CN107601681B - Composite microbial inoculum for repairing metamitron polluted water body - Google Patents
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- 239000002131 composite material Substances 0.000 title claims abstract description 20
- 239000002068 microbial inoculum Substances 0.000 title claims abstract description 20
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- SXRSQZLOMIGNAQ-UHFFFAOYSA-N Glutaraldehyde Chemical compound O=CCCCC=O SXRSQZLOMIGNAQ-UHFFFAOYSA-N 0.000 claims description 12
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- 238000004132 cross linking Methods 0.000 claims description 4
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- 238000010438 heat treatment Methods 0.000 claims description 4
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- 239000004721 Polyphenylene oxide Substances 0.000 description 1
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- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention belongs to the technical field of microorganisms, and discloses a composite microbial inoculum for repairing metamitron polluted water, which is prepared by the following steps: 1) uniformly inoculating the acid bacterium delphite seed solution, the pseudomonas aeruginosa seed solution, the bacillus atrophaeus seed solution and the pseudomonas niger seed solution into an inorganic salt domestication culture medium, and culturing to obtain a composite bacterial solution; 2) and putting the compound bacterial liquid into a carrier with twice mass to obtain the compound bacterial agent. The composite microbial inoculum can effectively restore the metamitron polluted water body, and has high removal rate and strong environmental friendliness.
Description
Technical Field
The invention belongs to the technical field of microorganisms, and particularly relates to a composite microbial inoculum for repairing metamitron polluted water.
Background
The chemical name of metamitron (metamitron) is 4-amino-3-methyl-6-phenyl-1, 2, 4-triazine-5 (4H) -ketone, which belongs to triazinone selective preemergence herbicide, the application range is wide, the herbicide is mainly absorbed by the roots of plants and then conveyed into leaves, and the herbicide plays a role in killing weeds by inhibiting the Hill reaction of photosynthesis. Can prevent and kill monocotyledonous and dicotyledonous weeds, is a herbicide applied to prevent and kill gramineous and broadleaf weeds in beet fields, is mainly used for field weeding of dry crops such as corn, beet and other crops, and can also prevent and kill various weeds such as datura, chickweed, wild sesame, bluegrass and the like.
The loss of metamitron pesticide into the environment will cause serious environmental pollution, sometimes even extremely dangerous consequences. The pesticide lost into the environment floats into the atmosphere through evaporation and transpiration, and the floating pesticide is adsorbed by dust in the air. And spread with the wind. Causing pollution to the atmospheric environment. The pesticides in the atmosphere flow into water through rainfall, so that the water environment is polluted, and harm is caused to people, livestock, particularly aquatic organisms (such as fish and shrimps). Meanwhile, the pesticide lost in the soil can cause soil hardening. The water body treatment of pesticide pollution is difficult, so far, most of the water body is treated by adopting physical and chemical methods, but the methods have high treatment cost and strict technical requirements, and are difficult to popularize in practical application.
The biological treatment method is to decompose aniline compounds by using the action of microorganisms. The biological treatment method has much lower cost than physical and chemical methods, no secondary pollution and stronger variability and adaptability of microorganisms, so the method becomes an ideal method for treating the polluted water body. At present, the research on the microbial inoculum for treating the water body polluted by the metamitron pesticide is less, and the degradation microbial inoculum is rare.
Disclosure of Invention
In order to overcome the defects of the prior art, the invention aims to provide the composite microbial inoculum for repairing the metamitron polluted water body.
The invention is realized by adopting the following technical scheme:
a composite microbial inoculum for repairing metamitron polluted water is prepared according to the following steps:
1) uniformly inoculating the acid bacterium delphite seed solution, the pseudomonas aeruginosa seed solution, the bacillus atrophaeus seed solution and the pseudomonas niger seed solution into an inorganic salt domestication culture medium, and culturing to obtain a composite bacterial solution;
2) and putting the compound bacterial liquid into a carrier with twice mass to obtain the compound bacterial agent.
