Study On The Separation And Purification And The Immobilization Of The Inulinase

Inulin is a polyfructan consisting of linear β-2,1-linked polyfructose chains terminated by a glucose ( residue attached through a sucrose-type linkage) ,it includes 2～60 fructosyl units. Inulinase( 2,1-β-D fructan fructanohydrolase ) ,can hydrolyze inulin into fructoses or fructooligosaccharides.One hydrolysate fructoses possess high sweet,low caloric, pretects from decayed teeth and diabetes and so on.The other hydrolysate fructooligosaccharides has a variety of physiological funtions such as selective utilization by intestinal bacteria, raising immunity ,improving lipid metabolism , reducing blood fatty an cholesterol, promoting the absorbing of minerals, benefiting to the incorporation of the vitamins.Because oftheir excellent functional performances, they have great applied valuation and broad prospects in food industry,medical project and the developing of the energy resource. The immobilized inulinase hydrolyzing inulin to produce FOS or HFOS can work reverslly,continuely, separate the products simply. what,s more, the processing is simple which is high effects,purie products and low cost .consequently the study of the immobilizationof the inulinaseto hydrolye inulin has realistic signification .Rencently there are some restricted factors to the industry production. They are the unpurified inulinase and the low transformility rate of the immobolization. The paper study the immobilization from the purification of the enzyme,the methods of the inulinase immobilization and the hydrolyze continuely of the inulin to find a method which is low cost,simplized work process,high transformility rate and high utilized rate . On the respect of the purification ,I select SephadexG-75 chromatography and DEAE-cellulose chromatography to separate and purify the inulinase ,the purpose is to reach idel effect Which can save resource and simplify steps for reducing the pruduction cost. On the other respect of the immobilization ,the paper utilize PVA-CA immobilize by embedding method,PVA-CA combine crosslinking immobilization and D201 macroporous resin immobilized by adsorb-crosslinking method which utilize the orthogonal collocation to obtain the optimizing operation . The result shows: 1,The DEAE-cellulose chromatography separate and purify the inulinase can purify 11.2 times than crude extracts and get 55.1% activity yield, the peak which shows activity consists only one protein bent in the PAGE so that it has reached electrophoretic . Through the comparative study to the three immobilization it shows that D201 macroporous resin immobilized by adsorb-crosslinking method is better than the other two methods. The conditions of immobilization were studied , and under the optimal conditions, the enzyme activity can reach 515.916u, the enzyme activity recorery of the immobilization inulinase was 67.4%, it is higher 50% than the other report. The main products are fructoses and little gluctoses from the analyse of the inulinase hydrolyze the inulin with the the paper chromatography. In an immobilized inulinase packed-bed reactor, 4.5% inulin soulution was completely hydrolyzed at 55 ℃and space velocity of 12ml/h, the ransforming rate is 80.2% after 40h.