基于脂质立方液晶相生物电催化体系的构筑及生物电子应用
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摘要
脂质立方液晶相是由一定浓度的两亲性物质在水相中自组装成含双连续水道结构的闭合脂质双层,脂质双分子层进而扭曲成具有类生物膜性质的一种三维立体结构。脂质立方液晶相具有粘性大、生物相容性好、在水相中能稳定存在的特点。这些特点使得脂质立方液晶相可作为包埋生物催化剂(酶或蛋白质)的基质,建立生物电催化体系,进而构建生物电子装置(生物传感器、生物燃料电池等)。本论文选择单油酸甘油酯(Monoolein,MO)制备脂质立方液晶相,并以此作为包埋蛋白质(血红蛋白)、酶(乙醇脱氢酶、漆酶)的基质,构筑生物电子装置。具体概括如下:
1、制备了脂质立方液晶相包埋氧化还原性蛋白质,血红蛋白(Hemoglobin,Hb)的修饰电极,考察了血红蛋白与电极间直接电子转移行为,进而构建了过氧化氢第三代生物传感器。结果表明,血红蛋白与电极间的电子转移动力学常数(Ks)为3.03 s-1,催化底物过氧化氢的表观米氏常数(ΚaMpp)为0.27 mM,所构建第三代过氧化氢传感器的线性范围为7~239μM,检测限为3.1μM。
2、制备了脂质立方液晶相包埋四氯苯醌(Tetrachlorobenzoquinone,TCBQ)的碳纳米管修饰电极(MO-TCBQ/SWNT/GC),考察了该电极对NADH的电催化性能,对NADH检测的线性范围为5~1650μM,检测限为0.82μM。进一步制备了TCBQ和乙醇脱氢酶(Alcohol Dehydrogenase,ADH)共包埋于脂质立方液晶相的生物电极(MO-TCBQ-ADH/SWNT/GC),进而构建了基于脱氢酶的乙醇生物传感器,该生物电极对乙醇检测的线性范围为0.2~13 mM,检测限为0.05 mM。
3、制备了甲苯胺蓝(Toluidine blue,TB)和乙醇脱氢酶(ADH)共包埋于脂质立方液晶相中的修饰电极(MO-TB-ADH/GC),基于该电极对乙醇良好的催化氧化性能构建了生物燃料电池的阳极。制备了漆酶(Laccase)包埋于脂质立方液晶相中的修饰电极(MO-Laccase/GC),基于该电极在ABTS溶液中对氧气良好的催化还原性能,构建了电池的阴极。基于此进一步构筑了基于脂质立方液晶相的无隔膜型乙醇/氧气生物燃料电池。研究表明,在室温和氧气饱和的条件下,电池的开路电位为0.38 V,在0.06 V的工作电压下,输出功率可达到0.2μW/cm~2。
Liquid-crystalline lipid cubic phase is formed by self-assembling of polar lipids in aqueous media.The internal structure consists of one congruent lipid bilayer, forming a three-dimensional and well-ordered structure interwoven by aqueous channels.The lipidic cubic phase is of high viscosity, good biocompatibility,and good stability in the presence of excess water.These special characteristics make it suitable for hosting the biocatalysts ( proteins or enzymes ) on electrode surface, and therefore establishing bioelectrocatalysis systems to build bioelectronic devices ( biosensors,biofuel cells etc. ).
In this thesis,liquid crystal cubic phase formed with monoolein has been used as immobilizing matrix to host protein ( hemoglobin ),enzyme ( alcohol dehydrogenase, laccase ) on glass carbon electrode surface to construct bioelectronic devices.The details are summarized as follows:
1. Liquid crystal cubic phase formed with monoolein has been used as immobilizing matrix to host redox protein hemoglobin. The direct electron transfer between hemoglobin and electrode was investigated and the third generation biosensor of hydrogen peroxide was constructed.Based on this system,the enhanced average kinetic electron transfer rate constant ks of 3.03 s?1 and the small apparent Michaelis-Menten constantΚaMpp of 0.27 mM were obtained. The proposed hydrogen peroxide biosensor showed a linear range of 7 ~239μM with a detection limit of 3.1μM.
2. A modified electrode, MO-TCBQ/SWNT/GC, was constructed utilizing the lipidic cubic phase to immobilize Tetrachlorobenzoquinone ( TCBQ ) on SWNT-based electrode. A NADH sensor was constructed based on the electrocatalytic activity of TCBQ and NADH. The proposed NADH sensor showed a linear range of 5~1650μM with a detection limit of 0.82μM.A biosensor for the detection of ethanol was further constructed based on a bioelectrode MO-TCBQ-ADH/SWNT/GC utilizing the lipidic cubic phase to co-immobilize TCBQ and ADH on SWNT-modified electrode.The biosensor exhibits a wide linear dynamic range ( from 0.2 to 13 mM ),and low detection limit of 0.05 mM.
