Ecofriendly Stability Indicating RP-UHPLC Method for Simultaneous Estimation of Amiloride and Hydrochlorothiazide in Bulk and pharmaceutical dosage form using AGREE and GAPI Tools

An easy, quick and reliable RP-UHPLC technique for the quantification of Amiloride (AML) and Hydrochlorothiazide (HCTZ) is established as well as validated in the present article using the green analytical chemistry (GAC) tools. The mobile phase consisted of a mixture of 0.1% formic acid with ethanol in a 55:45 % v/v ratio delivered at a flow rate of 0.5 mL/min through a 120x4.6mm Hypersil gold ODS endcapped reverse phase column having a particle size of 3 micron. AML and HCTZ were detected at 265 nm with respective retention periods of 2.982 and 5.654 min. This methodology was rigorously validated in alignment with ICH guidelines. The calibration plots exhibited linearity over the concentration ranges of 2.5–25 μg/mL for AML and 25-150 μg/mL for HCTZ. The detection limits for AML and HCTZ were 0.69 and 0.61 μg/mL whereas quantification limits were 2.10 and 1.84 μg/mL. At three different levels of a standard addition, the percentage recovery was determined to be between 99.37 and 100.7%. The precision of the method for both intra as well as inter-day fell under the acceptable range (RSD < 2). Both the bulk medication as well as its formulation were subjected to a forced degradation trial and the results indicate that the technique is stability indicating and can differentiate between the peaks of the active analytes and the degradants. The evaluation of the method’s environmental impact utilized green analytical chemistry tools such as GAPI and AGREE revealing a greenness rate exceeding 75%, which is excellent. Hence, the established chromatographic approach was straightforward, highly sensitive, quick, precise, cost effective as well as eco-friendly than the earlier reported methods making it suitable for routine quality control testing in industrial sector.