CN116694456A - Preparation device of bacteria immobilized carrier and application method thereof - Google Patents
Preparation device of bacteria immobilized carrier and application method thereof Download PDFInfo
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- CN116694456A CN116694456A CN202310527910.4A CN202310527910A CN116694456A CN 116694456 A CN116694456 A CN 116694456A CN 202310527910 A CN202310527910 A CN 202310527910A CN 116694456 A CN116694456 A CN 116694456A
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- bacteria
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- calcium chloride
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- 241000894006 Bacteria Species 0.000 title claims abstract description 144
- 238000000034 method Methods 0.000 title claims abstract description 25
- 238000002360 preparation method Methods 0.000 title claims abstract description 17
- 238000002156 mixing Methods 0.000 claims abstract description 51
- 239000001963 growth medium Substances 0.000 claims abstract description 49
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 46
- 238000004132 cross linking Methods 0.000 claims abstract description 45
- 238000000855 fermentation Methods 0.000 claims abstract description 40
- 230000004151 fermentation Effects 0.000 claims abstract description 40
- 238000010438 heat treatment Methods 0.000 claims abstract description 26
- 238000007789 sealing Methods 0.000 claims abstract description 21
- 239000004033 plastic Substances 0.000 claims abstract description 18
- 229920003023 plastic Polymers 0.000 claims abstract description 18
- 229910001220 stainless steel Inorganic materials 0.000 claims abstract description 18
- 239000010935 stainless steel Substances 0.000 claims abstract description 18
- 239000000523 sample Substances 0.000 claims abstract description 10
- 239000011550 stock solution Substances 0.000 claims abstract description 4
- 239000008188 pellet Substances 0.000 claims description 52
- 230000001580 bacterial effect Effects 0.000 claims description 50
- UXVMQQNJUSDDNG-UHFFFAOYSA-L Calcium chloride Chemical compound [Cl-].[Cl-].[Ca+2] UXVMQQNJUSDDNG-UHFFFAOYSA-L 0.000 claims description 49
- 230000002572 peristaltic effect Effects 0.000 claims description 40
- 239000000243 solution Substances 0.000 claims description 34
- 239000007788 liquid Substances 0.000 claims description 24
- IXPNQXFRVYWDDI-UHFFFAOYSA-N 1-methyl-2,4-dioxo-1,3-diazinane-5-carboximidamide Chemical compound CN1CC(C(N)=N)C(=O)NC1=O IXPNQXFRVYWDDI-UHFFFAOYSA-N 0.000 claims description 21
- 239000010865 sewage Substances 0.000 claims description 21
- 235000010413 sodium alginate Nutrition 0.000 claims description 21
- 239000000661 sodium alginate Substances 0.000 claims description 21
- 229940005550 sodium alginate Drugs 0.000 claims description 21
- 238000003756 stirring Methods 0.000 claims description 21
- 239000011259 mixed solution Substances 0.000 claims description 19
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 18
- 230000001105 regulatory effect Effects 0.000 claims description 18
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 claims description 16
- 241000589774 Pseudomonas sp. Species 0.000 claims description 16
- 229910052799 carbon Inorganic materials 0.000 claims description 16
- 239000012528 membrane Substances 0.000 claims description 16
- 230000001954 sterilising effect Effects 0.000 claims description 15
- 238000003860 storage Methods 0.000 claims description 13
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 12
- 238000009360 aquaculture Methods 0.000 claims description 12
- 244000144974 aquaculture Species 0.000 claims description 12
- 238000003760 magnetic stirring Methods 0.000 claims description 12
- 239000000203 mixture Substances 0.000 claims description 12
- VWDWKYIASSYTQR-UHFFFAOYSA-N sodium nitrate Chemical compound [Na+].[O-][N+]([O-])=O VWDWKYIASSYTQR-UHFFFAOYSA-N 0.000 claims description 12
- 238000005086 pumping Methods 0.000 claims description 11
- 238000001816 cooling Methods 0.000 claims description 10
- 229920002472 Starch Polymers 0.000 claims description 9
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 claims description 9
- 230000000694 effects Effects 0.000 claims description 9
- 229920001610 polycaprolactone Polymers 0.000 claims description 9
- 239000004632 polycaprolactone Substances 0.000 claims description 9
- 239000008107 starch Substances 0.000 claims description 9
- 238000004659 sterilization and disinfection Methods 0.000 claims description 8
- 239000005995 Aluminium silicate Substances 0.000 claims description 6
- 235000012211 aluminium silicate Nutrition 0.000 claims description 6
- LLSDKQJKOVVTOJ-UHFFFAOYSA-L calcium chloride dihydrate Chemical compound O.O.[Cl-].[Cl-].[Ca+2] LLSDKQJKOVVTOJ-UHFFFAOYSA-L 0.000 claims description 6
- 229940052299 calcium chloride dihydrate Drugs 0.000 claims description 6
- 229940041514 candida albicans extract Drugs 0.000 claims description 6
- 230000001276 controlling effect Effects 0.000 claims description 6
- NLYAJNPCOHFWQQ-UHFFFAOYSA-N kaolin Chemical compound O.O.O=[Al]O[Si](=O)O[Si](=O)O[Al]=O NLYAJNPCOHFWQQ-UHFFFAOYSA-N 0.000 claims description 6
- 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 claims description 6
- 229940061634 magnesium sulfate heptahydrate Drugs 0.000 claims description 6
- 239000002609 medium Substances 0.000 claims description 6
- 229910000402 monopotassium phosphate Inorganic materials 0.000 claims description 6
- 235000019796 monopotassium phosphate Nutrition 0.000 claims description 6
- PJNZPQUBCPKICU-UHFFFAOYSA-N phosphoric acid;potassium Chemical compound [K].OP(O)(O)=O PJNZPQUBCPKICU-UHFFFAOYSA-N 0.000 claims description 6
- 239000004317 sodium nitrate Substances 0.000 claims description 6
- 235000010344 sodium nitrate Nutrition 0.000 claims description 6
- 235000019698 starch Nutrition 0.000 claims description 6
- 239000000725 suspension Substances 0.000 claims description 6
- 239000012138 yeast extract Substances 0.000 claims description 6
- 238000012258 culturing Methods 0.000 claims description 5
- 229910002651 NO3 Inorganic materials 0.000 claims description 4
- NHNBFGGVMKEFGY-UHFFFAOYSA-N Nitrate Chemical compound [O-][N+]([O-])=O NHNBFGGVMKEFGY-UHFFFAOYSA-N 0.000 claims description 4
- 229960002713 calcium chloride Drugs 0.000 claims description 4
- 239000001110 calcium chloride Substances 0.000 claims description 4
- 229910001628 calcium chloride Inorganic materials 0.000 claims description 4
- DGAQECJNVWCQMB-PUAWFVPOSA-M Ilexoside XXIX Chemical compound C[C@@H]1CC[C@@]2(CC[C@@]3(C(=CC[C@H]4[C@]3(CC[C@@H]5[C@@]4(CC[C@@H](C5(C)C)OS(=O)(=O)[O-])C)C)[C@@H]2[C@]1(C)O)C)C(=O)O[C@H]6[C@@H]([C@H]([C@@H]([C@H](O6)CO)O)O)O.[Na+] DGAQECJNVWCQMB-PUAWFVPOSA-M 0.000 claims description 3
- 240000003183 Manihot esculenta Species 0.000 claims description 3
- 235000016735 Manihot esculenta subsp esculenta Nutrition 0.000 claims description 3
- IOVCWXUNBOPUCH-UHFFFAOYSA-M Nitrite anion Chemical compound [O-]N=O IOVCWXUNBOPUCH-UHFFFAOYSA-M 0.000 claims description 3
- 238000005273 aeration Methods 0.000 claims description 3
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 claims description 3
- 238000009835 boiling Methods 0.000 claims description 3
- 230000010261 cell growth Effects 0.000 claims description 3
- 238000010924 continuous production Methods 0.000 claims description 3
- 238000005265 energy consumption Methods 0.000 claims description 3
- 239000011521 glass Substances 0.000 claims description 3
- 230000007774 longterm Effects 0.000 claims description 3
- 239000001301 oxygen Substances 0.000 claims description 3
- 229910052760 oxygen Inorganic materials 0.000 claims description 3
- 229920000642 polymer Polymers 0.000 claims description 3
- 239000011734 sodium Substances 0.000 claims description 3
- 229910052708 sodium Inorganic materials 0.000 claims description 3
- 235000013619 trace mineral Nutrition 0.000 claims description 3
- 239000011573 trace mineral Substances 0.000 claims description 3
- 238000005406 washing Methods 0.000 claims description 3
- 230000001483 mobilizing effect Effects 0.000 claims description 2
- 239000002351 wastewater Substances 0.000 claims description 2
- 230000010354 integration Effects 0.000 claims 1
- 244000005700 microbiome Species 0.000 abstract description 4
