Method for enhancing removal of high-concentration antibiotics by coupling three-dimensional electrode with sulfur-mediated biological system

Abstract

The invention discloses a method for enhancing removal of high-concentration antibiotics by a three-dimensional electrode coupled sulfur-mediated biological system. The three-dimensional electrode structure is composed of a two-dimensional electrode and pig manure biochar, when the three-dimensional electrode structure is combined with a sulfur-mediated sewage biological treatment system for use, enrichment of functional microorganisms can be promoted, electron transfer and synergistic metabolism among the functional microorganisms can be improved, efficient synergistic removal of COD-sulfate-ammonia nitrogen-sulfamethoxazole can be promoted, and the effect of removing the COD-sulfate-ammonia nitrogen-sulfamethoxazole can be achieved. And a new technology is provided for treatment of wastewater containing carbon, nitrogen, sulfur and antibiotics. Especially in deep treatment of high-concentration antibiotics and pharmaceutical wastewater, the coupling technology can realize efficient removal of pollutants and long-term stable operation of the system, and a technical path with innovativeness and application potential is provided for treatment of complex refractory wastewater.

Description

Technical Field

[0001] This invention belongs to the field of wastewater biological treatment technology, specifically, it relates to a method for enhancing the removal of high-concentration antibiotics using a three-dimensional electrode coupled sulfur-mediated biological system. Background Technology

[0002] The widespread use of antibiotics has become a significant source of emerging pollutants in the aquatic environment. Because antibiotics are generally poorly absorbed by humans and animals, large quantities of parent drugs and their metabolites are excreted into wastewater treatment systems and ultimately enter environmental water bodies. Long-term accumulation of antibiotic pollution can lead to imbalances in aquatic ecosystems, the widespread dissemination of resistance genes, and potential risks to human health.

[0003] Sulfonamide antibiotics (such as sulfamethoxazole, SMX) are frequently detected in various water bodies due to their large usage, strong persistence in the environment, and significant ecotoxicity. Among numerous pollution sources, antibiotic production processes and pharmaceutical wastewater are among the main sources with the highest concentrations and strongest toxicity of antibiotics in the environment. This type of wastewater typically contains high concentrations of organic matter, residual antibiotics and their intermediate products, along with complex components such as high salt, high sulfate, and high ammonia nitrogen, exhibiting obvious characteristics of high concentration, strong toxicity, and complex pollution. If this type of wastewater is discharged directly without effective treatment, it will trigger a series of environmental risks, including eutrophication, the spread of antibiotic resistance genes, and the accumulation of toxic substances in aquatic organisms. Therefore, there is an urgent need to develop efficient, stable, and economical treatment technologies to address the problem of antibiotic wastewater pollution.

[0004] Currently, mainstream wastewater treatment processes are mainly based on the activated sludge process. Sulfur-mediated biological treatment technology based on the sulfur cycle has attracted much attention due to its ability to efficiently remove multiple pollutants such as carbon, nitrogen, and sulfur under low-carbon conditions. This technology relies on functional microorganisms such as sulfate-reducing bacteria, which maintain electron transfer and energy metabolism through the cyclic transformation between sulfur species of different valence states, and exhibits a certain degree of tolerance and degradation potential for some toxic organic compounds. Patent CN113845217A discloses a sulfur-mediated bioelectrochemical enhanced removal device for recalcitrant organic pollutants. This device uses an upflow sulfate-reducing anaerobic sludge bed reactor with an external power supply. The main functional bacteria in the reactor are sulfate-reducing bacteria, and the electrodes are placed vertically. The reaction process is under anaerobic conditions. Using this device for sulfur-mediated bioelectrochemical wastewater treatment can effectively remove typical recalcitrant organic pollutants from wastewater. For sulfamethoxazole at a concentration of 100 μg / L, the removal rate reaches over 40% within a hydraulic retention time of 6 hours. The article "Insights into the role of electrochemical stimulation on sulfur-driven biodegradation of antibiotics in wastewater treatment" points out that in an electrochemically stimulated sulfur-mediated bioreactor, the removal efficiency of sulfamethoxazole at a concentration of 1000 μg / L is 40.6±2.6%, which is much higher than that of a conventional sulfur-mediated bioreactor without electrochemical stimulation (30.4±2.3%). However, at high concentrations of sulfamethoxazole, it has a severe inhibitory effect on the sulfur-metabolizing microbial community in the sludge. The activity of microorganisms and the stability of the system will decrease significantly, manifested as inhibition of metabolic enzyme systems, obstruction of electron transport, and disorder of microbial community structure. This leads to a significant decrease in the treatment performance and operating efficiency of the system, which seriously restricts the practical application of sulfur-mediated biotechnology in the treatment of high-concentration antibiotic wastewater.

[0005] Therefore, developing a new wastewater treatment technology with high antibiotic concentrations that combines high efficiency, stability, and sustainability has become a key scientific and engineering problem that urgently needs to be solved. Summary of the Invention

[0006] The purpose of this invention is to overcome the above-mentioned defects and deficiencies in the prior art and to provide a method for enhancing antibiotic removal in a three-dimensional electrode coupled sulfur-mediated biological system.

