CN110218661A - A kind of Candida tropicalis and its application and a kind of production method of long-chain biatomic acid - Google Patents
A kind of Candida tropicalis and its application and a kind of production method of long-chain biatomic acid Download PDFInfo
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- CN110218661A CN110218661A CN201810172091.5A CN201810172091A CN110218661A CN 110218661 A CN110218661 A CN 110218661A CN 201810172091 A CN201810172091 A CN 201810172091A CN 110218661 A CN110218661 A CN 110218661A
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- 229910001651 emery Inorganic materials 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- XYIBRDXRRQCHLP-UHFFFAOYSA-N ethyl acetoacetate Chemical compound CCOC(=O)CC(C)=O XYIBRDXRRQCHLP-UHFFFAOYSA-N 0.000 description 1
- 230000008020 evaporation Effects 0.000 description 1
- 239000000284 extract Substances 0.000 description 1
- 239000006052 feed supplement Substances 0.000 description 1
- 239000006260 foam Substances 0.000 description 1
- 235000011389 fruit/vegetable juice Nutrition 0.000 description 1
- 238000012239 gene modification Methods 0.000 description 1
- 230000005017 genetic modification Effects 0.000 description 1
- 235000013617 genetically modified food Nutrition 0.000 description 1
- 239000004519 grease Substances 0.000 description 1
- IPCSVZSSVZVIGE-UHFFFAOYSA-N hexadecanoic acid Chemical compound CCCCCCCCCCCCCCCC(O)=O IPCSVZSSVZVIGE-UHFFFAOYSA-N 0.000 description 1
- 239000001257 hydrogen Substances 0.000 description 1
- 229910052739 hydrogen Inorganic materials 0.000 description 1
- FIKTURVKRGQNQD-UHFFFAOYSA-N icos-2-enoic acid Chemical compound CCCCCCCCCCCCCCCCCC=CC(O)=O FIKTURVKRGQNQD-UHFFFAOYSA-N 0.000 description 1
- VKOBVWXKNCXXDE-UHFFFAOYSA-N icosanoic acid Chemical compound CCCCCCCCCCCCCCCCCCCC(O)=O VKOBVWXKNCXXDE-UHFFFAOYSA-N 0.000 description 1
- 230000001939 inductive effect Effects 0.000 description 1
- FBAFATDZDUQKNH-UHFFFAOYSA-M iron chloride Chemical compound [Cl-].[Fe] FBAFATDZDUQKNH-UHFFFAOYSA-M 0.000 description 1
- YAQXGBBDJYBXKL-UHFFFAOYSA-N iron(2+);1,10-phenanthroline;dicyanide Chemical compound [Fe+2].N#[C-].N#[C-].C1=CN=C2C3=NC=CC=C3C=CC2=C1.C1=CN=C2C3=NC=CC=C3C=CC2=C1 YAQXGBBDJYBXKL-UHFFFAOYSA-N 0.000 description 1
- JEIPFZHSYJVQDO-UHFFFAOYSA-N iron(III) oxide Inorganic materials O=[Fe]O[Fe]=O JEIPFZHSYJVQDO-UHFFFAOYSA-N 0.000 description 1
- 239000010977 jade Substances 0.000 description 1
- 238000011031 large-scale manufacturing process Methods 0.000 description 1
- 239000000314 lubricant Substances 0.000 description 1
- 229910052748 manganese Inorganic materials 0.000 description 1
- 239000011572 manganese Substances 0.000 description 1
- 229940099596 manganese sulfate Drugs 0.000 description 1
- 239000011702 manganese sulphate Substances 0.000 description 1
- 235000007079 manganese sulphate Nutrition 0.000 description 1
- SQQMAOCOWKFBNP-UHFFFAOYSA-L manganese(II) sulfate Chemical compound [Mn+2].[O-]S([O-])(=O)=O SQQMAOCOWKFBNP-UHFFFAOYSA-L 0.000 description 1
- 230000004060 metabolic process Effects 0.000 description 1
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 description 1
- 230000002906 microbiologic effect Effects 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- WQEPLUUGTLDZJY-UHFFFAOYSA-N n-Pentadecanoic acid Natural products CCCCCCCCCCCCCCC(O)=O WQEPLUUGTLDZJY-UHFFFAOYSA-N 0.000 description 1
- 229920001778 nylon Polymers 0.000 description 1
- QIQXTHQIDYTFRH-UHFFFAOYSA-N octadecanoic acid Chemical compound CCCCCCCCCCCCCCCCCC(O)=O QIQXTHQIDYTFRH-UHFFFAOYSA-N 0.000 description 1
- OQCDKBAXFALNLD-UHFFFAOYSA-N octadecanoic acid Natural products CCCCCCCC(C)CCCCCCCCC(O)=O OQCDKBAXFALNLD-UHFFFAOYSA-N 0.000 description 1
- 238000011017 operating method Methods 0.000 description 1
- 238000010979 pH adjustment Methods 0.000 description 1
- 239000003973 paint Substances 0.000 description 1
- 239000002245 particle Substances 0.000 description 1
- 150000002978 peroxides Chemical class 0.000 description 1
- 239000004014 plasticizer Substances 0.000 description 1
- 239000001103 potassium chloride Substances 0.000 description 1
- 235000011164 potassium chloride Nutrition 0.000 description 1
- BUCIWTBCUUHRHZ-UHFFFAOYSA-K potassium;disodium;dihydrogen phosphate;hydrogen phosphate Chemical compound [Na+].[Na+].[K+].OP(O)([O-])=O.OP([O-])([O-])=O BUCIWTBCUUHRHZ-UHFFFAOYSA-K 0.000 description 1
- 230000001376 precipitating effect Effects 0.000 description 1
- 230000001105 regulatory effect Effects 0.000 description 1
- 229960002477 riboflavin Drugs 0.000 description 1
- RMAQACBXLXPBSY-UHFFFAOYSA-N silicic acid Chemical compound O[Si](O)(O)O RMAQACBXLXPBSY-UHFFFAOYSA-N 0.000 description 1
- 235000012239 silicon dioxide Nutrition 0.000 description 1
- 239000011684 sodium molybdate Substances 0.000 description 1
- 235000015393 sodium molybdate Nutrition 0.000 description 1
- TVXXNOYZHKPKGW-UHFFFAOYSA-N sodium molybdate (anhydrous) Chemical compound [Na+].[Na+].[O-][Mo]([O-])(=O)=O TVXXNOYZHKPKGW-UHFFFAOYSA-N 0.000 description 1
- 239000008117 stearic acid Substances 0.000 description 1
- 238000004659 sterilization and disinfection Methods 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 229960003495 thiamine Drugs 0.000 description 1
- DPJRMOMPQZCRJU-UHFFFAOYSA-M thiamine hydrochloride Chemical compound Cl.[Cl-].CC1=C(CCO)SC=[N+]1CC1=CN=C(C)N=C1N DPJRMOMPQZCRJU-UHFFFAOYSA-M 0.000 description 1
- 229910021654 trace metal Inorganic materials 0.000 description 1
- 238000012546 transfer Methods 0.000 description 1
- DXNCZXXFRKPEPY-UHFFFAOYSA-N tridecanedioic acid Chemical compound OC(=O)CCCCCCCCCCCC(O)=O DXNCZXXFRKPEPY-UHFFFAOYSA-N 0.000 description 1
- 230000003827 upregulation Effects 0.000 description 1
- 230000003519 ventilatory effect Effects 0.000 description 1
- 235000010374 vitamin B1 Nutrition 0.000 description 1
- 239000011691 vitamin B1 Substances 0.000 description 1
- 235000019164 vitamin B2 Nutrition 0.000 description 1
- 239000011716 vitamin B2 Substances 0.000 description 1
- 235000019154 vitamin C Nutrition 0.000 description 1
- 239000011718 vitamin C Substances 0.000 description 1
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- 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
- C12N1/14—Fungi; Culture media therefor
- C12N1/16—Yeasts; Culture media therefor
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- 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
- C12N1/14—Fungi; Culture media therefor
- C12N1/16—Yeasts; Culture media therefor
- C12N1/165—Yeast isolates
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- 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
- C12P7/64—Fats; Fatty oils; Ester-type waxes; Higher fatty acids, i.e. having at least seven carbon atoms in an unbroken chain bound to a carboxyl group; Oxidised oils or fats
- C12P7/6409—Fatty acids
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- 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
- C12R2001/645—Fungi ; Processes using fungi
- C12R2001/72—Candida
- C12R2001/74—Candida tropicalis
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Abstract
The invention discloses a kind of Candida tropicalis (Candida tropicalis) bacterial strain CAT H1614, including its microbial inoculum, the preparation method of the method and long-chain biatomic acid of application and a kind of fermenting and producing long-chain biatomic acid.Candida tropicalis bacterial strain CAT H1614 of the invention can keep high vigor in the environment of pH value is lower than 7.0 for a long time, thus produce long-chain biatomic acid especially suitable for bioanalysis, and fermentation process produces acid fastly, and yield is high, moreover it is possible to reduce fermentation period.Long-chain biatomic acid fermentation process provided by the invention and preparation method have significant cost advantage, and can effectively reduce the pressure to resource and environment, therefore have obviously industrial value advantage.
