Analytical Method Validation on Estimation of Rotigotine in Pharmaceutical Formulations

A stability-indicating analytical method was developed for quantifying Rotigotine in a pharmaceutical microsphere formulation using Reversed-Phase High-Performance Liquid Chromatography (RP-HPLC), following the ICH guidelines. The analysis was carried out on a C8 column (150 mm × 4.6 mm; 5 µm particle size) with the column oven maintained at 40 °C. The mobile phase consisted of two components: Mobile Phase A - orthophosphoric acid and triethylamine in purified water (1:1:1000), and Mobile Phase B - acetonitrile, applied in gradient mode. The chromatographic conditions included a flow rate of 1.0 mL/min, a sample temperature of 25 °C, and a diluent comprising water, acetonitrile, and triethylamine (500:500:1). The injection volume was set at 20 µL, with ultraviolet-visible (UV-Vis) detection at a wavelength of 278 nm and a total run time of 15 minutes. All system suitability parameters met the requirements specified by the FDA. The precision of the method was demonstrated through six replicate preparations, yielding assay values ranging from 99.34% to 100.27%. The calibration curve, plotted across 80% to 120% of the target concentration, exhibited a coefficient of determination (R²) of 0.9998. Recovery studies within the same concentration range showed recoveries between 100.10% and 100.13%. The method showed a limit of detection (LOD) of 0.010 µg/mL and a limit of quantification (LOQ) of 0.035 µg/mL for Rotigotine. To verify that the assay functions as a true stability-indicating method, forced degradation experiments were carried out to evaluate the stability of Rotigotine within the pharmaceutical microsphere formulation.