Sephadex LH-20 multiple-column chromatography for the simultaneous separation of progesterone, deoxycorticosterone and 17alpha-hydroxyprogesterone from small plasma samples.

A simple and convenient chromatographic method is described for the simultaneous and complete separation of the unconjugated steroids progesterone, deoxycorticosterone and 17 alpha-hydroxyprogesterone on 40-cm columns of Sephadex LH-20 using a water-saturated solvent system containing n-heptane-chloroform (1:1) plus 0.25% of ethanol. Manual operation of up to ten columns run in parallel is facilitated by the use of graduated, cylindrical, solvent reservoirs on top of the columns. Thus, negligible column-to-column, and only limited day-to-day, variations occur in the elution patterns, and constant high recoveries are obtained. When combined with a previously described 60-cm LH-20 chromatography, eight important corticosteroids can be isolated individually from 16 to 20 small plasma samples per day.