LC-MS-MS Determination of Oxcarbazepine and an Active Metabolite in Human Plasma for Clinical Application.

Abstract

A bioanalytical method for the simultaneous determination of oxcarbazepine (OXC) and its pharmacologically active metabolite, 10, 11-dihydro-10-hydroxycarbamazepine (HOXC), in human plasma was developed using a high-performance liquid chromatography with tandem mass spectrometry. After protein precipitation by acetonitrile, the analytes (OXC and HOXC) and a stable-labeled isotope of OXC as an internal standard (IS) were chromatographed on a Synergi Hydro-RP column (2.0 mm × 50 mm, 4 μm) with a gradient elution at a flow rate 0.5 mL/min. Detection was performed in electrospray ionization in the positive mode by monitoring the selected ion transitions at m/z 253.1 → 180.2, m/z 255.1 → 192.2 and m/z 257.2 → 184.2 for OXC, HOXC and the IS, respectively. The method was validated according to current bioanalytical method validation guidelines. The calibration standard curve ranged from 0.02 to 10 μg/mL for OXC and 0.1-50 μg/mL for HOXC using only 0.05 mL of plasma. No interferences were detected in blank plasma and hemolyzed plasma did not have any impacts on the assay. Accuracy and precision in the intra- and inter-batch reproducibility were within 15%. Neither cross-analytes inter-conversion nor matrix effects were observed. The method was successfully applied to determine plasma concentrations of OXC and HOXC to support a clinical study.