Glucuronidation of N-hydroxy heterocyclic amines by human and rat liver microsomes

Keith R. Kaderlik, Gerard J. Mulder, Robert J. Turesky, Nicholas P. Lang, Candee H. Teitel, M. Paul Chiarelli, Fred F. Kadlubar

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31 Scopus citations


The food-borne carcinogenic and mutagenic heterocyclic aromatic amines undergo bioactivation to the corresponding N-hydroxy (OH)-arylamines and the subsequent N-glucuronidation of these metabolites is regarded as an important detoxification reaction. In this study, the rates of glucuronidation for the N-OH derivatives of 2-amino-3-methylimidazo[4,5-f]-quinoline (IQ), 2-amino-l-methyl-6-phenylimidazo[4,5-b]- pyridine (PhIP), 2-amino-6-methyi-dipyrido[l,2-a:3',2'-d]imidazole (Glu-P-1) and 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MelQx) by liver microsomal glucuronosyltransferase were compared to that of the proximate human urinary bladder carcinogen, N-OH-aminobiphenyl (N-OH-ABP) and the proximate rat colon carcinogen N-OH-3,2'-dimethyl-4-aminobiphenyl (N-OH-DMABP). Human liver microsomes catalyzed the uridine 5'-diphosphoglucuronic acid (UDPGA)-dependent glucuronidation of N-OH-IQ, N-OH-PhIP, N-OH-Glu-P-1 and N-OH-MeIQx at rates of 59%, 42%, 35% and 27%, respectively, of that measured for N-OH-ABP (11.5 nmol/min/mg). Rat liver microsomes also catalyzed the UDPGA-dependent glucuronidation of N-OH-PhIP, N-OH-Glu-P-1 and N-OH-IQ at rates of 30%, 20% and 10%, respectively of that measured for N-OH-DMABP (11.2 nmol/min/mg); activity towards N-OH-MelQx was not detected. Two glucuronide(s) of N-OH-PhIP, designated I and II, were separated by HPLC. Conjugate II was found to be chromatographically and spectrally identical with a previously reported major biliary metabolite of PhlP in the rat, while conjugate I was identical with a major urinary metabolite of PhIP in the dog. Hepatic microsomes from rat, dog and human were found to catalyze the formation of both conjugates. The rat preferentially formed conjugate II (I to II ratio of 1:15), while the dog and human formed higher relative amounts of conjugate I (I to II ratio of 2.5:1.0 and 1.3:1.0 respectively). Fast atom bombardment mass spectrometry of conjugates I and II gave the corresponding molecular ions and showed nearly identical primary spectra. However, collision-induced spectra were distinct and were consistent with the identity of conjugates I and II as structural isomers. Moreover, the UV spectrum of conjugate I exhibited a λmax at 317 nm and was essentially identical to that of N-OH-PhIP, while conjugate II was markedly different with a λmax of 331 nm. Both conjugates were stable in 0.1 N HCI and were resistant to hydrolysis by rat, dog and human liver microsomal β-glucuronidases. In contrast, conjugate II, but not conjugate I, was readily cleaved by Esherichia coli β-glucuronidase to yield N-OH-PhIP. On comparison of these properties with those of the known N2- and N3-glucuronides of PhIP, conjugates I and II were judged to be N-OH-PhIP N2- glucuronide and N-OH-PhIP N3-glucuronide, respectively. These results indicate that both human and rat liver microsomes catalyze the glucuronidation of several N-OH heterocyclic amines and, in the case of N-OH-PhIP, two microsomal N-glucuronide metabolites were formed with distinct chemical and enzymatic properties.

Original languageEnglish (US)
Pages (from-to)1695-1701
Number of pages7
Issue number8
StatePublished - Aug 1994


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