[0007] The above-mentioned objective of this invention is achieved through the following technical solution:

[0008] This invention provides a method for enhancing antibiotic removal in a three-dimensional electrode-coupled sulfur-mediated biological system, the method comprising the following steps: S1. Activated sludge with sulfate-reducing bacteria as the dominant microbial community and pig manure biochar were filled between the anode / cathode two-dimensional electrodes of the bioelectrochemical-sulfur-mediated wastewater biological treatment system to prepare a three-dimensional electrode coupled sulfur-mediated biological system. S2. Wastewater is introduced into the three-dimensional electrode-coupled sulfur-mediated biological system in step S1 for treatment.

[0009] Three-dimensional electrode technology involves filling the space between traditional two-dimensional electrodes (anode / cathode) with a large number of conductive or electrocatalytic particles. Under an applied electric field, these particles become charged due to repolarization, forming countless tiny "third electrodes." This allows the electrochemical reaction to extend beyond the surface of the main electrode to the entire reactor space. This invention addresses the problems of low removal rates and poor system stability of sulfamethoxazole (SMX) in existing antibiotic and pharmaceutical wastewater treatment processes. It proposes a method to enhance antibiotic removal in wastewater using a three-dimensional electrode coupled with a sulfur-mediated biological wastewater treatment system. This involves adding pig manure biochar as a filling material between the two-dimensional electrodes (anode / cathode) of the existing bioelectrochemical-sulfur-mediated biological wastewater treatment system to form a third electrode. Activated sludge with sulfate-reducing bacteria (SRB) as the dominant flora is then inoculated. The sulfate-reducing bacteria, as functional bacteria in the treatment system, complete the sulfate reduction reaction, thereby achieving the removal of organic pollutants. The synergistic effect of biochar and external electric field allows the granular electrode (third electrode) of pig manure biochar to act as an electron donor and acceptor, promoting electron release and electron transfer efficiency from pollutants, and participating in the redox reaction process of pollutants. This strengthens the SRB activated sludge system and achieves efficient enrichment of functional microorganisms. Simultaneously, pig manure biochar possesses advantages such as a large specific surface area, abundant surface functional groups and rich in various nutrients, and excellent electrical conductivity. It can efficiently adsorb antibiotics, especially sulfamethoxazole (SMX), and also provides numerous attachment and growth sites for microorganisms. Furthermore, it can act as an electron mediator in interfacial reactions, compensating for the insufficient electron supply in traditional biological treatment systems. Therefore, combining pig manure biochar as the third electrode with a bioelectrochemical-sulfur-mediated wastewater biological treatment system to obtain a three-dimensional electrode coupled with a sulfur-mediated biological system can promote more efficient electron transfer and synergistic metabolism among functional microorganisms, thereby improving the system's resilience and operational stability, and significantly increasing the degradation rate of pollutants.

[0010] Furthermore, the bioelectrochemical-sulfur-mediated wastewater biological treatment system, referring to patent CN113845217A, includes a main body. The main body comprises a lower reaction zone and an upper overflow zone, separated by an overflow weir. The reaction zone is inoculated with activated sludge dominated by sulfate-reducing bacteria and pig manure biochar. A feeding port is provided above the overflow zone. An anode and a cathode are configured within the reaction zone. The anode is connected to the positive terminal of an external power supply, while the cathode is connected to the negative terminal. An inlet is located at the bottom of the reaction zone and connected to an inlet tank via an inlet pipe. A peristaltic pump is installed on the inlet pipe. A circulation port is located at the lower end of the overflow zone and connected to the inlet pipe via a circulation pipe and equipped with a circulation pump. An outlet is located in the middle of the overflow zone and connected to an outlet tank via an outlet pipe. An outlet pump is installed on the outlet pipe. The outer wall of the reaction zone is provided with an insulation layer.

[0011] Furthermore, the method for preparing the pig manure biochar is to dry the pig manure, heat it at 600-800℃ under anaerobic conditions for 1-3 hours, grind it, sieve it, and dry it to obtain the biochar.

[0012] Specifically, the method for preparing the pig manure biochar is as follows: S1. Place the pig manure in a 105℃ oven overnight to dry it, and obtain a dry pig manure sample; S2. The dried sample is loaded into a ceramic boat and transferred to the center of a tube furnace. Vacuum is drawn for calcination, and the sample is pyrolyzed at a preset heating rate of 10℃ / min to 700℃ for 2 hours. S3. The obtained pig manure biochar was sieved through a 100-mesh sieve and dried in an oven at 105℃ for 24 hours before use.

[0013] Preferably, the specific surface area of ​​the pig manure biochar is 200–400 m². 2 / g.

[0014] Furthermore, the final concentration (addition amount) of the pig manure biochar is 1–5 g / L. That is, the addition amount of pig manure biochar is 1–5% of the total volume of sludge added in the three-dimensional electrode coupled sulfur-mediated biological system.

[0015] Preferably, the amount of pig manure biochar added is 3 g / L.

[0016] Furthermore, the method for preparing the activated sludge involves aerating the sludge with nitrogen for 7–9 hours, then sieving it and acclimating the sludge to SO42-2000. 2- The COD removal rate and sludge concentration remain stable, which is the result.

[0017] Furthermore, the acclimation process involves placing the sludge in a sulfur-mediated biological wastewater treatment system and cultivating it through continuous flow. The influent nutrients include: sodium acetate as the sole carbon source, providing an electron donor, with a COD of approximately 1000 mg / L; anhydrous sodium sulfate as the electron acceptor; and nitrogen (N) and phosphorus (P) supplemented at a COD:N:P ratio of 100:5:1. Since the COD:N:P ratio in domestic and industrial wastewater is typically 100:5:1, the sludge is anaerobically acclimated using influent prepared at this ratio to obtain SRB-enriched sludge. Subsequently, during operation, progressively increasing SMX concentrations are applied to the system to screen and enrich SMX-tolerant functional microbial communities within the sludge, thereby improving the system's stability and treatment capacity under SMX-containing wastewater conditions.

