CN113960141B - Preparation method of immobilized microbial membrane for measuring BOD of seawater - Google Patents
Preparation method of immobilized microbial membrane for measuring BOD of seawater Download PDFInfo
- Publication number
- CN113960141B CN113960141B CN202111131062.2A CN202111131062A CN113960141B CN 113960141 B CN113960141 B CN 113960141B CN 202111131062 A CN202111131062 A CN 202111131062A CN 113960141 B CN113960141 B CN 113960141B
- Authority
- CN
- China
- Prior art keywords
- electrode
- seawater
- bod
- preparing
- steps
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
- 230000000813 microbial effect Effects 0.000 title claims abstract description 70
- 239000013535 sea water Substances 0.000 title claims abstract description 46
- 239000012528 membrane Substances 0.000 title claims abstract description 35
- 238000002360 preparation method Methods 0.000 title claims abstract description 16
- 238000002484 cyclic voltammetry Methods 0.000 claims abstract description 34
- 244000005700 microbiome Species 0.000 claims abstract description 21
- 230000004060 metabolic process Effects 0.000 claims abstract description 4
- 230000000379 polymerizing effect Effects 0.000 claims abstract description 3
- 239000000243 solution Substances 0.000 claims description 38
- 239000000725 suspension Substances 0.000 claims description 34
- 238000000034 method Methods 0.000 claims description 31
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 claims description 22
- 230000001580 bacterial effect Effects 0.000 claims description 22
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 claims description 20
- 239000010931 gold Substances 0.000 claims description 20
- 239000011259 mixed solution Substances 0.000 claims description 20
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 19
- 239000001963 growth medium Substances 0.000 claims description 17
- 229960000907 methylthioninium chloride Drugs 0.000 claims description 17
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 claims description 14
- 229910052737 gold Inorganic materials 0.000 claims description 14
- 239000000126 substance Substances 0.000 claims description 13
- 239000010936 titanium Substances 0.000 claims description 12
- 241000194108 Bacillus licheniformis Species 0.000 claims description 11
- 239000010802 sludge Substances 0.000 claims description 11
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 claims description 10
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 10
- HSFWRNGVRCDJHI-UHFFFAOYSA-N alpha-acetylene Natural products C#C HSFWRNGVRCDJHI-UHFFFAOYSA-N 0.000 claims description 10
- 239000007853 buffer solution Substances 0.000 claims description 10
- 239000008367 deionised water Substances 0.000 claims description 10
- 229910021641 deionized water Inorganic materials 0.000 claims description 10
- 238000001035 drying Methods 0.000 claims description 10
- 229910052757 nitrogen Inorganic materials 0.000 claims description 10
- 229920001197 polyacetylene Polymers 0.000 claims description 10
- 229920000123 polythiophene Polymers 0.000 claims description 10
- 238000000926 separation method Methods 0.000 claims description 10
- 238000009210 therapy by ultrasound Methods 0.000 claims description 10
- 241000589517 Pseudomonas aeruginosa Species 0.000 claims description 8
- BQCADISMDOOEFD-UHFFFAOYSA-N Silver Chemical compound [Ag] BQCADISMDOOEFD-UHFFFAOYSA-N 0.000 claims description 8
- RTAQQCXQSZGOHL-UHFFFAOYSA-N Titanium Chemical compound [Ti] RTAQQCXQSZGOHL-UHFFFAOYSA-N 0.000 claims description 8
- 229920000767 polyaniline Polymers 0.000 claims description 8
- -1 polyphenylene ethylene Polymers 0.000 claims description 8
- 229910052719 titanium Inorganic materials 0.000 claims description 8
- 241000222120 Candida <Saccharomycetales> Species 0.000 claims description 7
- 229920000015 polydiacetylene Polymers 0.000 claims description 6
- 241000193171 Clostridium butyricum Species 0.000 claims description 5
- XDTMQSROBMDMFD-UHFFFAOYSA-N Cyclohexane Chemical compound C1CCCCC1 XDTMQSROBMDMFD-UHFFFAOYSA-N 0.000 claims description 4
- 241000588724 Escherichia coli Species 0.000 claims description 4
- 241000588748 Klebsiella Species 0.000 claims description 4
- 239000003960 organic solvent Substances 0.000 claims description 4
- 239000002609 medium Substances 0.000 claims description 3
- 241000235648 Pichia Species 0.000 claims description 2
- RBTBFTRPCNLSDE-UHFFFAOYSA-N 3,7-bis(dimethylamino)phenothiazin-5-ium Chemical compound C1=CC(N(C)C)=CC2=[S+]C3=CC(N(C)C)=CC=C3N=C21 RBTBFTRPCNLSDE-UHFFFAOYSA-N 0.000 claims 3
- 241001052560 Thallis Species 0.000 claims 1
- 230000001737 promoting effect Effects 0.000 claims 1
- 238000005259 measurement Methods 0.000 abstract description 13
- 238000012258 culturing Methods 0.000 description 21
- 230000000052 comparative effect Effects 0.000 description 14
- 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 14
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 9
- 229920001817 Agar Polymers 0.000 description 8
- 239000008272 agar Substances 0.000 description 8
- 239000004020 conductor Substances 0.000 description 8
- 239000002504 physiological saline solution Substances 0.000 description 8
- 238000005406 washing Methods 0.000 description 8
- 239000004793 Polystyrene Substances 0.000 description 7
- 238000001514 detection method Methods 0.000 description 7
- SQHOAFZGYFNDQX-UHFFFAOYSA-N ethyl-[7-(ethylamino)-2,8-dimethylphenothiazin-3-ylidene]azanium;chloride Chemical compound [Cl-].S1C2=CC(=[NH+]CC)C(C)=CC2=NC2=C1C=C(NCC)C(C)=C2 SQHOAFZGYFNDQX-UHFFFAOYSA-N 0.000 description 7
- 239000006916 nutrient agar Substances 0.000 description 7
- 229920002223 polystyrene Polymers 0.000 description 7
- 241000894006 Bacteria Species 0.000 description 5
- 241000320412 Ogataea angusta Species 0.000 description 5
- 229920000265 Polyparaphenylene Polymers 0.000 description 5
- 239000000499 gel Substances 0.000 description 5
- 239000000648 calcium alginate Substances 0.000 description 4
- 235000010410 calcium alginate Nutrition 0.000 description 4
- 229960002681 calcium alginate Drugs 0.000 description 4
- OKHHGHGGPDJQHR-YMOPUZKJSA-L calcium;(2s,3s,4s,5s,6r)-6-[(2r,3s,4r,5s,6r)-2-carboxy-6-[(2r,3s,4r,5s,6r)-2-carboxylato-4,5,6-trihydroxyoxan-3-yl]oxy-4,5-dihydroxyoxan-3-yl]oxy-3,4,5-trihydroxyoxane-2-carboxylate Chemical compound [Ca+2].O[C@@H]1[C@H](O)[C@H](O)O[C@@H](C([O-])=O)[C@H]1O[C@H]1[C@@H](O)[C@@H](O)[C@H](O[C@H]2[C@H]([C@@H](O)[C@H](O)[C@H](O2)C([O-])=O)O)[C@H](C(O)=O)O1 OKHHGHGGPDJQHR-YMOPUZKJSA-L 0.000 description 4
- 230000035945 sensitivity Effects 0.000 description 4
- 239000007788 liquid Substances 0.000 description 3
- 238000006243 chemical reaction Methods 0.000 description 2
- 238000004132 cross linking Methods 0.000 description 2
- 230000007423 decrease Effects 0.000 description 2
- 238000013048 microbiological method Methods 0.000 description 2
- 238000012544 monitoring process Methods 0.000 description 2
- 230000008569 process Effects 0.000 description 2
- 238000001179 sorption measurement Methods 0.000 description 2
- 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 description 1
- DEXFNLNNUZKHNO-UHFFFAOYSA-N 6-[3-[4-[2-(2,3-dihydro-1H-inden-2-ylamino)pyrimidin-5-yl]piperidin-1-yl]-3-oxopropyl]-3H-1,3-benzoxazol-2-one Chemical compound C1C(CC2=CC=CC=C12)NC1=NC=C(C=N1)C1CCN(CC1)C(CCC1=CC2=C(NC(O2)=O)C=C1)=O DEXFNLNNUZKHNO-UHFFFAOYSA-N 0.000 description 1
- 241000193830 Bacillus <bacterium> Species 0.000 description 1
- 239000002028 Biomass Substances 0.000 description 1
- UXVMQQNJUSDDNG-UHFFFAOYSA-L Calcium chloride Chemical compound [Cl-].[Cl-].[Ca+2] UXVMQQNJUSDDNG-UHFFFAOYSA-L 0.000 description 1
- 239000004952 Polyamide Substances 0.000 description 1
- 239000004372 Polyvinyl alcohol Substances 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000015556 catabolic process Effects 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 230000008878 coupling Effects 0.000 description 1
- 238000010168 coupling process Methods 0.000 description 1
- 238000005859 coupling reaction Methods 0.000 description 1
- 239000003431 cross linking reagent Substances 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 238000006731 degradation reaction Methods 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 229920002521 macromolecule Polymers 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 235000015097 nutrients Nutrition 0.000 description 1
- 230000003204 osmotic effect Effects 0.000 description 1
- 229920002647 polyamide Polymers 0.000 description 1
- 229920002451 polyvinyl alcohol Polymers 0.000 description 1
- 238000011084 recovery Methods 0.000 description 1
- 230000029058 respiratory gaseous exchange Effects 0.000 description 1
- 239000010865 sewage Substances 0.000 description 1
- 229910052709 silver Inorganic materials 0.000 description 1
- 239000004332 silver Substances 0.000 description 1
- 239000000661 sodium alginate Substances 0.000 description 1
- 235000010413 sodium alginate Nutrition 0.000 description 1
- 229940005550 sodium alginate Drugs 0.000 description 1
- 239000011780 sodium chloride Substances 0.000 description 1
- 238000007711 solidification Methods 0.000 description 1
- 230000008023 solidification Effects 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 238000013112 stability test Methods 0.000 description 1
- 230000004083 survival effect Effects 0.000 description 1
- 230000008961 swelling Effects 0.000 description 1
- 230000002195 synergetic effect Effects 0.000 description 1
- 238000012546 transfer Methods 0.000 description 1
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N27/00—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
- G01N27/26—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating electrochemical variables; by using electrolysis or electrophoresis
- G01N27/28—Electrolytic cell components
- G01N27/30—Electrodes, e.g. test electrodes; Half-cells
- G01N27/327—Biochemical electrodes, e.g. electrical or mechanical details for in vitro measurements
- G01N27/3275—Sensing specific biomolecules, e.g. nucleic acid strands, based on an electrode surface reaction
-
- 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
- C12N1/14—Fungi; Culture media therefor
- C12N1/16—Yeasts; Culture media therefor
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
-
- 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
- C12N1/36—Adaptation or attenuation of cells
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/18—Water
- G01N33/1806—Biological oxygen demand [BOD] or chemical oxygen demand [COD]
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Wood Science & Technology (AREA)
- Zoology (AREA)
- Organic Chemistry (AREA)
- Genetics & Genomics (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biotechnology (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- Medicinal Chemistry (AREA)
- Biomedical Technology (AREA)
- Virology (AREA)
- Tropical Medicine & Parasitology (AREA)
- Molecular Biology (AREA)
- General Engineering & Computer Science (AREA)
- Microbiology (AREA)
- Physics & Mathematics (AREA)
- Mycology (AREA)
- Analytical Chemistry (AREA)
- General Physics & Mathematics (AREA)
- Pathology (AREA)
- Immunology (AREA)
- Food Science & Technology (AREA)
- Electrochemistry (AREA)
- Botany (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Spectroscopy & Molecular Physics (AREA)
- Emergency Medicine (AREA)
- Biodiversity & Conservation Biology (AREA)
- Cell Biology (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
- Immobilizing And Processing Of Enzymes And Microorganisms (AREA)
Abstract
The invention provides a preparation method of an immobilized microbial membrane for measuring seawater BOD, which comprises the following steps: s1, domesticating microorganisms suitable for high-salinity seawater; s2, generating a layer of electron mediator which promotes microorganism metabolism on the surface of the electrode; and S3, polymerizing the conductive solution doped with microorganisms on the surface of the electronic medium layer of the electrode by a cyclic voltammetry to form a microbial film with a double-layer structure on the surface of the electrode. The microbial film has the advantages of high measurement accuracy, good stability and long service life, and can realize the rapid and accurate measurement of the BOD of the high-salt seawater.
Description
Technical Field
The invention relates to the technical field of sewage treatment, in particular to a preparation method of an immobilized microbial membrane for measuring seawater BOD.
Background
Microbial membranes are a central part of rapid BOD measurement, and generally use single species or mixed species of microorganisms as biosensing elements. When BOD substances are added into water or degradation metabolism occurs, the change or conversion of the internal and external source respiration modes of microorganisms is caused, the intensity of the coupling output current is changed, the current value output by the sensor and the BOD concentration are in a linear relation under certain conditions, and the BOD concentration of a sample can be measured according to the linear relation between the current value and the BOD concentration.
The existing microorganism fixing method comprises an adsorption method, a crosslinking method and an embedding method, but the adsorption method has the problems that microorganisms are easy to fall off and the service life is influenced; the embedding method is not suitable for the reaction of macromolecular substances, and the embedding material is easy to introduce larger mass transfer resistance when in use; the survival rate of the immobilized microbial film by the crosslinking method is low, the preparation is complex, and the dosage of the crosslinking agent and the microbial biomass are mutually restricted.
The existing calcium alginate embedding method and polyvinyl alcohol embedding method have the defects of easy swelling, weak acid-base adaptability, no osmotic pressure resistance, easy absorption of colored substances, more flocs and the like to different degrees, and influence the detection stability and sensitivity. Therefore, a microbial membrane fixing method with strong stability and high sensitivity and capable of detecting BOD of high-salinity seawater needs to be developed.
Disclosure of Invention
In view of the above, the invention provides a microbial membrane with high sensitivity, strong stability, suitability for high salinity seawater, good repeatability and long service life and a preparation method thereof.
The technical scheme of the invention is realized as follows: the invention provides a preparation method of an immobilized microbial membrane for measuring seawater BOD, which comprises the following steps:
s1, domesticating microorganisms suitable for high-salinity seawater;
s2, generating a layer of electron mediator which promotes microorganism metabolism on the surface of the electrode;
s3, polymerizing the conductive solution doped with domesticated microorganisms on the surface of the electronic medium layer of the electrode by a cyclic voltammetry to form a microbial film with a double-layer structure on the surface of the electrode.
Based on the above technical solution, preferably, the operation steps of step S1 are as follows:
s11, taking microorganisms separated from seawater sludge, and carrying out domestication and 2-3 times of plate separation to obtain pure bacterial strains;
s12, the pure strain in the step S11 is subjected to shaking culture, is inoculated to a culture medium after being washed by a sterile buffer solution, and is prepared into bacterial suspension of 8X 109CFU/mL for standby when the concentration of the bacterial body reaches 10X 109 CFU/mL.
Based on the above technical solution, preferably, the operation steps of step S2 are as follows:
s21, sequentially carrying out ultrasonic treatment on the electrode in an organic solvent and deionized water for 5min, drying by nitrogen, immersing in 0.1-0.3mol/L sulfuric acid, and scanning by adopting an electrochemical cyclic voltammetry to obtain a working electrode;
and S22, doping the methylene blue on the surface of the working electrode in the step S21 by electrochemical cyclic voltammetry to obtain the electrode covered with the methylene blue film.
On the basis of the technical scheme, preferably, the organic solvent is one or a combination of more of ethanol, acetone and cyclohexane.
Based on the above technical solution, preferably, the operation steps of step S3 are as follows:
s31, dissolving a conductive substance in the bacterial suspension in the step S12 to prepare a mixed solution;
s32, immersing the electrode covered with the methylene blue film in the step S22 into the mixed solution in the step S31, and scanning by adopting electrochemical cyclic voltammetry to obtain the microbial film electrode covered with the double-layer structure.
On the basis of the technical scheme, preferably, the conductive substance is one or a combination of more of polyacetylene, polythiophene, polyaniline, polyphenylene, polystyrene and polydiacetylene, and the concentration is 0.01-0.03mol/L.
On the basis of the above technical scheme, preferably, the microorganism in the step S1 is one of bacillus licheniformis, candida, hansenula, klebsiella, escherichia coli, clostridium butyricum and pseudomonas aeruginosa.
On the basis of the above technical solution, preferably, the electrode in the step S2 is a disc electrode, and the diameter of the disc electrode is 1-5mm.
On the basis of the technical scheme, preferably, the disc electrode is one of a gold disc electrode, a silver disc electrode and a titanium disc electrode.
Compared with the prior art, the preparation method of the immobilized microbial film for measuring the BOD of the seawater has the following beneficial effects:
(1) The microbial film of the invention is used for measuring the BOD of seawater, and has the advantages of short measurement time and real-time monitoring on site; the classical 5 day method takes 5 days in the whole measurement process and cannot be detected in real time on site, and the sensor method of the microbiological method can finish measurement within 10 minutes.
(2) The microbial film of the invention has high measurement accuracy, and can reduce the error of the general BOD rapid detection method from +/-20% to +/-5%.
(3) The microbial film of the invention also has the advantages of good stability and long service life, and the stable service life exceeds 3 months, and the service life of the far-reaching calcium alginate gel is 1 month.
(4) The strain used in the invention is separated and domesticated from the seawater sludge, so that the rapid and accurate measurement of the BOD of the high-salt seawater can be realized.
Drawings
In order to more clearly illustrate the embodiments of the invention or the technical solutions in the prior art, the drawings that are required in the embodiments or the description of the prior art will be briefly described, it being obvious that the drawings in the following description are only some embodiments of the invention, and that other drawings may be obtained according to these drawings without inventive effort for a person skilled in the art.
FIG. 1 is a standard graph of an immobilized microbial membrane for measuring BOD of seawater according to the present invention;
FIG. 2 is a graph showing the results of a microbial film stability test of an immobilized microbial film for measuring BOD of seawater according to the present invention.
Detailed Description
The following description of the embodiments of the present invention will clearly and fully describe the technical aspects of 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, which can be made by those skilled in the art based on the embodiments of the present invention without making any inventive effort, are intended to fall within the scope of the present invention.
