Liquid chromatographic method development for quantification of inorganic nitrite and nitrate impurities from nitroglycerin drug substance by using ion-pair reagents with liquid-liquid extraction technique

Abstract

A large number of laboratory studies have reported Nitrite (NO2−) and Nitrate (NO3−) to be among the most common degradation products of the high-explosive Nitroglycerin drug substance. A novel, simple, robust and rapid reversed-phase high-performance liquid chromatography method has been developed for quantification of inorganic Nitrite and Nitrate impurities from Nitroglycerin drug substance. Successful separation was achieved in isocratic elution, using Inertsil C8-3, (250 × 4.6 mm, 5.0 μm) column, with mobile phase consisting of pH 7.0 tetrabutyl ammonium hydrogen sulfate buffer, methanol and acetonitrile (96:02:02, v/v/v). Flow rate was monitored at 2.0 mL min−1 and ultraviolet detection at 220 nm. The present work describes the role of an ion-pair reagent in the separation of polar compounds and liquid-liquid extraction technique for separation of polar and non-polar compounds. Nitroglycerin was subjected to various stress conditions to demonstrate the stability-indicating power of the method. The performance of the method was validated as per present International Council for Harmonisation (ICH) guidelines for specificity, linearity, accuracy, precision, ruggedness and robustness. The developed method can be a valuable alternative to the current ion-exchange chromatographic method mentioned in the literature. To the best of our knowledge, a rapid Liquid Chromatography (LC) method, which separates inorganic Nitrite and Nitrate impurities of Nitroglycerin, disclosed in this investigation was not published elsewhere.