New Nano-materials, Bio-electrochemical Study

Direct electrochemistry of redox enzymes and proteins has aroused great interest in biological and bioelectrochemical fields. Studies on direct electrochemistry of redox proteins/enzymes can be used to extract essential physicochemical data concerning the kinetics and energetics of protein redox reactions, provide mechanistic studies of electron exchange among proteins/enzymes in biological systems. Moreover, direct electron transfer between immobilized enzyme and underlying electrode can establish a foundation for fabricating new kinds of mediator-free biosensors, biofuel cells, bioreactors.In this work, the newly prepared novel nanomaterials, such as nanoparticles and one-dimensional nitrogen doping carbon nanotubes, can act as the substrates for protein immobilization and electron relays ("electrical nanoplugs"). These nanomaterials have been applied for assemblying the nano-interfaces with biomacro-molecules on the electrode surfaces. This modifying electrode surfaces on the molecular scale can allow efficient electron transfer between the electrode and redox proteins, and show high bioelectrocatalytic activity. In addition, bactericidal effect of ordered meso-TiO₂ and photoelectrochemical behavior of Au-TiO₂ composite nanoparticles have been preliminarily investigated. The main results and conclusions are summarized as follows:1. Nitrogen-doped carbon nanotubes (CN_X nanotubes) have been synthesized by catalytic method with diethylamine as carbon source and Fe-containing SAPO-5 molecular sieve as catalyst. This novel bamboo-like nitrogen-doping CN_X nanotubes are firstly used for the immobilization of hemoglobin (Hb) and their bioelectrochemical behaviors were studied.TEM displays that the nanotubes are multi-walled nanotubes with the diameter of 10-40 nm and the "bamboo" segment distance of ca. 20nm. The walls of the "bamboo" segments exhibit some disorder or defect sites. Energy-dispersive X-ray spectroscopy (EDX) analysis shows the presence of carbon, nitrogen and oxygen in the CN_X nanotubes, indicating N incorporation into the graphene lattice. The atomic ratio of C:N in the nanotubes estimated from EDX data is about 0.85:0.15. It can beclearly observed by transmission electron microscopy (TEM) and high resolution transmission electron microscopy (HRTEM) that Hb could be strongly adsorbed on the surface of CNX nanotubes. The CV results of Hb/ CNX nanotubes modified electrode shows a pair of well-defined, quasi reversible redox peaks centered at approximate -O.33V (vs.SCE) in pH7.0 phosphate buffer solution. It reflects the characteristic of Hb heme Fe(III)/ Fe(II) redox couple with fast heterogeneous electron transfer rate. The Hb incorporated into CNX nanotube films exhibits good bioelectrocatalytic activities for the reduction of hydrogen peroxide and nitrite. The results demonstrate that the CNX nanotubes may improve the protein loading with the retention of bioactivity and promote the direct electron transfer greatly. It can be attributed to the existence of a large amount of active sites on the surface of CNX nanotubes and a suitable biocompatible C-N microenvironment introduced by nitrogen doping.2. In order to further investigate the novel bamboo-like CNX nanotubes acting as the electron relay("electrical nanoplugs") for the direct electrochemistry of redox enzyme, this CNX nanotubes are used for the immobilization of a relatively large enzyme glucose oxidase (GOx) and its bioelectrochemical studies. The morphologies and adsorptions of GOx immobilization onto CNX nanotubes have been clearly observed by transmission electron microscopy (TEM) and high resolution transmission electron microscopy(HRTEM). Electrochemical impedance spectroscopy (EIS) is also used to feature the GOx adsorbed onto the surface of CNX nanotubes. The immobilized GOx incorporated into CNX nanotubes films exhibits a well-defined nearly reversible cyclic voltammetric peaks for the electroactive centers of GOx and a fast heterogeneous electron transfer rate with the rate constant (Ks) of 1.96 s'1. The immobilized GOx onto the CNX nanotubes exhibited its bioelectrocatalytic activity for the oxidation of glucose. The obtained results suggest that CNX nanotubes could not only facilitate the direct electron transfer between the enzyme and electrode, but also retain the high enzyme loading and the enzymatic bioactivity.3. The new type of nano-SnO2 composite films have been prepared by the sol-gel method, in which a natural organic polymer gelatin acted as dispersant. Horseradish peroxidase (HRP) was immobilized into a new type of sol-gel-derived nano-sized tin oxide / gelatin composite film (SnO2 composite film) using a sol-gel film /enzyme/sol-gel film "sandwich" configuration. HRP / Sn(>2 composite film diplays a pair of stable and quasi-reversible cyclic voltammetric peaks for the HRP Fe(LU) / HRP Fe(II) redox couple with a formal potential of about -0.25 V (vs. SCE ) in a pH6.0 phosphate buffer solution, showing the electron transfer between the enzyme and the underlying electrode is greatly enhanced in the microenvironment with nano-SnO2 particles and nanoporous structures. Morphologies and microstructures of the composite films and HRP/composite films are characterized with TEM, AFM. Electrochemical impedance spectroscopy (EIS) is also used to feature the HRP incorporated into composite films. FTIR and UV-Vis spectroscopy demonstrate that HRP in the composite film could retain its native secondary structure. With the advantages of organic-inorganic hybrid materials, the HRP/SnC>2 composite film modified electrode displays good stability and electrocatalytic activity to the reduction of H2O2 with the linear concentration of 0.05 l.Ommol/L and its apparent Michaelis-Menten constant( K^p) of 0.345mM, indicating a high affinity of HRP entrapped into the composite film toward H2O2.4. Considering the pore diameter of mesoporous sieve SBA-15 matches well with the size of biomacromolecules, ordered mesoporous sieve SBA-15 is used as a novel substrate matrix to immobilize the hemoglobin(Hb). FTIR and UV-Vis spectroscopy demonstrate that Hb can efficiently be incorporated into the SBA-15 and retain its native secondary structure. Immobilized Hb shows its redox peaks at -0.28Vvs. SCE , corresponding to the characteristic of Hb heme Fe(III)/Fe( II) redox couple. Achieving the direct electron transfer of Hb may be attributed to the suitable orientation of Hb molecules introduced by SBA-15 mesoporous structural effect.5. Ordered mesoporous TiCh have been synthesized using block copolymer EOa,PO7OEO2o(Pi23) as the template. The mesoporous material presents high surface area (BET surface area: 208m2/g) and high ordered structure. The photocatalytic bactericidal behaviors of this mesoporous TiC>2 are firstly studied. Experimental results show that E.coli can be killed by the mesoporous TiC<2 under UV irradiation efficiently, the cell viability is only 10 % of initial cell count after 60min illumination. The effect of different TiC<2 loading on the inactivation of E.coli is investigated, revealing the optimum effective TiC>2 loading concentration. Compared with other types of TiC>2 materials, ordered mesoporous TiC*2 shows its high photocatalytic...