CN105505830A - Acinetobacter strain for producing biosurfactant and application of acinetobacter strain - Google Patents
Acinetobacter strain for producing biosurfactant and application of acinetobacter strain Download PDFInfo
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- CN105505830A CN105505830A CN201610038100.2A CN201610038100A CN105505830A CN 105505830 A CN105505830 A CN 105505830A CN 201610038100 A CN201610038100 A CN 201610038100A CN 105505830 A CN105505830 A CN 105505830A
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- 239000003876 biosurfactant Substances 0.000 title claims abstract description 49
- 241000589291 Acinetobacter Species 0.000 title abstract description 8
- 238000000034 method Methods 0.000 claims abstract description 18
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 claims abstract description 10
- 229910052799 carbon Inorganic materials 0.000 claims abstract description 9
- 239000005862 Whey Substances 0.000 claims abstract description 8
- 102000007544 Whey Proteins Human genes 0.000 claims abstract description 8
- 108010046377 Whey Proteins Proteins 0.000 claims abstract description 8
- 239000000843 powder Substances 0.000 claims abstract description 8
- 239000003208 petroleum Substances 0.000 claims abstract description 6
- 238000010790 dilution Methods 0.000 claims abstract description 4
- 239000012895 dilution Substances 0.000 claims abstract description 4
- 238000012216 screening Methods 0.000 claims abstract description 4
- 239000002609 medium Substances 0.000 claims description 22
- 239000000243 solution Substances 0.000 claims description 15
- 239000013543 active substance 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
- 239000012043 crude product Substances 0.000 claims description 10
- 241000588624 Acinetobacter calcoaceticus Species 0.000 claims description 9
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 claims description 9
- 238000004519 manufacturing process Methods 0.000 claims description 9
- 230000001580 bacterial effect Effects 0.000 claims description 8
- 239000007788 liquid Substances 0.000 claims description 8
- 238000007747 plating Methods 0.000 claims description 8
- 229910019142 PO4 Inorganic materials 0.000 claims description 7
- ZLMJMSJWJFRBEC-UHFFFAOYSA-N Potassium Chemical compound [K] ZLMJMSJWJFRBEC-UHFFFAOYSA-N 0.000 claims description 7
- NBIIXXVUZAFLBC-UHFFFAOYSA-K phosphate Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 claims description 7
- 239000010452 phosphate Substances 0.000 claims description 7
- 229910052700 potassium Inorganic materials 0.000 claims description 7
- 239000011591 potassium Substances 0.000 claims description 7
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 7
- NLXLAEXVIDQMFP-UHFFFAOYSA-N Ammonia chloride Chemical compound [NH4+].[Cl-] NLXLAEXVIDQMFP-UHFFFAOYSA-N 0.000 claims description 6
- TWRXJAOTZQYOKJ-UHFFFAOYSA-L Magnesium chloride Chemical compound [Mg+2].[Cl-].[Cl-] TWRXJAOTZQYOKJ-UHFFFAOYSA-L 0.000 claims description 6
- 235000015097 nutrients Nutrition 0.000 claims description 6
- 229940001516 sodium nitrate Drugs 0.000 claims description 6
- 235000010344 sodium nitrate Nutrition 0.000 claims description 6
- 239000004317 sodium nitrate Substances 0.000 claims description 6
- 239000006228 supernatant Substances 0.000 claims description 6
- 239000002689 soil Substances 0.000 claims description 5
- 239000007787 solid Substances 0.000 claims description 5
- DBPRUZCKPFOVDV-UHFFFAOYSA-N Clorprenaline hydrochloride Chemical compound O.Cl.CC(C)NCC(O)C1=CC=CC=C1Cl DBPRUZCKPFOVDV-UHFFFAOYSA-N 0.000 claims description 4
- 230000004913 activation Effects 0.000 claims description 4
- BNIILDVGGAEEIG-UHFFFAOYSA-L disodium hydrogen phosphate Chemical compound [Na+].[Na+].OP([O-])([O-])=O BNIILDVGGAEEIG-UHFFFAOYSA-L 0.000 claims description 4
- 229910000397 disodium phosphate Inorganic materials 0.000 claims description 4
- 235000019800 disodium phosphate Nutrition 0.000 claims description 4
- 238000011081 inoculation Methods 0.000 claims description 4
- 239000002054 inoculum Substances 0.000 claims description 4
- 229940050906 magnesium chloride hexahydrate Drugs 0.000 claims description 4
- DHRRIBDTHFBPNG-UHFFFAOYSA-L magnesium dichloride hexahydrate Chemical compound O.O.O.O.O.O.[Mg+2].[Cl-].[Cl-] DHRRIBDTHFBPNG-UHFFFAOYSA-L 0.000 claims description 4
- 229910052751 metal Inorganic materials 0.000 claims description 4
- 239000002184 metal Substances 0.000 claims description 4
