Abstract: An ultra high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method for the analysis of biomarker 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-OHdG) in deoxyribonucleic acid (DNA) was developed. The artificial oxidation of 2′-deoxyguanosine (dG) at numerous stages in the sample preparation bring a challenge to the accurate measurement of 8-OHdG in DNA. To avoid the artificial oxidation during the enzymatic digestion, desferrioxamine mesylate as a protectant was added into the mixtures. By utilizing YM-3 Centricon membrane (3000 of relative molecular mass cut off), excess proteins in enzymatic solutions were effectively removed, allowing direct UPLC-MS/MS analysis. The UPLC-MS/MS analysis showed a linear relationship between the peak areas and the amounts of 8-OHdG in the range of 17.6~1400 fmol, and the correlation coefficient was 0.9998. By using the developed method, the content of 8-OHdG in calf thymus DNA (CT DNA) was estimated about 12.9±2.35 (calculated as 8-OHdG/106 dG), which was consistent with the previous work. This method can also be applicable for the detection of 8-OHdG in DNA under various oxidative stresses.

Key words: 2`-deoxyguanosine (dGuo), 8-oxo-7,8-dihydro-2`-deoxyguanosine (8-oxodGuo), calf thymus DNA (CT DNA), ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS)