Reduction of various ketones by red algae

Article abstract of DOI:10.1016/j.tetasy.2006.04.007

The reduction of acetophenone derivatives, (+)- and (-)-camphorquinones and steroidal ketones using red algae (Cyanidioschyzon merolae 10D and Cyanidium caldarium) was investigated. It was found that fluoro, chloro and bromo acetophenone derivatives 1a-i were reduced with good enantioselectivity. On the contrary, reduction of methyl and methoxy acetophenone 1j-o showed low enantioselectivity. The reduction followed Prelog's rule, giving the (S)-alcohols in all cases. Moreover, (+)- camphorquinone 5a was reduced to give (-)-3S- exo-hydroxycamphor 5d as the major product with high stereoselectivity in high yield. In addition, it was found that reduction of 5α-androstane-3,17-dione 8a gave the 3α-OH isomer (3α-OH/3β-OH = 76/24) with high stereoselectivity. Overall it was found that C. merolae and C. caldarium were able to reduce various substrates.

Full text of DOI:10.1016/j.tetasy.2006.04.007

Tetrahedron: Asymmetry 17 (2006) 1179–1185
Reduction of various ketones by red algae
a
b
a
Takamitsu Utsukihara, Osami Misumi, Nakahide Kato,
b
a,
*
Tsuneyoshi Kuroiwa and C. Akira Horiuchi
a
Department of Chemistry, Rikkyo (St. Paul’s) University, Nishi-Ikebukuro, Toshima-Ku, Tokyo 171-8501, Japan
Department of Life science, Rikkyo (St. Paul’s) University, Nishi-Ikebukuro, Toshima-Ku, Tokyo 171-8501, Japan
b
Received 22 February 2006; accepted 4 April 2006
Abstract—The reduction of acetophenone derivatives, (+)- and (ꢀ)-camphorquinones and steroidal ketones using red algae (Cyanidios-
chyzon merolae 10D and Cyanidium caldarium) was investigated. It was found that fluoro, chloro and bromo acetophenone derivatives
1
a–i were reduced with good enantioselectivity. On the contrary, reduction of methyl and methoxy acetophenone 1j–o showed low
enantioselectivity. The reduction followed Prelog’s rule, giving the (S)-alcohols in all cases. Moreover, (+)- camphorquinone 5a was
reduced to give (ꢀ)-3S- exo-hydroxycamphor 5d as the major product with high stereoselectivity in high yield. In addition, it was found
that reduction of 5a-androstane-3,17-dione 8a gave the 3a-OH isomer (3a-OH/3b-OH = 76/24) with high stereoselectivity. Overall it was
found that C. merolae and C. caldarium were able to reduce various substrates.
ꢀ
2006 Elsevier Ltd. All rights reserved.
1. Introduction
that the biotransformation of (+)-camphorquinone by
cyanobacteria gives (ꢀ)-3S-exo-hydroxycamphor with
3
Optically active compounds are important building blocks
for the synthesis of pharmaceuticals, pesticides, phero-
mones, flavors, fragrances and advanced materials such
as liquid crystals. It is known that the synthesis of chiral
alcohols can be achieved from the corresponding prochiral
ketones by asymmetric reduction. Recently, many biocata-
high stereoselectivity. Moreover, we reported the biotrans-
formation of (+)- and (ꢀ)-camphorquinones by Nicotiana
tabacum and Catharanthus roseus, and it was found that
(+)-camphorquinone was reduced with moderate stereo-
selectivity by C. roseus to provide (ꢀ)-(3S)-exo-hydroxy-
9
camphor. It is known that the plant cultured cells are
1
–6
lytic reductions of ketones have been reported.
Asym-
capable of regio- and stereoselective hydroxylation, oxida-
metric reduction by chemical methods involves the use of
expensive reagents, and heavy metals are often employed.
tion, reduction, hydrogenation, glycosylation and hydroly-
7
18
sis for various organic compounds.
Furthermore,
Acetophenone derivatives are very interesting model
compounds as foreign substrates for biotransformation,
because either enantiomer may be formed, which may be
determined easily. These compounds have been effectively
used as a building block for the asymmetric synthesis of
Synechococcus elongatus PCC 7942 (cyanobacteria) re-
duced both the endocyclic C@C of s-trans enones and the
exocyclic C@C of s-cis enones with high enantioselectivity
1
9,20
to afford optically active (S)-a-substituted ketones.
8
drugs. Reduction of the carbonyl group is performed
Here, we report a method for the asymmetric reduction
with photosynthetic microbes (Cyanidioschyzon merolae
(C. merolae) and Cyanidium caldarium (C. caldarium)).
