Validation of a Stability-Indicating Assay of Amprolium Hydrochloride in Water Soluble Powder Formulation using Hydrophilic Interaction Liquid Chromatography

A simple stability-indicating method was developed and validated to determine amprolium hydrochloride (HCl) in water soluble powder formulation. The method is based on zwitterionic hydrophilic interaction liquid chromatography (ZIC-HILIC) coupled with photo diode array detector (PDA). The separation was achieved on ZIC-HILIC column (250 mm × 4.6 mm, 5 µm) at 25°C temperature. The optimized mobile phase consisted of an isocratic solvent mixture of 200mM Ammonium acetate (NH4AC) solution and acetonitrile (ACN) (25:75; v/v) with pH adjusted to 5.7 by glacial acetic acid. The mobile phase flow was fixed at 0.5 ml/min and the analytes were monitored at 267 nm. The effects of the chromatographic conditions on the peak retention, peak USP tailing factor and column efficiency were systematically optimized. Forced degradation experiments were carried out by exposing amprolium HCl standard and the water soluble powder formulation for thermal, photolytic, oxidative and acid-base hydrolytic stress conditions. The degradation products were well-resolved from the main peak and the excipients thus proving that the method is a reliable and stability-indicating. The method was validated as per ICH and USP guidelines and found to be adequate for the routine quantitative determination of amprolium hydrochloride in commercially available water soluble powder dosage forms.