Determination of metoprolol and its two metabolites in human plasma and urine by high performance liquid chromatography with fluorescence detection and its application in pharmacokinetics

A simple, specific and sensitive HPLC method has been developed for the determination of metoprolol and its two metabolites in human plasma and urine. Separation of metoprolol, alpha-hydroxymetoprolol, O-desmethylmetoprolol and esmolol (internal standard) was achieved on an Agilent XDB-C18 column (150 mm x 4.6 mm, 5 mu m) using fluorescence detection with Ex 216 nm and Em 312 nm. The mobile phase consisted of ACN-H2O-0.1%TFA. The analysis was performed in less than 16 min with a flow rate of 0.8 mL/min. The assay was linear over the concentration range of 5-600 ng/mL and 2.5-300 ng/mL for metoprolol and its metabolites, respectively. The LOQ were 5.0 and 2.5 ng/mL for plasma and urine, respectively. Good precision and accuracy for metoprolol and its two metabolites were obtained. The extraction recoveries were found to be more than 86.91% both in plasma and urine. At the same time, the method was successfully applied to nine healthy volunteers who had been given an oral tablet of 100 mg metoprolol. Crown Copyright (c) 2013 Published by Elsevier B.V. All rights reserved.