In particular, the amount of the solvent to be used,
the microbial inoculum is prepared according to the following steps:
1) uniformly mixing the acid-eating bacterium delphite seed solution, the pseudomonas aeruginosa seed solution, the atrophic bacillus seed solution and the pseudomonas niger seed solution according to the volume ratio of 1-2:2-3:2-3:3-5, then inoculating the mixture into an inorganic salt domestication culture medium according to the inoculation amount of 10%, and performing shaking culture for 12 hours at the temperature of 30 ℃ and the speed of 150rpm to obtain a composite bacterial solution;
2) and (3) putting the compound bacterial liquid into a carrier with twice mass, stirring for 30min at 20 ℃, standing for 24 h at 4 ℃, gradually forming a flora biofilm on the biological carrier, and filtering to dry to obtain the microbial carrier.
Further, the air conditioner is provided with a fan,
the preparation method of the carrier comprises the following steps: soaking polyphenyl ether in saturated salt solution for 2 hours, then discharging liquid, soaking in 3wt% sodium hydroxide solution for 6 hours, discharging liquid, then washing with 75% ethanol to be neutral, then adding chitosan, glycerol and water, fully mixing uniformly, heating in a 60 ℃ water bath kettle for 1 hour, after vacuum filtration, adding 2% glutaraldehyde solution, crosslinking for 1 hour, washing with water to remove redundant glutaraldehyde, and then adding 0.5M CaCl2The aqueous solution was allowed to stand at room temperature for 12 hours, and vacuum-dried to obtain a carrier.
Preferably, the polyphenyl ether, the chitosan, the glycerol, the water, the glutaraldehyde solution and the CaCl2The proportion of the aqueous solution is as follows: 50-80g of 1-2kg, 5-7 g of 2-3kg, 5-7L of 3-5L.
Preferably, the preparation method of the inorganic salt acclimatization culture medium comprises the following steps:
to obtain (NH)4)SO42g，KH2PO41g，Na2HPO40.5g，MgSO40.2g, 1mL of trace element solution and 200mg of metamitron, adding water to a constant volume of 1000mL, adjusting the pH value to 7.0, and sterilizing at 121 ℃ for 20min for later use;
wherein, the preparation of the microelement solution comprises the following steps: FeCl30.5g，FeSO40.5g，MnSO40.2g，ZnSO40.1g，CoCl20.1g, adding water to a constant volume of 1000mL, and filtering and sterilizing to obtain the product.
Preferably, the acidovorax delbrueckii is ATCC 15668, the pseudomonas aeruginosa is ATCC 9027, the bacillus atrophaeus is ATCC9372, and the pseudomonas melanogaster is ATCC 19375.
Preferably, the concentrations of the acidovorax defueli seed solution, the pseudomonas aeruginosa seed solution, the bacillus atrophaeus seed solution and the pseudomonas nigricans seed solution are (1-5) multiplied by 107cfu/ml。
The beneficial effects of the invention mainly comprise the following aspects:
the composite microbial inoculum is put into a water body, and a bacterial strain forms a biological film on the surface of a biological carrier, so that metabolism is carried out, and metamitron in sewage is adsorbed and decomposed;
the invention adopts four strains, has reasonable compatibility and good synergistic performance, can mutually promote the proliferation and the enzyme activity, and has good repairing effect;
after the treatment of the composite microbial inoculum, the content of the metamitron is greatly reduced, the removal rate of a low-concentration group can reach 99.8 percent, the removal rate of a high-concentration group is 99.4 percent, the effect is greatly superior to that of a control group, and the comparative experiment shows that the strains can be symbiotically proliferated and have good synergistic performance.
According to the invention, through modification treatment of polyphenyl ether, the porosity and the specific surface area are improved, the adsorption effect is good, the strain adhesion is high, the removal rate of metamitron can reach 99.7%, the removal rate of COD can reach 94.8%, and compared with a conventional carrier, the removal rate of metamitron and COD can be obviously improved.
