CN110241111A - A kind of method of modified activated carbon immobilized cell enhancing bacterium chromium reducing power - Google Patents
A kind of method of modified activated carbon immobilized cell enhancing bacterium chromium reducing power Download PDFInfo
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Abstract
The invention discloses a kind of methods of modified activated carbon immobilized cell enhancing micrococcus luteus chromium reducing power, belong to environment remediation technical field.Immobilized spherule reagent major parameter: sodium alginate (3%, w/v), calcium chloride (4%, w/v), micrococcus luteus bacterium solution, modified activated carbon 0.1-0.5g, sodium alginate soln, micrococcus luteus bacterium solution and modified activated carbon are uniformly mixed, it instills calcium chloride solution and carries out cross-linking reaction, it after standing four hours at room temperature, is added in hexavalent chromium solution, carries out reduction reaction.Add the reduction effect of 0.1-0.5g modified activated carbon Cr VI compared with the reduction effect that modified activated carbon is not added, reduction rate improves 8.4%-13.4%.The present invention utilizes the absorption feature of active carbon, and immobilization bacterium can be promoted efficiently to restore the hexavalent chromium in water body, significantly improve micrococcus luteus to hexavalent chrome reduction efficiency in water body.
Description
Technical field
The invention belongs to environment remediation technical fields, and in particular to a kind of modified activated carbon immobilized cell enhancing bacterium chromium
The method of reducing power.
Background technique
Chromium is a kind of element that the content in natural resources is relatively high and distributed more widely.Chromium is mainly with naturally occurring and people
It is entered in various environment for two kinds of approach of activity, such as surface water, underground water, seawater, soil, rock and air, and with difference
Concentration and different combining forms are present in the earth's crust.Chromium is a kind of most common metal pollutant, be U.S. EPA generally acknowledge 129
One of kind emphasis pollution nuisance, while being also one of internationally recognized three kinds of carcinogenic substances.Chromium for Recent Years contamination accident is in China
Hunan, Yunnan etc. multiple situation mostly is presented, cause serious social influence.The thus extensive pass by scientists
Note, becomes the hot and difficult issue research topic of environment in recent years area research.The physical chemistry administering method of pollution of chromium exists
Various disadvantage.Therefore, domestic and international researcher starts goal in research being transferred to novel method -- on biological restoration.It is raw
Object repairing method is to pass through microorganism to the metabolism of the absorption of Cr (VI), enrichment or microorganism for high toxicity and Qiang Qian
The Cr (VI) of shifting property is converted into the Cr (III) of hypotoxicity and weak migration, and final realize reduces Cr (VI) ion contained in environment
Purpose.It is practical until just starting to be applied to after the 1980s that heavy metal pollution is administered using biological restoration.Biology
The advantages of repairing method, is that easily controllable, investment is smaller, efficiency is higher, at low cost and without secondary pollution, therefore in environment remediation
On have a good application prospect.But microorganism remediation has microorganism adaptability poor, vulnerable to external environment influence, efficiency
Low disadvantage is restricted its extensive use.In order to improve the tolerance of microorganism, and the metal of increase bacterial strain
Reduction efficiency, a large number of researchers have probed into various enhancement microbiologicals and have repaired chromium from multiple directions such as physics, chemistry, biologies
The method of pollution.
Immobilized microorganism technology obtains relatively broad in terms of handling toxic middle metal and organic pollution difficult to degrade
Application and have preferable achievement.Compared with traditional biologic treating technique, activity is combined using immobilized microorganism technology
It is hexavalent chromium polluted in the characterization of adsorption reduction water body of charcoal, there is apparent superiority.The present invention passes through addition modified activated carbon energy
The reduction efficiency of Cr VI is enough significantly increased, therefore achievement of the present invention has good practicability.
Summary of the invention
The purpose of the invention is to improve microorganism to the efficiency of hexavalent chrome reduction, it is solid a kind of modified activated carbon has been invented
Surely change the method that cell increases bacterium chromium reducing power, the present invention can significantly increase microorganism to the reducing power of Cr VI.
