DETERMINATION IN PLASMA OF FREE 17-HYDROXY AND 17-DESOXY CORTICOSTEROIDS AND THEIR GLUCURONICACID CONJUGATES*

Abstract

INTRODUCTION AS PART of a long-term investigation (1–4) on the systemic effects of both accidental and surgical trauma, it was decided to determine the blood levels of adrenocortical steroids. It is believed that the measurement of the degree and duration of adrenocortical stimulation can be more objectively evaluated by this means than by methods hitherto employed (e.g., urinary excretion studies). In 1952, Nelson and Samuels (5) reported a method for determining “17-hydroxycorticosteroids” following simple chloroform extraction of plasma or whole blood and chromatographic separation on Florisil, utilizing the Porter-Silber (6) reaction which determines only those steroids containing an alpha-ketol group with a hydroxyl group in the 17 position. In this laboratory it was not found possible to employ this method with any degree of reproducibility. In order to determine adrenocortical steroids reproducibly in peripheral blood plasma, it was found necessary to carry out a more thorough purification of the plasma extracts and to take certain essential technical precautions in the steroid separation and in column chromatography on Florisil or silica. A preliminary report of this method has been given (7); however, in presenting the details, no claims are made for any particular originality. Indeed, well tried extraction procedures were used, as described in the literature (8–11). The procedure of column chromatography employed was essentially that used by Nelson and Samuels, except that certain technically essential features were worked out as to the type of column and the purity and particle size of the Florisil.