- 230000002411 adverse Effects 0.000 abstract 1
- 238000005516 engineering process Methods 0.000 abstract 1
- 230000007613 environmental effect Effects 0.000 abstract 1
- 230000001546 nitrifying effect Effects 0.000 description 9
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 6
- GQPLMRYTRLFLPF-UHFFFAOYSA-N Nitrous Oxide Chemical compound [O-][N+]#N GQPLMRYTRLFLPF-UHFFFAOYSA-N 0.000 description 6
- 239000002002 slurry Substances 0.000 description 5
- 239000003795 chemical substances by application Substances 0.000 description 4
- 239000010440 gypsum Substances 0.000 description 3
- 229910052602 gypsum Inorganic materials 0.000 description 3
- 229910052757 nitrogen Inorganic materials 0.000 description 3
- 239000001272 nitrous oxide Substances 0.000 description 3
- 239000005431 greenhouse gas Substances 0.000 description 2
- 239000011159 matrix material Substances 0.000 description 2
- 230000000813 microbial effect Effects 0.000 description 2
- 230000007812 deficiency Effects 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 238000007599 discharging Methods 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 238000012851 eutrophication Methods 0.000 description 1
- 230000003100 immobilizing effect Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012544 monitoring process Methods 0.000 description 1
- QJGQUHMNIGDVPM-UHFFFAOYSA-N nitrogen group Chemical group [N] QJGQUHMNIGDVPM-UHFFFAOYSA-N 0.000 description 1
- 235000015097 nutrients Nutrition 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 238000004321 preservation Methods 0.000 description 1
- 239000003223 protective agent Substances 0.000 description 1
- 238000004064 recycling Methods 0.000 description 1
- 230000000241 respiratory effect Effects 0.000 description 1
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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
- C12M27/00—Means for mixing, agitating or circulating fluids in the vessel
- C12M27/02—Stirrer or mobile mixing elements
-
- C—CHEMISTRY; METALLURGY
- C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F3/00—Biological treatment of water, waste water, or sewage
- C02F3/34—Biological treatment of water, waste water, or sewage characterised by the microorganisms used
-
- 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
- C12M25/00—Means for supporting, enclosing or fixing the microorganisms, e.g. immunocoatings
- C12M25/16—Particles; Beads; Granular material; Encapsulation
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- C12M29/00—Means for introduction, extraction or recirculation of materials, e.g. pumps
- C12M29/20—Degassing; Venting; Bubble traps
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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- C12M29/00—Means for introduction, extraction or recirculation of materials, e.g. pumps
- C12M29/26—Conditioning fluids entering or exiting the reaction vessel
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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- C12M37/00—Means for sterilizing, maintaining sterile conditions or avoiding chemical or biological contamination
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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- C12M37/00—Means for sterilizing, maintaining sterile conditions or avoiding chemical or biological contamination
- C12M37/02—Filters
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- C12M41/00—Means for regulation, monitoring, measurement or control, e.g. flow regulation
- C12M41/12—Means for regulation, monitoring, measurement or control, e.g. flow regulation of temperature
- C12M41/18—Heat exchange systems, e.g. heat jackets or outer envelopes
- C12M41/24—Heat exchange systems, e.g. heat jackets or outer envelopes inside the vessel
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- C12M41/00—Means for regulation, monitoring, measurement or control, e.g. flow regulation
- C12M41/42—Means for regulation, monitoring, measurement or control, e.g. flow regulation of agitation speed
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- C12M41/00—Means for regulation, monitoring, measurement or control, e.g. flow regulation
- C12M41/44—Means for regulation, monitoring, measurement or control, e.g. flow regulation of volume or liquid level
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- C12M41/48—Automatic or computerized control
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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
- C12N1/20—Bacteria; Culture media therefor
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- C12N11/00—Carrier-bound or immobilised enzymes; Carrier-bound or immobilised microbial cells; Preparation thereof
- C12N11/02—Enzymes or microbial cells immobilised on or in an organic carrier
- C12N11/10—Enzymes or microbial cells immobilised on or in an organic carrier the carrier being a carbohydrate
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- C12N11/00—Carrier-bound or immobilised enzymes; Carrier-bound or immobilised microbial cells; Preparation thereof
- C12N11/14—Enzymes or microbial cells immobilised on or in an inorganic carrier
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- C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F2103/00—Nature of the water, waste water, sewage or sludge to be treated
- C02F2103/20—Nature of the water, waste water, sewage or sludge to be treated from animal husbandry
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- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
- C12R2001/38—Pseudomonas
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- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02W—CLIMATE CHANGE MITIGATION TECHNOLOGIES RELATED TO WASTEWATER TREATMENT OR WASTE MANAGEMENT
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Abstract
The invention belongs to the field of water treatment of pond culture, in particular to a bacteria immobilized carrier preparation device and a use method thereof, and the bacteria immobilized carrier preparation device comprises a first motor, a bacteria fermentation tank, a bacteria culture stock solution, a mixing tank and a carrier crosslinking bottle, wherein the bottom of the first motor is rotationally connected with a first stainless steel connecting rod, the side of the first stainless steel connecting rod is integrally connected with a first plastic blade, a bacteria culture medium is arranged outside the first plastic blade, the bacteria fermentation tank is tightly attached to the outside of the bacteria culture medium, a first heating rod is arranged on one side of the bacteria fermentation tank, a first thermometer probe is arranged on the other side of the bacteria fermentation tank, and a first sealing cover is tightly attached to the top of the bacteria fermentation tank; the invention is convenient to provide an optimized environment for the efficient operation of microorganisms through a microorganism immobilization technology, reduces the loss of strains, and improves the resistance of the strains to environmental adverse factors, thereby prolonging the working efficiency and the duration of denitrification microecological preparation products.
Description
Technical Field
The invention relates to the field of water treatment of pond culture, in particular to a preparation device of a bacterial immobilized carrier and a use method thereof.
Background
With the development of aquaculture industry, water eutrophication caused by the increase of the concentration of nitrogen-containing compounds in the aquaculture tail water is attracting attention. Because of the respiratory requirements of the fishes in the culture water, the dissolved oxygen in the culture water is usually kept above 5mg/L, and the aerobic condition inhibits the traditional process of converting nitrate into nitrogen by anaerobic denitrifying bacteria and discharging the nitrogen out of the culture system. The aerobic condition is unfavorable for the reduction of nitrous oxide greenhouse gases in the traditional denitrification process of anaerobic denitrifying bacteria, and the novel heterotrophic nitrification-aerobic denitrification can convert nitrous oxide into nitrogen under the aerobic condition, so that the emission of nitrous oxide greenhouse gases is reduced.
