Detection of 3,3′-Dichlorobenzidine in Water from the Waka River in Wakayama, Japan

Abstract

In a previous study, we found that water concentrate from the Waka River, which flows through an industrial area in Wakayama, Japan, showed significant mutagenicity in Salmonella typhimurium YG1024 without a mammalian metabolic system (S9 mix), and 4-amino-3,3′-dichloro-5,4′-dinitrobiphenyl (ADDB) was identified as a major direct-acting mutagenic constituent in the water concentrate. ADDB induced 428,000 revertants/μg in YG1024 without S9 mix, and this activity was 48 times as high as that with S9 mix. In this study, to clarify whether other mutagenic contaminants were present in the river, water concentrates were collected at three sites along the Waka River and examined for mutagenicity by the Ames test using YG1024 and YG1029 with and without S9 mix. All water concentrates showed potent mutagenicity in YG1024 with and/or without S9 mix. ADDB was detected in water concentrate that was extremely mutagenic without S9 mix. An indirect-acting mutagen, which accounted for more than 30% of the total mutagenicity of the water concentrates in YG1024 with S9 mix, was isolated from two extremely mutagenic water concentrates by HPLC separation. On the basis of spectral data and co-chromatography using authentic chemicals, this mutagen was identified as 3,3′-dichlorobenzidine (DCB). DCB showed strong mutagenicity in YG1024 with S9 mix, inducing 5,186 revertants/μg. DCB was detected in water concentrates collected on the other three sampling dates in the range from 15.8 to 28.5 μg/g of blue rayon. These results suggest that Waka River water might be continually contaminated with mutagens, and DCB was thought to be the major mutagenic constituent of the river water.