CN110747104B - Continuous in-situ separation device - Google Patents
Continuous in-situ separation device Download PDFInfo
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- CN110747104B CN110747104B CN201911173803.6A CN201911173803A CN110747104B CN 110747104 B CN110747104 B CN 110747104B CN 201911173803 A CN201911173803 A CN 201911173803A CN 110747104 B CN110747104 B CN 110747104B
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- 238000000926 separation method Methods 0.000 title claims abstract description 161
- 238000011065 in-situ storage Methods 0.000 title claims abstract description 70
- 239000007788 liquid Substances 0.000 claims abstract description 118
- 238000000855 fermentation Methods 0.000 claims abstract description 86
- 230000004151 fermentation Effects 0.000 claims abstract description 86
- 238000010992 reflux Methods 0.000 claims abstract description 14
- 238000005192 partition Methods 0.000 claims description 11
- 238000004891 communication Methods 0.000 claims description 8
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 7
- 230000000149 penetrating effect Effects 0.000 claims description 2
- 239000011521 glass Substances 0.000 claims 1
- 241000894006 Bacteria Species 0.000 abstract description 6
- 238000011109 contamination Methods 0.000 abstract description 4
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- 238000009776 industrial production Methods 0.000 abstract description 3
- HIWPGCMGAMJNRG-ACCAVRKYSA-N Sophorose Natural products O([C@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@H]1O)[C@H]1[C@H](O)[C@@H](O)[C@@H](O)[C@@H](CO)O1 HIWPGCMGAMJNRG-ACCAVRKYSA-N 0.000 description 21
- HIWPGCMGAMJNRG-UHFFFAOYSA-N beta-sophorose Natural products OC1C(O)C(CO)OC(O)C1OC1C(O)C(O)C(O)C(CO)O1 HIWPGCMGAMJNRG-UHFFFAOYSA-N 0.000 description 21
- 238000000034 method Methods 0.000 description 19
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- ZTOKUMPYMPKCFX-CZNUEWPDSA-N (E)-17-[(2R,3R,4S,5S,6R)-6-(acetyloxymethyl)-3-[(2S,3R,4S,5S,6R)-6-(acetyloxymethyl)-3,4,5-trihydroxyoxan-2-yl]oxy-4,5-dihydroxyoxan-2-yl]oxyoctadec-9-enoic acid Chemical compound OC(=O)CCCCCCC/C=C/CCCCCCC(C)O[C@@H]1O[C@H](COC(C)=O)[C@@H](O)[C@H](O)[C@H]1O[C@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](COC(C)=O)O1 ZTOKUMPYMPKCFX-CZNUEWPDSA-N 0.000 description 6
- WRMNZCZEMHIOCP-UHFFFAOYSA-N 2-phenylethanol Chemical compound OCCC1=CC=CC=C1 WRMNZCZEMHIOCP-UHFFFAOYSA-N 0.000 description 6
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- 238000005516 engineering process Methods 0.000 description 5
- 239000012530 fluid Substances 0.000 description 5
- XBDQKXXYIPTUBI-UHFFFAOYSA-N dimethylselenoniopropionate Natural products CCC(O)=O XBDQKXXYIPTUBI-UHFFFAOYSA-N 0.000 description 4
- 230000005764 inhibitory process Effects 0.000 description 4
- 238000003860 storage Methods 0.000 description 4
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- 240000004808 Saccharomyces cerevisiae Species 0.000 description 3
- 235000014680 Saccharomyces cerevisiae Nutrition 0.000 description 3
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 3
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- PZDOWFGHCNHPQD-VNNZMYODSA-N sophorose Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@H](C=O)O[C@@H]1O[C@H](CO)[C@@H](O)[C@H](O)[C@H]1O PZDOWFGHCNHPQD-VNNZMYODSA-N 0.000 description 3
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- 239000010696 ester oil Substances 0.000 description 2
- 150000002148 esters Chemical class 0.000 description 2
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- 239000007791 liquid phase Substances 0.000 description 2
- 238000000622 liquid--liquid extraction Methods 0.000 description 2
- 238000011068 loading method Methods 0.000 description 2
- 229920001451 polypropylene glycol Polymers 0.000 description 2
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- 238000001556 precipitation Methods 0.000 description 2
- 235000019260 propionic acid Nutrition 0.000 description 2
- IUVKMZGDUIUOCP-BTNSXGMBSA-N quinbolone Chemical compound O([C@H]1CC[C@H]2[C@H]3[C@@H]([C@]4(C=CC(=O)C=C4CC3)C)CC[C@@]21C)C1=CCCC1 IUVKMZGDUIUOCP-BTNSXGMBSA-N 0.000 description 2
- 238000000638 solvent extraction Methods 0.000 description 2
- 235000015112 vegetable and seed oil Nutrition 0.000 description 2
- 239000008158 vegetable oil Substances 0.000 description 2
- HVCOBJNICQPDBP-UHFFFAOYSA-N 3-[3-[3,5-dihydroxy-6-methyl-4-(3,4,5-trihydroxy-6-methyloxan-2-yl)oxyoxan-2-yl]oxydecanoyloxy]decanoic acid;hydrate Chemical compound O.OC1C(OC(CC(=O)OC(CCCCCCC)CC(O)=O)CCCCCCC)OC(C)C(O)C1OC1C(O)C(O)C(O)C(C)O1 HVCOBJNICQPDBP-UHFFFAOYSA-N 0.000 description 1
- 229930186217 Glycolipid Natural products 0.000 description 1
- 241000699670 Mus sp. Species 0.000 description 1
- 239000001888 Peptone Substances 0.000 description 1
- 108010080698 Peptones Proteins 0.000 description 1
- 241000186428 Propionibacterium freudenreichii Species 0.000 description 1
- 241001278052 Starmerella Species 0.000 description 1
