A methylation-assisted gas chromatography-mass spectrometry strategy for comprehensive monosaccharide profiling in Polygonatum polysaccharides: Overcoming limitations of conventional chromatographic techniques
Liang Xu , Jinjian Yao , Zhen Wang , Songzi Xie , Jinmei Ou , Bin Wang
{"title":"A methylation-assisted gas chromatography-mass spectrometry strategy for comprehensive monosaccharide profiling in Polygonatum polysaccharides: Overcoming limitations of conventional chromatographic techniques","authors":"Liang Xu , Jinjian Yao , Zhen Wang , Songzi Xie , Jinmei Ou , Bin Wang","doi":"10.1016/j.chroma.2025.466253","DOIUrl":null,"url":null,"abstract":"<div><div>Conventional methods for analyzing monosaccharides, such as 1-Phenyl-3-methyl-5-pyrazolone (PMP) derivatization coupled with high-performance liquid chromatography (HPLC), struggle to detect ketoses and acidic sugars. This limitation hinders the precise structural analysis of polysaccharides in traditional Chinese medicine (TCM). A methylation-assisted Gas Chromatography-Mass Spectrometry (GC–MS) approach was developed to address this issue in the current study. By optimizing the methylation process and using GC–MS with a BR-17 column and an Electron Ionization (EI) source, simultaneous detection of ten different monosaccharides was achieved within 20 min. GC–MS profiling of monosaccharides generally gives two peaks per analyte, except in the cases of fructose and galacturonic acid. Notably, fructose (Fru) was identified for the first time in the polysaccharides of <em>Polygonatum</em> species: <em>P. sibiricum</em> at 18.55 μg/mg, <em>P. cyrtonema</em> at 16.90 μg/mg, and <em>P. kingianum</em> at 14.35 μg/mg, overcoming the limitations of the inability to detect ketoses in the PMP-HPLC method. Data analysis revealed distinct species-specific markers: <em>P. kingianum</em> polysaccharides (PKP) uniquely contained <em>α</em>-galacturonic acid (α-Gal: 19.95 μg/mg), whereas <em>P. sibiricum</em> polysaccharides (PSP) featured six monosaccharides, including glucose (Glc: 135.45 μg/mg), mannose (Man: 78.95 μg/mg), and ribose (Rib: 26.90 μg/mg), with a total of 299.50 μg/mg. <em>P. cyrtonema</em> polysaccharides (PCP) showed elevated levels of glucuronic acid (GlcA, 38.40 μg/mg). This derivatization-enhanced GC–MS method effectively fingerprints TCM polysaccharides, enabling clear differentiation of <em>Polygonatum</em> species through analysis of monosaccharide composition and total sugar content. This approach establishes clear structural distinctions, overcoming the limitations of conventional chromatographic techniques and enhancing the quality control and authentication of TCM polysaccharides.</div></div>","PeriodicalId":347,"journal":{"name":"Journal of Chromatography A","volume":"1759 ","pages":"Article 466253"},"PeriodicalIF":4.0000,"publicationDate":"2025-07-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Journal of Chromatography A","FirstCategoryId":"1","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S0021967325005989","RegionNum":2,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"BIOCHEMICAL RESEARCH METHODS","Score":null,"Total":0}
引用次数: 0
Abstract
Conventional methods for analyzing monosaccharides, such as 1-Phenyl-3-methyl-5-pyrazolone (PMP) derivatization coupled with high-performance liquid chromatography (HPLC), struggle to detect ketoses and acidic sugars. This limitation hinders the precise structural analysis of polysaccharides in traditional Chinese medicine (TCM). A methylation-assisted Gas Chromatography-Mass Spectrometry (GC–MS) approach was developed to address this issue in the current study. By optimizing the methylation process and using GC–MS with a BR-17 column and an Electron Ionization (EI) source, simultaneous detection of ten different monosaccharides was achieved within 20 min. GC–MS profiling of monosaccharides generally gives two peaks per analyte, except in the cases of fructose and galacturonic acid. Notably, fructose (Fru) was identified for the first time in the polysaccharides of Polygonatum species: P. sibiricum at 18.55 μg/mg, P. cyrtonema at 16.90 μg/mg, and P. kingianum at 14.35 μg/mg, overcoming the limitations of the inability to detect ketoses in the PMP-HPLC method. Data analysis revealed distinct species-specific markers: P. kingianum polysaccharides (PKP) uniquely contained α-galacturonic acid (α-Gal: 19.95 μg/mg), whereas P. sibiricum polysaccharides (PSP) featured six monosaccharides, including glucose (Glc: 135.45 μg/mg), mannose (Man: 78.95 μg/mg), and ribose (Rib: 26.90 μg/mg), with a total of 299.50 μg/mg. P. cyrtonema polysaccharides (PCP) showed elevated levels of glucuronic acid (GlcA, 38.40 μg/mg). This derivatization-enhanced GC–MS method effectively fingerprints TCM polysaccharides, enabling clear differentiation of Polygonatum species through analysis of monosaccharide composition and total sugar content. This approach establishes clear structural distinctions, overcoming the limitations of conventional chromatographic techniques and enhancing the quality control and authentication of TCM polysaccharides.
期刊介绍:
The Journal of Chromatography A provides a forum for the publication of original research and critical reviews on all aspects of fundamental and applied separation science. The scope of the journal includes chromatography and related techniques, electromigration techniques (e.g. electrophoresis, electrochromatography), hyphenated and other multi-dimensional techniques, sample preparation, and detection methods such as mass spectrometry. Contributions consist mainly of research papers dealing with the theory of separation methods, instrumental developments and analytical and preparative applications of general interest.