Preparation Of Magnetized Immobilized Cellulase And Its Properties

Cellulase has been widely utilized in food, paper making, textile, energy production etc.However, the industrial appliction of cellulase is severely limited by its disadvantages of easyinactivition in reaction and difficult recovery, which results in high cost in production.Enzmye immobilization technology could not only overcome these shortcomings, but alsorealize enzyme industrialization.In this study, the main object was to immobilize cellulase on magnetic carrier. Thepreparation process and properties of magnetic chitosan microspheres carrier were bothinvestigated. Cellulase crosslinked with magnetic microspheres was researched andcombination of absorb/precipitation/aggregate and cross-linking technique to immobilizeCLEAs onto magnetic microspheres was also investigated. Characteristics of twoimmobilized enzyme were studied, the main results are listed as follows：1. Magnetic chitosan microspheres were synthesized by the method of reverse phasesuspension cross-linking technique with glutaraldehyde as cross-linker. The characteristics ofmorphology, particle distribution and magnetic responsiveness were analyzed by scanningelectron microscope, laser particle size analyzer and physical properites measurement system,respectively. The results showed the prepared magnetic microspheres were well-shapedsphere with average diameter of2-10μm in which the Fe3O4was dispersed uniformly; thevalue of saturation magnetization was16.15emu/g which owned good magnetic response.2. In the absorption-linkage method,50mg/mL cellulase solution was absorded on0.25gmagnetic carrier for2h, and then2%glutaraldehyde was added for cross-linkage for3h at20℃. The activity recovery and catalyse activity were35%and327.2U, respectively.3. CLEAs immobilized on magnetic carrier were prepared with ammonium sulfate asprecipitant and glutaraldehyde as cross-linker. The optimum factors to prepare CLEAs ontomagnetic carrier were as followed: the amount of12.5mg/mL cellulase was absorbed on0.2gmagnetic carrier for2h, then95%ammonium sulfate was added to deposit free enzyme for30min,3%glutaraldehyde was used to cross-link the enzyme for7h at20○C. The optimalactivity recovery and catalyse activity were50%and412.5U, respectively. 4. The optimum reaction temperature and pH of the CLEAs with magnetic carrier were60℃and4.0, while the conditions for the cellulase cross-linked onto magnetic carrier were60℃and5.0, respectively. The tolerance of the two immobilized enzymes to hightemperature and pH around5-7were both improved. The thermal and pH stability of CLEAswere superior to cellulase cross-linked onto magnetic carrier.5. Two immobilized enzymes had above50%relatively activity recovery retained afterstorage for28d, the stability was increased compared with free cellulase which was20%activity left. CLEAs with magnetic carrier kept30%activity after9times usage andmaintained the catalyzation ability after5times usage. The cellulase cross-linked ontomagnetic carrier had32%activity recovery after9times use.