- DPDMMXDBJGCCQC-UHFFFAOYSA-N [Na].[Cl] Chemical compound [Na].[Cl] DPDMMXDBJGCCQC-UHFFFAOYSA-N 0.000 claims description 3
- 235000019270 ammonium chloride Nutrition 0.000 claims description 3
- ZPWVASYFFYYZEW-UHFFFAOYSA-L dipotassium hydrogen phosphate Chemical compound [K+].[K+].OP([O-])([O-])=O ZPWVASYFFYYZEW-UHFFFAOYSA-L 0.000 claims description 3
- 239000012153 distilled water Substances 0.000 claims description 3
- RBTARNINKXHZNM-UHFFFAOYSA-K iron trichloride Chemical compound Cl[Fe](Cl)Cl RBTARNINKXHZNM-UHFFFAOYSA-K 0.000 claims description 3
- 229960002337 magnesium chloride Drugs 0.000 claims description 3
- 229910001629 magnesium chloride Inorganic materials 0.000 claims description 3
- 239000012074 organic phase Substances 0.000 claims description 3
- 239000006161 blood agar Substances 0.000 claims description 2
- 238000009629 microbiological culture Methods 0.000 claims description 2
- 238000012546 transfer Methods 0.000 claims description 2
- 239000011573 trace mineral Substances 0.000 claims 1
- 235000013619 trace mineral Nutrition 0.000 claims 1
- 238000000855 fermentation Methods 0.000 abstract description 12
- 230000004151 fermentation Effects 0.000 abstract description 12
- 239000000126 substance Substances 0.000 abstract description 7
- 239000013527 degreasing agent Substances 0.000 abstract description 5
- 238000005265 energy consumption Methods 0.000 abstract description 4
- 230000004060 metabolic process Effects 0.000 abstract description 4
- 235000013351 cheese Nutrition 0.000 abstract description 3
- 238000000605 extraction Methods 0.000 abstract description 3
- 239000003513 alkali Substances 0.000 abstract description 2
- 239000011248 coating agent Substances 0.000 abstract description 2
- 238000000576 coating method Methods 0.000 abstract description 2
- 238000003912 environmental pollution Methods 0.000 abstract description 2
- 239000001963 growth medium Substances 0.000 abstract 3
- 238000009395 breeding Methods 0.000 abstract 1
- 230000001488 breeding effect Effects 0.000 abstract 1
- 230000003247 decreasing effect Effects 0.000 abstract 1
- 238000000746 purification Methods 0.000 abstract 1
- 239000003921 oil Substances 0.000 description 33
- 238000010521 absorption reaction Methods 0.000 description 7
- 239000004094 surface-active agent Substances 0.000 description 7
- 229910000831 Steel Inorganic materials 0.000 description 6
- 230000000694 effects Effects 0.000 description 6
- 239000010959 steel Substances 0.000 description 6
- 238000005457 optimization Methods 0.000 description 5
- 230000008569 process Effects 0.000 description 5
- 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 4
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 4
- 229930186217 Glycolipid Natural products 0.000 description 4
- 238000004458 analytical method Methods 0.000 description 4
- HEDRZPFGACZZDS-UHFFFAOYSA-N chloroform Substances ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 description 4
- 150000002632 lipids Chemical class 0.000 description 4
- LZZYPRNAOMGNLH-UHFFFAOYSA-M Cetrimonium bromide Chemical compound [Br-].CCCCCCCCCCCCCCCC[N+](C)(C)C LZZYPRNAOMGNLH-UHFFFAOYSA-M 0.000 description 3
- 239000003153 chemical reaction reagent Substances 0.000 description 3
- 238000005237 degreasing agent Methods 0.000 description 3
- 238000005516 engineering process Methods 0.000 description 3
- 230000007613 environmental effect Effects 0.000 description 3
- 238000011160 research Methods 0.000 description 3
- 239000002699 waste material Substances 0.000 description 3
- 238000005033 Fourier transform infrared spectroscopy Methods 0.000 description 2
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 2
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 2
- XEEYBQQBJWHFJM-UHFFFAOYSA-N Iron Chemical compound [Fe] XEEYBQQBJWHFJM-UHFFFAOYSA-N 0.000 description 2
- 239000001888 Peptone Substances 0.000 description 2
- 108010080698 Peptones Proteins 0.000 description 2
- 239000002253 acid Substances 0.000 description 2
- 230000007423 decrease Effects 0.000 description 2
- 238000006073 displacement reaction Methods 0.000 description 2
- 238000002474 experimental method Methods 0.000 description 2
- 239000008103 glucose Substances 0.000 description 2
- JEIPFZHSYJVQDO-UHFFFAOYSA-N iron(III) oxide Inorganic materials O=[Fe]O[Fe]=O JEIPFZHSYJVQDO-UHFFFAOYSA-N 0.000 description 2
- 125000000325 methylidene group Chemical group [H]C([H])=* 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 229910052757 nitrogen Inorganic materials 0.000 description 2
