A method for degrading microplastics in aquaculture wastewater and its application

Abstract

The application relates to the technical field of wastewater treatment, and particularly discloses a degradation method and application of microplastics in aquaculture wastewater. The degradation method comprises the steps of preparation of a mesoporous structure magnetic immobilized material, pretreatment, adsorption-enzymolysis, microbial degradation, magnetic separation, precipitation and the like. The shell layer and the mesoporous structure of the mesoporous material are used to efficiently immobilize composite enzymes, so that enzyme loss is avoided, and the composite enzymes are used for targeted degradation of PET, PE and PP, and the total microplastic removal rate is more than 94.5%. The composite heat-resistant bacteria relay enzymolysis product is deepened and mineralized, and a phased treatment agent is matched to enhance mass transfer, so that the microbial activity is increased by 3 times. The mesoporous material is protected by the shell layer, the enzyme activity retention rate is more than 85%, the effluent COD is less than or equal to 50 mg / L, meets the discharge requirements, is suitable for microplastic pollution treatment in multiple scenes, and has the advantages of high efficiency, economy and environmental friendliness.

Description

Technical Field

[0001] This invention relates to the field of wastewater treatment technology, and more specifically to a method and application for the degradation of microplastics in aquaculture wastewater. Background Technology

[0002] With the rapid development of aquaculture, microplastic pollution in aquaculture wastewater has become an increasingly prominent problem. Due to their small particle size and large specific surface area, microplastics easily adsorb pollutants and accumulate in organisms, posing a potential threat to the ecological environment and human health. Currently used methods for microplastic treatment include physical adsorption, chemical oxidation, and biodegradation, but these methods suffer from low efficiency, high cost, and potential secondary pollution.

[0003] Biodegradation methods have attracted widespread attention due to their environmental friendliness and sustainability. Several methods for degrading microplastics using enzymes and microorganisms have been developed in the prior art; however, these methods are typically limited to specific types of microplastics, have limited degradation efficiency, and the enzymes and microorganisms are difficult to recycle.

[0004] Magnetic nanomaterials have shown great potential in environmental remediation due to their ease of separation and recycling. However, existing magnetic immobilized materials often suffer from problems such as low enzyme loading, poor stability, and unsatisfactory recycling efficiency. Furthermore, single types of enzymes or microorganisms are insufficient to address the complex and diverse microplastic pollution, especially the mixed pollution of multiple microplastics such as PET, PE, and PP commonly found in aquaculture wastewater.

[0005] Therefore, it is of great significance to develop a method for treating aquaculture wastewater that can efficiently degrade various microplastics and recycle materials. Summary of the Invention

[0006] The purpose of this invention is to provide a method and application for the degradation of microplastics in aquaculture wastewater, so as to solve the problems mentioned in the background art.

[0007] To achieve the above objectives, the present invention provides a method for degrading microplastics in aquaculture wastewater, comprising the following steps:

[0008] (1) Pre-treat the aquaculture wastewater to remove large particulate suspended solids;

[0009] (2) The pretreated aquaculture wastewater is introduced into the adsorption-enzymatic hydrolysis reaction zone, and mesoporous magnetic immobilization material is added. The microplastics are adsorbed and enzymatically hydrolyzed under the conditions of 35-40℃ and pH 6.5-7.5, and the wastewater is discharged to the next process.

[0010] (3) The effluent from step (2) is introduced into the microbial degradation zone and degraded by a composite heat-resistant bacterial agent at 50-55℃, and then discharged to the next process.

[0011] (4) The magnetic immobilization material of the effluent medium pore structure in step (3) is magnetically separated, recycled and reused to obtain the separated effluent;

[0012] (5) The effluent after separation is subjected to sedimentation treatment, and the supernatant is discharged or reused.

[0013] Preferably, in step (1), the pretreatment includes bar screen filtration and sedimentation. The bar screen aperture is 1-5 mm, the sedimentation time is 1-2 h, and the SS of the effluent after pretreatment is ≤50 mg / L. Large particulate impurities (such as feed residue, aquaculture excrement, silt, etc.) in the wastewater are separated by physical means (bar screen filtration + sedimentation), eliminating interference sources for subsequent reactions, preventing large particles from clogging the pores of the mesoporous magnetic material (ensuring the exposure of adsorption sites), competing for substrates for enzymes and microorganisms, and preventing particle abrasion of the material structure. The SS of the effluent after pretreatment is ≤50 mg / L, providing a clean reaction environment for adsorption-enzymatic hydrolysis and microbial degradation, and improving the stability of subsequent processes.

[0014] Preferably, in step (2), the mesoporous magnetic immobilization material comprises a magnetic iron oxide nanorod core, a mesoporous silica intermediate layer, and a composite enzyme-sodium alginate-chitosan microcapsule outer shell layer, and the dosage of the mesoporous magnetic immobilization material is 1-3 g / L. The mesoporous magnetic immobilization material performs a dual function of "adsorption + enzymatic hydrolysis": the large specific surface area of ​​the mesoporous silica intermediate layer adsorbs microplastics, the nanorod core ensures the material dispersion, and the composite enzyme directionally breaks down the plastic molecular chains.

[0015] The outer shell of the composite enzyme-sodium alginate-chitosan microcapsule integrates the functions of enzyme immobilization, mass transfer regulation, activity protection, structural support, and synergistic linkage.

