DNA polymerase β
    Masami Yamada
DNA polymerase β
    山田雅巳………………………………………………………………………………………………………115

Original Article
一般論文

Micronucleus test in rats after oral administration of methyl methanesulfonate for 4 weeks
     Shigenari Ozawa, Rika Takayama, Kazuo Kobayashi, Tsuyoshi Kitamura and Nobuo Shibata ………………125

11 th JEMS Annual Symposium “Chemical Mechanisms of Depression and Repression of Activity In Environmental Mutagenesis ”
第11 回公開シンポジウム「環境変異原の活性発現と抑制の化学的機構」

Organic chemistry plays critical roles in studies of environmental mutagens
     Hikoya Hayatsu
環境変異原研究における有機化学の役割
     早津彦哉………………………………………………………………………………………………………131

Does tamoxifen - induced DNA damage cause endometrial cancer?
     Shinya Shibutani ………………………………………………………………………………………………133

Formation of direct mutagen MX in water samples and induction of toxic effects
     Naohide Kinae, Chitose Sugiyama, Kayoko Shimoi
水中の直接変異原MX の生成と毒性発現の機構
     木苗直秀，杉山千歳，下位香代子…………………………………………………………………………141

Formation and development of mutagenic and/or carcinogenic compounds through free radical mechanisms in foods
     Kiyomi Kikugawa, Kazuyuki Hiramoto and Tetsuta Kato
フリーラジカルを経由する変異・発がん物質の生成と発現
     菊川清見，平本一幸，加藤哲太……………………………………………………………………………149

Electrochemical studies of quinone and nitroarene in generation and quenching of superoxide
     Kiyoshi Fukuhara and Naoki Miyata
化学物質による活性酸素の生成と消去
     福原 潔，宮田直樹……………………………………………………………………………………………155

Genotoic properties of alkylating agents ：mutagenicity and its modulating effects
     Kazuhiko Takahashi, Chiharu Kato, Hiroaki Kato
アルキル化剤の変異原性発現と抑制機構
      高橋和彦，加藤千晴，加藤宏明……………………………………………………………………………163

Activation mechanism of environmental mutagens by porphyrin iron model oxidation systems
     Keiko Inami, Eriko Okochi, Masataka Mochizuki
ポルフィリン・鉄モデル酸化系による変異原活性化の機構
      稲見圭子，大河内江里子，望月正隆………………………………………………………………………173
 
 

Does tamoxifen induced DNA damage cause endometrial cancer?
Shinya Shibutani
Laboratory of Chemical Biology, Department of Pharmacological Sciences,
State University of New York at Stony Brook,
Stony Brook, New York, USA 11794 - 8651
Summary
I have previously described genotoxicity of tamoxifen-derived DNA adducts in this journal
（Environ. Mutagen Res., 20 ：201- 211, 1998 ）. However, a large number of reports regarding this
issue have published in last two years. Here I review the recent findings from my laboratory and
others. Tamoxifen- DNA adducts are primarily formed via sulfation of α - hydroxy moieties of
tamoxifen, N- desmethyltamoxifen, and tamoxifen N- oxide. α -（N²- Deoxyguanosinyl ）tamoxifen
（dG- N²-TAM ）and α -（N²- deoxyguanosinyl ）- N- desmethyltamoxifen （dG- N²- N- desmethylTAM ）
were major adducts in the liver of rats and mice treated with tamoxifen. α -（N²- Deoxyguanosinyl ）
tamoxifen N- oxide （dG- N²-TAM N- oxide ）was also detected as a minor adduct in mouse liver.
Site-specific mutagenesis studies showed that dG- N²-TAM is a potential mutagenic lesion,
generating mainly G →T transversions, accompanied by fewer G →A transitions in mammalian
cells. This tamoxifen adduct can be removed from DNA by nucleotide excision repair. Significant
amounts of dG- N²- TAM adducts were detected in the endometrium of women treated with
tamoxifen; a marked inter-individual variation was observed in the level of tamoxifen - DNA adducts.
Such mutagenic tamoxifen- DNA adducts, if not repaired, may act as initiators, leading to
development of endometrial cancer. Genotoxicity of toremifene, a chlorinated tamoxifen analog, was
much lower than tamoxifen, due to the limited formation of DNA adducts induced by toremifene
metabolites. Use of toremifene, instead of tamoxifen, may reduce incidence of endometrial cancer in
women receiving breast cancer prevention treatment.
Keywords ：tamoxifen ，toremifene ，DNA adduct ，endometrial cancer, breast cancer
received ：October 27, 2000 accepted ：October 30, 2000
Environmental Mutagen Society of Japan
This paper was presented at the 11 th JEMS Annual Symposium at the ABC Hall, Tokyo, June 3 rd, 2000. The Symposium entitled
“Chemical Mechanisms of Depression and Repression of Activity In Environmental Mutagenesis ”，was organized by
Masataka Mochizuki and sponsored by the Japanese Environmental Mutagen Society.
 
