Anode for microbial fuel cell, microbial fuel cell and application

Abstract

The application belongs to the technical field of microbial fuel cells, and particularly relates to an anode for a microbial fuel cell, a microbial fuel cell and application. The anode for the microbial fuel cell comprises an electrode base and a modified material coated on the surface of the electrode base, and the modified material comprises nano-hydroxyapatite (nHA) and carbon nanotubes (CNTs). The nHA modification enables the electrode base to have better biocompatibility and surface area, is conducive to the formation of a dense biological membrane on the surface of the anode, and increases the electrocatalytic activity of the microbial fuel cell. The excellent conductivity of the CNTs can promote electron transfer and shorten the start-up time of the microbial fuel cell. The anode, a cathode and a proton exchange membrane can be assembled into a microbial fuel cell, and the microbial fuel cell can be used for degrading diclofenac and has a high degradation efficiency.

Description

Technical Field

[0001] This invention belongs to the field of microbial fuel cell technology, specifically relating to an anode for a microbial fuel cell, a microbial fuel cell, and its applications. Background Technology

[0002] Diclofenac, a nonsteroidal anti-inflammatory drug (NSAID), is widely used to relieve pain and reduce fever. However, diclofenac cannot be completely removed in urban wastewater treatment plants, making it a frequently detected substance in surface water. Diclofenac is also a persistent toxic organic pollutant that can cause toxic damage to terrestrial and various aquatic organisms and poses a potential risk to human health through accumulation in the food chain. Therefore, developing an effective method for removing diclofenac from wastewater is essential.

[0003] To date, research has shown that diclofenac can be effectively removed through methods such as Fenton oxidation, UV irradiation, ozone oxidation, and adsorption. For example, Chinese invention patent CN109942119A discloses a method for removing diclofenac from wastewater, utilizing activated carbon adsorption, photocatalytic degradation under the action of titanium dioxide, and autodegradation to reduce the content of organic pollutants in water. However, these technologies still have some significant drawbacks, such as causing secondary pollution, high cost, and producing toxic intermediates. In recent years, it has been discovered that diclofenac is biodegradable, with a mild reaction, no secondary pollution, low energy consumption, and environmental friendliness. However, it is difficult to screen diclofenac-degrading bacteria using traditional microbial methods, as microorganisms exhibit slow metabolic rates under diclofenac inhibition. Bioelectrochemical systems can utilize electroactive microorganisms to continuously provide electrons to effectively remove halogenated pollutants, overcoming the main shortcomings of purely biological processes. Microbial fuel cells are a typical example of a bioelectrochemical system.

[0004] Microbial fuel cells (MFCs) are an emerging environmentally friendly water treatment and biomass new energy technology that recovers energy by treating wastewater. The anode biofilm, rich in functional microorganisms, is the core of MFCs, enabling them to simultaneously degrade organic pollutants and generate electricity. As the carrier of the biofilm, the composition and structure of the anode are key factors affecting the performance of MFCs. Traditional carbon-based materials are widely used due to their good biocompatibility, conductivity, chemical stability, and low cost. However, these materials have relatively limited biocompatibility, surface area, and electrocatalytic activity, which are unfavorable for microbial attachment and growth, as well as extracellular electron transfer. Designing and fabricating nanomaterial-modified electrodes with superior performance is an effective method to improve the performance of MFCs.

[0005] Nano-hydroxyapatite (nHA) possesses excellent biocompatibility and bioactivity, a large specific surface area, and strong adsorption capacity, making it a promising biomaterial widely used in biomedical fields such as drug carriers, bone tissue repair materials, and anti-tumor composite materials. For example, Chinese invention patent CN 1772969A discloses a method for preparing a nano-hydroxyapatite / carbon nanotube composite coating. Using nano-sized HA and CNTs, the sintering temperature is significantly reduced, resulting in a composite coating with good adhesion and a dense structure. This significantly improves the bonding force between the biocomposite coating and medical metal materials, potentially making it suitable for use as an implant in weight-bearing parts of the human body. To date, there are no reports on the use of nHA / CNT nanocomposites for modifying the anodes of microbial fuel cells. Improving the biocompatibility, surface area, and electrocatalytic activity of microbial fuel cell anodes, and thus enhancing the ability of microbial fuel cells to degrade diclofenac, remains a problem requiring further research and development. Summary of the Invention

[0006] The purpose of this invention is to provide a microbial fuel cell to solve the problems of poor power generation performance and low diclofenac degradation efficiency in existing microbial fuel cells.