Example 1
The preparation process of immobilized microbial film for measuring BOD of sea water includes the following steps:
s1, preparing strain suspension; the strain is selected from bacillus licheniformis separated from seawater sludge, and the selected bacillus licheniformis is domesticated and subjected to plate separation for 2 times to obtain a pure strain. Shake culturing pure strain in culture medium solution, PBS solution and 0.8% sodium chloride salt water at 30deg.C and 140rpm for 24 hr, centrifuging at 25deg.C and 7000r/min for 20min, washing with equal volume of sterile buffer solution for 2 times to obtain Bacillus licheniformis suspension, inoculating Bacillus licheniformis to nutrient agar plate, culturing for 6 hr, and placing in refrigerator at 4deg.C. Before use, a loop of strain is taken from the agar plane and inoculated into 200mL of sterile culture medium again for culturing for 12h to ensure that the concentration of the strain reaches 10 multiplied by 10 9 CFU/mL, diluted with sterile physiological saline to obtain 8×10 9 The CFU/mL bacterial suspension is ready for use.
S2, preparing a working electrode, namely sequentially carrying out ultrasonic treatment on a gold plate electrode with the diameter of 1mm in ethanol and deionized water for 5min respectively, drying by nitrogen, then putting the gold plate electrode into a 0.1mol/L dilute sulfuric acid solution, and scanning for a plurality of times by adopting an electrochemical cyclic voltammetry within the range of-0.2-1.2V until the superposition of scanning curves is stable, and taking the gold plate electrode as the working electrode for standby.
S3, preparing a microbial membrane electrode; the methylene blue is doped on the surface of a working electrode by an electrochemical cyclic voltammetry to obtain a new methylene blue film NMB/Au; then preparing a conductive material containing 0.01mol/L and 8×10 9 A mixed solution of a CFU/mL concentration bacillus licheniformis suspension; and (3) immersing NMB/Au into the mixed solution, and obtaining the microbial membrane electrode with a double-layer structure by adopting an electrochemical cyclic voltammetry.
The conductive substances are polyacetylene, polythiophene and polystyrene, and polyacetylene: polythiophene: the mass ratio of the polyphenylene ethylene is 1:1:1.
Example 2
The preparation process of immobilized microbial film for measuring BOD of sea water includes the following steps:
s1, preparing strain suspension; the strain is selected from Hansenula polymorpha separated from seawater sludge, and the Hansenula polymorpha after selection is domesticated and subjected to plate separation for 3 times to obtain a pure strain. Shake culturing pure strain in culture medium solution, PBS solution and 0.8% sodium chloride salt water at 30deg.C and 140rpm for 24 hr, centrifuging cultured Hansenula polymorpha at 25deg.C and 7000r/min for 20min, washing with equal volume of sterile buffer solution for 2 times to obtain Hansenula polymorpha suspension, inoculating Hansenula polymorpha to nutrient agar plate, culturing for 6 hr, and placing in refrigerator at 4deg.C. Before use, a loop of strain is taken from the agar plane and inoculated into 200mL of sterile culture medium again for culturing for 12h to ensure that the concentration of the strain reaches 10 multiplied by 10 9 CFU/mL, diluted with sterile physiological saline to obtain 8×10 9 The CFU/mL bacterial suspension is ready for use.
S2, preparing a working electrode, namely sequentially carrying out ultrasonic treatment on a silver plate electrode with the diameter of 5mm in acetone and deionized water for 5min respectively, drying by nitrogen, then putting the silver plate electrode into a 0.3mol/L dilute sulfuric acid solution, and scanning for a plurality of times by adopting an electrochemical cyclic voltammetry within the range of-0.2-1.2V until the superposition of scanning curves is stable, and taking the silver plate electrode as the working electrode for standby.
S3, preparing a microbial membrane electrode; the methylene blue is doped on the surface of a working electrode by an electrochemical cyclic voltammetry to obtain a new methylene blue film NMB/Ag; then preparing a conductive material containing 0.03mol/L and 8×10 10 The mixed solution of the microbial bacteria solution with the concentration of CFU/mL; and immersing NMB/Ag into the mixed solution, and obtaining the microbial membrane electrode with a double-layer structure by adopting an electrochemical cyclic voltammetry.
The conductive substances are polyaniline and polyphenylene; polyaniline: the volume ratio of the polyphenylene is 1:1.
Example 3
The preparation process of immobilized microbial film for measuring BOD of sea water includes the following steps:
s1, preparing strain suspension; the strain is selected from colibacillus separated from seawater sludge, and the selected colibacillus is domesticated and subjected to plate separation for 2 times to obtain a pure strain. Shake culturing pure strain in culture medium solution, PBS solution and 0.8% sodium chloride salt water at 30deg.C and 140rpm for 24 hr, centrifuging at 25deg.C and 7000r/min for 20min, washing with equal volume of sterile buffer solution for 2 times to obtain Escherichia coli suspension, inoculating Escherichia coli into nutrient agar plate, culturing for 6 hr, and placing in refrigerator at 4deg.C. Before use, a loop of strain is taken from the agar plane and inoculated into 200mL of sterile culture medium again for culturing for 12h to ensure that the concentration of the strain reaches 10 multiplied by 10 9 CFU/mL, diluted with sterile physiological saline to obtain 8×10 9 The CFU/mL bacterial suspension is ready for use.
S2, preparing a working electrode, namely sequentially carrying out ultrasonic treatment on a titanium disk electrode with the diameter of 2mm in cyclohexane and deionized water for 5min respectively, drying by nitrogen, then putting the titanium disk electrode into a 0.1mol/L dilute sulfuric acid solution, and scanning for a plurality of times by adopting an electrochemical cyclic voltammetry within the range of-0.2-1.2V until the superposition of scanning curves is stable, and taking the titanium disk electrode as the working electrode for standby.
S3, preparing a microbial membrane electrode; the methylene blue is doped on the surface of a working electrode by an electrochemical cyclic voltammetry to obtain a new methylene blue film NMB/Ti; then preparing a conductive material containing 0.01mol/L and 8×10 9 The mixed solution of the microbial bacteria solution with the concentration of CFU/mL; and immersing NMB/Ti into the mixed solution, and obtaining the microbial membrane electrode with a double-layer structure by adopting an electrochemical cyclic voltammetry.
