OPTIMIZATION OF ROBUST HPLC APPROACH FOR ASSESSING PROCESS RELATED IMPURITIES OF LEMBOREXANT AND ELUCIDATION OF STRESS-INDUCED DEGRADATION PRODUCTS THROUGH LCMS/MS

This study primarily aimed to establish an uncomplicated yet highly responsive HPLC technique to effectively separate and quantify process-related impurities of Lemborexant. Additionally, it aimed to explore the forced degradation behavior of lemborexant through systematic assessments by utilizing LCMS. The chromatographic separation of drug substance, process related impurities and its degradation products (DPs) was achieved on Kinetex C18 (150×4.6 mm, 5 μm) column at that was maintained at 350C temperature using 10 mM ammonium formate buffer pH 4.2, acetonitrile and methanol in 65:25:10 (v/v) isocratic elution at 0.7 mL/min. Detection wavelength was selected as 265 nm. In the proposed conditions, lemborexant is identified at 4.06 and 6.19, 9.33 min and 1.60 min respectively for impurity 1, 2 and 3 min with acceptable system suitability and specificity. The method produces LOD at 0.009 for impurities with calibration range of 40–280 µg/mL for lemborexant and 0.04 - 0.28 µg/mL for impurities. The remaining validation parameters were observed to fall within acceptable ranges for both lemborexant and its impurities. The compound underwent exposure to various stress conditions (acid, base, peroxide, thermal, and UV light) as outlined in accordance with ICH Q1A (R2) guidelines. The degradation products formed during the stress study were detected and characterized using LCMS/MS in ESI positive mode. This involved a thorough comparison of collision-induced dissociation mass spectrometry data between the degradation products and lemborexant. Consequently, potential structures for five degradation compounds were proposed. The outcomes of supplementary validation investigations were equally satisfactory, confirming their appropriateness for the regular quantification of lemborexant and its related impurities in both bulk drug and pharmaceutical formulations. Furthermore, these findings have the potential to shed light on the mechanism of stress-induced degradation in lemborexant