Development and validation of a high-performance liquid chromatography with tandem mass spectrometry method for quantification of tofacitinib in human plasma

Abstract

Introduction. The use of tofacitinib as a pharmacological treatment for rheumatoid arthritis remains relevant in the light of the predicted increase in prevalence of the disease. At the same time, due to the withdrawal of several foreign pharmaceutical companies from the Russian pharmaceutical market, there has been a heightened demand for domestically produced medications, including generic formulations of tofacitinib. Registration of generic drugs necessitates the conduct of bioanalytical studies. Development and validation of a method for quantifying the analyte in biosamples remains to be a crucial part of the bioequivalence studies.Aim. The aim of this research is to develop and validate a method for the quantitative determination of tofacitinib in human plasma using high-performance liquid chromatography as the separation system coupled with a tandem mass spectrometer for detection purposes.Materials and methods. Biosample preparation was based on plasma proteins precipitation using acetonitrile. Baricitinib was selected as an internal standard. The analytical range of the method was 1.00 to 200.00 ng/mL and was further expanded to 0.30 to 200.00 ng/mL during the analytical phase of the study. The mobile phase consisted of water and acetonitrile, both acidified with formic acid (0.1 % v/v). The stationary phase was a Phenomenex Kinetex C18 column [100 × 3.0 mm, with a particle size of 5 µm (Phenomenex, USA)]. Sample separation and detection were carried out using high-performance liquid chromatography coupled with a tandem mass spectrometry (HPLC-MS/MS), operating in positive ion mode. The multiple reaction monitoring (MRM) transitions selected for the analyte and the internal standard were 313.30 to 173.00 m/z and 371.90 to 186.00 m/z, respectively.Results and discussion. The developed assay was validated in accordance with the current requirements of regulatory documentation from the EAEU (Eurasian Economic Union), FDA (US Food and Drug Administration), and EMA (European Medicines Agency) with the following parameters being evaluated: selectivity, specificity, carry-over, matrix effect, recovery, calibration curve, lower limit of quantitation, accuracy, precision, stability. The validated method was applied in the analytical part of a bioequivalence study of domestically produced generic tofacitinib.Conclusion. A method for the quantitative determination of tofacitinib in human blood plasma with an analytical range of 0.30–200.00 ng/mL was developed and validated. Application of the assay during the analytical phase of bioequivalence study of generic tofacitinib confirms the possibility of using the method in similar bioanalytical investigations.

Keywords

• tofacitinib
• validation
• lc-ms/ms
• bioequivalence
• pharmacokinetics
• bioanalytical study