CN106964647A - A kind of method of utilization composite bacteria agent remediating lead-contaminated soil - Google Patents
A kind of method of utilization composite bacteria agent remediating lead-contaminated soil Download PDFInfo
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- CN106964647A CN106964647A CN201710417799.8A CN201710417799A CN106964647A CN 106964647 A CN106964647 A CN 106964647A CN 201710417799 A CN201710417799 A CN 201710417799A CN 106964647 A CN106964647 A CN 106964647A
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- soil
- bacillus subtilis
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- bacillus
- lead
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B09—DISPOSAL OF SOLID WASTE; RECLAMATION OF CONTAMINATED SOIL
- B09C—RECLAMATION OF CONTAMINATED SOIL
- B09C1/00—Reclamation of contaminated soil
- B09C1/10—Reclamation of contaminated soil microbiologically, biologically or by using enzymes
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
Abstract
The invention belongs to microbial technology field, a kind of method of utilization composite bacteria agent remediating lead-contaminated soil is disclosed, it comprises the following steps：First that soil scarifying is smooth, spray water, holding soil moisture content is 30wt%, then sows microbial inoculum using the addition of 200g microbial inoculums according to soil per ton, the 3rd day, then soil of scarifying sowed microbial inoculum according to soil per ton using the addition of 100g microbial inoculums；8th day, detect lead content in soil.Operation is simple for the restorative procedure of the present invention, and rehabilitation cost is low, environment friendly and pollution-free, can be applied in large-scale lead-contaminated soil, has a extensive future.
Description
Technical field
The invention belongs to microbial technology field, and in particular to a kind of side of utilization composite bacteria agent remediating lead-contaminated soil
Method.
Background technology
Heavy metal generally refers to about 60 kind elements or proportion about 45 kind members more than 5.0 of the proportion more than 4.0
Element.It is significant that heavy metal in Study on environmental pollution is primarily referred to as the bio-toxicity such as Pb, Cd, Hg, Cr and metalloid As
Element, in addition to the element such as the heavy metal Zn with certain toxicity, Cu, Co, Ni and Sn.Some heavy metal elements be human body and
Element necessary to other organisms, but its concentration exceedes certain limit and will cause poisoning.Soil pollution by heavy metal refers to people
Class activity inputs heavy metal in soil, causes heavy metal content in soil apparently higher than background content, and cause ecological environment matter
Measure the phenomenon deteriorated.Investigation display, Pearl River Delta urban You Jin40% farmlands vegetable plot soil pollution by heavy metal is exceeded, wherein
10% category severe overweight.Decades can be typically remained in heavy metal in soil, wherein lead can be remained 100 years.
Lead belongs to one kind of heavy metal, is a kind of non-essential element of human body, and it can influence nerve, hematopoiesis, reproduction and hair
Educate, endocrine, sorts of systems and the organ such as immune, bone.About 5,000,000 tons of lead, wherein most are discharged every year on average in the whole world
Into soil, countries in the world soil is caused different degrees of lead contamination occur.Development and leaded chemical substance with industrial or agricultural
It is widely used, lead largely enters soil during exploitation of mineral resources, metallurgical, the waste processing disposal of plating etc. containing land pollutant
Environment, causes in world wide lead contamination phenomenon generally existing in soil.Harm of the lead to human health is very big, can to nerve,
Cardiovascular and endocrine etc. system causes harm, or even threat to life.Therefore the improvement and reparation of lead-contaminated soil are by more
Carry out more concerns.
Approach is mainly repaired in current Lead Pollution in Soil reparation three kinds：Peripheral doses, chemical remediation and biological prosthetic.Peripheral doses
Have the advantages that thorough, stable, but implement that quantities is big, investment cost is high, destroy soil texture, and will also be to swapping out dirt
Soil is stacked or handled.Chemical remediation is although simple and easy to apply, instant effect, but often renovation agent large usage quantity, and heavy metal member
Element is easily-activated.Biological prosthetic then to include phytoremediation and microorganism remediation, although wherein phytoremediation treats both principal and secondary aspect of disease, but repairs week
Phase is long, and investment is big.Although microorganism remediation processing cost is low, it is small that surrounding enviroment are disturbed, do not produce secondary pollution, suitable micro-
Living resources are few, and repairing effect is not obvious.
