An automated analytical method for the determination of felbamate in human plasma by robotic sample preparation and reversed-phase high performance liquid chromatography

Abstract An automated analytical method for the determination of felbamate in human plasma is described. Sample cleanup and preparation was performed by means of a Zymate II laboratory robot and consisted of a liquid—liquid extraction of felbamate and the internal standard, primidone, from human plasma to dichloromethane. The dichloromethane was evaporated and reconstituted in a phosphate buffer. Separation was performed by reversed-phase high performance liquid chromatography using a 5 μm Hypersil® ODS column (150 × 4.6 mm) and a mobile phase consisting of a mixture of phosphate buffer (pH = 6.5, 0.015 M) and acetonitrile (79:21, v/v). Quantitation was performed by measurement of the UV absorbance at a wavelength of 210 nm. The lower limit of quantitation was 0.100 μg ml −1 using 200 μl of plasma. The mean absolute analytical recovery of felbamate was 75.2% ( n = 28). The recovery of the internal standard, primidone was 74.7% ( n = 10). The within-day precision was below 3.8% at all concentration levels, except at the lower limit of quantitation (18.3%). The within-day accuracy varied between −3.7 and +7.4%. The between-day precision was below 5.0% at all concentration levels. The between-day accuracy of the method varied between −5.7 and +1.6%. The selectivity of the method towards several other anti-epileptic drugs has been demonstrated.