CN114150030A - Production method of hyaluronic acid - Google Patents
Production method of hyaluronic acid Download PDFInfo
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- CN114150030A CN114150030A CN202111609573.0A CN202111609573A CN114150030A CN 114150030 A CN114150030 A CN 114150030A CN 202111609573 A CN202111609573 A CN 202111609573A CN 114150030 A CN114150030 A CN 114150030A
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- KIUKXJAPPMFGSW-DNGZLQJQSA-N (2S,3S,4S,5R,6R)-6-[(2S,3R,4R,5S,6R)-3-Acetamido-2-[(2S,3S,4R,5R,6R)-6-[(2R,3R,4R,5S,6R)-3-acetamido-2,5-dihydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-2-carboxy-4,5-dihydroxyoxan-3-yl]oxy-5-hydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-3,4,5-trihydroxyoxane-2-carboxylic acid Chemical compound CC(=O)N[C@H]1[C@H](O)O[C@H](CO)[C@@H](O)[C@@H]1O[C@H]1[C@H](O)[C@@H](O)[C@H](O[C@H]2[C@@H]([C@@H](O[C@H]3[C@@H]([C@@H](O)[C@H](O)[C@H](O3)C(O)=O)O)[C@H](O)[C@@H](CO)O2)NC(C)=O)[C@@H](C(O)=O)O1 KIUKXJAPPMFGSW-DNGZLQJQSA-N 0.000 title claims abstract description 116
- 229920002674 hyaluronan Polymers 0.000 title claims abstract description 116
- 229960003160 hyaluronic acid Drugs 0.000 title claims abstract description 116
- 238000004519 manufacturing process Methods 0.000 title claims abstract description 37
- 238000000855 fermentation Methods 0.000 claims abstract description 271
- 230000004151 fermentation Effects 0.000 claims abstract description 270
- 238000000034 method Methods 0.000 claims abstract description 19
- ROWKJAVDOGWPAT-UHFFFAOYSA-N Acetoin Chemical compound CC(O)C(C)=O ROWKJAVDOGWPAT-UHFFFAOYSA-N 0.000 claims description 58
- 239000001963 growth medium Substances 0.000 claims description 47
- 239000007788 liquid Substances 0.000 claims description 40
- 238000011218 seed culture Methods 0.000 claims description 34
- GFAZHVHNLUBROE-UHFFFAOYSA-N hydroxymethyl propionaldehyde Natural products CCC(=O)CO GFAZHVHNLUBROE-UHFFFAOYSA-N 0.000 claims description 29
- 238000012258 culturing Methods 0.000 claims description 27
- 230000036284 oxygen consumption Effects 0.000 claims description 25
- 239000002609 medium Substances 0.000 claims description 23
- 238000005273 aeration Methods 0.000 claims description 19
- 239000001888 Peptone Substances 0.000 claims description 18
- 108010080698 Peptones Proteins 0.000 claims description 18
- 235000019319 peptone Nutrition 0.000 claims description 18
- 238000011081 inoculation Methods 0.000 claims description 17
- 235000015278 beef Nutrition 0.000 claims description 14
- 239000000284 extract Substances 0.000 claims description 14
- 230000003213 activating effect Effects 0.000 claims description 12
- 238000012807 shake-flask culturing Methods 0.000 claims description 12
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 claims description 8
- 241000120569 Streptococcus equi subsp. zooepidemicus Species 0.000 claims description 8
- 230000008569 process Effects 0.000 claims description 8
- 229920001817 Agar Polymers 0.000 claims description 4
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 claims description 4
- 239000008272 agar Substances 0.000 claims description 4
- 229940041514 candida albicans extract Drugs 0.000 claims description 4
- 239000008103 glucose Substances 0.000 claims description 4
- 229910052943 magnesium sulfate Inorganic materials 0.000 claims description 4
- 235000019341 magnesium sulphate Nutrition 0.000 claims description 4
- 229910000402 monopotassium phosphate Inorganic materials 0.000 claims description 4
- 235000019796 monopotassium phosphate Nutrition 0.000 claims description 4
- GNSKLFRGEWLPPA-UHFFFAOYSA-M potassium dihydrogen phosphate Chemical compound [K+].OP(O)([O-])=O GNSKLFRGEWLPPA-UHFFFAOYSA-M 0.000 claims description 4
- LWIHDJKSTIGBAC-UHFFFAOYSA-K potassium phosphate Substances [K+].[K+].[K+].[O-]P([O-])([O-])=O LWIHDJKSTIGBAC-UHFFFAOYSA-K 0.000 claims description 4
- 239000012138 yeast extract Substances 0.000 claims description 4
- PXEDJBXQKAGXNJ-QTNFYWBSSA-L disodium L-glutamate Chemical compound [Na+].[Na+].[O-]C(=O)[C@@H](N)CCC([O-])=O PXEDJBXQKAGXNJ-QTNFYWBSSA-L 0.000 claims description 3
- 239000006260 foam Substances 0.000 claims description 3
- 235000013923 monosodium glutamate Nutrition 0.000 claims description 3
- 229940073490 sodium glutamate Drugs 0.000 claims description 3
- 238000007747 plating Methods 0.000 claims 1
- 230000015572 biosynthetic process Effects 0.000 abstract description 3
- 238000003786 synthesis reaction Methods 0.000 abstract description 3
- 239000000243 solution Substances 0.000 description 41
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 33
- 239000000203 mixture Substances 0.000 description 9
- 230000000052 comparative effect Effects 0.000 description 6
- 238000000605 extraction Methods 0.000 description 4
- 230000000813 microbial effect Effects 0.000 description 4
- UJOBWOGCFQCDNV-UHFFFAOYSA-N 9H-carbazole Chemical compound C1=CC=C2C3=CC=CC=C3NC2=C1 UJOBWOGCFQCDNV-UHFFFAOYSA-N 0.000 description 2
