Deconjugation of N-glucuronide conjugated metabolites with hydrazine hydrate--biomarkers for exposure to the food-borne carcinogen 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP).
The metabolism of the carcinogen 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) has been investigated in rabbit liver S9. Two phase I metabolites, N(2)-OH-PhIP and 4'-OH-PhIP were identified based on UV and mass spectra and co-elution with reference standards. Fortification of the incubation with UDGPA resulted in a complete glucuronidation of PhIP and N(2)-OH-PhIP, while 4'-OH-PhIP was only partly glucuronidated. Also, the PhIP metabolite 5-OH-PhIP was completely glucuronidated by rabbit liver S9, while 5-OH-PhIP was a poor substrate for CYP mediated hydroxylation. The glucuronic acid conjugates of PhIP metabolites were unsusceptible to treatment with beta-glucuronidase indicating that these are N-glucuronides. Treatment of the conjugates with hydrazine hydrate, however, resulted in complete hydrolysis of the glucuronic acid conjugates as well as in reduction to the parent amine of metabolites hydroxylated in the exocyclic amino group. Urine was collected from a male volunteer following consumption of fried chicken. Treatment of the urine with beta-glucuronidase/sulfatase resulted in release of 4'-OH-PhIP, while treatment with hydrazine hydrate in addition resulted in release of substantial amounts of PhIP and 5-OH-PhIP. The data show that hydrazine hydrate can hydrolyse N-glucuronides of metabolites of PhIP, glucuronides that are unsusceptible to enzymatic hydrolysis. In addition the data indicate that humans metabolise a large fraction of ingested PhIP to genotoxic metabolites. The chemical hydrolysis of glucuronide conjugates of PhIP metabolites with hydrazine hydrate observed in this study may also be a useful approach in the development of biomarkers for exposure and effect of other xenobiotics.
Frandsen
H
H
Institute of Food Safety, Danish Institute for Food and Veterinary Research, Mørkhøj Bygade 19, DK-2600 Søborg, Denmark. hf@dfvf.dk <hf@dfvf.dk>
D000818:Animals; D000970:Antineoplastic Agents / Q000378:metabolism; D015415:Biomarkers / Q000032:analysis*; D001711:Biotransformation; D002273:Carcinogens / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D005506:Food Contamination / Q000032:analysis; D005965:Glucuronates / Q000378:metabolism; D005966:Glucuronidase / Q000378:metabolism; D020719:Glucuronides / Q000378:metabolism*; D006801:Humans; D006834:Hydrazines / Q000378:metabolism; D007093:Imidazoles / Q000378:metabolism* / Q000652:urine; D008099:Liver / Q000378:metabolism*; D008297:Male; D013058:Mass Spectrometry / Q000379:methods; D011817:Rabbits; D013056:Spectrophotometry, Ultraviolet / Q000379:methods
D000970:Antineoplastic Agents; D015415:Biomarkers; D002273:Carcinogens; D005965:Glucuronates; D020719:Glucuronides; D006834:Hydrazines; D007093:Imidazoles; C029424:hydrazine; C049584:2-amino-1-methyl-6-phenylimidazo(4,5-b)pyridine; D005966:Glucuronidase