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- 238000012136 culture method Methods 0.000 description 1
- 230000006837 decompression Effects 0.000 description 1
- DGLRDKLJZLEJCY-UHFFFAOYSA-L disodium hydrogenphosphate dodecahydrate Chemical compound O.O.O.O.O.O.O.O.O.O.O.O.[Na+].[Na+].OP([O-])([O-])=O DGLRDKLJZLEJCY-UHFFFAOYSA-L 0.000 description 1
- 239000006185 dispersion Substances 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 238000004945 emulsification Methods 0.000 description 1
- 239000006052 feed supplement Substances 0.000 description 1
- 238000012262 fermentative production Methods 0.000 description 1
- 238000005187 foaming Methods 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 239000007789 gas Substances 0.000 description 1
- 208000015181 infectious disease Diseases 0.000 description 1
- 238000011081 inoculation Methods 0.000 description 1
- WRUGWIBCXHJTDG-UHFFFAOYSA-L magnesium sulfate heptahydrate Chemical compound O.O.O.O.O.O.O.[Mg+2].[O-]S([O-])(=O)=O WRUGWIBCXHJTDG-UHFFFAOYSA-L 0.000 description 1
- 229940061634 magnesium sulfate heptahydrate Drugs 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 230000000813 microbial effect Effects 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 230000003020 moisturizing effect Effects 0.000 description 1
- 229910000402 monopotassium phosphate Inorganic materials 0.000 description 1
- 235000019796 monopotassium phosphate Nutrition 0.000 description 1
- 231100000956 nontoxicity Toxicity 0.000 description 1
- 235000019319 peptone Nutrition 0.000 description 1
- 238000005373 pervaporation Methods 0.000 description 1
- 239000003208 petroleum Substances 0.000 description 1
- GNSKLFRGEWLPPA-UHFFFAOYSA-M potassium dihydrogen phosphate Chemical compound [K+].OP(O)([O-])=O GNSKLFRGEWLPPA-UHFFFAOYSA-M 0.000 description 1
- LWIHDJKSTIGBAC-UHFFFAOYSA-K potassium phosphate Substances [K+].[K+].[K+].[O-]P([O-])([O-])=O LWIHDJKSTIGBAC-UHFFFAOYSA-K 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 230000009467 reduction Effects 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 150000003839 salts Chemical class 0.000 description 1
- 238000005070 sampling Methods 0.000 description 1
- 229930000044 secondary metabolite Natural products 0.000 description 1
- 238000005063 solubilization Methods 0.000 description 1
- 230000007928 solubilization Effects 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 230000001502 supplementing effect Effects 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 238000012546 transfer Methods 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
- 238000009736 wetting Methods 0.000 description 1
Classifications
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M29/00—Means for introduction, extraction or recirculation of materials, e.g. pumps
- C12M29/18—External loop; Means for reintroduction of fermented biomass or liquid percolate
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01D—SEPARATION
- B01D21/00—Separation of suspended solid particles from liquids by sedimentation
- B01D21/24—Feed or discharge mechanisms for settling tanks
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01D—SEPARATION
- B01D21/00—Separation of suspended solid particles from liquids by sedimentation
- B01D21/30—Control equipment
- B01D21/34—Controlling the feed distribution; Controlling the liquid level ; Control of process parameters
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M39/00—Means for cleaning the apparatus or avoiding unwanted deposits of microorganisms
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M47/00—Means for after-treatment of the produced biomass or of the fermentation or metabolic products, e.g. storage of biomass
- C12M47/10—Separation or concentration of fermentation products
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- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Chemical & Material Sciences (AREA)
- Zoology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Health & Medical Sciences (AREA)
- Organic Chemistry (AREA)
- Wood Science & Technology (AREA)
- Biotechnology (AREA)
- Biomedical Technology (AREA)
- Sustainable Development (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Microbiology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Apparatus Associated With Microorganisms And Enzymes (AREA)
Abstract
