A method for remediating waste-contaminated soil by co-immobilizing microorganisms and enzymes with magnetic plastic charcoal.

Abstract

This invention relates to the field of in-situ remediation technology for landfill-contaminated soil, and in particular to a method for remediating landfill-contaminated soil using co-immobilized magnetic plastic landfill char. The invention uses plastic waste as raw material to prepare magnetic plastic landfill char. Simultaneously, functional microorganisms with strong plastic-degrading capabilities, enriched and domesticated from the surface of plastic waste, are adsorbed and immobilized onto the plastic landfill char. Then, the functional microbial culture medium and enzymes are encapsulated and immobilized through the colloidal protection of water-soluble polymer compounds, resulting in co-immobilized magnetic plastic landfill char particles. When applied to the in-situ remediation of landfill-contaminated soil, this method can enhance the biocatalytic degradation of organic pollutants such as plastics and plastic additives in landfill-contaminated soil through co-immobilization, solving the problems of recalcitrant plastic degradation and the migration and removal of pollutants released from landfill-contaminated soil.

Description

Technical Field

[0001] This invention relates to the field of in-situ remediation technology for soil contaminated by waste, and in particular to a method for remediating soil contaminated by waste by co-immobilizing magnetic plastic waste char with bacteria and enzymes. Background Technology

[0002] The widespread use of plastics and their products has exacerbated the generation of plastic waste, and the soil pollution caused by the informal dumping or landfilling of plastic waste is becoming increasingly serious. Currently, most plastic waste treatment methods involve landfilling, incineration, and reprocessing, but these are costly and produce toxic and harmful gases and residues, causing secondary pollution. Furthermore, improper treatment methods can generate a new type of pollutant—microplastics. Long-term storage of plastic waste can lead to the release and migration of plastic additives such as bisphenols (BPs), phthalates (PAEs), and benzotriazole UV stabilizers (BUVSs), posing unpredictable ecological risks to the environment. In addition, an increasing number of new pollutants are being detected in waste-polluted environments, with polycyclic aromatic hydrocarbons (PAHs) and antibiotics appearing frequently. The removal of new pollutants from waste-polluted environments will become a major focus of future research in the field of waste pollution.

[0003] Current treatment technologies primarily target single plastic adsorption or single organic pollutant degradation and removal, with fewer technologies taking a comprehensive approach. Therefore, exploring an in-situ soil remediation technology that combines plastic degradation and organic pollutant removal is of great significance. At the same time, finding a technology that can simultaneously achieve green and environmentally friendly, efficient resource utilization of plastic waste and effective treatment of pollutants is also crucial. Summary of the Invention

[0004] Based on the above, this invention provides a method for remediating landfill-contaminated soil using co-immobilized magnetic plastic landfill charcoal. This invention prepares magnetic plastic landfill charcoal from plastic waste as raw material. Simultaneously, functional microorganisms with strong plastic-degrading capabilities, enriched and domesticated from the surface of plastic waste, are adsorbed and immobilized onto the plastic landfill charcoal. Then, the functional microbial culture medium and enzymes are encapsulated and immobilized through the colloidal protection of water-soluble polymer compounds, resulting in co-immobilized magnetic plastic landfill charcoal particles. These particles are then applied to the in-situ remediation of landfill-contaminated soil to address the problems of recalcitrant plastic degradation and the migration and removal of pollutants released from landfill-contaminated soil.

[0005] To achieve the above objectives, the present invention provides the following solution:

[0006] One of the technical solutions of this invention is a method for preparing microbial enzyme co-immobilized magnetic plastic waste charcoal, comprising the following steps:

[0007] Plastic waste fragments in soil contaminated by garbage are crushed to obtain plastic pellets;

[0008] The plastic particles were added to an alkaline solution, heated, and then pyrolyzed to obtain porous plastic waste charcoal.

[0009] The porous plastic waste charcoal was mixed with nano magnetic powder and placed in ammonia water for adsorption to obtain magnetic plastic waste charcoal.

[0010] Degrading bacteria on the surface of plastics in waste-polluted soil were enriched and cultured using LB liquid medium, and then transferred to inorganic salt basal medium. After 3-5 transfers and acclimatizations, functional bacterial solutions were obtained.

[0011] The functional bacterial culture was inoculated into LB liquid medium, and the magnetic plastic waste charcoal was added simultaneously for the expansion culture and adsorption immobilization of microorganisms. Then, solid and liquid were separated to obtain solid and culture medium. The solid was freeze-dried to obtain magnetic plastic waste charcoal immobilized microbial particles.

[0012] The culture medium was mixed with an enzyme and a water-soluble polymer solution to obtain a cross-linking agent-enzyme mixed buffer.

[0013] The cross-linking agent-enzyme mixed buffer solution was mixed with the magnetic plastic waste char immobilized microbial particles by shaking and then freeze-dried to obtain the bacterial-enzyme co-immobilized magnetic plastic waste char.

[0014] The second technical solution of the present invention is a kind of bacterial enzyme co-immobilized magnetic plastic waste char prepared according to the above preparation method.

[0015] The third technical solution of this invention is the application of the above-mentioned bacterial enzyme co-immobilized magnetic plastic waste charcoal in the remediation of waste-contaminated soil.

[0016] The fourth technical solution of the present invention is a method for remediating waste-contaminated soil, which achieves the remediation of waste-contaminated soil by mixing the above-mentioned microbial enzyme co-immobilized magnetic plastic waste charcoal with waste-contaminated soil.

