Low-temperature wastewater decolorizing and purifying process based on biological enzyme catalysis

Abstract

The invention discloses a low-temperature wastewater decolorizing and purifying process based on biological enzyme catalysis. The process comprises the steps of wastewater pretreatment, layered dosing, low-temperature catalytic reaction, carrier recovery and reutilization and the like. According to the process, a layered enzyme loading system is adopted, an upper-layer carrier, a middle-layer carrier and a lower-layer carrier respectively form independent reaction areas in a reaction device, enzyme carriers are directionally added through top, middle and bottom dosing ports, and multi-layer synergistic decolorization is realized. Before operation, the temperature and density of wastewater are detected, and the carrier density is adjusted according to the middle-layer target buoyancy, so that the carrier keeps stable suspension in different batches of wastewater; in the reaction process, buoyancy is adjusted through the floating increasing particles or the counterweight particles, and layered balance is achieved. After the reaction is finished, the carrier is cleaned, dried and subjected to buoyancy retest, and the carrier with good activity can be reused. The process is simple in structure, stable in operation and low in energy consumption, is suitable for decoloration and purification of printing and dyeing and chemical wastewater in a low-temperature environment, and solves the problems of unstable layering of a biological enzyme carrier and low low-temperature reaction efficiency in the prior art.

Description

Technical Field

[0001] This invention relates to the fields of wastewater treatment and biocatalysis, specifically a low-temperature wastewater decolorization and purification process based on bio-enzyme catalysis. Background Technology

[0002] With the rapid development of industries such as printing and dyeing, chemicals, papermaking, and pharmaceuticals, industrial wastewater commonly contains complex organic dyes and their intermediates that are difficult to degrade naturally. These substances often possess conjugated double bonds, aromatic rings, and azo structures, exhibiting not only high color intensity but also strong chemical stability. This results in low decolorization rates, long treatment cycles, and significant fluctuations in effluent quality during conventional biological treatment processes. While traditional oxidation, coagulation sedimentation, and adsorption methods can remove color to some extent, their operation requires large amounts of chemical reagents or high temperatures, leading to high energy consumption and potential secondary pollution, making them unsuitable for long-term continuous operation. Especially in winter or in cold regions, where wastewater temperatures often fall below 15°C, the metabolic activity of most microbial systems decreases, significantly slowing down biochemical reaction rates. Conventional processes struggle to maintain stable treatment effects under low-temperature conditions.

[0003] In recent years, decolorization technology based on bio-enzyme catalysis has gradually become a research hotspot. Laccase, lignin peroxidase, manganese peroxidase, and azo reductase can all catalyze the decomposition of azo bonds and aromatic ring structures in dye molecules under mild conditions, offering advantages such as high efficiency, low energy consumption, and no secondary pollution. However, free enzymes have poor stability in solution, are easily deactivated by pH, temperature, and heavy metal ions, and are difficult to recover and reuse after the reaction, resulting in high operating costs and limited engineering application. To improve enzyme stability and recyclability, existing technologies mostly employ immobilized enzymes or carrier encapsulation methods. However, conventional immobilized carriers have uniform density and pore size, making it difficult to form natural stratification in the reaction system, leading to significant differences in reaction intensity in different regions. As the reaction time increases, the carrier often floats or deposits, gradually destabilizing the enzyme reaction zone and thus reducing the overall decolorization efficiency.

[0004] Furthermore, the density and temperature of wastewater fluctuate with changing operating conditions, and traditional immobilized carriers cannot adaptively adjust their buoyancy accordingly. If the carrier density is too low, it easily floats to the surface and cannot fully contact the dye molecules; if the density is too high, it settles to the bottom, affecting the reaction interface and causing enzyme catalytic layer shift. Especially in low-temperature environments, the increased viscosity of wastewater and uneven distribution of dissolved oxygen make it even more difficult for the carrier to remain in the ideal reaction layer, resulting in stratification disorder and decreased reaction efficiency. Although existing technologies propose using magnetic particles for recovery or multi-layer carrier composite structures, a dynamic buoyancy matching mechanism for wastewater density differences has not yet been established, and the carrier distribution still requires manual adjustment, resulting in insufficient stability and continuity.

[0005] In view of the above problems, a low-temperature wastewater decolorization and purification process based on bio-enzyme catalysis is proposed. Summary of the Invention

[0006] The purpose of this invention is to provide a low-temperature wastewater decolorization and purification process based on bio-enzyme catalysis to solve the problems mentioned in the background art.

[0007] To achieve the above objectives, the present invention provides the following technical solution: a low-temperature wastewater decolorization and purification process based on bio-enzyme catalysis, comprising the following steps: S1. Wastewater pretreatment: Wastewater containing organic coloring substances is filtered, and its pH and dissolved oxygen are adjusted before entering the reaction system; S2, Low-temperature bio-enzyme reaction: The pretreated wastewater is introduced into a low-temperature bio-enzyme reaction device and a bio-enzyme catalytic decolorization reaction is carried out at 10-15℃. S3. Layered configuration and synergistic reaction: The reaction system is equipped with multiple layered reaction zones, and enzyme carriers are arranged in different reaction zones to form multi-layered catalytic regions at different depths in the wastewater. The middle reaction zone detects the buoyancy of the carrier and adjusts the buoyancy of the carrier particles to keep the carrier stably suspended in the middle zone, thereby achieving layered synergistic decolorization. S4. Layered drug delivery control: The reaction system is equipped with multiple drug delivery ports corresponding to each reaction layer, which are used to directionally add enzyme carriers or auxiliary reaction components to different reaction layers to adjust the reaction activity and catalytic efficiency of each layer. S5. Regeneration and Effluent Treatment: After the reaction cycle is completed, the reaction system is regenerated and maintained, and purified effluent is obtained through subsequent filtration or membrane separation processes.

[0008] Preferably, in step S1, the wastewater pretreatment step includes dynamically adjusting the pH of the wastewater to control the pH of the reaction system within the weakly acidic to weakly alkaline range of 6.5 to 8.0. During the operation of the reaction system, the pH is slightly corrected according to the buoyancy state of the intermediate carrier, and the adjustment usually cannot exceed ±0.5. This ensures the catalytic activity of the biological enzymes while maintaining the suspension stability of the carrier. Dissolved oxygen concentration is monitored simultaneously during the adjustment process to prevent a decrease in the oxidation capacity of the reaction system.

[0009] Preferably, in step S2, the low-temperature bio-enzyme reaction step is catalyzed by a low-temperature resistant biological oxidoreductase, wherein the biological enzyme includes any one or a combination of bacterial laccase, azoreductase, or lignin peroxidase; the reaction system is operated in continuous or quasi-continuous flow mode at 10–15°C, and the dissolved oxygen concentration (2.0–6.0 mg / L), fluid circulation rate (0.2–1.0 m / s), and stirring shear intensity (50–300 s⁻¹) of the reaction solution are controlled. -1 The settings and controls are configured to keep the enzyme catalytic conditions in each layered reaction zone constant. During the reaction process, the temperature and dissolved oxygen parameters of the reaction solution are monitored online, and the circulation flow rate is adjusted to maintain the stability of the enzyme catalytic environment.

[0010] Preferably, in step S3, a floating enzyme carrier is provided in the upper region of the layered reaction system. The floating enzyme carrier uses an inert porous material with a density lower than that of the reaction solution as a substrate. A biological enzyme with surface oxidation activity is loaded on the substrate. The biological enzyme includes bacterial laccase, peroxidase, or a complex enzyme system thereof. The floating enzyme carrier is introduced into the upper region of the reaction system after surface cross-linking and curing treatment to form a stable floating layer below the liquid surface. The reaction conditions in the upper region are controlled by an external gas distribution device or a liquid surface disturbance device to control oxygen distribution and flow stability.

[0011] Preferably, in step S3, a suspended enzyme carrier is provided in the middle layer region of the layered reaction system. The enzyme carrier is based on a gel or composite porous material with a density similar to that of the reaction solution. A biological enzyme that can maintain its activity at low temperature is loaded on the substrate. The biological enzyme includes bacterial laccase, azo reductase, lignin peroxidase or a combination thereof. The middle layer region is equipped with a buoyancy detection and adjustment device, which is used to detect the buoyancy state of the suspended enzyme carrier in the reaction solution and adjust the buoyancy of the carrier particles accordingly, so that the carrier remains stably suspended in the middle layer region. The formulation process is achieved by adjusting the physical or chemical parameters within the system, including minute adjustments to fluid shear strength, solution density, or electrolyte concentration.

[0012] Preferably, in step S3, a high-density enzyme carrier is provided in the lower region of the layered reaction system. The high-density carrier is based on a corrosion-resistant inorganic or composite material, and a biological enzyme with reducing or redox combined activity is immobilized on the surface of the matrix. The biological enzyme includes azo reductase, peroxidase, or a co-carrier system thereof. The lower layer region maintains a uniform distribution of the carrier through a bottom guide or stirring structure, and the carrier is redispersed in the lower layer region after periodic backwashing to maintain reactivity.

[0013] Preferably, in step S4, before a single batch of wastewater enters the reaction system, the temperature, density, and salinity parameters of the batch of wastewater are detected, and the buoyancy of the enzyme carriers to be added to each layer is pre-adjusted accordingly. The outer wall of the reaction system is provided with multiple independent dosing ports along the height direction, corresponding to the upper, middle and lower regions respectively. The buoyancy-pre-adjusted buoyancy enzyme carriers, mid-layer suspended enzyme carriers, and high-density enzyme carriers were directionally added to their respective reaction layers through their corresponding dosing ports. Each dosing port is connected to an external carrier supply device via tiered dosing pipelines, and is activated by the control system according to the detected parameters. This enables the layered deployment and initial distribution control of different buoyancy carriers.

[0014] Preferably, after the reaction cycle is completed, the enzyme carrier in the reaction system is separated and recovered. The separation process includes sequential solid-liquid separation and stratification. By setting up layered screen components or graded collection structures, the upper floating carrier, the middle suspended carrier and the lower high-density carrier can be recovered to their respective storage units. The separated reaction liquid is polished after being filtered by sand, membrane separation or other filtration units before being discharged; The recovered enzyme carriers are washed and tested before being reused in the corresponding layers to prevent carrier particles from entering the effluent system.

