| In this work, two kinds of self-assembled nano-structured artificial peroxidases without protein were obtained, these are Heme-histidine-imidazole ternary complex artificial peroxidase and Heme-histidine-sodium decyl sulfate complex micelles artificial peroxidase. Both artificial peroxidases were constructed successfully exploiting iron porphyrin Heme and some chemical materials, such as imidazole, histidine, and sodium decyl sulfate. Each of them was made through a self assembly process in an aqueous solution to form a homogeneous artificial peroxidase solution.In the first part of research work, the catalytic reaction rates, structural changes, enzyme kinetics parameters, suicide inactivation coefficients and metal ions or amino acids impacting on hematin-histidineimidazole ternary complex artificial peroxidase were evaluated using modern spectral technology, such as UV-visible spectroscopy and FTIR spectroscopy.The Heme, histidine and imidazole were self-assembled into nanostructured ternary complex artificial peroxidase?Atificial Peroxidase, AP? in phosphate buffer solution. The artificial peroxidase has catalytic properties of peroxidase and can catalyze better peroxidase substrate, guaiacol or 2, 2-linked N-bis?3-ethyl-benzothiazol-6 sulfonic acid? diammonium salt and hydrogen peroxide, through oxidation-reduction reaction. According to this feature which the artificial peroxidases catalytic double substrate reactants, the individual components of the artificial peroxidase, the concentrations of Heme, histidine and imidazole, external conditions, such as pH, metal ions and amino acids influencing on the artificial peroxidase were tested. The results showed that in a certain concentrations of Heme range, the artificial peroxidase apparent activity increased with increasing concentration; in a certain range of concentrations of histidine or imidazole, the artificial peroxidase apparent activity increased with increasing concentration of histidine or imidazole, and then decreased, respectively. At p H 9.0, the artificial peroxidase apparent activity reached the maximum value, indicating that the optimum pH value was pH 9.0. Metal ions Fe3+, Fe2+, Co2+ and amino acids Cys affect greatly on the artificial peroxidase apparent activity, which Fe3+ and Cys may destroy the structure of the artificial peroxidase, and Fe2+ and Co2+ affect the artificial peroxidase apparent activity by other factors.In pH 9.0 50 mmol·L-1 phosphate buffer solution, the artificial peroxidase's enzyme kinetics parameters were measured. Michaelis constant Km, catalytic rate constant Kcat and catalytic efficiency were 5.02 ?mol·L-1, 0.081 s-1, 0.0162 L·?mol-1s-1, respectively. The catalytic efficiency of artificial peroxidase was 22.3% of the natural horseradish peroxidase. In pH 7.0 10 mmol·L-1 H2O2, 50 mmol·L-1 phosphate buffer solution, the peroxide-based suicide inactivation rate constant ki of the artificial peroxidase was obtained to be 4.3813 min-1. Artificial peroxidases may be a alternative to natural enzymes and provide a theoretical basis for scientific research.In the second part of the research work, the electrochemical properties and electric catalytic properties of a micelle artificial peroxidase was tested by the electrochemical analysis methods, such as cyclic voltammetry and linear voltammetry.The Heme, histidine and anionic surfactant sodium decyl sulfate were self-assembled into micelle artificial peroxidase?MAP?. The MAP was confined with a chitosan film onto a hydroxylated fullerenes modified glassy carbon electrode. The cyclic voltammetry or linear voltammetry were used to detect properties of the modified glassy carbon electrode. Results shown that one single electron may transfer between MAP and the electrode effectively, the electron transfer rate ks was 1.96 s-1, the electroactive material surface density on the electrode surface was 9.10×10-9 mol·cm-2. The electrocatalytic ability of modified electrode was also detected, hydrogen peroxide was found with a well current response on the electrode. The linear response range was from 300 ?mol·L-1 to 1700 ?mol·L-1. The detection limit was 32 ?mol·L-1. The apparent Michaelis constant Km app of MAP on the electrode was 1.08 mmol·L-1, which may provide a theoretical basis for the research and development of artificial enzyme biosensors. |
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