- AEMOLEFTQBMNLQ-AQKNRBDQSA-N D-glucopyranuronic acid Chemical compound OC1O[C@H](C(O)=O)[C@@H](O)[C@H](O)[C@H]1O AEMOLEFTQBMNLQ-AQKNRBDQSA-N 0.000 description 2
- IAJILQKETJEXLJ-UHFFFAOYSA-N Galacturonsaeure Natural products O=CC(O)C(O)C(O)C(O)C(O)=O IAJILQKETJEXLJ-UHFFFAOYSA-N 0.000 description 2
- 239000007789 gas Substances 0.000 description 2
- 229940097043 glucuronic acid Drugs 0.000 description 2
- 239000002054 inoculum Substances 0.000 description 2
- 238000001819 mass spectrum Methods 0.000 description 2
- 241001522301 Apogonichthyoides nigripinnis Species 0.000 description 1
- 241000561734 Celosia cristata Species 0.000 description 1
- 229920002683 Glycosaminoglycan Polymers 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- OVRNDRQMDRJTHS-UHFFFAOYSA-N N-acelyl-D-glucosamine Natural products CC(=O)NC1C(O)OC(CO)C(O)C1O OVRNDRQMDRJTHS-UHFFFAOYSA-N 0.000 description 1
- OVRNDRQMDRJTHS-FMDGEEDCSA-N N-acetyl-beta-D-glucosamine Chemical compound CC(=O)N[C@H]1[C@H](O)O[C@H](CO)[C@@H](O)[C@@H]1O OVRNDRQMDRJTHS-FMDGEEDCSA-N 0.000 description 1
- MBLBDJOUHNCFQT-LXGUWJNJSA-N N-acetylglucosamine Natural products CC(=O)N[C@@H](C=O)[C@@H](O)[C@H](O)[C@H](O)CO MBLBDJOUHNCFQT-LXGUWJNJSA-N 0.000 description 1
- KWYUFKZDYYNOTN-UHFFFAOYSA-M Potassium hydroxide Chemical compound [OH-].[K+] KWYUFKZDYYNOTN-UHFFFAOYSA-M 0.000 description 1
- KEAYESYHFKHZAL-UHFFFAOYSA-N Sodium Chemical compound [Na] KEAYESYHFKHZAL-UHFFFAOYSA-N 0.000 description 1
- 239000004115 Sodium Silicate Substances 0.000 description 1
- 241001052560 Thallis Species 0.000 description 1
- 230000002378 acidificating effect Effects 0.000 description 1
- 230000004075 alteration Effects 0.000 description 1
- 239000002775 capsule Substances 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 210000001520 comb Anatomy 0.000 description 1
- 210000002808 connective tissue Anatomy 0.000 description 1
- 239000002537 cosmetic Substances 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 125000000600 disaccharide group Chemical group 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 238000005461 lubrication Methods 0.000 description 1
- 230000014759 maintenance of location Effects 0.000 description 1
- 244000005700 microbiome Species 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 229950006780 n-acetylglucosamine Drugs 0.000 description 1
- NTHWMYGWWRZVTN-UHFFFAOYSA-N sodium silicate Chemical compound [Na+].[Na+].[O-][Si]([O-])=O NTHWMYGWWRZVTN-UHFFFAOYSA-N 0.000 description 1
- 229910052911 sodium silicate Inorganic materials 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 239000011550 stock solution Substances 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 210000003954 umbilical cord Anatomy 0.000 description 1
- 210000004127 vitreous body Anatomy 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P19/00—Preparation of compounds containing saccharide radicals
- C12P19/26—Preparation of nitrogen-containing carbohydrates
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/38—Chemical stimulation of growth or activity by addition of chemical compounds which are not essential growth factors; Stimulation of growth by removal of a chemical compound
Abstract
The invention belongs to the field of fermentation production of hyaluronic acid, and relates to a production method of hyaluronic acid. The hyaluronic acid prepared by the method provided by the invention can promote the synthesis of hyaluronic acid, and the yield of hyaluronic acid in the obtained fermentation product is obviously improved.
Description
Technical Field
The invention belongs to the field of fermentation production of hyaluronic acid, and particularly relates to a production method of hyaluronic acid.
Background
Hyaluronic Acid (HA) is a linear acidic mucopolysaccharide composed of repeating tandem connection of N-acetylglucosamine (GlcNac) and glucuronic acid (GlcA) as disaccharide units, and is widely present in connective tissues and microbial capsules of higher animals. Hyaluronic acid has excellent moisture retention, viscoelasticity and lubrication, can be widely applied to a plurality of fields such as medicines, cosmetics and cosmetology, and has extremely high commercial value.
The hyaluronic acid can be prepared by conventional extraction method and microorganism fermentation method. In the traditional extraction method, hyaluronic acid is extracted from cockscomb, human umbilical cord and bulls-eye vitreous body, and the method has low extraction rate and high cost. At present, the production of hyaluronic acid is mainly carried out by a microbial fermentation method, and although the microbial fermentation method can overcome the defects of the traditional extraction method, the production of hyaluronic acid by the microbial fermentation method still has the problem that the yield is required to be improved. Therefore, it is required to develop a new fermentation process of hyaluronic acid to increase the fermentation yield of hyaluronic acid.
Disclosure of Invention
The invention aims to provide a production method capable of improving the yield of hyaluronic acid.
