Preparation, Characterization Of CuTAPc-Fe₃O₄ Nanoparticle Composite And Application On Laccase Immobilization

Biosensitive material is an important part of fiber optic biosensor. The material's physical and chemical characters influence the whole fiber optic biosensor greatly. Organic-inorganic nanaparticle composite is a novel interdiscipline, it has good application in the future because of it's small size, excellent characters. It has a wide application prospect in medical science, biological engineering , food industries and environment monitoring . Organic-inorganic nanaparticle composites is stable in water and organic liquor and it is innocuous, which meet the require as a carrier, they are applified for immobilization of biological substances.In this thesis, a new kind of carrier copper tetraaminophthalocyanine (CuTAPc)-Fe3O4 with excellent characters was successfully prepared and was used in laccase immobilization. The required immobilized laccase was sensitive material for biosensors.The work of this paper includes mainly three aspects as followed: (1) The CuTAPc-Fe3O4 nanoparticle composite was prepared by in situ composite. The composite were characterized by IR, FEG-SEM, XRD, X-ray photoelectron spectroscopy and particle size analyzer. The nanoparticle composites took the shape of roundish spheres with their mean diameter of about 50 run. The nanoparticle composites not only have the virtues of nana materials but also have good dispersion , excellent consistent and abundant active functions on the surface for immobilization of laccase.(2 ) The study of laccase activity is an important part in the study of fiber optic biosensor based on enzyme catalyse reaction. The activity of laccase was determined using 2,2'-azino-bis-(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) and Ferrocene (Fc-H) as enzyme substrates. When ABTS or Fc-H was used as the enzyme substrate, the best pH for the laccase activity and the best temperature for the laccase were studied, respectively. In order to determined compatibilities of different substrates, the Km of the laccase for these two kinds of substrates were determined, the Km of ABTS smaller than the Km of Fc-H. Form those results, we chose ABTS as the suitable substrate of laccase.(3) The laccase was immobilized on the surface of the composite by crosslinking with glutaraldehyde. The whole immobilize process and the effects of glutaraldehyde quantities , glutaraldehyde concentration, activating time and the amount of laccaseon the activity were studied. After immobilization, the best pH, temperature and Km of immobilized laccase catalyse ABTS were studied . Compared with free laccase , the immobilized laccase has better stabilities in storage and operation . Immobilized laccase has better characters and can be applied in more areas than free laccase. This work provided good basis for developing the fiber optic biosensor with excellent properties.