The conductive substances are polystyrene and polydiacetylene, and the polystyrene: the volume ratio of the polydiacetylene is 1:1.
Example 4
The preparation process of immobilized microbial film for measuring BOD of sea water includes the following steps:
s1, preparing strain suspension; the strain is selected from candida separated from seawater sludge, and the pure strain is obtained by domestication and 2-3 times of plate separation of the selected candida. Pure strain was grown in medium solution, PBS solution and 0.8% at 30℃and 140rpmAnd (3) carrying out shake culture on sodium chloride salt water for 24 hours, centrifuging the cultured candida at 25 ℃ and 7000r/min for 20 minutes, obtaining candida suspension after washing for 2 times by using an equal volume of sterile buffer solution, inoculating the candida to a nutrient agar plate, culturing for 6 hours, and placing in a refrigerator at 4 ℃. Before use, a loop of strain is taken from the agar plane and inoculated into 200mL of sterile culture medium again for culturing for 12h to ensure that the concentration of the strain reaches 10 multiplied by 10 9 CFU/mL, diluted with sterile physiological saline to obtain 8×10 9 The CFU/mL bacterial suspension is ready for use.
S2, preparing a working electrode, namely sequentially carrying out ultrasonic treatment on a gold plate electrode with the diameter of 3mm in acetone and deionized water for 5min respectively, drying by nitrogen, then putting the gold plate electrode into a 0.3mol/L dilute sulfuric acid solution, and scanning for a plurality of times by adopting an electrochemical cyclic voltammetry within the range of-0.2-1.2V until the superposition of scanning curves is stable, and taking the gold plate electrode as the working electrode for standby.
S3, preparing a microbial membrane electrode; the methylene blue is doped on the surface of a working electrode by an electrochemical cyclic voltammetry to obtain a new methylene blue film NMB/Au; then preparing a conductive material containing 0.01mol/L and 8×10 9 The mixed solution of the microbial bacteria solution with the concentration of CFU/mL; and (3) immersing NMB/Au into the mixed solution, and obtaining the microbial membrane electrode with a double-layer structure by adopting an electrochemical cyclic voltammetry.
The conductive liquid is polyacetylene, polythiophene and polyaniline, and polyacetylene: polythiophene: the volume ratio of polyaniline is 1:1:1.
Example 5
The preparation process of immobilized microbial film for measuring BOD of sea water includes the following steps:
s1, preparing strain suspension; the strain is selected from klebsiella isolated from seawater sludge, and pure strain is obtained by domestication and 2 times of plate separation of the selected klebsiella. Shake culturing pure strain in culture medium solution, PBS solution and 0.8% sodium chloride salt water at 30deg.C and 140rpm for 24 hr, centrifuging at 25deg.C and 7000r/min for 20min, washing with equal volume of sterile buffer solution for 2 times to obtain suspension, and inoculating the strain to nutrient agarThe plates were incubated for 6h and then placed in a refrigerator at 4 ℃. Before use, a loop of strain is taken from the agar plane and inoculated into 200mL of sterile culture medium again for culturing for 12h to ensure that the concentration of the strain reaches 10 multiplied by 10 9 CFU/mL, diluted with sterile physiological saline to obtain 8×10 9 The CFU/mL bacterial suspension is ready for use.
S2, preparing a working electrode, namely sequentially carrying out ultrasonic treatment on a titanium disk electrode with the diameter of 4mm in cyclohexane and deionized water for 5min respectively, drying by nitrogen, then putting the titanium disk electrode into a 0.1mol/L dilute sulfuric acid solution, and scanning for a plurality of times by adopting an electrochemical cyclic voltammetry within the range of-0.2-1.2V until the superposition of scanning curves is stable, and taking the titanium disk electrode as the working electrode for standby.
S3, preparing a microbial membrane electrode; the methylene blue is doped on the surface of a working electrode by an electrochemical cyclic voltammetry to obtain a new methylene blue film NMB/Ti; then preparing a conductive material containing 0.01mol/L and 8×10 9 The mixed solution of the microbial bacteria solution with the concentration of CFU/mL; and immersing NMB/Ti into the mixed solution, and obtaining the microbial membrane electrode with a double-layer structure by adopting an electrochemical cyclic voltammetry.
The conductive substances are polystyrene and polydiacetylene, and the polystyrene: the volume ratio of the polydiacetylene is 1:1.
Example 6
The preparation process of immobilized microbial film for measuring BOD of sea water includes the following steps:
s1, preparing strain suspension; the strain is selected from clostridium butyricum separated from seawater sludge, and the selected strain is domesticated and subjected to plate separation for 3 times to obtain a pure strain. Shake culturing pure strain in culture medium solution, PBS solution and 0.8% sodium chloride salt water at 30deg.C and 140rpm for 24 hr, centrifuging at 25deg.C and 7000r/min for 20min to obtain mixed strain, washing with equal volume of sterile buffer solution for 2 times to obtain clostridium butyricum suspension, inoculating clostridium butyricum into nutrient agar plate, culturing for 6 hr, and placing in refrigerator at 4deg.C. Before use, a loop of strain is taken from the agar plane and inoculated into 200mL of sterile culture medium again for culturing for 12h to ensure that the concentration of the strain reaches 10 multiplied by 10 9 CFU/mL, diluted with sterile physiological saline to obtain 8×10 9 The CFU/mL bacterial suspension is ready for use.
S2, preparing a working electrode, namely sequentially carrying out ultrasonic treatment on a silver plate electrode with the diameter of 5mm in acetone and deionized water for 5min respectively, drying by nitrogen, then putting the silver plate electrode into a 0.3mol/L dilute sulfuric acid solution, and scanning for a plurality of times by adopting an electrochemical cyclic voltammetry within the range of-0.2-1.2V until the superposition of scanning curves is stable, and taking the silver plate electrode as the working electrode for standby.
S3, preparing a microbial membrane electrode; the methylene blue is doped on the surface of a working electrode by an electrochemical cyclic voltammetry to obtain a new methylene blue film NMB/Ag; then preparing a conductive material containing 0.03mol/L and 8×10 10 The mixed solution of the microbial bacteria solution with the concentration of CFU/mL; and immersing NMB/Ag into the mixed solution, and obtaining the microbial membrane electrode with a double-layer structure by adopting an electrochemical cyclic voltammetry.
