A self-activation fenton-like composite repair material and a preparation method and application thereof

Abstract

The application relates to a Fenton-like composite material, in particular to a self-activated Fenton-like composite repair material and a preparation method and application thereof, the composite repair material is CaO2@Fe3O4@BIP-BC, a protein imprinting biochar is used as a matrix, magnetite serving as an activator and calcium peroxide serving as a slow-release oxidant are sequentially loaded on the surface. Compared with the prior art, the application has the advantages that (1) the biochar surface active groups and calcium peroxide improve the high efficiency and long-acting property of the Fenton-like process, (2) the protein imprinting improves the selectivity of the removal of target harmful bacteria and harmful bacteria genes, (3) the biochar simultaneously loads the calcium peroxide and the magnetite, the simultaneous addition of the activator and the oxidant is realized, and the use cost and the operation complexity are reduced. The method solves the problems that the traditional Fenton-like oxidation treatment has poor specific removal effect on harmful bacteria and harmful bacteria genes in the environment, the treatment time is short, the required dosage is high, secondary pollution is serious, and the activator and the oxidant need to be used separately.

Description

TECHNICAL FIELD

[0001] The application relates to a Fenton-like composite material, in particular to a self-activated Fenton-like composite repair material and a preparation method and application thereof. BACKGROUND

[0002] With the large use and improper disposal of antibiotics and other antibacterial drugs, a large amount of harmful bacteria and harmful bacteria genes such as antibiotic resistance bacteria (ARBs) and antibiotic resistance genes (ARGs) are generated in the environment, which seriously endanger people's health. At the same time, harmful bacteria and harmful bacteria genes are widely spread in the environment through sewage or other waste, which also threatens the ecological safety of soil and water. As a new type of pollutant, harmful bacteria and harmful bacteria genes have the characteristics of persistence, easy spread and easy outbreak. The current common physical, biological and chemical treatment technologies cannot effectively control the spread and diffusion of harmful bacteria genes. Therefore, it is of great significance to develop new high-efficiency removal materials and technologies to realize the remediation of soil and water contaminated by harmful bacteria and harmful bacteria genes and to protect the ecological safety of water and soil environment.

[0003] Fenton-like oxidation treatment is a mature soil and groundwater organic pollutant treatment technology, which is also gradually used for the removal of harmful bacteria and harmful bacteria genes in water and soil environment. Common activators in Fenton-like oxidation treatment include divalent iron, trivalent iron, zero-valent iron, magnetite and the like, and common oxidants include hydrogen peroxide, persulfate, periodate and the like. For example, a study uses pyrite (FeS2) to activate periodate (PI) to disinfect ARB, and the results show that the FeS2 / PI system can disinfect 1x10 7 CFU mL -1 Kanamycin-resistant E. coli is disinfected to below the detection limit. However, the traditional Fenton-like oxidation treatment method often has problems such as poor reactivity, low utilization rate, and Fe(III) / Fe(II) cycle limitation in actual application. At the same time, the oxidant and activator of the traditional Fenton-like method need to be added separately, which increases the operation complexity of the method, and a self-activated Fenton-like material needs to be developed to reduce the operation difficulty of Fenton-like. More importantly, the oxidation of the current Fenton-like material has broad-spectrum, which not only removes harmful bacteria and harmful bacteria genes, but also causes inactivation of beneficial bacteria in the environment, which can be solved by improving the specific removal effect of the material on harmful bacteria and harmful bacteria genes. In order to solve the above problems, it is necessary to develop a self-activated Fenton-like composite repair material which has high efficiency and long-term effect and can selectively remove harmful bacteria and harmful bacteria genes in water and soil environment. SUMMARY

[0004] The present application aims to solve at least one of the above problems by providing a self-activated Fenton-like composite repair material, a preparation method and application thereof. The present application has the following advantages: (1) the biochar surface active groups and calcium peroxide improve the efficiency and long-acting of Fenton-like; (2) the protein imprinting improves the selectivity of removing target harmful bacteria and harmful bacteria genes; (3) the biochar as a carrier simultaneously loads calcium peroxide and magnetite, realizes the simultaneous addition of activator and oxidant, and reduces the use cost and operation complexity. By using the above technical innovation, the problems of poor selectivity of removing harmful bacteria and harmful bacteria genes, short treatment time, high required dosage, serious secondary pollution, and the need to use activator and oxidant separately in the traditional Fenton-like oxidation treatment method are solved. The present application realizes the efficient and long-acting repair of soil and water bodies polluted by target harmful bacteria and harmful bacteria genes through Fenton-like reaction. The biochar, oxidized iron and oxidized calcium and other substances in the material after the reaction will not have a great impact on the properties of soil and water bodies even if they remain in the environment, and have good ecological and environmental safety.

[0005] The present inventors have found in years of research that calcium peroxide is more stable than hydrogen peroxide and is difficult to dissolve in water, has a certain slow-release effect, and is a good long-acting oxidant. Magnetite is common in the natural environment and contains Fe 2+ , Fe 3+ , has strong oxidant activation performance, but is prone to agglomeration when alone, which limits the effect. Biochar has a good pore structure and is an excellent carrier. The surface of biochar contains a large number of redox functional groups, which is conducive to the Fe 2+ / Fe 3+ cycle and promotes the efficiency of Fenton reaction. Protein imprinting technology has good biocompatibility and adsorption selectivity for target bacteria. After protein imprinting modification of biochar, the selectivity of biochar for target harmful bacteria is improved, which is conducive to improving the specific oxidation removal effect of the self-activated Fenton-like material based on biochar carrier on harmful bacteria and harmful bacteria genes contained therein. Therefore, it has strong feasibility and potential application value to prepare a self-catalytic Fenton-like composite material using protein imprinting biochar as a carrier and calcium peroxide as a slow-release oxidant to realize the selective and efficient and long-acting removal of harmful bacteria genes in water and soil environments.