Detailed Description
In order to make those skilled in the art better understand the technical solutions in the present application, the technical solutions in the present application will be clearly and completely described below with reference to specific embodiments of the present application, and it is obvious that the described embodiments are only a part of the embodiments of the present application, and not all embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
The experimental procedures in the following examples are conventional unless otherwise specified. The test materials used in the following examples were purchased from conventional biochemical sources, unless otherwise specified. The quantitative tests in the following examples, all set up three replicates and the results averaged.
Example 1
A composite microbial inoculum for repairing metamitron polluted water is prepared according to the following steps:
soaking polyphenyl ether in saturated salt solution for 2 hr, draining, soaking in 3wt% sodium hydroxide solution for 6 hr, draining, washing with 75% ethanol to neutrality, adding chitosan,Mixing glycerol and water, heating in 60 deg.C water bath for 1 hr, vacuum filtering, adding 2% glutaraldehyde solution, crosslinking for 1 hr, washing with water to remove excessive glutaraldehyde, and adding 0.5M CaCl2Standing the aqueous solution at room temperature for 12 hours, and drying in vacuum to obtain a carrier; the polyphenyl ether, the chitosan, the glycerol, the water, the glutaraldehyde solution and the CaCl2The proportion of the aqueous solution is as follows: 1kg of 50g of 5kg of 2kg of 5L of 3L; the pore diameter of the polyphenyl ether is 100-300 mu m, and the specific surface area is 40-90m2/g；
Respectively performing slant culture and seed culture on acidovorax delbrueckii ATCC 15668, pseudomonas aeruginosa ATCC 9027, bacillus atrophaeus ATCC9372 and pseudomonas niger ATCC 19375 to obtain acidovorax delbrueckii seed solution, pseudomonas aeruginosa seed solution, bacillus atrophaeus seed solution and pseudomonas niger seed solution, and controlling the concentration of the four seed solutions to be 1 × 107cfu/ml。
To obtain (NH)4)SO42g，KH2PO41g，Na2HPO40.5g，MgSO40.2g, 1mL of trace element solution and 200mg of metamitron, adding water to a constant volume of 1000mL, adjusting the pH value to 7.0, and sterilizing at 121 ℃ for 20min to prepare an inorganic salt acclimation culture medium for later use; wherein, the preparation of the microelement solution comprises the following steps: FeCl30.5g，FeSO40.5g，MnSO40.2g，ZnSO40.1g，CoCl20.1g, adding water to a constant volume of 1000mL, and filtering and sterilizing to obtain the product;
uniformly mixing the acid-eating bacterium delphite seed solution, the pseudomonas aeruginosa seed solution, the bacillus atrophaeus seed solution and the pseudomonas nigricans seed solution according to the volume ratio of 1:2:2:3, then inoculating the mixture into an inorganic salt acclimation culture medium according to the inoculation amount of 10%, and carrying out oscillation culture for 12 hours at the temperature of 30 ℃ and the speed of 150rpm to obtain a composite bacterial solution;
and (3) putting the compound bacterial liquid into a carrier with twice mass, stirring for 30min at 20 ℃, standing for 24 h at 4 ℃, gradually forming a flora biofilm on the biological carrier, and filtering to dry for later use.