[0015] Preferably, the buoyancy-reducing formulation system is used for density correction when the buoyancy of the enzyme-carrying particles is higher than the target value. This system includes iron oxide powder, barium sulfate powder, or fine-grained quartz sand as the main counterweight material, and a binder compatible with the enzyme carrier composite layer to form a stable structure. The binder can be a hydrophilic polymer such as polyvinyl alcohol, chitosan, or carboxymethyl cellulose, and its addition ratio is 1-5% of the total mass of the carrier. After mixing, it is allowed to stand and solidify at room temperature, so that the counterweight particles are evenly distributed inside or on the surface of the carrier. The composite carrier prepared by this formulation has a density increase of 0.03-0.10 g / cm³, which can effectively reduce the buoyancy of the carrier and make it stably stay at the target layer.

[0016] Preferably, the buoyancy-enhancing formulation system is used to lighten the enzyme-carrying particles when their buoyancy is lower than the target value. This system uses foamed polyethylene, hollow glass microspheres, foamed polypropylene microparticles, or expanded perlite as the main buoyancy-enhancing materials, supplemented with polyvinyl alcohol or chitosan as the binding matrix, and adds 0.1 to 0.3 parts of surfactant to enhance interfacial bonding. The buoyancy-enhancing materials account for 1 to 8% of the total mass of the carrier. After mixing, stirring, and cross-linking curing, a lightweight composite structure is formed. The density of the resulting carrier is reduced by 0.02 to 0.08 g / cm³, allowing it to remain stably suspended or rise slowly in the reaction solution to restore the suspension balance of the middle reaction zone.

[0017] Compared with existing technologies, the advantages of this invention are as follows: This invention uses a layered enzyme-carrying system, ensuring stable suspension of the upper, middle, and lower layers of carriers within different reaction depths, thereby achieving highly efficient catalysis through multi-point dosing. Before process operation, the carrier buoyancy is adjusted according to the wastewater temperature and density to maintain the suspension equilibrium of the middle layer carrier. During the reaction, the density is fine-tuned through buoyancy enhancement and weighting systems to prevent drift. After operation, the carrier can be reused after cleaning, drying, and re-measuring its buoyancy. This process operates stably under low-temperature conditions and has high decolorization efficiency, solving the problems of unstable enzyme carrier layering, low reaction rates, and difficulty in recycling in existing technologies. Detailed Implementation

[0018] The technical solution of the present invention will be clearly and completely described below with reference to the embodiments of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, and not all of them. All other embodiments obtained by those skilled in the art based on the embodiments of the present invention without creative effort are within the scope of protection of the present invention.

[0019] This invention provides a technical solution: a low-temperature wastewater decolorization and purification process based on bio-enzyme catalysis.

[0020] This embodiment provides a low-temperature wastewater decolorization and purification process based on bio-enzyme catalysis. The overall process flow includes a pretreatment stage, a low-temperature enzyme catalytic reaction stage, a stratified dosing and buoyancy control stage, a carrier separation and recovery stage, and an effluent polishing stage. The entire system operates continuously or semi-continuously in a closed reaction device. Inside the device, three reaction layers—upper, middle, and lower—naturally form according to different density ranges. Each layer independently corresponds to a different type of enzyme carrier to meet the decomposition requirements of dye-based organic pollutants under low-temperature conditions. Dosing ports for the top, middle, and bottom layers are respectively provided on the side walls of the reaction device for the directional addition of their respective enzyme carriers and auxiliary nutrients to different reaction layers. This ensures that the reaction proceeds spatially in stratified layers and temporally in steps, thereby improving the overall decolorization rate and stability of the system.

[0021] Before the process starts, the wastewater first enters the pretreatment unit for basic conditioning. The system selects whether to perform cooling treatment based on the actual influent temperature. When the influent temperature is around 15 degrees Celsius or below, it can directly enter the reaction stage without additional cooling. If the influent temperature is too high, it is slowly reduced to a suitable range of 10-15 degrees Celsius through heat exchange modules or natural cooling to ensure the spatial structure stability of enzyme molecules and prevent the active site from becoming unstable or the protein structure from becoming loose due to high temperature.

[0022] After temperature adjustment, the monitoring system detects and slowly adjusts the pH value to maintain the system's pH between 6.8 and 7.5. pH adjustment is performed using a step-by-step, small-dose dosing method to prevent local over-flushing; simultaneously, dissolved oxygen concentration is maintained within the range of 3–5 mg / L through micro-aeration or low-pressure aeration, thereby providing sufficient oxygen and a suitable environment for subsequent enzyme reactions.

[0023] After pretreatment, the wastewater enters the low-temperature enzyme reaction device. The system sequentially adds enzyme particles to different layers according to a preset program. The upper layer mainly contains enzymes with oxidizing properties, such as laccase or peroxidase; the middle layer contains composite enzyme carriers for deep degradation in a semi-suspended state; and the lower layer contains high-density carrier particles, primarily azoreductase, to treat deposited dyes and higher molecular weight intermediates. The fluid circulation rate, temperature, and dissolved oxygen concentration of each reaction layer are automatically regulated by the central control unit to maintain a dynamic balance between the three reaction zones.

[0024] Once the system reaches a stable operating stage, the wastewater undergoes thorough enzymatic decolorization and organic matter decomposition within the three-layer reaction zone, ultimately being discharged from the bottom and entering the carrier separation and recovery module. This module utilizes a combination of physical sieving and magnetic separation to recover the upper floating carrier, the middle suspended carrier, and the lower high-density carrier to their respective storage tanks. The recovered carrier can then be sent to the regeneration line for cleaning and buoyancy recalibration based on its enzyme activity test results, and reused after meeting the conditions for reuse.

[0025] The treated effluent then enters the polishing unit, where residual particles and colloidal substances are removed using sand filtration, membrane separation, or a combination of filtration methods, resulting in a stable and clear effluent. The entire process achieves highly efficient decolorization and purification under adjustable low-temperature conditions. Furthermore, the use of layered reactions and carrier recycling reduces reagent consumption and the risk of secondary pollution, demonstrating strong engineering adaptability and promotional value.

[0026] Specifically, before the wastewater enters the low-temperature enzyme reaction system, its initial pH needs to be dynamically adjusted to ensure that the enzyme molecules maintain their activity under stable conditions. In this embodiment, an automated pH control system is set up in the pretreatment stage to achieve slow adjustment. This system consists of a real-time detection probe, a micro-metering pump, and a conditioning liquid storage tank.

[0027] During operation, the detection probe measures the initial pH value of the wastewater in real time. When the pH is acidic (below 6.5), a weak alkaline solution is slowly added proportionally. Sodium hydroxide, sodium carbonate, or a compound alkaline solution can be used, with the concentration generally controlled between 1% and 3% by mass. For wastewater with a high pH (above 8.0), a diluted acid solution is used for neutralization. Diluted phosphoric acid, acetic acid, or citric acid solutions can be selected, with the concentration controlled between 0.5% and 2% by mass. All adjusting solutions are added intermittently and periodically, with each adjustment range controlled within the range of 0.1% to 0.3% to avoid sudden changes.

[0028] Once the system is running stably, the pH control unit and dissolved oxygen adjustment module work together. When the dissolved oxygen concentration is low, the pH value will slowly decrease, and the system will automatically delay the next acid-base adjustment cycle to prevent cumulative errors. Conversely, when the dissolved oxygen is too high, the acid-base adjustment frequency will increase accordingly to maintain the overall balance of the system.

[0029] For wastewater with strong buffering capacity (such as systems containing acetate or phosphate), a buffering component, such as a composite solution of sodium bicarbonate and sodium dihydrogen phosphate, can be added to the conditioning solution in a ratio controlled between 1:1 and 1:2 to form a weak buffering system, maintaining a stable pH within the range of 6.8 to 7.5 over a long period. Within this range, both oxidative and reductive enzymes can maintain high reaction efficiency, preventing irreversible inactivation of enzyme structures due to extreme pH.

[0030] After adjustment, the current pH and dissolved oxygen curve data are recorded by the central control unit as reference parameters for subsequent buoyancy adjustment and intermediate carrier compounding, so as to keep the initial conditions of the entire process consistent.

[0031] Specifically, in step S2, the bioenzyme is distributed in an immobilized form in each layered region of the reaction system and forms a stable bond with the carrier material; The reaction system operates in continuous or quasi-continuous flow mode at 10–15℃, with the linear velocity of the reaction liquid controlled at 0.2–1.5 cm / s, ensuring an average residence time of 20–90 minutes throughout the system. The dissolved oxygen concentration (2.0–6.0 mg / L), fluid circulation rate (0.2–1.0 m / s), and stirring shear intensity (50–300 s⁻¹) of the reaction liquid are controlled. -1 The settings and controls are configured to keep the enzyme catalytic conditions in each layered reaction zone constant. During the reaction process, the temperature and dissolved oxygen parameters of the reaction solution are monitored online, and the circulation flow rate is adjusted to maintain the stability of the enzyme catalytic environment.

[0032] In this embodiment, the low-temperature enzyme catalytic reaction stage is the core of the entire process. The key is to select enzymes that still have high catalytic activity under low-temperature conditions and to use an immobilization process to stably attach them to the carrier, thereby achieving continuous reaction and recycling.

[0033] In practical applications, the main enzymes available can be categorized into two main types: oxidative enzymes and reductant enzymes. Oxidative enzymes can include bacterial laccase or lignin peroxidase (LiP). The former has a strong decolorizing ability for various aromatic and azo dyes, while the latter is effective in degrading dye molecules containing hydroxybenzene rings and conjugated structures. Reductant enzymes, represented by azo reductases, are suitable for decomposing azo bonds in dark-colored or high-concentration dye wastewater. These enzymes can be used individually or in combination to form a synergistic system.

[0034] In a typical formulation, by weight, there are 10-20 parts laccase, 5-10 parts lignin peroxidase, and 10-15 parts azoreductase. Depending on the type of wastewater, a system that emphasizes oxidation (increasing the proportion of laccase) or a system that emphasizes reduction (increasing the proportion of azoreductase) can be selected to achieve a dynamic equilibrium in the redox reaction.

