Current trends in liquid chromatography-mass spectrometry analysis of small interfering RNAs: A short review

Togawa, Hiroyuki; Yamaguchi, Takao; Kawakami, Junji et al.

Journal of Chromatography A, 2026, 1766, 466550

Number of Access:212（2026-05-21 21:16 Counts）

Identifier to cite or link to this item: https://hdl.handle.net/11094/103568

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Title
Current trends in liquid chromatography-mass spectrometry analysis of small interfering RNAs: A short review
Creator
CreatorName
Togawa, Hiroyuki
NRID
1000080596601 1000080596601
ORCID
0000-0003-3180-0257 0000-0003-3180-0257
researchmap permalink
takao-yamaguchi takao-yamaguchi
Scopus Author ID
56322895500 56322895500
CreatorName
Yamaguchi, Takao
NRID
1000040289012 1000040289012
CreatorName
Kawakami, Junji
NRID
1000080243252 1000080243252
ORCID
0000-0002-6842-6812 0000-0002-6842-6812
researchmap permalink
read0014368 read0014368
Scopus Author ID
7003800550 7003800550
CreatorName
Obika, Satoshi
Subject
Liquid chromatography
Mass spectrometry
siRNA
Description
Togawa H., Yamaguchi T., Kawakami J., et al. Current trends in liquid chromatography-mass spectrometry analysis of small interfering RNAs: A short review. Journal of Chromatography A 1766, 466550 (2026); https://doi.org/10.1016/j.chroma.2025.466550.
Abstract
Small interfering RNA (siRNA) therapeutics represent one of the most rapidly advancing pharmaceutical modalities. Since the approval of the first siRNA-based drug in 2018, seven siRNA therapeutics have been approved to date. During manufacturing, “non-optimal duplexes” and “single-stranded impurities” can arise from synthetic byproducts such as shortmers and longmers, or from minor imbalances in strand quantification and mixing. Furthermore, phosphorothioate linkages introduce chirality, generating complex diastereomeric mixtures. The use of delivery systems, including lipid nanoparticles or N-acetylgalactosamine (GalNAc) conjugates, further increases the structural complexity and heterogeneity of the final product. Compared to single-stranded oligonucleotides, siRNA—being double-stranded—yields more complex chromatographic and mass spectral profiles, making analytical characterization particularly challenging. Therefore, the development of robust and reliable analytical methods is essential to ensure the quality of siRNA therapeutics. This review summarizes key analytical techniques for the quality evaluation of siRNA therapeutics. Liquid chromatography (LC) is one of the most widely used approaches, with separation modes such as ion-pair reversed-phase (IP-RP), anion exchange (AEX), size exclusion chromatography (SEC), and hydrophilic interaction chromatography (HILIC) being actively explored. Mass spectrometry (MS) is another powerful tool that provides molecular mass-based structural information not accessible via UV detection alone. The combination of LC and MS offers a highly effective platform for characterizing siRNA therapeutics, enabling both qualitative and quantitative assessment of impurities and isomers. Furthermore, tandem MS (MS/MS) can provide sequence-specific information for the identification of active pharmaceutical ingredients. In addition to quality assessment, the application of LC and MS techniques in the pharmacokinetic analysis of siRNA therapeutics is also briefly discussed.
Publisher
Elsevier B.V.
Source Title
Journal of Chromatography A
Volume (Issue)
1766
Page
466550
Date of Issued
2026-01-11
Language
English
Handle URL
https://hdl.handle.net/11094/103568
PISSN
00219673
eISSN
18733778
Relation.isIdenticalTo
DOI (Publisher Version)
https://doi.org/10.1016/j.chroma.2025.466550
Access Rights
open access
CopyRight
This article is licensed under a Creative Commons Attribution 4.0 International License.
oaire:version
Version of Record
Category
学術雑誌論文 Journal Article