Binding of chlorinated benzidines to the rat hepatic aromatic hydrocarbon receptor

Abstract

The binding of benzidine, 3,3′-dichlorobenzidine (3,3′-Cl 2BZ), and the asymmetrically-substituted chlorinated benzidines 3,5-dichlorobenzidine (3,5-Cl 2BZ) and 3,5,3′-trichlorobenzidine (Cl 3BZ) to the rat hepatic cytosolic aromatic hydrocarbon (Ah) receptor was measured, in order to assess the mechanism of P-450I induction by 3,3′-Cl 2BZ. Cl 3BZ is the most mutagenic benzidine derivative in the Ames assay. Binding affinity to the Ah receptor protein was determined by displacement of labelled 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD) from the receptor, measured with the sucrose density gradient centrifugation technique. The rank order of affinities and the apparent inhibitor constants were: Cl 3BZ (4 μM) > 3,5-Cl 2BZ (8.4 μM) > 3,3′-Cl 2BZ (10 μM). Benzidine did not displace TCDD from the receptor protein. 4-Aminobiphenyl a structural link between the benzidine and biphenyl series competed weakly with TCDD. The 50% inhibition concentration was about 150 μM. The results are consistent with the hypothesis that the induction of P-450 enzymes by 3,3′-Cl 2BZ in vivo is mediated by the Ah receptor.