9
–11
using various biocatalysts such as cultured cells
fresh-cut vegetables.
and
1
2
Recently, we have reported that the biotransformation of a
synthetic substance into a more useful substance by plant
cultured cells is an important reaction in synthetic chemis-
2. Results and discussion
1
3–17
try.
During the course of our studies, it was revealed
2.1. Reduction of acetophenone derivatives
Recently, a report by Kuroiwa et al. on unicellular red
*
Corresponding author. Fax: +81 3 3985 2397; e-mail addresses:
horiuchi@rikkyo.ac.jp; cahoriuchi@nifty.com
algae, C. merolae, provided information on the basic and
essential genes. The cells of C. merolae offer unique
2
1
0957-4166/$ - see front matter ꢀ 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetasy.2006.04.007

1180
T. Utsukihara et al. / Tetrahedron: Asymmetry 17 (2006) 1179–1185
advantages for the studies of mitochondrial and plastid
acetophenones 1a–i was high due to the ꢀI effect, except at
the meta-position. The enantioselectivity of the reduction
depends on the position of the substituent. The minor role
of the ꢀI effect was indicated by the reduction of methyl
acetophenones 1m–o. It was found that the electronic effect
of the substituents on the aromatic ring has a defined role
in the enantioselectivity of the reaction.
2
2,23
divisions
because they do not have a rigid cell wall
and contain just one nucleus, one mitochondrion and one
plastid, the divisions of which can be highly synchronized
2
4
by light/dark cycles. On the other hand, unicellular red
algae, C. caldarium, is considered to be one of the most
2
5
primitive algae. This algae has a simple cellular structure,
with each cell having one plastid and one nucleus plus
other organelles, such as Golgi bodies, endoplasmic reticu-
The reduction of C. merolae 10D and C. caldarium of sev-
eral aromatic ketones having alkyl chains of different
length shown in Table 2. It was found that compounds
3a and 3b were reduced after 3 days incubation in good
enantioselectivity, while longer alkanones, 3c–f, could not
be reduced. This result is apparently due to the size of
the alkyl side chains and their lower solubility in the reac-
tion medium.
2
6
lum and microbodies. C. merolae and C. caldarium are
extremely characteristic algae inhabiting sulfate-rich hot
springs.
A series of aromatic ketones were targeted for biotransfor-
mation using C. merolae and C. caldarium in Allen’s med-
ium. ortho-, meta- and para-substituted fluoro, chloro,
bromo, methyl and methoxy acetophenones 1a–o were re-
duced to the corresponding (S)-alcohols in all cases (Table
2.2. Effect of XAD on reduction of acetophenone derivatives
2
7
1
). The alcohol products had the (S)-configuration, which
TM
is consistent with Prelog’s rule. Reduction of fluoro, chloro
and bromo acetophenones provided good enantioselectiv-
ity (67–99%). The yields increased with increasing size of
the halogens. In contrast, reduction of methyl and methoxy
acetophenones was observed in low yields (7–38%) and
enantioselectivity (2–41%). In the case of methoxy groups,
there was a substantial drop in the enantioselectivity of the
product (Table 1, entries 10–12). In these cases, it seems
that the ꢀM effect of the substituents was superior to their
Recently, the hydrophobic polymer, Amberlite XAD, has
been used to reduce the substrate in microbial reduction,
28
and increased the chemical yield of the reduction. Amber-
TM
lite XAD, a hydrophobic polymer such as polystyrene or
polyacryl ester, is a commercially available adsorbent, and
has a large surface area. The hydrophobic polymer,
TM
Amberlite XAD, can control the stereoselectivity of the
reduction of ketones by the fungus, Geotrichum candi-
2
9
dam. Previously, Nakamura et al. reported that the use
ꢀ
I effect. The enantioselectivity of the halogen-substituted
of XAD-7 increases the enantioselectivity in the reduction
Table 1. Reduction of aryl ketones by red algae
Entry
Substrate
R
Product
C. merolae 10D
C. caldarium
Yield (%)
ee (%)
Config.
Yield (%)
ee (%)
Config.
