Biotransformation of (+)- and (-)-camphorquinones by plant cultured cells

Article abstract of DOI:10.1016/S0031-9422(01)00133-9

Biotransformation of (+)- and (-)-camphorquinones with suspension plant cultured cells of Nicotiana tabacum and Catharanthus roseus was investigated. It was found that the plant cultured cells of N. tabacum and C. roseus reduce stereoselectively the carbonyl group of (+)- and (-)-camphorquinones to the corresponding α-keto alcohols.

Full text of DOI:10.1016/S0031-9422(01)00133-9

Phytochemistry 57 (2001) 669–673
www.elsevier.com/locate/phytochem
Biotransformation of (+)- and (ꢀ)-camphorquinones
by plant cultured cells
a,
Wen Chaia, Hiroki Hamadab, Jumpei Suharaa, C.Akira Horiuchi *
aDepartment of Chemistry, Rikkyo (St. Paul’s) University, Nishi-Ikebukuro, Toshima-ku, Tokyo 171-8501, Japan
bDepartment of Applied Science, Faculty of Science, Okayama University of Science, Ridai-cho, Okayama 700-0005, Japan
Received 15 August 2000; received in revised form 23 January 2001
Abstract
Biotransformation of (+)- and (ꢀ)-camphorquinones with suspension plant cultured cells of Nicotiana tabacum and Cathar-
anthus roseus was investigated.It was found that the plant cultured cells of N. tabacum and C. roseus reduce stereoselectively the
carbonyl group of (+)- and (ꢀ)-camphorquinones to the corresponding a-keto alcohols. # 2001 Elsevier Science Ltd.All rights
reserved.
Keywords: Nicotiana tabacum; Solanaceae; Catharanthus roseus; Apocynaceae reduction; Biotransformation; (+)-Camphorquinone; (ꢀ)-Cam-
phorquinone; 2-exo-Hydroxyepicamphor; 2-endo-Hydroxyepicamphor; 3-exo-Hydroxycamphor; 3-endo-Hydroxycamphor
1. Introduction
(ꢀ)-camphorquinones (1a and 1b) by N. tabacum and
C. roseus yields the corresponding a-keto alcohols.
Asymmetric reductions of ketones by biocatalysts are
useful preparative methods to obtain chiral alcohols
(Sih and Chen, 1984).Chiral a-keto alcohols are useful
and important intermediates for the synthesis of opti-
cally active natural products having stereodirecting
groups.In previous work, microbes such as baker’s
yeast have been used widely to reduce camphorquinone.
(Pfrunder and Tamm, 1969; Chenevert and Thiboutot,
1988; Rebolledo et al., 1991; Miyazawa et al., 1995). In
the report of Miyazawa, (+)-camphorquinone (1a) was
stereoselectivity reduced by Glemerella cingulata to give
(ꢀ)-3S-exo-hydroxycamphor (4a).
2. Results and discussion
2.1. Biotransformation of (+)-camphorquinone (1a)
and (ꢀ)-camphorquinone (1b) by N. tabacum
It was found that cultured cells of N. tabacum reduce
substrates 1a and 1b into a-keto alcohols.( ꢀ)-Cam-
phorquinone (1b) afforded a mixture of diastereomeric
isomers of four a-keto alcohols: (ꢀ)-2S-exo-hydroxy
epicamphor (3b); (+)-3R-exo-hydroxycamphor (2b);
(ꢀ)-3S-endo-hydroxycamphor (4b) and (+)-2R-endo-
hydroxyepicamphor (5b).(+)-Camphorquinone ( 1a)
was bioselectively transformed to give three corre-
sponding enantiomers: (+)-2R-exo-hydroxyepicamphor
(3a); (ꢀ)-3S-exo-hydroxycamphor (4a) and (+)-3R-
On the other hand, plant cultured cells have been
studied for the ability to transform foreign substrates
into useful substances (Suga and Hirata, 1990).How-
ever, hitherto known biochemical reduction of the car-
bonyl group by plant culture cells is only NADH-
dependent reduction of the C–C double bond of a,b-
unsaturated ketones (Hirata et al., 1982; Shimoda et al.,
1998).There is very little information on the bio-
transformation of diketones by plant cultured cells.
Here, we report that the biotransformation of (+)- and
endo-hydroxycamphor (5a).However,
(
ꢀ)-2S-endo-
hydroxyepicamphor (2a) could not be formed from 1a
in this work (Schemes 1 and 2).Table 1 shows results of
time-course experiments.(+)-Camphorquinone ( 1a) was
easily reduced to obtain (ꢀ)-3S-exo-hydroxycamphor
(4a, 57%) and (+)-2R-exo-hydroxyepicamphor (3a,
37%) as the major products after 10 h incubation.
Substrate 1b was transformed to (ꢀ)-2S-exo-hydroxy
* Corresponding author.Fax: +81-3-5992-3434.
E-mail address: horiuchi@rikkyo.ac.jp (C.A. Horiuchi).
0031-9422/01/$ - see front matter # 2001 Elsevier Science Ltd.All rights reserved.
PII: S0031-9422(01)00133-9