The invention belongs to the technical field of fermentation, and particularly relates to a continuous in-situ separation device which comprises a shell and a shell inner cavity, wherein a baffle is arranged at the lower part of the shell inner cavity, and divides the shell inner cavity into an upper liquid separating bin and a lower standing bin; an outlet pipe and an inlet pipe are arranged on the shell at the corresponding position of the liquid separation bin, the outlet pipe is positioned at the upper part of the inlet pipe, at least one layer of liquid separation baffle is arranged in the liquid separation bin between the outlet pipe and the inlet pipe, each layer of liquid separation baffle inclines at an angle of 30-60 degrees relative to the horizontal plane, an overflow port and a reflux port are arranged on each layer of liquid separation baffle, and the reflux ports of the liquid separation baffles positioned at the bottommost layer from top to bottom are communicated with the standing bin through a reflux conduit; the flow guide channel arranged on the baffle plate is communicated with the liquid separating bin between the liquid separating baffle plate at the bottommost layer and the baffle plate; the bottom of the shell is provided with a bottom valve which is communicated with the standing bin. The device of the invention ensures that the fermentation liquor is always in a circulating state, reduces the loss of thalli, reduces the risk of bacteria contamination and is easy for industrial production.
Description
Technical Field
The invention belongs to the technical field of fermentation, and particularly relates to a continuous in-situ separation device.
Background
In-situ separation is also called as in-situ separation coupling technology, and is a fermentation technology which utilizes a separation and purification means to continuously separate a product from a culture medium in a fermentation or biological reaction process, so that the fermentation process and the separation process are synchronously carried out, the product generated in the culture medium is separated in time, the inhibitory effect of the product is further built, and the comprehensive yield and the yield of fermentation are integrally increased.
Different in-situ separation technologies according to separation means include adsorption method, liquid-liquid extraction method, gas stripping method, pervaporation method, membrane separation method and the like. Adsorption refers to the use of adsorbent materials such as macroporous resins, activated carbon, etc. to adsorb the products in the fermentation medium during fermentation, thereby reducing the inhibitory effect of the products in the fermentation process. For example, in-situ separation of propionic acid-coupled VB in the literature resin 12 Research on fermentation Process [ D]In the method, an in-situ separation mode of resin adsorption is adopted, so that the product inhibition of propionic acid to the propionibacterium freudenreichii is greatly reduced, and VB is improved 12 Is a product of the above process. The liquid-liquid extraction method is to add an extraction phase to separate the fermentation product from the fermentation liquid in the fermentation process in an extraction mode, so as to reduce the whole biological reactionProduct inhibition in the medium during the reaction. For example, in the synthesis of 2-phenylethanol by bioconversion in the water-polypropylene glycol two-phase system of the publication (Programme, 2011,11 (5): 782-785), 2-phenylethanol in the fermentation broth is extracted by polypropylene glycol double liquid phase, thereby increasing the yield of Saccharomyces cerevisiae 2-phenylethanol. The membrane separation is to separate the fermentation liquor product by adopting a membrane separation mode, for example, a method for separating ethanol in situ by using a membrane separation technology (CN 103695475A) in Chinese patent application utilizes a vapor permeation membrane to collect and concentrate the volatilized ethanol vapor at the upper part of a culture medium after vacuum decompression by adopting a membrane assembly mode, thereby reducing the ethanol content in the fermentation culture medium and increasing the production of bioethanol.
Sophorolipid (Sophorolipid) is a microbial secondary metabolite produced by candida by fermentation process under certain conditions with sugar, vegetable oil and the like as carbon sources. Sophorolipids are a type of glycolipid biosurfactant. The surfactant has the general performances of solubilization, emulsification, wetting, foaming, dispersion, surface tension reduction and the like of the conventional surfactant, and has the characteristics of no toxicity, 100% biodegradability, temperature resistance, high salt resistance, wide pH adaptation range, environmental friendliness and the like. The sophorolipid can be applied to the fields of petroleum, environmental protection, medicine, food, cosmetics, washing, home care, agriculture, feed and the like, and can be used for partially or completely replacing chemical synthetic surfactants. The sophorose ester has multiphase distribution in the fermentation process, which comprises an oil phase, a lactone type sophorose ester phase and a fermentation liquid phase, and the sophorose ester not only has the product inhibition condition in multiphase mixing, but also weakens dissolved oxygen mass transfer in a culture system. Therefore, the yield and the production efficiency of the sophorose ester can be effectively increased by adopting an in-situ separation means.