[0017] The present invention discloses the following technical effects:

[0018] Enriching and cultivating microbial communities with strong plastic-degrading capabilities on plastic surfaces can maximize the efficient decomposition of plastics. However, both microorganisms and enzymes face challenges such as difficulty in recycling and easy inactivation, limiting their practical applications. Immobilization technology can fix functional bacteria and enzymes onto adsorbent materials, and then effectively maintain the quantity and activity of microorganisms through encapsulation and cross-linking, improving and enhancing their stability. It also enables reusability, thereby reducing usage costs. Magnetic carbon possesses excellent magnetism and a high specific surface area, enabling effective adsorption and immobilization of enzymes. By immobilizing enzymes on magnetic carbon materials, not only can enzyme stability and reusability be improved, but application and recycling in soil can also be facilitated. Plastics are organic polymers with carbon as their main skeleton, characterized by low density, abundant porosity, and high specific surface area. Plastic waste with high carbon content can become an ideal carbonization material. By converting waste plastics into magnetic carbon materials, not only can waste be reused, but the properties of magnetic carbon, such as high surface area and magnetism, can also be used to immobilize enzymes and microorganisms.

[0019] Compared to physicochemical treatment technologies, microbial degradation and enzyme-catalyzed bioremediation technologies are safer, more environmentally friendly, stable, and more efficient, making them the most promising technologies for pollutant treatment. Enzymes are biocatalysts with highly efficient catalytic and specific reaction capabilities for organic matter. They can decompose recalcitrant organic pollutants such as plastics, cellulose, and plasticizers, producing more environmentally friendly decomposition products, thereby reducing their environmental impact. Co-remediation of bacteria and enzymes organically combines the catalytic degradation effects of functional microbial communities with those of enzyme preparations, improving degradation efficiency; it can also regulate the environmental microecological balance and increase the adaptability of the bacteria-enzyme complex in the environment. The advantage of this invention lies in improving the biocatalytic degradation of organic pollutants such as plastics and plastic additives in waste-contaminated soil through bacteria-enzyme co-immobilization, while also adsorbing microplastics in the contaminated soil. This technology is simple and easy to implement, highly reusable, and has high remediation efficiency, enabling the efficient degradation and resource utilization of waste plastics, and offering both environmental and economic benefits in solving plastic pollution and recycling. Attached Figure Description

[0020] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0021] Figure 1 This is a process flow diagram illustrating the preparation of magnetic plastic waste charcoal co-immobilized with bacteria and enzymes, and its use in the remediation of waste-contaminated soil. Detailed Implementation

[0022] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as a limitation of the present invention, but rather as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0023] It should be understood that the terminology used in this invention is merely for describing particular embodiments and is not intended to limit the invention. Furthermore, with respect to numerical ranges in this invention, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. Any stated value or intermediate value within a stated range, as well as each smaller range between any other stated value or intermediate value within said range, is also included in this invention. The upper and lower limits of these smaller ranges may be independently included or excluded from the range.

[0024] Unless otherwise stated, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. While only preferred methods and materials have been described herein, any methods and materials similar or equivalent to those described herein may be used in the implementation or testing of this invention. All references to this specification are incorporated by way of citation to disclose and describe methods and / or materials associated with those references. In the event of any conflict with any incorporated reference, the content of this specification shall prevail.

[0025] Various modifications and variations can be made to the specific embodiments described in this specification without departing from the scope or spirit of the invention, as will be apparent to those skilled in the art. Other embodiments derived from this specification will also be apparent to those skilled in the art. This specification and embodiments are merely exemplary.

[0026] The terms “include,” “including,” “have,” “contain,” etc., used in this article are all open-ended terms, meaning that they include but are not limited to.

[0027] The first aspect of this invention provides a method for preparing bacterial enzyme co-immobilized magnetic plastic waste charcoal, comprising the following steps:

[0028] Plastic waste fragments in soil contaminated by garbage are crushed to obtain plastic pellets;

[0029] The plastic particles were added to an alkaline solution, heated, and then pyrolyzed to obtain porous plastic waste charcoal.

[0030] The porous plastic waste charcoal was mixed with nano magnetic powder and placed in ammonia water for adsorption to obtain magnetic plastic waste charcoal.

[0031] Degrading bacteria on the surface of plastics in waste-polluted soil were enriched and cultured using LB liquid medium, and then transferred to inorganic salt basal medium. After 3-5 transfers and acclimatizations, functional bacterial solutions were obtained.

[0032] The functional bacterial culture was inoculated into LB liquid medium, and the magnetic plastic waste charcoal was added simultaneously for the expansion culture and adsorption immobilization of microorganisms. Then, solid and liquid were separated to obtain solid and culture medium. The solid was freeze-dried to obtain magnetic plastic waste charcoal immobilized microbial particles.

[0033] The culture medium was mixed with an enzyme and a water-soluble polymer solution to obtain a cross-linking agent-enzyme mixed buffer.

[0034] The cross-linking agent-enzyme mixed buffer solution was mixed with the magnetic plastic waste char immobilized microbial particles by shaking and then freeze-dried to obtain the bacterial-enzyme co-immobilized magnetic plastic waste char.