[0006] Based on the above theoretical analysis, the purpose of the present application is achieved by the following technical solutions:

[0007] The present application discloses a self-activated Fenton-like composite repair material. The composite repair material is CaO2@Fe3O4@BIP-BC, which uses protein imprinting biochar as a substrate and sequentially loads magnetite as an activator and calcium peroxide as a slow-release oxidant on the surface.

[0008] The composite material first specifically adsorbs target harmful bacteria or harmful bacteria genes on the surface of the biochar through protein imprinting, and then slowly releases H2O2 in the material by the reaction of calcium peroxide with moisture in the environment, and Fe 2+ / Fe 3+ Fenton-like reaction occurs, thereby achieving target harmful bacteria inactivation and harmful bacteria gene degradation by using the generated active oxygen free radicals.

[0009] Preferably, in the CaO2@Fe3O4@BIP-BC, the mass ratio of Fe3O4 to BIP-BC is 12-1:6.

[0010] Preferably, the protein imprinting biochar is obtained by binding target harmful bacteria on the surface of biochar through protein imprinting technology.

[0011] Preferably, the particle size of the composite repair material is 0.2-2.5mm.

[0012] The second aspect of the present application discloses a method for preparing the self-activated Fenton-like composite repair material as described in any of the above, comprising the following steps:

[0013] S1: Preparation of biochar: The crushed biomass material is pyrolyzed by a limited oxygen slow heating method, ground and sieved after cooling, soaked in an HCl solution, and then washed to obtain biochar;

[0014] S2: Preparation of protein imprinting biochar: inoculate target harmful bacteria in a LB medium containing protease, centrifuge the precipitate after shaking culture, resuspend the precipitate in a lysis extraction buffer and heat, centrifuge the supernatant again, add DNase and RNase to the supernatant and mix evenly, stand in ice water to remove nucleic acid impurities, add acidified acetone-ethanol mixture to the supernatant and stand in ice water to precipitate proteins, centrifuge and collect the precipitate and wash, remove methanol by vacuum filtration, resuspend the precipitate in a sample solubilization buffer, add the biochar obtained in step S1 to the resuspension and mix and wash, and dry to obtain protein imprinting biochar BIP-BC;

[0015] S3: Preparation of Fe3O4@BIP-BC, which is carried out by co-precipitation, the specific steps being: prepare ferric chloride solution and ferrous chloride solution respectively, disperse the protein imprinting biochar obtained in step S2 in the ferric chloride solution, add ferrous chloride solution to it after a certain period of contact, then add an excess of ammonia water solution and continue stirring, separate and wash the precipitate, and freeze-dry to obtain Fe3O4@BIP-BC particles;

[0016] S4: Preparation of CaO2@Fe3O4@BIP-BC, which is prepared by a co-precipitation method, and the specific steps are as follows: a calcium chloride solution is prepared, the Fe3O4@BIP-BC particles obtained in step S3 are dispersed in the calcium chloride solution, then a neutralizing agent and a stabilizer are added and stirred uniformly, hydrogen peroxide is added dropwise into the mixture, the precipitate is separated and washed, and the composite repair material is obtained after freeze-drying.

[0017] Preferably, in step S1:

[0018] The biomass material includes straw, wood chips, fruit peels, and other residues and the like;

[0019] The pyrolysis process is as follows: preheating at 100℃ for 1h under a nitrogen atmosphere, then heating to 300-700℃ at a rate of 10℃ / min and pyrolyzing for 2h;

[0020] The sieving is performed using a 100-mesh screen;

[0021] The concentration of the HCl solution is 10-15wt%;

[0022] The soaking time is 12-24h.

[0023] Preferably, in step S2:

[0024] The target harmful bacteria include, but are not limited to, gram-negative bacteria such as Escherichia coli and gram-positive bacteria such as Staphylococcus aureus;

[0025] The LB medium containing protease is an LB medium with a trypsin concentration of 0.5-2.5%;

[0026] The shaking culture is a continuous shaking culture at 37℃ at a speed of 150-180rpm for 8-10h;

[0027] The lysis extraction buffer is a mixture of pH=7, 20mM HEPES, 2-5wt% sodium dodecyl sulfate, and protease inhibitors, and the precipitate is resuspended in the lysis extraction buffer at a wet cell weight of 10μL / mg;

[0028] The heating is at 95℃ for 3-5min;

[0029] The amount of DNase added is 1mg / mL, and the amount of RNase added is 0.5mg / mL;

[0030] The concentration of the acidified acetone-ethanol mixture is 1.5mM;

[0031] The sample solubilization buffer is a mixture of 8-10 M urea, 2-4 M thiazolinone, 2-4 wt% CHAPS, 1-2 wt% TCEP-HCl and 2-5% v / v carrier ampholyte 3-10 NL;

[0032] The biochar and the wet weight of the bacteria in the resuspension are added in equal weight.

[0033] Preferably, in step S3:

[0034] Ferric chloride: protein imprinting biochar: ferrous chloride = 0.324 g: 0.8 g: 0.126 g;

[0035] The aqueous ammonia solution is added in excess.

[0036] The contact time is 30-60 min, and the duration of stirring is 1 h.

[0037] Preferably, in step S4:

[0038] Calcium chloride: Fe3O4@BIP-BC particles: stabilizer: hydrogen peroxide = 1.078 g: 0.3 g: 5 mL: 5 mL;

[0039] The neutralizing agent is aqueous ammonia, and the stabilizer is PEG200.