After intensive and extensive research, the inventor of the invention finds that in the fermentation process of hyaluronic acid, when the oxygen consumption rate in the hyaluronic acid fermentation liquid reaches 12-20 mmol/L.h for the first time, acetoin is supplemented to promote the synthesis of hyaluronic acid, and the content of hyaluronic acid in the obtained fermentation product can be obviously improved. Based on this, the present invention has been completed.
In order to achieve the purpose, the invention adopts the following technical scheme: a production method of hyaluronic acid comprises the step of adding acetoin into hyaluronic acid fermentation liquor when the oxygen consumption rate in the hyaluronic acid fermentation liquor reaches 12-20 mmol/L.h for the first time in the fermentation culture process.
In a preferred embodiment, during the fermentation culture process, when the oxygen consumption rate in the hyaluronic acid fermentation liquor reaches 14-18 mmol/L.h for the first time, adding acetoin into the hyaluronic acid fermentation liquor.
In a preferred embodiment, the acetoin is added in a proportion of 0.1-0.5 wt% of the hyaluronic acid fermentation liquor.
In a preferred embodiment, the hyaluronic acid-producing strain is Streptococcus zooepidemicus (Streptococcus zooepidemicus).
In a preferred embodiment, the fermentation is terminated when the residual sugar content of the fermentation broth is < 5 g/L.
In a preferred embodiment, the method for producing hyaluronic acid comprises the steps of:
(1) activating strains: under the aseptic condition, streaking a hyaluronic acid production strain in a plate culture medium, and culturing at the temperature of 32-38 ℃ for 18-24 h;
(2) seed culture: picking single colonies from a cultured plate culture medium, inoculating the single colonies into a shake flask culture medium, then placing the shake flask culture medium into a shaking table, and culturing for 8-18 h under the conditions of the temperature of 32-38 ℃ and the rotating speed of 160-220 rpm to obtain a seed culture solution; optionally, inoculating the obtained seed culture solution into a seed tank filled with a shake flask culture medium in an inoculation amount of 0.5-5%, and culturing for 5-10 h at a temperature of 32-38 ℃, an aeration ratio of 0.3-1 VVM, a rotation speed of 150-300 rpm and a tank pressure of 0.03-0.05 MPa to obtain a seed solution;
(3) fermentation culture: inoculating the cultured seed culture solution or the seed solution into a fermentation tank filled with a fermentation culture medium by 1-10% of inoculation amount, carrying out fermentation culture at the temperature of 32-38 ℃, the aeration ratio of 0.5-1.5 VVM, the rotation speed of 80-300 rpm, the tank pressure of 0.04-0.06 MPa and the pH of 6.5-8.0, adding acetoin into the fermentation liquid when the oxygen consumption rate in the fermentation liquid reaches a preset value for the first time, continuing the culture, and ending the fermentation when the content of residual sugar in the fermentation liquid is less than or equal to 5 g/L.
In a preferred embodiment, in step (1), the plate medium is a beef extract peptone agar medium.
In a preferred embodiment, in step (2), the shake flask culture medium is a beef extract peptone medium.
In a preferred embodiment, in the step (3), the pH value of the fermentation medium is 6.5-7.5, and each liter of the fermentation medium contains the following components: 60-80 g of glucose, 5-15 g of peptone, 2-6 g of yeast extract, 0.5-2 g of sodium glutamate, 0.1-1 g of monopotassium phosphate, 0.1-1 g of magnesium sulfate and 0.1-1 g of foam killer.
In a preferred embodiment, the fermentation tank used for the fermentation culture has a volume of 30L-100 m3。
The hyaluronic acid prepared by the method provided by the invention can obviously improve the content of the hyaluronic acid in the obtained fermentation product.
Detailed Description
In the fermentation culture process of hyaluronic acid, when the oxygen consumption rate (OUR) in the hyaluronic acid fermentation liquor reaches 12-20 mmol/L.h (such as 12, 13, 14, 15, 16, 17, 18, 19 and 20 mmol/L.h) for the first time, acetoin is added into the hyaluronic acid fermentation liquor. Preferably, in the fermentation culture process, when the oxygen consumption rate in the hyaluronic acid fermentation liquor reaches 14-18 mmol/L.h for the first time, acetoin is added into the hyaluronic acid fermentation liquor. In the invention, the oxygen consumption rate (OUR) is measured by detecting the fermentation tail gas by a mass spectrum analyzer. According to the invention, acetoin is added when OUR reaches 12-20 mmol/L.h for the first time, preferably 14-18 mmol/L.h in the fermentation process, so that the synthesis of hyaluronic acid can be promoted, and the content of hyaluronic acid in the fermentation product can be increased. In addition, the addition ratio of the acetoin is preferably 0.1-0.5 wt% of the hyaluronic acid fermentation broth, such as 0.1 wt%, 0.2 wt%, 0.3 wt%, 0.4 wt%, 0.5 wt% and the like.
The main improvement of the invention is that acetoin is added in the fermentation culture process, and the specific species and obtaining mode of the hyaluronic acid producing strain, culture conditions of each stage and the like can be selected conventionally in the field, for example, the hyaluronic acid producing strain can be Streptococcus zooepidemicus (Streptococcus zooepidemicus).
In the present invention, fermentation is generally terminated when the residual sugar content in the fermentation broth is less than or equal to 5 g/L.
