Abstract
Simple and precise spectrophotometric methods for quantitative assay of a mixture of hydrocortisone acetate (HCA) and clioquinol (CL) were developed and validated through different mathematical manipulation pathways. The developed methods utilized ratio spectra for resolving binary mixtures including absorbance subtraction, ratio subtraction coupled with spectrum subtraction, constant multiplication, constant value, and derivative ratio. The proposed methods were proved to be specific by analysing the laboratory-prepared mixtures and were applied for the assay of topical preparation successfully. The methods were validated using ICH guidelines where accuracy, repeatability and intermediate precision were within the acceptable limits. The linearity range was found to be 2-22 for HCA and 1.5-7 µg/mL for CL in all proposed methods and 2-7 µg/mL for HCA and CL in absorbance subtraction method through using a unified regression equation. The findings were statistically evaluated with respect to the official and reported methods, demonstrating that there was no significant difference.
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