CN111115841B - Method for reinforcing coupling and passivating nano zero-valent iron system by iron reducing bacteria - Google Patents
Method for reinforcing coupling and passivating nano zero-valent iron system by iron reducing bacteria Download PDFInfo
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- CN111115841B CN111115841B CN201911299838.4A CN201911299838A CN111115841B CN 111115841 B CN111115841 B CN 111115841B CN 201911299838 A CN201911299838 A CN 201911299838A CN 111115841 B CN111115841 B CN 111115841B
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Abstract
The invention relates to a method for coupling and passivating a nano zero-valent iron system by using reinforced iron reducing bacteria, which specifically comprises the following steps: step 1, completing preparation of the graphene oxide loaded nano zero-valent iron, wherein: preparing graphene oxide by adopting a Hummers method, adding 200mg of prepared graphene oxide into a 250mL three-neck flask, dissolving 0.496g of ferrous sulfate heptahydrate into a mixed solution of 60mL of water and 20mL of ethanol, adding the mixed solution into the three-neck flask, and carrying out ultrasonic treatment on the mixed solution for 30min at 50kHz on an ultrasonic instrument; 2, degrading 4-chlorophenol by using graphene oxide loaded nano zero-valent iron; step 3, performing artificial culture of Shewanella; step 4, finishing the degradation of 4-chlorophenol by reducing and passivating nano zero-valent iron by graphene oxide mediated Shewanella; the graphene oxide loaded passivated nano zero-valent iron coupled Shewanella is used for strengthening a coupling system, reducing the formation of a passivation layer, improving the reaction activity of the system and overcoming the defect that nano zero-valent iron is easy to inactivate through the coupling effect of iron reducing bacteria Shewanella and materials.
Description
Technical Field
The invention relates to the technical field of material coupling bacteria for enhancing reaction activity, in particular to preparation of a graphene oxide loaded nano zero-valent iron composite material and construction of a graphene oxide mediated reinforced Shewanella coupling passivated nano zero-valent iron system method.
Background
Zero-valent iron has attracted much attention as an effective dehalogenation reducing agent since the 80's in the 20 th century. In recent years, it has been found that nano zero-valent iron, which is a specific form of zero-valent iron, has the characteristics of small particles and large specific surface area, and can perform reductive dehalogenation on a plurality of organic halides, including Trichloroethylene (TCE), 4-chlorophenol, pentachlorophenol (PCP), tetrabromobisphenol a (tbbpa), Hexabromocyclododecane (HBCD), and the like.
However, the nano zero-valent iron has high surface activity and is easy to oxidize, an oxide film is generated on the surface of the nano zero-valent iron, the main component of the nano zero-valent iron is ferric oxide or ferroferric oxide, and a small amount of other iron oxides such as green rust and the like hinder electron transfer, so that the reaction activity is reduced. Once the traditional chelating agent such as EDTA (ethylenediaminetetraacetic acid) and NTA (nitrilotriacetic acid) is chelated with the nano zero-valent iron, the secondary pollution problem in the environment can be caused after the chelating agent is widely used, and experts and scholars also propose to add the dissimilatory iron reducing bacteria into the nano zero-valent iron reactor to solve the surface oxidation film of the nano zero-valent iron on the premise that the dissimilatory iron reducing bacteria can reduce iron oxide under the anaerobic condition. The dissimilatory iron reducing bacteria take Fe (III) oxide on the surface of nano zero-valent iron as a final electron acceptor to perform anaerobic respiration, and form secondary minerals (goethite, hematite, magnetite and the like) through biological reduction.
However, since the dissimilatory iron reducing bacteria and the nano zero-valent iron are not directly contacted through the mediator in the coupling process, fine particles of the nano zero-valent iron can rapidly agglomerate and increase the particle size, and the reaction activity can be reduced, so that the electron transfer between electrons and pollutants is hindered. When the iron oxide and the iron reducing bacteria can not be in direct contact, electrons generated on the respiratory chain of the iron reducing bacteria can not be transferred to the surface of the iron oxide, so that the reduction of the iron oxide is realized.
