Bioelectrochemical method for efficiently defluorinating and degrading perfluorinated compounds and application of bioelectrochemical method

Abstract

The invention relates to a bioelectrochemical method for efficiently removing fluorine and degrading perfluorinated compounds and application of the bioelectrochemical method, and belongs to the technical field of pollutant treatment. According to the method disclosed by the invention, the sulfate reducing flora (SRB) which takes Clostridiumsensuricto as a main dominant bacterium (the relative abundance is 46.06%) is enriched. The SRB rapidly grows on the surface of the cathode by using electrons provided by the cathode, and ferrous sulfide with conductivity is synthesized on the surface of the cathode, so that the resistance is further reduced, and electron transfer between the SRB and the cathode is promoted. The reacted cathode forms a biological membrane with Tissierella (the relative abundance is 41.32%) as a main dominant bacterium, and compared with an electrochemical method without a biological membrane, the bioelectrochemical method has the advantage that the degradation rate and the defluorination rate of perfluorinated compounds are remarkably improved. In addition, by optimizing the composition of the electrolyte, the degradation rate and the defluorination rate can be further improved, and the defluorination rate can reach 90% or above.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of pollutant treatment, and particularly relates to a bioelectrochemical method for efficiently defluorinating and degrading perfluorinated compounds and application thereof. BACKGROUND

[0002] Perfluorinated compounds (PFAS) are a kind of toxic compounds, which can be used as a moisture-proof agent of electronic products and an auxiliary agent of special powder fire-fighting foam, and have special surface activity, and have an impact on aquatic plants and the like in water environment. PFAS has been widely present in the environment, especially in the soil and water near the municipal sewage treatment plant and fluorine chemical plant. PFAS not only has environmental persistence and can be bioaccumulated, but also has a toxic effect, which poses a serious threat to human health. The carbon-fluorine bond of PFAS has high bond energy, so it has good stability and will not decompose or decompose very slowly in the natural environment. The goal of PFAS degradation is not only removal, but also mineralization, that is, in addition to the process of converting organic carbon into CO2 by breaking the carbon-carbon bond, the process of converting organic fluorine into inorganic fluoride ion (F – ) by breaking the carbon-fluorine bond, that is, defluorination, to avoid the production of toxic short-chain intermediates.

[0003] At present, the degradation technology of PFAS mainly focuses on energy-intensive physical and chemical methods, such as high-temperature pyrolysis technology and electrochemical reduction technology. Electrochemical reduction technology can theoretically break the carbon-fluorine bond through direct electron transfer or produce active substances to mediate the breaking of the carbon-fluorine bond, and can realize effective defluorination and even mineralization of PFAS in a short time. However, the electrochemical degradation defluorination technology needs a high voltage (for example, -1.8 V vs. Ag / AgCl reference electrode) and has high energy consumption, and it is difficult to realize mineralization, generally needs to modify the electrode material or combine with photocatalysis technology, and the operation is complex and cumbersome. PFAS is an anionic compound, and there is electrostatic repulsion between the cathode, which makes the electron transfer efficiency between the pollutant and the cathode poor and leads to poor degradation and defluorination effect.

[0004] Compared with energy-intensive degradation technology, microbial degradation does not require high temperature and high energy, and is a green, in-situ and sustainable method for removing pollutants, which has a strong application prospect. However, there are few reports on the effective microbial degradation of PFAS, and the reported microbial degradation of PFAS often has the problems of low degradation rate, long degradation time and low defluorination rate. Microorganisms are difficult to utilize chemically stable PFAS for dehalogenation respiration, and there are problems of limited electron transfer in the dehalogenation process and insufficient enzyme co-factor.

[0005] The dehalogenation (e.g. dechlorination, defluorination and debromination) of microorganisms is often limited by the lack of electron donors and low electron transfer efficiency. The electrochemical cathode as a stable, controllable and continuous electron source provides sufficient electron donors for microbial growth and metabolism. The combination of electrochemically active functional microorganisms and electrochemical cathode can make up for the lack of electron donors in the dehalogenation process of microorganisms. In the electrochemical system, the enzymatic reaction is enhanced due to the shortening of the electron transfer chain and the improvement of the electron transfer efficiency. The combination of microorganisms and electrochemistry can promote the transfer of electrons from electrochemically active functional microorganisms to organic fluorides, effectively promoting the degradation and defluorination of organic fluorides. SUMMARY

[0006] The present application aims to overcome the shortcomings of the prior art and provide a bioelectrochemical method for efficient defluorination and degradation of perfluorinated compounds and its application.