[0035] In a preferred embodiment of the present invention, the LB liquid culture medium is composed of: 10 g / L tryptone, 5 g / L yeast extract, 10 g / L sodium chloride and 1000 mL water, with a pH of 7.2-7.4.

[0036] In a preferred embodiment of the present invention, the inorganic salt basal culture medium is composed of: 5-10 g / L plastic waste fragments, 1.0 g / L ammonium nitrate, 0.4 g / L potassium dihydrogen phosphate, 1.2 g / L sodium chloride, 0.2 g / L calcium carbonate, 0.2 g / L magnesium sulfate heptahydrate, 0.1 g / L compound vitamins, and 1000 mL water, with a pH of 7.2-7.4.

[0037] In a preferred embodiment of the present invention, plastic waste fragments in soil contaminated by garbage are crushed and passed through a 50-200 mesh sieve to obtain plastic particles.

[0038] In a preferred embodiment of the present invention, the mass-to-volume ratio of the plastic particles to the alkaline solution is 1:1.5-4 (w / v); the alkaline solution is a 10% KOH solution; the heating treatment specifically involves ultrasonic heating at 45-60℃ for 1-2 hours; the heating treatment followed by filtration, washing, and drying steps are included before pyrolysis; the pyrolysis specifically involves pyrolysis at 350-650℃ in an N2 environment for 1-4 hours; the pyrolysis is followed by cooling and grinding through a 50-200 mesh sieve.

[0039] The main functions of alkaline solutions are to remove impurities from the surface of plastic particles, promote irregular and slight corrosion of the particle surface (increasing the specific surface area of ​​the particles), and enhance the structural stability of plastic particles. If the amount of alkaline solution added exceeds the range described above, the alkaline solution will cause significant damage to the surface morphology and chemical properties of the plastic, resulting in partial degradation or severe embrittlement of the plastic; or the surface modification effect of the plastic particles will be unsatisfactory.

[0040] The purpose of ultrasonic heating is to accelerate the reaction and promote uniform corrosion on the surface of plastic particles.

[0041] The purpose of pyrolysis is to promote the carbonization of plastic particles, increase the pore structure of the material, and further improve the stability of the material structure.

[0042] In a preferred embodiment of the present invention, the mass concentration of the ammonia water is 5%;

[0043] Ammonia, as a weakly alkaline solution, acts as a catalyst. Excessive ammonia concentration will damage the magnetic structure; insufficient concentration will result in weak catalysis and a magnetically unsound product. Other commonly used alkaline substances in this field, such as sodium hydroxide, have poor catalytic effects and are not suitable for this invention.

[0044] In a preferred embodiment of the present invention, the mass ratio of the porous plastic waste charcoal to the nano-magnetic powder is 1-3:1; the nano-magnetic powder is at least one of zero-valent Fe, Fe2O3 and Fe3O4; the adsorption is specifically performed at 25°C and 180 r / min for 8-12 h; the adsorption is further performed by ultrasonic dispersion for 1-5 min before adsorption.

[0045] The reason why the mass ratio of porous plastic waste charcoal to nano-magnetic powder is limited to 1-3:1 is that if too little nano-magnetic powder is added, the magnetic properties of the synthesized material will be insufficient, affecting the later repair and recycling; if too much nano-magnetic powder is added, the adsorption and catalytic degradation performance of the synthesized material will be significantly reduced.

[0046] In a preferred embodiment of the present invention, the enrichment culture conditions are 30°C, 180 r / min, 24 h; the bacterial solution obtained by enriching and culturing the degrading bacteria on the plastic surface of the garbage-contaminated soil using LB liquid medium is transferred to an inorganic salt basal medium at a volume concentration of 1%-5%; the inorganic salt basal medium uses plastic garbage fragments at a mass concentration of 0.5%-1% as the sole carbon source; the plastic garbage fragments are from the same source as the plastic used in enriching and culturing the degrading bacteria on the plastic surface of the garbage-contaminated soil; the acclimatization conditions are 30°C, 180 r / min, 5-10 days / time.

[0047] In a preferred embodiment of the present invention, the functional bacterial solution is inoculated into LB liquid culture medium at a volume concentration of 5%-10%; the magnetic plastic waste charcoal has a mass concentration of 5%-15% in the LB liquid culture medium; and the conditions for the expansion culture and adsorption fixation of the microorganisms are 25-30℃, 180r / min, and 18-24h.

[0048] The reason why the conditions for the large-scale culture and adsorption fixation of microorganisms are limited to the above parameter range is that exceeding the above range will lead to unstable microbial particle structure or unsatisfactory adsorption-degradation effect in magnetic plastic waste char.

[0049] In a preferred embodiment of the present invention, the enzyme is at least one selected from laccase, catalase, protease, cellulase, and ligninase; the concentration of the enzyme in the culture medium is 0.5-10 mg / mL; the culture medium is ultrasonically disrupted in an ice-water bath for 3-5 min before adding the enzyme; the water-soluble polymer solution is a 1.5-3 g / L water-soluble polymer PBS buffer solution; the water-soluble polymer is one selected from polyacrylamide, polyethyleneimine, polyethylene glycol, carboxymethyl cellulose, and polyvinyl alcohol; the volume ratio of the culture medium to the water-soluble polymer solution is 1:2-4; the mass concentration of the magnetic plastic waste carbon immobilized microbial particles in the crosslinking agent-enzyme mixed buffer is 3%-12%; the microbial particles are ultrasonically dispersed for 3-5 min before oscillation; the oscillation conditions are 25°C, 180 r / min, and oscillation for 2-8 h; the process after oscillation and before freeze-drying includes a filtration and washing step.

