Browsing by Subject "Nanospray MS"

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    Formation, Persistence, and Repair of 1,2,3,4-Diepoxybutane-Induced bifunctional DNA adducts in tissues of rodents exposed to 1,3-Butadiene by Inhalation.
    (2009-09) Goggin, Melissa Mary
    1, 3-butadiene (BD) is an industrial chemical and environmental pollutant. It is also classified as a probable human carcinogen based on animal tumorigenesis studies and human epidemiologic evidence. BD is metabolized to three reactive epoxides, of which 1,2,3,4-diepoxybutane (DEB) is the most mutagenic, likely due to its bifunctional electrophilic structure that enables it to form DNA-DNA cross-links, 1,4-bis-(guan-7- yl)-2,3-butanediol (bis-N7G-BD) and 1-(guan-7-yl)-4-(aden-1-yl)-2,3-butanediol (N7GN1A- BD), and 1,N6-(2-hydroxy-3-hydroxymethyl-1,3-propanodiyl)-dA (1,N6-HMHPdA) exocyclic DNA adducts. The purpose of this research is to identify mechanismbased biomarkers of exposure to BD. Our laboratory has identified DNA-DNA crosslinks and exocyclic deoxyadenosine adducts formed from DEB that could potentially be used as biomarkers of exposure. Highly specific and sensitive isotope dilution HPLCESI+- MS/MS methods have been developed to analyze DEB-induced DNA-DNA crosslinks and exocyclic DEB-dA adducts in DNA extracted from laboratory animals exposed to BD by inhalation. bis-N7G-BD was the most abundant adduct, N7G-N1ABD was ~10 times less abundant than bis-N7G-BD, and 1,N6-HMHP-dA was the least abundant (~4 fold lower than N7G-N1A-BD). The quantitative isotope dilution mass spectrometry methods developed in this work were employed to investigate the dose dependent formation, persistence, and repair of the BD-induced DNA adducts in laboratory rodents. Species, gender, and tissue differences in adduct levels were observed. Mouse DNA contained a greater number of DEB-specific adducts than rat DNA at identical exposures. Adduct levels were also higher in female as compared to male rodents, and in liver DNA (compared to lung, brain, kidney, and thymus). Although bis-N7G-BD was most abundant DEB-DNA adduct immediately following exposure, it did not persist in mouse or rat liver while the other adducts did. We did not observe repair of the bifunctional lesions by BER or NER, however, in vitro studies suggest that 1,N6-HMHP-dA is repaired by AlkB. The research presented in this thesis is consistent with animal inhalation studies where mice were more susceptible to tumor formation. The data also suggests that these differences are due to species differences in the extent of BD metabolism to DEB. Two of the adducts, N7G-N1A-BD and 1,N6- HMHP-dA were persistent in DNA and may be responsible for the mutagenicity of BD.