DEVELOPMENT AND VALIDATION OF AN RP-HPLC METHOD FOR THE DETERMINATION OF CHLORHEXIDINE AND P-CHLOROANILINE IN VARIOUS PHARMACEUTICAL FORMULATIONS

Abstract

□ A simple, rapid and sensitive isocratic reversed-phase (RP) high-performance liquid chromatography (HPLC) method was developed and validated for the simultaneous determination of chlorhexidine (CHX) and p-chloroaniline (CAL) in various pharmaceutical formulations. Compound separation was achieved in less than 10 min with an XBridge C18 column that was maintained at 40°C and a mobile phase consisting of 32:68 (v/v) of acetonitrile and a pH 3.0 phosphate buffer solution (a 0.05 M monobasic sodium phosphate solution containing 0.2% of triethylamine). Analyses were performed at a flow rate of 2 mL min−1 and at a detection wavelength of 239 nm. The method was shown to be selective, linear, accurate, and precise in intra-day and inter-day analyses. The robustness of the method was shown by slightly changing the flow rate, column oven temperature, and proportion of acetonitrile in the mobile phase. The method was found, however, to be very sensitive to the pH of the mobile phase buffer. The method was successfully validated following the guidelines of the International Conference on Harmonization (ICH). This validation proved that the method was suitable for the determination of CHX and CAL in toothpaste, mouthwash, wound cleanser, and skin and hand disinfectants.