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

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Abstract
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 87 °C with high concentrations of β-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.