[0018] Preferably, the specific composition of the influent nutrients includes: 1281.26 mg / L CH3COONa, 1213 mg / L Na2SO4, 26 mg / L CaCl2, 38.9 mg / L MgCl2, 191 mg / L NH4Cl, 1.44 mg / L KH2PO4, and 5.04 mg / L K2HPO4.

[0019] More preferably, the trace element stock solution for the influent nutrient solution comprises: 0.2 mg / L KI, 5 mg / L FeCl3·6H2O, and 0.5 mg / L H3BO3. 3、 0.5mg / L CuSO4, 0.63 mg / L MnSO4·H2O, 0.83 mg / L ZnSO4·7H2O, 0.5mg / L CoCl2·6H2O.

[0020] Furthermore, the amount of activated sludge added is 13-17 g-MLSS / L, and MLVSS / MLSS = 0.5-0.7.

[0021] Preferably, the amount of activated sludge added is 15 g-MLSS / L, and MLVSS / MLSS=0.6.

[0022] Furthermore, the anode (9) and cathode (10) are made of carbon brush material.

[0023] Furthermore, the carbon brush material is a carbon fiber brush with a diameter of 50 mm and a height of 100 mm.

[0024] Furthermore, the carbon brush material is first soaked in acetone for 24 hours before use, then washed with ultrapure water, and heat-treated in a muffle furnace at 400–500°C for 2 hours to remove surface impurities. Finally, it is connected to a DC power supply using 0.5 mm titanium wires. This forms a three-dimensional conductive microenvironment with the pig manure biochar.

[0025] Preferably, the power supply is a constant voltage DC power supply with a voltage of 0.2 to 1 V.

[0026] Furthermore, the reaction temperature of the three-dimensional electrode coupled sulfur-mediated biological system is 28–30 °C.

[0027] Furthermore, the antibiotic is sulfamethoxazole.

[0028] Furthermore, the concentration of sulfamethoxazole in the wastewater is 0.003–1000 μg / L.

[0029] Furthermore, the wastewater may also contain 300–50,000 mg / L of COD, 100–5,000 mg / L of sulfate, and / or 100–5,000 mg / L of ammonia nitrogen.

[0030] According to the above method, this invention determined the removal rate of SMX in wastewater containing 100 μg / L, 500 μg / L, and 1000 μg / L SMX by the three-dimensional electrode coupled sulfur-mediated biological system provided by this invention. The results showed that when 3 g / L of pig manure biochar was added, the SMX removal rates reached 95.9±0.3%, 83.3±3.6%, and 73.7±0.5%, respectively, improving the degradation rate; at the same time, it improved the removal rates of conventional pollutants such as carbon, nitrogen, and sulfur. Secondly, the sludge activity and electron transport activity in the three-dimensional electrode coupled sulfur-mediated biological system were investigated. The system of this invention has a significant effect on improving sludge activity and system electron transport capacity. For systems dependent on sulfate reduction, efficient nutrient metabolism can also simultaneously promote sulfate reduction, improving the system's SMX removal efficiency, shock resistance, and long-term stability, reducing operating costs, and providing a new solution for the treatment of related high SMX pollution or complex pollutants such as SMX, COD, ammonia nitrogen, and sulfur.

[0031] Compared with the prior art, the present invention has the following beneficial effects: This invention provides a method for enhancing the removal of high-concentration antibiotics using a three-dimensional electrode coupled with a sulfur-mediated biological system. The three-dimensional electrode structure comprises a two-dimensional electrode and pig manure biochar. When used in conjunction with a sulfur-mediated wastewater biological treatment system, it promotes the enrichment of functional microorganisms, enhances electron transfer and synergistic metabolism among these microorganisms, and thus facilitates the efficient synergistic removal of COD, sulfate, ammonia nitrogen, and sulfamethoxazole. This provides a new technology for treating wastewater containing carbon, nitrogen, sulfur, and antibiotics. Especially in the deep treatment of high-concentration antibiotic and pharmaceutical wastewater, this coupling technology can achieve efficient pollutant removal and long-term stable system operation, providing an innovative and promising technical approach for the treatment of complex and recalcitrant wastewater. Attached Figure Description

[0032] Figure 1 This is a schematic diagram of the three-dimensional electrode-coupled sulfur-mediated biological system of the present invention. In the diagram, 1-inlet tank, 2-peristaltic pump, 3-circulation pump, 4-external power supply, 5-insulation layer, 6-inlet, 7-feeding port, 8-reaction zone, 9-anode, 10-cathode, 11-outlet pump, 12-outlet tank, 13-overflow zone, 14-overflow weir, 15-inlet pipe, 16-circulation port, 17-outlet, 18-outlet pipe.

[0033] Figure 2 The adsorption rates of different concentrations of pig manure biochar on different concentrations of SMX were determined.

[0034] Figure 3 The removal rates of different concentrations of pig manure biochar for different concentrations of SMX were determined.

[0035] Figure 4 The removal rates of COD, sulfate, and SMX in a sulfur-mediated biological system and a three-dimensional electrode coupled sulfur-mediated biological system were determined.