In the known literature (Dolman, b.m., kaismann, c., martin, p.j., & winter burn, j.b. (2017). Integrated sophorolipid production and gravity separation.process Biochemistry,54,162-171.Doi: 10.1016/j.procbio.2016.12.021), an in-situ separation technique for producing sophoroesters by fermentation and a gravity separation device are disclosed, in the disclosed technique, sophoroesters are performed by adopting a gap operation mode, that is, fermentation liquid in fermentation is pumped into the gravity separation device, and after separation interface between a lactone type sophoroester layer and a hair is clear, the fermentation liquid is pumped back into a reactor through an outlet at the upper part or the lower part of the device for continuous fermentation, and meanwhile, the lactone type sophoroester product is separated and collected. The publications (Liu, z., tian, x., chen, y., lin, y., mohsin, a., & Chu, j. (2018) & Efficient sophorolipids production via a novel in situ separation technology by Starmerella b omicola. Process biochemistry. Doi:10.1016/j. Procbio.2018.12.005) modified the one sophorose ester in situ separation process on the basis of the disclosed in situ separation protocol, which adds a preseparation unit prior to the gravity separation unit.
In the known in situ separation scheme for the fermentative production of sophoroesters, the in situ separation process is carried out intermittently, i.e. after transferring part of the fermentation broth into a separation device, standing for gravity separation, so that the lactone-type sophoroesters layer can be separated from the fermentation broth, then discharging the lactone-type sophoroesters from the separation device, then discharging the fermentation broth and inputting the fermentation broth back into the fermentation tank, and after the separation device is emptied, the next separation can be carried out. The prior in-situ separation technical scheme of the sophorose ester has the following problems:
(1) Intermittent batch operation is unfavorable for continuous industrial production, and is difficult to be applied in industrial production.
(2) Intermittent batch operation increases a significant amount of operational effort.
(3) The separation device communicated with the fermentation system needs to be emptied for a plurality of times, so that the probability of bacteria infection is greatly increased.
(4) The stationary gravity separation process requires a longer stationary delamination time.
(5) The thalli can also be precipitated in the standing gravity separation process, and the thalli loss can be caused in the step of emptying the separation device, thereby influencing the production efficiency.
Disclosure of Invention
Aiming at the problems in the prior art, the invention provides a continuous in-situ separation device which can be applied to fermentation production of sophoroesters in an in-situ separation mode and fermentation processes of similar products (such as mice Li Tangzhi) so as to reduce product inhibition in the fermentation process and improve production efficiency.
The technical scheme of the invention is that the continuous in-situ separation device comprises a shell (1) and a shell inner cavity wrapped by the shell (1), wherein a baffle (6) is arranged at the lower part of the shell inner cavity, the baffle (6) divides the shell inner cavity into a liquid separating bin (9) at the upper part and a standing bin (8) at the lower part, a flow guide channel is arranged on the baffle (6), and the flow guide channel is communicated with the liquid separating bin (9) and the standing bin (8);
an outlet pipe (2) and an inlet pipe (3) are arranged on a shell (1) at the corresponding position of a liquid separating bin (9), the outlet pipe (2) is positioned at the upper part of the inlet pipe (3), at least one layer of liquid separating baffle (5) is arranged in the liquid separating bin (9) between the outlet pipe (2) and the inlet pipe (3), the liquid separating baffle (5) divides the liquid separating bin (9) into more than two independent spaces, each layer of liquid separating baffle (5) is parallel to each other, each layer of liquid separating baffle (5) inclines at an angle of 30-60 degrees relative to the horizontal plane, an overflow port (5.1) and a reflux port (5.2) are arranged on each layer of liquid separating baffle (5), the overflow port (5.1) is higher than the reflux port (5.2), and the reflux ports (5.2) of the liquid separating baffles (5) positioned at the bottommost layer from top to bottom are communicated with the bin (8) through a reflux guide pipe (5.3) penetrating through a partition plate (6); the diversion channel arranged on the baffle plate (6) is communicated with a liquid separation bin (9) between the liquid separation baffle plate (5) at the bottommost layer and the baffle plate (6);
the bottom of the shell (1) is provided with a bottom valve (4), and the bottom valve (4) is communicated with a standing bin (8).
