Development of a Reliable Purity Method for Ataluren in a Sachet Formulation Through the Analysis of Its Stress Degradation Behavior Using HPLC, LC‐MS, and 2D‐NMR

ABSTRACT

Ataluren is a medication commonly used for Duchenne muscular dystrophy. Currently, it is available in granules for oral suspension. A unique reverse‐phase HPLC approach with a UV detector was established to detect and quantify ataluren and its degradation components in medicinal products. This technique utilized a phenyl hexyl stationary phase column with a mixture of Eluent‐A (0.03% trifluoroacetic acid) and Eluent‐B (acetonitrile and methanol—20:80 %v/v) as the mobile phase. To ensure ataluren's inherent stability, it underwent rigorous stress testing in a wide range of degrading circumstances, such as aqueous, acidic, alkaline hydrolysis, humidity, thermal, oxidative, and photolytic environments. The medication was determined to be degradable in both methanolic acidic and alkaline environments, revealing the presence of two distinct degradation products. The fractions of degradation products were collected and purified using flash chromatography. The structures were confirmed using 2D‐NMR and LC‐MS. A resolution above 2.0 was achieved, effectively separating all components of the degradation. The developed approach is highly sensitive, enabling the quantification of ataluren and its degradation products at concentrations as low as 0.04% of the test concentration. The approach demonstrated high precision, linearity, accuracy, and specificity following its development and rigorous testing. It also fulfilled all the relevant regulatory requirements.