Quaternary ammonium-linked glucuronidation of 1-substituted imidazoles: studies of human UDP-glucuronosyltransferases involved and substrate specificities.
A series of eight 1-substituted imidazoles was investigated as model substrates for glucuronidation at an aromatic tertiary amine of polyaza heterocyclic ring systems. The human UDP-glucuronosyltransferases (UGTs) involved and substrate specificities were investigated. Nine expressed enzymes (UGT1A1, UGT1A3, UGT1A4, UGT1A6, UGT1A7, UGT1A9, UGT1A10, UGT2B7, and UGT2B15) were examined, but only UGT1A4 catalyzed the formation of a quaternary ammonium-linked glucuronide metabolite for six of the substrates. UGT1A3 also catalyzed the glucuronidation of the previously investigated 1-phenylimidazole but none of the newly investigated compounds. No glucuronidation was observed with 1-(4-nitrophenyl)imidazole, the compound with the 4-phenyl substituent with the largest electron withdrawing effect. The incubation conditions for the determination of the kinetic constants for UGT1A4 catalysis of six substrates were optimized and included incubation at pH 7.4 with alamethicin at 10 microg/mg of protein. Latency disrupting agents, including alamethicin and sonication, enhanced glucuronidation 1.25-fold at most. There were 17.5- and 2.2-fold variations in the apparent K(m) (range, 0.18-3.15 mM) and V(max) values (range, 0.16-0.35 nmol/min/mg of protein). Linear correlation analyses between UGT1A4 kinetics and substrate physicochemical parameters showed significant correlation between V(max) and both the partition coefficient (log P, n-octanol/water) and pK(a) and between K(m) and pK(a), thereby indicating that the lipophilicity and the ease of availability of the tertiary amine lone pair of electrons of the substrate are important with respect to enzyme catalysis.
Vashishtha
S C
SC
Drug Metabolism and Drug Disposition Group, College of Pharmacy and Nutrition, University of Saskatchewan, Saskatoon, Canada.
Hawes
E M
EM
McKay
G
G
McCann
D J
DJ
D002384:Catalysis; D002851:Chromatography, High Pressure Liquid; D020719:Glucuronides / Q000378:metabolism; D014453:Glucuronosyltransferase / Q000378:metabolism*; D006801:Humans; D007093:Imidazoles / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D000644:Quaternary Ammonium Compounds / Q000378:metabolism; D013379:Substrate Specificity; D000099148:UGT1A1 Enzyme
D020719:Glucuronides; D014453:Glucuronosyltransferase; D007093:Imidazoles; D000644:Quaternary Ammonium Compounds; D000099148:UGT1A1 Enzyme