[0050] The reason for limiting the amount of enzyme and the amount of magnetic plastic waste char immobilized microbial particles to the above parameters is that exceeding the above range will lead to structural instability of the microbial-enzyme co-immobilized magnetic plastic waste char or unsatisfactory adsorption-degradation effect.

[0051] A second aspect of the present invention provides a microbial enzyme co-immobilized magnetic plastic waste char prepared according to the above-described preparation method.

[0052] The third aspect of this invention provides the application of the above-mentioned bacterial enzyme co-immobilized magnetic plastic waste charcoal in the remediation of waste-contaminated soil.

[0053] The fourth aspect of this invention provides a method for remediating waste-contaminated soil, which achieves the remediation of waste-contaminated soil by mixing the aforementioned microbial enzyme-immobilized magnetic plastic waste charcoal with the waste-contaminated soil.

[0054] In a preferred embodiment of the present invention, the amount of the bacterial enzyme co-immobilized magnetic plastic waste charcoal added is 0.5%-5% of the mass of the waste-contaminated soil; the remediation cycle is 2-6 months.

[0055] Unless otherwise specified, the "%" mentioned in this invention refers to a percentage by mass.

[0056] Unless otherwise specified, all raw materials used in the embodiments and comparative examples of this invention can be obtained through commercial channels.

[0057] The LB liquid culture medium used in this embodiment of the invention consists of: 10 g / L tryptone, 5 g / L yeast extract, 10 g / L sodium chloride and 1000 mL water, with a pH of 7.2-7.4.

[0058] The inorganic salt basal culture medium used in this embodiment of the invention consists of: 5-10 g / L plastic waste fragments, 1.0 g / L ammonium nitrate, 0.4 g / L potassium dihydrogen phosphate, 1.2 g / L sodium chloride, 0.2 g / L calcium carbonate, 0.2 g / L magnesium sulfate heptahydrate, 0.1 g / L compound vitamins, and 1000 mL water, with a pH of 7.2-7.4.

[0059] The particle size of the nano Fe3O4 magnetic powder used in the embodiments of the present invention is 20-50 nm.

[0060] The particle size of the nano-Fe2O3 magnetic powder used in the embodiments of the present invention is 20-50 nm.

[0061] The nano-zero valent iron used in the embodiments of the present invention has a particle size of 20-30 nm.

[0062] The waste-contaminated topsoil and waste-contaminated soil mentioned in the embodiments and comparative examples of this invention are all waste-contaminated soils from the same source.

[0063] The process flow diagram for preparing magnetic plastic waste charcoal using bacterial enzyme co-immobilization and utilizing it for remediation of waste-contaminated soil is shown below. Figure 1 As shown.

[0064] Example 1

[0065] (1) Collect the topsoil of the garbage-contaminated soil (thickness ≤30cm), collect the larger plastic garbage fragments in the contaminated soil, wash and dry them, crush them, and pass them through a 100-mesh sieve to obtain plastic particles.

[0066] In addition, severely aged and damaged plastic waste from contaminated soil was selected and enriched with plastic-degrading bacteria using LB liquid medium (30℃, 180r / min, 24h). The bacteria were then transferred at 5% (v / v) to an inorganic salt basal medium containing 1% (w / v) washed and dried plastic waste fragments as the sole carbon source. This process was repeated to obtain a functional bacterial solution with strong plastic-degrading ability (30℃, 180r / min, 7d / time, 5 times of transfer and acclimatization).

[0067] (2) The above plastic particles were added to a 10% KOH solution at a ratio of 1:3 (w / v), ultrasonically heated at 45°C for 2 hours, filtered, washed with water, and dried at 60°C. The dried plastic particles were then pyrolyzed at 400°C for 2.5 hours in a N2 environment, cooled, ground, and sieved (100 mesh) to prepare porous plastic waste charcoal.

[0068] (3) The porous plastic waste char obtained above is mixed with nano Fe3O4 magnetic powder at a mass ratio of 1:1, and then ultrasonically dispersed in a 5% ammonia solution for 5 min. After mixing and adsorption at 25℃ and 180 r / min for 8 h, magnetic plastic waste char is obtained by magnetic recovery.

[0069] (4) The functional bacterial solution obtained from domestication in step (1) was inoculated into LB liquid medium at 10% (v / v), and 10% (w / v) magnetic plastic waste charcoal was added at the same time to carry out the microbial expansion culture and adsorption fixation (30℃, 180r / min, 24h). Solid and liquid were separated to obtain solid and culture medium. The solid was freeze-dried to obtain magnetic plastic waste charcoal immobilized microbial particles.

[0070] (5) After the culture medium obtained in step (4) was sonicated in an ice-water bath for 3 min, 8 mg / mL laccase was added, and then mixed with 2 g / L polyacrylamide PBS buffer solution (0.1 M, pH 7) at a volume ratio of 1:3 to obtain a cross-linking agent-enzyme mixed buffer. 10% (w / v) of magnetic plastic landfill char immobilized microbial particles were added to the above cross-linking agent-enzyme mixed buffer, sonicated for 5 min, and then shaken at 25℃ and 180 r / min for 8 h. After filtration and washing with water, the microbial enzyme co-immobilized magnetic plastic landfill char was obtained and stored at 4℃ for later use.

