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研究生： 陳信安
Chen, Xin-An
論文名稱： 綠色合成氨基化之石墨烯量子點應用於疫苗檢測和生物影像
Green Synthesis Amino-functionalized Graphene Quantum Dots Applied in Inspect of Vaccine and Bioimaging
指導教授： 施士塵
Shi, Shih-Chen
學位類別： 碩士
Master
系所名稱： 工學院 - 機械工程學系
Department of Mechanical Engineering
論文出版年： 2021
畢業學年度： 109
語文別： 中文
論文頁數： 119
中文關鍵詞： 綠色合成 、石墨烯量子點 、氨基化 、疫苗檢測 、生物影像
外文關鍵詞： Green synthesis, GQDs, Amino-fucntionalized, Inspection of vaccine, Bioimaging
相關次數： 點閱：172  下載：0 
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    • 石墨烯量子點(Graphene quantum dots, GQDs)為新興的奈米材料，其光電性能和生物毒性皆有非常優異的表現，但材料特性上仍存在需要克服之缺點，如合成的產量低、量子產率低、無法精準控制尺寸和窄的發光光譜。
    本研究首先將纖維素透過破壞結晶段並利用固體酸催化劑選擇性的酸水解成葡萄糖，再將葡萄糖透過水熱法合成出石墨烯量子點，過程中透過實驗設計，調整葡萄糖合成石墨烯量子點的各項參數進行合成產率和結構缺陷之最佳化，使合成產率由12.35 %提升至24.67 %，而結構缺陷(ID/IG)由0.853減少至0.778。由MTT assay顯示由天然的生物質(纖維素)合成出的石墨烯量子點具有更低的生物毒性，(48 h , 400 μg/mL): Cell Viability > 100 %。
    本實驗透過綠色還原劑(抗壞血酸)和水熱法進行量子點的還原，有效移除量子點表面環氧官能基團並增加表面和邊緣羧基(-COOH)的比率，使量子產率提升38倍。最後進行量子點之氨基化，氨基化後，其發光性能、化學穩定性和生物毒性都大幅度的提升，比起原先的量子點，量子產率提升89倍，且經過半年時間仍維持85 %的PL強度。石墨烯量子點表面的氨基能誘導細胞自噬，使細胞活性提升。
    將氨基化石墨烯量子點應用於疫苗檢測的顯示劑和生物影像的螢光分子，在疫苗檢測上其檢測極限為2 pg/mL，線性回歸之R2 = 0.9982，且不同濃度下之變異數皆 < 0.04 %；在生物影像中，透過免疫螢光染色將氨基化石墨烯量子點標記於細胞膜之位置，較高的光穩定度，使此產品比起其他螢光染劑更適合用於螢光探針。

    關鍵字: 綠色合成、石墨烯量子點、氨基化、疫苗檢測、生物影像

    Nowadays Graphene quantum dots (GQDs) is the novel nano-materials which have the excellent performances in photoelectric and cytotoxic. Whereas there have some disadvantages of material’s character should be overcome. For example: Low product’s yield、Low quantum yield、Size uncontrollable and Narrow emission spectra. In this study, first we transfer cellulose into glucose by destroyed it’s crystallization section and selectively hydrolysis. Second we use carbonization method to synthesize GQDs from glucose molecular, and then optimize the product’s yield and defects of structure via the regulation of process parameters. Owing to epoxide groups will affect photoluminescent of GQDs, so we use two-step reduction to remove it and increase the carboxyl groups at the same time. Finally, use ammonia to functionalized GQDs. We found that both performance of photoelectric and chemical stability were increased through functionalizing. For example: PL QYs 89 times higher than further one、Maintain 85 % PL intensity after 6 months. Owing to amine groups can induce autophagy. In MTT assay, cell viability of amino-functionalized graphene quantum dots (af-GQDs) will 52% higher than GQDs which conditions are 800 μg/mL and incubated 48 h. Applied the af-GQDs to inspect of vaccine. The limit of detection is 2 pg/mL and the coefficient of variation lower than 0.04 % in each concentration of secondary antibody. In bioimaging, we can use immunohistochemistry to labeling the specific location in cell by af-GQDs. Because af-GQDs have excellent performance on PL stability, it is much more suitable than another fluorophore dye to applied in bioimaging.