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W. Chai et al. / Phytochemistry 57 (2001) 669–673
Scheme 1.Biotransformation of (+)-camphorquinone ( 1a) by plant cultured cells.
Scheme 2.Biotransformation of ( ꢀ)-camphorquinone (1b) by plant cultured cells.
Table 1
Biotransformation of camphorquinone by plant cultured cells
Substrate
Plant cultured cells
T/ha
Total yield/%b
Product ratio/%
(+)-Camphorquinone (1a)
2a
/
3a
11
37
4a
66
57
5a
23
6
C. roseus
N. tabacum
3
10
96
82
/
(ꢀ)-Camphorquinone (1b)
2b
25
8
3b
5
4b
29
37
5b
41
6
C. roseus
N. tabacum
10
18
93
98
49
a
T is reaction time.
Total yield is the yield of product isolated.
b
epicamphor (3b, 49%) and (ꢀ)-3S-endo-hydro-
xycamphor (4b, 37%) after 18 h.(Figs.1 and 2).
samples (Thoren, 1970).From the 1H NMR spectral
data of the reaction mixtures, the presence of 2a-
hydroxycamphor (2) and 3a-hydroxycamphor (5) has
been shown (Table 2).Then, the structures and relative
yields of the products were determined on the basis of
the H NMR spectral peak areas of the proton at C2 or
C3 position.The transformation products and their
yields are shown in Table 1.
In this paper, we have used plant suspension cells to
transform camphorquinone.(+)-Camphorquinone ( 1a)
was stereoselectively reduced by C. roseus and N. taba-
cum to give (ꢀ)-3S-exo-hydroxycamphor (4a) over a
short time period (3–10 h).It was found that it is more
rapid than the reduction of 1a by Glemerella cingulata
and Mucor mucedo (Miyazawa et al., 1995) with the
same stereoselectivity (9–24 h).
2.2. Biotransformation of (+)-camphorquinone (1a)
and (ꢀ)-camphorquinone (1b) by C. roseus
1
Incubation of substrates 1a and 1b with cultured cells
of C. roseus, also gave the corresponding a-keto alco-
hols.Compound 1a rapidly disappeared and was unde-
tectable after 3 h incubation.Compound
1a was
converted to (ꢀ)-3S-exo-hydroxy-camphor (4a, 66%) as
the major product.Compound 1b was transformated to
(+)-2R-endo-hydroxyepicamphor (5b, 41%) as the
major product at the end of 10 h.
2.3. Discussion
After incubation, the column mixtures of a-keto alco-
hols were applied to a silica gel.Elution with n-hexane–
Et2O (3:1) gave two products: 2b-hydroxycamphor (3)
and 3b-hydroxycamphor (4) were identified by their IR,
GC–MS and NMR spectral data.In addition, the 1H
NMR spectral data of the proton at C2 or C3 position
of products 3 and 4 agreed with those of the authentic
3. Experimental
3.1. Analytical and substrates
IR: Jasco FT–IR 230; GC–MS: Shimadzu GCMS-
QP5050 (EI–MS 70 eV) using DB1 (0.25 mm ꢁ 30 m,

W. Chai et al. / Phytochemistry 57 (2001) 669–673
671
Fig.1. Biotransformation of (1 S)-(+)-camphorquinone (1a) using Nicotiana tabacum.
Fig.2. Biotransformation of (1 R)-(ꢀ)-camphorquinone (1b) using Nicotiana tabacum.

