Determination of hepatic uridine 5′-diphosphoglucuronic acid concentration by conjugation with diethylstilbestrol

J. B. Watkins, C. D. Klaassen

Research output: Contribution to journalArticle

34 Citations (Scopus)

Abstract

A sensitive and reliable assay for uridine 5′-diphosphoglucuronic acid (UDPGA) was developed that involved conjugation of diethylstilbestrol (DES) in vitro. This conjugation reaction is solely dependent upon UDPGA concentration. The assay uses 0.13 M Tris-HCl, pH 7.4, 6.7 mM MgCl2, 0.05% Brig 58, 0.25 mg guinea pig liver microsomal protein, 0.13 mM 3H-DES (0.2 μCi/ml), and 200 μl of boiled 10% liver homogenate in a total volume of 0.5 ml. After a 60-min incubation at 37°C, unconjugated DES is extracted into 5 ml of chloroform and the residual metabolized 3H-DES in the aqueous phase is determined by liquid scintillation spectrometry. After addition of ß-glucuronidase to the aqueous phase, about 90% of the radioactivity could be extracted into chloroform, demonstrating that DES-glucuronic acid is the primary metabolite. Thus, this method easily permits quantitation of UDPGA in rat liver in the 1-10 nmol range.

Original languageEnglish (US)
Pages (from-to)145-151
Number of pages7
JournalJournal of Pharmacological Methods
Volume7
Issue number2
DOIs
StatePublished - 1982
Externally publishedYes

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Uridine Diphosphate Glucuronic Acid
Diethylstilbestrol
Uridine
Acids
Liver
Chloroform
Assays
Glucuronic Acid
Magnesium Chloride
Glucuronidase
Radioactivity
Scintillation
Metabolites
Spectrometry
Rats
Spectrum Analysis
Guinea Pigs
Liquids
Proteins

Keywords

  • Glucuronidation of diethylstilbestrol
  • Glucuronyltransferase
  • Uridine 5′-diphosphoglucuronic acid determination

ASJC Scopus subject areas

  • Pharmacology

Cite this

Determination of hepatic uridine 5′-diphosphoglucuronic acid concentration by conjugation with diethylstilbestrol. / Watkins, J. B.; Klaassen, C. D.

In: Journal of Pharmacological Methods, Vol. 7, No. 2, 1982, p. 145-151.

Research output: Contribution to journalArticle

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