In chinese patent 201810142003.7, the embodiment of the present invention provides a method for immobilizing nitrifying bacteria using gypsum as a carrier, comprising the steps of: 1) Mixing embedding agent with nutrient solution, stirring uniformly at 20-100deg.C to obtain crosslinked slurry; 2) Mixing the protective agent with nitrifying bacteria bacterial liquid, and uniformly stirring to obtain nitrifying bacteria slurry; 3) Cooling the crosslinked slurry to below 45 ℃, mixing the crosslinked slurry with nitrifying bacteria slurry, uniformly stirring, mixing with gypsum powder, and continuously and uniformly stirring to obtain a carrier matrix containing nitrifying bacteria bacterial agents; 4) Granulating a carrier matrix containing the nitrifying bacterial agent, and drying to obtain the immobilized nitrifying bacterial agent taking gypsum as a carrier. The immobilized nitrifying bacteria prepared by the method solves the problems of high cost or low activity of nitrifying bacteria in the traditional method.
The existing preparation of the bacteria immobilized carrier has the problems that organic carbon in pond culture sewage is insufficient, traditional liquid microbial bacteria liquid is splashed into a culture water body, the efficiency of denitrification of the splashed microorganisms is low due to the lack of organic carbon sources in the culture water body, and efficient strains are easily lost from a culture system in the rainfall and water changing processes.
Therefore, a bacterial immobilization carrier preparation apparatus and a method of using the same have been proposed in order to solve the above-mentioned problems.
Disclosure of Invention
In order to make up the deficiency of the prior art, the problems that organic carbon in the pond culture sewage is insufficient, the traditional liquid microbial liquid is splashed into the culture water body, the efficiency of denitrification of the splashed microorganisms is low due to the lack of organic carbon sources in the culture water body, and efficient strains are easily lost from a culture system in the rainfall and water changing processes are solved.
The invention solves the technical problems by adopting the following technical scheme: the invention relates to a preparation device of a bacteria immobilized carrier and a use method thereof, comprising a first motor, a bacteria fermentation tank, a bacteria culture stock solution, a mixing tank and a carrier crosslinking bottle, wherein the bottom of the first motor is rotationally connected with a first stainless steel connecting rod, the side of the first stainless steel connecting rod is integrally connected with a first plastic blade, the outside of the first plastic blade is provided with a bacteria culture medium, the bacteria fermentation tank is tightly attached to the outside of the bacteria culture medium, one side of the bacteria fermentation tank is provided with a first heating rod, the other side of the bacteria fermentation tank is provided with a first thermometer probe, the top of the bacteria fermentation tank is tightly attached with a first sealing cover, the side of the first sealing cover is communicated with a first rubber pipe, the top of the first rubber pipe is provided with a first peristaltic pump, the bacteria culture reserve liquid is communicated with the end part of the first rubber tube, the outside of the bacteria culture reserve liquid is tightly attached with a culture medium reserve bottle, the side of the bacteria culture medium is communicated with a second rubber tube, the top of the second rubber tube is provided with a second peristaltic pump, the mixing tank is communicated with the end part of the second rubber tube, the inside of the mixing tank is tightly attached with the mixed liquid, the inside of the mixed liquid is communicated with a third rubber tube, the top of the third rubber tube is provided with a third peristaltic pump, the carrier crosslinking bottle is communicated with the end part of the third rubber tube, the inside of the carrier crosslinking bottle is tightly attached with a bacteria crosslinking calcium chloride solution, the bottom of the bacteria crosslinking calcium chloride solution is provided with a magnetic stirring rod, the bottom of the carrier crosslinking bottle is tightly attached with a heating pad, the bottom of the heating pad is provided with a rotating speed display, and a magnetic stirring speed regulating knob is arranged at the bottom of the rotating speed display.
Preferably, the opposite side intercommunication of first sealed lid has first blast pipe, and the top of first blast pipe closely laminates there is first aseptic filter membrane, the top intercommunication of first heating rod has first temperature controller, the top electric connection of second peristaltic pump has programmable logic controller, the inside of mixed liquor closely laminates there is the second plastic vane, and the top fixedly connected with second stainless steel connecting rod of second plastic vane, and the top rotation of second stainless steel connecting rod is connected with the second motor, the second heating rod is installed to one side of blending tank, and the opposite side of blending tank installs the second thermometer probe, and the top electric connection of second heating rod has second temperature controller, the top of blending tank closely laminates there is the second sealed lid, and the inside of second sealed lid has seted up the dog-house, the avris of dog-house is provided with the second blast pipe, and the top of second blast pipe closely laminates there is the second aseptic bottle, the top intercommunication of carrier crosslinked has the third blast pipe, and the top of third closely laminates there is the third aseptic filter membrane, the fourth cross-linked membrane has the inside of fourth cross-linked calcium chloride to install the water valve of rubber end, the fourth cross-linked membrane.
Preferably, the method comprises the following steps:
s1, preparing heterotrophic nitrification-aerobic denitrification bacteria expansion culture solution, sterilizing at high temperature and high pressure, and placing the culture solution in a sterile culture medium storage bottle, wherein the heterotrophic nitrification-aerobic denitrification bacteria expansion culture solution comprises the following components: 0.3g/L ethanol, 0.1g/L sodium nitrate, 0.03g/L potassium dihydrogen phosphate, 0.02g/L magnesium sulfate heptahydrate, 0.02g/L calcium chloride dihydrate, 0.01g/L yeast extract, and pH was adjusted to 7.0 using 5M sodium hydroxide solution.
S2, the volume of the mixing tank is 10L, 8L of 40g/L sodium alginate, 10g/L kaolin and 10g/L starch mixed solution are prepared each time through a feeding port, a second temperature controller monitors the temperature in the mixing tank, a second heating rod is regulated and controlled to be heated to 100 ℃, heating is stopped, a second motor is used for stirring the mixed solution for 1h, the solution in the mixing tank is uniformly mixed, and the mixed solution is reduced to the room temperature level.
S3, opening a water stop valve, adding 8L of sterile 39-4% bacteria crosslinked calcium chloride solution into the carrier crosslinked bottle through a fourth rubber tube, and closing the water stop valve, wherein the volume of the carrier crosslinked bottle is 20L.
S4, regulating and controlling an ultraviolet sterilizing lamp by the programmable logic controller to perform ultraviolet sterilization on the liquid in the culture medium storage bottle and the mixing tank for 30 minutes.
S5, inoculating heterotrophic nitrification-aerobic denitrification strain Pseudomonas sp.DM02 which is separated from the laboratory and has high-efficiency denitrification capability for low-nutrition aquaculture sewage into a bacterial fermentation tank, wherein the heterotrophic nitrification-aerobic denitrification strain Pseudomonas sp.DM02 is only required to be inoculated once in one continuous production period.
S6, controlling a first peristaltic pump by a programmable logic controller, pumping sterilized bacteria expansion culture solution in a culture medium storage bottle into a bacteria fermentation tank, driving a first plastic blade to stir the bacteria culture medium by a first motor through a first stainless steel connecting rod, collecting the temperature of the bacteria culture medium in the bacteria fermentation tank in real time by a first temperature controller through a first thermometer probe, regulating the temperature in the bacteria fermentation tank to 37 ℃ by a first heating rod, ensuring pressure balance and aerobic environment in the bacteria fermentation tank through a first exhaust pipe, and preventing bacteria in air from entering the bacteria fermentation tank by a first sterile filter membrane.
