 3.38 (s, 3H),
843 cmꢂ1; 1H NMR (500 MHz, CDCl3)
d
1.64 (d, J¼6.5 Hz, 3H), 3.38
3.47e3.83 (m, PEG-methylenes); 13C NMR (125 MHz, CDCl3)
d
41.6,
(s, 3H), 3.46e3.83 (m, PEG-methylenes), 6.09 (q, J¼6.5 Hz, 1H), 6.64
(br s,1H), 7.22e7.56 (m, 7H), 7.72 (dd, J1¼3.0 Hz, J2¼5.5 Hz,1H), 7.85
58.9, 70.1, 70.4 (PEG), 70.6, 71.8, 72.9.
Under an argon atmosphere, (ꢀ)-2a (269.2 mg, 1.211 mmol),
DCC (261.6 mg, 1.268 mmol) and DMAP (73.2 mg, 0.599 mmol)
were added to a solution of 6 (2.000 g, 0.4001 mmol) in CH2Cl2
(13 mL), and the solution was stirred overnight at room tempera-
ture. After evaporation, the residue was poured into Et2O to pre-
cipitate the compound (ꢀ)-7a as a white solid in 95% yield (1.985 g;
purity, ca. 84%); IR (KBr) 2886, 1734, 1653, 1466, 1342, 1281, 1242,
(dd, J1¼3.0 Hz, J2¼5.5 Hz, 1H); 13C NMR (125 MHz, CDCl3)
d 22.1,
59.0, 64.5, 68.7, 70.4, 70.7, 71.8, 73.7, 126.1, 127.9, 128.4, 128.8, 129.0,
130.8, 131.1, 131.7, 132.3, 141.2, 166.4, 167.6; ESI-TOF MS m/z 2478 Da
(2476 Da calcd for [CH3(OCH2CH2)105C16H14NO3$2Na]2þ).
4.3.7. Compound (ꢀ)-7g. Yield 91% (purity, ca. 80%) from 7 with
(ꢀ)-2g; IR (KBr) 2884, 1734, 1651, 1468, 1342, 1281, 1242, 1115, 962,
1113, 964, 843 cmꢂ1; 1H NMR (500 MHz, CDCl3)
d
1.53 (d, J¼6.5 Hz,
843 cmꢂ1; 1H NMR (500 MHz, CDCl3)
d 0.03 (s), 0.041 (s), 0.047 (s),
3H), 2.44 (dt, J1¼7.0 Hz, J2¼15.0 Hz, 1H), 2.49 (dt, J1¼7.0 Hz,
J2¼15.0 Hz, 1H), 2.65 (td, J1¼7.0 Hz, J2¼17.0 Hz, 1H), 2.73 (td,
J1¼7.5 Hz, J2¼17.0 Hz, 1H), 3.38 (s, 3H), 3.40e3.46 (m, 2H),
3.47e3.82 (m, PEG-methylenes), 5.87 (q, J¼6.5 Hz, 1H), 6.50 (br s,
0.057 (s) and 0.062 (s) (6H), 0.79 (s), 0.826 (s), 0.829 (s) and 0.84 (s)
(9H), 1.54 (d, J¼6.5 Hz, 3H), 2.31e2.65 (m, 4H), 3.38 (s, 3H),
3.43e3.85 (m, PEG-methylenes), 4.49e4.58 (m, 1H), 5.87 (q,
J¼6.5 Hz,1H), 6.55 (br s,1H), 7.21e7.39 (m, 5H); 13C NMR (125 MHz,
1H), 7.23e7.38 (m, 5H); 13C NMR (125 MHz, CDCl3)
d
22.2, 29.7, 30.7,
CDCl3)
d
ꢂ5.07, ꢂ5.02, ꢂ4.96, ꢂ4.94, 17.76, 17.79, 22.2, 22.3, 25.59,
39.1, 58.9, 69.7, 70.0, 70.4 (PEG), 70.6, 71.8, 72.4, 125.9, 127.7, 128.3,
141.5, 171.3, 172.0; ESI-TOF MS m/z 2454 Da (2452 Da calcd for
[CH3(OCH2CH2)105C12H14NO3$2Na]2þ).
