A highly sensitive method for the quantification of fludrocortisone in human plasma using ultra-high-performance...

A simple and high sensitive ultra-high-performance liquid chromatography tandem mass spectrometry method for the determination of fludrocortisone in human plasma was developed and validated as per guidelines. The analyte and internal standard (IS), fludrocortisone-d5 , were extracted from human plasma via liquid-liquid extraction using tert-butyl methyl ether. The chromatographic separation was achieved on a Chromolith RP18e column using a mixture of acetonitrile and 2 mm ammonium formate (70:30, v/v) as the mobile phase at a flow rate of 0.7 mL/min. Quantitation was performed on a triple quadrupole mass spectrometer employing electrospray ionization technique, operating in multiple reaction monitoring and positive ion mode. The precursors to product ion transitions monitored for fludrocortisone and IS were m/z 381.2 → 343.2 and 386.2 → 348.4, respectively. The assay was validated with linear range of 40-3000 pg/mL. The intra- and inter-day precisions (relative standard deviation) were within 0.49-7.13 and 0.83-5.87%, respectively. The proposed method was successfully applied to pharmacokinetic studies in humans.