The conductive substances are polyaniline and polyphenylene; polyaniline: the volume ratio of the polyphenylene is 1:1.
Example 7
The preparation process of immobilized microbial film for measuring BOD of sea water includes the following steps:
s1, preparing strain suspension; the strain is selected from Pseudomonas aeruginosa separated from seawater sludge, and the selected Pseudomonas aeruginosa is domesticated and subjected to plate separation for 2 times to obtain a pure strain. Shake culturing pure strain in culture medium solution, PBS solution and 0.8% sodium chloride salt water at 30deg.C and 140rpm for 24 hr, centrifuging cultured Pseudomonas aeruginosa at 25deg.C and 7000r/min for 20min, washing with equal volume of sterile buffer solution for 2 times to obtain Pseudomonas aeruginosa suspension, inoculating Pseudomonas aeruginosa into nutrient agar plate, culturing for 6 hr, and placing in refrigerator at 4deg.C. Before use, a loop of strain is taken from the agar plane and inoculated into 200mL of sterile culture medium again for culturing for 12h to ensure that the concentration of the strain reaches 10 multiplied by 10 9 CFU/mL, diluted with sterile physiological saline to obtain 8×10 9 The CFU/mL bacterial suspension is ready for use.
S2, preparing a working electrode, namely sequentially carrying out ultrasonic treatment on a gold plate electrode with the diameter of 3mm in ethanol and deionized water for 5min respectively, drying by nitrogen, then putting the gold plate electrode into a 0.1mol/L dilute sulfuric acid solution, and scanning for a plurality of times by adopting an electrochemical cyclic voltammetry within the range of-0.2-1.2V until the superposition of scanning curves is stable, and taking the gold plate electrode as the working electrode for standby.
S3, preparing a microbial membrane electrode; the methylene blue is doped on the surface of a working electrode by an electrochemical cyclic voltammetry to obtain a new methylene blue film NMB/Au; then preparing a conductive material containing 0.01mol/L and 8×10 9 The mixed solution of the pseudomonas aeruginosa suspension with the concentration of CFU/mL; and (3) immersing NMB/Au into the mixed solution, and obtaining the microbial membrane electrode with a double-layer structure by adopting an electrochemical cyclic voltammetry.
The conductive substances are polyacetylene and polythiophene, polyacetylene: the volume ratio of polythiophene was 1:1.
Comparative example 1
Comparative example 1, doped with the absence of methylene blue, comprises the following specific steps:
s1, preparing strain suspension; the strain is selected from bacillus licheniformis separated from seawater sludge, and the selected bacillus licheniformis is domesticated and subjected to plate separation for 2 times to obtain a pure strain. Shaking culture of pure strain in culture medium solution, PBS solution and NaCl with volume fraction of 0.8% at 30deg.C and 140rpm for 24 hr, centrifuging at 25deg.C and 7000r/min for 20min, washing with equal volume of sterile buffer solution for 2 times to obtain Bacillus licheniformis suspension, inoculating Bacillus licheniformis to nutrient agar plate, culturing for 6 hr, and placing in refrigerator at 4deg.C. Before use, a loop of strain is taken from the agar plane and inoculated into 200mL of sterile culture medium again for culturing for 12h to ensure that the concentration of the strain reaches 10 multiplied by 10 9 CFU/mL, diluted with sterile physiological saline to obtain 8×10 9 The CFU/mL bacterial suspension is ready for use.
S2, preparing a working electrode, namely sequentially carrying out ultrasonic treatment on the gold plate electrode in ethanol and deionized water for 5min respectively, drying by nitrogen, then putting the gold plate electrode in a 0.1mol/L dilute sulfuric acid solution, and scanning for a plurality of times by adopting an electrochemical cyclic voltammetry within the range of-0.2-1.2V until the superposition of scanning curves is stable, and taking the gold plate electrode as the working electrode for standby.
S3, preparing a microbial membrane electrode; then preparing a conductive material containing 0.01mol/L and 8×10 9 CFU/mL concentration bacillus licheniformisThe mixed solution of the bacterial suspension; and immersing the working electrode into the mixed solution, and obtaining the microbial membrane electrode covered with microorganisms by adopting an electrochemical cyclic voltammetry. The conductive substances are polyacetylene, polythiophene and polystyrene, and polyacetylene: polythiophene: the mass ratio of the polyphenylene ethylene is 1:1:1.
Comparative example 2
The existing calcium alginate gel embedding method is taken as a comparative example 2, and the specific steps are as follows:
and (3) dripping sodium alginate solution and bacterial liquid on the surface of a working electrode, immersing in calcium chloride solution for solidification, and covering a polyamide membrane on the surface of cell gel to prevent the embedded cells from falling off. The prepared sensor is used for BOD rapid detection. The electrode and the bacterial liquid are the same as in the first embodiment.
Determination of BOD in seawater Using microbial films prepared in examples and comparative examples as sensors 5 The content is as follows. Drawing standard curves of the biomembrane modified electrodes prepared in the above examples and comparative examples, and determining BOD in seawater according to the standard curves 5 Concentration.
FIG. 1 is a standard curve of example 1, and FIG. 1 shows that in this example 1, BOD is used 5 The concentration of (2) is the abscissa, the current delta mA is the ordinate, and a linear relation can be obtained by drawing a standard curve of the microbial membrane modified electrode: y= 0.0521x-0.027, r 2 =0.9983。
The bacterial suspensions of examples 1 to 7 and comparative examples 1 to 2 were diluted to 10X 10 in order 8 CFU/mL、10×10 7 CFU/mL、10×10 6 CFU/mL、10×10 5 CFU/mL、10×10 4 CFU/mL; preparing microbial films on the surfaces of the electrodes by using the obtained bacterial suspensions with different concentrations, measuring the cyclic voltammetry curves of the microbial films in sulfuric acid solution with the concentration of 0.1mol/L after rinsing by distilled water, wherein the scanning voltage is-0.2-1.2V, and the scanning rate is 50mV/s; and drawing a standard curve of the microbial membrane modified electrode by taking the logarithm of the bacterial concentration as an abscissa and the peak current of the obtained cyclic voltammogram at 0.2V as an ordinate. The microbial films of the present invention were verified for sensitivity and accuracy by taking samples of the same concentration and measuring the colony count using the microbial films prepared in example 1 and comparative examples 1-2, and each example or comparative example was repeatedly examined 5 times to obtain a flatAnd (5) an average value.
