Chromatographic Separations in Primary and Orthogonal Methods for Antifungal Drugs Using Statistical Approach

Abstract

Chromatographic separation of complex mixtures is often challenging, it needs an organized approach to accomplish the anticipated separation in a limited amount of time. The current work demonstrates an efficient strategy via software assisted method development using S-Matrix fusion software for screening, optimizing, and selecting the optimal parameters for the separation of three antifungal drugs Itraconazole, Ketoconazole and Econazole and its related impurities. The developed platform implements dissimilar chromatographic systems and adapts diverse set of columns with enormous selectivity differences, evaluating wide range of pH for the aqueous mobile phase and the organic modifiers for the screening experiments that offered variety in the selectivity to ensure all the components are revealed and resolved. Combination of these dissimilar systems in assistance with automated screening statistical software advances the speed, efficiency, and understanding of all the critical parameters influencing the separation. The outcome of the automated screening was a mass compatible Ultra high-performance liquid chromatography (UHPLC) stability indicating primary method that resolved all the impurities. Multivariate robustness for the critical parameters like buffer concentration, flow rate, column temperature, and gradient were performed to find the method operable design space. Moreover, specificity of the primary method was demonstrated with an orthogonal method to ensure there is no coelution of the impurities with the degradation compound and with the active peak of intrest. The work emphasizes and summarizes the need for an orthogonal method in chromatographic separations for pharmaceutical analysis. It enlists strategies that can be adopted for the development of an orthogonal method and the various advances in the metrics for orthogonality. Orthogonal approach with the use of Ecofriendly Supercritical fluid chromatography method to separate four stereoisomers of itraconazole stereoisomers was developed.