Cortisol and cortisone analysis in serum and plasma by atmospheric pressure photoionization tandem mass spectrometry

Objectives: Cortisol metabolism is controlled by 11β-hydroxysteroid dehydrogenase (11β-HSD) isoenzymes, which interconvert cortisol and cortisone. Accurate measurement of the cortisol and cortisone concentrations and their ratio provide useful information about 11β-HSD activity. Methods: Cortisol and cortisone were extracted with methyl-tert-butyl ether from 100 μl of serum or plasma. The extract was evaporated, reconstituted with mobile phase, and analyzed by tandem mass spectrometry using a photoionization interface. The transitions monitored were: m/z 363 to 121 and 363 to 97 for cortisol, 361 to 163 and 361 to 105 for cortisone. Results: Within-run and between-run coefficients of variation were less than 6% and 12%; 14% and 22%; 11% and 21% for cortisol, cortisone, and their ratio, respectively. The limit of detection was 1 μg/l for cortisol and 5 μg/l for cortisone. Normal ranges for cortisol and cortisone concentration and for their ratio in plasma (n = 120) determined as the central 95% were 33–246 μg/l for cortisol, 8–27 μg/l for cortisone, and 0.081–0.301 for the cortisone/cortisol ratio. Conclusions: We developed a simple sensitive method for cortisol and cortisone analysis in plasma and serum that uses a small sample volume. The method is very specific, fast, does not have any known interference, and is useful for diagnosis of variety of disease and pathologic conditions.