Other MPEG5000-supported compounds were synthesized by
the same procedure.
25.61, 39.0, 41.9, 42.1, 43.9, 44.1, 58.9, 66.55, 66.59, 69.7, 70.1, 70.4
(PEG), 70.5, 70.6, 71.8, 72.5, 125.98, 126.04, 127.8, 128.4, 141.39,
141.43, 170.07, 170.13, 170.8, 170.9; ESI-TOF MS m/z 2525 Da (2524
Da calcd for [CH3(OCH2CH2)105C19H30NO4Si$2Na]2þ).
4.3.8. Compound (ꢀ)-16a. Yield 95% (purity, ca. 89%) from 6 with
(ꢀ)-13a; IR (KBr) 2886, 1732, 1651, 1468, 1342, 1281, 1242, 1111, 962,
4.3.2. Compound (ꢀ)-7b. Yield 71% (purity, ca. 70%) from 6 with
(ꢀ)-2b; IR (KBr) 2886, 1732, 1651, 1468, 1342, 1281, 1242, 1113, 962,
843 cmꢂ1
;
1H NMR (500 MHz, CDCl3)
d
0.87 (t, J¼7.5 Hz, 3H),
843 cmꢂ1
;
1H NMR (500 MHz, CDCl3)
d
1.53 (d, J¼6.5 Hz, 3H),
1.75e1.97 (m, 2H), 2.45 (dt, J1¼7.0 Hz, J2¼15.5 Hz, 1H), 2.48 (dt,
J1¼7.0 Hz, J2¼15.5 Hz, 1H), 2.66 (td, J1¼7.0 Hz, J2¼17.0 Hz, 1H), 2.76
(td, J1¼7.5 Hz, J2¼17.0 Hz, 1H), 3.38 (s, 3H), 3.39e3.45 (m, 2H),
3.46e3.83 (m, PEG-methylenes), 5.65 (t, J¼7.0 Hz, 1H), 6.23 (br s,
1.90e2.00 (m, 2H), 2.16e2.23 (m, 2H), 2.35e2.43 (m, 2H), 3.37 (s,
3H), 3.39e3.45 (m, 2H), 3.46e3.81 (m, PEG-methylenes), 5.88 (q,
J¼6.5 Hz, 1H), 6.12 (br s, 1H), 7.18e7.38 (m, 5H); 13C NMR (125 MHz,
CDCl3)
d
20.7, 22.2, 33.6, 35.2, 39.0, 58.9, 69.7, 70.0, 70.4 (PEG), 70.6,
1H), 7.20e7.38 (m, 5H); 13C NMR (125 MHz, CDCl3)
d 9.8, 29.2, 29.7,
71.8, 72.1, 125.9, 127.7, 128.4, 141.6, 172.1, 172.3; ESI-TOF MS m/z
30.7, 39.1, 58.9, 69.7, 70.1, 70.4 (PEG), 70.6, 71.8, 77.4, 126.3, 127.6,
128.2, 140.4, 171.3, 172.1; ESI-TOF MS m/z 2461 Da (2459 Da calcd
for [CH3(OCH2CH2)105C13H16NO3$2Na]2þ).