- QJGQUHMNIGDVPM-UHFFFAOYSA-N nitrogen group Chemical group [N] QJGQUHMNIGDVPM-UHFFFAOYSA-N 0.000 description 2
- 231100000252 nontoxic Toxicity 0.000 description 2
- 230000003000 nontoxic effect Effects 0.000 description 2
- 235000019319 peptone Nutrition 0.000 description 2
- 239000012071 phase Substances 0.000 description 2
- 238000002203 pretreatment Methods 0.000 description 2
- 239000002994 raw material Substances 0.000 description 2
- DAJSVUQLFFJUSX-UHFFFAOYSA-M sodium;dodecane-1-sulfonate Chemical compound [Na+].CCCCCCCCCCCCS([O-])(=O)=O DAJSVUQLFFJUSX-UHFFFAOYSA-M 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- FRXSZNDVFUDTIR-UHFFFAOYSA-N 6-methoxy-1,2,3,4-tetrahydroquinoline Chemical compound N1CCCC2=CC(OC)=CC=C21 FRXSZNDVFUDTIR-UHFFFAOYSA-N 0.000 description 1
- GUBGYTABKSRVRQ-XLOQQCSPSA-N Alpha-Lactose Chemical compound O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@H]1O[C@@H]1[C@@H](CO)O[C@H](O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-XLOQQCSPSA-N 0.000 description 1
- 241000894006 Bacteria Species 0.000 description 1
- 241000196324 Embryophyta Species 0.000 description 1
- 230000005526 G1 to G0 transition Effects 0.000 description 1
- GUBGYTABKSRVRQ-QKKXKWKRSA-N Lactose Natural products OC[C@H]1O[C@@H](O[C@H]2[C@H](O)[C@@H](O)C(O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@H]1O GUBGYTABKSRVRQ-QKKXKWKRSA-N 0.000 description 1
- 244000046052 Phaseolus vulgaris Species 0.000 description 1
- 235000010627 Phaseolus vulgaris Nutrition 0.000 description 1
- 241000589516 Pseudomonas Species 0.000 description 1
- UIIMBOGNXHQVGW-UHFFFAOYSA-M Sodium bicarbonate Chemical compound [Na+].OC([O-])=O UIIMBOGNXHQVGW-UHFFFAOYSA-M 0.000 description 1
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 description 1
- 229930006000 Sucrose Natural products 0.000 description 1
- CKUAXEQHGKSLHN-UHFFFAOYSA-N [C].[N] Chemical compound [C].[N] CKUAXEQHGKSLHN-UHFFFAOYSA-N 0.000 description 1
- 230000032683 aging Effects 0.000 description 1
- 150000001413 amino acids Chemical class 0.000 description 1
- BFNBIHQBYMNNAN-UHFFFAOYSA-N ammonium sulfate Chemical compound N.N.OS(O)(=O)=O BFNBIHQBYMNNAN-UHFFFAOYSA-N 0.000 description 1
- 229910052921 ammonium sulfate Inorganic materials 0.000 description 1
- 229940044197 ammonium sulfate Drugs 0.000 description 1
- 235000011130 ammonium sulphate Nutrition 0.000 description 1
- 239000007864 aqueous solution Substances 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 238000009835 boiling Methods 0.000 description 1
- 239000007853 buffer solution Substances 0.000 description 1
- 239000006227 byproduct Substances 0.000 description 1
- 229940041514 candida albicans extract Drugs 0.000 description 1
- 125000002915 carbonyl group Chemical group [*:2]C([*:1])=O 0.000 description 1
- 150000007942 carboxylates Chemical group 0.000 description 1
- 239000003093 cationic surfactant Substances 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 125000003636 chemical group Chemical group 0.000 description 1
- 238000001311 chemical methods and process Methods 0.000 description 1
- 239000007795 chemical reaction product Substances 0.000 description 1
- 238000004140 cleaning Methods 0.000 description 1
- 239000002131 composite material Substances 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 235000009508 confectionery Nutrition 0.000 description 1
- 238000011109 contamination Methods 0.000 description 1
- 239000010779 crude oil Substances 0.000 description 1
- 238000013461 design Methods 0.000 description 1
- 150000002016 disaccharides Chemical class 0.000 description 1
- 238000004945 emulsification Methods 0.000 description 1
- 230000001804 emulsifying effect Effects 0.000 description 1
- 125000004185 ester group Chemical group 0.000 description 1
- 238000011156 evaluation Methods 0.000 description 1
- 150000002191 fatty alcohols Chemical class 0.000 description 1
- 238000001914 filtration Methods 0.000 description 1
- 150000004676 glycans Chemical class 0.000 description 1
- 235000011187 glycerol Nutrition 0.000 description 1
- -1 glycolipid compound Chemical class 0.000 description 1
- 239000004519 grease Substances 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 238000009776 industrial production Methods 0.000 description 1
- 238000002329 infrared spectrum Methods 0.000 description 1
- 229910017053 inorganic salt Inorganic materials 0.000 description 1
- 229910052742 iron Inorganic materials 0.000 description 1
- 239000008101 lactose Substances 0.000 description 1