[0007] The second objective of this invention is to provide an application of microbial fuel cells in the degradation of diclofenac, in order to solve the problems of low degradation efficiency of diclofenac in the prior art.

[0008] The third objective of this invention is to provide an anode for microbial fuel cells to address the problems of low biocompatibility, low surface area, and low electrocatalytic activity in existing microbial fuel cell anodes.

[0009] To achieve the above objectives, the technical solution adopted by the present invention is as follows:

[0010] A microbial fuel cell includes an anode, a cathode, and a proton exchange membrane. The anode includes an electrode substrate and a modifying material coated on the surface of the electrode substrate. The modifying material includes nano-hydroxyapatite and carbon nanotubes, with a mass ratio of nano-hydroxyapatite to carbon nanotubes of 1:4. The anode is cultured in an anode nutrient solution containing anaerobic sludge to complete the acclimation of the anode biofilm.

[0011] The microbial fuel cell of this invention uses an nHA / CNT nanocomposite material as the anode modification material. The nHA modification gives the electrode matrix good biocompatibility and a large specific surface area, which can provide more sites and space for the attachment and growth of microorganisms, and is conducive to the formation of a dense biofilm on the anode surface. The excellent conductivity of CNTs can promote the extracellular electron transfer between microorganisms and electrodes, and shorten the start-up time of the microbial fuel cell. The synergistic effect of the two gives the microbial fuel cell of this invention high electrocatalytic performance, which can be used to degrade diclofenac with high degradation efficiency.

[0012] To facilitate rapid start-up of the microbial fuel cell, preferably, the anode nutrient solution uses phosphate buffer solution as the solvent, and the solutes are glucose, vitamins and minerals; the cathode solution corresponding to the cathode is potassium ferricyanide solution.

[0013] Preferably, the anolyte nutrient solution is 50 mmol / L. -1 Using phosphate buffer as a solvent, glucose, vitamin solution, and mineral solution were added to achieve concentrations of 1 g / L. -1 12.5 mL -1 and 12.5 mL L -1 The concentration of vitamin H in the vitamin solution is 2.0 mg / L. -1 The folic acid concentration is 2.0 mg / L. -1 The concentration of vitamin B6 was 10.0 mg / L. -1 The concentration of vitamin B2 was 5.0 mg / L. -1 The concentration of vitamin B1 was 5.0 mg / L. -1 The concentration of vitamin B3 was 5.0 mg / L. -1 The concentration of vitamin B5 was 5.0 mg / L. -1 The concentration of vitamin B12 was 0.1 mg / L. -1 The concentration of MgSO4 in the mineral solution is 3.0 mg / L. -1 The concentration of MnSO4 is 0.5 mg / L. -1 The concentration of NaCl is 1.0 mg / L. -1 The concentration of FeSO4·7H2O is 0.1 mg / L. -1 The concentration of CaCl2·2H2O is 0.1 mg / L. -1 The concentration of CoCl2·6H2O is 0.1 mg / L. -1 The concentration of ZnCl2 was 0.13 mg / L. -1 The concentration of CuSO4·5H2O is 0.01 mg / L. -1The concentration of AlK(SO4)·12H2O was 0.01 mg / L. -1 The concentration of H3BO3 was 0.01 mg / L. -1 The concentration of Na₂MoO₄ was 0.025 mg / L. -1 The concentration of NiCl2·6H2O was 0.024 mg / L. -1 The concentration of Na₂WO₄·2H₂O is 0.025 mg / L. -1 The potassium ferricyanide solution has a concentration of 50 mmol / L. -1 .

[0014] Preferably, the electrode substrate is a carbon-based electrode material; the carbon-based electrode material is a carbon brush; and the amount of the modifying material attached to the electrode substrate is 42–50 mg / g. -1 .