[0035] The choice of carrier has a significant impact on the stability of enzyme activity. In this embodiment, the carrier materials can be of the following types: porous natural materials: such as bentonite, calcium alginate gel, attapulgite, etc., with a surface pore size of 0.5-2 micrometers, which can adsorb enzyme molecules and provide appropriate mechanical support; composite polymers: polyvinyl alcohol (PVA), chitosan, carboxymethyl cellulose, etc., which are solidified by cross-linking agents to form flexible carriers; magnetic composite particles: with iron oxide or magnetic silicon as the core and an outer PVA-chitosan composite layer, used for subsequent magnetic separation and recovery.

[0036] A preferred immobilization formulation (by weight) may be: 15 parts laccase, 10 parts azo reductase, 30 parts PVA solution (10% by weight), 20 parts chitosan solution (5% by weight), 1-2 parts glutaraldehyde crosslinking agent, and deionized water to make up to 100 parts.

[0037] In preparation, various enzymes are first dissolved in buffer solution, then mixed thoroughly with PVA and chitosan solutions. A cross-linking agent is slowly added dropwise at room temperature, allowing the system to gradually form gel-like microspheres. After formation, the microspheres are incubated at 4°C for 12 hours to allow the enzyme molecules to be stably embedded in the network structure. The resulting enzyme-loaded particles have a smooth surface, a diameter of approximately 1–3 mm, and their density can be controlled by adjusting the solid-liquid ratio.

[0038] When preparing carriers for use at different layers, the component ratios can be fine-tuned in the basic formulation: (1) The proportion of chitosan and the proportion of PVA can be appropriately reduced in the upper layer carrier so that its density is slightly lower than that of water. The middle layer carrier formula remains unchanged; (2) The lower carrier is increased by 1 to 3 parts of magnetic iron oxide powder or silicon filler to increase the density and maintain stable sedimentation.

[0039] After the immobilized enzyme carrier is cleaned to remove free enzymes from its surface, it can be stored for later use. Its activity can be maintained for more than two weeks, making it suitable for long-term use under continuous operating conditions.

[0040] During the reaction, the immobilized enzyme carrier is evenly distributed in each reaction zone, and a slow flow is maintained by a circulating pump to ensure full contact between the enzyme and dye molecules. The system maintains a temperature of 10–15 degrees Celsius and a dissolved oxygen concentration of 3–5 mg / L through a central control unit, and automatically adjusts the flow rate and residence time to ensure stable decolorization reaction.

[0041] Specifically, in step S3, the floating enzyme carrier is introduced into the upper region of the reaction system after surface cross-linking and curing treatment to form a stable floating layer below the liquid surface. The reaction conditions in the upper region are controlled by an external gas distribution device or a liquid surface disturbance device to control oxygen distribution and flow stability.

[0042] This embodiment corresponds to the floating enzyme carrier used in the upper reaction zone. Its main function is to utilize the dissolved oxygen conditions at the water surface to rapidly oxidize and degrade the conjugated double bonds and aromatic ring structures in dye molecules under low-temperature conditions. To adapt to the high-oxygen and light environment of the upper layer, the carrier needs to have low density, good hydrophobic-hydrophilic balance, and strong enzyme immobilization stability.

[0043] In a typical preparation process, the carrier matrix solution is first prepared. By weight, 30–40 parts of polyvinyl alcohol (PVA), 15–20 parts of chitosan, 5–10 parts of glycerol, 1–2 parts of glutaraldehyde (a crosslinking agent), and deionized water are added to bring the total to 100 parts. PVA serves as the main film-forming material, chitosan provides amino active sites to facilitate enzyme binding, and glycerol acts as a flexible plasticizer to improve the mechanical properties of the carrier. To improve flocculation performance, 3–6 parts of foamed microspheres or closed-cell polypropylene microparticles can be added to the mixture to control the final density between 0.8 and 0.95 g / cm³.

[0044] The enzyme formulation mainly uses oxidative biological enzymes, such as laccase, lignin peroxidase, and manganese peroxidase, either alone or in combination. A typical ratio is 10-15 parts laccase, 5-10 parts lignin peroxidase, and 3-5 parts stabilizer (such as trehalose or ethylene glycol), with phosphate buffer added to adjust the pH to around 7.0.

[0045] In the preparation process, the carrier matrix solution is first heated to 80°C and stirred to fully dissolve the PVA. After cooling to 40°C, chitosan solution and glycerol are added and mixed evenly. Then, a crosslinking agent is slowly added and stirring continues for about 15 minutes to form a viscous colloid. When the temperature drops below 25°C, a pre-prepared enzyme solution is slowly mixed into the colloid, forming a homogeneous enzyme-polymer mixture under continuous stirring. Subsequently, particles are formed using droplet molding or micro-molding curing methods, and then dropped into a curing solution containing boric acid or calcium chloride for crosslinking and curing. The curing time is approximately 30–60 minutes.

[0046] After molding, the enzyme-carrying particles are washed with deionized water to remove free enzymes and cross-linking residues, and then allowed to stand at 4°C for 12 hours to stabilize their structure. The resulting particles have a diameter of approximately 1–2 mm, float on the water surface or in the upper layer of the water body, and are tough to the touch and not easily broken. Density testing shows that the average density of the particles is approximately 0.05 g / cm³ lower than the density of the reaction solution, allowing them to naturally aggregate in the upper region of the reaction apparatus to form a stable reaction layer.

[0047] During addition, the dye is directly added through the dosing port at the top of the device, with the quantity controlled at 3-5% of the reaction volume by mass. The upper carrier mainly performs the function of oxidation and decolorization in the system, and the high dissolved oxygen environment promotes the breakage of dye molecular structures. To maintain layer stability, a liquid surface turbulence or mild aeration device can be installed to prevent particle overflow. After each operating cycle, the upper carrier is recovered through a separation screen, and can be reused after cleaning.

[0048] Specifically, in step S3, a suspended enzyme carrier is provided in the middle layer region of the layered reaction system. The enzyme carrier uses a gel or composite porous material with a density similar to that of the reaction solution as a matrix. The matrix is ​​loaded with a biological enzyme that can maintain its activity at low temperature. The biological enzyme includes bacterial laccase, azo reductase, lignin peroxidase or a combination thereof. The middle layer region is equipped with a buoyancy detection and adjustment device, which is used to detect the buoyancy state of the suspended enzyme carrier in the reaction solution and adjust the buoyancy of the carrier particles accordingly, so that the carrier remains stably suspended in the middle layer region. The formulation process is achieved by adjusting the physical or chemical parameters within the system, including the fluid shear strength (20–300 s⁻¹). -1 Solution density (0.9–1.2 g / cm³) 3 ) or carrier density (0.95~1.25 g / cm³) 3 The fine adjustment of the fluid shear strength by 10–60 s further modulates the fluid shear strength. -1 The settings were adjusted to control the density of the reaction solution within ±0.005 to ±0.020 g / cm³. 3 Fine-tuning, or adjusting the electrolyte concentration by 5–50 mg / L, can be performed to maintain the stability of the carrier suspension.

[0049] This embodiment corresponds to the suspended enzyme-carrying particles in the middle reaction zone. Its core design lies in achieving stable suspension of the particles in the reaction solution, preventing them from floating to the surface or settling to the bottom, thereby creating a continuous and uniform enzyme reaction space in the middle region. The buoyancy of the middle carrier is adjusted through external detection and premixing ratio control, without on-site adjustment within the reaction system.

[0050] In preparation, the basic enzyme carrier matrix is ​​first prepared. By weight, take 25-35 parts polyvinyl alcohol (PVA), 10-15 parts chitosan, 10-20 parts starch or sodium alginate, 1-2 parts glutaraldehyde (a cross-linking agent), and deionized water to bring the total to 100 parts. PVA and chitosan form the main network structure, while starch or sodium alginate acts as a flexible regulating component, contributing to particle density control and pore structure formation. To improve structural strength, 1-3 parts of nano-silica or montmorillonite powder can be added to the solution to improve mechanical stability.

[0051] The basic enzyme system is a mixture of laccase and azoreductase, selected based on the characteristics of the wastewater. A typical ratio is 10 parts laccase, 10 parts azoreductase, 3-5 parts glycerol or ethylene glycol as an enzyme stabilizer, and 10-15 parts buffer solution (pH 7.0). After mixing, the mixture is thoroughly stirred with the matrix solution to obtain an enzyme-carrier hybrid system. Subsequently, it is shaped using a droplet method or a molding method, and cross-linked by being dropped into a calcium chloride curing solution. The temperature is controlled at 20-25℃, and the curing time is approximately 40 minutes. The resulting basic particles have a diameter of 1.5-3 mm and a density close to that of water (approximately 0.98-1.02 g / cm³), providing a benchmark for subsequent buoyancy adjustments.

[0052] During the external buoyancy adjustment process, the temperature and density of the wastewater batch are first tested. If the average density of the base particles is greater than the density of the wastewater, the particles will slowly sink during the static test. In this case, low-density buoyancy-enhancing particles are added to the mixing tank. These particles can be closed-cell polypropylene microparticles, microbubbly silica gel balls, or foamed glass microspheres, and the addition ratio is generally 2-8% of the base particle mass. If the density of the base particles is lower than the density of the wastewater and they float, high-density counterweight particles are added. These can be magnetic iron oxide powder, glass sand, or metallic silicon powder, and the addition ratio is controlled at 1-5%. The two types of adjusting particles are premixed with the base particles by mechanical stirring, and a small amount of crosslinking agent or gel binder is added for surface coating to maintain their combined state during subsequent operation.

[0053] After each batch is prepared, a "static chromatography test" is performed for verification. The prepared carrier sample is placed in a transparent water column and allowed to stand for 3 minutes, during which its main distribution layer is observed. If approximately 80% or more of the particles remain in the middle layer of the water column and do not significantly rise or sink over time, buoyancy matching is considered successful. This batch of enzyme-loaded particles is then numbered and recorded for use as the middle layer additive.

[0054] During addition, the particles are introduced through a dedicated dosing port in the middle layer of the reaction apparatus, with the dosage accounting for approximately 5-10% of the total system volume. The particles remain stably suspended under fluid circulation, eliminating the need for buoyancy adjustments during the reaction. After operation, the middle-layer particles are independently recovered and stored for later use via a staged recovery system.

