Stability Indicating RP-HPLC Method for Quantitative Estimation of Levetiracetam and its Impurities in Pharmaceutical Dosage Form

Abstract

Objectives: A simple, precise, accurate, robust and selective stability-indicating reverse phase high performance liquid chromatographic method for the separation and quantification of Levetiracetam and its impurities in Levetiracetam liquid dosage formulations. Materials and Methods: The analysis of improved RP-HPLC method for the separation and quantification of Levetiracetam and its impurities are described. Samples are analysed by means of reverse phase (RP-HPLC) using an Inertsil ODS-3V, 150 x 4.6 mm, 3μm and the mobile phase consists of two channels A and B. Channel-A: pH 5.50 phosphate buffer : acetonitrile (950:50 v/v) and channel-B: acetonitrile: water (90:10 v/v). The flow rate is 1.0 ml/min. The column temperature was maintained at 40°C and sample temperature was maintained at 25°C, injection volume 10μL and wavelength fixed at 205 nm. Results: For selectivity, the chromatograms were recorded for standard and sample solutions of Levetiracetam and its related substances. Selectivity studies reveal that the peak is well separated from each other. Therefore the method is selective for the determination of related substances in Levetiracetam. There is no interference of diluent and placebo at Levetiracetam and impurities peaks. The elution order and the retention times of impurities and Levetiracetam obtained from individual standard preparations and mixed standard preparations are comparable. The limit of detection (LOD) and limit of quantitation (LOQ) for Levetiracetam standard 0.0023% and 0.0070%, impurity-A 0.0049% and 0.0147%, impurity-C 0.0024% and 0.0074% and Levetiracetam RC-A 0.0091% and 0.0277% respectively. The linearity results for Levetiracetam and all the impurities in the specified concentration range are found satisfactory, with a correlation coefficient greater than 0.99.Calibration curve was plotted and correlation co-efficient for Levetiracetam and its impurities found to be 1.000, 0.9999, 1.000 and 0.9994 respectively. The accuracy studies were shown as %recovery for Levetiracetam and its impurities at specification level. The limit of % recovered shown is in the range of 80 and 120% and the results obtained were found to be within the limits. Hence the method was found to be accurate. For precision studies six replicate injections were performed. %RSD was determined from the peak areas of Levetiracetam and its impurities. The acceptance limit should be not more than 10, and the results were found to be within the acceptance limits. Conclusion: The developed LC method was validated with respect to specificity, precision, linearity, ruggedness and robustness. Validation study compared as per ICH guideline.