類核黃素修飾電極分析與應用; 電化學分析

Abstract

本實驗室曾對Phenothiazine化合物的物理化學性質進行探討，發現這類化合物，如toluidine blue，具有類似黃核素的電子傳遞特性以及作為蛋白質吸附劑的應用潛力，並可藉由電化學技巧修將之飾於電極表面。根據這些性質，本論文藉由phenothiazine以及電化學技巧如交流阻抗分析法（Electrochemical impedance spectroscopy，簡稱EIS）與電化學石英振盪天平（Electrochemical quartz crystal microbalance，簡稱EQCM），對蛋白質進行定量分析。實驗結果顯示：若將toluidine blue固定於導電玻璃電極上後，再以Fe(CN)63-/4-作為分子探針，分子探針在電極表面的電子傳遞阻抗（Charge transfer resistance）會因蛋白質添加而逐漸上升，其中以葡萄糖氧化脢的影響最鉅。EQCM分析也顯示類似結果，與EIS所得結果極為吻合。此外，我們也利用原子力顯微鏡（Atomic force microscope，簡稱AFM）與掃瞄式穿隧顯微鏡（Scanning tunneling microscope，簡稱STM）觀察蛋白質在電極表面的吸附行為。根據實驗結果，我們證實Phenothiazine化合物具有作為蛋白質分子膠的應用潛力，也證實交流阻抗與電化學石英振盪天平技巧是分析蛋白質含量的有效工具之一。Phenothiazines, such as toluidine blue, are useful model compounds for mimicking the electron transfer in riboflavin system. Since they are also effective molecular glues for proteins, we carried out studies in this thesis to investigate their application potential in protein analysis. Analyses with the electrochemical impedance spectroscopic (EIS) and electrochemical quartz crystal microbalance techniques (EQCM) revealed that the electron transfer associated with Fe(CN)63-/4- would be imposed with increasing impedance at the toluidine blue-modified electrodes during the incorporation of proteins like glucose oxidase (GOx), giving rise to increasing charge-transfer resistance and increasing mass for the electrodes. Among the tested proteins, GOx stood out to be the one with the highest affinity for the phenothiazine-modified electrodes. For the adsorption behavior, we also analyzed the electrodes with atomic force microscopic (AFM) and scanning tunneling microscopic (STM) technique. Proteins were spotted on the electrodes, fully supporting the hypotheses made based on EIS and EQCM. According to these results, we conclude tat phenothiazine compounds are effective molecular glues for proteins and that EIS and EQCM are useful techniques for protein analysis.