Sulfate esters of hydroxymethyl-methyl-benz[a]anthracenes as active metabolites of 7,12-dimethylbenz[a]anthracene.

7-Hydroxymethyl-12-methylbenz[a]anthracene (7-HMBA) and 12-hydroxymethyl-7-methylbenz[a]anthracene (12-HMBA), carcinogenic major metabolites of 7,12-dimethylbenz[a]anthracene (DMBA) in untreated rat liver, showed high mutagenic activities toward Salmonella typhimurium TA 98 after preincubation with a sulfotransferase-PAPS system consisting of ATP, sodium sulfate, and a post-mitochondrial fraction (S-9) or a soluble supernatant fraction (S-105) from untreated rat liver. The 7- and 12-HMBAs themselves induced His+ mutation in TA 98 only slightly after preincubation with S-9 in the presence of an NADPH-generating system. Mutagenicity of DMBA toward TA 98 after preincubation with S-9 in the presence of the NADPH-generating system was remarkably enhanced by the addition of ATP and sodium sulfate. The active metabolites, 7-HMBA sulfate and 12-HMBA sulfate, were isolated from these preincubation systems and identified by comparison with the corresponding synthetic specimens. The sulfuric acid ester conjugates were potent mutagens toward TA 98 in the absence of rat liver subcellular fractions. The conjugates bound covalently at significant rates to calf-thymus DNA as well as to S-105 proteins at 37 degrees and pH 7.4 through the 7- or 12-methylene carbon with concomitant loss of their sulfate group. In the presence of S-105, glutathione inhibited the mutagenicity of the metabolically formed or exogenously added 7- and 12-HMBA sulfates. The non-mutagenic glutathione conjugates were isolated from the incubation mixtures and identified as S-(12-methylbenz[a]anthracen-7-yl)methylglutathione from 7-HMBA or its sulfate and S-(7-methylbenz[a]anthracen-12-yl)methylglutathione from 12-HMBA or its sulfate.