In the in-situ separation fermentation process of sophorose ester, fermentation liquor in a fermentation tank is input from an inlet pipe (3) of an in-situ separation device, passes through an overflow port (5.1) of a multi-layer liquid separation baffle plate (5) and a space between the liquid separation baffle plates, and finally continuously flows back to the fermentation tank through an outlet pipe (2) arranged at the upper part of the separation device. The in-situ separation device and the fermentation tank form a closed continuous external circulation system. In the fermentation process, circulating fermentation liquid is subjected to polymerization in a narrow space between the liquid separation baffles, dispersed lactone-type sophoroesters are gradually polymerized into large droplets, the lactone-type sophoroesters in a fermentation liquid system have higher specific gravity than a culture medium, and the polymerized lactone-type sophoroesters can flow downwards along the inclined liquid separation baffles, are concentrated at the slope bottom of each layer of liquid separation baffles and flow back to the lower layer through a backflow port (5.2) arranged at the bottom of the liquid separation baffles, and the backflow port (5.2) of the liquid separation baffle at the bottommost layer is directly connected with a backflow conduit (5.3), so that the finally concentrated lactone-type sophoroesters flow back to a standing bin (8) at the bottom of the separation device through the backflow conduit (5.3). During the fermentation process, the liquid layer height of the lactone-type sophorose ester can be observed through a liquid level window (7) arranged at the bottom. After the liquid layer of lactone type sophoroester is filled up to a high degree in the still standing bin, part of sophoroester can be discharged through the bottom valve, and the operation is simple and convenient. A small amount of lactone type sophoroester liquid layer can be reserved in the operation process, and the lactone type sophoroester liquid layer also plays a role in isolating fermentation liquor, so that the risk of system bacterial contamination in the process of collecting sophoroester by liquid drainage is greatly reduced.
The space inside the in-situ separation device is divided into a liquid separating bin (9) for continuously and circularly flowing fermentation liquor and a relatively static standing bin (8) by the partition plate (6), and the lactone type sophorose ester which is collected and refluxed through aggregation can be further subjected to standing delamination in the standing bin. The fermentation liquid water phase is distributed on the upper part of the lactone type sophorose ester layer, overflows back into the liquid separating bin through a communicating hole (6.1) arranged on the partition plate (6) or a diversion port of a chimney type diversion port (6.2), and returns to the continuous circulation of the whole fermentation liquid.
The viewing mirror (7) is arranged on the shell corresponding to the standing bin (8), so that the situation in the standing bin (8) can be observed in time.
The flow guide channels arranged on the partition plate (6) are a plurality of communication holes (6.1) and/or chimney type flow guide ports (6.2). That is, the flow guide port may be a plurality of communication holes (6.1), a chimney-type flow guide port (6.2), or a plurality of communication holes (6.1) and a chimney-type flow guide port (6.2) which are present at the same time, and preferably a plurality of communication holes (6.1) or a plurality of communication holes (6.1) and a chimney-type flow guide port (6.2) which are present at the same time.
The spacing between each layer of liquid separation baffle plates (5) is 5-20cm, preferably 8-15cm, and more preferably 10cm. The number of layers of the liquid separation baffle (5) is preferably 2-5 layers, and as an embodiment, 3-4 layers are adopted, and according to the size of an actual separation device, the number of layers can also be 5, 6, 7, 8, 9, 10 layers or more. Each layer of liquid separation baffle is preferably inclined at an angle of 45 degrees relative to the horizontal plane.
The overflow port (5.1) arranged on each layer of liquid separation baffle (5) is positioned at the highest position of the liquid separation baffle (5), and the reflux port (5.2) is positioned at the lowest position of the liquid separation baffle (5).
An overflow drainage cover is fixed on the overflow port (5.1), and is positioned on one surface of the liquid separation baffle (5) close to the upper end liquid separation baffle (5), the overflow drainage cover can be a side opening or a top opening, preferably a side opening, and the side opening has higher turbulence condition and longer fluid flow path, thereby being beneficial to the occurrence of the sophorose ester oil phase polymerization process.
An oil-water separation device is fixed at the side opening or the top opening of the overflow drainage cover so as to improve the oil-water separation efficiency. The oil-water separation device is a laminated grid net or a laminated corrugated plate, so that the sophorose polymerization and separation effect is improved, each layer of overflow port (2) is filled with corrugated plates or a plurality of layers of grid nets to serve as coalescents, the oil drop polymerization process of the lactone sophorose ester oil phase is increased, particularly, the corrugated plate filler and the plurality of layers of grid nets provide tortuous channels for fluid to flow back and forth on the corrugated plate filler and the plurality of layers of grid nets, and the polymerization process of the lactone sophorose ester liquid drops is accelerated when fermentation liquor passes through the overflow ports.
The equivalent diameter of the grids in the grid net is 3-5mm. By equivalent diameter is meant that if the mesh openings of the grid mesh are circular, the equivalent diameter should be that circular diameter. If the mesh of the grid net is square, the equivalent diameter is the side length; if the mesh of the grid net is of other special shape, the annual diameter is the side length of a square with the same area corresponding to the mesh area, namely D=S 1/2 S is the mesh area and D is the equivalent diameter.
