DOI: 10.1002/rcm.70126 ISSN: 0951-4198

Determination of Residual Nitrilase in Brivaracetam Using Characteristic Peptides by Liquid Chromatography–Mass Spectrometry

Xueting Dai, Kongliang Xu, Tianji Zhang, Ming Li, Peize Wu, Jinlan Cheng, Hanrui Chen
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ABSTRACT

Rationale

Nitrilases are widely used as biocatalysts in pharmaceutical manufacturing because of their high stereoselectivity and mild reaction conditions. However, residual nitrilase in final drug products may compromise product quality and safety, highlighting the need for sensitive and reliable analytical methods for its monitoring and control.

Methods

In this study, a liquid chromatography–tandem mass spectrometry (LC–MS/MS) method was developed for the determination of residual nitrilase in pharmaceutical formulations.

Results

Recombinant nitrilase expressed in Escherichia coli was purified by Ni 2+ affinity chromatography and lyophilized prior to analysis. Following systematic optimization of digestion and chromatographic conditions, the method achieved a detection limit of 0.17 μg/g. The LC–MS/MS method was successfully applied to the analysis of brivaracetam samples at a concentration of 2000 μg/g, with residual nitrilase levels consistently below 85 μg/g, meeting relevant quality control requirements.

Conclusions

In this study, a sensitive and reliable LC–MS/MS method was developed for the determination of residual nitrilase in brivaracetam. The proposed method may serve as a general analytical strategy for quality control of nitrilase‐related pharmaceutical products.

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