[0055] Specifically, in step S3, a high-density enzyme carrier is provided in the lower region of the layered reaction system. The high-density carrier is based on a corrosion-resistant inorganic or composite material. The surface of the substrate is immobilized with a biological enzyme with reducing or redox combined activity. The biological enzyme includes azo reductase, peroxidase, or a co-carrier system thereof. The lower layer region maintains a uniform distribution of the carrier through a bottom guide or stirring structure, and the carrier is redispersed in the lower layer region after periodic backwashing to maintain reactivity.

[0056] The lower carrier described in this embodiment is located in the bottom sedimentation area of ​​the reaction device and is used to continuously carry out deep decolorization and organic matter decomposition reactions in a high-density environment. Its structure consists of three parts: a magnetic core, a supporting framework, and an enzyme immobilization outer layer. The design of the carrier focuses on balancing density stability, long-lasting enzyme activity, and mechanical abrasion resistance to ensure that it remains sedimented without caking during long-term operation and can be redispersed by liquid flow disturbance when necessary. The carrier uses magnetic iron oxide microparticles as the core, with a particle size controlled between 100 and 300 micrometers. After being washed with deionized water and coated with silica sol to form a thin silica layer, a hard, inert supporting framework with good adhesion properties is obtained. Subsequently, the treated magnetic powder is dispersed in an enzyme immobilization composite solution composed of polyvinyl alcohol and chitosan. The typical mass ratio of this solution is: 30 parts polyvinyl alcohol, 12 parts chitosan, 1.5 parts crosslinking agent, 3 parts glycerol, and buffer solution to make up to 100 parts. At room temperature, 12 parts azo reductase, 6 parts peroxidase, and 3 parts laccase were added to fully disperse the enzyme molecules in the composite sol system and allow them to non-covalently bind to amino or hydroxyl sites. The resulting suspension was then formed into wet particles by drop casting or roller coating, and immediately dropped into a crosslinking solution containing calcium chloride for curing for 30 to 40 minutes. After curing, the particles were removed and briefly immersed in a low-concentration crosslinking solution to increase the degree of crosslinking of the outer layer and form a dense protective skin. Finally, they were washed with cold water to remove free enzymes and residual crosslinking agents, and cured at 4 degrees Celsius for 12 hours to stabilize their three-dimensional network structure. The resulting carrier particles had a wet density of approximately 1.3 g / cm³, a diameter controlled between 2 and 3 mm, a smooth surface, a firm feel, and good shear resistance.

[0057] In terms of equipment layout, the reactor bottom adopts a shallow conical flow guiding structure with multiple liquid distribution holes in the center, enabling uniform flow field distribution during dosing and operation. The carrier is slowly added through a dedicated bottom dosing port, relying on its own weight to settle at the bottom and form a stable material layer. During system operation, the central control unit controls the reflux pump to start periodically based on bottom pressure difference or flow velocity monitoring data, generating short-term pulsating flow to disperse local accumulation areas, ensuring uniform distribution of the lower carrier layer and maintaining the active interface from clogging. Every two to four hours, the system automatically performs a 15 to 30 second pulsating process to ensure that the bottom area remains unobstructed. If a small number of particles float to the surface due to liquid entrainment during operation, they are immediately captured and recovered by the magnetic grid assembly at the outlet, preventing them from entering the upper reaction zone. When the equipment is shut down for maintenance, magnetic rods or magnetic trapping units can be used to centrally adsorb and recover the bottom carrier, followed by cleaning, drying, and enzyme activity testing. If the test results show that the enzyme activity is still within the set threshold range, the carrier weight ratio can be finely adjusted according to the density parameters of the new batch of wastewater before reuse. The formulation and process of this lower carrier can be achieved under normal conditions. The preparation process is simple and easy to control, with low cost, strong wear resistance and magnetic recovery. It can operate stably in actual engineering environments and achieve long-term recycling.

[0058] Specifically, in step S4, before a single batch of wastewater enters the reaction system, the temperature, density, and salinity parameters of the wastewater are detected, and the buoyancy of the enzyme carriers to be added to each layer is pre-adjusted accordingly. The outer wall of the reaction system is provided with multiple independent dosing ports along the height direction, corresponding to the upper, middle and lower regions respectively. After buoyancy pre-adjustment, the buoyant enzyme carrier, the intermediate suspended enzyme carrier, and the high-density enzyme carrier are respectively directed into their respective reaction layers through their corresponding dosing ports. Each dosing port is connected to an external carrier supply device through a stratified dosing pipeline, and is opened by the control system according to the detected parameters, thereby realizing the stratified delivery and initial distribution control of different buoyancy carriers.

[0059] The stratified dosing equipment and system structure described in this embodiment is based on an integrated reaction device. Internally, it naturally forms three reaction spaces—upper, middle, and lower—according to different density ranges. Each layer has an independent dosing channel, fluid circulation pipeline, and parameter monitoring nodes, thus achieving stratified carrier operation in its physical structure. The entire device adopts a vertical cylindrical structure, with a height of approximately 2.5 to 3 meters and a diameter of 0.8 to 1.2 meters. The material can be corrosion-resistant stainless steel or polypropylene composite material. Internally, it contains three layers of flow dividers with uniformly distributed fluid through-holes to form buffer transition areas. While there is fluid communication between the layers, a clear density interface is formed through differences in pore size, allowing the reaction liquid to automatically form a relatively stable stratified state during operation. The top of the cylinder is equipped with a micro-aeration distribution ring and a gas-liquid guide hood to control the dissolved oxygen environment of the upper layer and prevent carrier overflow; the middle section is equipped with a slow-flow rectifying net and sampling interface for monitoring the distribution of suspended carriers; and the bottom is equipped with a conical guide bottom and a multi-point liquid distributor for the uniform distribution of the lower layer carriers and pulsating backwashing.

[0060] The stratified dosing system consists of three independent dosing units, corresponding to the upper, middle, and lower layers. The upper dosing unit, installed at the top of the reactor, is equipped with precision electrically controlled valves and flow meters, and quantitatively delivers floating enzyme-carrying particles into the upper reaction zone via an airtight delivery pipe. The middle dosing unit, located in the middle section of the reactor, is connected to an independent branch with a slow-flow pipe and check valve, used to slowly inject the suspended carrier, adjusted by external buoyancy, into the middle layer. The bottom dosing unit is connected to the center of the guide bottom and is equipped with a high-pressure anti-clogging nozzle, which can be directly switched to a recirculation mode after dosing to maintain a stable flow field at the bottom. Each dosing unit is connected to a central control unit. The system opens the corresponding valves sequentially according to instructions from different reaction stages, achieving sequential dosing and stratification stabilization.

[0061] A buoyancy adjustment and external mixing device is installed around the equipment for carrier density calibration and batch pretreatment. This device includes a premixing tank with stirring function, a density detection probe, a temperature and conductivity detection module, and a micro-dosing unit. A propeller-type stirrer is installed inside the mixing tank, with a stirring speed adjustable between 30 and 80 revolutions per minute. During operation, the basic enzyme-carrying particles are first added to the premixing tank. Based on the detected wastewater density and temperature, the system automatically calculates the dosage of buoyancy-enhancing or counterweight particles, which are then slowly added by the micro-dosing unit. After thorough mixing, samples are taken for testing. The detection module is equipped with a transparent colorimetric tube or optical sensor cell to determine the suspension level of the particles in the water sample in real time. Once the target buoyancy range is confirmed, the entire batch of carrier is transported via sealed pipeline to the corresponding dosing unit storage tank for dosing.

[0062] The entire system's circulation pipeline consists of three channels: an upper micro-aeration circulation channel to maintain dissolved oxygen; a middle slow-flow circulation channel to maintain uniform distribution of suspended particles; and a lower dual-mode pipeline with reflux and pulsation for periodic backflushing. Each channel is equipped with an independent flow meter and regulating valve, and can be linked and adjusted via a central control unit. The system also features multi-point monitoring modules for temperature, pH, dissolved oxygen, and conductivity, with at least two sets of probes per layer. Monitoring data is fed back to the control unit in real time, enabling fully automated operation and layer parameter correction.

[0063] The equipment casing is also equipped with an inspection window, an emergency discharge port, and a filter recovery interface. After the reaction is complete, the reaction liquid can be introduced into the separation and recovery module through the bottom discharge port, and then processed through magnetic capture, screen classification, and membrane filtration to achieve complete separation of the carrier and water. Each layer of carrier is automatically transported to its respective regeneration tank through the recovery system. The tank can be cleaned, dried, and reconstituted with buoyancy for the next addition. The entire set of equipment has a compact structure and high functional integration, which can adapt to continuous operation as well as batch operation conditions, and can achieve automatic stratified dosing and buoyancy control, ensuring the controllability and stability of the stratified reaction from both structural and process perspectives.

[0064] Specifically, after the reaction cycle is completed, the enzyme carriers in the reaction system are separated and recovered.

[0065] The separation process includes sequential solid-liquid separation and stratified sieving. By setting up stratified sieve components or graded collection structures, the upper floating carrier, the middle suspended carrier, and the lower high-density carrier are recovered to their respective storage units. The separated reaction solution is polished after sand filtration, membrane separation, or other filtration units before being discharged. The recovered enzyme carrier is washed and tested before being reused in its corresponding stratum to prevent carrier particles from entering the effluent system.

[0066] This embodiment further relates to a method for precisely adjusting and compounding the buoyancy of the intermediate carrier, which is a key step in achieving layered stability in this process. The intermediate carrier mainly undertakes the decolorization and organic matter decomposition tasks in the main reaction zone and needs to be suspended in the middle layer of the reaction solution for a long time. Therefore, it must be externally adjusted to make its buoyancy similar to but slightly smaller than that of the water to maintain a slow floating equilibrium. The structure of the intermediate carrier is usually a composite particle with a lightweight porous core and an enzyme immobilization layer. It has high internal porosity and a large specific surface area, making it suitable for immobilizing various oxidases and co-catalysts. In a typical formulation, the lightweight core material can be hollow glass microspheres, expanded perlite, or modified polypropylene particles. The enzyme immobilization layer adopts a chitosan and polyvinyl alcohol composite system, supplemented with appropriate amounts of crosslinking agent, buffer salt, and stabilizer. The ratio can be: 20 to 30 parts lightweight core, 10 parts chitosan, 20 parts polyvinyl alcohol, 1 part crosslinking agent, and buffer solution to make up to 100 parts. Depending on specific needs, 5 to 10 parts of azo reductase, 3 to 5 parts of laccase, and an appropriate amount of oxidoreductase can be added to enhance the catalytic effect.

