Abstract

METHOD DEVELOPMENT AND VALIDATION FOR THE ESTIMATION OF METHOTREXATE BY RP-HPLC METHOD

Palleni Girija Lakshmi, Dr. M. Sathish Kumar*, Dr. A. Mallik, Dr. N. Jyothi*

ABSTRACT

A simple, Accurate, precise method was developed for the reversed-phase high performance liquid chromatography method was developed and validated for verification and validation of Methotrexate. The following is achieved by using column C₁₈ column (250 mm x 4.6mm), 5μ using anhydrous monobasic sodium phosphate (buffer) and acetonitrile in the ratio of 1:19 at ambient temperature. The pH was adjusted to 6.0. The flow rate of 1.0MI/min. Detection was carried using a UV detector at 280 nm. The method was validated. The total run time was found to be 40 min. The retention time was found to be 10 mins. The method was validated for system suitability, specificity, accuracy, precision, linearity, method precision, intermediate precision (analyst - 1, analyst-2), recovery. The calibration curve was found to be linear with correlation 0.99995. The percentage recovery for methotrexate was found to be in the range of 99.74-100.53 % and %RSD was found to be Not less than 2.0%. The proposed method was found to be fast, Accurate and precise for the validation of methotrexate in API.

Keywords: Methotrexate, RP-HPLC, Validation, Accuracy, Robustness.