CN115710570B - Trichloroethylene degrading bacterium and application thereof - Google Patents
Trichloroethylene degrading bacterium and application thereof Download PDFInfo
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- XSTXAVWGXDQKEL-UHFFFAOYSA-N Trichloroethylene Chemical group ClC=C(Cl)Cl XSTXAVWGXDQKEL-UHFFFAOYSA-N 0.000 title claims abstract description 121
- UBOXGVDOUJQMTN-UHFFFAOYSA-N trichloroethylene Natural products ClCC(Cl)Cl UBOXGVDOUJQMTN-UHFFFAOYSA-N 0.000 title claims abstract description 120
- 230000000593 degrading effect Effects 0.000 title claims abstract description 38
- 241000894006 Bacteria Species 0.000 title claims abstract description 36
- 244000063299 Bacillus subtilis Species 0.000 claims abstract description 31
- 235000014469 Bacillus subtilis Nutrition 0.000 claims abstract description 29
- 238000006731 degradation reaction Methods 0.000 claims abstract description 26
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- 238000000034 method Methods 0.000 claims abstract description 24
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- 238000009629 microbiological culture Methods 0.000 claims abstract description 6
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- 230000001580 bacterial effect Effects 0.000 claims description 18
- 239000001301 oxygen Substances 0.000 claims description 18
- 229910052760 oxygen Inorganic materials 0.000 claims description 18
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- GWEVSGVZZGPLCZ-UHFFFAOYSA-N Titan oxide Chemical compound O=[Ti]=O GWEVSGVZZGPLCZ-UHFFFAOYSA-N 0.000 description 2
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Abstract
The invention belongs to the field of bioengineering, and provides a trichloroethylene degrading bacterium and application thereof. The trichloroethylene degrading bacteria belong to gram positive bacteria, are identified as bacillus subtilis by molecular level, are preserved in China general microbiological culture Collection center (CGMCC) 24319, can grow by using trichloroethylene as a unique carbon source, do not depend on a co-metabolism matrix for degradation, can be directly degraded by being put into a sewage treatment system containing trichloroethylene, further can prevent the strain from entering soil or groundwater environment to cause long-term pollution, can avoid the defects of high treatment cost, secondary pollution and the like by adopting a physical and chemical method, has no side effect on operators, and provides a new germplasm resource for the research of directly treating the trichloroethylene under the biological aerobic condition, and has good application value in actual production.
Description
Technical Field
The invention belongs to the field of bioengineering, and provides a trichloroethylene degrading bacterium and application thereof.
Background
Trichloroethylene (TCE) is a common chemical raw material, but is one of the common organic pollutants in environmental water bodies due to unreasonable disposal and emission. It has potential carcinogenic, teratogenic and mutagenic toxicity, and poses a great threat to the ecological environment and human body. In the atmosphere, TCE can be degraded in a period of one week, but the TCE has strong adsorption property and stability, and the half life of the TCE in soil is about 0.5-1.5 years; the half-life in groundwater is about 1-4.5 years. TCE conversion between atmospheric and surface water environments relies primarily on the entry of soil into the groundwater body, thus TCE, once it enters the soil and groundwater environment, causes extensive long-term pollution. So it is important to effectively reduce the TCE pollution in the above-ground environment in time.
TCE can be removed by physical, chemical and biological methods. The physicochemical method has the advantages that more methods such as adsorption method, membrane separation method, stripping method and the like are used, the cost is high, and only pollutants are transferred, so that the pollutants cannot be changed into harmless products, and secondary pollution is easy to cause. The chemical method is currently researched by a TiO2 photocatalysis method, a zero-valent iron reduction method and a bimetal reduction method. Chemical methods are relatively expensive and there is a constant search for more cost effective methods. The TCE is converted into a nontoxic substance through the action of microorganisms, so that the method is economical, effective and free from secondary pollution, and becomes a research hot spot in recent years.
Microorganisms are the main body of the TCE biodegradation process, and metabolism of pollutants by microorganisms is the core of the microbial degradation process. The biological method has the advantages of high treatment efficiency, thorough degradation, no secondary pollution and the like, and is widely applied to the repair and treatment engineering of refractory organic matters. TCE is an organic matter with strong toxicity after all, most experiments prove that high-concentration TCE has strong toxicity to microorganisms, most aerobic bacteria in the research on trichloroethylene degrading bacteria degrade by aerobic-co-metabolism, a growth matrix, namely a first matrix, is used as an electron donor, microorganisms convert non-growth matrixes (trichloroethylene) by using the first matrix as a carbon source and an energy source, the number of the isolated TCE direct degrading bacteria is limited at present, the pollution site is complex, and the TCE can not be effectively removed by only relying on the TCE direct degrading bacteria.
