CN101294179A - Method for preparing small-molecular weight hyaluronic acid by adding hyaluronate lyase in course of fermentation - Google Patents
Method for preparing small-molecular weight hyaluronic acid by adding hyaluronate lyase in course of fermentation Download PDFInfo
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- CN101294179A CN101294179A CNA2008101233959A CN200810123395A CN101294179A CN 101294179 A CN101294179 A CN 101294179A CN A2008101233959 A CNA2008101233959 A CN A2008101233959A CN 200810123395 A CN200810123395 A CN 200810123395A CN 101294179 A CN101294179 A CN 101294179A
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- hyaluronic acid
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- unidasa
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Abstract
The invention discloses a method for producing hyaluronic acid with small molecular weight by adding hyaluronidase in fermentation process, which belongs to the technology field of the application of enzymolysis reaction in optimization of fermentation process. In the method, hyaluronidase (0.05 to 0.25 g/L) is added in the fermentation process (8 h) to degrade hyaluronic acid, so that the viscosity of a fermentation broth decreases from 300 cp of a reference sample (without hyaluronidase) to 50 to 150 cp, the dissolved oxygen concentration is improved from 0 to 1% of the reference sample to 10 to 15%, the output of hyaluronic acid is improved from 5.0 g/L of the reference sample to 6.0 to 7.0 g/L, and the molecular weight of the hyaluronic acid decreases from 1,300 kDa of the reference sample to 20 to 100 kDa. The inventive method is simple and efficient, and has important industrial application value. The decrease of the viscosity of the fermentation broth results in the improvement of downstream purification efficiency and the reduction of production cost. The hyaluronic acid with small molecular weight produced by the method can be applied to the fields of medicines and healthy food.
Description
Technical field
A kind of Unidasa that adds is during the fermentation produced the hyaluronic method of small molecular weight, belongs to the technical field of enzyme digestion reaction in the optimizing fermentation application facet.
Background technology
Hyaluronic acid is the straight chain line style negatively charged ion mucopolysaccharide that is connected into β-1,3 and β-1,4 glycosidic link by glucuronic acid and N-acetylglucosamine, is separated from bovine vitreous body by Meyer and Palmer in 1934.Hyaluronic acid is the important component in extracellular matrix, knuckle synovia, cartilaginous tissue, the umbilical cord, and the important physical function is arranged in human body, as the chemical constitution of regulating extracellular fluid, lubricated and promotion wound healing; Regulate the activation of adhesion, growth, motion and specific cell etc.
The size of hyaluronan molecule amount and its physiological function are closely related, and for example, hyaluronic acid stimulates the effect and the inversely proportional relation of hyaluronic molecular weight of chondrocyte and lymphoblast differentiation: molecular weight is 2 * 10
5～4 * 10
5Hyaluronic acid hormesis in the Da scope is the strongest, and molecular weight is greater than 10
6Hyaluronic acid then have no stimulation; High molecular weight hyaluronic acid can suppress the phagocytic activity of scavenger cell, and low-molecular-weight hyaluronic acid can impel scavenger cell to express the factor of some and inflammation-related.Compare with the high-molecular weight hyaluronic acid, low-molecular-weight hyaluronic acid can show different biological functions: low-molecular-weight hyaluronic acid can stimulate blood vessel hyperplasia, induces the expression of inflammation regulatory factor, the propagation of inhibition tumour and anti-microbial effect etc.In addition, low-molecular-weight hyaluronic acid also is widely used aspect protective foods, in Japan and China Taiwan the listing of hyaluronic acid aesthetic health care food is arranged all, and therefore, the preparation of research low-molecular-weight hyaluronic acid has important use and is worth and medical value.
The Production by Microorganism Fermentation hyaluronic acid belongs to high viscosity fermentation, and oxyty is the bottleneck that hyaluronic acid volume of production improves.The factor that influences fermentation broth viscosity is a lot, and wherein hyaluronic molecular weight influence is very big, and molecular weight is big more, and viscosity is high more.Therefore,, then can reduce viscosity and improve dissolved oxygen, finally improve hyaluronic output if can degrade hyaluronic molecular weight during the fermentation.The present invention improves dissolved oxygen and then improves hyaluronic acid volume of production by adding hyaluronic acid enzyme liberating hyaluronic acid during the fermentation, does not see bibliographical information as yet.
Summary of the invention
The purpose of this invention is to provide a kind of strategy that adds Unidasa during the fermentation and produce the hyaluronic method of small molecular weight, utilize the present invention can produce the hyaluronic acid of molecular weight between 20～100kDa.
Technical scheme of the present invention: a kind of Unidasa that adds is during the fermentation produced the hyaluronic method of small molecular weight, add Unidasa during the fermentation, when fermentation 8h, add Unidasa 0.05～0.25g/L degraded hyaluronic acid, reduce fermentation broth viscosity, improve dissolved oxygen and hyaluronic acid volume of production, the hyaluronic molecular weight of the small molecular weight of being produced is between 20～100kDa.
Beneficial effect of the present invention: the present invention adopts (8h) interpolation Unidasa (0.05～0.25g/L) degraded hyaluronic acid during the fermentation, the viscosity of fermented liquid drops to 50～150cp by the 300cp of reference examples (not adding Unidasa), oxyty brings up to 10%～15% by 0～1% of reference examples, hyaluronic acid volume of production also improves 6.0～7.0g/L by the 5.0g/L of reference examples, hyaluronic molecular weight is by 1 of reference examples, and 300kDa drops to 20～100kDa.The inventive method is simple, efficient, has important industrial application value, because fermentation broth viscosity reduces, the purification efficiency in downstream improves, and production cost reduces.The small molecular weight hyaluronic acid that the inventive method is produced can be applied to medicine and field of health care food.In addition, the optimizing fermentation of degradable polymer for other has certain guidance and edifies meaning.