- CXKWCBBOMKCUKX-UHFFFAOYSA-M methylene blue Chemical compound [Cl-].C1=CC(N(C)C)=CC2=[S+]C3=CC(N(C)C)=CC=C3N=C21 CXKWCBBOMKCUKX-UHFFFAOYSA-M 0.000 description 1
- 229960000907 methylthioninium chloride Drugs 0.000 description 1
- 239000000693 micelle Substances 0.000 description 1
- 230000000813 microbial effect Effects 0.000 description 1
- 230000007269 microbial metabolism Effects 0.000 description 1
- 244000005700 microbiome Species 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 230000007935 neutral effect Effects 0.000 description 1
- 229920001542 oligosaccharide Polymers 0.000 description 1
- 150000002482 oligosaccharides Chemical class 0.000 description 1
- 235000010482 polyoxyethylene sorbitan monooleate Nutrition 0.000 description 1
- 229920001184 polypeptide Polymers 0.000 description 1
- 229920001282 polysaccharide Polymers 0.000 description 1
- 239000005017 polysaccharide Substances 0.000 description 1
- 229920000053 polysorbate 80 Polymers 0.000 description 1
- 102000004196 processed proteins & peptides Human genes 0.000 description 1
- 108090000765 processed proteins & peptides Proteins 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- 238000004451 qualitative analysis Methods 0.000 description 1
- 238000011084 recovery Methods 0.000 description 1
- 230000009467 reduction Effects 0.000 description 1
- 238000005067 remediation Methods 0.000 description 1
- 229920006395 saturated elastomer Polymers 0.000 description 1
- 239000009671 shengli Substances 0.000 description 1
- 238000001228 spectrum Methods 0.000 description 1
- 239000005720 sucrose Substances 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
- 239000012138 yeast extract Substances 0.000 description 1
Classifications
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
- C12N1/205—Bacterial isolates
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P1/00—Preparation of compounds or compositions, not provided for in groups C12P3/00 - C12P39/00, by using microorganisms or enzymes
- C12P1/04—Preparation of compounds or compositions, not provided for in groups C12P3/00 - C12P39/00, by using microorganisms or enzymes by using bacteria
-
- C—CHEMISTRY; METALLURGY
- C25—ELECTROLYTIC OR ELECTROPHORETIC PROCESSES; APPARATUS THEREFOR
- C25D—PROCESSES FOR THE ELECTROLYTIC OR ELECTROPHORETIC PRODUCTION OF COATINGS; ELECTROFORMING; APPARATUS THEREFOR
- C25D5/00—Electroplating characterised by the process; Pretreatment or after-treatment of workpieces
- C25D5/34—Pretreatment of metallic surfaces to be electroplated
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
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- Engineering & Computer Science (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Genetics & Genomics (AREA)
- Biotechnology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Health & Medical Sciences (AREA)
- Microbiology (AREA)
- General Engineering & Computer Science (AREA)
- Biochemistry (AREA)
- Virology (AREA)
- Biomedical Technology (AREA)
- Tropical Medicine & Parasitology (AREA)
- Medicinal Chemistry (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Electrochemistry (AREA)
- Materials Engineering (AREA)
- Metallurgy (AREA)
- Mycology (AREA)
- General Chemical & Material Sciences (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
- Emulsifying, Dispersing, Foam-Producing Or Wetting Agents (AREA)
Abstract
The invention provides an acinetobacter strain for producing a biosurfactant and an application of the acinetobacter strain. The acinetobacter strain is obtained from a petroleum foul land through enrichment culture, dilution coating, screening and purification and is authenticated as acinetobacter. The invention discloses an enrichment culture medium and a fermentation culture medium for screening the strain, and the properties of the biosurfactant produced from the strain are analyzed. A carbon source used in the culture media is whey powder which is leftover remained after the extraction of cheese and is low in price and cost; the strain is high in breeding speed and metabolism speed, the biosurfactant produced through the metabolism can act as a biological degreaser at the room temperature, and the strain is environmentally friendly, toxic-free, pollution-free and good in biodegradability; a high-temperature strong-alkali environment required by a traditional chemical method oil removing is avoided, the energy consumption and the operation danger coefficient are decreased, and the secondary environmental pollution is avoided; and the strain is stable in performance and has good industrial application value.