The liquid separation baffle (5) is characterized in that more than two independent spaces into which the liquid separation bin (9) is divided are respectively provided with laminated corrugated plates so as to provide stronger turbulence disturbance and a more tortuous fluid flow path, thereby greatly increasing the separation efficiency of the in-situ separation device.
The backflow port (5.2) is fixedly provided with a backflow drainage cover, the backflow drainage cover is positioned on one surface of the liquid separation baffle (5) far away from the upper end liquid separation baffle (5), and the backflow drainage cover can be a side opening or a bottom opening, and is preferably provided with the bottom opening.
The bottom valve (4) is communicated with the storage tank (10), the storage tank (10) is a sterilized sealed tank body, sophorose ester separated in situ in the fermentation process is discharged into the storage tank (10) through the bottom valve (4), and the risk of bacteria contamination is further reduced.
Compared with the technical scheme of the intermittent in-situ separation device in the prior art (Dolman, B.M., kaisermann, C., martin, P.J., & Winterburn, J.B. (2017) Integrated sophorolipid production and gravity separation.Process Biochemistry,54,162-171.doi:10.1016/j.procbio.2016.12.021), the continuous in-situ separation device provided by the invention has the following remarkable technical effects:
(1) Unlike the existing in-situ separation scheme, which needs repeated liquid loading, standing separation and evacuation of the separation device, the technical scheme provided by the invention does not need a complicated operation process. The whole fermentation liquor is always in a circulating state, and only the separated lactone sophorolipid is discharged through a bottom valve at regular intervals.
(2) Compared with the existing in-situ separation scheme, in the technical scheme provided by the invention, the fermentation liquor is always in a dynamic circulation process and has no standing process, so that the bacterial loss in the in-situ separation process caused by cell precipitation is greatly less than that in the prior art.
Compared with the prior art, the technical scheme provided by the invention does not need to empty the separation device for multiple times, and has lower bacteria contamination risk.
Drawings
FIG. 1 is a schematic cross-sectional view of an in-situ separation apparatus according to example 1.
FIG. 2 is a schematic diagram of the structure of a closed circulation system among a fermentation tank, an in-situ separation device and a storage tank.
FIG. 3 shows the distribution and layering of vegetable oil, fermentation broth, and lactone-type sophoroesters during fermentation in example 2 (fermentation broth sampling layering for 1 to 10 days from left to right, wherein the lactone-type sophoroesters layer is always submerged in the lowest part of the liquid separation layer).
FIG. 4 is a schematic view of an in-situ separation device in which a plurality of communication holes are provided in a partition plate in the in-situ separation device of example 3.
FIG. 5 is a schematic diagram of the type and structure of overflow holes filled in the side-opened overflow port of example 4 (side-opened overflow holes, overflow holes filled in a multi-layer grid, and overflow holes filled in a corrugated plate, respectively from left to right)
Detailed Description
Example 1 continuous in situ separation fermentation separation device of sophorose ester and use method
The in-situ separation device adopted in the embodiment is shown in figure 1, and comprises a shell 1, an outlet pipe 2, an inlet pipe 3, a bottom valve 4, a liquid separation baffle 5, a baffle 6 and a liquid level window 7; the bottom valve 4 is arranged at the bottom of the shell 1, the inner cavity of the shell is divided into two independent bin bodies by the partition plate 6, namely a standing bin 8 at the lower part and a liquid separating bin 9 at the upper part, a liquid level window 7 is arranged on the shell 1 where the standing bin 8 is located, the inlet pipe 3 is arranged at the upper part of the partition plate 6, the outlet pipe 2 is arranged at the upper part of the shell 1, three layers of liquid separating baffles 5 are arranged between the outlet pipe 2 and the inlet pipe 3, each layer of liquid separating baffles 5 are obliquely arranged in a cavity between the outlet pipe 2 and the inlet pipe 3 at an included angle of 45 degrees with the horizontal, each layer of liquid separating baffles 5 are parallel to each other, the interval between each layer of liquid separating baffles 5 is 10cm, meanwhile, overflow ports 5.1 and backflow ports 5.2 are arranged on each layer of liquid separating baffles 5, and backflow ports 5.2 arranged on the liquid separating baffles 5 at the bottommost layer are directly drained into the standing bin 8 through one backflow conduit 5.3. Wherein the overflow port 5.1 is opened in a side opening (as shown in fig. 1 and 4), and the reflux port 5.2 is opened in a straight opening (as shown in fig. 1 and 4). In the in-situ separation fermentation process of the sophorose ester, fermentation liquor is input from an inlet pipe 3 of the in-situ separation device, and continuously flows back to the fermentation tank through a space between an overflow port 5.1 of a multi-layer liquid separation baffle and the liquid separation baffle by an outlet pipe 2 arranged at the upper part of the separation device. The in-situ separation device and the fermentation tank form a closed continuous external circulation system. In the fermentation process, circulating fermentation liquor generates polymerization in a narrow space between the liquid separation baffles, dispersed lactone-type sophoroesters are gradually polymerized into large drops, and as the specific gravity of the lactone-type sophoroesters in a fermentation liquor system is higher than that of a culture medium, the polymerized lactone-type sophoroesters can flow downwards along the inclined liquid separation baffles, are concentrated at the slope bottom of each layer of liquid separation baffles, flow back to the lower layer through a backflow port 5.2 arranged at the bottom of the liquid separation baffles, and the backflow port 5.2 of the liquid separation baffle at the bottommost layer is directly connected with a backflow conduit 5.3, and finally the concentrated lactone-type sophoroesters flow back to a standing bin 8 at the bottom of the separation device through the backflow conduit. During the fermentation process, the liquid layer height of the lactone-type sophorose ester can be observed through a liquid level window 7 arranged at the bottom. And after the internal ester liquid layer is filled up, the ester can be discharged through a bottom valve. The peristaltic pump is used as power in the liquid conveying process between the fermentation tank and the in-situ separation device.