[0036] Figure 5 The removal rates of ammonia nitrogen, nitrate, and nitrite in a three-dimensional electrode coupled sulfur-mediated biological system were determined. Detailed Implementation

[0037] The present invention will be further described below with reference to the accompanying drawings and specific embodiments, but the embodiments do not limit the present invention in any way. Unless otherwise specified, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in this technical field.

[0038] Unless otherwise specified, all reagents and materials used in the following examples are commercially available.

[0039] Example 1: Three-dimensional electrode coupled sulfur-mediated biological system for removing complex pollutants like Figure 1As shown, this embodiment provides a three-dimensional electrode-coupled sulfur-mediated biological system for removing complex pollutants. The appearance of the device is similar to that of an upflow anaerobic sludge blanket, and the height and diameter of the reactor can be designed according to actual conditions. Referring to patent CN113845217A, the specific configuration is as follows: The device includes a main body, which includes a lower reaction zone 8 and an upper overflow zone 13 within the reactor, separated by an overflow weir 14. A feed port 7 is provided above the overflow zone 13 to add solid material (pig manure biochar) into the reactor. The reaction zone 8 is inoculated with activated sludge dominated by sulfate-reducing bacteria (SRB) and pig manure biochar. The pig manure biochar can form a three-dimensional electrode with the anode 9 and cathode 10. The outer wall is provided with a heat insulation layer 5 to maintain a suitable reaction temperature for microorganisms. The anode 9 and cathode 10 are arranged in the reaction zone 8. The cathode 10 and anode 9 are made of carbon brush material. The anode 9 is connected to the positive terminal of the external power supply 4 through a resistor, while the cathode 10 is connected to the negative terminal of the external power supply 4 to form a bioelectrochemical circuit. The bottom of the reaction zone 8 is provided with a water inlet 6, which is connected to the water inlet tank 1 through a water inlet pipe 15. A peristaltic pump 2 is installed on the water inlet pipe 15 to control the water inlet flow rate. The overflow zone 13 is provided with a circulation port 16 at the lower end. The circulation port 16 is connected to the inlet pipe 15 through a circulation pipe and is equipped with a circulation pump 3 to realize liquid circulation and enhance mass transfer within the system. The overflow zone 13 is provided with an outlet 17 in the middle, which is connected to the outlet tank 12 through an outlet pipe 18. An outlet pump 11 is provided on the outlet pipe to control the discharge of water.

[0040] When using the aforementioned device to treat wastewater, sulfate-reducing bacteria, as functional bacteria in the reactor, complete the sulfate reduction reaction, thereby achieving the removal of organic pollutants. Under the stimulation of an external electric field, the electrodes can act as electron donors and acceptors, promoting the release and transfer of electrons from pollutants and participating in the redox reaction process, thus strengthening the SRB activated sludge system. However, under the stress of high-concentration antibiotics, the activity of sulfur-metabolizing microorganisms and the stability of the system will significantly decrease, manifested as inhibition of metabolic enzyme systems, obstruction of electron transfer, and disorder of microbial community structure, leading to a decrease in reaction rate and poorer operational stability. These adverse effects severely restrict the practical application of sulfur-mediated biotechnology in the treatment of high-concentration antibiotic wastewater. As a three-dimensional electrode, pig manure biochar granular electrodes have advantages such as large specific surface area, rich surface functional groups and rich in various nutrients, and excellent conductivity. They can efficiently adsorb antibiotics, especially sulfamethoxazole (SMX), making up for the deficiency of insufficient electron supply in traditional biological treatment systems, promoting more efficient electron transfer and synergistic metabolism among functional microorganisms, thereby improving the system's stress resistance and operational stability, and significantly increasing the degradation rate of pollutants.

[0041] Example 2: Cultivation of Sulfur-Mediated Sludge The sludge described in this embodiment is anaerobic activated sludge obtained through acclimatization and cultivation. The activated sludge originates from the secondary sedimentation tank of a wastewater treatment plant. The composition of the simulated wastewater is as follows: The initial concentration of the activated sludge is 10.0 ± 1.0 g-MLSS / L, and it is pretreated before acclimatization and cultivation. The method is as follows: (1) Pretreatment method of activated sludge: Aerate the collected activated sludge from the secondary sedimentation tank with nitrogen for 8 hours, then pass it through a 60-mesh sieve. After it has settled, pour off the supernatant to complete the pretreatment of activated sludge.

[0042] (2) Acclimation and cultivation method: The pretreated activated sludge was placed in the reactor of Example 1. At 25°C, the influent HRT (hydraulic retention time) was set to 12 h and COD (chemical oxygen demand) was set to 500 mg / L. After stabilizing for 24 h, the HRT was gradually shortened to 8 h and the COD was set to 1000 mg / L.

[0043] Specifically, COD was increased from 500 mg / L to 1000 mg / L during the start-up phase; after entering the SMX acclimatization / stable operation phase, COD remained at 1000 mg / L. SMX was acclimatized in stages, with the influent SMX concentration increasing from 100 μg / L to 500 μg / L, and finally to 1000 μg / L; each stage was run for at least 2 SRTs (SRT = 20 d), meaning the acclimatization time for each stage was no less than 40 d.

[0044] SMX acclimation concentration: Stage 1 (Startup Phase): 0; Stage 2 (Startup Phase): 0; Stage 3: 100 μg / L; Stage 4: 500 μg / L; Stage 5: 500 μg / L (with BC = 3 g / L); Stage 6: 1000 μg / L (with BC = 3 g / L); Stage 7: 1000 μg / L (add BC=3 g / L, Voltage=1 V).