Therefore, the application of the strain with high tolerance and degradation capability to the bioremediation and bioaugmentation technology of TCE is particularly important.
Disclosure of Invention
Aiming at the problems in the prior art, the invention aims to provide the trichloroethylene degrading bacteria with strong environmental adaptability so as to solve the problems of poor biodegradability and difficult degradation of the trichloroethylene.
Firstly, the trichloroethylene degrading bacteria are obtained by collecting samples from a sewage treatment system of a petrochemical sewage plant, enriching, and then carrying out gradient enrichment, separation and purification in an inorganic salt culture medium with trichloroethylene as the only carbon source. Meanwhile, fujiwara test experiments are further carried out, and it is confirmed that the bacteria can grow by taking trichloroethylene as the only carbon source.
The trichloroethylene degrading bacterium is bacillus subtilis Bacillus subtilis and is preserved in China general microbiological culture Collection center (CGMCC) with the preservation number of CGMCC No.24319.
The bacillus subtilis is a gram-positive bacterium, folds on the surface of a bacterial colony are rough and opaque, the bacterial colony is white or yellowish, the edge is irregular, and wire drawing is lifted.
The invention also aims to protect the application of the bacillus subtilis (Bacillus subtilis) in the aspect of trichloroethylene degradation. The specific application range comprises the biological strengthening treatment of waste water, waste gas and soil containing trichloroethylene.
The specific application method comprises the following steps: the bacillus subtilis or the microbial inoculum thereof is put into a treatment system, so that the impact of trichloroethylene pollutants can be effectively resisted, and organic matters in a biochemical system can be utilized as self-grown nutrient substances to rapidly degrade the organic matters. Preferably, the effective viable count of the initial bacillus subtilis of the treatment system is ensured to be 1-8 multiplied by 10 7 cfu/mL, the strain can be quickly adapted to the environment, the trichloroethylene is used as a carbon source for metabolic growth, and the trichloroethylene degradation effect of the strain is effectively exerted.
Furthermore, the trichloroethylene degrading bacterium can be continuously cultured and applied in trichloroethylene sewage, and the degrading effect is stable.
As a further scheme, the invention provides a microbial inoculum of the trichloroethylene degrading bacteria, which is prepared by fermentation:
the fermentation medium comprises the following components in percentage by mass: 5.5 to 6.5 percent of bean pulp, 6 to 6.5 percent of corn starch, 0.3 to 0.5 percent of glucose, 0.3 to 0.5 percent of calcium carbonate, 0.1 to 0.5 percent of corn steep liquor dry powder, 0.1 to 0.3 percent of dipotassium hydrogen phosphate, 0.05 to 0.15 percent of magnesium sulfate, 0.1 to 0.2 percent of sodium chloride, 0.01 to 0.03 percent of manganese sulfate, and the balance of water with the pH value of 7 to 7.5. Sterilizing at 121deg.C for 30 min.
The trichloroethylene degrading bacteria are subjected to high-density fermentation culture for 48-72 hours at the temperature of 30-37 ℃ and are subjected to fermentation to obtain a liquid microbial inoculum, wherein the effective viable count of the microbial inoculum is 5 multiplied by 10 8～2×1010 cfu/mL. The preparation method can be further prepared into solid microbial inoculum and the like, and the preparation method adopts conventional technology.
Further, the fermentation temperature is 33+/-1 ℃, the dissolved oxygen is about 30 percent, the culture is carried out for 48 to 72 hours, the dissolved oxygen is increased, the pH value is reduced, and the fermentation is stopped.
Preferably, the liquid microbial inoculum obtained by the method is applied to a treatment system containing trichloroethylene, the addition amount (by volume) of the microbial inoculum is 1 to 5 per mill of the volume of a biochemical system, and preferably, the biochemical system is at a temperature of between 25 and 38 ℃, the pH value is between 6.0 and 8.0, and the dissolved oxygen is more than or equal to 2mg/L.
