Preparation And Specific Adsorption Of Protein Imprinted Crystalline Calcium Alginate Hydrogel Microspheres

With the development of science and technology,studies on protein become the core work of post-genomic era gradually.However,traditional technologies of protein separation have been difficult to meet the development of modern society.Protein imprinting polymers have imprinted cavities and sites which match with template molecules in size,shape and conformation.They can be used as new materials for protein adsorption and separation because of the specific recognization.In light of the properties of abundant resources,low prices and good biocompatibility,alginate hydrogel has been applied to prepare protein imprinted materials and been studied exhaustively.But it is easy to swell in rebinding process,which will reduce the rebinding capabilities and specific recognition properties.In this work,with the aim of controlling the swelling ratios of alginate hydrogel,polyurethane grafted alginate hydrogel beads(PU-g-CaA)were prepared by radical polymerization.The product was used to prepare protein imprinted hydrogel material with bovine serum albumin(BSA)as template.Firstly,the modified products were characterized by FTIR,13C NMR,optical microscopy,scanning electron microscopy(SEM),thermogravimetry(TG),differential scanning calorimetry(DSC),X-ray diffraction(XRD).The swelling ratios of the hydrogel beads with and without modification were compared.The results showed that the grafting polymerization occurred between PU and alginate.The surface morphologies of materials were changed after modification.The thermal stability was improved and the swelling ratio was decreased.Secondly,the optimum preparation strategy of imprinted microspheres was obtained by investigating the influences of some factors on the adsorption capacity and imprinting efficiency,such as pre-assembling pH,protein concentration,eluant component and pH as well as the grafting ratio of modified product.Researches indicated that the modified product exhibited the optimum rebinding and recognizing properties when the imprinted substrate was with grafting ratio of 20%,pH value was controlled at 4.1,protein concentration was 2 mg/mL,Tris-HCl buffer solution with pH = 7.5 and Ca2+(0.09 mol/L)was chosen to elute template molecules.The elution process was lasted 750 min.At the same time,the impacts of environmental conditions on the imprinting effect were investigated.Results demonstrated that the best imprinting effect can be achieved when the adsorption temperature was 20?,the BSA solution contained 0.09 mol/L Ca2 + and the pH value was 4.6.Kinetic studies showed that the adsorption rates of imprinted microspheres decreased with increase of adsorption time and reached equilibrium in 120 min.Thermodynamics studies showed that the adsorption capacities of MIPs could not increase indefinitely with the increasing concentration of protein solution.Thermodynamic equilibrium was reached when the protein concentration was 4 mg/mL.Finally,the specific adsorption ability for different proteins and reusability of MIPs were investigated.The differences between PU-g-CaA and CaA MIPs were compared.The results indicated that the selectivity of modified MIPs had been improved.Besides,the selectivity can be maintained well after repeated elution and adsorption process.