A hydrogen peroxide biosensor has been presented in this paper. Gold nanoparticles (GNs) was self-assembled onto a platinum electrode to form a negatively charged surface, and cationic dye-thionine (Thio) was linked by Au—S and Au—N covalent bond and electrostatic adsorption as an electron transfer mediator. The mixture of GNs and horseradish peroxidase (HRP) was assembled on the electrode at the same procedure, and chitosan (Chit) was dropped onto the electrode surface to fabricate a novel hydrogen peroxide biosensor. The effects of applied potential, pH and temperature on the response currents have been investigated. The surface configuration, electrochemical impedance, reproducibility and stability have also been researched. Activation energy for enzymatic reaction of the biosensor was 12.4 kJ/mol, and the appar-ent Michaelis-Menten constant was 6.5×10^－4 mo/L. And the linear range of the biosensor was from 5.6×10^－5 to 2.6×10^－3 mol/L with a detection limit of 1.5×10^－5 mol/L. The method has been applied to construct a bienzyme-based biosensor of HRP and glucose oxidase (GOD) for the determination of glucose.

Key words: gold nanoparticle, thionine, chitosan, hydrogen peroxide, glucose, biosensor