Formation of 2-amino-alpha-carbolines in pan-fried poultry and P-32-postlabeling analysis of DNA adducts

Amino-alpha-carbolines are food-derived genotoxic heterocyclic aromatic ami nes (HAAs) that have been shown to be carcinogenic in rodents, to induce pr eneoplastic foci in rat liver and to be active in a number of genotoxicity tests, inducing gene mutations, DNA strand breaks and cell transformation. 2-Amino-alpha-carboline (A alpha C) has been estimated to account for about 20% of the carcinogenic HAAs that are ingested with the diet. Using the ta ndem cartridge solid-phase extraction procedure we were able to show that i n pan-fried poultry the proportion of A alpha C and its 3-methyl derivative (MeA alpha C) account for 15% of the HAA fraction. Covalent DNA adducts ar e considered to be the primary lesions in chemical carcinogenesis. Both A a lpha C and MeA alpha C have been shown to induce covalent DNA adducts in vi tro and in vivo. Formation of DNA adducts was investigated by P-32-postlabe lling analysis in vitro in primary rat hepatocytes treated with 10-500 mu M A alpha C or MeA alpha C. While for MeAnC-derived adducts the highest resp onse was obtained with the nuclease P1 enhancement procedure, AaC adducts w ere most efficiently detected upon P-32-postlabelling preceded by a cartrid ge enrichment step. Adduct levels detected were dose dependent but decrease d for the higher doses due to cytotoxic effects. We have characterised the major DNA adducts of A alpha C and MeA alpha C as deoxyguanosine derivative s with the amino-alpha-carboline bound via the 2-amino group to the C8 posi tion of guanine.