Identification and characterization of related substances in pomalidomide by hyphenated LC–MS techniques

•13 related substances (RSs) were characterized in pomalidomide (PML).•Incorporation of LC–TOF–MS, LC–MS–MS and NMR to characterize these RSs.•The forced degradation study was performed.•Most of these RSs were characterized as the hydroxylation products of PML.

The current study dealt with the separation, identification and characterization of related substances in pomalidomide by hyphenated techniques. Complete separation was obtained with an Inertsil ODS-SP column (250 mm × 4.6 mm, 5 μm) by linear gradient elution using a mobile phase consisting of 0.1% formic acid solution and acetonitrile. They were characterized by hyphenated chromatographic techniques with the accurate mass determination using high resolution LC–TOF–MS methods as well as the product MS spectra determination and elucidation. The degradation behaviors of pomalidomide under ICH prescribed stress conditions were also conducted. Pomalidomide was found to be labile to degrade under acid, alkaline, oxidative and thermal stress conditions, while it was relatively stable to photolytic stress. 13 related substances were detected and identified to be 10 degradation products and three process related substances. The hyphenated LC–MS method with high resolution accurate mass determination facilitated the qualitative analysis of the unknown compounds than that of the conventional HPLC–UV. The related compounds identified are valuable for pomalidomide manufacturing process optimization and quality control.

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