CN102517234A - Klebsiella strain and method for treating chlorpyrifos pesticide residue on surface of soil, fruits and vegetables by using Klebsiella strain - Google Patents
Klebsiella strain and method for treating chlorpyrifos pesticide residue on surface of soil, fruits and vegetables by using Klebsiella strain Download PDFInfo
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Abstract
The invention provides a microbial new strain, namely Klebsiellasp. Dsp-B, capable of degrading chlorpyrifos pesticide residue and a chlorpyrifos pesticide residue degrading inoculum developed by the degrading strain. The Genbank accession number is HQ836365. The degrading inoculum is used for degrading chemical pesticide by using a microbial technology and belongs to the technical field of environmental microbial restoration. In the production process, inclined surface seeds, shake flask seed liquid, a seed tank and products are provided. The preparation process is simple, low in production cost and easy in large-area popularization and application. The degrading inoculum can be directly sprayed and is very convenient during application, can degrade the residual chlorpyrifos on the surfaces of water, soil, fruits and vegetables quickly and efficiently during agricultural production after spraying so as to solve the problem that residue exceeds the standard, can ensure that green non-toxic pollution-free agricultural products are produced, and effectively improves quality and economic value of the agricultural products.
Description
Technical field
The invention belongs to environmental microorganism recovery technique field, be specifically related to the method for Cray Bai Shi dsp-B bacterial strain and processing soil and fruit and vegetable surfaces chlorpyrifos pesticide residue.
Background technology
Fruit, vegetables are the requisite daily necessities of people, and agricultural chemicals is to prevent disease pest and weed in the fruits and vegetables production process, guarantee the requisite selection of its quality.But; In recent years; The poisoning that remains of pesticide in the quality security problem of vegetables, particularly vegetables and fruits causes gets into people's the visual field in succession, like Foochow " malicious plum " incident, Anxi, Fujian " malicious mater convolvulus " incident, Hainan " malicious cowpea " incident, Guangxi " malicious Chinese cabbage " incident.Pesticide residue in the fruits and vegetables are not only endangering people's health, and have seriously restricted the export of farm produce of China.2005, the China's Agricultural outlet reached 271.8 hundred million dollars, increases by 17.7% on a year-on-year basis.But the influence that China has 90% agricultural and food export enterprise to receive external green barrier measure, about 9,000,000,000 dollars of annual loss meanwhile.Therefore, seek efficient, safe, economic agricultural-food pesticide residue processing and become the scientific research proposition that researcher needs to be resolved hurrily.The degraded of pesticide residue and waste water thereof mainly contains modes such as microbiological deterioration, chemical degradation and photodegradation.Mikrobe can be converted into CO with the organic pollutant in the bad border through various biochemical actions
2And H
2The material that O etc. are nontoxic or toxicity is less.Compare simple to operate, advantages such as degraded is thorough, non-secondary pollution that microbiological deterioration has with physics, chemical degradation mode.Therefore utilize microbial technique to handle pesticide residue, and contaminated soil and water body are carried out biological prosthetic is a kind of efficient ways.Chlorpyrifos 94 is that new and effective, the low toxicity of highly effective pesticides such as SRA-5172 and parathion-methyl substitutes kind, as a kind of broad spectrum type organophosphate insecticides, is widely used in the control of agricultural and urban health insect.Because its a large amount of frequent uses and its persistence detect chlopyrifos residue in a large number in environmental samples such as water body, soil and plant.Chlorpyrifos 94 belongs to moderate toxicity sterilant, can suppress the human body cholinesterase activity, accumulate after neural system, cause feeling sick, dizzy even clouded in mind, high density exposes can cause respiratory paralysis and death.Therefore, seek the focus that a kind of method of removing environment death by poisoning tick pesticide residue highly effective and safe becomes concern, and the improvement that utilizes the Chlorpyrifos 94 high efficiency degradation bacterial agent to be applied to its contaminate environment has important practical significance and application prospect.
Summary of the invention
The objective of the invention is practical problems and needs, a kind of chlorpyrifos pesticide degradation bacteria is provided to pesticide residual contamination in the present agriculture prodn.Another object of the present invention has provided uses chlorpyrifos degrading bacteria to prepare the method for water-in-oil microbial inoculum; Thereby slowly keep the supply of bacterial classification nutritive substance through slow releasing function; Prolong the quality guaranteed period of product greatly, use this microbial inoculum product can make the chlopyrifos residue amount reduce about 85%, and low production cost; Easy to use, have vast market prospect.
