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研究生:龔仁方
研究生(外文):GONG,REN-FANG
論文名稱:以Au/SBA-15固定化纖維水解酵素之初探
論文名稱(外文):The preliminary exploration of immobilization of cellulase on Au/SBA-15
指導教授:張原謀張原謀引用關係
指導教授(外文):Chang,Yuan-Mou
口試委員:劉世鈞王貴弘
口試委員(外文):Liu,Shi-JunWang,Gui-Hong
口試日期:2014-07-24
學位類別:碩士
校院名稱:國立臺南大學
系所名稱:生態科學與技術學系環境生態碩士班
學門:環境保護學門
學類:環境資源學類
論文種類:學術論文
論文出版年:2014
畢業學年度:102
語文別:中文
論文頁數:54
中文關鍵詞:中孔洞氧化矽奈米金固定化酵素纖維水解酵素
外文關鍵詞:mesoporous silicagold nanoparticlesenzyme immobilizationcellulase
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  • 點閱點閱:125
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  • 下載下載:1
  • 收藏至我的研究室書目清單書目收藏:0
本研究首先製備中孔氧化矽,之後利用3-胺丙基三甲氧基矽烷(3-Aminopropyltrimethoxysilane,APTMS )來進行氮官能基表面改質,增加金奈米顆粒在其上的均勻散佈。接著使用螯合劑並以微波還原方式合成,將金奈米顆粒負載在中孔氧化矽表面。之後再利用金—中孔洞氧化矽來對纖維水解酵素(Trichoderma reesi)進行固定化。以不修飾金奈米粒子,純粹由金奈米粒子和酵素上的官能基進行鍵結的方式,固定化纖維水解酵素。研究較佳的固定化條件,固定化酵素濃度為4wt%、固定化溫度為40℃及固定化pH值為5.0的條件之下。利用最佳條件去做固定化酵素穩定性分析,固定化酵素的表現不如預期良好。在高溫下固定化酶穩定性較游離酶差,而pH值安定性的部分也沒有明顯穩定,經過3次CMC水解反應後固定化酵素相對比活性已在40%以下,重複利用率也不高。此次實驗於固定酵素方法仍需要相當的改進,未來需朝向在金奈米顆粒和酵素固定之間加入交聯劑並重新探討固定化條件,以提升酵素穩定性。

In this study, the mesoporous silica was synthesized by the use of poly-(alkylene oxide) block copolymer. The preparation of mesoporous silica was synthesized by the use of poly-(alkylene oxide) block copolymer. To distribute the Au nanoparticles uniformly on the surface of mesoporous silica, its surfaces were functionalized with amine group prior to addition of APTMS by post-synthesis. To increasing gold nanoparticles evenly spread on mesoporous silica. Then, the surface of mesoporous silica was supported gold nanoparticles. The surface morphology of mesoporous silica after microwave synthesis and the crystal structure of metal nanoparticles. The enzyme, Trichoderma reesi cellulose was immobilized on gold nanoparticles. Gold nanoparticles with no modification, purely functional groups of gold nanoparticles and enzymes were binding on the way, the immobilized enzyme hydrolysis fibers. Study preferred immobilization conditions, the concentration of the immobilized enzyme 4wt%, under the condition of fixed temperature and 40 ℃ immobilized pH of 5.0. Under the optimal immobilization conditions to do the enzyme stability analysis, the immobilized enzyme is expected to perform as well. Stability at high temperature , the enzyme immobilized poor than free enzyme and part of the pH stability is also not well. The efficiency of immobilized cellulose was examined for carboxymethyl cellulose (CMC) hydrolysis for the recycling use , after 3 times the relatively specific activity just only for 40% and recycling rate is not high. The experimental method is still in the immobilization of enzymes requires considerable improvement in the future should face between gold nanoparticles and enzymes fixed crosslinking agent and re-examine the immobilization conditions to improve enzyme stability.
摘要 i
Abstract ii
誌謝 iii
目錄 iv
表目錄 vii
圖目錄 viii
第一章 緒論 1
1.1 研究背景 1
1.2 研究動機與目的 3
第二章 文獻回顧 4
2.1 纖維酒精 4
2.1.1 木質纖維素 5
2.2 酵素 9
2.3 纖維水解酵素 9
2.3.1 內切型纖維水解酵素 9
2.3.2 外切型纖維水解酵素 10
2.3.3 纖維雙醣水解酵素 10
2.4 酵素的固定化 13
2.5 中孔洞氧化矽材料發展及其研究 18
2.5.1 中孔洞分子篩之性質與應用 18
2.5.2 中孔洞二氧化矽負載金屬奈米顆粒 19
2.6 金奈米粒子 20
第三章 研究材料與方法 21
3.1 實驗材料與設備 21
3.1.1 化學藥品與試劑 21
3.2 實驗方法與步驟 22
3.2.1 中孔洞氧化矽合成步驟 23
3.2.2 中孔洞氧化矽表面官能基改質 23
3.2.3 微波還原法製備黃金顆粒 23
3.2.4 纖維水解酵素在Au / SBA-15上的固定化 24
3.3 實驗分析方法 24
3.3.1 還原醣測定 24
3.3.2 蛋白質濃度的測定 25
3.3.3 纖維水解酵素之活性分析 26
3.4 儀器分析 26
3.4.1 氮氣等溫吸附/脫附量測儀 26
3.4.2 高解析傅立葉紅外線吸收光譜 28
3.4.3 高解析穿透式電子顯微鏡 28
第四章 結果與討論 33
4.1 中孔洞氧化矽實驗合成分析結果 33
4.2 MPS表面官能基改質 33
4.3 奈米金屬粒子Au負載 35
4.4 固定化酵素之參數探討 39
4.4.1 酵素濃度對固定化活性 39
4.4.2 固定化溫度對活性之影響 41
4.4.3 固定化pH對活性之影響 42
4.4.4 固定在不同Au含量下對活性之影響 44
4.5 固定化酵素穩定性分析 45
4.5.1 固定化酵素之熱穩定性 45
4.5.2 固定化酵素之pH值穩定性 45
4.5.3 固定化酵素重複使用 47
第五章 結論 48
參考文獻 49
附錄 55
附錄一 還原醣檢量線 55
附錄二 蛋白質檢量線 55

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