Example 2
A composite microbial inoculum for repairing metamitron polluted water is prepared according to the following steps:
soaking polyphenyl ether in saturated salt solution for 2 hours, then discharging liquid, soaking in 3wt% sodium hydroxide solution for 6 hours, discharging liquid, then washing with 75% ethanol to be neutral, then adding chitosan, glycerol and water, fully mixing uniformly, heating in a 60 ℃ water bath kettle for 1 hour, after vacuum filtration, adding 2% glutaraldehyde solution, crosslinking for 1 hour, washing with water to remove redundant glutaraldehyde, and then adding 0.5M CaCl2Standing the aqueous solution at room temperature for 12 hours, and drying in vacuum to obtain a carrier; the polyphenyl ether, the chitosan, the glycerol, the water, the glutaraldehyde solution and the CaCl2The proportion of the aqueous solution is as follows: 50-80g of 1-2kg, 5-7 g of 2-3kg, 5-7L of 3-5L; the pore diameter of the polyphenyl ether is 100-300 mu m, and the specific surface area is 40-90m2/g；
Respectively performing slant culture and seed culture on acidovorax delbrueckii ATCC 15668, pseudomonas aeruginosa ATCC 9027, bacillus atrophaeus ATCC9372 and pseudomonas niger ATCC 19375 to obtain acidovorax delbrueckii seed solution, pseudomonas aeruginosa seed solution, bacillus atrophaeus seed solution and pseudomonas niger seed solution, and controlling the concentration of the four seed solutions to be 3 x 107cfu/ml。
To obtain (NH)4)SO42g，KH2PO41g，Na2HPO40.5g，MgSO40.2g, 1mL of trace element solution and 200mg of metamitron, adding water to a constant volume of 1000mL, adjusting the pH value to 7.0, and sterilizing at 121 ℃ for 20min for later use; wherein, the preparation of the microelement solution comprises the following steps: FeCl30.5g，FeSO40.5g，MnSO40.2g，ZnSO40.1g，CoCl20.1g, adding water to a constant volume of 1000mL, and filtering and sterilizing to obtain the product;
uniformly mixing the acid-eating bacterium delphite seed solution, the pseudomonas aeruginosa seed solution, the bacillus atrophaeus seed solution and the pseudomonas nigricans seed solution according to the volume ratio of 2:3:3:5, then inoculating the mixture into an inorganic salt acclimation culture medium according to the inoculation amount of 10%, and carrying out oscillation culture for 12 hours at the temperature of 30 ℃ and the speed of 150rpm to obtain a composite bacterial solution;
and (3) putting the compound bacterial liquid into a carrier with twice mass, stirring for 30min at 20 ℃, standing for 24 h at 4 ℃, gradually forming a flora biofilm on the biological carrier, and filtering to dry for later use.
Example 3
Test one:
selecting a water body sample: the metamitron polluted water body comes from an agricultural planting area in Hangzhou areas, and the concentration of the metamitron in the water body is set to be 100mg/L and 1000 mg/L;
grouping: example 1 the complex microbial inoculum is an experimental group; control group 1: the rest of example 1 was the same as that of example 1 without adding acid bacteria of Delftia sp; control group 2: the rest of the process is the same as the example 1 without adding pseudomonas aeruginosa; control group 3: the procedure of example 1 was repeated except that Bacillus atrophaeus was not added; control group 4: the procedure of example 1 was otherwise the same as that of example 1;
the using method comprises the following steps: intercepting with fence to remove solid impurities, adjusting pH of water body to 6.5-7.2, and adjusting pH to
The compound microbial inoculum is added in an amount of 3g per cubic meter, and the treatment time is 72 hours. The content of metamitron pollutants in the discharged water after the treatment of each group is shown in table 1:
TABLE 1
| Index (I) | 100mg/L | 1000mg/L |
| Experimental group | 0.2 | 3.9 |
| Control group 1 | 7.7 | 43.4 |
| Control group 2 | 5.6 | 31.7 |
| Control group 3 | 6.8 | 27.9 |
| Control group 4 | 4.3 | 31.4 |
And (4) conclusion: after the treatment of the composite microbial inoculum, the content of the metamitron is greatly reduced, the removal rate of a low-concentration group can reach 99.8 percent, the removal rate of a high-concentration group is 99.4 percent, the effect is greatly superior to that of a control group 1-4, and the synergistic effect is good as can be seen from a comparison experiment, and the strains can be symbiotically proliferated.
