Detection of Extracellular 8-Oxo-7,8-dihydro-2 '-Deoxyguanosine As A Biomarker of Oxidative Damage in X-Irradiated Fibroblast Cultures: Optimization of Analytical Procedure

We have developed a simple methodology, based on single-step solid-phase extraction followed by isocratic high-performance liquid chromatography coupled with electrochemical detection (HPLC-ECD), to determine extracellular 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG) in culture supernatants of normal human dermal fibroblasts. A standard addition method, using externally added 8-oxodG (0.5 and 1 pmol) was employed to eliminate matrix effects arising from the chemically complex, protein-rich medium. Secondly, applying this procedure to X-ray irradiated fibroblasts, we report a significant twofold increase in the levels of 8-oxodG at the radiobiologically relevant dose of 6 Gy. This suggests that extracellular 8-oxodG might be a useful biomarker for oxidative stress following moderate doses of X-irradiation.更多