Evaluation of Hydrolytic Products of 17-Ketosteroids by Means of Gas-Liquid Chromatography

In the course of evaluating urinary 17-ketosteroid (17-KS) gas-liquid chromatographic patterns, a number of peaks with retention times shorter than that of 17-KS were constantly observed. This backgroundmaterial was found to be degradation products of 17-KS produced by HCI hydrolysis. A study with pure 17-KS as a mixture and as individual steroids was used to identify the steroids responsible for most of the 9 compounds with retention times less than etiocholanolone. Hydrolysis in ethyl acetate containing perchloric acid produced smaller amounts of degradation products than HCI-hydrolysis. Enzymatic hydrolysisappears to be the most acceptable system for preparation of the extracts from urine for this system of assay. HE HYDROLYSIS OF conjugated urinary 17-ketosteroids (17-KS) by boiling or heating with mineral acids results in the formation of degrada-tion products (1-9). In this regard, the most labile and thoroughly studied 17-KS is dehydroepiandrosterorie (DHA) (1-3, 6-9). Degrada-tion of steroids by acid hydrolysis does not cause serious decrease in the total amount of 17-KS when measured by the Zimmerman reaction (6) and does not assume importance unless an individual 17-KS is to be identified. It is presumed that most of the degradation products react with the Zimmerman reagent. However, Sheath (7) was aI)le to detect losses of DHA and androsterone when these steroids were added to urine, and the specimen then subjected to acid hydrolysis. Evidence exists showing that the glucuronoside-conjugated 17-KS is adequately hydrolyzed by enzymatic systems (10), although the problem of inhibitors in urine remains unsolved. The use of sulfatases for split-ting steroids conjugated with sulfate is considered unsatisfactory, since inhibitors greatly reduce the effectiveness of this enzyme. For hydrolysis of sulfates, Lieberman et a!. (9) introduced the method of solvolysi