And S7, after the bacteria fermentation tank ferments for 18 hours, the programmable logic controller regulates and controls the second peristaltic pump to pump 0.5L of the bacteria culture medium rich in pseudomonasp.DM02 into the mixing tank, and simultaneously, the programmable logic controller regulates and controls the first peristaltic pump to pump 0.5L of the bacteria culture medium into the bacteria fermentation tank, and the second motor drives the second plastic blade to stir for 30 minutes through the second stainless steel connecting rod.
S8, regulating and controlling a third peristaltic pump to pump the mixture into a carrier crosslinking bottle (38) by a programmable logic controller, regulating and controlling the speed of the third peristaltic pump to enable the mixture to drop into 39-4% of bacteria crosslinking calcium chloride solution in a continuous drop shape, and immediately carrying out crosslinking reaction on the sodium alginate mixture rich in pseudomonadssp.DM02 bacteria and the 39-4% of bacteria crosslinking calcium chloride solution to generate carrier pellets. The magnetic stirring speed regulating knob regulates the speed of the magnetic stirrer to be 100rpm, so that the carrier pellets rotate along with 39-4% of the bacteria crosslinked calcium chloride solution to prevent the carrier pellets from agglomerating, and the rotating speed display displays the regulated rotating speed.
And S9, in the operation process, calculating the completion time t of the mixed liquid pump according to the volume of the mixed liquid in the mixing tank and the speed of the third peristaltic pump, stopping the third peristaltic pump after the time t passes by the programmable logic controller, and adding 8L of sodium alginate-kaolin-starch mixed liquid into the mixing tank again.
S10, opening a water stop valve, pumping the carrier and 39-4% of bacteria crosslinked calcium chloride solution generated in the carrier crosslinked bottle to a container with a proper volume through a fourth rubber tube, preserving the carrier in the environment of 4 ℃ for continuous crosslinking for 24 hours, separating the carrier balls from the 39-4% of bacteria crosslinked calcium chloride solution, preserving the carrier balls in a bacteria expansion culture medium of 4 ℃, recycling the 39-4% of bacteria crosslinked calcium chloride solution, adding a proper amount of calcium chloride, and re-injecting the carrier balls into the carrier crosslinked bottle for use.
S11, placing the produced carrier pellets into the heterotrophic nitrification-aerobic denitrification bacteria expansion culture solution, and placing the carrier pellets at a low temperature of 4 ℃ for long-term storage.
S12, before the heterotrophic nitrification-aerobic denitrification carrier pellets are used, placing the carrier pellets into a blue cap bottle containing the sterilized heterotrophic nitrification-aerobic denitrification bacteria expansion medium, and culturing for 18 hours at 37 ℃ by magnetic stirring at 100rpm to recover the activity of pseudomonasp.
S13, adding the carrier pellets and the high molecular carbon source into a sewage treatment reactor of the circulating water culture system according to a ratio of 1:1.
S14, the aquaculture wastewater enters from the bottom of the reactor through a water pump, and the treated aquaculture tail water flows back to the aquaculture barrel from the top of the reactor.
Preferably, the preparation of the aerobic denitrifying bacteria carrier pellet comprises the following steps:
s1, adding 500mL of a bacteria expansion culture medium into a 1L blue cap bottle, wherein the bacteria expansion culture medium comprises the following components: 0.3g/L ethanol, 0.1g/L sodium nitrate, 0.03g/L potassium dihydrogen phosphate, 0.02g/L magnesium sulfate heptahydrate, 0.02g/L calcium chloride dihydrate, 0.01g/L yeast extract, and pH was adjusted to 7.0 using 5M sodium hydroxide solution.
S2, sealing the blue cap bottle, placing the blue cap bottle in an autoclave for sterilization at 121 ℃ for 21 minutes, taking out, and cooling to room temperature.
S3, adding 100 mu.l of Pseudomonas sp.DM02 bacterial suspension into the blue cap bottle.
S4, sealing with a 0.22 mu m breathable film, and placing in a shaking table at a constant temperature of 37 ℃ for 18h at 100rpm.
S5, slowly adding 8g of sodium alginate, 2g of kaolin and 2g of tapioca starch into a mixing beaker containing 200mL of boiling pure water, stirring while adding, and cooling to room temperature after fully and uniformly stirring.
S6, adding the bacterial suspension in the 10mL blue cap bottle into a mixing beaker containing the sodium alginate mixed solution, and fully stirring and uniformly mixing.
S7, placing the beaker containing the 4% calcium chloride solution on a magnetic stirrer, placing a sterile magnetic stirrer, and opening the magnetic stirrer to 100rpm.
And S8, pumping the sodium alginate mixture mixed with bacteria into a 4% calcium chloride solution by using a peristaltic pump, and mobilizing the peristaltic pump speed to enable the sodium alginate mixture containing aerobic denitrifying bacteria bacterial liquid to be dripped into the 4% calcium chloride solution in continuous liquid drops, wherein the sodium alginate mixture immediately undergoes a crosslinking reaction with the 4% calcium chloride solution to generate carrier pellets.
S9, transferring the carrier pellets and part of 4% calcium chloride solution into a 100mL blue cap bottle, sealing the bottle by using a breathable filter membrane, and placing the bottle and the bottle in a refrigerator at 4 ℃ for crosslinking for 24 hours.
S10, placing the crosslinked carrier pellets in a sterile cell expansion culture medium for storage at 4 ℃.
Preferably, the aerobic denitrifying bacteria carrier pellets and a high molecular organic carbon source (polycaprolactone) are placed in a column reactor, the cultivation sewage is pumped in from the top of the column, and water flowing out from the bottom is returned to the cultivation system, so that the ectopic cultivation sewage treatment is realized, and the method comprises the following steps:
s1, disposing 2.5L of heterotrophic nitrification-aerobic denitrification bacteria expansion culture medium in a 5L blue cap bottle.
S2, sealing the blue cap bottle, placing the blue cap bottle in an autoclave for sterilization at 121 ℃ for 21 minutes, taking out, and cooling to room temperature.
S3, taking 2L of carrier pellets containing the Pseudomonas sp.DM02 strain stored at 4 ℃, placing the carrier pellets in a blue cap bottle containing the sterilized heterotrophic nitrification-aerobic denitrification bacteria expansion medium, placing the blue cap bottle on a magnetic stirrer, and culturing the blue cap bottle for 18 hours at 37 ℃ by using the magnetic stirrer at 100rpm to recover the activity of the Pseudomonas sp.DM02 bacteria in the carrier pellets.
S4, washing the carrier pellets with the recovered activity for 2 times by using a PBS solution, wherein the PBS solution comprises the following formula:
100mg/LKH 2 PO 4
6.6mg/LNaCl
8.2mg/LMgSO 4 ·7H 2 O
13.4mg/LKCl
118mg/LNaHCO 3
1mL trace element
The formula of the microelement solution is as follows:
10g/LFeSO 4 ·7H 2 O
10g/LFeCl 3 ·7H 2 O
2g/LZnSO 4 ·7H 2 O
4g/LCuSO 4 ·7H 2 O
0.5g/LNaMoO 4 ·2H 2 O
0.1g/LMnCl 2 ·4H 2 O
0.1g/LH 3 BO 4
0.3g/LNa 2 SeO 3
10g/L citric acid
S5, adding the carrier pellets and the high molecular carbon source polycaprolactone into a reactor column according to a ratio of 1:1, wherein the reactor column is an organic glass column with an inner diameter of 20cm and a height of 50 cm.
