在分析過程中，樣品的前處理技術不僅影響實驗的結果且左右著整體實驗上時間及金錢的消耗，故如何去選擇一個快速、環保及符合經濟成本的前處理技術仍為現今最炙手可熱之話題，在此本論文提供兩種微萃取的技術分別連接於大氣壓基質輔助雷射脫離子阱附質譜儀及毛細管電泳儀。
第一個部分我們開發一新穎的一滴溶劑萃取一滴樣品之超微量萃取法(DDSME)結合大氣壓基質輔助雷射脫附離子阱質譜(AP-MALDI-Ion Trap)以快速分析水中、尿中及血中的尼福密酸(Niflumic Acid)，其中對於影響萃取的因子如有機溶劑、基質(α-cyano-4-hydroxy cinnamic acid, α-CHCA)濃度、萃取時間、水溶液的pH 值及鹽類添加效應逐一進行探討，在選用辛烷做為萃取溶劑、5000ppm 的α-CHCA、萃取時間十分鐘、pH 值為8.5 及不添加任何鹽類的最佳化的條件下，我們可測得水中、尿中及血中的偵測極限分別為0.1ng/mL、1ng/mL 及40ng/mL，此技術不僅提供一快速且高靈敏度的新萃取技術，並可作為藥物濃縮探針的有效方法。
第二部分我們利用一滴溶劑微萃取法(SDME)結合毛細管電泳儀對於不同基質中的三環系列藥物進行鑑別，萃取過程中欲分析物經由一滴溶劑微萃取法的有機溶劑萃取獲得，再藉由氮氣將溶劑吹乾並將欲分析物以毛細電泳所需緩衝溶液回溶，最後將樣品進樣以進行分析，其中萃取前水樣先經由氨水鹼化，藉此促進分析物萃取效率。在本研究中亦對五個影響萃取效率的因子(溶劑種類、萃取時間、磁石轉速、水溶液的pH 值及鹽類添加效應)分別進行探討，與傳統的液相-液相萃法取及固相萃取法相較之下此技術不僅操作簡易且符合經濟成本，此外完全無樣品殘留之顧慮，有效節約實驗成本及降低實驗過程中產生的對實驗人員的傷害和對環境的污染。

Excellent sample preparation techniques not only concern the fruitful results but also time and cost of experiments. Thus, it is a big challenge to develop a rapid and low cost of sample preparation method. There are two types of microextraction techniques developed in this thesis to couple with atmospheric pressure matrix assisted laser desorption mass spectrometry (AP-MALDI/MS) and capillary electrophoresis (CE) for analysis of drugs. The first one is a novel microextraction technique termed drop to drop solvent microextraction (DDSME) to couple to AP-MALDI/MS for the determination of niflumic acid in water, human urine and plasma. The effects of extraction solvent, concentration of α-cyano-4-hydroxy cinnamic acid, sampling time, pH and salt addition on the extraction efficiency were investigated and optimized. The best optimum conditions for the DDSME method were using octanol as extraction solvent, 5000 ppm of α-cyano-4-hydroxy cinnamic acid, 10 min of sampling time, pH of 8.5 and without salt addition. The limits of detection (LOD) of this method were in the range of 0.1 to 40 ng/ml. This method is not only a fast and sensitivity technique but also can be used as concentrating probe of drug. The second project is to combine the single drop microextraction (SDME) with capillary electrophoresis for the analysis of tricyclic antidepressant drugs in various matrixes including water and urine. These samples were extracted into organic solvent suspended in a stirred aqueous solution from the tip of a moicrosyringe needle. Nitrogen gas was used to remove the organic solvent, and then the buffer solution was used in capillary electrophoresis to re-dissolve samples. Prior to SDME, the aqueous solution was treated with ammonium hydroxide in order to promote extraction efficiency. The effects of extraction solvent, sampling time, sample agitation rate, pH and salt addition on the extraction efficiency were examined and optimized. This technique is simpler and cheaper than the traditional liquid-liquid extraction and solid phase extraction techniques.