[0045] The influent nutrients include: sodium acetate as the sole carbon source, providing an electron donor, with a COD of approximately 1000 mg / L; anhydrous sodium sulfate as the electron acceptor, supplemented with nitrogen (N) and phosphorus (P) elements in a COD:N:P ratio of 100:5:1. The specific composition includes: 1281.26 mg / L CH3COONa, 1213 mg / L Na2SO4, 26 mg / L CaCl2, 38.9 mg / L MgCl2, 191 mg / L NH4Cl, 1.44 mg / L KH2PO4, and 5.04 mg / L K2HPO4. In addition, trace elements are added to meet the growth needs of microorganisms. The trace element stock solution includes: 0.2 mg / L KI, 5 mg / L FeCl3·6H2O, and 0.5 mg / L H3BO4. 3、 0.5mg / L CuSO4, 0.63 mg / L MnSO4·H2O, 0.83 mg / L ZnSO4·7H2O, 0.5 mg / L CoCl2·6H2O.

[0046] Water samples were taken from the outlet at different times, up to SO4 levels. 2- The COD removal rate remained stable, thus obtaining sulfur-mediated sludge. The sludge was tested, and the mixed liquor suspended solids (MLSS) and mixed liquor volatile suspended solids (MLVSS) concentrations were 15 ± 0.5 g / L and 9 ± 0.5 g / L, respectively.

[0047] Since the COD:N:P ratio in domestic sewage and industrial wastewater is mostly 100:5:1, anaerobic acclimation of sludge is carried out using influent with a COD:N:P ratio of 100:5:1 to obtain SRB-enriched sludge. Subsequently, during operation, the SMX concentration is applied to the system in progressively increasing steps to screen and enrich the SMX-tolerant functional microbial community in the sludge, thereby improving the stability and treatment capacity of the system under SMX-containing wastewater conditions.

[0048] Example 3: Effect of biochar from different raw material sources on SMX adsorption efficiency I. Experimental Methods 1. Biochar preparation To evaluate the impact of biochar from different raw material sources on the removal of complex pollutants by a three-dimensional electrode coupled sulfur-mediated biological system, this embodiment compares biochar prepared from pig manure and straw. Pig manure and rice straw were dried at 105°C to obtain dried pig manure and rice straw samples. The obtained materials were placed in a ceramic boat and then placed in a tube furnace. The furnace was heated for 2 hours under anaerobic conditions at a preset heating rate of 10°C / min to 700°C. After cooling, the samples were removed, ground using an agate grinder, and then passed through a 100-mesh sieve. The sieved biochar was then dried again (105°C for 24 hours) and stored in a wide-mouth bottle for later use.

[0049] 2. SMX adsorption effect of different biochars 0.5 g (5 g / L) of pig manure and rice straw biochar were weighed and placed in 100 mL blue-capped bottles. 100 mL of artificially simulated wastewater containing 100, 500, and 1000 μg / L SMX was added to each bottle, respectively. The solutions were shaken at 200 r / min at 25℃, maintaining the initial pH unchanged. After 24 h, 1 mL of the supernatant was filtered through a 0.22 μm filter membrane. The SMX concentration of the filtrate was determined using ultra-high performance liquid chromatography (UHPLC). One blank was included, and each treatment was repeated in triplicate.

[0050] II. Experimental Results The results are shown in Table 1. Compared with straw biochar, pig manure biochar has better adsorption performance for SMX. Therefore, pig manure biochar was selected for subsequent experiments.

[0051] Table 1. Effects of different biochars on SMX adsorption efficiency

[0052] Example 4: Effect of pig manure biochar on SMX adsorption and removal I. Experimental Methods (1) Effect of pig manure biochar addition on SMX adsorption: Batch biodegradation experiments were conducted to clarify the effect of pig manure biochar addition on the biodegradation process and to distinguish the contributions of adsorption and biodegradation. 0.1, 0.3, and 0.5 g of biochar samples were weighed into 100 mL blue-capped bottles at different amounts (1, 3, and 5 g / L), and 100 mL of artificially simulated wastewater containing 100, 500, and 1000 μg / L SMX were added. The solution was shaken at 200 r / min at 25℃, with the initial pH unchanged. Sampling times were set at 0 min, 30 min, 1 h, 2 h, 4 h, 6 h, 8 h, 12 h, and 24 h. Samples were taken at the set times, and 1 mL of the supernatant was filtered through a 0.22 μm filter membrane. The SMX concentration of the filtrate was determined using ultra-high performance liquid chromatography.

[0053] (2) Effect of pig manure biochar addition on SMX removal: 0.1, 0.3, and 0.5 g of biochar samples were weighed into 100 mL blue-capped bottles at different pig manure biochar addition amounts (1, 3, and 5 g / L), and 100 mL of artificially simulated wastewater containing 1 g-VSS / L SRB sludge and 100, 500, and 1000 μg / L SMX were added. The solution was shaken at 200 r / min at 25℃ to maintain the initial pH of 7. Sampling times were set at 0 h, 2 h, 4 h, 8 h, 12 h, 24 h, 36 h, 48 h, 60 h, 72 h, 96 h, and 120 h. Samples were taken at the set times, and 1 mL of the supernatant was filtered through a 0.22 μm filter membrane. The SMX concentration of the filtrate was determined by ultra-high performance liquid chromatography.