Technical solution of the present invention is summarized as follows:
A kind of method that modified activated carbon immobilized cell increases bacterium chromium reducing power, comprising the following steps:
1. it is 200 mesh or more that active carbon, which is broken into granule size,；
2. being added to 5g active carbon containing 50ml 70%HNO3Flask in, and at room temperature with 170rpm stir 1h
To remove the impurity in active carbon, it is washed with deionized water to neutrality, it is last sufficiently dry；Active carbon after drying is added
In 100ml 10% (w/v) PEI (polyetherimide) methanol solution, at room temperature with 170rpm stirring 24 hours, immediately after will
Active carbon is transferred in 200ml 1% (w/v) glutaraldehyde solution and is crosslinked, and is finally washed with deionized to obtain modified active
Charcoal；
3. bacterium source is separated in laboratory screening, Selective agar medium is improvement LB liquid medium, and formula is prepared: albumen
Peptone 10gL-1, sodium chloride 5gL-1, yeast extract 5gL-1, dipotassium hydrogen phosphate 0.05gL-1, epsom salt 0.2g
L-1, if solid LB media, then plus agar 1.5~2.0%, pH 8.5~9.0 sterilize；
4. isolated micrococcus luteus will be screened containing shaking flask culture is carried out in LB culture medium, condition of culture is initial
PH 8.0-9.0,30 DEG C of temperature, shaking speed 170rpm；
5. the condition of immobilized spherule are as follows: sodium alginate 3g, calcium chloride 4g, modified activated carbon 0.1-0.5g, by alginic acid
Sodium and modified activated carbon are dissolved in 90ml distilled water, sterilizing；
6. cultured micrococcus luteus in step (4) is filtered, is collected by centrifugation, and press inoculum concentration OD600Value is 1.5, is used
Sterile water is made into 10ml and is uniformly mixed with mixed liquor in step (4), be slowly dropped into mass concentration be 4%w/v calcium chloride solution in,
After crosslinking four hours, it is added and contains in 120mg/L Cr (VI) LB liquid medium, room temperature shaker culture 84h.
7. configuring diphenylcarbazide solution, 0.2g diphenylcarbazide is dissolved in 50ml acetone, moves to 100ml volumetric flask,
It is diluted with water to graduation mark, is shaken up, sequentially adds the dense H of 12.5ml slowly into refrigerant liquid2SO4With 12.5ml concentrated phosphoric acid, it is protected from light
It saves.
In the step (3), sample 10g is taken to be added in the sterilized liquid LB culture medium of 90ml improvement, with 10 times of ladders
Degree is diluted, and respectively draws 0.2ml, is coated on the solid improvement LB culture medium of Cr containing 200mg/L (VI), will with spreading rod
Bacterium solution smoothens, and cultivates 24 hours in 30 DEG C of constant incubators and grows to bacterium colony, identifies isolated bacterial strain, is micrococcus luteus.
To diphenylcarbazide solution obtained by step (7), using in diphenyl carbazide spectrophotometry measurement sample six
The concentration of valence chromium, the calculation formula of Cr (VI) concentration are y=0.6031x+0.0065, the coefficient R of standard curve2=
0.0997。
Modified active Carbon Materials are each to the removal of pollutant advantageous, modified self structure (specific surface area, porosity
Deng) or performance optimized, it is more preferable to the repairing effect of heavy metal pollution.Method of the invention passes through improvement active carbon and control
The dosage (0.3g) of preparing active carbon can significantly improve micrococcus luteus and improve to hexavalent chrome reduction efficiency, reduction rate in water body
13.4%.
Detailed description of the invention
Fig. 1 is the hexavalent chromium concentration trend graph at any time of embodiment 1；
Fig. 2 is the hexavalent chromium concentration trend graph at any time of embodiment 2；
Fig. 3 is the hexavalent chromium concentration trend graph at any time of embodiment 3；
Fig. 4 is the hexavalent chromium concentration trend graph at any time of embodiment 4；
Fig. 5 is the hexavalent chromium concentration trend graph at any time of embodiment 5；
Fig. 6 is the display figure without adding modified activated carbon immobilized spherule；
Fig. 7 is the display figure for adding modified activated carbon immobilized spherule.
Specific embodiment
The purpose of following specific embodiments or embodiment is in order to further illustrate the present invention, rather than to limit of the invention
It is fixed.