The content of the invention
In order to overcome the defect of biological prosthetic lead-contaminated soil in the prior art, compound bacteria is utilized the invention provides one kind
The method of agent remediating lead-contaminated soil.
The present invention adopts the following technical scheme that realization
A kind of method of utilization composite bacteria agent remediating lead-contaminated soil, it comprises the following steps：
First that soil scarifying is smooth, spray water, holding soil moisture content is 30wt%, then uses 200g according to soil per ton
The addition of microbial inoculum sows microbial inoculum, and the 3rd day, then soil of scarifying used the addition of 100g microbial inoculums according to soil per ton
Sow microbial inoculum；8th day, detect lead content in soil；Complete the reparation of soil.
Specifically, the microbial inoculum is prepared according to following technique：
By bacillus pumilus seed liquor（1×108cfu/ml）With bacillus subtilis seed liquor（1×108cfu/ml）According to
1-2:2-3 volume ratio mixing, then gone to according to 10% inoculum concentration and 24h cultivated in fermentation tank, obtain bacillus pumilus-
Bacillus subtilis mixed fermentation liquid；
Preferably, the Fermenter Medium Component is：According to mass percent, glucose 8%, yeast extract 5%, corn steep liquor 3%,
Urea 0.1%, ferrous sulfate 0.02%, magnesium sulfate 0.02%, potassium dihydrogen phosphate 0.01%, dipotassium hydrogen phosphate 0.01%, pH 6.5.
By bacillus pumilus-bacillus subtilis mixed fermentation liquid, Gordonia bronchialis zymotic fluid, acinetobacter calcoaceticus hair
Zymotic fluid, sphingol pseudomonad zymotic fluid and into brilliant pole fermented liquid according to 5-7:3-4:2-3:1-2:1-2 volume ratio
It is well mixed, obtain liquid bacterial agent；
Preferably, Gordonia bronchialis zymotic fluid, acinetobacter calcoaceticus zymotic fluid, sphingol pseudomonad zymotic fluid and Cheng Jingjie
The concentration of bacillus fermentation liquid is controlled 1 × 1010cfu/ml；
Wheat stalk is crushed with pulverizer, 100 mesh sieves is then crossed, powder of straw is obtained, by powder of straw and diatomite according to 1:1
Mass ratio, which is mixed evenly, obtains carrier；
Liquid bacterial agent is added in the carrier of equivalent weight, then 200rpm stirring 3min carry out low temperature drying, drying temperature
For 15-20 DEG C, water content is 6-10% after drying, and packaging is produced.
Preferably, the bacillus pumilus is bacillus pumilus (Bacillus pumilus) CGMCC NO.7126；
The bacillus subtilis is bacillus subtilis（Bacillus subtilis）CGMCC No：0954；The Gordonia bronchialis is
Gordonia bronchialis (Gordonia amicalis) CGMCC No.4794；The acinetobacter calcoaceticus is acinetobacter calcoaceticus
(Acinetobactercalcoaceticus) CGMCC No.7334；The sphingol pseudomonad is sphingol pseudomonad
（Sphingomonas sp.）CGMCC No.4589；It is described to be into brilliant arthrobacterium (Arthrobacter into brilliant arthrobacterium
crystallopoietes)ATCC 15481。
Strain of the present invention can be from CGMCC and American Type Culture collection warehousing（ATCC）Etc. commercial sources purchase
It can buy.Each strain fermentation culture of the present invention is the cellar culture mode of this area, is not innovative point of the present invention, unknown herein
State.
The beneficial effect that the present invention is obtained mainly includes but is not limited to the following aspects：
Operation is simple for the restorative procedure of the present invention, and rehabilitation cost is low, environment friendly and pollution-free, can be applied to large-scale lead contamination
In soil, have a extensive future.
