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Immobilization of mannase and application of mannase in enzyme reactor

A mannanase and enzyme reactor technology, applied in the directions of immobilized enzymes, glycosylases, enzymes, etc., can solve the problems of complicated preparation, complicated separation of substrates and enzymes, lower relative enzyme activity, etc., and reduce production. cost, realize the reuse of enzymes, and solve the effect of high viscosity

Pending Publication Date: 2022-02-18
承德康尔润食品有限公司 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Patent CN105274163A provides a method for preparing mannose by mixing black tea leaves with Trichoderma spores, which is complex and does not mention reaction batches; patent CN105524911A provides a method for preparing immobilized mannanase using nanofiber materials The method of mannan oligosaccharides uses the immobilized enzyme to react with the substrate, the separation of the substrate and the enzyme is complicated, and the actual use batch is not mentioned, which cannot reflect the advantages of the immobilized enzyme; patent CN101492667A provides a method using The method of immobilizing enzyme-producing microbial cells using microporous membranes, using the immobilized cells to carry out 4-8 consecutive reactions, the relative enzyme activity was reduced from the initial 200% to 50%, which greatly limited its practical application; Patent CN102586219A provides a kind of immobilized mannanase prepared by adsorbing macroporous resin, and applied to the degradation of aloe polysaccharides, but this method does not mention the application in polysaccharides such as guar gum, and does not mention use batch

Method used

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  • Immobilization of mannase and application of mannase in enzyme reactor
  • Immobilization of mannase and application of mannase in enzyme reactor
  • Immobilization of mannase and application of mannase in enzyme reactor

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0047] Example 1 Carrier Screening

[0048] Choose different carriers (Xi'an Lanxiao Technology New Materials Co., Ltd.) LX-103B, LX-1000HA(A), LX-HFA001, LX1000HFA, LX-1000EA, (Aimee Kejian China Biomedical Co., Ltd.) LKZ126, LKZ618, (Tianjin Nankai Hecheng Technology Co., Ltd.) Each 2g of ES-101 and ESR-1 was divided into 150ml Erlenmeyer flasks, and 2g of mannanase was added to each Erlenmeyer flask, and 50mL of 0.1M pH7 phosphate buffer was added. , sealed, placed on a shaker, 25°C, 15rpm, immobilized for 48h. Use a Buchner funnel to extract the liquid, retain the immobilized enzyme, and measure the activity of the immobilized enzyme.

[0049] Measure the immobilized enzyme activity of different carriers, the results are shown in Table 2 and figure 2 :

[0050] Table 2 The results of enzyme activity determination of mannanase immobilized on different carriers

[0051]

[0052] From the above results, it can be seen that LX-HFA001 has the highest enzyme activity, an...

Embodiment 2

[0053] Embodiment 2 optimal immobilization conditions

[0054] The optimal immobilization conditions were determined by single factor variable method.

[0055] (1) Effects of different concentrations of phosphate on the activity of immobilized enzymes: 0.05M, 0.1M, 0.2M phosphate buffers were selected as buffers, and immobilization conditions were adopted: mannanase (Beijing Guarrun Technology Co., Ltd.) Dry powder 2g, LX-HFA001 carrier 2g, 25°C, pH 7, shake at 150rpm for 48h. Determine the optimal buffer concentration by measuring the activity of the immobilized enzyme.

[0056] The effects of different buffer concentrations on the activity of immobilized enzymes are shown in Table 3.

[0057] Table 3 Effect of different phosphate concentrations on immobilization

[0058]

[0059] It can be seen from the data in Table 3 that different concentrations of phosphate have a greater impact on the enzyme activity of the immobilized enzyme. The optimum concentration is 0.1M. ...

Embodiment 3

[0081] Embodiment 3 immobilized enzyme reactor prepares mannan oligosaccharide

[0082] Enzyme reactor assembly: take an appropriate amount (selected according to the size of the chromatographic column) of immobilized mannanase prepared under optimal conditions, and fill it into the chromatographic column (the filling volume is 2 / 3 of the column volume); Figure 7 The enzyme reactor is assembled as shown.

[0083] Detection of mannan oligosaccharides: Use the DNS method to measure the amount of reducing sugar in the reaction solution: take 200 μL of the samples taken and place them in 10 mL centrifuge tubes, then put 200 μL of acetic acid-sodium acetate buffer into the centrifuge tubes solution, then add 500 μL of DNS reagent, shake slightly to mix evenly, plug the cap of the centrifuge tube tightly, heat it in a boiling water bath for 5 minutes, take it out immediately, and quickly cool it to room temperature with flowing cold water, then add deionized Dilute the water to 2....

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Abstract

The invention discloses immobilization of mannase and application of the mannase in an enzyme reactor. The preparation method comprises the following steps: taking materials according to the mass ratio of mannase to a carrier of 1:1, adding 0.1M of a phosphate buffer solution with pH of 5, and carrying out immobilization reaction at 25DEG C for 24 hours. A resin immobilization carrier is employed for immobilization treatment on mannase so as to investigate the optimal immobilization condition of the mannase; the immobilized enzyme is used for preparing the enzyme reactor, and the reactor is used for preparing mannan oligosaccharide, so that the enzyme is repeatedly utilized (15 batches or more), the defect of high viscosity of a reaction substrate is overcome, and a technical foundation is laid for reducing the production cost of the mannan oligosaccharide.

Description

technical field [0001] The invention belongs to the technical field of mannan oligosaccharide preparation, and in particular relates to the immobilization of mannanase and its application in an enzyme reactor. Background technique [0002] Mannan oligosaccharides refer to oligosaccharides connected by 2-10 mannose through β-1,4 glycosidic bonds. It is a new type of antigenic active substance extracted from the cell wall of yeast culture at the earliest. It is widely found in konjac flour and melon Bean gum, kale gum and various microbial cell walls. Mannan oligosaccharide has the characteristics of stable properties, low calorie, no dental caries, and lower blood sugar, so it is widely used in the food industry as a new generation of functional food ingredients. [0003] At present, mannan oligosaccharides are prepared from mannan such as konjac flour, locust bean gum, coffee grounds and other raw materials. The manufacturing method is enzymatic decomposition, chemical met...

Claims

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Application Information

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IPC IPC(8): C12N11/089C12P19/14C12P19/12C12P19/04C12P19/00C12N9/24
CPCC12N11/089C12P19/14C12P19/12C12P19/04C12P19/00C12N9/2491C12Y302/01025
Inventor 刘力强张伟谷雅文曹毅武建刘燕静袁江宏袁润
Owner 承德康尔润食品有限公司
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