CN107201383A - It is a kind of to improve the D-ALPHA-Hydroxypropionic acid production method that D-ALPHA-Hydroxypropionic acid produces intensity - Google Patents
It is a kind of to improve the D-ALPHA-Hydroxypropionic acid production method that D-ALPHA-Hydroxypropionic acid produces intensity Download PDFInfo
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- CN107201383A CN107201383A CN201610156757.9A CN201610156757A CN107201383A CN 107201383 A CN107201383 A CN 107201383A CN 201610156757 A CN201610156757 A CN 201610156757A CN 107201383 A CN107201383 A CN 107201383A
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- 238000004519 manufacturing process Methods 0.000 title claims abstract description 34
- JVTAAEKCZFNVCJ-UWTATZPHSA-N D-lactic acid Chemical compound C[C@@H](O)C(O)=O JVTAAEKCZFNVCJ-UWTATZPHSA-N 0.000 title claims description 31
- 238000000855 fermentation Methods 0.000 claims abstract description 81
- 230000004151 fermentation Effects 0.000 claims abstract description 81
- 238000011218 seed culture Methods 0.000 claims abstract description 35
- 239000002253 acid Substances 0.000 claims abstract description 30
- 239000002609 medium Substances 0.000 claims abstract description 24
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 claims abstract description 18
- 241000193830 Bacillus <bacterium> Species 0.000 claims abstract description 18
- 238000011534 incubation Methods 0.000 claims abstract description 18
- 238000007747 plating Methods 0.000 claims abstract description 17
- 239000001963 growth medium Substances 0.000 claims abstract description 11
- 239000012531 culture fluid Substances 0.000 claims abstract description 9
- 239000002054 inoculum Substances 0.000 claims abstract description 9
- 229910052757 nitrogen Inorganic materials 0.000 claims abstract description 9
- 230000003204 osmotic effect Effects 0.000 claims abstract description 9
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 claims description 26
- 229940041514 candida albicans extract Drugs 0.000 claims description 24
- 239000012138 yeast extract Substances 0.000 claims description 24
- 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 23
- 239000008103 glucose Substances 0.000 claims description 23
- VTYYLEPIZMXCLO-UHFFFAOYSA-L Calcium carbonate Chemical compound [Ca+2].[O-]C([O-])=O VTYYLEPIZMXCLO-UHFFFAOYSA-L 0.000 claims description 16
- 229960003237 betaine Drugs 0.000 claims description 12
- KWIUHFFTVRNATP-UHFFFAOYSA-N glycine betaine Chemical compound C[N+](C)(C)CC([O-])=O KWIUHFFTVRNATP-UHFFFAOYSA-N 0.000 claims description 12
- DHMQDGOQFOQNFH-UHFFFAOYSA-N Glycine Chemical compound NCC(O)=O DHMQDGOQFOQNFH-UHFFFAOYSA-N 0.000 claims description 10
- HDTRYLNUVZCQOY-UHFFFAOYSA-N α-D-glucopyranosyl-α-D-glucopyranoside Natural products OC1C(O)C(O)C(CO)OC1OC1C(O)C(O)C(O)C(CO)O1 HDTRYLNUVZCQOY-UHFFFAOYSA-N 0.000 claims description 8
- 229920001817 Agar Polymers 0.000 claims description 8
- 239000001888 Peptone Substances 0.000 claims description 8
- 108010080698 Peptones Proteins 0.000 claims description 8
- VMHLLURERBWHNL-UHFFFAOYSA-M Sodium acetate Chemical compound [Na+].CC([O-])=O VMHLLURERBWHNL-UHFFFAOYSA-M 0.000 claims description 8
- HDTRYLNUVZCQOY-WSWWMNSNSA-N Trehalose Natural products O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-WSWWMNSNSA-N 0.000 claims description 8
- 239000008272 agar Substances 0.000 claims description 8
- HDTRYLNUVZCQOY-LIZSDCNHSA-N alpha,alpha-trehalose Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-LIZSDCNHSA-N 0.000 claims description 8
- 229910000019 calcium carbonate Inorganic materials 0.000 claims description 8
- 229910052943 magnesium sulfate Inorganic materials 0.000 claims description 8
- 235000019341 magnesium sulphate Nutrition 0.000 claims description 8
- 229910000402 monopotassium phosphate Inorganic materials 0.000 claims description 8
- 235000019796 monopotassium phosphate Nutrition 0.000 claims description 8
- 235000019319 peptone Nutrition 0.000 claims description 8
