CN104560936A - Compound immobilization method for fructosyl transferase - Google Patents
Compound immobilization method for fructosyl transferase Download PDFInfo
- Publication number
- CN104560936A CN104560936A CN201510000076.9A CN201510000076A CN104560936A CN 104560936 A CN104560936 A CN 104560936A CN 201510000076 A CN201510000076 A CN 201510000076A CN 104560936 A CN104560936 A CN 104560936A
- Authority
- CN
- China
- Prior art keywords
- fructosyl transferase
- enzyme
- immobilization
- thalline
- liquid
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Landscapes
- Immobilizing And Processing Of Enzymes And Microorganisms (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
The invention discloses a compound immobilization method for fructosyl transferase, and belongs to the field of biotechnologies. The technology has the advantages that fructosyl transferase extraction technological equipment is simple; the fructosyl transferase is embedded by calcium alginate after chitosan covalence immobilization; the problems of forming and integer of chitosan covalence immobilized enzyme can be solved; the immobilization efficiency of the enzyme is high (enzyme activity recovery rate reaches above 70%); the enzyme can be used repeatedly for more than 85 times during batch reaction.
Description
Technical field
A compound process for fixation for fructosyl transferase, belongs to biological technical field.
Background technology
Oligofructose refers to that fructosyl in sucrose is by β-1,2 glycosidic links are combined formed kestose, the general name of GF3 and GF4 and composition thereof with 1 ~ 3 D-Fructose, also known as Nutriflora P, FOS etc., molecular formula is G-F-Fn(G is glucose, F is fructose, n=1 ~ 3).Oligofructose is a kind of prebiotics, and it has and improves intestinal microflora, suppresses enteron aisle corrupt, relaxes bowel, promote the absorption of mineral substance, improve lipid metabolism, the physiological functions such as strengthening immunity.At present, the many employings of lot of domestic and international enterprise are that the enzyme transforming process of raw material is to produce oligofructose with sucrose.
It is be substrate with sucrose that enzyme transforming process produces the principle of oligofructose, utilizes the katalysis of fructosyl transferase, carries out intermolecular fructosyl shift reaction and obtain oligofructose.Enzyme transforming process produces oligofructose can be divided into again intermittent production and consecutive production two kinds of modes.Intermittent production has another name called submerged fermentation production, refers to and is directly mixed with 50% ~ 60% sucrose solution by fructosyl transferase, and under certain temperature and pH, oligofructose is produced in reaction.Its advantage is that processing unit is simple, but enzyme cannot realize recycling, and oligofructose output is also lower, and production cost is higher.Therefore adopt the mode of consecutive production both at home and abroad, i.e. immobilized enzyme method more.
The extraction of industrial fructosyl transferase adopts machinery broken wall law more, and this method equipment used is expensive.Comparatively speaking, natural cracking process cost of the present invention is lower.Nature cracking process refers to be cultivated bacterial strain in suitable substratum, and treat later stage metabolic waste accumulation, living environment worsens, but thalline is not yet dead, time still uncracked, filter, washs thalline with sterilized water, is then transferred to by thalline in sterilized water and continues to cultivate.Prolongation in time, thalline can dead cracking, and content is released, but fructosyl transferase is not yet hydrolyzed or only has a small amount of hydrolysis, now filters, and obtains enzyme liquid.
Chinese patent 200910026901.7 discloses a kind of with the method for large pore anion resin immobilized fructosyltransferase, and it comprises the preparation of free fructosyl transferase and immobilization two portions of fructosyl transferase.This invention for carrier with large pore anion resin D380, D296R or D301-III, with glutaraldehyde as cross linker, adopts and first adsorbs being fixed of method be cross-linked afterwards.Although this method enzyme is lived, the rate of recovery can reach 81.2%, recycles number of times only 10 times, recycles number of times less.In this method, a part of enzyme electrostatic adhesion is on resin, and a part of covalent cross-linking is on resin.In immobilized enzyme and sucrose reaction process, the enzyme be adsorbed on resin by electrostatic interaction is likely replaced by other electronegative ions, thus departs from resin, causes loss, causes immobilized enzyme to decline.
