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研究生:沈豊鈞
研究生(外文):Li-Jun Shen
論文名稱:類沸石咪唑骨架材料 封裝大腸桿菌之相關研究
指導教授:謝發坤
指導教授(外文):Fa-Kuen Shieh
學位類別:碩士
校院名稱:國立中央大學
系所名稱:化學學系
學門:自然科學學門
學類:化學學類
論文出版年:2020
畢業學年度:108
語文別:中文
論文頁數:66
中文關鍵詞:金屬有機骨架材料類沸石咪唑骨架材料-8類沸石咪唑骨架材料-90大腸桿菌生物複合材料
外文關鍵詞:MOFZIF-8ZIF-90E. coliBiocomposites
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通過將功能性奈米材料與生命系統整合而成的奈米生物雜化物已經成為橫跨材料工程和生物科學界的一項新興領域,人們嘗試透過結合功能性材料與生命系統期望改善適應環境壓力的能力、增強生物系統中現有的功能或實現全新的功能,其中新興孔洞金屬有機骨架材料 (MOFs),因為可藉由調整金屬離子與有機配體組合的提供多樣選擇性,依需求設計,且具有一致的孔洞性質、高比表面積、優異的熱及化學穩定性、有催化功能的開放金屬位點等,因此近期有一系列著重在金屬有機骨架材料與生物系統結合的相關研究。
本研究先以類沸石咪唑骨架材料-8 (ZIF-8) 塗層於大腸桿菌表菌表面 (E. coli⊂ZIF-8),並藉由抗生素來驗證文獻中曾提到ZIF-8塗層可能存在著缺陷,進而使抗生素會透過缺陷造成大腸桿菌的死亡,並進一步利用本實驗室於2015年發表於JACS利用類沸石咪唑骨架材料-90 (ZIF-90) 封裝酵素之技術,擴展到原核生物-大腸桿菌,形成微米級ZIF-90封裝大腸桿菌 (E. coli@ZIF-90) 來提供完整的保護,在經歷培養於含抗生素的環境後,移除材料的大腸桿菌依然可以回復其生長狀態,我們藉由掃描式電子顯微鏡及共聚焦顯微鏡來觀察兩者 (ZIF-8/ZIF-90) 封裝之差異,並嘗試藉由zeta電位之量測與紅外光譜儀的數據來了解造成兩者差異之原因,希望能將此一研究成果應用於生物系統之運輸與儲存、細胞固定化、生物傳感等領域。
Nanobiohybrids, synthesized by integrating functional nanomaterials with living systems, have emerged as a novel branch of research at the interface of materials engineering and biological science. Nanobiohybrids use synthetic nanomaterials to overcome harsh environmental problems and impart organisms with emergent properties. Among all materials, Metal-organic Frameworks (MOFs) is a novel material made from metal ions and organic linkers. MOFs have consistent pore properties, large surface area, thermo-chemical stability, catalyzing, etc through the adjustment of the precursors. Therefore, the development of the combination of Metal-organic frameworks and living systems is rapidly emerging.
The prokaryotic living cell Escherichia coli (E. coli) for the first time is encapsulating into a single crystalline particle of Metal-organic Frameworks via de novo approach to create a novel biocomposite of “cyborgcells”. In the de novo approach, E. coli is entirely armored with single-crystalline ZIF-90 microporous in a single macro-size particle for the completely size-shielding protection, and therefore E. coli can be survived even lethal antibiotic treatment.
By comparison with ZIF-8, the poly-crystalline multiple-nanoparticles are formed in surrounding on E. coli surface as their higher zeta potential differences, but with poor antibiotic protection. We hope that this model may pave the way to new avenues for the transportation and storage of living systems, cell immobilization, biosensing.
中文摘要 i
Abstract ii
目錄 iii
圖目錄 vi
表目錄 viii
第一章 緒論 1
1-1 奈米生物雜化物 (Nanobiohybrids) 1
1-2 金屬有機骨架材料 2
1-2-1 金屬有機骨架材料 2
1-2-2 類沸石咪唑骨架材料 4
1-2-3 類沸石咪唑骨架材料-8/-90 5
1-3 微生物 7
1-3-1 微生物 7
1-3-2 大腸桿菌 (Escherichia coli, E. coli) 7
1-3-3 質體 (Plasmid) 9
1-4 研究動機與目的 11
第二章 實驗 12
2-1 實驗藥品及材料 12
2-2 實驗儀器 14
2-2-1 紫外光可見光分光光譜儀(UV/VIS Spectrophotometer) 15
2-2-2 X射線粉末繞射儀 (Power X-ray Diffractometer, PXRD) 15
2-2-3 掃描式電子顯微鏡 (Scanning Electron Microscope, SEM) 17
2-2-4 攜帶式分光光度計 (Ultrospec 10 cell Density Meter) 18
2-2-5 螢光顯微鏡 (Fluorescence Microscopy) 19
2-2-6 共軛焦雷射掃描顯微鏡 (Confocal Laser Scanning Microscope, CLSM) 20
2-2-7 聚合酶連鎖反應 (Polymerase Chain Reaction) 21
2-2-8 瓊脂糖凝膠電泳 (Agarose Gel Electrophoresis) 22
2-2-9 Zeta電位分析儀 (Zeta Potential Analyzer) 23
2-2-10 傅立葉轉換紅外線光譜儀 (Fourier-transform Infrared Spectroscopy, FTIR) 24
2-3 實驗步驟 26
2-3-1 培養大腸桿菌相關步驟 26
2-3-2奈米級類沸石咪唑骨架材料-8塗層於大腸桿菌 (E. coli⊂ZIF-8) 之相關實驗步驟 27
2-3-3 微米級類沸石咪唑骨架材料-90封裝大腸桿菌 (E. coli@ZIF-90) 之相關實驗步驟 27
2-3-4 類沸石咪唑骨架材料-8/-90封裝大腸桿菌 (E. coli⊂ZIF-8/E. coli@ZIF-90) 承受抗生素之活性實驗步驟 28
2-3-5 大腸桿菌鑑定之相關實驗步驟 28
第三章 結果與討論 30
3-1 奈米級類沸石咪唑骨架材料-8塗層於大腸桿菌/微米級類沸石咪唑骨架材料-90封裝大腸桿菌 (E. coli⊂ZIF-8/E. coli@ZIF-90) 之材料相關鑑定 30
3-1-1 X射線粉末繞射儀的鑑定結果 30
3-1-2 掃描式電子顯微鏡的鑑定結果 31
3-1-3 螢光顯微鏡與共軛焦螢光顯微鏡的鑑定結果 32
3-1-4 紅外光光譜儀的結果探討 35
3-1-5 Zeta電位的結果探討 38
3-2 奈米級類沸石咪唑骨架材料-8塗層於大腸桿菌/微米級類沸石咪唑骨架材料-90封裝大腸桿菌 (E. coli⊂ZIF-8/E. coli@ZIF-90) 之抗生素耐受測試活性探討與鑑定 40
3-2-1 奈米級類沸石咪唑骨架材料-8塗層於大腸桿菌/微米級類沸石咪唑骨架材料-90封裝大腸桿菌 (E. coli⊂ZIF-8/E. coli@ZIF-90) 之抗生素耐受活性測試 40
3-2-2 抗生素耐受測試後之菌種鑑定 41
第四章 結論 44
第五章 參考文獻 45
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