Technical scheme
The present invention provides a kind of chlorpyrifos degrading bacteria, and this bacterial strain is the Klebsiella (Klebsiella sp.) of Gram-negative, and main biological characteristics is G
-, facultative anaerobe, thalline is the club shape, and single, paired or short chain shape is arranged, long 0.8-1.2 μ m, wide 2.0-2.4 μ m does not move, and pod membrane is arranged, and does not form gemma.Colony diameter at LB culture medium culturing 12h is 0.5mm, is semisphere, and is translucent, moistening, smooth surface, and electromicroscopic photograph is as shown in Figure 1; Can utilize citrate and dextrose as sole carbon source, ammonia is as nitrogenous source; Glucose fermentation produces sour aerogenesis; The methyl red experiment is negative; The VP reaction shows positive; Three pool iron agar tests do not produce hydrogen sulfide.The Genbank accession number of this bacterial strain 16SrDNA is HQ836365.Can utilize Chlorpyrifos 94 to grow for sole carbon source and the energy, be CO2 and water and simple mineral compound with its thorough mineralising.In the laboratory condition shake flask test, the chlorpyrifos degrading rate reaches more than 85%.The righttest growth pH of this bacterium is 7.0-7.2, and optimum growth temperature is 35 ℃.This degradation bacteria can be produced through the general fermentation equipment of fermentation industry, and with low cost, microbial inoculum directly sprays easy to use, is easy to promote large-scale application;
The technical process of using above-mentioned chlorpyrifos degrading bacteria to produce the microbial inoculum product is: inclined-plane kind → medicine bottle seed liquor → seeding tank → fermentor tank → microbial inoculum product;
Detailed implementation step of the present invention is:
(1) single colony inoculation of picking chlopyrifos residue degradation bacteria Klebsiella sp .dsp-B is in the test tube slant;
(2) the inclined-plane kind with chlopyrifos residue degradation bacteria dsp-B is inoculated in the LB culture media shaking vase, and shaking culture is to logarithmic phase;
(3) change above-mentioned cultured bacterium liquid over to seeding tank by 15% volume inoculum size, continue to be cultured to logarithmic phase;
The seed liquor that (4) will reach logarithmic phase is produced cultivation in the jar by 15% inoculum size access;
(5) fermentation is processed the water-in-oil formulation with inoculum after accomplishing, and is distributed into the liquid dosage form product with plastic barrel or packing bottle;
The culture medium prescription of LB described in the step 2 is: yeast extract paste 5g/L, peptone 10g/L, NaCl 10g/L, pH:7.0-7.2;
The substratum of seeding tank and fermentor tank is identical in the step 3 and 4, fills a prescription to be: glucose 5g/L, NH
4NO
310g/L, K
2HPO
42 g/L, MgSO
47H
2O 0.5 g/L, Na
2HPO
42 g/L, CaCO
33g/L, NaCl 1.0 g/L, yeast extract paste 0.5 g/L, pH:7.0-7.2;
The air flow of sterile air is 1:0.8-1.5 in the culturing process of seeding tank and fermentor tank, and stirring velocity is 150-220 rev/min, and culture temperature is 28-35 ℃, and omnidistance incubation time is 48-72 hour, finishes 4,000,000,000/ml of back thalline;
The oil phase that adds in the water-in-oil formulation that step 5 is developed is No. 10 white oils, and emulsifying agent is span80, and solubility promoter is a tween 80.Fermented liquid is processed the nutritive substance that can make behind the water-in-oil wherein to be wrapped in around the degradation bacteria suspending stabilizedly; Provide the nutrition substrate to keep the activity of bacterial classification through slowly-releasing; Prevent the deposition of degradation bacteria; And oil phase can provide suitable moist environment to degradation bacteria, thereby prolongs the quality guaranteed period of microbial inoculum product greatly.Directly be sprayed onto fruit and vegetable surfaces during use, thereby, improve the residual degradation rate of fruit and vegetable surfaces farming, also can prevent the generation of phenomenons such as fruits and vegetables dehydration, brown stain simultaneously because water in oil water retention characteristic can make nutraceutical slow release keep the activity of bacterium.
