CN107083406B - Method for producing (R) -3-hydroxybutyric acid - Google Patents
Method for producing (R) -3-hydroxybutyric acid Download PDFInfo
- Publication number
- CN107083406B CN107083406B CN201710396141.3A CN201710396141A CN107083406B CN 107083406 B CN107083406 B CN 107083406B CN 201710396141 A CN201710396141 A CN 201710396141A CN 107083406 B CN107083406 B CN 107083406B
- Authority
- CN
- China
- Prior art keywords
- hydroxybutyric acid
- fermentation
- corynebacterium glutamicum
- producing
- cgmcc
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
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
- C12P7/00—Preparation of oxygen-containing organic compounds
- C12P7/40—Preparation of oxygen-containing organic compounds containing a carboxyl group including Peroxycarboxylic acids
- C12P7/42—Hydroxy-carboxylic acids
-
- 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
- C12N1/20—Bacteria; Culture media therefor
- C12N1/205—Bacterial isolates
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
- C12R2001/15—Corynebacterium
Landscapes
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Genetics & Genomics (AREA)
- Biotechnology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- Microbiology (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- Biomedical Technology (AREA)
- Virology (AREA)
- Tropical Medicine & Parasitology (AREA)
- Medicinal Chemistry (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
The invention discloses a method for producing (R) -3-hydroxybutyric acid by microbial one-step fermentation, which adopts non-pathogenic microbes to directly convert a carbon source and a nitrogen source into (R) -3-hydroxybutyric acid by fermentation, wherein the produced (R) -3-hydroxybutyric acid can be further prepared into salts such as sodium salt, potassium salt, magnesium salt, calcium salt and the like, can be used as a pharmaceutical active ingredient or a nutritional supplement, and has wide industrial application prospect. The microorganism used in the method comprises corynebacterium glutamicum which is constructed by genetic engineering and is preserved in the China general microbiological culture Collection center with the preservation number of CGMCC No. 13957.
Description
Technical Field
The invention belongs to the field of bioengineering, and particularly relates to a method for producing (R) -3-hydroxybutyric acid and salts thereof through microbial fermentation.
Background
(R) -3-hydroxybutyric acid ((R) -3-Hydroxybutyrate, 3HB) is an optically active chiral compound with CAS number 625-72-9. (R) -3-hydroxybutyric acid is a compound produced in mammals by the metabolism of long-chain fatty acids in the liver, is present in plasma and peripheral tissues as the main ketone body, and can be used as an energy source in most tissues of the body. (R) -3-hydroxybutyric acid has, in addition to its nutritional function, a role in the treatment of a number of diseases, including: 1. many diseases that benefit from elevated ketone body levels (e.g., neurological disorders including epilepsy and myoclonus and neurodegenerative diseases including alzheimer's disease and dementia, etc.); 2. reducing free radical damage (e.g., ischemia) by oxidizing coenzyme Q; 3. enhancing metabolic efficiency (improving training efficiency and athletic performance, treating inadequate nutrition, angina pectoris, myocardial infarction, etc.); 4. treatment of diseases associated with, for example, cancer, particularly brain cancer (e.g., astrocytoma, etc.); 5. has good therapeutic effect on glucose metabolism disorder (such as type I diabetes, type II diabetes, hypoglycemia and hypoketosis); 6. can be used for preventing and treating osteopenia, osteoporosis, severe osteoporosis and related fracture. Based on these functions, (R) -3-hydroxybutyric acid and its salts can be used as food additives and drugs, and have great health and medicinal value.
The preparation of (R) -3-hydroxybutyric acid is mainly a chemical method, including direct chemical synthesis of (R) -3-hydroxybutyric acid, production of (R) -3-hydroxybutyric acid by enzymatic degradation of poly-3-hydroxybutyrate using poly-3-hydroxybutyrate depolymerizing enzyme. The method for chemically synthesizing (R) -3-hydroxybutyric acid requires high-temperature, high-pressure reaction conditions, expensive chiral metal catalysts and the like; the enzymatic degradation of poly-3-hydroxybutyrate to produce (R) -3-hydroxybutyrate needs to consume a large amount of organic solvent, has long reaction time, needs high-purity poly-3-hydroxybutyrate as a starting material, and is easy to cause racemization, but the method is still at a laboratory level at present, and if the method is used for industrial mass production, the problems of high cost, low yield and the like still exist.
