Determination of prednisolone in human adipose tissue incubation medium using LC–MS/MS to support the measurement of 11β-hydroxysteroid dehydrogenase activity

A method for the determination of prednisolone in human adipose tissue incubation medium has been developed, validated and used to support studies designed to measure the activity of 11β-hydroxysteroid dehydrogenase in human adipose tissue. After incubation, samples (80 μL) were extracted using Oasis HLB μElute SPE plates and the resulting extracts were analyzed using reversed-phase chromatography coupled to an Applied Biosystems Sciex PE API-4000 mass spectrometer with a TurboIonSpray® interface (400 °C). The method was validated over the calibration range of 0.5–100 ng/mL. Intraday precision and accuracy were 6.1% R.S.D. or less and within 6.3%, respectively. Interday precision and accuracy were 4.2% R.S.D. or less and within 3.6%, respectively. Extraction recovery of prednisolone was greater than 84% over the range of low to high quality control sample concentrations. The validated assay was used to support studies designed to estimate ex vivo 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) enzyme activity in human adipose tissue.