Binding of 3-hydroxybenzo[a]pyrene to bovine hemoglobin and albumin.
Previous studies examined the bioavailability and first-pass biotransformation of 3-hydroxy[(3)H]benzo[a]pyrene ([(3)H]-3-OHBaP) in an isolated perfused catfish intestinal model. This work showed that 3-OHBaP, or a metabolite formed in intestine, bound covalently to blood protein. In this study, the blood adducts were characterized in vitro by incubating bovine ferric hemoglobin or albumin with [(3)H]-3OHBaP under various conditions. Incubation of 2 microM [(3)H]-3-OHBaP with hemoglobin for 1 h resulted in 7.49 pmol bound/mg protein, while albumin binding was 1.37 pmol/mg protein. Mild acid hydrolysis released only 5% of the radioactivity from 3-OHBaP-hemoglobin adducts. After gel filtration, the 3-OHBaP-hemoglobin adducts were examined by HPLC analysis. A single peak of radioactivity was detected at the same retention time as the heme component of hemoglobin. Unbound 3-OHBaP was oxidized to BaP-3,6-dione during incubation with ferric hemoglobin. Treatment of hemoglobin with ascorbic acid decreased the formation of hemoglobin adducts by 33%, while hydrogen peroxide treatment increased adduct formation by 44%. Incubation of [(3)H]-BaP-3-beta-D-glucuronide (BaP-3G) with hemoglobin and beta-glucuronidase resulted in greater binding to hemoglobin than incubation with [(3)H]-3-OHBaP alone. The hemoglobin adduct obtained from [(3)H]-BaP-3G also co-migrated with heme. These results indicate that an oxidative process is involved in formation of the heme adduct and that 3-OHBaP or BaP-3G might be a precursor of the bound metabolite.
Sugihara
Narumi
N
Faculty of Pharmacy and Pharmaceutical Sciences, Fukuyama University, Sanzou, Gakuen-cho, Fukuyama, Hiroshima, Japan.
James
Margaret O
MO
D000818:Animals; D001205:Ascorbic Acid / Q000737:chemistry / Q000378:metabolism; D001580:Benzopyrenes / Q000737:chemistry / Q000378:metabolism*; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D005966:Glucuronidase / Q000378:metabolism; D006454:Hemoglobins / Q000737:chemistry / Q000378:metabolism*; D006861:Hydrogen Peroxide / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D012709:Serum Albumin / Q000737:chemistry / Q000378:metabolism*; D013997:Time Factors; D014316:Tritium
D001580:Benzopyrenes; D006454:Hemoglobins; D012709:Serum Albumin; D014316:Tritium; C021338:3-hydroxybenzo(a)pyrene; D006861:Hydrogen Peroxide; D005966:Glucuronidase; D001205:Ascorbic Acid