Example 1
The present embodiment the method mainly sequentially includes the following steps:
(1) active carbon is broken into particle size is 200 mesh or more；
(2) 5g active carbon is added to containing 50ml 70%HNO3Flask in, and at room temperature with 170rpm stir 1h
To remove the impurity in active carbon, it is washed with deionized water to neutrality, it is last sufficiently dry；Active carbon after drying is added
In 100ml 10% (w/v) PEI (polyetherimide)/methanol solution, at room temperature with 170rpm stirring 24 hours, immediately after will
Active carbon is transferred in 200ml 1% (w/v) glutaraldehyde solution and is crosslinked, and is finally washed with deionized to obtain modified active
Charcoal；
(3) LB liquid medium is configured, shaking flask culture micrococcus luteus to logarithmic growth phase takes OD600The bacterium solution of value about 1.5,
It is collected by centrifugation, is configured to 10ml bacterial suspension with sterile water；
(4) 3g sodium alginate and 0.1g modified activated carbon are weighed, 90ml is added and distills aqueous systems, sterilizing will be in step (2)
10ml bacterial suspension is mixed evenly with it, is uniformly instilled in 4%w/v calcium chloride solution, is stood four hours at room temperature, will be washed
Bead afterwards is poured into containing in 120mg/L Cr (VI) LB liquid medium, and condition of culture is initial pH 8.0,30 DEG C of temperature, is shaken
Bed revolving speed 170rpm, cultivates 84h；
Conclusion: as shown in Figure 1, the hexavalent chrome reduction rate of the modified activated carbon of addition 0.1g is 72.5%, modification is not added
The hexavalent chrome reduction rate of active carbon is 64.1%, and hexavalent chrome reduction rate improves 8.4%.
Embodiment 2
The present embodiment the method mainly sequentially includes the following steps:
(1) active carbon is broken into particle size is 200 mesh or more；
(2) 5g active carbon is added to containing 50ml 70%HNO3Flask in, and at room temperature with 170rpm stir 1h
To remove the impurity in active carbon, it is washed with deionized water to neutrality, it is last sufficiently dry；Active carbon after drying is added
In 100ml 10% (w/v) PEI (polyetherimide)/methanol solution, at room temperature with 170rpm stirring 24 hours, immediately after will
Active carbon is transferred in 200ml 1% (w/v) glutaraldehyde solution and is crosslinked, and is finally washed with deionized to obtain modified active
Charcoal；
(3) LB liquid medium is configured, shaking flask culture micrococcus luteus to logarithmic growth phase takes OD600The bacterium solution of value about 1.5,
It is collected by centrifugation, is configured to 10ml bacterial suspension with sterile water；
(4) 3g sodium alginate and 0.2g modified activated carbon are weighed, 90ml is added and distills aqueous systems, sterilizing will be in step (2)
10ml bacterial suspension is mixed evenly with it, is uniformly instilled in 4%w/v calcium chloride solution, is stood four hours at room temperature, will be washed
Bead afterwards is poured into containing in 120mg/L Cr (VI) LB liquid medium, and condition of culture is initial pH 8.0,30 DEG C of temperature, is shaken
Bed revolving speed 170rpm, cultivates 84h；
Conclusion: as shown in Fig. 2, the hexavalent chrome reduction rate of the modified activated carbon of addition 0.2g is 73.3%, modification is not added
The hexavalent chrome reduction rate of active carbon is 64.1%, and hexavalent chrome reduction rate improves 9.2%.
Embodiment 3
The present embodiment the method mainly sequentially includes the following steps:
(1) active carbon is broken into particle size is 200 mesh or more；
(2) 5g active carbon is added to containing 50ml 70%HNO3Flask in, and at room temperature with 170rpm stir 1h
To remove the impurity in active carbon, it is washed with deionized water to neutrality, it is last sufficiently dry；Active carbon after drying is added
In 100ml 10% (w/v) PEI (polyetherimide)/methanol solution, at room temperature with 170rpm stirring 24 hours, immediately after will
Active carbon is transferred in 200ml 1% (w/v) glutaraldehyde solution and is crosslinked, and is finally washed with deionized to obtain modified active
Charcoal；
(3) LB liquid medium is configured, shaking flask culture micrococcus luteus to logarithmic growth phase takes OD600The bacterium solution of value about 1.5,
It is collected by centrifugation, is configured to 10ml bacterial suspension with sterile water；
(4) 3g sodium alginate and 0.3g modified activated carbon are weighed, 90ml is added and distills aqueous systems, sterilizing will be in step (2)
10ml bacterial suspension is mixed evenly with it, is uniformly instilled in 4%w/v calcium chloride solution, is stood four hours at room temperature, will be washed
Bead afterwards is poured into containing in 120mg/L Cr (VI) LB liquid medium, and condition of culture is initial pH 8.0,30 DEG C of temperature, is shaken
Bed revolving speed 170rpm, cultivates 84h；
Conclusion: as shown in figure 3, the hexavalent chrome reduction rate of the modified activated carbon of addition 0.3g is 77.5%, modification is not added
The hexavalent chrome reduction rate of active carbon is 64.1%, and hexavalent chrome reduction rate improves 13.4%.
