High-Speed Simultaneous Determination of Nine Antiepileptic Drugs Using Liquid Chromatography-Mass Spectrometry

Therapeutic drug monitoring of antiepileptic drugs (AEDs) is important and widely practiced. However, simultaneous AED assays usually concentrate only on old or new AEDs. A new simultaneous assay was developed to monitor both older and newer AEDs in the same sample. This assay measures zonisamide (ZNS), lamotrigine (LTG), topiramate (TPM), phenobarbital (PB), phenytoin (PHT), carbamazepine (CBZ), carbamazepine-10,11-diol (CBZ-Diol), 10-hydroxycarbamazepine (MHD), and carbamazepine-10,11 -epoxide (CBZ-E). Sample pretreatment consisted of a single solid-phase extraction (SPE) for all AEDs in a 100-μL plasma sample. HPLC separation was achieved on a Shimdazu Shimpack XR-ODS (4.6 id X 50 mm, 2.2-μm) column with a gradient mobile phase of acetate buffer, methanol, acetonitrile, and tetrahydrofuran. Four internal standards were used. Detection was achieved by atmospheric pressure chemical ionization mass spectrometry (APCI-MS) in selected-ion monitoring (SIM) mode with constant polarity switching. High recovery (88%-96%) was obtained for all compounds by SPE. Linearity was observed throughout an 80-fold concentration range, with correlation coefficient (r2) values higher than 0.99 for all AEDs. For the standards, the accuracy ranged from 89.3% to 111.8%. The within-run coefficient of variation (CVw) value was ≤9.7%, the between-run coefficient of variation (CVb) was ≤16.2%, and the total variability (CVt) was ≤16.8%. For the quality controls (QCs), accuracy ranged from 89.3% to 108.8%, CVw was ≤9.6%, CVb was ≤14.1%, and CVt was ≤15.1%. The correlation r2 values for comparison of this assay with existing validated assays in our laboratory (GC-MS or LC-MS) were 0.95, 0.91, 0.87, 0.95, and 0.95 for PHT, LTG, CBZ, CBZ-E, and CBZ-Diol, respectively.