Formation of hepatic DNA adducts was studied in rats following intraperitoneal administration of a hepatocarcinogen, 3-methoxy-4-aminoazobenzene (3-MeO-AAB) and a non-hepatocarcinogen, 2-methoxy-4-aminoazobenzene (2-MeO-AAB). The 32P-post-labeling assay revealed 3-MeO-AAB to give more than 20-fold higher amounts of DNA adducts than did 2-MeO-AAB. Furthermore, five adducts, one of which accounted for over 70% of the total modified bases, were found in DNA from 3-MeO-AAB-treated rats, whereas only one adduct was apparent in 2-MeO-AAB-treated DNA. Our data thus suggested that the difference in hepatocarcinogenic activity between 3-MeO-AAB and 2-MeO-AAB might be, at least in part, dependent on quantitative and qualitative differences in their azo dye-DNA adduct formation in the rat liver.