Beneficial effect of the present invention does:
1 the present invention provides the degradation bacteria of a kind of organophosphorus pesticide Chlorpyrifos 94, and laboratory biological degradation experiment and field experiment result show, the degradation rate of Chlorpyrifos 94 is reached about 85%;
2 use these methods to produce the chlorpyrifos degrading microbial inoculum to have a production cost low, easy to use, the advantage of good degrading effect is adapted at grain vegetables fruit production export base and pollution-free food production base and uses on a large scale.This invention is for preserving the ecological environment, and the protection people health increases additional value of farm products and has great significance.
Description of drawings:
Fig. 1:The electromicroscopic photograph of klebsiella dsp-B;
Fig. 2:The chlorpyrifos degrading curve of degradation bacteria strains dsp-B in the soil different treatment.
Embodiment:
Embodiment 1
Gather field, the peach garden topsoil that farming Quan Yi district, Chengdu uses chlorpyrifos pesticide for successive years.The pesticidal contamination soil sample 5g that takes by weighing collection is added to 100ml and contains in the sterilization basis salt culture medium of 100mg/l Chlorpyrifos 94, and the prescription of basic salt culture medium is (being g/L): NH
4NO
31.0, MgSO
47H
2O 0.5, (NH
4)
2SO
40.5, KH
2PO
40.5 NaCl 0.5, K
2HPO
41.5, PH:7.0-7.2, in 37 ℃, 180r/min shaking culture 7d.Insert in the salt culture medium of corresponding 100ml basis by 5% volume ratio with primary enrichment culture liquid, Chlorpyrifos 94 concentration is increased to 200mg/L simultaneously, under same condition, cultivates.Get 5ml bacterium liquid behind the 7d and insert 37 ℃ of cultivations down in the same substratum again, the concentration of the process that goes down to posterity death by poisoning tick is progressively brought up to 1000mg/l, goes down to posterity 4 times again, and approximately 7d goes down to posterity 1 time.The pesticide degradation bacteria bacterium liquid of enrichment is made 10 times of gradients (10 with sterilized water
-4, 10
-5, 10
-6) dilution, get 0.1 ml bacteria suspension respectively and coat on the inorganic salt solid medium that contains 50 mg/L Chlorpyrifos 94 37 ℃ of cultivations; When bacterium colony appears in flat board, with single bacterium colony on inorganic salt agricultural chemicals flat board, rule separation, purifying bacterial strain.Single bacterium colony inclined-plane that growth is fast, bacterium colony is regular, go down to posterity stable is preserved.With the 5 strain bacterium activation that primary dcreening operation obtains, insert in the salt culture medium of 100mg/l Chlorpyrifos 94 basis, 37 ℃, 180r/min cultivates 10d, surveys its degradation rate.Through multiple sieve, obtain the highest bacterial strain B of degradation rate.Through the 16SrDNA sequential analysis, find that the sequence similarity of bacterial strain B and Klebsiella 2009I6 is 99%, with the sequence similarity of Klebsiella sp. MS6 also be 99%, GenBank accession number HQ836365.Therefore B should belong to Klebsiella, and main biological characteristics is G
-, facultative anaerobe, thalline is the club shape, and single, paired or short chain shape is arranged, long 0.8-1.2 μ m, wide 2.0-2.4 μ m does not move, and pod membrane is arranged, and does not form gemma.Colony diameter at LB culture medium culturing 12h is 0.5mm, is semisphere, and is translucent, moistening, smooth surface; Can utilize citrate and dextrose as sole carbon source, ammonia is as nitrogenous source; Glucose fermentation produces sour aerogenesis; The methyl red experiment is negative; The VP reaction shows positive; Three pool iron agar tests do not produce hydrogen sulfide;
Embodiment 2
The inclined-plane of getting the klebsiella of Gram-negative among the embodiment 1 is inoculated in the 100mlLB substratum, and LB substratum (g/l) is filled a prescription as follows: peptone 10, yeast extract 5; NaCl 10; PH 7.0-7.2, constant-temperature shaking culture obtains bacterium liquid to logarithmic phase.Above-mentioned cultured bacterium liquid is changed over to 121 ℃ of high pressure moist heat sterilizations and is cooled to the seeding tank about 35 ℃ by 15% volume inoculum size, and the seed tank culture based formulas is: glucose 5g/L, NH