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
1a
1b
1c
1d
1e
1f
o-F
m-F
p-F
2a
2b
2c
2d
2e
2f
40
54
50
70
78
71
87
84
92
30
32
13
16
7
91
71
74
98
73
94
99
77
98
2
6
6
37
39
16
S
S
S
S
S
S
S
S
S
S
S
S
S
S
S
34
58
50
70
68
79
80
90
85
38
29
9
92
75
67
98
75
96
99
75
98
4
S
S
S
S
S
S
S
S
S
S
S
S
S
S
S
o-Cl
m-Cl
p-Cl
o-Br
m-Br
p-Br
o-OCH
m-OCH
p-OCH
o-CH
m-CH
p-CH
1g
1h
1i
2g
2h
2i
1
1
1
1
1
1
1j
3
2j
1k
1l
3
2k
2l
7
3
12
27
41
16
1m
1n
1o
3
2m
2n
2o
28
9
31
3
3
36
Reaction conditions: Substrate (20 mg), red algae (dry weight 1 g/L) and Allen’s medium (100 mL) were employed for 3 days (42 ꢁC, pH 2.5, and 2000 lx).
Table 2. Reduction of aryl ketones by red algae
Entry
Substrate
R
Product
C. merolae 10D
C. caldarium
Yield (%)
ee (%)
Config.
Yield (%)
ee (%)
Config.
1
2
3
4
5
6
3a
3b
3c
3d
3e
3f
CH
CH
CH
CH
CH
CH
3
3
3
3
3
3
4a
4b
—
—
—
—
28
53
0
0
0
80
78
—
—
—
—
S
24
46
0
0
0
82
74
—
—
—
—
S
CH
2
S
S
(CH
(CH
(CH
(CH
2
2
2
2
)
3
)
4
)
5
)
6
—
—
—
—
—
—
—
—
0
0
Reaction conditions: Substrate (20 mg), red algae (dry weight 1 g/L) and Allen’s medium (100 mL) were employed for 3 days (42 ꢁC, pH 2.5, and 2000 lx).

T. Utsukihara et al. / Tetrahedron: Asymmetry 17 (2006) 1179–1185
1181
of simple aliphatic and aromatic ketones.30 Thus, we exam-
ined the efficiency of using XAD-7 in terms of the
enantioselectivity.
addition, the reduction of octanophenone 3f is the first re-
port of the use of biocatalysts for this substrate (Scheme 2).
R
O
Cyanidioschyzon merolae
or
R
OH
ortho-, meta- and para-substituted fluoro, chloro, bromo,
methyl and methoxy acetophenones 1a–o were reduced to
the corresponding (S)-alcohols in all cases (Scheme 1).
The yield and stereoselectivity did not change using
XAD-7. Moreover, we investigated the effect of the size
of the alkyl group for 1-phenylalkanones 3a–f by C. mero-
lae and C. caldarium. These results are shown in Table 3.
From these results, it was found that increasing the length
of the alkyl chain (Table 3, entries 3–6) decreased the
enantioselectivity for the corresponding (S)-alcohols. In
Cyanidium caldarium
3a-g
4
a-g
Substrate
R
3a
CH3
3b
CH CH2
3
3
c
CH
CH
3
(CH
(CH
2
)
3
3d
3
2
)
4
3
e
CH (CH )
3 2 5
O
Cyanidioschyzon merolae
or
OH
3f
CH (CH )
3 2 6
Cyanidium caldarium
Scheme 2.
R
R
The reduction of asymmetrical ketones by cyanobacteria
S. elongatus PCC 7942) as biocatalysts under light condi-
tions gave the corresponding (S)-alcohols with excellent
(
1a-m
2
a-m
3
1–33
Substrate
R
enantioselectivities.
Such results suggest that the
reduction of aliphatic ketones by red algae will be very
useful for the practical preparation of (S)-alcohols with,
for example, S. elongatus PCC 7942.
1
a
o-F
m-F
p-F
1b
1
c
2.3. Reduction of (+)-and (ꢀ)-camphorquinones
1d
o-Cl
m-Cl
p-Cl
1
e
Biotransformation of (+)-camphorquinone 5a afforded a
mixture of diastereoisomers of three a-keto alcohols: (+)-
1
f
(
2R)-exo-hydroxyepicamphor 5b, (ꢀ)-(3S)-exo-hydroxy-
1g
o-Br
m-Br
p-Br
camphor 5d and (+)-(3R)-endo-hydroxycamphor 5e. How-
ever, (ꢀ)-(2S)-endo-hydroxyepicamphor 5c could not be
obtained from (+)-camphorquinone 5a using C. merolae
and C. caldarium (Scheme 3).
1h
1
i
1
j
o-OCH3
m-OCH3
(
ꢀ)-Camphorquinone 6a was transformed to give the four
1k
corresponding isomers: (ꢀ)-(2S)-exo-hydroxyepicamphor
6b, (+)-(2R)-endo-hydroxyepicamphor 6c, (+)-(3R)-exo-
hydroxycamphor 6d and (ꢀ)-(3S)-endo-hydroxycamphor
1
l
p-OCH3
o-CH3
1m
6
e (Scheme 4).