EXAMPLE 2 continuous in situ separation fermentation of sophoroesters
Since the lactone-type sophoroesters are separated from the bottom valve of the continuous in-situ separation apparatus when fermentation is performed by the continuous in-situ separation method provided in example 1, it is necessary to control the specific gravity of the fermentation medium and the fermentation broth to be lower than that of the lactone-type sophoroesters throughout the whole fermentation period. The embodiment is carried out by adopting the following culture method: candida ATCC22214 was selected as a production strain, and was inoculated into a 1L shake flask (200 mL of medium in which glucose was 2% w/v, peptone was 2% w/v, yeast powder was 1% w/v, pH 5.0) and shake-cultured at 30 ℃ for 24 hours as a seed solution. Taking a 5L fermentation tank as a culture tank body, wherein the liquid loading amount is 3L (fermentation medium: glucose 6%w/v; rapeseed oil 1%w/v; yeast powder 0.3%w/v; disodium hydrogen phosphate dodecahydrate 0.1%w/v; monopotassium phosphate 0.1%w/v; magnesium sulfate heptahydrate 0.05%w/v; initial pH is 5.0), inoculating seed liquid into the tank according to the inoculation amount of 1%, controlling the dissolved oxygen to be 50%, and controlling the dissolved oxygen to be linked with the rotating speed (the upper and lower limits of the rotating speed are 300-800 rpm), wherein the fermentation temperature is 25 ℃, and continuously supplementing the rapeseed oil according to 3.75g/h during the fermentation; feed supplement pipe moisturizing rate: 3.35mL/min, while glucose solution (at a concentration of 800 g/L) was continuously fed at a rate of 10 g/L/day. The experimental group used in situ separation fermentation with the in situ separation device shown in example 1, peristaltic pump set peristaltic delivery rate of 100mL/h. And meanwhile, the same fermentation conditions are installed in a fermentation tank fermentation device without an in-situ separation device to perform fermentation as a comparison. As a result, according to the culture conditions of the present embodiment, the specific gravity of the sophorose ester liquid layer is higher than that of the fermentation liquid (or fermentation medium) in the whole fermentation period (see FIG. 3), so that the lactone-type sophorose ester liquid can be settled at the bottom of the standing area of the in-situ separation device, and can be smoothly separated from the bottom valve of the in-situ separation device. Wherein the total yield of the lactone-type sophoroesters of the experimental group which adopts the in-situ separation device reaches 82g/L, and the lactone-type sophoroesters which can be collected in the culture medium without adopting the in-situ separation device are only 41g/L.
Example 3 improvement of the in-situ separation apparatus of example 1
In the continuous in-situ separation apparatus of example 1, the partition 6 divides the in-situ separation apparatus into a stationary compartment 9 and a liquid separation compartment 8. Wherein the liquid in the liquid separation bin is always in a motion circulation state, and the liquid in the standing bin is relatively static, and can flow back into the liquid separation bin only through a chimney type flow guide port 6.2 arranged on the baffle plate 6. Because the liquid in the liquid separation bin is relatively static, part of bacteria can be settled and the loss of part of bacteria is illuminated. As shown in fig. 4, the chimney-like flow-guiding openings 6.2 in example 1 are thus changed to a plurality of openings 6.1 provided in the partition. The improvement increases the fluid communication between the standing bin 9 and the liquid separating bin 8 at the boundary, and can reduce the bacterial precipitation by about 30-40%.