To be that compound immobilization is namely first crosslinked embed again for the immobilization technology of present method.Take chitosan as carrier, the immobilized enzyme that glutaraldehyde as cross linker obtains is fragmental, though be conducive to enzymatic reaction, not easy to be recycled, there is shaping and problem that is shaping.The basis of this immobilized enzyme then can solve shaping and problem that is integer preferably through calcium alginate gel embedding again.Through the immobilized enzyme enzyme rate of recovery alive high (more than 70%) that this method obtains, during batch reactions, can Reusability more than 85 times.
summary of the invention:
The present invention is directed to the deficiencies in the prior art, provide a kind of compound process for fixation of fructosyl transferase.The present invention adopts nature cracking process, by simple processing unit, is effectively extracted fructosyl transferase; Efficiently solve the shaping of chitosan covalent immobilization enzyme and integer problem; Overcome the shortcoming of liquid submerged fermentation, achieve recycling of enzyme; Obtain enzyme to live high, the Reusability immobilized fructosyltransferase often of the rate of recovery.
Technical solution of the present invention adopts following steps:
A compound process for fixation for fructosyl transferase, comprises the steps:
(1) bacterium producing multi enzyme preparation is inoculated in fermention medium, temperature 28-32 DEG C, air flow 500-1500ml/min, stirring velocity 80-150r/min condition under, cultivate 4-6 days, obtained fermented liquid;
Above-mentioned fermention medium component is as follows, is all weight percentage:
Sucrose 5%-15%, dregs of beans 5%-10%, excess water, initial pH6.5-7.5;
(2) by fermentation liquor thalline broken wall obtained for step (1), filtration, obtained fructosyl transferase crude enzyme liquid;
(3) fructosyl transferase crude enzyme liquid is carried out vacuum-concentrcted, concentrate the enzyme activity of rear feed liquid with high effective liquid chromatography for measuring, control the enzyme work of concentrated solution at more than 100U/ml;
(4) concentrated rear fructosyl transferase enzyme liquid is carried out compound immobilization process with chitosan, sodium alginate and calcium chloride respectively, finally obtained immobilized fructosyltransferase.
Preferred according to the present invention, the bacterium producing multi enzyme preparation in described step (1) is aspergillus niger or aspergillus oryzae.
Preferred according to the present invention, the method adopted of breaking of the thalline in described step (2) is nature cracking process.
Preferred according to the present invention, in described step (2), condition is as follows: carried out filtering and collecting thalline by the thalline after step (1) fermentation culture with filter cloth, thalline is washed 3-5 time with sterilized water, after continuing to cultivate thalline 3-5d in the sterilized water of access and fermented liquid same volume, cross and filter broken thalline, obtain fructosyl transferase enzyme liquid.
Preferred according to the present invention, in described step (4), compound immobilization comprises covalent immobilization and embedded immobilization two step.
Preferred according to the present invention, the reaction conditions of covalent immobilization is: select chitosan to be covalent immobilization carrier, after fructosyl transferase is mixed in the ratio of enzyme 100-200U/g carrier alive with chitin carrier, in 0.5%-1%(w/w) ratio add linking agent glutaraldehyde, adjust ph 6.5-7.5, covalent immobilization temperature 30-40 DEG C, stirs 5-10h with the speed of 100-160r/min.
Preferred according to the present invention, the reaction conditions of embedded immobilization is: after fully being washed by the fructosyl transferase of covalent immobilization, centrifuge dripping.Fully mixed under 10-15 DEG C of condition with the fructosyl transferase through covalent immobilization by sodium alginate soln, blending ratio is the fructosyl transferase adding 1-5g covalent immobilization in every 100ml sodium alginate soln.Feed liquid after mixing is slowly instilled CaCl
2in solution, form dripping pill.Finally the dripping pill of formation is put into the CaCl of same concentration
2in solution after hardening treatment 10-20h, namely obtain the immobilized fructosyl transferase of compound.
Preferred according to the present invention, described sodium alginate soln concentration is 2%-4%(w/w).