[0071] (6) The above-mentioned microbial enzyme-immobilized magnetic plastic waste char was mixed with the waste-contaminated topsoil at an application ratio of 3.5% (w / w), and thoroughly mixed using a soil turning device. The waste-contaminated soil remediation was carried out on a 3-month cycle. The treatment effect of the contaminated soil was evaluated from aspects such as microplastic size, additive concentration, and soil improvement. The results are shown in Table 1.

[0072] Example 2

[0073] (1) Same as (1) in Example 1.

[0074] (2) The above plastic particles were added to a 10% KOH solution at a ratio of 1:4 (w / v), ultrasonically heated at 50°C for 1.5 h, filtered, washed with water, and dried at 60°C. The dried plastic particles were then pyrolyzed at 600°C for 1.5 h in a N2 environment, cooled, ground, and sieved (100 mesh) to prepare porous plastic waste charcoal.

[0075] (3) The porous plastic waste char obtained above is mixed with nano zero-valent iron powder at a mass ratio of 2:1, placed in a 5% ammonia solution and ultrasonically dispersed for 5 min, then mixed and adsorbed at 25℃ and 180 r / min for 8 h, and magnetic plastic waste char is obtained by magnetic recovery.

[0076] (4) The subsequent steps are the same as in Example 1.

[0077] Example 3

[0078] (1)-(2) are the same as (1)-(2) in Example 1.

[0079] (3) The porous plastic waste char obtained above is mixed with nano Fe2O3 magnetic powder at a mass ratio of 2:1, and then ultrasonically dispersed in a 5% ammonia solution for 5 min. After mixing and adsorption at 25℃ and 180 r / min for 10 h, magnetic plastic waste char is obtained by magnetic recovery.

[0080] (4) The functional bacterial solution obtained from domestication in step (1) was inoculated into LB liquid medium at 10% (v / v), and 10% (w / v) magnetic plastic waste charcoal was added at the same time to carry out the microbial expansion culture and adsorption fixation (30℃, 180r / min, 24h). Solid and liquid were separated to obtain solid and culture medium. The solid was freeze-dried to obtain magnetic plastic waste charcoal immobilized microbial particles.

[0081] (5) After the culture medium obtained in step (4) was ultrasonically broken in an ice-water bath for 3 min, 10 mg / mL cellulase was added, and then mixed with 2 g / L polyvinyl alcohol PBS buffer solution (0.1 M, pH 7) at a volume ratio of 1:4. 10% (w / v) magnetic plastic landfill char immobilized microbial particles were added, ultrasonically dispersed for 5 min, and then shaken at 25℃ and 180 r / min for 8 h. After filtration and washing with water, the microbial enzyme co-immobilized magnetic plastic landfill char was obtained and stored at 4℃ for later use.

[0082] (6) The steps are the same as (6) in Example 1.

[0083] Example 4

[0084] (1)-(3) are the same as (1)-(3) in Example 1.

[0085] (4) The functional bacterial solution obtained from domestication in step (1) was inoculated into LB liquid medium at 7% (v / v), and 14% (w / v) magnetic plastic waste charcoal was added at the same time to carry out the microbial expansion culture and adsorption fixation (30℃, 180r / min, 24h). Solid and liquid were separated to obtain solid and culture medium. The solid was freeze-dried to obtain magnetic plastic waste charcoal immobilized microbial particles.

[0086] (5) After the culture medium obtained in step (4) was sonicated in an ice-water bath for 3 min, 5 mg / mL laccase was added, and then mixed with 2 g / L polyacrylamide PBS buffer solution (0.1 M, pH 7) at a volume ratio of 1:2 to obtain a cross-linking agent-enzyme mixed buffer. 10% (w / v) of magnetic plastic landfill char immobilized microbial particles were added to the above cross-linking agent-enzyme mixed buffer, sonicated for 5 min, and then shaken at 25℃ and 180 r / min for 6 h. After filtration and washing with water, the microbial enzyme co-immobilized magnetic plastic landfill char was obtained and stored at 4℃ for later use.

[0087] (6) The above-mentioned bacterial enzyme co-immobilized magnetic plastic waste char was mixed with the waste-contaminated topsoil at an application ratio of 1.5% (w / w), and thoroughly mixed using a soil turning device. Waste-contaminated soil remediation was carried out in a cycle of 5 months.

[0088] The results are shown in Table 1.

[0089] Comparative Example 1 (without laccase)

[0090] The only difference from Example 1 is that in step (5), the above magnetic plastic waste char immobilized microbial particles are added at 10% (w / v) to a PBS buffer solution containing 2 g / L polyacrylamide (0.1 M, pH 7), ultrasonically dispersed for 5 min, and then shaken at 25°C and 180 r / min for 8 h. After filtration and washing with water, the microbial immobilized magnetic plastic waste char is obtained by freeze drying and stored at 4°C for later use.

[0091] The remaining steps were the same as in Example 1. The results are shown in Table 1.

[0092] Comparative Example 2 (No bacteria added)

[0093] (1) Collect the topsoil of the garbage-contaminated soil (thickness ≤30cm), collect the larger plastic garbage fragments in the contaminated soil, wash and dry them, crush them, and pass them through a 100-mesh sieve to obtain plastic particles.

