COMPLETE HYDROLYSIS OF STEROID CONJUGATES AND QUANTITATIVE EXTRACTION OF FREE STEROIDS FROM URINE

A simple procedure for the serial hydrolysis and extraction of 5-ml urine samples is described. The samples are incubated at pH 4.45 for 24 hours at 37[degree]C with high concentrations of [beta]-glucuronidase and sulfatases from Helix pomatia. The incubated samples are freeze-dried and the residual solids are repeatedly extracted with a total of 15 ml of chloroform-methanol (1:2, v/v). The overall efficiency is over 99%. A method for the systematic estimation of urinary steroids based on the combined thin-layer and gas-liquid chromatographic analysis of total extracts is outlined.