[0015] Preferably, the coating involves spraying a dispersion containing a modifying material onto the surface of the electrode substrate; the dispersion is obtained by mixing nano-hydroxyapatite, carbon nanotubes, a dispersant, and a binder. Spraying can increase the surface roughness of the electrode substrate, providing more sites and space for microbial attachment and growth, improving microbial film formation, and the uniform mixing of the modifying material and binder can also prevent biofilm detachment.

[0016] Preferably, the dispersant is anhydrous ethanol, and the amount of anhydrous ethanol used is 10 mL for every 6 mg of nano hydroxyapatite; the binder is a 5 wt% Nafion solution, and the amount of 5 wt% Nafion solution used is 200 mg for every 6 mg of nano hydroxyapatite.

[0017] An application of a microbial fuel cell, as described above, in the degradation of diclofenac.

[0018] The microbial fuel cell of this invention uses nHA / CNTs nanocomposite material as the anode modification material. During the degradation of diclofenac, the good biocompatibility of nano-hydroxyapatite is conducive to enriching a relatively abundant microbial population and giving it better microbial activity. This is beneficial for the domestication of functional microbial groups such as electroactive microorganisms and organic matter degrading bacteria. Furthermore, in the environment of the modification material with better bioactivity, the microorganisms have better tolerance to the impact of diclofenac. At the same time, CNTs can promote electron transfer. The synergistic effect of the two makes the microbial fuel cell of this invention have a high diclofenac degradation efficiency.

[0019] An anode for a microbial fuel cell includes an electrode substrate and a modifying material coated on the surface of the electrode substrate; the modifying material includes nano-hydroxyapatite and carbon nanotubes, wherein the mass ratio of nano-hydroxyapatite to carbon nanotubes is 1:4.

[0020] The anode for the microbial fuel cell of the present invention uses nHA / CNTs nanocomposite material as a modifying material. nHA modification gives the electrode matrix good biocompatibility and a large specific surface area, which can provide more sites and space for the attachment and growth of microorganisms, and is conducive to the formation of a dense biofilm on the anode surface. The excellent conductivity of CNTs can promote the extracellular electron transfer between microorganisms and electrodes, shorten the start-up time of the microbial fuel cell, and increase the maximum output voltage and maximum output power density of the microbial fuel cell.

[0021] Preferably, the electrode substrate is a carbon-based electrode material; the carbon-based electrode material is a carbon brush; and the amount of the modifying material attached to the electrode substrate is 42–50 mg / g. -1 .

[0022] Preferably, the coating involves spraying a dispersion containing a modifying material onto the surface of the electrode substrate; the dispersion is obtained by mixing nano-hydroxyapatite, carbon nanotubes, a dispersant, and a binder. Spraying can increase the surface roughness of the electrode substrate, providing more sites and space for microbial attachment and growth, improving microbial film formation, and the uniform mixing of the modifying material and binder can also prevent biofilm detachment.

[0023] Preferably, the dispersant is anhydrous ethanol, and the amount of anhydrous ethanol used is 10 mL for every 6 mg of nano hydroxyapatite; the binder is a 5 wt% Nafion solution, and the amount of 5 wt% Nafion solution used is 200 mg for every 6 mg of nano hydroxyapatite. Attached Figure Description

[0024] Figure 1 The output voltage curves of MFCs (external resistance 1000Ω) are shown, where (a) is the output voltage curve of nHA / CNTs / CB-MFCs and (b) is the output voltage curve of CB-MFCs.

[0025] Figure 2 The removal rate curves of diclofenac in nHA / CNTs / CB-MFCs, CB-MFCs and nHA / CNTs / CB-MFCs-open circuit over 48 hours are shown.

[0026] Figure 3The images show scanning electron microscope (SEM) images of the anolyte biofilm of MFCs after long-term operation. Among them, (a) is a scanning electron microscope image of the anolyte biofilm of nHA / CNTs / CB-MFCs after long-term operation at 1000x magnification, (b) is a scanning electron microscope image of the anolyte biofilm of CB-MFCs after long-term operation at 1000x magnification, (c) is a scanning electron microscope image of the anolyte biofilm of nHA / CNTs / CB-MFCs after long-term operation at 10000x magnification, and (d) is a scanning electron microscope image of the anolyte biofilm of CB-MFCs after long-term operation at 10000x magnification.

