研究生: |
黃義澧 Huang, Yi-Li |
---|---|
論文名稱: |
利用LC-MS鑑定哺乳類聚醣中差異性鏈結單醣殘基 Identifying specific and differentially linked glycosyl residues in mammalian glycans by targeted LC-MS analysis |
指導教授: |
陳頌方
Chen, Sung-Fang |
學位類別: |
碩士 Master |
系所名稱: |
化學系 Department of Chemistry |
論文出版年: | 2017 |
畢業學年度: | 105 |
語文別: | 中文 |
論文頁數: | 61 |
中文關鍵詞: | 醣鏈結分析 、部分甲基化醣醇 、液相層析串聯式質譜 |
英文關鍵詞: | glycans linkage analysis, partially methylated alditol,, LC-MS |
DOI URL: | https://doi.org/10.6345/NTNU202202826 |
論文種類: | 學術論文 |
相關次數: | 點閱:87 下載:77 |
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聚醣是由數個單醣藉由糖苷鍵連結而成,由於醣蛋白中的聚醣結構具有高度複雜性,因此了解聚醣中單糖的組成與鍵結就成為重要的工作。在本實驗中開發一個方法,將聚醣經過一連串反應並水解形成 ”部份甲基化糖醇partially O-methylated alditols (PMAs)” 結合液相層析搭配串聯式質譜儀進行分析。實驗流程方面,取得聚醣後將聚醣進行還原,還原端的單醣會被還原而開環,接著進行聚醣的全甲基化,會將聚醣上所有free hydroxyl group的氫置換成甲基(CH3),甲基化聚醣以MALDI-TOF分析確認甲基化反應是否完全,最後進行聚醣的酸水解將聚醣的糖苷鍵破壞形成單醣,此時單醣在醣苷鍵位置會保留OH,最後將所有單醣進行還原得到目標分析物PMA。本實驗分析兩種聚醣的PMA,Fetuin N-glycan與Lewisa所產生的PMA都能經由LC-MS偵測。總而言之,本實驗可以鑑定出聚醣中不同鍵結的單醣,且此方法不須進行額外的衍生化實驗,與LC-MS平台有很好的相容性。
Glycan is a compound consisting a large number of monosaccharides linked glycosidically. Due to its high complexity of glycan structures on glycoproteins, assessing the configuration and position of glycosidic linkages of a glycan is in great demand. In this study, a method via partially O-methylated alditols (PMAs) from glycan combined with LC-MS analysis is developed. N-glycans were first per-methylated with methyl iodide, and levels of methylation were further confirmed with MALDI-TOF. PMAs were then produced via totally hydrolysis and reduction. PMAs from Fetuin N-glycan and Lewisa were successfully detected by LC-MS analysis. In conclusion, this proposed glycan linkage analysis can be performed without additional derivatization step for GC analysis, and should be suitable for the LC-MS-based platform.
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