S6, pumping the culture sewage in the culture cylinder from the bottom of the reactor through a peristaltic pump, and returning the culture sewage to the culture cylinder after flowing out of the top of the reactor, wherein the reactor does not need aeration, so that the energy consumption is saved. After the water quality of the effluent of the reactor is domesticated for 1 week, the nitrite of the effluent is less than 0.05mg/L, the nitrate of the effluent is less than 1mg/L, each reactor containing aerobic denitrifying bacteria carrier balls can ensure the stable water quality of 2 to 4 cultivation cylinders for one cultivation period (3 to 12 months), the carrier does not need to be replaced, and the high polymer organic carbon source polycaprolactone in the reactor can be cleaned and recycled after one cultivation period and can be continuously used in the second cultivation period.
Drawings
In order to more clearly illustrate the embodiments of the invention or the technical solutions in the prior art, the drawings which are used in the description of the embodiments or the prior art will be briefly described, it being obvious that the drawings in the description below are only some embodiments of the invention, and that other drawings can be obtained from these drawings without inventive faculty for a person skilled in the art.
FIG. 1 is a schematic view of an apparatus for preparing a bacterial immobilization carrier according to the present invention;
FIG. 2 is a schematic diagram of a simple generation device for preparing aerobic denitrifying bacteria carrier pellets according to the present invention;
FIG. 3 is a schematic view of aerobic denitrifying bacteria carrier pellets prepared by the apparatus of the present invention;
FIG. 4 is a schematic view showing the application of the preparation of the bacterial immobilization carrier of the present invention to the treatment of tail water of cultivation.
In the figure: 1. a first motor; 2. a first exhaust pipe; 3. a first sterile filter membrane; 4. a first sealing cover; 5. a bacterial culture medium; 6. a first heating rod; 7. a bacterial fermenter; 8. a first plastic blade; 9. a first thermometer probe; 10. a first rubber tube; 11. a first stainless steel connecting rod; 12. a first temperature controller; 13. a first peristaltic pump; 14. a second rubber tube; 15. a second peristaltic pump; 16. bacterial culture stock; 17. a medium storage bottle; 18. a programmable logic controller; 19. an ultraviolet sterilizing lamp; 20. a mixing tank; 21. a second plastic blade; 22. a second heating rod; 23. a mixed solution; 24. a second sealing cover; 25. a second motor; 26. a second exhaust pipe; 27. a second sterile filter membrane; 28. a second stainless steel connecting rod; 29. a feed port; 30. a second temperature controller; 31. a third rubber tube; 32. a second thermometer probe; 33. a third peristaltic pump; 34. a third exhaust pipe; 35. a third sterile filter membrane; 36. a water stop valve; 37. a fourth rubber tube; 38. a carrier cross-linking bottle; 39. a calcium chloride solution; 40. a magnetic stirrer; 41. a magnetic stirring speed regulating knob; 42. a rotational speed display; 43. and (3) heating the pad.
Detailed Description
The following description of the technical solutions in the embodiments of the present invention will be made clearly and completely with reference to the accompanying drawings in the embodiments of the present invention, and it is apparent that the described embodiments are only some embodiments of the present invention, not all embodiments. All other embodiments, based on the embodiments in the invention, which a person of ordinary skill in the art would obtain without making any inventive effort, are within the scope of the invention.
Example 1
Referring to fig. 1 and 3, a bacterial immobilization carrier preparation device and a use method thereof, the preparation of aerobic denitrification bacterial carrier pellets comprises the following steps:
s1, adding 500mL of a bacteria expansion culture medium into a 1L blue cap bottle, wherein the bacteria expansion culture medium comprises the following components: 0.3g/L ethanol, 0.1g/L sodium nitrate, 0.03g/L potassium dihydrogen phosphate, 0.02g/L magnesium sulfate heptahydrate, 0.02g/L calcium chloride dihydrate, 0.01g/L yeast extract, and pH was adjusted to 7.0 using 5M sodium hydroxide solution.
S2, sealing the blue cap bottle, placing the blue cap bottle in an autoclave for sterilization at 121 ℃ for 21 minutes, taking out, and cooling to room temperature.
S3, adding 100 mu.l of Pseudomonas sp.DM02 bacterial suspension into the blue cap bottle.
S4, sealing with a 0.22 mu m breathable film, and placing in a shaking table at a constant temperature of 37 ℃ for 18h at 100rpm.
S5, slowly adding 8g of sodium alginate, 2g of kaolin and 2g of tapioca starch into a mixing beaker containing 200mL of boiling pure water, stirring while adding, and cooling to room temperature after fully and uniformly stirring.
S6, adding the bacterial suspension in the 10mL blue cap bottle into a mixing beaker containing the sodium alginate mixed solution 23, and fully stirring and uniformly mixing.
S7, placing a beaker containing the 4% calcium chloride solution on a magnetic stirrer, placing a sterile magnetic stirrer 40, and opening the magnetic stirrer to 100rpm.
And S8, pumping the sodium alginate mixed solution 23 mixed with bacteria into a 4% calcium chloride solution by using a peristaltic pump, and regulating the rate of the peristaltic pump to ensure that the sodium alginate mixed solution 23 containing aerobic denitrifying bacteria bacterial solution is dripped into the 4% calcium chloride solution in continuous liquid drops, and the sodium alginate mixed solution 23 and the 4% calcium chloride solution are immediately subjected to a crosslinking reaction to generate carrier pellets.
S9, transferring the carrier pellets and part of 4% calcium chloride solution into a 100mL blue cap bottle, sealing the bottle by using a breathable filter membrane, and placing the bottle and the bottle in a refrigerator at 4 ℃ for crosslinking for 24 hours.
S10, placing the crosslinked carrier pellets in a sterile cell expansion culture medium for storage at 4 ℃.
Example two
Referring to fig. 1 and 4, a bacterial immobilized carrier preparation device and a use method thereof are disclosed, wherein aerobic denitrifying bacterial carrier pellets and a high molecular organic carbon source (polycaprolactone) are placed in a column reactor, cultivation sewage is pumped in from the top of the column, and water flowing out from the bottom is returned to a cultivation system, so that the treatment of ectopic cultivation sewage is realized, and the method comprises the following steps:
s1, disposing 2.5L of heterotrophic nitrification-aerobic denitrification bacteria expansion culture medium in a 5L blue cap bottle.
S2, sealing the blue cap bottle, placing the blue cap bottle in an autoclave for sterilization at 121 ℃ for 21 minutes, taking out, and cooling to room temperature.
S3, taking 2L of carrier pellets containing the Pseudomonas sp.DM02 strain stored at 4 ℃, placing the carrier pellets in a blue cap bottle containing the sterilized heterotrophic nitrification-aerobic denitrification bacteria expansion medium, placing the blue cap bottle on a magnetic stirrer, and culturing the blue cap bottle for 18 hours at 37 ℃ by using the magnetic stirrer at 100rpm to recover the activity of the Pseudomonas sp.DM02 bacteria in the carrier pellets.
