title: Rapid and sensitive quantification of urinary N7-methylguanine by isotope-dilution liquid chromatography/electrospray ionization tandem mass spectrometry with on-line solid-phase extraction.
abstract: A rapid, highly specific and sensitive isotope-dilution liquid chromatography/tandem mass spectrometry (LC/MS/MS) method coupled with an on-line solid-phase extraction (SPE) system was developed to measure N7-methylguanine (N7-MeG) in urine. 15N5-Labeled N7-MeG was synthesized to serve as an internal standard, and an on-line SPE cartridge was used for on-line sample cleanup and enrichment. The urine sample can be directly analyzed within 15 min without prior sample purification. The detection limit for this method was estimated as 8.0 pg/mL (4.8 pmol) on-column. This method was further applied to study exposure to methylating agents arising from cigarette smoke. Sixty-seven volunteers were recruited, including 32 regular smokers and 35 nonsmokers. Urinary cotinine, a major metabolite of nicotine, was also determined using an isotope-dilution LC/MS/MS method. The results showed that urinary levels of N7-MeG observed in smokers (4215 +/- 1739 ng/mg creatinine) were significantly (P < 0.01) higher than those in nonsmokers (3035 +/- 720 ng/mg creatinine). It was further noted that the urinary level of N7-MeG was found to be correlated with that of cotinine for smokers, implying that cigarette smoking resulted in increased DNA methylation, followed by depurination and excretion of N7-MeG in urine. As a result of the on-line extraction system, this method is capable of routine high-throughput analysis and accurate quantitation of N7-MeG, and could be a useful tool for health surveillance of methylating agent exposure.
authors.affiliation: Department of Occupational Safety and Health, Chung Shan Medical University, Taichung, Taiwan.
authors.forename: Mu-Rong
authors.initials: MR
authors.lastname: Chao
authors.forename: Chien-Jen
authors.initials: CJ
authors.lastname: Wang
authors.forename: Hsi-Hsien
authors.initials: HH
authors.lastname: Yang
authors.forename: Louis W
authors.initials: LW
authors.lastname: Chang
authors.forename: Chiung-Wen
authors.initials: CW
authors.lastname: Hu
mesh_terms: D000328:Adult; D002853:Chromatography, Liquid / Q000379:methods*; D003367:Cotinine / Q000652:urine; D006147:Guanine / Q000031:analogs & derivatives* / Q000652:urine; D006801:Humans; D008297:Male; D008875:Middle Aged; D012015:Reference Standards; D012680:Sensitivity and Specificity; D012907:Smoking; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D013997:Time Factors
chemical_list: D006147:Guanine; C008450:7-methylguanine; D003367:Cotinine