TABLE 1 colony count measurement results
Table 1 shows that the concentration of the bacterial suspension measured in example 1 of the present invention was substantially the same as that measured by the plate count method, and the relative standard deviation was lower than that of the plate colony count method. The invention has better accuracy, and the counting result is superior to the traditional plate colony technology method and calcium alginate gel embedding method. The accuracy and the service life of the microbial film without being doped with methylene blue are lower than those of the microbial film with being doped with methylene blue, which shows that the methylene blue and the microorganisms have synergistic effect, and the detection accuracy and the service life can be increased. The accuracy of example 2 is the lowest and the relative standard deviation is the greatest. The service life of the microbial films in examples 1-7 reaches 98-110d and is far longer than that of the microbial films in comparative examples 1-2, which shows that the biological film has the advantages of high accuracy and long service life. The microorganisms of examples 1 to 7 have strong adsorptivity on the electrode surface and do not fall off; the gel of example 2 swells and the microorganisms are easily dispersed and have weak adsorptivity, thereby affecting the detection result and the service life.
To verify the stability of the immobilization of the biofilm of the invention, the biological membranes were immobilized with a specific BOD 5 Seawater with concentration as detection object, BOD of 0-105d is tested 5 The concentration and the result are shown in FIG. 2. FIG. 2 shows the BOD of seawater detected in 105d using the biofilms of example 1 and comparative example 2 5 The current at the initial detection of seawater (at 0 d) was 10mA, and the concentration was consistent with that of example and comparative example 2. The current of example 1 was always between 9.5-10.4mA at 0-105d, the maximum and minimum values differing by 0.9mA; the current of comparative example 2 was between 8.0 and 11.8mA at 0-35d, the maximum and minimum values were 3.8mA, and there was a continuous decrease after 35d, with no sign of recovery, and the current was 4.9mA at 105 d. The microbial film has the advantages of high measurement accuracy and good stability, can reduce the error from +/-20% to within +/-5%, and also shows that the comparative example 2 detects after the service life is longerThe result is in a continuous decline trend and cannot be recovered, and needs to be replaced in time to detect.
The microbial film of the invention is used for measuring the BOD of seawater, and has the advantages of short measurement time and real-time monitoring on site; the classical 5-day method takes 5 days in the whole measurement process and cannot be detected in real time on site, and the sensor method of the microbiological method can finish measurement within 10 minutes.
The foregoing description of the preferred embodiments of the invention is not intended to be limiting, but rather is intended to cover all modifications, equivalents, alternatives, and improvements that fall within the spirit and scope of the invention.
Claims (6)
1. The preparation method of the immobilized microbial membrane for measuring seawater BOD is characterized by comprising the following steps of: the method comprises the following steps:
s1, domesticating microorganisms suitable for high-salinity seawater, wherein the operation steps are as follows:
s11, taking microorganisms separated from seawater sludge, and carrying out domestication and 2-3 times of plate separation to obtain pure bacterial strains;
s12, the pure strain in the step S11 is subjected to shaking culture, is washed by a sterile buffer solution and is inoculated in a culture medium, and when the concentration of thalli reaches 10 multiplied by 10 9 CFU/mL was prepared into 8X 10 9 The CFU/mL bacterial suspension is reserved;
s2, generating a layer of electron mediator for promoting microorganism metabolism on the surface of the electrode, wherein the operation steps are as follows:
s21, sequentially carrying out ultrasonic treatment on the electrode in an organic solvent and deionized water for 5min, drying by nitrogen, immersing in 0.1-0.3mol/L sulfuric acid, and scanning by adopting an electrochemical cyclic voltammetry to obtain a working electrode;
s22, doping methylene blue on the surface of the working electrode in the step S21 by electrochemical cyclic voltammetry to obtain an electrode covered with a methylene blue film;
s3, polymerizing a conductive solution doped with domesticated microorganisms on the surface of an electronic medium layer of the electrode by a cyclic voltammetry to form a microbial film with a double-layer structure on the surface of the electrode, wherein the operation steps are as follows:
s31, dissolving a conductive substance in the bacterial suspension in the step S12 to prepare a mixed solution;
s32, immersing the electrode covered with the methylene blue film in the step S22 into the mixed solution in the step S31, and scanning by adopting electrochemical cyclic voltammetry to obtain the microbial film electrode covered with the double-layer structure.
2. The method for preparing the immobilized microbial membrane for measuring seawater BOD as claimed in claim 1, wherein the method comprises the following steps: the organic solvent is one or more of ethanol, acetone and cyclohexane.
3. The method for preparing the immobilized microbial membrane for measuring seawater BOD as claimed in claim 2, wherein: the conductive substance is one or more of polyacetylene, polythiophene, polyaniline, polyphenylene ethylene and polydiacetylene, and the concentration is 0.01-0.03mol/L.
4. The method for preparing the immobilized microbial membrane for measuring seawater BOD as claimed in claim 1, wherein the method comprises the following steps: the microorganism in the step S1 is one of bacillus licheniformis, candida, hansenula, klebsiella, escherichia coli, clostridium butyricum and pseudomonas aeruginosa.
5. The method for preparing the immobilized microbial membrane for measuring seawater BOD as claimed in claim 1, wherein the method comprises the following steps: the electrode in the step S2 is a disc electrode, and the diameter of the disc electrode is 1-5mm.
6. The method for preparing the immobilized microbial membrane for measuring BOD of seawater according to claim 5, wherein the method comprises the following steps: the electrode is one of a gold plate electrode, a silver plate electrode and a titanium plate electrode.
