Method for preparing microbe carrier for immobilization of Flavobacterium through refrigerating-defrosting twice

A technology for immobilizing microorganisms and Flavobacteria, which is applied in the direction of immobilization on/in the organic carrier, can solve the problems of poor diffusion and mass transfer performance, low mechanical strength, and small elasticity, and achieve arbitrary control of shape and size, mechanical High strength and excellent elasticity

Inactive Publication Date: 2006-07-05
SHENYANG INST OF APPLIED ECOLOGY - CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Chemically cross-linked immobilized products are generally regular spherical particles, but the chemical reaction is intense during the granulation process, the loss of microbial activity is large, and operating parameters need to be strictly controlled, resulting in a long process
Physical cross-linking is generally achieved by freezing. After long-term low-temperature freezing of PVA and Na Alg, the carrier is easy to be brittle and loses elasticity. Although the shape of the carrier can be controlled arbitrarily, the process is short, the reaction is mild, and the loss of microbial activity is small, but the product’s Poor diffusion and mass transfer performance, uneven pores, low mechanical strength, small elasticity, low microbial density after proliferation and culture, and easy loss, and immobilized products are easily broken after long-term use
[0004] When using the above-mentioned physical cross-linking method to fix Flavobacterium sp., because Flavobacterium colonies are easy to slide and move, bacteria loss often occurs, and because the metabolites of Flavobacterium are acidic, they can react with Na Alg, resulting in Partial dissolution of Na Alg leads to failure of immobilization

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] (1) Incline culture of Flavobacterium sp.

[0022] Inject Flavobacterium sp. into a test tube of beef peptone basal medium (mass percentage: 0.3% beef extract, 1.0% peptone, 0.5% NaCl, 1.5-2.0% agar, pH=7.0-7.2), place at 28°C-30 Cultivate in a constant temperature incubator for 24 h.

[0023] (2) Flavobacterium sp. liquid seed preparation

[0024] Pick up 2 loops of bacterial lawn on the slant medium with an inoculation loop, and insert it into the liquid seed medium (mass percentage: 1.25% glucose, 0.25% yeast extract, 0.1% NH NO , 0.02% MgSO 7H O, 0.02% KCl, pH =7.0-7.2), cultured for 16-20 hours under the condition of shaker rotation speed 135rpm-145rpm, 28°C-30°C, to obtain liquid seeds.

[0025] (3) Preparation of gel solution

[0026] Weigh 10.5% PVA powder, 1.0% Na Alg, 6.0% AC, and 1.5% zeolite powder according to the mass percentage, mix them evenly, soak them in tap water with a mass percentage of 81% for 24 hours, and prepare 200ml of gel liquid; damp hea...

Embodiment 2

[0034]The difference from Example 1 is:

[0035] When preparing the gel solution, weigh 9.8% PVA powder, 0.3% Na Alg, 4.5% AC, and 0.5% zeolite powder according to the mass percentage, mix them evenly, soak them in tap water with a mass percentage of 84.9% for 24 hours, and prepare 200ml of the gel solution ; The preparation mass concentration of the reinforcement solution is 7.5% Na 2 SO 4 ; When preparing the carrier, the first freezing temperature is minus 10°C, freezing for 15 hours, the second freezing temperature is minus 10°C, freezing for 15 hours, and soaking in the reinforcing agent for 15 hours. The mechanical strength of the immobilized carrier prepared according to the above steps is 15.2kg / cm 2 . After the regeneration treatment, the bacteria reproduced normally, and the mechanical strength of the carrier was 15.0kg / cm 2 .

Embodiment 3

[0037] The difference from Example 1 is:

[0038] When preparing the gel solution, weigh 10.0% PVA powder, 0.5% Na Alg, 5% AC, and 1% zeolite powder according to the mass percentage, mix them evenly, soak them in tap water with a mass percentage of 84.5% for 24 hours, and prepare 200ml of the gel solution ; The preparation mass concentration of the reinforcement solution is 8.0% Na 2 SO 4 . When preparing the carrier, the first freezing temperature is minus 13°C, freezing for 16 hours, the second freezing temperature is minus 11°C, freezing for 16 hours, and soaking in the reinforcing agent for 18 hours. The mechanical strength of the immobilized carrier prepared according to the above steps is 18.6kg / cm 2 . After the regeneration treatment, the bacteria reproduced normally, and the mechanical strength of the carrier was 18.4kg / cm 2 .

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Abstract

This invention describes a process for preparing a microbe carrier prepared by two times freezing-defreezing of flavorbacteria, comprising the steps of: adding appropriate amounts of activated carbon, zeolite powders and Tween 80 into the PVA- NaíñAlg gel; uniformly mixing the gel and the flavorbacteria seed liquid, and then pouring the mixed liquid into a culture plate that is covered with two layers of non-woven fabrics at 35-48 deg.C; freezing the culture plate in a refrigerator at 10-15 deg.C for 15-18h, defreezing, and repeating the procedure once again; solidifying the mixed liquid with the solification agent, immersing in sterilized water for a time, and then cutting for enrichment culture. The carrier in this invention has good elasticity, diffusion and mechanical properties. The flavorbacteria immobilized have good activities without any leakage.

Description

technical field [0001] The invention relates to bacteria immobilized biological preparations, in particular to a preparation method of sodium alginate (Na·Alg)-polyvinyl alcohol (PVA)-non-woven fabric double freezing-thawing cross-linked immobilized microbial carrier. Background technique [0002] PVA and Na Alg are two kinds of immobilization carriers with excellent performance, which have good biocompatibility and molding controllability, and have a good embedding effect on most microorganisms. It has been widely used in the fields of development and chemical synthesis. [0003] There are two traditional methods of embedding and fixing PVA and Na Alg: chemical cross-linking and physical cross-linking. Chemically cross-linked immobilized products are generally regular spherical particles, but the chemical reaction during the granulation process is intense, the loss of microbial activity is large, and operating parameters need to be strictly controlled, resulting in a long ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N11/04
Inventor 李海波李培军鞠京丽张轶许华夏
Owner SHENYANG INST OF APPLIED ECOLOGY - CHINESE ACAD OF SCI
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