Rapid measurement of 8-oxo-7,8-dihydro-2'-deoxyguanosine in human biological matrices using ultra-high-performance liquid chromatography-tandem mass spectrometry.
journal contributionposted on 24.10.2012, 08:55 by Patricia M. W. Lam, Vilas Mistry, Timothy H. Marczylo, Justin C. Konje, Mark D. Evans, Marcus S. Cooke
Interaction of reactive oxygen species with DNA results in a variety of modifications, including 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG), which has been extensively studied as a biomarker of oxidative stress. Oxidative stress is implicated in a number of pathophysiological processes relevant to obstetrics and gynecology; however, there is a lack of understanding as to the precise role of oxidative stress in these processes. We aimed to develop a rapid, validated assay for the accurate quantification of 8-oxodG in human urine using solid-phase extraction and ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) and then investigate the levels of 8-oxodG in several fluids of interest to obstetrics and gynecology. Using UHPLC-MS/MS, 8-oxodG eluted after 3.94 min with an RSD for 15 injections of 0.07%. The method was linear between 0.95 and 95 nmol/L with LOD and LOQ of 5 and 25 fmol on-column, respectively. Accuracy and precision were 98.7-101.0 and <10%, respectively, over three concentrations of 8-oxodG. Recovery from urine was 88% with intra- and interday variations of 4.0 and 10.2%, respectively. LOQ from urine was 0.9 pmol/ml. Rank order from the greatest to lowest 8-oxodG concentration was urine>seminal plasma>amniotic fluid>plasma>serum>peritoneal fluid, and it was not detected in saliva. Urine concentrations normalized to creatinine (n=15) ranged between 0.55 and 1.95 pmol/μmol creatinine. We describe, for the first time, 8-oxodG concentrations in human seminal plasma, peritoneal fluid, amniotic fluid, and breast milk, as well as in urine, plasma, and serum, using a rapid UHPLC-MS/MS method that will further facilitate biomonitoring of oxidative stress.