Identification of biliary metabolites of m-dichlorobenzene in rats

At least 12 metabolites were observed in the bile of rats administered with m-dichlorobenzene in the preliminary experiments by HPLC. Some of them were assumed to be conjugates containing phenyl and dihydro-hydroxyphenyl moieties with glutathione or cysteine by enzymatic and thermal reactions. To isolate these metabolites, the bile collected from 20 rats administered with 500 mg/kg of m-dichlorobenzene and 300 mg/kg of cysteine was separated into five fractions by HPLC. Eighteen metabolites were isolated from these fractions by TLC and HPLC. The chemical structure of the major metabolite excreted was determined as trans-2,4-dichloro-6-(glutathion-S-yl)cyclohexa-2,4-dien+ ++-1-ol by 13C-NMR, 1H-NMR, FAB-MS, and some reaction experiments. The metabolite excreted in the secondary largest amount was identified as its positional isomer, trans-3,5-dichloro-6-(glutathion-S-yl)cyclohexa-2,4-dien+ ++-1-ol. Their diastereomers were also observed in the bile. trans-2,4-Dichloro-6-(cystein-S-yl)cyclohexa-2,4-dien- 1-ol and its positional isomer, which were possibly derived from glutathione conjugates above, were also identified. 3,5-Dichlorophenyl conjugates with glutathione or cysteine and 3,5-dichlorophenyl mercapturic acid, and their 2,4-dichlorophenyl isomers, were excreted. Three monochlorophenol conjugates, of which chemical structures were still not established, were present in the bile.