Description
Technical field
The present invention relates to the application of the acinetobacter calcoaceticus of biological technical field, be specifically related to acinetobacter calcoaceticus strain and the application thereof of a strain biosurfactant production.
Background technology
Bio-surfactant is the amphipathic compound that microorganism oozy class in metabolic process contains hydrophilic radical (as amino acid or polypeptide, disaccharides, oligosaccharides or polysaccharide etc.) and lipophilic group (as saturated or unsaturated fatty alcohol or lipid acid etc.).Bio-surfactant, except the feature with chemical surfactant, also has following features: (1) has higher surfactivity at oil-water interface; (2) nontoxic, safety, readily biodegradable, environmental friendliness; (3) there is very strong grease emulsifying ability; (4) reaction product can introduce the chemical group of novel type, and wherein some group is that chemical process is difficult to synthesis, and molecular structure type is various; (5) just can using biosurfactant under normal temperature, condition of normal pressure; (6) raw material that fermentation strain produces bio-surfactant extensively to exist and inexpensive at nature, is typical " green " technique.
Bio-surfactant mainly comprises glycolipid, lipopeptid, phosphatide, lipid acid, polysaccharide-composite of lipid and neutral lipid derivative etc., and known bio-surfactant is based on glycolipid.Bio-surfactant is due to features such as its good solubilising, emulsification, reduction tensio-active agent and environment friendly, obtain in environment remediation on the improvement of oily(waste)water, petroleum pollution ground, oil recovery industry, the field such as clean and apply widely, and infiltrate to other field.Research shows, the oil displacement efficiency of bio-surfactant than chemical surfactant oil displacement efficiency height 3.5-8 doubly, and price is only 30% of chemical surfactant.Bio-surfactant substituted chemistry tensio-active agent gradually becomes a kind of new research tendency.
The oil removing of the normal nulling part of metal-plated pre-treatment and rust cleaning, be the important procedure of plating, the quality of electroplated component treatment before plating quality, directly affects electrolytic coating quality.According to statistics, the major cause of plating piece 90% quality accident causes because treatment before plating is improper.The traditional technology of oil removing before metal current plating, mainly contains thermokalite pyrochemistry washing out method, ultrasonic oil removal, chemical reagent except wet goods; Wherein high temperature alkaline process energy consumption is large, seriously polluted is eliminated; Ultrasonic oil removal is large because of facility investment, limits its application in practice; Chemical reagent oil removing is the deoiling method that application is maximum at present, but, there is following shortcoming in traditional chemical reagent oil removing: under normal temperature, deoiling effect is poor, must heat during use (mostly more than 70 DEG C), deoiling effect dies down gradually along with the aging of tank liquor, not only waste resource but also caused the secondary pollution of greasy dirt, body refuse, also there is the problems such as high spumescence, high alkalinity and seriously corroded.Therefore a kind of less energy-consumption, safety and environmental protection, easily degraded, high efficiency degreaser is badly in need of in industrial production.
Summary of the invention
The object of the invention is the bacterial strain by filtering out a strain biosurfactant production, the bio-surfactant utilizing this bacterial strain to produce, reaches normal temperature oil removing, and biodegradable, free of contamination effect, be applied to Metal plating pre-treatment.
Technical scheme of the present invention is: adopt enrichment culture, dilution spread, purifying to obtain strain X H-2 from petroleum pollution, through identifying that this bacterium is acinetobacter (Acinetobactersp.XH-2).And submitted to China General Microbiological culture presevation administrative center to carry out culture presevation.
Enrichment medium used is (w/v, %): magnesium chloride 2, dipotassium hydrogen phosphate 6, potassium primary phosphate 3.8, iron trichloride 0.5, ammonium chloride 10, and additional cosmoline carries out enrichment culture as sole carbon source.
Fermention medium used is (w/v, %): whey powder 0.1, SODIUMNITRATE 0.1, potassium primary phosphate 0.2, Sodium phosphate dibasic 0.6, magnesium chloride hexahydrate 0.02, sodium-chlor 2, micro-1mL/L, Tween801, pH are 6, and distilled water is prepared.
