Nontargeted Analysis of DNA Adducts by Mass-Tag MS: Reaction of <i>p</i>‑Benzoquinone with DNA

Using a method in which DNA adducts are discovered based on their conversion in a nucleotide form to phosphorimidazolides with isotopologue benzoylhistamines (or <i>p</i>-bromobenzoylhistamine) prior to detection by MALDI-TOF-MS, we have profiled the adducts that form when calf thymus DNA is reacted in vitro with <i>p</i>-benzoquinone (BQ). We find, as relative values normalized to 100% of adducts observed, 79% BQ-dCMP, 21% BQ-methyl-dCMP (a new DNA adduct), and trace amounts of BQ-dAMP and BQ-dGMP. Because mC is 5% of C in this DNA, the reaction of BQ with DNA in vitro is about five times faster at methyl-C than C. When equal amounts of dCMP and methyl-dCMP are reacted with BQ, equal amounts of the corresponding adducts are observed. Thus, the microenvironment of methyl-C in DNA enhances its reactivity relative to C with BQ. In a prior, similar study, but based on analysis by ³²P-postlabeling, the second most abundant adduct was assigned to BQ-A, apparently because of comigration of the BQ-A and BQ-methyl-C adducts (as bisphosphates) in the chromatographic step. Because the calf thymus DNA (used as received) was contaminated with RNA, we also detected the ribonucleotide adduct, BQ-CMP