Characterization of Oligonucleotide Products and Impurities

This technical report outlines a method developed for characterizing oligonucleotide products and their impurities using Insight Biologics software. The software facilitates data processing for oligonucleotide therapeutics, particularly for sequence confirmation and impurity analysis. The report details the application of LC-MS/MS methods to analyze a crude phosphorothioate 20mer oligonucleotide sample and a purified version. It describes the workflow for target sequence confirmation and impurity profile analysis, including the use of high-resolution mass spectrometry and LC-UV data streams. The findings indicate that the software can provide a comprehensive list of impurity identifications along with sequence confirmation and abundance calculations. Additionally, the report discusses the importance of secondary purification steps for confirming impurities below a certain threshold. The results demonstrate the effectiveness of the method in detecting and quantifying impurities in oligonucleotide samples, emphasizing the analytical challenges posed by closely related structural impurities.