A Superoxide Sensor Based on a Multilayer Cytochrome c Electrode

Abstract

A novel multilayer cytochrome c electrode for the quantification of superoxide radical concentrations is introduced. The electrode consists of alternating layers of cytochrome c and poly(aniline(sulfonic acid)) on a gold wire electrode. The formation of multilayer structures was proven by SPR experiments. Assemblies with 2-15 protein layers showed electrochemical communication with the gold electrode. For every additional layer, a substantial increase in electrochemically active cytochrome c (cyt. c) was found. For electrodes of more than 10 layers, the increase was more than 1 order of magnitude as compared to monolayer electrode systems. Thermodynamic and kinetic parameters of the electrodes were characterized. The mechanism of electron transfer within the multilayer assembly was studied, with results suggesting a protein-protein electron-transfer model. Electrodes of 2-15 layers were applied to the in vitro quantification of enzymatically generated superoxide, showing superior sensitivity as compared to a monolayer-based sensor. An electrode with 6 cyt. c/PASA layers showed the highest sensitivity of the systems studied, giving an increase in sensitivity of half an order of magnitude versus the that of the monolayer electrode. The stability of the system was optimized using thermal treatment, resulting in no loss in sensor signal or protein loading after 10 successive measurements or 2 days of storage.

Electron-Transfer Rate in Potential-Modulated Redox Reactions with Electro-Active Optical Waveguides

“…Usually electron-transfer kinetics of redox species needs be investigated under a variety of interfacial conditions [1][2][3] and electrochemical impedance spectroscopy have been widely deployed for those tasks. [4][5][6][7] However, electrochemical techniques based purely on electrical measurements invariably exhibit inadequate sensitivity to detect small faradaic currents from low concentrations of redox species on the electrode surface. [8][9][10] Non-faradaic currents from ions dissolved in the electrolyte solution typically create a strong background current (i.e., the capacitive electric double-layer current) that largely dominates the measured electrical signal.…”

Electrochemical DNA-sensor for evaluation of total antioxidant capacity of flavours and flavoured waters using superoxide radical damage

“…The immobilization of the enzyme can be carried out via short-chain thiol modified gold electrodes, long-chain thiol (mercaptoundecanoic acid), mixed-thiol, long-chain mixed thiol (mercaptoundecanoic acid/mercaptoundecanol) and hemin modified electrode (McNeil et al, 1995;Gobi and Mizutani, 2000;Ignatov et al, 2002). However, the performance of many of these types of devices is interfered by hydrogen peroxide, uric acid and some communication interference between the protein and the electrode (Chen et al, 2000;Beissenhirtz et al, 2004;Endo et al, 2002;Campanella et al, 2004;Emregül, 2005). The protective effect of antioxidants at a cellular level could only be achieved by monitoring the DNA integrity (Barroso et al, 2011).…”

Sensores eletroquímicos: considerações sobre mecanismos de funcionamento e aplicações no monitoramento de espécies químicas em ambientes microscópicos

“…Por esta razão, um aumento crescente no número de artigos publicados na literatura envolvendo a fabricação de microeletrodos para determinação destas purinas em tempo real tem sido evidenciado [131][132][133] . Outros compostos gerados como respostas a estímulos às célu-las também têm sido determinados com sensores eletroquímicos e, dentre eles, podem-se citar peróxido de hidrogênio 134 , lactato [135][136][137] e superóxidos 138,139 . A possibilidade de monitoramento de mais de um composto na superfície do cérebro com um dispositivo único constituído por conjunto de microeletrodos de disco separados foi discutida por Michael e colaboradores 125 .…”