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  • 學位論文

硼酸緩衝溶液毛細電泳質譜分析雙糖異構物靈敏度提升之探討

Sensitivity Improvement of disaccharide analysis by Capillary Electrophoresis/ Electrospray Ionization Mass Spectrometry Using Nonvolatile Borate Buffer

指導教授 : 何國榮

摘要


硼酸於雙糖異構物的毛細電泳分離佔據非常重要的角色,因此在雙糖的分離上,硼酸根離子是不可或缺的。質譜偵測雙糖時,為了提升其質譜靈敏度需將雙糖進行衍生化,衍生化的方式可分為閉環標記與開環標記(還原胺化)。由於閉環標記可保有分析結構資訊的優點,因此我們首先嘗試以閉環標記鍵結/組成異構物,然而此條件下無法達成有效分離。當以開環標記時,實驗結果發現可增加其分離解析度,然而卻喪失分析物的結構資訊。對於組態/組成異構物的分離時,閉環標記可提供基線分離且保有結構資訊。因此在本研究中,鍵結異構物以開環標記而組態異構物以閉環標記。本研究中以銨鹽取代鈉鹽以減少鈉離子對分析物的訊號抑制,並搭配實驗室先前開發的低流速鞘流毛細電泳/質譜界面。我們在實驗結果中發現,以低流速鞘流界面分析雙糖異構物時,其靈敏度相較於之前使用的傳統界面有明顯的提升(分析物濃度下降20倍)。原因為低流速鞘流界面中使用小口徑的電灑噴頭,具有低稀釋倍數、小口徑較抗鹽等特質。為了更進一步增加雙糖的靈敏度,我們嘗試以液體接合式-低流速鞘流界面以減少硼酸根離子的訊號。我們結合毛細管塗佈與添加有機相的方式使硼酸根離子的電泳淌度小於銜接管中的電滲流淌度,因此硼酸根會留在液體接合槽中,藉此減少分析物的訊號抑制而達成靈敏度的提升。實驗結果發現,使用液體接合式-低流速鞘流界面相較於低流速鞘流界面對於雙糖的分析可達到一個數量級靈敏度的提升。

並列摘要


Due to the interaction between borate anion and diols of the disaccharides, borate became a popular buffer in the analysis of disaccharides especially for isomer separation. Borate buffer offers good separation efficiency, however, it was found to be incompatible with CE-ESI-MS due to ion suppression. Using ammonium salt instead of sodium salt could improve sensitivity; however, the problem of borate anion suppression still exists. In order to alleviate the problem resulting from non-volatile borate buffer, capillary coating was employed to reduce the electroosmotic flow (EOF). Under a low EOF, borate anion stays on liquid junction reservoir. However, analytes still migrate toward the ESI source, so borate would not interfere with the signal generated by analytes. This strategy was applied to a liquid junction-low flow interface. With the proposed approach, which maintaining a good separation efficiency, the interference due to borate can be reduced.

並列關鍵字

disaccharide CE-MS Borate Capillary coating

參考文獻


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