[0016] Enzyme immobilization: Prevents enzyme loss and ensures continuous enzymatic hydrolysis. Sodium alginate molecules contain a large number of carboxyl groups, which can form an "egg-box" gel network with calcium ions to encapsulate complex enzymes such as polyethylene terephthalate hydrolase (PETase) and methyl terephthalate monohydroxyethyl ester hydrolase (MHETase). The amino groups of the outer chitosan can form electrostatic interactions with the carboxyl groups of sodium alginate. At the same time, the gel structure is strengthened by glutaraldehyde cross-linking, forming a dual immobilization mechanism to prevent enzyme loss during wastewater flow. The enzyme encapsulation rate reaches over 90%.

[0017] Mass transfer regulation: Balancing substrate contact and structural protection to improve enzymatic hydrolysis efficiency. By controlling the thickness and pore size of the outer shell, efficient mass transfer between microplastics and enzymes is achieved while avoiding mesoporous layer blockage. The microcapsule particle size is optimized to 50-200 μm, corresponding to an outer shell thickness of only 5-20 μm. Combined with the 8-15 nm pore size of the mesoporous silica intermediate layer, a three-tiered mass transfer channel is formed: outer shell sieving – mesoporous layer adsorption – enzyme active site reaction. The outer shell allows microplastics and small molecule substrates to enter while blocking large particulate impurities (such as pretreatment residual suspended matter), preventing mesoporous blockage.

[0018] Activity Protection: The amino groups (basic) of chitosan and the carboxyl groups (acidic) of sodium alginate form a buffer pair, stabilizing the pH inside the microcapsules at 6.5-7.5. This prevents enzyme denaturation caused by pH fluctuations in aquaculture wastewater. The low thermal conductivity of the outer shell buffers the temperature difference between the adsorption-enzymatic reaction zone and the microbial degradation zone, preventing high temperatures from the heat-resistant bacterial zone from reaching the enzyme active site and reducing the impact of temperature fluctuations on the enzyme. The cationic properties of chitosan can adsorb heavy metal ions (such as Cu²⁺ and Zn²⁺ in aquaculture wastewater), reducing their damage to the enzyme active site.

[0019] Structural support: Extends service life and reduces costs. The cross-linked structure of sodium alginate-chitosan can encapsulate the mesoporous silica intermediate layer and Fe3O4 nanorod core, preventing structural damage during mechanical stirring under magnetic separation, while also resisting the impact of ultrasonic cleaning. The support material can be recycled more than 15 times.

[0020] Synergistic Integration: Promoting Substrate Transfer in Enzymatic Hydrolysis-Microbial Degradation. Fragments such as methyl terephthalate (MHET) and short-chain hydrocarbons produced by enzymatic hydrolysis can be slowly released through the pores of the outer shell, avoiding excessively high local product concentrations that inhibit enzyme activity. At the same time, it continuously provides a carbon source for thermostable bacteria such as Thermobifida fusca in the microbial degradation zone, resolving the contradiction between "accumulation of enzymatic hydrolysis products" and "insufficient microbial substrate".

[0021] Preferably, in step (2), the composite enzyme comprises PETase, MHETase, keratinase, PE degrading enzyme, and PP degrading enzyme, with an enzyme activity of 500-1000 U / g carrier, and the hydraulic retention time in the adsorption-enzymatic reaction zone is 3-5 h. The composite enzyme initially decomposes recalcitrant microplastics such as PET, PE, and PP into small molecule fragments (such as MHET and short-chain hydrocarbons), solving the problem of low efficiency in the direct degradation of intact microplastics by microorganisms.

[0022] Preferably, in step (2), the preparation method of the mesoporous magnetic immobilization material includes:

[0023] Magnetic iron oxide nanorods with a particle size of 30-80 nm were dispersed in an ethanol solution, and a silane coupling agent (APTES) was added. The mixture was reacted at 50-70 °C for 3-5 h. After centrifugation and washing, tetraethyl orthosilicate and hexadecyltrimethylammonium bromide were added. The mixture was reacted at 60-80 °C for 6-10 h. The template agent was removed by calcination to form a mesoporous silica intermediate layer with a pore size of 8-15 nm, thus obtaining the modified magnetic particles.

[0024] The modified magnetic particles were added to the composite enzyme solution and stirred at 35-40℃ for 1-3 hours for adsorption.

[0025] The adsorbed magnetic particles were added to a sodium alginate solution with a weight concentration of 2-3%, and a calcium chloride solution with a weight concentration of 3-6% was added dropwise to form microcapsules with a particle size of 50-200 μm.

[0026] The microcapsules were transferred into a mixed solution containing 2-3% chitosan and 0.3-0.8% glutaraldehyde by weight, crosslinked and cured for 1-3 hours, washed and dried to obtain the mesoporous magnetic immobilized material.

[0027] Preferably, in step (3), the compound thermoresistant bacterial agent comprises *Thermobifida fusca*, *Geobacillus thermodenitrificans*, *Bacillus thermophilus*, and *Anoxybacillus flavithermus*, in a ratio of (30-40):(20-30):(15-25):(15-25), with an addition amount of (1-9.5) × 10⁻⁶. 9 CFU / L, hydraulic retention time 5-7h. The composite heat-resistant bacterial agent uses small molecule fragments generated by enzymatic hydrolysis as a carbon source, and undergoes metabolic mineralization at 50-55℃ (the suitable temperature for heat-resistant bacteria). The enzymatically hydrolyzed fragments are further converted into CO2, water, and small molecule organic matter, completely eliminating microplastic pollution; a hydraulic retention time of 5-7h ensures the mineralization rate.

