ELLAGIC ACID INHIBITS DNA-BINDING OF BENZO(A)PYRENE ACTIVATED BY DIFFERENT MODES

The possible mechanisms of inhibition of benzo(alpha)pyrene (BaP) bind ing to DNA in vivo and in vitro by ellagic acid (EA) were investigated . In the in vitro studies BaP was activated by cytochrome P-450-depend ent mixed function oxidase system, ascorbate-dependent lipid peroxidat ion, and peroxidation of linoleic acid. Reactions were performed in th e dark at 37 degrees C for 20 to 30 min in buffered aqueous solutions with 2mg DNA and 40 nmol tritiated BaP. The levels of BaP-DNA adducts were determined by assay of the radioactivity. BaP-DNA adduct levels r anged from 146 to 170 fmol/mg DNA in ascorbate and NADPH-dependent rea ctions and the level was 8.25+/-0.425 pmol/mg DNA in the hematin-media ted linoleic acid reaction. Addition of EA to the reaction mixtures re sulted in a significant inhibition in BaP-DNA adduct formation, depend ing upon the concentration of EA; e.g., 500 mu M EA resulted in 42 to 53% inhibition of binding in the three systems of carcinogen activatio n. Similarly, EA feeding to male NMRI mice at a concentration of 12 mu g/ml drinking water significantly decreased the levels of carcinogen- DNA adducts in the lungs. Reactions performed in vitro, in which DNA w as preincubated overnight at 4 degrees C with EA, revealed a decrease of 20% in adduct formation. MDA formation during the process of lipid peroxidation, stimulated by NADPH and ascorbate in the liver microsome s of mouse and by hematin in linoleic acid, was taken as an index for the free radical reactions. Addition of EA to in vitro systems inhibit ed the MDA formation with an IC50 (concentration for 50% inhibition) o f 480, 400, and 400 mu M for NADPH, ascorbate, and hematin-mediated fr ee radical reactions, respectively.