Description
Technical field
The present invention relates to fermentation arts, and in particular to a kind of Candida tropicalis and its application in fermentation arts,
And a kind of production method of long-chain biatomic acid.
Background technique
The general formula of long-chain biatomic acid (LCDA) is (HOOC (CH2) nCOOH, n >=7), have very extensive purposes, with length
Chain binary acid is that raw material can synthesize extraordinary nylon, fine perfumery, high-grade hot melt adhesive, cold resistant plasticizer, senior lubricant, advanced
Antirust agent, advanced paint and coating etc..Long-chain biatomic acid can usually be synthesized with chemical method or bioanalysis.Chemical method synthesizes road
Wire length, reaction need high temperature and pressure, it is harsher to catalyst requirement, thus long-chain biatomic acid kind at industrial scale compared with
Lack, only a small number of kinds such as 12 carbon long-chain biatomic acids.And bioanalysis is to be obtained using long chain alkane as substrate by microorganism conversion
It arrives, production process room temperature, normal pressure, it can be with large-scale production such as a variety of long-chain biatomic acids from C9 to C18.
Bioanalysis production long-chain biatomic acid has been investigated for many years.Scholar screens from oil field earliest can produce length
The bacterial strain of chain binary acid carries out mutagenesis to bacterial strain to improve the yield of long-chain biatomic acid.Some scholars also studied long-chain binary
Key enzyme in sour synthesis process.The document of foreign study long-chain biatomic acid is to carry out genetic modification to bacterial strain mostly, passes through resistance
Disconnected or reduction fatty acid beta oxidation related enzyme systems strengthen the enzyme system of fatty acid α-ω oxidation to improve the yield of product.It is open
Patent such as CN 1071951A, CN 1067725C, CN 1259424C, 200410018255.7,200610029784.6 etc. all
The method of Production of Long Chain Dicarboxylic Acids Through Micro-Biological Fermen is provided, these methods require during the fermentation, are especially producing acid
The pH value of fermentation liquid is adjusted to 7.0 or more by the phase, main reasons is that: first, fermentation strain candida tropicalis is in meta-alkali
There are higher enzyme activity, therefore alkaline condition high catalytic efficiency under property environment；Second, under alkaline environment, long-chain can be made
Binary acid exists with the salt form for being dissolved in the binary acid of water, so that keeping good mass transfer in fermentation process.Therefore, existing
It in the method for conventional fermenting and producing long-chain biatomic acid, requires in fermentation, especially produces sour phase (transition phase), fermentation system
PH value is greater than 7.0.Also, it during the fermentation, needs to add in a large amount of alkali as the accumulation of long-chain biatomic acid increases
With the long-chain biatomic acid of generation.In the fermentation liquid that fermentation finishes, long-chain biatomic acid is present in fermentation liquid in a salt form,
It needs that a large amount of acid is added to convert long-chain biatomic acid for the salt of long-chain biatomic acid in system again during extraction purification binary acid
Product, cause salt in fermentation process liquid system at concentrations up to 50000-70000ug/ml, these high-salt wastewaters are difficult to handle, right
It is very important challenge for environment, has seriously affected the development of bioanalysis long-chain biatomic acid industry.
Patent US 6569670B2 report in the environment of pH value 5.8 with fatty acid be raw material production long-chain biatomic acid
Method is particularly easy to blister, cause the main reason is that fatty acid can exist in the form of fatty acid salt in 7.0 or more pH value
Fermentation can not carry out, so must ferment at lower ph, but bacterial strain metabolism is slow under low ph value, production efficiency
It is low.
Summary of the invention
To overcome defect present in existing fermenting and producing long-chain biatomic acid technique, the first purpose of this invention is to mention
It is also highly active in the fermentation system below of pH value 7.0 for a kind of novel Candida tropicalis, it is applicable in very much
In fermenting and producing long-chain biatomic acid.
A second object of the present invention is to provide a kind of microbial inoculums including the Candida tropicalis.
Third object of the present invention is to provide a kind of applications of Candida tropicalis.
Fourth object of the present invention is to provide a kind of method of fermenting and producing long-chain biatomic acid.
Fifth object of the present invention is to provide a kind of preparation methods of long-chain biatomic acid.
Sixth object of the present invention is to provide a kind of long-chain biatomic acids.
Candida tropicalis (Candida tropicalis) bacterial strain CAT H1614 provided by the invention, preservation are compiled
Number be CCTCC NO:M 2013143.
The present invention also provides the microbial inoculums including Candida tropicalis bacterial strain CAT H1614 above-mentioned.
The present invention also provides Candida tropicalis bacterial strain CAT H1614 above-mentioned to prepare in long-chain biatomic acid in fermentation
Application.
The method of fermenting and producing long-chain biatomic acid provided by the invention uses Candida tropicalis bacterial strain CAT above-mentioned
H1614 carries out fermenting and producing long-chain biatomic acid.
In the method for fermenting and producing long-chain biatomic acid provided by the invention, the fermentation process includes the strain growth phase and turns
The change phase, wherein in the transition phase, the pH value for controlling fermentation system is 7.0 or less；Preferably 4.0~6.8；More preferably 5.0
~6.5.
Thallus in the method for fermenting and producing long-chain biatomic acid provided by the invention, after the strain growth to 30 times of dilution
Optical density OD620When greater than 0.5, the pH value for controlling fermentation system is 7.0 or less；Preferably 4.0~6.8；More preferably 5.0~
6.5。
In the method for fermenting and producing long-chain biatomic acid provided by the invention, the substrate for the fermentation process includes alkane
Hydrocarbon preferably includes the normal alkane of C9~C22；It more preferably include the normal alkane of C9~C18；Most preferably include C10, C11, C12,
The normal alkane of C13, C14, C15 or C16；
And/or the culture medium for the fermentation process includes following component (w/v): glucose 1%~5%, corn pulp
0.1%~0.9%, yeast extract 0.1%~0.5%, potassium nitrate 0.05%~1.2%, potassium dihydrogen phosphate 0.05%~1.0%,
Urea 0.05%~0.3%, ammonium sulfate 0.05%~0.3% and sodium chloride 0.05%~0.2%；Or including following component
(w/v): glucose 1%~5%, potassium nitrate 0.05%~0.6%, potassium dihydrogen phosphate 0.02%~0.6%, ammonium sulfate 0.05%
~0.3% and magnesium sulfate 0.05%~0.3%.
In the method for fermenting and producing long-chain biatomic acid provided by the invention, in the fermentation process, the inoculation of the bacterial strain
Amount is 10%~30%；
And/or temperature is 28~32 DEG C；
And/or air quantity is 0.3~0.7vvm；
And/or pressure is 0.05~0.14MPa；
And/or the strain growth is interim, the pH value of fermentation system is not less than 3.0, preferably 3.5~6.5.
The preparation method of long-chain biatomic acid provided by the invention includes:
S1: it ferments and long-chain biatomic acid fermentation liquid is made；And
S2: purification process is extracted to resulting long-chain biatomic acid fermentation liquid, the long-chain biatomic acid is made；
Wherein, the method system that the step S1 uses the described in any item fermenting and producing long-chain biatomic acids of preceding solution
Obtain the long-chain biatomic acid fermentation liquid.
In the preparation method of long-chain biatomic acid provided by the invention, the extraction purification processing of the step S2 includes: by institute
The long-chain biatomic acid fermentation liquid acidification obtained separates to obtain solid content, then in organic solvent by solid content dissolution, separates clearly
Liquid, crystallization, thus obtains long-chain biatomic acid.
In the preparation method of long-chain biatomic acid provided by the invention, the pH value of the acidification is 2.5~5, preferably 3~4；
And/or the isolated method is centrifugation or filtering；
And/or the organic solvent includes: the one or more of alcohol, acid, ketone and ester；Wherein, the alcohol include methanol,
One of ethyl alcohol, isopropanol and n-butanol are a variety of；The acid includes acetic acid；The ketone includes acetone；The ester includes second
Acetoacetic ester and/or butyl acetate；
And/or after the solid content is dissolved in organic solvent, decoloration, then clear liquid is separated to obtain, the method for the decoloration is preferred
Active carbon decoloring；The additive amount of the active carbon is no more than the 5% of liquor capacity；The temperature of the decoloration is 85~100 DEG C；
The time of the decoloration is 15~165min；
And/or the crystallization is decrease temperature crystalline；The decrease temperature crystalline is the following steps are included: be cooled to 65~80 DEG C, heat preservation
After 1~2 hour, then 25~35 DEG C are cooled to, crystallized；
And/or after the crystallization, crystal is isolated, long-chain biatomic acid is thus obtained；The isolated method is centrifugation point
From.
The present invention also provides the preparation methods of such as described in any item long-chain biatomic acids of aforementioned techniques technical solution to be made
Long-chain biatomic acid.
The present invention screens to have obtained new Candida tropicalis bacterial strain CAT H1614, can be lower than 7.0 in pH value
High vigor can be kept under environment for a long time, thus produces long-chain biatomic acid especially suitable for bioanalysis, produces acid fastly, yield is high, also
Fermentation period can be reduced.
