Enantioselective reduction of γ-hydroperoxy-α,β-unsaturated carbonyl compounds catalyzed by lipid-coated peroxidase in organic solvents

Article abstract of DOI:10.1016/S0957-4166(03)00436-1

The reduction of racemic γ-hydroperoxy-α,β-unsaturated carbonyl compounds in the presence of lipid-coated horseradish peroxidase as a homogenious catalyst in organic solvents such as toluene, benzene and chlororform containing a small amount of water enantioselectively afforded optical active (R)-alcohols and (S)-hydroperoxides, respectively.

Full text of DOI:10.1016/S0957-4166(03)00436-1

TETRAHEDRON:
ASYMMETRY
Pergamon
Tetrahedron: Asymmetry 14 (2003) 2339–2350
Enantioselective reduction of g-hydroperoxy-a,b-unsaturated
carbonyl compounds catalyzed by lipid-coated peroxidase in
organic solvents
Hidetaka Nagatomo, Yoh-ichi Matsushita,* Kazuhiro Sugamoto and Takanao Matsui
Faculty of Engineering, Miyazaki University, Gakuen-Kibanadai-Nishi, Miyazaki 889-2155, Japan
Received 26 March 2003; revised 29 May 2003; accepted 30 May 2003
Abstract—The reduction of racemic g-hydroperoxy-a,b-unsaturated carbonyl compounds in the presence of lipid-coated
horseradish peroxidase as a homogenious catalyst in organic solvents such as toluene, benzene and chlororform containing a small
amount of water enantioselectively afforded optical active (R)-alcohols and (S)-hydroperoxides, respectively.
©
2003 Elsevier Ltd. All rights reserved.
1
1–15
1. Introduction
cation for syntheses of several natural products.
Since racemic g-hydroperoxy-a,b-unsaturated carbonyl
compounds are easily synthesized by our method, we
next investigated separating them into optically active
g-hydroperoxy and g-hydroxy compounds. Several
methods for the resolution of alkyl hydroperoxides by
the use of chromatographic or enzymatic methods have
been reported. The peracetalization of racemic alkyl
hydroperoxides with chiral vinyl ether gave
diastereomeric peracetals, which were separated by liq-
Naturally occurring compounds having a g-hydroper-
oxy- or a g-hydroxy-a,b-unsaturated carbonyl moiety
are known, for example, 4-hydroperoxy-2-alkenal and
4
-hydroxy-2-alkenal as cytotoxic products during lipid-
1–4
peroxidation in the living body,
cyototoxic (E)-13-
hydroxy-10-oxo-11-octadecenoic acid and its lactone
5
–8
from a water extract of corn, and several macrocyclic
9
10
antibiotics such as brefeldin A and patulolide C. We
have reported novel and direct methods for conversion
of a,b,g,d-unsaturated carbonyl compounds into the
corresponding g-hydroperoxy-a,b-unsaturated ones by
cobalt(II) porphyrin-catalyzed reduction–oxygenation
with oxygen and triethylsilane (Scheme 1) and its appli-
uid chromatography followed by deprotection to afford
16,17
each enantiomer.
Although this is an excellent and
general method for the preparation of enantiomerically
pure hydroperoxides, the chiral vinyl ether used therein
is not easily available and both protection and depro-
tection steps are needed. Dussault et al. reported that
lipoxygenase-catalyzed oxygenation of linoleic acid fol-
lowed by protection of the hydroperoxy group and
oxidative cleavage of the olefin moiety gave (4S)-per-
18–20
oxy-2-alkenal enantioselectively in high yield.
How-
ever, this chemoenzymatic method is enable to afford
the corresponding (R)-enantiomer. The lipase-catalyzed
resolution of racemic hydroperoxides in organic sol-
21,22
vents was reported.
Since the hydroperoxides were
enantioselectively acylated by the lipase and then disin-
tegrated to ketone, the remaining hydroperoxides had
optical activity with an excess of the (S)-enantiomer.
Unfortunately, the corresponding (R)-hydroperoxides
were not obtained by this enzymatic method. Recently,
a-hydroperoxyalkylbenzenes and/or alkyl a-hydroper-
oxyalkanoates have been found to be enantioselectively
Scheme 1.
*
Corresponding author. E-mail: matusita@cc.miyazaki-u.ac.jp
0
957-4166/$ - see front matter © 2003 Elsevier Ltd. All rights reserved.
doi:10.1016/S0957-4166(03)00436-1

2
340
H. Nagatomo et al. / Tetrahedron: Asymmetry 14 (2003) 2339–2350
2
3–25
15
reduced with aryl methyl sulfide by chloroperoxidase,
ate or good yield (Scheme 3). The lipid-HRP was
26–30
with guaiacol by peroxidase,
and with thiophenol
prepared according to the method of Goto et al. (Fig.
1). The mixture of both buffer solution (pH 5.5–7.0)
of HRP and benzene solution of dioleyl N-D-gluconog-
lutamate (DOGG) was emulsified at 16,000 rpm and
then lyophilized, giving the lipid-HRP as light-brownish
powder. The molar ratio of DOGG to HRP was
changed between 330/1 and 2500/1.
3
1
44
by seleno-subtilisin in an aqueous medium, to give
enantiomerically enriched hydroperoxides and alcohols,
respectively, in moderate or good enantiomeric excess
as shown in Scheme 2. We expected that the enantiose-
lective reduction by peroxidase was applicable to the
racemic g-hydroperoxy-a,b-unsaturated carbonyl com-
pounds. Since the racemic g-hydroperoxy-a,b-unsatu-
rated carbonyl compounds are slightly soluble in water,
their reduction was considered to be favorable in an
organic solvent. Recently, lipid-coating techniques to
solubilize an enzyme in homogeneous organic solvents
44–63
32–43
were reported by Okahata et al.
and Goto et al.
The lipid-coated enzymes prepared from several
enzymes and amphiphilic lipids (surfactants) were
found to show high catalytic activity in organic media.
In particular, Goto et al. reported that lipid-coated
horseradish peroxidase (lipid-HRP) acted as a homoge-
neous catalyst in anhydrous benzene for the oxidation
of o-phenylenediamine using H O or t-BuOOH as an
2
2
6
0
oxidant. Herein we report an enantioselective reduc-
tion of racemic g-hydroperoxy-a,b-unsaturated car-
bonyl compounds in the presence of the lipid-HRP in
organic media containing small amounts of water.
Scheme 3.
Figure 1. Preparation of lipid-coated horseradish peroxidase
(
lipid-HRP).
2
.2. Determination of enantiomeric excesses of the
products obtained from the lipid-HRP-catalyzed reduc-
tion of racemic ethyl 4-hydroperoxy-2(E)-hexenoate,
rac-1b
The enzymatic reduction of 1.0 mmol of racemic ethyl
4
-hydroperoxy-2(E)-hexenoate
rac-1b
with
(syringol)
3,5-
(1.0
dimethoxy-4-hydroxybenzaldehyde
mmol) was first carried out in the presence of the
−
4
lipid-HRP (2×10 mmol) in 20 ml of benzene contain-
ing 1 v/v% of 0.1 M phosphate buffer (pH 6.5) at 20°C
for 3 h (Scheme 4). The solvent was evaporated in
vacuo, and the residue was chromatographed on silica
gel with hexane–EtOAc (9/1) as an eluent, to give the
alcohol (R)-2b (Y: 39%) and the hydroperoxide (S)-1b
(Y: 48%). After (R)-2b and (S)-1b were, respectively,
converted into (R)- and (S)-enantiomers of methyl
Scheme 2.
