Journal Article

Detection of 1,<i>N</i><sup>6</sup>-ethenodeoxyadenosine and 3,<i>N</i><sup>4</sup>-ethenodeoxycytidine by immunoaffinity/<sup>32</sup>P-post-labelling in liver and lung DNA of mice treated with ethylcarbamate (urethane) or its metabolites

R.Charles Fernando, Jegadeesan Nair, Alain Barbin, James A. Miller and Helmut Bartsch

in Carcinogenesis

Volume 17, issue 8, pages 1711-1718
Published in print August 1996 | ISSN: 0143-3334
e-ISSN: 1460-2180 | DOI:
Detection of 1,N6-ethenodeoxyadenosine and 3,N4-ethenodeoxycytidine by immunoaffinity/32P-post-labelling in liver and lung DNA of mice treated with ethylcarbamate (urethane) or its metabolites

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The capacity of the chemical carcinogen ethyl carbamate (EC, urethane) and its metabolites vinyl carbamate (VC) and vinyl carbamate epoxide (VCO) to form ethenobases was studied in liver and lung DNA of 12-day-old and adult CD-1, B6C3F1, C3H/HeJ and C57BL/6J mice. Following single and multiple doses of EC, VC or VCO, the formation of 1,N6-ethenodeoxyadenosine (ɛdA) and 3,N4-ethenodeoxycytidine (ɛdC) was quantified by an immunoaffinity chromatography/32P-postlabelling technique. Both etheno adducts were detected in untreated control DNA samples from liver and lung in the range of 2–15 adducts/109 parent nucleotides. Following five repeated injections of 250 or 280 nmol/g body wt VC to adult mice, 51, 57 and 78 ɛdA/109 dA and 28, 42 and 42 ɛdC/109 dC (means of duplicate analyses) were detected in liver DNA of CD-1, C3H/HeJ and C57BL/6J mice respectively. In lung DNA of these VC-treated mice, the levels were 87, 49 and 58 (ɛdA/109 dA) and 64, 39 and 43 (ɛdC/109 dC) respectively. Under similar dose regimens, lower levels of etheno adducts were detected in B6C3F1 mice. Etheno-DNA adducts were also formed in liver and lung upon treatment with EC in adult mice, but at 3-fold lower levels as compared with VC. In 12-day-old C3H/HeJ and C57BL/6J mice, 2- to 3-fold higher etheno adduct levels were detected in liver DNA, when compared with adults, upon a single treatment with 250 nmol/g body wt VC, suggesting that young animals are more susceptible to adduct formation. Combined analysis of adduct formation in adult CD-1, C3H/HeJ and C57BL/6J mice at the higher dose showed a statistically significant increase in etheno adduct formation in the order EC > VC. The results demonstrate that EC and its activated intermediates bind to liver and lung DNA to form ɛdA and ɛdC, and the differences in DNA binding further support the hypothesis that metabolic activation of EC to VC is involved. Preliminary data also suggest that background levels of ɛdA and ɛdC in DNA are affected by the type of diet given to the animals.

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Subjects: Clinical Cytogenetics and Molecular Genetics

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