Reasonable compatibility between each strain in composite bacteria agent of the present invention, symbiosis is coordinated, mutually not antagonism, and activity is high, can be effective
Ground handles lead-contaminated soil；
Relative to single fermentation mode, the synergy between mixed fermentation bacillus subtilis and bacillus pumilus causes bacterium
The activity of SOD and dehydrogenase is greatly improved in strain, improves heavy metal repair ability；
The present invention, as carrier, not only strengthens the time-to-live of bacterial strain and the field planting energy in soil using diatomite and powder of straw
Power, heavy metal also has stronger adsorptivity, can also be supplied nutrients for soil.
Embodiment
In order that those skilled in the art more fully understand the technical scheme in the application, have below in conjunction with the application
Body embodiment, the technical scheme to the application is clearly and completely described, it is clear that described embodiment is only this Shen
Please a part of embodiment, rather than whole embodiment.Based on the embodiment in the application, those of ordinary skill in the art are not having
There is the every other embodiment made and obtained under the premise of creative work, should all belong to the scope of protection of the invention.
Embodiment 1
A kind of method of utilization composite bacteria agent remediating lead-contaminated soil, it comprises the following steps：
First that soil scarifying is smooth, spray water, holding soil moisture content is 30wt%, then uses 200g according to soil per ton
The addition of microbial inoculum sows microbial inoculum, and the 3rd day, then soil of scarifying used the addition of 100g microbial inoculums according to soil per ton
Sow microbial inoculum；8th day, detect lead content in soil；Complete the reparation of soil.
The microbial inoculum is prepared according to following technique：
By bacillus pumilus seed liquor（1×108cfu/ml）With bacillus subtilis seed liquor（1×108cfu/ml）According to
1:2 volume ratio mixing, then goes to according to 10% inoculum concentration and 24h is cultivated in fermentation tank, obtain bacillus pumilus-withered grass
Bacillus mixed fermentation liquid；
The Fermenter Medium Component is：According to mass percent, glucose 8%, yeast extract 5%, corn steep liquor 3%, urea
0.1%, ferrous sulfate 0.02%, magnesium sulfate 0.02%, potassium dihydrogen phosphate 0.01%, dipotassium hydrogen phosphate 0.01%, pH 6.5.
By bacillus pumilus-bacillus subtilis mixed fermentation liquid, Gordonia bronchialis zymotic fluid, acinetobacter calcoaceticus hair
Zymotic fluid, sphingol pseudomonad zymotic fluid and into brilliant pole fermented liquid according to 5:3:2:1:1 volume ratio is well mixed, and is obtained
To liquid bacterial agent；Gordonia bronchialis zymotic fluid, acinetobacter calcoaceticus zymotic fluid, sphingol pseudomonad zymotic fluid and Cheng Jingjie
The concentration of bacillus fermentation liquid is controlled 1 × 1010cfu/ml；
Wheat stalk is crushed with pulverizer, 100 mesh sieves is then crossed, powder of straw is obtained, by powder of straw and diatomite according to 1:1
Mass ratio, which is mixed evenly, obtains carrier；
Liquid bacterial agent is added in the carrier of equivalent weight, then 200rpm stirring 3min carry out low temperature drying, drying temperature
For 15 DEG C, water content is 10% after drying, and packaging is produced.
Bacterial strain is selected from following：Bacillus pumilus (Bacillus pumilus) CGMCC NO.7126（Reference can be made to
CN104152377A）, bacillus subtilis（Bacillus subtilis）CGMCC No：0954（Reference can be made to CN1554744A）、
Gordonia bronchialis (Gordonia amicalis) CGMCC No.4794（Reference can be made to CN102250801A）, acinetobacter calcoaceticus
(Acinetobactercalcoaceticus) CGMCC No.7334（Reference can be made to CN 103266073A）, the false unit cell of sphingol
Bacterium（Sphingomonas sp.）CGMCC No.4589（Reference can be made to CN102168054A）, into brilliant arthrobacterium (Arthrobacter
crystallopoietes)ATCC 15481。
Embodiment 2
A kind of method of utilization composite bacteria agent remediating lead-contaminated soil, it comprises the following steps：
First that soil scarifying is smooth, spray water, holding soil moisture content is 30wt%, then uses 200g according to soil per ton
The addition of microbial inoculum sows microbial inoculum, and the 3rd day, then soil of scarifying used the addition of 100g microbial inoculums according to soil per ton
Sow microbial inoculum；8th day, detect lead content in soil；Complete the reparation of soil.