Currently, most of 3-hydroxybutyric acid is marketed as racemate, i.e., an equal amount of a mixture of (R) -3-hydroxybutyric acid and (S) -3-hydroxybutyric acid. Although studies have shown that (S) -3-hydroxybutyric acid is not physiologically active, racemic 3-hydroxybutyric acid and its salts, especially the sodium salt, are still currently the predominant commercial form and accepted by a large number of consumers. It is expected that (R) -3-hydroxybutyric acid and its salts, which are single optically active, will be the mainstream of the consumer market in the future, replacing the optically inactive 3-hydroxybutyric acid and its salts.
Compounds of natural or biological origin are generally regarded as safer and more "natural" is sought for the origin of pharmaceutical, food and cosmetic ingredients. For marketing purposes, pharmaceutical, food, and cosmetic manufacturers prefer to use products of biological origin instead of the same substances synthesized chemically. The search for a biological method to replace the chemical method for producing (R) -3-hydroxybutyric acid as a widely used natural compound has been a research hotspot.
Disclosure of Invention
In order to realize a biological method for producing safe and nontoxic (R) -3-hydroxybutyric acid, the inventor explores a microbial one-step fermentation method, selects an nonpathogenic microorganism as a host, utilizes a genetic engineering technology to modify, weakens a branch metabolic pathway by enhancing the expression of genes related to the biosynthesis of the (R) -3-hydroxybutyric acid, and screens a production strain with high yield of the (R) -3-hydroxybutyric acid. The (R) -3-hydroxybutyric acid producing bacteria can effectively accumulate (R) -3-hydroxybutyric acid in the fermentation process, and have wide industrial application prospects.
Accordingly, a first object of the present invention is to provide a method for producing (R) -3-hydroxybutyric acid.
The second object of the present invention is to provide food grade (R) -3-hydroxybutyric acid and its salts.
The third purpose of the invention is to provide food-grade racemate 3-hydroxybutyric acid and salts thereof so as to meet the market demand.
The fourth object of the present invention is to provide a (R) -3-hydroxybutyric acid-producing bacterium.
The invention comprises the following technical scheme:
a method for producing (R) -3-hydroxybutyric acid by directly converting a carbon source and a nitrogen source into (R) -3-hydroxybutyric acid by fermentation using an avirulent microorganism, wherein the (R) -3-hydroxybutyric acid produced in the microorganism is secreted into a fermentation broth, and wherein (R) -3-hydroxybutyric acid is purified from the fermentation broth,
the microorganism is selected from corynebacterium glutamicum, bacillus subtilis, lactobacillus fermentation bacillus brevis, brachyspira sporangii, bacillus brevis flavus and bacillus brevis ammoniagenes, and the microorganism has the following sequential biotransformation functions: converting the carbon source and coenzyme a to acetyl-coenzyme a; converting acetyl-coa to acetoacetyl-coa; converting acetoacetyl-coa to (R) -3-hydroxybutyryl-coa; (R) -3-hydroxybutyryl-CoA is converted into (R) -3-hydroxybutyric acid.
Preferably, the microorganism overexpresses one or more enzymes selected from the group consisting of: Succinyl-CoA transferase (Succinyl-CoA transferase), acetoacetyl-CoA synthetase (acetoacetyl-CoA synthase), acetoacetyl-CoA reductase (acetoacetyl-CoA reductase, thioesterase (thioesterase).
More preferably, the microorganism overexpresses acetoacetyl-coa synthetase and acetoacetyl-coa reductase.
Preferably, the microorganism inhibits or down-regulates the expression of β -ketothiolase (β -ketothiolase).
In a preferred embodiment, the microorganism is Corynebacterium glutamicum, which is deposited in the China general microbiological culture Collection center with the collection number of CGMCC No. 13957.
During fermentation, different media can be used for different microorganisms, wherein the carbon source can be selected from glucose, sucrose, maltose, molasses, starch and glycerol. The nitrogen source in the fermentation medium can be selected from organic nitrogen sources and inorganic nitrogen sources, wherein the organic nitrogen sources are selected from corn steep liquor, bran hydrolysate, bean cake hydrolysate, yeast extract, yeast powder, peptone and urea; the inorganic nitrogen source is selected from ammonium sulfate, ammonium nitrate or ammonia water.