Preferred according to the present invention, described CaCl
2strength of solution is 3%-5%(w/w).
beneficial effect
1, the extraction of current industrial fructosyl transferase adopts machinery broken wall law more, and this method equipment used is expensive, technical sophistication, cost are very high.And in the present invention fructosyl transferase extraction adopt be nature cracking process.Nature cracking process refers to the thalline phase after incubation, during not yet dead, cracking, filters and wash thalline, and is transferred to by thalline in sterilized water and continues to cultivate until thalline natural death cracking, the process that content and enzyme liquid discharge voluntarily.This kind cracking process method is novel, use equipment is simple, cost is very low, and pole is suitable for the utilization of industrialization scale operation.
2, through the fructosyl transferase that resin absorption is fixing, in batchwise reaction procedure, enzyme molecule easily comes off, and the number of times that utilizes of this immobilized enzyme is reduced; Through the fructosyl transferase that chitosan covalency is fixing, how in fragmental, this immobilization enzyme-to-substrate can fully contact and react, but not easy to be recycled, needs integer just to can be used for continuous seepage oligofructose.And fructosyl transferase is first fixing again with calcium alginate embedded through chitosan covalency in the present invention, enzyme molecule difficult drop-off and efficiently solve the shaping of chitosan covalent immobilization enzyme and integer problem, is more conducive to suitability for industrialized production oligofructose.
3, utilize the technology of the present invention to make enzyme immobilizatio efficiency high (the enzyme rate of recovery alive reaches more than 70%), during batch reactions, this immobilized enzyme can Reusability more than 85 times.
Embodiment
In order to better set forth technical scheme of the present invention, below by embodiment, the present invention will be further described, but institute of the present invention protection domain is not limited thereto.
example one
A compound process for fixation for fructosyl transferase, comprises the steps:
(1) bacterium producing multi enzyme preparation-aspergillus niger is inoculated in fermention medium, temperature 28-32 DEG C, air flow 500ml/min, stirring velocity 150r/min condition under, cultivate 6 days, obtained fermented liquid;
Above-mentioned fermention medium component is as follows, is all weight percentage:
Sucrose 5%, dregs of beans 10%, excess water, initial pH6.5;
(2) by fermentation liquor thalline broken wall obtained for step (1), filtration, obtained fructosyl transferase crude enzyme liquid; Wherein, thalline breaks and adopts nature cracking process, concrete steps comprise: carried out filtering and collecting thalline by the thalline after step (1) fermentation culture with filter cloth, thalline is washed 3 times with sterilized water, after continuing to cultivate thalline 3d in the sterilized water of access and fermented liquid same volume, cross and filter broken thalline, obtain fructosyl transferase enzyme liquid.
(3) fructosyl transferase crude enzyme liquid is carried out vacuum-concentrcted, concentrate the enzyme activity of rear feed liquid with high effective liquid chromatography for measuring, control the enzyme work of concentrated solution at 120U/ml;
(4) concentrated rear fructosyl transferase enzyme liquid is carried out compound immobilization process with chitosan, sodium alginate and calcium chloride respectively, finally obtained immobilized fructosyltransferase.Wherein, compound immobilization comprises covalent immobilization and embedded immobilization two step.
In the present embodiment, the reaction conditions of covalent immobilization is: select chitosan to be covalent immobilization carrier, after fructosyl transferase is mixed in the ratio of enzyme 200U/g carrier alive with chitin carrier, in 1%(w/w) ratio add linking agent glutaraldehyde, adjust ph 6.5, covalent immobilization temperature 30 DEG C, stirs 10h with the speed of 100r/min.
In the present embodiment, the reaction conditions of embedded immobilization is: after fully being washed by the fructosyl transferase of covalent immobilization, centrifuge dripping.Be 2%(w/w by strength of solution) sodium alginate soln fully mix under 10 DEG C of conditions with the fructosyl transferase through covalent immobilization, blending ratio is the fructosyl transferase adding 1g covalent immobilization in every 100ml sodium alginate soln.By feed liquid after mixing, slowly to instill strength of solution be 5%(w/w) CaCl2 solution in, form dripping pill.After finally the dripping pill of formation being put into the CaCl2 solution hardening treatment 10h of same concentration, namely obtain the immobilized fructosyl transferase of compound.