Disclosure of Invention
The invention aims to solve the problems that nano zero-valent iron is easy to agglomerate and oxidize and a technical method for coupling reinforced iron reducing bacteria and passivated nano zero-valent iron. The graphene oxide mediated Shewanella is researched to passivate the surface of the passivated nano zero-valent iron so as to degrade and remove the persistent organic pollutants.
The technical scheme of the invention provides a method for reinforcing coupling and passivating a nano zero-valent iron system by using iron reducing bacteria, which specifically comprises the following steps:
Wherein: preparing graphene oxide by adopting a Hummers method, adding 200mg of prepared graphene oxide into a 250mL three-neck flask, dissolving 0.496g of ferrous sulfate heptahydrate into a mixed solution of 60mL of water and 20mL of ethanol, adding the mixed solution into the three-neck flask, and carrying out ultrasonic treatment on the mixed solution for 30min at 50kHz on an ultrasonic instrument;
under the conditions of stirring of a mechanical stirrer and nitrogen protection, 25mL of sodium borohydride solution is dropwise added into a three-neck flask by using a peristaltic pump, and the reaction time is 30 min; after the reaction, continuously introducing nitrogen and stirring for 20min to ensure that the mixed solution completely reacts; centrifuging at 5000rpm for 5min, collecting the product, sequentially washing with deionized water and anhydrous ethanol for 3 times, and drying in a vacuum drying oven at 60 ℃ to obtain graphene oxide loaded nano zero-valent iron;
Carrying out a reaction of graphene oxide loaded nano zero-valent iron and 4-chlorophenol, and carrying out a degradation batch experiment reaction in a 100mL serum bottle sealed by a butyronitrile eversion rubber plug in a dark place, wherein the initial concentration of the 4-chlorophenol is 10 mg/L; after the reaction is finished, centrifugally collecting a passivated sample, drying the passivated sample in a vacuum drying oven at 60 ℃, storing the dried sample in a 10mL PE tube for later use, and obtaining a final sample which is oxidized graphene loaded passivated nano zero-valent iron;
Culturing Shewanella in LB liquid culture medium, performing shake culture at 30 deg.C and 200rpm, centrifuging for 16h at late logarithmic growth stage, collecting bacterial suspension, and washing with inorganic salt culture solution for 3 times to obtain final enriched bacterial liquid;
The experiment of degrading 4-chlorophenol by reducing and passivating nano zero-valent iron through graphene oxide mediated Shewanella is carried out in a 100mL serum bottle sealed by nitrile rubber in a dark place, and the concentration of 4-chlorophenol is controlled to be 20 mu g/L;
adding 150mg of graphene oxide loaded passivated nano zero-valent iron into a serum bottle, taking 20mM sodium lactate as an electron donor, adding 3mL of enriched Shewanella bacteria, taking 20mM HEPES as a biological buffer to keep the pH value within the range of 6.9-7.1, filling the residual volume with an inorganic salt culture medium, and taking the graphene oxide, the passivated nano zero-valent iron and the Shewanella bacteria together as a coupling system; sodium lactate as the only carbon source, and the concentration of the bacterial suspension is controlled at OD600＝1.2。
Further, in the step 3, the rotating speed of the low-temperature centrifuge is 8000rpm, the temperature is 4 ℃, and the working time of the low-temperature centrifuge is 5 min.
Further, in the step 4, the serum bottle is sealed by nitrile rubber and then placed on a constant temperature shaking table at the temperature of 30 ℃ for reaction at the speed of 200 r/min.
The invention has the beneficial effects that:
(1) the graphene oxide loaded nano zero-valent iron prepared by the method overcomes the defect that nano zero-valent iron is easy to agglomerate through the large specific surface area and pores of graphene oxide.