Embodiment
Comparative examples: the used bacterial classification of the present invention is streptococcus zooepidemicus Streptococcus zooepidemicusWSH-24, (or being designated as H-24) (Wen Qi, Liu Dengru, Chen Jian, stifled state become " add tensio-active agent and promote streptococcus zooepidemicus high yield hyaluronic acid ", " chemical industry progress " 2006,25 (9), 1089～1094; Chinese patent 200610098195.3 is open), changeing the inclined-plane earlier cultivated 12 hours in 37 ℃ of incubators, carried out shake flask fermentation then 14 hours, 37 ℃ of temperature, rotating speed 200rpm fermented in the 7L fermentor tank 16 hours at last, and fermentation condition is: pH 7.0, mixing speed 200rpm, Ventilation Rate 1.0vvm.Fermention medium is (g/L): sucrose: 70; Yeast powder: 25; MgSO
47H
2O:2.0; Na
2HPO
4: 6.3; K
2SO
4: 7.0.Hyaluronic molecular weight is 1,300,000, and output is 5.0g/L.
Embodiment 1: ferment tank adopts Unidasa to add strategy, the same comparative examples of all the other conditions.When fermentation 8h, add Unidasa (0.05g/L).Hyaluronic acid volume of production is brought up to 6.0g/L during fermentation ends, and molecular weight is 100kDa.
Embodiment 2: ferment tank adopts Unidasa to add strategy, the same comparative examples of all the other conditions.When fermentation 8h, add Unidasa (0.15g/L).Hyaluronic acid volume of production is brought up to 6.5g/L during fermentation ends, and molecular weight is 60kDa.
Embodiment 3: ferment tank adopts Unidasa to add strategy, the same comparative examples of all the other conditions.When fermentation 8h, add Unidasa (0.25g/L).Hyaluronic acid volume of production is brought up to 7.0g/L during fermentation ends, and molecular weight is 20kDa.
Claims (1)
1. one kind is added the hyaluronic method of Unidasa production small molecular weight during the fermentation, it is characterized in that adding during the fermentation Unidasa, when fermentation 8h, add Unidasa 0.05～0.25g/L degraded hyaluronic acid, reduce fermentation broth viscosity, improve dissolved oxygen and hyaluronic acid volume of production, the hyaluronic molecular weight of the small molecular weight of being produced is between 20～100kDa.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNA2008101233959A CN101294179A (en) | 2008-05-26 | 2008-05-26 | Method for preparing small-molecular weight hyaluronic acid by adding hyaluronate lyase in course of fermentation |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNA2008101233959A CN101294179A (en) | 2008-05-26 | 2008-05-26 | Method for preparing small-molecular weight hyaluronic acid by adding hyaluronate lyase in course of fermentation |
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| CN101294179A true CN101294179A (en) | 2008-10-29 |
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102898545A (en) * | 2012-11-15 | 2013-01-30 | 山东众山生物科技有限公司 | Method for reducing viscosity of hyaluronic acid |
| CN104120158A (en) * | 2014-07-01 | 2014-10-29 | 江南大学 | Method for improving fermentation yield of low-molecular hyaluronic acid (HA) by adding hyaluronidase |
| CN104342420A (en) * | 2013-07-30 | 2015-02-11 | 惠觅宙 | Recombinant long-acting human hyaluronidase, and encoding gene, production method and application thereof |
| CN106367459A (en) * | 2016-09-30 | 2017-02-01 | 江南大学 | Method for preparing oligomeric hyaluronic acid with different molecular weights |
| CN112553272A (en) * | 2020-12-24 | 2021-03-26 | 华熙生物科技股份有限公司 | Method for improving hyaluronic acid yield |
| CN112662718A (en) * | 2021-01-13 | 2021-04-16 | 山东众山生物科技有限公司 | Preparation method of oligomeric sodium hyaluronate |
| CN117535363A (en) * | 2023-12-27 | 2024-02-09 | 常州加源医药科技研发有限公司 | Production method of functional hyaluronic acid |
-
2008
- 2008-05-26 CN CNA2008101233959A patent/CN101294179A/en active Pending
Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102898545A (en) * | 2012-11-15 | 2013-01-30 | 山东众山生物科技有限公司 | Method for reducing viscosity of hyaluronic acid |
| CN102898545B (en) * | 2012-11-15 | 2015-06-10 | 山东众山生物科技有限公司 | Method for reducing viscosity of hyaluronic acid |
| CN104342420A (en) * | 2013-07-30 | 2015-02-11 | 惠觅宙 | Recombinant long-acting human hyaluronidase, and encoding gene, production method and application thereof |
| CN104120158A (en) * | 2014-07-01 | 2014-10-29 | 江南大学 | Method for improving fermentation yield of low-molecular hyaluronic acid (HA) by adding hyaluronidase |
| CN106367459A (en) * | 2016-09-30 | 2017-02-01 | 江南大学 | Method for preparing oligomeric hyaluronic acid with different molecular weights |
| CN112553272A (en) * | 2020-12-24 | 2021-03-26 | 华熙生物科技股份有限公司 | Method for improving hyaluronic acid yield |
| CN112553272B (en) * | 2020-12-24 | 2022-08-09 | 华熙生物科技股份有限公司 | Method for improving hyaluronic acid yield |
| CN112662718A (en) * | 2021-01-13 | 2021-04-16 | 山东众山生物科技有限公司 | Preparation method of oligomeric sodium hyaluronate |
| CN117535363A (en) * | 2023-12-27 | 2024-02-09 | 常州加源医药科技研发有限公司 | Production method of functional hyaluronic acid |
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Open date: 20081029 |