[0067] During the buoyancy adjustment process, a reference buoyancy value F0 is first measured based on the temperature and density of the wastewater in that batch. The base carrier sample with the closest buoyancy value is then selected as the core batch. If the test results show that the buoyancy of the base carrier is too high, meaning it remains in the upper layer of the water sample, a small amount of counterweight particles is added for correction. The counterweight particles can be fine-grained quartz sand, barium sulfate, or trace amounts of metal oxide powder, and their addition ratio is generally controlled within 0.5 to 3% by mass to increase the overall density of the carrier to the target range. Conversely, if the buoyancy of the base carrier is too low, meaning it sinks to the bottom of the water sample, lighter buoyancy particles are added for supplementation. The lighter buoyancy particles can be hydrophobically treated foamed polyethylene or microporous hollow glass beads, and the addition amount also does not exceed 3% by mass. All addition operations are completed in an independent mixing tank, and the stirring speed is maintained at 40 to 60 revolutions per minute during the mixing process to avoid enzyme molecule inactivation due to violent shearing. After preparation, take a sample for observation. The particles should rise slowly in the standard specific gravity liquid and eventually remain in the middle layer. Stable suspension for more than three minutes is considered as qualified preparation.

[0068] To improve the controllability and recovery efficiency of the carrier, this embodiment also introduces a magnetic compounding method, that is, adding a very small amount of magnetically responsive microparticles to the carrier system, so that they can be identified and quickly recovered by the magnetic capture unit in the subsequent separation stage. The amount of magnetic microparticles added generally does not exceed 0.2% of the total mass, which will not significantly change the buoyancy of the carrier, but can significantly simplify the recovery process. Through the above buoyancy measurement and compounding steps, the suspension depth of the middle layer carrier in the reaction system can be flexibly controlled according to the density, temperature and composition changes of different batches of wastewater, so that the enzyme reaction zone is always maintained at the ideal distribution layer.

[0069] After the reaction is complete, the middle-layer carrier enters the separation and recovery system with the liquid flow. It is separated through a combination of sieve filtration and magnetic capture. After removing moisture, its buoyancy and enzyme activity can be retested. If the enzyme activity is still within a usable range, the particles are sent to the buoyancy adjustment unit, fine-tuned according to the new wastewater density parameters, and then re-added for recycling. This method eliminates the need for complex control within the reaction system, relying entirely on external physical adjustment to achieve a stable suspension effect. This ensures the stability of the reaction layer and avoids interference between the middle-layer carrier and the upper and lower layers, thus forming a clearly structured and highly efficient layered system.

[0070] Specifically, the buoyancy adjustment method for the intermediate suspended enzyme carrier includes the following steps: Before a single batch of wastewater enters the reaction system, its temperature and density parameters are measured. Using basic enzyme-carrying particles as the main body, high-density counterweight particles and low-density buoyancy-enhancing particles were selected as adjustment materials.

[0071] When the buoyancy of the basic enzyme-carrying particles is found to be greater than the target buoyancy, high-density counterweight particles are added to the mixing system.

[0072] When the buoyancy of the basic enzyme-carrying particles is less than the target buoyancy, low-density buoyancy-enhancing particles are added to the mixing system.

[0073] After thorough mixing and bonding solidification, an enzyme-carrying particle system is formed with buoyancy matching the density of the batch of wastewater, which is then used in the middle layer of the reaction system.

[0074] This embodiment further illustrates the process steps for recovering and reusing enzyme-carrying particles after the reaction has completed, particularly focusing on the regeneration treatment of particles with remaining enzyme activity to reduce operating costs and maintain the long-term stability of the system. After the reaction, the device introduces the treated wastewater and suspended particles into the separation and recovery module through a bottom discharge pipe. This module consists of four parts: primary screening, magnetic separation, washing and drying, and buoyancy compounding. First, the particles are filtered through multiple layers of screens to remove inert sludge and impurities with a diameter smaller than the set value, retaining the main enzyme-carrying particles. The lower layer of magnetic carrier can be directly adsorbed and collected by the magnetic separation device during this process, while the middle and upper layers of non-magnetic carrier are collected by stratified sedimentation or flotation through a density differential flow channel.

[0075] All recovered particles were placed in a washing tank and gently washed with neutral buffer or diluted ethanol solution to remove surface deposits and residual organic matter, preventing contamination during reuse. After washing, the particles were dehydrated through a filter and transferred to a drying chamber where they were dried at a constant temperature of 40 to 50 degrees Celsius for 6 to 8 hours to ensure the enzyme structure was not damaged. Samples of the dried particles were taken to test their enzyme activity using colorimetric or redox rate assays. When the enzyme activity remained above 60% of its initial value, the particle was considered a reusable carrier.

[0076] For reusable carriers, the system re-measures buoyancy, comparing the actual buoyancy of the dried particles with the density of the current batch of wastewater. If the particle buoyancy is higher than the target value, an appropriate amount of counterweight particles is added to the external mixing tank and gently stirred to slightly increase its density; if the buoyancy is lower, a small amount of lighter particles is added to adjust it. This method can quickly restore the carrier's stratification stability in the reaction system. After recombination, the carrier's buoyancy stability is tested again to ensure that it can remain suspended in the middle or designated layer in the standard specific gravity liquid for more than three minutes before being reintroduced.

[0077] To improve recycling efficiency and automation, the system features a centralized recycling circulation channel that automatically distributes carriers from different layers to their corresponding storage tanks. Each tank is equipped with stirring and temperature control devices, allowing for separate cleaning, drying, and buoyancy-based recombination operations based on the carrier type. The entire recycling process is automatically executed by the central control unit according to the reaction cycle, requiring no manual intervention. Severely deactivated or structurally damaged particles can be removed through grading and magnetic screening before being sent to a waste treatment unit for unified processing.

[0078] This recycling and reuse process effectively reduces the consumption of carriers and enzymes, enabling continuous operation across multiple batches. Simultaneously, through buoyancy recalibration and magnetic-assisted separation, the re-added carrier maintains an ideal stratified state in the new batch of wastewater. The entire process is mild, controllable, and highly repeatable, making it suitable for large-scale engineering operations of bio-enzyme catalytic decolorization and purification systems, demonstrating significant economic and environmental advantages.

[0079] Specifically, the enzyme-carrying particles that are separated and recovered after the reaction are reused. The reuse process includes washing, drying, and detecting enzyme activity in the enzyme-carrying particles. When the test results show that the enzyme activity of the enzyme-loaded particles reaches the set usage threshold, The buoyancy of the particle was re-measured, and the buoyancy was adjusted based on the measurement results: When the buoyancy of the particles is higher than the target buoyancy range, add counterweight particles to adjust the balance. When the buoyancy of the particles is lower than the target buoyancy range, add buoyancy-enhancing particles for adjustment. The adjusted enzyme-carrying particles are then reintroduced into the corresponding layers of the reaction system for reuse.

[0080] This embodiment further illustrates the operation, maintenance, and long-term circulation strategy of the enzyme-carrying system to ensure the stability of enzyme activity and layered structure under continuous or multi-batch operation conditions. During operation, the central control unit monitors dissolved oxygen, pH, temperature, and flow rate parameters of each layer in real time, and automatically adjusts the reaction zone to maintain it within the optimal environmental range. The system employs a modular pipeline design: a micro-aeration distribution ring in the upper layer, a slow-flow rectification mechanism in the middle layer, and a pulsed backflow device in the lower layer, thus forming independent yet continuous fluid circulation channels between the three layers. The operating pressure of the upper aeration system is maintained at 0.05 to 0.1 MPa, creating a stable oxidation environment through microbubbles, which is beneficial for the catalysis of oxidative enzymes such as laccase and peroxidase; the middle slow-flow zone maintains a low-turbulence state, allowing the suspended carrier to be evenly distributed under weak disturbance; and the lower layer maintains particle looseness through intermittent pulsed backflow to avoid localized stagnation or dead zones.

[0081] In continuous operation mode, the system automatically executes a five-step cycle of "running—pulsation—detection—enzyme replenishment—recovery" according to a set period. Every 6 to 8 hours of operation, the bottom pulsation device automatically activates once, lasting 15 to 30 seconds, to break up localized accumulations. Every 24 hours, the system performs online detection, including the dissolved oxygen value, redox potential, and the rate of color decrease in the reaction solution for each layer. If the detection results indicate that the enzyme activity of the middle layer carrier has decreased to a set threshold (e.g., below 70% of the initial activity), the system automatically initiates the enzyme replenishment program. The enzyme replenishment program injects a certain amount of enzyme solution or pre-activated carrier particles into the suspension zone through the middle layer dosing port to replenish the reaction activity. This operation is completed without stopping the main system, ensuring the stability of the continuous reaction.

[0082] To prevent structural aging or enzyme layer shedding of the carriers after long-term operation, the system has a periodic maintenance procedure. Every 30 to 45 days of operation, the central control unit issues a maintenance command, sequentially transferring each carrier layer into the recovery channel and initiating the cleaning and regeneration process. Cleaning uses low-speed stirring and neutral buffer rinsing to remove attached contaminants and residual dyes. After cleaning, the carriers are dried at low temperature and their buoyancy and enzyme activity are retested. Those that pass the test are reused, while those that fail are discarded and replaced. Through this periodic regeneration mechanism, the system can maintain a highly efficient reaction state for a long time without frequent carrier replacement.

[0083] In addition, the automated control strategy of this system enables the operating parameters to be dynamically adjusted according to the characteristics of the wastewater. When the influent temperature approaches the optimal reaction temperature of the enzyme, the system can automatically shut down the heating or cooling module to save energy consumption; if it is detected that the pH of the wastewater deviates from the optimal range (such as being acidic or alkaline), a neutralization buffer solution is automatically added to keep the reaction solution within the optimal range of enzyme activity (pH 6.5 to 7.5). The system also has a data recording function, which can archive the water quality parameters, carrier dosage, and operating efficiency of each batch of treatment for long-term tracking analysis and optimization of process conditions.