[0094] (2) The above plastic particles were added to a 10% KOH solution at a ratio of 1:3 (w / v), ultrasonically heated at 45°C for 2 hours, filtered, washed with water, and dried at 60°C. The dried plastic particles were then pyrolyzed at 400°C for 2.5 hours in a N2 environment, cooled, ground, and sieved (100 mesh) to prepare porous plastic waste charcoal.

[0095] (3) The plastic waste char obtained above is mixed with nano Fe3O4 magnetic powder at a mass ratio of 1:1, and then ultrasonically dispersed in a 5% ammonia solution for 5 min. After mixing and adsorption at 25℃ and 180 r / min for 8 h, magnetic plastic waste char is obtained by magnetic recovery.

[0096] (4) The above magnetic plastic waste char particles were mixed at 10% (w / v) with a PBS buffer solution containing 2 g / L polyacrylamide (0.1M, pH7) with 8 mg / mL laccase added. After ultrasonic dispersion for 5 min, the mixture was shaken at 25°C and 180 r / min for 8 h. After filtration and washing with water, the immobilized enzyme magnetic plastic waste char was obtained by freeze drying and stored at 4°C for later use.

[0097] (5) The above-mentioned immobilized enzyme magnetic plastic waste charcoal was mixed with the waste-contaminated topsoil at an application ratio of 3.5% (w / w), and the remaining steps were the same as (6) in Example 1. The results are shown in Table 1.

[0098] Comparative Example 3 (without added bacterial enzymes)

[0099] (1) Collect the topsoil of the garbage-contaminated soil (thickness ≤30cm), collect the larger plastic garbage fragments in the contaminated soil, wash and dry them, crush them, and pass them through a 100-mesh sieve to obtain plastic particles.

[0100] (2) The above plastic particles were added to a 10% KOH solution at a ratio of 1:3 (w / v), ultrasonically heated at 45°C for 2 hours, filtered, washed with water, and dried at 60°C. The dried plastic particles were then pyrolyzed at 400°C for 2.5 hours in a N2 environment, cooled, ground, and sieved (100 mesh) to prepare porous plastic waste charcoal.

[0101] (3) The plastic waste char obtained above is mixed with nano Fe3O4 magnetic powder at a mass ratio of 1:1, and then ultrasonically dispersed in a 5% ammonia solution for 5 min. After mixing and adsorption at 25℃ and 180 r / min for 8 h, magnetic plastic waste char is obtained by magnetic recovery.

[0102] (4) The above magnetic plastic waste char particles were mixed with PBS buffer solution containing 2 g / L polyacrylamide (0.1 M, pH 7) at 10% (w / v), ultrasonically dispersed for 5 min, and then shaken at 25℃ and 180 r / min for 8 h. After filtration and washing with water, the mixture was freeze-dried to obtain freeze-dried magnetic plastic waste char and stored at 4℃ for later use.

[0103] (5) The freeze-dried magnetic plastic waste charcoal was mixed with the waste-contaminated topsoil at an application ratio of 3.5% (w / w), and the remaining steps were the same as (6) in Example 1. The results are shown in Table 1.

[0104] Comparative Example 4

[0105] The only difference from Example 1 is that the preparation of magnetic plastic waste charcoal is omitted, and the magnetic plastic waste charcoal is replaced with biocharcoal. The preparation method of the biocharcoal is as follows: waste biomass wood chips are crushed, calcined at 500°C for 2 hours in a nitrogen atmosphere, and then ground and sieved to obtain wood chip biocharcoal with a particle size of <100μm.

[0106] Comparative Example 5 (with added bacterial enzymes, but without immobilization)

[0107] The only difference from Example 1 is that:

[0108] (4) The functional bacterial solution obtained in step (1) was inoculated into LB liquid medium at 10% (v / v), cultured at 30℃, 180r / min for 24h, and then freeze-dried and mixed with 10% (w / v) magnetic plastic waste charcoal to obtain a bacterial-magnetic plastic waste charcoal mixture.

[0109] (5) Add 8 mg / mL laccase to the bacterial-magnetic plastic waste charcoal mixture obtained in step (4), mix at 25°C and 180 r / min for 8 h, filter and wash with water, freeze dry to obtain bacterial-enzyme-magnetic plastic waste charcoal, and store at 4°C for later use.

[0110] The remaining steps are the same as in Example 1.

[0111] Comparative Example 6

[0112] (1) Collect a number of plastic wastes made of PE, PP, PVC and other materials, and enrich the microorganisms on the surface of the plastics; enrich functional bacterial groups with plastic degradation ability in a selective inorganic salt culture medium with at least one of PE, PP and PVC as the sole carbon source, and reduce the weight of the degraded plastic by more than 10%. After centrifuging the above microbial liquid at 5000 rpm for 10 min, resuspend the bacterial cells in sterile deionized water to obtain a bacterial suspension (OD600 of 0.8).

[0113] (2) After crushing the waste biomass wood chips, calcining them at 500°C for 2 hours in a nitrogen atmosphere, and then grinding and sieving them to obtain wood chip biochar with a particle size of <100μ0.