(2) The graphene oxide loaded passivated nano zero-valent iron coupled Shewanella prepared by the invention strengthens a coupling system, reduces the formation of a passivation layer, improves the reaction activity of the system and overcomes the defect that nano zero-valent iron is easy to inactivate through the coupling effect of iron reducing bacteria Shewanella and materials.
(3) The iron reducing bacteria related by the invention are wide in distribution, and the coupling method is simple and easy to implement.
Brief description of the drawings
FIG. 1 is a Scanning Electron Microscope (SEM) image of the graphene oxide loaded with nano zero-valent iron prepared by the invention
FIG. 2 is a graph showing the effect of graphene oxide loaded nano zero-valent iron on degradation of 4-chlorophenol
FIG. 3 is a graph showing the effect of graphene oxide mediated Shewanella reduction passivation nano zero-valent iron degradation of 4-chlorophenol
FIG. 4 is an XRD representation diagram of graphene oxide loaded passivated nano zero-valent iron, passivated zero-valent iron reduced by Shewanella and graphene oxide loaded passivated zero-valent iron reduced by Shewanella
FIG. 5 is a Scanning Electron Microscope (SEM) image of collected graphene oxide loaded passivated nano zero-valent iron coupled Shewanella after degradation reaction.
Detailed Description
The technical means of the present invention will be described in detail with reference to the following embodiments.
The embodiment provides a method for coupling and passivating a nano zero-valent iron system by using reinforced iron reducing bacteria, which specifically comprises the following steps:
Preparing graphene oxide by adopting a Hummers method, adding 200mg of prepared graphene oxide into a 250mL three-neck flask, dissolving 0.496g of ferrous sulfate heptahydrate into a mixed solution of 60mL of water and 20mL of ethanol, adding the mixed solution into the three-neck flask, and carrying out ultrasonic treatment on the mixed solution for 30min at 50kHz on an ultrasonic instrument;
under the stirring of a mechanical stirrer and the protection of nitrogen, 25mL of sodium borohydride solution is added dropwise into a three-neck flask by using a peristaltic pump, and the reaction time is 30 min. After the reaction, nitrogen gas was continuously introduced and stirred for 20min to complete the reaction of the mixed solution. Centrifuging at 5000rpm for 5min to collect product, washing with deionized water and anhydrous ethanol for 3 times, and oven drying at 60 deg.C in vacuum drying oven.
The collected sample is freshly prepared graphene oxide loaded with nano zero-valent iron.
FIG. 1 is a Scanning Electron Microscope (SEM) representation of graphene oxide loaded with nanoscale zero-valent iron, and the representation shows that graphene oxide has a rugate and pore structure as a mediating carrier, and Fe is present on the surface of graphene oxide due to negative charges2+Can be easily adsorbed on the surface or form a complex with the surface oxygen-containing functional group. With soluble Fe2The nano zero-valent iron is formed on the surface layer of the graphene oxide, wherein the agglomeration of the nano zero-valent iron is prevented by the low mobility attached to the surface of the graphene oxide.
And (3) reacting the prepared graphene oxide loaded nano zero-valent iron with 4-chlorophenol after the reaction in the step (1). The degradation batch experiment reaction is carried out in a 100mL serum bottle sealed by a butyronitrile rubber eversion plug in a dark place, and the initial concentration of 4-chlorophenol is 10 mg/L. And after the reaction is finished, centrifugally collecting a passivated sample, drying the passivated sample in a vacuum drying oven at 60 ℃, storing the dried sample in a 10mL PE tube for later use, and obtaining the final sample which is the graphene oxide loaded passivated nano zero-valent iron.
FIG. 2 is a graph showing the effect of graphene oxide loaded nano zero-valent iron on degradation of 4-chlorophenol, and shows that the degradation reaction is substantially balanced in about 60 hours. In the reaction time, the degradation efficiency of the nano zero-valent iron to the 4-chlorophenol is only 26.2%, the surface of the nano zero-valent iron is gradually oxidized to form a passivation layer in the degradation reaction process, so that the reaction is prevented from further proceeding, the graphene oxide loaded with the nano zero-valent iron improves the degradation effect of the nano zero-valent iron to the 4-chlorophenol, and the degradation efficiency reaches 37.8%.