Described strain X H-2 is fermentation culture 48h in described fermention medium, 4 DEG C of centrifugal 20min of 10000rpm, get supernatant liquor 6mol/LHCl and adjust pH to 2.0, after extracting 2 times with isopyknic ethyl acetate solution, merge organic phase concentrated on Rotary Evaporators, concentrated solution is poured in beaker, after naturally volatilizing, namely obtain the crude product of tensio-active agent.
In order to study the characteristic of this bio-surfactant further, this crude product is used to carry out composition analysis, the mensuration of micelle-forming concentration and stability analysis.
The bio-surfactant main component that the present invention announces, is tentatively judged as glycolipid class through qualitative analysis and the results of FT-IR.
After measured, the micelle-forming concentration of this bio-surfactant is 200mg/L, significantly lower than conventional chemical surfactant as sodium laurylsulfonate (2120mg/L), cetyl trimethylammonium bromide (1300mg/L).
Be applied to the removal that length is respectively 4cm, 2cm and 0.3cm steel disc surface anticorrosion oil, deoiling effect is good, and the time is short, only needs 16min.
The advantage that the present invention has and positively effect are:
Applicant screens the bacterial strain of a strain biosurfactant production from petroleum pollution, called after acinetobacter calcoaceticus XH-2, adopt the method for biological fermentation, to extract the byproduct whey powder after cheese, SODIUMNITRATE, inorganic salt for raw material, acinetobacter calcoaceticus XH-2 is utilized to produce Glycolipids Biosurfactants via, this bio-surfactant, under normal temperature condition, can remove the cosmoline of metallic surface.
Carbon source used in the present invention is tankage whey powder remaining after extracting cheese, and cheap, cost is low; Microbial reproduction is fast, metabolism is fast, using the bio-surfactant of microbial metabolism generation as biological degreasing agent, namely it play a role at normal temperatures, product environmental protection, nontoxic pollution-free, biological degradability are good, avoid the high temperature strong alkali environment needed for conventional chemical methods oil removing, reduce energy consumption and operational hazards coefficient, avoid secondary environmental pollution; Bio-surfactant made by the present invention is after the high Inversion phenomenon of difference, pH process, and surface tension variations is little, and after this bio-surfactant boils 10min through 100 DEG C, its oil removal efficiency does not reduce, and has good industrial application value.
Accompanying drawing explanation
Fig. 1 is the Determination of Critical Micelle Concentration figure of strain X H-2 of the present invention institute biosurfactant production
Fig. 2 is the infrared absorpting light spectra of bio-surfactant of the present invention
Fig. 3 is the design sketch of bio-surfactant of the present invention for oil removing before metal plating
Embodiment
The screening of acinetobacter calcoaceticus (Acinetobactersp.XH-2) bacterial strain of embodiment 1 biosurfactant production.
Sample from by the soil of petroleum pollution, the sample that the present embodiment uses is on-the-spot by the soil sample of crude oil pollution for Shandong region Shengli Oil Field recovers the oil, take 5g soil sample at 50mL enrichment medium (w/v, %: magnesium chloride 2, dipotassium hydrogen phosphate 6, potassium primary phosphate 3.8, iron trichloride 0.5, ammonium chloride 10, additional cosmoline is as sole carbon source) in cultivate, take out 1mL nutrient solution to be forwarded to again in enrichment medium, transfer after 3 times; Dilution spread, on blood agar, carries out line purifying to there being the bacterial strain of transparent circle on LB solid medium.
By single colony inoculation good for purifying in the LB liquid nutrient medium of 5mL, fermention medium (the w/v do not optimized is transferred to after activation 12h, %: glucose 1, peptone 1, potassium primary phosphate 0.2, Sodium phosphate dibasic 0.6, magnesium chloride hexahydrate 0.02, micro-1mL/L) in, after fermentation culture 2d, thalline abandoned by centrifugal fermented liquid, do the experiment of oil extraction circle with supernatant liquor and utilize JK9913 type Full-automatic tension instrument to adopt suspension ring method chart surface tension, and carrying out oil removing experiment.
Its oil extraction loop diameter of the strain X H-2 screened is 4.5cm, and surface tension is 32.58mN/m, has certain deoiling effect.
Select strain X H-2 as the original strain of biological degreasing agent, through identifying that this Pseudomonas is in acinetobacter.
The fermentation technology optimization of embodiment 2 strain X H-2.
With surface tension and oil removing time for index, the fermention medium of strain X H-2 and fermentation condition are optimized.
From LB solid plate, the mono-colony inoculation of picking strain X H-2 is in the LB liquid nutrient medium of 5mL, and after activation 12h, be transferred in fermention medium, the rotating speed of shaking table is 160rpm, and temperature is set to 30 DEG C.
