【摘要】： 血紅蛋白(Hemoglobin, Hb)是脊椎動(dòng)物紅細(xì)胞內(nèi)的呼吸蛋白,由珠蛋白和血紅素組成,是血液中分子氧的載體。研究血紅蛋白的電化學(xué)行為,不僅可以探索生物大分子之間的長(zhǎng)程電子傳遞的機(jī)理,進(jìn)一步認(rèn)識(shí)生物大分子的結(jié)構(gòu)與功能之間的關(guān)系,而且有助于我們了解生命體系內(nèi)的物質(zhì)代謝和能量交換的過(guò)程及生命過(guò)程的氧化還原機(jī)理,從而更有效地開發(fā)和研制具有高催化性能的電化學(xué)生物傳感器。 本論文以實(shí)現(xiàn)血紅蛋白的直接電子傳遞為目的,分別選取具有良好生物相容性的碳納米材料、高分子材料等仿生材料作為血紅蛋白的固定材料,并采用適當(dāng)?shù)姆椒▽⒀t蛋白固定于電極表面,基于這些材料與血紅蛋白之間的特殊相互作用,實(shí)現(xiàn)了血紅蛋白與基底電極之間的直接電子傳遞。這些固定材料不但為血紅蛋白保持自然結(jié)構(gòu)和生物活性提供了適宜的微環(huán)境,而且顯著提高了血紅蛋白與電極間的電子傳遞速率�；诖酥苽涞幕瘜W(xué)修飾電極能夠催化還原過(guò)氧化氫(H_2O_2)、一氧化氮(NO)等小分子。本論文的主要工作有: (1)首次將乙炔黑與離子液體1-丁基-3-甲基咪唑六氟磷酸鹽(BMIMPF_6)組合制得復(fù)合膜,并成功將Hb固定在電極表面,為Hb的電化學(xué)研究建立了新的平臺(tái)。用紫外-可見(jiàn)光譜法和電化學(xué)交流阻抗對(duì)Hb/AB-BMIMPF_6復(fù)合膜進(jìn)行了表征,結(jié)果表明Hb在AB-BMIMPF_6復(fù)合膜中保持其生物活性,同時(shí)該膜促進(jìn)了Hb與電極間電子的傳遞。該修飾電極對(duì)H_2O_2、NO的還原具有良好的催化作用。 (2)采用一種新型合成的高分子材料-纖維素季銨鹽,利用靜電吸附作用,采用層層自組裝法將荷負(fù)電的血紅蛋白包埋于荷正電的纖維素季銨鹽內(nèi),并固定在電極表面,從而實(shí)現(xiàn)了血紅蛋白的直接電化學(xué)。研究表明,固定在纖維素季銨鹽膜內(nèi)的血紅蛋白沒(méi)有變性,能催化還原H_2O_2�；诖�,我們構(gòu)建了一種無(wú)媒介體的新型過(guò)氧化氫傳感器。 (3)結(jié)合亞甲基藍(lán)和納米活性炭微球的優(yōu)異性能,制備出一種復(fù)合膜,并成功實(shí)現(xiàn)Hb的包埋固定。Hb在此復(fù)合膜內(nèi)表現(xiàn)出優(yōu)良的直接電化學(xué)行為。原子力顯微鏡、電化學(xué)交流阻抗、紫外-可見(jiàn)光譜研究表明,該復(fù)合膜與Hb之間存在著靜電作用,它改變Hb的結(jié)構(gòu)取向,因而更利于電子傳遞。固定于亞甲基藍(lán)-納米活性炭微球復(fù)合膜內(nèi)的Hb也表現(xiàn)出了良好的生物催化活性,能催化還原H_2O_2。 (4)采用β-環(huán)糊精和離子液體1-丁基-3-甲基咪唑四氟硼酸鹽(BMIMBF_4)制得修飾膜,利用它們優(yōu)異的生物相容性包埋固定血紅蛋白,實(shí)現(xiàn)了血紅蛋白在電極表面的直接電子傳遞。紫外-可見(jiàn)光譜實(shí)驗(yàn)表明,固定于β-環(huán)糊精-BMIMBF_4復(fù)合膜內(nèi)的血紅蛋白仍然能維持其原始構(gòu)象和生物活性。該復(fù)合膜修飾的玻碳電極能有效地催化H_2O_2還原,同時(shí)響應(yīng)快、穩(wěn)定性高。
[Abstract]:Hemoglobin (Hemoglobin, Hb) is a respiratory protein in erythrocytes of vertebrates. It is composed of globin and heme, and is the carrier of molecular oxygen in blood. The study of the electrochemical behavior of hemoglobin can not only explore the mechanism of long-range electron transport among biomolecules, but also further understand the relationship between the structure and function of biological macromolecules. It also helps us to understand the process of substance metabolism and energy exchange and the mechanism of redox in the life process, so that the electrochemical biosensor with high catalytic performance can be developed and developed more effectively. In order to realize the direct electron transfer of hemoglobin, carbon nanomaterials with good biocompatibility and biomimetic materials such as polymer materials were selected as the immobilized materials of hemoglobin. The direct electron transfer between hemoglobin and substrate electrode was realized based on the special interaction between hemoglobin and hemoglobin. These immobilization materials not only provide a suitable microenvironment for hemoglobin to maintain its natural structure and biological activity, but also significantly increase the electron transfer rate between hemoglobin