[0027] Figure 4 The relative abundance of microbial communities at the phylum level in three groups of samples: anaerobic sludge inoculum (G0), nHA / CNT / CB-MFCs (G3), and CB-MFCs (G5);

[0028] Figure 5 Heatmaps of the microbial community structure at the genus level for three groups of samples: anaerobic sludge inoculum (G0), nHA / CNT / CB-MFCs (G3), and CB-MFCs (G5). Detailed Implementation

[0029] The technical solution of the present invention will be further described below with reference to the accompanying drawings and specific embodiments.

[0030] The reagents used in the embodiments and experimental examples of this invention are as follows:

[0031] Materials: Carbon brushes, Nafion 117 proton exchange membrane (Shanghai Hesen Electric Co., Ltd.)

[0032] Reagents: Nano-hydroxyapatite (purchased from Bailingwei Technology), multi-walled carbon nanotubes XFM31 (purchased from Nanjing Xianfeng Nanomaterials Technology Co., Ltd.), diclofenac sodium (DS, purchased from Bailingwei Technology).

[0033] I. Examples of the Microbial Fuel Cell of the Present Invention

[0034] Example 1

[0035] The microbial fuel cell in this embodiment is a dual-chamber microbial fuel cell, including an anode, a cathode, and a proton exchange membrane. The anode includes an electrode substrate and a modification material coated on the surface of the electrode substrate. The modification material includes nano-hydroxyapatite and carbon nanotubes, and the amount of modification material attached to the electrode substrate is 42 mg / g. -1 (Under parallel experimental conditions, the amount of modified material attached to the electrode substrate was 42–50 mg g) -1 ).

[0036] The specific construction and operation process of the microbial fuel cell in this embodiment is as follows:

[0037] (1) Preparation of nHA / CNTs modified carbon brush: 6 mg of nano hydroxyapatite and 24 mg of carbon nanotubes (mass ratio of the two is 1:4, total 30 mg) were added to 10 mL of anhydrous ethanol, and 200 mg of 5 wt% Nafion (binder) was added and ultrasonically dispersed for 20 minutes to obtain a dispersion. Then, the dispersion was sprayed onto the carbon brush surface in small amounts multiple times using a spray gun under infrared dryer. After that, it was placed in the air for at least 24 hours to dry completely to obtain nHA / CNTs modified carbon brush.

[0038] (2) Assemble MFCs: Use the nHA / CNTs modified carbon brush prepared in step (1) as the anode, ordinary bare carbon brush as the cathode, and Nafion 117 proton exchange membrane as the separator to assemble microbial fuel cells, denoted as nHA / CNTs / CB-MFCs;

[0039] (3) Operation of MFCs: The anodes were cultured in an anode nutrient solution containing anaerobic sludge (mixed bacterial source). The anode nutrient solution was prepared at a concentration of 50 mmol / L. -1 Using phosphate buffer as a solvent, glucose, vitamin solution, and mineral solution were added to achieve concentrations of 1 g / L. -1 12.5 mL -1 and 12.5 mL L -1 ; with 50 mmol L -1 Potassium ferricyanide was used as the cathode solution to start and run nHA / CNTs / CB-MFCs.

[0040] In this example, 50 mmol L -1 The solutes and concentrations of each solute in the phosphate buffer solution, vitamin solution, and mineral solution are shown in Tables 1-3.

[0041] Table 1 50mmol L -1 Solutes and concentrations of each solute in the PBS buffer solution

[0042]

[0043] Table 2. Solutes and concentrations of each solute in the vitamin solution.

[0044]

[0045]

[0046] Table 3. Solutes and concentrations of each solute in the mineral solution.

[0047]

[0048] II. Comparative Example

[0049] Comparative Example 1

[0050] The only difference between the microbial fuel cell in this comparative example and the microbial fuel cell in Example 1 is that the anode and cathode of the microbial fuel cell in this comparative example are both bare carbon brushes, referred to as CB-MFCs.