And (2) test II:
selecting a water body sample: the metamitron polluted water body comes from an agricultural planting area in Hangzhou areas, and the concentration of the metamitron in the water body is set to be 100mg/L, COD and 560 mg/L;
grouping: the complex microbial inoculum of the embodiment 2 is an experimental group; control group 1: the carrier is the conventional diatomite carrier, and the rest is the same as the example 2; control group 2: the carrier adopts conventional turfy soil: the sawdust is a composite carrier of 1:1, and the rest is the same as the embodiment 2;
the using method comprises the following steps: intercepting with fence to remove solid impurities, adjusting pH of water body to 6.5-7.2, and adjusting pH to
The compound microbial inoculum is added in an amount of 3g per cubic meter, and the treatment time is 72 hours. The content of metamitron pollutants in the discharged water after the treatment of each group is shown in table 2:
TABLE 2
| Index (I) | Metamitron mg/L | COD mg/L |
| Experimental group | 0.3 | 29.3 |
| Control group 1 | 7.1 | 74.8 |
| Control group 2 | 6.9 | 83.6 |
As shown in Table 2, the modified polyphenylene oxide improves the porosity and the specific surface area, has good adsorption effect and high strain adhesion, achieves 99.7% of metamitron removal rate and 94.8% of COD removal rate, and can significantly improve the removal rates of metamitron and COD compared with the conventional carrier.
Although embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that changes, modifications, substitutions and alterations can be made in these embodiments without departing from the principles and spirit of the invention, the scope of which is defined in the appended claims and their equivalents.
Claims (1)
1. The composite microbial inoculum for repairing the metamitron polluted water body is characterized by being prepared according to the following steps:
1) uniformly mixing the acid-eating bacterium delphite seed solution, the pseudomonas aeruginosa seed solution, the atrophic bacillus seed solution and the pseudomonas niger seed solution according to the volume ratio of 1-2:2-3:2-3:3-5, then inoculating the mixture into an inorganic salt domestication culture medium according to the inoculation amount of 10%, and performing shaking culture for 12 hours at the temperature of 30 ℃ and the speed of 150rpm to obtain a composite bacterial solution; the acidovorax defueli is ATCC 15668, the pseudomonas aeruginosa is ATCC 9027, the bacillus atrophaeus is ATCC9372, and the pseudomonas nigricans is ATCC 19375;
2) placing the compound bacterial liquid into a carrier with twice mass, stirring for 30min at 20 ℃, standing for 24 h at 4 ℃, gradually forming a flora biofilm on the carrier, and filtering to dry to obtain the microbial carrier liquid;
the preparation method of the carrier comprises the following steps: soaking polyphenyl ether in saturated salt solution for 2 hours, then discharging liquid, soaking in 3wt% sodium hydroxide solution for 6 hours, discharging liquid, then washing with 75% ethanol to be neutral, then adding chitosan, glycerol and water, fully mixing uniformly, heating in a 60 ℃ water bath kettle for 1 hour, after vacuum filtration, adding 2% glutaraldehyde solution, crosslinking for 1 hour, washing with water to remove redundant glutaraldehyde, and then adding 0.5M CaCl2Standing the aqueous solution at room temperature for 12 hours, and drying in vacuum to obtain a carrier; the polyphenyl ether, the chitosan, the glycerol, the water, the glutaraldehyde solution and the CaCl2The proportion of the aqueous solution is as follows: 50-80g of 1-2kg, 5-7 g of 2-3kg, 5-7L of 3-5L;
the preparation method of the inorganic salt acclimatization culture medium comprises the following steps:
to obtain (NH)4)SO42g，KH2PO41g，Na2HPO40.5g，MgSO40.2g, 1mL of trace element solution and 200mg of metamitron, adding water to a constant volume of 1000mL, adjusting the pH value to 7.0, and sterilizing at 121 ℃ for 20min for later use;
the preparation method of the trace element solution comprises the following steps: FeCl30.5g，FeSO40.5g，MnSO40.2g，ZnSO40.1g，CoCl20.1g, adding water to a constant volume of 1000mL, and filtering and sterilizing to obtain the product;
the concentrations of the acid bacterium delphite seed liquid, the pseudomonas aeruginosa seed liquid, the atrophic bacillus seed liquid and the black pseudomonas seed liquid are all (1-5)107cfu/ml。
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| CN115094014B (en) * | 2022-08-25 | 2022-11-25 | 江苏朴厚环境工程有限公司 | Ochrobactrum pallidum, microbial inoculum thereof and application of Ochrobactrum pallidum in pesticide wastewater treatment |
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| "除草剂2，"4-滴微生物降解研究进展";韩丽珍;《农药》;20121031;第51卷(第10期);第710-719页 * |
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