[0040] The neutralizing agent is used to adjust the reaction environment to be alkaline.

[0041] The reaction occurring in step S4 is CaCl2+H2O2→CaO2+HCl.

[0042] The third aspect of the application discloses an application of the self-activated Fenton-like composite repair material as described in any of the above in removing harmful bacteria and harmful genes in the environment, especially for treating antibiotic-resistant bacteria and antibiotic resistance genes in soil and groundwater.

[0043] Preferably, the CaO2@Fe3O4@BIP-BC is mixed in water or water-containing soil, wherein the dosage of Fe3O4@BIP-BC is controlled to be 10-40 mg / L, and / or the concentration of CaO2 is 0.5-3 mM.

[0044] More specifically, when the composite repair material is used, the following can be used:

[0045] Deep soil in-situ injection-high pressure rotary jet method: insert the grouting pipe into the deepest contaminated soil layer, mix the self-activated Fenton-like composite remediation material with water at a certain solid-liquid ratio, and then inject it into the contaminated deep soil under the action of high-pressure grouting pump and compressed air along the rotary jet grouting pipe. At the same time, the grouting pipe with nozzle is gradually lifted at a certain interval to ensure that the material can cover all the pollution depth. The composite remediation material can fully contact with the contaminated soil due to the high pressure and large jet range of high pressure rotary jet, so as to achieve the purpose of in-situ removal of harmful bacteria and harmful genes in soil.

[0046] Surface soil in-situ plowing method: evenly spread the self-activated Fenton-like composite remediation material on the surface of the contaminated soil, and then plow the soil to mix the remediation material with the soil evenly. During the remediation process, the soil should have a certain moisture content to promote the contact between calcium peroxide and water to release hydrogen peroxide. Through the self-activated Fenton-like effect and physical adsorption effect of the material, the purpose of in-situ removal of harmful bacteria and harmful genes in the surface soil is achieved.

[0047] Gravity addition method: add the self-activated Fenton-like material at a certain proportion into the water pump suction pipe or the water suction horn of the water suction well by gravity. Use the water pump impeller to mix the reagent with the contaminated water uniformly, and achieve the effect of removing harmful bacteria and harmful genes in the water environment through the adsorption and self-activated Fenton-like effect of the material.

[0048] Pressure addition method: mix the self-activated Fenton-like reagent with water at a certain proportion, and then add the reagent into the water pipe or sewage pool by using the water pump or water jet. Then the reagent is mixed uniformly with the contaminated water in the water pipe or pool, and the effect of removing harmful bacteria and harmful genes in the water environment is achieved through the adsorption and self-activated Fenton-like effect of the material.

[0049] Among them,

[0050] In the deep soil in-situ injection-high pressure rotary jet method, the addition amount of the composite remediation material is 0.1-2.0% of the dry weight of the soil, the solid-liquid ratio of the material and water in the grouting slurry is 1:10-1:1000, the grouting pipe is inserted to the same depth as the maximum depth of the contaminated soil, the lifting interval of the grouting pipe is 0.5-2.5m, the rotary jet pressure is 0.5-2.5MPa, and the rotary jet angle is 120-350°.

[0051] In the surface soil in-situ plowing method, the spreading amount of the composite remediation material is 0.05-2.5% of the dry weight of the contaminated soil, the plowing depth of the soil is generally 15-50cm, the soil moisture content after spreading the composite remediation material is adjusted to 40-80% of the maximum field water holding capacity, and the treatment time of the remediation material is preferably 10-20d.

[0052] In the gravity dosing method and the pressure dosing method, the dosage of the medicament is 15-20 mg / L, and the hydraulic retention time is 2-3 h.

[0053] Compared with the prior art, the present application has the following beneficial effects:

[0054] In the common Fenton-like oxidation treatment method, the oxidant and the activator are added into the water or soil environment respectively, and there is no selectivity, and the bacteria in the environment are not selectively inactivated, and there are the following disadvantages: the contact effect of the activator and the oxidant is poor, the reaction efficiency is low, secondary pollution is prone to occur, the reaction time is short, and the reaction effect is difficult to control.

[0055] The self-activated Fenton-like material prepared in the present application is different from other Fenton-like oxidation treatment methods, and the material can utilize the selective adsorption capacity of the protein blotting biochar, the slow-release oxidation effect of calcium peroxide, and the activation effect of magnetite, and innovatively realize the purpose of self-catalytic slow release of active oxygen free radicals of the repair material, effectively improve the efficiency, long-acting property and operability of the Fenton-like advanced oxidation technology, and the selective adsorption of the target bacteria reduces the influence of the beneficial microorganisms in the environment in the repair process, and has the advantages of long action period, good treatment effect, stable structure, low cost, convenient use and environmental friendliness:

[0056] 1) The biochar contains rich oxygen-containing functional groups and π-π structures, so that it has strong electron transfer effect, can accelerate the circulation rate of Fe 3+ / Fe 2+ , and improve the inactivation effect of the Fenton-like reaction on harmful bacteria and the oxidation degradation effect on harmful bacteria genes.

[0057] 2) The repair material has Fenton-like long-acting property through the slow-release H2O2 effect of calcium peroxide, and the reaction time is relatively long, and there is no need for secondary dosing.

[0058] 3) The biochar is modified by the protein blotting method, so that it has selective adsorption effect on the target harmful bacteria, improves the removal selectivity of the target harmful bacteria and harmful bacteria genes in the water and soil environment, and reduces the killing effect of the Fenton-like oxidation on the beneficial bacteria in the water and soil environment.

