A Stability‐Indicating RP‐HPLC Method for the Simultaneous Analysis of a Novel Synthetic Decapeptide and Six Related Substances

Abstract

Currently, the therapeutic potential of complex biomolecules such as peptides has significantly enhanced the demand for analytical method development and validation for their quality control. Peptides are more complex to analyze than small molecules because of their larger size and unique physical–chemical properties, which create different challenges for analytical method development. Reversed‐phase high‐performance liquid chromatography (RP‐HPLC) with ultraviolet or mass spectrometry is the most widely used technique for analyzing peptides and related substances. In the present study, a decapeptide and its six related possible impurities were synthesized in‐house and identified. A stability‐indicating RP‐HPLC method has been developed for the estimation of the decapeptide and its related substances. The developed method was validated according to the International Conference on Harmonization guidelines for specificity, linearity, limit of detection, quantification, accuracy, precision, and robustness. The stability‐indicating capability of the proposed method was studied under the stress conditions of acid, base, oxidative, thermal, humidity, and photolytic degradation. The decapeptide peak and its degradant were found to be homogeneous and pure in the studied stress trials, with mass balance for all types of degradation exceeding 95%. It confirms the reliability and appropriateness of determining the integrity and purity of the decapeptide and its related substances.