title: Development of a sensitive and specific liquid chromatography/mass spectrometry method for the determination of tenofovir in human plasma.
abstract: A sensitive and specific method for the quantitation of tenofovir (TFV) in human plasma by liquid chromatography/electrospray ionization mass spectrometry was developed and validated. Plasma samples were prepared by solid-phase extraction performed on Waters Oasis cation-exchange cartridges (30 mg). Chromatographic separation was performed isocratically on a reversed-phase Waters Atlantis dC18 column (2.0x100 mm, 3 microm). The mobile phase consisted of a hydroxylamine/acetic acid buffer (pH 6.75) and methanol (93:7, v/v). The acquisition was performed in selected ion monitoring mode for the protonated molecular ions [M+H]+ of m/z 288.2 for TFV and 212.2 for the internal standard, zalcitibine. The method was fully validated to determine its specificity, recovery, linearity and sensitivity, accuracy and precision. The analytical range was set at 1-750 ng/mL using a 200 microL plasma sample, with a mean coefficient of determination (r2) of 0.9969. The mean accuracies for the calibration standards ranged from -5.0 to 4.3%, while the precisions were within 1.2 and 6.4%. Intra-assay and inter-assay mean accuracies for three quality control concentrations (2, 60, and 600 ng/mL) ranged from -6.1 to 10.7%, while the precisions were within 1.3 and 9.1%. TFV was shown to be stable under normal storage and assay conditions; no degradation was seen when stored at -20 degrees C or -80 degrees C for up to 6 months, and after 16 h at room temperature in the injection matrix. The present method provides an accurate, precise, and sensitive tool for TFV quantitation and was successfully applied to an external proficiency-testing program and pharmacokinetic analysis.
authors.affiliation: The University of Alabama at Birmingham School of Medicine, Division of Clinical Pharmacology, Birmingham, Alabama 35294-0019, USA.
authors.forename: Chantelle
authors.initials: C
authors.lastname: Bennetto-Hood
authors.forename: Mary C
authors.initials: MC
authors.lastname: Long
authors.forename: Edward P
authors.initials: EP
authors.lastname: Acosta
mesh_terms: D000225:Adenine / Q000031:analogs & derivatives* / Q000097:blood / Q000737:chemistry / Q000493:pharmacokinetics; D002138:Calibration; D002853:Chromatography, Liquid / Q000379:methods*; D004355:Drug Stability; D005615:Freezing; D006801:Humans; D015394:Molecular Structure; D063065:Organophosphonates / Q000097:blood* / Q000737:chemistry / Q000493:pharmacokinetics; D012015:Reference Standards; D015203:Reproducibility of Results; D018894:Reverse Transcriptase Inhibitors / Q000097:blood* / Q000737:chemistry / Q000493:pharmacokinetics; D012680:Sensitivity and Specificity; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D013696:Temperature; D000068698:Tenofovir; D013997:Time Factors; D016047:Zalcitabine / Q000737:chemistry
chemical_list: D063065:Organophosphonates; D018894:Reverse Transcriptase Inhibitors; D016047:Zalcitabine; D000068698:Tenofovir; D000225:Adenine