[0084] This embodiment further illustrates the operation maintenance and recycling regeneration strategy of the enzyme-loaded system to ensure the stability of the layered structure and enzyme activity under continuous or multi-batch operation conditions. The entire reaction system is managed in real time by a central control unit, which monitors key parameters such as temperature, pH, dissolved oxygen, and flow rate of each layer, and achieves stable operation through automatic adjustment. The internal of the system adopts a vertical layered flow field design, with a micro-aeration distribution ring configured in the upper layer, a slow-flow rectification mechanism configured in the middle layer, and a pulsating reflux device configured in the lower layer, thus forming three independent and connected reaction areas. The upper layer maintains an oxidation environment through microbubbles, the middle layer forms a stable suspension area, and the lower layer is periodically backflushed to prevent accumulation.

[0085] Before each batch of wastewater enters the reaction system, the system first automatically detects basic parameters such as the temperature, density, and conductivity of this batch of wastewater, and calculates the buoyancy reference value based on this. Subsequently, the middle-layer carrier is batch-compounded by an external buoyancy adjustment unit. The compounding process is completely carried out outside the reaction system. The specific steps are as follows: According to the detected reference buoyancy, select the basic carrier sample with the closest density in the current storage tank, and measure its suspension layer position through an external buoyancy meter. When the measured buoyancy is greater than the reference value, a small amount of weighting particles (such as fine quartz sand or trace metal oxide powder) are automatically added, and after stirring evenly, it is measured again; when the measured buoyancy is less than the reference value, a small amount of light particles (such as hydrophobically modified expanded polyethylene or hollow glass microspheres) are automatically added until the carrier slowly rises in the standard water column and stably stays in the middle layer area, and it is considered qualified if it does not shift for more than three minutes.

[0086] The core innovation of this process lies in the real-time matching of buoyancy adjustment with wastewater parameters. Based on the detected buoyancy difference in the middle layer, the system automatically adjusts the ratio of light to heavy particles to ensure the carrier density is always compatible with the batch of wastewater. After adjustment, the system records the adjustment results as the batch's operating data for automatic reference in subsequent batches. The adjusted middle layer carrier is then sealed and transported to the dosing unit's storage tank, where it is sequentially added to the middle reaction zone of the reaction device, forming a stable three-layer reaction system together with the upper buoyant carrier and the lower magnetic carrier. Through this pre-adjusted buoyancy matching method, the carrier can maintain stable stratification even when wastewater density changes, preventing drift or mixing due to water quality differences, thus achieving adaptive stratification under different operating conditions.

[0087] During system operation, the central control unit automatically executes a five-step cycle of "running—pulsation—detection—enzyme replenishment—recovery" according to a pre-set program. Every 6 to 8 hours of operation, the bottom pulsation device automatically activates once, lasting 15 to 30 seconds, to break up localized buildup. Every 24 hours, the system automatically detects the parameters and reaction efficiency of each layer. If a shift in buoyancy in the middle layer or instability in the carrier layer is detected, a fine-tuning program is initiated, injecting a small amount of light or weight particles into the reaction zone through a bypass circulation to rebalance the buoyancy distribution without interrupting the reaction. If enzyme activity is detected to have dropped to a set threshold, the system automatically replenishes pre-activated carrier or enzyme solution through the dosing port to restore reaction activity.

[0088] After continuous operation reaches the set cycle, the system issues a maintenance command, sequentially introducing each layer of carrier into the recycling module to complete screening, washing, drying, and buoyancy readjustment. Reusable carriers, after their buoyancy is re-measured, are re-mixed according to the test data of the new batch of wastewater, while deactivated or structurally damaged particles are automatically screened out. Through this cyclical method, the carriers can not only be reused repeatedly but also adaptively adjust their buoyancy according to the characteristics of each batch of wastewater, always maintaining a stable suspended state in the middle layer. The entire process is completed at room temperature and with low energy consumption, achieving long-term continuous operation without the need for complex mechanical stratification devices.

[0089] Specifically, the buoyancy-reducing formulation system is used for density correction when the buoyancy of the enzyme-carrying particles exceeds the target value. This system includes iron oxide powder, barium sulfate powder, or fine-grained quartz sand as the main counterweight materials, and a binder compatible with the enzyme carrier composite layer to form a stable structure. The binder can be a hydrophilic polymer such as polyvinyl alcohol, chitosan, or carboxymethyl cellulose, and its addition ratio is 1-5% of the total mass of the carrier. After mixing, it is allowed to stand and solidify at room temperature, so that the counterweight particles are evenly distributed inside or on the surface of the carrier. The composite carrier prepared by this formulation has a density increase of 0.03-0.10 g / cm³, which can effectively reduce the buoyancy of the carrier and make it stably stay at the target layer.

[0090] In this embodiment, the buoyancy reduction system is mainly used for density correction when the buoyancy of the enzyme-carrying particles is higher than the target value and they exhibit an upward trend in the reaction system. Its core design involves introducing high-density inorganic filler particles inside the carrier and combining them with a hydrophilic polymer matrix to form a stable structure. This moderately increases the overall density without affecting enzyme activity, allowing the particles to return to their predetermined middle or lower layer positions. The system uses iron oxide powder, barium sulfate powder, and fine-grained quartz sand as the main weighting materials. These three materials can be used individually or in combination according to the density adjustment range. Iron oxide powder has good magnetic responsiveness, enabling rapid magnetic capture in the later recovery stage; barium sulfate powder has high density and is chemically inert, not reacting adversely with enzyme molecules; and quartz sand has a uniform particle size distribution, providing structural support and preventing localized accumulation.

[0091] In a typical formulation, 2-5 parts by weight of iron oxide powder, 1-3 parts by weight of barium sulfate powder, and 2 parts by weight of quartz sand are selected as high-density aggregates. Simultaneously, 1.5-3 parts by weight of polyvinyl alcohol solution (10% by weight) and 1-2 parts by weight of chitosan solution (5% by weight) are added as a binding matrix. If necessary, 0.1 parts by weight of glycerol or ethylene glycol are added to improve flexibility. After adding each component to the mixing tank in proportion, the mixture is stirred at low speed at 25-35 degrees Celsius to ensure uniform distribution of the weighting particles. The stirring time is controlled at 15-20 minutes to avoid local coagulation or particle sedimentation. Subsequently, 0.2-0.5 parts by weight of glutaraldehyde, a cross-linking agent, are added to allow the matrix to gradually cross-link and solidify during stirring, forming a composite with a certain strength. After the mixed system is allowed to stand for 30 minutes, a binder slurry with suitable fluidity is formed. This slurry can be coated onto the surface of the enzyme-carrying particles to be adjusted by roller coating or impregnation, or it can be directly mixed with the carrier and re-granulated; both methods achieve stable bonding.

[0092] When the density adjustment range is large, a two-layer structure of "internal filling + external wrapping" can be adopted. First, some weight particles are mixed with the enzyme-carrying matrix to form the core, and then a secondary weighting is achieved through an outer adhesive coating. After the solidified carrier is left to dry for 4 hours, its density increases by approximately 0.03 to 0.10 grams per cubic centimeter compared to the original particles. Buoyancy tests show that the treated carrier, under the same temperature and wastewater density conditions, can slowly sink from the water surface and remain in the target middle or lower layer, maintaining a stable suspension without significant drift over time.

[0093] To ensure enzyme activity remains unaffected, the entire preparation process is conducted at room temperature, and a biocompatible hydrophilic polymer system is selected as the binder. For use in the middle layer carrier, a mixed formulation containing a small amount of iron oxide can be chosen to balance magnetic responsiveness and density adjustment precision; for use in the lower layer carrier, the proportion of barium sulfate or quartz sand is appropriately increased to achieve a density of 1.2 to 1.4 g / cm³ to ensure sedimentation stability. After preparation, the carrier is allowed to stand for 12 hours to allow the internal binder to fully solidify before being introduced into the reaction system.

[0094] Using the above formulation and process, buoyancy can be precisely adjusted without altering the carrier enzyme layer structure, achieving accurate layer control. This method is simple to operate, uses widely available and inexpensive counterweight materials, and is suitable for layer stability regulation in various low-temperature enzyme catalytic reaction systems.

[0095] Specifically, the buoyancy-enhancing formulation system is used to lighten enzyme-carrying particles when their buoyancy is lower than the target value. This system uses foamed polyethylene, hollow glass microspheres, foamed polypropylene microparticles, or expanded perlite as the main buoyancy-enhancing materials, supplemented with polyvinyl alcohol or chitosan as the binding matrix, and adds 0.1 to 0.3 parts of surfactant to enhance interfacial bonding. The buoyancy-enhancing materials account for 1 to 8% of the total mass of the carrier. After mixing, stirring, and cross-linking curing, a lightweight composite structure is formed. The density of the resulting carrier is reduced by 0.02 to 0.08 g / cm³, allowing it to remain stably suspended or rise slowly in the reaction solution to restore the suspension balance of the middle reaction zone.

[0096] In this embodiment, the buoyancy-enhancing formulation system is suitable for lightening enzyme-carrying particles with excessive density that tend to settle in the reaction system. Its core design involves introducing low-density hollow materials or foamed particles into the carrier, creating a microcavity structure within the particles, thereby reducing the overall density while maintaining the integrity of the enzyme layer and reaction permeability. This system uses hollow glass microspheres, foamed polyethylene, foamed polypropylene, or expanded perlite as the main buoyancy-enhancing materials, combined with hydrophilic polymers such as polyvinyl alcohol and chitosan to form a flexible coating structure. This allows lightweight particles to be uniformly distributed on the surface or inside the enzyme carrier, constructing a porous, low-density composite carrier.

