Biodegradation of bisphenol A by cultured cells of Caragana chamlagu.

Article abstract of DOI:10.1271/bbb.67.218

The biological degradation of 2,2-bis(4-hydroxyphenol)propane (1; bisphenol A, BPA), a representative endocrine disruptor, was studied with plant-cultured cells of Caragana chamlagu. An initial BPA concentration of 425 microM in an aqueous solution was degraded by C. chamlagu at 25 degrees C for 2 days in the dark, and two intermediates were then completely dissipated after 10 days.

Full text of DOI:10.1271/bbb.67.218

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Biodegradation of Bisphenol A by Cultured Cells of
Caragana chamlagu
Wen CHAIa, Hiroshi SAKAMAKIb, Susumu KITANAKAc, Michihiko SAITOd & C. Akira
HORIUCHIa
a Department of Chemistry, Rikkyo (St. Paul's) University 3-34-1 Nishi-Ikebukuro,
Toshima-ku, Tokyo 171-8501, Japan
b College of Science and Technology, Nihon University 7-24-1 Funabashi-shi, Chiba
274-8501, Japan
c College of Pharmacy, Nihon University 7-7-1 Funabashi-shi, Chiba 274-8501, Japan
d Food Packaging Lab., National Food Research Institute 2-1-12 Kannondai, Tsukuba-shi,
Ibaraki 305-8642, Japan
Published online: 22 May 2014.
To cite this article: Wen CHAI, Hiroshi SAKAMAKI, Susumu KITANAKA, Michihiko SAITO & C. Akira HORIUCHI (2003)
Biodegradation of Bisphenol A by Cultured Cells of Caragana chamlagu , Bioscience, Biotechnology, and Biochemistry,
67:1, 218-220, DOI: 10.1271/bbb.67.218
To link to this article: http://dx.doi.org/10.1271/bbb.67.218
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Biosci. Biotechnol. Biochem., 67 (1), 218–220, 2003
Preliminary Communication
Biodegradation of Bisphenol A by Cultured Cells of Caragana chamlagu
4
Wen CHAI,1 Hiroshi SAKAMAKI,2 Susumu KITANAKA,3 Michihiko SAITO
and C. Akira HORIUCHI
,
1,
†
1Department of Chemistry, Rikkyo (St. Paul's) University, 3-34-1 Nishi-Ikebukuro, Toshima-ku,
Tokyo 171-8501, Japan
2College of Science and Technology, Nihon University, 7-24-1 Funabashi-shi, Chiba 274-8501, Japan
3College of Pharmacy, Nihon University, 7-7-1 Funabashi-shi, Chiba 274-8501, Japan
4Food Packaging Lab., National Food Research Institute, 2-1-12 Kannondai, Tsukuba-shi,
Ibaraki 305-8642, Japan
Received July 29, 2002; Accepted October 24, 2002
(98 atom
z
D) was purchased from Aldrich Chemical
The biological degradation of 2,2-bis(4-hydroxy-
phenol)propane (1; bisphenol A, BPA), a representative
endocrine disruptor, was studied with plant-cultured
cells of Caragana chamlagu. An initial BPA concentra-
(Milwaukee, WI, U.S.A.). The Murashige and Skoog
medium was purchased from Wako Pure Chemical
Industries (Osaka, Japan), as were 2,4-dichloro-
phenoxyacetic acid (Ti grade
z
À98 ) and sucrose.
tion of 425
C. chamlagu at 25
m
M
in an aqueous solution was degraded by
C for 2 days in the dark, and two
The callus tissue induced from the stem of
Caragana chamlagu (Leguminosae) that was used in
our previous study6) was also used in this investiga-
tion, this callus tissue of C. chamlagu having been
maintained for approximately 6 years. The callus tis-
sue of C. chamlagu was transferred to a freshly pre-
pared Murashige-Skoog medium7) (containing 1 ppm
9
intermediates were then completely dissipated after 10
days.
Key words: bisphenol
A;
plant-cultured
cells;
Caragana chamlagu; biological degrada-
tion; endocrine disrupter
of 2,4-dichlorophenoxyacetic acid as auxin, 3
crose, and 0.8
dark.
The cultured cells of C. chamlagu (10 g) were
transplanted to the MS medium (200 ml) containing
1 ppm of 2,4-dichlorophenoxyacetic acid, and the
suspension cells were then incubated while shaking
z
su-
z agar), and then grown at 259C in the
2,2-Bis(4-hydroxyphenyl)propane (bisphenol A or
BPA, 1) is generally used as a starting material for
polymers including polycarbonates, epoxy resins,
phenol resins, polyesters, and polyacrylates. This
compound is commonly suspected to act as an endo-
crine disruptor.1) Some examples of the biological
degradation of BPA by bacteria2) and basidiomy-
cetes3) have recently been reported. However, these
biological methods to eliminate the pollutant in an a-
queous solution cannot completely decompose the
total organic carbon.
There is growing interest in the ability of plant-cul-
tured cells. One example is the convertion of foreign
substances in the stereo- and regioselective control of
organic synthesis.4,5) However, to our knowledge,
there has been no study on the decomposition of
BPA by means of plant-cultured cells. We have been
investigating oxidation by using plant-cultured cells.
In particular, the plant-cultured cells of Caragana
chamlagu had high ability for oxidation.6) We report
here the total degradation of BPA in an aqueous so-
lution by using C. chamlagu in the biocatalytic reac-
tion.
(110 rpm) at 25
20 mg, 425 ) was then added to the suspension
9C in the dark for 5 days. BPA (1;
m
M
cells which were further incubated. After the indicat-
ed incubation period, the cultured cells were removed
by ˆltration, and the ˆltrate was extracted with
distilled ether. The amount of residual BPA was
measured by gas chromatography-mass spectrometry
(GC-MS: GCMS-QP5050, Shimadzu, Tokyo,
Japan). GC was programmed to raise the oven
temperature from 509C to 2509C at 189C min, and
W
MS was conducted at 70 eV (EI) or 300 eV (CI). The
reaction products were isolated by column chro-
matography on silica gel 60 (Merck, Darmstadt,
Germany), using n-hexane-Et2O (3:1) as the eluent.
The chemical structures of the reaction products were
determined by GC-MS, IR spectra (Jasco FT-IR 230
spectrometer, Japan), and H and 13C-NMR spectra
1
which were measured with a Jeol GSX 400 spectrom-
eter (Nihon Denshi, Tokyo, Japan) in deu-
BPA (GC grade
À99z) was purchased from
Tokyo Kasei (Tokyo, Japan), and BPA-d16
†
To whom correspondence should be addressed. Tel Fax: +81-3-3985-2397; E-mail: horiuchi
＠
rikkyo.ac.jp
W

