Simple and validated HPLC–UV analysis of levetiracetam in deproteinized plasma of patients with epilepsy

Abstract

We present a simple, fast and validated method for the determination of the new generation antiepileptic drug (AED) levetiracetam (LEV) in plasma of patients with epilepsy using high performance liquid chromatography (HPLC) with UV detection. Plasma sample (500 μL) pretreatment was based on simple deproteinization by methanol spiked with the internal standard (I.S.). HPLC analysis was carried out on a Synergi 4-μm Hydro-RP, 150 mm × 4 mm I.D. column. The mobile phase was a mixture of potassium dihydrogen phosphate buffer (50 mM, pH 4.5) and acetonitrile (94:6, v/v) at an isocratic flow rate of 1.5 mL/min. The UV detector was set at 205 nm. Calibration curves were linear (mean correlation coefficient = 0.999) over a range of 4–80 μg/mL. The quantitation limit was 2 μg/mL and the absolute recovery was >90% for LEV and the I.S. Both intra and interassay precision and accuracy were lower than 7.5%. The chromatographic run lasted 13 min. The present procedure omitting expensive solid phase or time-consuming liquid–liquid extraction and drying steps is cheaper, faster and simpler than mostly published analytical methods for levetiracetam. Applied to a large population of patients with epilepsy this assay proved very practical in our therapeutic drug monitoring setting (TDM).