Long-chain biatomic acid fermentation process provided by the invention and preparation method can ferment under acidic environment, on the one hand
A large amount of precipitatings particle (including a large amount of binary acid crystals) are directly obtained, lye in fermentation process on the one hand can be effectively reduced
The dosage of acid, simplifies the extraction process of dicarboxylic acid product when dosage and subsequent long-chain biatomic acid extract, and keeps long-chain biatomic acid raw
The amount of the salt generated in production. art is greatly reduced and (reduces to original and produce 1/10th or lower of salt content).In addition, due to
Candida tropicalis CAT H1614 has high catalysis activity, and process of the invention, which also has, to be shortened fermentation time, improves
Produce acid amount, reduce culture medium dosage, suitable for multiple types long-chain biatomic acid production many advantages, such as.With existing production technology phase
Than not only there is significant cost advantage, but also can effectively reduce the pressure to resource and environment, therefore have clearly
Industrial value advantage.
Detailed description of the invention
Fig. 1 is the preparation flow figure of candida tropicalis CAT H1614 bacterial strain.
Specific embodiment
The first aspect of the invention provides a kind of Candida tropicalis (Candida tropicalis) bacterial strain CAT
H1614, is deposited in China typical culture collection center (CCTCC), address Wuhan, China, Wuhan University, and deposit number is
CCTCC NO:M 2013143.
Candida tropicalis CAT H1614 provided by the invention can be by candida tropicalis CCTCC NO:M 203052
It is obtained after mutagenesis, screening, mutagenesis and screening process are as shown in Figure 1.
Detailed process can be with are as follows: accesses 203052 (starting strain) glycerol tube of candida tropicalis CCTCC NO:M and fills
Have in the shaking flask of YPD culture solution and be added 0.001%~0.02% 5 FU 5 fluorouracil, 28~32 DEG C, 200~250rpm concussion
Culture 16~for 24 hours.Take the medium centrifugal grown collect thallus and with brine several times, finally use equivalent LiCl
Aqueous solution suspension cell.Appropriate cell suspending liquid is taken to be placed in sterilized petri dishes using ultraviolet light irradiation mutagenesis, ultraviolet lamp power
15W, 20~30cm of irradiation distance, 60~120s of irradiation time.Cell suspending liquid dilution spread after taking mutagenesis contains YPD culture
The plate of base, 28~30 DEG C of 48~72h of culture, the well-grown monoclonal of picking (are wrapped to primary dcreening operation culture medium is equipped in culture medium
Include substrate long chain alkane) orifice plate, initial ph value is 6.0~6.8 or so, in the process with the accumulation of long-chain biatomic acid, pH value
Decline, does not add alkali in the process, and is to maintain nature pH, and generally 4.5~6.8, fermentation carries out at 28~32 DEG C, and shaking table turns
Speed 200~250rpm, fermentation period 60~72 hours, after fermentation, therefrom picking with respect to high yield long-chain biatomic acid bacterial strain,
Then these bacterial strains are subjected to secondary screening through orifice plate again, the bacterial strain of secondary screening is using 500ml shake flask fermentation secondary screening, 500ml shaking flask hair
Ferment carries out at 28~32 DEG C, 200~250rpm of shaking speed, and fermentation process pH chooses naturally, fermentation period 96~120 hours
The Candida tropicalis strain for selecting high yield long-chain biatomic acid, is stored in glycerol tube.
YPD culture medium is following component: 20g/L glucose, 10g/L yeast extract, 20g/L peptone and 2% fine jade
Rouge.
Primary dcreening operation and the culture medium of secondary screening include following component: glucose 1%~5%, corn pulp 0.1%~0.9%, yeast
Cream 0.1%~0.5%, potassium nitrate 0.05%~1.2%, potassium dihydrogen phosphate 0.05%~1.0%, urea 0.05%~0.3%,
Ammonium sulfate 0.05%~0.3% and sodium chloride 0.05%~0.2% (w/v).Primary dcreening operation and secondary screening can be used this field common
The specific ingredient of operating method, culture medium can also be adjusted or be selected according to process actual conditions by those skilled in the art.
Candida tropicalis bacterial strain CAT H1614 of the invention still can be protected for a long time in the environment of pH is lower than 7.0
High enzyme activity is held, thus produces long-chain biatomic acid especially suitable for bioanalysis, produces acid fastly, yield is high, moreover it is possible to reduce fermentation period.
The second aspect of the invention provides the microbial inoculum including the Candida tropicalis bacterial strain CAT H1614.It should
It may also include all kinds of nutriments of the conventional guarantee Candida tropicalis strain activity in this field in microbial inoculum.
The third aspect of the invention provides the application of the Candida tropicalis bacterial strain CAT H1614.The application
The application in long-chain biatomic acid is prepared in fermentation for Candida tropicalis bacterial strain CAT H1614.
The fourth aspect of the invention provides a kind of method of fermenting and producing long-chain biatomic acid, uses the above-mentioned torrid zone
Candida bacteria strain CAT H1614 ferments to produce long-chain biatomic acid.
In an embodiment of the method for fermenting and producing long-chain biatomic acid according to the present invention, the process of fermentation can be wrapped
Include strain growth phase and transition phase, wherein in the transition phase of fermentation, the pH value for controlling fermentation system is 7 hereinafter, preferably
4.0~6.8, more preferably 5.0~6.5.
Or can be, when strain growth to thallus optical density (OD620) when being greater than 0.5 (dilution 30 times), control fermentation body
The pH value of system is 7.0 hereinafter, it is preferred that 4.0~6.8, more preferably 5.0~6.5；Specific pH value example can be with are as follows: and 3,3.1,
3.2,3.3,3.4,3.5,3.6,3.7,3.8,3.9,4.0,4.1,4.2,4.3,4.4,4.5,4.6,4.7,4.8,4.9,5.0,
5.1,5.2,5.3,5.4,5.5,5.6,5.7,5.8,5.9,6.0,6.1,6.2,6.3,6.4,6.5,6.6,6.7,6.8,6.9,
7.0.The adjusting of pH value or control mode are not particularly limited, and can be a certain pH value of permanent control, do not control pH value, not less than certain pH
Value, not higher than certain pH value, up-regulation pH value, downward ph adjustment, outside range control enter or naturally into the pH value range
One of mode or a variety of combinations.The present invention for pH value regulation method also without limitation, fermentation arts can be used
The lye of debita spissitudo is such as added in conventional means.
In an embodiment of the method for fermenting and producing long-chain biatomic acid according to the present invention, microbe conversion process can
It is carried out in fermentation medium or buffer solution.Specifically, working as Candida tropicalis (Candida tropicalis) bacterium
OD after strain CAT H1614 is cultivated to mycelium dilution thirtyfold620Greater than 0.5, start at this point, long chain alkane substrate can be initially added into
Microbe conversion.When microbe conversion process carries out in fermentation medium, substrate can directly be added into culture medium, it can also be by bacterium
Strain is transferred in fermentor, adds fermentation medium and substrate carries out microbe conversion；When microbe conversion process is in buffer solution
When middle progress, first the thallus that culture obtains can be separated, gone in buffer solution, and substrate is added and carries out microbe conversion.
In an embodiment of the method for fermenting and producing long-chain biatomic acid according to the present invention, the substrate of fermentation includes
Alkane, preferably includes the normal alkane of C9~C22, more preferably includes the normal alkane of C9~C18, most preferably include C10, C11, C12,
The normal alkane of C13, C14, C15 or C16.
In an embodiment of the method for fermenting and producing long-chain biatomic acid according to the present invention, when in fermentation medium
When middle progress microbe conversion, the ingredient for the culture medium used that ferments optionally includes carbon source, nitrogen source, inorganic salts, trophic factors etc..
Wherein, carbon source can be Candida fermentable sugar, comprising: glucose, sucrose, maltose etc. it is one or more, it is described
The additive amount of carbon source can be 1%~10% (w/v)；Nitrogen source can be organic nitrogen and/or inorganic nitrogen, and organic nitrogen includes: yeast
One of cream, peptone, corn pulp are a variety of, and inorganic nitrogen includes: one of urea, ammonium sulfate, potassium nitrate or a variety of, institute
The additive amount for stating nitrogen source can be 0.1%~3% (w/v)；Inorganic salts include: potassium dihydrogen phosphate, potassium chloride, magnesium sulfate, chlorination
One of calcium, iron chloride, copper sulphate are a variety of, and the additive amount of the inorganic salts can be 0.1%~1.5% (w/v)；Nutrition
The factor includes: vitamin B1, vitamin B2, one of vitamin C, biotin or a variety of, the additive amount of the trophic factors
It can be 0%~1% (w/v).
In a preferred embodiment of the method for fermenting and producing long-chain biatomic acid according to the present invention, one kind can be selected
Water-soluble liquid culture medium (hereinafter referred to as " fermentation medium 1 ") comprising following component: glucose 1%~5%, corn pulp 0.1%
~0.9%, yeast extract 0.1%~0.5%, potassium nitrate 0.05%~1.2%, potassium dihydrogen phosphate 0.05%~1.0%, urea
0.05%~0.3%, ammonium sulfate 0.05%~0.3% and sodium chloride 0.05%~0.2% (w/v).Fermentation medium 1 can be with
Suitable for the shaking flask from tens milliliters to the fermenting and producing of several hundred tons of fermentor scales.
