Development and Validation of a Stability‐Indicating RS HPLC Method for Leuprolide Acetate and Its Related Substances: Characterization of Degradation Products With LC‐MS

ABSTRACT A new stability‐indicating related substance method was developed and validated for leuprolide acetate (LPA) and its related impurities and established the degradation profile for the LPA drug that was exposed to various stress conditions. During the stress study, the drug showed substantial degradation in acidic (14.34%), basic (7.47%), and thermal (5.70%) conditions. The high‐performance liquid chromatography method was developed as an isocratic method, mobile phase A (triethylamine pH 3.0), and mobile phase B (acetonitrile/n‐propanol [300:200]). The chromatographic separation was attained by using an analytical HPLC column Inertsil ODS 3; 100 × 4.6 mm 3 μ. The degradation products (DPs) were characterized by using the LC‐MS with an electron spray ionization technique. The method was validated according to ICH guidelines. The method showed a good limit of detection (0.092–0.274 μg/mL) and quantification limits (0.244–0.829 μg/mL) for LPA and its known impurities. During the linearity test, the method showed correlation coefficient ( R 2 ) values between 0.9999 and 0.9990. The method showed acceptable precision % of RSD values (0.01–5.70) and accuracy values in terms of % of recovery (95.10–109.60) for LPA and its related impurities.