A New LC-MS/MS Method for the Quantification of Endogenous and Vinyl Chloride-Induced 7-(2-Oxoethyl)Guanine in Sprague-Dawley Rats

作者：Mutlu Esra; Jeong Yo Chan; Collins Leonard B; Ham Amy Joan L; Upton Patricia B; Hatch Gary; Winsett Darrell; Evansky Paul; Swenberg James A^*

来源：Chemical Research in Toxicology, 2012, 25(2): 391-399.

DOI：10.1021/tx200447w

Vinyl chloride (VC) is an industrial chemical that is known to be carcinogenic to animals and humans. VC primarily induces hepatic angiosarcomas following high exposures (>= 50 ppm). VC is also found in Superfund sites at ppb concentrations as a result of microbial metabolism of trichloroethylene and perchloroethylene. Here, we report a new sensitive LC-MS/MS method to analyze the major DNA adduct formed by VC, 7-(2-oxoethylguanine) (7-OEG). We used this method to analyze tissue DNA from both adult and weanling rats exposed to 1100 ppm [C-13(2)]-VC for 5 days. After neutral thermal hydrolysis, 7-OEG was derivatized with O-t-butyl hydroxylamine to an oxime adduct, followed by LC-MS/MS analysis. The limit of detection was 1 fmol, and the limit of quantitation was 1.5 fmol on the column. The use of stable isotope VC allowed us to demonstrate for the first time that endogenous 7-OEG was present in tissue DNA. We hypothesized that endogenous 7-OEG was formed from lipid peroxidation and demonstrated the formation of [C-13(2)]-7-OEG from the reaction of calf thymus DNA with [C-13(18)]-ethyl linoleate (EtLa) under peroxidizing conditions. The concentrations of endogenous 7-OEG in liver, lung, kidney, spleen, testis, and brain DNA from adult and weanling rats typically ranged from 1.0 to 10.0 adducts per 10(6) guanine. The exogenous 7-OEG in liver DNA from adult rats exposed to 1100 ppm [C-13(2)]-VC for 5 days was 104.0 +/- 23.0 adducts per 10(6) guanine (n = 4), while concentrations in other tissues ranged from 1.0 to 39.0 adducts per 10(6) guanine (n = 4). Although endogenous concentrations of 7-OEG in tissues in weanling rats were similar to those of adult rats, exogenous [C-13(2)]-7-OEG concentrations were higher in weanlings, averaging 300 adducts per 10(6) guanine in liver. Studies on the persistence of [C-13(2)]-7-OEG in adult rats sacrificed 2, 4, and 8 weeks postexposure to [C-13(2)]-VC demonstrated a half-life of 7-OEG of 4 days in both liver and lung.

• 出版日期2012-2