研究生: |
陳怡君 |
---|---|
論文名稱: |
毛細管電泳技術對葡萄糖偵測法的開發與研究 |
指導教授: | 林震煌 |
學位類別: |
碩士 Master |
系所名稱: |
化學系 Department of Chemistry |
論文出版年: | 2010 |
畢業學年度: | 98 |
語文別: | 中文 |
中文關鍵詞: | 葡萄糖 、電泳 、拉曼 |
論文種類: | 學術論文 |
相關次數: | 點閱:1830 下載:0 |
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毛細管電泳(capillary electrophoresis, CE)雖然具有高分離效率、能提供各種分離模式、溶劑消耗量低…等多項優點,但是此方法對於不具UV吸收性質或非螢光性物質的分析,仍有不足之處。本研究以葡萄糖(D-glucose)做為測試樣品,分別探討當毛細管電泳結合拉曼光譜法、螢光衍生法及間接吸收光譜法時,對葡萄糖分離與偵測的效果。
實驗結果發現,以綠光半導體雷射(500 mW, 532 nm)為激發光源,藉由CCD偵測器可同時觀測到葡萄糖具有多處拉曼特徵峰。其中以1129 cm-1的譜峰最適合做為電泳時的觀測峰。 在pH=12.1的氫氧化鈉溶液中,添加0.5 mM的界面活性劑-CTAB做為管壁修飾劑,以負電的模式進行電泳,可直接偵測到葡萄糖的單一譜峰。
為了提高偵測的靈敏度,本實驗以2-氨基吖啶酮(2-aminoacridone)為衍生試劑,對葡萄糖進行螢光化反應。衍生物以10 mM硼酸鹽溶液稀釋後,配製於10 mM硼酸鹽及100 mM界面活性劑-SDS所組成的緩衝溶液中。在此條件之下,以藍光半導體雷射(100 mW, 473 nm)為激發光源,進行毛細管電泳分析。在觀測CCD偵測器的即時光譜時,發現衍生試劑(2-氨基吖啶酮)的最大螢光波長為520 nm,但經螢光衍生後則紅光位移至580 nm。此螢光標識法可將葡萄糖的偵測極限改良至~ 1.1 × 10-5 M。上述方法可提供蜂蜜純度的鑑定或快速分析糖尿病患檢體等研究。
由於間接的方式無法得知分析物的光譜或質譜,本實驗嘗試將基質輔助雷射脫附游離飛行式質譜法(MALDI-TOFMS)中,經常使用到的基質:氰基-4-羥基肉桂酸(α-cyano-4-hydroxycinnamic acid, CHCA),直接添加於電泳溶液(同前述條件)中,作為為背景溶液,以間接吸收法進行偵測。當以氙燈經截光器後取出波長337 nm的紫外線作為吸收光源,且CHCA的濃度為10 mM時,葡萄糖的偵測極限為~1.3 × 10-3 M,線性範圍10-1 M ~10-3 M。
我們使用三種方法偵測葡萄糖,其中拉曼光譜法可以進行定量以及定性分析,未來如果克服了靈敏度的限制,拉曼光譜法將會是非常便利的偵測方式。
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