S4, washing the carrier pellets with the recovered activity for 2 times by using a PBS solution, wherein the PBS solution comprises the following formula:
100mg/LKH 2 PO 4
6.6mg/LNaCl
8.2mg/LMgSO 4 ·7H 2 O
13.4mg/LKCl
118mg/LNaHCO 3
1mL trace element
The formula of the microelement solution is as follows:
10g/LFeSO 4 ·7H 2 O
10g/LFeCl 3 ·7H 2 O
2g/LZnSO 4 ·7H 2 O
4g/LCuSO 4 ·7H 2 O
0.5g/LNaMoO 4 ·2H 2 O
0.1g/LMnCl 2 ·4H 2 O
0.1g/LH 3 BO 4
0.3g/LNa 2 SeO 3
10g/L citric acid
S5, adding the carrier pellets and the high molecular carbon source polycaprolactone into a reactor column according to a ratio of 1:1, wherein the reactor column is an organic glass column with an inner diameter of 20cm and a height of 50 cm.
S6, pumping the culture sewage in the culture cylinder from the bottom of the reactor through a peristaltic pump, and returning the culture sewage to the culture cylinder after flowing out of the top of the reactor, wherein the reactor does not need aeration, so that the energy consumption is saved. After the water quality of the effluent of the reactor is domesticated for 1 week, the nitrite of the effluent is less than 0.05mg/L, the nitrate of the effluent is less than 1mg/L, each reactor containing aerobic denitrifying bacteria carrier balls can ensure the stable water quality of 2 to 4 cultivation cylinders for one cultivation period (3 to 12 months), the carrier does not need to be replaced, and the high polymer organic carbon source polycaprolactone in the reactor can be cleaned and recycled after one cultivation period and can be continuously used in the second cultivation period.
Working principle: firstly preparing heterotrophic nitrification-aerobic denitrification bacteria expansion culture solution, sterilizing at high temperature and high pressure, and placing the culture solution in a sterile culture medium storage bottle 17, wherein the heterotrophic nitrification-aerobic denitrification bacteria expansion culture solution comprises the following components: 0.3g/L ethanol, 0.1g/L sodium nitrate, 0.03g/L potassium dihydrogen phosphate, 0.02g/L magnesium sulfate heptahydrate, 0.02g/L calcium chloride dihydrate, 0.01g/L yeast extract, adjusting pH to 7.0 by using 5M sodium hydroxide solution, mixing tank 20 volume to 10L, preparing 8L of 40g/L sodium alginate, 10g/L kaolin, 10g/L starch mixture 23 each time through a feed port 29, monitoring the temperature in mixing tank 20 by a second temperature controller 30, adjusting and controlling the temperature of second heating rod 22 to 100 ℃, stopping heating after heating, stirring the mixture 23 by a second motor 25 for one hour, uniformly mixing the solution in mixing tank 20, reducing the mixture 23 to room temperature level, opening a water stop valve 36, adding 8L of sterile 39-4% bacteria cross-linked calcium chloride solution 39 into a carrier cross-linking bottle 38 by a fourth rubber tube 37, and closing the water stop valve 36, and the carrier cross-linking bottle 38 volume to 20L.
The programmable logic controller 18 controls the ultraviolet sterilizing lamp 19 to sterilize the liquid in the culture medium reserve bottle 17 and the mixing tank 20 for 30 minutes, the heterotrophic nitrification-aerobic denitrification strain pseudoomonassp.DM02 separated in the laboratory and having high-efficient denitrification capability for low-nutrition aquaculture sewage is inoculated into the bacterial fermentation tank 7, only one time in one continuous production period is needed to be inoculated, the programmable logic controller 18 controls the first peristaltic pump 13 to pump sterilized bacterial expansion culture liquid in the culture medium reserve bottle 17 into the bacterial fermentation tank 7, the first motor 1 drives the first plastic blade 8 to stir the bacterial culture medium 5 through the first stainless steel connecting rod 11, the first temperature controller 12 collects the temperature of the bacterial culture medium 5 in the bacterial fermentation tank 7 in real time through the first thermometer probe 9, the temperature in the bacterial fermentation tank 7 is controlled to be 37 ℃ through the first heating rod 6, the pressure balance and the good oxygen environment in the bacterial fermentation tank 7 are ensured through the first exhaust pipe 2, and the first sterile filter membrane 3 prevents bacteria in air from entering the bacterial fermentation tank 7.
After the bacteria fermentation tank 7 ferments for 18h, the programmable logic controller 18 regulates and controls the second peristaltic pump 15 to pump 50.5L of the bacteria culture medium rich in pseudonassp.DM02 into the mixing tank 20, meanwhile, the programmable logic controller 18 regulates and controls the first peristaltic pump 13 to pump 0.5L of the bacteria culture medium 5 into the bacteria fermentation tank 7, the second motor 25 drives the second plastic blade 21 through the second stainless steel connecting rod 28 to stir for 30 min, the programmable logic controller 18 regulates and controls the third peristaltic pump 33 to pump the mixed solution 23 into the carrier crosslinking bottle 38, the speed of the third peristaltic pump 33 is regulated and controlled, the mixed solution 23 drops into 39-4% of bacteria crosslinking calcium chloride solution 39 in a continuous liquid drop shape, and the sodium alginate mixed solution 23 rich in pseudonassp.DM02 bacteria immediately undergoes a crosslinking reaction with 39-4% of bacteria crosslinking calcium chloride solution 39, so that carrier pellets are generated. The magnetic stirring speed regulating knob 41 regulates the speed of the magnetic stirrer 40 to 100rpm, so that the carrier pellets rotate along with the 39-4% bacteria crosslinked calcium chloride solution 39 to prevent the carrier pellets from agglomerating, the rotating speed display 42 displays the regulated rotating speed, the pumping time t of the mixed liquor 23 is calculated according to the volume 8L of the mixed liquor 23 in the mixing tank 20 and the speed of the third peristaltic pump 33 in the running process, the third peristaltic pump 33 is stopped after the time t passes by the programmable logic controller 18, and the 8L sodium alginate-kaolin-starch mixed liquor 23 is added into the mixing tank 20 again.
The carrier produced in the carrier cross-linking bottle 38 and 39-4% bacteria cross-linking calcium chloride solution 39 are pumped to a container with a proper volume through a fourth rubber tube 37, after the carrier cross-linking bottle is preserved in an environment of 4 ℃ for 24 hours, the carrier balls are separated from the 39-4% bacteria cross-linking calcium chloride solution 39, the carrier balls are preserved in a bacteria expanding culture medium of 4 ℃, a proper amount of calcium chloride is added after the 39-4% bacteria cross-linking calcium chloride solution 39 is recycled and is injected into the carrier cross-linking bottle 38 again for use, the produced carrier balls are placed in the heterotrophic nitrification-aerobic denitrification bacteria expanding culture medium, the carrier balls are placed in a blue cap bottle containing the sterilized heterotrophic nitrification-aerobic denitrification bacteria expanding culture medium for long-term preservation at a low temperature of 4 ℃, the carrier balls are placed in a blue cap bottle containing the sterilized heterotrophic nitrification-aerobic denitrification bacteria expanding culture medium for continuous cross-linking for 24 hours through magnetic stirring, the carrier balls are cultivated for 18 hours at a temperature of 100rpm, pseudomonassp.DM02 bacteria in the carrier balls and a high molecular carbon source are added into a sewage treatment reactor of a circulating water aquaculture system according to 1:1, and the sewage treatment reactor is fed into a water pump from the bottom of the water treatment reactor from the bottom to the water tank.
The foregoing has outlined and described the basic principles, features, and advantages of the invention. It will be understood by those skilled in the art that the present invention is not limited to the embodiments described above, and that the embodiments and descriptions described herein are merely illustrative of the principles of the invention, and various changes and modifications may be made without departing from the spirit and scope of the invention, which is defined by the appended claims.