4NO
310g/L, K
2HPO
42 g/L, MgSO
47H
2O 0.5 g/L, Na
2HPO
42 g/L, CaCO
33g/L, NaCl1.0 g/L, yeast extract paste 0.5 g/L, pH:7.0-7.2; Continue to be cultured to logarithmic phase, the air flow of sterile air is 1:1.5 in the culturing process, and stirring velocity is 200 rev/mins, and culture temperature is 35 ℃.The seed liquor that will reach logarithmic phase is produced a jar cultivation by 15% inoculum size access; The used substratum of fermentor tank is identical with seeding tank; Postvaccinal leavening temperature is controlled at 28-35 ℃, and the air flow of sterile air is 1:0.8-1.5, and stirring velocity is 150-220 rev/min; Omnidistance fermented incubation time is 48-72 hour, and thalline quantity reaches 4,000,000,000/ml after the fermentation ends.Fermentation is processed water-in-oil formulation with inoculum according to certain oil-water ratio after accomplishing under the effect of emulsifying agent, be distributed into liquid dosage form with plastic barrel or packing bottle then;
Application examples 1
Shake under bottle condition in the laboratory, the liquid degradation microbial inoculum in the application implementation example 2, it is in the minimal medium of 100mg/l that the inoculum size by 5% is inoculated in 50ml Chlorpyrifos 94 concentration, at 35 ℃; PH=7.2,180r/min cultivate down, after cultivating the 3rd; 6,9, its chlopyrifos residue concentration is surveyed in the 12d sampling.Contrast is not for adding degradation bacteria.The degradation rate of Chlorpyrifos 94 is 85.5% behind the 12d, and the degradation rate of contrast has only 5.2% (seeing table 1);
Table 1. klebsiella shakes under bottle condition degraded to Chlorpyrifos 94 in the laboratory
| Processing mode | Contrast | Handle |
| Pesticide residue concentration (mg/L) | 95.7 | 15.6 |
| Degradation rate | 11.1% | 85.5% |
Application examples 2
4 processing are established in soil degrading potted plant experiment in laboratory altogether, sterilization soil+Chlorpyrifos 94, non-sterilization soil+Chlorpyrifos 94, sterilization soil+Chlorpyrifos 94+dsp-B, non-sterilization soil+Chlorpyrifos 94+dsp-B, and Chlorpyrifos 94 concentration is 100mg/kg, the inoculum size of degradation bacteria dsp-B is 10
8Cfu/Kg, each handles repetition 2 times.In repairing back the 3rd, 6,9,12, its chlopyrifos residue concentration is surveyed in the 15d sampling, and the result sees Fig. 2;
Can know by Fig. 2; After cultivating 15d; Chlorpyrifos degrading microbial inoculum of the present invention meets first _ order kinetics equation four kinds of different treatment degradation in soil situation; Corresponding degradation half life is respectively 198.04,42.52,8.76 and 6.99h, and the degradation rate of soil Chlorpyrifos 94 has evident difference in different treatment.In sterile soil, the degradation rate of Chlorpyrifos 94 has only 5.6%, and in non-sterile soil, the degradation rate of Chlorpyrifos 94 is 22.1%, and obviously faster than sterile soil, this phenomenon explanation mikrobe plays an important role in the degradation process of soil Chlorpyrifos 94.The degradation rate that adds degradation bacteria dsp-B in the sterile soil is 67.1%, is higher than no sterilizing soil, explains that the ability of bacterial strain dsp-B chlorpyrifos degradation is higher than indigenous mikrobe in the soil.What degradation rate was the highest appears in the non-sterile soil that has added bacterial strain dsp-B, is 76.2%, and this phenomenon explanation bacterial strain dsp-B has good degradation capability to the Chlorpyrifos 94 in the soil, and and soil in indigenous microorganism between stable group's relation is arranged;
Application examples 3
2 mu of fields choosing dragon's fountain plantation peach are experimental subjects; Wherein one mu of conduct contrast; Another mu is the degradation bacterial agent treatment group, at the week back collection soil sample sample of spraying insecticide, adopts the gas chromatography combined with mass spectrometry method to measure chlorpyrifos pesticide residue content (being measured by Sichuan University's life science institute Biological resources and test center of key lab of the ecotope Board of Education); Treatment group is sprayed the water-in-oil degradation bacterial agent by development among the embodiment 2 then, and concentration is 3 * 10
