Construction And Characterization Of A Nanostructured Biocatalyst Consisting Of Immobilized Enzyme On Montmorillonite And Its Application In Biodiesel

Nanostructured enzyme catalyst refers to the immobilization of enzymes on a nanostructured materials.Enzyme immobilization is an efficient route to improving the biocatalytic process economics by the reuse of enzyme and the enhancement of enzyme stability,thereby permitting the catalysis under harsh reaction conditions at an industrial scale.In this study,lipase/?-Amylase was immobilized on amphiphilic modified montmorillonite/sodium-montmorillonite support via physically adsorbed and covalently bound.Employing different supports,we acquired three new nanobiocatalytic systems,i.e.,Mt-?-Amylase hybrid nanobiocatalytic system,lipase-Mt nanobiocatalytic system and lipase@APTES-Mt microcapsules.The properties of the immobilized lipases were assayed and compared with those of the free enzyme.Based on the above relevant experimental research,the following conclusions were drawn.?1?This chapter report a rational design of Mt-?-Amylase hybrid nanobiocatalytic system based on allosteric effect and an explanation of the increase in catalytic activity when certain enzymes are immobilized in specific nanomaterials.Employing different supports,we acquired three new nanobiocatalytic systems,i.e.,?-Amylase/Ca-Mt??-Amylase/Na-Mt and?-Amylase/Ca-Mt-NaCl.The supports as well as the novel immobilized biocatalysts were characterized by a combination of techniques,namely XRD and FT-IR.Through studying enzymatic performance of these systems and free?-Amylase with/without Ca²⁺.The results show that it is feasible to improve the activity and stability of?-amylase by replacing Ca²⁺in the Ca-Mt interlayer.As a bonus,the new system we devised was found to enjoy higher stability and durability than free?-amylase.?2?In this chapter,lipase from Aspergillus oryzae was immobilized on 3-aminopropyltriethoxysilane?APTES?amphiphilic functionalized montmorillonite?Mt?support via 1-?3-Dimethylaminopropyl?-3-ethyl-carbodiimide hydrochloride?EDC?spacer.The APTES-Mt and APTES-Mt based nanostructured biocatalyst?lipase-Mt?were characterized by XRD,FT-IR,TEM,SEM and contact angle analysis.Optimal immobilization conditions were determined considering parameters of EDC concentration,initial lipase concentration,pH,and reaction time.The results indicated that the immobilization efficiency that could be achieved under the optimal conditions was 85.32%with the average loaded amount of 426.6 mg/g APTES-Mt.The specific activity of lipase-Mt was 40.65 U/mg,which is nearly 4-fold higher than that displayed by free lipase.Further,the interfacial activation by amphiphilic surface of APTES-Mt and enlarged catalytic interface contributed to the improved activity and storage stability of lipase-Mt.?3?The microstructure of the prepared lipase@APTES-Mt microcapsules was spheroidicity,the particle size distribution ranged from about 10⁷0?m,after crosslinking and drying by the crosslinking agent,the morphology and structure of the microcapsules could still be remained intactly.The lipase could be encapsulated well within the prepared microcapsules,which ensured the stability of reaction system during the process of biocatalysis experiments.The infrared spectroscopic characterization showed that the infrared structural groups peaks of microcapsules were consistent with the infrared characteristic peaks of the used raw materials.Optimal prepartion conditions were determined considering parameters of R_o/w,lipase loading,and temperature.The results showed that the activity of lipase@APTES-Mt reached the maximum when R_o/w was 20,the optimal reaction temperature was higher than that of free lipase.In addition,the thermal stability was improved,lipase@APTES-Mt retained86%of its original activity after 20 days of storage at 4?.?4?In this chapter,lipase-Mt was used to produce biodiesel from olive oil via transesterification reaction.A high biodiesel yield?91.3%?was obtained under the optimal reaction conditions and the yield of the prepared biodiesel were tested by GC.Moreover,the catalyst of lipase-Mt could be removed easily by centrifugation and recycled at least 5 times.As a new kind of support,the APTES-Mt could be a promising alternative for the carriers of immobilizing enzymes used for the green and efficient production of biodiesel.