Improvement Of Immobilized Glucose Isomerase Performance And Study On New Immobilized Technology

High Fructose Syrup(HFCS),the world's second most consumed sugar,is widely used in food,beverage,medicine and other industries due to its characteristics of crisp taste,low price,High solubility and good moisture retention.Glucose Isomerase(GI)for the production of HFCS is the world's largest and most commercially successful immobilized enzyme,with annual consumption of more than 500 tons worldwide.In industrial production of HFCS,the step of glucose isomerization into fructose by GI is considered to be the key step in HFCS production.The production of HFCS is a long and continuous process,with the use cycle of the immobilized GI lasting about 10months after each column installation.At present,the HFCS production enterprises in the process of using immobilized GI mainly by the activity of enzymes suppliers give parameters as standard,the production process more extensive use in such a way that loss of activity of immobilized GI faster,most of the time,give less than enzyme preparation suppliers give waste enzyme activity of enzyme activity efficiency of the ideal value.The loss of enzyme activity can be slowed down by strictly controlling the factors affecting enzyme activity and regularly maintaining the use condition of GI during the use of immobilized GI.Therefore,the study and optimization of various physical and chemical factors affecting the immobilized GI reaction system in the production of HFCS can slow down the activity loss of immobilized GI,thus improving the use efficiency of the enzyme and prolonging the use cycle.At present,most of the GI used in HFCS production in China depends on imports.The development of commercialized domestic immobilized GI can gradually replace imported enzymes,which can reduce the production cost of domestic HFCS enterprises and enhance the technical competitiveness.Nano-metal-organic framework structures(MOFs),as a new kind of materials,not only exhibit the rigidity of metals but also retain some properties of organic ligands.It has shown good application value in hydrogen storage,gas adsorption and separation,drug release,catalytic reaction and other fields.MOFs can not only adsorb and fix enzymes with their own porous structure,but also coordinate with amino and carboxyl groups at the internal unsaturated metal sites,and firmly fix free enzymes in the form of covalent bonds.MOFs will be widely used in the field of immobilization in the future.In this study,a single factor experiment was used to select the range of factors in affecting the activity of immobilized GI,such as activation time,activation temperature,substrate concentration,bed height,reaction temperature,p H and ion concentration of two commonly used immobilized GI activities.Then,taking immobilized glucose isomerase A as an example,the reaction system conditions of glucose conversion process were optimized by response surface methodology.The optimal conditions were determined after response surface optimization:activation time:3.5h;Reaction temperature:70?;PH value:7.5;Bed height:96%;Substrate concentration:48%;SO₃^2-Concentration:21 m M;Mg²⁺concentration:28 m M;The maximum concentration of fructose at the exit of the simulated heterogeneous column was 601.2 mg/m L when the flow rate of feed and liquid was 1.42 m L/min per column volume/3 h.Then the experiment of GI pressure regulation in simulated heterogeneous column was carried out.In the simulation experiment,the feed liquid backwashing was used to adjust the pressure of GI in the isomeric column.The results showed that the activity of GI decreased after a long time of use.The backwashing was used to adjust the pressure of isomeric column,and the activity of the enzyme was significantly improved.The results showed that the activity of glucose isomerase could be improved by regulating the pressure of isomerase column at regular intervals during the use of glucose isomerase.According to the experimental results and with reference to GB/T 23533-2009 put forward the GI quality criteria and the matters that should be paid attention to when using,in order to facilitate enterprises to make appropriate production parameters adjustment for different batches of different quality GI,to avoid the unnecessary loss caused by GI quality problems.GI crude enzyme was extracted from Serratia marcescens and its activity was determined to be 58.7U.The The glucose isomerase immobilized with MOF-525 was synthesized and characterized by TEM,IR and UV-Vis absorption spectra.The results showed that the immobilized carrier MOF-525 was correct.Then MOF-525 was used as the immobilized carrier to immobilize the extracted GI by adsorption and covalent coordination.The loading rate of the enzyme was 37.5%,and the performance of the immobilized GI was studied.The results showed that MOF-525 immobilized GI was more stable under acidic conditions and the best activity was found at p H 7.0.After MOF-525 immobilization,the activity of GI decreased by 13%after six reuses,showing good reusability.It lays a foundation for future research on immobilized GI which has application potential.