After testing, the compound immobilization efficiency of fructosyl transferase is high, and the enzyme rate of recovery alive reaches 75%; During batch reactions, can Reusability 90 times.
example two
A compound process for fixation for fructosyl transferase, comprises the steps:
(1) bacterium producing multi enzyme preparation-aspergillus niger is inoculated in fermention medium, temperature 28-32 DEG C, air flow 1000ml/min, stirring velocity 110r/min condition under, cultivate 5 days, obtained fermented liquid;
Above-mentioned fermention medium component is as follows, is all weight percentage:
Sucrose 10%, dregs of beans 7%, excess water, initial pH7.0;
(2) by fermentation liquor thalline broken wall obtained for step (1), filtration, obtained fructosyl transferase crude enzyme liquid; Wherein, thalline breaks and adopts nature cracking process, concrete steps comprise: carried out filtering and collecting thalline by the thalline after step (1) fermentation culture with filter cloth, thalline is washed 4 times with sterilized water, after continuing to cultivate thalline 4d in the sterilized water of access and fermented liquid same volume, cross and filter broken thalline, obtain fructosyl transferase enzyme liquid.
(3) fructosyl transferase crude enzyme liquid is carried out vacuum-concentrcted, concentrate the enzyme activity of rear feed liquid with high effective liquid chromatography for measuring, control the enzyme work of concentrated solution at 150U/ml;
(4) concentrated rear fructosyl transferase enzyme liquid is carried out compound immobilization process with chitosan, sodium alginate and calcium chloride respectively, finally obtained immobilized fructosyltransferase.Wherein, compound immobilization comprises covalent immobilization and embedded immobilization two step.
In the present embodiment, the reaction conditions of covalent immobilization is: select chitosan to be covalent immobilization carrier, after fructosyl transferase is mixed in the ratio of enzyme 150U/g carrier alive with chitin carrier, in 0.75%(w/w) ratio add linking agent glutaraldehyde, adjust ph 7.0, covalent immobilization temperature 35 DEG C, stirs 7h with the speed of 130r/min.
In the present embodiment, the reaction conditions of embedded immobilization is: after fully being washed by the fructosyl transferase of covalent immobilization, centrifuge dripping.Be 3%(w/w by strength of solution) sodium alginate soln fully mix under 12 DEG C of conditions with the fructosyl transferase through covalent immobilization, blending ratio is the fructosyl transferase adding 3g covalent immobilization in every 100ml sodium alginate soln.By feed liquid after mixing, slowly to instill strength of solution be 4%(w/w) CaCl2 solution in, form dripping pill.After finally the dripping pill of formation being put into the CaCl2 solution hardening treatment 15h of same concentration, namely obtain the immobilized fructosyl transferase of compound.
After testing, the compound immobilization efficiency of fructosyl transferase is high, and the enzyme rate of recovery alive reaches 79%; During batch reactions, can Reusability 87 times.
example three
A compound process for fixation for fructosyl transferase, comprises the steps:
(1) bacterium producing multi enzyme preparation-aspergillus niger is inoculated in fermention medium, temperature 28-32 DEG C, air flow 1500ml/min, stirring velocity 80r/min condition under, cultivate 4 days, obtained fermented liquid;
Above-mentioned fermention medium component is as follows, is all weight percentage:
Sucrose 15%, dregs of beans 5%, excess water, initial pH7.5;
(2) by fermentation liquor thalline broken wall obtained for step (1), filtration, obtained fructosyl transferase crude enzyme liquid; Wherein, thalline breaks and adopts nature cracking process, concrete steps comprise: carried out filtering and collecting thalline by the thalline after step (1) fermentation culture with filter cloth, thalline is washed 5 times with sterilized water, after continuing to cultivate thalline 5d in the sterilized water of access and fermented liquid same volume, cross and filter broken thalline, obtain fructosyl transferase enzyme liquid.
(3) fructosyl transferase crude enzyme liquid is carried out vacuum-concentrcted, concentrate the enzyme activity of rear feed liquid with high effective liquid chromatography for measuring, control the enzyme work of concentrated solution at 200U/ml;
(4) concentrated rear fructosyl transferase enzyme liquid is carried out compound immobilization process with chitosan, sodium alginate and calcium chloride respectively, finally obtained immobilized fructosyltransferase.Wherein, compound immobilization comprises covalent immobilization and embedded immobilization two step.
