Propylthiouracil quantification in human plasma by high-performance liquid chromatography coupled with electrospray tandem mass spectrometry: Application in a bioequivalence study

• This HPLC-MS-MS method for measuring propylthiouracil in human plasma offers advantages over those previously reported, in terms of both a simple liquid–liquid extraction without clean-up procedures, as well as a faster run time.
• The assay performance results indicate that the method is precise and accurate enough for the routine determination of the propylthiouracil in human plasma.
• The test formulation with and without food was bioequivalent to reference formulation. Food administration increased the Tmax and decreased the bioavailability (Cmax and AUC).

A rapid, sensitive and specific method for quantifying propylthiouracil in human plasma using methylthiouracil as the internal standard (IS) is described. The analyte and the IS were extracted from plasma by liquid–liquid extraction using an organic solvent (ethyl acetate). The extracts were analyzed by high performance liquid chromatography coupled with electrospray tandem mass spectrometry (HPLC–MS/MS) in negative mode (ES−). Chromatography was performed using a Phenomenex Gemini C18 5 μm analytical column (4.6 mm × 150 mm i.d.) and a mobile phase consisting of methanol/water/acetonitrile (40/40/20, v/v/v) + 0.1% of formic acid. For propylthiouracil and I.S., the optimized parameters of the declustering potential, collision energy and collision exit potential were −60 (V), −26 (eV) and −5 (V), respectively. The method had a chromatographic run time of 2.5 min and a linear calibration curve over the range 20–5000 ng/mL. The limit of quantification was 20 ng/mL. The stability tests indicated no significant degradation. This HPLC–MS/MS procedure was used to assess the bioequivalence of two propylthiouracil 100 mg tablet formulations in healthy volunteers of both sexes in fasted and fed state. The geometric mean and 90% confidence interval CI of Test/Reference percent ratios were, without and with food, respectively: 109.28% (103.63–115.25%) and 115.60% (109.03–122.58%) for Cmax, 103.31% (100.74–105.96%) and 103.40% (101.03–105.84) for AUClast. Conclusion: This method offers advantages over those previously reported, in terms of both a simple liquid–liquid extraction without clean-up procedures, as well as a faster run time (2.5 min). The LOQ of 20 ng/mL is well suited for pharmacokinetic studies. The assay performance results indicate that the method is precise and accurate enough for the routine determination of the propylthiouracil in human plasma. The test formulation with and without food was bioequivalent to reference formulation. Food administration increased the Tmax and decreased the bioavailability (Cmax and AUC).