Monitoring polymerization events leading to the discovery of new high-molecular weight (MW) impurities is challenging during chemical syntheses of active pharmaceutical ingredients. Employing reversed-phase chromatography (RPC) stationary phases (SPs) in size-exclusion chromatography (SEC) mode could be a potential solution given their high efficiency, sensitivity, and extensive solvent compatibility. However, there is a lack of generalized means for trace polymeric impurities across a wide range of physicochemical properties. Herein, we developed a SEC-based approach with a C18 SP for screening such high-MW impurities. Seven polymer standards presenting a variety of functional groups, consisting of hydrophobic, heterocyclic, ionic, and neutral hydrophilic moieties, were utilized as model impurities to establish the screening conditions. Nine mobile phases (tetrahydrofuran-based, buffered methanol, and buffered acetonitrile) were proposed to cover all model polymers and a majority of potential high-MW impurities in small molecule chemical syntheses. The established screening system demonstrated a linearity of 0.05-1.0 % w/w (R2>0.99) for the selected model impurities with proper elution conditions. Two real high-MW impurities, BMT-041910 (polymeric degradation) and poly(phenyl thiirane) (by-product polymerization), were identified from the proposed high-MW impurity screening. The successful conditions yielded a quantitative limit better than 0.1 % w/w in both cases. We believe the developed screening platform is applicable to the analysis of a wide variety of unknown high-MW impurities of low abundance potentially generated during drug substance development.
Keywords: C18 stationary phase; Pharmaceutical high-molecular weight impurity; Screening platform; Size-exclusion chromatography.
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