目錄
目錄………………………………………………………………………壹
圖表目錄…………………………………………………………………伍
其他研究成果……………………………………………………………捌
第一章開發一滴溶劑萃取一滴樣品之超微量萃取法連結大氣壓基質
輔助雷射脫附質譜
1.1、導論…………………………………………………………………1
1.1.1、樣品前處理技術…………………………………………………1
1.1.1.1、液相-液相萃取法……………………………………………2
1.1.1.2、固相微萃取法…………………………………………………3
1.1.1.3、溶劑微萃取法…………………………………………………4
1.1.1.4、一滴溶劑微萃取法……………………………………………5
1.1.1.5、離子液體………………………………………………………6
1.1.1.6、中空纖維-液相微萃取法……………………………………7
1.1.2、大氣壓基質輔助雷射脫附游離法.……………………………8
1.1.2.1、離子形成機制…………………………………………………9
1.1.2.2、基質的特性與功能……………………………………………9
1.1.3、研究目的………………………………………………………10
1.2、實驗………………………………………………………………11
1.2.1、藥品……………………………………………………………11
1.2.2、儀器及參數設定………………………………………………11
1.2.3、一滴溶劑萃取一滴樣品之超微量萃取步驟…………………12
1.3、結果與討論………………………………………………………13
1.3.1、一滴溶劑萃取一滴樣品之超微量萃取法條件最佳化………13
1.3.1.1、溶劑選擇……………………………………………………13
1.3.1.2、α-CHCA 濃度………………………………………………14
1.3.1.3、水溶液pH 值…………………………………………………14
1.3.1.4、萃取時間……………………………………………………15
1.3.1.5、鹽類添加濃度………………………………………………15
1.3.2、不同基質樣品之定性表現……………………………………16
1.4、結論………………………………………………………………17
1.5、參考資料…………………………………………………………38
第二章一滴溶劑微萃取法連結毛細管電泳的應用
2.1、導論………………………………………………………………40
2.1.1、前言……………………………………………………………40
2.1.2、樣品前處理技術………………………………………………41
2.1.2.1、液相-液相-液相微萃取法…………………………………41
2.1.2.2、液相-液相-液相微萃取法基本原理………………………42
2.1.2.3、動態三相微萃取法…………………………………………43
2.1.2.4、動態三相微萃取法基本原理………………………………43
2.1.2.5、水性頂空液相微萃取法……………………………………44
2.1.3、電泳的發展.……………………………………………………45
2.1.3.1、電泳基本原理………………………………………………46
2.1.3.2、電滲流原理…………………………………………………47
2.1.3.3、毛細管電泳的分離模式……………………………………48
2.1.3.3.a 、毛細管區帶電泳法………………………………………49
2.1.3.3.b、微胞電動毛細管層析法…………………………………49
2.1.3.3.c、毛細管等電聚焦法………………………………………50
2.1.3.3.d、毛細管凝膠電泳法………………………………………50
2.1.3.3.e、毛細管等速電泳法………………………………………51
2.1.3.3.f、毛細管電層析法…………………………………………52
2.1.4、研究目的………………………………………………………52
2.2、實驗………………………………………………………………53
2.2.1、藥品……………………………………………………………53
2.2.2、儀器及實驗條件………………………………………………54
2.2.3、一滴溶劑微萃取法萃取步驟…………………………………54
2.3、結果與討論………………………………………………………55
2.3.1、毛細管電泳分離條件最佳化…………………………………55
2.3.1.1、緩衝溶液濃度………………………………………………55
2.3.1.2、緩衝溶液pH 值………………………………………………56
2.3.1.3、界面活性劑濃度……………………………………………56
2.3.1.4、有機溶劑添加百分比………………………………………57
2.3.1.5、分離電壓……………………………………………………58
2.3.2、一滴溶劑微萃取法之萃取條件最佳化………………………58
2.3.2.1、溶劑選擇……………………………………………………59
2.3.2.2、水溶液pH 值…………………………………………………59
2.3.2.3、萃取時間……………………………………………………60
2.3.2.4、攪拌子轉速…………………………………………………60
2.3.2.5、鹽類添加濃度………………………………………………61
2.3.3、不同基質樣品之定性表現……………………………………62
2.4、結論………………………………………………………………63
2.5、參考資料…………………………………………………………85
圖表目錄
Fig 1-1. Side view of the first commercial SPME device made by Supelco………………………………………………………………18
Figure 1-2. Side view of the single-drop microextraction system used by Jeannot and Cantwell in 1996…………………19
Figure 1-3. Side view of organic drop attached to the needle tip used by Y. He and H. K. Lee in 1997 ……………20
Figure 1-4. Side views of dynamic SDME within the microsyringe used by Y. He and H. K. Lee in 1997……………21
Figure 1-5. Side views of the apparatus used for headspace single-drop solvent microextraction used by Michael A. Jeannot in 2001………………………………………………………22
Figure 1-6. Side views of the two different versions of single-drop solvent microextraction used by Gui-bin Jiang in 2003…………………………………………………………………23
Figure 1-7. Side views of hollow fiber-protected LPME system used by G.Shen and H. K. Lee in 2002…………………24
Figure 1-8. Side views of dynamic LPME/HF within the hollow fiber used by L. Zhao and H. K. Lee in 2002…………………25
Figure 1-9. Structures of niflumic acid and alpha-cyano-4-hydroxy cinnamic acid………………………………………………26
Figure 1-10. Side view of the commericially available AP-MALDI source from Mass Technologies ……………………………27
Figure 1-11. Schematic of drop to drop solvent microextraction ………………………………………………………28
Figure 1-12. AP-MALDI mass spectra of niflumic acid obtained (A)without DDSME (B) with DDSME by using octanol as extraction solvent………………………………………………29
Figure 1-13. DDSME/AP-MALDI/MS/MS mass spectra of niflumic acidat m/z 283…………………………………………………………30
Figure 1-14. Proposed mechanism for fragmentation of the ion at m/z 283.07……………………………………………………31