In another preferred embodiment of the method for fermenting and producing long-chain biatomic acid according to the present invention, according to fermentation
The water-soluble liquid culture medium (hereinafter referred to as " fermentation medium 2 ") of low nutrition object concentration also can be selected in the characteristics of pH value of system,
Including following component: glucose 1%~5%, potassium nitrate 0.05%~0.6%, potassium dihydrogen phosphate 0.02%~0.6%, sulfuric acid
Ammonium 0.05%~0.3% and magnesium sulfate 0.05%~0.3% (w/v).Fermentation medium 2 reduces the ingredient of some complexity such as
The ingredient of corn pulp, yeast extract etc., culture medium is more clear, can preferably control fermentation index and product quality.Its available water
It prepares, sterilize 20min at 121 DEG C, is cooled to suitable temperature, uses as fermented and cultured.
In an embodiment of the method for fermenting and producing long-chain biatomic acid according to the present invention, during microbe conversion
Fermentation conversion rate can also be further promoted by adding sugar aqueous solution as carbon source.The mode that sugar aqueous solution is added can be batch
It is secondary to add, or continuous flow adds.Sugar can be common sucrose or glucose, and the concentration for adding sugar aqueous solution can be
10%~70% (w/v).By adding sugar aqueous solution, the sugared concentration 0.1%~1% (w/v) of microbe conversion system can control.
In an embodiment of the method for fermenting and producing long-chain biatomic acid according to the present invention, when microbe conversion process
When carrying out in buffer solution, buffer solution can be phosphate buffer solution.In a preferred embodiment, buffer solution
It can be potassium dihydrogen phosphate-disodium hydrogen phosphate buffer solution.
In an embodiment of the method for fermenting and producing long-chain biatomic acid according to the present invention, fermentation condition can be controlled
Be made as: the inoculum concentration of Candida tropicalis bacterial strain CAT H1614 can be 10%~30%；In another embodiment,
Temperature can be 28~32 DEG C；In another embodiment, air quantity can be 0.3~0.7vvm；In another embodiment
In, pressure can be 0.05~0.14MPa；In another embodiment, dissolved oxygen is not less than 10% during microbe conversion.
In an embodiment of the method for fermenting and producing long-chain biatomic acid according to the present invention, in the strain growth phase,
When strain culturing is grown, the pH value of system is not less than 3.0, preferably 3.5~6.5.
In a preferred embodiment of the method for fermenting and producing long-chain biatomic acid according to the present invention, it may include following
Step:
Seed bottle culture process:
The glycerol tube strain of candida tropicalis is taken to be inoculated in the seed bottle equipped with YPD culture solution, pH is naturally, 28~32
At DEG C, 200~250rpm shaking table culture 1~2 day.
Seed tank culture technique:
Take shake-flask seed access equipped in the seeding tank of seed culture medium, inoculum concentration is 10%~30%, is inoculated with post-fermentation
The initial ph value of system is at 6.0~6.8,28~32 DEG C, and 0.3~0.7vvm of ventilation quantity, tank presses 0.05~0.14MPa, is kept
The DO of certain mixing speed, control seed culture process is not less than 10%, cultivates 15~30h, cultivates the standard of mature seed
OD after being 30 times of dilution620For greater than 0.5, more excellent is OD620It is 0.5~1.0.
Include following ingredient it is preferable to use aqueous solution seed culture medium: sucrose 1%~3%, corn pulp 0.15%~1%,
Yeast extract 0.2%~1.5%, KH2PO40.4%~1.5%, urea 0.05%~0.5%.
Microbe conversion technique:
It will be inoculated into the fermentor containing fermentation medium in the resulting seed liquor of seed tank culture, inchoate aspect after inoculation
Product is 4~6L, inoculum concentration 10%~30% (v/v, opposite ferment initial volume), and fermentation starting addition 0~10% (v/v, relatively
Ferment initial volume) alkane, 28~32 DEG C of fermentation processes temperature, ventilation quantity is about 0.3~0.7vvm, tank pressure (gauge pressure)
About 0.05~0.14MPa, keeps certain mixing speed, and control dissolved oxygen is not less than 10%.Control the pH value of fermentation liquid, fermentation
Originating pH is about 5.0~6.8, and with the growth of microorganism, the pH of fermentation liquid gradually declines, and control pH is not less than 3.0, to thallus
Optical density (OD620) when being greater than 0.5 (dilution 30 times), control pH value is 7.0 hereinafter, preferably about 4.0~6.8, more preferably
5.0~6.5, until fermentation ends.Start batch when fermentation period is 10~20 hours and alkane is added, controls alkane in fermentation liquid
Hydrocarbon content is no more than 10%, and total fermentation period is about 100~180 hours.Optionally, by adding sugar aqueous solution in fermentation process
Fermentation liquid sugar concentration is controlled in 0.1%~1% (w/v).
Or be centrifuged the resulting seed liquor of seed tank culture, the conversion tank containing phosphate buffer solution is added in thallus
In, thallus dosage 10%~30% (v/v, opposite initial volume of fermenting), alkane is added in batch in the process, controls alkane in conversion fluid
Hydrocarbon is no more than 10%, controls 28~32 DEG C of temperature, and ventilation quantity is about 0.3~0.7vvm, and tank pressure (gauge pressure) is about 0.05~
0.14MPa, keeps certain mixing speed, and control dissolved oxygen is not less than 10%.Add 10%~40% liquid alkaline control conversion fluid
PH value be 7.0 hereinafter, preferably 4.0~6.8, more preferably 5.0~6.5, until conversion terminates, total transformation period is about
100~180 hours.
The fifth aspect of the invention provides a kind of preparation method of long-chain biatomic acid, comprising the following steps:
S1: it ferments and long-chain biatomic acid fermentation liquid is made；And
S2: purification process is extracted to resulting long-chain biatomic acid fermentation liquid, the long-chain biatomic acid is made；
Wherein, step S1 is made described using the method for the fermenting and producing long-chain biatomic acid of the fourth aspect of the invention
Long-chain biatomic acid fermentation liquid.
In the preparation method of long-chain biatomic acid of the invention, existing mention can be used to resulting long-chain biatomic acid fermentation liquid
Purifying process is taken final long-chain biatomic acid product is made.
In an embodiment of the preparation method of long-chain biatomic acid according to the present invention, at the extraction purification of step S2
Reason may include following procedure: resulting long-chain biatomic acid fermentation liquid being acidified, separates to obtain solid content, then the solid content is dissolved
In organic solvent, clear liquid is separated to obtain, crystallizes, thus obtains long-chain biatomic acid.
In one preferred embodiment, the pH value of acidification can be 2.5~5, preferably can be 3~4；At another
In preferred embodiment, the separate mode that solid content is separated after acidification can be the modes such as centrifugation, filtering；It is preferred at another
Embodiment in, for dissolved solid organic solvent can for alcohol, acid, ketone and ester one or more, alcohol therein
Including but not limited to methanol, ethyl alcohol, isopropanol, n-butanol etc., acid therein includes but is not limited to acetic acid etc., and ketone therein includes
But be not limited to acetone etc., ester therein includes but is not limited to ethyl acetate, butyl acetate etc.；In another preferred embodiment
In, solid content decolourizes after being dissolved in organic solvent, then isolated clear liquid, and it is preferable to use active carbons to take off for the method for decoloration
Color, the additive amount of active carbon are no more than the 5% of liquor capacity, and the temperature of decoloration is 85~100 DEG C, the time of decoloration is 15~
165min；In another preferred embodiment, after separating clear liquid, decrease temperature crystalline, decrease temperature crystalline can comprise the following steps that head
After being first cooled to 65~80 DEG C, heat preservation 1~2 hour, then 25~35 DEG C are cooled to, crystallized；In another preferred embodiment
In, after crystallization, gained crystal is isolated, long-chain biatomic acid is thus obtained, the mode for separating crystal can be centrifuge separation.
The sixth aspect of the invention provides long-chain biatomic acid product made from the preparation method of foregoing long-chain binary acid.
Long-chain biatomic acid (LCDA of the present invention；Also referred to as long chain dicarboxylic acid and long chain diacid) it include chemical formula HOOC
(CH2) nCOOH binary acid, wherein n >=7；Preferably, 20 >=n >=7；It is highly preferred that 16 >=n >=7.It is of the present invention
The example of LCDA includes:: azelaic acid (HOOC (CH2)7COOH), decanedioic acid (HOOC (CH2)8COOH), eleven carbon diacids
(HOOC(CH2)9COOH, 1,9- nine carbon dicarboxylic acids or 1,11- eleven carbon diacids, the present invention in label be DC11 "), 12 carbon
Binary acid (HOOC (CH2)10COOH, 1,10- ten carbon dicarboxylic acids or 1,12- dodecanedicarboxylic acid, the present invention in label for
" DC12 "), tridecanyldicarboxylic acid (HOOC (CH2)11COOH, 1,11- ten one carbon dicarboxylic acids or 1,13- tridecanyldicarboxylic acid, this hair
Bright middle label is DC13 "), tetradecane diacid (HOOC (CH2)12COOH, 1,12- ten two carbon dicarboxylic acids or 1,14 carbon of 14-
Binary acid, the present invention in label be DC14 "), pentadecane binary acid (HOOC (CH2)13COOH, 1,13- ten three carbon dicarboxylic acids or
Label is DC15 "), 16-dicarboxylic acid (HOOC (CH in 1,15- pentadecane binary acid, the present invention2)14COOH, 1,14- ten four
Carbon dicarboxylic acids or 1,16- 16-dicarboxylic acid, the present invention in label be DC16 "), seventeen carbon diacids (HOOC (CH2)15COOH, 1,15- pentadecane dicarboxylic acids or 1,17- seventeen carbon diacids, the present invention in label be DC17 "), octadecane diacid
(HOOC(CH2)16COOH, 1,16- ten six carbon dicarboxylic acids or 1,18- octadecane diacid, label is DC18 " in the present invention) etc..
