A METHOD FOR THE SEPARATION OF FOUR PHENOLIC ESTROGENS IN HUMAN URINE BY PAPER CHROMATOGRAPHY AND AN EVALUATION OF BIOLOGICAL ASSAY OF THESE COMPOUNDS

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Abstract
A method for clean separation of four phenolic estrogens in human urine is presented. This method involves enzymatic hydrolysis, ether extraction, paper chromatographic separation, purification and finally bioassay. The specificity and reliability of the bioassay of estrogens have been critically appraised. Biological assays of estrone and estradiol-17[beta] are satisfactory. Assay of two impeded estrogens, estriol and 16-epiestriol are less reliable, but multiple level assay can increase the accuracy. In recovery experiments in which estrogens were added to urine, not all of these added compounds were recovered, the deficit amounting to 30 to 50% of the amount added. It seems that the loss occurred during the extraction procedure. Washing with NaHCO3 may remove amounts of estriol and 16-epiestriol. All results of urinary excretion of estrogens described in this paper were obtained from the urine following enzyme hydrolysis. The portion following further acid hydrolysis has not been discussed here. The actual values will therefore be higher.