- PJNZPQUBCPKICU-UHFFFAOYSA-N phosphoric acid;potassium Chemical compound [K].OP(O)(O)=O PJNZPQUBCPKICU-UHFFFAOYSA-N 0.000 claims description 8
- 239000000843 powder Substances 0.000 claims description 8
- 235000015099 wheat brans Nutrition 0.000 claims description 8
- 240000008042 Zea mays Species 0.000 claims description 7
- 235000005824 Zea mays ssp. parviglumis Nutrition 0.000 claims description 7
- 235000002017 Zea mays subsp mays Nutrition 0.000 claims description 7
- 229940040526 anhydrous sodium acetate Drugs 0.000 claims description 7
- 235000005822 corn Nutrition 0.000 claims description 7
- 239000004471 Glycine Substances 0.000 claims description 5
- DWNBOPVKNPVNQG-LURJTMIESA-N (2s)-4-hydroxy-2-(propylamino)butanoic acid Chemical compound CCCN[C@H](C(O)=O)CCO DWNBOPVKNPVNQG-LURJTMIESA-N 0.000 claims 1
- 235000015177 dried meat Nutrition 0.000 claims 1
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N lactic acid Chemical compound CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 abstract description 10
- 239000004310 lactic acid Substances 0.000 abstract description 5
- 235000014655 lactic acid Nutrition 0.000 abstract description 5
- 241000186660 Lactobacillus Species 0.000 abstract description 3
- 229940039696 lactobacillus Drugs 0.000 abstract description 3
- 150000007513 acids Chemical class 0.000 abstract 1
- 238000011081 inoculation Methods 0.000 abstract 1
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 12
- 239000000306 component Substances 0.000 description 9
- 238000000034 method Methods 0.000 description 9
- FYYHWMGAXLPEAU-UHFFFAOYSA-N Magnesium Chemical compound [Mg] FYYHWMGAXLPEAU-UHFFFAOYSA-N 0.000 description 6
- NINIDFKCEFEMDL-UHFFFAOYSA-N Sulfur Chemical compound [S] NINIDFKCEFEMDL-UHFFFAOYSA-N 0.000 description 6
- 239000005864 Sulphur Substances 0.000 description 6
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 6
- 239000011777 magnesium Substances 0.000 description 6
- 229910052749 magnesium Inorganic materials 0.000 description 6
- 239000012533 medium component Substances 0.000 description 6
- 239000013028 medium composition Substances 0.000 description 6
- 239000001301 oxygen Substances 0.000 description 6
- 229910052760 oxygen Inorganic materials 0.000 description 6
- 239000003002 pH adjusting agent Substances 0.000 description 6
- ONIBWKKTOPOVIA-UHFFFAOYSA-N Proline Natural products OC(=O)C1CCCN1 ONIBWKKTOPOVIA-UHFFFAOYSA-N 0.000 description 5
- 239000003795 chemical substances by application Substances 0.000 description 5
- ONIBWKKTOPOVIA-BYPYZUCNSA-N L-Proline Chemical compound OC(=O)[C@@H]1CCCN1 ONIBWKKTOPOVIA-BYPYZUCNSA-N 0.000 description 4
- 239000003223 protective agent Substances 0.000 description 4
- 241000894006 Bacteria Species 0.000 description 3
- 239000000047 product Substances 0.000 description 3
- ODHCTXKNWHHXJC-VKHMYHEASA-N 5-oxo-L-proline Chemical compound OC(=O)[C@@H]1CCC(=O)N1 ODHCTXKNWHHXJC-VKHMYHEASA-N 0.000 description 2
- FRXSZNDVFUDTIR-UHFFFAOYSA-N 6-methoxy-1,2,3,4-tetrahydroquinoline Chemical compound N1CCCC2=CC(OC)=CC=C21 FRXSZNDVFUDTIR-UHFFFAOYSA-N 0.000 description 2
- 239000003513 alkali Substances 0.000 description 2
- 235000011114 ammonium hydroxide Nutrition 0.000 description 2
- 239000003814 drug Substances 0.000 description 2
- 230000002363 herbicidal effect Effects 0.000 description 2
- 239000004009 herbicide Substances 0.000 description 2
- 230000008595 infiltration Effects 0.000 description 2
- 238000001764 infiltration Methods 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 229910021645 metal ion Inorganic materials 0.000 description 2
- 239000000575 pesticide Substances 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- QCVGEOXPDFCNHA-UHFFFAOYSA-N 5,5-dimethyl-2,4-dioxo-1,3-oxazolidine-3-carboxamide Chemical compound CC1(C)OC(=O)N(C(N)=O)C1=O QCVGEOXPDFCNHA-UHFFFAOYSA-N 0.000 description 1
- 239000002028 Biomass Substances 0.000 description 1
- 241000195493 Cryptophyta Species 0.000 description 1
- 229930182843 D-Lactic acid Natural products 0.000 description 1
- 102000002322 Egg Proteins Human genes 0.000 description 1