Claims (5)
1. A preparation device of a bacterial immobilization carrier is characterized in that: including first motor (1), bacterial fermentation jar (7), bacterial culture stock solution (16), blending tank (20), carrier cross-linking bottle (38), the bottom rotation of first motor (1) is connected with first stainless steel connecting rod (11), and the avris integration of first stainless steel connecting rod (11) is connected with first plastic vane (8), and the outside of first plastic vane (8) is provided with bacterial culture medium (5), bacterial fermentation jar (7) closely laminate in the outside of bacterial culture medium (5), and one side of bacterial fermentation jar (7) installs first heating rod (6), and first thermometer probe (9) are installed to the opposite side of bacterial fermentation jar (7), the top of bacterial fermentation jar (7) closely laminates first sealed lid (4), and the avris intercommunication of first sealed lid (4) has first rubber tube (10), and the top of first rubber tube (10) has first peristaltic pump (13), bacterial stock solution (16) intercommunication in the tip of first rubber tube (10), and bacterial culture medium (14) are installed at the second side of bacterial fermentation jar (7) closely laminate, and the second rubber tube (14) is installed at the lateral side of second rubber tube (14), the utility model discloses a mixing tank, including first rubber tube (14), second rubber tube (14), mixing tank (20), mixing tank (23) are closely laminated to inside, mixing tank (23) are inside to be linked together has third rubber tube (31), and third peristaltic pump (33) are installed at the top of third rubber tube (31), carrier crosslinked bottle (38) intercommunication and the tip of third rubber tube (31), and the inside of carrier crosslinked bottle (38) is closely laminated with bacterium crosslinked calcium chloride solution (39), and the bottom of bacterium crosslinked calcium chloride solution (39) is provided with magnetic stirring piece (40), heating pad (43) are closely laminated to the bottom of carrier crosslinked bottle (38), and rotational speed display (42) are installed to the bottom of heating pad (43), and magnetic stirring speed regulation and control knob (41) are installed to the bottom of rotational speed display (42).
2. The apparatus for producing a bacterial immobilization carrier according to claim 1, wherein: the other side of the first sealing cover (4) is communicated with a first exhaust pipe (2), the top of the first exhaust pipe (2) is tightly attached with a first sterile filter membrane (3), the top of the first heating rod (6) is communicated with a first temperature controller (12), the top of the second peristaltic pump (15) is electrically connected with a programmable logic controller (18), the interior of the mixed liquid (23) is tightly attached with a second plastic blade (21), the top of the second plastic blade (21) is fixedly connected with a second stainless steel connecting rod (28), the top of the second stainless steel connecting rod (28) is rotationally connected with a second motor (25), one side of the mixing tank (20) is provided with a second heating rod (22), the other side of the mixing tank (20) is provided with a second thermometer probe (32), the top of the second heating rod (22) is electrically connected with a second temperature controller (30), the top of the mixing tank (20) is tightly attached with a second sealing cover (24), the interior of the second sealing cover (24) is provided with a charging port (29), the top of the second sealing cover (24) is provided with a second exhaust pipe (27), the top of the second exhaust pipe (27) is provided with a sterile filter membrane (38), and the top of the third exhaust pipe (34) is tightly attached with a third sterile filter membrane (35), a fourth rubber pipe (37) is communicated with the inside of the bacteria crosslinked calcium chloride solution (39), and a water stop valve (36) is arranged at the end part of the fourth rubber pipe (37).
3. The apparatus for preparing a bacterial immobilization carrier and the method of using the same according to claims 1-2, characterized by comprising the steps of:
s1, preparing heterotrophic nitrification-aerobic denitrification bacteria expansion culture solution, sterilizing at high temperature and high pressure, and placing the culture solution in a sterile culture medium storage bottle (17), wherein the heterotrophic nitrification-aerobic denitrification bacteria expansion culture solution comprises the following components: 0.3g/L ethanol, 0.1g/L sodium nitrate, 0.03g/L potassium dihydrogen phosphate, 0.02g/L magnesium sulfate heptahydrate, 0.02g/L calcium chloride dihydrate, 0.01g/L yeast extract, and pH was adjusted to 7.0 using 5M sodium hydroxide solution.
S2, the volume of the mixing tank (20) is 10L, 8L of 40g/L sodium alginate, 10g/L kaolin and 10g/L starch mixed solution are prepared each time through a feeding port (29), a second temperature controller (30) monitors the temperature in the mixing tank (20), a second heating rod (22) is regulated and controlled to be heated to 100 ℃, heating is stopped, a second motor (25) agitates the mixed solution (23) for 1h, the solution in the mixing tank (20) is uniformly mixed, and the mixed solution (23) is reduced to the room temperature level.
S3, opening a water stop valve (36), adding 8L of sterile 39-4% bacteria crosslinked calcium chloride solution (39) into a carrier crosslinking bottle (38) through a fourth rubber tube (37), and closing the water stop valve (36), wherein the volume of the carrier crosslinking bottle (38) is 20L.
S4, regulating and controlling an ultraviolet sterilizing lamp (19) by a programmable logic controller (18) to perform ultraviolet sterilization on the liquid in the culture medium storage bottle (17) and the mixing tank (20) for 30 minutes.
S5, inoculating heterotrophic nitrification-aerobic denitrification strain Pseudomonas sp.DM02 separated from the laboratory and having high-efficiency denitrification capability for low-nutrition aquaculture sewage into a bacterial fermentation tank (7), wherein the heterotrophic nitrification-aerobic denitrification strain Pseudomonas sp.DM02 is only required to be inoculated once in one continuous production period.
S6, a programmable logic controller (18) controls a first peristaltic pump (13), sterilized bacteria in a culture medium storage bottle (17) are pumped into a bacteria fermentation tank (7), a first motor (1) drives a first plastic blade (8) to stir a bacteria culture medium (5) through a first stainless steel connecting rod (11), a first temperature controller (12) collects the temperature of the bacteria culture medium (5) in the bacteria fermentation tank (7) in real time through a first thermometer probe (9), the temperature in the bacteria fermentation tank (7) is regulated and controlled to be 37 ℃ through a first heating rod (6), pressure balance and a good oxygen environment in the bacteria fermentation tank (7) are guaranteed through a first exhaust pipe (2), and bacteria in air are prevented from entering the bacteria fermentation tank (7) through a first sterile filter membrane (3).
S7, after the bacteria fermentation tank (7) ferments for 18h, the programmable logic controller (18) regulates and controls the second peristaltic pump (15) to pump 0.5L of the bacteria culture medium (5) rich in Pseudomonas sp.DM02 into the mixing tank (20), meanwhile, the programmable logic controller (18) regulates and controls the first peristaltic pump (13) to pump 0.5L of the bacteria culture medium (5) into the bacteria fermentation tank (7), and the second motor (25) drives the second plastic blade (21) to stir for 30 minutes through the second stainless steel connecting rod (28).
S8, a programmable logic controller (18) regulates and controls a third peristaltic pump (33) to pump the mixed solution (23) into a carrier crosslinking bottle (38), regulates and controls the speed of the third peristaltic pump (33) to enable the mixed solution (23) to drop into 39-4% bacteria crosslinking calcium chloride solution (39) in a continuous liquid drop shape, and the sodium alginate mixed solution (23) rich in Pseudomonas sp.DM02 bacteria immediately undergoes crosslinking reaction with the 39-4% bacteria crosslinking calcium chloride solution (39) to generate carrier pellets. The magnetic stirring speed regulating knob (41) regulates the speed of the magnetic stirring rod (40) to be 100rpm, so that the carrier pellets rotate along with the 39-4% bacteria cross-linked calcium chloride solution (39), the carrier pellets are prevented from agglomerating, and the rotary speed display (42) displays the regulated rotary speed.