[0028] Preferably, the treatment agents are added in stages in steps (2) and (3): rhamnolipin 50-80 mg / L is added in the enzymatic hydrolysis stage, and sophorolipid 20-50 mg / L is added in the microbial degradation stage. Rhamnolipin is added in the enzymatic hydrolysis stage (to enhance the hydrophilicity of microplastics and promote enzyme-substrate contact), and sophorolipid is added in the microbial stage (to activate the metabolic enzymes of the microbial agent and accelerate the utilization of fragments), with the dosage adapted to the needs of the two stages.

[0029] Preferably, in step (4), the magnetic separation uses a high-gradient magnetic separator with a magnetic field strength of 1.0-1.5T; the recovered material is reused after ultrasonic cleaning and enzyme activity regeneration. The high-gradient magnetic separator (1.0-1.5T) uses the strong magnetic responsiveness of Fe3O4 nanorods to separate the material; ultrasonic cleaning (300-500W, 5-15min) removes impurities and residual fragments attached to the material surface, and enzyme activity regeneration restores enzyme activity.

[0030] Preferably, in step (5), polyaluminum chloride is added during sedimentation at a dosage of 3-8 mg / L, with a sedimentation time of 0.5-2 h, resulting in an effluent COD ≤ 50 mg / L. The addition of polyaluminum chloride flocculates residual fine suspended solids, a small amount of unseparated material, and microbial metabolites, achieving solid-liquid separation after sedimentation for 0.5-2 h. The effluent COD ≤ 50 mg / L meets the "Fishery Water Quality Standard" (GB 11607-1989) and can be directly discharged or reused for aquaculture replenishment; the amount of sedimented sludge is small (≤ 0.1 g / L), resulting in low subsequent treatment costs.

[0031] On the other hand, the present invention also discloses the application of the above method in the treatment of aquaculture wastewater. Specifically, aquaculture wastewater can be wastewater from pig farms, cattle farms, chicken farms, and aquaculture, etc.

[0032] The beneficial effects of this invention are as follows:

[0033] Through the synergistic effect of mesoporous magnetic immobilization materials (Fe3O4 core, mesoporous SiO2 layer, composite enzyme-sodium alginate-chitosan shell), five composite enzymes, four composite thermoresistant bacteria, and staged treatment agents, significant beneficial effects are achieved: the shell layer and mesoporous structure of the mesoporous material efficiently immobilize the composite enzymes, preventing enzyme loss. Combined with the targeted degradation of PET, PE, and PP by the composite enzymes, the total microplastic removal rate reaches over 94.5%; the composite thermoresistant bacteria relay the enzymatic hydrolysis products to deepen mineralization, and the staged treatment agents enhance mass transfer, increasing microbial activity by 3 times; the mesoporous material, protected by the shell layer, retains over 85% of enzyme activity, and the effluent COD ≤ 50 mg / L meets discharge requirements. It is suitable for microplastic pollution treatment in multiple scenarios, combining high efficiency, economy, and environmental friendliness. Detailed Implementation

[0034] The following provides a detailed description of specific embodiments of the present invention. It should be understood that the specific embodiments described herein are for illustrative and explanatory purposes only and are not intended to limit the scope of the invention.

[0035] It should be noted that all reagents and raw materials used in this invention are commercially available, and the reagents are of analytical grade.

[0036] Example 1 (The following components are in parts by weight)

[0037] (1) Preparation of mesoporous magnetic immobilization materials:

[0038] One part of magnetic iron oxide nanorods with an average particle size of 50 nm was dispersed in 100 parts of 95% ethanol solution, 0.7 parts of APTES were added, and the mixture was reacted at 60 °C for 4 h. After centrifugation and washing, 100 parts of deionized water, 2 parts of tetraethyl orthosilicate and 0.7 parts of hexadecyltrimethylammonium bromide were added, and the mixture was reacted at 70 °C for 8 h. The template agent was removed by calcination at 550 °C for 4 h, forming a mesoporous silica intermediate layer with an average pore size of 12 nm, thus obtaining the modified magnetic particles.

[0039] The modified magnetic particles were added to 50 parts of a compound enzyme solution (total concentration 1.5 mg / mL, PETase:MHETase:keratase (EC 3.1.1.74):PE degrading enzyme (manganese peroxidase):PP degrading enzyme (glutathione S-transferase) = 3:2:2:1:1 (weight ratio)). The mixture was stirred and adsorbed at 38 °C for 2 h. After the adsorption was completed, the mixture was placed in a centrifuge and centrifuged at 8000 rpm for 10 min. The lower precipitate was collected to obtain the adsorbed magnetic particles.

[0040] The adsorbed magnetic particles were added to 50 parts of a 2% sodium alginate solution and then dropped into a 5% calcium chloride solution using electrostatic spraying (voltage 18kV, flow rate 0.8mL / h, distance 12cm) to form microcapsules with an average particle size of 120μm.

[0041] Microcapsules were transferred to 50 parts of a mixed solution containing 2% chitosan and 0.5% glutaraldehyde by weight, crosslinked and cured for 2 hours, washed and dried to obtain a mesoporous magnetic immobilized material.

[0042] (2) Pretreatment: Take 1 m³ of wastewater from a pig farm, filter the wastewater through a 3 mm screen, let it settle for 1.5 h, and the effluent SS = 42 mg / L;

[0043] (3) Adsorption-enzymatic hydrolysis: The pretreated aquaculture wastewater is introduced into the adsorption-enzymatic hydrolysis reaction zone, and mesoporous magnetic immobilization material (addition amount 2g / L) is added. Microplastics are adsorbed and enzymatically hydrolyzed under the conditions of 38℃ and pH 7.0, with a hydraulic retention time of 4h, and rhamnolipid (addition amount 60mg / L) is added.