- 108010000912 Egg Proteins Proteins 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 240000007594 Oryza sativa Species 0.000 description 1
- 235000007164 Oryza sativa Nutrition 0.000 description 1
- 241001052560 Thallis Species 0.000 description 1
- 239000000159 acid neutralizing agent Substances 0.000 description 1
- 239000003905 agrochemical Substances 0.000 description 1
- GZCGUPFRVQAUEE-SLPGGIOYSA-N aldehydo-D-glucose Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@@H](O)C=O GZCGUPFRVQAUEE-SLPGGIOYSA-N 0.000 description 1
- 239000002585 base Substances 0.000 description 1
- 229940022769 d- lactic acid Drugs 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 230000018044 dehydration Effects 0.000 description 1
- 238000006297 dehydration reaction Methods 0.000 description 1
- 235000014103 egg white Nutrition 0.000 description 1
- 210000000969 egg white Anatomy 0.000 description 1
- 238000005265 energy consumption Methods 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 239000012530 fluid Substances 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 231100000053 low toxicity Toxicity 0.000 description 1
- 229910052751 metal Inorganic materials 0.000 description 1
- 239000002184 metal Substances 0.000 description 1
- 230000002906 microbiologic effect Effects 0.000 description 1
- 239000000178 monomer Substances 0.000 description 1
- 150000007524 organic acids Chemical class 0.000 description 1
- 235000020265 peanut milk Nutrition 0.000 description 1
- 239000012466 permeate Substances 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- 238000004321 preservation Methods 0.000 description 1
- 230000001737 promoting effect Effects 0.000 description 1
- 235000009566 rice Nutrition 0.000 description 1
- 238000004904 shortening Methods 0.000 description 1
- 239000001632 sodium acetate Substances 0.000 description 1
- 235000017281 sodium acetate Nutrition 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
Classifications
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- 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
- C12P7/00—Preparation of oxygen-containing organic compounds
- C12P7/40—Preparation of oxygen-containing organic compounds containing a carboxyl group including Peroxycarboxylic acids
- C12P7/56—Lactic acid
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- Organic Chemistry (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Zoology (AREA)
- Life Sciences & Earth Sciences (AREA)
- Wood Science & Technology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Microbiology (AREA)
- General Chemical & Material Sciences (AREA)
- Biotechnology (AREA)
- Health & Medical Sciences (AREA)
- Biochemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention discloses a kind of D production method of lectic acid for improving D production of lactic acid intensity, comprise the following steps：Flat board culture：Bacillus BS1 5 is seeded to plating medium Anaerobic culturel, 30~45 DEG C of cultivation temperature, 20~48h of incubation time；Seed culture：The bacillus of flat board culture is seeded to seed culture medium Anaerobic culturel, 30~45 DEG C of cultivation temperature, 12~24h of incubation time；Fermentation and acid：The seed culture fluid of acquisition is seeded in fermentation medium and carries out fermentation and acid; inoculum concentration is 3%~15%; 30~45 DEG C of fermentation temperature; it is passed through the environment that nitrogen keeps its anaerobism; the pH controls of fermentation system are added by the raising of osmotic pressure in osmoprotectant control fermentation tank 4.5~7.0 using nertralizer.During D lactobacillus inoculation fermentation and acids; due to there is the pH in the continuous generation fermentation tank of principal product lactic acid gradually to reduce; and osmoprotectant can be good at alleviating the problem of osmotic pressure is too high in environment; therefore appropriate osmoprotectant is added in fermentation process, so as to improve production intensity.
