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研究生: 洪銘聰
Ming-Tsung Hung
論文名稱: 多頻道碳六十固定化酵素石英壓電生化感測器之研製與應用
Preparation and Application of Multi-channel Piezoelectric Crystal Bio-sensor Based on Immobilized Fullerene C60-Enzyme for Bio-Species
指導教授: 施正雄
Shih, Jeng-Shong
學位類別: 碩士
Master
系所名稱: 化學系
Department of Chemistry
論文出版年: 2006
畢業學年度: 94
語文別: 中文
論文頁數: 133
中文關鍵詞: 石英壓電感測器碳六十固定化酵素葡萄糖氧化酵素尿素分解酵素過氧化氫分解酵素
英文關鍵詞: QCM, Fullerene, Immobilized Enzyme, Glucose Oxidase, Urease, Catalase
論文種類: 學術論文
相關次數: 點閱:210下載:2
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  • 葡萄糖、尿素、過氧化氫皆為人體內重要的物種中的其中三種。本研究中,一自製震盪線路及微電腦介面的多頻道石英壓電生化感測器已成功地被建立,實驗室自行撰寫的訊號擷取程式亦可分別用來偵測待測樣品物種(葡萄糖酸、銨根離子、氧氣)之頻率訊號。此多頻道生化感測器裝置包括了一個四頻道的壓電系統(包括兩工作電極以及兩個參考電極)及一氧電極系統。
    葡萄糖氧化酶、尿素分解酶及過氧化氫分解酶皆被成功地固定化在碳六十上,而後塗佈在一多孔性材質(SiO2)上,經固定化後之酵素依然保有其催化之活性,特別是在此多頻道石英壓電系統中。將碳六十及碳六十/大環胺醚塗佈於晶片上分別作為葡萄糖酸及銨根離子的吸附劑,藉由晶片上吸附劑吸附催化後的產物,則會造成震盪頻率的下降,進而可以推得產物吸附的多寡,之後換算目標待測物的注入濃度。
    此多頻道生化感測器對於個別的目標待測物種有相當好的選擇性。模擬血液中干擾物(半乳糖、半胱胺酸、尿酸等)的存在,對整個壓電系統影響也不至於太大。經固定化之後的酵素,其活性在15天中皆可保持在90%以上。此自製多頻道偵測系統中,亦能有不錯的線性關係,R2值皆可在0.978以上,而最佳偵測下限為12.37μg/mL。利用自製的多頻道石英壓電感測系統可以用來作混合物之定性分析,利用簡單線性迴歸及多重線性迴歸(MLR)之分析方法,亦可成功的將單一頻道及多頻道之樣品作定量分析工作。

    Glucose, urea and hydrogen peroxide are important bio-species in the human body. In this study, a multi-channel piezoelectric crystal bio-sensor with the home-made computer interface for data processing was prepared and employed to detect separately various bio-species, e.g. glucose, urea and H2O2. The multi-channel PZ system includes two working electrodes and two reference electrodes and one oxygen electrode. Various immobilized fullerene C60-enzymes, e.g. C60-glucose oxidase, C60-Urease and C60-catalase, were synthesized and applied to catalyze oxidation of glucose, decomposition of urea and H2O2, respectively in this piezoelectric bio-sensor system. The C60 and C60-Cryptand-22 coated piezoelectric crystals were used to detect various products of these enzyme-catalyzed reactions. The oscillated frequency decreased due to adsorption of product catalyzed by immobilized C60-enzyme on the coating material. The C60-enzyme coated PZ bio-sensor exhibited quiet good selectivity for these bio-species. Other interfering compounds e.q. cysteine, tyrosine, uric acid and galactose in sample solutions showed nearly no interference with these C60-enzyme coated PZ bio-sensor. The activity of immobilized C60-enzyme can be hold over 90% in 15 days. The home-made multi-channels Piezoelectric crystal bio-sensor showed good detection limit of 12.37μg/mL and R2 values of calibration curve were more than 0.978. The piezoelectric crystal bio-sensor system can distinguish individual components in the mixed samples. Furthermore, multivariate linear regression (MLR) was employed to make quantitative analysis for these biospecies.

