Simultaneous determination of cortisol and cortisone from human serum by liquid chromatography-tandem mass spectrometry

A fast,sensitive,and selective liquid chromatography-tandem mass spectrometry (lc-ms/ms) method was validated and then the levels of cortisol and cortisone from sera of healthy adults were determined by the lc-ms/ms method. one hundred μl of serum sample was directly extracted by adding 2 ml ethyl acetate,followed by chromatographic separation on a c18 column with a mobile phase consisting of 5 mm ammonium acetate and methanol (25: 75,v/v). the precision,accuracy,and average recovery of the method were 1.5-5.3%,95.4-102.5%,and 96.4% for cortisol,and 1.9-6.0%,89.2-98.8%,and 79.9% for cortisone,respectively. the method was linear from 1.0 to 500.0 ng/ml r 2 = 0.999 for cortisol and 2.5 to 100.0 ng/ml r 2 = 0.998 for cortisone. the limits of detection (lod) and quantification (loq) were 0.2 and 1.0 ng/ml for cortisol,and 1.0 and 2.5 ng/ml for cortisone,respectively. the average cortisol concentration (133.9 ± 63.7 ng/ml) of samples collected between 9:00 and 11:00 a.m. was higher approximately 4.4 times than that of cortisone (30.5 ± 10.7 ng/ml) p < 0.0001. the average cortisone/cortisol ratio was 0.225. therefore,the lc-ms/ms method may be useful for the diagnosis of some adrenal diseases and the assessment of 11β-hydroxysteroid dehydrogenase (11β-hsd) activity in clinical laboratories. © 2014 sanghoo lee et al.