研究生: |
蔡志鑫 Chih-Hsin Tsai |
---|---|
論文名稱: |
線上濃縮技術在非水相毛細管電泳與毛細管電泳/表面增強拉曼法上的應用 Applications of on-line sample concentration techniques on nonaqueous capillary electrophoresis (NACE) and capillary electrophoresis/surface-enhanced Raman spectroscopy (CE/SERS) |
指導教授: |
林震煌
Lin, Cheng-Huang |
學位類別: |
博士 Doctor |
系所名稱: |
化學系 Department of Chemistry |
論文出版年: | 2007 |
畢業學年度: | 95 |
語文別: | 中文 |
論文頁數: | 195 |
中文關鍵詞: | 毛細管電泳 、掃集 、蛇根鹼 、紫光LED 、低溫槽 、安非他命 、非水相電泳 、堆積 、拉曼 、孔雀石綠 、結晶紫 |
英文關鍵詞: | capillary electrophoresis, sweeping, reserpine, violet light-emitting diode, Low-temperature bath, 3,4-Methylenedioxymethamphetamine, Nonaqueous capillary electrophoresis, Stacking, Raman spectroscopy, Malachite green, Crystal violet |
論文種類: | 學術論文 |
相關次數: | 點閱:195 下載:8 |
分享至: |
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本研究成功的發展了三種新的毛細管電泳分析技術。首先是成功的開拓了LED (發光二極體)在毛細管電泳分析領域的適用性。這是以市售紫光LED (405 nm) 為螢光激發光源,對血壓平(reserpine)及衍生物進行螢光偵測。使用CZE-stacking濃縮技術偵測極限可達1.6 × 10-8 M。若使用sweeping-MEKC (微胞掃集法)及CSEI-sweep-MEKC (陽離子選擇完全注射掃集MEKC法)濃縮技術時,其偵測極限分別可以達到2.1 × 10-9 M及2.1 × 10-10 M。另外藉由NDA (naphthalene-2,3-dicarboxaldehyde)做為螢光標識試劑,與多巴胺進行衍生反應以後,以螢光偵測結合MEKC及sweeping-MEKC濃縮技術進行測量,其偵測極限可達6.3 × 10-6 M及3.0 × 10-8 M。
其次,本研究首先發展以低溫-非水相毛細管電泳的新方法。對其光學異構物±3,4-methylenedioxymethamphetamine (±3,4-MDMA)可以獲得良好的分離效果。本文詳細探討各種最佳的電泳條件,包括使用各種不同的低溫槽及毛細管內最佳化的高導電度的緩衝溶液。在CZE模式下偵測極限可以達到4.7 × 10-6 M,再結合低溫/非水相堆積線上濃縮技術(LTB/NACZE-stacking),偵測極限更可以達到5.0 × 10-9 M。此外為了增加樣品進樣量以及能夠有更窄的樣品區帶,在樣品區帶和電泳背景溶液之間加入一段高導區帶,造成溶液之間有不同的導電梯度,使得樣品進樣量相對增加。利用這些技術,亦成功的應用在真實樣品3,4-MDMA的分析上。
最後,本研究對於非螢光性物質的偵測,亦成功的發展出新的方法。傳統上毛細管電泳法對非螢光性物質的偵測方法不外乎使用間接法,或是將非螢光性物質加以螢光衍生劑衍生後加以偵測。本研究選用非螢光性物質孔雀石綠為測試樣品,並以波長532 nm 雷射(Nd:YAG的第二倍頻波)為拉曼激發光源。在孔雀石綠定量分析上,以單光器(有效寬度0.4 nm)以及拉曼波數1616 cm-1作為收光範圍。 在毛細電泳/共振拉曼的模式下,孔雀石綠在CZE和MEKC模式下的偵測極限為1.6 × 10-5 M 和 1.1 × 10-5 M。當結合線上濃縮技術stacking及sweeping時,偵測極限可以達到3.4 × 10-7 M和5.3 × 10-9 M。而在毛細電泳/表面增強拉曼模式下,再結合線上濃縮技術stacking及sweeping,偵測極限甚至可以分別高達到4.4 × 10-8 M和1.1 × 10-9 M。本方法亦有效的應用在真實樣品的偵測上。
Several new methods for capillary electrophoresis were developed in the study. The first work represents the applications of violet LED-induced fluorescence detection in CE separations and this suggests that the violet LED has great potential for use as a new light source in CE separations, not only for naturally fluorescent compounds (excited in violet region) but for derivatives as well. The detection limit of reserpine was determined to be 1.6 × 10-8 M by CZE-stacking and this was improved to 2.1 × 10-9 and 2.1 × 10-10 when sweeping-MEKC and CSEI-sweep- MEKC techniques were applied. In addition, dopamine labeled-NDA (naphthalene-2,3-dicarboxaldehyde) was selected as the model compound. The detection limit of dopamine was determined to be 6.3 × 10-6 and 3.0 × 10-8 by means of a MEKC and sweeping-MEKC.
On the other hand, a low-temperature and ambient-temperature nonaqueous-stacking techniques in capillary electrophoresis are described for the first time. The low temperature bath was also applied to the separation of isomers ±3,4-methylenedioxymethamphetamine (±3,4-MDMA). 3,4-MDMA was determined at a concentration of 4.7 × 10-6 M by normal nonaqueous capillary electrophoresis (NACE) and this was improved to 5.0 × 10-9 M, when the low temperature bath/nonaqueous capillary electrophoresis-stacking (LTB/NACE-stacking) techniques was applied. Furthermore, in an attempt to increase the amount of sample injected, as well as to focus them onto a small zone, two novel buffer systems are proposed. One of these employs an “ultra-high conductivity zone”, which was inserted between the sample zone and background solution to build an unequal conductivity gradient. As a result, a large volume of sample injection can be achieved. Using these techniques, the content of 3,4-MDMA in an illicit drug and a suspect urine sample was readily detected.
Finally, if the analyte can not emit or absorb UV/visible radiation, alternate types of detection, such as indirect fluorescence/absorbance and derivative can be used. A non-fluorescent compound (malachite green, MG) and a doubled Nd:YAG laser (532 nm, 300 mW) were selected as the model compound and light source. In order to carry out a qualitative analysis of MG, a monochromator (effective bandwidth, 0.4 nm) was used to collect the specific Raman line at 1616 cm-1. On the capillary electrophoresis-resonance Raman spectroscopy (CE-RRS) mode, the limits of detection for MG were 1.6 × 10-5 M and 1.1 × 10-5 M, respectively for CZE and MEKC modes. When the stacking and sweeping modes were applied, the LOD could be improved to 3.4 × 10-7 M and 5.3 × 10-9 M. On the capillary electrophoresis/surface-enhanced Raman spectroscopy (CE/SERS) mode, the limit of detection for MG could be improved to 4.4 × 10-8 M and 1.1 × 10-9 M, when the stacking and sweeping modes were applied, respectively. The method was also extended to the determination of MG in an actual sample.
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