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

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...

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Published in:Chemical research in toxicology 2012-02, Vol.25 (2), p.391-399
Main Authors: Mutlu, Esra, Jeong, Yo-Chan, Collins, Leonard B, Ham, Amy-Joan L, Upton, Patricia B, Hatch, Gary, Winsett, Darrell, Evansky, Paul, Swenberg, James A
Format: Article
Language:English
Subjects:
Animals
Brain - metabolism
Chromatography, Liquid
DNA Adducts - analysis
DNA Adducts - metabolism
Guanine - analogs & derivatives
Guanine - analysis
Guanine - metabolism
Kidney - metabolism
Liver - metabolism
Lung - metabolism
Male
Rats
Rats, Sprague-Dawley
Spleen - metabolism
Tandem Mass Spectrometry
Testis - metabolism
Vinyl Chloride - pharmacokinetics
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Summary: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 [13C2]-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 [13C2]-7-OEG from the reaction of calf thymus DNA with [13C18]-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 106 guanine. The exogenous 7-OEG in liver DNA from adult rats exposed to 1100 ppm [13C2]-VC for 5 days was 104.0 ± 23.0 adducts per 106 guanine (n = 4), while concentrations in other tissues ranged from 1.0 to 39.0 adducts per 106 guanine (n = 4). Although endogenous concentrations of 7-OEG in tissues in weanling rats were similar to those of adult rats, exogenous [13C2]-7-OEG concentrations were higher in weanlings, averaging 300 adducts per 106 guanine in liver. Studies on the persistence of [13C2]-7-OEG in adult rats sacrificed 2, 4, and 8 weeks postexposure to [13C2]-VC demonstrated a half-life of 7-OEG of 4 days in both liver and lung.
ISSN:0893-228X
1520-5010
DOI:10.1021/tx200447w