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HPLC-ESI+-MS/MS analysis of N7-guanine-N7-guanine DNA cross-links in tissues of mice exposed to 1,3-butadiene.
- Source :
-
Chemical research in toxicology [Chem Res Toxicol] 2007 May; Vol. 20 (5), pp. 839-47. Date of Electronic Publication: 2007 Apr 25. - Publication Year :
- 2007
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Abstract
- 1,3-butadiene (BD) is a major industrial chemical used in rubber and plastics production and is recognized as an animal and human carcinogen. Although the exact mechanism of BD-induced carcinogenesis is unknown, chemical reactions of epoxide metabolites of BD with DNA to form nucleobase adducts are likely to contribute to multistage carcinogenesis. Among BD-derived epoxy metabolites, 1,2:3,4-diepoxybutane (DEB) appears to be the most genotoxic and carcinogenic, probably because of its bifunctional nature. Initial DNA alkylation by DEB produces N7-(2'-hydroxy-3',4'-epoxybut-1'-yl)guanine monoadducts, which can then be hydrolyzed to N7-(2',3',4'-trihydroxy-1'-yl)guanine or can react with another site in double-stranded DNA to form 1,4-bis(guan-7-yl)-2,3-butanediol (bis-N7G-BD) cross-links. While (2',3',4'-trihydroxy-1'-yl)guanine lesions have been previously quantified in vivo, they cannot be used as a biomarker of DEB because the same lesions are also formed by another, more prevalent BD metabolite, 1,2-epoxy-3,4-butanediol. In contrast, bis-N7G-BD can only be formed from DEB, potentially providing a specific biomarker of DEB formation. We have developed a quantitative HPLC-ESI+-MS/MS method for measuring racemic and meso forms of bis-N7G-BD in DNA extracted from tissues of BD-exposed laboratory animals. In our approach, bis-N7G-BD adducts are released from DNA as free bases by neutral thermal hydrolysis, purified by solid-phase extraction, and subjected to HPLC-ESI+-MS/MS analysis. Selected reaction monitoring is performed by following the loss of a guanine moiety from protonated molecules of bis-N7G-BD and the formation of protonated guanine under collision-induced dissociation. Quantitative analysis of racemic and meso forms of bis-N7G-BD is based on isotope dilution with the corresponding 15N-labeled internal standards. The lower limit of quantification of our current method is 10-20 fmol/0.1 mg of DNA. The accuracy and precision of the new method were determined by spiking control mouse liver DNA with racemic and meso forms of bis-N7G-BD (10 fmol each), followed by sample processing and HPLC-ESI+-MS/MS analysis. Calculated amounts of racemic and meso forms of bis-N7G-BD were within 20% of the theoretical value (9.7 +/- 2 and 9.2 +/- 1.9 fmol, respectively, N = 4). DNA extracted from liver and lung tissues of mice exposed to 625 ppm butadiene for 5 days contained 3.2 +/- 0.4 and 1.8 +/- 0.5 racemic adducts per 10(6) guanines, respectively, while the amounts of meso-bis-N7G-BD were below the detection limits of our method (1 per 10(7) guanines). Control animals did not contain either bis-N7G-BD lesion. Sensitive and specific quantitative methods for bis-N7G-BD analysis developed in this work provide a unique biomarker of DEB-induced DNA alkylation following exposure to BD.
- Subjects :
- Animals
Butadienes chemistry
Butadienes metabolism
Carcinogens, Environmental chemistry
Carcinogens, Environmental metabolism
Cross-Linking Reagents chemistry
DNA Adducts chemistry
DNA Adducts metabolism
Guanine chemistry
Guanine metabolism
Liver chemistry
Liver drug effects
Liver metabolism
Lung chemistry
Lung drug effects
Lung metabolism
Mice
Reproducibility of Results
Butadienes toxicity
Carcinogens, Environmental toxicity
Chromatography, High Pressure Liquid
DNA Adducts drug effects
Guanine analogs & derivatives
Spectrometry, Mass, Electrospray Ionization methods
Subjects
Details
- Language :
- English
- ISSN :
- 0893-228X
- Volume :
- 20
- Issue :
- 5
- Database :
- MEDLINE
- Journal :
- Chemical research in toxicology
- Publication Type :
- Academic Journal
- Accession number :
- 17455958
- Full Text :
- https://doi.org/10.1021/tx700020q