DNA adduct formation in human hepatoma cells treated with 3-nitrobenzanthrone: analysis by the 32P-postlabeling method

Abstract

3-Nitrobenzanthrone (3-nitro-7 H-benz[ d, e]anthracen-7-one, 3-NBA) is a powerful mutagen and a suspected human carcinogen existing in diesel exhaust and airborne particulates. Recently, one of the major presumed metabolites of 3-NBA, 3-aminobenzanthrone (3-ABA), was detected in human urine samples. Here we analyzed DNA adducts formed in 3-NBA-exposed human hepatoma HepG2 cells by a 32P-postlabeling/thin layer chromatography (TLC) method and a 32P-postlabeling/polyacrylamide gel electrophoresis (PAGE) method. With HepG2 cells exposed to 3-NBA (0.36–36.4 μM) for 3 h, we obtained three spots or bands corresponding to adducted nucleotides. Two were assigned as 2-(2′-deoxyadenosin- N 6-yl)-3-aminobenzanthrone-3′-phosphate (dA3′p- N 6-C2-ABA) and 2-(2′-deoxyguanosin- N 2-yl)-3-aminobenzanthrone-3′-phosphate (dG3′p- N 2-C2-ABA), with identical mobilities to those of synthetic standards on PAGE analysis. The chemical structure of the substance corresponding to the other spot or band could not be identified. Quantitative analyses revealed that the major adduct was dA3′p- N 6-C2-ABA and its relative adduct labeling (RAL) value at 36.4 μM of 3-NBA was 200.8 ± 86.1/10 8 nucleotide.