[0044] (4) Microbial degradation: The effluent from step (3) is introduced into the microbial degradation zone, and a compound thermostable bacterial agent (Thermobifida fusca: Geobacillus thermodenitrificans: Bacillus thermophilus: Anoxybacillus flavithermus = 35:25:20:20 (quantity ratio), with a total dosage of 2.4 × 10⁻⁶) is added. 9 The enzymatically hydrolyzed microplastics were degraded at 52℃ with a hydraulic retention time of 6 hours, and sophiopogonin (dosage 30 mg / L) was added.

[0045] (5) Magnetic separation: The magnetic material in the effluent from step (4) is recovered by a high gradient magnetic separator (magnetic field strength 1.2T, time 10min); the recovered material is then ultrasonically cleaned (400W, 10min).

[0046] (6) Sedimentation: Add polyaluminum chloride (5 mg / L) to the separated effluent, and let it settle for 1 hour. The effluent COD is 43 mg / L.

[0047] Example 2 (The following components are in parts by weight)

[0048] (1) Preparation of mesoporous magnetic immobilization materials:

[0049] One part of magnetic iron oxide nanorods with an average particle size of 30 nm was dispersed in 100 parts of 95% ethanol solution, 0.5 parts of APTES were added, and the mixture was reacted at 50 °C for 3 h. After centrifugation and washing, 100 parts of deionized water, 1 part of tetraethyl orthosilicate and 0.5 parts of hexadecyltrimethylammonium bromide were added, and the mixture was reacted at 60 °C for 6 h. The template agent was removed by calcination at 550 °C for 4 h, forming a mesoporous silica intermediate layer with an average pore size of 8 nm, thus obtaining the modified magnetic particles.

[0050] The modified magnetic particles were added to 50 parts of a compound enzyme solution (total concentration 1 mg / mL, PETase:MHETase:keratase (EC 3.1.1.74):PE degrading enzyme (manganese peroxidase):PP degrading enzyme (glutathione S-transferase) = 3:2:2:1:1). The mixture was stirred and adsorbed at 35℃ for 1 h. After the adsorption was completed, the mixture was placed in a centrifuge and centrifuged at 8000 rpm for 10 min. The lower precipitate was collected to obtain the adsorbed magnetic particles.

[0051] The adsorbed magnetic particles were added to 50 parts of a 2% sodium alginate solution and then dropped into a 3% calcium chloride solution using electrostatic spraying (voltage 15kV, flow rate 0.5mL / h, distance 10cm) to form microcapsules with an average particle size of 50μm.

[0052] Microcapsules were transferred to 50 parts of a mixed solution containing 2% chitosan and 0.3% glutaraldehyde by weight, crosslinked and cured for 1 hour, washed and dried to obtain a mesoporous magnetic immobilized material.

[0053] (2) Pretreatment: Take 1 m³ of wastewater from a chicken farm, filter the wastewater through a 1 mm screen, let it settle for 1 h, and the effluent SS = 48 mg / L;

[0054] (3) Adsorption-enzymatic hydrolysis: The pretreated aquaculture wastewater is introduced into the adsorption-enzymatic hydrolysis reaction zone, and mesoporous magnetic immobilization material (addition amount 1g / L) is added. Microplastics are adsorbed and enzymatically hydrolyzed under the conditions of 35℃ and pH 6.5, with a hydraulic retention time of 3h, and rhamnolipin (addition amount 50mg / L) is added.

[0055] (4) Microbial degradation: The effluent from step (3) is introduced into the microbial degradation zone, and a compound thermostable bacterial agent (Thermobifida fusca: Geobacillus thermodenitrificans: Bacillus thermophilus: Anoxybacillus flavithermus = 30:20:15:15 (quantity ratio) is added at a dosage of 1.2 × 10⁻⁶. 9 The microplastics after enzymatic hydrolysis were degraded at 50℃ with a hydraulic retention time of 5 hours, and then sophorolipid (dosage 20 mg / L) was added.

[0056] (5) Magnetic separation: The magnetic material in the effluent from step (4) is recovered by a high gradient magnetic separator (magnetic field strength 1.0T, time 8min), with a separation efficiency of 97.3%; the recovered material is then ultrasonically cleaned (300W, 5min).

[0057] (6) Sedimentation: Add polyaluminum chloride (3 mg / L) to the separated effluent, and let it settle for 0.5 h. The effluent COD is 46 mg / L.

[0058] Example 3 (The following components are in parts by weight)

[0059] (1) Preparation of mesoporous magnetic immobilization materials:

[0060] One part of magnetic iron oxide nanorods with an average particle size of 80 nm was dispersed in 100 parts of 95% ethanol solution, one part of APTES was added, and the reaction was carried out at 70 °C for 5 h. After centrifugation and washing, 100 parts of deionized water, 3 parts of tetraethyl orthosilicate and 1 part of hexadecyltrimethylammonium bromide were added, and the reaction was carried out at 80 °C for 10 h. The template agent was removed by calcination at 550 °C for 4 h, forming a mesoporous silica intermediate layer with an average pore size of 15 nm, and the modified magnetic particles were obtained.

[0061] The modified magnetic particles were added to 50 parts of a compound enzyme solution (total concentration 2 mg / mL, PETase:MHETase:keratase (EC 3.1.1.74):PE degrading enzyme (manganese peroxidase):PP degrading enzyme (glutathione S-transferase) = 3:2:2:1:1). The mixture was stirred and adsorbed at 40℃ for 3 h. After the adsorption was completed, the mixture was placed in a centrifuge and centrifuged at 8000 rpm for 10 min. The lower precipitate was collected to obtain the adsorbed magnetic particles.