Biodegradation of Bisphenol A
219
Fig. 1. Biocatalyic Degradation of Bisphenol A by C. chamlagu
.
Table 1. Main Fragment Ion (m z) of the Reaction Compounds by a GC-MS Analysis (EI)
W
teriochloroform solutions with tetramethylsilane as
an internal standard.
110.1, 115.8, 127.4, 142.8, 155.9; EIMS m z: 134
W
(100) [M]+, 119 (89) [M-CH3]+, 91 (53), 77 (27), 65
The activity of C. chamlagu in degrading BPA in
vivo was investigated. First, the degradation of BPA
by the suspension cells of C. chamlagu was exam-
ined. The results of time-course experiments are
shown in Fig. 1. It can be seen from Fig. 1 that BPA
disappeared after 2 days of incubation, and then
intermediates 1a and 1b disappeared after 10 days.
Compound 1b was isolated by column chro-
matography and followed by GC-MS, IR, and NMR
analysis. The spectral data of 1b are as follow: IR
nmax (KBr) cm„1: 3405; NMR dH (CDCl3): 2.12 (3H,
(42) and 51 (47); CIMS m z: 135 (100) [M+H]+. The
W
spectra of 1b show that the structure is in agreement
with that of 4-isopropenylphenol. 1a was deduced to
be 4-(2-propanol)phenol by its GC-MS spectrum. In
order to clarify that their origin was not in the cells,
the biodegradation of BPA-d16 by C. chamlagu was
carried out. The results of GC-MS spectra show that
the deuterides of 1a and 1b (1a-d10 and 1b-d9) were
obtained as intermediates, proving that 1a and 1b
were intermediates of the biodegradation of BPA.
The results of the time-course experiments indicate
that oxidative cleavage of the C-C bond had oc-
curred. In order to identify the pathway for the bio-
＝
8.8 Hz)
s), 4.98 (1H, s), 5.27 (1H, s), 6.79 (2H, d,
J
and 7.36 (2H, d, 8.8 Hz); NMR C (CDCl3): 29.2,
J
＝
d