In some embodiments of the method for fermenting and producing long-chain biatomic acid according to the present invention, in 10L ferment tank
In technique, production acid the amount at least eleven carbon diacids of 110mg/g, at least dodecanedicarboxylic acid of 150mg/g, at least can produce
The pentadecane binary acid of the tetradecane diacid of the tridecanyldicarboxylic acid of 130mg/g, at least 150mg/g, at least 140mg/g or
At least 16-dicarboxylic acid of 130mg/g.
In some embodiments of the method for fermenting and producing long-chain biatomic acid according to the present invention, in 10L ferment tank
In technique, compared with traditional handicraft, at least 60% base amount can be saved；In other embodiments, 90% or so can be saved
Base amount.
In some embodiments of the method for fermenting and producing long-chain biatomic acid according to the present invention, converted in 10L fermentor
In technique, the production acid amount at least dodecanedicarboxylic acid of 150mg/g can produce, and reach 92% or more weight conversion rate (w/
W, alkane are converted to the weight percent of binary acid).
In some embodiments of the method for fermenting and producing long-chain biatomic acid according to the present invention, in 200M3Fermentor hair
In ferment technique, the production acid amount at least dodecanedicarboxylic acid of 150mg/g can produce, and reach 92% or more weight conversion rate
(w/w, alkane are converted to the weight percent of binary acid).
It, can be secondary by adding in some embodiments of the method for fermenting and producing long-chain biatomic acid according to the present invention
Carbon source improves fermentation conversion rate, and can achieve 95% or more weight conversion rate, (w/w, alkane are converted to the weight of binary acid
Percentage).
In some embodiments of the method for fermenting and producing long-chain biatomic acid according to the present invention, in 200M3Fermentor hair
In ferment technique, the production acid amount at least tridecanyldicarboxylic acid of 140mg/g can produce, and reach 85% or more weight conversion rate
(w/w, alkane are converted to the weight percent of binary acid).
In some embodiments of the method for fermenting and producing long-chain biatomic acid according to the present invention, in 450M3Fermentor hair
In ferment technique, the production acid amount at least dodecanedicarboxylic acid of 150mg/g can produce, and reach 90% or more weight conversion rate
(w/w, alkane are converted to the weight percent of binary acid).
Below by embodiment, the present invention is described in detail, so that the features and advantages of the present invention become apparent from.But it answers
This points out that for embodiment for understanding design of the invention, the scope of the present invention is not limited only to reality listed herein
Apply example.
It is such as not particularly illustrated, according to fermentation arts common sense, heretofore described percentage is mass volume ratio, it may be assumed that w/
v；% indicates g/100mL.
The embodiment of the present invention and comparative example use technology well known to those skilled in the art, such as Chinese patent ZL
Measuring method disclosed in 95117436.3 measures the binary acid concentration in fermentation liquid, specific continuous mode are as follows: with hydrochloric acid solution tune
The pH to 3.0 of fermentation liquid is saved, then plus 100mL ether is for the binary acid in extractive fermentation liquid, then uses evaporation to remove second
Ether obtains binary acid powder, in ethanol by the dissolution of obtained binary acid powder, and is titrated with the NaOH solution of 0.1mol/L,
Finally obtain the binary acid titer in fermentation liquid.
The specific continuous mode of the fermentation treatment fluid salt content of the embodiment of the present invention and comparative example are as follows: set fermentation treatment fluid
In drying in the glass evaporating dish after constant weight, water bath method.If residue has a color, hydrogen peroxide is added dropwise to bubble collapse, then
Water bath method is handled several times repeatedly, and until color bleaches or colour stable is constant, the evaporating dish being evaporated is dried to perseverance
Weight, weighing calculate salt content.
The acquisition of 1 candida tropicalis CAT H1614 bacterial strain of embodiment
The access of 203052 glycerol tube of starting strain CCTCC NO:M is equipped in the shaking flask of YPD culture solution and is added
0.01% 5 FU 5 fluorouracil, 30 DEG C, 200rpm shake culture 16~for 24 hours.The medium centrifugal grown is taken to collect thallus simultaneously
Several times with brine, the LiCl aqueous solution suspension cell of equivalent is finally used.Appropriate cell suspending liquid is taken to be placed in sterile flat
Ultraviolet light irradiation mutagenesis, ultraviolet lamp power 15W, irradiation distance 20cm, irradiation time 90s are used in ware.By the bacterial strain of mutagenesis
It is cultivated on the plate for including YPD culture medium, 29 DEG C of cultivation temperature, incubation time 50h, the well-grown Dan Ke of picking
The grand orifice plate extremely equipped with primary dcreening operation culture medium (including substrate long chain alkane in culture medium), ferments, does not add alkali in the process, and
It is to maintain natural ph, fermentation carries out at 29 DEG C, shaking speed 220rpm, fermentation period 70 hours, after fermentation, carries out
Primary dcreening operation.Therefrom then these bacterial strains are carried out secondary screening, the bacterium of secondary screening through orifice plate again with respect to the bacterial strain of high yield long-chain biatomic acid by picking
Strain is using 500ml shake flask fermentation secondary screening, and 500ml shake flask fermentation carries out at 29 DEG C, shaking speed 220rpm, fermentation process pH
Naturally, fermentation period 110 hours.Stable, high-yielding bacterial strain is finally obtained, CAT H1614 is named as, is stored in glycerol tube.Process
Referring to Fig.1.
2~7 candida tropicalis CAT H1614 of embodiment produces different binary acid in 10L fermentation cylinder for fermentation
The glycerol tube strain of candida tropicalis CAT H1614 is taken to be inoculated in equipped with 30ml YPD fluid nutrient medium (grape
Sugar 2%, yeast extract 1%, peptone 2%) seed bottle in, pH is naturally, at 29 DEG C, 220rpm shaking table culture 1 day.Take shaking flask kind
Son access is equipped with 5L seed culture medium (sucrose 2%, corn pulp 0.3%, yeast extract 0.5%, KH2PO40.8%, urea 0.3%)
Seeding tank in, inoculum concentration 10%, after inoculation the initial ph value of system be 6.0,29 DEG C at, ventilation quantity 0.4vvm, tank pressure
0.08MPa cultivates 18h, and pH drops to 3, OD naturally in incubation620It is long to 0.7 when be inoculated into containing 6L fermentation medium 1
(glucose 4%, corn pulp 0.5%, yeast extract 0.4%, potassium nitrate 1%, potassium dihydrogen phosphate 0.1%, urea 0.12%, sulfuric acid
Ammonium 0.06% and sodium chloride 0.1%) fermentor in, after inoculation initial volume be 5L, inoculum concentration 20%, fermentation processes
30 DEG C of temperature, ventilation quantity is about 0.4vvm, and tank pressure (gauge pressure) is about 0.12MPa, and control dissolved oxygen is not less than 20%.Add 30%
Liquid alkaline controls the pH value of fermentation liquid, and earlier fermentation is mainly based on thalli growth, and fermentation starting pH is about 6.5, with microorganism
Growth, the pH of fermentation liquid gradually declines, and control pH is not less than 3.0, to thallus optical density (OD620) it is greater than 0.5 (30 times of dilution)
When, control pH is about 5.5, until fermentation ends, start batch when fermentation period is 10~20 hours and alkane, control hair is added
Determination of Alkane Content is no more than 10% in zymotic fluid, and total fermentation period is 152 hours.
Embodiment 2~7, which is respectively as follows:, (is implemented hendecane hydrocarbon (embodiment 2), 12 carbon alkanes by above-mentioned zymotechnique
Example 3), triclecane (embodiment 4), tetradecane hydrocarbon (embodiment 5), pentadecane alkane (embodiment 6), hexadecane hydrocarbon
(embodiment 7) carries out fermentation and corresponding long-chain biatomic acid, yield result such as table 1 is made.
The fermentation results of 1 candida tropicalis CAT H1614 of table
As seen from the results in Table 1, the candida tropicalis CAT H1614 of the invention long chain alkane that ferments at low ph values can be made
Standby corresponding long-chain biatomic acid, and yield is higher, adapts to the preparation of a variety of long-chain biatomic acids.
