Impedance spectroscopy and conductometric biosensing for probing catalase reaction with cyanide as ligand and inhibitor

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• 作者：Naima Bouyahia ; Mohamed Larbi Hamlaoui ; Mouna Hnaien ; Florence Lagarde ; Nicole Jaffrezic-Renault
• 关键词：Cyanide ; Catalase ; Inhibition ; Conductometric biosensor ; Impedimetric biosensor ; I₅₀
• 刊名：Bioelectrochemistry
• 出版年：2011
• 出版时间：February 2011
• 年：2011
• 卷：80
• 期：2
• 页码：155-161
• 全文大小：680 K

文摘

In this work, a new biosensor was prepared through immobilization of bovine liver catalase in a photoreticulated poly (vinyl alcohol) membrane at the surface of a conductometric transducer. This biosensor was used to study the kinetics of catalase–H₂0₂ reaction and its inhibition by cyanide. Immobilized catalase exhibited a Michaelis–Menten behaviour at low H₂0₂ concentrations (< 100 mM) with apparent constant K_M^app = 84 ± 3 mM and maximal initial velocity V_M^app = 13.4 μS min^− 1. Inhibition by cyanide was found to be non-competitive and inhibition binding constant K_i was 13.9 ± 0.3 μM. The decrease of the biosensor response by increasing cyanide concentration was linear up to 50 μM, with a cyanide detection limit of 6 μM. In parallel, electrochemical characteristics of the catalase/PVA biomembrane and its interaction with cyanide were studied by cyclic voltammetry and impedance spectroscopy. Addition of the biomembrane onto the gold electrodes induced a significant increase of the interfacial polarization resistance R_P. On the contrary, cyanide binding resulted in a decrease of Rp proportional to KCN concentration in the 4 to 50 μM range. Inhibition coefficient I₅₀ calculated by this powerful label-free and substrate-free technique (24.3 μM) was in good agreement with that determined from the substrate-dependent conductometric biosensor (24.9 μM).