Preparation Of Layer-by-layer Assembly Polyelectrolyte Films And Hybrid Microcapsules For Enzyme Immobilization

The layer-by-layer (LbL) assembly technique is a simple method with mildoperating conditions. LbL films possess features of a certain mechanical strength andthermal stability, and have shown potential applications in the fields of separationmembranes, catalysis and controlled drug release. To adopt a simpler and moreeffective method for enzyme immobilization and provide for appropriate physical andchemical micro-environment, LbL method was used to prepare functional filmssupport. Inspired by the structure of diatom cell, the organic-inorganic compositemicrocapsule support was fabricated by a synergy of LbL and biomimeticmineralization methods, thus using for-glucosidase and catalase immobilization.The main details in this study were summarized as follows:Firstly, PDDA and G were self-assembled on SPES membrane to fabricate theSPES-PDDA-G-PDDA enzyme membrane. Support preparation and enzymeimmobilization were realized simultaneously in this process. The preparationconditions of enzyme membranes were optimized, and effect of enzyme locations onleakage ratio of enzyme, enzymatic activity and stability was also examined. Theresults indicated that compared to free enzyme, the stabilities of immobilized enzymehave been significantly enhanced, and enzyme located in the inner sandwiched layerexhibited a lower enzyme leakage ratio and higher stabilities. At70℃, free G onlyremained5%relative activity while immobilized G still kept above60%relativeactivity. After storing for40days, immobilized G still retained65%of its initialactivity, while free G lost about70%of its initial activity.Secondly, dopamine-modified alginate was chemically synthesized by theEDC/NHS method, and then the functional (PEI/DA-Alg)nmultilayers film wasformed on PC membrane by LbL method. Catalase was then immobilized by thecovalent binding of catechol groups and enzyme molecules. There was not only theelectrostatic force between the multilayers, but also the adhesive attraction induced byDA-Alg, thereby improving the stability of multilayers. The assembly of PEI andDA-Alg on the quartz substrate was investigated under different assembly conditions.On that basis, CAT immobilized on the (PEI/DA-Alg)nmultilayers film was studied interms of enzyme activity and stability. It was found that the immobilized CAT showedimproved stabilities. The covalently immobilized CAT retained above50%of its initial activity after7batches. After storing for39days, immobilized CAT still kept69%of its initial activity, while free CAT nearly lost95%of its initial activity.Thirdly, inspired by the structure of diatom cell, DA-Alg and PEI wereself-assembled on CaCO3by a synergy of LbL and biomimetic mineralizationmethods to fabricate (PEI/DA-Alg)n/PEI/Silica hybrid microcapsule. CAT was thenencapsulated in microcapsules. On one hand, the bifunctional polymer of PEI exertedits shape-forming ability to fabricate microcapsule. On the other hand, PEI exerted itsbiomimetic mineralization inducing ability to form a dense silica inorganic shell. Themechanical stability of microcapsule, enzyme activity and stability of immobilizedCAT were systematically studied. Results of PSS osmotic experiment indicated thatthe mechanical stability of hybrid microcapsule was higher than that of organicmicrocapsule. Compared to free CAT, incapsulated CAT showed enhanced stabilities.CAT incapsulated in hybrid microcapsule retained above65%of its initial activityafter7batches, while the enzymatic activity of CAT incapsulated in organicmicrocapsule decreased to30%or less. After storing for45days, CAT incapsulated inhybrid microcapsule retained above78%of its initial activity, while CAT incapsulatedin organic microcapsule only remained21%of its initial activity.