Styrene/maleic Anhydride Copolymer Nanoparticles For The Immobilization Of Enzymes

Enzymes are widely used in the field of textiles,fermentation,food,new energy etc.Because of the vulnerability of protein,enzymes have the defects of easy to inactivate,difficult to recover,and costly,which has limited theirs applications in industry.Immobilization of enzymes on carriers can effectively enhance their stabilities,realize the resuability,and reduce the cost.Currently,various methods have been employed to immobilize enzymes,including absorption,covalent binding,encapsulation,and cross-linking.Specially,covalent binding is the most widely used approach due to the solid immobilization and well selectivity.However,it exhibited significant deficiencies,like low immobilization efficiency,complicated procedures,and inactivation.Base on these consideration,we developed a new method for highly efficient enzyme immobilization using highly reactive styrene maleic anhydride copolymer nanoparticles as carriers.The main results obtained as followed.1.Cellulase was immobilized on Styrene/maleic anhydride copolymer nanoparticles(SMN)prepared by self-stable precipitation polymerization.Firstly,Styrene/maleic anhydride copolymer nanoparticles were synthesized by self-stable precipitation polymerization in isoamyl acetate with AIBN as the initiator.Secondly,cellulase was covalently bond on the surface of SMN by the interaction of cellulase and anhydride groups at room temperature.By this strategy,immobilization yield of cellulase is 78.5%,and the binding amount reached 167 mg/g.CMC was used to determine the activity of cellulase,and there was no obviously changes in activity for immobilized cellulase with temperature ranges from 30 ? to 80 ? comparing with free cellulase.The immobilized cellulase maintained 80%of primary activity as the hydrolysis conducted at pH 8.0,which showed 66%higher than that of native cellulase.Additionally,the immobilized cellulase exhibited an excellent reusability and no loss of activity was detected after 6 cycles usages,and just 20%of primary activity lost after 10 cycles.This work provides a mild and facile strategy to immobilize cellulase by SMN,and a good reusability was achieved after immobilization,which demonstrated its value for the production of bioethanol to overcome the energy crisis.2.Papain was covalently bound to styrene/maleic anhydride copolymer nanoparticles(SMN)prepared by self-stable precipitation polymerization.The reaction time,pH and the amount of papain during the immobilization were investigated.With the optimum reaction time of 2 h,the optimum pH of 7.4,and the optimal enzyme addition amount of 8 mg/mL,the maximum immobilization yield reached 63%,and the best loading amount was 126.6 mg/g.Casein was employed as the substrate to determine the reusability of immobilized papain,which showed that 70%of primary activity maintained after 5 cycles usages.The immobilization of papain was achieved by covalently bond on SMNs,with the stability of immobilized papain improved and the cost reduced,benefiting to its industrial application.