Stability-indicating assay method for acotiamide: Separation, identification and characterization of its hydroxylated and hydrolytic degradation products along with a process-related impurity by ultra-high-performance liquid chromatography/electrospray ionization quadrupole time-of-flight tandem mass spectrometry.

RATIONALE: The presence of impurities and degradation products will affect the pharmacokinetic, pharmacodynamic properties and alter the safety of a drug. Hence, the development of a stability-indicating assay method is an integral part of quality product development and is crucial for the regulatory approval of drug products.

METHODS: Acotiamide was subjected to stress degradation under hydrolytic, oxidative, photo and thermal stress conditions. The resulted degradation products (DPs), as well as a process-related impurity (IMP), were selectively separated from the drug on a Waters Acquity HSS cyano column (100 × 2.1 mm, 1.8 μm) with a mobile phase containing a gradient mixture of 0.1% formic acid and acetonitrile (ACN) at a flow rate of 0.25 mL min-1 .

RESULTS: The drug was found to degrade under hydrolytic (acidic and basic), oxidative and photolytic stress while it remained stable under neutral hydrolytic and thermal stress conditions. The seven degradation products (DPs) and one process-related impurity (IMP) were observed. All the DPs and process-related IMP were well separated by the developed ultra-high-performance liquid chromatography (UHPLC) method and subsequently characterized by UHPLC/electrospray ionization quadrupole time-of-flight tandem mass spectrometry (ESI-QTOF-MS/MS). The proposed UHPLC method was validated with respect to specificity, linearity, accuracy, precision and robustness as per ICH guideline, Q2 (R1).

CONCLUSIONS: All the observed DPs were new and formed by hydrolysis of an amide bond, phenyl ring hydroxylation and hydrolysis of the methoxy group of the phenyl ring. The despropyl process-related impurity was observed and well separated from the drug. The proposed UHPLC mass spectrometric method has greater utility in the identification of DPs in much less time with excellent selectivity.