Studies On Immobilized L-asparaginase Based On Polyacrylamide/N,N'-methylenebisacrylamide

Studies on immobilized enzyme based on synthesized polymers are being widely carried out nowadays in the world, which has not yet been deeply studied in China. In this paper, L-asparaginase immobilized in poiy (acrylamide/N, N'-methylenebisacrylamide) copolymer gels was synthesized and its properties were studied. The kinetic behavior of the immobilized L-asparaginase was studied. Mathematical models of catalytic process of immobilized L-asparaginase for batch reaction and continuous reaction systems were established and simulations were carried out.In study on polymer carrier for enzyme immobilization, poly (acrylamide/N, N'-methylenebisacrylamide) copolymer gel was prepared in an aqueous buffer medium by a free-radical solution polymerization of acrylamide I N, N '-methylenebisacrylamide / ammonium persulfate / N, N, N', N'-tetramethylethylenediamine I polyethylene glycol. The result shows that the diluent (PEG) is useful for control of polymerization rate and for improvement of the tenacity of the gel, which make poly (acrylamide/N, N '-methylenebisacrylamide) copolymer gel more suitable for enzyme immobilization.The effects of the concentration of monomer and cross-linker on the swelling ability and rigidity of the gel were investigated. Optimization of the gel for enzyme entrapment was given. The results show that for fixed cross-linker concentration the swelling ratio of the gel decreases with the increase of monomer concentration, and that for fixed monomer concentration the swelling ratio decreases with the increase of cross-linker concentration. The result also shows that for fixed cross-linker concentration the rigidity of the gel increases with the increase of monomer concentration, and that at low monomer concentration the rigidity of the gel increases with the increase of cross-linker concentration, while at high monomer concentration the effect of cross-linker concentration on the rigidity of the gel is not significant.In the study on immobilized L-asparaginase, the enzyme loading, reaction temperature, pH, operation and storage stability were investigated. The results show that with the increase of enzyme loading amount the activity per gram immobilized enzyme increases, while the activity per enzyme loading decreases. Macro-kineticsi ^jIü ± ^'Vconstant K m(⁾ decreases and Vm^j increases with the increase of enzyme loading amount. The optimal medium pH for immobilized L-asparaginase catalysis reaction is 8.O(O.OIMI'ris-HCl). High activity of immobilized L-asparaginase was observed after six times use or 42 days' storage.In study on the kinetic behavior of the immobilized L-asparaginase, the effects of reaction temperature, enzyme loading amount and diffusion resistance on the catalytic reaction rate was investigated. The results show that the catalytic reaction rate increases with the increase of temperature or enzyme loading amount and that the catalytic reaction rate was smaller than theoretical prediction at the beginning stage of the reaction due to diffusion resistance.Mathematical models of catalytic process of immobilized L-asparaginase for batch reaction and continuous reaction systems were established and the model prediction is in good agreement with experimental data. The effects of enzyme loading, effective diffl.tsion coefficient, particle size and rate of flow of substrate feed on the substrate concentration profiles within gel particle and on the catalytic reaction rate were simulated. The simulation results show that the effects of enzyme loading, effective diffusion coefficient, particle size and rate of flow of substrate feed on the substrate concentration profile within the gel particle and on the catalytic reaction rate are significant.