Development and Validation of a Novel Method for the Analysis of Impurities in Gemcitabine Hydrochloride Using RP-HPLC

Abstract

This study outlines a highly efficient method for estimating unknown impurities in Gemcitabine Hydrochloride API using gradient reverse-phase high-performance liquid chromatography (RP-HPLC). The mobile phase composition comprised 0.1% Orthophosphoric acid in water (A) and 100% Methanol (B) with a C18 (250 X 4.6) mm, 5µm stationary phase. The developed stability indicator by RP-HPLC utilized a gradient composition over 40 minutes at a 210 nm wavelength. The method successfully met validation parameters, demonstrating linearity, accuracy, and precision. This approach proves to be a straightforward, rapid, precise, accurate, and robust tool for estimating Gemcitabine Hydrochloride impurities.

Introduction:

This study addresses regulatory guidelines emphasizing impurity profiling for API, with a specific focus on Gemcitabine Hydrochloride. Gemcitabine is a widely-used antimetabolite drug in chemotherapy, necessitating rigorous impurity analysis due to its impact on drug quality and safety. Regulatory bodies such as ICH and FDA stress transparency in identifying impurities in APIs. The molecular structure of Gemcitabine Hydrochloride is presented.

Methods:

The analytical method involved HPLC with a C18 column, UV/PDA detector, and specific chromatographic conditions. Standard and impurity stock solutions were meticulously prepared. System suitability, specificity, linearity, and recovery were thoroughly assessed. Impurity identification, robustness, stability indication, and a comparison with a reference method were conducted.

Results and Discussion:

System suitability studies indicated the method's suitability, meeting all acceptance criteria. Impurity detection confirmed the presence of impurities in the Gemcitabine sample. The method demonstrated specificity with no observed interference. Linearity studies indicated a high correlation. Limit of Quantification (LOQ) and Limit of Detection (LOD) tests met acceptance criteria, with favourable recovery values. Impurity identification enhanced understanding of drug composition. Robustness testing showed the method's stability under varied conditions. Stability indication studies confirmed the accurate detection and quantification of Gemcitabine Hydrochloride and impurities over time.

Summary and Conclusion:

In response to limitations in existing monograph methods, this study developed a robust analytical method for Gemcitabine Hydrochloride API. The method's specificity, precision, and linearity were conclusively demonstrated through rigorous validation, surpassing the limitations of current monograph methods. This method aligns with international regulatory standards, offering a valuable tool for routine quality control in the pharmaceutical industry.