Salimi Abdollah, Hallaj Rahman, Soltanian Saied
Department of Chemistry, University of Kurdistan, P.O.Box 416, Sanandaj, Iran.
Biophys Chem. 2007 Nov;130(3):122-31. doi: 10.1016/j.bpc.2007.08.004. Epub 2007 Aug 24.
Cyclic voltammetry at potential range -1.1 to 0.5 V from aqueous buffer solution (pH 7) containing CoCl(2) produced a well defined cobalt oxide (CoOx) nanoparticles deposited on the surface of glassy carbon electrode. The morphology of the modified surface and cobalt oxide formation was examined with SEM and cyclic voltammetry techniques. Hemoglobin (Hb) was successfully immobilized in cobalt-oxide nanoparticles modified glassy carbon electrode. Immobilization of hemoglobin onto cobalt oxide nanoparticles have been investigated by cyclic voltammetry and UV-visible spectroscopy. The entrapped protein can take direct electron transfer in cobalt-oxide film. A pair of well defined, quasi-reversible cyclic voltammetric peaks at about -0.08 V vs. SCE (pH 7), characteristic of heme redox couple (Fe(III)/Fe(II)) of hemoglobin, and the response showed surface controlled electrode process. The dependence of formal potential (E(0')) on the solution pH (56 mV pH(-1)) indicated that the direct electron transfer reaction of hemoglobin was a one-electron transfer coupled with a one proton transfer reaction process. The average surface coverage of Hb immobilized on the cobalt oxide nanoparticles was about 5.2536x10(-11) mol cm(-2), indicating high loading ability of nanoparticles for hemoglobin entrapment. The heterogeneous electron transfer rate constant (k(s)) was 1.43 s(-1), indicating great of facilitation of the electron transfer between Hb and electrodeposited cobalt oxide nanoparticles. Modified electrode exhibits a remarkable electrocatalytic activity for the reduction of hydrogen peroxide and oxygen. The Michaels-Menten constant K(m) of 0.38 mM, indicating that the Hb immobilized onto cobalt oxide film retained its peroxidases activity. The biosensor exhibited a fast amperometric response <5 s, a linear response over a wide concentration range 5 microM to 700 microM and a low detection limit 0.5 microM. According to the direct electron transfer property and enhanced activity of Hb in cobalt oxide film, a third generation reagentless biosensor without using any electron transfer mediator or specific reagent can be constructed for determination of hydrogen peroxide in anaerobic solutions.
在含有CoCl₂的水缓冲溶液（pH 7）中，于-1.1至0.5 V的电位范围内进行循环伏安法，可在玻碳电极表面产生定义明确的氧化钴（CoOx）纳米颗粒。采用扫描电子显微镜（SEM）和循环伏安法技术研究了修饰表面的形态和氧化钴的形成情况。血红蛋白（Hb）成功固定在氧化钴纳米颗粒修饰的玻碳电极上。通过循环伏安法和紫外可见光谱法研究了血红蛋白在氧化钴纳米颗粒上的固定情况。被包埋的蛋白质能够在氧化钴膜中进行直接电子转移。在相对于饱和甘汞电极（SCE）约-0.08 V（pH 7）处出现一对定义明确的准可逆循环伏安峰，这是血红蛋白血红素氧化还原对（Fe(III)/Fe(II)）的特征峰，且响应显示为表面控制的电极过程。形式电位（E(0')）对溶液pH的依赖性（56 mV pH⁻¹）表明，血红蛋白的直接电子转移反应是一个单电子转移与一个单质子转移反应过程。固定在氧化钴纳米颗粒上的血红蛋白平均表面覆盖度约为5.2536×10⁻¹¹ mol cm⁻²，表明纳米颗粒对血红蛋白包埋具有较高的负载能力。异相电子转移速率常数（k(s)）为1.43 s⁻¹，表明血红蛋白与电沉积氧化钴纳米颗粒之间的电子转移得到了极大促进。修饰电极对过氧化氢和氧气的还原表现出显著的电催化活性。米氏常数K(m)为0.38 mM，表明固定在氧化钴膜上的血红蛋白保留了其过氧化物酶活性。该生物传感器呈现出快速的安培响应<5 s，在5 μM至700 μM的宽浓度范围内具有线性响应，检测限低至0.5 μM。基于血红蛋白在氧化钴膜中的直接电子转移特性和增强的活性，可构建一种无需使用任何电子转移介质或特定试剂的第三代无试剂生物传感器，用于测定厌氧溶液中的过氧化氢。