2
. Results and discussion
2
.1. Preparation of racemic g-hydroperoxy-a,b-unsatu-
rated carbonyl compounds and lipid-HRP
4
-hydroxy-2(E)-hexenoate by the procedure described
Racemic g-hydroperoxy-a,b-unsaturated carbonyl com-
pounds rac-1a–f used for substrates were prepared
from a,b,g,d-unsaturated carbonyl compounds by the
in Scheme 5 of Section 2.5, the absolute configurations
were determined by comparison of their specific rota-
tion values with that of methyl (2E,4S)-4-hydroxy-2-
II
64
Co (tdcpp)-catalyzed reduction–oxygenation in moder-
hexenoate reported in the literature. The enantiomeric

H. Nagatomo et al. / Tetrahedron: Asymmetry 14 (2003) 2339–2350
2341
Scheme 4. The lipid-HRP was prepared from a mixture of HRP
−
4
(
20 mg, 4.5×10 mmol) in 5.0 ml of 0.1 M phosphate buffer
(
pH 6.5) and DOGG (338 mg, 0.4 mmol) in 10 ml of benzene.
Figure 2. HPLC chromatograms of the products obtained from
the lipid-HRP-catalyzed reduction of rac-1b. (A) Authentic
racemic sample of the alcohol rac-2b and the hydroperoxide
rac-1b. (B) The crude products obtained from the reduction of
rac-1b for 3 h. Conditions: Shimadzu LC-9A with SPD-10A
UV detector at 254 nm; column, Daicel Chiralpak AS (0.46×25
cm); eluent, 10% 2-propanol in hexane; flow rate, 0.5 ml/min;
injected sample, 20 ml; column temp., rt.
phosphate buffer (pH 5.5–7.0) at 20°C for 3 h (Table 1).
In the absence of the lipid, the yield of the (R)-alcohol
(
R)-2b was low (entry 1). The production of (R)-2b
increased markedly with an increase in the molar ratio
of DOGG to HRP, and 900/1 ratio afforded the best
yield (entries 2–4). The molar ratio of DOGG to HRP
slightly influenced on the ee of (R)-2b (entries 1–4). The
pH of buffer affected to both the yield and the ee of
(
R)-2b (entries 3 and 5–7). The yield and the ee of (R)-2b
were found to be highest at the range of pH 6.0–6.5.
Table 1. Influences of the molar ratio of DOGG to HRP
for preparation of the lipid-HRP and pH of buffer on the
reduction of rac-1b
Scheme 5.
excesses (ee) of both the alcohol (R)-2b and the hydroper-
oxide (S)-1b were determined based on the ratio of areas
corresponding to the two enantiomers in the chro-
matogram analyzed by chiral HPLC on Daicel Chiralpak
AS (Fig. 2).
Entry
Molar ratio
pH of buffer
Alcohol (R)-2b
Yield (%) Ee (%)
DOGG/HRP
1
2
3
4
5
6
7
0/1
330/1
900/1
2500/1
900/1
900/1
900/1
6.0
6.0
6.0
6.0
5.5
6.5
7.0
10
30
39
27
14
40
30
92
91
91
90
85
94
70
2
.3. Influences of molar ratio of DOGG to HRP for
preparation of the lipid-HRP and pH of buffer on the
lipid-HRP-catalyzed reduction of rac-1b
The catalytic activity of the lipid-HRP prepared under
different conditions was evaluated on reduction of rac-1b
with syringol in benzene containing 1 v/v% of 0.1 M

2
342
H. Nagatomo et al. / Tetrahedron: Asymmetry 14 (2003) 2339–2350
The lipid-HRP-catalyzed reduction of rac-1b was fol-
lowed by the measurement of consumption of the
hydroperoxide 1b and production of the alcohol 2b by
the chiral HPLC, and the time courses of the reduction
are shown in Figure 3. The hydroperoxide 1b was
decreased in the reduction progress and its ee was
gradually increased: The (S)-enantiomer of 1b was on
the increase. On the other hand, the ee of the alcohol 2b
was unchanged at ca. 90% during the reaction course
and the (R)-enantiomer of 2b was predominantly pro-
duced. Since the reduction of (R)-enantiomer of 1b
proceeds extremely fast compared with its (S)-enan-
tiomer, the kinetic resolution of 1b takes place to
advantage on the reduction of rac-1b in benzene con-
taining of a small amount of the buffer.
2
.4. Effects of reaction conditions on the lipid-HRP-
catalyzed reduction of rac-1b
In order to optimize the reaction conditions, effects of
water content, the amount of syringol, solvent, and
temperature were examined on the lipid-HRP-catalyzed
reduction of rac-1b with syringol in solvent containing
0
2
–1.0 v/v% of 0.1 M phosphate buffer (pH 6.5). Table
summarizes the effects of the reaction conditions. No
reduction of rac-1b took place in anhydrous benzene
entry 1). The reduction was observed to proceed in the
(
presence of a small amount of the buffer and the yield
of the (R)-alcohol (R)-2b became constant at 41% in
benzene containing 0.66 v/v% of buffer or above
(
entries 2–4). The water content was not influenced on
the ee of (R)-2b. The effect of water content on the
reactivity of enzymes in non-aqueous media has been a
focus of recent research.
65–70
Although Goto et al. have
noted that the lipid-coated HRP was active in anhy-
drous benzene for the oxidation of o-phenylenediamine
60
using H O2 or t-BuOOH, we found that a small
2
amount of water was required for the enhancement of
the reactivity of the lipid-HRP on the reduction of the
hydroperoxide 1b. On the other hand, the molar ratio
of syringol to rac-1b had no influence on the yield and
the ee of (R)-2b (entries 4–6), and the use of 1.0 equiv.
of syringol is sufficient for the reduction. We next tried
to use chloroform, dichloromethane, or toluene instead
Figure 3. Time courses of the reduction of rac-1b with
syringol (1.0 equiv.) in the presence of the lipid-HRP (0.02
mol%) in benzene containing 1 v/v% of 0.1 M phosphate
buffer (pH 6.0) at 20°C: the alcohol 2b (ꢀ), the hydroperox-
ide 1b (ꢁ).
Table 2. Effects of buffer content, the amount of syringol, solvent, and temperature on the lipid-HRP-catalyzed reduction
Entry
Buffer (v/v%)
Syringol (equiv.)
Solvent
Temp. (°C)
Alcohol (R)-2b
Yield (%) Ee (%)
1
2
3
4
5
6
7
8
9
0
1.0
1.0
1.0
1.0
2.0
4.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0
Benzene
Benzene
Benzene
Benzene
Benzene
Benzene
CH Cl
20
20
20
20
20
20
20
20
20
10
15
25
30
No reaction
–
0.33
0.66
1.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0
21
41
41
41
40
27
42
48
36
43
48
46
94
94
94
94
94
92
94
95
93
94
92
92
2
2
CHCl3
Toluene
Toluene
Toluene
Toluene
Toluene
10
11
12
13

H. Nagatomo et al. / Tetrahedron: Asymmetry 14 (2003) 2339–2350
2343
of benzene on the reduction (entries 7–9): hexane,
diethyl ether, ethyl acetate, and alcohols were not used
because the lipid-HRP was insoluble in these solvents.
When using dichloromethane instead of benzene, both
the yield and the ee of (R)-2b were decreased. The
reactivity of the lipid-HRP in chloroform was similar to
that in benzene, and that in toluene was better than
that in benzene. Benzene, chloroform and toluene
showed almost the same enantioselectivity. Finally, the
effect of the reaction temperature was examined (entries
that of methyl (2E,4S)-4-hydroxy-2-hexenoate reported
64
in the literature. The reduction of racemic alkyl 4-
hydroperoxy-2-alkenoates proceeded enantioselectively,
to give the (R)-alcohols and the (S)-hydroperoxides in
good yields except for t-butyl ester rac-1d. The reduc-
tion progress for rac-1d was very slow and ca. 55% of
the hydroperoxide remained after 72 h, so that the ee of
the hydroperoxide S-1d was somewhat low (ee 82%).
