A Stepwise Strategy Employing Automated Screening for Reversed-Phase Chromatographic Separation of Itraconazole and Its Impurities

Abstract

The development of methods for the chromatographic separation of complex mixtures is often challenging, and an organized approach is required to rapidly achieve the desired separation. Herein, software (S-Matrix Fusion)-assisted platform is used to effectively screen, optimize, and select optimal parameters for the separation of itraconazole and its 15 impurities. The employed platform exploits dissimilar chromatographic systems, uses a diverse set of columns with enormous selectivity differences, and evaluates a wide range of aqueous mobile phase pH and organic modifiers for screening experiments. The wide range of selectivity ensures the identification and resolution of critical analytes, while the combination of dissimilar systems and automated screening statistical software increases the speed, efficiency, and level of understanding of critical parameters influencing separation. Automated screening using the described platform allows the establishment of a mass-compatible ultra-high-performance liquid chromatography method capable of separating all impurities. Multivariate robustness of critical parameters such as buffer concentration, flow rate, column temperature, and gradient is evaluated to identify the operational range of the method. Moreover, the optimized method is validated following the International Conference on Harmonization (ICH) guidelines, evaluated in terms of specificity, analysis repeatability, intermediate precision, linearity, accuracy, limit of quantification, limit of detection, solution stability, and range, and concluded to be fit for purpose in the quality control laboratories for release, stability, and investigation purposes.