[0062] The adsorbed magnetic particles were added to 50 parts of a 3% sodium alginate solution and then dropped into a 6% calcium chloride solution using electrostatic spraying (voltage 20kV, flow rate 1mL / h, distance 15cm) to form microcapsules with an average particle size of 200μm.

[0063] Microcapsules were transferred into 50 parts of a mixed solution containing 3% chitosan and 0.8% glutaraldehyde by weight, crosslinked and cured for 3 hours, washed and dried to obtain a mesoporous magnetic immobilized material.

[0064] (2) Pretreatment: Take 1m³ of wastewater from a cattle farm, filter the wastewater through a 5mm screen, let it settle for 2 hours, and the effluent SS=35mg / L;

[0065] (3) Adsorption-enzymatic hydrolysis: The pretreated aquaculture wastewater is introduced into the adsorption-enzymatic hydrolysis reaction zone, and mesoporous magnetic immobilization material (addition amount 1g / L) is added. Microplastics are adsorbed and enzymatically hydrolyzed under the conditions of 40℃ and pH 7.5, with a hydraulic retention time of 5h, and rhamnolipid (addition amount 80mg / L) is added.

[0066] (4) Microbial degradation: The effluent from step (3) is introduced into the microbial degradation zone, and a compound thermostable bacterial agent (Thermobifida fusca: Geobacillus thermodenitrificans: Bacillus thermophilus: Anoxybacillus flavithermus = 40:30:25:25 (quantity ratio), dosage 3.6×10 9The microplastics after enzymatic hydrolysis were degraded at 55℃ with a hydraulic retention time of 7 h, and sophiopogonin (dosage 50 mg / L) was added.

[0067] (5) Magnetic separation: The magnetic material in the effluent from step (4) is recovered by a high gradient magnetic separator (magnetic field strength 1.5T, time 9min); the recovered material is then ultrasonically cleaned (500W, 15min).

[0068] (6) Sedimentation: Add polyaluminum chloride (8 mg / L) to the separated effluent, and let it settle for 2 hours. The effluent COD is 38 mg / L.

[0069] Example 4 (The following components are in parts by weight)

[0070] (1) Preparation of mesoporous magnetic immobilization materials:

[0071] One part of magnetic iron oxide nanorods with an average particle size of 60 nm was dispersed in 100 parts of 95% ethanol solution, 0.8 parts of APTES were added, and the reaction was carried out at 65 °C for 4.5 h. After centrifugation and washing, 100 parts of deionized water, 2.5 parts of tetraethyl orthosilicate and 0.8 parts of hexadecyltrimethylammonium bromide were added, and the reaction was carried out at 75 °C for 9 h. The template agent was removed by calcination at 550 °C for 4 h, forming a mesoporous silica intermediate layer with an average pore size of 10 nm, and the modified magnetic particles were obtained.

[0072] The modified magnetic particles were added to 50 parts of a complex enzyme solution (total concentration 1.8 mg / mL, PETase:MHETase:keratase (EC 3.1.1.74):PE degrading enzyme (manganese peroxidase):PP degrading enzyme (glutathione S-transferase) = 3:2:2:1:1). The mixture was stirred and adsorbed at 37℃ for 2.5 h. After the adsorption was completed, the mixture was placed in a centrifuge and centrifuged at 8000 rpm for 10 min. The lower precipitate was collected to obtain the adsorbed magnetic particles.

[0073] The adsorbed magnetic particles were added to 50 parts of a 2.5% sodium alginate solution and then dropped into a 4.5% calcium chloride solution using electrostatic spraying (voltage 19kV, flow rate 0.9mL / h, distance 14cm) to form microcapsules with an average particle size of 150μm.

[0074] Microcapsules were transferred to 50 parts of a mixed solution containing 2.5% chitosan and 0.6% glutaraldehyde by weight, crosslinked and cured for 2.5 h, washed and dried to obtain a mesoporous magnetic immobilized material.

[0075] (2) Pretreatment: Take 1 m³ of aquaculture wastewater, filter the wastewater through a 3 mm screen, let it settle for 1.5 h, and the effluent SS = 40 mg / L;

[0076] (3) Adsorption-enzymatic hydrolysis: The pretreated aquaculture wastewater was introduced into the adsorption-enzymatic hydrolysis reaction zone, and mesoporous magnetic immobilization material (addition amount 2g / L) was added. Microplastics were adsorbed and enzymatically hydrolyzed under the conditions of 37℃ and pH 7.2, with a hydraulic retention time of 4h, and rhamnolipid (addition amount 70mg / L) was added.

[0077] (4) Microbial degradation: The effluent from step (3) is introduced into the microbial degradation zone, and a compound thermostable bacterial agent (Thermobifida fusca: Geobacillus thermodenitrificans: Bacillus thermophilus: Anoxybacillus flavithermus = 38:28:22:22 (quantity ratio) is added at a dosage of 3.0 × 10⁻⁶. 9 The enzymatically hydrolyzed microplastics were degraded at 53℃ with a hydraulic retention time of 6 hours, and sophiopogonin (dosage 40 mg / L) was added.

[0078] (5) Magnetic separation: The magnetic material in the effluent from step (4) is recovered by a high gradient magnetic separator (magnetic field strength 1.3T, time 12min); the recovered material is then ultrasonically cleaned (350W, 13min).

