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Method for producing hyaluronic acid

A technology of hyaluronic acid and conditions, applied in the field of hyaluronic acid production, can solve the problems of unfavorable bacterial growth or product accumulation at constant pH, unfavorable hyaluronic acid synthesis, unfavorable production amplification and the like

Pending Publication Date: 2022-02-01
HEC PHARM
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  • Application Information

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Problems solved by technology

However, the optimal pH for the growth of hyaluronic acid cells is often not the optimal pH value for product synthesis, and constant pH is not conducive to cell growth or product accumulation
At present, there are also patent reports on technical solutions for pH change regulation, such as patent CN 1978658 A, which discloses a method for increasing the yield of hyaluronic acid produced by fermentation by an intermittent high pH co-forced strategy. The yield of hyaluronic acid can be increased from 5.0g / L to 6.7g / L, however, the cumbersome process control of this technical scheme is not conducive to production scale-up, and maintaining a high pH for a long time in this technical scheme will affect the viability of the bacteria, which will affect the growth of the bacteria to a large extent, which is not conducive to the synthesis of hyaluronic acid

Method used

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  • Method for producing hyaluronic acid
  • Method for producing hyaluronic acid
  • Method for producing hyaluronic acid

Examples

Experimental program
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Effect test

Embodiment 1

[0034] Take out the glycerin tube strain from the ultra-low temperature refrigerator, melt it in a 37°C water bath or in the palm of your hand, inoculate it into the seed medium at a volume ratio of 1%, and cultivate it at 200-220 rpm at 36.5°C for 17.5-18 hours. Before the end of the culture, perform a Gram staining microscope. After there is no abnormality, inoculate a 15L fermenter with a ventilation rate of 0.5-1vvm, stirring at 100-600rpm, and ferment at 37°C for 30h.

[0035] The fermenter adopts the following pH control strategy:

[0036] Early stage of fermentation (0-6 hours): After inoculation, the pH of the fermentation broth naturally drops to 6.7, and the pH of the fermentation broth is maintained at 6.7 by 20-30% NaOH alkali control. In the middle stage of fermentation (6-12h), the pH is increased by 0.1 every 1h from 6h until the pH rises to 7.4, and the pH is constantly controlled to 7.4. In the middle and late stages of fermentation (12 to 30 hours), the pH o...

Embodiment 2

[0038] The fermenter adopts the following pH control strategy:

[0039] Early stage of fermentation (0-6h): After inoculation, the pH of the fermentation broth naturally drops to 7.2, and the pH of the fermentation broth is maintained at 7.2 by alkali control. In the middle stage of fermentation (6-12h), the pH is increased by 0.2 every 1h from 6h until the pH rises to 7.6, and the pH is constantly controlled to 7.6. In the middle and late stages of fermentation (12-30h), by changing the parameters of the alkali control and automatic control program, the constant rate of the peristaltic pump and the opening time of the solenoid valve, the pH of the fermentation process is kept in a state of continuous fluctuation. Finish. pH control curve such as figure 1 shown. The results showed that the bacterial cell concentration (OD660) of the 12h intermediate sample was 5.42, and the output of hyaluronic acid in the tank was 8.52g / L.

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Abstract

The invention provides a method for producing hyaluronic acid. The method comprises the following steps that streptococcus zooepidemicus is subjected to fermentation treatment so as to obtain hyaluronic acid, and the fermentation treatment is carried out under the following conditions: within 0-6 hours after the fermentation treatment is started, the fermentation treatment is carried out under the condition that the pH value is 6.7-7.2; the fermentation treatment is carried out under the condition that the pH is 7.4 to 7.6 within 6 to 12 hours after the fermentation treatment is started; and the fermentation treatment is carried out under the condition that the pH value is 6.5-9.0 after 12 hours after the fermentation treatment is started. Compared with the prior art, the method has the advantage that the yield of hyaluronic acid is obviously increased.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular, the invention relates to a method for producing hyaluronic acid. Background technique [0002] Hyaluronic acid (hyaluronic acid or hyaluronan, HA) is a polysaccharide composed of disaccharide repeating units of glucuronic acid and glucosamine, widely distributed in the dermis and epidermis of cartilage tissue, joint fluid, and skin tissue, and in which It plays physiological functions such as moisturizing, nourishing, repairing and preventing damage. The production methods of hyaluronic acid include animal tissue extraction method and fermentation method. Due to the high preparation cost and complicated separation and purification of animal tissue extraction method, it is gradually replaced by fermentation method. [0003] Fermentation yield is the key to cost control of hyaluronic acid produced by fermentation. During the production of hyaluronic acid by microbial fermentation, pH i...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P19/26C12N1/20C12R1/46
CPCC12P19/26C12N1/20
Inventor 石江水肖文龙马雷磊封海生李峰谢文平
Owner HEC PHARM
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