[0051] III. Experimental Examples

[0052] After the successful start-up of nHA / CNTs / CB-MFCs and CB-MFCs, the effects of long-term diclofenac exposure on the power generation performance, microbial community structure and degradation performance of microbial fuel cells were studied by adding a certain concentration (5 mg / L, 10 mg / L) of diclofenac to the anolyte for nearly 300 days.

[0053] Experimental Example 1: Power Generation Performance

[0054] Representative dynamic changes in the output voltage of nHA / CNTs / CB-MFCs and CB-MFCs throughout the entire operation of the microbial fuel cell are as follows: Figure 1 As shown. By Figure 1 It can be seen that when the anodes of microbial fuel cells were inoculated and cultured in an anode nutrient solution containing anaerobic sludge for approximately 29 and 41 days, the maximum output voltage of nHA / CNTs / CB-MFCs and CB-MFCs reached approximately 680 mV, respectively, and maintained a relatively stable voltage output for several consecutive cycles, indicating successful anode biofilm acclimation, i.e., successful battery startup. It is worth noting that although the maximum voltages of nHA / CNTs / CB-MFCs and CB-MFCs after successful startup are similar, the startup time of nHA / CNTs / CB-MFCs is significantly shorter. nHA modification gives the carbon brushes good biocompatibility and a large specific surface area, which is conducive to the attachment and growth of more microorganisms on the electrode surface. At the same time, the excellent electrochemical activity of CNTs facilitates extracellular electron transfer between microorganisms and the electrode. These are all favorable factors for the shorter startup time of nHA / CNTs / CB-MFCs.

[0055] After the microbial fuel cell was successfully started, 5 mg·L⁻¹ was added to the anolyte of nHA / CNTs / CB-MFCs and CB-MFCs, respectively. -1 Diclofenac, from Figure 1It can be observed that the addition of diclofenac at this lower concentration caused a decrease in the voltage of both nHA / CNTs / CB-MFCs and CB-MFCs. The maximum output voltage of nHA / CNTs / CB-MFCs decreased slightly for 2-3 consecutive cycles, reaching 630±15mV; the voltage of CB-MFCs decreased more significantly, dropping to a minimum of 550±20mV. However, the voltage of both microbial fuel cells gradually recovered in the following cycles. After the voltage recovered, 10 mg·L⁻¹ of diclofenac was added to the anolyte of both nHA / CNTs / CB-MFCs and CB-MFCs. -1 Diclofenac, from Figure 1 It can be seen that under the influence of high concentrations of diclofenac, the voltage output of nHA / CNTs / CB-MFCs and CB-MFCs is greatly suppressed, dropping to 150-200 mV. Subsequently, during long-term acclimatization, the output voltage of both microbial fuel cells slowly recovered. This indicates that lower concentrations of diclofenac (<5 mg·L⁻¹) are more effective. -1 The effect of high concentrations (>10 mg·L⁻¹) on the energy output of MFC is relatively small, while high concentrations (>10 mg·L⁻¹) have a smaller impact. -1 The significant impact indicates that high concentrations of diclofenac can have a substantial impact on the anolyte biofilm, affecting the activity of electrogenic bacteria. However, with long-term domestication, the microorganisms can gradually adapt to the environment of diclofenac, and the output voltage gradually recovers. The voltage of nHA / CNTs / CB-MFCs eventually stabilized at 650±20mV, essentially recovering to the level before the addition of diclofenac; while the voltage of CB-MFCs only stabilized at 580±50mV, and the voltage output was relatively unstable. This is because the better biocompatibility of nano-hydroxyapatite facilitates the attachment and growth of more diverse microorganisms, promoting the domestication of functional microbial communities such as electroactive microorganisms and organic matter-degrading bacteria. Furthermore, in the microenvironment of the modified material with better bioactivity, the microorganisms have a better tolerance to the diclofenac impact, and their growth and metabolism are not significantly inhibited or affected.

[0056] Experimental Example 2: Diclofenac Removal Rate

[0057] In the removal rate experiment, an open-circuit experiment was set up to exclude the removal rate of diclofenac due to adsorption by various materials contained in the anode chamber of the microbial fuel cell. The microbial fuel cell in the open-circuit experiment has the same structure as nHA / CNTs / CB-MFCs, the only difference being that the anode and cathode are disconnected to form an open circuit, denoted as nHA / CNTs / CB-MFCs-open circuit.

