QUALITATIVE AND QUANTITATIVE INFLUENCES OF ORTHO-CHLORINE SUBSTITUENTS ON THE MICROSOMAL METABOLISM OF 4-TOLUIDINES

The influence of ortho chlorine substituents on 4-toluidine rat liver microsomal metabolism was investigated with 4-toluidine, 2-chloro-4-to luidine, and 2,6-dichloro-4-toluidine as substrates. Microsomal metabo lic products were identified and quantified by HPLC, chemical assays, and synthesized reference compounds. Metabolites identified include pr oducts from aromatic ring hydroxylation, side-chain C-hydroxylation (b enzyl alcohols and benzaldehydes), N-hydroxylation (hydroxylamines, ni trosotoluenes, azoxy, azo, and hydrazo derivatives), and two new types of microsomal metabolites: secondary amines [ie. (halogenated) N-(4'- aminobenzyl)-4-toluidines] and an imine [ie. 5'-dichlorobenzylidene)-2 ,6-dichloro-4-toluidine]. The secondary amine appeared to be a major m icrosomal metabolite for all three 4-toluidines studied. Quantificatio n of the metabolite patterns demonstrated several influences of the ch lorine substituents on metabolism of the 4-toluidine derivatives. The total conversion increased significantly in the order: 4-toluidine < 2 -chloro-4-toluidine < 2,6-dichloro-4-toluidine, especially because of a marked increase in microsomal side-chain C-hydroxylation with increa sing number of ortho chlorine substituents. The rate of N-hydroxylatio n varied much less. Aromatic ring hydroxylation was observed to a sign ificant extent only for nonchlorinated 4-toluidine. Additional data fr om microsomal binding studies and molecular orbital calculations provi ded insight in possible mechanisms underlying the observed changes in metabolic profiles with increasing number of chlorine substituents at an ortho position with respect to the amine group.