2461 Da (2459 Da calcd for [CH3(OCH2CH2)105C13H16NO3$2Na]2þ).
4.3.3. Compound (ꢀ)-7c. Yield 90% (purity, ca. 61%) from 6 with
(ꢀ)-2c; IR (KBr) 2886, 1732, 1651, 1468, 1342, 1281, 1242, 1115, 964,
4.3.9. Compound (ꢀ)-16b. Yield 83% (purity, ca. 76%) from 6 with
(ꢀ)-13b; IR (KBr) 2886, 1732, 1647, 1466,1342, 1281, 1242,1113, 964,
843; 1H NMR (500 MHz, CDCl3)
d
1.02 (d, J¼6.5 Hz, 3H), 1.53 (d,
J¼6.5 Hz, 3H), 2.16e2.46 (m, 5H), 3.38 (s, 3H), 3.39e3.83 (m, PEG-
843 cmꢂ1
;
1H NMR (500 MHz, CDCl3)
d
0.88 (t, J¼7.5 Hz, 3H),
methylenes), 5.88 (q, J¼6.5 Hz, 1H), 7.09 (br s, 1H), 7.22e7.39 (m,
1.75e1.86 (m, 1H), 1.87e2.00 (m, 3H), 2.15e2.24 (m, 2H), 2.33e2.48
(m, 2H), 3.38 (s, 3H), 3.40e3.46 (m, 2H), 3.47e3.82 (m, PEG-
5H); 13C NMR (125 MHz, CDCl3)
d 19.51, 19.55, 22.12, 22.14, 27.3,

8066
M. Okudomi et al. / Tetrahedron 66 (2010) 8060e8067
methylenes), 5.66 (t, J¼7.0 Hz, 1H), 6.28 (br s, 1H), 7.18e7.39 (m,
5H); 13C NMR (125 MHz, CDCl3)
9.8, 20.8, 29.2, 33.5, 35.2, 39.0,
4.5. Typical experimental procedure for enantioselective
hydrolysis of MPEG-supported substrates
d
58.9, 69.7, 70.1, 70.4 (PEG), 70.5, 70.6, 71.8, 77.2, 126.4, 127.7, 128.3,
140.4, 172.0, 172.4; ESI-TOF MS m/z 2468 Da (2466 Da calcd for
[CH3(OCH2CH2)105C14H18NO3$2Na]2þ).
To a test tube containing 125 mg of (ꢀ)-7a (sub. concn, 5 mM)
was added 4.5 mL of hexane and 0.5 mL of 0.1 M phosphate buffer
(pH 6.5). To the mixture was added 20 mg of Novozym 435 (2.0 U/
mg, using tributyrin at pH 8.0 and 40 ꢁC), and the solution was
stirred for 24 h at 30 ꢁC. After the organic layer was separated and
evaporated in vacuo, the ee of the resulting (R)-1 (98% ee) in the
residue was determined by GC analysis. On the other hand, the
aqueous layer was diluted with CH2Cl2, and dried over Na2SO4.
After evaporation, the residue was hydrolyzed with 2 M NaOH aq
(1.0 mL) in MeOH (4.0 mL). The products were extracted with
hexane (ꢃ3), and the organic layer was dried over Na2SO4. After
evaporation, the ee of the corresponding (S)-1 (55% ee) was de-
termined by GC analysis.
4.3.10. Compound (ꢀ)-17a. Yield 94% (purity, ca. 74%) from 6 with
(ꢀ)-14a; IR (KBr) 2886, 1734, 1655, 1468, 1342, 1281, 1242, 1115, 962,
843 cmꢂ1
;
1H NMR (500 MHz, CDCl3)
d
1.21 (d, J¼6.0 Hz, 3H),
2.39e2.47 (m, 2H), 2.53e2.69 (m, 2H), 2.76 (dd, J1¼7.0 Hz, J2¼13.5 Hz,
1H), 2.92 (dd, J1¼6.5 Hz, J2¼13.5 Hz, 1H), 3.38 (s, 3H), 3.41e3.46 (m,
2H), 3.48e3.82 (m, PEG-methylenes), 5.11 (qt, J1¼6.0 Hz, J2¼6.5 Hz,
1H), 6.25 (br s, 1H), 7.15e7.33 (m, 5H); 13C NMR (75 MHz, CDCl3)
d
19.2, 29.7, 30.7, 39.2, 42.0, 58.9, 69.7, 70.0, 70.4, 70.6, 71.6, 71.8,
126.3, 128.2, 129.3, 137.4, 171.4, 172.2; ESI-TOF MS m/z 2460 Da (2459
Da calcd for [CH3(OCH2CH2)105C13H16NO3$2Na]2þ).
Other reactions were carried out by the same procedure.
