Automatic identification of oligonucleotide metabolites in complex biological samples using ultra high-performance liquid chromatography high-resolution mass spectrometry combined with the molecule profiler software

IF 2.8 3区医学 Q2 BIOCHEMICAL RESEARCH METHODS

Journal of Chromatography B Pub Date : 2025-02-04 DOI:10.1016/j.jchromb.2025.124505

Xinyuan Ye , Lili Jiang , Ying Xiong , Cui Zhang , Chenxi Wang , Meiling Qi , Chenyue Zhu , Yi Chen , Zhifeng Du , Zhongzhe Cheng , Hongliang Jiang

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引用次数: 0

Abstract

Oligonucleotide (ON) is one of the rapidly developing fields in biotherapeutics. Ultra high-performance liquid chromatography high-resolution mass spectrometry (UHPLC-HRMS) has been widely used for the identification of metabolites due to its high sensitivity, high resolution, and ability to provide structural information. The identification of ON metabolites in matrix has been reported by UHPLC-HRMS, however, manual data processing is time-consuming. In this study, an analytical strategy based on UHPLC-QTOF-MS/MS and molecule profiler software was established and employed for the automatic identification of metabolites of ONs. Fomivirsen (FMVS), a 21-mer antisense oligonucleotide with 20 phosphorothioate linkages, was selected as proof of concept. Firstly, the sample preparation and UHPLC-QTOF-MS/MS condition were optimized. Secondly, the feasibility of the automatic identification of ON metabolites by this strategy was verified using enzymatic digests of FMVS. Finally, in vitro and in vivo metabolites of FMVS were identified. As a result, the enzymatic digests of FMVS were successfully identified by the established strategy, and a total of 17 metabolites were identified from serum, plasma and tissues. FMVS was mainly metabolized by 3′-exonuclease in plasma, liver and kidney. This is the first study on metabolites identification of FMVS, and the proposed strategy would simplify the identification of ON metabolites, thus could be used for other ON metabolites.

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来源期刊

Journal of Chromatography B 医学-分析化学

CiteScore

5.60

自引率

3.30%

发文量

306

审稿时长

44 days

期刊介绍： The Journal of Chromatography B publishes papers on developments in separation science relevant to biology and biomedical research including both fundamental advances and applications. Analytical techniques which may be considered include the various facets of chromatography, electrophoresis and related methods, affinity and immunoaffinity-based methodologies, hyphenated and other multi-dimensional techniques, and microanalytical approaches. The journal also considers articles reporting developments in sample preparation, detection techniques including mass spectrometry, and data handling and analysis. Developments related to preparative separations for the isolation and purification of components of biological systems may be published, including chromatographic and electrophoretic methods, affinity separations, field flow fractionation and other preparative approaches. Applications to the analysis of biological systems and samples will be considered when the analytical science contains a significant element of novelty, e.g. a new approach to the separation of a compound, novel combination of analytical techniques, or significantly improved analytical performance.