In some embodiments of the invention, the method for producing hyaluronic acid comprises the steps of:
(1) activating strains: under the aseptic condition, streaking a hyaluronic acid production strain in a plate culture medium, and culturing at the temperature of 32-38 ℃ for 18-24 h;
(2) seed culture: picking single colonies from a cultured plate culture medium, inoculating the single colonies into a shake flask culture medium, then placing the shake flask culture medium into a shaking table, and culturing for 8-18 h under the conditions of the temperature of 32-38 ℃ and the rotating speed of 160-220 rpm to obtain a seed culture solution; optionally, inoculating the obtained seed culture solution into a seed tank filled with a shake flask culture medium in an inoculation amount of 0.5-5%, and culturing for 5-10 h at a temperature of 32-38 ℃, an aeration ratio of 0.3-1 VVM, a rotation speed of 150-300 rpm and a tank pressure of 0.03-0.05 MPa to obtain a seed solution;
(3) fermentation culture: inoculating the cultured seed culture solution or the seed solution into a fermentation tank filled with a fermentation culture medium by 1-10% of inoculation amount, carrying out fermentation culture at the temperature of 32-38 ℃, the aeration ratio of 0.5-1.5 VVM, the rotation speed of 80-300 rpm, the tank pressure of 0.04-0.06 MPa and the pH of 6.5-8.0, adding acetoin into the fermentation liquid when the oxygen consumption rate in the fermentation liquid reaches a preset value for the first time, continuing the culture, and ending the fermentation when the content of residual sugar in the fermentation liquid is less than or equal to 5 g/L.
The invention has no special limitation on the types of plate culture medium, shake flask culture medium and fermentation culture medium, and can be various existing culture media suitable for growth of thalli at various stages. For example, the plate medium and the shake flask medium may each independently be a beef extract peptone agar medium. In a preferred embodiment, the pH value of the fermentation medium is 6.5-7.5, and each liter of the fermentation medium contains the following components: 60-80 g glucose, 5-15 g peptone, 2-6 g yeast extract, 0.5E monosodium glutamate2g, 0.1-1 g of monopotassium phosphate, 0.1-1 g of magnesium sulfate and 0.1-1 g of foam killer. In addition, the agent for adjusting the pH during the fermentation culture may be specifically selected from a sodium hydroxide solution or a potassium hydroxide solution. In addition, the volume of a fermentation tank used for the fermentation culture can be 30L-100 m3。
The following detailed description of embodiments of the invention is intended to be illustrative of the invention and is not to be construed as limiting the invention. The examples do not specify particular techniques or conditions, and are performed according to the techniques or conditions described in the literature in the art or according to the product specifications. The reagents or instruments used are not indicated by the manufacturer, and are all conventional products commercially available.
The production process of fermented hyaluronic acid is further illustrated in the following examples and comparative examples using Streptococcus zooepidemicus (strain No.: ATCC39920) as a production strain.
The relevant equipment referred to in the following examples and comparative examples comprises: 50L fermenter, 1m3Fermentation tank, 15m3Fermenter and 100m3And (4) fermenting the mixture.
In the following examples and comparative examples:
(1) oxygen consumption rate (OUR) is obtained by detecting fermentation tail gas by a mass spectrum analyzer;
(2) the content of hyaluronic acid is measured by adopting a carbazole method of Bitter-Muir;
(3) the content of residual sugar in the fermentation liquor is measured by a Sielman biosensing analyzer.
Example 1: a production method of hyaluronic acid, wherein fermentation is carried out by using a 50L fermentation tank, comprises the following steps:
(1) activating strains: under aseptic conditions, streaking the streptococcus zooepidemicus seed frozen stock solution into a beef extract peptone agar culture medium, and culturing at 35 ℃ for 22 h;
(2) seed culture: picking single colony from a cultured plate culture medium, inoculating the single colony into a 2L shake flask filled with a beef extract peptone culture medium, then placing the shake flask into a shaking table, and culturing for 12h at the temperature of 35 ℃ and the rotation speed of 200rpm to obtain a seed culture solution;
(3) fermentation culture: inoculating the cultured seed culture solution into a fermentation tank filled with a fermentation culture medium by 5 percent of inoculation amount for fermentation culture, wherein the fermentation culture temperature is 35 ℃, the pH value is controlled to be 6.6-7.6 by using a sodium hydroxide solution in the whole fermentation process, the OUR value of the fermentation liquid is monitored in the whole fermentation process, the aeration ratio is controlled to be 1.0-1.5 VVM in the fermentation process, the rotating speed is controlled to be 200-300 rpm, and the tank pressure is controlled to be 0.04-0.05 MPa. When the oxygen consumption rate (OUR) in the hyaluronic acid fermentation liquor is up to 12 mmol/L.h for the first time, adding 0.3 wt% of acetoin into the hyaluronic acid fermentation liquor, continuously culturing until the residual sugar is less than or equal to 5g/L, and placing in a tank. Wherein, each liter of fermentation medium contains the following components: 70g of glucose, 10g of peptone, 5g of yeast extract, 1g of sodium glutamate, 0.5g of monopotassium phosphate, 0.5g of magnesium sulfate and 0.5g of sodium silicate, and adjusting the pH value to 7.0 by using a sodium hydroxide solution. When the fermentation culture period reaches 20 hours, the residual sugar in the fermentation liquid is less than or equal to 5g/L, the fermentation is stopped, and the content of the hyaluronic acid in the fermentation liquid is 12.8 g/L.
