UHPLC-MS/MS method for simultaneous determination of carbamazepine and its seven major metabolites in serum of epileptic patients

Carbamazepine (CBZ) was considered as the drug of choice in the treatment for various forms of epilepsy, yet the non-negligible adverse effects of CBZ suspend as the major concern for rational and efficient clinical medication. This study developed a method for the simultaneous determination of CBZ and its seven metabolites acridine (AI), iminostilbene (IM), acridone (AO), carbamazepine‑10,11‑epoxide (CBZ‑EP), 10,11‑dihydro‑10,11‑trans‑dihy‑droxy‑carbamazepine (CBZ‑DiOH), 2‑hydroxycarbamazepine (2‑OH‑CBZ) and 3‑hydroxycarbamazepine (3‑OH‑CBZ) with phenacetin as the internal standard (IS) using ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). Plasma samples were purified with the one-step 96-well protein precipitation plate. Chromatographic separation was achieved on an Agilent Eclipse XDB-C18 (1.8 μm, 2.1 mm × 50 mm) chromatography column carrying the mobile phrase of acetonitrile and 0.1% formic acid in a gradient elution at a flow rate 0.3 mL/min. Agilent G6460 MS/MS in the positive MRM mode using electrospray ionization (ESI) was applied for quantification of target compounds. CBZ and its seven metabolites showed good linear correlation coefficient (r > 0.990). Intra-day and inter-day precision (CV) were no >15%. This method was successfully accomplished within 5 min which was especially applicable for therapeutic drug monitoring (TDM) of CBZ and its metabolites in epileptic patients, and it provided insights for toxicological studies to achieve a rational and effective individualized administration in clinical practice.