Development and validation of a solid phase extraction-HPLC method for the determination of carbamazepine and its metabolites, carbamazepine epoxide and carbamazepine trans-diol, in plasma

A solid phase extraction-HPLC method has been developed and validated for the rapid analysis of carbamazepine and its two metabolites, carbamazepine epoxide and carbamazepine trans-diol, in human plasma. The analysis was performed using a C18 Bakerbond-BDC analytical column (250 mm×4.6 mm i.d., particle size 5 μm). The optimal conditions for the separation were established with the mobile phase acetonitrile – 10 mM phosphate buffer, pH 7.0 (30:70, v/v) at a flow rate of 1.5 mL min-1 and temperature of 35 °C, with UV detection at 210 nm. The total run time was about 8 minutes. The SPE procedure for the extraction of the analytes from a plasma sample was developed using Oasis HLB cartridges and subsequently, the eluate was injected into the HPLC system for analysis. Afterwards, the SPE-HPLC method was subjected to validation. Linearity was obtained over the concentration range of 0.2–25 μg mL-1 for carbamazepine, carbamazepine epoxide and carbamazepine trans-diol, with correlation coefficients higher than 0.995. The method showed good intra-day and inter-day precision with a relative standard deviation below 7.96 %, while the accuracy ranged from 92.09 to 108.5 % for all analytes. Finally, the method was successfully applied to the analysis of the plasma samples of epileptic patients in monoand polytherapy.