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Magnetic crosslinking lipase aggregation as well as preparation method and application thereof

A technique of lipase, aggregates, applied in the direction of immobilization on or in inorganic carriers, immobilization on/in organic carriers, etc.

Inactive Publication Date: 2013-09-25
INST OF PROCESS ENG CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, magnetic nanoparticles are rarely used to prepare magnetically cross-linked lipase aggregates, and there are few reports, especially the preparation of magnetic cross-linked lipase aggregates using functionalized magnetic nanoparticles with dendritic polyamine groups on the surface. No literature report

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] (1) Weigh 15 mg of self-made functionalized magnetic nanoparticles with dendritic amine groups on the surface (preparation method: magnetic nanopolymers with surface amination (see: Guobin Shan, Huizhou Liu, Jianmin Xing, Guandong Zhang, Zhiya) Ma,Xianqiao Liu,Junguo Liu.Surface Modification of Nano-magnetic Fe3O4 Particles and its Applications.China-EU Forum on Nanosized Technology,2002,Beijing,China.P68-76.) as the core, with ethylenediamine and methyl acrylate As a monomer, the dendritic material is synthesized by the divergent synthesis method well known to those skilled in the art.) Placed in a triangular flask containing 15ml of 100mM phosphate buffer with pH=8, under ultrasonic conditions with a frequency of 40KHz and a power of 240w Ultrasound in an ice bath for 20 minutes, and the dispersion is even;

[0042] (2) Weigh 250mg of lipase (in terms of protein content), place it in 250ml of 100mM phosphate buffer, stir magnetically for 30 minutes at room temperature to...

Embodiment 2

[0047] (1) Weigh 12 mg of functionalized magnetic nanoparticles with dendritic amine groups on the surface and place them in a triangular flask containing 12 ml of 100 mM phosphate buffer at pH=8. Under ultrasonic conditions with a frequency of 40KHz and a power of 240w Ultrasound in ice bath for 20min, uniform dispersion;

[0048] (2) Weigh 150mg of lipase (based on protein content) in 150ml of 100mM phosphate buffer, stir it magnetically for 30min at room temperature to fully dissolve the enzyme to form a homogeneous solution, and pour it into the magnetic dispersion obtained in (1) In the particle system, shake at room temperature for 40 minutes;

[0049] (3) In (2), add 150ml ethanol dropwise, then stir at 4℃ for 30min;

[0050] (4) Add 5 ml of 25% glutaraldehyde to (3), shake at room temperature for 8 hours, after magnetic separation, take the solid, wash thoroughly with distilled water, and freeze-dry in vacuum for 36 hours to obtain magnetic cross-linked lipase aggregates.

[...

Embodiment 3

[0053] (1) Weigh 10 mg of functionalized magnetic nanoparticles with dendritic amine groups on the surface and place them in a triangular flask containing 10 ml of 100 mM phosphate buffer at pH=8. Under ultrasonic conditions with a frequency of 40KHz and a power of 240w Ultrasound in ice bath for 30min, uniform dispersion;

[0054] (2) Weigh 120mg of lipase (calculated as protein content) in 120ml of 100mM phosphate buffer, stir magnetically for 30min at room temperature to fully dissolve the enzyme to form a homogeneous solution, and pour it into the magnetic dispersion prepared in (1) In the particle system, shake at room temperature for 40 minutes;

[0055] (3) Add 150ml of acetonitrile dropwise in (2), then stir at 4°C for 30min;

[0056] (4) Add 4ml of 25% glutaraldehyde to (3), shake at room temperature for 8 hours, after magnetic separation, take the solid, wash thoroughly with distilled water, and freeze-dry in vacuum for 36 hours to obtain magnetic cross-linked lipase aggre...

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Abstract

The invention discloses a magnetic crosslinking lipase aggregation as well as a preparation method and application thereof. The magnetism crosslinking lipase aggregation is prepared from functional group magnetic nanoparticles with dendritic polyamine on the surface. The preparation method integrates a crosslinking enzyme aggregation technology and a magnetic immobilized enzyme technology, not only are the enzyme immobilization efficiency and enzyme activity recovery rate of the magnetism immobilized enzyme improved, but also the problem of the poor operational stability of the crosslinking enzyme aggregation is also greatly improved. The prepared magnetic crosslinking lipase aggregation has high activity and excellent operational stability, and can be used as a biocatalyst widely.

Description

Technical field [0001] The invention belongs to the field of cross-linked enzyme aggregates, and specifically relates to a magnetic cross-linked lipase aggregate prepared by using functionalized magnetic nanoparticles with dendritic amine groups on the surface, and a preparation method and application thereof. Background technique [0002] As a catalyst, biological enzymes are widely used in medicine, food, fragrance, oil, cosmetics and pesticide industries because of their high efficiency, specificity, mild reaction conditions, and environmental friendliness. (Reetz, MT, 2002. Lipases as practical biocatalysts.Curr.Opin.Chem.Biol.6,145–150). Among them, lipase has a wide range of substrates due to its action, showing a broader application prospect. However, the natural lipase has poor operational stability, is easy to inactivate, is not easy to reuse, and is difficult to achieve process continuity and automation. After the reaction is mixed into the product, it often causes dif...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N11/14C12N11/08
Inventor 刘春朝刘英郭晨
Owner INST OF PROCESS ENG CHINESE ACAD OF SCI
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