A liquid chromatographic assay with mass-spectrometric detection was developed for the quantitative determination of the cytochrome P450 3A phenotyping probe midazolam in human plasma. Sample pretreatment involved a one-step extraction of 600 μl aliquots with ethyl acetate. Midazolam and the internal standard, lorazepam, were separated on a column (150 mm×4.6 mm, i.d.) packed with 5 μm Zorbax Eclipse XDB-C8 material, using a mobile phase composed of methanol and 10 mM aqueous ammonium acetate (60:40, v/v). Column effluents were analyzed using mass-spectrometry with an atmospheric pressure chemical ionization source. Calibration curves were linear in the concentration range of 1.00-200 ng/ml. The accuracy and precision ranged from 92.8 to 112% and 0.056 to 13.4%, respectively, for four different concentrations of quality control samples analyzed in triplicate on eight separate occasions. The developed method was subsequently applied to study the pharmacokinetics of midazolam in a group of 35 human subjects at a single dose of 25 μg/kg.

Midazolam
dx.doi.org/10.1016/j.jchromb.2004.04.003, hdl.handle.net/1765/64413
Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences
Department of Medical Oncology

Lepper, E.R, Hicks, J.K, Verweij, J, Zhai, G, Figg, W.D, & Sparreboom, A. (2004). Determination of midazolam in human plasma by liquid chromatography with mass-spectrometric detection. Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences, 806(2), 305–310. doi:10.1016/j.jchromb.2004.04.003