According to a preferred embodiment of the invention, when the microorganism is Corynebacterium glutamicum, the fermentation medium consists of: 75g/L glucose, 25-30g/L corn steep liquor and (NH)4)2SO4 20g/L、KH2PO4 1.5g/L、MgSO4·7H2O0.5g/L, urea 1.0g/L, histidine 30mg/L, molasses 25g/L, biotin 100 mu g/L, and antifoaming agent 0.2 g/L.
Preferably, when feeding is performed during fermentation, i.e. in a fed-batch culture, the feed medium used consists of: 500g/L ammonium sulfate and 650g/L glucose.
The purity of the (R) -3-hydroxybutyric acid prepared by the method of the invention is more than 95%, preferably more than 96%, more than 97%, more than 98%, more preferably more than 99%.
The (R) -3-hydroxybutyric acid prepared by the above method is free from bacterial endotoxins and free from chemical offensive odors such as bitter taste and solvent residue.
(R) -3-hydroxybutyric acid is an acid and may form salts with bases. Alternatively, (R) -3-hydroxybutyric acid of the present invention is in the form of a salt, such as sodium, potassium, magnesium, calcium; sodium salts are preferred. These salts may also be optically active compounds.
In view of the fact that racemic 3-hydroxybutyric acid and its sodium salt have been conventionally accepted by food and drug manufacturers, as well as consumers, racemic 3-hydroxybutyric acid and its salts can be prepared by racemization of (R) -3-hydroxybutyric acid and its salts. For example, racemization can be achieved by heating and holding (R) -3-hydroxybutyric acid in an alkaline solution such as sodium hydroxide solution for a certain period of time.
The racemic 3-hydroxybutyric acid salt may be sodium 3-hydroxybutyrate, potassium 3-hydroxybutyrate, magnesium 3-hydroxybutyrate, calcium 3-hydroxybutyrate or a mixture thereof.
The (R) -3-hydroxybutyric acid and the salts thereof prepared by the method do not contain bacterial endotoxin and toxic chemical substances, so that the food safety of the (R) -3-hydroxybutyric acid and the salts thereof can be guaranteed. In addition, since it does not contain chemical residues or chemical reaction impurities, (R) -3-hydroxybutyric acid of the present invention does not contain chemical odors such as bitterness, and can be directly used for manufacturing pharmaceuticals and health products.
Corynebacterium glutamicum is preserved in China general microbiological culture Collection center (CGMCC) with the preservation number of CGMCC No. 13957.
The (R) -3-hydroxybutyric acid producing strain constructed by the invention can realize the effective accumulation of (R) -3-hydroxybutyric acid in fermentation liquor in the fermentation process to obtain food-grade (R) -3-hydroxybutyric acid, thereby having wide industrial application prospect.
The Latin of the (R) -3-hydroxybutyric acid high-yield genetic engineering bacteria constructed by the invention has the chemical name of Corynebacterium glutamicum, the Chinese name of Corynebacterium glutamicum or Corynebacterium glutamicum, and is preserved in the common microorganism center of China Committee for culture Collection of microorganisms, the preservation date is 3 months and 30 days in 2017, the preservation address is the institute of microbiology of China academy of sciences No. 3 of the national institute of Western No.1 of North Chen of the sunward area in Beijing, and the preservation number is CGMCC No. 13957.
Detailed Description
The present invention will be described in further detail with reference to specific examples. It should be understood that the following examples are illustrative only and are not intended to limit the scope of the present invention.
The addition amount, content and concentration of various substances are referred to herein, wherein the percentage refers to the mass percentage unless otherwise specified.
In the examples of the present invention, if no specific description is made about the operation temperature, the temperature is usually room temperature (15 to 30 ℃).
As used herein, the terms "Corynebacterium glutamicum", "Corynebacterium glutamicum" and "strain CGMCC No. 13957" have the same meaning and refer to a strain constructed by the inventors for the fermentative production of (R) -3-hydroxybutyric acid.
