New Stability Indicating RP‐HPLC Methods for the Impurity Profiling of Amifampridine Phosphate

This article presents the development and validation of two new stability‐indicating RP‐HPLC methods for the determination of two related substances (RS) and the assay of amifampridine phosphate (AMP) via forced degradation and mass balance studies. In RS method, AMP was resolved adequately from its two RS, namely, 4‐aminopyridine and 3,4,5‐triaminopyridine on YMC Triart C18 (250 × 4.6 mm, 3 µm) column. Ammonium acetate solution (2.3 g/L) with its pH adjusted to 9.0 ± 0.05 was used as buffer. Gradient elution was implemented with the use of mobile phase A (buffer) and mobile phase B (buffer and acetonitrile in 80:20 v/v). The flow rate, injection volume, column temperature, and wavelengths were maintained as 0.7 mL/min, 20 µL, 40°C, and 240 nm, respectively. Additionally, an assay method for AMP was also developed using the same column. Both methods underwent validation in accordance with ICH guidelines. In the RS method, the limit of detection and limit of quantitation values were 0.010% and 0.031%, respectively. The correlation coefficient values exceeded 0.995 in both methods. The recoveries ranged from 93.5% to 100.7% for the RS method and from 100.1% to 101.1% for the AMP assay method.