A NOVEL HPLC PROCEDURE FOR THE ANALYSIS OF 8-OXOGUANINE IN DNA

The chromatographic quantitation of 8-oxoguanine adducts in DNA is wid espread in the literature, although results obtained by HPLC of 8-oxod eoxyguanosine do not always agree with levels determined by GC-MS. To help explain this discrepancy, here we describe a novel procedure for the analysis of 8-oxoguanine adducts in DNA. Although it proved diffic ult to directly quantitate 8-oxoguanine in the presence of high levels of endogenous guanine using conventional reversed-phase HPLC, a simpl e preincubation of DNA acid hydrolysates with guanase allowed such ana lyses. The assay relied on our observation that 8-oxoguanine was not a substrate for guanase, and on sensitive electrochemical detection. Th e limit of detection for 8-oxoguanine was 5 nM or 250 fmol on column. Using this procedure, the background level of 8-oxoguanine in commerci ally available calf thymus DNA was 0.4 nmol/mg DNA or 3.2 mol/10(5) mo l guanine.