In vitro biosynthesis of radioactive estradiol-17 beta, 17-glucosiduronate by rhesus monkey liver.

A method for the preparation of radioactive estradiol-17 beta, 17-glucosiduronate by incubating 3H-estradiol with rhesus monkey liver microsomal preparation in the presence of uridine diphosphoglucuronic acid is described. Small but significant amounts of the conjugate were also obtained from the 150,00 pellet and cytosol fractions. The addition of NADPH to the incubation media increased the yield of radioactive-estradiol-17 beta, 17-glucosiduronate perhaps by preserving the integrity of the C-17-hydroxyl group. As expected, the effect of the reduced nucleotide was more pronounced in the fractions other than the microsome. The biosynthesized conjugate was isolated and purified by multiple column chromatography and the structure was confirmed by derivative formation, enzyme hydrolysis and crystallization of the aglycone.