1
n
m-CH3
p-CH3
From these results, it was found that only hydroxy ketones
were formed and diols were not obtained. (+)-Camphor-
quinone 5a was reduced stereoselectively by C. merolae
1
o
Scheme 1.
Table 3. Reduction of aryl ketones by red algae using XAD
Entry
Substrate
R
Product
C. merolae 10D
C. caldarium
Yield (%)
ee (%)
Config.
Yield (%)
ee (%)
Config.
1
2
3
4
5
6
3a
3b
3c
3d
3e
3f
CH
CH
CH
CH
CH
CH
3
3
3
3
3
3
4a
4b
4c
4d
4e
4f
22
48
69
69
71
44
79
82
83
54
47
42
S
S
S
S
S
S
36
50
74
66
45
55
87
75
80
59
54
37
S
S
S
S
S
S
CH
2
(CH
(CH
(CH
(CH
2
2
2
2
)
3
)
4
)
5
)
6
Reaction conditions: Substrate (20 mg), red algae (dry weight 1 g/L), XAD-7 (250 mg) and Allen’s medium (100 mL) were employed at 42 ꢁC (5 days,
pH 2.5 and 2000 lx).

1182
T. Utsukihara et al. / Tetrahedron: Asymmetry 17 (2006) 1179–1185
OH
O
H
O
O
H
OH
H
O
OH
O
O
H
OH
5
a
5
b
5
c
5
d
5e
Scheme 3. Reduction of (+)-camphorquinone 5a by red algae.
1
0D and C. caldarium to give (ꢀ)-(3S)-exo-hydroxycam-
selectivity in the reduction of steroidal ketones, andro-
stane-3,17-dione 7a and D -androstene-3,17-dione 8a were
4
phor 5d in 83% and 84% yield, respectively. These results
indicate that the reduction activities are higher than those
of plant cultured-cells N. tabacum and C. roseus.
investigated as model compounds. These results are shown
in Table 5.
9
In the case of (ꢀ)-camphorquinone 6a using C. merolae
O
1
0D and C. caldarium, (+)-(3R)-exo-hydroxycamphor 6d
was obtained in 64% yield, respectively (Table 4).
a
Table 4. Reduction of camphorquinone 5a and 6a by red algae
O
b
c
Run Substrate Red algae
Yield
%)
Product ratio
(%)
4
-androstene-3,17-dione 8a
(
O
O
1
2
3
4
5a
5a
6a
6a
C. merolae 92
C. caldarium 91
C. merolae 93
C. caldarium 92
5b (16) 5c (ꢀ) 5d (83) 5e (1)
5b (15) 5c (ꢀ) 5d (84) 5e (1)
6b (19) 6c (12) 6d (64) 6e (5)
6b (18) 6c (13) 6d (64) 6e (5)
a
Reaction conditions: Substrate (20 mg) and red algae (70 mL) were
cultivated with shaking at 25 ꢁC for 2 days.
Isolated yields.
O
HO
3α-hydroxy-5α-androstan-17-one 9a
5
α-androstane-3,17-dione 7a
b
c
1
Relative intensities by H NMR signals.
Scheme 5. Reduction of steroidal ketones 7a and 8a by red algae.
The growth rate of red algae is faster than that of other
typical microbes. Some of the problems using plant cul-
tured cells as biocatalysts are that they are usually grown
very slowly and require a large amount of the catalyst.
Reduction of (+)- and (ꢀ)-camphorquinones 5a and 6a
by N. tabacum and C. roseus needs 15 g plant cultured cells
to biotransform 20 mg of the substrates; however, only
a
Table 5. Reduction of steroidal ketones 7a and 8a by red algae
b
b
Run
Substrate Red algae
Product ratio 3a-OH/ Conv.
c
(
%)
3b-OH
1
2
3
4
7a
7a
8a
8a
C. merolae
C. caldarium
C. merolae
9a (80) 75/25
9a (78) 76/24
7a (27) 9a (45) 77/23
80
78
72
66
C. caldarium 7a (24) 9a (42) 76/24
1
00 mg (dry weight) of red algae is required to biotrans-
a
form stereoselectively.
Reaction conditions: Substrate (20 mg) and red algae (70 mL) were
cultivated with shaking at 42 ꢁC for 7 days.
b
c
2
.4. Reduction of steroidal ketones
Determined by GC–MS peak ratio.
Relative intensities by H NMR signals.
