Polythiophene supported MnO 2 nanoparticles as nano-stabilizer for simultaneously electrostatically immobilization of d-amino acid oxidase and hemoglobin as efficient bio-nanocomposite in fabrication of dopamine bi-enzyme biosensor.

A novel, highly selective and sensitive voltammetric bi-enzyme biosensor for sensing dopamine (DA), in presence of H2 O2 which resulted as d-alanine enzymatic oxidation, was fabricated on the basis of simultaneously electrostatically immobilization of d-amino acid oxidase (DAAO) and hemoglobin (Hb) on MnO2 nanoparticles (MnO2 NPs) enriched poly thiophene (PTh). Cyclic voltammetry (CV), technique was applied for electropolymerization of thiophene on glassy carbon electrode (GCE) surface and MnO2 NPs dispersed on PTh network by soaking PTh/GCE in potassium permanganate (KMnO4 ) solution. Excellent catalytic properties and large surface area of MnO2 NPs/PTh composite caused it was used as an enzymes immobilization host in this developed bi-enzyme biosensor. The developed biosensor was characterized by scanning electron microscopy (SEM), energy dispersive X-ray analysis (EDX), and CV. The performance of the modified bi-enzyme biosensor was investigated in terms of its response time, detection limit, sensitivity, stability and selectivity in a lab environment. The composite of DAAO-Hb/MnO2 NPs/PTh to construct a bi-enzyme biosensor in this study showed a linear response with DA in the concentration range of 0.04-9.0μM with R-squared value of 0.994 (for S/N=3) and its sensitivity and detection limite were about 12.801μA/μM and 41nM respectively. Also this bi-enzyme biosensors exhibited high selectivity, rapid response (5s) and long-term stability (42days). At the end, the proposed biosensor was applied successfully in human serum as real sample.