Identification of an acetaldehyde adduct in human liver DNA and quantitation as N²-ethyldeoxyguanosine

Acetaldehyde, an ubiquitous mutagen and carcinogen, could be involved in human cancer etiology. Because DNA adducts are important in carcinogenesis, we have used liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) to explore the presence in human liver DNA of the major acetaldehyde DNA adduct, N²-ethylidenedeoxyguanosine (1). DNA was isolated and enzymatically hydrolyzed in the presence of NaBH₃CN, which quantitatively converts adduct 1 to N²-ethyldeoxyguanosine (N²-ethyl-dGuo, 2). [¹⁵N₅]N²-Ethyl- dGuo was synthesized and used as an internal standard. Adduct 2 was enriched from the hydrolysate by solid phase extraction and analyzed by LC-ESI-MS/MS. Clear peaks were observed for adduct 2 in analyses of human liver DNA, calf thymus DNA, and rat liver DNA. These peaks were not observed, or were much smaller, when the NaBH₃CN step was omitted. When the DNA was subjected to neutral thermal hydrolysis prior to NaBH₃CN treatment, adduct 2 was not observed. Control experiments using [¹³C ₂] acetaldehyde demonstrated that adducts 1 and 2 were not formed as artifacts during DNA isolation and analysis. These results strongly indicate that adduct 1 is present in human liver DNA and demonstrate that it can be quantified as adduct 2. Levels of adduct 2 measured in 12 human liver samples were 534 ± 245 fmol/μmol dGuo (mean ± SD). The results of this study establish the presence of an acetaldehyde adduct in human liver DNA and suggest that it is a commonly occurring endogenous DNA adduct.