1
4
Androstane-3,17-dione 7a and D -androstene-3,17-dione
a are commercially available steroids which have been
used for partial synthesis of many steroidal hormones,
including testosterone. Steroidal ketones are known as
important substrates for stereoselectivity and regioselectiv-
ity. It is known that the biotransformation for steroidal
8
The position of the hydroxyl group was established by
1
3
chemical shifts in the C NMR spectra. The stereochemis-
try of the secondary alcohols is due to the multiplicity of
1
the relevant H NMR signal (18-H and 19-H methyl
signals). In addition, the 3b-H resonance showed a charac-
teristic downfield shift (Dd 0.4 ppm) compared to 3a-H for
3b-OH.
3
5
,17-dione gives the C -b hydroxy derivatives (Scheme
17
3
4,35
).
In order to examine the stereoselectivity and regio-
O
O
O
H
OH
H
OH
H
O
O
OH
H
HO
O
6
a
6
b
6
c
6d
6
e
Scheme 4. Reduction of (ꢀ)-camphorquinone 6a by red algae.

T. Utsukihara et al. / Tetrahedron: Asymmetry 17 (2006) 1179–1185
1183
It was found that the reduction of steroidal ketones 7a and
a occurs at the C -position preferentially, rather than the
was treated with a shaker (120 rpm) at 42 ꢁC in the light
(2000 lx). At the end of the reaction, XAD-7 was filtered
from the medium. The filtrate was extracted with ether
(2 · 50 mL), washed with water (2 · 20 mL) and dried.
XAD-7 was washed with ether (50 mL) and the extract
was concentrated into an oily residue. All the products
8
3
3
6
C -ketone (Table 5). Moreover, the more unstable C -a
1
7
3
hydroxy compounds are obtained preferentially, rather
than less hindered C -b hydroxy derivatives.
3
1
were determined by IR, H NMR and gas chromato-
3. Conclusion
graphic analyses.
27
D
In summary, the above studies have demonstrated the
biotransformation of a broad assortment of structurally
different acetophenone derivatives, (+)- and (ꢀ)-camphor-
quinones and steroidal ketones. In the case of acetophen-
one derivatives 3a–f, increasing the size of the flanking
groups by incorporating hydrophobic groups (increasing
the alkyl chain length) led to low enantioselectivity. The
presence of electron-withdrawing substituents in the phenyl
ring 1a–i improved the enantioselectivity of the reduction
product. Moreover, in the reduction of steroidal ketones
Compound 2c.39 Colourless oil; ½aꢁ ¼ ꢀ40:0 (c 0.9,
ꢀ
1
1
CHCl ); IR 3353, 1076 and 831 cm
;
H NMR: d
3
(ppm) 1.48 (d, J = 6.0 Hz, 3H), 1.83 (br s, 1H), 4.88 (q,
J = 6.0 Hz, 1H) and 7.00–7.34 (m, 4H); MS m/z: 140
(M , 18.9), 125 (100), 122 (11.9), 101 (6.97), 96 (24.1), 83
(2.60), 75 (19.0), 63 (4.39), 57 (7.95) and 51 (21.3).
+
Compound 2f.37,39 Colourless oil; ½aꢁ ¼ ꢀ45:0 (c 0.9,
27
D
ꢀ1
1
CHCl ); IR 3360, 1050, 898 and 828 cm
; H NMR:
3
d (ppm) 1.47 (d, J = 6.0 Hz, 3H), 1.88 (br s, 1H), 4.86 (q,
J = 6.0 Hz, 1H) and 7.22–7.30 (m, 4H); MS m/z: 156
(M , 13.2), 141 (60.3), 138 (7.39), 121 (10.2), 113 (27.1),
7
a and 8a, the C -a hydroxy compounds are obtained pref-
3
+
erentially, rather than C -b hydroxy derivatives. We have
3
established a convenient and environmentally benign stereo-
103 (9.88), 87 (1.42), 77 (100), 63 (6.40) and 51 (41.8).
selective reduction system employing red algae C. merolae
Compound 2i.39,41 Yellow oil; ½aꢁ ¼ ꢀ37:3 (c 1.1, CHCl );
27
1
0D and C. caldarium.
D
3
ꢀ
1
1
IR 3326, 1078, 896 and 825 cm
;
H NMR: d (ppm)
1.47 (d, J = 6.0 Hz, 3H), 1.87 (br s, 1H), 4.86 (q,
4. Experimental
J = 6.0 Hz, 1H) and 7.23–7.45 (m, 4H); MS m/z: 200
+
(M , 10.7), 184 (7.46), 182 (6.05), 157 (20.7), 121 (17.1),
The racemic alcohols were obtained by the reduction of the
corresponding ketones with sodium borohydride or
authentic samples were purchased. XAD-7 was washed
with 1 M hydrochloric acid, 1 M sodium hydroxide, water,
methanol and ether. C. caldarium was obtained from the
Institute of Applied Microbiology Culture Collection
107 (0.58), 77 (100), 63 (7.93) and 51 (53.6).