Example 2: a production method of hyaluronic acid, wherein fermentation is carried out by using a 50L fermentation tank, comprises the following steps:
(1) activating strains: the same as example 1;
(2) seed culture: the same as example 1;
(3) fermentation culture: inoculating the cultured seed culture solution into a fermentation tank filled with a fermentation culture medium by 5 percent of inoculation amount for fermentation culture, wherein the fermentation culture temperature is 35 ℃, the pH value is controlled to be 6.6-7.6 by using a sodium hydroxide solution in the whole fermentation process, the OUR value of the fermentation liquid is monitored in the whole fermentation process, the aeration ratio is controlled to be 1.0-1.5 VVM in the fermentation process, the rotating speed is controlled to be 200-300 rpm, and the tank pressure is controlled to be 0.04-0.05 MPa. When the oxygen consumption rate (OUR) in the hyaluronic acid fermentation liquor is first 14 mmol/L.h, adding 0.3 wt% of acetoin into the hyaluronic acid fermentation liquor, and continuously culturing until the residual sugar is less than or equal to 5 g/L. The composition per liter of fermentation medium is the same as in example 1. When the fermentation culture period reaches 20 hours, the residual sugar in the fermentation liquid is less than or equal to 5g/L, the fermentation is stopped, and the content of the hyaluronic acid in the fermentation liquid is 13.0 g/L.
Example 3: a production method of hyaluronic acid, wherein fermentation is carried out by using a 50L fermentation tank, comprises the following steps:
(1) activating strains: the same as example 1;
(2) seed culture: the same as example 1;
(3) fermentation culture: inoculating the cultured seed culture solution into a fermentation tank filled with a fermentation culture medium by 5 percent of inoculation amount for fermentation culture, wherein the fermentation culture temperature is 35 ℃, the pH value is controlled to be 6.6-7.6 by using a sodium hydroxide solution in the whole fermentation process, the OUR value of the fermentation liquid is monitored in the whole fermentation process, the aeration ratio is controlled to be 1.0-1.5 VVM in the fermentation process, the rotating speed is controlled to be 200-300 rpm, and the tank pressure is controlled to be 0.04-0.05 MPa. When the oxygen consumption rate (OUR) in the hyaluronic acid fermentation liquor is firstly 18 mmol/L.h, adding 0.3 wt% of acetoin into the hyaluronic acid fermentation liquor, and continuously culturing until the residual sugar is less than or equal to 5 g/L. The composition per liter of fermentation medium is the same as in example 1. When the fermentation culture period reaches 20 hours, the residual sugar in the fermentation liquid is less than or equal to 5g/L, the fermentation is stopped, and the content of the hyaluronic acid in the fermentation liquid is 12.7 g/L.
Example 4: a production method of hyaluronic acid, wherein fermentation is carried out by using a 50L fermentation tank, comprises the following steps:
(1) activating strains: the same as example 1;
(2) seed culture: the same as example 1;
(3) fermentation culture: inoculating the cultured seed culture solution into a fermentation tank filled with a fermentation culture medium by 5 percent of inoculation amount for fermentation culture, wherein the fermentation culture temperature is 35 ℃, the pH value is controlled to be 6.6-7.6 by using a sodium hydroxide solution in the whole fermentation process, the OUR value of the fermentation liquid is monitored in the whole fermentation process, the aeration ratio is controlled to be 1.0-1.5 VVM in the fermentation process, the rotating speed is controlled to be 200-300 rpm, and the tank pressure is controlled to be 0.04-0.05 MPa. When the oxygen consumption rate (OUR) in the hyaluronic acid fermentation liquor is firstly 20 mmol/L.h, adding 0.3 wt% of acetoin into the hyaluronic acid fermentation liquor, and continuously culturing until the residual sugar is less than or equal to 5 g/L. The composition per liter of fermentation medium was the same as in example 1. When the fermentation culture period reaches 20 hours, the residual sugar in the fermentation liquid is less than or equal to 5g/L, the fermentation is stopped, and the content of the hyaluronic acid in the fermentation liquid is 12.3 g/L.
Example 5: a production method of hyaluronic acid, wherein fermentation is carried out by using a 50L fermentation tank, comprises the following steps:
(1) activating strains: the same as example 1;
(2) seed culture: the same as example 1;
(3) fermentation culture: inoculating the cultured seed culture solution into a fermentation tank filled with a fermentation culture medium by 5 percent of inoculation amount for fermentation culture, wherein the fermentation culture temperature is 35 ℃, the pH value is controlled to be 6.6-7.6 by using a sodium hydroxide solution in the whole fermentation process, the OUR value of the fermentation liquid is monitored in the whole fermentation process, the aeration ratio is controlled to be 1.0-1.5 VVM in the fermentation process, the rotating speed is controlled to be 200-300 rpm, and the tank pressure is controlled to be 0.04-0.05 MPa. When the oxygen consumption rate (OUR) in the hyaluronic acid fermentation liquor is first 16 mmol/L.h, adding 0.1 wt% of acetoin into the hyaluronic acid fermentation liquor, and continuously culturing until the residual sugar is less than or equal to 5 g/L. The composition per liter of fermentation medium is the same as in example 1. When the fermentation culture period reaches 20 hours, the residual sugar in the fermentation liquid is less than or equal to 5g/L, the fermentation is stopped, and the content of the hyaluronic acid in the fermentation liquid is 12.9 g/L.
