The Preparation Of Magnetic Composite Microspheres And Application In Protein Immobilization

Fe₃O₄ nanoparticles were synthesized by precipitation method. The surfaces of Fe₃O₄ nanoparticles were modified by oleic acid to obtain hydrophobic magnetic fluid. The properties of the hydrophybic Fe₃O₄ nanoparticles, such as structures, diameter and diameter distribution, morphologies and magnetic properties were characterized by Fourier transformation infrared (FTIR), X-ray diffraction(XRD), transmission electron microscopy (TEM) and Viberating sample magnetism (VSM) separately. The results showed that the nanoparticles with oleic acid on their surface were superparamagetism, and the diameter was between 10-15nm.Poly(methyl methacrylate)-co-poly(glycidyl methacrylate) (PMMA-co-PGMA) and polystyrene-co-poly(2-acrylanmido-2-methyl propanesulfonic acid) (PSt-co-PAMPS) magnetic composite microspheres were obtained by miniemulsion polymerization using sodium lauryl sulfate (SDS) as emulsfier and divinylbenzene (DVB) as crosslinker in the presence of magnetic fluid. The epoxy groups were modified to form amino groups on the surfaces of magnetic composite microspheres. The amount of amino groups was 431μmol/g, which was quantitatively determined by linear potentiometric titration method. The structures, morphologies, diameter and diameter distribution, magnetic properties and magnetism contents of the microspheres were characterized by FTIR, TEM, VSM and TG respectivly. The PSt-co-PAMPS and PMMA-co-PGMA were both uniform sized and the mean sizes of the microspheres were 170nm. The magnetism contents of PMMA-co-PGMA microspheres were controlled in the range of 20%-50%. Both of them were superparamagetism.PMMA-co-PGMA-NH2 microspheres could immobilizatize bovine serum albumin (BSA) by covalent adsorption after modified by glutaraldehyde (GA). However, PSt-co-PAMPS microspheres could immobilizatize BSA by physical adsorption. The immobilization behavior of microspheres was compared. The amount of immobilized BSA on PMMA-co-PGMA-CHO microspheres was more than that on PSt-co-PAMPS microspheres and PMMA-co-PGMA-CHO microspheres need less time to reach immobilization equilibrium. It means that covalent interaction had an advantage on BSA immobilization.As the PMMA-co-PGMA-CHO microspheres were preferable in respect of protein immobilization. They were chosen as carrier of the papain. The reaction condition for immobilization of enzyme was optimized and the difference of immobilized enzyme and free enzyme was investigated.