In order to satisfy the pursuit of consumers and pharmaceutical and food manufacturers for a "natural" or "biological" source of (R) -3-hydroxybutyric acid and its salts, the inventors studied a method for producing (R) -3-hydroxybutyric acid by microbial fermentation. The inventor screens microorganism varieties for constructing the genetic engineering strains, abandons common but potentially pathogenic microorganisms such as colibacillus, selects nonpathogenic corynebacterium glutamicum, bacillus subtilis, lactobacillus fermentation brevibacterium, brachypodium sporangium, yellow brevibacterium and ammonia producing brevibacterium as hosts, constructs the genetic engineering strains, and obtains a plurality of strains capable of producing (R) -3-hydroxybutyric acid by one-step fermentation through screening. The strains can not generate endotoxin which can cause harm to most people in the fermentation process, and the design concept of 'no toxicity and no harm' is met.
In the fermentation process, in order to obtain higher yield of (R) -3-hydroxybutyric acid, conditions such as dissolved oxygen, temperature, pH and the like can be adjusted and controlled as necessary.
Preferably, fermentation is carried out with a constant dO2The content is controlled to be 15-25 percent. For example, the fermentation can be carried out under the following conditions: the air flow rate is about 1vvm, where vvm (aeration ratio) is the ratio of aeration per minute to the actual feed volume of the tank (for example, 1vvm equals 30L/min for a fermentor containing 30 liters of fermentation broth and 1vvm equals 5L/min for a fermentor containing 5 liters of fermentation broth).
Preferably, the temperature is controlled to be 30-32 ℃ when fermentation is carried out; in the later stage of fermentation, the temperature can be increased to 34-37 ℃ so as to facilitate the synthesis and discharge of (R) -3-hydroxybutyric acid into the fermentation liquor.
Preferably, when fermentation is carried out, the pH is controlled to be 6.0-8.0, preferably 6.5-7.0; in the later stage of fermentation, the pH value can be controlled to be 6.8-7.0, so that the synthesis and discharge of (R) -3-hydroxybutyric acid into fermentation liquor are facilitated.
The term "late fermentation stage" as used above means the stage from the growth plateau to the death stage of the microorganism. For example using OD600nmValue monitoring of microbial concentration, OD600nmThe value no longer rises and tends to fallAnd (5) reducing.
The residual sugar in the whole fermentation process is controlled to be about 1 to 3.0 percent, and more preferably 1.5 to 2.5 percent.
After the fermentation is finished, the fermentation liquor needs to be recovered, and the (R) -3-hydroxybutyric acid is extracted from the fermentation liquor. For example, a supernatant from which the cells have been removed is obtained by centrifugation, and the supernatant is concentrated if necessary; (R) -3-hydroxybutyric acid is isolated by post-treatment means such as drying and purification. Or removing thallus and macromolecular substances by filtration (including ultrafiltration and nanofiltration), to obtain filtrate, and concentrating the filtrate if necessary; and then (R) -3-hydroxybutyric acid is separated by post-treatment means such as drying, purification and the like. Or centrifuging to obtain supernatant, removing macromolecular substances by ultrafiltration or nanofiltration to obtain filtrate, and concentrating the filtrate if necessary; and then (R) -3-hydroxybutyric acid is separated by post-treatment means such as drying, purification and the like.
In the preparation of (R) -3-hydroxybutyric acid salts such as sodium, potassium, magnesium and calcium salts, the reaction temperature of equivalent amounts of (R) -3-hydroxybutyric acid with the corresponding base or metal oxide such as sodium hydroxide is controlled to be 30 ℃ or less, preferably 25 ℃ or less, more preferably 20 ℃ or less, to avoid racemization as much as possible.
Because no organic solvent is used in the preparation process, the obtained (R) -3-hydroxybutyric acid and salt products thereof do not contain chemical peculiar smell such as bitter taste, and can be directly used for manufacturing medicines and health care products.
Examples
Example 1: preculture and fermentation
Thawing the preservation solution of strain CGMCC No.13957 in glycerin tube, inoculating to seed culture medium (glucose 75g/L, corn steep liquor 25-30g/L, (NH)4)2SO4 20g/L、KH2PO4 1.5g/L、MgSO4·7H2O0.5g/L, urea 1.0g/L, histidine 30mg/L, molasses 25g/L, biotin 100. mu.g/L, pH 7.0), in a 5000mL Erlenmeyer flask, the liquid was filled in 500mL, cultured at 30 ℃ for 18 hours, and shaken when the absorbance value. DELTA. OD was 0.4-0.5, to obtain a seed culture solution. The detection result shows that the product is free from mixed bacteria.