Compound 2l.39,40 Colourless oil; ½aꢁ ¼ ꢀ4:2 (c 0.9,
27
D
ꢀ1
1
CHCl ); IR 3378, 1079 and 831 cm
;
H NMR: d
3
(ppm) 1.47 (d, J = 6.0 Hz, 3H), 1.87 (br s, 1H), 3.80 (s,
3H), 4.85 (q, J = 6.0 Hz, 1H) and 6.9–7.3 (m, 4H); MS
+
(
IAMCC). Gas chromatographic analysis was performed
m/z: 152 (M , 15.6), 134 (100), 119 (64.2), 103 (5.69), 91
using chiral GC-column (Chirasil-DEX CB; 25 m)
equipped on Shimadzu GC-17A. Gas chromatography/
mass spectrometry analysis was Shimadzu GCMS-
QP5050 (EI-MS 70 eV) using DB1 (0.25 mm · 30 m,
(77.5), 77 (38.1), 65 (63.6) and 51 (36.7).
Compound 2o.37 Colourless oil; ½aꢁ ¼ ꢀ13:0 (c 0.4,
27
D
ꢀ
1 1
CHCl ); IR 3367, 1078, 896 and 811 cm ; H NMR: d
3
0
.25 lm) capillary column GC; GC: GC-17A. The IR spec-
(ppm) 1.48 (d, J = 6.0 Hz, 3H), 1.80 (br s, 1H), 2.34 (s,
tra were measured on a Jasco FT-IR 230. The NMR spec-
tra were measured on a JEOL GSX 400 spectrometer.
CDCl3 with tetramethylsilane as the internal standard
was used.
3H), 4.86 (q, J = 6.0 Hz, 1H) and 6.84–7.30 (m, 4H); MS
m/z: 136 (M , 32.3), 121 (100), 118 (35.3), 117 (43.9), 115
(19.8), 103 (5.47), 93 (93.6), 91 (95.7), 79 (3.25), 77 (59.3),
65 (39.5), 63 (22.6) and 51 (34.4).
+
Compound 4a.37–39 Colourless oil; ½aꢁ ¼ ꢀ43:7 (c 0.9,
27
D
4
.1. Preparation of microbial culture
Allen’s medium was prepared by mixing (NH ) SO
4
ꢀ
1 1
CHCl ); IR 3350, 1060, 749 and 732 cm ; H NMR: d
(ppm) 1.48 (d, J = 6.0 Hz, 3H), 1.90 (br s, 1H), 4.88 (q,
3
4
2
(
(
2.64 g), KH PO4 (0.544 g), MgSO4 (0.492 g), CaCl2
J = 6.0 Hz, 3H) and 7.2–7.4 (m, 5H); EIMS m/z: 122
(M , 27.0), 107 (77.3), 104 (7.31), 79 (100), 77 (61.0) and
2
+
0.148 g) and P4 metal (4 mL) in distilled H O (1 L). The
2
medium was adjusted to pH 2.2–2.5 and sterilized.
51 (47.5).
Compound 4b.37–41 Colourless oil; ½aꢁ ¼ ꢀ38:1 (c 1.0,
27
D
P4 metal solution was FeCl (196 mg), MnCl (36 mg),
3
2
ꢀ
1 1
ZnSO (22.2 mg), CoCl (4 mg), Na MoO (2.5 mg) and
CHCl ); IR 3355, 1093, 750 and 692 cm ; H NMR: d
4
2
2
4
3
Na EDTA (1000 mg) dissolved in distilled H O (1 L).
(ppm) 0.90 (t, J = 6.2 Hz, 3H), 1.7–1.9 (m, 2H), 1.95 (br
2
2
s, 1H), 4.6 (t, J = 6.0 Hz, 1H) and 7.2–7.4 (m, 5H); EIMS
+
4
.2. General procedure of biotransformation conditions
m/z: 136 (M , 11.5), 118 (4.70), 117 (7.53), 107 (100), 91
(6.55), 79 (97.9), 77 (54.9) and 51 (47.5).
