A LIQUID CHROMATOGRAPHIC-TANDEM MASS SPECTROMETRIC METHOD FOR THE QUANTITATIVE ANALYSIS OF DEXAMETHASONE IN HUMAN PLASMA

Abstract

ABSTRACT A simple and specific assay for the rapid quantification of the plasma concentration of dexamethasone from 0.250 to 250 ng/mL has been developed and validated using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The method used methyl-t-butyl ether (MTBE) to extract dexamethasone and the internal standard (IS) beclomethasone from alkalinized human plasma sample. The chromatographic separation was on a C18 column (50 × 3 mm, 5-µm) using acetonitrile-water-formic acid (35 : 65 : 0.1, v/v) mobile phase at a flow rate of 0.500 mL/min. The retention times were approximately 1.8 min for dexamethasone and 2.0 min for the internal standard. The tandem mass spectrometric detection was by monitoring singly charged precursor→production transitions 393→373 (m/z) for dexamethasone and 409→391 (m/z) for the internal standard. The validated method used 0.5-mL plasma. The linear correlation coefficients of the calibration curves were greater than, or equal to, 0.9972. The run time for each sample injection was 3.0 min. The recoveries of the extraction were found to be 88–91% for dexamethasone at three concentration levels and 84.5% for the IS, respectively. The intra-day precision and accuracy of the control samples were of <4.8% relative standard deviation (RSD) (n=6) and −7.6 to 5.6% relative errors (RE). The inter-day precision and accuracy were <5.1% RSD (n=18) and −6.9 to −0.3% RE. The partial volume experiments showed excellent dilution integrity for high concentration level control samples. The stability data illustrated that this compound was stable in plasma at least for three freeze/thaw cycles, or keeping plasma samples at ambient temperature for 24 hours, or storing extracted samples at room temperature for 48 hours, and or storing frozen samples at approximately −20°C for up to three months.