The microbial inoculum is prepared according to following technique：
By bacillus pumilus seed liquor（1×108cfu/ml）With bacillus subtilis seed liquor（1×108cfu/ml）According to
2:3 volume ratio mixing, then goes to according to 10% inoculum concentration and 24h is cultivated in fermentation tank, obtain bacillus pumilus-withered grass
Bacillus mixed fermentation liquid；
The Fermenter Medium Component is：According to mass percent, glucose 8%, yeast extract 5%, corn steep liquor 3%, urea
0.1%, ferrous sulfate 0.02%, magnesium sulfate 0.02%, potassium dihydrogen phosphate 0.01%, dipotassium hydrogen phosphate 0.01%, pH 6.5.
By bacillus pumilus-bacillus subtilis mixed fermentation liquid, Gordonia bronchialis zymotic fluid, acinetobacter calcoaceticus hair
Zymotic fluid, sphingol pseudomonad zymotic fluid and into brilliant pole fermented liquid according to 7:4:3:2:2 volume ratio is well mixed, and is obtained
To liquid bacterial agent；Gordonia bronchialis zymotic fluid, acinetobacter calcoaceticus zymotic fluid, sphingol pseudomonad zymotic fluid and Cheng Jingjie
The concentration of bacillus fermentation liquid is controlled 1 × 1010cfu/ml；
Wheat stalk is crushed with pulverizer, 100 mesh sieves is then crossed, powder of straw is obtained, by powder of straw and diatomite according to 1:1
Mass ratio, which is mixed evenly, obtains carrier；
Liquid bacterial agent is added in the carrier of equivalent weight, then 200rpm stirring 3min carry out low temperature drying, drying temperature
For 20 DEG C, water content is 8% after drying, and packaging is produced.
Bacterial strain is selected from following：Bacillus pumilus (Bacillus pumilus) CGMCC No.7126, bacillus subtilis
（Bacillus subtilis）CGMCC No：0954th, Gordonia bronchialis (Gordonia amicalis) CGMCC No.4794, second
Sour calcium acinetobacter calcoaceticus (Acinetobacter calcoaceticus) CGMCC No.7334, sphingol pseudomonad
（Sphingomonas sp.）CGMCC No.4589, into brilliant arthrobacterium (Arthrobacter crystallopoietes) ATCC
15481。
Embodiment 3
The repairing test of lead-contaminated soil：
Sample is selected：
Lead tolerance is that 200mg/kg is lead-contaminated soil sample；
Group：
Test group：1 group of embodiment；Control group 1（Without bacillus pumilus-bacillus subtilis mixed fermentation liquid, other are same
Embodiment 1）, control group 2（Without Gordonia bronchialis zymotic fluid, other be the same as Examples 1）, control group 3（It is false single without sphingol
Born of the same parents' fermented liquid, other be the same as Examples 1）, control group 4（Bacillus pumilus and bacillus subtilis are using the side separately fermented
Formula, other be the same as Examples 1）；
Operating process：
Soil after each group is handled is dried at 65 DEG C, accurately weighs 1g, uses BCR continuous extractions, and extracting can hand in soil
Change the lead of state.Step is as follows：The accurate air-dried pedotheque 1g weighed by 100 mesh sieves, plus 40mL 0.1mol/L HAc, put
Continuous oscillation 16h at 24 ± 1 DEG C, then centrifuges 15min under 5000rpm in constant temperature oscillator.Supernatant is taken, ICP-AES is used
Determine Pb2+Content.Concrete outcome is shown in Table 1.
Table 1
| Group | Test group | Control group 1 | Control group 2 | Control group 3 | Control group 4 |
| Lead concentration（mg/kg） | 4.86 | 41.53 | 23.41 | 27.96 | 16.47 |
Conclusion：Each bacterial strain is mutually cooperateed with composite bacteria agent of the present invention, and compatibility rationally, except lead ability preferably, has a extensive future.