[0059] 4) The calcium peroxide and magnetite are sequentially loaded on the protein blotting biochar to produce a self-activated Fenton-like reaction, so that the activator and the oxidant can be simultaneously dosed when the repair material is used, the problem that the activator and the oxidant need to be separately dosed in the traditional Fenton-like reaction is solved, and the use cost and the operation complexity of the material are reduced.

[0060] More specifically:

[0061] (1) The application takes biochar as a matrix, magnetite as an activator, and calcium peroxide as an H2O2 slow-release agent, uses a two-step co-precipitation method to first load magnetite on the surface of biochar, then adds polyethylene glycol PEG200 as a protective agent and ammonia to create an alkaline environment to neutralize the generated HCl, and through co-precipitation, loads CaO2 on the surface of the material, to prepare a self-activated Fenton-like composite repair material, thereby innovatively achieving the purpose of self-catalyzing the production of active free radicals. It can solve the problem that the oxidant and the activator need to be prepared and used separately, effectively improve the operability of the Fenton-like advanced oxidation technology, and reduce the difficulty of actual repair work.

[0062] (2) The common harmful bacteria and harmful gene removal method lacks selectivity, threatens the beneficial bacteria in the environment, and destroys the microbial diversity in the environment. The Western blotting technology realizes selective adsorption of target harmful bacteria by combining bacterial fragment substances with biochar, and reduces the oxidative damage to beneficial microorganisms in the environment.

[0063] (3) The Fenton-like oxidation treatment method for removing harmful bacteria and harmful genes has been applied, but is often limited by the Fe 2+ / Fe 3+ cycle step. The application uses biochar as a matrix to load magnetite for activating hydrogen peroxide, and the biochar not only has an activating effect on hydrogen peroxide in the reaction, but also is beneficial to the Fe 2+ / Fe 3+ cycle, thereby improving the Fenton reaction efficiency. The method is used for the control and removal of harmful bacteria and harmful genes in the environment, and has not been reported in the past pollution repair material research and development work. At the same time, the material preparation method is simple to operate, the raw material cost is low, there is no secondary pollution, and has strong practical application prospect.

[0064] (4) The current Fenton-like oxidation treatment method has a fast reaction rate, and if the harmful bacteria and harmful genes are to be effectively removed, the oxidant dosage is large and the reaction time is short, resulting in a high risk of revival and regeneration of harmful bacteria. Calcium peroxide slowly reacts with water in the environment to generate hydrogen peroxide, thereby creating a sustained oxidative environment, which can effectively remove harmful bacteria and reduce the risk of revival and regeneration.

[0065] (5) The Western blotting and two-step co-precipitation method used to prepare the Western blotting self-activated Fenton-like composite material are simple and feasible, the raw materials used are cheap and easy to obtain, environmentally friendly, and suitable for mass production, and have great application potential in the field of harmful bacteria and harmful gene contaminated soil and groundwater remediation. BRIEF DESCRIPTION OF DRAWINGS

[0066] Figure 1 The adsorption isotherm of Fe3O4@BIP-BC and Fe3O4@BC in Example 2;

[0067] Figure 2 Adsorption kinetics of Fe3O4@BIP-BC and Fe3O4@BC in Example 2;

[0068] Figure 3 is a competitive adsorption test in Example 2, wherein a) monoprotocol competitive adsorption of Fe3O4@BIP-BC and Fe3O4@BC, b) binary system competitive adsorption of Fe3O4@BIP-BC;

[0069] Figure 4 Inactivation effect of CaO2@Fe3O4@BIP-BC on harmful bacteria under different mass ratios of Fe3O4 to BIP-BC in Example 3;

[0070] Figure 5 Inactivation effect of CaO2@Fe3O4@BIP-BC on harmful bacteria under different CaO2 concentrations in Example 3;

[0071] Figure 6 Inactivation effect of CaO2@Fe3O4@BIP-BC on harmful bacteria under different Fe3O4@BIP-BC dosages in Example 3;

[0072] Figure 7 is the inactivation effect of CaO2@Fe3O4@BIP-BC on harmful bacteria under different water matrix conditions in Example 4, wherein a) different HA concentrations, b) different pH, c) different salinity. DETAILED DESCRIPTION

[0073] The present application will be described in detail below with reference to the accompanying drawings and specific examples.

[0074] In the following examples, if not specifically stated, the reagents used are conventional commercially available products, and the methods used are conventional means in the art.

[0075] In the following examples, the mass percentage or mass concentration is used.

[0076] Example 1

[0077] A preparation method of a self-activated Fenton-like composite repair material for selectively removing harmful bacteria and harmful genes, the specific steps are as follows:

[0078] (I) Raw material pretreatment and preparation of protein blotting biochar

[0079] (1) Preparation and pretreatment of biochar:

[0080] (i) The collected corn stalk biomass material was washed, cut into small pieces of 3-5 cm, dried at 75 °C, and ground into powder using a grinder. The dried biomass material was then loaded into a crucible, compacted, covered, and placed in a muffle furnace under a nitrogen atmosphere. The pyrolysis process was carried out using a limited oxygen slow heating method, first preheating at 100 °C for 1 h, then heating to 500 °C for 2 h. After cooling to room temperature, the material was ground in a mortar and sieved through a 100 mesh screen, then sealed in a brown jar for storage.

[0081] (ii) The obtained initial corn biochar powder was first soaked in an HC1 solution (10 wt%) for 12 h, and after standing, the supernatant was discarded. The powder was then washed with deionized water, and the sample was dried for later use.