Acetophenone derivatives (20 mg) with XAD-7 (250 mg)
were added to the culture of C. merolae and C. caldarium
Compound 4c.37 Colourless oil; ½aꢁ ¼ ꢀ39:3 (c 0.5,
27
D
ꢀ
1 1
(
100 mL, 1 g/L as dry weight) in Allen’s medium and
CHCl ); IR 3361, 1110, 754 and 694 cm ; H NMR: d
3
shaken for the periods shown in the tables. The mixture
(ppm) 0.90 (t, J = 7.0 Hz, 3H), 1.2–1.8 (m, 6H), 2.10 (br

1184
T. Utsukihara et al. / Tetrahedron: Asymmetry 17 (2006) 1179–1185
s, 1H), 4.60 (t, J = 6.0 Hz, 1H) and 7.2–7.4 (m, 5H); EIMS
m/z: 164 (M , 0.59), 146 (2.20), 128 (0.50), 117 (8.56), 115
10. Hamada, H.; Tomi, R.; Asada, Y.; Furuya, T. Tetrahedron
Lett. 2002, 4087–4089.
+
1
1
1
1
1. Shimoda, K.; Kubota, N.; Hamada, H.; Hamada, H.
(
4.73), 107 (100), 104 (3.79), 91 (5.87), 79 (65.4), 77 (32.5)
Tetrahedron Lett. 2006, 1541–1544.
2. Yadav, J. S.; Nanda, S.; Reddy, P. T.; Rao, A. B. J. Org.
Chem. 2002, 67, 3900–3903.
3. Matsuura, Y.; Chai, W.; Endoh, E.; Suhara, J.; Hamada, T.;
Horiuchi, C. A. Phytochemistry 2002, 61, 669–673.
4. Sakamaki, H.; Kitanaka, S.; Chai, W.; Hayashida, Y.;
Takagi, Y.; Horiuchi, C. A. J. Nat. Prod. 2001, 64, 630–631.
and 51 (15.5).
Compound 4d.4 Colourless oil; ½aꢁD ¼ ꢀ15:6 (c 1.2,
27
ꢀ
1 1
CHCl ); IR 3349, 1060, 755 and 694 cm ; H NMR: d
3
(
ppm) 0.88 (t, J = 7.0 Hz, 3H), 1.4–1.8 (m, 8H), 1.9 (br s,
1
H), 4.62 (t, J = 6.0 Hz, 1H) and 7.2–7.4 (m, 5H); EIMS
+
m/z: 178 (M , 2.84), 160 (1.91), 131 (0.68), 117 (9.47),
15 (4.77), 107 (100), 104 (6.74), 91 (6.64), 79 (53.6), 77
27.2) and 51 (11.1).
15. Chai, W.; Sakamaki, H.; Kitanaka, S.; Horiuchi, C. A. Bull.
Chem. Soc. Jpn. 2003, 76, 177–182.
1
(
1
6. Sakamaki, H.; Ito, K.; Chai, W.; Hayashida, Y.; Kitanaka,
S.; Horiuchi, C. A. J. Mol. Catal. B: Enzym. 2004, 27, 177–
181.
Compound 4e.4 Colourless oil; ½aꢁD ¼ ꢀ17:6 (c 1.2,
27
ꢀ
1
1
17. Sakamaki, H.; Ito, K.; Taniai, T.; Kitanaka, S.; Takagi, Y.;
CHCl ); IR 3343, 1035, 755 and 694 cm ; H NMR: d
3
Chai, W.; Horiuchi, C. A. J. Mol. Catal. B: Enzym. 2005, 32,
(
ppm) 0.90 (t, J = 6.2 Hz, 3H), 1.2–1.8 (m, 10H), 2.0 (br
1
03–106.
s, 1H), 4.62 (t, J = 6.0 Hz, 1H) and 7.2–7.4 (m, 5H); EIMS
1
8. Ishihara, K.; Hamada, H.; Hirata, T.; Nakajima, N. J. Mol.
Catal. B: Enzym. 2003, 23, 145–170.
+
m/z: 192 (M , 1.71), 174 (1.99), 145 (0.16), 131 (0.73), 117
(
7
10.2), 115 (4.97), 107 (100), 104 (10.0), 91 (7.57), 79 (46.7),
7 (23.1) and 51 (8.58).
19. Shimoda, K.; Kubota, N.; Hamada, H.; Kaji, M.; Hirata, T.
Tetrahedron: Asymmetry 2004, 15, 1677–1679.
