Assay of human plasma cortisone by liquid chromatography: normal plasma concentrations (between 8 and 10 a.m.) of cortisone and corticosterone

Abstract

A high-performance liquid chromatographic method using ultraviolet detection to quantitate human plasma concentrations of cortisone simultaneously with cortisol and corticosterone is described. The method is based on the use of an octadecyl silica column (100 mm x 2 mm I.D., 3 microns), an ultraviolet absorbance detector (242 nm) with a 10 mm path length flow-cell, and a mobile phase composed of water-tetrahydrofuran-acetonitrile (82:10:8, v/v) containing 5 ml/l triethylamine and citric acid to adjust the pH of the buffer to 6.5. Flumethasone is used as the internal standard. The detection limit of the method for the three steroids is 300 ng/l using a 1-ml sample. The average inter-assay coefficient of variation for cortisone is 3.3% and the average recovery is 100.8%. Possible interferences from common drugs and endogenous and exogenous steroids in the method have been studied. Plasma concentrations (drawn from 8 to 10 a.m.) of cortisone and corticosterone for 43 normal volunteers have been determined.