title: METHODS FOR AROMATIC AND HETEROCYCLIC AMINE CARCINOGEN-DNA ADDUCT ANALYSIS BY LIQUID CHROMATOGRAPHY-TANDEM MASS SPECTROMETRY.
abstract: Xenobiotic-DNA adducts are used as biomarkers to assess the genotoxic effects of carcinogens. Rats were dosed with 4-aminobiphenyl (4-ABP), 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx), or 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP). DNA was isolated from the colons of vehicle and carcinogen-treated rats and digested using different nucleases and alkaline phosphatase. Deoxyribonucleoside adducts were quantified by capillary liquid chromatography-tandem mass spectrometry (LC-MS/MS) using isotope dilution methods with deuterated internal standards. Major adducts were those bound to the C8 position of deoxyguanosine. 3'- and 5'-Exonucleases were the most efficient nucleases at isolating dG-C8-ABP adducts. However, bulky adducts such as dG-C8-MeIQx and dG-C8-PhIP were better isolated using nuclease P1 rather than a combination of micrococcal nuclease and spleen phosphodiesterase. The use of DNase I enhanced the detection of all three adducts. We describe LC-MS/MS methods for DNA adduct detection and support the testing of different nucleases that increase DNA digestion efficiency and make available more DNA adducts for detection.
authors.affiliation: Department of Pharmacology & Toxicology and James Graham Brown Cancer Center, University of Louisville School of Medicine, Louisville, KY 40292.
authors.forename: Jason R
authors.initials: JR
authors.lastname: Neale
authors.forename: Ned B
authors.initials: NB
authors.lastname: Smith
authors.forename: William M
authors.initials: WM
authors.lastname: Pierce
authors.forename: David W
authors.initials: DW
authors.lastname: Hein