Ultrasensitive Simultaneous Quantification of 1,N2-Etheno-2′-deoxyguanosine and 1,N2-Propano-2′-deoxyguanosine in DNA by an Online Liquid Chromatography-Electrospray Tandem Mass Spectrometry Assay

Exocyclic DNA adducts produced by exogenous and endogenous compounds are emerging as potential tools to study a variety of human diseases and air pollution exposure. A highly sensitive method involving online reverse-phase high performance liquid chromatography with electrospray tandem mass spectrometry detection in the multiple reaction monitoring mode and employing stable isotope-labeled internal standards was developed for the simultaneous quantification of 1,N-2-etheno-2'-deoxyguanosine (1,N-2-epsilon dGuo) and 1,N-2-propano-2'-deoxyguanosine (1,N-2-propanodGuo) in DNA. This methodology permits direct online quantification of 2'-deoxyguanosine and ca. 500 amol of adducts in 100 mu g of hydrolyzed DNA M the same analysis. Using the newly developed technique, accurate determinations of 1,N-2-etheno-2'-deoxyguanosine and 1,N2-propano-2'-deoxyguanosine levels in DNA extracts of human cultured cells (4.01 +/- 0.32 1,N-2-epsilon dGuo/10(8) dGuo and 3.43 +/- 0.33 1,N-2-propanodGuo/10(8) dGuo) and rat tissue (liver, 2.47 +/- 0.61 1,N-2-epsilon dGuo/10(8) dGuo and 4.61 +/- 0.69 1,N-2-propanodGuo/108 dGuo; brain, 2.96 +/- 1.43,N-2-epsilon dGuo/10(8) dGuo and 5.66 +/- 3.70 1,N-2-propanoclGuo/10(8) dGuo; and lung, 0,87 +/- 0.34 1,N-2-edGuo/ 10(8) dGuo and 2.25 +/- 1.72 1,N-2-propanodGuo/10(8) dGuo) were performed. The method described herein can be used to study the biological significance of exocyclic DNA adducts through the quantification of different adducts in humans and experimental an with pathological conditions and after air pollution exposure.