Discrepancies in the measurement of UVC-induced 8-oxo-2 '-deoxyguanosine: Implications for the analysis of oxidative DNA damage

Ultraviolet (UV)light-induced indirect, oxidative damage to DNA has receive d increasing attention with respect to the mutagenic and carcinogenic effec ts of solar radiation. An oxidative lesion that has raised particular inter est because of its qualitative and quantitative importance is 8-oxo-2'-deox yguanosine. This deoxynucleoside lesion is most frequently measured by high performance liquid chromatography with electrochemical detection (HPLC-EC) following enzymatic hydrolysis of DNA or as the base equivalent, 8-oxoguan ine, by gas chromatography-mass spectrometry (GC-MS) following acid hydroly sis of DNA. We have noted a discrepancy in the literature whereby the level s of 8-oxo-2'-deoxyguanosine measured by HPLC-EC in WC-irradiated DNA are s ignificantly higher than when 8-oxoguanine is measured by GC-MS, By making use of the availability of both RPLC-IEC and stable-isotope dilution GC-RIS methodologies in our laboratory we have confirmed the discrepancy noted in the literature by parallel analysis of the same WC-irradiated calf thymus DNA samples. Furthermore, analysis of the WC-induced product by UV-visible spectrophotometry, voltammetry and its detection by a monoclonal antibody w hich recognises 8-oxo-2'-deoxyguanosine strongly suggests that the product is indeed 8-oxo-2'-deoxyguanosine. Partial explanation for this discrepancy could be an inordinate resistance of WC-irradiated DNA to formic acid hydr olysis. However, we cannot completely exclude the possibility that there is a formic acid-labile species which co-elutes with 8-oxo-2'-deoxyguanosine in enzymatically digested UVC-irradiated DNA. Whether this phenomenon is un ique to UV-irradiation damage or occurs with other systems that cause oxida tive damage to DNA awaits further investigation. Irrespective of the exact mechanism, there will be significant implications for the analysis of oxida tive DNA damage. (C) 1999 Academic Press.