透過原位創新合成法封裝微生物於類沸石咪唑骨架材料(ZIF-90)之相關研究

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研究生：	王思儒 Szu-Ju Wang
論文名稱：	透過原位創新合成法封裝微生物於類沸石咪唑骨架材料(ZIF-90)之相關研究
指導教授：	謝發坤 Fa-Kuen Shieh
口試委員:
學位類別：	碩士 Master
系所名稱：	理學院 - 化學學系 Department of Chemistry
論文出版年：	2018
畢業學年度：	106
語文別：	中文
論文頁數：	75
中文關鍵詞：	金屬有機骨架材料、大腸桿菌、生物複合材料、類沸石咪唑骨架材料
相關次數：	點閱：11 下載：0
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近年來，許多功能性細菌或微生物已經被研究並用於監測或改善環境問題，而當微生物遭遇紫外線、極端溫度、酸鹼環境或抗生素等威脅，將會使細胞生命力降低甚至死亡，而影響微生物的工作效率，所以常利用具生物相容性的材料塗層於微生物加強其抵抗惡劣環境的能力，目前已推進至“Cyborg Cell”的時代，常用的材料包括金屬奈米顆粒、二氧化矽、碳酸鈣等無機物，提供微生物堅固的保護層。與其他材料相比金屬有機骨架材料 (MOFs) 因具規則排列的孔洞以及其晶體大小的可調控性和水熱穩定性，為近年來新興材料之一。本研究將MOFs與微生物結合成生物複合材料，利用ZIF-90作為外殼保護大腸桿菌，探討封裝於微米級單晶ZIF-90內的大腸桿菌抵擋惡劣環境的能力。結果顯示，利用掃描式電子顯微鏡鑑定其粒徑大小約7-8微米；更進一步以共軛焦雷射掃描顯微鏡證實大腸桿菌成功地被封裝於金屬有機骨架材料中(E. coli@ZIF-90)。將E. coli@ZIF-90置於抗生素—氨苄青黴素的環境測試，證明藉由ZIF-90的孔洞 (3.5 Å) 選擇性抵抗外部的氨苄青黴素 (4.2×7.1×10.9 Å3) 保護大腸桿菌。此外也使用奈米級多晶ZIF-90或ZIF-8塗層於大腸桿菌表面，並從掃描式電子顯微鏡的影像進行觀察鑑定，但因合成環境嚴苛使大腸桿菌的存活率極低，未來仍需要改善合成環境及探討其存活率，藉以比較微米級單晶封裝與奈米級多晶塗層之差異。

In recent years, functional bacteria or microorganisms have been studied and used as biosensor to overcome environmental problems. Microorganisms have been armored by biocompatible materials with functionalization to resist harsh environments such as ultraviolet radiation, extreme temperatures, acid or base and antibiotics, and pushed the application of microorganism into the era of the “cyborg cells”. Comparing Metal Organic Frameworks (MOFs) with common materials like metal nanoparticles, silica or calcium carbonate, the MOFs has regular pore structure, tunable crystal size and high hydrothermal stability. Herein, we based on our previous reported that a de novo approach, i.e., synthesis under water and mild conditions, for encapsulating the Escherichia coli (E. coli) into ZIF-90 single crystals, E. coli@ZIF-90, as protective structure against harsh conditions. Spectral analysis such as scanning electron microscopy (SEM) images indicated that the morphology of biocomposites are uniform crystals with particle size of 7−8 μm. Moreover, the confocal microscopy was carried out to confirm that the E. coli were embedded in ZIF-90 crystals which provided shelter for the biocomposites against antibiotics. Consequently, E. coli could readily regain vitality after the removal of MOF protection. In addition, making a comparison between single crystals of E. coli@ZIF-90 and E. coli biocomposites coated by nanoscaled polycrystalline ZIF-90/ZIF-8; however, the survival rate of E. coli was low probably due to the harsh condition for coating by polycrystalline MOFs. Thus, the optimal synthetic condition for enhancing the vitality of E. coli which is protected by polycrystalline ZIF-90/ZIF-8 need to be further studied in the near future.

中文摘要                                I
Abstract                        II
圖目錄                                VI
表目錄                                IX
第一章 緒論                        1
1-1 金屬有機骨架材料                1
1-2 類沸石咪唑骨架材料                3
1-3 類沸石咪唑骨架材料-90                4
1-4 微生物                        6
1-5 大腸桿菌 (Escherichia coli)        8
1-6 質體 (Plasmid)                9
1-7 賽博格 (Cyborg)                11
1-8 生物複合金屬有機骨架材料        12
1-9 研究動機與目的                13
 第二章 實驗部分                        14
2-1 實驗藥品及材料                14
2-2 實驗儀器                        16
2-3 實驗儀器與方法                17
2-3-1 粉末X光繞射儀 (Powder X-Ray Diffraction, PXRD)  
                                    17
2-3-2 掃描式電子顯微鏡 (Scanning Electron Microscope, SEM)     
                                    18
2-3-3 熱重分析儀 (Thermogravimetric Analyzer, TGA)     
                                    19
2-3-4 螢光顯微鏡 (Fluorescence Microscopy)      20
2-3-5 共軛焦雷射掃描顯微鏡 (Confocal Microscopy)      21
2-3-6 菌落聚合酶鏈鎖反應 (Colony Polymerase Chain Reaction, PCR)                        22
2-3-7 DNA膠體電泳確認目標基因片段        24
2-3-8 攜帶型分光光度計 (Ultrospec 10 Cell Density Meter)     
                                    24
2-4 實驗步驟                        27
2-4-1 培養大腸桿菌                27
2-4-2 類沸石咪唑骨架材料-90封裝大腸桿菌相關之實驗步驟 (E. coli@ZIF-90)                        31
2-4-3 奈米級多晶類沸石咪唑骨架材料塗層於大腸桿菌表面之相關實驗步驟                                35
 第三章 結果與討論                39
3-1 微米級類沸石咪唑骨架材料-90封裝大腸桿菌  (E. coli@ZIF-90) 之相關鑑定                        39
3-1-1 粉末X光繞射鑑定結果                39
3-1-2 掃描式電子顯微鏡鑑定結果        40
3-1-3 熱重分析鑑定結果                41
3-1-4 螢光顯微鏡與共軛焦顯微鏡鑑定結果 42
3-1-5 鹽酸與乙二胺四乙酸對大腸桿菌之生長測試 44
3-1-6 Ampicillin對大腸桿菌之生長測試      45
3-1-7 固態培養基 (Agar Plate) &瓊脂膠體 (Agarose Gel) 之菌落測試                                     46
3-2 奈米級多晶類沸石咪唑骨架材料塗層於大腸桿菌表面之相關鑑定     
                                         49
3-2-1 粉末X光繞射鑑定結果                     49
3-2-2 掃描式電子顯微鏡鑑定結果             50
3-2-3 奈米級多晶類沸石咪唑骨架材料-90與奈米級多晶類沸石咪唑骨架材料-8塗層於大腸桿菌表面之活性鑑定     52
3-2-4 調控甘油與叔丁醇濃度合成不同尺度之奈米級類沸石咪唑骨架材料-90之討論     53
 第四章 結論     56
 第五章 參考文獻     57




                                

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