Embodiment 4
The present embodiment the method mainly sequentially includes the following steps:
(1) active carbon is broken into particle size is 200 mesh or more；
(2) 5g active carbon is added to containing 50ml 70%HNO3Flask in, and at room temperature with 170rpm stir 1h
To remove the impurity in active carbon, it is washed with deionized water to neutrality, it is last sufficiently dry；Active carbon after drying is added
In 100ml 10% (w/v) PEI (polyetherimide)/methanol solution, at room temperature with 170rpm stirring 24 hours, immediately after will
Active carbon is transferred in 200ml 1% (w/v) glutaraldehyde solution and is crosslinked, and is finally washed with deionized to obtain modified active
Charcoal；
(3) LB liquid medium is configured, shaking flask culture micrococcus luteus to logarithmic growth phase takes the bacterium that OD600 value is about 1.5
Liquid is collected by centrifugation, and is configured to 10ml bacterial suspension with sterile water；
(4) 3g sodium alginate and 0.4g modified activated carbon are weighed, 90ml is added and distills aqueous systems, sterilizing will be in step (2)
10ml bacterial suspension is mixed evenly with it, is uniformly instilled in 4%w/v calcium chloride solution, is stood four hours at room temperature, will be washed
Bead afterwards is poured into containing in 120mg/L Cr (VI) LB liquid medium, and condition of culture is initial pH 8.0,30 DEG C of temperature, is shaken
Bed revolving speed 170rpm, cultivates 84h；
Conclusion: as shown in figure 4, the hexavalent chrome reduction rate of the modified activated carbon of addition 0.4g is 75.83%, modification is not added
The hexavalent chrome reduction rate of active carbon is 64.1%, and hexavalent chrome reduction rate improves 11.73%.
Embodiment 5
The present embodiment the method mainly sequentially includes the following steps:
(1) active carbon is broken into particle size is 200 mesh or more；
(2) 5g active carbon is added to containing 50ml 70%HNO3Flask in, and at room temperature with 170rpm stir 1h
To remove the impurity in active carbon, it is washed with deionized water to neutrality, it is last sufficiently dry；Active carbon after drying is added
In 100ml 10% (w/v) PEI (polyetherimide)/methanol solution, at room temperature with 170rpm stirring 24 hours, immediately after will
Active carbon is transferred in 200ml 1% (w/v) glutaraldehyde solution and is crosslinked, and is finally washed with deionized to obtain modified active
Charcoal；
(3) LB liquid medium is configured, shaking flask culture micrococcus luteus to logarithmic growth phase takes OD600The bacterium solution of value about 1.5,
It is collected by centrifugation, is configured to 10ml bacterial suspension with sterile water；
(4) 3g sodium alginate and 0.5g modified activated carbon are weighed, 90ml is added and distills aqueous systems, sterilizing will be in step (2)
10ml bacterial suspension is mixed evenly with it, is uniformly instilled in 4%w/v calcium chloride solution, is stood four hours at room temperature, will be washed
Bead afterwards is poured into containing in 120mg/L Cr (VI) LB liquid medium, and condition of culture is initial pH 8.0,30 DEG C of temperature, is shaken
Bed revolving speed 170rpm, cultivates 84h；
Conclusion: as shown in figure 5, the hexavalent chrome reduction rate of the modified activated carbon of addition 0.5g is 75%, modified work is not added
Property charcoal hexavalent chrome reduction rate be 64.1%, hexavalent chrome reduction rate improves 10.9%.