9Cfu/ml, totally 20 L are sprayed in the peach soil equably, collection soil is returned the content that Sichuan University surveys its Chlorpyrifos 94 behind the degraded 10d, control group with clear water as contrast.Through measuring; The content of the dead tick of soil poisoning is 8.7mg/kg before the degraded, and the content of degradation bacterial agent processed group degraded back chlopyrifos is 1.6mg/kg, and its degradation rate is 81.6%; A little more than the potted plant experiment degradation rate of soil; Analysis maybe be relevant with factor of natural environment such as light degradation, and the degradation rate of control group chlopyrifos is 27.6% behind the 10d, can find out that degradation bacterial agent is remarkable to the residual degraded of the dead tick of soil poisoning;
Table 2. klebsiella in field experiment to the residual degraded of the dead tick of soil poisoning
| Processing mode | Contrast | Handle |
| Pesticide residue concentration (mg/L) | 6.3 | 1.6 |
| Degradation rate | 27.6% | 81.6% |
Application examples 4:
Liquid degradation microbial inoculum among the investigation embodiment 2 is to the residual removal effect of fruit and vegetable surfaces Chlorpyrifos 94 farming.Choosing the Chinese cabbage of dragon's fountain in ploughing is experimental subjects, adopts the Chlorpyrifos 94 of 100mg/L concentration that the Chinese cabbage blade face is sprayed, treat that the agricultural chemicals drop disappears on the blade after, its Chlorpyrifos 94 content is surveyed in sampling.Behind the spray medicine, used OD at a distance from one day
600nm=0 (clear water contrast), spray in the sub-district that the water-in-oil degradation bacteria liquid that fermented liquid concentration OD=1.0 processes is crossed the spray medicine, and 3 replicated plots, 1m are established in each processing
2/ sub-district, the sub-district random alignment.Spray medicine and spray bacterium amount all are 20mL/m
2, the time of spraying is selected in carries out during 4-5 afternoon, sprays back 24 hours no rain.Through 7 days degraded, its Chlorpyrifos 94 content was surveyed by the serpentine method Chinese cabbage 5g that gathers in every sub-district.The content of Chlorpyrifos 94 is 6.20mg/kg on the degraded frontvane, and the content that sprays the experimental group degraded back Chlorpyrifos 94 of water-in-oil degradation bacteria liquid is 1.21mg/kg, and degradation rate is 80.5%, and sprays OD
600nmThe control group chlopyrifos residue amount of=0 bacterium liquid is 5.02mg/kg, degradation rate only 19.3%;
Utilizing microbiological deterioration and handle agricultural-food, is frontier, new way that pesticide residue control is handled, and many developed countries and enterprise of major company drop into huge fund to be engaged in the research and development of pesticide residue efficient degrading bacteria.Laboratory and real application research all show and utilize the mikrobe pesticide residue in the fruits and vegetables of degrading, and can effectively improve the quality and the economic worth of agricultural-food.
Claims (4)
1. a degrading chlorpyrifos pesticide residue bacterium is characterized in that this mikrobe is the dsp-B bacterial strain of the Klebsiella (Klebsiella sp.) of Gram-negative, and main biological characteristics is G
-, facultative anaerobe, thalline is the club shape, and single, paired or short chain shape is arranged, long 0.8-1.2 μ m, wide 2.0-2.4 μ m does not move, and pod membrane is arranged, and does not form gemma; Colony diameter at LB culture medium culturing 12h is 0.5mm, is semisphere, and is translucent, moistening, smooth surface; Can utilize citrate and dextrose as sole carbon source, ammonia is as nitrogenous source; Glucose fermentation produces sour aerogenesis; The methyl red experiment is negative; The VP reaction shows positive; Three pool iron agar tests do not produce hydrogen sulfide; The Genbank accession number of this bacterial strain 16SrDNA is HQ836365.
