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Simultaneous separation of small interfering RNA and lipids using ion-pair reversed-phase liquid chromatography
Journal article   Peer reviewed

Simultaneous separation of small interfering RNA and lipids using ion-pair reversed-phase liquid chromatography

Li Li, Joe P Foley and Roy Helmy
Journal of chromatography. A, v 1601, pp 145-154
13 Sep 2019
DOI: https://doi.org/10.1016/j.chroma.2019.04.061
PMID: 31072602
Featured in Collection :   UN Sustainable Development Goals @ Drexel

Abstract

Chemistry Techniques, Analytical - methods Chromatography, Reverse-Phase Drug Stability Lipids - isolation & purification Nanoparticles - chemistry RNA Interference RNA, Small Interfering - isolation & purification
RNA interference offers a novel approach for the development of new therapeutics for targets that are otherwise "undruggable" using traditional modalities. The safety and efficacy of siRNA-based therapy mainly rely on lipid or polymer-based nanocarriers to overcome inherent barriers to a systemic delivery of siRNA. A multicomponent lipid nanoparticle (LNP) system is a promising delivery platform, typically consisting of a cationic lipid, phospholipid, PEG-containing short-chain lipid, and cholesterol. Characterization and chemical analysis of the LNP formulation is important to assure drug product stability, a key consideration for chemistry, manufacturing and control strategy. Here we report an ion-pair reversed phase UHPLC method capable of simultaneously separating both siRNA and functional lipids in LNPs with a minimal retention gap for two classes of biologically essential yet chemically distinct molecules. Key chromatographic parameters critical to the separation are discussed, including the structure of the ion-pair agent, stationary phase chemistry, column temperature and an organic additive. The results showed that the retention time of siRNA is tunable by using various ion-pair reagents. The retention factor of the siRNA exhibited a first order relationship with the number of carbons in the alkyl chain of the ion-pair reagents. In contrast, the type of ion-pair reagent has no significant impact on the separation of phospholipids. Separations using a BEH phenyl column and dibutylammonium acetate as the ion-pair reagent showed satisfactory selectivity for a range of double-stranded siRNAs and phospholipids, key components for lipid nanoparticle formulations. Furthermore, the method was applied to the separation of an experimental LNP formulation, demonstrating good selectivity for siRNA, functional lipids and their potential degradation products.

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Collaboration types
Industry collaboration
Domestic collaboration
Web of Science research areas
Biochemical Research Methods
Chemistry, Analytical
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