Determination of the Oxidative Damage Markers of Nucleic Acids 8-Hydroxyguanosine and 8-Hydroxy-2'-Deoxyguanosine in Urine Using High-Performance Liquid Chromatography with Tandem Mass-Spectrometric Detection

Abstract

Products of oxidative damage to nucleic acids are considered as relatively stable biomarkers in the diagnosis of negative consequences of oxidative stress. The concentrations of biomarkers of oxidative degradation of DNA (8-hydroxy-2'-deoxyguanosine, 8-OHdG) and RNA (8-hydroxyguanosine, 8-OHG) in biofluids increased when the body was exposed to toxic compounds, radiation, and other negative factors associated with oxidative stress. Urine is considered as a priority matrix for biomonitoring the consequences of oxidative stress because of noninvasive sampling and higher concentrations of the target analytes. We developed a procedure for the combined determination of 8-OHdG and 8-OHG in urine using HPLC–MS/MS analysis. A structurally similar exogenous compound, 8-[(1-hydroxybutan-2-yl)amino]-1,3,7-trimethyl-1-purine-2,6(3H,7H)-dione, was selected as an internal standard. The measurement range from 1 to 50 ng/mL was set for both analytes. A solid-phase extraction procedure on a hydrophilic–lipophilic balance (HLB) sorbent in the analyte retention mode was optimized to prepare biological samples for analysis. With the use of high-resolution HPLC–MS/MS technique, the error of analysis did not exceed 25% over the entire analytical range. A total of 130 urine samples of chemical plant workers aged 20 to 70 years without diagnosed systemic diseases were analyzed. The 8-OHdG and 8-OHG contents of the urine samples ranged from 1 to 20 and from 2 to 12 ng/mL, respectively. The dependence of the concentrations of both biomarkers in urine on the age of the workers was established.