Isolation and Identification of a New 2-Phenylbenzotriazole-Type Mutagen (PBTA-3) in the Nikko River in Aichi, Japan

• Published in: Chemical research in toxicology 2000-07, Vol.13 (7), p.535-540
• Main Authors: Shiozawa, Tatsushi, Tada, Atsuko, Nukaya, Haruo, Watanabe, Tetsushi, Takahashi, Yoshifumi, Asanoma, Masaharu, Ohe, Takeshi, Sawanishi, Hiroyuki, Katsuhara, Taka, Sugimura, Takashi, Wakabayashi, Keiji, Terao, Yoshiyasu
• Format: Article
• Language: English
• Subjects: 2-Phenylbenzotriazole; Chromatography, High Pressure Liquid; Freshwater; Japan; Japan, Nikko R; Magnetic Resonance Spectroscopy; Mutagens - analysis; Mutagens - chemical synthesis; Mutagens - isolation & purification; Triazoles - analysis; Triazoles - chemical synthesis; Triazoles - isolation & purification; Water Pollutants, Chemical - analysis; Water Pollutants, Chemical - chemical synthesis; Water Pollutants, Chemical - isolation & purification
• ISSN: 0893-228X; 1520-5010
• DOI: 10.1021/tx0000264

We have previously determined the chemical structures of two 2-phenylbenzotriazole mutagens (PBTA-1 and PBTA-2) in blue cotton-adsorbed material from the Nishitakase River in Kyoto, Japan. In the present study, further analysis of mutagenic substances in the Nikko River, which flows through Aichi Prefecture in Japan, allowed the isolation of a new mutagen. Material (2.2 g) adsorbed on blue cotton (3 kg) at a site below the sewage plant on the Nikko River was purified by various column chromatographies, and a mutagen (120 μg) accounting for 11% of the total mutagenicity was isolated. On the basis of data from UV, mass, and 1H NMR spectra of the mutagen, the compound was deduced to be a PBTA-1 analogue. As with PBTA-1, the mutagen was able to be synthesized from the azo dye 2-[(2-bromo-4,6-dinitrophenyl)azo]-4-methoxy-5-[(2-hydroxyethyl)amino]acetanilide by reduction and chlorination. Since all spectra of the mutagen isolated from the river water were the same as those of the synthesized form, the structure was concluded to be 2-[2-(acetylamino)-4-[(2-hydroxyethyl)amino]-5-methoxyphenyl]-5-amino-7-bromo-4-chloro-2H-benzotriazole (PBTA-3). PBTA-3 is a potent mutagen, inducing 81 000 and 3 000 000 revertants per microgram of Salmonella typhimurium TA 98 and YG1024 respectively, in the presence of an S9 mix. In addition to its detection in the water of the Nikko River, PBTA-3 was detected in water samples from three other rivers flowing through regions where dyeing industries have been developed. Like PBTA-1 and PBTA-2, PBTA-3 might have also been produced from azo dyes during industrial processes in dyeing factories and/or through treatment at sewage plants.