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Method for immobilizing D-psicose3-epimerase through polydopamine-magnetic Fe3O4 nanoparticles

An epimerase, psicose technology, applied in the directions of isomerase, immobilized on or in an inorganic carrier, immobilized on/in an organic carrier, etc., can solve the problem of reduced magnetic separation efficiency, chemical Problems with high activity, loss of magnetism and dispersibility

Active Publication Date: 2019-10-25
山东星光首创生物科技有限公司 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Fe 3 o 4 Due to the high surface energy caused by the large specific surface area, magnetic nanoparticles tend to agglomerate. The characteristics of high chemical activity and easy oxidation will lead to the loss of magnetism and dispersion, resulting in a decrease in magnetic separation efficiency and affecting the adsorption effect of immobilization.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030]Embodiment 1 A kind of polydopamine-magnetic Fe 3 o 4 Method for immobilizing D-psicose 3-epimerase on nanoparticles

[0031] (1) Preparation of D-psicose 3-epimerase: Inoculate the recombinant bacteria Bacillus subtilis 1A751 / pUB-P43dpe-dal into LB liquid medium and culture overnight at 37°C. Transfer to LB medium with 3% inoculum size, and culture at 37°C for 14h. Collect the fermentation broth, wash the cells with a pH 7.5, 50 mmol / L Tris-HCl buffer solution after refrigerated centrifugation, and resuspend the cells with a pH 7.5, 50 mmol / L Tris-HCl buffer solution. Add 20mg / mL lysozyme, and keep warm in a 37°C water bath for 30min. The cells were sonicated for 10 minutes, centrifuged at 4°C for 20 minutes, and the supernatant was taken. Filter through a 0.45 μm filter membrane, freeze and vacuum-dry the crude enzyme solution to make crude enzyme powder.

[0032] (2) Polydopamine-magnetic Fe 3 o 4 Preparation of nanoparticles: prepare 50mL 0.2mol / L FeCl respect...

Embodiment 2

[0034] Embodiment 2 A kind of polydopamine-magnetic Fe 3 o 4 Method for immobilizing D-psicose 3-epimerase on nanoparticles

[0035] (1) Preparation of D-psicose 3-epimerase: Inoculate the recombinant bacteria Bacillus subtilis 1A751 / pUB-P43dpe-dal into LB liquid medium and culture overnight at 37°C. Transfer to LB medium with 3% inoculum size, and culture at 37°C for 14h. Collect the fermentation broth, wash the cells with a pH 7.5, 50 mmol / L Tris-HCl buffer solution after refrigerated centrifugation, and resuspend the cells with a pH 7.5, 50 mmol / L Tris-HCl buffer solution. Add 20mg / mL lysozyme, and keep warm in a 37°C water bath for 30min. The cells were sonicated for 10 minutes, centrifuged at 4°C for 20 minutes, and the supernatant was taken. Filter through a 0.45 μm filter membrane, freeze and vacuum-dry the crude enzyme solution to make crude enzyme powder.

[0036] (2) Polydopamine-magnetic Fe 3 o 4 Preparation of nanoparticles: prepare 50mL 0.2mol / L FeCl respec...

Embodiment 3

[0038] Embodiment 3 A kind of polydopamine-magnetic Fe 3 o 4 Method for immobilizing D-psicose 3-epimerase on nanoparticles

[0039] (1) Preparation of D-psicose 3-epimerase: Inoculate the recombinant bacteria Bacillus subtilis 1A751 / pUB-P43dpe-dal into LB liquid medium and culture overnight at 37°C. Transfer to LB medium with 3% inoculum size, and culture at 37°C for 14h. Collect the fermentation broth, wash the cells with a pH 7.5, 50 mmol / L Tris-HCl buffer solution after refrigerated centrifugation, and resuspend the cells with a pH 7.5, 50 mmol / L Tris-HCl buffer solution. Add 20mg / mL lysozyme, and keep warm in a 37°C water bath for 30min. The cells were sonicated for 10 minutes, centrifuged at 4°C for 20 minutes, and the supernatant was taken. Filter through a 0.45 μm filter membrane, freeze and vacuum-dry the crude enzyme solution to make crude enzyme powder.

[0040] (2) Polydopamine-magnetic Fe 3 o 4 Preparation of nanoparticles: prepare 50mL 0.2mol / L FeCl respec...

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Abstract

The invention discloses a method for immobilizing D-psicose3-epimerase through polydopamine-magnetic Fe3O4 nanoparticles, and belongs to the technical field of food processing. The method includes thesteps of preparing the D-psicose3-epimerase and polydopamine-magnetic Fe3O4 nanoparticles, and immobilizing the D-psicose3-epimerase through the polydopamine-magnetic Fe3O4 nanoparticles. The polydopamine-magnetic Fe3O4 nanoparticles have strong magnetic responsibility, can be effectively controlled by magnetic force under the effect of an external magnetic field, can enhance the immobilizing effect through high-activity phenolic hydroxyl groups and other functional groups on the surface of polydopamine, and improves the stability of a carrier and the capacity of an enzyme, and the enzyme activity of the immobilized enzyme can reach 450.45 U per gram of carrier or above.

Description

technical field [0001] The invention relates to a polydopamine-magnetic Fe 3 o 4 The invention discloses a method for immobilizing D-psicose 3-epimerase on nanoparticles, belonging to the technical field of food processing. Background technique [0002] Traditional sweeteners such as sucrose are high in calories, and excessive intake can lead to chronic diseases such as diabetes, obesity, and cardiovascular disease. Therefore, the development of new low-calorie functional sweeteners has become a research hotspot in international food science. At present, functional sweeteners such as xylitol, fructooligosaccharide, and erythritol have been widely used in developed countries, but due to some gaps with sucrose in terms of taste and sweetness, they cannot completely replace sucrose. As a rare sugar, D-psicose is about 70% as sweet as sucrose and contains almost no calories. In addition, D-psicose has physiological functions that glucose and fructose do not have. For example,...

Claims

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Application Information

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IPC IPC(8): C12N11/14C12N11/08C12P19/24C12P19/02H01F1/00
CPCC12N9/90C12N11/08C12N11/14C12P19/02C12P19/24H01F1/0054
Inventor 江波曹永兴潘永胜金辉李宁周焕霞代艺伟张涛
Owner 山东星光首创生物科技有限公司
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