[0114] (3) The above-mentioned sawdust biochar and the above-mentioned bacterial suspension were mixed at a mass ratio of 1:10 and mixed and adsorbed by shaking at 150 r / min for 8 h at 15 °C. After freezing, drying, and grinding, microbial immobilized biochar (particle size <100 μ0) was obtained. The above-mentioned microbial immobilized biochar (1%, w / w) was added to a mixed solution containing 1.5% (w / w) carboxymethyl cellulose and 1.5% (w / w) sodium alginate, stirred evenly, and then slowly added dropwise to a 2% calcium chloride solution. After crosslinking for 16 h, immobilized particles were obtained. The above-mentioned immobilized particles were washed three times with deionized water and dried at low temperature to obtain biochar-based microbial immobilized particles.

[0115] (4) Experiment on the remediation effect of landfill-contaminated soil: The above-mentioned biochar-based microbial immobilized particles were uniformly added to the landfill-contaminated soil at a ratio of 1.5% (w / w), thoroughly mixed using a soil turning device, and left to stand for 3 months. The remediation effect was evaluated by detecting the abundance ratio of microplastics of different particle sizes in the soil using density flotation and microscopic observation and analysis. The results are shown in Table 1.

[0116] Table 1. Remediation effects of different embodiments on waste-contaminated soil

[0117]

[0118] Table 1 shows that the bacterial enzyme co-immobilized plastic waste charcoal material prepared from waste plastics can effectively degrade plastic fragments and reduce the amount of microplastics in the soil. The test results in Table 1 show that different embodiments have significant degradation effects on plastics, reducing plastic fragments (<1cm) by a small amount. 2 The proportion of larger plastic fragments (1-5cm) increased from 30.26% to 38.85%-40.45%. 2 and 5-10cm 2 The proportion of microplastics also decreased significantly, from 24.20% and 15.62% before treatment to 20.52%-21.35% and 12.98%-13.25%, respectively. This demonstrates that the microbial-enzyme co-immobilized plastic waste charcoal of the present invention can degrade residual plastic fragments in waste-contaminated soil. Furthermore, the number of microplastics in the contaminated soil also decreased, from 3645 / kg before treatment (blank control) to 2641-2794 / kg, indicating that the microbial-enzyme co-immobilized plastic waste charcoal of the present invention can also adsorb microplastics in contaminated soil.

[0119] Furthermore, this invention can effectively treat organic pollutants in waste-contaminated soil. The results of the embodiments show that the application of bacterial enzyme co-immobilized plastic waste charcoal can achieve a degradation rate of 82.32%-89.46% for plastic additives after 3 months, which is more than 80% higher than that of untreated contaminated soil (blank control (3 months)). It can also effectively degrade antibiotics and polycyclic aromatic hydrocarbons, with degradation rates of 71.52%-75.64%, which is also more than 65% higher than that of untreated contaminated soil (blank control (3 months)).

[0120] Compared to Comparative Example 3 (without added bacterial enzymes), whether it was adding enzymes alone (Comparative Example 2), adding bacteria alone (Comparative Example 1), or even adding bacterial enzymes but without immobilization (Comparative Example 5), the immobilized plastic waste charcoal material obtained by this invention can improve the degradation of plastic fragments and the adsorption of microplastics. Simultaneously, the degradation rate of organic pollutants is also increased by more than 60% compared to Comparative Example 3. The application of bacterial enzyme co-immobilized plastic waste charcoal (Examples 1-4) shows significant effects on both plastic degradation and adsorption, and organic pollutant degradation compared to Comparative Example 3, especially the organic pollutant degradation rate, which is increased by up to 70%. This demonstrates that this invention can simultaneously and effectively achieve the degradation of plastics and the removal of organic pollutants from waste-contaminated soil.

[0121] Regarding the degradation of organic matter, the sum of the degradation rates of Comparative Example 1 (single added bacteria) and Comparative Example 2 (single added enzyme) was less than the degradation rate in Example 1; this indicates that the effect of the present invention is not simply the additive effect of bacteria + laccase, but also has a synergistic enhancement effect.

[0122] Compared with Comparative Example 6, the degradation effect of the present invention on plastics is basically the same, but the degradation efficiency of plastic additives, antibiotics and polycyclic aromatic hydrocarbons is greatly improved.

[0123] Example 5

[0124] The difference from Example 1 is that in step (6), after 3 months of contaminated soil remediation, immobilized plastic waste char is recovered: Contaminated soil is mixed with water (1.5 times the volume of soil), and stirred repeatedly with a magnetic rod until no adsorption occurs. The recovered immobilized plastic waste char particles are then used again for the remediation of contaminated soil from the same source, following the same application ratio and steps. The remediation steps are repeated 5 times. The remediation effects are compared, and the results are shown in Table 2.

[0125] Table 2. Detection results of indicators for multiple remediation of landfill-contaminated soil using microbial enzyme co-immobilized plastic waste char.

[0126]

[0127]

[0128] As shown in Table 2, the treatment effect decreases with increasing recycling frequency. However, the bacterial-enzyme co-immobilized plastic waste charcoal still achieves a certain degree of plastic degradation and adsorption after five recycling cycles, with an organic pollutant degradation rate exceeding 50%. Furthermore, the recovery rate of solid particles reaches 95.15% initially and 78.94% after five uses. This indicates that the bacterial-enzyme co-immobilized plastic waste charcoal prepared in this invention can be recycled and reused multiple times.