[0082] (2) Preparation of protein blotting biochar: 100 mL of LB medium solution containing trypsin at a concentration of 0.5-2.5% was prepared according to the published proportion, E. coli HB101 bacteria were inoculated, and 50 ug / mL of streptomycin was added to the medium, then placed in a 37 °C, 180 rpm condition for overnight shaking culture. After the culture, the solution was centrifuged at 6000-8000 rpm for 5 min, and the supernatant was discarded. The precipitate was resuspended in lysis extraction buffer (pH = 7, 20 mM HEPES, 5 wt% sodium dodecyl sulfate (SDS), and a mixture of protease inhibitors) at a proportion of 10 μL / mg wet cell weight, and heated at 95 °C for 5 minutes. Centrifuged at 4000 rpm for 15 minutes at 4 °C, the supernatant was separated from the cell material. After adding DNase (1 mg / mL) and RNase (0.5 mg / mL) to the collected supernatant and mixing well with a vortex mixer, nucleic acid impurities were removed by standing in ice water for 15 minutes. Then 1.5 mM of acidified acetone-ethanol mixture was added to the supernatant, and the protein was precipitated by standing in ice water for 30 minutes. The precipitate was centrifuged at 10000 rpm for 20 minutes at 4 °C, washed with methanol three times, and resuspended in sample solubilization buffer (10 M urea, 4 M thiazolinone, 4 wt% CHAPS, 2 wt% TCEP-HCl, and 5% v / v carrier ampholyte 3-10 NL) by rapid vacuum treatment to remove methanol. The resuspended solution was added to the biochar obtained in step S1, and then the mixture was mixed at 180 rpm for 16 h, washed with normal saline three times, and finally dried to obtain protein blotting biochar BIP-BC for later use.

[0083] (B) Preparation of self-activated Fenton-like composite repair material

[0084] (1) Co-precipitation preparation of Fe3O4@BIP-BC material: Fe3O4@BIP-BC was prepared by co-precipitation method. The specific operation method is as follows: 0.435 g of ferric chloride was dissolved in 20 mL of deionized water, and 1.2 g of protein blotting biochar was added to the ferric chloride solution and contacted for 30 min. Then 0.126 g of ferrous chloride was weighed according to the molar ratio of 2:1, dissolved in 20 mL of deionized water and added to the ferric chloride solution, then 10 mL of ammonia solution was quickly added to the solution, stirred for 1 h, then the precipitate was washed with deionized water and anhydrous ethanol, and then dried by freeze dryer to obtain Fe3O4@BIP-BC particles with a mass ratio of 1:4, which were stored at room temperature for use.

[0085] (2) Co-precipitation loading of CaO2@Fe3O4@BIP-BC material: CaO2@Fe3O4@BIP-BC was prepared by co-precipitation method. The specific operation method is as follows: 1.078 g of CaCl2 particles was weighed and dissolved in deionized water, 0.3 g of Fe3O4@BIP-BC particles was dispersed in the CaCl2 solution, stirred for 15 min, then 5 mL of ammonia water and 5 mL of polyethylene glycol PEG200 were added, wherein the ammonia water can neutralize the HCl generated in the reaction, and the polyethylene glycol PEG200 is mainly used as a stabilizer. After mixing evenly, 5 mL of H2O2 solution was slowly added, and it was observed that the solution gradually changed from black to black gray, indicating that the co-precipitation reaction occurred and the CaO2@Fe3O4@BIP-BC composite material loaded with calcium peroxide was formed. The suspension was centrifuged and washed with anhydrous ethanol several times, and then dried in a freeze dryer to obtain CaO2@Fe3O4@BIP-BC material with a mass ratio of 8:1:4, which was stored at room temperature for use.

[0086] Example 2

[0087] The Fe3O4@BIP-BC particles (carrying protein blotting) prepared in Example 1 and the Fe3O4@BC particles (without protein blotting) prepared were used to test the adsorption performance of the target bacteria.

[0088] Among them, the preparation method of Fe3O4@BC particles is basically the same as that of Fe3O4@BIP-BC particles, the difference is that the apparent protein blotting is not performed in the preparation process of Fe3O4@BC particles (i.e. substep (2) in step (1)). No CaO2 is added to prevent the occurrence of Fenton-like reaction to inactivate the bacteria and affect the accuracy of the test results of the adsorption performance.

[0089] (1) Adsorption isotherm

[0090] Fe3O4@BIP-BC material and Fe3O4@BC material were mixed with E. coli HB101 bacterial solution at a concentration of 0.2 mg / mL in a 200 mL system, mixed in an incubator for 2 h, and the initial OD600 of the bacterial suspension was 0.02, 0.04, 0.06, 0.08, 0.1 and 0.12. The bacterial-nanoparticle complex was separated under the action of a magnetic field, and the concentration of E. coli HB101 cells in the supernatant was detected.

[0091] (2) Adsorption kinetics

[0092] Fe3O4@BIP-BC material and Fe3O4@BC material were mixed with E. coli HB101 bacterial solution at a concentration of 0.2 mg / mL in a 200 mL system, mixed in an incubator for 2 h. At 0, 20, 40, 60, 80, 100, 120 min, sample the mixed system, separate the bacterial-nanoparticle complex under the action of a magnetic field, and detect the concentration of E. coli HB101 cells in the supernatant.

[0093] (3) Adsorption selectivity

[0094] To study the adsorption selectivity of Fe3O4@BIP-BC material and Fe3O4@BC material, Staphylococcus aureus, Shewanella CN32 and E. coli DH5a were selected as competitive bacteria.