Preferably, the initial concentration of the trichloroethylene in the biochemical system is below 900mg/L, more preferably, the initial concentration of the trichloroethylene in the biochemical system is below 300 mg/L; most preferably, the initial concentration of trichloroethylene in the biochemical system is below 150 mg/L.
Compared with the prior art, the invention has the beneficial effects that:
The invention provides a degradation bacterium capable of directly taking trichloroethylene as a unique carbon source, which can rapidly degrade the trichloroethylene under the condition of taking the trichloroethylene as the unique carbon source and energy source, has the characteristics of simplicity and high efficiency in application, has strong toxicity tolerance capability to the trichloroethylene, has wide application range to conditions such as temperature, pH and the like, and has good application prospect in biological purification of industrial wastewater and waste gas.
Preservation information
Preservation time: 2022, 1 month 17;
Preservation unit name: china general microbiological culture Collection center (China Committee for culture Collection);
Preservation number: CGMCC No.24319;
deposit unit address: the dynasty district North Star, department 1, hospital 3 in Beijing;
The classified name is Bacillus subtilis (Bacillus subtilis).
Drawings
FIG. 1 is a graph showing the comparison of biodegradation of trichloroethylene by Bacillus subtilis YJY22-10 at different temperatures in example 4;
FIG. 2 is a graph showing the comparison of biodegradation of trichloroethylene by Bacillus subtilis YJY22-10 under different pH conditions in example 4;
FIG. 3 is a graph showing the degradation of various initial concentrations of trichloroethylene by Bacillus subtilis YJY22-10 in example 4.
Detailed Description
The above-described aspects of the present invention will be described in further detail with reference to the following detailed description, but it should not be construed that the scope of the above-described subject matter of the present invention is limited to the following examples. All techniques realized based on the above description of the present invention are within the scope of the present invention, and the following examples are accomplished by conventional techniques unless otherwise specified.
The term "dissolved oxygen" in the microbial inoculum fermentation process in the following examples refers to air saturation (%), and is a general expression method of dissolved oxygen concentration in the fermentation industry.
The "dissolved oxygen" in the aeration process of sewage treatment in the following examples refers to the absolute concentration of dissolved oxygen in water, expressed in mg/L, and is a common dissolved oxygen expression method for sewage treatment in the environmental protection industry.
EXAMPLE 1 enrichment, screening and separation of trichloroethylene degrading bacteria
And (3) collecting sludge near a sewage treatment system of a petrochemical and agricultural sewage treatment plant area for enrichment and screening treatment, and enriching trichloroethylene degrading bacteria by adopting a method for gradually increasing the concentration of trichloroethylene.
The specific operation method comprises the following steps: taking 10mL of activated sludge sample, inoculating the activated sludge sample into 100mL of enrichment culture medium (initial concentration of trichloroethylene is 10 mg/L), carrying out shaking culture at 30 ℃ for 2-5d by using a shaking table at 160r/min to carry out enrichment culture, taking out 10mL of bacterial liquid, inoculating the bacterial liquid into fresh 100mL of inorganic salt culture medium (concentration of trichloroethylene is 20 mg/L) when the content of trichloroethylene in a sampling detection system is lower than 1mg/L, and repeating the steps by taking 50, 75, 100, 125, 150 and 200mg/L as gradients to gradually increase the concentration of trichloroethylene in the culture medium to 500mg/L.
The final enrichment culture solution is coated on an inorganic salt culture medium plate with trichloroethylene as the only carbon source after gradient dilution, and is cultured for 2-3d in a 30 ℃ incubator. Single colonies with different forms which are grown on the plate are respectively streaked on an inorganic salt culture medium plate for purification for at least 3 times, the single colonies are picked up after the purification and inoculated in 10mL of LB liquid culture medium, the single colonies are respectively inoculated in 100mL of inorganic salt liquid culture medium containing 50mg/L of trichloroethylene with the inoculation amount of 2% (V/V) in the next day, the culture is carried out at 30 ℃ and 160r/min for 24 hours, and then the gas chromatography is adopted for sampling and detecting the content of the trichloroethylene.
10 Strains of bacteria are separated by the method, wherein the strain with the number of 45 can completely degrade 100mg/L of trichloroethylene within 24 hours and store the trichloroethylene.