500mL of the seed culture was inoculated into a 7-liter fermentor and the liquid content was 5 liters. The medium composition is the same as the seed medium, and the pH is 6.4 to 6.7 (after sterilization). A supplemented medium: 500g/L ammonium sulfate and 650g/L glucose. The culture temperature was 30 ℃, the pot pressure was 0.05MPa, and the initial aeration ratio was 1 vvm. Stirring was carried out at 600 rpm. Controlling the pH value to be about 6.5 during fermentation; in the later stage of fermentation, the pH is controlled at about 6.7, and the temperature is raised to about 35 ℃. The ventilation rate and the stirring speed are flexibly controlled according to dissolved oxygen, and the dissolved oxygen constant dO2The content is controlled to be 15-25%. Controlling residual sugar: when the initial sugar is reduced to about 3.0 percent, feeding materials is started, and the residual sugar in the whole fermentation process is controlled to about 1.5 to 2.0 percent. The total fermentation time is 72 hours, and the concentration of the product (R) -3-hydroxybutyric acid is 11.5 g/L.
Example 2: separation of fermentation liquor and extraction and purification of (R) -3-hydroxybutyric acid
5.2 liters of the fermentation solution obtained in example 1 were centrifuged at 4500rpm, and the cells were discarded. The supernatant was filtered through 1% celite. Then adding 1% active carbon, stirring and decoloring for 30 minutes, and filtering to obtain clear filtrate.
Nanofiltration treatment is carried out on the filtrate by using a nanofiltration membrane, the obtained filtrate passes through 732 cation exchange resin, and the penetration liquid is collected. Concentrated to 1000g/L as an oil, which was poured out while hot to give 55.9g of (R) -3-hydroxybutyric acid in 91% yield. And (R) -3-hydroxybutyric acid purity is determined by high performance liquid chromatography. The chromatographic column was a Shim-pack Vp-ODSC18 column (150 L.times.4.6), the mobile phase was acetonitrile: water (v/v) ═ 15: 85, ultraviolet detection wavelength of 210nm, sample injection amount of 20 microlitre, flow rate of 1mL/min and column temperature of 10 ℃. The detection shows that the purity of the (R) -3-hydroxybutyric acid is 99.2 percent, and the specific rotation: [ alpha ] to]D 20＝-25.1°(C＝6％，H2O)。
Example 3: preparation of sodium (R) -3-hydroxybutyrate
5g of (R) -3-hydroxybutyric acid from example 2 was slowly added to an equivalent of 2N NaOH solution to perform neutralization reaction at a temperature of 25 ℃ or lower. Concentrating by rotary evaporation until solid is separated out, standing for 2 hours, filtering and collecting the solid, drying by blowing at 60 ℃ to obtain 4.8 g of (R) -3-sodium hydroxybutyrate powder, and collectingThe ratio was 79%. Melting point 152 ℃, specific optical rotation: [ alpha ] to]D 20＝-14.2°(C＝10％，H2O)。
Example 4: racemization of (R) -3-hydroxybutyric acid
10g of (R) -3-hydroxybutyric acid was slowly added to 100mL of 2N NaOH solution and heated to 60 ℃ for 4 hours. The temperature was lowered to room temperature and the optical rotation was measured by a polarimeter to be 0 °. Neutralizing with hydrochloric acid to pH 7.0, concentrating by rotary evaporation to precipitate solid, standing for 2 hr, and vacuum filtering to collect solid to obtain sodium 3-hydroxybutyrate powder 10.8g, yield 89%, and optical rotation of 0 ° (C10%, H) as determined by polarimeter2O). The results show that racemic sodium 3-hydroxybutyrate was obtained.
In conclusion, the (R) -3-hydroxybutyric acid genetic engineering bacteria constructed by the invention can realize the effective accumulation of (R) -3-hydroxybutyric acid in fermentation liquor in the fermentation process, produce safe and nontoxic food-grade (R) -3-hydroxybutyric acid and have wide industrial application prospects.
Claims (4)
1. Corynebacterium glutamicum is preserved in China general microbiological culture Collection center (CGMCC) with the preservation number of CGMCC No. 13957.