[0054] (3) Effect of pig manure biochar on the removal of SMX at different concentrations: Pig manure biochar was accurately weighed at a dosage of 3.0 g / L into a 100 mL blue-capped bottle, and 100 mL of artificially simulated wastewater containing 100, 500, and 1000 μg / L SMX and 1 g-VSS / L sulfate-reduced sludge was added. The solution was shaken at 200 r / min at 25℃ to maintain the initial pH of 7. A blank control consisted of artificially simulated wastewater, sulfate-reduced sludge, SMX, and SRB inhibitor. Sampling times were set at 0 h, 2 h, 4 h, 8 h, 12 h, 24 h, 36 h, 48 h, 60 h, 72 h, 96 h, and 120 h. Samples were taken at the set times, and 1 mL of the supernatant was filtered through a 0.22 μm filter membrane. The SMX concentration of the filtrate was determined using ultra-high performance liquid chromatography.

[0055] II. Experimental Results Depend on Figure 2 The results show that pig manure biochar with added amounts of 3 g / L and 5 g / L exhibits good adsorption performance for SMX, and from Figure 2 It can be seen that the adsorption performance of 3 g / L pig manure biochar for SMX is better than that of 5 g / L pig manure biochar. For economic reasons, 3 g / L pig manure biochar is selected for addition.

[0056] Depend on Figure 3 The results show that pig manure biochar at concentrations of 3 g / L and 5 g / L exhibits good biological removal performance for SMX, and from Figure 3 It can be seen that 3 g / L pig manure biochar has better SMX removal performance than 5 g / L pig manure biochar. For economic reasons, 3 g / L pig manure biochar is selected for addition.

[0057] As shown in Table 2, pig manure biochar exhibits good SMX removal performance under high concentrations (500 μg / L and 1000 μg / L).

[0058] Table 2. Removal efficiency of 3 g / L pig manure biochar for different concentrations of SMX

[0059] Example 5: Removal effect of a three-dimensional electrode coupled with a sulfur-mediated biological system on compound pollution. I. Experimental Methods 1. Method of using a three-dimensional electrode-coupled sulfur-mediated biological system Sulfate-reduced sludge was inoculated into the reactor, and the mixed liquor suspended solids concentration (MLSS) was controlled at 15±0.5 g / L and the mixed liquor volatile suspended solids concentration (MLVSS) at 9±0.5 g / L. The reactor was continuously operated under the following conditions: hydraulic retention time (HRT) of 8 h, sludge retention time (SRT) of 20 d, influent flow rate (Q) of 4.05 L / d, and internal circulation flow rate of 3Q. Pig manure biochar granules (biochar mass m = 3 (g / L) × effective reactor volume V (L)) were directly added to the reactor as granular electrodes for the three-dimensional electrode, so that they were uniformly dispersed in the reactor and fully contacted with the sludge. The reactor was maintained under an external electric field with an applied voltage of 1 V, an external resistance of 50 Ω, and a system current of 10 mA, with the HRT maintained at 8 h during the external electric field stage.

[0060] 2. Long-term operation of a three-dimensional electrode coupled sulfur-mediated bioreactor After an external voltage was applied to the reactor, it was initially operated continuously for 80 days, during which time the SMX concentration in the influent was maintained at 1000 μg / L and the pig manure biochar concentration at 3 g / L. To achieve start-up, biofilm formation, and biological acclimatization of the bioelectrochemical reactor, the external voltage was increased in several stages: 0.2 V, 0.4 V, 0.8 V, and 1 V. After start-up, it was operated continuously for 40 days, maintaining the SMX concentration in the influent at 1000 μg / L, the pig manure biochar concentration at 3 g / L, and the external electric field voltage at 1 V.

[0061] 3. Antibiotic assay methods Antibiotics were analyzed using ultra-high performance liquid chromatography (UHPLC). The assay parameters were set as follows: flow rate 0.3 mL / min, injection volume 25 μL, column temperature 30℃, and detection wavelength 275 nm. The mobile phase consisted of acetonitrile (phase A) and 0.1% formic acid aqueous solution (phase B), with a volume ratio of 35:65 (v / v). The detection method was the external standard method.

[0062] II. Experimental Results Depend on Figure 4and Figure 5 This demonstrates that the three-dimensional electrode-coupled sulfur-mediated biological system can effectively remove high concentrations of organic matter, nitrogen, sulfur nutrients, and antibiotics, especially SMX at high concentrations. The three-dimensional electrode-coupled sulfur-mediated biological system of this invention can achieve a removal rate of 70% at an SMX concentration of 1000 μg / L, far exceeding the approximately 45% removal rate of ordinary sulfur-mediated biological systems (according to background art, at an SMX concentration of 1000 μg / L, electrochemical stimulation increases SMX removal efficiency by about 10% compared to ordinary sulfur-mediated biological systems; while the three-dimensional electrode-coupled sulfur-mediated biological system of this invention increases SMX removal efficiency by about 25% compared to ordinary sulfur-mediated biological systems). This demonstrates the practicality and optimizability of this system in the removal of complex pollutants, providing a new solution for related water treatment technologies.

[0063] Example 6: Sludge activity and electron transport activity in a three-dimensional electrode coupled sulfur-mediated biological system I. Experimental Methods 1. Sludge MLVSS determination method (testing sludge stability) The gravimetric method was used. The weight of the foil cup and the high-temperature resistant filter membrane was weighed and recorded as m0. V mL of the mud-water mixture was filtered through the filter membrane and placed into the foil cup. The foil cup and the mud-containing filter membrane were placed together in a multi-functional drying oven at 105℃ and heated for 24 hours. After constant weight, the weight was recorded as m1. Then, it was placed in a muffle furnace at 550℃ and heated for 2 hours. After cooling, it was removed, and the total weight of the sludge and the foil cup was recorded as m2. The calculation formula is as follows: .

