Enrichment and recovery of oligonucleotide impurities by N-Rich twin-column continuous chromatography

https://doi.org/10.1016/j.jchromb.2022.123439Get rights and content
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Highlights

  • Isolation of oligonucleotide impurities by N-Rich, a twin-column chromatography process.

  • N-Rich process principles and experimental design.

  • Broader accumulation of several impurities simultaneously.

  • Process comparison for the collection of 1 mg oligonucleotide impurity via N-Rich, preparative chromatography and HPLC.

Abstract

N-Rich is a twin-column continuous chromatography technology well suited for small-scale isolation and the enrichment of product related impurities. For the first time, N-Rich was used for impurity isolation from a double-stranded RNA (dsRNA) therapeutic synthetic oligonucleotide (ON), produced by solid-phase synthesis. By employing the N-Rich process, where the desired impurities are recycled and selectively enriched, and interfering substances are depleted, it was possible to obtain substantial amounts of high purity marginal impurities with a reproducible, automatized, and productive method. The productivity-purity tradeoff inherent to traditional impurity isolation methods, i.e., analytical chromatography, was effectively alleviated. Using N-Rich, satisfactory purity values and mass recoveries of several low-concentrated impurities could be obtained simultaneously.

A performance comparison demonstrated an up to 15-fold increase for purity values and up to 20-fold mass impurity isolation and concentration with the N-Rich technology in comparison to conventional isolation procedures, drastically reducing processing times, manual handling, and waste production.

Keywords

Oligonucleotide impurities
Analytical characterization
Continuous chromatography
Preparative chromatography
N-Rich

Data availability

Data will be made available on request.

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