    英文摘要 I 中文摘要 III 目錄 IV 圖目錄 VIII 表目錄 XII 表目錄 XII 第一章 緒論 1 1-1 碳六十化學 1 1-1-1 碳六十的發現 1 1-1-2 碳六十的命名 2 1-1-3 碳六十的基本性質 2 1-1-4 碳六十的化學反應 6 1-1-5 碳六十的應用 8 1-2 冠狀醚的簡介與應用 10 1-2-1冠狀醚的簡介 10 1-2-2 冠狀醚的應用 13 1-3 壓電晶體之壓電性 16 1-3-1 壓電性 16 1-3-2 壓電材料 18 1-3-2.1 石英(Quartz) 18 1-3-2.2 羅爾賽鹽(Rochelle salt) 22 1-3-2.3 鈦酸鋇瓷 23 1-3-3 石英震盪器 25 1-3-4 AT cut石英晶體的特性 26 1-3-5 石英震盪器的線路 27 1-3-6 震盪頻率的量測 29 1-3-7 石英微量天平 31 1-3-8 液相石英壓電感測器之應用 36 1-4 酵素化學 39 1-4-1 酵素本質 39 1-4-2 酵素的發現 40 1-4-3 酵素的分類 40 1-4-4 酵素的特性 42 1-4-5 酵素溫度效應 45 1-4-6 酵素pH 值效應 46 1-4-7 溶劑對酵素的影響 47 1-4-8 固定化酵素 48 1-4-8 酵素於生化感測器之應用 53 1-5 迴歸分析 56 1-6 研究目的及動機 57 第二章 實驗部分 58 2-1 藥品及器材 58 2-2 碳六十/大環胺醚化合物之合成 58 2-3 實驗系統 60 2-3-1 多頻道液相壓電系統 60 2-3-1-1 石英震盪線路 60 2-3-1-2 多頻道線路設計 63 2-3-1-3 多頻道靜相生化感測系統 64 2-4 石英晶片的處理 65 2-4-1 配製表面塗佈液 66 2-4-2 表面塗佈法 66 2-4-2-1旋轉塗佈法(Spin coating) 66 2-4-2-2浸漬塗佈法(Dip-coating) 68 2-5 實驗裝置及步驟 69 2-5-1 碳六十與酵素間的作用力關係 69 2-5-2 碳六十/酵素的合成 71 2-5-3 碳六十/酵素固定化方式 71 第三章 實驗結果與討論 73 3-1 碳六十與酵素間的作用力探討 73 3-1-1 碳六十與酵素間的吸附與脫附現象 73 3-2 碳六十/大環胺醚的鑑定 77 3-3 碳六十固定化酵素 77 3-3-1 碳六十/固定化酵素的鑑定 77 3-3-2 碳六十/固定化酵素之活性及訊號確定 78 3-3-3 碳六十/固定化酵素多次使用之可行性 86 3-3-4 碳六十/固定化酵素的各項評估 86 3-3-4-1 不同pH值對碳六十/酵素之影響 86 3-3-4-2 不同溫度對碳六十/酵素之影響 91 3-3-4-3 保存期限之評估 91 3-4 多頻道生化感測系統 98 3-4-1 石英壓電感測系統各頻道之感測 98 3-4-2 多頻道生化感測器之穩定性 101 3-4-3 石英壓電感測器之脫附訊號 102 3-4-4 石英壓電系統晶片塗佈物之塗佈量效應 104 3-4-5 多頻道生化感測系統之分析物濃度效應與再現性 104 3-4-6 生化感測器之定性分析 112 3-4-7 血液其他成分對葡萄糖及尿素偵測之干擾研究 114 3-4-8 混合生化樣本之定量分析 114 第四章 結論 125 參考文獻 126 附錄一 130 附錄二 133

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