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W. Chai et al. / Phytochemistry 57 (2001) 669–673
Table 2
1H NMR spectraa of the a-ketols
b
Compound
CDCl3: ꢂ C2 or C3
Compound
CDCl3: ꢂ C2 or C3
2a-Hydroxycamphor
2b-Hydroxycamphor
3b-Hydroxycamphor
3a-Hydroxycamphor
3.87 (s, b, 1H)
3.58 (s, 1H)
3.72 (s, 1H)
2
3.86 (s, b, 1H)
3.54 (s, 1H)
3.75 (s, 1H)
3 (isolated)
4 (isolated )
5
4.28 (d, J=5 Hz, 1H)
4.21 (d, J=5 Hz, 1H)
a
Shifts are given in ppm downfield from the TMS signal; s=singlet; d=doublet; b=broadened.
Thoren (1970).
b
0.25 mm) capillary column GC; GC: GC-17A at a col-
extracted with EtOAc–Et2O (1:1).The resulting residue
was applied to a silica gel column, that was eluted with
n-hexane–Et2O (3:1) to give: 2b-hydroxycamphor (3),
(IR (KBr): ꢁ 3448, 1751 and 1098 cmꢀ1; CI–MS m/z:
umn temp.of 80–200 ꢂC at 10ꢂC/min; 1H and 13C NMR:
Jeol GSX 400 spectrometer.
(1S)-(+)-Camphorquinone was purchased from
25
Aldrich Chem.Co.(mp 200–202 ꢂC, ½ꢀꢃD +100ꢂ (C6H5
[M+H]+ 169; H NMR (CDCl3): ꢂ (ppm) 0.94 (s, 3H),
1
CH3; c 1.9)). (1R)-(ꢀ)-Camphorquinone was purchased
1.03 (s, 3H), 1.04 (s, 3H), 1.18–1.94 (m, 4H), 2.18 (d,
J=7 Hz, 1H), 2.65 (s, b, 1H) and 3.54 (s, 1H); 13C
NMR (CDCl3): ꢂ (ppm) 10.3, 18.9, 20.3, 21.2, 33.9, 46.6,
49.2, 58.6, 79.5 and 218.7) and 3b-hydroxycamphor (4),
(IR (KBr): ꢁ 3448, 1751 and 1123 cmꢀ1; CI–MS m/z:
from Tokyo Kasei Kogyo Co., Ltd (mp 200ꢂC).
3.2. Cultivation of suspension cells of N. tabacum and
C. roseus
1
[M+H]+ 169; H NMR(CDCl3): ꢂ (ppm) 0.94 (s, 3H),
The callus tissues of N. tabacum were transferred to
freshly prepared MS medium (Murashige and Skoog,
1962) containing 1 ppm of 2,4-dichlorophenoxyacetic
acid as auxin and 3% sucrose, and then were grown with
continuous shaking (110 rpm) for 8 days at 25ꢂC under
the light.The callus tissues of C. roseus were transferred
to freshly prepared SH medium (Schenk and Hildeb-
randt, 1976) containing 2 ppm of 2,4-dichlorophenox-
yacetic acid as auxin and 3% sucrose, and then were
grown with continuous shaking (110 rpm) for 8 days at
25ꢂC under the light.
0.95 (s, 3H), 1.00 (s, 3H), 1.20–2.00 (m, 4H), 2.10 (d,
J=5 Hz, 1H), 2.59 (s, b, 1H) and 3.75 (s, 1H); 13C
NMR(CDCl3): ꢂ (ppm) 9.03, 20.1, 21.0, 25.2, 28.6, 48.2,
49.2, 57.0, 77.4 and 220.0) (Thoren, 1970)
The ratios of the isomers were calculated from the
peak areas of 1H NMR spectral data: ꢂ 3.86 (s, b, 1H for
2), 3.54 (s, 1H for 3), 3.75 (s, 1H for 4) and 4.21 (d, J=
5 Hz, 1H for 5).
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