Quantitative detection of N-7-(2-hydroxyethyl)guanine adducts in DNA usinghigh-performance liquid chromatography/electrospray ionization tandem massspectrometry

High-performance liquid chromatography (HPLC) was combined with electrospra y ionization tandem mass spectrometry (ESI-MS/MS) to develop a sensitive an d selective method for the quantitative measurement of N-7-(2-hydroxyethyl) guanine (N-7-HEG) adducts in DNA obtained from ethylene oxide-exposed biolo gical samples. Selected reaction monitoring (SRM) was used as the detection mode while the fragmentation product ion at m/z 152 generated from the pre cursor protonated N7-HEG (m/z 196) was monitored. The detection limits for N7-HEG were estimated by twofold serial dilution and determined to be 4 fmo l in neat standard solution and 16 fmol when a matrix effect is considered. When the mass spectrometer was operated in the selected ion monitoring mod e using only the first quadrupole (without MS/MS function), the detection l imits increased to 128 fmol and 1 pmol (when matrix effect is considered), respectively. A good linear correlation (R-2 = 0.999) was observed for sign al intensities obtained by injecting 16 fmol-33 pmol of N-7-HEG into the HP LC/ESI-MS/MS system. Hep G2 cells were incubated for 8 h with medium contai ning various concentrations of ethylene oxide (ranging from 0.05 to 5.0 mM) . A dose-response relationship was established, indicating that the adduct formation increases with the exposure level. The method shows potential, al though the detection limit needs to be lowered for practical applications, for use in monitoring N7-HEG formation in other biological systems. Copyrig ht (C) 2001 John Wiley & Sons, Ltd.