Example 4 improvement of the in situ separation device of example 1
The efficiency of the coalescence of small oil droplets (i.e., lactone-type sophorose ester droplets) of the fermentation broth during circulation determines the in situ separation device of the whole device. The use of stacked corrugated plates or multi-layered grid mesh is effective to provide a more turbulent flow disturbance and a more tortuous fluid flow path to greatly increase the separation efficiency of the in situ separation device. As shown in fig. 5, this embodiment provides an improved way of in-situ separation device, that is, the position of the overflow port of the in-situ separation device is filled with laminated corrugated plates or multi-layer grid mesh to be used as an oil-water separator, so as to improve the separation efficiency of lactone type sophoroesters.
The foregoing is merely a preferred embodiment of the present invention and it should be noted that modifications and adaptations to those skilled in the art may be made without departing from the principles of the present invention, which are intended to be comprehended within the scope of the present invention.
Claims (10)
1. The continuous in-situ separation device is characterized by comprising a shell (1) and a shell inner cavity wrapped by the shell (1), wherein a baffle (6) is arranged at the lower part of the shell inner cavity, the baffle (6) divides the shell inner cavity into a liquid separating bin (9) at the upper part and a standing bin (8) at the lower part, a flow guide channel is arranged on the baffle (6), and the flow guide channel is communicated with the liquid separating bin (9) and the standing bin (8); an outlet pipe (2) and an inlet pipe (3) are arranged on a shell (1) at the corresponding position of a liquid separating bin (9), the outlet pipe (2) is positioned at the upper part of the inlet pipe (3), at least one layer of liquid separating baffle (5) is arranged in the liquid separating bin (9) between the outlet pipe (2) and the inlet pipe (3), the liquid separating baffle (5) divides the liquid separating bin (9) into more than two independent spaces, each layer of liquid separating baffle (5) is parallel to each other, each layer of liquid separating baffle (5) inclines at an angle of 30-60 degrees relative to the horizontal plane, an overflow port (5.1) and a reflux port (5.2) are arranged on each layer of liquid separating baffle (5), the overflow port (5.1) is higher than the reflux port (5.2), and the reflux ports (5.2) of the liquid separating baffles (5) positioned at the bottommost layer from top to bottom are communicated with the bin (8) through a reflux guide pipe (5.3) penetrating through a partition plate (6); the diversion channel arranged on the baffle plate (6) is communicated with a liquid separation bin (9) between the liquid separation baffle plate (5) at the bottommost layer and the baffle plate (6); the bottom of the shell (1) is provided with a bottom valve (4), the bottom valve (4) is communicated with a standing bin (8), and fermentation liquid is continuously input through an inlet pipe (3) in the fermentation process and continuously flows back to the fermentation tank through an outlet pipe (2).
2. The continuous in-situ separation device according to claim 1, wherein the housing corresponding to the standing bin (8) is provided with a sight glass (7) so as to facilitate timely observation of the situation in the standing bin (8).
3. Continuous in-situ separation device according to claim 1, characterized in that the flow guiding channels provided on the partition (6) are a plurality of communication holes (6.1) and/or chimney-like flow guiding ports (6.2).
4. Continuous in-situ separation device according to claim 1, characterized in that the spacing between each layer of liquid separation baffles (5) is 5-20cm.
5. Continuous in-situ separation device according to claim 1, characterized in that the number of layers of the liquid separation baffle (5) is 2-5.
6. Continuous in-situ separation device according to claim 1, characterized in that the overflow port (5.1) provided on each layer of the liquid separation baffle (5) is located at the highest position of the liquid separation baffle (5), and the reflux port (5.2) is located at the lowest position of the liquid separation baffle (5).
7. The continuous in-situ separation device according to claim 1, wherein an overflow drainage cover is fixed on the overflow port (5.1), the overflow drainage cover is positioned on one surface of the liquid separation baffle (5) close to the upper liquid separation baffle (5), and the overflow drainage cover is provided with a side opening or a bottom opening; the backflow port (5.2) is fixedly provided with a backflow drainage cover, the backflow drainage cover is positioned on one surface of the liquid separation baffle (5) far away from the upper end liquid separation baffle (5), and the backflow drainage cover is provided with a side opening or a bottom opening.
8. The continuous in-situ separation device according to claim 7, wherein an oil-water separation device is fixed at the side opening or the top opening of the overflow drainage cover, and the oil-water separation device is a laminated grid mesh or a laminated corrugated plate.
9. The continuous in-situ separation device according to claim 1, characterized in that the liquid separation baffle (5) is provided with laminated corrugated plates in more than two independent spaces into which the liquid separation bin (9) is divided.
10. Continuous in-situ separation device according to claim 1, characterized in that the bottom valve (4) communicates with a tank (10).
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