Quantitative Estimation and Identification of Estrogens in Bovine Urine

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Abstract
This work was conducted to further develop methodology for the chemical assay of estrogen in bovine urine. The method includes use of C14 internal standards, a combination of enzyme and acid hydrolysis, paper chromatography, and fluorimetry for the assay of 4 estrogens in bovine pregnancy urine. Enzyme hydrolysis liberated approximately 90% of the 17[alpha]-estradiol but only about 10% of the estrone as compared with enzyme followed by acid hydrolysis. Approximately equal amounts of 17[beta]-estradiol and estriol-like compound were liberated by enzyme and acid hydrolysis. Three of the compounds, 17[alpha]-estradiol, 17[beta]-estradiol and estrone, were positively identified by paper chromatographic mobilities, sulfuric acid and absorption spectra, and biological potencies. Additional proof of structure for 17[alpha]-estradiol was provided by the Kagi-Miescher and Haenni-Iron-Kober tests. Chromatographic studies suggest the presence of 16, 17-epiestriol, but inadequate quantities of the hormone prevented identification.