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標題: | 支撐性離子液體於毛細管電層析神經降壓素與腦啡肽的應用 Supported Ionic Liquid for the CEC Separation of Enkephalins and Neurotensins |
作者: | Huei-Chi Chen 陳慧綺 |
指導教授: | 劉春櫻(Chuen-Ying Liu) |
關鍵字: | 離子液體,毛細管電層析, ionic liquid,CEC, |
出版年 : | 2007 |
學位: | 碩士 |
摘要: | 本研究以共價鍵結的方式,將咪唑陽離子鍵結於毛細管的內壁,作為開管式毛細管電層析的靜相,用於分離結構相似的神經降壓素 (neurotensin,NT)與腦啡肽(enkephalin)。首先以3-(chloropropyl)- trimethoxysilane (CPTMS)將毛細管矽烷化,再將咪唑溶於甲苯或DMF,與CPTMS的活性基反應,繼而以ㄧ溴辛烷進行烷化反應。所製備的管柱經電滲流的測定,發現以DMF為溶劑之鍵結量優於甲苯,顯示溶解度增加與吡啶的催化,可改進咪唑鍵結量,使管柱在pH 4~7的條件下,電滲透流皆為逆向。而製備管柱通入磷酸緩衝溶液時,管壁 dialkylimidazolium與磷酸會形成離子對 (ion-pair)或產生反應,使靜相較易脫落,因而改用醋酸緩衝溶液及Tris作為移動相。由研究發現,以電壓-20 kV,添加10%(v/v) 正丙醇於醋酸緩衝溶液 (pH 4, 100 mM)時,兩者可於十分鐘內達到基線分離,RSD值均 < 3.31% (n=5);而以電壓-20 kV,添加10 % (v/v)甲醇於醋酸緩衝溶液 (pH 5.0, 10 mM)為最佳分離條件時,可使(Gln4)-NT、(D-Trp11)-NT、(D-Tyr11)-NT與(L-Tyr11)-NT出現三支吸收峰,僅掌性異構物無法分離,RSD值均 < 7.8 % (n=5)。此外,本研究亦比較咪唑鍵結管柱與不同碳鏈長度之咪唑陽離子靜相對分離的影響,發現咪唑鍵結管柱與靜相之作用力較強,且分析物在異相系統中擴散性較差,波峰拖尾較嚴重;而咪唑陽離子碳鏈較短 (C4)時,滯留效果不佳,分析物無法分離;碳鏈長度較長(C8, or C12)時,分析物滯留效果較佳,且隨碳鏈長度增長分析物的滯留時間亦無明顯的延長。綜上所述,此離子液體不僅提供管柱表面的正電荷,使其具逆向電滲透流,還可減少分析物在管柱表面的吸附,且靜相具凡德瓦力 (分散力與偶極力)、π-π電子作用力、氫鍵與靜電交互作用力等機制,故即使構造甚為相似,電荷/質量比雷同的分析物均可達成功的分離。 Two peptides enkephalins and neurotensins were carried out with supported-ionic liquid open-tubular columns. To prepare ionic liquid columns, the capillary was first silanzed with 3-(chloropropyl)- trimethoxysilane (CPTMS). Imidazole was then subjected to be bond covalently to the CPTMS in the presence of toluene or DMF as solvent. Finally imidazole was linked with 1-bromooctane to form a positively charged diakylimidazolium layer for the generation of reversed electroosmotic flow (EOF) over a pH range of 4 to 7. Using phosphate as background electrolyte (BEG) the strong ion-pair interaction between diakylimidazolium and phosphate caused the bleeding of stationary phase. With running buffer of acetate (pH 4, 100 mM)/10% (v/v) 1-propanol, [Leu]-enkephalin and [Met]-enkephalin were baseline separated within 10 min with RSD < 3.31% (n=5). With running buffer of acetate (pH 5, 10 mM)/10% (v/v) methanol, four closely related neurotensins showed three absorbed peaks within 15 min with RSD < 7.8% (n=5). In this study, the migration pattern of imidazole coated column was also compared with those of different carbon length of substituent on ionic liquid column. Because of the strong hydrogen bond interaction, the peaks of imidazole coated column showed more tailing. For C8 or C12 of substituent of ionic liquid, the hydrophobic force and hydrogen bond interaction were stronger, and the column efficiency was better than the short carbon chain length (C4) . |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/29764 |
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