Immobilization of enzymes on the nano-Au film modified glassy carbon electrode for the determination of hydrogen peroxide and glucose

A new enzyme-based amperometric biosensor for hydrogen peroxide was developed relying on the efficient immobilization of horseradish peroxidase (HRP) to a nano-scaled particulate gold (nano-Au) film modified glassy carbon electrode (GC). The nano-Au film was obtained by a chitosan film which was first formed on the surface of GC. The high affinity of chitosan for nano-Au associated with its amino groups resulted in the formation of nano-Au film on the surface of GC. The film formed served as an intermediator to retain high efficient and stable immobilization of the enzyme. H2O2,was detected using hydroquinone as an electron mediator to transfer electrons between the electrode and HRP. The HRP immobilized on nano-Au film maintained excellent electrocatalytical activity to the reduction of H2O2. The experimental parameters such as the operating potential of the working electrode, mediator concentration and pH of background electrolyte were optimized for best analytical performance of amperometry. The linear range of detection for H2O2 is from 6.1 x 10(-6) to 1.8 x 10(-3) mol L-1 with a detection limit of 6.1 mumol L-1 based on signal/noise = 3. The proposed HRP enzyme sensor has the features of high sensitivity (0.25 Almol(-1)cm(-2)), fast response time (t(90%) less than or equal to 10 s) and a long-term stability (> 1 month). As an extension, glucose oxidase (GOD) was chemically bound to HRP-modified electrode. A GOD/HRP bienzyme-modified electrode formed in this way can be applied to the determination of glucose with satisfactory performance.