[0079] (6) Sedimentation: Add polyaluminum chloride (6 mg / L) to the separated effluent, and let it settle for 1.5 h. The effluent COD is 41 mg / L.

[0080] Comparative Example 1

[0081] Step (1) is omitted. Spherical iron oxide is used to replace the mesoporous magnetic immobilization material in Example 1. The remaining components and steps are the same as in Example 1.

[0082] Comparative Example 2

[0083] In step (1), the complex enzyme is replaced with the single enzyme PETase, and the remaining components and steps are the same as in Example 1.

[0084] Comparative Example 3

[0085] In step (4), the composite heat-resistant bacterial agent is replaced with a single bacterium, Thermobifida fusca, and the remaining components and steps are the same as in Example 1.

[0086] Comparative Example 4

[0087] In step (3), rhamnolipin is not added, and in step (4), sophorolipid is not added. The remaining components and steps are the same as in Example 1.

[0088] The effluent from Examples 1-4 and Comparative Examples 1-4 was subjected to relevant tests. The test standards and methods are as follows, and the test results are recorded in Table 1.

[0089] 1. Microplastic removal rate

[0090] Testing standard: GB / T 39304-2020 Determination of microplastics in water by microscopic counting method.

[0091] Detection method: Take wastewater samples before and after treatment, and enrich microplastics by negative pressure filtration through a 0.45μm filter membrane; place the filter membrane under a microscope, identify and count the number of PET, PE and PP microplastics according to their morphology (particulate, fibrous, etc.); calculate the removal rate of each type and the total removal rate according to "(number before treatment - number after treatment) / number before treatment × 100%".

[0092] 2. Enzyme activity

[0093] Testing standard: GB / T 23195-2008 "Determination of Amylase Value in Honey by Ultraviolet Spectrophotometry".

[0094] Detection method: Take a sample of mesoporous magnetic immobilized material, add the corresponding substrate (PET powder, PP emulsion, etc.), and react at 38℃ and pH 7.0 for 1 h; centrifuge and collect the supernatant, and use a UV spectrophotometer to detect the absorbance of the product (such as terephthalic acid, short-chain fatty acid) at the characteristic wavelength (240-280 nm); calculate the enzyme activity by referring to the enzyme activity standard curve, and express it as "U / g carrier".

[0095] 3. Testing Standard: GB 4789.2-2016 "National Food Safety Standard - Microbiological Examination of Food: Determination of Total Colony Count"

[0096] Detection method: Take water samples from the microbial degradation zone, serially dilute them, spread them on LB solid medium, and incubate them in a constant temperature incubator at 50-55℃ for 24 hours; count the number of colonies, convert them to "CFU / mL" according to the dilution factor, and characterize the number of viable bacteria and microbial activity.

[0097] 4. Magnetic separation efficiency

[0098] Testing standard: Refer to HJ / T 201-2005 "Technical Requirements for Environmental Labeling Products - Magnetic Materials" (related to magnetic separation performance).

[0099] Test method: Record the mass of magnetic material added before magnetic separation (m1), collect the material after separation, dry and weigh it (m2); calculate the separation efficiency according to "m2 / m1×100%", and at the same time, observe the number of residual material particles in the effluent through a microscope to assist in the verification.

[0100] 5. Mass transfer efficiency

[0101] Testing standards: Refer to "Preparation and Application Technology of Immobilized Enzymes" (Industry-standard methods)

[0102] Detection method: Fluorescently labeled microplastic fragments were used as a simulated substrate and added to the adsorption-enzymatic reaction system; the fluorescence intensity inside the material was measured at regular intervals, and the rate of substrate diffusion into the material was calculated; the mass transfer efficiency was expressed in "cm / s", reflecting the contact rate between the substrate and the enzyme.

[0103] 6. Effluent COD

[0104] Testing Standard: HJ 828-2017 "Determination of Chemical Oxygen Demand in Water - Dichromate Method"

[0105] Detection method: Take the supernatant after precipitation treatment, add potassium dichromate digestion solution, and digest at 165℃ for 2 hours; after cooling, titrate with ferrous ammonium sulfate standard solution, calculate the COD value based on the volume consumed, and express the degree of organic pollution in the effluent in "mg / L".

[0106] Table 1 Test Results

[0107] index Example 1 Example 2 Example 3 Example 4 Comparative Example 1 Comparative Example 2 Comparative Example 3 Comparative Example 4 PET removal rate (%) 97.2 95.8 98.7 97.9 90.5 96.3 88.2 92.1 PE removal rate (%) 96.3 94.5 97.5 96.6 82.3 65.1 70.3 89.5 PP removal rate (%) 95.1 93.2 96.9 95.7 79.8 62.4 67.5 87.2 Total removal rate (%) 96.2 94.5 97.7 96.7 84.2 74.6 75.3 89.6 Enzyme activity retention rate (%) 88.5 85.4 90.3 89.1 58.7 87.9 88.4 86.3 Microbial activity (CFU / mL) <![CDATA[2.8×10 9 ]]> <![CDATA[2.2×10 9 ]]> <![CDATA[3.5×10 9 ]]> <![CDATA[3.0×10 9 ]]> <![CDATA[2.6×10 9 ]]> <![CDATA[2.7×10 9 ]]> <![CDATA[8.2×10 8 ]]> <![CDATA[2.4×10 9 ]]> Mass transfer efficiency (cm / s) <![CDATA[1.2×10 -4 ]]> <![CDATA[1.0×10 -4 ]]> <![CDATA[1.5×10 -4 ]]> <![CDATA[1.3×10 -4 ]]> <![CDATA[0.5×10 -4 ]]> <![CDATA[1.1×10 -4 ]]> <![CDATA[1.2×10 -4 ]]> <![CDATA[0.7×10 -4 ]]> Magnetic separation efficiency (%) 98.5 97.3 99.2 98.8 96.2 98.3 98.1 98.4 Effluent COD (mg / L) 43 46 38 41 51 58 63 49

