以四氯金酸和L-半胱氨酸为原料,合成了L-半胱氨酸功能化纳米金粒子,将此纳米粒子修饰在金盘电极表面并共价偶联漆酶分子.以循环伏安法研究了此固定漆酶电极在无氧磷酸盐缓冲液中电化学行为和催化氧还原能力,并进一步评估了其作为氧传感器使用的性能:以计时电流法测定其对氧气的检测限,与氧气的亲和力(以米氏常数表征),研究了传感器的长期使用性、热稳定性和pH-催化电流关系.结果表明,此固定漆酶电极可以实现漆酶活性中心T2与导电纳米粒子间的直接电子迁移而无需任何外加电子中介体(氧化还原峰的式电位为192.5 mVvsAgCl/Ag),并在接近漆酶活性中心T3氧化还原式电位(780 mV vs NHE)附近发生氧还原；测得的固酶电极与氧气的亲和力较高(米氏常数为216.4 μmol/L)且检测限低达0.22 μmol/L,在4℃下保存60d后活力仍然保持初始活力的大约78％.但这种电极的热稳定性较差,受pH值影响较明显,在pH值近于生理条件时几乎完全丧失活力.%L-Cysteine functionalized nanogold particles were synthesized with hydrogen tetrachloroaurate tetrahydrate and L-cysteine as raw materials. The functionalized nanogold particles were attached to the surface of gold disk electrode and then were covalently coupled with laccase. Electrochemical behavior of this laccase-based electrode in deaerated phosphate buffer solution and its electrocatalytic effect for oxygen reduction were investigated by cyclic voltammetry. Furthermore, the performance of this laccase based cathode as oxygen sensor was evaluated: oxygen detection limit of this sensor and its affinity towards oxygen ( characterized with Michaelis-Menten constant KM ) were measured with chronoamperometry. The long-term usability, thermal stability of O2 sensor together with the relationship between pH value and catalytic current were also studied. The results indicate that a direct electron transfer between the redox active centre T2 of laccase and the conductive functionalized nanogold particle occurs without any additional electron relay ( formal potential of redox peaks; 192. 5 mV vs AgCl/Ag) , and oxygen reduction occurs at a potential similar to that of T3 redox centre in laccase (780 mV vs NHE). The affinity of the laccase based sensor towards O2 was relatively high (KM =216.4 |xmol/L) and the detection limit of oxygen was 0.22 p,mol/L. Catalytic current for oxygen reduction can still retain approximately 78% of the initial value after storage at 4 t refrigerator for 60 days. However, this laccase based O2 sensor displayed inferior thermal stability, and its catalytic current corresponding to O2 reduction was strongly influenced by pH value in buffer solution; when pH value of the buffer solution comes close to physiological condition, near all catalytic activity towards O2 was lost.
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