[0058] Removal rate test method: During different cycles and times of operation of the microbial fuel cell (0h, 4h, 8h, 12h, 24h, 36h, 48h), a small amount of anolyte was taken using a sterile syringe, filtered twice consecutively through a 0.22μm microporous filter, and then analyzed by high-performance liquid chromatography (HPLC). The diclofenac removal rate was calculated. The HPLC system (Agilent Technologies 1260 Infinity) used a C18 column (150mm × 4.6mm, 5μm), with a mobile phase of methanol-1% acetic acid (75:25, v / v) and a flow rate of 1.0 mL / min. -1 The column temperature was 30℃, the detection wavelength was 275nm, and the injection volume was 10μL.

[0059] The removal rate of diclofenac varies at different stages of operation in a microbial fuel cell. With increasing operating cycles, the removal efficiency gradually increases and then stabilizes. The stable removal rates of diclofenac in different types of microbial fuel cell reactors are as follows: Figure 2 As shown, the removal rates of diclofenac in nHA / CNTs / CB-MFCs, CB-MFCs, and nHA / CNTs / CB-MFCs-open circuit were (74.62±8.56)%, (46.51±2.80)%, and (21.05±2.51)%, respectively. Firstly, the removal rates of nHA / CNTs / CB-MFCs and CB-MFCs were significantly higher than those of nHA / CNTs / CB-MFCs-open circuit, indicating that microbial degradation and electrochemical degradation are the main factors for the enhanced removal of diclofenac in MFCs, while simple physical adsorption is a secondary factor. Secondly, diclofenac achieved a higher degradation efficiency in nHA / CNTs / CB-MFCs, which is attributed to the anode modification materials of the MFCs. nHA is conducive to enriching a richer population of functional microorganisms and giving them better microbial activity, while CNTs can promote electron transfer. The synergistic effect of these two materials enabled diclofenac to achieve high microbial and electrochemical degradation efficiency in microbial fuel cells.

[0060] Experimental Example 3: Morphology of Anodic Microorganisms

[0061] The morphology of the anolyte biofilms of nHA / CNTs / CB-MFCs and CB-MFCs after 322 days of long-term operation was observed using scanning electron microscopy. Figure 3 As shown. By Figure 3 As shown in (a) and (b), both types of microbial fuel cells have a certain amount of microorganisms attached to their anode carbon brushes. However, the nHA / CNTs / CB-MFCs sample has significantly more microorganisms attached to its carbon brush, resulting in a denser biofilm, while the CB-MFCs sample has a sparser biofilm. For microbial morphology, see [link to documentation]. Figure 3(c) and (d) Both biofilm samples showed the observation of various forms of microorganisms, including cocci, bacilli and filamentous bacteria.

[0062] Experiment Example 4: Microbial Community Structure Analysis

[0063] To gain a more detailed understanding of the functional microbial community in the anode biofilm samples of microbial fuel cells after long-term operation, high-throughput sequencing technology was used to identify and compare the microbial community structure of anaerobic sludge inoculum (AS), nHA / CNT / CB-MFCs, and CB-MFCs after 322 days of operation.

[0064] The Shannon, Simpson, Sobs, Chao, and Coverage alpha diversity indices of the three groups of samples—anaerobic sludge inoculum, nHA / CNT / CB-MFCs, and CB-MFCs—are shown in Table 4. First, the microbial coverage rates in the anaerobic sludge inoculum, nHA / CNT / CB-MFCs, and CB-MFCs reached 99.80%, 99.73%, and 99.75%, respectively, and the sequencing results accurately reflect the sample information. Second, regarding diversity indices, the anaerobic sludge inoculum sample showed the highest microbial community diversity and richness, indicating that microbial community diversity decreased after long-term operation of the microbial fuel cell. This is because the current generated during MFC operation has a selective effect on the microorganisms at the anode, and the addition of diclofenac further enhances the selective pressure, inhibiting some microorganisms that cannot tolerate diclofenac or its metabolites, thus leading to a decrease in microbial community diversity and richness. Meanwhile, for both the nHA / CNT / CB-MFCs and CB-MFCs biofilm samples, the nHA / CNT / CB-MFCs sample showed a higher microbial diversity index and a richer variety of microorganisms. This is because nHA has excellent biocompatibility, which is more conducive to the growth and attachment of a wider range of microbial communities. This finding is also consistent with previous scanning electron microscopy results.