220
W. CHAI et al.
with dioxygenases, which is similar to the reaction of
BPA by KO2. This is the ˆrst time that a BPA con-
centration of 425 in an aqueous solution has been
completely dissipated by plant-cultured cells of C.
chamlagu
m
M
.
Acknowledgment
This work was ˆnancially supported by the Endo-
crine Disrupters Project of the Ministry of Agricul-
ture, Forestry, and Fisheries, Japan.
Scheme 1. Veriˆcation Experiments of Pathway for BPA Bio-
degradation.
References
1) Alexander, H. C., Dill, D. C., Smith, L. W., Guiney,
P. D., and Dorn, P., Bisphenol A: Acute aquatic
toxicity. Environmental Toxicol. Chem., 7, 19–26
(1988).
2) Spivack, J., Leib, T. K., and Lobus, J. H., Novel
pathway for bacterial metabolism of bisphenol A. J.
Biol. Chem., 269, 7323–7329 (1994).
3) Hirano, T., Honda, Y., Watanabe, T., and Kuwahara,
M., Degradation of bisphenol A by the lignin-degrad-
ing enzyme, manganese peroxidase, produced by the
Scheme 2. Pathway for BPA Biodegradation by Cultured Cells
of C. chamlagu
.
white-rot basidiomycete, Pleurotus ostreatus
Biotechnol. Biochem., 64, 1958–1962 (2000).
. Biosci.
degradation, we investigated the oxidation of the su-
peroxide anion radical [O„2 ] with BPA (1). A typical
procedure was the reaction of potassium superoxide
(20.0 mmol), 18-crown 6-ether (10.0 mmol), BPA (1,
4) Suga, T., and Hirata, T., Review article number 55,
biotransformation of exogenous substrates by plant
cell cultures. Phytochemisty, 29, 2393–2406 (1990).
5) Chai, W., Hamada, H., Suhara, J., and Horiuchi, C.
A., Biotransformation of (+)- and („)-camphorqui-
nones by plant cultured cells. Phytochemistry, 57,
669–673 (2001).
2.0 mmol) and DMF (60 ml) at 25
The resulting mixture was extracted with distilled
ether and gave 4-isopropenylphenol (1b, 30 ).
9
C for 35 hours.
z
In the biological degradation of BPA with the bac-
terium or basidiomycete, the reaction pathway can be
explained by the work of the lignin-degrading en-
zyme.3) However, it is not believed that the degrada-
tion pathway of BPA using plant-cultured cells is
similar to the reaction of the lignin-degrading en-
zyme. On the basis of the foregoing results, it is pos-
sible that the biological degradation of BPA proceed-
ed by oxidation of the superoxide anion radical [O„2 ]
6) Sakamaki, H., Kitanaka, S., Chai, W., Hayasida, Y.,
Takagi, Y., and Horiuchi, C. A., Biotransformation
of thujopsene by Caragana chamlagu. J. Nat. Prod.
64, 630–631 (2001).
,
7) Murashige, T., and Skoog, F., A resised medium for
rapid growth and bio assays with tobacco tissue cul-
tures. Physiol. Plant., 15, 473–497 (1962).