8 candida tropicalis CAT H1614 of embodiment produces long-chain biatomic acid in 10L fermentation cylinder for fermentation
The glycerol tube strain of candida tropicalis CAT H1614 is taken to be inoculated in equipped with 25mLYPD fluid nutrient medium (glucose
2%, yeast extract 1%, peptone 2%) seed bottle in, pH is naturally, at 30 DEG C, 230rpm shaking table culture 2 days.Take shake-flask seed
Access is equipped with 6L seed culture medium (sucrose 2%, corn pulp 0.3%, yeast extract 0.5%, KH2PO40.8%, urea 0.3%)
In 10L seeding tank, inoculum concentration 20%, the initial ph value of system is ventilation quantity 0.5vvm at 6.2,30 DEG C, tank pressure after inoculation
0.1MPa cultivates 20h, and pH drops to 3 naturally in incubation.OD620It is long to 0.6 when be inoculated into containing 1 (Portugal of fermentation medium
Grape sugar 2%, corn pulp 0.2%, yeast extract 0.2%, potassium nitrate 0.08%, potassium dihydrogen phosphate 0.3%, urea 0.2%, ammonium sulfate
0.1% and sodium chloride 0.1%) fermentor in, after inoculation initial volume be 6L, inoculum concentration 15%, fermentation starting addition 6%
12 carbon alkanes of (v/v, opposite initial volume of fermenting), 28 DEG C of fermentation processes temperature, ventilation quantity is about 0.4vvm, tank pressure
(gauge pressure) is about 0.11MPa, and control dissolved oxygen is not less than 20%, adds the pH value of 32% liquid alkaline control fermentation liquid.Earlier fermentation
Mainly based on thalli growth, fermentation starting pH is about 6.6, and with the growth of microorganism, the pH of fermentation liquid gradually declines, control
PH is not less than 3.0, to thallus optical density (OD620) when being greater than 0.5 (dilution 30 times), pH 5.0 is controlled, until fermentation ends,
Fermentation period starts batch and alkane is added when being 10~20 hours, control Determination of Alkane Content in fermentation liquid and be no more than 10%.
1 candida tropicalis CCTCC NO:M 203052 (starting strain) of comparative example is produced in 10L fermentation cylinder for fermentation to be grown
Chain binary acid
Candida tropicalis CCTCC NO:M 203052 (starting strain) replacement bacterial strain CAT H1614 also uses embodiment 8
Same process fermentation.
2 candida tropicalis CCTCC NO:M 203052 (starting strain) of comparative example is in 10L fermentor using traditional work
Skill fermenting and producing long-chain biatomic acid
Candida tropicalis CCTCC NO:M 203052 (starting strain) is fermented using existing traditional handicraft: seeding tank
The mature seed of culture is inoculated into containing fermentation medium (glucose 3%, potassium dihydrogen phosphate 0.5%, yeast extract 0.2%, corn pulp
0.15%, urea 0.25%, sodium chloride 0.2%, potassium nitrate 0.7%) fermentor in, C12 alkane and feed supplement sugar disappear.In 29
It is cultivated under the conditions of DEG C ventilatory capacity 0.5vvm, tank pressure 0.1Mpa.Preceding 20 hours pH ferment naturally, based on thalli growth, works as thallus
Grow optical density (OD620) it is greater than 0.6, start batch and add C12 alkane, hereafter adds once, controlled in fermentation liquid for every eight hours
Alkane concentration maintains the left and right 5% (V:V), while adjusting pH to 6.5, after 48 hours, adjusts pH with NaOH solution within every 4 hours
To 7.0,48~72 hours, pH to 7.5 is adjusted with NaOH solution within every 4 hours, 72~120 hours, every 4 hours with NaOH solution tune
PH to 7.8 is saved, to tank is put, adjusts pH to 8.0 with NaOH solution within every 4 hours within 120 hours.Fermentation to 24,48,72 hours batch are mended
Add the glucose of 1% (W:V).
The fermentation results of above-mentioned fermentation process are shown in Table 2.
Fermentation results of 2 different strains of table under different process
As shown in Table 2, fermentation results are substantially better than routine to candida tropicalis CAT H1614 of the invention at low ph values
Technique and existing bacterial strain, and high catalysis activity can be kept for a long time.
9 candida tropicalis CAT H1614 of embodiment ferments DC12 in 10L tank
The glycerol tube strain of candida tropicalis CAT H1614 is taken to carry out seed culture according to 8 the method for embodiment.
OD620It is long to 0.8 when, seed liquor is centrifuged, thallus is added in the conversion tank containing 6L phosphate buffer solution, thallus dosage
25% (v/v, opposite initial volume of fermenting), 12 carbon alkanes are added in batch in the process, control alkane in conversion fluid and are no more than
10%, 29 DEG C of temperature are controlled, ventilation quantity is about 0.5vvm, and tank pressure (gauge pressure) is about 0.12MPa, and control dissolved oxygen is not less than 10%,
The pH value 6.2 of 28% liquid alkaline control conversion fluid is added, until conversion terminates, total transformation period is 154 hours, produces acid 152mg/
G consumes lye 180mL, and binary acid is to alkane weight conversion rate 92.4%.
10 candida tropicalis CAT H1614 of embodiment is in 200M3Fermentation cylinder for fermentation DC12
The glycerol tube strain for taking candida tropicalis CAT H1614 carries out seed culture according to 8 the method for embodiment,
PH drops to 3 naturally in incubation.OD620It is long to 0.8 when be inoculated into containing fermentation medium 2 (glucose 4%, potassium nitrate
0.1%, potassium dihydrogen phosphate 0.1%, ammonium sulfate 0.1% and magnesium sulfate 0.1%) fermentor in, initial volume is after inoculation
6L, inoculum concentration 22%, 12 carbon alkanes of fermentation starting addition 4% (v/v, opposite initial volume of fermenting), fermentation processes
28 DEG C of temperature, ventilation quantity is about 0.6vvm, and tank pressure (gauge pressure) is about 0.10MPa, and control dissolved oxygen is not less than 20%.Adding concentration is
The pH value of 33% liquid alkaline control fermentation liquid, for earlier fermentation mainly based on thalli growth, fermentation starting pH is about 6.7, with
The pH of the growth of microorganism, fermentation liquid gradually declines, and control pH is not less than 3.0, to thallus optical density (OD620) (dilute greater than 0.5
Release 30 times) when, control pH is about 6.0, starts stream plus concentration as 25% sugar juice, controls fermentation liquid sugar concentration 0.5%.It is sending out
The ferment period starts batch and alkane is added when being 10~20 hours, control Determination of Alkane Content in fermentation liquid and be no more than 10%.Total fermentation week
Phase 122h produces acid 182.3mg/g, and binary acid is to alkane weight conversion rate 100.4%, and 2.5 tons of caustic dosage.
11 candida tropicalis CAT H1614 of embodiment is in 200M3Fermentation cylinder for fermentation DC13
The glycerol tube strain for taking candida tropicalis CAT H1614 carries out seed culture according to 8 the method for embodiment,
PH drops to 3 naturally in incubation.OD620It is long to 0.8 when be inoculated into containing fermentation medium 2 (glucose 3.8%, potassium nitrate
0.12%, potassium dihydrogen phosphate 0.12%, ammonium sulfate 0.12% and magnesium sulfate 0.12%) fermentor in, inoculum concentration 25%, hair
The triclecane of ferment starting addition 4% (v/v, opposite initial volume of fermenting), 28 DEG C of fermentation processes temperature, ventilation quantity is about
For 0.6vvm, tank pressure (gauge pressure) is about 0.10MPa, and control dissolved oxygen is not less than 20%.Add the liquid alkaline control hair that concentration is 30%
The pH value of zymotic fluid, earlier fermentation is mainly based on thalli growth, and fermentation starting pH is about 6.6, with the growth of microorganism, fermentation
The pH of liquid gradually declines, and control pH is not less than 3.0, to thallus optical density (OD620) when being greater than 0.5 (dilution 30 times), control pH is about
It is 6.5, until fermentation ends, start batch when fermentation period is 10~20 hours and alkane is added, control alkane in fermentation liquid
Content is no more than 10%.Total fermentation period 132h, produces acid 147.3mg/g, and binary acid adds alkali to alkane weight conversion rate 85.4%
3.7 tons of amount.
12 candida tropicalis CAT H1614 of embodiment is in 450M3Fermentation cylinder for fermentation DC12
The glycerol tube strain for taking candida tropicalis CAT H1614 carries out seed culture according to 8 the method for embodiment,
PH drops to 3 naturally in incubation.OD620It is long to 0.8 when be inoculated into containing fermentation medium 2 (glucose 3.3%, potassium nitrate
0.15%, potassium dihydrogen phosphate 0.15%, ammonium sulfate 0.15% and magnesium sulfate 0.15%) fermentor in, inoculum concentration 23%, hair
12 carbon alkanes of ferment starting addition 5% (v/v, opposite initial volume of fermenting), 28 DEG C of fermentation processes temperature, ventilation quantity is about
For 0.5vvm, tank pressure (gauge pressure) is about 0.10MPa, and control dissolved oxygen is not less than 20%.Add the liquid alkaline control hair that concentration is 33%
The pH value of zymotic fluid, earlier fermentation is mainly based on thalli growth, and fermentation starting pH is about 6.5, with the growth of microorganism, fermentation
The pH of liquid gradually declines, and control pH is not less than 3.0, to thallus optical density (OD620) when being greater than 0.5 (dilution 30 times), control pH is about
It is 6.2, until fermentation ends, start batch when fermentation period is 10~20 hours and alkane is added, control alkane in fermentation liquid
Content is no more than 10%.Total fermentation period 142h, produces acid 160.8mg/g, and binary acid adds alkali to alkane weight conversion rate 93.8%
5 tons of amount.