[0108] As shown in the table above, Examples 1-4, based on the synergistic system of "mesoporous magnetic immobilization material + composite enzyme + composite bacteria + staged treatment agent," exhibit excellent microplastic degradation performance and process stability. The total microplastic removal rate of each example reached over 94.5%, with Example 3 achieving a total removal rate of 97.7% due to the highest material dosage and sufficient reaction time. The synergistic effect of the composite enzyme and composite bacteria enabled the removal rate of PE and PP to exceed 93%, solving the problem of weak degradation of polyolefin plastics by traditional methods. Simultaneously, the structural protection of the mesoporous material maintained the enzyme activity retention rate at 85.4%-90.3%, the magnetic separation efficiency remained stable at ≥97.3%, and the effluent COD was ≤46 mg / L, fully achieving the technical goals of "high-efficiency degradation - material recycling - water quality compliance."

[0109] Comparative Example 1, due to the use of conventional spherical magnetic particles (without a mesoporous silica interlayer and microcapsule shell), exhibits a significantly lower core performance compared to the Example 1. The mass transfer efficiency is only 0.5 × 10⁻⁻⁻⁴. 4The flow rate was less than half that of Example 1. Due to the lack of a large specific surface area and pore size channels in the mesoporous structure, the probability of contact between microplastics and enzymes decreased by 40%. The enzyme activity retention rate dropped from 88.5% (Example 1) to 58.7%. Due to the absence of a protective outer shell, the enzyme loss rate during circulation increased to over 35%. Although the PET removal rate still reached 90.5% (dependent on the direct action of PETase), the PE and PP removal rates were only 82.3% and 79.8%, respectively, with a total removal rate of 84.2%, 12 percentage points lower than in Example 1. The effluent COD increased to 51 mg / L, due to insufficient enzymatic hydrolysis leading to increased organic residue. The core difference lies in the lack of the crucial supporting role of the mesoporous structure in the "mass transfer-enzyme immobilization-structural protection" process.

[0110] Comparative Example 2, using only PETase, showed insufficient degradation capacity for PE and PP, becoming a performance bottleneck. The PET removal rate was 96.3%, close to that of Example 1 (97.2%), demonstrating the specific degradation activity of PETase; however, the PE and PP removal rates were only 65.1% and 62.4%, respectively, 31.2% and 32.7% lower than Example 1, with a total removal rate of 74.6%, less than the 78% of Example 1. This difference directly stems from the functional deficiencies of the composite enzyme system: the surface barrier-breaking effect of keratinase and the specific cleavage function of the PE / PP degrading enzyme were not demonstrated, resulting in the ineffective decomposition of polyolefin plastics, leaving a large amount remaining in the water; simultaneously, due to the lack of MHETase for subsequent degradation, the PET enzymatic hydrolysis product MHET accumulated, indirectly increasing the effluent COD to 58 mg / L. It is evident that the synergistic network of "barrier breaking-specific degradation-relay refinement" of the composite enzyme is the core for achieving comprehensive degradation of multiple types of microplastics.

[0111] Comparative Example 3, due to the addition of only a single Thermobifida fusca inoculant, showed severely insufficient synergistic mineralization capacity in the microbial degradation process. Microbial activity decreased from 2.8 × 10⁻⁶. 9 The CFU / mL (Example 1) dropped sharply to 8.2 × 10⁻⁶. 8 The CFU / mL concentration was low because a single bacterium cannot adapt to multi-substrate environments and lacks metabolic complementarity with other bacteria (such as the degradation ability of *Anoxybacillus flavithermus* for PE / PP fragments). The total microplastic removal rate was 75.3%, 20.9 percentage points lower than in Example 1. The removal rates for PE and PP were only 70.3% and 67.5%, respectively, because *Thermobifida fusca* primarily targets PET fragments and has extremely low utilization of polyolefin fragments. Furthermore, the single bacterium's metabolites were limited, leading to an effluent COD of 63 mg / L, the highest among all test groups. This confirms that the "functional specialization-synergistic metabolism" characteristics of the compound bacterial agent are key to deepening the mineralization of enzymatic hydrolysis products and reducing effluent pollution.

[0112] Comparative Example 4, lacking the addition of rhamnolipids and sophorolipids, showed a significant impact on both mass transfer and degradation efficiency. The mass transfer efficiency decreased from 1.2 × 10⁻ ... 4 cm / s (Example 1) decreased to 0.7 × 10⁻ 4 The microplastic removal rate was 89.6%, 6.6 percentage points lower than in Example 1, due to insufficient hydrophilicity caused by the absence of rhamnolipids and sophorolipids, making it difficult for them to fully contact enzymes and microorganisms. The total microplastic removal rate was 89.6%, 6.6 percentage points lower than in Example 1, with the most significant decreases in PE and PP removal rates (6.8% and 7.9% lower, respectively), because the non-polar characteristics of polyolefin plastics make them more dependent on rhamnolipids and sophorolipids. Although the enzyme activity retention rate (86.3%) and magnetic separation efficiency (98.4%) fluctuated relatively little, the decrease in surface contact efficiency directly limited the overall degradation performance, and the effluent COD increased to 49 mg / L. The difference indicates that the staged addition of rhamnolipids and sophorolipids can effectively improve the synergistic efficiency of enzymatic hydrolysis and microbial degradation by "enhancing hydrophilicity and promoting mass transfer".

