Analytical Assay Method Development and Validation of Itraconazole in Itraconazole Ointment By Reverse Phase-HPLC

This study presents the analytical assay method development and validation of Itraconazole in an ointment dosage formulation by using reverse phase HPLC. The assay method development was carried out by using C18 column. In the beginning, the study was carried on Itraconazole API by taking USP method as a reference. By changing few chromatographic parameters, a symmetrical peak and a satisfactory result could able to achieve. Changes done as per USP chapter “allowable adjustment to United States Pharmacopeia method”.  The aim was to achieve an advanced chromatographic conditions on HPLC system with C18 column (150 × 4.6 mm, 5µm particle size) using mobile phase composed of acetonitrile and neutral buffer. The separation was achieved at different gradient flows for alternative methods.  The ideal wavelength was calibrated by using PDA detector and the same wavelength was used for UV- visible detector. The assay method developed was also validated with full agreement with present regulatory guidelines by applying well developed analytical method validation techniques and means which includes the parameters such as linearity, accuracy, method precision, specificity with force degradation, robustness, solution stability, system suitability. The method shows linearity over a concentration range of 10µg/ml to 250µg/ml with r²=0.999 and thus this represents the method is efficient to provide good detector response. The lower limit of detection and quantification was achieved by carrying out the studies like LOD and LOQ and the results were found to be 10µg/ml and 5µg/ml respectively.