3. A bioelectrode, MO-TB-ADH/GC,was prepared based on lipid cubic phase to co-entrap alcohol dehydrogenase and toluidine blue ( TB ).A bioanode was constructed based on the good electrocatalytic oxidation activity of the prepared bioelectrode toward ethanol.A biocathode,MO-Laccase/GC,was prepared with a similar methode.The electrode with Laccase as biocatalyst to reduce oxygen reduction.A new type of membrane-less ethanol/O2 biofuel cell has been constructed.At room temperature and under oxygen-saturation condition,the biofuel cell was found to have a open potential of 0.38 V and a power output of 0.2μW/cm2 under the work potential of 0.06 V.
1、制备了脂质立方液晶相包埋氧化还原性蛋白质,血红蛋白(Hemoglobin,Hb)的修饰电极,考察了血红蛋白与电极间直接电子转移行为,进而构建了过氧化氢第三代生物传感器。结果表明,血红蛋白与电极间的电子转移动力学常数(Ks)为3.03 s-1,催化底物过氧化氢的表观米氏常数(ΚaMpp)为0.27 mM,所构建第三代过氧化氢传感器的线性范围为7~239μM,检测限为3.1μM。
2、制备了脂质立方液晶相包埋四氯苯醌(Tetrachlorobenzoquinone,TCBQ)的碳纳米管修饰电极(MO-TCBQ/SWNT/GC),考察了该电极对NADH的电催化性能,对NADH检测的线性范围为5~1650μM,检测限为0.82μM。进一步制备了TCBQ和乙醇脱氢酶(Alcohol Dehydrogenase,ADH)共包埋于脂质立方液晶相的生物电极(MO-TCBQ-ADH/SWNT/GC),进而构建了基于脱氢酶的乙醇生物传感器,该生物电极对乙醇检测的线性范围为0.2~13 mM,检测限为0.05 mM。
3、制备了甲苯胺蓝(Toluidine blue,TB)和乙醇脱氢酶(ADH)共包埋于脂质立方液晶相中的修饰电极(MO-TB-ADH/GC),基于该电极对乙醇良好的催化氧化性能构建了生物燃料电池的阳极。制备了漆酶(Laccase)包埋于脂质立方液晶相中的修饰电极(MO-Laccase/GC),基于该电极在ABTS溶液中对氧气良好的催化还原性能,构建了电池的阴极。基于此进一步构筑了基于脂质立方液晶相的无隔膜型乙醇/氧气生物燃料电池。研究表明,在室温和氧气饱和的条件下,电池的开路电位为0.38 V,在0.06 V的工作电压下,输出功率可达到0.2μW/cm~2。
Liquid-crystalline lipid cubic phase is formed by self-assembling of polar lipids in aqueous media.The internal structure consists of one congruent lipid bilayer, forming a three-dimensional and well-ordered structure interwoven by aqueous channels.The lipidic cubic phase is of high viscosity, good biocompatibility,and good stability in the presence of excess water.These special characteristics make it suitable for hosting the biocatalysts ( proteins or enzymes ) on electrode surface, and therefore establishing bioelectrocatalysis systems to build bioelectronic devices ( biosensors,biofuel cells etc. ).
In this thesis,liquid crystal cubic phase formed with monoolein has been used as immobilizing matrix to host protein ( hemoglobin ),enzyme ( alcohol dehydrogenase, laccase ) on glass carbon electrode surface to construct bioelectronic devices.The details are summarized as follows:
1. Liquid crystal cubic phase formed with monoolein has been used as immobilizing matrix to host redox protein hemoglobin. The direct electron transfer between hemoglobin and electrode was investigated and the third generation biosensor of hydrogen peroxide was constructed.Based on this system,the enhanced average kinetic electron transfer rate constant ks of 3.03 s?1 and the small apparent Michaelis-Menten constantΚaMpp of 0.27 mM were obtained. The proposed hydrogen peroxide biosensor showed a linear range of 7 ~239μM with a detection limit of 3.1μM.
2. A modified electrode, MO-TCBQ/SWNT/GC, was constructed utilizing the lipidic cubic phase to immobilize Tetrachlorobenzoquinone ( TCBQ ) on SWNT-based electrode. A NADH sensor was constructed based on the electrocatalytic activity of TCBQ and NADH. The proposed NADH sensor showed a linear range of 5~1650μM with a detection limit of 0.82μM.A biosensor for the detection of ethanol was further constructed based on a bioelectrode MO-TCBQ-ADH/SWNT/GC utilizing the lipidic cubic phase to co-immobilize TCBQ and ADH on SWNT-modified electrode.The biosensor exhibits a wide linear dynamic range ( from 0.2 to 13 mM ),and low detection limit of 0.05 mM.
3. A bioelectrode, MO-TB-ADH/GC,was prepared based on lipid cubic phase to co-entrap alcohol dehydrogenase and toluidine blue ( TB ).A bioanode was constructed based on the good electrocatalytic oxidation activity of the prepared bioelectrode toward ethanol.A biocathode,MO-Laccase/GC,was prepared with a similar methode.The electrode with Laccase as biocatalyst to reduce oxygen reduction.A new type of membrane-less ethanol/O2 biofuel cell has been constructed.At room temperature and under oxygen-saturation condition,the biofuel cell was found to have a open potential of 0.38 V and a power output of 0.2μW/cm2 under the work potential of 0.06 V.
引文
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