Determination of 2-hydroxyflutamide in human plasma by liquid chromatography-tandem mass spectrometry (LC-MS/MS): application to a bioequivalence study on Chinese volunteers.

A sensitive, simple and rapid ultra fast liquid chromatography (UFLC)-ESI-MS/MS method was developed for the determination of 2-hydroxyflutamide in human plasma using tegafur as the internal standard. The plasma sample was pretreated with methanol for protein precipitation and the analytes were separated on an Ultimate C18 column (5 microm, 2.1 mm x 50 mm, MD, USA) with the mobile phase consisted of acetonitrile and water (2:1, v/v). Detection was performed on a triple-quadrupole tandem mass spectrometer under a negative multiple reaction-monitoring mode (MRM). The mass transition ion-pair was followed as m/z 290.90-204.8 for 2-hydroxyflutamide and 198.9-128.8 for tegafur. Linear calibration curves were obtained in the concentration range of 1.742-1452 ng/ml with a lower limit of quantification of 1.742 ng/ml. The intra- and inter-batch precision values were less than 8.1% and 5.6%, respectively. The established method was successfully applied to a bioequivalence study of two flutamide preparations (250 mg) in 20 healthy male volunteers.