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Cells in the metabolic processes continue to produce a variety of reactive oxygen species (reactive oxygen species, ROS) such as superoxide anion radical (O2-), hydroxyl radical (· OH), lipid radicals (ROO-), peroxide of hydrogen ( H202 H202), wherein the most stable one ROS. Study found that the hydrogen peroxide in the process of cell metabolism plays an important role in regulating it and intracellular biological macromolecules response, causing lipid peroxidation, intracellular proteins and enzymes denatured DNA damage, eventually leading to cell death , tissue damage, cardiovascular disease, cancer and neuron degeneration and other diseases. Therefore the establishment of a simple and rapid method for the detection of H202 concentration on the prevention, diagnosis and monitoring of clinical disease is of great significance. This thesis Nace hydroxyapatite, horseradish peroxidase - the nanometer hydroxyapatite (HRP-HAP) Composites and for Determination of H202 cells. The main contents are as follows: Preparation and characterization of nano hydroxyapatite (HAP). Ca (OH) 2 and H3PO4 raw materials, using a coprecipitation method of preparing a nano hydroxyapatite; using X-ray diffraction (XRD), transmission electron microscopy (TEM), scanning electron microscope (SEM), infrared spectroscopy (FTIR) its characterization. XRD results showed that the prepared HAP hexagonal; TEM showed that the prepared HAP needle structure, length of 160-180 nm, a width of about 20-25 nm, and the grain is well dispersed, no serious agglomeration; SEM showed that the HAP having a good three-dimensional porous structure. Horseradish peroxidase (HRP) in the surface of the nano-hydroxyapatite (HAP) fixed, direct electrochemical, and H202 restore power catalysis. HRP molecular modification to the HAP surface preparation HRP-HAP composite infrared spectroscopy (FTIR), ultraviolet spectroscopy (UV-vis) and electrochemical methods (CV) to characterize the results show that the HRP adsorbed on HAP surface. HRP-HAP modified GC electrode surface prepared electrode HRP-HAP/GC, voltammetry of the HRP-HAP/GC electrode electrochemical properties. HRP fixed after the HAP surface can be effectively direct electron transfer, the cyclic voltammetry curves exhibited a pair of good, nearly symmetrical redox peaks, oxidation, reduction peak potential of -338 mV and - 401mV (vs. SCE), the formal potential E0 'of -370 mV, and almost does not change with the change of the scan rate, the apparent electron transfer rate constant (ks) of 3.77 ± 0.78 s-HAP nanomaterials can provide a good the micro-environment in order to accelerate electron transfer of HRP. Voltammetry and chronoamperometry study the electrode HRP-HAP/GC H202 Reduction Electrocatalysis. The results showed that, in the potential for the -400 mV when the electrode HRP-HAP/GC H202 having the best response, response time of about 2s, catalytic current with H2O2 concentration was linear in the range of 0.82 mmol / L in 5 micromol / L to correlation (correlation coefficient of 0.998), the lowest detection limit of 0.1 micromol / L. In addition, the electrode can also be used for the detection of hydrogen peroxide in the actual sample. Horseradish peroxidase - nano-hydroxyapatite (HRP-HAP) modified electrode used for the detection of RAW264.7 murine macrophage cells H202. By voltammetry the electrode HRP-HAP/GC RAW264.7 cells H202 Determination intracellular H202 concentration of about 8 nmol / L, according to the response current relationship between the concentration curve drawn. The same time, the electrode of the N-formyl-L-methionyl-L-bright aminoacyl-L-phenylalanine tripeptide (fMLP) stimulation of RAW264.7 cells H202 release, results show that the after fMLP (0.3μmol / L), with time, H202 cells release is gradually increased, reached a maximum at 30 min (approximately 17nmol / L), after which the concentration gradually decreases. Show that the method can be used for H202 release from intracellular dynamic processes.
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