In the present embodiment, the reaction conditions of covalent immobilization is: select chitosan to be covalent immobilization carrier, after fructosyl transferase is mixed in the ratio of enzyme 100U/g carrier alive with chitin carrier, in 0.5%(w/w) ratio add linking agent glutaraldehyde, adjust ph 7.5, covalent immobilization temperature 40 DEG C, stirs 5h with the speed of 160r/min.
In the present embodiment, the reaction conditions of embedded immobilization is: after fully being washed by the fructosyl transferase of covalent immobilization, centrifuge dripping.Be 4%(w/w by strength of solution) sodium alginate soln fully mix under 15 DEG C of conditions with the fructosyl transferase through covalent immobilization, blending ratio is the fructosyl transferase adding 5g covalent immobilization in every 100ml sodium alginate soln.By feed liquid after mixing, slowly to instill strength of solution be 3%(w/w) CaCl2 solution in, form dripping pill.After finally the dripping pill of formation being put into the CaCl2 solution hardening treatment 20h of same concentration, namely obtain the immobilized fructosyl transferase of compound.
After testing, the compound immobilization efficiency of fructosyl transferase is high, and the enzyme rate of recovery alive reaches 73%; During batch reactions, can Reusability 92 times.
example four
A compound process for fixation for fructosyl transferase, comprises the steps:
(1) bacterium producing multi enzyme preparation-aspergillus oryzae is inoculated in fermention medium, temperature 28-32 DEG C, air flow 500ml/min, stirring velocity 150r/min condition under, cultivate 4 days, obtained fermented liquid;
Above-mentioned fermention medium component is as follows, is all weight percentage:
Sucrose 5%, dregs of beans 10%, excess water, initial pH6.5;
(2) by fermentation liquor thalline broken wall obtained for step (1), filtration, obtained fructosyl transferase crude enzyme liquid; Wherein, thalline breaks and adopts nature cracking process, concrete steps comprise: carried out filtering and collecting thalline by the thalline after step (1) fermentation culture with filter cloth, thalline is washed 3 times with sterilized water, after continuing to cultivate thalline 5d in the sterilized water of access and fermented liquid same volume, cross and filter broken thalline, obtain fructosyl transferase enzyme liquid.
(3) fructosyl transferase crude enzyme liquid is carried out vacuum-concentrcted, concentrate the enzyme activity of rear feed liquid with high effective liquid chromatography for measuring, control the enzyme work of concentrated solution at 200U/ml;
(4) concentrated rear fructosyl transferase enzyme liquid is carried out compound immobilization process with chitosan, sodium alginate and calcium chloride respectively, finally obtained immobilized fructosyltransferase.Wherein, compound immobilization comprises covalent immobilization and embedded immobilization two step.
In the present embodiment, the reaction conditions of covalent immobilization is: select chitosan to be covalent immobilization carrier, after fructosyl transferase is mixed in the ratio of enzyme 100U/g carrier alive with chitin carrier, in 0.5%(w/w) ratio add linking agent glutaraldehyde, adjust ph 7.5, covalent immobilization temperature 30 DEG C, stirs 5h with the speed of 160r/min.
In the present embodiment, the reaction conditions of embedded immobilization is: after fully being washed by the fructosyl transferase of covalent immobilization, centrifuge dripping.Be 2%(w/w by strength of solution) sodium alginate soln fully mix under 10 DEG C of conditions with the fructosyl transferase through covalent immobilization, blending ratio is the fructosyl transferase adding 5g covalent immobilization in every 100ml sodium alginate soln.By feed liquid after mixing, slowly to instill strength of solution be 3%(w/w) CaCl2 solution in, form dripping pill.After finally the dripping pill of formation being put into the CaCl2 solution hardening treatment 20h of same concentration, namely obtain the immobilized fructosyl transferase of compound.