Example 6: a production method of hyaluronic acid, wherein fermentation is carried out by using a 50L fermentation tank, comprises the following steps:
(1) activating strains: the same as example 1;
(2) seed culture: the same as example 1;
(3) fermentation culture: inoculating the cultured seed culture solution into a fermentation tank filled with a fermentation culture medium by 5 percent of inoculation amount for fermentation culture, wherein the fermentation culture temperature is 35 ℃, the pH value is controlled to be 6.6-7.6 by using a sodium hydroxide solution in the whole fermentation process, the OUR value of the fermentation liquid is monitored in the whole fermentation process, the aeration ratio is controlled to be 1.0-1.5 VVM in the fermentation process, the rotating speed is controlled to be 200-300 rpm, and the tank pressure is controlled to be 0.04-0.05 MPa. When the oxygen consumption rate (OUR) in the hyaluronic acid fermentation liquor is first 16 mmol/L.h, adding 0.5 wt% of acetoin into the hyaluronic acid fermentation liquor, and continuously culturing until the residual sugar is less than or equal to 5 g/L. The composition per liter of fermentation medium is the same as in example 1. When the fermentation culture period reaches 20 hours, the residual sugar in the fermentation liquid is less than or equal to 5g/L, the fermentation is stopped, and the content of the hyaluronic acid in the fermentation liquid is 13.7 g/L.
Example 7: a process for the production of hyaluronic acid,1m for fermentation3A fermenter comprising the steps of:
(1) activating strains: the same as example 1;
(2) seed culture: picking single colony from the cultured plate culture medium, inoculating into a 500ml shake flask filled with beef extract peptone culture medium, then placing into a shaking table, and culturing for 12h at 35 ℃ and 200rpm to obtain seed culture solution. Inoculating the seed culture solution into a 50L seed tank filled with beef extract peptone culture medium at an inoculation amount of 0.5%, and culturing for 6h under the conditions of 35 ℃, 200rpm of rotation speed, 0.05MPa of tank pressure and 0.8VVM of aeration ratio to obtain an expanded culture seed solution;
(3) fermentation culture: inoculating the cultured seed solution into a fermentation tank filled with a fermentation culture medium by 5 percent of inoculation amount for fermentation culture, wherein the fermentation culture temperature is 35 ℃, the pH value is controlled to be 6.6-7.6 by using a sodium hydroxide solution in the whole fermentation process, the OUR value of the fermentation liquid is monitored in the whole fermentation process, the aeration ratio is controlled to be 1.0-1.5 VVM in the fermentation process, the rotating speed is controlled to be 150-200 rpm, and the tank pressure is controlled to be 0.04-0.05 MPa. When the oxygen consumption rate (OUR) in the hyaluronic acid fermentation liquor is first 16 mmol/L.h, adding 0.3 wt% of acetoin into the hyaluronic acid fermentation liquor, and continuously culturing until the residual sugar is less than or equal to 5 g/L. The composition per liter of fermentation medium is the same as in example 1. When the fermentation culture period reaches 20 hours, the residual sugar in the fermentation liquid is less than or equal to 5g/L, the fermentation is stopped, and the content of the hyaluronic acid in the fermentation liquid is 14.3 g/L.
Example 8: a method for producing hyaluronic acid by fermenting 15m3A fermenter comprising the steps of:
(1) activating strains: the same as example 1;
(2) seed culture: picking single colony from the cultured plate culture medium, inoculating the single colony into a 2000ml shake flask filled with beef extract peptone culture medium, then placing the shake flask into a shaking table, and culturing for 12h at the temperature of 35 ℃ and the rotation speed of 200rpm to obtain a seed culture solution. Inoculating the seed culture solution into 1m with an inoculation amount of 0.5 percent3Culturing in a seed tank containing beef extract peptone culture medium at 35 deg.C, rotation speed of 200rpm, tank pressure of 0.05MPa and aeration ratio of 0.6VVM for 6h to obtain expanded culture seed solution;
(3) fermentation culture: inoculating the cultured seed solution into a fermentation tank filled with a fermentation culture medium by 5 percent of inoculation amount for fermentation culture, wherein the fermentation culture temperature is 35 ℃, the pH value is controlled to be 6.6-7.6 by using a sodium hydroxide solution in the whole fermentation process, the OUR value of the fermentation liquid is monitored in the whole fermentation process, the aeration ratio is controlled to be 0.8-1.2 VVM in the fermentation process, the rotating speed is controlled to be 100-150 rpm, and the tank pressure is controlled to be 0.04-0.05 MPa. When the oxygen consumption rate (OUR) in the hyaluronic acid fermentation liquor is first 16 mmol/L.h, adding 0.3 wt% of acetoin into the hyaluronic acid fermentation liquor, and continuously culturing until the residual sugar is less than or equal to 5 g/L. The composition per liter of fermentation medium is the same as in example 1. When the fermentation culture period reaches 20 hours, the residual sugar in the fermentation liquid is less than or equal to 5g/L, the fermentation is stopped, and the content of the hyaluronic acid in the fermentation liquid is 14.0 g/L.
Example 9: a method for producing hyaluronic acid by fermenting with 100m3A fermenter comprising the steps of:
(1) activating strains: the same as example 1;
(2) seed culture: picking single colony from the cultured plate culture medium, inoculating the single colony into a 2000ml shake flask filled with beef extract peptone culture medium, then placing the shake flask into a shaking table, and culturing for 12h at the temperature of 35 ℃ and the rotation speed of 200rpm to obtain a seed culture solution. Inoculating the seed culture solution into a 500L seed tank containing beef extract peptone culture medium at an inoculum size of 0.5%, culturing at 35 deg.C, rotation speed of 200rpm, tank pressure of 0.05MPa and aeration ratio of 0.8VVM for 6h to obtain first-stage expanded culture seed solution, inoculating into 5m seed solution at an inoculum size of 1%3Culturing in a seed tank containing beef extract peptone culture medium at 35 deg.C, rotation speed of 150rpm, tank pressure of 0.05MPa and aeration ratio of 0.5VVM for 5 hr to obtain secondary propagation seed liquid;
(3) fermentation culture: inoculating the cultured seed solution into a fermentation tank filled with a fermentation culture medium by 5 percent of inoculation amount for fermentation culture, wherein the fermentation culture temperature is 35 ℃, the pH value is controlled to be 6.6-7.6 by using a sodium hydroxide solution in the whole fermentation process, the OUR value of the fermentation liquid is monitored in the whole fermentation process, the aeration ratio is controlled to be 0.6-1.0 VVM in the fermentation process, the rotating speed is controlled to be 80-120 rpm, and the tank pressure is controlled to be 0.04-0.05 MPa. When the oxygen consumption rate (OUR) in the hyaluronic acid fermentation liquor is first 16 mmol/L.h, adding 0.3 wt% of acetoin into the hyaluronic acid fermentation liquor, and continuously culturing until the residual sugar is less than or equal to 5 g/L. The composition per liter of fermentation medium is the same as in example 1. When the fermentation culture period reaches 20 hours, the residual sugar in the fermentation liquid is less than or equal to 5g/L, the fermentation is stopped, and the content of the hyaluronic acid in the fermentation liquid is 13.4 g/L.
