Benzo[a]pyrene quinone (BPQ) is an important cytochrome P-450 (CYP) metabolite of polycyclic aromatic hydrocarbons (PAHs), which are ubiquitous environmental pollutants found in tobacco smoke and car exhaust. BPQ can enter a futile redox-cycle in the presence of NAD(P)H and amplify the production of reactive oxygen species (ROS) including H2O2 and superoxide through this process. We hypothesized that in the presence of transition metal ions such as Cu+, hydroxyl radicals would be generated from H2O2 through Fenton chemistry. The hydroxyl radicals would then initiate lipid peroxidation of polyunsaturated fatty acids (PUFAs). Lipid-hydroperoxides undergo homolytic decomposition to give rise to bifunctional electrophiles including malondialdehyde (MDA), 4-hydroxy-2-nonenal (4-HNE), 4-oxo-2-nonenal (4-ONE) and 4,5-epoxy-2(E)-decenal (4,5-EDE). These endogenous bifunctional electrophiles can modify DNA bases such as 2’-deoxyguanosine (dGuo) to form M1G-2’-deoxyribose, 1,N2-propano-dGuo, heptanone-etheno-dGuo and etheno-dGuo, respectively. Using a stable isotope dilution liquid chromatography (LC)/mass spectrometry (MS) method that we developed, we were able to quantify these adducts in the selected reaction-monitoring (SRM) mode. Our experiments demonstrated that the profiles of the four types of DNA adducts derived from non-enzymatic lipid peroxidation of arachidonic acid were very distinct for dGuo bases and cellular DNA. When dGuo bases were modified, the major adducts were etheno-dGuo and heptanone-etheno-dGuo, with none M1G-2’-deoxyribose and 1,N2-propano-dGuo detected. On the contrary, when the experiment was conducted with cellular DNA, the major adduct was 1,N2-propano-dGuo, followed by etheno-dGuo, trace amount of heptanone-etheno-dGuo and undetectable M1G-2’-deoxyribose. Introduction of the redox cycling of BPQ into the system promoted lipid peroxidation and elevated lipid hydroperoxide derived dGuo adducts at low to moderate concentrations of BPQ (1-10 μM) for reactions with both dGuo bases and cellular DNA; however, the profiles of the adducts generated remain unchanged. Adduct levels declined when a high concentration of BPQ (100 μM) was used, which suggested that further oxidation of the initially formed adducts had occurred. Etheno-dGuo is mutagenic in mammalian cells and so this study raises the possibility that mutations can result through lipid hydroperoxide-mediated damage as well as through direct BPQ adduct formation or oxidative DNA damage. Therefore, it will be important in the future to assess the relative importance of these three pathways of DNA damage. Supported by NIH CA 92537.

[Proc Amer Assoc Cancer Res, Volume 46, 2005]