Simultaneous determination of alfentanil and midazolam in human plasma using liquid chromatography and tandem mass spectrometry

A fast, sensitive and selective liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the determination of alfentanil and midazolam in human plasma has been developed and validated. Alfentanil and midazolam were extracted from plasma using a mixed-mode cation exchange solid phase extraction method, with recoveries of both compounds greater than 80% at 3 different concentrations (1, 10 and 100 ng/ml). Compounds were analyzed on a C-18 column with a water and methanol mobile phase gradient with acetic acid as an additive, at a flow rate of 0.3 ml/min. The working assay range was linear from 0.25 to 100 ng/ml for each compound. The signal to noise ratio was 80 and 40 for alfentanil and midazolam, respectively, at the lowest concentration calibration standard, with less than 10% matrix suppression by human plasma at this concentration. Alfentanil and midazolam were stable in human plasma during storage at -80 degrees C, processing, and analysis. The procedure was validated and applied to the analysis of plasma samples from healthy human subjects administered oral and intravenous alfentanil and midazolam. (C) 2011 Elsevier B.V. All rights reserved.