Adsorption Of BSA Based On Glutamic Acid Modified Magnetic Particles

Bovine serum protein (BSA) is the most abundant protein in the bovine blood, which account for more than 50% of the serum total protein. It has many important physiological functions. As an excellent protein model, it is largely used in biological engineering, chemical, pharmaceutical, food engineering and other fields. It not only could be used as medical supplies, but also as a healthy food, spices and so on with huge market demand. Therefore, the rapid and efficient separation and purification of BSA has been one of the hot spots in the research field. The method of separating protein from magnetic material is a new separation technology, which has the advantages of easy operation, high efficiency, low cost and easy regeneration. Chitosan has a variety of functions by chemical modification due to containing more hydroxyl and amino groups as basic polysaccharide. Magnetic chitosan microspheres which made by chitosan and magnetic particles are non-toxic and biological. It could increase its adsorption capacity by connecting glutamate with magnetic chitosan microspheres. In the study, the aim was the preparation of magnetic chitosan microspheres which is modified by glutamate and its adsorption capcitablity to BSA. It could provide a technical reference and theoretical basis for separation and purification technology of BSA.This study mainly research about the effect of magnetic chitosan microspheres connected with glutamate on BSA adsorption capacity. It consists of four parts, including preparation of Fe3O4 nano-particles with chitosan by chemical co-precipitation and reduction method, optimization of magnetic chitosan microspheres preparation conditions, glutamate-modified magnetic chitosan microspheres and its separation result of the BSA in aqueous solution, the effect of glutamate magnetic chitosan microspheres on separation result of BSA in bovine serum, which providing a theoretical basis and technology roadmap for separation and purification technology of the BSA.1. Preparation of Fe3O4 nano-particles with chitosan by chemical co-precipitation and reduction method. The main study dosed influence on the particle size and magnetic properties of reduction reaction, ammonia volume, curing time, curing temperature on preparation of magnetic Fe3O4 nanoparticles influence the particle size and magnetic properties. The results showed that the magnetic Fe3O4 magnetic nanoparticles could increased and later decreased with reduction increased. The magnetic Fe3O4 magnetic nanoparticles can be slightly increased when the ammonia water volume from 20 mL to 35 mL, while the magnetic nanoparticles Fe3O4 particle size quickly widened when the volume more than 40 mL. after magnetic nanoparticles Fe3O4 particle size increases as the amount of ammonia quickly widened. the magnetic force of the magnetic nanoparticles Fe3O4 increased and then changed little with curing time, curing temperature increased. The optimal results of the single factor experiment showed that Fe3O4 nano-particles were spherical shape, the average particle diameter was 0.7?m, the content of Fe was 92.6%, and saturation magnetization was 38.63 emu/g.2. The preparation conditions of chitosan magnetic microsphere were studied using reverse suspension method with liquid paraffin as oil phase, glutaraldehyde as crosslinking agent and Span-80 as the surfactant. The results showed that the particle size of chitosan magnetic microsphere increases with the oil phase volume increase and it decreases as the stirring speed accelerated. Meanwhile, the additive amount between 10?30 mg of Chitosan has little influence on the particle size. At the same reaction time, the reaction velocity increased as the rise of temperature and product particle size also increased, but the color of product deepen is not conducive to the function. The appearance of Chitosan magnetic microspheres prepared under ideal conditions was cylindrical spherical and average particle diameter of 23.65 ?m, saturation magnetization of 18.59 emu/g.3. Chitosan glutamate Functional Magnetic Microspheres used for separating aqueous BSA. Magnetic adsorbent was constructed by using glutamic acid crosslinked by epichlorohydrin magnetic chitosan microspheres. Using the magnetic sorbent to separate BSA aqueous solution, and adsorption properties were evaluated. The results showed that the solution pH, ionic strength, the initial concentration of BSA adsorption rate on adsorption time of BSA significantly; maximum adsorption pH value was 5.0; BSA concentration, adsorption time and BSA adsorption capacity was positively correlated, and ionic strength concentrations on the contrary; the adsorption kinetics of quasi-two actions were consistent with the model and Langmuir isotherm model; about 40 min adsorption reached equilibrium and the maximum adsorption capacity was 94.1 mg/g under ideal conditions. Using NaCl solution 3g/100g as eluent, the elution rate was 97.2%, after 5 adsorption-elution cycles, elution rate is greater than 90%, no significant decline of repeated adsorption.4. Using constructed magnetic adsorbent to separate BSA bovine serum.The results of SDS-PAGE electroresis showed that the purity of BSA was about 90% through th experiment of adsorption separation from bovine serum BSA which estimated recovery was 28,85%.It proved that Magnetic adsorbent was a effective method for BSA separation.