In the description of this specification, reference term " one embodiment ", " some embodiments ", " example ", " specifically show
The description of example " or " some examples " etc. means to combine specific features, structure, material or the spy that the embodiment or example are described
Point is contained at least one embodiment of the present invention or example.In this manual, to the schematic representation of above-mentioned term not
Identical embodiment or example must be directed to.Although embodiments of the invention have been shown and described above, Ke Yili
Solution, above-described embodiment is exemplary, it is impossible to be interpreted as limitation of the present invention, one of ordinary skill in the art is at this
Above-described embodiment can be changed, changed in the range of invention, replaced and modification.
Claims (5)
1. a kind of method of utilization composite bacteria agent remediating lead-contaminated soil, it comprises the following steps：
First that soil scarifying is smooth, spray water, holding soil moisture content is 30wt%, then uses 200g according to soil per ton
The addition of microbial inoculum sows microbial inoculum, and the 3rd day, then soil of scarifying used the addition of 100g microbial inoculums according to soil per ton
Sow microbial inoculum；8th day, detect lead content in soil.
2. according to the method described in claim 1, it is characterised in that the composite bacteria agent is prepared according to following technique：
1）By bacillus pumilus-bacillus subtilis mixed fermentation liquid, Gordonia bronchialis zymotic fluid, acinetobacter calcoaceticus fermentation
Liquid, sphingol pseudomonad zymotic fluid and into brilliant pole fermented liquid according to 5-7:3-4:2-3:1-2:1-2 volume ratio is mixed
Close uniform, obtain liquid bacterial agent；
2）Wheat stalk is crushed with pulverizer, 100 mesh sieves is then crossed, powder of straw is obtained, by powder of straw and diatomite according to 1:1
Mass ratio be mixed evenly and obtain carrier；
3）Liquid bacterial agent is added in the carrier of equivalent weight, then 200rpm stirring 3min carry out low temperature drying, dry temperature
Spend for 15-20 DEG C, water content is 6-10wt% after drying, packaging is produced.
3. method according to claim 2, it is characterised in that the bacillus pumilus-bacillus subtilis mixing hair
Zymotic fluid is prepared in accordance with the following steps：Bacillus pumilus seed liquor and bacillus subtilis seed liquor are pressed into 1-2:2-3's
Volume ratio is mixed, and is then gone to according to 10% inoculum concentration and 24h is cultivated in fermentation tank, obtain bacillus pumilus-bacillus subtilis
Bacterium mixed fermentation liquid.
4. method according to claim 3, it is characterised in that the Fermenter Medium Component is：According to quality percentage
Than glucose 8%, yeast extract 5%, corn steep liquor 3%, urea 0.1%, ferrous sulfate 0.02%, magnesium sulfate 0.02%, potassium dihydrogen phosphate
0.01%, dipotassium hydrogen phosphate 0.01%, pH 6.5.
5. method according to claim 2, it is characterised in that the bacillus pumilus is bacillus pumilus
(Bacillus pumilus)CGMCC NO.7126；The bacillus subtilis is bacillus subtilis（Bacillus
subtilis）CGMCC No：0954；The Gordonia bronchialis is Gordonia bronchialis (Gordonia amicalis) CGMCC
No.4794；The acinetobacter calcoaceticus is acinetobacter calcoaceticus (Acinetobactercalcoaceticus) CGMCC
No.7334；The sphingol pseudomonad is sphingol pseudomonad（Sphingomonas sp.）CGMCC No.4589；Institute
State into brilliant arthrobacterium and be into brilliant arthrobacterium (Arthrobacter crystallopoietes) ATCC 15481.
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| CN107617639A (en) * | 2017-11-03 | 2018-01-23 | 裘正荣 | It is a kind of to utilize microorganism and the method for mineral matter combined governance pollution by polychlorinated biphenyles soil |
| CN108611099A (en) * | 2018-04-19 | 2018-10-02 | 郭婷 | A kind of soil-repairing agent that resource utilization is high |
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| CN110628674A (en) * | 2019-09-29 | 2019-12-31 | 山东农业大学 | Bacillus pumilus with functions of improving acid soil and removing potassium and preparation and application of microbial inoculum thereof |
| CN115125174A (en) * | 2022-07-19 | 2022-09-30 | 广东省科学院动物研究所 | Lead-resistant bacillus pumilus and application thereof |
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