ANALYSIS OF 4-AMINOBIPHENYL-DNA ADDUCTS IN HUMAN URINARY-BLADDER AND LUNG BY ALKALINE-HYDROLYSIS AND NEGATIVE-ION GAS-CHROMATOGRAPHY MASS-SPECTROMETRY

Analysis of carcinogen-DNA adducts has been regarded as a useful means of assessing human exposure to chemical carcinogens. We have es?tabli shed a method for quantitation of 4-aminobiphenyl (4-ABP)-DNA adducts by alkaline hydrolysis and gas chromatography with negative ion chemic al ionization mass spectrometry (GC-NICI-MS). Aliquots of DNA (typical ly 100 mu g/ml) were spiked with an internal standard, d(g)-4-ABP, and were hydrolyzed in 0.05 N NaOH at 130 degrees C overnight. The libera ted 4-ABP was extracted with hexane and derivatized using pentafluorop ropionic anhydride in trimethylamine for 30 min at room temperature pr ior to GC-NICI-MS. With in vitro [H-3]N-hydroxy-4-ABP modified DNA sta ndards, we observed 59 +/- 7% (n = 9) recovery of the 4-ABP and a line ar correlation between hydrolyzed 4-ABP and the adduct levels ranging from about 1 in 10(8) to 1 in 10(4) nucleotides (r = 0.999, n = 9). Th e method was further validated by comparison of the results with that obtained by the P-32-postlabeling method. There was excellent agreemen t (r = 0.994, p<0.001) between the two methods for quantitation of the adduct in eight samples of Salmonella typhimurium DNA treated with 4- ABP and rat liver S9, although the P-32-postlabeling method gave sligh tly higher values. The DNA adducts in 11 human lung and 8 urinary blad der mucosa specimens were then determined by our GC-NICI-MS method. Th e adduct levels were found to be <0.32 to 49.5 adducts per 10(8) nucle otides in the lungs and <0.32 to 3.94 adducts per 10(8) nucleotides in the bladder samples. Our results indicate that the alkaline hydrolysi s/GC-NICI-MS method is sensitive, structure-selective, and accurate, a nd will be useful for molecular dosimetry of human exposure to this ca rcinogen.