    Key words : Green synthesis, GQDs , Amino-functionalized, Inspection of vaccine, Bioimaging

    口試合格證明 I 摘要 II 致謝 XIII 總目錄 XIV 表目錄 XVII 圖目錄 XVIII 第1章 緒論 1 1-1 前言 1 1-2 文獻回顧 3 1-2-1 纖維素與葡萄糖 3 1-2-2 纖維素水解技術 5 1-2-3 量子點 8 1-2-4 碳點（Carbon dots , CDs） 12 1-2-5 石墨烯量子點之光致發光機制 16 1-2-6 石墨烯量子點之光致發光特性 21 1-2-7 石墨烯量子點之合成方法 24 1-2-8 石墨烯量子點之還原 29 1-2-9 石墨烯量子點之官能化 30 1-2-10 石墨烯量子點對於細胞之作用 33 1-2-11 高壓釜水熱法與PID控制器 38 1-2-12 比色法 41 1-2-13 酵素結合免疫吸附分析法 44 1-2-14 儀器原理 45 第2章 研究動機、歷程與目標 53 2-1 研究動機 53 2-2 研究歷程與目標 54 2-2-1 纖維素水解為葡萄糖 54 2-2-2 調整水熱參數進行產率和缺陷之最佳化 55 2-2-3 氨基化石墨烯量子點 55 2-2-4 MTT檢測量子點氨基化前後之細胞活性 56 2-2-5 氨基化石墨烯量子點應用於疫苗檢測 57 2-2-6 氨基化石墨烯量子點應用於生物影像 58 第3章 實驗內容 59 3-1 纖維素選擇性水解為葡萄糖 59 3-1-1 製備固體酸催化劑 59 3-1-2 配製DNS試劑 59 3-1-3 纖維素以機械力破壞結晶段 59 3-1-4 纖維素選擇性水解為葡萄糖 60 3-2 葡萄糖水熱合成石墨烯量子點之最佳化 60 3-2-1 葡萄糖水熱合成為石墨烯量子點 60 3-2-2 過濾與透析 61 3-3 石墨烯量子點之官能化 62 3-3-1 還原石墨烯量子點 62 3-3-2 氨基化石墨烯量子點 62 3-4 MTT比色法評估官能化石墨烯量子點對細胞之影響 63 3-4-1 製備培養基和MTT試劑 63 3-4-2 細胞培養與接種 63 3-4-3 氨基化前後之石墨烯量子點與細胞共培養 63 3-4-4 MTT 呈色 64 3-5 氨基化石墨烯量子點應用於疫苗檢測 64 3-6 氨基化之石墨烯量子點應用於細胞成像 65 第4章 結果與討論 66 4-1 纖維素水解為葡萄糖 66 4-1-1 機械力破壞纖維素結晶段 66 4-1-2 固體酸催化劑 66 4-1-3 纖維素選擇性水解為葡萄糖 68 4-2 葡萄糖水熱合成石墨烯量子點之最佳化 70 4-2-1 水熱環境 70 4-2-2 水熱合成溫度 74 4-2-3 水熱合成時間 77 4-3 石墨烯量子點之官能化 81 4-3-1 還原石墨烯量子點 81 4-3-2 氨基化石墨烯量子點 88 4-4 MTT評估氨基化石墨烯量子點對細胞之影響 100 4-5 氨基化石墨烯量子點應用於疫苗檢測 102 4-6 氨基化石墨烯量子點應用於生物影像 105 第5章 總結 107 5-1 結論 107 5-2 未來展望 109 參考文獻 110

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