To further verify that the isolated species was growth metabolised with trichloroethylene as the sole carbon and energy source, a Fujiwara test experiment was performed to detect the concentration of free polychlorinated hydrocarbons in the system. TCE was treated with pyridine in an alkaline environment and then the absorbance of the aqueous phase was measured by spectrophotometry at 470nm, when the formation of pink color indicated as Fujiwara test positive.
The trichloroethylene content in the inorganic salt culture medium is 100mg/L, the experimental group is inoculated with the selected bacteria with the number of 45 by 2% (V/V), the control group is added with sterile water with corresponding content, after culturing for 24 hours at 30 ℃ and 160r/min, sampling is carried out for detection, no pink color of the water phase is detected after the treatment of the experimental group, and the control group is pink and positive. The experiment shows that the strain can take trichloroethylene as the only carbon source for growth metabolism.
Each 1L of the inorganic salt culture medium contains the following components ：K2HPO4 1g,KH2PO4 1g,NaCl 0.5g,NH4Cl 1g,MgSO4·7H2O 0.2g,CaCl2 15mg,FeSO4·7H2O 2mg,CuSO4·5H2O 0.4mg,KI 1mg,MnSO4·H2O 4mg,H3BO3 5mg,CoCl2·6H2O 1mg,Na2MoO4·2H2O 2mg,NiCl2·6H2O2mg,pH 7.0±0.2,, the pH is natural, the inorganic salt culture medium is prepared by distilled water, the temperature is 121 ℃, the sterilization is carried out for 20min, the agar is added into the solid culture medium for 20g, and trichloroethylene is added before the use.
Adding trichloroethylene into the solid culture medium, attaching a piece of sterile filter paper on the cover of the culture dish, dripping the trichloroethylene on the filter paper sheet, and culturing the thallus by using the steam of the trichloroethylene.
The enrichment culture medium is prepared by adding water to 1.0L according to 10.0g of peptone, 5.0g of yeast powder and 10.0g of sodium chloride, regulating the pH value to 7.0, adding agar to the solid culture medium to 20.0g, and sterilizing at 121 ℃ for 20min.
Example 2 identification of species and colony characteristics of Trichloroethylene degrading bacteria
And (3) strain identification: the trichloroethylene degrading bacterium with the number of 45 obtained in the example 1 is subjected to 16S rDNA identification, the NCBI database is used for comparing the 16S rDNA sequences, and the identification result on the molecular level shows that the bacterium belongs to bacillus subtilis, and the nucleotide sequence of the 16S rDNA is shown as SEQ ID in a sequence table.
The trichloroethylene degrading bacterium bacillus subtilis is gram positive bacterium, folds are rough and opaque on the surface of a bacterial colony, the bacterial colony is white or yellowish, the edge is irregular, and wire drawing is lifted. The biological sample is named YJY22-10 and is subjected to biological preservation, and the biological preservation number is CGMCC No.24319
EXAMPLE 3 preparation of trichloroethylene degrading bacteria
The trichloroethylene degrading bacterial agent prepared by the strain of the invention is prepared by the following specific preparation method:
firstly, transferring a test tube inclined plane strain stored on a nutrient agar culture medium at 4 ℃ to a room temperature (20-25 ℃) for activation for 4-8 hours, wherein the culture medium is not required to be additionally added during the activation;
Step two, inoculating the single colony obtained in the step one into LB liquid medium (100 ml), culturing at 35 ℃ and 185r/min overnight to obtain primary seed liquid, inoculating the primary seed liquid into LB medium according to 10% (V/V), culturing for 16 hours under the same conditions to obtain secondary seed liquid, and storing at 4 ℃;
and thirdly, inoculating the secondary seed liquid obtained in the second step into a fermentation culture medium according to 1 per mill (V/V) for fermentation culture, controlling the fermentation temperature to be 33+/-1 ℃ and the dissolved oxygen to be about 30%, culturing for 20-30h, raising the dissolved oxygen, lowering the pH value, and stopping fermentation to obtain the trichloroethylene degrading bacterial agent.
The viable count of the trichloroethylene degrading bacteria obtained in this example was 1.3X10 10 cfu/mL.
The fermentation medium comprises the following components in percentage by mass: 6% of soybean meal, 6% of corn starch, 0.5% of glucose, 0.3% of calcium carbonate, 0.1% of corn steep liquor dry powder, 0.2% of dipotassium hydrogen phosphate, 0.1% of magnesium sulfate, 0.1% of sodium chloride, 0.02% of manganese sulfate, 0.1% of polyether defoamer and pH7-7.5; sterilizing at 121deg.C for 30 min.