After testing, the compound immobilization efficiency of fructosyl transferase is high, and the enzyme rate of recovery alive reaches 76%; During batch reactions, can Reusability 88 times.
example five
A compound process for fixation for fructosyl transferase, comprises the steps:
(1) bacterium producing multi enzyme preparation-aspergillus oryzae is inoculated in fermention medium, temperature 28-32 DEG C, air flow 1500ml/min, stirring velocity 80r/min condition under, cultivate 6 days, obtained fermented liquid;
Above-mentioned fermention medium component is as follows, is all weight percentage:
Sucrose 15%, dregs of beans 5%, excess water, initial pH7.5;
(2) by fermentation liquor thalline broken wall obtained for step (1), filtration, obtained fructosyl transferase crude enzyme liquid; Wherein, thalline breaks and adopts nature cracking process, concrete steps comprise: carried out filtering and collecting thalline by the thalline after step (1) fermentation culture with filter cloth, thalline is washed 5 times with sterilized water, after continuing to cultivate thalline 3d in the sterilized water of access and fermented liquid same volume, cross and filter broken thalline, obtain fructosyl transferase enzyme liquid.
(3) fructosyl transferase crude enzyme liquid is carried out vacuum-concentrcted, concentrate the enzyme activity of rear feed liquid with high effective liquid chromatography for measuring, control the enzyme work of concentrated solution at 150U/ml;
(4) concentrated rear fructosyl transferase enzyme liquid is carried out compound immobilization process with chitosan, sodium alginate and calcium chloride respectively, finally obtained immobilized fructosyltransferase.Wherein, compound immobilization comprises covalent immobilization and embedded immobilization two step.
In the present embodiment, the reaction conditions of covalent immobilization is: select chitosan to be covalent immobilization carrier, after fructosyl transferase is mixed in the ratio of enzyme 200U/g carrier alive with chitin carrier, in 1%(w/w) ratio add linking agent glutaraldehyde, adjust ph 6.5, covalent immobilization temperature 40 DEG C, stirs 10h with the speed of 100r/min.
In the present embodiment, the reaction conditions of embedded immobilization is: after fully being washed by the fructosyl transferase of covalent immobilization, centrifuge dripping.Be 4%(w/w by strength of solution) sodium alginate soln fully mix under 15 DEG C of conditions with the fructosyl transferase through covalent immobilization, blending ratio is the fructosyl transferase adding 1g covalent immobilization in every 100ml sodium alginate soln.By feed liquid after mixing, slowly to instill strength of solution be 5%(w/w) CaCl2 solution in, form dripping pill.After finally the dripping pill of formation being put into the CaCl2 solution hardening treatment 10h of same concentration, namely obtain the immobilized fructosyl transferase of compound.
After testing, the compound immobilization efficiency of fructosyl transferase is high, and the enzyme rate of recovery alive reaches 81%; During batch reactions, can Reusability 92 times.
Claims (9)
1. a compound process for fixation for fructosyl transferase, is characterized in that, comprise the steps:
(1) bacterium producing multi enzyme preparation is inoculated in fermention medium, temperature 28-32 DEG C, air flow 500-1500ml/min, stirring velocity 80-150r/min condition under, cultivate 4-6 days, obtained fermented liquid;
Above-mentioned fermention medium component is as follows, is all weight percentage:
Sucrose 5%-15%, dregs of beans 5%-10%, excess water, initial pH6.5-7.5;
(2) by fermentation liquor thalline broken wall obtained for step (1), filtration, obtained fructosyl transferase crude enzyme liquid;
(3) fructosyl transferase crude enzyme liquid is carried out vacuum-concentrcted, concentrate the enzyme activity of rear feed liquid with high effective liquid chromatography for measuring, control the enzyme work of concentrated solution at more than 100U/ml;
(4) concentrated rear fructosyl transferase enzyme liquid is carried out compound immobilization process with chitosan, sodium alginate and calcium chloride respectively, finally obtained immobilized fructosyltransferase.
2. the method for claim 1, is characterized in that, the bacterium producing multi enzyme preparation in described step (1) is aspergillus niger or aspergillus oryzae.
3. the method for claim 1, is characterized in that, the thalline in described step (2) break adopt method be nature cracking process.
4. the method for claim 1, is characterized in that, in described step (2), condition is as follows:
With filter cloth, the thalline after step (1) fermentation culture is carried out filtering and collecting thalline, thalline is washed 3-5 time with sterilized water, after continuing to cultivate thalline 3-5d in the sterilized water of access and fermented liquid same volume, cross and filter broken thalline, obtain fructosyl transferase enzyme liquid.