Example 10
Hyaluronic acid was prepared according to example 5, except that acetoin was added to the fermentation broth at a ratio of 0.05 wt% when the oxygen consumption rate (OUR) in the fermentation broth of hyaluronic acid was first 16 mmol/l.h, and the fermentation was stopped when the fermentation culture period reached 20h, at which time the residual sugar in the fermentation broth was not more than 5g/L, and the hyaluronic acid content in the fermentation broth was 11.3g/L, as in example 5.
Example 11
Hyaluronic acid was prepared according to example 6, except that acetoin was added to the fermentation broth at a ratio of 0.55 wt% when the oxygen consumption rate (OUR) in the fermentation broth of hyaluronic acid was first 16 mmol/l.h, and under the same conditions as in example 6, when the fermentation culture period reached 20h, the residual sugar in the fermentation broth was no greater than 5g/L, and the fermentation was stopped, the hyaluronic acid content in the fermentation broth was 11.5 g/L.
Comparative example 1
Preparing hyaluronic acid according to the method of example 1, except that acetoin is not added in the fermentation culture process, and the other conditions are the same as example 1, when the fermentation culture period reaches 20 hours, the residual sugar in the fermentation liquid is less than or equal to 5g/L, the fermentation is stopped, and the content of hyaluronic acid in the fermentation liquid is 9.5 g/L.
Comparative example 2
Hyaluronic acid was prepared according to example 1, except that acetoin was added to the fermentation broth at a ratio of 0.3 wt% when the oxygen consumption rate (OUR) in the fermentation broth of hyaluronic acid was first 10 mmol/l.h, and under the same conditions as in example 1, when the fermentation culture period reached 20h, the residual sugar in the fermentation broth was no greater than 5g/L, and the fermentation was stopped, the hyaluronic acid content in the fermentation broth was 11.0 g/L.
Comparative example 3
Hyaluronic acid was prepared according to example 4, except that acetoin was added to the fermentation broth at a ratio of 0.3 wt% when the oxygen consumption rate (OUR) in the fermentation broth of hyaluronic acid was first 22 mmol/l.h, and under the same conditions as in example 4, when the fermentation culture period reached 20h, the residual sugar in the fermentation broth was no greater than 5g/L, and the fermentation was stopped, the hyaluronic acid content in the fermentation broth was 10.9 g/L.
Although embodiments of the present invention have been shown and described above, it is understood that the above embodiments are exemplary and should not be construed as limiting the present invention, and that variations, modifications, substitutions and alterations can be made in the above embodiments by those of ordinary skill in the art without departing from the principle and spirit of the present invention.
Claims (10)
1. The production method of hyaluronic acid is characterized by comprising the step of adding acetoin into hyaluronic acid fermentation liquor when the oxygen consumption rate in the hyaluronic acid fermentation liquor reaches 12-20 mmol/L.h for the first time in the fermentation culture process.
2. The production method of hyaluronic acid according to claim 1, characterized in that during fermentation culture, when the oxygen consumption rate in the hyaluronic acid fermentation broth reaches 14-18 mmol/L-h for the first time, acetoin is added to the hyaluronic acid fermentation broth.
3. The production method of hyaluronic acid according to claim 1, wherein the acetoin is added in an amount of 0.1-0.5 wt% based on the hyaluronic acid fermentation broth.
4. The method for producing hyaluronic acid according to claim 1, wherein the strain producing hyaluronic acid is Streptococcus zooepidemicus (Streptococcus zooepidemicus).
5. The method for producing hyaluronic acid according to claim 1, wherein the fermentation is terminated when the residual sugar content in the fermentation broth is less than or equal to 5 g/L.
6. The method for producing hyaluronic acid according to any of claims 1-5, wherein the method for producing hyaluronic acid comprises the steps of:
(1) activating strains: under the aseptic condition, streaking a hyaluronic acid production strain in a plate culture medium, and culturing at the temperature of 32-38 ℃ for 18-24 h;
(2) seed culture: picking single colonies from a cultured plate culture medium, inoculating the single colonies into a shake flask culture medium, then placing the shake flask culture medium into a shaking table, and culturing for 8-18 h under the conditions of the temperature of 32-38 ℃ and the rotating speed of 160-220 rpm to obtain a seed culture solution; optionally, inoculating the obtained seed culture solution into a seed tank filled with a shake flask culture medium in an inoculation amount of 0.5-5%, and culturing for 5-10 h at a temperature of 32-38 ℃, an aeration ratio of 0.3-1 VVM, a rotation speed of 150-300 rpm and a tank pressure of 0.03-0.05 MPa to obtain a seed solution;
(3) fermentation culture: inoculating the cultured seed culture solution or the seed solution into a fermentation tank filled with a fermentation culture medium by 1-10% of inoculation amount, carrying out fermentation culture at the temperature of 32-38 ℃, the aeration ratio of 0.5-1.5 VVM, the rotation speed of 80-300 rpm, the tank pressure of 0.04-0.06 MPa and the pH of 6.5-8.0, adding acetoin into the fermentation liquid when the oxygen consumption rate in the fermentation liquid reaches a preset value for the first time, continuing the culture, and ending the fermentation when the content of residual sugar in the fermentation liquid is less than or equal to 5 g/L.