[0113] The preferred embodiments of the present invention have been described in detail above. However, the present invention is not limited to the specific details of the above embodiments. Within the scope of the technical concept of the present invention, various simple modifications can be made to the technical solutions of the present invention, and these simple modifications all fall within the protection scope of the present invention. Furthermore, it should be noted that the various specific technical features described in the above embodiments can be combined in any suitable manner without contradiction. To avoid unnecessary repetition, the present invention will not further describe the various possible combinations.

[0114] Furthermore, various different embodiments of the present invention can be combined in any way, as long as they do not violate the spirit of the present invention, they should also be regarded as the content disclosed by the present invention.

Claims

1. A method for degrading microplastics in aquaculture wastewater, characterized by, The method comprises the following steps: (1) pretreating the aquaculture wastewater to remove large particle suspended solids; (2) introducing the pretreated aquaculture wastewater into an adsorption-enzymolysis reaction zone, adding mesoporous structure magnetic immobilized material, adsorbing and enzymolyzing microplastics at 35-40 DEG C and pH 6.5-7.5, and simultaneously adding rhamnolipid, and discharging water to the next process; The mesoporous structure magnetic immobilized material comprises a magnetic ferroferric oxide nanorod core, a mesoporous silica intermediate layer and a composite enzyme-sodium alginate-chitosan microcapsule shell layer, the composite enzyme comprises polyethylene terephthalate hydrolase, p-toluene hydroxyethyl hydrolyze, cutinase, PE degradation enzyme and PP degradation enzyme, and the enzyme activity is 500-1000 U / g carrier; (3) introducing the water discharged in step (2) into a microbial degradation zone, degrading the enzymolyzed microplastics by a composite heat-resistant bacterial agent at 50-55 DEG C, and simultaneously adding sophorolipid, and discharging water to the next process; The composite heat-resistant bacterial agent comprises Thermobifida fusca, Geobacillus thermodenitrificans, Bacillus caldovelox and Bacillus caldolyticus, and the quantity ratio is (30-40):(20-30):(15-25):(15-25); (4) magnetically separating and recycling the mesoporous structure magnetic immobilized material in the water discharged in step (3) to obtain separated water; (5) precipitating the separated water, and discharging or recycling the supernatant.

2. The method of claim 1, wherein the microplastics are degraded in the aquaculture wastewater. In step (1), the pretreatment comprises grid filtration and sedimentation, the grid aperture is 1-5 mm, the sedimentation time is 1-2 h, and the SS of the water after pretreatment is less than or equal to 50 mg / L.

3. The method of claim 2, wherein the microplastics are degraded in the aquaculture wastewater. In step (2), the mesoporous structure magnetic immobilized material is added in an amount of 1-3 g / L.

4. The method of claim 3, wherein the microplastics are degraded in the aquaculture wastewater. In step (2), the hydraulic retention time of the adsorption-enzymolysis reaction zone is 3-5 h.

5. The method of claim 4, wherein the microplastics are degraded in the aquaculture wastewater. In step (2), the mesoporous structure magnetic immobilized material is prepared by the following method: The magnetic ferroferric oxide nanorod with a particle size of 30-80 nm is dispersed in an ethanol solution, a silane coupling agent is added, and the mixture is reacted at 50-70 DEG C for 3-5 h, then centrifuged and washed, and then tetraethyl orthosilicate and cetyltrimethylammonium bromide are added, and the mixture is reacted at 60-80 DEG C for 6-10 h, and then the template agent is removed by calcination to form a mesoporous silica intermediate layer with a pore size of 8-15 nm, thereby obtaining the modified magnetic particles; The modified magnetic particles are added into a composite enzyme solution, and stirred at 35-40 DEG C for 1-3 h to adsorb; The adsorbed magnetic particles are added into a sodium alginate solution with a weight concentration of 2-3%, and a calcium chloride solution with a weight concentration of 3-6% is dropped into the mixture to form microcapsules with a particle size of 50-200 mu m; The microcapsules are transferred into a mixed solution containing chitosan with a weight concentration of 2-3% and glutaraldehyde with a weight concentration of 0.3-0.8%, and crosslinked and solidified for 1-3 h, and then washed and dried to obtain the mesoporous structure magnetic immobilized material.

6. The method of claim 5, wherein the microplastics are degraded in the aquaculture wastewater. In step (3), the adding amount of the complex heat-resistant bacterial agent is (1-9.5)×10 9 CFU / L, and the hydraulic retention time is 5-7 h.

7. The method of claim 1, wherein the microplastics are degraded in the aquaculture wastewater. In steps (2) and (3), the rhamnolipid is added in an amount of 50-80 mg / L, and the sophorolipid is added in an amount of 20-50 mg / L.

8. The method of degradation of microplastics in aquaculture wastewater according to claim 1, characterized in that: In step (4), the magnetic separation uses high gradient magnetic separator with magnetic field strength of 1.0-1.5T; the recovered material is reused after ultrasonic cleaning and enzyme activity regeneration. 9.The method of claim 1, wherein the microplastics are degraded in the aquaculture wastewater. In step (5), polyaluminum chloride is added for precipitation treatment with dosage of 3-8mg / L, precipitation time of 0.5-2h, and effluent COD≤50mg / L.

10. Application of the degradation method of any one of claims 1-9 in aquaculture wastewater treatment.