VALIDATED LIQUID CHROMATOGRAPHIC ULTRAVIOLET METHOD, FOR THE QUANTIZATION OF LEVETIRACETAM IN HUMAN PLASMA USING LIQUID – LIQUID EXTRACTIONAbstract
A simple, sensitive and rapid high performance liquid chromatography method with UV detection (215nm) was developed and validated for the quantification of Levetiracetam in human plasma. Following a single-step liquid–liquid extraction with diethyl ether/dichloromethane (70/30 v/v), the analyte and internal standard (Zonisamide) were separated using an isocratic mobile phase, mixture of buffer (5mM Di-Potassium hydrogen phosphate anhydrous, pH7.2): Methanol (85:15 v/v) on reverse phase C-8 Kromasil column. The lower limit of quantization was 1µg/mL, with a relative standard deviation of less than 20%. A linear range of 1µg/mL to 40µg/mL was established. This HPLC method was validated with between and within-batch precision of 5.6–8.9% and 3.9-5.3%, respectively. The between and within batch accuracy was 99.9-106.3% and 96.2-102.0%, respectively. Frequently co-administered drugs did not interfere with the described methodology. Stability of Levetiracetam in plasma was >90%, with no evidence of degradation during sample processing (autosampler) and 60 days storage in a freezer. This validated method is sensitive and simple with between-batch precision of <10% and was used in pharmacokinetic studies.