[0065] Table 4. Microbial diversity indices of Shannon, Simpson, Sobs, and Chao.

[0066] Table1 Shannon, Simpson and Chao index of bacterial community

[0067]

[0068] (1) Analysis of microbial community structure at the phylum level

[0069] Figure 4This study shows the phylum-level microbial community structure of three anode biofilm samples—anaerobic sludge inoculum, nHA / CNT / CB-MFCs, and CB-MFCs—after 322 days of operation of a microbial fuel cell. Significant differences were found in the composition and abundance of dominant phyla among the three biofilm samples. In the anaerobic sludge inoculum, Chloroflexigate, Desulfobacterota, Modullibacteria, and Bacteroidete were the dominant bacterial groups, accounting for 25.37%, 15.63%, 12.11%, and 9.79% of all sequences, respectively. Firmicutes were present in the original inoculum sludge at an abundance of only 2.73%. Compared to the anaerobic sludge inoculum, the community structure of nHA / CNT / CB-MFCs and CB-MFCs changed significantly after long-term operation.

[0070] First, from the perspective of bacterial species involved in diclofenac degradation, the abundance of Proteobacteria, Actinobacteriota, and Firmicutes was higher than that of the inoculated sludge. In the nHA / CNT / CB-MFC biofilm samples, the abundance of Proteobacteria, Actinobacteriota, and Firmicutes reached 25.52%, 10.01%, and 9.93%, respectively. In the CB-MFCs biofilm samples, the abundance of Proteobacteria and Firmicutes increased to 50.60% and 9.54%, respectively. Proteobacteria and Firmicutes are bacterial types frequently found in anaerobic environments that process chlorine-containing compounds. Furthermore, as Gram-positive bacteria, Firmicutes play a crucial role in the degradation of complex substances by converting large organic molecules into smaller ones while generating electrons. This indicates that the high proportion of Proteobacteria and Firmicutes in the anode biofilms of nHA / CNT / CB-MFCs and CB-MFCs under long-term diclofenac exposure is related to the long-term acclimatization and enhanced degradation of diclofenac.

[0071] Secondly, from the perspective of microbial species related to electricity generation, the phyla Proteobacteria, Actinobacteriota, and Firmicutes are rich in electroactive microorganisms. These microorganisms can efficiently transfer electrons from electroactive microorganisms to electrodes via cytochrome C, thereby achieving good electricity generation performance. Meanwhile, Desulfobacterota are also electroactive microorganisms, playing an important role in extracellular electron transfer. Compared to the inoculated sludge, the increased abundance of these dominant microbial communities can well explain why the two microbial fuel cells in the experiment could simultaneously achieve good electricity generation performance while degrading diclofenac. In particular, the abundance of Proteobacteria was significantly increased in nHA / CNT / CB-MFCs and CB-MFCs, reaching 25.52% and 50.60%, respectively, compared to 1.35% in the inoculated anaerobic sludge, indicating that the MFC system is conducive to the enrichment of Proteobacteria.

[0072] (2) Analysis of microbial community structure at the genus level

[0073] To gain a deeper understanding of the changes in functional microorganisms and microbial communities among three anode biofilm samples—anaerobic sludge inoculum, nHA / CNT / CB-MFCs, and CB-MFCs—further analysis was conducted on the differences at the genus level among the three samples. A heatmap was generated by clustering the top 50 species by total abundance at the taxonomic level, as shown below. Figure 5 As shown, the heatmap data can be divided into three groups based on the clustering results: the first group is the inoculum anaerobic granular sludge, the second group is the nHA / CNT / CB-MFCs biofilm samples, and the third group is the CB-MFCs biofilm samples. At the genus level, the dominant bacterial species in the inoculum anaerobic granular sludge are mainly concentrated in Moduliflexaceae, Syntrophobacter, Ardenticatenales, Syntrophales, and Anaerolineaceae. However, in the nHA / CNT / CB-MFCs and CB-MFCs samples, the proportion of these groups decreased sharply, indicating that most microorganisms in the activated sludge inoculum are sensitive and intolerant to the current generated by MFCs, and the microbial community structure of the biofilm has changed significantly after long-term operation of the inoculum and MFCs.

