Based On Inorganic / Bio-organic Nano-hybrid Membrane Electrode Sensor, Inhibition Kinetics And Its Application

1. Based on nano-calcium carbonate to build a type of polyphenol oxidase inhibition biosensor research and applicationIn this present work, a simple highly reversible and sensitive amperometric biosensor, based on the immobilization of tyrosinase (Tyro) by calcium carbonate nano-materials (nano-CaCO3), was applied for determination of food preservative, benzoic acid. The detection of benzoic acid was performed via its inhibiting action on the Tyro/nano-CaCO3 modified glassy carbon electrode. The effects of enzyme substrate type and substrate concentration on the inhibitory have been investigated in detail. A potential value of–0.20V versus SCE, and a constant catechol concentration of 6μM were selective to carry out the amperometric inhibition measurement. The inhibitor biosensor had a fast response to benzoic acid (<5s) with a wide linear range of 5.6×10-7 to 9.2×10-5 M and a high sensitivity of 1061.4 mA M-1 cm-2. The inhibiting action of benzoic acid on the Tyro/nano-CaCO3 electrode was highly reversible (100%) and of the typical competitive type, with an apparent inhibition constant of 17μM. This inhibitor biosensor was successfully applied for the determination of benzoic acid in some real beverage sample, such as Coca-Cola, Pepsi-Cola, sprite and Yoghurt. Results were compared to those obtained using high performance liquid chromatography, showing a good agreement.2. Based on self-assembled monolayer laponite/CHT in horseradish peroxidase of the direct electrochemical and applicationA novel method for fabricating hydrogen peroxide (H2O2) sensor has been presented based on the self-assembly of laponite nanoparticles with heme proteins on functional glassy carbon electrode. Cyclic voltammetry of HRP/laponite/CHT/GCE showed a pair of stable and quasi-reversible peaks for HbFe(III)/Fe(II) redox couple at about -0.09V versus saturated calomel electrode (SCE) in pH 7.0 phosphate buffer, at the scan rate of 0.1V s-1. The electrochemical reaction of HRP entrapped on the laponite/chitosan self-assembled film exhibited a surface-controlled electrode process. The formal potential of Hb heme Fe(III)/Fe(II) couple varied linearly with the increase of pH over the range of 4.0-9.0 with a slope of -60 mV pH-1, which implied that an electron transfer was accompanied with single proton transfer in the electrochemical reaction. As a result, a kind of novel thirdgeneration biosensors for H2O2 was developed. The resultant HRP electrode exhibited fast amperometric response to H2O2, excellent stability, long-term life and good reproducibility.3. Based on chitosan / laponite composite materials to build a glucose oxidase electrochemical sensorNanocomposite matrix based on chitosan/laponite was successfully utilized to construct a new type of amperometric glucose biosensor. This hybrid material combined the merits of organic biopolymer, chitosan, and synthesized inorganic clay, laponite. Glucose oxidase (GOD) immobilized in the material maintained its activity well as the usage of glutaraldehyde was avoided. The composite films were characterized by Fourier transform infrared (FT-IR) and atomic force microscopy (AFM). The parameters affecting the fabrication and experimental conditions of biosensors were optimized. The sensitivity of the proposed biosensor (33.9 mA M-1 cm-2) permitted the determination of glucose in the concentration range of 1×10-6 - 5×10-5 M with a detection limit of 0.3μM based on S/N=3. The apparent Michaelis-Menten constant (KMapp) for the sensor was found to be 15.8 mM.