STABILITY INDICATING METHOD DEVELOPMENT AND VALIDATION OF SIMULTANEOUS ESTIMATION OF NIVOLUMAB AND RELATLIMAB IN BULK AND PHARMACEUTICAL DOSAGE FORM BY USING RP-HPLC

A simple, specific, sensitive, rapid, precise, accurate, robust, and stability indicating reverse phase high performance liquid chromatography method has been developed for the quantitative analysis of Nivolumab and Relatlimab in pharmaceutical dosage form. Chromatographic separation of Nivolumab and Relatlimab was achieved on Waters Alliance-e2695 by using Waters Waters X-Terra RP-18 (250x 4.6mm, 5µ) column and the mobile phase containing Acetonitrile: 0.1% Perchloric acid in the ratio of 40:60% v/v. The flow rate was 1.0 ml/min; detection was carried out by absorption at 231nm using a photodiode array detector at ambient temperature. The number of theoretical plates and tailing factor for Nivolumab and Relatlimab were NLT 2000 and should not more than 2 respectively. % Relative standard deviation of peak areas of all measurements always less than 2.0 respectively. The validation was carried out an optimized method and obtained results were within the limits of acceptance criteria for all the parameters like System suitability, Specificity, Linearity, Accuracy, Precision, LOD, LOQ, Robustness and stability of the method has been checked by the forced degradation studies. Based on the above information the method can be used for the daily routine analysis.