氧化鐵磁性奈米顆粒(Fe3O4)因具高比表面積、超順磁性、可回收、無毒性等特點，常作為固定化酵素的載體。本研究利用化學共沉法合成 Fe3O4時，以富含羧酸基聚合物ES-COOH修飾其表面，再經由碳二醯胺(EDC)與N-羥基硫代琥珀亞胺(NHS)活化，使其與脂肪酶(lipase)產生共價鍵結，達到酵素固定化的目的。於材料鑑定與特性分析中，藉由 XRD 確認合成出來的材料是Fe3O4，再由 TGA 與 FTIR 證實了成功的製備得 Fe3O4-ES複材，進而以SQUID 得知複材依然保有超順磁性，且粒徑分析則證明包覆了ES-COOH的Fe3O4粒徑會變小。最後，將Fe3O4-ES複材應用於脂肪酶之固定化，結果顯示最適化條件為:當酵素添加量越少固定化效果越好，選用 18.9 U 為最佳添加量，載體添加量低於 0.1 g 載體與固定酵素量成等比例成長，EDC 濃度 0.025 g/mL 且在偏酸的環境(pH 4-6)中會有比較好的固定化效果。以最適化的固定參數去比較有、無表面修飾的影響，得知Fe3O4-ES 磁性奈米複材的固定化效率可達約 42%，但未修飾的Fe3O4則僅有 16%；最後，本研究亦證明了固定化酵素具有重複使用的催化活性。

Nano-magnetite particle (Fe3O4) has been commonly used as the carrier for enzyme immobilization because of its high specific surface area, superparamagnetism, easy re-covery and non-toxicity. In this study nano-Fe3O4 composites (Fe3O4-ES) were firstly prepared by using the chemical co-precipitation method and in-situ surface modification with a carboxyl group enriching polymer ES-COOH. And then lipase was covalently bounded to the nanocomposite being activated by carbodiimide (EDC) and N-hydroxysuccinimide (NHS). The structure and magnetic property of the resultant nanocomposites were characterized by X-ray diffraction and the superconducting quan-tum interference device magnetometer. The attachment of ES-COOH to Fe3O4 was con-firmed by thermogravimetric analysis and Fourier transform infrared spectroscopy. In addition, data from dynamic light scattering measurement indicate the particle size of Fe3O4-ES become smaller than plain Fe3O4. For lipase immobilization, results show that optimal conditions are carrier 0.1 g, enzyme loading 18.9 U, EDC concentration 0.025 g/mL and in acidic condition (pH 4-6). Under these conditions the immobilization effi-ciencies for Fe3O4-ES and Fe3O4 are 42% and 16%, respectively. Moreover, the cova-lently immobilized enzyme on Fe3O4-ES also demonstrated an excellent operating sta-bility.

中文摘要 i
Abstract ii
誌謝 iii
目錄 iv
表目錄 vi
圖目錄 vii
第一章緒論 1
1.1 奈米特性 1
1.2 研究動機與目的 2
第二章文獻回顧 5
2.1 磁性奈米複材 5
2.2 磁性奈米粒子載體的選擇與應用 5
2.3 脂肪酶 (lipase) 6
2.4 酵素固定化 12
2.5 氧化鐵微粒於固定化酶之應用 14
2.6 固定化脂肪酶 16
2.7 酵素反應動力學方程式 19
第三章材料與實驗方法 20
3.1 實驗藥品 20
3.2 實驗儀器 20
3.2.1 測量儀器與設備 20
3.2.2 其他設備 21
3.3 實驗架構 22
3.4 Fe3O4-ES奈米複材製備 23
3.5 材料特性分析 23
3.5.1 FTIR 特性分析 23
3.5.2 TGA 熱重分析 23
3.5.3 Zeta Potential 表面電位 23
3.5.4 DLS 粒徑分析 24
3.5.5 X-ray 繞射分析 24
3.5.6 SQUID 磁性分析 24
3.6 脂肪酶之活性分析 24
3.7 固定化變因探討 25
3.8 固定化脂肪酶之重複使用性 25
3.9 脂肪酶反應動力學之探討 25
第四章結果與討論 27
4.1. 材料特性分析 27
4.1.1 FTIR 特性分析 27
4.1.2 TGA 熱重分析 29
4.1.3 Zeta Potential 表面電位 31
4.1.4 DLS 粒徑分析 33
4.1.5 X-ray 繞射分析 35
4.1.6 SQUID 磁性分析 37
4.2. 固定化變因探討 39
4.2.1 酵素的添加濃度 39
4.2.2 固定化 pH 值 41
4.2.3 奈米粒子添加量 43
4.2.4 EDC 添加量 46
4.2.5 固定化時間 48
4.2.6 ES-COOH添加量之影響 50
4.3 重複使用性 52
4.4. 酵素之反應動力學 54
第五章結論 56
參考文獻 57

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