[0097] In a typical formulation, by weight, 3-6 parts of hollow glass microspheres, 2-4 parts of foamed polyethylene granules, 10 parts of chitosan solution (5% by weight), 20 parts of polyvinyl alcohol solution (10% by weight), 0.3 parts of crosslinking agent glutaraldehyde, 0.5 parts of glycerol, and deionized water are added to a total of 100 parts. After uniformly mixing the polyvinyl alcohol and chitosan solution, the foamed polyethylene granules and hollow glass microspheres are slowly added, and the mixture is stirred at low speed at approximately 30 degrees Celsius for 20 minutes to ensure the lightweight particles are uniformly dispersed in the matrix. Subsequently, the crosslinking agent is added dropwise and stirring continues for 5 minutes to allow the system to gradually solidify into a gel-like suspension. To enhance interfacial adhesion, 0.1 to 0.3 parts of surfactant, such as polysorbate (Tween-80) or alkyl glycosides, can be added to improve the adhesion between the lightweight particles and the polymer matrix.

[0098] The resulting mixture was granulated by spraying or molding, and after curing, it was soaked in cold water for 30 minutes to remove residual crosslinking agent, followed by natural drying for 4 hours. Density testing showed that the adjusted carrier density decreased by 0.02 to 0.08 g / cm³ compared to the original value, resulting in a significant increase in buoyancy. When placed in the reaction solution, the particles slowly rose and remained stable in the middle layer without drifting or floating. For the lightweight modification of the upper carrier, the proportion of foamed polyethylene can be appropriately increased to 5-8 parts to ensure it floats on the liquid surface without overflowing; while the lightweight adjustment of the middle carrier mainly uses hollow glass microspheres, achieving precise control of suspension depth through subtle changes in proportion.

[0099] To ensure that enzyme molecules are not blocked by the polymer matrix, this embodiment pre-binds the enzyme with a portion of the chitosan solution during mixing, and then blends it with lightweight materials. This immobilizes the enzyme in the inner layer of the matrix network, while the outer layer is a porous, lightweight filler layer, thus balancing reactivity and buoyancy control. Through this "inner enzyme, outer vesicle" structural layout, the carrier forms a stable microporous buoyancy structure in water, maintaining sufficient contact area while effectively preventing sedimentation.

[0100] If the carrier density is detected to be too high during reuse, a secondary buoyancy enhancement process can be performed using the same formula. The recovered dried particles are then added back into the above formula system, gently stirred, and allowed to solidify at room temperature to restore the original buoyancy. This process does not alter the enzyme immobilization layer of the original carrier, exhibiting high repeatability and stability.

[0101] This lightweight formulation and process enables flexible control of the buoyancy of enzyme-carrying particles, ensuring that the carriers at different layers remain stably suspended in the low-temperature reaction system. This guarantees long-term uniform distribution and high efficiency of the multilayer enzyme reaction zones. The method is simple to operate, operates under mild conditions, and does not damage the enzyme, making it suitable for continuous operation and batch reuse under industrial conditions.

[0102] Comparative Example 1: Conventional low-temperature enzymatic decolorization process without stratified buoyancy control To verify the effectiveness of the process of this invention, this comparative example uses a conventional low-temperature enzyme decolorization method without stratified buoyancy control and without stratified addition of enzyme-carrying particles as a control, while keeping other conditions as consistent as possible with those of this invention.

[0103] 1. Experimental conditions Wastewater type and initial indicators Simulated dyeing and printing wastewater, initial color: 180 times; Initial COD: 320 mg / L; Initial pH: 6.8.

[0104] Reaction apparatus and dosing method A single stirred reactor was used, with a reaction volume of 10L; The enzyme-carrying particles are not subject to buoyancy stratification control and are directly and uniformly added to the reactor in one go; Total amount of enzyme-carrying particles added: 7.5% of the reaction system volume (approximately 0.75L particle volume).

[0105] Running parameters Reaction temperature: 12.5℃ (low temperature conditions, but without stratification optimization); Adjust the pH to 7.0–7.2; Dissolved oxygen concentration should be controlled at 3–5 mg / L; Stirring speed: 40 r / min; Reaction time: 120 min, samples were taken at 0, 30, 60 and 120 min for testing.

[0106] Testing items Water color at various time points; COD of effluent at various time points; Observe the distribution of enzyme-carrying particles during the reaction process (whether there is obvious floating or sedimentation).

[0107] 2. Experimental Results Color change and decolorization rate

[0108] It can be seen that the decolorization rate is about 70% at 120 minutes, the overall decolorization speed is relatively slow, and the color reduction is not obvious in the first 60 minutes.

[0109] COD Changes and Removal Rate

[0110] At 120 minutes, the COD removal rate was about 28%, which showed some effect in removing organic matter, but the overall reduction was limited.

[0111] Observation of enzyme-carrying particle distribution About 20 to 30 minutes after the reaction begins, most of the enzyme-carrying particles gradually float to the vicinity of the liquid surface, and the number of particles in the bottom area is significantly reduced. After running for 60 minutes, a thick particle enrichment layer formed in the upper part of the reactor, while the lower part was basically clear liquid, exhibiting a typical floating and unstable stratification phenomenon. As particles aggregate on the liquid surface, the deep wastewater does not come into sufficient contact with the enzyme, resulting in limited overall decolorization efficiency.

[0112] This comparative example did not adopt the layered buoyancy control and mid-layer stable suspension addition method proposed in this invention. Although it can achieve a certain degree of decolorization and COD removal under the same low temperature conditions (12.5℃), the enzyme-carrying particles are prone to floating and agglomeration, and the reaction zone is unevenly distributed. At 120 min, the decolorization rate is only about 70% and the COD removal rate is about 28%, and the overall treatment efficiency is low.

[0113] Example 1: This embodiment uses the layered buoyancy control method proposed in this invention to verify whether increasing the amount of enzyme-carrying particles in the middle layer (10°C) can significantly improve the decolorization effect under the lowest temperature condition (10°C).

[0114] 1. Experimental conditions 1.1 Initial wastewater parameters Consistent with the comparative example: Initial chromaticity: 180x Initial COD: 320 mg / L Initial pH: 6.8 1.2 Layered enzyme-carrying particle addition method (core of this invention) Upper carrier dosage: 3% Mid-layer carrier dosage: 10% (high dose) Substrate dosage: 3% After adjustment, the densities of the three types of carriers were stably distributed in the upper, middle, and lower layers, respectively.

[0115] 1.3 Operating Parameters Reaction temperature: 10℃ (minimum temperature) pH: Adjust to 7.0–7.2 Dissolved oxygen: 3-5 mg / L Stirring speed: 40 r / min Total reaction time: 120 min 2. Experimental Results 2.1 Color change and decolorization rate

[0116] Compared to the control group (only 70%), increasing the concentration of the middle layer carrier under low temperature conditions keeps the middle reaction zone active and significantly improves the decolorization rate.

[0117] 2.2 COD Changes and Removal Rate

[0118] The COD removal rate increased by about 5 percentage points compared to the control group (28%), which is consistent with the improvement brought about by the increase in reactive surface area.

[0119] 2.3 Stratification stability and particle suspension state At 10°C, the enzyme reaction rate decreases due to the low temperature, but the 10% high-concentration middle-layer particles remain stably suspended in the liquid depth range of 30-60%. The upper, middle and lower reaction zones are clearly distributed, with no floating accumulation or sinking dead zones; During the experiment, the three layers of particles maintained a relatively stable distribution structure, ensuring that the catalytic reaction in each layer was carried out synchronously.

[0120] At 10°C, this embodiment achieved higher enzyme catalytic activity by increasing the dosage of the middle-layer enzyme-carrying particles (10%), resulting in a decolorization rate of 77.8% after 120 minutes, significantly better than the comparative example (70%) without the structure of this invention. This demonstrates that the present invention can maintain high processing performance at lower temperatures, verifying the upper limit of the dosage range and the effectiveness of the layered structure.

[0121] Example 2: This embodiment is used to verify whether the best overall treatment effect can be obtained by using a moderate amount of medium-layer enzyme particles (7.5%) under the intermediate temperature conditions (12.5°C) recommended by the present invention.

[0122] 1. Experimental conditions 1.1 Initial wastewater parameters (same as Example 1 and Comparative Example) Initial chromaticity: 180x Initial COD: 320 mg / L Initial pH: 6.8 1.2 Layered dosing structure (this invention) Upper carrier dosage: 3% Mid-layer carrier dosage: 7.5% (medium dose) Substrate dosage: 3% The three types of enzyme-carrying particles were set to floating, neutral suspension, and high-density sinking densities, respectively, to form a stable three-layer reaction zone.

[0123] 1.3 Operating Parameters Reaction temperature: 12.5℃ (preferred value for this invention) pH: 7.0–7.2 Dissolved oxygen: 3-5 mg / L Stirring speed: 40 r / min Total reaction time: 120 min 2. Experimental Results 2.1 Color change and decolorization rate

[0124] The three-layer reaction zone is most catalytically active at this temperature, and the decolorization rate is significantly faster than in Example 1 in the first 60 minutes.

[0125] 2.2 COD Changes and Removal Rate

[0126] The COD removal rate was improved by approximately 3.1% compared to Example 1, which was significantly better than the 28% of the comparative example.

[0127] 2.3 Stratification Stability and Particle Suspension The enzyme-carrying particles in the middle layer remain a highly stable suspension at a liquid depth of 35–55%; The upper particles simultaneously complete the oxidation and decomposition of surface pigments; The lower-layer particles adhere stably to the bottom and continuously participate in the degradation of deep organic matter; The overall distribution is the most uniform, and it is the group with the most stable structure among the three sets of embodiments; The reactor interior has virtually no floating or sinking dead zones, exhibiting optimal three-dimensional contact efficiency.

[0128] At 12.5℃, the combination with a middle layer enzyme particle dosage of 7.5% exhibited the best treatment performance.

[0129] The decolorization rate reached 83.9% and the COD removal rate reached 35.9% after 120 minutes, which were significantly improved compared with Example 1 and the comparative example. This indicates that when the temperature is at the optimal intermediate value, the enzyme activity, mass transfer efficiency and particle suspension stability reach the optimal balance.

[0130] Example 3: This embodiment is used to verify whether a good decolorization effect can still be maintained when the amount of enzyme-carrying particles added to the middle layer is reduced to 5% under the upper temperature limit (15°C), and to further illustrate the effectiveness of the upper limit of the parameter range.

