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Cross-linked microsphere as well as preparation method and application thereof

A technology of cross-linking microspheres and microspheres, which is applied in the fields of biochemical equipment and methods, microorganisms, artificial cell structures, etc., can solve the problems of high viscosity, unsatisfactory size, and narrow application range of chitosan.

Pending Publication Date: 2020-06-12
苏州新丝原生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] In order to solve the problem that existing microcarriers contain toxic cross-linking agent residues and the size does not meet the requirements, chitosan alone has a narrow application range and high viscosity, gelatin alone has poor stability, and microspheres prepared by spray freezing alone are fragile , low stability, etc., the present invention combines spray freeze-drying and chemical cross-linking for the first time, that is, prepares chitosan-gelatin physical mixed microspheres by spray freeze-drying for the first time, and then suspends them in the cross-linking system. Chitosan-gelatin microcarriers with a diameter of 100-400 μm were prepared by chemical cross-linking while maintaining their structural integrity

Method used

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  • Cross-linked microsphere as well as preparation method and application thereof
  • Cross-linked microsphere as well as preparation method and application thereof
  • Cross-linked microsphere as well as preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0094] Chitosan powder (1.2g, degree of deacetylation is 98.7%, viscosity is 29mPa.s, purchased from Zhejiang Golden Shell Pharmaceutical Co., Ltd.) was added to 2% (w / v) glacial acetic acid aqueous solution (60mL), Dissolve at 65° C. for 5 h, and let stand for 24 h to obtain a bubble-free, uniform and transparent 2% (w / v) chitosan solution (60 mL).

[0095] Add type A gelatin (2.4g) with 250 freezing strength (double freezing strength) into deionized water (60mL), dissolve at 65°C, and let it stand for 24h to obtain stable and transparent 4% (w / v) gelatin solution (60 mL).

[0096] Mix the chitosan solution and the gelatin solution, and use a 13000rpm high-speed homogenizer to homogenize for 10 minutes to make them mix uniformly to obtain a chitosan-gelatin mixed solution with a solid content of 3.0%.

[0097] The chitosan-gelatin mixed solution was placed in the atomizer of the spray freezing tower, and sprayed into the cooling tower pre-cooled to -60 ° C, the frozen micros...

Embodiment 2

[0100] Chitosan powder (2.0g, degree of deacetylation is 90%, viscosity is 112mPa.s, purchased from Zhejiang Golden Shell Pharmaceutical Co., Ltd.) was added to 2% (w / v) glacial acetic acid aqueous solution (100mL), Dissolve at 65° C. for 5 h, and let stand for 24 h to obtain a bubble-free, uniform and transparent 2% (w / v) chitosan solution (100 mL).

[0101] Add type A gelatin (1.0g) with 250 freezing strength (double freezing strength) into deionized water (25mL), dissolve at 65°C, and let stand for 24h to obtain stable, transparent 4% (w / v) gelatin solution (25 mL).

[0102] Mix the chitosan solution and the gelatin solution, and use a 13000rpm high-speed homogenizer to homogenize for 10 minutes to make them mix uniformly to obtain a chitosan-gelatin mixed solution with a solid content of 2.4%.

[0103] The chitosan-gelatin mixed solution was placed in the atomizer of the spray freezing tower, and sprayed into the cooling tower pre-cooled to -60 ° C, the frozen microsphere...

Embodiment 3

[0106] Chitosan powder (1.2g, degree of deacetylation is 98.7%, viscosity is 29mPa.s, purchased from Zhejiang Golden Shell Pharmaceutical Co., Ltd.) was added to 2% (w / v) glacial acetic acid aqueous solution (60mL), Dissolve at 65° C. for 5 h, and let stand for 24 h to obtain a bubble-free, uniform and transparent 2% (w / v) chitosan solution (60 mL).

[0107] Add type A gelatin (2.4g) with 250 freezing strength (double freezing strength) into deionized water (60mL), dissolve at 65°C, and let it stand for 24h to obtain stable and transparent 4% (w / v) gelatin solution (60 mL).

[0108] Mix the chitosan solution and the gelatin solution, and use a 13000rpm high-speed homogenizer to homogenize for 10 minutes to make them mix uniformly to obtain a chitosan-gelatin mixed solution with a solid content of 3.0%.

[0109] Put the chitosan-gelatin mixed solution in the atomizer of the spray freezing tower, and spray it into the cooling tower pre-cooled to -80°C, collect the frozen micros...

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Abstract

The invention belongs to the field of biological tissue engineering cytoskeleton materials, and relates to a cross-linked microsphere as well as a preparation method and application thereof. Accordingto the method, chitosan-gelatin physical mixed microspheres are prepared in a spray freeze drying mode, and then amino and carboxyl in the physical mixed microspheres are subjected to chemical crosslinking, so that crosslinked microspheres are prepared. The problem that the microsphere structure is unstable due to a single spray freezing mode is solved; meanwhile, the cell adhesion characteristicof the microsphere can be improved by combining chitosan and gelatin, cell culture efficiency can be improved by using the microsphere as a microcarrier, and two-dimensional static culture and three-dimensional dynamic culture can be achieved. The chitosan and gelatin raw materials are low in cost, the large-scale production cost of the physically mixed microspheres and the chemically cross-linked microspheres is low, toxic substance residues are avoided, large-scale cell culture can be conducted, and great application prospects are achieved in the aspect of in-vivo injection.

Description

technical field [0001] The invention belongs to the field of biological tissue engineering cell scaffold materials, and relates to a preparation method of chitosan-gelatin crosslinked microspheres, the microspheres prepared by the preparation method, and the application of the microspheres as cell culture microcarriers. Background technique [0002] In recent years, in the treatment of diseases such as diabetes, keratopathy, and cartilage tissue damage, a large number of research results have been obtained around cell therapy, and some of them have been applied clinically, with great application prospects. Microcarrier culture technique is an effective means to achieve large-scale culture of adherent cells. At present, some solid or porous microcarriers have been commercialized abroad, such as the Cytopore of GE Healthcare. TM , Cytodex TM , Cytoline TM and other series of microspheres, as well as Corning's (CORNING) soluble microcarriers (DMC), etc. At the same time, ...

Claims

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Application Information

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IPC IPC(8): C08J3/24C08J3/12C08J9/28C08L89/00C08L5/08C12N5/071
CPCC08J3/246C08J3/122C08J9/28C12N5/0062C08J2389/00C08J2405/08C08J2305/08C08J2489/00C08J2201/0484C12N2533/72C12N2533/54
Inventor 孙振华左丽娜施坤宁丁升祥杨慧莹蒋雪峰
Owner 苏州新丝原生物科技有限公司
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