The preparation of 13 long-chain biatomic acid product of embodiment
It by the sulphur acid for adjusting pH value to 3 of fermentation liquid made from embodiment 2, is acidified, is centrifugated to obtain solid content, then will
Solid content is dissolved in acetic acid, and the active carbon for being no more than supernatant volume 5% is added, and decolourize 60min under the conditions of 90 DEG C, filtering point
From clear liquid is obtained, clear liquid is cooled to 80 DEG C of heat preservation 1.5h, then be cooled to 30 DEG C, crystallizes, be centrifugated to obtain dicarboxylic acid product.
The preparation of 14 long-chain biatomic acid product of embodiment
It by the sulphur acid for adjusting pH value to 3.5 of fermentation liquid made from embodiment 3, is acidified, is centrifugated to obtain solid content, then
In ethanol by solid content dissolution, the active carbon for being no more than supernatant volume 5% is added, decolourize 75min under the conditions of 95 DEG C, filtering
Clear liquid is separated to obtain, clear liquid is cooled to 80 DEG C of heat preservation 1h, then be cooled to 35 DEG C, crystallizes, is centrifugated to obtain dicarboxylic acid product.
The preparation of 15 long-chain biatomic acid product of embodiment
It by the sulphur acid for adjusting pH value to 3.2 of fermentation liquid made from embodiment 11, is acidified, filters to obtain solid content, then will
Solid content is dissolved in acetic acid, and the active carbon for being no more than supernatant volume 5% is added, and decolourize 70min under the conditions of 85 DEG C, filtering point
From clear liquid is obtained, clear liquid is cooled to 65 DEG C of heat preservation 1h, then be cooled to 35 DEG C, crystallizes, be centrifugated to obtain dicarboxylic acid product.
Candida tropicalis CAT H1614 utilizes present invention process fermentation and candida tropicalis CCTCC NO:M
203052 (starting strains) utilize the comparison of salt content in the fermentation liquid fermentation treatment fluid of traditional handicraft fermentation
The fermentation liquid of Example 8 and comparative example 2 adds sulphur acid for adjusting pH to 3.0, removes solid content, obtain supernatant (fermentation
Treatment fluid), supernatant is placed in the glass evaporating dish after drying constant weight, water bath method.If residue has color, peroxide is added dropwise
Change hydrogen to bubble collapse, then water bath method, is handled several times, until color bleaches or colour stable is constant repeatedly.It will be evaporated
Evaporating dish drying to constant weight, weighing.Calculate salt content.It the results are shown in Table 3.
Salt content under 3 different strains difference pH of table in fermentation gained fermentation treatment fluid
| Group | Strain | Technique | Salt content (ppm) in fermentation treatment fluid |
| Embodiment 8 | Candida tropicalis CAT H1614 | Present invention process | 6800 |
| Comparative example 2 | Starting strain M 203052 | Existing traditional handicraft | 68750 |
As seen from the results in Table 3, candida tropicalis CAT H1614 of the invention ferments under conditions of pH value is lower than 7, institute
In fermentation liquid salt content have and obviously decline, can significantly mitigate subsequent purifying process, thus bring production cost and
The reduction of environmental pressure.
3 candida tropicalis H5343ALK2-1 fermenting and producing DC13 of comparative example
Culture medium (it include: glucose 27.0g/L, ammonium sulfate 7.0g/L, potassium dihydrogen phosphate 5.1g/L, magnesium sulfate 0.5g/L,
Calcium chloride 0.1g/L, citric acid 0.06g/L, ferric trichloride 0.023g/L, biotin 0.0002g/L, trace metal: boric acid
0.0009g/L, copper sulphate 0.00007g/L, potassium iodide 0.00018g/L, ferric trichloride 0.00036g/L, manganese sulfate
0.00072g/L, sodium molybdate 0.00036g/L, zinc sulfate 0.00072g/L, SAG471 anti-foaming agent 0.8ml, water surplus) with appropriate
Mode carry out heat sterilization to avoid occurring precipitation reaction, be added in the fermentor of sterilizing after cooling.It is nonvaccinated complete
Full culture medium is found very limpid and is shallow bale of straw, not strong smell.Slight muddiness is generated after anti-foaming agent is added.
Above-mentioned culture medium is aseptically utilized, is cultivated by being stirred, ventilating to the fermentor that original liquid volume is 12L
Candida tropicalis H5343ALK2-1.The aseptic culture medium is vaccinated connecing for 5% candida tropicalis H5343ALK2-1
Kind object, and stir culture about 10 is small under conditions of 35 DEG C, pH5.8 and Ventilation Rate are enough that dissolved oxygen is made to be maintained at 20% or more
When.When the culture terminates exponential growth, dissolved oxygen is begun to ramp up, and transition phase starts in the following manner: with 0.7g/L/
The raw material flow that the rate of hour starts continuously to add Exxon Developmental Fluid137 is (a kind of to contain about 94.4% ten
The hydrocarbon of three alkane, remaining be mainly dodecane), the raw material flow and fermentation auxiliary element 1.25%Emersol 267 (one
The oleic acid of kind industrial grade) and a kind of 1.25%Emery 2203 (methyl grease (tallowate) of industrial grade) mixing.With
This simultaneously, the temperature in fermentor is down to 30 DEG C by 35 DEG C, and Ventilation Rate is down to 0.4vvm, and is applied with 0.4 bar to tank
Counter-pressure.PH is maintained between 5.8-5.9 using the KOH of 6N in growth period and transition phase.When biomass density reaches about
When 10g/L, start continuously to add glucose to fermentor with the rate of 1.58g glucose/L/ hours.According to microscopical daily
Observation result and the assessment steeped to the storage accumulated in yeast cells, the rate of adding of glucose described in transition phase is reduced to
0-15%.PPG (polypropylene glycol) anti-foaming agent of 7ml is added to fermentor in transition phase to control slight foam.Transition phase
After carrying out 50 hours, 1, the 13- tridecandioic acid of 41.5g/Kg (that is: 41.5mg/g) is contained in whole fermentation liquids in fermentor
(1,13-tridecanedioicacid)。
4 candida tropicalis H5343HDC23-3 fermenting and producing hybrid long chain dicarboxylic acid of comparative example
Culture medium (it include: glucose 27.0g/L, potassium dihydrogen phosphate 4.9g/L, magnesium sulfate 0.6g/L, calcium chloride 0.1g/L,
Citric acid 0.06g/L, ferric trichloride 0.023g/L, biotin 0.000012g/L, trace meter: copper sulphate 0.00007g/L, sulphur
Sour manganese 0.00432g/L, zinc sulfate 0.00072g/L, citrate 0.00708g/L, SAG471 anti-foaming agent 0.6ml, water surplus)
It is sterilized in the right way to avoid occurring any precipitation reaction, is then added in the fermentor of sterilizing.It is not inoculated with
Complete medium be found very limpid and be shallow bale of straw, not strong smell.Aseptically utilize above-mentioned training
Base is supported, cultivates candida tropicalis by being stirred, ventilating to the fermentor that original liquid volume is 12L
H5343HDC23-3.The aseptic culture medium be vaccinated the inoculum of 3% candida tropicalis H5343HDC23-3 and
35 DEG C are enough stir culture about 12 hours under conditions of making dissolved oxygen be maintained at 20% or more with Ventilation Rate.Pass through addition 6N's
NH4The pH in growth period is adjusted and is maintained 5.8-5.9, the NH by OH4OH is also inorganic nitrogen-sourced in culture medium.When the training
When supporting object and terminating exponential growth, dissolved oxygen is begun to ramp up, transition phase by add a kind of starting of inducing substance and at the same time start with
2.0g/L/ hours rates continuously add the high fatty acid sunflower of oil content raw material flow (containing 84.4% oleic acid, 5.2%
Linoleic acid, 4.7% stearic acid, 3.9% palmitinic acid, and contain a small amount of arachic acid (20: 0), eicosaenoicacid
(20: 1), pentadecanoic acid, the moon silicic acid, remaining substance of tetradecanoic acid).At the same time, the temperature in fermentor is down to by 35 DEG C
30 DEG C, Ventilation Rate is down to 0.4vvm, and pH is regulated and controled reagent N H4OH changes NaOH into.Using 6N NaOH by the pH of transition phase
Adjust and maintain 5.8-5.9.When biomass density reaches about 10g/L, start the speed with 1.22g glucose/L/ hours
Rate continuously adds glucose to fermentor.According to microscopical observation result daily and to the storage bubble accumulated in yeast cells
Assessment, is reduced to 0-45% for the rate of adding of glucose described in transition phase.Anti-foaming agent is not added in transition phase.Transition phase
After carrying out 50 hours, total dicarboxylic acids of 71g/Kg (that is: 71mg/g) is contained in whole fermentation liquids in fermentor.
