QUANTITATION OF 8-OXOGUANINE AND STRAND BREAKS PRODUCED BY 4 OXIDIZING-AGENTS

Reactive oxygen species, produced endogenously or by exposure to envir onmental chemicals and ionizing radiation, induce a wide range of DNA lesions. The variety of chemistries associated with different oxidants suggests that each will produce a unique spectrum of DNA damage produ cts. To extend our efforts to relate genotoxin chemistry to DNA damage , we measured both strand breaks and 8-oxoguanine (8-oxoG) in DNA afte r exposure to gamma-radiation, Fe(II)-EDTA/H2O2, Cu(II)H2O2, and perox ynitrite at concentrations approaching physiological relevance. We fou nd that the ratio of 8-oxoG to strand breaks varied more than 10-fold depending on the oxidizing agent: similar to 0.4 for Cu(II)H2O2 and pe roxynitrite and similar to 0.03 for Fe(II)-EDTA/H2O2 and gamma-radiati on. In the case of Cu(II)H2O2, the relative proportion of 8-oxoG and s trand breaks was found to vary more than 2-fold (0.14-0.37) for differ ent Cu(II) concentrations, consistent with other studies. We were able to detect 8-oxoG formation by peroxynitrite by using low peroxynitrit e concentrations in conjunction with a sensitive immunoaffinity/KPLC-E CD methodology. The level of 8-oxoG relative to strand breaks produced by peroxynitrite was higher than that produced by Fe(II)-EDTA/H2O2 an d gamma-radiation, which is consistent with the altered reactivity or accessibility of a non-hydroxyl radical species produced by peroxynitr ite.