4.3.11. Compound (ꢀ)-17b. Yield 95% (purity, ca. 83%) from 6 with
(ꢀ)-14b; IR (KBr) 2886,1734,1653,1466,1342,1281,1242,1115, 964,
4.6. Typical preparative scale reaction of MPEG-supported
substrates
843 cmꢂ1
;
1H NMR (500 MHz, CDCl3)
d
1.23 (d, J¼6.5 Hz, 3H),
1.83e1.94 (m, 2H), 2.10 (t, J¼7.5 Hz, 2H), 2.30 (dt, J1¼3.5 Hz,
J2¼7.5 Hz, 2H), 2.77 (dd, J1¼6.5 Hz, J2¼13.5 Hz, 1H), 2.89 (dd,
J1¼7.0 Hz, J2¼13.5 Hz, 1H), 3.38 (s, 3H), 3.41e3.46 (m, 2H),
3.47e3.83 (m, PEG-methylenes), 5.14 (qt, J1¼6.0 Hz, J2¼6.5 Hz, 1H),
To a recovery flask containing 3.0 g of (ꢀ)-7b (sub. concn, 5 mM)
was added 108 mL of hexane and 12 mL of 0.1 M phosphate buffer.
To the mixture was added 479.5 mg of Novozym 435, and the so-
lution was stirred for 24 h at 30 ꢁC. After the organic layer was
separated and evaporated in vacuo, the residue was purified by
preparative TLC (hexane/AcOEt¼3/1) to give (R)-1 (17.7 mg, 25%,
6.08 (br s, 1H), 7.15e7.34 (m, 5H); 13C NMR (125 MHz, CDCl3)
d 19.5,
20.7, 33.6, 35.1, 39.0, 42.1, 58.9, 69.8, 70.1, 70.4, 70.6, 71.2, 71.8,126.3,
128.2, 129.3, 137.5, 172.1, 172.5; ESI-TOF MS m/z 2468 Da (2466 Da
calcd for [CH3(OCH2CH2)105C14H18NO3$2Na]2þ).
20
20
99% ee, [
a
]
þ34.5 (c 1.77, MeOH)); lit.8
[a
]
þ45 (c 5.15, MeOH).
D
D
On the other hand, the aqueous layer was diluted with CH2Cl2, and
dried over Na2SO4. After the solution was evaporated in vacuo, the
residue was poured into Et2O to precipitate the compound (S)-7b as
a white solid. To a solution of (S)-7b in MeOH (96 mL) was added
2 M NaOH aq (24 mL), and the mixture was stirred for 1 h at room
temperature. The products were extracted with ether (ꢃ3), and the
organic layer was washed with brine and dried over Na2SO4. After
evaporation, the residue was purified by column chromatography
4.3.12. Compound (ꢀ)-18a. Yield 80% (purity, ca. 67%) from 6 with
(ꢀ)-15a; IR (KBr) 2884,1732, 1676,1468,1342, 1281, 1242, 1113, 962,
843 cmꢂ1
;
1H NMR (500 MHz, CDCl3)
d
1.24 (d, J¼6.0 Hz, 3H),
1.76e1.95 (m, 2H), 2.41e2.49 (m, 2H), 2.55e2.63 (m, 2H), 3.38 (s,
3H), 3.40e3.82 (m, PEG-methylenes), 4.47 (d, J¼12.0 Hz, 1H), 4.48
(d, J¼12.0 Hz, 1H), 5.08 (q, J¼6.5 Hz, 1H), 6.46 (br s, 1H), 7.22e7.38
(m, 5H); 13C NMR (75 MHz, CDCl3)
d 20.1, 29.7, 30.7, 35.9, 39.3, 58.9,
66.5, 68.7, 69.6, 70.2, 70.5, 70.8, 71.8, 72.9, 127.5, 127.6, 128.2, 138.3,
171.6, 172.2; ESI-TOF MS m/z 2482 Da (2481 Da calcd for
[CH3(OCH2CH2)105C15H20NO4$2Na]2þ).
(hexane/EtOAc¼2/1) on silica gel to give the corresponding (S)-1
21
(16.8 mg, 24%, 94% ee, [
a
]
ꢂ21.0 (c 1.68, MeOH)).