Description
Technical field
It is more particularly to a kind of to utilize fermentation control means to improve the invention belongs to technical field of bioengineering
The method of D-ALPHA-Hydroxypropionic acid production efficiency.
Background technology
Lactic acid is one of big organic acid in the world three, and it can be widely used in food, medicine, chemical industry and agriculture
The fields such as industry.Special D-ALPHA-Hydroxypropionic acid can be widely used in medicine, chemical industry and agriculture field.With D-ALPHA-Hydroxypropionic acid
The PLA synthesized for monomer is (such as transparent with its good biodegradable properties and other excellent use characteristics
Property, thermoplasticity, Product safety etc.) turned into 21 century green material most with prospects.In addition
The herbicide such as the biological pesticide " galloping horse " manufactured with high-purity D-ALPHA-Hydroxypropionic acid and " prestige despot ", as new low
Toxicity agricultural chemicals, can effectively herbicide pesticide it is not only economical but also reliable.In the market, the D-ALPHA-Hydroxypropionic acid price of ad eundem
5~10 times more expensive than Pfansteihl.Therefore the production and purifying of D-ALPHA-Hydroxypropionic acid are gradually taken seriously.
The preparation method of lactic acid has fermentation method, chemical synthesis and enzyme process.Wherein, fermentation method and chemistry are closed
Industrial application is obtained into method, and China mainly uses fermentation method.Compared with Pfansteihl, current D-ALPHA-Hydroxypropionic acid
Still there is larger gap in terms of the diversity and fermentation level of fermented bacterium, therefore further improve
The production strain of D-ALPHA-Hydroxypropionic acid, the fermenting property of raising strain are for promoting the fermentation of D-ALPHA-Hydroxypropionic acid industry and expanding D-
Lactic acid is using significant.
In D-ALPHA-Hydroxypropionic acid strain fermentation acid process, due to having in the continuous generation fermentation tank of principal product lactic acid
PH can gradually reduce, in order to maintain the optimal pH of thalli growth, need during the fermentation by continuous
Mend alkali and maintain pH, with the continuous inflow of alkali in fermentation process, the concentration of metal ions in zymotic fluid is not
Disconnected rise necessarily causes the rise of osmotic pressure, and too high osmotic pressure will influence growth and the production acid of thalline.And permeate
Pressure protective agent such as glycine betaine, trehalose, proline, glycine isosmoticity protective agent can be good at alleviating
Appropriate osmoprotectant is added in the problem of osmotic pressure is too high in environment, therefore fermentation process, so as to improve
Production intensity.Patent document CN101457243 discloses a kind of new work for improving Pidolidone fermentation production rate
Skill, the method that the content of the invention mainly with the addition of glycine betaine, proline in the fermentation medium, to effectively improve
The fermentation production rate of Pidolidone.It yet there are no the hair for being applied to D-ALPHA-Hydroxypropionic acid using osmoprotectant both at home and abroad
In ferment production.
The content of the invention
The purpose of the present invention is that the defect for being directed to prior art can improve D-ALPHA-Hydroxypropionic acid production intensity there is provided one kind
D-ALPHA-Hydroxypropionic acid production method.
To achieve these goals, the present invention uses following technical scheme：One kind can improve D-ALPHA-Hydroxypropionic acid production
The D-ALPHA-Hydroxypropionic acid production method of intensity, comprises the following steps：
(1) flat board culture：Bacillus BS1-5 is seeded to plating medium Anaerobic culturel, cultivation temperature
30~45 DEG C, 20~48h of incubation time；
(2) seed culture：Bacillus through step (1) flat board culture is seeded to seed culture medium anaerobism
Culture, 30~45 DEG C of cultivation temperature, 12~24h of incubation time；
(3) fermentation and acid：The seed culture fluid that step (2) is obtained, which is seeded in fermentation medium, to be fermented
Production acid, inoculum concentration is 3%~15%, and 30~45 DEG C of fermentation temperature is passed through the environment that nitrogen keeps its anaerobism, adopted
With nertralizer by the pH controls of fermentation system 4.5~7.0, add and permeated in osmoprotectant control fermentation tank
The raising of pressure.