[0129] Magnetic particle recovery rate = (mass of recovered microbial enzyme-immobilized plastic waste charcoal particles / mass of microbial enzyme-immobilized plastic waste charcoal particles before use) × 100%;

[0130] The percentage of plastic fragments by size = (number of plastic fragments / total number of plastic fragments) × 100%;

[0131] Degradation rate of plastic additives = (concentration of plastic additives before treatment - concentration of plastic additives after treatment) / concentration of plastic additives before treatment × 100%;

[0132] Antibiotic degradation rate = (antibiotic concentration before treatment - antibiotic concentration after treatment) / antibiotic concentration before treatment × 100%;

[0133] Polycyclic aromatic hydrocarbon (PAH) degradation rate = (PAH concentration before treatment - PAH concentration after treatment) / PAH concentration before treatment × 100%.

[0134] The embodiments described above are merely preferred embodiments of the present invention and are not intended to limit the scope of the present invention. Various modifications and improvements made by those skilled in the art to the technical solutions of the present invention without departing from the spirit of the present invention should fall within the protection scope defined by the claims of the present invention.

Claims

1. A method for preparing microbial enzyme co-immobilized magnetic plastic waste charcoal, characterized in that, Includes the following steps: Plastic waste fragments in soil contaminated by garbage are crushed to obtain plastic pellets; The plastic particles were added to an alkaline solution, heated, and then pyrolyzed to obtain porous plastic waste charcoal. The porous plastic waste charcoal was mixed with nano-magnetic powder and placed in ammonia water for adsorption to obtain magnetic plastic waste charcoal. Degrading bacteria on the surface of plastics in waste-polluted soil were enriched and cultured using LB liquid medium, and then transferred to inorganic salt basal medium. After 3-5 transfers and acclimatizations, functional bacterial solutions were obtained. The functional bacterial culture was inoculated into LB liquid medium, and the magnetic plastic waste charcoal was added simultaneously for the expansion culture and adsorption immobilization of microorganisms. Then, solid and liquid were separated to obtain solid and culture medium. The solid was freeze-dried to obtain magnetic plastic waste charcoal immobilized microbial particles. The culture medium was mixed with an enzyme and a water-soluble polymer solution to obtain a cross-linking agent-enzyme mixed buffer. The cross-linking agent-enzyme mixed buffer solution was mixed with the magnetic plastic waste char immobilized microbial particles by shaking and then freeze-dried to obtain the bacterial-enzyme co-immobilized magnetic plastic waste char.

2. The preparation method according to claim 1, characterized in that, The mass-to-volume ratio of the plastic particles to the alkaline solution is 1:1.5-4 (w / v); the alkaline solution is a 10% KOH solution; the heating treatment is specifically ultrasonic heating at 45-60℃ for 1-2 hours; the pyrolysis is specifically pyrolysis at 350-650℃ for 1-4 hours under N2 environment.

3. The preparation method according to claim 1, characterized in that, The ammonia water has a mass concentration of 5%; the mass ratio of the porous plastic waste charcoal to the nano-magnetic powder is 1-3:1; the nano-magnetic powder is at least one of zero-valent Fe, Fe2O3, and Fe3O4; the adsorption is specifically performed at 25℃ and 180r / min for 8-12h.

4. The preparation method according to claim 1, characterized in that, The enrichment culture conditions are 30℃, 180r / min, 24h; the culture is transferred to an inorganic salt basal medium at a volume concentration of 1%-5%; the inorganic salt basal medium uses plastic waste fragments at a mass concentration of 0.5%-1% as the sole carbon source; the acclimatization conditions are 30℃, 180r / min, 5-10 days / time.

5. The preparation method according to claim 1, characterized in that, The functional bacterial solution was inoculated into LB liquid medium at a volume concentration of 5%-10%; the magnetic plastic waste charcoal had a mass concentration of 5%-15% in the LB liquid medium; the conditions for the expansion culture and adsorption fixation of the microorganisms were 25-30℃, 180r / min, and 18-24h.

6. The preparation method according to claim 1, characterized in that, The concentration of the enzyme in the culture medium is 0.5-10 mg / mL; the enzyme is at least one of laccase, catalase, protease, cellulase, and ligninase; the water-soluble polymer solution is a 1.5-3 g / L water-soluble polymer PBS buffer solution; the water-soluble polymer is one of polyacrylamide, polyethyleneimine, polyethylene glycol, carboxymethyl cellulose, and polyvinyl alcohol; the volume ratio of the culture medium to the water-soluble polymer solution is 1:2-4; the mass concentration of the magnetic plastic waste char immobilized microbial particles in the crosslinking agent-enzyme mixed buffer is 3%-12%; the mixture is ultrasonically dispersed for 3-5 min before shaking and mixing; the shaking and mixing conditions are 25℃, 180 r / min, and shaking and mixing for 2-8 h.

7. A microbial enzyme co-immobilized magnetic plastic waste charcoal prepared by the preparation method according to any one of claims 1-6.

8. The application of the bacterial enzyme co-immobilized magnetic plastic waste charcoal as described in claim 7 in the remediation of waste-contaminated soil.

9. A method for remediating waste-contaminated soil, characterized in that, The remediation of waste-contaminated soil is achieved by mixing the microbial enzyme co-immobilized magnetic plastic waste charcoal as described in claim 7 with the waste-contaminated soil.

10. The method for remediating waste-contaminated soil according to claim 9, characterized in that, The amount of the bacterial enzyme co-immobilized magnetic plastic waste charcoal added is 0.5%-5% of the mass of the waste-contaminated soil.

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