Abstract
A sensitive and rapid bioanalytical method based on the LC-triple-stage fragmentation (LC-MS(3)) strategy on a hybrid triple quadrupole-linear ion trap mass spectrometer in combination with protein precipitation extraction for sample pretreatment has been developed and validated for the simultaneous determination of the antiepileptic drug oxcarbazepine (OXC) and its main active metabolite (MHD) in human serum. The separation was performed on a Waters XBridge BEH C18 column (2.5 µm, 2.1 × 50 mm) in isocratic elution with 0.1% formic acid in water and methanol (50:50, v:v) as the mobile phase. The run time for each sample was 2.0 min. The calibration curves ranging from 25 to 1600 ng/mL for OXC and from 0.5 to 32 μg/mL for MHD showed correlation coefficients (r) better than 0.99. All of the validation data, such as precision, accuracy and other parameters, fit the requirements of the current bioanalytical method validation guidelines. The LC-MS(3) method for quantitation of OXC and MHD was compared with the LC-MRM based method. Passing-Bablok regression coefficients and Bland-Altman plots showed that the developed LC-MS(3) method is a reliable method for quantitative analysis of OXC and MHD. The proposed LC-MS(3) method was successfully applied to determine the serum concentrations of OXC and MHD to support a clinical study.