A simple and fast LC-MS/MS method for the simultaneous determination of tenofovir alafenamide and tenofovir in human plasma.

A simple and fast liquid chromatography-tandem mass spectrometry method was established and validated for the simultaneous determination of tenofovir alafenamide (TAF) and tenofovir (TNF) in human plasma. A simple protein precipitation procedure was employed to extract analytes from plasma. Chromatographic separation was performed on an Eclipse Plus C18 column utilizing a fast gradient elution starting with 2% of 2 mM ammonium acetate-formic acid (100/0.1, v/v) followed by increasing the percentages of acetonitrile. Detection was performed on a tandem mass spectrometer equipped with an electrospray ionization source operated in the positive ionization mode, using the transitions of m/z 477.2 → m/z 346.1 for TAF, and m/z 288.1 → m/z 176.1 for TNF. TAF-d5 and TNF-d7 were used as the internal standard of TAF and TNF, respectively. The method was validated in concentration ranges of 1.25-500 ng/mL for TAF and 0.300-15.0 ng/mL for TNF with acceptable accuracy and precision.