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 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 [(13)C(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 [(13)C(2)]-7-OEG from the reaction of calf thymus DNA with [(13)C(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 [(13)C(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 [(13)C(2)]-7-OEG concentrations were higher in weanlings, averaging 300 adducts per 10(6) guanine in liver. Studies on the persistence of [(13)C(2)]-7-OEG in adult rats sacrificed 2, 4, and 8 weeks postexposure to [(13)C(2)]-VC demonstrated a half-life of 7-OEG of 4 days in both liver and lung.
Department of Environmental Sciences and Engineering, University of North Carolina at Chapel Hill , Chapel Hill, North Carolina 27599, United States.
Esra
E
Mutlu
Yo-Chan
YC
Jeong
Leonard B
LB
Collins
Amy-Joan L
AJ
Ham
Patricia B
PB
Upton
Gary
G
Hatch
Darrell
D
Winsett
Paul
P
Evansky
James A
JA
Swenberg
D000818:Animals; D001921:Brain / Q000378:metabolism; D002853:Chromatography, Liquid; D018736:DNA Adducts / Q000032:analysis* / Q000378:metabolism; D006147:Guanine / Q000031:analogs & derivatives* / Q000032:analysis / Q000378:metabolism; D007668:Kidney / Q000378:metabolism; D008099:Liver / Q000378:metabolism; D008168:Lung / Q000378:metabolism; D008297:Male; D051381:Rats; D017207:Rats, Sprague-Dawley; D013154:Spleen / Q000378:metabolism; D053719:Tandem Mass Spectrometry; D013737:Testis / Q000378:metabolism; D014752:Vinyl Chloride / Q000493:pharmacokinetics
D018736:DNA Adducts; D006147:Guanine; C032220:7-N-(2-oxoethyl)guanine; D014752:Vinyl Chloride