Analytical Method Development, Validation and Optimization of Fluconazole Drug Using RP- HPLC

This study reported the development and validation of a simple, precise, and cost-effective Reverse Phase High-Performance Liquid Chromatography (RP-HPLC) method for the quantitative estimation of Fluconazole. Method development was systematically performed through nine optimization trials by varying the mobile phase composition, flow rate, and column type to achieve a sharp, symmetrical, and well-resolved peak with minimal tailing. The final optimized chromatographic conditions comprised a Kromasil C18 column (250 mm × 4.6 mm, 5 µm) with a mobile phase of Water: Acetonitrile (60:40 v/v) at a flow rate of 1.5 mL/min, which yielded a retention time of 2.231 minutes. The method exhibited excellent specificity, with no interfering peaks observed at the retention time of Fluconazole. Validation was performed in accordance with ICH guidelines, confirming the method’s reliability for routine quality control applications. System suitability parameters, including tailing factor, theoretical plates, and %RSD, were found to be within acceptable limits. The method demonstrated excellent linearity across 20–150% of the target concentration (r² = 0.999), along with high precision and accuracy. The assay of the drug substance was determined to be 99.49%, which was within the acceptable range. Overall, the developed RP-HPLC method proved to be simple, rapid, accurate, precise, robust, sensitive, and specific, making it suitable for the routine estimation of Fluconazole in pharmaceutical formulations.