• 1,4-Diamino-2,3-dicyano-1,4-bis (2-aminophenylthio) butadiene (U0126);
  • Liquid chromatography-tandem mass spectrometry;
  • Rat plasma;
  • Validation

A simple, robust, and rapid LC-MS/MS method was developed for the quantitation of U0126 and validated in rat plasma. Plasma samples (20 μL) were deproteinized using 200 μL ACN containing 30 ng/mL of chlorpropamide, internal standard. Chromatographic separation performed on an Agilent Poroshell 120 EC-C18 column (4.6 × 50 mm, 2.7 μm particle size) with an isocratic mobile phase consisting of a 70:30 v/v mixture of ACN and 0.1% aqueous formic acid. Each sample was run at 0.6 mL/min for a total run time of 2 min per sample. Detection and quantification were performed using a mass spectrometer in selected reaction-monitoring mode with positive ESI at m/z 381 [RIGHTWARDS ARROW] 123.9 for U0126 and m/z 277 [RIGHTWARDS ARROW] 175 for the internal standard. The standard curve was linear over a concentration range of 20–5000 ng/mL with correlation coefficients greater than 0.9965. Precision, both intra- and interday, was less than 10.1% with an accuracy of 90.7–99.4%. No matrix effects were observed. U0126 in rat plasma degraded approximately 41.3% after 3-h storage at room temperature. To prevent degradation, sample handling should be on an ice bath and all solutions kept at 4°C. This method was successfully applied to a pharmacokinetic study of U0126 at various doses in rats.