 

水中の直接変異原MX の生成と毒性発現の機構
木苗 直秀，杉山 千歳，下位 香代子
静岡県立大学食品栄養科学部 〒422- 8526 静岡市谷田52 - 1
Formation of direct mutagen MX in water samples and induction of toxic effects
Naohide Kinae, Chitose Sugiyama, Kayoko Shimoi
School of Food and Nutritional Sciences, University of Shizuoka, 52 - 1, Yada, Shizuoka 422 - 8526, Japan
Summary
A potent direct mutagen, 3- chloro -4 -dichloromethyl -5 -hydroxy- 2 （5H ）-furanone （MX ），has been
detected from tap water, swimming pool water and domestic effluent after chlorine disinfection. MX induced
glandular stomach cancer on Wistar rats in two stage carcinogenesis test with N- methy- Nﾕ- nitro - N-nitrosoguanidine.
After intragastrically administration of MX to Wistar rats, the liver and stomach were
applied to ³²P- postlabeling method, DNA- adducts were formed with and without 2- amino -3 -methyl
imidazo -［4, 5 - f ］- quinoxaline （MeIQx ）which is found in broiled meats and fish. To detect the precursors
of MX contained in domestic effluent, several materials were chlorinated and analyzed by GC- MS- SIM
method. Polyphenolic compounds containing （±）catechin and diosmin were identified. These results show
that MX may give adverse effect on aquatic organisms and also on human beings as a potent mutagen and
carcinogen in the water samples. Therefore, the risk assessment and risk management of MX will be
important task in near future.
Keywords ：3- chloro -4 -dicholoromethyl -5 -hydroxy- 2 （5H ）-furanone （MX ），mutagenicity, carcinogenicity ，DNA- adduct ，MX precursor
受付：2000 年9 月25 日 受理：2000 年9 月25 日
日本環境変異原学会
This paper was presented at the 11 th JEMS Annual Symposium at the ABC Hall, Tokyo, June 3 rd, 2000. The Symposium entitled
“Chemical Mechanisms of Depression and Repression of Activity In Environmental Mutagenesis ”，was organized by
Masataka Mochizuki and sponsored by the Japanese Environmental Mutagen Society.
 

フリーラジカルを経由する変異・発がん物質の生成と発現
菊川 清見，平本 一幸，加藤 哲太
東京薬科大学薬学部 〒192- 0392 東京都八王子市堀之内1432- 1
Formation and development of mutagenic and/or carcinogenic compounds through free radical mechanisms in foods
Kiyomi Kikugawa, Kazuyuki Hiramoto and Tetsuta Kato
School of Pharmacy, Tokyo University of Pharmacy and Life Science,
1432 - 1, Horinouchi, Hachioji, Tokyo 192 - 0392, Japan
Summary
Free radicals including carbon - centered, oxygen - centered and pyrazine cation radicals are involved in the
DNA damage and the formation of mutagens in foods. Carbon- centered radicals generated from
arylbenzenediazonium salts modified DNA bases. Reactive oxygen species generated from fragrant
furanones or pyranones produced by Maillard reaction in many foodstuffs or hydroxyhydroquinone in
roasted coffee beans broke the DNA strand. Scavenging of the free radicals by ethanol or phenolic
antioxidants prevented the DNA damage. Pyrazine cation radical produced in the Maillard reaction
participated in the formation of heterocyclic amine mutagens. Scavenging of the pyrazine cation radicals by
ascorbate or erythorbate or decreasing the pyrazine cation radical by controlling sugar content prevented
the formation of the mutagens. The mutagenicty of hamburger was effectively reduced by addition of
ascorbate or erythorbate and by addition of a large amount of sugars or onion.
Keywords ：carbon- centered radical ，reactive oxygen species ，pyrazine cation radical ，Maillard reaction ，DNA damage ，heterocyclic amine.
受付：2000 年7 月10 日 受理：2000 年7 月12 日
日本環境変異原学会
This paper was presented at the 11 th JEMS Annual Symposium at the ABC Hall, Tokyo, June 3 rd, 2000. The Symposium entitled
“Chemical Mechanisms of Depression and Repression of Activity In Environmental Mutagenesis ”，was organized by
Masataka Mochizuki and sponsored by the Japanese Environmental Mutagen Society.
 
 

化学物質による活性酸素の生成と消去
福原 潔，宮田 直樹
国立医薬品食品衛生研究所 〒158- 8501 世田谷区上用賀1-18 -1
Electrochemical studies of quinone and nitroarene in generation and quenching of superoxide
Kiyoshi Fukuhara and Naoki Miyata
National Institute of Health Sciences Setagaya, Tokyo 158 - 8501, Japan
Summary
Quinones are common in several natural products and endogeneous biochemicals or generated through
metabolism of aromatic hydrocarbons. Some quinones are potent redox active compounds which can
undergo enzymatic redox cycling with their corresponding semiquinone anion radicals and as a result
generate superoxide anion radicals.
In this paper, we review the catalytic activity of quinones and nitroarenes, as mediators in the reductive
activation of molecular oxygen and in the oxidative quenching of superoxide, examined by electrochemical
method.
Keywords ：quinone ，nitroarene ，superoxide ，electrochemistry
受付：2000 年8 月11 日 受理：2000 年8 月12 日
日本環境変異原学会
This paper was presented at the 11 th JEMS Annual Symposium at the ABC Hall, Tokyo, June 3 rd, 2000. The Symposium entitled
“Chemical Mechanisms of Depression and Repression of Activity In Environmental Mutagenesis ”，was organized by
Masataka Mochizuki and sponsored by the Japanese Environmental Mutagen Society.
 
 

アルキル化剤の変異原性発現と抑制機構
高橋 和彦，加藤 千晴，加藤 宏明
名古屋市立大学薬学部 〒467- 8603 名古屋市瑞穂区田辺通3-1
Genotoic properties of alkylating agents ：mutagenicity and its modulating effects
Kazuhiko Takahashi, Chiharu Kato, Hiroaki Kato
Faculty of Pharmaceutical Sciences, Nagoya City University
3 - 1 Tanabe - dori, Mizuho-ku, Nagoya 467 - 8603, Japan
Summary
Our study is focused on the relationships between chemical reactivity and genotoxic properties.
Mutational specificities of N- methyl - N- nitrosourea （MNU ）and methyl methanesulfonate （MMS ）were
analysed using ada - and/or ogt - deficient strains of Escherichia coli, which encode O⁶- methylguanine - DNA
methyltransferases. SN 1 - type methylating agents, MNU, induced predominantly GC →AT transitions in all
strains. In contrast, SN 2 - type methylating agents, MMS, showed wide mutation spectrum. MMS effeciently
induced GC →AT and AT →GC transitions in both wild type and ogt - strains. In ada - strain, GC →TA
transversions were strongly induced. However, GC →AT transitions were predominantly induced in ada -ogt- strain.
The capacities of methylating agents in inducing the adaptive response were also examined. Exposure of
E. coli cells to a low concentration of methylating agents increases their resistance to both mutagenic and
cytotoxic effects of alkylating agents. This process is known as the adaptive response and is triggered by
formation of methylphosphotriesters in the DNA. Methyl iodide, a typical SN 2 - type methylating agents and
a very weak mutagen, induced the adaptive response in E. coli to a similar extent to those induced by potent
mutagenic methylating agents such as MNU and N- methyl- N'- nitro - N- nitrosoguanidine （MNNG ）．The
mechanism of induction of the adaptive response involve direct methylation of Ada protein itself.
Keywords ：methylating agents ，mutation spectrum, adaptive response ，O⁶- methylguanine- DNA methyltransferase
受付：2000 年8 月18 日 受理：2000 年8 月18 日
日本環境変異原学会
This paper was presented at the 11 th JEMS Annual Symposium at the ABC Hall, Tokyo, June 3 rd, 2000. The Symposium entitled
“Chemical Mechanisms of Depression and Repression of Activity In Environmental Mutagenesis ”，was organized by
Masataka Mochizuki and sponsored by the Japanese Environmental Mutagen Society.
 

