Stability Indicating Reverse Phase Ultra Performance Liquid Chromatography Method Development and Validation for Amiloride and Hydrochlorothiazide Tablet in the Presence of Degradation Products and Application to Green Analytical Principles

Abstract

Amiloride (AML) and hydrochlorothiazide (HCTZ) combination is used alone or with other medicines to treat high blood pressure (hypertension). The present article provides the development and validation of an reverse phase ultra performance liquid chromatography (RP-UPLC) approach for the quantification of AML, HCTZ, and related impurities in pharmaceutical dosage forms. The analytical column used for the separation of impurities is the Waters X bridge C18 3.5 μm (50 mm X 4.6 mm). Used 0.1% formic acid and ethanol as MP in an isocratic mode with 0.3 mL/min as flow rate and 3 μL as injection volume, at 254 nm as detection in UV and 12 minutes as total run time. The samples were prepared specifically to undergo forced degradation, which involved subjecting them to hydrolysis, oxidation, thermal, and photolytic conditions. The technique underwent validation by the principles set forth by the ICH Q2 guidelines. The validation confirmed that the method possesses specificity, linearity, ruggedness, robustness and accuracy. The methodology employed exhibited a linear relationship extending from 10 to 150% for all impurities. The recovery analysis was conducted throughout a range of concentrations, starting from 10 to 150% concentration. The average recovery value was determined to be within acceptable limits. The evidence of degradation and the findings of the verified study suggest that the nature of the subject under investigation is stable. Hence, this approach might be employed within the domains of pharmaceutical research and development and quality control departments. Green analytical chemistry tools are used to assess the method’s greenness and calculated using GAPI, AGREE and ecological-scale and found excellent green of >75%.