The reactivity of alkyl 4-hydroxy-2-hexenoates was
decreased with the increase in bulkyness of the ester.
Adam et al. revealed that HRP catalyze efficiently the
kinetic resolution of secondary hydroperoxides in
buffer (Scheme 3) and the replacement of the alkyl side
chain by a longer alkyl or a bulky branched one
9
–13). The reaction at 10 or 15°C proceeded slower
than that at 20°C. The ee of (R)-2b was somewhat
decreased at 25°C and above. Thus, the reduction with
1
.0 equiv. of syringol in toluene containing 0.1 M
2
6–30
resulted in the decrease in the ee or the reactivity.
phosphate buffer (pH 6.5) at 20°C was found to be the
No conversion of racemic ethyl 4-hydroperoxy-2-
octenoate rac-1e is presumably due to chain lengthen-
ing. The reactivity of the lipid-HRP for the
hydroperoxides in toluene is similar to that of HRP in
aqueous solution described by Adam et al.
most suitable.
2
.5. The lipid-HRP-catalyzed reduction of racemic
g-hydroperoxy-a,b-unsaturated carbonyl compounds
To demonstrate the applicability of the lipid-HRP-cata-
lyzed reduction, several alkyl 4-hydroperoxy-2-
alkenoates were allowed to react with syringol (1.0
equiv.) in the presence of the lipid-HRP (0.02 mol%) in
toluene containing 1.0 v/v% of 0.1 M phosphate buffer
The kinetic resolution of racemic (E)-4-hydroperoxy-
hexenal rac-1f by the lipid-HRP was next attempted
(
Scheme 6). After the reduction of rac-1f was carried
out for 0.5 or 1.0 h, the alcohol (R)-2f and the
hydroperoxide (S)-1f were isolated by silica gel column
chromatography. Because the hydroperoxide rac-1f and
the alcohol rac-2f were unseparable under the chiral
HPLC conditions, the reaction time for 50% conversion
of rac-1f could not determined. The hydroperoxide
(
pH 6.5) at 20°C (Table 3). The reaction was termi-
nated at a conversion rate of about 50% which was
detected by HPLC. The alcohol and the hydroperoxide
were isolated by silica gel column chromatography, and
then converted by the procedure shown in Scheme 5
into (R)- and (S)-enantiomers of methyl 4-hydroxy-
(
S)-6 was reduced with trimethylphosphite to the corre-
sponding alcohol (S)-2f. The ee of the alcohols (R)-2f
and (S)-2f were determined by chiral capillary GC on
Ultra Alloy-CD (Fig. 4). The absolute configurations of
the products were determined compatible with those of
2(E)-hexenoate, (S)-2a and (R)-2a, respectively. Their
absolute configurations were determined by comparison
of the specific rotation values of (S)-2a and (R)-2a with
(
R)- and (S)-enantiomers of 2f derived from R-2b and
Table 3. The lipid-HRP-catalyzed reduction of racemic
S-1b according to the procedure described in Scheme 7.
Table 4 shows the results of the reduction of rac-1f.
When rac-1f was allowed to react for 0.5 h, (R)-2f and
a
alkyl (E)-4-hydroperoxy-2-alkenoates rac-1a–e
(
S)-1f were obtained, respectively, in 35% yield and ee
3% and in 60% yield and ee 73%. On the other hand,
the reduction of rac-1f for 1.0 h afforded (R)-2f (ee
2%) in 58% yield and (S)-2f (ee 97%) in 25% yield.
9
7
Thus, we could prepare the (S)-hydroperoxide (S)-1f
and both (R)- and (S)-enantiomers of the alcohol 2f in
high enantioselectivity. 4-hydroperoxy- and 4-hydroxy-
Entry Hydroperoxide Time (h)
Yield/% (ee/%)
R)-2 (S)-1
(
1
2
3
4
5
rac-1a
rac-1b
rac-1c
rac-1d
rac-1e
1.5
3.0
5.0
72
24
39 (97)
45 (97)
43 (98)
33 (95)
44 (93)
40 (>99)
40 (98)
50 (82)
b
No reaction
–
a
Yield is isolated one, and ee was determined by chiral HPLC.
Not determined.
b
Scheme 6.

2
344
H. Nagatomo et al. / Tetrahedron: Asymmetry 14 (2003) 2339–2350
Figure 4. GC chromatograms of the products obtained from
the lipid-HRP-catalyzed reduction of racemic (E)-4-hydroxy-
2-hexenal rac-1f. (A) Authentic racemic sample of rac-2f. (B)
The alcohol 2f obtained from the reduction of rac-1f for 0.5
h. Conditions: column, Ultra Alloy-CD No. 2; flow gas, He;
column temp., 90°C; detector, FID.
2
-hexenal are important for the lipid-peroxidation
3,4
products in fish, but their syntheses in optically active
forms have not been reported.
Scheme 7. The abbreviations used are as follows: DHP, 3,4-
dihydro-2H-pyran, PPTS, pyridinium p-toluenesulfonate,
DIBAL-H, diisobutyl aluminum hydride.
3. Conclusion
The lipid-HRP was found to catalyze the enantioselec-
tive reduction of g-hydroperoxy-a,b-unsaturated car-
bonyl compounds in organic solvent containing a small
amount of water, to afford the (R)-alcohol and (S)-
hydroperoxide, respectively.
Optical rotation was recorded on a Jasco DIP-100MU
(
Japan) with CHCl3 as a solvent. The enantiomeric
excesses (ee) of alkyl (E)-4-hydroperoxy-2-alkenoate
and (E)-4-hydroxy-2-alkenoate were determined by chi-
ral HPLC: Shimadzu LC-9A (Japan) with SPD-10A
UV detector at 254 nm; column, Daicel Chiralpak AS
column (0.46×25 cm) with Daicel Chiralpak AS pre-
column (0.46×5 cm) (Japan); eluent, hexane-2-propanol
4. Experimental
(
90/10); flow rate, 0.5 ml/min. The ee of (E)-4-hydroxy-
4
.1. General
2-hexenal was determined by chiral GC: Shimadzu
GC-14B (Japan) with FID detector; column, Ultra
Alloy-CD No. 2 (stationary phase, heptakis(2,3,6-tri-O-
pentyl)-b-cyclodextrin; 0.25 mm×45 m; Frontier Lab,
Japan); flow gas, He; flow rate, 1.61 ml/min; injection
temp., 300°C; detector temp., 300°C; column temp.,
90°C.
Horseradish peroxidase (HRP) (Activity; 180 units/mg)
was purchased from Wako Pure Chemicals Industries,
Ltd. (Japan) and used without further purification. All
commercial chemicals were analytical grade quality. All
purchased solvents were of high purity and were redis-
tilled before use. Column chromatography was exe-
cuted with BW-300 purchased from Fuji Silysia Co.,
Ltd. (Japan).
4.2. Preparation of lipid-coated horseradish peroxidase,
lipid-HRP
1
13
H and C NMR spectra were recorded on a AC-250P
A mixture of 5 ml of 0.1 M phosphate buffer (pH 6.5)
−
4
(
Bruker, USA) in CDCl solution with TMS (l=0.00)
solution of HRP (20 mg, 4.5×10 mmol) and 10 ml of
benzene solution of dioleyl N- -gluconoglutamate
3
as an internal standard. IR spectra were measured in
D
CHCl solution on a Hitachi IR 270-30 spectrometer
(DOGG) (338 mg, 0.41 mmol) was emulsified with
Ultra Turrax T25 homogenizer (IKA Labortechick,
German) at 16,000 rpm for 5 min to give W/O emul-
sions. The emulsions were lyophilized by use of FDU-
3
(
Japan). High resolution mass spectrometry (HRMS)
was performed on a Hitachi M-2000AM mass spec-
trometer (Japan) in electron impact mode at 70 eV.