The LB culture medium is prepared from 10.0g of peptone, 5.0g of yeast powder and 10.0g of sodium chloride by adding water to 1.0L, adjusting the pH value to 7.0, and sterilizing at 121 ℃ for 20min.
Example 4 detection of degradation Properties of trichloroethylene degrading bacteria
2ML of the trichloroethylene degrading bacterial agent obtained in the example 3 is inoculated into 50mL of an inorganic salt culture medium, wherein the trichloroethylene is used as the only carbon source and energy source, and the degradation capacity is detected. In order to reduce volatilization of trichloroethylene and ensure dissolved oxygen of the strain, the following degradation experiments are carried out in a glass bottle with a 250mL frosted opening.
1. Biological degradation characteristics of bacillus subtilis YJY22-10 on trichloroethylene at different temperatures
The biological degradation characteristic experiment of the bacillus subtilis YJY22-10 on trichloroethylene at different temperatures shows that the bacillus subtilis YJY22-10 has better degradation capability on trichloroethylene at the temperature of more than 30 ℃, and the degradation capability of the bacillus subtilis YJY22-10 at the temperature of 33 ℃ is optimal from the practical application, and the specific implementation steps are as follows:
setting 8 experimental groups, repeating 3 times each group, adopting inorganic salt culture medium (liquid), adding trichloroethylene according to 150mg/L, naturally adjusting pH, respectively setting different culture temperatures after inoculation, setting the experimental temperatures to 20-40 ℃ (20 ℃,25 ℃,28 ℃,30 ℃,33 ℃,35 ℃,38 ℃ and 40 ℃), placing in a shaking table for 160r/min shake culture, sampling after 24 hours, and performing headspace gas phase detection to obtain the optimal action temperature of 25-40 ℃, shown in figure 1 in detail;
2. biological degradation characteristics of bacillus subtilis YJY22-10 on trichloroethylene under different pH conditions
7 Experimental groups are set, 3 repeats of each group are carried out, an inorganic salt culture medium is adopted as a culture medium, trichloroethylene is added according to 150mg/L, different pH values are respectively set, the experimental range of the pH values is set to be 4-10 (respectively set to be 4,5,6,7,8,9 and 10), after inoculation, shake culture is carried out in a shaking table at 30 ℃ and 160r/min, sampling extraction is carried out after 24 hours for detection, the application range is wider, and the optimal application pH value of the culture medium is obtained to be 6.5-8.0, and is shown in figure 2 in detail.
3. Degradation of different initial concentrations of trichloroethylene by Bacillus subtilis YJY22-10
Setting 5 experimental groups, setting three repetitions of each group, adding 50mL of inorganic salt culture medium into a grinding triangular flask, taking trichloroethylene as a unique carbon source, wherein the initial concentration of the trichloroethylene in each experimental group is 146 mg/L, 293 mg/L, 586 mg/L, 879 mg/L and 1172mg/L respectively, culturing at the pH of natural and 30 ℃ for 160r/min, sampling and detecting the residual amount of the trichloroethylene after 24 hours, and when the initial concentration is below 900mg/L, the degradation rate of the trichloroethylene in 24 hours can reach more than 80 percent; when the initial concentration is below 150mg/L, the degradation rate of the catalyst in 24 hours can reach about 100 percent. The strain has strong toxicity tolerance to trichloroethylene, and the detail is shown in figure 3.
Example 5 simulation application of trichloroethylene degrading bacteria in trichloroethylene sewage treatment
A trichloroethylene degrading bacterial agent was prepared in the same manner as in example 3, the culture temperature was 33.+ -. 1 ℃ and the dissolved oxygen was controlled to be 30%, and the viable count of the obtained bacterial agent was 5X 10 8 cfu/mL.
Simulation application: the source of the treated water is the water entering an aerobic section of a pesticide sewage treatment plant, the treatment capacity is 5L, the addition amount of the trichloroethylene degrading bacteria is added according to 0.3% (V/V), the aeration treatment is carried out at room temperature to ensure that the dissolved oxygen is more than 2mg/L, the pH of the sewage is 6.0-8.0, the COD of the raw sewage is about 4000mg/L, the ammonia nitrogen is about 380mg/L, the organic amine is about 200mg/L, the trichloroethylene concentration is artificially added to 100mg/L, the trichloroethylene content of the sewage is less than 1mg/L after 24 hours, and the degradation rate reaches more than 99.9%.
