Simple and Rapid Quantification of Pioglitazone and Hydroxy Pioglitazone in Human Plasma Using Liquid Chromatography Coupled with Tandam Mass Spectrometry

A liquid chromatographic-tandem mass spectrometric (LC-MS/MS) method was developed for the simultaneous determination of Pioglitazone (PIO) and Hydroxy Pioglitazone (PIO M IV) in human plasma. Analytes were separated from plasma by protein precipitation extraction technique with 0.1% Formic Acid in acetonitrile as precipitant. Chromatographic separation was performed on a Thermo Hypurity 50 x 4.6 mm, 5μ-C18 column with the mobile phase consisted of 0.10% Formic acid in acetonitrile: 10mm Ammonium acetate (70:30). A tandem mass spectrometer equipped with atmospheric pressure chemical ionization source was used as detector and operated in the positive ion mode. Quantification was performed using multiple reaction monitoring (MRM) of the transitions m/z 357.2, 134.1 and m/z 373.1, 149.9 for PIO and PIO M IV respectively. The method showed a good linearity in a concentration range of 20.48 - 40000 ng/mL for PIO, and 10.29 - 2000 ng/mL for PIO M IV. The intra and inter-day precision was less than 15% and the absolute recovery was above 95%. This method was selective and rapid, sensitive for investigating drug concentrations in clinics