[0064] 2. Dehydrogenase (DHA) assay method DHA activity was determined by quantifying the amount of reduced form of 2,3,5-triphenyltetrazol chloride (TTC). The specific steps were as follows: 2 mL of 0.18% Na2SO3, 3 mL of Tris-HCl buffer (pH=7.6), 4 mL of sludge sample, and 1 mL of 0.4% TTC were added sequentially to a 50 mL centrifuge tube. After shaking well, the tube was immediately placed in a 37℃ constant temperature and light-protected water bath and gently shaken for 30 minutes. Then, 1 drop of concentrated sulfuric acid was added to terminate the reaction, and the tube was centrifuged at 4000 rpm for 10 minutes. The supernatant was discarded, 10 mL of acetone was added, and the tube was thoroughly shaken and allowed to stand for 5 minutes for extraction. Finally, the supernatant was colorimetrically measured at a wavelength of 485 nm, and the DHA concentration was calculated using the TTC reduction standard curve.

[0065] 3. Method for measuring adenosine triphosphate (ATP) The ATP content was detected using an enhanced ATP assay kit. The specific steps are as follows: Take an appropriate amount of sludge sample into a centrifuge tube, centrifuge at 5000 g for 5 min, and discard the supernatant; add 1 mL of ATP extraction solution to the remaining sludge, let stand for 15 minutes, and collect the supernatant (keep the experiment on ice throughout); add 180 μL of ATP detection solution to the microplate, then add 20 μL of the extracted sample, and measure the chemiluminescence value of the sample; calculate the ATP content in the sample according to the ATP standard curve.

[0066] 4. Methods for determining extracellular polymeric substances (EPS) (1) EPS extraction: EPS was extracted using a modified thermal extraction method. The specific steps are as follows: Take an appropriate amount of mud-water mixture sample into a centrifuge tube and centrifuge at 4500 rpm and 4℃ for 15 minutes; discard the supernatant and add 5% NaCl solution to the original mark in the centrifuge tube. Shake well and keep warm in a 60℃ water bath for 30 minutes; then centrifuge at 12000 rpm and 4℃ for 15 minutes. Filter the supernatant through a 0.22 μm aqueous phase filter membrane to obtain the original EPS.

[0067] (2) EPS component analysis: Protein (PN) and humic substances (HA) quantification: The modified Lowry method was used for both. Reagent preparation: Reagent A was a mixture of 143 mM NaOH and 270 mM Na2CO3, Reagent B was 57 mM CuSO4, Reagent C was 124 mM sodium tartrate, Reagent D was a mixture of Reagent A:B:C = 98:1:1, and Reagent E was Folin-Ciocalteu reagent. PN determination: Add 4.2 mL of reagent D to 3 mL of sample, shake well and let stand for 10 minutes, then add 0.6 mL of reagent E, shake well again and let stand for 45 minutes. Measure the absorbance using a blank solution at 750 nm wavelength as a reference, and calculate the concentration using a standard curve prepared with bovine serum albumin as a standard. HA determination: Add 4.2 mL of reagent A to 3 mL of sample, shake well and let stand for 10 minutes, then add 0.6 mL of reagent E, shake well and let stand for 45 minutes. Measure the absorbance using a blank solution at 735 nm wavelength as a reference, and calculate the concentration using a standard curve prepared with humic substances as a standard.

[0068] 5. Electron Supply Capacity (EDC) Measurement Method The determination procedure is as follows: 6 mL of 0.1 M PBS was injected into the experimental cell, and the applied voltage was set to the desired potential Eh = -0.49 V, with electrochemical oxidation Eh = +0.61 V (relative to the standard hydrogen electrode). Once the current reached a steady state, 130 μL of electron transfer medium and 10 mM 2,2'-Azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS) were added. ABTS mediates the electrochemical oxidation of EPS, generating an oxidation current. After the current stabilized, 1 mL of EPS was added, and the changes in the current peak during the reaction were observed and recorded.

[0069] 6. Electron Acceptance Capability (EAC) Measurement Method The measurement procedure is as follows: 6 mL of 0.1 M PBS was injected into the experimental cell. The applied voltage was set to the desired potential: Eh = -0.49 V, and the electrochemical oxidation potential was set to Eh = +0.61 V (relative to the standard hydrogen electrode). Once the current reached a steady state, 130 μL of electron transfer medium and 10 mM diquat (DQ) were added. DQ mediates the electrochemical reduction of EPS, generating a reduction current. After the current stabilized, 1 mL of EPS was added, and the changes in the current peak during the reaction were observed and recorded.

[0070] 7. Cytochrome C (Cyt C) assay method During each stable operation phase, an appropriate amount of reactor sludge sample was taken and resuspended in 1 mL Tris-HCl buffer (50 mM, pH=8.0, containing 2 mM EDTA, 100 mM NaCl, and 0.1% Triton X-100). Then, 1 mg of lysozyme and protease inhibitor were added, and the mixture was incubated at 37°C for 30 min. After incubation, the mixture was sonicated at 4°C for 30 min, followed by centrifugation for 20 min (4°C, 10000 g), and the supernatant was collected for later use. The Cyt C concentration was determined using a double-antibody sandwich enzyme-linked immunosorbent assay (ELISA). Furthermore, to analyze the direct electron transport characteristics of Cyt C, a chronoamperometry method was used in a standard three-electrode system (glassy carbon working electrode, platinum wire counter electrode, and Ag / AgCl reference electrode), and the electron transport capacity of Cyt C was estimated by the integrated current peak area.