Priority Applications (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN202111131062.2A CN113960141B (en) | 2021-09-26 | 2021-09-26 | Preparation method of immobilized microbial membrane for measuring BOD of seawater |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN202111131062.2A CN113960141B (en) | 2021-09-26 | 2021-09-26 | Preparation method of immobilized microbial membrane for measuring BOD of seawater |
Publications (2)
Publication Number | Publication Date |
---|---|
CN113960141A CN113960141A (en) | 2022-01-21 |
CN113960141B true CN113960141B (en) | 2023-10-20 |
Family
ID=79462611
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN202111131062.2A Active CN113960141B (en) | 2021-09-26 | 2021-09-26 | Preparation method of immobilized microbial membrane for measuring BOD of seawater |
Country Status (1)
Country | Link |
---|---|
CN (1) | CN113960141B (en) |
Citations (7)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN101105470A (en) * | 2006-07-10 | 2008-01-16 | 北京化工大学 | Biosensor enzyme sensitive film and its preparation method |
CN102320687A (en) * | 2011-06-18 | 2012-01-18 | 山东大学 | The preparation method of a kind of polyaniline-mikrobe combined electrode |
CN103512938A (en) * | 2012-06-15 | 2014-01-15 | 中国科学院理化技术研究所 | Electrochemical sensor and device for immediately monitoring and detecting water body biotoxicity |
CN104007154A (en) * | 2013-02-22 | 2014-08-27 | 中国科学院理化技术研究所 | Redox hydrosol, preparation method and application of integrated microbial electrode for rapid detection of water body biotoxicity |
CN106981668A (en) * | 2017-05-04 | 2017-07-25 | 天津天狮学院 | A kind of microorganism electrode and its preparation method and application |
CN108535343A (en) * | 2018-04-01 | 2018-09-14 | 桂林理工大学 | The preparation method and applications of methylene blue-gold composite nano particle modified electrode |
CN111398382A (en) * | 2020-04-21 | 2020-07-10 | 普洱学院 | Polymethylene blue modified glassy carbon electrode and preparation method and application thereof |
-
2021
- 2021-09-26 CN CN202111131062.2A patent/CN113960141B/en active Active
Patent Citations (7)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN101105470A (en) * | 2006-07-10 | 2008-01-16 | 北京化工大学 | Biosensor enzyme sensitive film and its preparation method |
CN102320687A (en) * | 2011-06-18 | 2012-01-18 | 山东大学 | The preparation method of a kind of polyaniline-mikrobe combined electrode |
CN103512938A (en) * | 2012-06-15 | 2014-01-15 | 中国科学院理化技术研究所 | Electrochemical sensor and device for immediately monitoring and detecting water body biotoxicity |
CN104007154A (en) * | 2013-02-22 | 2014-08-27 | 中国科学院理化技术研究所 | Redox hydrosol, preparation method and application of integrated microbial electrode for rapid detection of water body biotoxicity |
CN106981668A (en) * | 2017-05-04 | 2017-07-25 | 天津天狮学院 | A kind of microorganism electrode and its preparation method and application |
CN108535343A (en) * | 2018-04-01 | 2018-09-14 | 桂林理工大学 | The preparation method and applications of methylene blue-gold composite nano particle modified electrode |
CN111398382A (en) * | 2020-04-21 | 2020-07-10 | 普洱学院 | Polymethylene blue modified glassy carbon electrode and preparation method and application thereof |
Also Published As
Publication number | Publication date |
---|---|
CN113960141A (en) | 2022-01-21 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
Niyomdecha et al. | A novel BOD biosensor based on entrapped activated sludge in a porous chitosan-albumin cryogel incorporated with graphene and methylene blue | |
WO2011022921A1 (en) | Reactor-type rapid measuring instrument for biochemical oxygen demand (bod) and measurement method thereof | |
CN102796660B (en) | For proofing unit and the on-line water quality monitoring method of monitoring water quality on line | |
CN113358722B (en) | Method for realizing rapid detection of water toxicity based on suspended electrochemical active microorganisms | |
Liu et al. | Immobilized multi-species based biosensor for rapid biochemical oxygen demand measurement | |
AU2007226332B2 (en) | A bacterium consortium, bio-electrochemical device and a process for quick and rapid estimation of biological oxygen demand | |
Estevez‐Canales et al. | Silica immobilization of Geobacter sulfurreducens for constructing ready‐to‐use artificial bioelectrodes | |
Qian et al. | A disposable biofilm-modified amperometric biosensor for the sensitive determination of pesticide biotoxicity in water | |
Hu et al. | Development of a mediator-type bioelectrochemical sensor based on polypyrrole immobilized ferricyanide and microorganisms for biochemical oxygen demand fast detection | |
CN113960141B (en) | Preparation method of immobilized microbial membrane for measuring BOD of seawater | |
Liu et al. | The fabrication and the use of immobilized cells as test organisms in a ferricyanide‐based toxicity biosensor | |
Zhang et al. | Direct electrochemistry and electrocatalysis of hemoglobin immobilized in a polymeric ionic liquid film | |
CN116818866A (en) | Method and system for detecting water quality biotoxicity in real time | |
CN112114019B (en) | Potentiometric antifouling sensor based on indole active substance release and application thereof | |
CN105842315B (en) | A kind of electrode sensitive layer for biochemical oxygen demand (BOD) detection | |
CN103773753A (en) | Microbial film for biosensor and preparation method thereof | |
Karube | Possible developments in microbial and other sensors for fermentation control | |
Renneberg et al. | A salt-tolerant yeast-based microbial sensor for 24 hour community wastewater monitoring in coastal regions | |
Cheng et al. | Fabrication of Tyrosinase Biosensor Based on Multiwalled Carbon Nanotubes‐Chitosan Composite and Its Application to Rapid Determination of Coliforms | |
RU133126U1 (en) | BIOSENSOR FOR DETERMINATION OF METHYLAMINE, DIMETHYLAMINE AND TRIMETHYLAMINE IN AQUEOUS MEDIA | |
JP4875367B2 (en) | Stock solution of nitrifying bacteria immobilization membrane | |
CN110923130B (en) | Microbial sensor and preparation method and application thereof | |
Ohki et al. | Biological oxygen demand sensor using an arsenic resistant bacterium | |
Dhall et al. | Selection of an apt support for the immobilization of microbes for the development of a BOD biosensor | |
JP3882677B2 (en) | Preservation solution for nitrifying bacteria-immobilized membrane and ammonia-oxidizing bacteria-immobilized membrane |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
PB01 | Publication | ||
PB01 | Publication | ||
SE01 | Entry into force of request for substantive examination | ||
SE01 | Entry into force of request for substantive examination | ||
GR01 | Patent grant | ||
GR01 | Patent grant |