[0007] The present application uses sulfate-reducing bacteria as microorganisms, uses the cathode to provide sufficient electron sources for microorganisms, uses bioelectrochemistry to form a sulfate-reducing bacteria biofilm on the cathode, uses the electrically conductive ferrous sulfide synthesized by sulfate-reducing bacteria to improve the electrical conductivity of the cell membrane of the sulfate-reducing bacteria group and reduce the interfacial resistance, promotes the electron transfer between the sulfate-reducing bacteria group and the cathode, and significantly promotes the microbial degradation and defluorination of fluorochlorooctanoic acid. By adding vitamin B12, the efficiency of microbial degradation and defluorination of fluorochlorooctanoic acid is further improved. At the same time, by optimizing the concentration of Fe 2+ , the sulfate-reducing bacteria group is attached to the cathode, so that the defluorination rate is greatly improved.

[0008] The FeS nanoparticles synthesized in situ by sulfate-reducing bacteria (SRB) can be attached to the cell surface and the periplasmic layer of the cell, improve the electrical conductivity of the cell membrane, reduce the interfacial resistance, and improve the electron transfer efficiency. In addition, the extracellular polymeric substance (EPS) secreted by the microorganism itself contains quinone groups and also has the effect of electron shuttle to improve the electron transfer efficiency, and the proteins in the EPS are conducive to the combination with pollutants and the adsorption of pollutants by the biofilm, and the polysaccharide component is conducive to the formation of the cathode biofilm. The formation of the cathode biofilm helps to adsorb pollutants and enhance the electron transfer between the microbial cells and the cathode.

[0009] To achieve the above object, the technical scheme adopted by the present application is as follows: In a first aspect, the present application provides a bioelectrochemical method for efficient defluorination and degradation of perfluorinated compounds, comprising the following steps: S1, constructing a bioelectrochemical reactor, adding a sulfate-reducing bacteria group culture solution and an electrolyte into the bioelectrochemical reactor, and conducting an electric reaction to form a sulfate-reducing bacteria group cathode biofilm on the cathode and complete the start-up of the bioelectrochemical reactor; S2, adding perfluorinated compounds to the cathode of the started bioelectrochemical reactor and powering the reaction, so that the perfluorinated compounds can be degraded and defluorinated; The dominant bacteria of the sulfate-reducing bacteria group in the sulfate-reducing bacteria group culture solution at the genus level include Clostridium_sensu_stricto, Desulfotomaculum, Sporomusa, unclassified_Lutispora, Tissierella, Anaerocolumna, Caproiciproducens, uncultured_Firmicutes_bacterium, and the relative abundances thereof are 46.06%, 15.27%, 10.99%, 6.88%, 2.55%, 3.08%, 2.07%, and 1.17%, respectively.

[0010] The present application uses the sulfate-reducing bacteria group as microorganisms, provides sufficient electron sources for the microorganisms by using a cathode, forms a sulfate-reducing bacteria group biofilm on the cathode by using bioelectrochemistry, improves the conductivity of the cell membrane of the sulfate-reducing bacteria group by using the ferrous sulfide with conductivity synthesized by the sulfate-reducing bacteria, reduces the interface resistance, promotes the electron transfer between the sulfate-reducing bacteria group and the cathode, and significantly promotes the microbial degradation and defluorination of fluorochloro-octanoic acid.

