title: The involvement of primary and secondary metabolism in the covalent binding of 1,2- and 1,4-dichlorobenzenes.
abstract: The microsomal oxidation of 1,2-[14C]- and 1,4-[14C]dichlorobenzene (DICB) was investigated with special attention for possible differences in biotransformation that might contribute to the isomer-specific hepatotoxicity. Major metabolites of both isomers were dichlorophenols (2,5-DICP for 1,4-DICB and 2,3- and 3,4-DICP for 1,2-DICB, respectively) and dichlorohydroquinones. The formation of polar dihydrodiols appeared to be a major route for 1,2-DICB but not 1,4-DICB. Both the hepatotoxic 1,2-DICB and the non-hepatotoxic 1,4-DICB were oxidized to metabolites that covalently interacted with protein and only to a small extent with DNA. Protein binding could be inhibited by the addition of the reducing agent ascorbic acid with a concomitant increase in the formation of hydroquinones and catechols, indicating the involvement of reactive benzoquinone metabolites in protein binding. However, in the presence of ascorbic acid, a substantial amount of protein-bound metabolites of 1,2-DICB was still observed, in contrast to 1,4-DICB where binding was nearly completely inhibited. This latter effect was ascribed to the direct formation of reactive benzoquinone metabolites in a single P450-mediated oxidation of para-substituted dichlorophenols (such as 3,4-DICP) in the case of 1,2-DICB. In contrast, the major phenol isomer derived from 1,4-DICB (i.e. 2,5-DICP) is oxidized to its hydroquinone derivative, which needs prior oxidation in order to generate the reactive benzoquinone species. Residual protein binding in the presence of ascorbic acid could also indicate the involvement of reactive arene oxides in the protein binding of 1,2-DICB, but not of 1,4-DICB. However, MO computer calculations did not provide indications for differences in chemical reactivity and/or stability of the various arene oxide/oxepin tautomers that can be formed from either 1,2-DICB or 1,4-DICB. In conclusion, reactive intermediates in the secondary metabolism of 1,2-DICB lead to more covalent binding than those derived from 1,4-DICB, which correlates very well with their reported hepatotoxic potency.
authors.affiliation: Department of Toxicology, Agricultural University, Wageningen, The Netherlands.
authors.forename: C
authors.initials: C
authors.lastname: den Besten
authors.forename: M
authors.initials: M
authors.lastname: Ellenbroek
authors.forename: M A
authors.initials: MA
authors.lastname: van der Ree
authors.forename: I M
authors.initials: IM
authors.lastname: Rietjens
authors.forename: P J
authors.initials: PJ
authors.lastname: van Bladeren
mesh_terms: D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism; D002417:Cattle; D002722:Chlorobenzenes / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D004247:DNA / Q000378:metabolism; D066298:In Vitro Techniques; D008297:Male; D008862:Microsomes, Liver / Q000378:metabolism; D010084:Oxidation-Reduction; D051381:Rats; D017208:Rats, Wistar
chemical_list: D002352:Carrier Proteins; D002722:Chlorobenzenes; C004726:2-dichlorobenzene; D004247:DNA; C018511:4-dichlorobenzene