Phenobarbital N-glucosylation by human liver microsomes
- 79 Downloads
Glucosylation of xenobiotics in mammals has been observed for a limited number of drugs. Generally, these glucoside conjugates are detected as urinary excretion products with limited information on their formation. An in vitro assay is described for measuring the formation of the phenobarbital N-glucoside diasteriomers ((5R)-PBG, (5S)-PBG) using human liver microsomes. Human livers (n=18) were screened for their ability to N-glucosylate PB. Cell viability, period of liver storage, prior drug exposure, serum bilirubin levels, age, sex and ethnicity did not appear to influence the specific activities associated with the formation of the PB N-glucosides. The average rate of formation for both PB N-glucoside was 1.42±1.04 (range 0.11–4.64) picomole/min/mg-protein with an (5S)-PBG/(5R)-PBG ratio of 6.75±1.34. The apparent kinetic constants, Km and Vmax, for PB N-glucosylation for eight of the livers ranged from 0.61–20.8 mM and 2.41–6.29 picomole/min/mg-protein, respectively. The apparent Vmax/Km ratio for PB exhibited a greater than 20 fold variation in the ability of the microsomes to form the PB N-glucosides. It would appear that the formation of these barbiturate N-glucoside conjugates in vitro are consistent with the amount of barbiturate N-glucosides formed and excreted in the urine in prior drug disposition studies.
KeywordsPhenobarbital metabolism N-Glucosylation liver microsomes humans
Unable to display preview. Download preview PDF.
- 5.Kalow W., Tang B.K., Kadar D., and Inaba T. (1978): Distinctive patterns of amobarbital metabolites in man. Clin. Pharmac. Ther., 24, 576–582.Google Scholar
- 6.Kalow W., Tang B.K., Kadar D., Endrenyi L., and Chan F.Y. (1979): A method of studying drug metabolism in populations: Racial differences in amobarbital metabolism. Clin. Pharmac. Ther., 26, 766–776.Google Scholar
- 7.Kalow W., Kadar D., and Inaba T., and Tang B.K. (1977): A case of deficiency of N-hydroxylation of amobarbital. Clin. Pharmac. Ther., 21, 530–535.Google Scholar
- 19.Radominska A., Little J., Pyrek, J.S., Drake R.R., Igari Y., Fournel-Gigleux S., Magdalou J., Burchell B., Elbein A.D., Siest G., and Lester R. (1993): A Novel UDP-Glc-Specific Glucosyltransferase Catalyzing The Biosynthesis of 6-O-Glucosides of Bile Acids in Human Liver Microsomes. J. Biol. Chem., 268, 15127–15135.PubMedGoogle Scholar
- 21.Drake R.R., Igari Y., Lester R., Elbein A.D., and Radominske A. (1992): Application of 5-azido-UDP-glucose and 5-azido-UDP-glucuronic acid Photoaffinity Probes for the Determination of the Active Site Orientation of Microsomal UDP-glucosyltransferases and UDP-glucuronosyltransferases. J. Biol. Chem., 267, 11360–11365.PubMedGoogle Scholar
- 24.Segel I.H. (1976): Biochemical Calculations. New York, John Wiley & Sons, p. 222.Google Scholar
- 25.Howell S.R., Hazelton G.A., and Klaassen C.A. (1986): Depletion of Hepatic UDP-Glucuronic Acid by Drugs that are Glucuronidated. J. Pharmac. Exp. Ther., 236, 610–614.Google Scholar
- 27.Clarke’s Isolation and Identification of Drugs In pharmaceuticals, Body Fluids and Post-mortem Materials, 2nd Edn., Moffat, A.C. Editor, The Pharmaceutical Press, London; 1986, pp. 883–884.Google Scholar
- 29.Neighbors S.M. and Soine W.H. (1995): Identification of Phenobarbital N-Glucuronides as Urinary Metabolites of Phenobarbital in Mice. Drug Metab. Disp., 23, 548–552.Google Scholar