2
2
0. Shimoda, K.; Kubota, N.; Hamada, H.; Yamane, S.; Hirata,
T. Bull. Chem. Soc. Jpn. 2004, 77, 2269–2272.
Compound 4f.42 Colourless oil; ½aꢁD ¼ ꢀ18:3 (c 0.8,
27
ꢀ
1 1
CHCl ); IR 3347, 1060, 757 and 698 cm ; H NMR: d
ppm) 0.87 (t, J = 6.2 Hz, 3H), 1.2–1.8 (m, 10H), 2.0 (br
s, 1H), 4.62 (t, J = 6.0 Hz, 1H) and 7.2–7.4 (m, 5H); EIMS
m/z: 206 (M , 1.39), 188 (2.06), 131 (0.85), 117 (11.9), 115
1. Matsuzaki, M.; Misumi, O.; Shin-I, T.; Maruyama, S.;
Takahara, M.; Miyagishima, S.; Mori, T.; Nishida, K.;
Yagisawa, F.; Nishida, K.; Yoshida, Y.; Mishimura, Y.;
Nakao, S.; Kobayashi, T.; Momoyama, Y.; Higashiyama, T.;
Minoda, A.; Sano, M.; Nomoto, H.; Oishi, K.; Hayayashi,
H.; Ohta, F.; Mishizawa, S.; Haga, S.; Miura, S.; Morishita,
T.; Kabeya, Y.; Terasawa, K.; Suzuki, Y.; Ishii, Y.; Asakawa,
S.; Takano, H.; Ohta, N.; Kuroiwa, H.; Tanaka, K.; Shimizu,
N.; Sugano, S.; Sato, N.; Nozaki, H.; Ogasawara, N.;
Kohara, Y.; Kuroiwa, T. Nature 2004, 428, 653–657.
2. Kuroiwa, T. Bioessays 1998, 20, 2344–2354.
3
(
+
(
5.48), 107 (100), 104 (14.4), 91 (8.76), 79 (40.6), 77 (20.2)
and 51 (6.78).
Compound 5d.43 White crystal; ½aꢁD ¼ ꢀ78:4 (c 1.5,
27
ꢀ1
1
CHCl ); IR 3448, 1751 and 1123 cm
;
H NMR: d
3
2
2
(
(
3
ppm) 0.94 (s, 3H), 0.95 (s, 3H), 1.00 (s, 3H), 1.20–2.00
m, 4H), 2.10 (d, J = 5.0 Hz, 1H), 2.59 (br s, 1H) and
.75 (s, 1H); EIMS m/z: 168 (M , 13.2), 153 (1.21), 140
4.02), 125 (22.8), 107 (6.47), 100 (11.6), 83 (86.9), 69
39.8), 55 (100) and 53 (15.1).
3. McFadden, G. I.; Ralph, S. A. Proc. Natl. Acad. Sci. U.S.A.
2003, 100, 3357–3359.
+
2
4. Terui, S.; Suzuki, K.; Talahashi, H.; Itoh, R.; Kuroiwa, T. J.
Phycol. 1995, 31, 958–961.
(
(
2
2
5. Geitler, L. Arch. Hydrobiol. Suppl. 1933, 12, 622–634.
6. Kuroiwa, T.; Kawazu, T.; Takahashi, H.; Suzuki, K.; Ohta,
N.; Kuroiwa, H. Cytologia 1994, 59, 149–158.
Acknowledgements
27
D
27
D
2
7. Product 2a: ½aꢁ ¼ ꢀ43:0 (c 0.87, CHCl
3
); 2b: ½aꢁ ¼ ꢀ25:1
2
D
7
(
½
c 1.1, CHCl
3
); 2d: ½aꢁ ¼ ꢀ44:7 (c 0.85, CHCl
3
); 2e:
2
7
27
D
This work was partially supported by Frontier Project
Environmental Changes and Life Adaptation Strategies’
and a Grant-in-Aid for Scientific Research (No. 15550140).
aꢁ ¼ ꢀ39:5 (c 1.6, CHCl ); 2g: ½aꢁ ¼ ꢀ42:0 (c 1.7, CHCl );
D
3
3
2
7
27
‘
2h: ½aꢁ ¼ ꢀ28:0 (c 1.7, CHCl ); 2j: ½aꢁ ¼ ꢀ1:7 (c 0.88,
D
3
D
2
7
27
D
3
).
CHCl ); 2k: ½aꢁ ¼ ꢀ2:7 (c 0.88, CHCl ); 2m: ½aꢁ ¼ ꢀ18:2
3
3
D
2
D
7
(
c 0.62, CHCl
3
); 2n: ½aꢁ ¼ ꢀ25:8 (c 0.62, CHCl
2
8. Anderson, B. A.; Hansen, M. M.; Harkness, A. R.; Henry, C.
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2
D
7
36. Product 9a: ½aꢁ ¼ þ80:3 (c 0.63, CHCl
3
).
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