[0131] 1. Experimental conditions 1.1 Initial wastewater parameters (Same as the previous two examples) Color intensity: 180x COD: 320 mg / L pH: 6.8 1.2 Layered dosing structure Upper carrier: 3% Middle layer carrier: 5% (low dose) Lower carrier: 3% Density control ensures that the three-layered particles form stable strata in the upper, middle, and lower regions.

[0132] 1.3 Operating Parameters Reaction temperature: 15℃ (maximum temperature) pH: 7.0–7.2 DO: 3-5 mg / L Stirring speed: 40 r / min Total reaction time: 120 min 2. Experimental Results 2.1 Color change and decolorization rate

[0133] Although the dosage was low, the decolorization rate still reached 80% due to the higher temperature (which accelerated the enzyme reaction), which was significantly better than the 70% of the control group.

[0134] 2.2 COD Changes and Removal Rate

[0135] The COD removal rate was slightly lower than that of Example 2 (35.9%), but higher than that of Example 1 (32.8%), consistent with the expectation of low dosage.

[0136] 2.3 Stratification Stability and Particle Suspension At 15℃, the water convection velocity increases slightly, and the distribution height of mid-layer particles shifts slightly upward. Due to the low dosage, the thickness of the intermediate suspended layer is relatively thin, but it can still maintain effective catalysis; The upper particle reaction zone has enhanced activity, and some pigment degradation processes are completed earlier in the upper layer. The three-layer structure can still maintain the boundary, but the "reduction in middle layer reaction load" caused by the reduction in the number of middle layer particles is more obvious. The overall reaction remained stable, with no accumulation or sedimentation dead zones observed.

[0137] At 15°C, reducing the dosage of the enzyme-carrying particles in the middle layer to 5% still resulted in an 80% decolorization rate and a 34.4% COD removal rate within 120 minutes, significantly better than the comparative examples (70% and 28%). This demonstrates that even at the lower limit of the dosage range of this invention, the process can still maintain stable and effective processing capacity.

[0138] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.

Claims

1. A low temperature wastewater decolorization and purification process based on biological enzyme catalysis, characterized in that, The method comprises the following steps: S1, wastewater pretreatment: the wastewater containing organic coloring substances is filtered, the pH and dissolved oxygen are adjusted, and then the wastewater is introduced into a reaction system; S2, low-temperature biological enzyme reaction: the pretreated wastewater is introduced into a low-temperature biological enzyme reaction device, and a biological enzyme catalytic decolorization reaction is performed at 10-15 DEG C; S3, layered configuration and synergistic reaction: a plurality of layered reaction zones are arranged in the reaction system, and enzyme carriers are arranged in different reaction zones to form a plurality of catalytic layers at different depths of the wastewater; The middle layer reaction zone is kept stable suspension by detecting the buoyancy state of the carrier and adjusting the buoyancy of the carrier particles, so that the layered synergistic decolorization is realized; S4, layered drug delivery control: the reaction system is provided with a plurality of drug delivery ports corresponding to each reaction layer, which is used for directional delivery of enzyme carriers or auxiliary reaction components to different reaction layers to adjust the reaction activity and catalytic efficiency of each layer; S5, regeneration and effluent treatment: the reaction system is regenerated and maintained after the reaction operation cycle is completed, and the purified effluent is obtained through subsequent filtration or membrane separation process.

2. The low-temperature wastewater decolorization and purification process based on biological enzyme catalysis according to claim 1, characterized in that: In step S1, the wastewater pretreatment step includes dynamically adjusting the pH of the wastewater, so that the pH of the reaction system is controlled in the weak acid to weak base range of 6.5-8.0, and the pH is slightly corrected during the operation of the reaction system according to the buoyancy state of the middle layer carrier, and the adjustment usually does not exceed ±0.5, so that the biological enzyme catalytic activity is ensured while the carrier is kept stable suspension, and the dissolved oxygen concentration is monitored simultaneously during the adjustment to prevent the oxidation capacity of the reaction system from decreasing.

3. The process for decolorization and purification of low temperature wastewater based on bio-enzyme catalysis according to claim 1, characterized in that: In step S2, the low-temperature biological enzyme reaction step uses low-temperature resistant biological oxidation-reduction enzyme for catalysis, and the biological enzyme includes any one or a combination of bacterial laccase, azo reductase or lignin peroxidase; The reaction system was operated in continuous flow or quasi-continuous flow mode at 10-15 ℃, and the enzyme catalytic conditions of each layered reaction zone were kept constant by setting and controlling the dissolved oxygen concentration (2.0-6.0 mg / L), fluid circulation rate (0.2-1.0 m / s), and stirring shear intensity (50-300 s -1 ) of the reaction solution. During the reaction operation, the temperature and dissolved oxygen parameters of the reaction liquid are monitored online, and the circulation flow is adjusted to maintain the stability of the enzyme catalytic environment.

4. The process for decolorization and purification of low temperature wastewater based on bio-enzyme catalysis according to claim 1, characterized in that: In step S3, the upper layer region of the layered reaction system is provided with a floating enzyme carrier, the floating enzyme carrier takes an inert porous material with a density lower than the reaction liquid as a substrate, the substrate is loaded with biological enzymes with surface oxidation activity, and the biological enzymes include bacterial laccase, peroxidase or a complex enzyme system thereof; The floating enzyme carrier is put into the upper layer region of the reaction system after surface cross-linking and solidification treatment, so as to form a stable floating layer below the liquid surface, and the reaction conditions of the upper layer region are controlled by an external gas distribution device or a liquid surface disturbance device to control the oxygen distribution and flow stability.

5. The process for decolorization and purification of low temperature wastewater based on bio-enzyme catalysis according to claim 1, characterized in that: In step S3, the middle layer region of the layered reaction system is provided with a suspended enzyme carrier, the enzyme carrier takes a gel or a composite porous material with a density close to the reaction liquid as a substrate, the substrate is loaded with biological enzymes that can remain active at low temperature, and the biological enzymes include bacterial laccase, azo reductase, lignin peroxidase or a combination thereof; The middle layer region is provided with a buoyancy detection and adjustment device for detecting the buoyancy state of the suspended enzyme carrier in the reaction liquid and adjusting the buoyancy of the carrier particles accordingly, so that the carrier is kept stable suspension in the middle layer region; The process of formulation is achieved by adjusting physical or chemical parameters within the system, including the fluid shear intensity (20-300 s -1 ), the solution density (0.9-1.2 g / cm 3 ) or the carrier density (0.95-1.25 g / cm 3 ) in small adjustments.

6. The process for decolorization and purification of low temperature wastewater based on bio-enzyme catalysis according to claim 1, characterized in that: In step S3, the lower region of the layered reaction system is provided with high-density enzyme carriers, which are based on corrosion-resistant inorganic or composite materials, and the surface of the substrate is fixed with biological enzymes with reducing or redox complex activity, including azo reductase, peroxidase or co-loading system thereof; The lower region maintains uniform distribution of the carriers through the bottom flow guide or stirring structure, and the carriers are re-dispersed in the lower region after periodic backwashing to maintain reaction activity.

7. The low-temperature wastewater decolorization and purification process based on bio-enzyme catalysis according to claim 1, characterized in that: In step S4, before the single batch of wastewater enters the reaction system, the temperature, density and salinity parameters of the batch of wastewater are detected, and the required enzyme carriers of each layer are respectively pre-adjusted according to the buoyancy; The outer wall of the reaction system is provided with multiple independent dosing ports in the height direction, respectively corresponding to the upper region, the middle region and the lower region; The floating enzyme carriers, the middle suspended enzyme carriers and the high-density enzyme carriers pre-adjusted by buoyancy are respectively directed into the corresponding reaction layer through the corresponding dosing ports, Each dosing port is connected with an external carrier supply device through a layered supply pipeline, and is opened according to the corresponding parameters detected by the control system, so as to realize the layered placement and initial distribution control of different buoyancy carriers.

8. The process for decolorization and purification of low temperature wastewater based on bio-enzyme catalysis according to claim 1, characterized in that: After the reaction operation cycle ends, the enzyme carriers in the reaction system are separated and recovered, The separation process includes solid-liquid separation and layer position screening in sequence, By setting a layered screen assembly or a hierarchical collection structure, the upper floating carriers, the middle suspended carriers and the lower high-density carriers are respectively recovered to the corresponding storage units; The reaction liquid after separation is polished by sand filtration, membrane separation or other filtration units, and then discharged; The recovered enzyme carriers are used again in the corresponding layer after cleaning and detection to avoid the entry of carrier particles into the effluent system.

9. The low-temperature wastewater decolorization and purification process based on biological enzyme catalysis according to claim 5, characterized in that: The density correction system is used when the buoyancy of the enzyme-loaded particles is higher than the target value, which includes a stable structure formed by using iron oxide powder, barium sulfate powder or fine quartz sand as the main weight material, and a binder compatible with the enzyme carrier composite layer; the binder can be selected from hydrophilic polymers such as polyvinyl alcohol, chitosan or carboxymethyl cellulose, and the addition ratio is 1-5% of the total mass of the carrier, and the mixed material is placed at room temperature for solidification, so that the weight particles are uniformly distributed in the carrier interior or surface; the composite carrier prepared by this formula has a density increase of 0.03-0.10 grams per cubic centimeter, which can effectively reduce the buoyancy of the carrier and make it stably stay in the target layer.

10. The low-temperature wastewater decolorization and purification process based on biological enzyme catalysis according to claim 5, characterized in that: The increase of buoyancy of the preparation system is used for lightening when the enzyme-carrying particle buoyancy is lower than the target value, the system uses foamed polyethylene, hollow glass microbead, foamed polypropylene particle or expanded perlite as the main buoyancy-increasing material, is supplemented with polyvinyl alcohol or chitosan as the bonding matrix, and adds 0.1-0.3 parts of surfactant to enhance the interface bonding; the buoyancy-increasing material accounts for 1-8% of the total mass of the carrier, after mixing, stirring and cross-linking solidification, a light composite structure is formed, the density of the obtained carrier is reduced by 0.02-0.08 g / cm3, and the carrier can be stably suspended or slowly rises in the reaction liquid to restore the suspended equilibrium of the middle layer reaction zone.

Images