Stability-Indicating HILIC Method for the Determination of Citicoline and Characterization of its Degradation Products by LC–MS/TOF, 1H and 13C NMR

Abstract

A stability-indicating HILIC method was developed for analysis of Citicoline in the presence of its degradation products. It was performed using Atlantis HILIC Si column (50 × 4.6 mm, 3 µm particle size) and a mobile phase constituted by 70% of acetonitrile in formate buffer 0.02 M (v/v) adjusted to pH 3. The column’s thermostat compartment was adjusted at 30 °C. Citicoline was subjected to oxidative, acidic, basic, hydrolytic, thermal and UV light stress conditions. The column effluents were monitored at 270 nm by photodiode array and mass detectors. Depending on the stress, three degradation products appeared; one after acidic or photolytic action, the second by NaOH reaction and the third with oxidative stress. LC–ESI–MS/TOF, 1H and 13C NMR were used to characterize them. The most probable formation mechanisms of the identified impurities were proposed. The developed method was validated in terms of specificity, limit of detection and quantification, linearity, accuracy and precision as per ICH guidelines. The proposed method was successfully applied for the determination of Citicoline in dosage forms.