[0095] First, single bacterial solution was used for E. coli HB101 bacteria and competitive bacteria adsorption experiment to explore the adsorption selectivity of Fe3O4@BIP-BC material and Fe3O4@BC material; in short, 20 mg of Fe3O4@BIP-BC and Fe3O4@BC were added to 100 mL of two bacterial solutions (OD600=0.1), mixed at 25°C at a speed of 180 rpm for 2 hours, then the supernatant was taken to count the bacteria in the solution, and the amount of bacteria adsorbed by the material was calculated.

[0096] Staphylococcus aureus, Shewanella CN32 and E. coli DH5a were selected as competitive cells to study the adsorption selectivity of Fe3O4@BIP-BC in a binary system. In short, 20 mg of Fe3O4@BIP-BC was added to a bacterial solution containing the same concentration of E. coli HB101 and competitive bacteria (OD600=0.1). The mixture was placed in a shaking incubator at 25°C and 180 rpm for 2 hours, and then the remaining bacteria in the supernatant were counted. The effect of competitive bacteria was evaluated by displacement of target bacteria, and the calculation method was as follows:

[0097] Substitution rate = (HB101 adsorption amount - HB101 adsorption amount in competitive adsorption) / HB101 adsorption amount.

[0098] The results of the adsorption experiment are shown in Table 3, which shows that the biochar material prepared with the protein imprint has a higher adsorption amount of the target bacteria than the original biochar. In combination with the results of the competitive adsorption experiment, it can be found that the biochar carrying the protein imprint has a much higher adsorption amount of the target bacteria than other bacteria, and the adsorption of the target bacteria by the material is almost not affected in the presence of other bacteria. Figure 1

[0099] Example 3

[0100] 1) The effect of the composite repair material prepared by using the CaO2@Fe3O4@BIP-BC material prepared in Example 1 on the treatment of water harmful bacteria and harmful genes under different activator ratios (Fe3O4:BIP-BC).

[0101] The composite repair material was prepared according to the steps of Example 1.

[0102] 100 mL of E. coli HB101 bacterial solution with an initial concentration of 10 6 CFU / mL was respectively configured in 5 250 mL glass conical flasks, and the composite material was prepared according to the mass ratio of Fe3O4:BIP-BC as 2:1, 1:1, 1:2, 1:4, and 1:6, wherein the dosage of Fe3O4@BIP-BC was controlled as 30 mg / L, and the concentration of CaO2 in the 200 mL reaction system was 1 mM. After the repair material and the polluted water sample were fully contacted and uniformly mixed, the mixed system was placed in a shaking incubator under the condition of 25°C and 180 rpm for 3 h, and the residual bacteria number in the water body was calculated at a fixed time point, and the removal rate was calculated, and the results are shown in Table 4. Figure 4 As can be seen from Table 4, the removal rates of E. coli HB101 by different ratio activator materials are different, and the effects of Fe3O4:BIP-BC ratios of 1:4 and 1:6 are the best. In the actual engineering use process, the best ratio should be determined in combination with the material preparation and application cost. Figure 4

[0103] 2) The effect of the composite repair material prepared by using the CaO2@Fe3O4@BIP-BC material prepared in Example 1 on the treatment of water harmful bacteria and harmful genes under different CaO2 concentrations.

[0104] The composite repair material was prepared according to the steps of Example 1.

[0105] 100 mL of E. coli HB101 bacterial solution with an initial concentration of 10 6 ​​100 mL of E. coli HB101 bacteria solution with a CFU of 10 Figure 5 As can be seen from Figure 5 different CaO2 concentrations have different effects on the removal rate of E. coli HB101, and the effect is best when the CaO2 concentration is 1 mM. In actual engineering use, the best ratio should be determined in combination with the cost of material preparation and application.

[0106] 3) The effect of CaO2@Fe3O4@BIP-BC material prepared in Example 1 on the effect of composite repair material on harmful bacteria and harmful genes in water under different Fe3O4@BIP-BC dosages.

[0107] The composite repair material was prepared according to the steps of Example 1.

[0108] 100 mL of E. coli HB101 bacteria solution with a CFU of 10 6 CFU / ml were prepared and placed in 5 250 mL glass conical flasks, and composite materials were prepared according to the mass ratio of Fe3O4:BIP-BC of 1:4, wherein the dosages of Fe3O4@BIP-BC were controlled to be 10, 20, 30 and 40 mg / L respectively, and the concentration of CaO2 in the 200 mL reaction system was 1 mM. After the repair material was fully mixed with the contaminated water sample, the mixed system was placed in a shaking incubator at 25°C and 180 rpm for 3 h. Water samples were taken at fixed time points to calculate the residual bacteria in the water body and the removal rate, and the results are shown in Figure 6 As can be seen from Figure 6 different Fe3O4@BIP-BC dosages have different effects on the removal rate of E. coli HB101, and the effects are best when the dosages of Fe3O4@BIP-BC are 30 mg / L and 40 mg / L. In actual engineering use, the best ratio should be determined in combination with the cost of material preparation and application.

[0109] Example 4

[0110] The inactivation performance of CaO2@Fe3O4@BIP-BC material prepared in Example 1 on harmful bacteria under different water matrix conditions was tested, and the bacteria solution was prepared according to Example 1.

[0111] In the experiment considering the influence of solution pH, the initial solution pH was adjusted to 3, 5, 7, 9 by adding HCl or NaOH, and the harmful bacteria inactivation experiment was carried out;

[0112] In the experiment considering the influence of solution salinity, the solution ionic strength was adjusted by adding different amounts of NaCl suspension in the reaction system, and the harmful bacteria inactivation experiment was carried out under the conditions of ionic concentration of 0 mM, 15 mM, 75 mM, 150 mM and 300 mM;

[0113] In order to determine the effect of symbiotic humic acid (HA), HA was added to the disinfection system, and the harmful bacteria removal experiment was carried out under the conditions of the final concentration of HA being 0, 0.1, 1 and 10 mg / L.

