| The free lipase is not favored in industrial developments because of its easy deactivation, low stability and difficulty to recover for reuse. However, through adopting by immobilization on the surface of inorganic support, the molecules of lipase are protected by supports, so the conformations of enzymes tend to stable. At the same time, it is easy to recover biocatalyst from reaction systems and overcome the obstacles caused by homogeneous catalyst. So the technology of immobilized enzyme becomes more and more popular. LDHs is a kind of inorganic laminated nano-materials, and this material can be synthesized easily and cheaply. Moreover, LDHs is nontoxic (or little toxic) and can provide large, open and accessible two-dimensional surfaces for enzyme adsorption. Metal elements formed LDHs sheets can be regulated, which can change the charge of the surface; interlayer anions are varied and the surface of LDHs can be modified, which can improve the polarity of LDHs. These advantages have made LDHs popular in the field of biochemistry.In this thesis, MgAl-LDHs (Layered double hydroxides) precursor was prepared together with sodium dodecyl sulfate (SDS) as an intercalated anion by the coprecipitation method, and then modification on this kind of MgAl-SDS-LDHs with y-aminopropyl triethoxysilane (KH550) was carried out. This intercalated and surface modified MgAl-LDHs can act as an enzyme immobilization support, which is used for immobilization Candida lipolytic lipase. It is well known that the catalytic activity of lipase is prefer to happen on the hydrophilicity/hydrotalcite interface, so supports with a certain hydrophobicity are beneficial to lipase immobilization owing to the interfacial activation between lipase and the support. Intercalation with SDS and modification with KH550can effectively improve the hydrophilicity/hydrotalcite property of LDHs, which will be in favor of Candida lipolytic lipase playing a catalytic role on the hydrophilicity/hydrotalcite interface. Moreover, amino group is benefical to immobilization of this lipase.In order to obtain immobilized enzyme with better catalytical effect, the procedure of lipase molecule immobilized on surface of the modified MgA1-LDHs was carried out under different conditions to determine the optimum conditions of immobilization. Under the optimum conditions, including enzyme dosage of103.0mg/g, immobilization time6.0h, immobilization pH value of7.6, immobilization temperature30℃, and the molar ratio of Mg2+to Al3+about3.0, an immobilization efficiency of69.2%and an activity recovery over56.4%were obtained based on the immobilization of Candida lipolytic lipase on the above supports.In addition, the optimum catalytic temperature of immobilized lipase increased, and the optimum catalytic pH value range of immobilized lipase with higher catalytic activity retention was wider than the free lipase, which indicates that lipase is more stable after immobilized on the supports. Moreover, the immobilized lipase exhibited higher stability against heat than its free soluble form. So, the immobilized enzyme was found to have higher temperature resistance, wider pH value and better thermostability in reactive activity. This kind of immobilized enzyme catalysis can easily recover for reuse by centrifugation and the enzyme activity can be retain. This can overcome the obstacles caused by homogeneous catalysts (the free lipase is difficulty to recover for reuse), which make it possible to apply in industry. |
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