2. A method for producing (R) -3-hydroxybutyric acid by directly converting a carbon source and a nitrogen source into (R) -3-hydroxybutyric acid by fermentation using the Corynebacterium glutamicum of claim 1, wherein the (R) -3-hydroxybutyric acid produced in the Corynebacterium glutamicum is secreted into a fermentation broth, and the (R) -3-hydroxybutyric acid is purified from the fermentation broth.
3. The method of claim 2, wherein the carbon source is selected from the group consisting of glucose, sucrose, maltose, molasses, starch and glycerol; the nitrogen source is selected from organic nitrogen sources and inorganic nitrogen sources, and the organic nitrogen sources are selected from corn steep liquor, bran hydrolysate, bean cake hydrolysate, yeast extract, yeast powder, peptone and urea; the inorganic nitrogen source is selected from ammonium sulfate, ammonium nitrate or ammonia water.
4. A product prepared by the process of claim 2 or 3, which is free of bacterial endotoxin and has a purity of greater than 95% for (R) -3-hydroxybutyric acid.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710396141.3A CN107083406B (en) | 2017-05-27 | 2017-05-27 | Method for producing (R) -3-hydroxybutyric acid |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710396141.3A CN107083406B (en) | 2017-05-27 | 2017-05-27 | Method for producing (R) -3-hydroxybutyric acid |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN107083406A CN107083406A (en) | 2017-08-22 |
| CN107083406B true CN107083406B (en) | 2021-02-02 |
Family
ID=59608670
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201710396141.3A Active CN107083406B (en) | 2017-05-27 | 2017-05-27 | Method for producing (R) -3-hydroxybutyric acid |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN107083406B (en) |
Families Citing this family (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| AU2018250146B2 (en) | 2017-04-04 | 2020-11-05 | Nanjing Nutrabuilding Bio-Tech Co., Ltd. | Preparation of (R)-3-hydroxybutyric acid or its salts by one-step fermentation |
| CN110862316A (en) * | 2018-08-27 | 2020-03-06 | 浙江华睿生物技术有限公司 | Crystal form of (R) -3-hydroxybutyric acid and application thereof |
| CN111944853A (en) * | 2019-05-17 | 2020-11-17 | 南京纽邦生物科技有限公司 | Method for producing (R) -3-hydroxybutyric acid by microbial fermentation |
| CN110904161A (en) * | 2019-12-27 | 2020-03-24 | 浙江英玛特生物科技有限公司 | Method for producing high-purity (R) - (-) -3-hydroxybutyric acid by adopting enzyme method |
| CN111304140A (en) * | 2020-03-05 | 2020-06-19 | 清华大学 | Recombinant intestinal bacterium for producing (R) -3-hydroxybutyric acid and construction method thereof |
| CN111534561A (en) * | 2020-05-19 | 2020-08-14 | 南京纽邦生物科技有限公司 | Preparation method of (R) -3-hydroxybutyric acid |
Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1429912A (en) * | 2002-01-04 | 2003-07-16 | 清华大学 | Method of producing D-(-)-3-hydroxy butanoic acid |
| CN101969769A (en) * | 2008-01-04 | 2011-02-09 | 伊希斯创新有限公司 | Ketone bodies and ketone body esters as blood lipid lowering agents |
| CN103890187A (en) * | 2011-10-18 | 2014-06-25 | 昭和电工株式会社 | Production method for organic acid using coa-transferase |
| CN105050594A (en) * | 2013-03-19 | 2015-11-11 | 南佛罗里达大学 | Compositions and methods for producing elevated and sustained ketosis |
| CN105950517A (en) * | 2016-06-30 | 2016-09-21 | 江南大学 | Corynebacterium glutamicum and application thereof |
| CN106148440A (en) * | 2016-08-10 | 2016-11-23 | 洛阳华荣生物技术有限公司 | Fermentative Production gamatine |
| WO2017016902A1 (en) * | 2015-07-29 | 2017-02-02 | Evonik Degussa Gmbh | Production of 3-hydroxybutyrate |
| WO2017066498A1 (en) * | 2015-10-13 | 2017-04-20 | Lanzatech New Zealand Limited | Genetically engineered bacterium comprising energy-generating fermentation pathway |
-
2017
- 2017-05-27 CN CN201710396141.3A patent/CN107083406B/en active Active