5. the method for claim 1, is characterized in that, in described step (4), compound immobilization comprises covalent immobilization and embedded immobilization two step.
6. method as claimed in claim 5, it is characterized in that, the reaction conditions of covalent immobilization is:
Chitosan is selected to be covalent immobilization carrier, after fructosyl transferase is mixed in the ratio of enzyme 100-200U/g carrier alive with chitin carrier, in 0.5%-1%(w/w) ratio add linking agent glutaraldehyde, adjust ph 6.5-7.5, covalent immobilization temperature 30-40 DEG C, stirs 5-10h with the speed of 100-160r/min.
7. method as claimed in claim 5, it is characterized in that, the reaction conditions of embedded immobilization is: after fully being washed by the fructosyl transferase of covalent immobilization, centrifuge dripping; Fully mixed under 10-15 DEG C of condition with the fructosyl transferase through covalent immobilization by sodium alginate soln, blending ratio is the fructosyl transferase adding 1-5g covalent immobilization in every 100ml sodium alginate soln; Solution after mixing is slowly instilled CaCl
2in solution, form dripping pill; Finally the dripping pill of formation is put into the CaCl of same concentration
2in solution after hardening treatment 10-20h, namely obtain the immobilized fructosyl transferase of compound.
8. method as claimed in claim 7, it is characterized in that, described sodium alginate soln concentration is 2%-4%(w/w).
9. method as claimed in claim 7, is characterized in that, described CaCl
2strength of solution is 3%-5%(w/w).
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201510000076.9A CN104560936A (en) | 2015-01-02 | 2015-01-02 | Compound immobilization method for fructosyl transferase |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201510000076.9A CN104560936A (en) | 2015-01-02 | 2015-01-02 | Compound immobilization method for fructosyl transferase |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN104560936A true CN104560936A (en) | 2015-04-29 |
Family
ID=53078016
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201510000076.9A Pending CN104560936A (en) | 2015-01-02 | 2015-01-02 | Compound immobilization method for fructosyl transferase |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN104560936A (en) |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105112306A (en) * | 2015-09-21 | 2015-12-02 | 山东百龙创园生物科技有限公司 | Aspergillus oryzae as well as culture method and application thereof |
| CN107245479A (en) * | 2017-07-12 | 2017-10-13 | 广西驰胜农业科技有限公司 | A kind of FOS production method |
| CN107699558A (en) * | 2017-11-23 | 2018-02-16 | 湖北文理学院 | Immobilised enzymes, its preparation method and its application in atrazine-contaminated soil is repaired |
| CN110172407A (en) * | 2018-12-11 | 2019-08-27 | 青岛蔚蓝生物集团有限公司 | One plant of aspergillus oryzae for producing transfructosylase and its application |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1335402A (en) * | 2001-08-12 | 2002-02-13 | 广西大学 | Production process of cane-fruit oligosaccharide with immobilized fructose-base transferase |
| CN101368195A (en) * | 2008-08-22 | 2009-02-18 | 江门量子高科生物工程有限公司 | Preparation method for high purity fructo-oligosaccharide |
| CN101979526A (en) * | 2010-11-05 | 2011-02-23 | 山西三盟实业发展有限公司 | Method for preparing complex enzyme preparation for vinegar |
| CN102618601A (en) * | 2012-04-17 | 2012-08-01 | 广西大学 | Method for preparing sucrose-6-ethyl ester by using biological fermentation and immobilized enzyme methods |
-
2015
- 2015-01-02 CN CN201510000076.9A patent/CN104560936A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1335402A (en) * | 2001-08-12 | 2002-02-13 | 广西大学 | Production process of cane-fruit oligosaccharide with immobilized fructose-base transferase |
| CN101368195A (en) * | 2008-08-22 | 2009-02-18 | 江门量子高科生物工程有限公司 | Preparation method for high purity fructo-oligosaccharide |
| CN101979526A (en) * | 2010-11-05 | 2011-02-23 | 山西三盟实业发展有限公司 | Method for preparing complex enzyme preparation for vinegar |