[0011] The present application uses bioelectrochemistry to degrade and defluorinate perfluorinated compounds, and after degradation and defluorination (after reaction), the dominant bacteria of the sulfate-reducing bacteria group on the cathode biofilm at the genus level include Tissierella, Clostridium_sensu_stricto, Bacillaceae_bacterium_mt6, uncultured_Firmicutes_bacterium, Lachnoclostridium, Sedimentibacter, Anaerocolumna, and Caproiciproducens, and the relative abundances thereof are 41.32%, 15.67%, 9.83%, 8.44%, 6.70%, 3.24%, 1.70%, and 1.45%, respectively. After the reaction, the dominant bacteria group of the sulfate-reducing bacteria group changes, and a biofilm mainly formed by Tissierella (with a relative abundance of 41.32%) is formed.

[0012] As a preferred embodiment of the bioelectrochemical method, in step S1, the electrolyte is a sulfate-reducing bacteria culture medium; the sulfate-reducing bacteria culture medium contains ferrous ions (Fe 2+ ).

[0013] As a preferred embodiment of the bio-electrochemical method of the present application, the concentration of ferrous ions (Fe 2+ ) in the sulfate-reducing bacteria culture medium is 1-100 mg / L.

[0014] As a preferred embodiment of the bio-electrochemical method of the present application, the concentration of ferrous ions (Fe 2+ ) in the sulfate-reducing bacteria culture medium is 5-50 mg / L. The sulfate-reducing bacteria culture medium with the Fe 2+ concentration in this range has better effect on degradation and defluorination of perfluorinated compounds as electrolyte.

[0015] As a preferred embodiment of the bio-electrochemical method of the present application, the concentration of ferrous ions (Fe 2+ ) in the sulfate-reducing bacteria culture medium is 10-20 mg / L. The sulfate-reducing bacteria culture medium with the Fe 2+ concentration in this range has better effect on degradation and defluorination of perfluorinated compounds as electrolyte.

[0016] As a preferred embodiment of the bio-electrochemical method of the present application, the sulfate-reducing bacteria culture medium in step S1 further contains vitamin B12.

[0017] As a preferred embodiment of the bio-electrochemical method of the present application, the concentration of vitamin B12 in the sulfate-reducing bacteria culture medium is 100 μg / L.

[0018] As a preferred embodiment of the bio-electrochemical method of the present application, the sulfate-reducing bacteria culture medium comprises water, 0.5 g / L KH2PO4, 1.0 g / L NH4Cl, 0.5-1.5 g / L Na2SO4, 0.05 g / L CaCl2, 2.0 g / L MgCl2·6H2O, 0.1 g / L ascorbic acid, 0.1 g / L sodium mercaptoacetate, 0.005-0.5 g / L FeSO4·7H2O and 1.0 g / L yeast extract.

[0019] As a preferred embodiment of the bio-electrochemical method of the present application, the preparation method of the sulfate-reducing bacteria culture medium in step S1 is as follows: the soil source is cultured with the sulfate-reducing bacteria culture medium, and the medium is changed after the medium turns black (5 days of culture), and the sulfate-reducing bacteria culture medium is obtained after 5 days of culture.

[0020] As a preferred embodiment of the bio-electrochemical method of the present application, the soil source is petroleum hydrocarbon contaminated soil.

[0021] As a preferred embodiment of the bio-electrochemical method of the present application, the culture of the soil source includes any one of a, b and c: a, the soil of the bacteria source is dried and ground into powder, and then a suspension is prepared with normal saline, and then cultured with the sulfate-reducing bacteria culture medium; b, the soil of the bacteria source is directly cultured with the sulfate-reducing bacteria culture medium; c, the soil of the bacteria source is prepared into a suspension with sterile water, and then cultured with the sulfate-reducing bacteria culture medium.

[0022] As a preferred embodiment of the bioelectrochemical method, the sulfate-reducing bacteria culture medium comprises water, 0.5 g / L KH2PO4, 1.0 g / L NH4Cl, 1 g / L Na2SO4, 0.05 g / L CaCl2, 2.0 g / L MgCl·6H2O, 0.1 g / L ascorbic acid, 0.1 g / L sodium mercaptoacetate, 0.5 g / L FeSO4·7H2O and 1.0 g / L yeast extract.

[0023] As a preferred embodiment of the bioelectrochemical method, in the preparation method of the sulfate-reducing bacteria culture medium, the temperature of the culture is 28-37℃, and the culture time is 3-7 days.