Patent Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1429912A (en) * | 2002-01-04 | 2003-07-16 | 清华大学 | Method of producing D-(-)-3-hydroxy butanoic acid |
| CN101969769A (en) * | 2008-01-04 | 2011-02-09 | 伊希斯创新有限公司 | Ketone bodies and ketone body esters as blood lipid lowering agents |
| CN103890187A (en) * | 2011-10-18 | 2014-06-25 | 昭和电工株式会社 | Production method for organic acid using coa-transferase |
| CN105050594A (en) * | 2013-03-19 | 2015-11-11 | 南佛罗里达大学 | Compositions and methods for producing elevated and sustained ketosis |
| WO2017016902A1 (en) * | 2015-07-29 | 2017-02-02 | Evonik Degussa Gmbh | Production of 3-hydroxybutyrate |
| WO2017066498A1 (en) * | 2015-10-13 | 2017-04-20 | Lanzatech New Zealand Limited | Genetically engineered bacterium comprising energy-generating fermentation pathway |
| CN105950517A (en) * | 2016-06-30 | 2016-09-21 | 江南大学 | Corynebacterium glutamicum and application thereof |
| CN106148440A (en) * | 2016-08-10 | 2016-11-23 | 洛阳华荣生物技术有限公司 | Fermentative Production gamatine |
Non-Patent Citations (1)
| Title |
|---|
| Efficient (R)-3-hydroxybutyrate production using acetyl CoA-regenerating pathway catalyzed by coenzyme A transferase;Ken’ichiro Matsumoto;《Appl Microbiol Biotechnol》;20130131;第97卷(第1期);205-210 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN107083406A (en) | 2017-08-22 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| US11198890B2 (en) | Preparation of (R)-3-hydroxybutyric acid or its salts by one-step fermentation | |
| CN107083406B (en) | Method for producing (R) -3-hydroxybutyric acid | |
| JP4359349B2 (en) | Production method of vanillin | |
| EP2164975B1 (en) | Process for preparing a concentrated aqueous glucose solution from corn | |
| CN112481336B (en) | Method for biosynthesis of compounds using lignocellulose derivatives | |
| DE102005056668A1 (en) | Fermentative preparation of organic compounds, useful for making e.g. amino acids or enzymes, comprises using a starch source from which non-starch solids have not been removed, hydrolyzed and mixed with sugars | |
| RU2451081C2 (en) | Fermentative production of organic compounds | |
| JP6861202B2 (en) | Improved production of vanillin by fermentation | |
| CN106148440B (en) | Production of agmatine by fermentation method | |
| EP2850213B1 (en) | Strain producing turanose and uses thereof | |
| TW201142032A (en) | Utilization of agro residual substrates for fermentative production of L-arginine | |
| US5686277A (en) | Fermentation process for preparing xylitol using mutant cells | |
| CN115715328A (en) | Fermentation process | |
| Papagianni | Microbial bioprocesses | |
| CN114807286B (en) | Method for refining prednisone acetate based on resting cell transformation | |
| EP4133095B1 (en) | Process for preparing phenylacetic acid | |
| JP4870442B2 (en) | Method for producing lactic acid | |
| CN114395590A (en) | Method for producing levodopa by microbial fermentation and application | |
| CN111944853A (en) | Method for producing (R) -3-hydroxybutyric acid by microbial fermentation | |
| CN110684810A (en) | Method for preparing powdery gamma-aminobutyric acid from fermentation liquor rich in gamma-aminobutyric acid | |
| US20110262979A1 (en) | Novel composition of matter for use in producing co-enzyme q10 and a novel method for producing co-enzyme q10 |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| TA01 | Transfer of patent application right |
Effective date of registration: 20180102 Address after: 313000 Huzhou, Huzhou City, Zhejiang, No. 6, Hongfeng Road, No. 1366, floor 502, west side Applicant after: Zhejiang Huari Biotechnology Co., Ltd. Address before: 313000 Hongfeng Road, Huzhou, Huzhou, Zhejiang, 3, 2 floors and 205 rooms Applicant before: Huzhou Heng Rui Nutrition Health Technology Co., Ltd. |
|
| TA01 | Transfer of patent application right | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |