Determination of Potential Genotoxic Impurity, 5-Amino-2-Chloropyridine, in Active Pharmaceutical Ingredient Using the HPLC-UV System.

Abstract

A novel analytical method, based on high-performance liquid chromatography with a UV (HPLC-UV) detection system for the sensitive detection of a genotoxic impurity (GTI) 5-amino-2-chloropyridine (5A2Cl) in a model active pharmaceutical ingredient (API) tenoxicam (TNX), has been developed and validated. The HPLC-UV method was used for the determination of GTI 5A2Cl in API TNX. The compounds were separated using a mobile phase composed of water (pH 3 adjusted with orthophosphoric acid): MeOH, (50:50: v/v) on a C18 column (150×4.6mm i.d., 2.7μm) at a flow rate of 0.7mLmin-1. Detection was carried out in the 254nm wavelength. Column temperature was maintained at 40°C during the analyses and 10μL volume was injected into the HPLC-UV system. The method was validated in the range of 1-40μgmL-1. The obtained calibration curves for the GTI compound was found linear with equation, y=40766x-1125,6 (R2=0.999). The developed analytical method toward the target compounds was accurate, and the achieved limit of detection and limit of quantification values for the target compound 5A2Cl were 0.015 and 0.048μgmL-1, respectively. The recovery values were calculated and found to be between 98.80 and 100.03%. The developed RP-HPLC-UV analytical method in this research is accurate, precise, rapid, simple and appropriate for the sensitive analysis of target GTI 5A2Cl in model API TNX.