Ozone-Induced DNA Damage: A Pandora's Box of Oxidatively Modified DNA Bases

Ozone is a major component of air pollution and carries potentially mutagenic and harmful affects to health. The oxidation of isolated calf thymus DNA (CT-DNA) led to the nearly quantitative loss of normal DNA 2'-deoxyribonucleosides in the following order: T > G > C ≫ A. The major modification of pyrimidines (T, C, and 5-methylcytosine (5mC)) was the corresponding 5-hydroxyhydantoin derivative after complete digestion of DNA to its component 2'-deoxyribonucleosides. The oxidation of 5mC was 2.5-fold more susceptible than C considering the relative mole fraction of 5mC to C in CT-DNA. Other common oxidation products of pyrimidines (e.g., 5,6-dihydroxy-5,6-dihydropyrimidines, the so-called pyrimidine 5,6-glycols) were formed with a lower yield than 5-hydroxyhydantoin derivatives. In addition, several common oxidation products of G were observed (e.g., 8-oxo-7,8-dihydroguanine (8oxoG)) albeit with relatively minor yields. The sum of individual products was notably less than the loss of 2'-deoxyribonucleosides from which they were derived. In a search for additional products, we discovered the formation of pyrimidine ring fragments, predominantly N-formamide and N-urea, which were measured as a dinucleotide next to a nonmodified nucleotide upon partial digestion of oxidized DNA. Interestingly, the latter fragments were also observed in dinucleotides containing 8oxoG, indicating the formation of tandem lesions during ozonolysis of DNA. The oxidation of DNA upon exposure to ozone can be explained by reactions of an intermediate ozonide. These studies underline the complexity of ozone-induced DNA damage and provide valuable information to assess the formation of this damage in cellular DNA.