Using glucose, sucrose, lactose, whey powder, glycerine, sweet oil, deep-fried twisted dough sticks waste oil as carbon source, using peptone, SODIUMNITRATE, ammonium sulfate, yeast extract (ferment is carried), dregs of beans as nitrogenous source, after fermentation 2d, carry out the mensuration of surface tension and oil removing time, the results are shown in Table 1 and 2, mainly determine the suitableeest carbon nitrogen source according to the oil removing time; And then carrying out concentration optimization, carbon source concentration is set to (w/v, %) 0.1,0.5,1.5,3, is set to nitrogen concentration (w/v, %) and is set to 0.1,0.2,1,3; Determine that carbon source is 0.1% whey powder, nitrogenous source is 0.1% SODIUMNITRATE.
Initial pH is set to 5,6,7,8,9,10, additional 1%Tween80 and SDS tensio-active agent.
Fermention medium (w/v, %) after optimization is: whey powder 0.1, SODIUMNITRATE 0.1, potassium primary phosphate 0.2, Sodium phosphate dibasic 0.6, magnesium chloride hexahydrate 0.02, sodium-chlor 2, micro-1mL/L, Tween801, pH are 6, and distilled water is prepared.
Fermentation condition comprises inoculum size and plants the optimization in age, and inoculum size (v/v, %) is set to 1,2,3,4,8; By be in early growth period (6h), exponential phase (12h), stationary phase (20h), decline phase (30h) bacterial strain be transferred in the fermention medium optimized, determining optimum inoculation amount 4% and planting age is 12h.
Table 1 different carbon source kind to the metallic surface oil removing time and produce the surface tension table look-up of tensio-active agent
Table 2 different nitrogen sources kind to the metallic surface oil removing time and produce the surface tension table look-up of tensio-active agent
Embodiment 3 utilizes the strain X H-2 bio-surfactant produced that ferments to prepare biological degreasing agent.
The mono-bacterium colony of strain X H-2 on picking LB solid plate is after 5mLLB liquid nutrient medium activation 12h, be transferred in the fermention medium of embodiment 2 optimization by 4% inoculum size, at 30 DEG C, 160rpm shaking table top fermentation cultivation 2d, get in supernatant liquor culture dish holding after fermented liquid is centrifugal, by long 4cm, wide 2cm, high 0.3cm steel disc is placed in one, and observes and records the time needed for oil removing.The results are shown in A in accompanying drawing 3, figure and be depicted as steel disc surface coverage oil film, in figure, B is depicted as the oil film on steel disc surface by Ex-all, and oil removing shortest time is, after for some time placed by steel disc after C is depicted as oil removing in 16min, figure, occur iron rust, show that steel disc oil film is divided.
Embodiment 4 utilizes strain X H-2 fermentation culture to prepare bio-surfactant, and carries out the research of the mensuration of micelle-forming concentration, composition preliminary evaluation and stability.
The fermented liquid of strain X H-2 fermentation culture 2d is obtained according to the method for embodiment 3, in 4 DEG C, the centrifugal 20min of rotating speed of 10000rpm, get supernatant liquor 6mol/LHCl and adjust pH to 2.0, after extracting 2 times with isopyknic ethyl acetate solution, merge organic phase concentrated on Rotary Evaporators, poured into by concentrated solution in beaker, after naturally volatilizing, namely obtain the crude product of tensio-active agent, output is 3.1g/L.
Obtained crude product is made into 10,20,50,100,200,300,400,500, the aqueous solution of 1000mg/L, measure its surface tension respectively and be depicted as curve, seeing shown in accompanying drawing 1.When bio-surfactant solution is from 0-50mg/L, surface tension drops to 38.58mN/m very soon from 75mN/m; Later decline is slow; When surfactant concentration reaches 200mg/L, surface tension reduces to 33.12mN/m; Sample concentration is higher than after 200mg/L, and the surface tension of solution almost no longer declines; Therefore, the CMC value of this tensio-active agent is 200mg/L, is starkly lower than conventional chemical surfactant sodium laurylsulfonate (SDS, 2120mg/L), cetyl trimethylammonium bromide (CTAB, 1300mg/L), its good surfactivity is more conducive to practical application.
Methylene blue-chloroform test is adopted to identify the charging property of this bio-surfactant.In methylene blue-chloroform test, above-mentioned tensio-active agent crude product is made into the solution of 0.3g/L, gets 5mL surfactant soln, add 10mL methylene blue and 5mL chloroform successively, fully after mixing, observe after leaving standstill several minutes.Experimental result is that water layer darkens, and shows that this bio-surfactant is cationic surfactant.