Claims (4)
1. a kind of method of modified activated carbon immobilized cell enhancing bacterium chromium reducing power, it is characterised in that including following step
It is rapid:
(1) active carbon is broken into granule size is 200 mesh or more；
(2) 5g active carbon is added to containing 50ml 70%HNO3Flask in, and room temperature with 170rpm stir 1h, removal live
Property charcoal in impurity, be washed with deionized water to neutrality, it is last sufficiently dry；100ml 10%w/ is added in active carbon after drying
In the methanol solution of v PEI, at room temperature with 170rpm stirring 24 hours, active carbon is transferred to 200ml 1%w/v immediately after
Glutaraldehyde solution in be crosslinked, be finally washed with deionized to obtain modified activated carbon；
(3) bacterium source is separated in laboratory screening, and Selective agar medium is improvement LB liquid medium, and formula is prepared: peptone
10g·L-1, sodium chloride 5gL-1, yeast extract 5gL-1, dipotassium hydrogen phosphate 0.05gL-1, epsom salt 0.2gL-1, if solid LB media, then plus agar 1.5~2.0%, pH 8.5~9.0 sterilize；
(4) isolated micrococcus luteus will be screened containing shaking flask culture is carried out in LB culture medium, condition of culture is initial pH
8.0-9.0,30 DEG C of temperature, shaking speed 170rpm；
(5) condition of immobilized spherule are as follows: sodium alginate 3g, calcium chloride 4g, modified activated carbon 0.1-0.5g, by sodium alginate and
Modified activated carbon is dissolved in 90ml distilled water, sterilizing；
(6) cultured micrococcus luteus in step (4) is filtered, is collected by centrifugation, and press inoculum concentration OD600Value is 1.5, with nothing
Bacterium water is made into 10ml and is uniformly mixed with mixed liquor in step (4), and being slowly dropped into mass concentration is to hand in 4%w/v calcium chloride solution
Connection was added and contains in 120mg/L Cr (VI) LB liquid medium after four hours, room temperature shaker culture 84h；
(7) diphenylcarbazide solution is configured, 0.2g diphenylcarbazide is dissolved in 50ml acetone, moves to 100ml volumetric flask, add
Water is diluted to graduation mark, shakes up, and sequentially adds the dense H of 12.5ml slowly into refrigerant liquid2SO4With 12.5ml concentrated phosphoric acid, it is protected from light guarantor
It deposits.
2. the method for modified activated carbon immobilized cell enhancing bacterium chromium reducing power according to claim 1, feature
It is that step (3) is described: sample 10g is taken to be added in the sterilized liquid LB culture medium of 90ml improvement, is carried out with 10 times of gradients dilute
It releases, respectively draws 0.2ml, be coated on the solid improvement LB culture medium of Cr containing 200mg/L (VI), smoothened bacterium solution with spreading rod,
It cultivates 24 hours in 30 DEG C of constant incubators and is grown to bacterium colony, identify isolated bacterial strain, be micrococcus luteus.
3. the method for modified activated carbon immobilized cell enhancing bacterium chromium reducing power according to claim 1 or 2, special
Sign is: the material of immobilized spherule is sodium alginate, calcium chloride, modified activated carbon, physiological saline, sterile water and bacterium solution.
4. the method for modified activated carbon immobilized cell enhancing bacterium chromium reducing power according to claim 4, feature
It is to diphenylcarbazide solution obtained by step (7), using Cr VI in diphenyl carbazide spectrophotometry measurement sample
Concentration, the calculation formula of Cr (VI) concentration is y=0.6031x+0.0065, the coefficient R of standard curve2=0.0997.
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| CN113149230A (en) * | 2021-02-26 | 2021-07-23 | 中南大学 | Treatment method for reducing hexavalent chromium by wood charcoal electron transfer enhanced microorganisms |
| CN114686377A (en) * | 2020-12-31 | 2022-07-01 | 中国石油化工股份有限公司 | Preparation and preservation method of nitrifying bacteria agent |
| CN115286119A (en) * | 2022-07-20 | 2022-11-04 | 西南科技大学 | Microorganism strengthening agent using mineral/biomass as carrier for removing hexavalent chromium and preparation method thereof |
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