Quantitating exposure to chemical carcinogens: in vivo alkylation of hemoglobin by benzo[a]pyrene

Mild acid hydrolysis of globin preparations from erythrocytes of mice, previously exposed topically to benzo[a]pyrene (BaP), releases tetrols which are detectable by HPLC/fluorescence analysis. If the mouse is exposed to radiolabelled BaP, radioactivity can be found in the acid-releasable tetrols. Treatment of the globin preparations prior to acid hydrolysis with proteolytic enzymes, but not enzymes that degrade nucleic acids, followed by dialysis, reduces the amount of tetrols that can be detected. Because the procedure used for the isolation of globin preparations from mouse blood precludes the presence of non-covalently bound BaP or its cellular metabolites, it is concluded that prior to acid hydrolysis, the tetrols were covalently attached to the hemoglobin, most probably as a result of the metabolic conversion of the applied carcinogen to the chemically reactive anti-diol epoxide. There is a dose response relationship between the amount of BaP applied to the skin of the mouse and the occurrence, 24 h later, of BaP adducts to hemoglobin, while the adduct, once formed, disappears with a half-life of 6 days. The amount of anti-benzo[a]pyrene diol epoxide (anti-BaPDE) binding to DNA and hemoglobin at various doses of BaP appears to be qualitatively similar.