32POSTLABELLING ANALYSIS OF URINARY MUTAGENS FROM SMOKERS OF BLACK TOBACCO IMPLICATES 2-AMINO-1-METHYL-6-PHENYLIMIDAZO[4,5-B]PYRIDINE (PHIP) AS A MAJOR DNA-DAMAGING AGENT

When mutagens extracted from the urine of two smokers of black tobacco were reacted with DNA in vitro in the presence of a metabolic activation system, several DNA adducts were detected by P-32-postlabelling analysis. Some of these adducts were also visible, but only faintly, on the autoradiogram for a non-smoker's urine. DNA adducts produced in vitro by 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline or 2-amino-1-methyl-6-phenylimidazo[3,5-b]pyridine could not account for the adduct pattern produced by the urinary mutagens. However, three or four 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP)-related DNA adducts were present among the five or six adducts observed for smokers in the autoradiograms of urinary mutagen-adducted nucleotides. Mutagenicity testing combined with HPLC fractionation of urinary extracts also supported the postlabelling data which implicates PhIP as a mutagen in the urine of smokers of black tobacco.