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Epsilon-polylysine fermentation method adopting cell immobilization and off-site and intermittent adsorption

A technology of polylysine and fermentation method, which is applied in the field of bioengineering applications, and can solve the problems of unfavorable improvement of the yield of ε-polylysine, low fermentation yield, disadvantages, etc.

Inactive Publication Date: 2018-12-14
NINGDE NORMAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Reports show that ε-polylysine fermentation mainly uses free cells, but this is not conducive to increasing the cell concentration in the fermentation broth, which is an important factor for the low fermentation yield of ε-polylysine; in addition, ε-polylysine As a secondary metabolite, ε-polylysine accumulates to a certain level in the fermentation process, and it inhibits the further synthesis of cells, which is not conducive to improving the yield of ε-polylysine. Therefore, ε-polylysine fermentation The above two significant unfavorable factors must be overcome in the development of the method, which will further increase the fermentation yield of ε-polylysine

Method used

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  • Epsilon-polylysine fermentation method adopting cell immobilization and off-site and intermittent adsorption

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Effect test

Embodiment 1

[0020] 1. Spore Suspension Preparation

[0021] The ε-polylysine-producing bacteria was Streptomyces anhydrous ( Streptomyces ahygroscopicus ) GIM8, inoculate the fresh slant strains onto the oat agar medium (preparation method as follows) plate, incubate upside down at 30°C for 7 days, wash with sterile water to collect spores, prepare spore suspension, and make the spore concentration about 5 × 10 8 Spores / mL, as inoculum for preparing liquid seeds;

[0022] Preparation of oat agar medium (taking 1 L as an example): Weigh 40 g of oats, wash with tap water, boil in 1200 mL of deionized water for 10 min, filter and discard the residue, add 18 g of agar to the filtrate, heat to dissolve completely, replenish water To 1 L, sterilize at 121°C for 30 min for plate preparation.

[0023] 2. Liquid Seed Preparation

[0024] Inoculate 10 mL of spore suspension into 500 mL of fermentation medium (packed in a 1 L Erlenmeyer flask) at a temperature of 30 °C and a rotation speed of ...

Embodiment 2

[0036] 1. The preparation of spore suspension, liquid seed, and fermentation medium formula are the same as in Example 1

[0037] 2. Immobilization carrier installation and fixation

[0038] Use the sponge as the cell immobilization carrier (cut into length, width and thickness about 30 cm×10 cm×1.5 cm), and fix it to the stirrer, shaft and baffle plate of a 10 L fermenter with fine iron wire, and the dosage is 8 pieces.

[0039] 3. Sterilization, inoculation and fermentation

[0040] Pour 6.7 L of fermentation medium into the fermenter and sterilize at 121°C for 30 minutes; after cooling, use 12.5% ​​ammonia water to adjust the initial pH of the medium to 6.8; add liquid seeds according to the inoculation amount of 5% (v / v); ferment The conditions were ventilation flow rate 1.0 vvm, temperature 30°C, rpm = 0.

[0041] 4. pH control, nutrient supplement, feeding and defoaming

[0042] During the fermentation process, when the pH of the fermentation liquid dropped to 3.8, th...

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Abstract

The invention provides an epsilon-polylysine fermentation method adopting cell immobilization and off-site and intermittent adsorption. Loofah sponge or sponge is used as a cell immobilizing carrier,and is mounted and fixed on a stirrer, a shaft and a baffle plate in a fermenting tank, then a fermentation medium is added, sterilization and cooling are performed, and a liquid seed is introduced. Cells are adsorbed to the immobilizing carrier in the fermentation process to realize cell immobilization; a sterilized organic nitrogen source solution is intermittently added in the epsilon-polylysine synthesis stage in a fed-batch manner; and off-site separation of epsilon-polylysine is carried out by ion exchange resin when the concentration epsilon-polylysine in a fermentation solution is accumulated to 3.0 g / L, and the separation process is repeated when the concentration of the fermentation solution is afresh accumulated to 3.0 g / L. Cell immobilization and off-site separation technologies are used in the epsilon-polylysine fermentation in a combination manner, and are also combined with fed-batch addition of the organic nitrogen source, so cell growth and product synthesis are effectively promoted, the synthesis period is prolonged, the fermentation yield of epsilon-polylysine is significantly improved, and the method has a practical application prospect.

Description

technical field [0001] The invention belongs to the application field of bioengineering, and relates to a fermentation method for separating ε-polylysine by adopting cell immobilization, dislocation and intermittent adsorption. Background technique [0002] ε-polylysine is an extracellular L-lysine polymer synthesized by Streptomyces, and its lysine residues are linked by dehydration condensation of α-carboxyl and ε-amino groups. It is a special-shaped peptide with a molecular weight of usually For 3500-4000. ε-polylysine has a very broad antibacterial spectrum, and has antibacterial activity against bacteria, yeast, mold and virus, and its antibacterial concentration is low. The antibacterial mechanism is that positively charged ε-polylysine is adsorbed to the surface of negatively charged microbial cells through electrostatic interaction, destroying the cytoplasmic membrane and causing the leakage of intracellular substances, thereby inhibiting or killing microorganisms. ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P13/02C12N11/02
CPCC12N11/02C12P13/02
Inventor 刘盛荣张维瑞孙端方杨小鹃
Owner NINGDE NORMAL UNIV
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