Tensio-active agent crude product KBr pressed disc method is carried out infrared spectra (FT-IR) analysis.The results are shown in accompanying drawing 2, as can be seen from the figure, 3390cm
-1near there is strong and wide absorption peak, show to there is a large amount of-OH in this molecule; 2924cm
-1and 2854cm
-1absorption vibration belong to the absorption peak of aliphatics C-H, and 1460cm
-1neighbouring absorption peak can belong to continuous print C-H vibration on molecule carbon chain to be caused; 1671cm
-1the absorption peak at place from the stretching vibration of carbonyl, 1728cm
-1near have the comparatively strong and absorption peak of point, show the existence having ester group; 1074cm
-1with 1025cm
-1the absorption peak at place, from the stretching vibration of C-O-C key, shows to there is carboxylate group in this molecule.With the glycolipid class infrared spectrogram contrast delivered, tentatively determine that this bio-surfactant is glycolipid compound.
Above-mentioned tensio-active agent crude product is made into the solution of 0.3g/L, carries out the stability analysis of this bio-surfactant under differing temps, pH and salinity process.
(1) temperature stability: this bio-surfactant solution is after (4 DEG C, room temperature, 60 DEG C, 100 DEG C) place 30min at different temperature, chart surface tension;
(2) pH stability: this bio-surfactant crude product different pH (2,4,6,8,10,12) buffer solution, after room temperature places 12h, chart surface tension;
(3) salinity stability: the solution of this bio-surfactant crude product different salinity (0-9%) is made into the solution that concentration is 0.3g/L, after room temperature places 12h, chart surface tension.
The results are shown in following table 3, under differing temps, pH and salinity process, surface tension variations is little, and this bio-surfactant has good stability and tolerance.The supernatant liquor of fermented liquid is after boiling water bath process 10min, and the impact of its oil removal efficiency is little.
Table 3 temperature, pH and salinity are on the impact of bio-surfactant stability
Above one embodiment of the present of invention have been described in detail, but described content being only preferred embodiment of the present invention, can not being considered to for limiting practical range of the present invention.All equalizations done according to the present patent application scope change and improve, and all should still belong within patent covering scope of the present invention.
Claims (5)
1. the acinetobacter calcoaceticus strain (Acinetobactersp.XH-2) of a strain biosurfactant production, is characterized in that: can produce bio-surfactant, and submits to China General Microbiological culture presevation administrative center to carry out culture presevation.
2. the acinetobacter calcoaceticus strain of a strain biosurfactant production according to claim 1, it is characterized in that: the screening method of described bacterial strain is: sample from by the soil of petroleum pollution, take 5g soil sample in 50mL enrichment medium, described enrichment medium (w/v, %) be: magnesium chloride 2, dipotassium hydrogen phosphate 6, potassium primary phosphate 3.8, iron trichloride 0.5, ammonium chloride 10 that additional cosmoline is as sole carbon source; Take out 1mL nutrient solution to be forwarded in enrichment medium, transfer after 3 times, dilution spread, on blood agar, carries out line purifying to there being the bacterial strain of transparent circle on LB solid medium again.
3. utilize the acinetobacter calcoaceticus strain of the strain biosurfactant production described in claim 1 to produce the method for bio-surfactant, it is characterized in that: from LB solid plate, picking list colony inoculation is in the LB liquid nutrient medium of 5mL, after activation 12h, be transferred in fermention medium, inoculum size 4%, fermention medium (w/v, %) be: whey powder 0.1, SODIUMNITRATE 0.1, potassium primary phosphate 0.2, Sodium phosphate dibasic 0.6, magnesium chloride hexahydrate 0.02, sodium-chlor 2, trace element 1mL/L, Tween801, pH is 6, distilled water is prepared, shaking speed is 160rpm, temperature is set to 30 DEG C, cultivate 2d, in 4 DEG C, the centrifugal 20min of 10000rpm, get supernatant liquor 6mol/LHCl and adjust pH to 2.0, after extracting 2 times with isopyknic ethyl acetate solution, merge organic phase concentrated on Rotary Evaporators, concentrated solution is poured in beaker, namely the crude product of tensio-active agent is obtained after naturally volatilizing.
4. method obtains bio-surfactant according to claim 3.
5. the application of bio-surfactant according to claim 4 before metal plating in oil removing.
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| CN111394271A (en) * | 2019-09-12 | 2020-07-10 | 浙江大学 | Acinetobacter for producing surfactant and application thereof |
| CN114075511A (en) * | 2021-11-22 | 2022-02-22 | 山东科环环境管理咨询有限公司 | Petroleum-polluted soil remediation agent and preparation process thereof |
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| CN111394271A (en) * | 2019-09-12 | 2020-07-10 | 浙江大学 | Acinetobacter for producing surfactant and application thereof |
| CN114075511A (en) * | 2021-11-22 | 2022-02-22 | 山东科环环境管理咨询有限公司 | Petroleum-polluted soil remediation agent and preparation process thereof |
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