H. Nagatomo et al. / Tetrahedron: Asymmetry 14 (2003) 2339–2350
2345
a
Table 4. The lipid-HRP-catalyzed reduction of racemic (E)-4-hydroperoxy-2-hexenal rac-1f
Entry
Time (h)
Product
(
R)-2f
(S)-2f
2
0
b
20
D
Yield/% (ee/%)
[h]
Yield /% (ee/%)
[h]
D
1
2
0.5
1.0
35 (93)
58 (72)
–52.3
–
60 (70)
25 (97)
–c
+55.0
c
a
Yield was isolated one, ee was determined by chiral GC, and [h] 2D 0 was measured in CHCl3.
Based on the yield of the intermediate 1f.
Not determined.
b
c
5
06 freeze-dryer (EYELA, Japan) for 6 h to give lipid-
4.3.3. Isopropyl (E)-4-hydroperoxy-2-hexenoate, rac-1c.
HRP (406 mg) as light-brown powder.
Yield 54%; colorless oil; R (hexane–EtOAc 4:1) 0.19;
f
−
1
wmax (CHCl )/cm 3550, 3400, 3050, 3000, 2975, 2900,
3
4
.3. Preparation of alkyl (E)-4-hydroperoxy-2-
1
1
720, 1670, 1470, 1375, 1320, 1300, 1260, 1200, 1140,
alkenoates and (E)-4-hydroperoxy-2-hexenal
110, 1040, and 990; l (250 MHz, CDCl ) 0.96 (3H, t,
H
3
J 7.6 Hz, CH ), 1.26, 1.29 (6H, s, OCHCH ), 1.53–1.76
3
3
Alkyl (2E,4E)-2,4-alkadienoate or (2E,4E)-2,4-hexa-
(
2H, m, CH ), 4.45 (1H, dtd, J 0.9, 6.5 and 6.8 Hz,
2
dienal (40 mmol) and 5,10,15,20-tetrakis (2,6-
CHOOH), 5.06 (1H, m, OCH), 6.00 (1H, d, J 16.0 Hz,
II
dichlorophenyl)porphinatocobalt(II) (Co (tdcpp)) (37.3
CHꢀCH-CO), 6.81 (1H, dd, J 6.8 and 16.0 Hz,
mg, 0.04 mmol) were dissolved in 100 ml of 2-
propanol–dichloromethane (1/1) in a 200 ml kjeldahl
flask equipped with three-way stop-cock. The atmo-
sphere in the flask was replaced with oxygen by bub-
bling for 5 min, and then an oxygen balloon was
attached to the flask through the three-way stopcock.
Triethylsilane (9.3 ml, 60 mmol) was added to the
solution at 28°C. The reaction was terminated when the
substrate was completely consumed by checking on
TLC. After removing the solvent under reduced pres-
sure, the residue was purified by silica gel column
chromatography with hexane–ethyl acetate to afford
CHꢀCH-CO), and 9.53 (1H, br s, OOH); l (63 MHz,
C
CDCl ) 9.35, 21.65, 25.11, 68.17, 85.52 (C-OOH),
3
1
23.36, 146.41, and 166.11; m/z (HRMS) 189.1280
+
(MH ; C H O requires MH, 189.1325).
9 17 4
4
.3.4. t-Butyl (E)-4-hydroperoxy-2-hexenoate, rac-1d.
Yield 80%; colorless oil; R (hexane–EtOAc 4:1) 0.19;
f
−1
wmax (CHCl )/cm 3550, 3400, 3050, 3000, 2975, 2900,
3
1
1
720, 1670, 1470, 1375, 1320, 1300, 1260, 1200, 1140,
110, 1040, and 990; l (250 MHz, CDCl ) 0.95 (3H, t,
H
3
J 7.4 Hz, CH ), 1.49 (9H, s, OCCH ), 1.55–1.72 (2H,
3
3
m, CH ), 4.41 (1H, dtd, J 0.9, 6.6, 6.9 Hz, CHOOH),
2
alkyl
(E)-4-hydroperoxy-2-alkenoate
or
(E)-4-
5
.98 (1H, dd, J 0.9, 16.0 Hz, CHꢀCH-CO), 6.79 (1H,
hydroperoxy-2-hexenal.
dd, J 6.9 and 16.0 Hz, CHꢀCH-CO) and 9.21 (1H, br
s, OOH); l (63 MHz, CDCl ) 9.44, 25.17, 27.99,
C
3
4
.3.1. Methyl (E)-4-hydroperoxy-2-hexenoate, rac-1a.
8
0.95(C-OOH), 85.68, 124.91, 145.29, and 165.82; m/z
+
10 19 4
Yield 58%; colorless oil; R (hexane–EtOAc, 4:1) 0.20;
f
(
HRMS) 203.1205 (MH ; C H O requires MH,
−
1
wmax (CHCl )/cm 3550, 3400, 3050, 3000, 2975, 2900,
3
2
03.1895).
1
1
720, 1670, 1470, 1375, 1320, 1300, 1260, 1200, 1140,
110, 1040, and 990; l (250 MHz, CDCl ) 0.95 (3H, t,
H
3
4
.3.5. Ethyl (E)-4-hydroperoxy-2-octenoate, rac-1e.
J 7.4 Hz, CH ), 1.53–1.76 (2H, m, CH ), 3.73 (3H, s,
3
2
Yield 63%; colorless oil; R (hexane–EtOAc 4:1) 0.25;
f
OCH ), 4.45 (1H, ddt, J 0.9, 6.8 and 6.9 Hz, CHOOH),
3
−1
wmax (CHCl )/cm 3550, 3400, 3030–2860, 1720, 1660,
3
6
.04 (1H, d, J 16.0 Hz, CHꢀCH-CO), 6.89 (1H, dd, J
1
475 and 1375; l (250 MHz, CDCl ) 0.90 (3H, t, J 7.2
H 3
6
.8 and 16.0 Hz, CHꢀCH-CO), and 9.11 (1H, br s,
Hz, CH ), 1.30 (3H, t, J 7.2 Hz, OCH CH ), 1.30–1.68
3
2
3
OOH); l (63 MHz, CDCl ) 9.38, 26.17, 85.59 (C-
C
3
(
6H, m, 3×CH ), 4.19 (2H, q, J 7.3 Hz, OCH ), 4.52
2
2
OOH), 122.67, 146.86, and 166.88; m/z (HRMS)
+
(1H, dt, J 6.7 and 6.8, CHOOH), 6.04 (1H, d, J 16.0
1
61.1078 (MH ; C H O requires MH, 161.1112).
7 13 4
Hz, CHꢀCH-CO), 6.90 (1H, dd, J 6.8 and 16.0 Hz,
CHꢀCH-CO), and 8.84 (1H, s, OOH); l (63 MHz,
4
.3.2. Ethyl (E)-4-hydroperoxy-2-hexenoate, rac-1b.
C
CDCl ) 13.87, 14.19, 22.56, 27.23, 31.84, 60.79, 84.64,
Yield 68%; colorless oil; R (hexane–EtOAc 4:1) 0.19;
3
f
−
1
123.14, 146.88, and 166.53; m/z (HRMS) 203.1280
wmax (CHCl )/cm 3550, 3400, 3050, 3000, 2975, 2900,
3
+
(
MH ; C H O requires MH, 203.1283).