The plating medium includes following components：10~30g/L of glucose, 1~3g/L of yeast extract, nothing
Water 1~4g/L of sodium acetate, 0.1~0.4g/L of anhydrous magnesium sulfate, 1~3g/L of potassium dihydrogen phosphate, 15~25g/L of agar.
The seed culture medium includes following components：10~40g/L of glucose, 1~3g/L of yeast extract, egg
White peptone 1~3g/L, 0.1~0.4g/L of anhydrous magnesium sulfate, 10~30g/L of calcium carbonate.
The fermentation medium includes following components：150~180g/L of glucose, 4~8g/L of yeast extract, it is beautiful
Rice ＆ peanut milk 4~6g/L of dry powder, 0.4~0.6g/L of magnesium sulfate, 8~12g/L of wheat bran.
The osmoprotectant glycine betaine, trehalose, proline or glycine, ferment in D-ALPHA-Hydroxypropionic acid
It is added to during the beginning or in fermentation process in fermentation tank, addition is 0.2~5g/L.
Strain used in the present invention produces bacterium for the D-ALPHA-Hydroxypropionic acid of a height growth rate of producing acid, in 2012
It is preserved in China typical culture collection center (China, Wuhan, Wuhan University) on December 6, in, it is classified
Lactobacillus (Sporolactobacillussp.) BS1-5 is named as, preservation registration number is CCTCC NO.
M2012516, it is more specifically categorized as synanthrin lactobacillus (Sporolactobacillussp.inulinus).
The method that the present invention improves D-ALPHA-Hydroxypropionic acid fermentation production strength is mainly when D-ALPHA-Hydroxypropionic acid ferments beginning or hair
A small amount of osmoprotectant is added in ferment hair is continuously added due to metal base to alleviate in the fermentation medium
The raising of the osmotic pressure brought in fermentation tank, the raising of osmotic pressure can influence growth and the production acid of D-ALPHA-Hydroxypropionic acid strain.
The addition of osmoprotectant, cell density improves, while shortening fermentation period by a relatively large margin.
Its principle is：Fermented according to microbiological anaerobic and produce the characteristic of D-ALPHA-Hydroxypropionic acid, with the addition of in fermentation system appropriate
Osmoprotectant, and these protective agents are mostly hydrophilic materials, they can enter cell, maintain bacterium
Osmotic pressure of the body cell in high substrate, high product and high metal ion concentration, prevents thalline from being pressed through because of infiltration
It is high and dehydration is dead.Higher biomass ensure that raising of the concentration of D-ALPHA-Hydroxypropionic acid with production intensity by a relatively large margin,
So as to reduce the energy consumption in D-ALPHA-Hydroxypropionic acid production process, production efficiency is improved.
Embodiment
With following embodiment, the invention will be further described, but it is necessary to note that following examples are only used
Further illustrated in the content of the invention, do not constitute limiting the scope of the invention.
Embodiment 1
(1) flat board culture：Bacillus BS1-5 is seeded to plating medium Anaerobic culturel, cultivation temperature
30 DEG C, incubation time 48h；
(2) seed culture：Bacillus through step (1) flat board culture is seeded into seed culture medium to detest
Oxygen culture, 30 DEG C of cultivation temperature, incubation time 24h；
(3) fermentation and acid：The seed culture fluid that step (2) is obtained, which is seeded in fermentation medium, to be sent out
Ferment production acid, inoculum concentration is 15%, 30 DEG C of fermentation temperature, the environment that nitrogen keeps its anaerobism is passed through, in
The pH of fermentation system is controlled 6.0 with agent.
Plating medium composition is (g/L)：Glucose 10, yeast extract 1, anhydrous sodium acetate 1, anhydrous sulphur
Sour magnesium 0.1, potassium dihydrogen phosphate 1, agar 15.
Seed culture based component is (g/L)：Glucose 10, yeast extract 1, peptone 1, anhydrous magnesium sulfate
0.1, calcium carbonate 10.