and electrode. The chemically modified electrode can catalyze the reduction of hydrogen peroxide (H_2O_2), nitric oxide (NO) and other small molecules. The main work of this thesis is as follows: (1) the composite membrane was prepared by the combination of acetylene black and ionic liquid 1 Ding Ji 3 methyl imidazolium hexafluorophosphate (BMIMPF_6) for the first time, and Hb was successfully immobilized on the electrode surface. A new platform for electrochemical research of Hb is established. The Hb/AB-BMIMPF_6 composite film was characterized by UV-Vis spectroscopy and electrochemical impedance spectroscopy. The results showed that Hb kept its biological activity in the AB-BMIMPF_6 composite film and promoted the electron transfer between the Hb and the electrode. The modified electrode has a good catalytic effect on the reduction of H _ 2O _ 2no. (2) A new synthetic polymer material, cellulose quaternary ammonium salt, is used for electrostatic adsorption. The hemoglobin with negative charge was encapsulated in the positively charged quaternary ammonium cellulose salt and immobilized on the surface of the electrode by means of layer by layer self-assembly method, thus realizing the direct electrochemistry of hemoglobin. The results showed that hemoglobin immobilized in cellulose quaternary ammonium salt film had no denaturation and could catalyze the reduction of H _ 2O _ 2. Based on this, we constructed a novel hydrogen peroxide sensor without medium. (3) A composite membrane was prepared by combining the excellent properties of methylene blue and nano-activated carbon microspheres. The encapsulation and fixation of Hb were successfully realized. Hb showed excellent direct electrochemical behavior in the composite film. Atomic force microscopy (AFM) electrochemical impedance spectroscopy (EIS) and UV-Vis spectroscopy show that there is electrostatic interaction between the composite film and Hb which changes the structural orientation of Hb and is more favorable for electron transfer. The Hb immobilized in the composite membrane of methylene blue and nano-activated carbon also showed good biocatalytic activity. The modified membrane was prepared from 尾 -cyclodextrin and ionic liquid 1-Ding Ji -3-methyl imidazolium tetrafluoroborate (BMIMBF_4), and hemoglobin was immobilized by their excellent biocompatibility. The direct electron transfer of hemoglobin on the electrode surface was realized. UV-Vis spectra showed that hemoglobin immobilized in 尾 -cyclodextrin-BMIMBF_4 composite membrane could still maintain its original conformation and biological activity. The modified glassy carbon electrode can effectively catalyze the reduction of H_2O_2 with fast response and high stability.
【學(xué)位授予單位】：湘潭大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2010
【分類號(hào)】：R341

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