Development and Validation of HPLC Method for the estimation of Triamcinolone Acetonide and 5-Fluorouracil

A sensitive HPLC method has been developed and validated for the determination of Triamcinolone Acetonide and 5-Fluorouracil in topical formulations. The developed method is found to be specific, reproducible and stability indicating. The Phenomenex C18, 250 × 4.6 mm, 5µm particle size column was used and mobile phase contains two solvents: Solvent A contains 10mM phosphate buffer pH 3.0 and solvent B was Acetonitrile. The detector linearity was established from concentrations ranging from 1 μg/ml to 10 μg/ml for both Triamcinolone Acetonide and 5-Fluorouracil with a correlation coefficient of 0.999. The limit of detection(LOD) and limit of quantification(LOQ) for Triamcinolone Acetonide was found to be 0.04 and 0.13 respectively and 5-Fluorouracil was found to be 0.01 and 0.03 respectively. The method was proved to be robust with respect to changes in flow rate, column temperature, mobile phase ratio and detector wavelength. Keywords: Triamcinolone Acetonide, 5-fluorouracil, Validation, HPLC method, topical formulations