Fermentation medium components are (g/L):Glucose 150, yeast extract 4, Dried Corn Steep Liquor Powder 4, magnesium sulfate
0.4, wheat bran 8.
The fermentation and acid stage, table 1 is addition and has been not added with infiltration using sodium hydroxide as pH adjusting agent
Press the experimental result of protective agent glycine betaine：
Table 1 is not added with the experimental result with addition glycine betaine
Embodiment 2
(1) flat board culture：Bacillus BS1-5 is seeded to plating medium Anaerobic culturel, cultivation temperature
45 DEG C, incubation time 20h；
(2) seed culture：Bacillus through step (1) flat board culture is seeded into seed culture medium to detest
Oxygen culture, 45 DEG C of cultivation temperature, incubation time 12h；
(3) fermentation and acid：The seed culture fluid that step (2) is obtained, which is seeded in fermentation medium, to be sent out
Ferment production acid, inoculum concentration is 3%, 45 DEG C of fermentation temperature, the environment that nitrogen keeps its anaerobism is passed through, using neutralization
Agent controls the pH of fermentation system 7.0.
Plating medium composition is (g/L)：Glucose 30, yeast extract 3, anhydrous sodium acetate 4, anhydrous sulphur
Sour magnesium 0.4, potassium dihydrogen phosphate 3, agar 25.
Seed culture based component is (g/L)：Glucose 40, yeast extract 3, peptone 3, anhydrous magnesium sulfate
0.4, calcium carbonate 30.
Fermentation medium components are (g/L):Glucose 150, yeast extract 8, Dried Corn Steep Liquor Powder 6, magnesium sulfate 0.6,
Wheat bran 12.
The fermentation and acid stage, osmoprotectant was glycine betaine using sodium hydroxide as pH adjusting agent, under
Table is addition and the experimental result for being not added with osmoprotectant：
Table 2 is not added with the experimental result with addition glycine betaine
Embodiment 3
(1) flat board culture：Bacillus BS1-5 is seeded to plating medium Anaerobic culturel, cultivation temperature
37 DEG C, incubation time 24h；
(2) seed culture：Bacillus through step (1) flat board culture is seeded into seed culture medium to detest
Oxygen culture, 37 DEG C of cultivation temperature, incubation time 20h；
(3) fermentation and acid：The seed culture fluid that step (2) is obtained, which is seeded in fermentation medium, to be sent out
Ferment production acid, inoculum concentration is 10%, 37 DEG C of fermentation temperature, the environment that nitrogen keeps its anaerobism is passed through, in
The pH of fermentation system is controlled 6.5 with agent.
Plating medium composition is (g/L)：Glucose 20, yeast extract 2, anhydrous sodium acetate 2, anhydrous sulphur
Sour magnesium 0.3, potassium dihydrogen phosphate 2, agar 20.
Seed culture based component is (g/L)：Glucose 20, yeast extract 2, peptone 2, anhydrous magnesium sulfate
0.3, calcium carbonate 20.
Fermentation medium components are (g/L):Glucose 150, yeast extract 6, Dried Corn Steep Liquor Powder 5, magnesium sulfate
0.5, wheat bran 10.
The fermentation and acid stage, osmoprotectant was glycine betaine, sea using sodium hydroxide as pH adjusting agent
Algae sugar, proline or glycine, table 3 below, 4,5,6 are the reality for adding and being not added with osmoprotectant
Test result：
Table 3 is not added with the experimental result with addition glycine betaine
Table 4 is not added with the experimental result with addition glycine
Table 5 is not added with the experimental result with addition trehalose
Table 6 is not added with the experimental result with addition proline
Embodiment 4
(1) flat board culture：Bacillus BS1-5 is seeded to plating medium Anaerobic culturel, cultivation temperature
37 DEG C, incubation time 24h；
(2) seed culture：Bacillus through step (1) flat board culture is seeded into seed culture medium to detest
Oxygen culture, 37 DEG C of cultivation temperature, incubation time 20h；
(3) fermentation and acid：The seed culture fluid that step (2) is obtained, which is seeded in fermentation medium, to be sent out
Ferment production acid, inoculum concentration is 10%, 37 DEG C of fermentation temperature, the environment that nitrogen keeps its anaerobism is passed through, in
The pH of fermentation system is controlled 6.5 with agent.
