Chromatographic Characterization of Hemoglobin Benzo[a]pyrene-7,8-diol-9,10-epoxide Adducts

The formation of hemoglobin-carcinogen adducts has been detectedin carcinogen-treated animals and in human populations. Althoughpolynuclear aromatic hydrocarbons are ubiquitous in the humanenvironment and DNA-aromatic hydrocarbon adducts have been detectedin human tissue, the occurrence of hemoglobin-polynuclear aromatichydrocarbon adducts in humans has not been thoroughly described.In this study we examined the effects of reaction conditionson the extent of in vitro reaction of human hemoglobin and (+)[³H]benzo[a]pyrene-7,8-dol-9,10-epoxide (anti)(BPDE), a metabolitethought to be largely responsible for the carcinogenic effectof benzo[a]pyrene. The chromatographic properties of the resultinghemoglobin-BPDE adducts were examined by conventional DEAE-celluloseion exchange liquid chromatography and by reversed phase highperformance liquid chromatography. Several adducts were formedwhich were chromatographically resolved from hemoglobin andfrom the individual globins. Some adducts were basic and someacidic relative to unaltered hemoglobin, suggesting adduct formationby reaction at carboxyl and basic nitrogen groups, respectively.Alteration of the ion - chromatographic properties of the adductsby an ionic sulfhydryl reagent, together with only a moderateeffect of pH on the extent of adduct formation, indicated thatthe adducts were not formed via reaction with the ß₉₃cysteine sulfhydryl group. The chromatographic techniques employedmay be applicable for the characterization and analysis of otherhemoglobin-carcinogen adducts.