Example 6 application of Trichloroethylene degrading bacteria in Trichloroethylene wastewater treatment
A trichloroethylene degrading bacterial agent was prepared in the same manner as in example 3, wherein the culturing temperature was 35.+ -. 1 ℃ and the dissolved oxygen was controlled to be 30%, and the viable count of the obtained bacterial agent was 1.5X10 10 cfu/mL.
The source of the treated water is the water inlet of an aerobic tank of a petrochemical sewage treatment plant, the treatment capacity is 10L, the addition amount of the trichloroethylene degrading bacteria is 0.2% (V/V), the dissolved oxygen is more than 2mg/L, the pH of sewage is 6.0-8.0 by aeration treatment at room temperature, the trichloroethylene concentration is less than 5mg/L, the COD is about 2200mg/L, the volatile phenol is about 240mg/L, the ammonia nitrogen is about 100mg/L, the sulfide is about 10mg/L, water inlet and outlet are carried out once every 24 hours, the concentration of the trichloroethylene in the water inlet and outlet is detected, the system is continuously used, and the bacteria can be seen to have continuous degradation capacity according to the tracking condition.
Claims (9)
1. A trichloroethylene degrading bacterium is characterized in that the strain is bacillus subtilis (Bacillus subtilis) and is preserved in China general microbiological culture Collection center (CGMCC) with the preservation number of CGMCC No.24319.
2. The application of the bacillus subtilis in the aspect of trichloroethylene degradation is characterized in that the bacillus subtilis is preserved in China general microbiological culture collection center (CGMCC) with the preservation number of CGMCC No.24319.
3. The use of bacillus subtilis according to claim 2 in the degradation of trichloroethylene, characterized in that the specific application range comprises: and (3) carrying out biological strengthening treatment on waste water, waste gas and soil containing trichloroethylene.
4. The application of bacillus subtilis in the aspect of trichloroethylene degradation according to claim 2, which is characterized in that the specific application method is as follows: the bacillus subtilis or the microbial inoculum thereof is put into a treatment system, so that the effective viable count of the initial bacillus subtilis of the treatment system is ensured to be 1-8 multiplied by 10 7 cfu/mL.
5. The use of bacillus subtilis according to claim 2 for the degradation of trichloroethylene, characterized in that it is used in continuous culture in trichloroethylene sewage.
6. The trichloroethylene degrading bacterial agent is characterized by being prepared by fermentation:
The fermentation medium comprises the following components in percentage by mass: 5.5 to 6.5 percent of bean pulp, 6 to 6.5 percent of corn starch, 0.3 to 0.5 percent of glucose, 0.3 to 0.5 percent of calcium carbonate, 0.1 to 0.5 percent of corn steep liquor dry powder, 0.1 to 0.3 percent of dipotassium hydrogen phosphate, 0.05 to 0.15 percent of magnesium sulfate, 0.1 to 0.2 percent of sodium chloride, 0.01 to 0.03 percent of manganese sulfate, the balance of water and pH7 to 7.5; autoclaving at 121deg.C for 30 min;
Culturing trichloroethylene degrading bacteria at 30-37deg.C for 48-72 hr to obtain liquid microbial inoculum with effective viable count of 5×10 8~2×1010 cfu/mL;
The trichloroethylene degrading bacteria are bacillus subtilis (Bacillus subtilis) and are preserved in China general microbiological culture Collection center (CGMCC) with the preservation number of CGMCC No.24319.
7. The trichloroethylene degrading bacterial agent according to claim 6, wherein the fermentation temperature is 33+ -1 ℃, dissolved oxygen is about 30%, and the fermentation is stopped after culturing for 48-72 hours with rising dissolved oxygen and lowering pH.
8. The method for using the trichloroethylene degrading bacterial agent as defined in claim 6, wherein the obtained liquid bacterial agent is applied to a treatment system containing trichloroethylene, the bacterial agent is added in an amount of 1-5 per mill of the volume of a biochemical system, the biochemical system condition is that the temperature is 25-38 ℃, the pH is 6.0-8.0, and the dissolved oxygen is more than or equal to 2mg/L.
9. The method of claim 8, wherein the initial concentration of trichloroethylene in the biochemical system is less than 900 mg/L.
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