The enzymatic synthesis of aldosterone glucuronide and its determination by high performance liquid chromatography

Abstract

This thesis describes the development of a possible procedure for the synthesis of aldosterone-18-glucuronide. Aldosterone is a steroid which is clinically important. Aldosterone-18-glucuronide is not available commercially. Without a pure standard, it is difficult to isolate the synthesized product using HPLC. The various procedure for glucuronide formation in the literature were studied and used in this research. Addosterone and UDPGA from rabbit liver in Tris buffer at pH 7.4 incubated at 37°C for one hour gave a positive result. To separate the products from the reactants, HPLC was used. HPLC separated the reactants from the newly found product. The newly found product was collected from the HPLC at its retention time and found that it was an acid-labile conjugate of aldosterone because it released aldosterone when hydrolyzed at pH 1.0 at room temperature. The UVmax of this compound was found to be different from aldosterone and glucuronic acid. However, the test for glucuronic acid after hydrolysis of the synthesized aldosterone conjugate using HPLC gave reasons to think that the synthesized product might be a Tris complex of aldosterone-18-glucuronide. Suggestions for further work are discussed.