Plating medium composition is (g/L)：Glucose 20, yeast extract 2, anhydrous sodium acetate 2, anhydrous sulphur
Sour magnesium 0.3, potassium dihydrogen phosphate 2, agar 20.
Seed culture based component is (g/L)：Glucose 20, yeast extract 2, peptone 2, anhydrous magnesium sulfate
0.3, calcium carbonate 20.
Fermentation medium components are (g/L):Glucose 160, yeast extract 6, Dried Corn Steep Liquor Powder 5, magnesium sulfate
0.5, wheat bran 10.
The fermentation and acid stage, osmoprotectant was trehalose, and following table is using ammoniacal liquor as pH adjusting agent
Add and be not added with the experimental result of osmoprotectant：
Table 7 is not added with the experimental result with addition trehalose
Embodiment 5
(1) flat board culture：Bacillus BS1-5 is seeded to plating medium Anaerobic culturel, cultivation temperature
37 DEG C, incubation time 24h；
(2) seed culture：Bacillus through step (1) flat board culture is seeded into seed culture medium to detest
Oxygen culture, 37 DEG C of cultivation temperature, incubation time 20h；
(3) fermentation and acid：The seed culture fluid that step (2) is obtained, which is seeded in fermentation medium, to be sent out
Ferment production acid, inoculum concentration is 10%, 37 DEG C of fermentation temperature, the environment that nitrogen keeps its anaerobism is passed through, in
The pH of fermentation system is controlled 6.5 with agent.
Plating medium composition is (g/L)：Glucose 20, yeast extract 2, anhydrous sodium acetate 2, anhydrous sulphur
Sour magnesium 0.3, potassium dihydrogen phosphate 2, agar 20.
Seed culture based component is (g/L)：Glucose 20, yeast extract 2, peptone 2, anhydrous magnesium sulfate
0.3, calcium carbonate 20.
Fermentation medium components are (g/L):Glucose 180, yeast extract 6, Dried Corn Steep Liquor Powder 5, magnesium sulfate
0.5, wheat bran 12.
The fermentation and acid stage, osmoprotectant was trehalose using sodium hydroxide as pH adjusting agent, under
Table is addition and the experimental result for being not added with osmoprotectant：
Table 8 is not added with the experimental result with addition trehalose
Embodiment 6
(1) flat board culture：Bacillus BS1-5 is seeded to plating medium Anaerobic culturel, cultivation temperature
37 DEG C, incubation time 24h；
(2) seed culture：Bacillus through step (1) flat board culture is seeded into seed culture medium to detest
Oxygen culture, 37 DEG C of cultivation temperature, incubation time 20h；
(3) fermentation and acid：The seed culture fluid that step (2) is obtained, which is seeded in fermentation medium, to be sent out
Ferment production acid, inoculum concentration is 10%, 37 DEG C of fermentation temperature, the environment that nitrogen keeps its anaerobism is passed through, in
The pH of fermentation system is controlled 6.5 with agent.
Plating medium composition is (g/L)：Glucose 20, yeast extract 2, anhydrous sodium acetate 2, anhydrous sulphur
Sour magnesium 0.3, potassium dihydrogen phosphate 2, agar 20.
Seed culture based component is (g/L)：Glucose 20, yeast extract 2, peptone 2, anhydrous magnesium sulfate
0.3, calcium carbonate 20.
Fermentation medium components are (g/L):Glucose 150, yeast extract 6, Dried Corn Steep Liquor Powder 5, magnesium sulfate
0.5, wheat bran 10.