2. utilize the described Cray Bai Shi of claim 1 degradation bacteria strains to be developed into the chlopyrifos residue degradation bacterial agent, it is characterized in that, concrete preparation technology is following:
(1) single colony inoculation of picking chlopyrifos residue degradation bacteria Klebsiella sp .dsp-B is in the test tube slant;
(2) the inclined-plane kind with chlopyrifos residue degradation bacteria dsp-B is inoculated in the LB culture media shaking vase, and shaking culture is to logarithmic phase;
(3) change above-mentioned cultured bacterium liquid over to seeding tank by 10-15% volume inoculum size, continue to be cultured to logarithmic phase; The used culture medium prescription of seeding tank is: glucose 5g/L, NH
4NO
310g/L, K
2HPO
42 g/L, MgSO
47H
2O 0.5 g/L, Na
2HPO
42 g/L, CaCO
33g/L, NaCl 1.0 g/L, yeast extract paste 0.5 g/L, pH:7.0-7.2;
The seed liquor that (4) will reach logarithmic phase is inoculated in the production jar by the inoculum size of 10-15% and is cultivated; It is the same with seeding tank to produce jar used culture medium prescription;
(5) air flow of sterile air is 1:0.8-1.5 in the culturing process of seeding tank and production jar, and stirring velocity is 150-220 rev/min, and culture temperature is 28-35 ℃, and omnidistance incubation time is 48-72 hour, finishes 4,000,000,000/ml of back thalline;
(6) fermentation is processed the water-in-oil formulation with inoculum after accomplishing; Oil phase preparation: get 6 parts of span-80 and 94 parts of No. 10 white oil mixing autoclavings are subsequent use; Water preparation: the sterilization tween 80 of adding 4% and fully dissolving in the good ferment product of activation; Emulsification preparation: earlier the oil phase impouring is put in the vertical colloid mill; Under stirring at low speed, slowly add water (oil phase and water are in the ratio thorough mixing of 1:3); High-speed stirring 3min-5min then, quick emulsification becomes the water-in-oil shape, is distributed into the water-in-oil liquid dosage form with plastic barrel or packing bottle.
3. the preparation method of chlopyrifos residue high efficiency degradation bacterial agent according to claim 2; It is characterized in that: when the said fermented liquid of step (6) is processed the water-in-oil product; Need to add a certain proportion of span80 as emulsifying agent; Its HLB value is 4.3, can make immiscible profit two miscible each other, and forms homodisperse invert emulsion system; Microbiobacterial agent receives such environmental effects and loss of activity after being administered in the environment easily, the most important thing is the supply of nutritive element and moisture in these factors; Fermented liquid is processed the nutritive substance that can make behind the water-in-oil wherein to be wrapped in around the degradation bacteria suspending stabilizedly; Provide the nutrition substrate to keep the activity of bacterial classification through slowly-releasing; Prevent the deposition of degradation bacteria; And oil phase can provide suitable moist environment to degradation bacteria, thereby prolongs the quality guaranteed period of microbial inoculum product greatly; Directly be sprayed onto fruit and vegetable surfaces during use, thereby, improve the residual degradation rate of fruit and vegetable surfaces farming, also can prevent the generation of phenomenons such as fruits and vegetables dehydration, brown stain simultaneously because water in oil water retention characteristic can make nutraceutical slow release keep the activity of bacterium.
4. klebsiella dsp-B as claimed in claim 1 is in the application of handling on water body, soil and the fruit and vegetable surfaces chlorpyrifos pesticide residue.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103484393A (en) * | 2013-06-04 | 2014-01-01 | 四川大学 | Klebsiella inoculum capable of biodegrading residual chlorpyrifos pesticide on surfaces of garden stuffs and soil |
| CN106399101A (en) * | 2016-06-14 | 2017-02-15 | 西南交通大学 | Anaerobic bacterium fermentation cultivation method based on magnetic-response Pickering emulsion |
-
2011
- 2011-12-27 CN CN2011104433107A patent/CN102517234A/en active Pending
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103484393A (en) * | 2013-06-04 | 2014-01-01 | 四川大学 | Klebsiella inoculum capable of biodegrading residual chlorpyrifos pesticide on surfaces of garden stuffs and soil |
| CN106399101A (en) * | 2016-06-14 | 2017-02-15 | 西南交通大学 | Anaerobic bacterium fermentation cultivation method based on magnetic-response Pickering emulsion |
| CN106399101B (en) * | 2016-06-14 | 2019-12-20 | 西南交通大学 | Anaerobic bacteria fermentation culture method based on magnetic response Pickering emulsion |
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Application publication date: 20120627 |