Shewanella (Shewanella putrefeaciens) in this example was artificially cultured. Shewanella is cultured in LB liquid culture medium, shaking culture is carried out at 30 ℃ and 200rpm, bacterial suspension is collected by centrifugation at 16h logarithmic growth end stage (low temperature centrifuge 8000rpm, 4 ℃, 5min), and washing is carried out for 3 times by using inorganic salt culture solution (pH is 6.9-7.1) to obtain final enriched bacterial liquid.
The experiment of degrading 4-chlorophenol by reducing and passivating nano zero-valent iron through graphene oxide mediated Shewanella is carried out in a 100mL serum bottle sealed by nitrile rubber in a dark place, and the concentration of 4-chlorophenol is controlled to be 20 mu g/L. Adding 150mg of graphene oxide loaded passivated nano zero-valent iron into a serum bottle, taking 20mM sodium lactate as an electron donor, adding 3mL of enriched Shewanella bacteria, taking 20mM HEPES as a biological buffer to keep the pH value within the range of 6.9-7.1, filling the residual volume with an inorganic salt culture medium, filling the graphene oxide and passivated nano zero-valent iron into the serum bottle, and filling the rest volume with the graphene oxide and passivated nano zero-valent ironThe valence iron and the Shewanella are combined into a coupling system. Sodium lactate as the only carbon source, and the concentration of the bacterial suspension is controlled at OD6001.2. The serum bottle is sealed by nitrile rubber and then placed on a constant temperature shaking table at the temperature of 30 ℃ for reaction at the speed of 200 r/min. The control groups are three groups, namely a blank group added with 4-chlorophenol independently, Shewanella bacteria liquid subjected to high-temperature sterilization and single passivated nano zero-valent iron (not loaded with graphene oxide, the addition amount is 50 mg).
FIG. 3 is a Scanning Electron Microscope (SEM) image of collected graphene oxide loaded passivated nano zero-valent iron coupled Shewanella after degradation reaction. Characterization of shewanella morphology by scanning electron microscopy required an overnight pretreatment by addition of 2.5% glutaraldehyde for fixation. The graphene oxide is used as a mediator, and is subjected to reaction with nano zero-valent iron, then the nano zero-valent iron with the original spherical surface is passivated, and then is subjected to biological reduction by Shewanella to form a secondary mineral, and the graphene oxide is a needle-like material, namely the secondary mineral containing Fe (II).
The results show that: the graphene oxide loaded nano zero-valent iron can perform a good coupling effect with Shewanella, the pore structure of the graphene oxide is obvious, and Shewanella is convenient to adhere to the surface to react with the passivated nano zero-valent iron. Fig. 4 is an XRD characterization diagram, where the nanometer zero-valent iron is passivated by reduction of shewanella, and coupled with shewanella after the nanometer zero-valent iron is loaded with graphene oxide, green embroidery and cyanite are generated, and it can be seen from fig. 4 that the passivated nanometer zero-valent iron is converted into cyanite after reduction of shewanella under the mediation of graphene oxide, and characteristic peaks thereof mainly appear at 11.83 ° and 23.66 °, and the cyanite contains fe (ii) in a binding state, and has a certain degradation effect on chlorinated organic pollutants. FIG. 5 is a graph showing the effect of degrading 4-chlorophenol by reducing and passivating nano zero-valent iron with graphene oxide mediated Shewanella, and shows that the degradation efficiency of 4-chlorophenol reaches 47.3% after 12 days of reaction without graphene oxide; in a system with graphene oxide, the degradation effect of 4-chlorophenol is remarkably improved, and the final degradation efficiency is 89.5 percent
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 claims and their equivalents.
