Analysis of 7-methylbenz[a]anthracene-DNA adducts formed in SENCAR mouse epidermis by 32P-postlabeling

The present study has analysed the DNA adducts formed in SENCAR mouseepidermis following topical application of 7-methylbenz[a]anthracene (7-MBA). Mice were treated with 400 nmol of 7-MBA, which represents aninitiating dose of this hydrocarbon for SENCAR mice. DNA adducts wereanalysed 24 h after topical application of the hydrocarbon by 32P-postlabeling coupled with either HPLC analysis or an improved TLC proceduregiving better resolution of DNA adducts through the use of a D6 solvent[isopropanol:4N NH4OH (1:1)] following D5. Twenty-four hours after topicalapplication of 400 nmol 7-MBA, the level of total covalent binding was 0.37+/- 0.07 pmol/mg DNA as determined by 32P- postlabeling. This level ofbinding correlated well with the relative tumor initiating activity of thishydrocarbon compared to 7,12- dimethylbenz[a]anthracene (6.4 +/- 0.01pmol/mg DNA) and dibenz[a,j]anthracene (0.03 +/- 0.01 pmol/mg DNA).Analysis of the 32P- labeled 3',5'-diphosphodeoxyribonucleosides by HPLCand TLC revealed the presence of deoxyguanosine (dGuo) and deoxyadenosine(dAdo) adducts formed from both the anti- and syn-bay-region diol-epoxidesof 7-MBA (anti- and syn-7-MBADEs). The major DNA adduct derived from 7-MBAin mouse epidermis was tentatively identified as (+) anti-7-MBADE-trans-N2-dGuo. In addition, a minor dGuo adduct derived from the bay-region syn-diol-epoxide of 7-MBA was detected as well as a minor dAdo adduct from thisdiol-epoxide. Another minor dAdo adduct was also detectably present whicharose from either the anti- or syn-diol epoxide. Furthermore, severalunidentified DNA adducts were present in both HPLC and TLC chromatograms ofDNA samples from 7-MBA-treated mice. These results are discussed in termsof the role of specific 7-MBA-DNA adducts in tumor initiation by thishydrocarbon.