[0114] The test results are shown in Figure 7, and the results show that the pH has little effect on the bacteria inactivation effect, the inactivation speed is slow when the pH is high, and it may be because the reaction of CaO2 with water is inhibited when the pH is high; the HA concentration has a great effect on the bacteria inactivation effect, and the bacteria inactivation effect is poor when the HA concentration is high because of the competition with bacteria; the salinity has little effect on the bacteria inactivation effect, and the bacteria inactivation rate is different under different salinity conditions, but the bacteria are all inactivated finally.

[0115] Application Example 1

[0116] Deep soil in-situ injection-high pressure rotary jet method:

[0117] The grouting pipe is inserted into the deepest part of the contaminated soil layer, and the self-activated Fenton-like composite repair material of Example 1 is mixed with water in a certain solid-liquid ratio, and then injected into the contaminated deep soil under the action of high-pressure grouting pump and compressed air along the rotary jet grouting pipe with nozzle, and the grouting pipe with nozzle is gradually lifted at a certain interval to ensure that the material can cover all the pollution depth. The composite repair material can fully contact with the contaminated soil due to the high pressure and large jet range of high pressure rotary jet, so as to achieve the purpose of in-situ removal of harmful bacteria and harmful genes in soil.

[0118] Among them, the dosage of the composite repair material is controlled to be 0.1-2.0% of the dry weight of the contaminated soil, the solid-liquid ratio of the material and water in the grouting slurry is controlled to be 1:10-1:1000, the initial insertion depth of the grouting pipe is consistent with the maximum depth of the contaminated soil, the lifting interval of the grouting pipe can be 0.5-2.5 m, the rotary jet pressure can be 0.5-2.5 MPa, and the rotary jet angle can be 120-350°.

[0119] In the application example, the composite remediation material is added in an amount of 1.0% of the dry weight of the contaminated soil, the solid-liquid ratio of the material and water in the grouting slurry is controlled to be 1:100, the initial insertion depth of the grouting pipe is consistent with the maximum depth of the contaminated soil, the lifting interval of the grouting pipe can be 1.0 m, the rotary jet pressure can be 2.0 MPa, and the rotary jet angle can be 270°.

[0120] Application Example 2

[0121] Surface soil in-situ plowing method:

[0122] The self-activated Fenton-like composite remediation material of Example 1 is uniformly applied on the surface of the contaminated soil, and then the soil is plowed to mix the remediation material with the soil uniformly. During the remediation process, the soil is required to have a certain moisture content to promote the contact between calcium peroxide and water to release hydrogen peroxide, so as to achieve the purpose of in-situ removal of harmful bacteria and harmful genes in the surface soil through the self-activated Fenton-like effect and physical adsorption effect of the material.

[0123] In the application example, the composite remediation material is applied in an amount of 1.5% of the dry weight of the contaminated soil, the plowing depth of the soil is about 20-30 cm, the moisture content of the soil after applying the composite remediation material is adjusted to about 60% of the maximum field water holding capacity, and the treatment time of the remediation material is preferably 15 days.

[0124] In the application example, the composite remediation material is applied in an amount of 1.5% of the dry weight of the contaminated soil, the plowing depth of the soil is about 20-30 cm, the moisture content of the soil after applying the composite remediation material is adjusted to about 60% of the maximum field water holding capacity, and the treatment time of the remediation material is preferably 15 days.

[0125] Application Example 3

[0126] Gravity addition method:

[0127] The self-activated Fenton-like material of Example 1 is added in a certain proportion into the water suction pipe of the water pump or the water suction horn of the water suction well by gravity, the impeller of the water pump is used to mix the medicament with the contaminated water uniformly, and the adsorption effect and the self-activated Fenton-like effect of the material on harmful bacteria and harmful genes are used to remove harmful bacteria and harmful genes in the water environment.

[0128] In the application example, the medicament dosage is 18 mg / L, and the hydraulic retention time is 2.5 h.

[0129] In the application example, the medicament dosage is 18 mg / L, and the hydraulic retention time is 2.5 h.

[0130] Application Example 4

[0131] Pressure addition method:

[0132] The self-activated Fenton reagent of example 1 is mixed with water in a certain proportion, and then the reagent is added into a water pipe or a sewage pool by using a water pump or a water jet, and then the reagent is uniformly mixed with the polluted water in the water pipe or the pool, and the harmful bacteria and the harmful genes in the water environment are removed through the adsorption of the material to the harmful bacteria and the harmful genes and the self-activated Fenton effect.

[0133] In the formula, the dosage of the reagent is 15-20 mg / L, and the hydraulic retention time is 2-3 h.

[0134] In the application example, the dosage of the reagent is 16 mg / L, and the hydraulic retention time is 3 h.

[0135] In summary, the self-activated Fenton composite repair material prepared by using the biochar material with good adsorption capacity as a matrix, combining the Western blotting technology and the co-precipitation method, and using the magnetite as an activator and the calcium peroxide as a slow-release oxidant, is used for removing the harmful bacteria and the harmful genes in the water / soil environment.

[0136] The calcium peroxide slowly reacts with water in the environment to slowly release H2O2, so that the oxidation environment is maintained for a long time to completely inactivate the harmful bacteria and degrade the harmful genes.

[0137] The calcium peroxide and the magnetite are simultaneously loaded on the biochar to realize one-time addition of the activator and the oxidant, simplify the operation steps of the traditional Fenton treatment, improve the application efficiency, and reduce the raw material and labor costs. The biochar modified by the Western blotting is used as a matrix material, can selectively adsorb the target harmful bacteria in the environment, and promotes the reduction of Fe 3+ to improve the Fenton reaction efficiency.