D
4.7. Several data of alcohols
4.3.13. Compound (ꢀ)-18b. Yield 74% (purity, ca. 69%) from 6 with
(ꢀ)-15b; IR (KBr) 2884,1730, 1668,1466,1342,1281, 1242,1111, 962,
4.7.1. 1-Phenylethanol (1). The spectral data were in full agreement
with that of a commercial source. GC conditions: column, CP-Cy-
clodextrin-B-236-M19 (Chrompack), 0.25 mmꢃ50 m; injection,
160 ꢁC; detection, 160 ꢁC; oven, 140 ꢁC; carrier gas, He; head pres-
sure, 2.4 kg/cm2; retention time, 8.9 (R) and 9.2 (S) min.
843 cmꢂ1
;
1H NMR (500 MHz, CDCl3)
d
1.23 (d, J¼6.0 Hz, 3H),
1.75e2.00 (m, 4H), 2.21 (t, J¼7.5 Hz, 2H), 2.30 (t, J¼7.5 Hz, 2H), 3.38
(s, 3H), 3.42e3.83 (m, PEG-methylenes), 4.47 (d, J¼12.0 Hz, 1H),
4.48 (d, J¼12.0 Hz, 1H), 5.08 (q, J¼6.5 Hz, 1H), 6.33 (br s, 1H),
7.22e7.38 (m, 5H); 13C NMR (125 MHz, CDCl3)
d 20.2, 20.9, 33.6,
35.1, 35.9, 39.2, 58.9, 66.4, 68.4, 69.6, 70.1, 70.4, 70.8, 71.8, 72.9,
127.5, 127.6, 128.3, 138.2, 172.3, 172.5; ESI-TOF MS m/z 2489 Da
(2488 Da calcd for [CH3(OCH2CH2)105C16H22NO4$2Na]2þ).
4.7.2. 1-Phenyl-1-propanol (10). The spectral data were in full
agreement with that of a commercial source. GC conditions: column,
CP-Cyclodextrin-B-236-M19 (Chrompack), 0.25 mmꢃ50 m; injec-
tion, 140 ꢁC; detection, 140 ꢁC; oven, 120 ꢁC; carrier gas, He; head
pressure, 2.4 kg/cm2; retention time, 22.6 (R) and 23.4 (S) min. The
4.4. Screening of enzymes
authentic sample (R)-10: [
a
]
D
30 þ41.4 (c 1.20, CHCl3) (94% ee); lit.9 (S)-
In the screening test, we used the following enzymes: Lipase
immobilized from C. antarctica (Novozym 435, BioChemika), Li-
pase from porcine pancreas (PPL; Type II), Lipase Type VII from
Candida cylindracea (Sigma), Lipase A, Lipase D, Lipase D-360,
Lipase PS, Newlase F, PLE (Amano Enzyme, Inc.), Lipase OF (Meito
Sangyo Co., Ltd.), Esterase SNSM-87, Lilipase (Nagase & Co., Ltd.),
form, [
a]
20 ꢂ47.0 (c 1.00, CHCl3).
D
4.7.3. 1-Phenyl-2-propanol (11). The spectral data were in full
agreement with that of a commercial source. GC conditions: column,
CP-Cyclodextrin-B-236-M19 (Chrompack), 0.25 mmꢃ50 m; injec-
tion, 130 ꢁC; detection, 130 ꢁC; oven, 110 ꢁC; carrier gas, He; head
a
-Chymotrypsin (E. Merck). To a test tube containing 62.5 mg of
pressure, 2.4 kg/cm2; retention time, 27.1 (R) and 27.5 (S) min. The
30
(ꢀ)-9a (sub. concn, 5 mM) was added 2.5 mL of 0.1 M phosphate
buffer (pH 6.5). To the mixture was added 10 mg of Enzyme, and
the solution was incubated for 24 h at 30 ꢁC. The products were
extracted with 2.5 mL of Et2O, and detected by TLC (hexane/
AcOEt¼3/1). The ee of the product was determined by GC
analysis.
authentic sample (R)-11: [
a]
ꢂ32.8 (c 0.67, CHCl3) (99% ee); lit.10
D
(R)-form, [
a]
25 ꢂ37.57 (c 5.00, CHCl3).