Claims (1)
1. A method for reinforcing coupling and passivating a nano zero-valent iron system by iron reducing bacteria specifically comprises the following steps:
step 1, completing preparation of graphene oxide loaded nano zero-valent iron
Wherein: preparing graphene oxide by adopting a Hummers method, adding 200mg of prepared graphene oxide into a 250mL three-neck flask, dissolving 0.496g of ferrous sulfate heptahydrate into a mixed solution of 60mL of water and 20mL of ethanol, adding the mixed solution into the three-neck flask, and carrying out ultrasonic treatment on the mixed solution for 30min at 50kHz on an ultrasonic instrument;
under the conditions of stirring of a mechanical stirrer and nitrogen protection, 25mL of sodium borohydride solution is dropwise added into a three-neck flask by using a peristaltic pump, and the reaction time is 30 min; after the reaction, continuously introducing nitrogen and stirring for 20min to ensure that the mixed solution completely reacts; centrifuging at 5000rpm for 5min, collecting the product, sequentially washing with deionized water and anhydrous ethanol for 3 times, and drying in a vacuum drying oven at 60 ℃ to obtain graphene oxide loaded nano zero-valent iron;
the surface of the graphene oxide has negative charges, Fe2+Can be easily adsorbed on the surface or form a complex with the surface oxygen-containing functional group; with soluble Fe2Forming nano zero-valent iron on the surface layer of the graphene oxide, wherein the agglomeration of the nano zero-valent iron is prevented by the low mobility attached to the surface of the graphene oxide;
step 2, degrading 4-chlorophenol by using graphene oxide loaded nano zero-valent iron
Carrying out a reaction of graphene oxide loaded nano zero-valent iron and 4-chlorophenol, and carrying out a degradation batch experiment reaction in a 100mL serum bottle sealed by a butyronitrile eversion rubber plug in a dark place, wherein the initial concentration of the 4-chlorophenol is 10 mg/L; after the reaction is finished, centrifugally collecting a passivated sample, drying the passivated sample in a vacuum drying oven at 60 ℃, storing the dried sample in a 10mL PE tube for later use, and obtaining a final sample which is oxidized graphene loaded passivated nano zero-valent iron;
step 3, performing artificial culture of Shewanella
Culturing Shewanella in LB liquid culture medium, performing shake culture at 30 deg.C and 200rpm, centrifuging for 16h at late logarithmic growth stage, collecting bacterial suspension, and washing with inorganic salt culture solution for 3 times to obtain final enriched bacterial liquid; the rotation speed of the low-temperature centrifuge is 8000rpm, the temperature is 4 ℃, and the working time of the low-temperature centrifuge is 5 min;
step 4, finishing the degradation of 4-chlorophenol by reducing and passivating nano zero-valent iron by graphene oxide mediated Shewanella
The experiment of degrading 4-chlorophenol by reducing and passivating nano zero-valent iron through graphene oxide mediated Shewanella is carried out in a 100mL serum bottle sealed by nitrile rubber in a dark place, and the concentration of 4-chlorophenol is controlled to be 20 mu g/L;
adding 150mg of graphene oxide loaded passivated nano zero-valent iron into a serum bottle, taking 20mM sodium lactate as an electron donor, adding 3mL of enriched Shewanella bacteria, taking 20mM HEPES as a biological buffer to keep the pH value within the range of 6.9-7.1, filling the residual volume with an inorganic salt culture medium, and taking the graphene oxide, the passivated nano zero-valent iron and the Shewanella bacteria together as a coupling system; sodium lactate as the only carbon source, and the concentration of the bacterial suspension is controlled at OD600= 1.2; sealing the serum bottle with nitrile rubber, and reacting on a constant temperature shaking table at 30 ℃ at 200 r/min; the control groups are three groups, namely a blank group which is independently added with 4-chlorophenol, Shewanella bacteria liquid which is sterilized at high temperature and independent passivated nano zero-valent iron.
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