Formation of DNA adducts by the food mutagen 2-amino-3,4,8-trimethyl-3h-imidazo [4,8-f]quinoxaline (4,8-dimeiqx) in vitro and in vivo. identification of a N-(2'-deoxyguanosin-8-yl)-4,8-dimeiqx adduct

The covalent binding of the mutagenic N²-hydroxy metaboilte of the food mutagen 2-amino-3,4,8-trimethyl-3H-lmldazo[4,5-f]quinoxaline (4,8-DiMeIQx) to 2'-deoxy-nudeosides and DNA was investigated in vitro and in vivo. N²-Hydroxy-4,8-DiMeIQx reacted to a small extent spontaneously with 2-deoxyguanosine. However, acetylatlon of N²-hydroxy-4,8-DiMeIqx with acetic anhydride to form the N²-acetoxy derivative prior to reaction with 2-deoxyguanosine resulted in much higher yield of adduct. N²-Acetoxy-4,8-DiMeIQx did not form adducts with 2'-deoxy- adenosine, 2'-deoxycytldlne or 2'-deoxythymldlne. The adduct formed between the N metabolite of 4,8- DiMeIQx and 2-deoxyguanosine was analysed by mass spectrometry and NMR spectroscopy and the structure of the adduct was shown to be N²-Acetoxy-4,8-DiMeIQx. N²-Acetoxy-4,8-DiMeIQx. reacted with calf thymus DNA and formed a covalently bound 4,8-DiMeIQx residue, which could not be removed by repeated precipita tions or solvent extractions. The 4,8-DiMeIQx-DNA was hydrolysed enzymatically with nuclease P1/acid phosphat ase and HPLC analysis showed that 70% of the bound mutagen was recovered as N²-Acetoxy-4,8-DiMeIQx. An additional minor adduct accounting for ∼15% of the bound mutagen showed UV spectral characteristics similar to N²-(2'-deoxyguanosin-8-yl)-4,8-DiMeIQx and is probably an undigested oligomer. ³²P-Postlabelling analysis of calf thymus DNA modilied with 4,8-DiMeIQx in vitro and liver DNA from rats dosed with 50 mg/kg 4,8-DiMeIQx showed a similar adduct pattern. In both samples N²-(2'- deoxyguanosln-8-yl)-4,8-DiMeIQx accounted for 60-70% of the bound mutagen. Thus, these results show that 4,8-DiMeIQx similar to other heterocyclic amines form adducts with C-8 of guanine both in vitro and in vivo via Its N²-OH metabolite.