7. The method for producing hyaluronic acid according to claim 6, wherein in step (1), the plating medium is beef extract peptone agar medium.
8. The method for producing hyaluronic acid according to claim 6, wherein in step (2), the shake flask culture medium is beef extract peptone culture medium.
9. The method for producing hyaluronic acid according to claim 6, wherein in step (3), the fermentation medium has a pH of 6.5-7.5, and each liter of fermentation medium contains the following components: 60-80 g of glucose, 5-15 g of peptone, 2-6 g of yeast extract, 0.5-2 g of sodium glutamate, 0.1-1 g of monopotassium phosphate, 0.1-1 g of magnesium sulfate and 0.1-1 g of foam killer.
10. The method for producing hyaluronic acid according to claim 6, wherein the fermentation tank used for fermentation culture has a volume of 30L-100 m3。
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Citations (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS62257901A (en) * | 1986-05-02 | 1987-11-10 | Seikagaku Kogyo Co Ltd | Production of hyaluronic acid |
| US20020102577A1 (en) * | 2000-07-31 | 2002-08-01 | Raillard Sun Ai | Nucleotide incorporating enzymes |
| CN1826059A (en) * | 2003-06-30 | 2006-08-30 | 大塚制药株式会社 | Composition containing lactic acid bacterium producing equol |
| US20110081683A1 (en) * | 2008-01-25 | 2011-04-07 | Battelle Energy Alliance, Llc | Methods of combined bioprocessing and related microorganisms, thermophilic and/or acidophilic enzymes, and nucleic acids encoding said enzymes |
| CN102057052A (en) * | 2008-04-14 | 2011-05-11 | 巴斯夫欧洲公司 | Method for manufacturing an aqueous glucose solution |
| CN104619652A (en) * | 2012-06-15 | 2015-05-13 | 微视生物技术有限公司 | Novel biocatalyst compositions and processes for use |
| CN109468355A (en) * | 2018-10-16 | 2019-03-15 | 河北科技大学 | A method of improving fermenting and producing hyaluronan molecule amount |
| CN111939100A (en) * | 2020-08-19 | 2020-11-17 | 广州黛诗莎化妆品有限公司 | Water-supplementing hyaluronic acid combined water level fermentation whitening liquid extraction process |
| CN112522232A (en) * | 2020-12-07 | 2021-03-19 | 内蒙古金达威药业有限公司 | Synthetic method of nicotinamide ribokinase and nicotinamide mononucleotide |
| CN115927143A (en) * | 2022-09-05 | 2023-04-07 | 江南大学 | Method and strain for improving N-acetylglucosamine yield through multi-path optimization |
-
2021
- 2021-12-24 CN CN202111609573.0A patent/CN114150030A/en active Pending
Patent Citations (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS62257901A (en) * | 1986-05-02 | 1987-11-10 | Seikagaku Kogyo Co Ltd | Production of hyaluronic acid |
| US20020102577A1 (en) * | 2000-07-31 | 2002-08-01 | Raillard Sun Ai | Nucleotide incorporating enzymes |
| CN1826059A (en) * | 2003-06-30 | 2006-08-30 | 大塚制药株式会社 | Composition containing lactic acid bacterium producing equol |
| US20110081683A1 (en) * | 2008-01-25 | 2011-04-07 | Battelle Energy Alliance, Llc | Methods of combined bioprocessing and related microorganisms, thermophilic and/or acidophilic enzymes, and nucleic acids encoding said enzymes |
| CN102057052A (en) * | 2008-04-14 | 2011-05-11 | 巴斯夫欧洲公司 | Method for manufacturing an aqueous glucose solution |
| CN104619652A (en) * | 2012-06-15 | 2015-05-13 | 微视生物技术有限公司 | Novel biocatalyst compositions and processes for use |
| CN109468355A (en) * | 2018-10-16 | 2019-03-15 | 河北科技大学 | A method of improving fermenting and producing hyaluronan molecule amount |
| CN111939100A (en) * | 2020-08-19 | 2020-11-17 | 广州黛诗莎化妆品有限公司 | Water-supplementing hyaluronic acid combined water level fermentation whitening liquid extraction process |
| CN112522232A (en) * | 2020-12-07 | 2021-03-19 | 内蒙古金达威药业有限公司 | Synthetic method of nicotinamide ribokinase and nicotinamide mononucleotide |
| CN115927143A (en) * | 2022-09-05 | 2023-04-07 | 江南大学 | Method and strain for improving N-acetylglucosamine yield through multi-path optimization |
Non-Patent Citations (3)
| Title |
|---|
| TING-FENG WU1等: "Proteomic investigation of the impact of oxygen on the protein profiles of hyaluronic acid-producing Streptococcus zooepidemicus", 《PROTEOMICS》, vol. 9, pages 4512 - 4516 * |
| 方柏山著: "《生物技术过程模型化与控制》", 暨南大学出版社, pages: 19 * |
| 郭学平等: "透明质酸及其发酵生产概述", 《中国生化药物杂志》, no. 4, pages 209 - 212 * |
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