D
4.7.4. 4-Benzyloxy-2-butanol (12). The spectral data were in full
agreement with those reported.5b HPLC conditions: column,
CHIRALCEL OD-H (Daicel Chemical Industries, Ltd.); eluent, hexane/

M. Okudomi et al. / Tetrahedron 66 (2010) 8060e8067
8067
Tetrahedron 2007, 63, 1721; (j) Gotor, V.; Alfonso, I.; Garcia-Urdiales, E. Asym-
metric Organic Synthesis with Enzymes; Wiley-VCH: Weinheim, 2008.
2. (a) Gravert, D. J.; Janda, K. D. Chem. Rev. 1997, 97, 489; (b) Wentworth, P.; Janda,
K. D. Chem. Commun. 1999, 1917; (c) Dickerson, T. J.; Reed, N. N.; Janda, K. D.
Chem. Rev. 2002, 102, 3325; (d) Bergbreiter, D. E. Chem. Rev. 2002, 102, 3345.
3. (a) Benaglia, M.; Celentano, G.; Cozzi, F. Adv. Synth. Catal. 2001, 343, 171; (b)
Pozzi, G.; Cavazzini, M.; Quici, S.; Benaglia, M.; Dell’Anna, G. Org. Lett. 2004, 6,
441; (c) Oikawa, M.; Tanaka, T.; Kusumoto, S.; Sasaki, M. Tetrahedron Lett. 2004,
45, 787; (d) Reed, N. N.; Dickerson, T. J.; Boldt, G. E.; Janda, K. D. J. Org. Chem.
2005, 70, 1728; (e) Li, J.-H.; Hu, X.-C.; Liang, Y.; Xie, Y.-X. Tetrahedron 2006, 62,
31; (f) Hong, S. H.; Grubbs, R. H. Org. Lett. 2007, 9, 1955; (g) Tocco, G.; Fais, A.;
Meli, G.; Begala, M.; Podda, G.; Fadda, M. B.; Corda, M.; Attanasi, O. A.;
Filippone, P.; Berretta, S. Bioorg. Med. Chem. 2009, 19, 36; (h) Tocco, G.; Car-
nonaro, C. M.; Meli, G.; Podda, G. Molecules 2009, 14, 1044; (i) Zimmer, R.;
Dekaris, V.; Knauer, M.; Schefzig, L.; Reissig, H.-U. Synth. Commun. 2009, 39,1012.
4. (a) Veronese, F. M.; Morpurgo, M. Il Farmaco 1999, 54, 497; (b) Greenwald, R. B.
J. Controlled Release 2001, 74, 159.
2-propanol¼90/10; flow rate, 0.5 mL/min; 254 nm; temperature,
25 ꢁC; retention time, 13 (S) and 14 (R) min. The authentic sample
29
27
(R)-12: [
a
]
ꢂ14.9 (c 1.08, MeOH)(97% ee); lit.5b (S)-form, [
a]
D
D
þ19.0 (c 0.95, MeOH).
Acknowledgements
This work was partially supported by a Grant-in-Aid for Scien-
tific Research (No. 21550051) from the Ministry of Education,
Culture, Sports, Science and Technology, Japan. We thank Collabo-
rative Research Center of Meisei university for NMR analysis, and
Mr. Noriyuki Kataoka (Meisei University) for helpful discussions.
5. (a) Shimojo, M.; Matsumoto, K.; Nogawa, M.; Nemoto, Y.; Ohta, H. Tetrahedron
Lett. 2004, 45, 6769; (b) Nogawa, M.; Shimojo, M.; Matsumoto, K.; Okudomi, M.;
Nemoto, Y.; Ohta, H. Tetrahedron 2006, 62, 7300.
References and notes
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10. Masuda, S.; Nakajima, T.; Suga, S. Bull. Chem. Soc. Jpn. 1983, 56, 1089.
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A
Textbook, 5th ed.; Springer: Berlin-Heidelberg-New York, NY, 2004; (d)
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E.; Nógrádi, E.; Kozma, D.; Egri, G.; Pálovics, E.; Kiss, V. Org. Biomol. Chem. 2006,
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