title: Mass Spectrometric Characterization of an Acid-Labile Adduct Formed with 2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine and Albumin in Humans.
abstract: 2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) is a carcinogenic heterocyclic aromatic amine formed during the high-temperature cooking of meats. The cytochrome P450-mediated N-hydroxylation of the exocyclic amine group of PhIP produces 2-hydroxyamino-1-methyl-6-phenylimidazo[4,5-b]pyridine, an electrophilic metabolite that forms adducts with DNA and proteins. Previous studies conducted by our laboratory showed that the reaction of N-oxidized PhIP metabolites with human albumin in vitro primarily occurs at the Cys34residue, to produce an acid-labile linked sulfinamide adduct. On the basis of these findings, we developed a sensitive ultraperformance liquid chromatography-mass spectrometry method to measure acid-labile albumin-PhIP adducts in human volunteers administered a dietary-relevant dose of14C-labeled PhIP [Dingley, K. H., et al. (1999) Cancer Epidemiol., Biomarkers Prev. 8, 507-512]. Mild acid treatment of albumin (0.1 N HCl, 37 °C for 1 h) or proteolytic digestion with Pronase [50 mM ammonium bicarbonate buffer (pH 8.5) at 37 °C for 18 h] released similar amounts of covalently bound PhIP, which was characterized by multistage scanning and quantified by Orbitrap mass spectrometry. The amount of [14C]PhIP recovered by acid treatment of albumin 24 h following dosing accounted for 7.2-21.3% of the [14C]PhIP bound to albumin based on accelerator mass spectrometry measurements. 2-Amino-1-methyl-6-(5-hydroxy)phenylimidazo[4,5-b]pyridine, a hydrolysis product of the Cys34S-N linked sulfenamide adduct of PhIP, was not detected in either acid-treated or protease-treated samples. These findings suggest that a portion of the PhIP bound to albumin in vivo probably occurs as an acid-labile sulfinamide adduct formed at the Cys34residue.
authors.forename: Yi
authors.initials: Y
authors.lastname: Wang
authors.forename: Peter W
authors.initials: PW
authors.lastname: Villalta
authors.affiliation: School of Food Science and Engineering, Wuhan Polytechnic University , ChangQing Garden, Hankou, Wuhan 430023, P. R. China.
authors.forename: Lijuan
authors.initials: L
authors.lastname: Peng
authors.affiliation: Biosciences and Biotechnology Division, Center for Accelerator Mass Spectrometry, Lawrence Livermore National Laboratory , Livermore, California 94550, United States.
authors.forename: Karen
authors.initials: K
authors.lastname: Dingley
authors.affiliation: Biosciences and Biotechnology Division, Center for Accelerator Mass Spectrometry, Lawrence Livermore National Laboratory , Livermore, California 94550, United States.
authors.forename: Michael A
authors.initials: MA
authors.lastname: Malfatti
authors.affiliation: Biosciences and Biotechnology Division, Center for Accelerator Mass Spectrometry, Lawrence Livermore National Laboratory , Livermore, California 94550, United States.
authors.forename: K W
authors.initials: KW
authors.lastname: Turteltaub
authors.forename: Robert J
authors.identifier: 0000-0001-7355-9903
authors.initials: RJ
authors.lastname: Turesky
mesh_terms: D000418:Albumins / Q000378:metabolism*; D002138:Calibration; D002853:Chromatography, Liquid / Q000379:methods; D006801:Humans; D006868:Hydrolysis; D007093:Imidazoles / Q000378:metabolism*; D013058:Mass Spectrometry / Q000379:methods*
chemical_list: D000418:Albumins; D007093:Imidazoles; C049584:2-amino-1-methyl-6-phenylimidazo(4,5-b)pyridine