Analysis of betamethasone in rat plasma using automated solid-phase extraction coupled with liquid chromatography–tandem mass spectrometry. Determination of plasma concentrations in rat following oral and intravenous administration

A method is described for the determination of betamethasone in rat plasma by liquid chromatography–tandem mass spectrometry (LC–MS–MS). The analyte was recovered from plasma by solid-phase extraction and subsequently analyzed by LC–MS–MS. A Packard Multiprobe II, an automated liquid handling system, was employed for the preparation and extraction of a 96-well plate containing unknown plasma samples, standards and quality control samples in an automated fashion. Prednisolone, a structurally related steroid, was used as an internal standard. Using the described approach, a limit of quantitation of 2 ng/ml was achieved with a 50 μl aliquot of rat plasma. The described level of sensitivity allowed the determination of betamethasone concentrations and subsequent measurement of kinetic parameters of betamethasone in rat. Combination of automated plasma extraction and the sensitivity and selectivity of LC–MS–MS offers a valuable alternative to the methodologies currently used for the quantitation of steroids in biological fluids.