| CN102618601A (en) * | 2012-04-17 | 2012-08-01 | 广西大学 | Method for preparing sucrose-6-ethyl ester by using biological fermentation and immobilized enzyme methods |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105112306A (en) * | 2015-09-21 | 2015-12-02 | 山东百龙创园生物科技有限公司 | Aspergillus oryzae as well as culture method and application thereof |
| CN105112306B (en) * | 2015-09-21 | 2018-12-18 | 山东百龙创园生物科技股份有限公司 | One Aspergillus oryzae and its cultural method and application |
| CN107245479A (en) * | 2017-07-12 | 2017-10-13 | 广西驰胜农业科技有限公司 | A kind of FOS production method |
| CN107699558A (en) * | 2017-11-23 | 2018-02-16 | 湖北文理学院 | Immobilised enzymes, its preparation method and its application in atrazine-contaminated soil is repaired |
| CN110172407A (en) * | 2018-12-11 | 2019-08-27 | 青岛蔚蓝生物集团有限公司 | One plant of aspergillus oryzae for producing transfructosylase and its application |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN104560936A (en) | Compound immobilization method for fructosyl transferase | |
| Li et al. | On-site cellulase production and efficient saccharification of corn stover employing cbh2 overexpressing Trichoderma reesei with novel induction system | |
| Caudan et al. | Multiple EPS interactions involved in the cohesion and structure of aerobic granules | |
| CN103343117B (en) | Preparation method of immobilized cephalosporin C acylase | |
| CN110295209B (en) | Technological method for extracting alginate oligosaccharides by enzymatic hydrolysis of gulfweed | |
| CN104846035B (en) | A kind of method that sesbania enzyme process prepares galactomannan oligosaccharide | |
| CN112760311B (en) | Enzyme solution with better enzyme activity ratio of beta-mannase to alpha-galactosidase, and preparation method and application thereof | |
| CN105431534A (en) | beta-1,3-glucanase, polynucleotide, recombinant vector, transformant, production method for beta-1,3-glucanase, enzyme preparation, and production method for paramylon having reduced molecular weight | |
| CN108085353B (en) | Method for producing N-acetylglucosamine by degrading solid chitin with insect chitinase | |
| CN107312515B (en) | A kind of multi-element biologic compound oil displacement agent system and its injection technology | |
| CN1425696A (en) | Preparation of crust oligosaccharide and use | |
| CN103114113A (en) | Preparation method for k-carrageenan oligosaccharide with low polymerization degree | |
| CN100383153C (en) | Method for decoloring digest of poly sialic acid | |
| JP7141774B2 (en) | Method for preparing purified solution of poplar xylooligosaccharide, purified solution of poplar xylooligosaccharide prepared by the method, solid xylooligosaccharide and use thereof | |
| CN110128489B (en) | Method for preparing galactomannan-oligosaccharide by autohydrolysis | |
| CN106222160B (en) | The preparation method and applications of the compound alginate fixation support of molasses colloid | |
| RU2010124696A (en) | SYSTEMS AND METHODS FOR CHANGING THE SPEED OF ENZYMATIC PROCESSES | |
| CN103146784A (en) | Method for preparing chitosan oligosaccharide through degradation of chitosan by thermobifida fusca internally tangent beta-1, 4-glucanase | |
| CN111534556B (en) | Method for preparing high-concentration monosaccharide solution by using poplar enzyme method | |
| CN102286414B (en) | Chitin-degrading bacterial strain and method for preparing chitooligosaccharide by utilizing same | |
| CN108017155A (en) | A kind of preparation method of microalgae sewage-treating agent | |
| CN104560774B (en) | Method for preparing block oligosaccharide containing rich rhamnose sulfate from Enteromorpha polysaccharide | |
| CN106867990B (en) | Preparation method of immobilized beta-glucosidase | |
| CN105238770A (en) | Method for preparing endoinulinase through in-situ orientation dissolution and process for preparing fructo-oligosaccharide | |
| CN104450665A (en) | Method for immobilizing phaffia rhodozyma and preparing neokestose and application of method |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| WD01 | Invention patent application deemed withdrawn after publication | ||
| WD01 | Invention patent application deemed withdrawn after publication |
Application publication date: 20150429 |