1
1
720, 1670, 1470, 1375, 1320, 1300, 1260, 1200, 1140,
10 19
4
110, 1040, and 990; l (250 MHz, CDCl ) 0.95 (3H, t,
H
3
4
.3.6. (E)-4-Hydroperoxy-2-hexenal, rac-1f. Yield 52%;
J 7.5 Hz, CH ), 1.30 (3H, t, J 7.2 Hz, OCH CH ),
3
2
3
colorless oil; Rf (hexane–EtOAc 4:1) 0.14;
w
1
4
.56–1.70 (2H, m, CH ), 4.22 (2H, q, J 7.2 Hz, OCH ),
.45 (1H, dtd, J 0.9, 6.7 and 6.8 Hz CHOOH), 6.04
max
2
2
−
1
(CHCl )/cm 3550, 3400, 3030–2830, 1700, 1640, 1465,
3
and 1390; l (250 MHz, CDCl ) 0.99 (3H, t, J 7.5 Hz,
(
1H, d, J 15.8 Hz, CHꢀCH-CO) and 6.89 (1H, dd, J 6.8
H
3
CH ), 1.59–1.78 (2H, m, CH ), 4.60 (1H, dt, J 6.4 and
and 15.8 Hz, CHꢀCH-CO) 9.34 (1H, br s, OOH); l
3
2
C
6
.6 Hz, CHOOH), 6.31 (1H, dd, J 7.8 and 16.0 Hz,
(63 MHz, CDCl ) 9.33, 14.02, 25.13, 60.69, 85.55 (C-
3
CHꢀCH-CHO), 6.83 (1H, dd, J 6.4 and 16.0 Hz,
CHꢀCH-CHO), 8.90 (1H, br s, OOH), and 9.78 (1H, d,
OOH), 122.99, 146.60, and 175.37; m/z (HRMS)
+
1
75.0922 (MH ; C H O requires MH, 175.0970).
8 15 4

2
346
H. Nagatomo et al. / Tetrahedron: Asymmetry 14 (2003) 2339–2350
J 7.8 Hz, CHO); lC (63 MHz, CDCl ) 9.50, 25.23,
Methyl (2E,4R)-4-hydroxy-2-hexenoate, (R)-2a: color-
20
3
8
5.52, 133.09, 155.45, and 193.91; m/z (HRMS)
31.0702 (MH ; C H O requires MH, 131.0708).
less oil; [h] =−27.8 (c 2.10 in CHCl ); R (hexane–
D 3 f
−1
+
1
EtOAc, 4:1) 0.18; wmax (CHCl )/cm 3550, 3400, 3050,
3
6
11
3
3
1
000, 2975, 2900, 1720, 1670, 1470, 1375, 1320, 1300,
260, 1200, 1140, 1110, 1040, and 990; lH (250 MHz,
4
.4. The lipid-HRP-catalyzed reduction of ethyl (E)-4-
CDCl ) 0.95 (3H, t, J 7.5 Hz, CH ), 1.56–1.70 (2H, m,
CH ), 3.73 (3H, s, OCH ), 2.63 (1H, br s, OH), 4.45
hydroperoxy-2-hexenoate rac-1b in benzene
3
3
2
3
(
1H, dt, J 6.7 and 6.8 Hz, CHOH), 6.04 (1H, d, J 15.8
Hz, CHꢀCH-CO) and 6.89 (1H, dd, J 6.7 and 15.8 Hz,
CHꢀCH-CO); l (63 MHz, CDCl ) 9.29, 29.30, 51.43,
To a mixture of racemic ethyl (E)-4-hydroperoxy-2-
hexenoate rac-1b (175 mg, 1.0 mmol) and syringol (154
mg, 1.0 mmol) in benzene (20 ml) containing 1 v/v% of
C
3
7
1.96 (C-OH), 119.55, 150.52, and 167.08; m/z (HRMS)
+
7 13 3
0
.1 M phosphate buffer (pH 6.5) was added the lipid-
−4
144.0568 (MH ; C H O requires MH, 144.0897).
HRP (200 mg, 2.0×10 mmol), and the reaction mix-
ture was stirred at 20°C for 3 h. The solvent was
removed under reduced pressure and the residue was
purified by column chromatography (hexane–ethyl ace-
tate, 10/1–6/1) to give ethyl (2E,4R)-4-hydroxy-2-
hexenoate (R)-2b (62 mg, ee 90%) in 39% yield and
ethyl (2E,4S)-4-hydroperoxy-2-hexenoate (S)-1b (84
mg, ee 82%) in 48% yield. The ee of (R)-2b and (S)-1b
were determined by the chiral HPLC under the condi-
tions described above.
Methyl (2E,4S)-4-hydroperoxy-2-hexenoate, (S)-1a: col-
2
0
orless oil; [h] =−15.1 (c 3.00 in CHCl ); IR, NMR
D
3
and mass spectral data were coincided with those of
racemic one.
Methyl (2E,4S)-4-hydroxy-2-hexenoate, (S)-2a: color-
2
0
64
20
D
less oil; [h] =+26.3 (c 2.81 in CHCl ); lit. [h] =+24
D
3
(
c 3.1 in CHCl for ee 95%).
3
4
.6.2. The reduction of ethyl (E)-4-hydroperoxy-2-
4
.5. Measurement of the reaction progress on the lipid-
hexenoate rac-1b. To a mixture of rac-1b (700 mg, 4.0
mmol) and syringol (615 mg, 4.0 mmol) in toluene (80
ml) containing 1 v/v% of 0.1 M phosphate buffer (pH
HRP-catalyzed reduction of rac-1b
To a mixture of rac-1b (35 mg, 0.2 mmol) and syringol
−
4
6
.5) was added the lipid-HRP (800 mg, 8.0×10 mmol),
(
32 mg, 0.2 mmol) in benzene (4.0 ml) containing 1
and the reaction mixture was stirred at 20°C for 3 h.
The solvent was removed under reduced pressure and
the residue was purified by column chromatography
v/v% of 0.1 M phosphate buffer (pH 6.0) was added the
−
5
lipid-HRP (41 mg, 4.0×10 mmol), and the reaction
mixture was stirred at 20°C. At hourly intervals, 20 ml
of the reaction mixture was withdrawn and analyzed by
the chiral HPLC under the conditions described above:
The consumption of the hydroperoxide 1b, the produc-
tion of the alcohol 2b, and the ee of 1b and 2b were
determined at the same time.
(
4
hexane–ethyl acetate, 10/1–6/1) to give ethyl (2E,4R)-
-hydroxy-2-hexenoate, (R)-2b (317 mg, ee 97%) in 45%
yield and ethyl (2E,4S)-4-hydroperoxy-2-hexenoate,
S)-1b (252 mg, ee >99%) in 40% yield.
(
Ethyl (2E,4S)-4-hydroperoxy-2-hexenoate (S)-1b (252
mg) were dissolved in ether (5.0 ml) and subsequently
trimethyl phosphite (0.3 ml) was added to the solution.
The reaction mixture was stirred at 0°C for 1 h and
then at rt for 24 h. The solvent was removed under
reduced pressure and the residue was purified by
column chromatography (hexane–ethyl acetate, 8/1–6/
1) to give ethyl (2E,4S)-4-hydroxy-2-hexenoate (S)-2b
(222 mg, ee >99%) in 97% yield.