The fermentation and acid stage, osmoprotectant was glycine betaine, and following table is using ammoniacal liquor as pH adjusting agent
Add and be not added with the experimental result of osmoprotectant：
Table 9 is not added with the experimental result with addition glycine betaine
Claims (5)
1. a kind of improve the D-ALPHA-Hydroxypropionic acid production method that D-ALPHA-Hydroxypropionic acid produces intensity, it is characterised in that comprises the following steps：
Flat board culture：Bacillus BS1-5 is seeded to plating medium Anaerobic culturel, 30~45 DEG C of cultivation temperature, incubation time
20~48h；
Seed culture：Bacillus through step (1) flat board culture is seeded to seed culture medium Anaerobic culturel, cultivation temperature
30~45 DEG C, 12~24h of incubation time；
Fermentation and acid：The seed culture fluid that step (2) is obtained, which is seeded in fermentation medium, carries out fermentation and acid, and inoculum concentration is
3%~15%, 30~45 DEG C of fermentation temperature is passed through the environment that nitrogen keeps its anaerobism, using nertralizer by the pH of fermentation system
Control adds the raising of osmotic pressure in osmoprotectant control fermentation tank 4.5~7.0.
2. production method according to claim 1, it is characterised in that：The plating medium includes following components：Glucose
10~30g/L, 1~3g/L of yeast extract, 1~4g/L of anhydrous sodium acetate, 0.1~0.4g/L of anhydrous magnesium sulfate, potassium dihydrogen phosphate 1~3
G/L, 15~25g/L of agar.
3. production method according to claim 1, it is characterised in that：The seed culture medium includes following components：Glucose
10~40g/L, 1~3g/L of yeast extract, 1~3g/L of peptone, 0.1~0.4g/L of anhydrous magnesium sulfate, 10~30g/L of calcium carbonate.
4. production method according to claim 1, it is characterised in that：The fermentation medium includes following components：Glucose
150~180g/L, 4~8g/L of yeast extract, 4~6g/L of Dried Corn Steep Liquor Powder, 0.4~0.6g/L of magnesium sulfate, 8~12g/L of wheat bran.
5. production method according to claim 1, it is characterised in that：The osmoprotectant glycine betaine, trehalose, dried meat
Propylhomoserin or glycine, are added in fermentation tank, addition is 0.2~5g/L when D-ALPHA-Hydroxypropionic acid ferments and originated or in fermentation process.
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN110862314A (en) * | 2018-08-27 | 2020-03-06 | 中国石化扬子石油化工有限公司 | Method for separating and extracting D-lactic acid from D-sodium lactate fermentation broth |
| CN110862313A (en) * | 2018-08-27 | 2020-03-06 | 中国石化扬子石油化工有限公司 | Method for extracting D-lactic acid from D-ammonium lactate fermentation liquor |
| CN111363708A (en) * | 2020-04-26 | 2020-07-03 | 驻马店华中正大有限公司 | Method for improving butyric acid fermentation level |
| CN118599735A (en) * | 2024-07-12 | 2024-09-06 | 南京工业大学 | A kind of lactic acid bacteria resistant to low pH and producing D-lactic acid and its application |
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| CN101457243A (en) * | 2009-01-06 | 2009-06-17 | 天津科技大学 | Novel process for improving L-glutamic acid fermentation production rate |
| CN104419657A (en) * | 2013-09-11 | 2015-03-18 | 中国石油化工股份有限公司 | D-lactic acid producing strain with high growth rate and acid producing velocity and application thereof |
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Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101457243A (en) * | 2009-01-06 | 2009-06-17 | 天津科技大学 | Novel process for improving L-glutamic acid fermentation production rate |
| CN104419657A (en) * | 2013-09-11 | 2015-03-18 | 中国石油化工股份有限公司 | D-lactic acid producing strain with high growth rate and acid producing velocity and application thereof |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110862314A (en) * | 2018-08-27 | 2020-03-06 | 中国石化扬子石油化工有限公司 | Method for separating and extracting D-lactic acid from D-sodium lactate fermentation broth |
| CN110862313A (en) * | 2018-08-27 | 2020-03-06 | 中国石化扬子石油化工有限公司 | Method for extracting D-lactic acid from D-ammonium lactate fermentation liquor |
| CN111363708A (en) * | 2020-04-26 | 2020-07-03 | 驻马店华中正大有限公司 | Method for improving butyric acid fermentation level |
| CN118599735A (en) * | 2024-07-12 | 2024-09-06 | 南京工业大学 | A kind of lactic acid bacteria resistant to low pH and producing D-lactic acid and its application |
| CN118599735B (en) * | 2024-07-12 | 2025-01-24 | 南京工业大学 | A kind of lactic acid bacteria resistant to low pH and producing D-lactic acid and its application |
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