Simultaneous determination of ebastine and its three metabolites in plasma using liquid chromatography-tandem mass spectrometry.

We developed a method for determining ebastine, a new generation of antihistamines, and its three metabolites (hydroxyebastine, carebastine and desalkylebastine) in plasma simultaneously using LC/MS/MS. Four compounds and terfenadine, an internal standard, were extracted from plasma using a mixture of diethylether and dichloromethane in the presence of 1 M HCl. After drying the organic layer, the residue was reconstituted in mobile phase (acetonitrile:5 mM ammonium acetate, 50:50, v/v) and injected onto a reversed-phase C(18) column. The isocratic mobile phase was eluted at 0.2 ml/min. The ion transitions monitored in multiple reaction-monitoring mode were m/z 470.7-->167.1, 486.7-->167.1, 500.6-->167.1, 268.4-->167.1 and 472.7-->436.0 for ebastine, hydroxyebastine, carebastine, desalkylebastine and terfenadine, respectively. The coefficient of variation of the assay precision was less than 12.5%, and the accuracy exceeded 88%. The limit of detection was 0.5 ng/ml for desalkylebastine; 0.2 ng/ml for ebastine, hydroxyebastine and carebastine, respectively. This method was used to measure the plasma concentration of ebastine and its three metabolites from healthy subjects after a single 20 mg oral dose of ebastine. This analytic method is a very simple, sensitive, and accurate to determine the pharmacokinetic profiles of ebastine including its metabolites.