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A convenient solid phase method for extracting and fractionating steroid hormones from serum or urine is presented. The influence of the solvent used for extraction, of the body fluid to be extracted, and of the degree of sample dilution on the efficiency of extraction was studied for progesterone and cortisol. The potency of fractionation was demonstrated by the separation of the steroid pairs, deoxycortisol--cortisol and oestradiol--oestriol, from a urine sample. Influence of ionic strength, pH and temperature on the reproducibility was assessed for deoxycortisol in undiluted urine. With respect to practicability and efficiency, this method has proved to be considerably superior to conventional liquid-liquid techniques.

The usefulness of recrystallization in establishing the radiochemical purity of steroids is widely recognized, but the potential limitations of the technique have received little attention. The current study reports the failure of standard recrystallization procedures using methanol/water as the solvent pair to separate contaminating 14C-17-hydroxyprogesterone (17-hydroxy-4-pregnene-3, 20-dione) from 3H- and 14C-labeled 11-deoxycortisol (17,21-dihydroxy-4-pregnene-3,20-dione) despite ten serial crystallizations. The standard criteria of radiochemical purity were met despite gross impurity of the crystals as evidenced by thin layer chromatography. Thus, recrystallization may, under certain conditions, yield misleading results when employed as the only method for identifying radioactive steroids. These observations illustrate the importance of an optimal choice of solvent and crystallization conditions, and emphasize the need for confirmation by derivative formation and chromatography.