And S9, in the operation process, calculating the pumping completion time t of the mixed liquor (23) according to the volume (8L) of the mixed liquor in the mixing tank (20) and the speed of the third peristaltic pump (33), stopping the third peristaltic pump (33) after the time t passes by the programmable logic controller (18), and adding 8L of the sodium alginate-kaolin-starch mixed liquor into the mixing tank (20) again.
S10, opening a 36 water stop valve, pumping the carrier generated in the carrier crosslinking bottle (38) and 39-4% bacteria crosslinking calcium chloride solution (39) to a container with a proper volume through a fourth rubber tube (37), preserving the container in a 4 ℃ environment for continuous crosslinking for 24 hours, separating the carrier ball from the 39-4% bacteria crosslinking calcium chloride solution (39), preserving the carrier ball in a 4 ℃ bacteria expansion culture medium, and adding a proper amount of calcium chloride into the carrier crosslinking bottle (38) for use after the 39-4% bacteria crosslinking calcium chloride solution (39) can be recovered.
S11, placing the produced carrier pellets into the heterotrophic nitrification-aerobic denitrification bacteria expansion culture solution, and placing the carrier pellets at a low temperature of 4 ℃ for long-term storage.
S12, before the heterotrophic nitrification-aerobic denitrification carrier pellets are used, placing the carrier pellets into a blue cap bottle containing the sterilized heterotrophic nitrification-aerobic denitrification bacteria expansion medium, and culturing for 18 hours at 37 ℃ by magnetic stirring at 100rpm to recover the activity of Pseudomonas sp.DM02 bacteria in the carrier pellets.
S13, adding the carrier pellets and the high molecular carbon source into a sewage treatment reactor of the circulating water culture system according to a ratio of 1:1.
S14, the aquaculture wastewater enters from the bottom of the reactor through a water pump, and the treated aquaculture tail water flows back to the aquaculture barrel from the top of the reactor.
4. A bacteria immobilized carrier preparation apparatus and a use method thereof according to claims 1-3, wherein the preparation of the aerobic denitrifying bacteria carrier pellet comprises the steps of:
s1, adding 500mL of a bacteria expansion culture medium into a 1L blue cap bottle, wherein the bacteria expansion culture medium comprises the following components: 0.3g/L ethanol, 0.1g/L sodium nitrate, 0.03g/L potassium dihydrogen phosphate, 0.02g/L magnesium sulfate heptahydrate, 0.02g/L calcium chloride dihydrate, 0.01g/L yeast extract, and pH was adjusted to 7.0 using 5M sodium hydroxide solution.
S2, sealing the blue cap bottle, placing the blue cap bottle in an autoclave for sterilization at 121 ℃ for 21 minutes, taking out, and cooling to room temperature.
S3, adding 100 mu l of the Pseudomonas sp.DM02 bacterial suspension to the blue cap bottle.
S4, sealing with a 0.22 mu m breathable film, and placing in a shaking table at a constant temperature of 37 ℃ for 18h at 100rpm.
S5, slowly adding 8g of sodium alginate, 2g of kaolin and 2g of tapioca starch into a mixing beaker containing 200mL of boiling pure water, stirring while adding, and cooling to room temperature after fully and uniformly stirring.
S6, adding the bacterial suspension in the 10mL blue cap bottle into a mixing beaker containing the sodium alginate mixed solution, and fully stirring and uniformly mixing.
S7, placing the beaker containing the 4% calcium chloride solution on a magnetic stirrer, placing a sterile magnetic stirrer, and opening the magnetic stirrer to 100rpm.
And S8, pumping the sodium alginate mixture mixed with bacteria into a 4% calcium chloride solution by using a peristaltic pump, and mobilizing the peristaltic pump speed to enable the sodium alginate mixture containing aerobic denitrifying bacteria bacterial liquid to be dripped into the 4% calcium chloride solution in continuous liquid drops, wherein the sodium alginate mixture immediately undergoes a crosslinking reaction with the 4% calcium chloride solution to generate carrier pellets.
S9, transferring the carrier pellets and part of 4% calcium chloride solution into a 100mL blue cap bottle, sealing the bottle by using a breathable filter membrane, and placing the bottle and the bottle in a refrigerator at 4 ℃ for crosslinking for 24 hours.
S10, placing the crosslinked carrier pellets in a sterile cell expansion culture medium for storage at 4 ℃.
5. The apparatus for preparing a bacterial immobilization carrier and the method of using the same according to claim 1 to 4, characterized in that: the aerobic denitrifying bacteria carrier pellets and a high molecular organic carbon source (polycaprolactone) are placed in a column reactor, the cultivation sewage is pumped in from the top of the column, and water flowing out from the bottom is returned to the cultivation system, so that the ectopic cultivation sewage treatment is realized, and the method comprises the following steps:
s1, disposing 2.5L of heterotrophic nitrification-aerobic denitrification bacteria expansion culture medium in a 5L blue cap bottle.
S2, sealing the blue cap bottle, placing the blue cap bottle in an autoclave for sterilization at 121 ℃ for 21 minutes, taking out, and cooling to room temperature.
S3, taking 2L of carrier pellets containing the Pseudomonas sp.DM02 strain stored at 4 ℃, placing the carrier pellets in a blue cap bottle containing the sterilized heterotrophic nitrification-aerobic denitrification bacteria expansion medium, placing the blue cap bottle on a magnetic stirrer, and culturing the blue cap bottle for 18 hours at 37 ℃ by using the magnetic stirrer at 100rpm to recover the activity of the Pseudomonas sp.DM02 bacteria in the carrier pellets.
S4, washing the carrier pellets with the recovered activity for 2 times by using a PBS solution, wherein the PBS solution comprises the following formula:
100mg/L KH 2 PO 4
6.6mg/LNaCl
8.2mg/LMgSO 4 ·7H 2 O
13.4mg/LKCl
118mg/LNaHCO 3
1mL trace element
The formula of the microelement solution is as follows:
10g/LFeSO 4 ·7H 2 O
10g/LFeCl 3 ·7H 2 O
2g/LZnSO 4 ·7H 2 O
4g/L CuSO 4 ·7H 2 O
0.5g/LNaMoO 4 ·2H 2 O
0.1g/L MnCl 2 ·4H 2 O
0.1g/L H 3 BO 4
0.3g/LNa 2 SeO 3
10g/L citric acid
S5, adding the carrier pellets and the high molecular carbon source polycaprolactone into a reactor column according to a ratio of 1:1, wherein the reactor column is an organic glass column with an inner diameter of 20cm and a height of 50 cm.
S6, pumping the culture sewage in the culture cylinder from the bottom of the reactor through a peristaltic pump, and returning the culture sewage to the culture cylinder after flowing out of the top of the reactor, wherein the reactor does not need aeration, so that the energy consumption is saved. After the water quality of the effluent of the reactor is domesticated for 1 week, the nitrite of the effluent is less than 0.05mg/L, the nitrate of the effluent is less than 1mg/L, each reactor containing aerobic denitrifying bacteria carrier balls can ensure the stable water quality of 2 to 4 cultivation cylinders for one cultivation period (3 to 12 months), the carrier does not need to be replaced, and the high polymer organic carbon source polycaprolactone in the reactor can be cleaned and recycled after one cultivation period and can be continuously used in the second cultivation period.
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| CN117361749A (en) * | 2023-12-07 | 2024-01-09 | 中建环能科技股份有限公司 | Preparation method of sewage denitrification carrier |
| CN117361749B (en) * | 2023-12-07 | 2024-03-15 | 中建环能科技股份有限公司 | Preparation method of sewage denitrification carrier |
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