Based on previous embodiment and comparative example it is found that relative to existing zymotechnique, candidiasis provided by the invention
When for long-chain biatomic acid fermentation, fermentation time is short, and quantity of alkali consumption is few, and binary acid yield is high, and subsequent extracted technique acid dosage is big
Big to reduce, salt content substantially reduces in waste water, and the production cost of long-chain biatomic acid, and environment friend can be greatly reduced on the whole
It is good.
It will be apparent to one skilled in the art that under the premise of without departing substantially from scope and spirit of the present invention, it can
It is carry out various modifications and is changed, the combination between above-mentioned items technical characteristic and other skills for being completed according to above content
Art scheme, which changes, belongs to the scope of the invention.
Claims (12)
1. a kind of Candida tropicalis (Candida tropicalis) bacterial strain CAT H1614, deposit number CCTCC
NO:M 2013143.
2. including the microbial inoculum of Candida tropicalis bacterial strain CAT H1614 as described in claim 1.
3. Candida tropicalis bacterial strain CAT H1614 as described in claim 1 prepares answering in long-chain biatomic acid in fermentation
With.
4. a kind of method of fermenting and producing long-chain biatomic acid, which is characterized in that using the false silk ferment in the torrid zone as described in claim 1
Female bacteria strain CAT H1614 carries out fermenting and producing long-chain biatomic acid.
5. method as claimed in claim 4, which is characterized in that the fermentation process includes strain growth phase and transition phase,
In, in the transition phase, the pH value for controlling fermentation system is 7.0 or less；Preferably 4.0~6.8；More preferably 5.0~6.5.
6. method as claimed in claim 5, which is characterized in that the thallus optical density after the strain growth to 30 times of dilution
OD620When greater than 0.5, the pH value for controlling fermentation system is 7.0 or less；Preferably 4.0~6.8；More preferably 5.0~6.5.
7. such as the described in any item methods of claim 4-6, which is characterized in that the substrate for the fermentation process includes alkane
Hydrocarbon preferably includes the normal alkane of C9~C22；It more preferably include the normal alkane of C9~C18；Most preferably include C10, C11, C12,
The normal alkane of C13, C14, C15 or C16；
And/or the culture medium for the fermentation process includes following component (w/v): glucose 1%~5%, corn pulp
0.1%~0.9%, yeast extract 0.1%~0.5%, potassium nitrate 0.05%~1.2%, potassium dihydrogen phosphate 0.05%~1.0%,
Urea 0.05%~0.3%, ammonium sulfate 0.05%~0.3% and sodium chloride 0.05%~0.2%；Or including following component
(w/v): glucose 1%~5%, potassium nitrate 0.05%~0.6%, potassium dihydrogen phosphate 0.02%~0.6%, ammonium sulfate 0.05%
~0.3% and magnesium sulfate 0.05%~0.3%.
8. such as the described in any item methods of claim 4~7, which is characterized in that in the fermentation process, the inoculation of the bacterial strain
Amount is 10%~30%；
And/or temperature is 28~32 DEG C；
And/or air quantity is 0.3~0.7vvm；
And/or pressure is 0.05~0.14MPa；
And/or the strain growth is interim, the pH value of fermentation system is not less than 3.0, preferably 3.5~6.5.
9. a kind of preparation method of long-chain biatomic acid, comprising:
S1: it ferments and long-chain biatomic acid fermentation liquid is made；And
S2: purification process is extracted to resulting long-chain biatomic acid fermentation liquid, the long-chain biatomic acid is made；
Wherein, the long-chain biatomic acid fermentation liquid is made using the described in any item methods of claim 4~8 in the step S1.
10. preparation method as claimed in claim 9, which is characterized in that the extraction purification processing of the step S2 includes: by institute
The long-chain biatomic acid fermentation liquid acidification obtained separates to obtain solid content, then in organic solvent by solid content dissolution, separates clearly
Liquid, crystallization, thus obtains long-chain biatomic acid.
11. preparation method as claimed in claim 10, which is characterized in that the pH value of the acidification be 2.5~5, preferably 3~
4；
And/or the isolated method is centrifugation or filtering；
And/or the organic solvent includes: the one or more of alcohol, acid, ketone and ester；Wherein, the alcohol include methanol, ethyl alcohol,
One of isopropanol and n-butanol are a variety of；The acid includes acetic acid；The ketone includes acetone；The ester includes ethyl acetate
And/or butyl acetate；
And/or after the solid content is dissolved in organic solvent, decoloration, then clear liquid is separated to obtain, the method for the decoloration is preferably active
Carbon decoloring；The additive amount of the active carbon is no more than the 5% of liquor capacity；The temperature of the decoloration is 85~100 DEG C；It is described
The time of decoloration is 15~165min；
And/or the crystallization is decrease temperature crystalline；The decrease temperature crystalline is the following steps are included: be cooled to 65~80 DEG C, heat preservation 1~2
After hour, then 25~35 DEG C are cooled to, crystallized；
And/or after the crystallization, crystal is isolated, long-chain biatomic acid is thus obtained；The isolated method is centrifuge separation.
12. long-chain biatomic acid made from preparation method as described in claim 10 or 11.
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111748480A (en) * | 2020-06-03 | 2020-10-09 | 上海凯赛生物技术股份有限公司 | Candida virginiana and application thereof |
| CN112680368A (en) * | 2019-10-18 | 2021-04-20 | 上海凯赛生物技术股份有限公司 | Strain for producing long-chain dicarboxylic acid and fermentation method thereof |
| CN112680369A (en) * | 2019-10-18 | 2021-04-20 | 上海凯赛生物技术股份有限公司 | Bacterial strain for high yield of long-chain dicarboxylic acid and fermentation method thereof |
| CN112852893A (en) * | 2021-03-17 | 2021-05-28 | 青岛智库生物技术有限公司 | Method for producing long-chain dicarboxylic acid by biological fermentation |
| CN113061629A (en) * | 2021-03-17 | 2021-07-02 | 青岛智库生物技术有限公司 | Method for producing long-chain dicarboxylic acid by biological fermentation |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20050084940A1 (en) * | 2003-01-14 | 2005-04-21 | Anderson Kevin W. | Method for controlling biooxidation reactions |
| US20110028679A1 (en) * | 2007-11-26 | 2011-02-03 | Cognis Ip Management Gmbh | Polyamides Prepared From Long-Chain Dicarboxylic Acids and Methods for Making the Polyamides |
| CN105154483A (en) * | 2015-10-13 | 2015-12-16 | 齐鲁工业大学 | Application of candida tropicalis to preparation of dodecanedioic acid by utilizing unsaturated grease |
-
2018
- 2018-03-01 CN CN202311329545.2A patent/CN117210343A/en active Pending
- 2018-03-01 CN CN201810172091.5A patent/CN110218661A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20050084940A1 (en) * | 2003-01-14 | 2005-04-21 | Anderson Kevin W. | Method for controlling biooxidation reactions |
| US20110028679A1 (en) * | 2007-11-26 | 2011-02-03 | Cognis Ip Management Gmbh | Polyamides Prepared From Long-Chain Dicarboxylic Acids and Methods for Making the Polyamides |
| CN105154483A (en) * | 2015-10-13 | 2015-12-16 | 齐鲁工业大学 | Application of candida tropicalis to preparation of dodecanedioic acid by utilizing unsaturated grease |
Non-Patent Citations (1)
| Title |
|---|
| 葛明华等: ""十一碳二元酸发酵技术研究"", 《中国石油和化工标准与质量》 * |
Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112680368A (en) * | 2019-10-18 | 2021-04-20 | 上海凯赛生物技术股份有限公司 | Strain for producing long-chain dicarboxylic acid and fermentation method thereof |
| CN112680369A (en) * | 2019-10-18 | 2021-04-20 | 上海凯赛生物技术股份有限公司 | Bacterial strain for high yield of long-chain dicarboxylic acid and fermentation method thereof |
| WO2021073011A1 (en) * | 2019-10-18 | 2021-04-22 | 上海凯赛生物技术股份有限公司 | Strain for producing long-chain dicarboxylic acids and fermentation method therefor |
| CN112680369B (en) * | 2019-10-18 | 2024-05-03 | 上海凯赛生物技术股份有限公司 | Strain for high-yield long-chain dibasic acid and fermentation method thereof |
| CN112680368B (en) * | 2019-10-18 | 2024-05-07 | 上海凯赛生物技术股份有限公司 | Strain for producing long-chain dibasic acid and fermentation method thereof |
| CN111748480A (en) * | 2020-06-03 | 2020-10-09 | 上海凯赛生物技术股份有限公司 | Candida virginiana and application thereof |
| CN111748480B (en) * | 2020-06-03 | 2022-06-24 | 上海凯赛生物技术股份有限公司 | Candida virginiana and application thereof |
| CN112852893A (en) * | 2021-03-17 | 2021-05-28 | 青岛智库生物技术有限公司 | Method for producing long-chain dicarboxylic acid by biological fermentation |
| CN113061629A (en) * | 2021-03-17 | 2021-07-02 | 青岛智库生物技术有限公司 | Method for producing long-chain dicarboxylic acid by biological fermentation |
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