4
.6. The lipid-HRP-catalyzed reduction of alkyl (E)-4-
hydroperoxy-2-alkenoates
4.6.1. The reduction of methyl (E)-4-hydroperoxy-2-
hexenoate rac-1a. To a mixture of methyl (E)-4-
hydroperoxy-2-hexenoate rac-1a (206 mg, 1.25 mmol)
and syringol (193 mg, 1.25 mmol) in toluene (25 ml)
containing 1 v/v% of 0.1 M phosphate buffer (pH 6.5)
−
4
was added the lipid-HRP (250 mg, 2.5×10 mmol), and
the reaction mixture was stirred at 20°C for 1.5 h. The
solvent was removed under reduced pressure and the
residue was purified by column chromatography (hex-
ane–ethyl acetate, 10/1–6/1) to give methyl (2E,4R)-4-
hydroxy-2-hexenoate, R-2a (70 mg, ee 97%) in 39%
yield and methyl (2E,4S)-4-hydroperoxy-2-hexenoate,
Ethyl (2E,4R)-4-hydroxy-2-hexenoate, (R)-2b: colorless
2
0
oil; [h] =−24.2 (c 2.01 in CHCl ); R (hexane–EtOAc,
D
3
f
−
1
4:1) 0.17; wmax (CHCl )/cm 3630, 3500, 3030, 3000,
3
2950, 2900, 1720, 1660, 1460, 1380, 1300, 1280, 1240,
1180, 1140, 1100, and 1040; l (250 MHz, CDCl ) 0.97
(3H, t, J 7.0 Hz, CH ), 1.29 (3H, t, J 7.0 Hz,
OCH CH ), 1.52–1.75 (2H, m, CH ), 2.04 (1H, br s,
H
3
3
2
3
2
(
S)-1a (90 mg, ee 93%) in 44% yield.
OH), 4.21 (2H, q, J 7.0 Hz, OCH ), 4.22 (1H, m,
2
CHOH), 6.02 (1H, dd, J 1.1 and 15.8 Hz, CHꢀCH-CO)
Methyl (2E,4S)-4-hydroperoxy-2-hexenoate (S)-1a (90
mg) was dissolved in ether (5.0 ml) and subsequently
trimethyl phosphite (0.2 ml) was added to the solution.
The reaction mixture was stirred at 0°C for 1 h and
then at rt for 24 h. The solvent was removed under
reduced pressure and the residue was purified by
column chromatography (hexane–ethyl acetate, 8/1–6/
and 6.96 (1H, dd, J 5.3 and 15.8 Hz, CHꢀCH-CO); l
C
(63 MHz, CDCl ) 9.33, 14.02, 25.13, 60.69, 72.55 (C-
3
OH), 122.99,+ 146.60, and 175.37; m/z (HRMS)
159.0863 (MH ; C H O requires MH, 159.0065).
8
15
3
Ethyl (2E,4S)-4-hydroperoxy-2-hexenoate, (S)-1b: color-
2
0
less oil; [h] =−20.3 (c 1.00 in CHCl ); IR, NMR and
D
3
1
(
) to give methyl (2E,4S)-4-hydroxy-2-hexenoate (S)-2a
80 mg, ee 93%) in 98% yield.
mass spectral data were coincided with those of racemic
one.

H. Nagatomo et al. / Tetrahedron: Asymmetry 14 (2003) 2339–2350
2347
Ethyl (2E,4S)-4-hydroxy-2-hexenoate, (S)-2b: colorless
t-Butyl (2E,4S)-4-hydroperoxy-2-hexenoate (S)-1d (101
mg) were dissolved in ether (5.0 ml) and subsequently
trimethylphosphite (0.3 ml) was added to the solution.
The reaction mixture was stirred at 0°C for 1 h and
then at rt for 24 h. The solvent was removed under
reduced pressure and the residue was purified by
column chromatography (hexane–ethyl acetate, 8/1–6/
1) to give t-butyl (2E,4S)-4-hydroxy-2-hexenoate (S)-2d
(60 mg, ee 82%) in 64% yield.
2
0
oil; [h] =+25.0 (c 1.83 in CHCl ).
D
3
4.6.3. The reduction of isopropyl (E)-4-hydroperoxy-2-
hexenoate, rac-1c. To a mixture of isopropyl (E)-4-
hydroperoxy-2-hexenoate rac-1c (590 mg, 3.1 mmol)
and syringol (477 mg, 3.1 mmol) in toluene (50 ml)
containing 1 v/v% of 0.1 M phosphate buffer (pH 6.5)
−
4
was added lipid-HRP (600 mg, 6.0×10 mmol). The
reaction mixture was stirred at 20°C for 4 h. The
solvent was removed under reduced pressure and the
residue was purified by column chromatography (hex-
ane–ethyl acetate, 10/1–6/1) to give isopropyl (2E,4R)-
t-Butyl (2E,4R)-4-hydroxy-2-hexenoate, (R)-2d: color-
2
0
less oil; [h] =−15.0 (c 1.00 in CHCl ); R (hexane–
D
3
f
−
1
EtOAc, 4:1) 0.17; wmax (CHCl )/cm 3550, 3400, 3050,
3
4
-hydroxy-2-hexenoate (R)-2c (234 mg, ee 98%) in 43%
yield and isopropyl (2E,4S)-4-hydroperoxy-2-hexenoate
S)-1c (222 mg, ee 98%) in 40% yield.
3000, 2975, 2900, 1720, 1670, 1470, 1375, 1320, 1300,
1260, 1200, 1140, 1110, 1040, and 990; lH (250 MHz,
CDCl ) 0.97 (3H, t, J 7.4 Hz, CH ), 1.30 (9 H, s,
(
3
3
OCCH ), 1.55–1.67 (2H, m, CH ), 2.62 (1H, br s, OH),
3
2
Isopropyl (2E,4S)-4-hydroperoxy-2-hexenoate (S)-1c
222 mg) were dissolved in ether (5.0 ml) and subse-
quently trimethyl phosphite (0.3 ml) was added to the
solution. The reaction mixture was stirred at 0°C for 1
h and then at rt for 24 h. The solvent was removed
under reduced pressure and the residue was purified by
column chromatography (hexane–ethyl acetate, 8/1–6/
4.19 (1H, dt, J 5.3 and 5.4 Hz, CHOH), 5.94 (1H, dd,
J 1.5 and 15.7 Hz, CHꢀCH-CO), and 6.82 (1H, dd, J
5.3 and 15.7 Hz, CHꢀCH-CO); l (63 MHz, CDCl )
(
C
3
9.46, 28.02, 29.49, 72.22(C-OH), 80.40, 121.99, 148.86,
+
and 165.97; m/z (HRMS) 186.1356 (MH ; C H O
1
0
19
3
requires MH, 186.2122).
1
) to give isopropyl (2E,4S)-4-hydroxy-2-hexenoate
t-Butyl (2E,4S)-4-hydroperoxy-2-hexenoate, (S)-1d: col-
20
(
S)-2c (202 mg, ee 98%) in 97% yield.
orless oil; [h] =−15.7 (c 2.00 in CHCl ); IR, NMR
D 3
and mass spectral data were coincided with those of
racemic one.
Isopropyl (2E,4R)-4-hydroxy-2-hexenoate, (R)-2c: color-
2
0
less oil; [h] =−25.2 (c 1.00 in CHCl ); R (hexane–
D
3
f
−
1
EtOAc 4:1) 0.17; wmax (CHCl )/cm 3550, 3400, 3050,
t-Butyl (2E,4S)-4-hydroxy-2-hexenoate, (S)-2d: color-
3
20
3
1
000, 2975, 2900, 1720, 1670, 1470, 1375, 1320, 1300,
260, 1200, 1140, 1110, 1040, and 990; lH (250 MHz,
less oil; [h] =+12.8 (c 2.01 in CHCl ).
D 3
CDCl ) 0.97 (3H, t, J 7.4 Hz, CH ), 1.26 (6H, t, J 6.3
4.7. Conversion of alkyl (2E,4R)- and (2E,4S)-4-
hydroxy-2-hexenoate to the corresponding methyl esters
3
3
Hz, OCH(CH ) ), 1.56–1.69 (2H, m, CH ), 2.46 (1H, br
3
2
2
s, OH), 4.22 (1H, dt, J 5.7 and 5.8 Hz, CHOH), 5.06
1H, m, OCH), 6.00 (1H, dd, J 1.5 and 15.8 Hz,
(
4.7.1. Conversion of ethyl (2E,4R)- and (2E,4S)-4-
hydroxy-2-hexenoate. A mixture of ethyl (2E,4R)-4-
hydroxy-2-hexenoate R-2b (100 mg) and 2 M NaOH
(5.0 ml) was stirred at room temperature for 2 h. 2 M
HCl (8.0 ml) was added to the reaction mixture and the
mixture was extracted with diethyl ether (3×10 ml).
After the organic layer was dried over Na SO , the
CHꢀCH-CO), 6.91 (1H, dd, J 5.7 and 15.8 Hz,
CHꢀCH-CO), and; l (63 MHz, CDCl ) 9.42, 21.75,
C
3
2
9.46, 67.75, 72.23 (C-OH), 120.73, 149.69, and 166.14;
+
m/z (HRMS) 172.1103 (MH ; C H O requires MH,
9
17
3
172.1634).
2
4
Isopropyl (2E,4S)-4-hydroperoxy-2-hexenoate, (S)-1c:
solvent was removed under reduced pressure. The
residue was dissolved in diethyl ether (10 ml) and
subsequently an excess of diazomethane in diethyl ether
was added to the solution at 0°C. The solvent was
removed under reduced pressure and the residue was
purified by column chromatography (hexane–ethyl ace-
tate, 12/1) to give methyl (2E,4R)-4-hydroxy-2-
hexenoate, (R)-2a (86 mg, ee 97%) in 94% yield;
2
0
colorless oil; [h] =−19.8 (c 1.00 in CHCl ); IR, NMR
D
3
and mass spectral data were coincided with those of
racemic one.
Isopropyl (2E,4S)-4-hydroxy-2-hexenoate, (S)-2c: color-
2
0
less oil; [h] =+25.1 (c 1.30 in CHCl ).
D
3
20
4.6.4. The reduction of t-butyl (E)-4-hydroperoxy-2-
colorless oil; [h] =−27.1 (c 1.00 in CHCl ).
D 3
hexenoate, rac-1d. To a mixture of t-butyl (E)-4-
hydroperoxy-2-hexenoate rac-1d (202 mg, 1.0 mmol)
and syringol (155 mg, 1.0 mmol) in toluene (20 ml)
containing 1 v/v% of 0.1 M phosphate buffer (pH 6.5)
was added the lipid-HRP (200 mg, 2.0×10 mmol).
The reaction mixture was stirred at 20°C for 72 h. The
solvent was removed under reduced pressure and the
residue was purified by column chromatography (hex-
ane–ethyl acetate, 12/1–6/1) to give t-butyl (2E,4R)-4-
hydroxy-2-hexenoate (R)-2d (60 mg, ee 95%) in 33%
yield and t-butyl (2E,4S)-4-hydroperoxy-2-hexenoate
Ethyl (2E,4S)-4-hydroxy-2-hexenoate (S)-2b (100 mg)
was similarly converted into methyl (2E,4S)-4-hydroxy-
2-hexenoate (S)-2a (87 mg, ee >99%) in 95% yield;
−
4
20
colorless oil; [h] =+28.1 (c 1.00 in CHCl ).
D
3
4.7.2. Conversion of isopropyl (2E,4R)- and (2E,4S)-4-
hydroxy-2-hexenoate. A mixture of isopropyl (2E,4R)-
4-hydroxy-2-hexenoate (R)-2c (100 mg) and 2 M
NaOH (5.0 ml) was stirred at room temperature for 2
h. 2 M HCl (8.0 ml) was added to the reaction mixture
and the mixture was extracted with diethyl ether (3×10
(
S)-1d (101 mg, ee 82%) in 50% yield.

2
348
H. Nagatomo et al. / Tetrahedron: Asymmetry 14 (2003) 2339–2350
ml). After the organic layer was dried over Na SO , the
(2E,4S)-4-Hydroxy-2-hexenal, (S)-2f: colorless oil;
20
2
4
solvent was removed under reduced pressure. The
residue was dissolved in diethyl ether (10 ml) and
subsequently an excess of diazomethane in diethyl ether
was added to the solution at 0°C. The solvent was
removed under reduced pressure and the residue was
purified by column chromatography (hexane–ethyl ace-
tate, 12/1) to give methyl (2E,4R)-4-hydroxy-2-
hexenoate (R)-2a (79 mg, ee 98%) in 90% yield;
[h] =+55.0 (c 0.73 in CHCl ); R (hexane–EtOAc, 4:1)
D
3
f
−
1
0.12; wmax (CHCl )/cm 3620, 3500, 3050, 3010, 2970,
3
2920, 2860, 2780, 1700, 1640, 1480, 1400, 1230, 1130,
1060, 1020, and 980; l (250 MHz, CDCl ) 0.98 (3H, t,
H
3
J 7.4 Hz, CH ), 1.59–1.75 (2H, m, CH ), 2.63 (1H, br s,
3
2
OH), 4.38 (1H, ddt, J 1.5, 4.7, and 6.7 Hz, CHOH),
6.31 (1H, ddd, J 1.5, 7.9 and 15.7 Hz, CHꢀCH-CHO),
6.85 (1H, dd, J 4.7 and 15.7 Hz, CHꢀCH-CHO), and
9.57 (1H, d, J 7.9 Hz, CHO); l (63 MHz, CDCl )
2
0
colorless oil; [h] =−27.4 (c 1.00 in CHCl ).
D
3
C
3
9
.50, 29.45, 72.22 (C-OH), 130.82, 159.27, and 193.90;
+
Isopropyl (2E,4S)-4-hydroxy-2-hexenoate (S)-2c (100
mg) was similarly converted into methyl (2E,4S)-4-
hydroxy-2-hexenoate (S)-2a (77 mg, ee 98%) in 89%
m/z (HRMS) 114.0621 (MH ; C H O requires MH,
6 11 2
114.0329).
2
0
yield; colorless oil; [h] =+27.3 (c 1.00 in CHCl ).
4.8.2. Preparation of enantiomerically enriched (2E,4R)-
-hydroxy-2-hexenal (R)-2f. To a mixture of (E)-4-
D
3
4
hydroperoxy-2-hexenal rac-1f (131 mg, 1.0 mmol) and
syringol (154 mg, 1.0 mmol) in toluene (20 ml) contain-
ing 1 v/v% of 0.1 M phosphate buffer (pH 6.5) was
4
.7.3. Conversion of t-butyl (2E,4R)- and (2E,4S)-4-
hydroxy-2-hexenoate. To methanol solution (5 ml) of
t-butyl (2E,4R)-4-hydroxy-2-hexenoate (R)-2d 60 mg
was added one drop of sulfuric acid, and the reaction
mixture was stirred at 60°C for 20 h. The solvent was
removed under reduced pressure and the residue was
purified by column chromatography (hexane–ethyl ace-
tate, 12/1) to give methyl (2E,4R)-4-hydroxy-2-
hexenoate (R)-2a (53 mg, ee 95%) in 80% yield;
−
4
added the lipid-HRP (200 mg, 2.0×10 mmol), and the
reaction mixture was stirred at 20°C for 0.5 h. The
solvent was removed under reduced pressure and the
residue was purified by column chromatography (hex-
ane–ethyl acetate, 12/1–6/1) to give (2E,4R)-4-
hydroperoxy-2-hexenal, (R)-2f (40 mg, ee 93%) in 35%
yield, and (2E,4S)-4-hydroperoxy-2-hexenal (S)-1f (95
mg, ee 73%) in 60% yield.
2
0
colorless oil; [h] =−27.3 (c 1.12 in CHCl ).
D
3
t-Butyl (2E,4S)-4-hydroxy-2-hexenoate (S)-2d (60 mg)
was similarly converted into methyl (2E,4S)-4-hydroxy-
(
2E,4R)-4-Hydroxy-2-hexenal, (R)-2f: colorless oil;
20
D 3
[
h] =−52.3 (c 1.21 in CHCl ).
2
-hexenoate (S)-2a (56 mg, ee 82%) in 90% yield;
20
colorless oil; [h] =+23.0 (c 1.19 in CHCl ).
D
3
4
.9. Synthesis of (2E,4R)- and (2E,4S)-4-hydroxy-2-
hexenal from ethyl (2E,4R)- and (2E,4S)-4-hydroxy-2-
4
.8. The lipid-HRP-catalyzed reduction of (E)-4-
hexenoate
hydroperoxy-2-hexenal rac-1f
Ethyl (2E,4R)-4-hydroxy-2-hexenoate (R)-2b (270 mg,
1.7 mmol, ee 97%) and 3,4-dihydro-2H-pyran (286 mg,
3.4 mmol) were dissolved in dichloromethane (5.0 ml).
To the solution was added pyridinium p-toluene-
sulphonate (43 mg, 0.17 mmol), and the reaction mix-
ture was stirred at room temperature for 3 h. The
solvent was removed under reduced pressure and the
residue was purified by column chromatography (hex-
ane–ethyl acetate, 12/1) to give ethyl (2E,4R)-4-tetra-
hydropyranyloxy-2-hexenoate (R)-3 (380 mg, 93%).
After 5.0 ml of 1.0 M diisobutyl aluminium hydride in
hexane was added to dichloromethane solution of (R)-3
(380 mg) at −78°C, the reaction mixture was stirred for
3 h. To the reaction mixture was added 1.8 ml of water,
and the solution was stirred at room temperature for 15
min. The reaction mixture was dried over Na SO and
then the solvent was removed under reduced pressure.
The residue was purified by column chromatography
(hexane–ethyl acetate, 5/1) to give (2E,4R)-4-tetra-
hydropyranyloxy-2-hexenol (R)-4 (209 mg, 67%). A
mixture of (R)-4 (209 mg, 1.0 mmol) and manganese
dioxide (2.1 g, 10 wt. equiv.) in dichloromethane (5.0
ml) was stirred at room temperature for 2 h. The active
manganese dioxide was removed from the reaction
mixture by filtration and the solvent was removed
under reduced pressure. The residue was purified by
column chromatography (hexane–ethyl acetate, 8/1) to
give (2E,4R)-4-tetrahydropyranyloxy-2-hexenal (R)-5
4
.8.1. Preparation of enantiomerically enriched (2E,4S)-
-hydroperoxy- and (2E,4S)-4-hydroxy-2-hexenal. To a
4
mixture of (E)-4-hydroperoxy-2-hexenal rac-1f (131
mg, 1.0 mmol) and syringol (154 mg, 1.0 mmol) in
toluene (20 ml) containing 1 v/v% of 0.1 M phosphate
buffer (pH 6.5) was added the lipid-HRP (200 mg,
−
4
2
2
.0×10 mmol). The reaction mixture was stirred at
0°C for 1 h. The solvent was removed under reduced
pressure and the residue was purified by column chro-
matography (hexane–ethyl acetate, 12/1–6/1) to give
(
2E,4R)-4-hydroperoxy-2-hexenal (R)-2f (56 mg, ee
2%) in 58% yield and (2E,4S)-4-hydroperoxy-2-hexe-
nal (S)-1f (32 mg) in 25% yield.
7
(
2E,4S)-4-hydroperoxy-2-hexenal (S)-1f (32 mg) were
2
4
dissolved in ether (5.0 ml) and subsequently
trimethylphosphite (0.1 ml) was added to the solution.
The reaction mixture was stirred at 0°C for 1 h and
then at room temperature for 24 h. The solvent was
removed under reduced pressure and the residue was
purified by column chromatography (hexane–ethyl ace-
tate, 10/1–6/1) to give (2E,4S)-4-hydroxy-2-hexenal (S)-
2
f (23 mg, ee 97%) in 82% yield.
(
2E,4S)-4-Hydroperoxy-2-hexenal, (S)-1f: colorless oil;
20
D 3
[
h] =−24.4 (c 1.56 in CHCl ); IR, NMR and mass
spectral data coincided with those of racemic one.

H. Nagatomo et al. / Tetrahedron: Asymmetry 14 (2003) 2339–2350
2349
(
4
181 mg, 87%). To a THF (5.0 ml) solution of (2E,4R)-
-tetrahydropyranyloxy-2-hexenal (R)-5 (181 mg, 0.9
19. Dassault, P.; Lee, I. Q. J. Org. Chem. 1992, 57, 1952–
1954.
mmol) was added 0.5 M HCl (2.5 ml), and the reaction
mixture was stirred at room temperature for 3 h. The
reaction mixture was quenched with 1 M K CO (2.5
20. Dussault, P.; Sahli, A.; Westermeyer, T. J. Org. Chem.
1993, 58, 5469–5474.
21. Baba, N.; Mimura, M.; Hiratake, J.; Uchida, K.; Oda, J.
Agric. Biol. Chem. 1988, 52, 2685–2687.
22. Baba, N.; Takeno, K.; Iwasa, J.; Oda, J. Agric. Biol.
Chem. 1990, 54, 3349–3350.
23. Fu, H.; Kondo, H.; Ichikawa, Y.; Look, G. C.; Wong,
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2
3
ml) and then extracted with dichloromethane (3×10
ml). The organic layer was dried over Na SO and the
2
4
solvent was removed under reduced pressure. The
residue was purified by column chromatography (hex-
ane–ethyl acetate, 8/1) to give (2E,4R)-4-hydroxy-2-
hexenal (R)-2f (57 mg, ee 97%) in 48% yield; colorless
2
2
2
4. Adam, W.; Hoch, U.; Saha-M o¨ ller, C. R.; Schreier, P.
Angew. Chem., Int. Ed. Engl. 1993, 32, 1737–1739.
5. Hu, S.; Hager, L. P. J. Am. Chem. Soc. 1999, 121,
2
0
oil; [h] =−55.1 (c 1.96 in CHCl ).
D
3
Ethyl (2E,4S)-4-hydroxy-2-hexenoate (S)-2b (240 mg,
.5 mmol, ee >99%) was similarly converted into
2E,4S)-4-hydroxy-2-hexenal (S)-2f (39 mg, ee >99%)
872–973.
1
6. Adam, W.; Hoch, U.; Lazarus, M.; Saha-Moller, C. R.;
(
Schreier, P. J. Am. Chem. Soc. 1995, 117, 11898–11901.
2
0
in 39% yield from (S)-2b; colorless oil; [h] =+56.3 (c
1
D
27. H o¨ ft, E.; Hamann, H.-J.; Kunath, A.; Adam, W.; Hoch,
U.; Saha-M o¨ ller, C. R.; Schreier, P. Tetrahedron: Asym-
metry 1995, 6, 603–608.
.65 in CHCl3).
2
8. Hoch, U.; Adam, W.; Fell, R.; Saha-M o¨ ller, C. R.;
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9. Adam, W.; Lazarus, M.; Hoch, U.; Korb, M. N.; Saha-
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