Figure 1-15. Effect of organic solvent on the extraction efficiency(A)Hexane (B)Toluene (C)Dichloromethane (D)Octanol …………………………………………………………………32
Figure 1-16. Effect of matrix concentration on the extraction efficiency(A)1000ppm (B)2000ppm (C)5000ppm (D)10000ppm by using α-CHCA as matrix for 50ppm of niflumic acid in aqueous solution …………………………………………33
Figure 1-17. Effect of sample pH on the extraction efficiency (A) pH=4 (B)pH=6 (C)pH=8.5 (DDW) (D)pH=11………34
Figure 1-18. Effect of extraction time on the extraction efficiency (A)1min (B)5min (C)10 min (D)15 min………………35
Figure 1-19. Effect of addition of salt on the extraction efficiency (A)no salt (B)5% (C)10% (D)20%……………………36
Figure 1-20. DDSME/AP-MALDI mass spectra of 50ppm of niflumic acid obtained from different matrix (A) without DDSME in human urine (B) with DDSME in human urine (C) with DDSME in human plasma………………………………………………37
Figure 2-1. Side view of the support liquid membrane extraction used by S. Pálmarsdóttir in 1997…………………64
Figure 2-2. Side view of the LLLME extraction unit used by S. Pedersen-Bjergaard and K. E. Rasmussen in 1999…………65
Figure 2-3. Side view of principle of the LLLME used by S.
Pedersen-Bjergaard and K. E. Rasmussenin 1999………………66
Figure 2-4. Side view of the dynamic LLLME used by L. Hou and H. K.Lee in 2003…………………………………………………67
Figure 2-5. Side view of the dynamic LLLME within the hollow fiber used by L. Hou and H. K. Lee in 2003…………68
Figure 2-6. Side view of headspace water-based liquid phase
microextraction used by J. Zhang, T. Su and H. K. Lee in 2005………………………………………………………………………69
Figure 2-7. Side view of CZE used by D. N. Heiger in 1992………………………………………………………………………70
Figure 2-8. Side view of MEKC used by D. N. Heiger in 1992………………………………………………………………………70
Figure 2-9. Structures of tricyclic drugs……………………71
Figure 2-10. Schematic of single-drop solvent microextraction………………………………………………………72
Figure 2-11. Effect of buffer concentration on the separation of (1) Nortriptyline, (2) Trimeprazine, (3) Promethazine, (4) Iminodibenzyl.Concentration of each tricyclic drug was 62.5µg /ml……………………………………73
Figure 2-12. Effect of buffer pH on the separation of (1) Nortriptyline, (2) Trimeprazine, (3) Promethazine, (4) Iminodibenzyl. Concentration of each tricyclic drug was 62.5µg /ml.……………………………………………………………74
Figure 2-13. Effect of SDS concentration on the separation of (1) Nortriptyline, (2) Trimeprazine, (3) Promethazine, (4) Iminodibenzyl.Concentration of each tricyclic drug was 62.5µg /ml. …………………………………………………………75
Figure 2-14. Effect of methanol percentage on the separation of (1) Nortriptyline, (2) Trimeprazine, (3) Promethazine, (4) Iminodibenzyl.Concentration of each tricyclic drug was 62.5µg /ml. …………………………………76
Figure 2-15. Effect of separation voltage on separation of (1) Nortriptyline, (2) Trimeprazine, (3) Promethazine, (4) Iminodibenzyl.Concentration of each tricyclic drug was 62.5µg /ml. …………………………………………………………77
Figure 2-16. Relative extraction efficiencies of hexane, toluene,cyclohexane and chloroform for the extraction of tricyclic drug from spiked pure water sample.………………78
Figure 2-17. Effect of sample pH on the extraction efficiency………………………………………………………………79
Figure 2-18. Extraction time profiles for the selection of sampling time…………………………………………………………80
Figure 2-19. Effect of sample agitation rate on extraction efficiency………………………………………………………………81
Figure 2-20. Effect of addition of salt on the extraction efficiency.……………………………………………………………82
Figure 2-21. Electropherogram of a spiked water sample obtained by SDME-CE: CE buffer, 20mM ammonium acetate (pH=9) ; SDS concentration, 10mM ; Methanol %, 50% ; inject voltage, 5kV/ 5s ; separation voltage, 28kV ; capillary, 70-cm (effective length) × 75µm ; detection, UV at 254nm. Peak : (1) Nortriptyline (0.06µg/ml), (2) Trimeprazine (0.015µg/ml), (3) Promethazine (0.015µg/ml), (4)Iminodibenzyl (0.06µg/ml).………………………………………83
Table 1. Within-day repeatability (n=6) for SDME-CE of tricyclic drugs from pure water and urine……………………84
其他研究成果
其他研究成果…………………………………………………………………………87

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