[0071] 8. Abundance of genes involved in sulfur metabolism The abundance of sulfur metabolism functional genes was characterized using metagenomic analysis: sludge samples were collected during the stable operation phase of the reactor, total DNA was extracted, and metagenomic sequencing was performed. The sequencing data were assembled to construct a non-redundant gene set. Subsequently, functional annotation and pathway analysis were performed on the gene set based on the KEGG database to screen key functional genes in the sulfate metabolism pathway within the sulfur metabolism pathway. Gene abundance was calculated using RPKM (Reads per kilobase per million reads). Furthermore, the abundance of all sulfate metabolism-related genes within the KEGG pathway was summed and standardized. The standardized results were used to characterize the relative abundance of key sulfate metabolism functional genes, thereby reflecting the changing trends of sulfate reduction-related metabolic capacity in sulfur-mediated biological systems.

[0072] II. Experimental Results As shown in Table 3, the introduction of the three-dimensional electrode structure significantly enhances sludge activity (increasing the abundance of sulfur metabolism functional genes) and system electron transport capacity (increasing DHA activity, ATP content, Cyt C content, EPS content, and EPS electron accepting / supply capacity). For systems dependent on sulfate reduction, efficient nutrient metabolism can simultaneously promote sulfate reduction, reduce sulfide pollution, and improve efficiency. In other words, by utilizing the three-dimensional electrode structure of this invention to regulate microbial nutrient utilization, under high-concentration antibiotic stress, the pollutant removal efficiency (significantly improving its adsorption and removal efficiency for high-concentration SMX in wastewater), shock resistance, and long-term stability of the three-dimensional electrode coupled sulfur-mediated bioreactor system can be improved, reducing operating costs. This system can be applied to the treatment of livestock wastewater and antibiotic production wastewater.

[0073] Table 3. Effects of 3 g / L pig manure biochar on relevant indicators

Claims

1. A method for enhancing antibiotic removal in a three-dimensional electrode-coupled sulfur-mediated biological system, characterized in that, The method includes the following steps: S1. Activated sludge with sulfate-reducing bacteria as the dominant microbial community and pig manure biochar were filled between the anode / cathode two-dimensional electrodes of the bioelectrochemical-sulfur-mediated wastewater biological treatment system to prepare a three-dimensional electrode coupled sulfur-mediated biological system. S2. Wastewater is introduced into the three-dimensional electrode-coupled sulfur-mediated biological system in step S1 for treatment.

2. The method according to claim 1, characterized in that, The bioelectrochemical-sulfur-mediated wastewater biological treatment system includes a main body; the interior of the main body includes a lower reaction zone (8) and an upper overflow zone (13), which are separated by an overflow weir (14). The reaction zone (8) is inoculated with activated sludge and pig manure biochar with sulfate-reducing bacteria as the dominant bacteria. A feed inlet (7) is provided above the overflow zone (13). An anode (9) and a cathode (10) are arranged in the reaction zone (8). The anode (9) is connected to the positive terminal of an external power source (4), while the cathode (10) is connected to... The negative terminal of the external power supply (4); the bottom of the reaction zone (8) is provided with a water inlet (6) and connected to the water inlet tank (1) through a water inlet pipe (15), and a peristaltic pump (2) is installed on the water inlet pipe (15); the lower end of the overflow zone (13) is provided with a circulation port (16), which is connected to the water inlet pipe (15) through a circulation pipe and is equipped with a circulation pump (3); the middle part of the overflow zone (13) is provided with a water outlet (17), which is connected to the water outlet tank (12) through a water outlet pipe (18), and a water outlet pump (11) is installed on the water outlet pipe (18); the outer wall of the reaction zone (8) is provided with a heat insulation layer (5).

3. The method according to claim 1 or 2, characterized in that, The method for preparing the pig manure biochar is as follows: after drying the pig manure, heat it at 600-800℃ under anaerobic conditions for 1-3 hours, then grind, sieve, and dry it to obtain the biochar.

4. The method according to claim 1 or 2, characterized in that, The final concentration of the pig manure biochar is 1–5 g / L.

5. The method according to claim 1 or 2, characterized in that, The method for preparing the activated sludge is to aerate the sludge with nitrogen for 7-9 hours, then sieve it and acclimate the sludge to SO42-200. 2- The COD removal rate and sludge concentration remain stable, which is the result.

6. The method according to claim 1 or 2, characterized in that, The amount of activated sludge added is 13-17 g-MLSS / L, and MLVSS / MLSS = 0.5-0.

7.

7. The method according to claim 1 or 2, characterized in that, The anode (9) and cathode (10) are made of carbon brush material.

8. The method according to claim 1, characterized in that, The antibiotic in question is sulfamethoxazole.

9. The method according to claim 1, characterized in that, The concentration of sulfamethoxazole in the wastewater was 0.003–1000 μg / L.

10. The method according to claim 1, characterized in that, The wastewater may also contain 300–50,000 mg / L of COD, 100–5,000 mg / L of sulfate, and / or 100–5,000 mg / L of ammonia nitrogen.

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