Simultaneous electrochemical determination of superoxide anion radical and nitrite using Cu,ZnSOD immobilized on carbon nanotube in polypyrrole matrix

A novel highly sensitive biosensor for the direct and simultaneous determination of superoxide anion radical (O-2(center dot-)) and nitrite (NO2-) was developed by incorporation of carbon nanotube (CNT) solubilized in nafion in polypyrrole (PPy) matrix on Pt electrode followed by immobilization of Cu,ZnSOD (SOD1) on it. The CNT/PPy nanocomposite electrode enhanced the immobilization of SOD1 and promoted the electron transfer of SOD1 minimizing its fouling effect. The surface morphological images of PPy and CNT-PPy nanocomposite on Pt electrode were obtained by scanning electron microscopy exhibiting highly microporous structures. The electrochemical behavior of the biosensor investigated by cyclic voltammetry revealed that the SOD1 immobilized electrode showed characteristic of SOD1 quasi-reversible redox peaks with a formal potential of +0.065 V vs. Ag/AgCl. The biosensor exhibited a linear response over the concentration range from 0.1 to 750 mu M, with a detection limit of 0.1 +/- 0.03 mu M for O-2(center dot-) and a corresponding linear range of 0.5-2000 mu M, with a detection limit of 0.5 +/- 0.025 mu M for NO2-. In addition, the biosensor exhibited high sensitivity, good reproducibility and retained stability over 30 days. This modified electrode was quite effective not only in detecting O-2(center dot-) and NO2- independently but also determining the concentration of O-2(center dot-) and NO2- simultaneously in vitro and from cancer cells. (C) 2010 Elsevier B.V. All rights reserved.