[0138] The above description of the embodiments is for facilitating the understanding and use of the application by ordinary skilled persons in the art. Those skilled in the art can easily make various modifications to the embodiments, and apply the general principles described herein to other embodiments without creative labor. Therefore, the application is not limited to the above embodiments, and the improvements and modifications made by those skilled in the art according to the disclosure of the application without departing from the scope of the application should be within the protection scope of the application.

Claims

1. A self-activated Fenton-like composite repair material, characterized in that, The composite repair material is CaO2@Fe3O4@BIP-BC, which takes the protein imprinting biochar as a substrate, and magnetite and calcium peroxide as an activator and a slow-release oxidant, respectively, which are sequentially loaded on the surface.

2. The self-activated Fenton-like composite repair material according to claim 1, characterized in that, In the CaO2@Fe3O4@BIP-BC, the mass ratio of Fe3O4 to BIP-BC is 12-1:

6.

3. The self-activated Fenton-like composite repair material according to claim 1, characterized in that, The protein imprinting biochar is obtained by binding target harmful bacteria on the surface of biochar through a protein imprinting technology.

4. The self-activated Fenton-like composite repair material according to claim 1, characterized in that, The particle size of the composite repair material is 0.2-2.5 mm.

5. A method for preparing the self-activation Fenton-like composite repair material according to any one of claims 1-4, characterized in that, The method comprises the following steps: S1: Preparation of biochar: pulverized biomass material is pyrolyzed by a limited oxygen slow heating method, and after cooling, it is ground and sieved, and then the sieved powder is soaked in an HCl solution, and then washed to obtain biochar; S2: Preparation of protein imprinting biochar: inoculate target harmful bacteria in a LB medium containing protease, and after oscillation culture, centrifuge to obtain the precipitate, resuspend the precipitate in a lysis extraction buffer and heat, centrifuge to obtain the supernatant, add DNase and RNase to the supernatant and mix uniformly, stand in ice water to remove nucleic acid impurities, add acidified acetone-ethanol mixture to the supernatant and stand in ice water to precipitate proteins, centrifuge to collect the precipitate and wash, remove methanol by vacuum filtration, resuspend the precipitate in a sample solubilization buffer, add the biochar obtained in step S1 to the resuspension, mix and wash, and dry to obtain protein imprinting biochar BIP-BC; S3: Preparation of Fe3O4@BIP-BC by co-precipitation, the specific steps are as follows: prepare ferric chloride solution and ferrous chloride solution respectively, disperse the protein imprinting biochar obtained in step S2 in the ferric chloride solution, add ferrous chloride solution to the ferric chloride solution after a certain period of contact, then add an excess of ammonia water solution and continue to stir, separate and wash the precipitate, and freeze-dry to obtain Fe3O4@BIP-BC particles; S4: Preparation of CaO2@Fe3O4@BIP-BC by co-precipitation, the specific steps are as follows: prepare calcium chloride solution, disperse the Fe3O4@BIP-BC particles obtained in step S3 in the calcium chloride solution, then add a neutralizing agent and a stabilizer and stir uniformly, add hydrogen peroxide to the mixture, separate and wash the precipitate, and freeze-dry to obtain the composite repair material.

6. The preparation method of a self-activated Fenton-like composite repair material according to claim 5, characterized in that, In step S1: The pyrolysis process is as follows: first, preheat at 100℃ for 1h under a nitrogen atmosphere, then heat to 300-700℃ for 2h; The sieving is performed using a 100-mesh screen; The concentration of the HCl solution is 10-15wt%; The soaking time is 12-24h.

7. The preparation method of a self-activated Fenton-like composite repair material according to claim 5, characterized in that, In step S2: The target harmful bacteria include gram-negative bacteria and gram-positive bacteria; The LB medium containing protease is an LB medium containing 0.5-2.5% trypsin; The oscillation culture is continuous oscillation culture at 37℃ at a speed of 150-180rpm for 8-10 hours; The lysis extraction buffer is a mixture of pH=7, 20mM HEPES, 2-5wt% sodium dodecyl sulfate and protease inhibitors, and the precipitate is reconstituted in the lysis extraction buffer at 10μL / mg wet cell weight; The heating is heating at 95℃ for 3-5 minutes; The addition amount of the DNase is 1mg / mL, and the addition amount of the RNase is 0.5mg / mL; The concentration of the acidified acetone-ethanol mixture is 1.5mM; The sample solubilization buffer is a mixture of 8-10M urea, 2-4M thiazolinone, 2-4wt% CHAPS, 1-2wt% TCEP-HCl and 2-5% v / v carrier ampholyte 3-10NL; The biochar is added in equal weight with the wet weight of the bacteria in the resuspension.

8. The preparation method of a self-activated Fenton-like composite repair material according to claim 5, characterized in that, In step S3: Ferric chloride: protein blotting biochar: ferrous chloride = 0.324g: 0.8g: 0.126g; The contact time is 30-60min, and the duration of stirring is 1h.

9. The preparation method of a self-activated Fenton-like composite repair material according to claim 5, characterized in that, In step S4: Calcium chloride: Fe3O4@BIP-BC particles: stabilizer: hydrogen peroxide = 1.078g: 0.3g: 5mL: 5mL; The neutralizing agent is ammonia water, and the stabilizer is PEG200.

10. The use of the self-activated Fenton-like composite repair material according to any one of claims 1-4 for removing harmful bacteria and harmful genes in the environment.

Images