[0074] The dominant taxa in the second group of nHA / CNT / CB-MFCs biofilm samples were *Geobacter* (24.46%), *Sphaerochaeta* (15.62%), *Rhodopseudomonas* (9.47%), *Actinomyces* (6.20%), and hydrogen- and acetic-producing bacteria (Petrimonas, 4.13%). The dominant taxa in the third group of CB-MFCs biofilm samples were *Escherichia-Shigella* (32.8%), *Trichlorobacter* (15.2%), *Geobacter* (9.13%), *Dechlorosoma* (6.15%), and *Rhodopseudomonas* (4.36%).

[0075] Geobacter is a reported electrogenic microorganism, and due to its adaptation to low redox potential, it has become one of the most representative electroactive microorganisms in MFC research. The relative abundance of Geobacter in the anolyte biofilm of nHA / CNT / CB-MFCs was not only significantly higher than that inoculated sludge, but also much higher than its abundance of 9.13% in CB-MFC samples.

[0076] On the one hand, as a typical electroactive microorganism, experimental results show that *Geobacter* has a strong tolerance to diclofenac. This explains why microbial fuel cells exposed to diclofenac for a long time can maintain good power output after domestication. Furthermore, the better power generation performance of nHA / CNT / CB-MFCs compared to CB-MFCs may also be due to the higher abundance of *Geobacter*. On the other hand, *Geobacter* has the ability to degrade organic pollutants. In studies on the degradation of chlorophenols, it has been found that it can transfer electrons to chlorophenols via flagella for dechlorination reactions. Studies have shown that the degradation of diclofenac in modified anode MFCs is also a dechlorination process. Therefore, the high abundance of *Geobacter* is also an important reason why nHA / CNT / CB-MFCs can achieve enhanced degradation of diclofenac in this study. In summary, the modified anode of nHA / CNT / CB-MFCs is conducive to the enrichment of Geobacter, and the presence of higher abundance of Geobacter endows the microbial fuel cell with better power generation performance and stronger diclofenac degradation ability.

[0077] Furthermore, Dechlorosoma is a species of bacteria that has been reported to have degradation functions. Previous researchers have also identified this genus in their studies on the degradation of chlorophenol. In both the nHA / CNT / CB-MFCs and CB-MFCs samples of this invention, Dechlorosoma showed a certain abundance, indicating that Dechlorosoma is also a dominant bacterium obtained through long-term domestication of diclofenac and plays an important role in the degradation of diclofenac.

Claims

1. The application of microbial fuel cells in the degradation of diclofenac, characterized by: The microbial fuel cell includes an anode, a cathode, and a proton exchange membrane. The anode includes a carbon brush and a modifying material coated on the surface of the carbon brush. The modifying material includes nano-hydroxyapatite and carbon nanotubes, with a mass ratio of nano-hydroxyapatite to carbon nanotubes of 1:

4. The anode is cultured in an anode nutrient solution containing anaerobic sludge to complete the acclimatization of the anode biofilm.

2. The application according to claim 1, characterized in that: The anolyte uses phosphate buffer solution as a solvent, and the solutes are glucose, vitamins and minerals; the cathode solution corresponding to the cathode is potassium ferricyanide solution.

3. The application according to claim 1, characterized in that: The amount of the modifying material adhering to the carbon brush is 42-50 mg / g. -1 .

4. The application according to claim 1, characterized in that: The coating involves spraying a dispersion containing a modifying material onto the surface of a carbon brush; the dispersion is obtained by mixing nano-hydroxyapatite, carbon nanotubes, a dispersant, and a binder.

5. The application according to claim 4, characterized in that: The dispersant is anhydrous ethanol, and the amount of anhydrous ethanol used is 10 mL for every 6 mg of nano hydroxyapatite; the binder is a 5 wt% Nafion solution, and the amount of 5 wt% Nafion solution used is 200 mg for every 6 mg of nano hydroxyapatite.

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