ABSTRACT
A fast and reliable HPLC method is reported for the separation and quantification of betamethasone and prednisolone in urine samples using Chromolith Performance RP-18e [100 mm x 4.6 mm] column. The separation and detection was achieved using an isocratic mobile phase composed of methanol:water [44:56 v/v] at 2.0 mL/min and wavelength of 254 nm. After successful optimisation of method parameters, it was applied to the urine samples. Solid phase extraction technique was used to clean the sample before analysis. The developed method was validated for the system suitability, precision and accuracy. The limits of detection for the prednisolone and betamethasone are 0.11 ng and 0.075 ng/10 microL injection, respectively allowing their determination in human urine samples. Recovery for spiked urine samples was in the range of 97-103%. The method offers a valuable alternative to the methodologies currently employed for separation and quantification of prednisolone and betamethasone in urine samples