ポルフィリン・鉄モデル酸化系による変異原活性化の機構
稲見 圭子，大河内 江里子，望月 正隆
共立薬科大学 〒105- 8512 東京都港区芝公園1-5-30
Activation mechanism of environmental mutagens by porphyrin iron model oxidation systems
Keiko Inami, Eriko Okochi, Masataka Mochizuki
Kyoritsu College of Pharmacy, Minato - ku, Tokyo 105 - 8512, Japan
Summary
Many environmental carcinogens damage DNA at an initiation step in carcinogenesis, and Ames test is a
short term screening of possible carcinogens. Most carcinogens require enzymatic activation through
oxidation by cytochrome P 450, which metabolizes promutagens to active species in the mutation assay. Iron
（III ）porphyrin and an oxidant catalyze the oxidation mimicking the metabolism by cytochrome P 450. In
this study, some carcinogens were tested in order to generalize the use of the chemical models. We
investigated the best conditions for mutation assay with the chemical models. The activation of 2-aminofluorene,
benzo ［a ］pyrene and tryptophan pyrolysates （Trp -P -1 and Trp -P -2 ）and 2-acetylaminofluorene
（AAF ）in mutation assay using Fe （F5 P ）Cl and three oxidants ；tert - butyl
hydroperoxide （t - BuOOH ），m- chloroperoxybenzoic acid （mCPBA ），and magnesium monoperoxy-phthalate
（MPPT ）．Aromatic amines displayed higher levels of mutagenicity with t - BuOOH, whereas the
polycyclic hydrocarbon was preferentially activated when mCPBA was used. All promutagens were
mutagenic in the presence of Fe （F₅ P ）Cl/MPPT. Our results show that this activation system will be
generally useful as an alternative of the conventional assay with S 9 mix, and a type of mutagens will be able
to be estimated from the difference of activity in different models used.
On the other hands, the mechanism of metabolic activation of AAF was investigated with Fe （F₅ P ）Cl/t -BuOOH.
A major product obtained from this reaction was 2 - nitro -9 -fluorenone （NO₂ F ＝O ），which
showed mutagenicity on Salmonella typhimurium TA1538. During the oxidation of AAF by the chemical
model, the formation of NO₂ F ＝O attended with a decrease of AAF, which was analyzed by HPLC. In order
to estimate the contribution of NO₂ F ＝O to the total mutagenicity after chemical model oxidation,
mutagenicity was assayed using Salmonella typhimurium YG7131 which is deficient in nitroreductase, and
the result was compared to that in a parent strain Salmonella typhimurium TA1538. The mutagenicity of
AAF decreased significantly in Salmonella typhimurium YG7131, which suggested that most of the
mutagenicity derived from AAF in the presence of the chemical model was due to NO₂ F ＝O formed.
Furthermore, the amount of NO₂ F ＝O formed in the incubation mixture accounted for the mutagenicity,
and suggested further that the mutagen formed in the models was NO₂ F ＝O.
Keywords ：Cytochrome P 450 model ，metalloporphyrin ，metabolic activation ，oxidation ，2 - acetylaminofluorene
受付：2000 年7 月10 日 受理：2000 年7 月10 日
日本環境変異原学会
This paper was presented at the 11 th JEMS Annual Symposium at the ABC Hall, Tokyo, June 3 rd, 2000. The Symposium entitled
“Chemical Mechanisms of Depression and Repression of Activity In Environmental Mutagenesis ”，was organized by
Masataka Mochizuki and sponsored by the Japanese Environmental Mutagen Society.