[0024] As a preferred embodiment of the bioelectrochemical method, in the preparation method of the sulfate-reducing bacteria culture medium, the temperature of the culture is 30℃, and the culture time is 5 days.

[0025] As a preferred embodiment of the bioelectrochemical method, in step S1, the volume ratio of the sulfate-reducing bacteria culture medium to the electrolyte is 1-5:100; preferably, the volume ratio is 2:100.

[0026] As a preferred embodiment of the bioelectrochemical method, in steps S1 and S2, the bioelectrochemical reactor is a double-chamber reactor.

[0027] As a preferred embodiment of the bioelectrochemical method, the double-chamber reactor comprises a cathode chamber and an anode chamber, and the cathode chamber and the anode chamber are separated by a cation exchange membrane.

[0028] As a preferred embodiment of the bioelectrochemical method, the cathode chamber is provided with a working electrode and a reference electrode; and the anode chamber is provided with a counter electrode.

[0029] As a preferred embodiment of the bioelectrochemical method, in step S2, the voltage of the power supply is -0.6 to -1.0 V; preferably, the voltage is -0.85 V.

[0030] As a preferred embodiment of the bioelectrochemical method, in step S2, the perfluorinated compound is chlorofluoro octanoic acid (CTFE4).

[0031] As a preferred embodiment of the bio-electrochemical method of the present application, in step S2, the amount of the perfluorinated compound added is: the concentration of the perfluorinated compound in the electrolyte is not higher than 50 mg / L.

[0032] In a second aspect, the present application provides the use of the above-mentioned bio-electrochemical method in defluorination and degradation of perfluorinated compounds in sewage and / or contaminated soil.

[0033] As a preferred embodiment of the use of the present application, the perfluorinated compound is fluorochloro octanoic acid (CTFE4).

[0034] Compared with the prior art, the present application has the following beneficial effects: The present application enriches the sulfate-reducing bacteria group (SRB) with Clostridium_sensu_stricto as the main dominant bacteria (relative abundance of 46.06%). The SRB utilizes the cathode to provide sufficient electron source for microorganisms, and forms a sulfate-reducing bacteria group biofilm on the cathode through bio-electrochemistry. The ferrous sulfide synthesized by the SRB has conductivity, which improves the conductivity of the cell membrane of the sulfate-reducing bacteria group and reduces the interfacial resistance, thereby promoting the electron transfer between the SRB and the cathode. The cathode after the reaction forms a biofilm with Tissierella (relative abundance of 41.32%) as the main dominant bacteria. Compared with the electrochemical method without biofilm, the bio-electrochemical method significantly improves the degradation rate and defluorination rate of the perfluorinated compound. By adding vitamin B12 to optimize the composition of the electrolyte, the degradation and defluorination efficiency of fluorochloro octanoic acid is further improved. At the same time, by optimizing the concentration of Fe 2+ in the electrolyte, the sulfate-reducing bacteria group is promoted to adhere to the cathode, the electron transfer efficiency is improved, and the degradation efficiency and defluorination rate are greatly improved, and the defluorination rate can reach more than 90%. BRIEF DESCRIPTION OF DRAWINGS

[0035] Figure 1 It is the appearance of the sulfate-reducing bacteria group culture solution in Example 1 of the present application; Figure 2 It is the X-ray diffraction pattern of the black solid in Example 1 of the present application; Figure 3 It is the classification pie chart of the bacterial cells of the SRB culture solution at the genus level in Example 1 of the present application; Figure 4 It is the degradation and defluorination results of fluorochloro octanoic acid by pure culture of the sulfate-reducing bacteria group culture solution in Example 2 of the present application; Figure 5 It is a structural schematic diagram of the double-chamber bio-electrochemical reactor of the present application; Figure 6The degradation and defluorination results of fluocinolone acetonide using the sulfate-reducing bacteria culture medium as the electrolyte in Example 3 of this invention are shown. Figure 7 The results of the analysis of bacterial cells at the phylum level in SRB culture medium, cathode biofilm, and cathode electrolyte in Example 3 of this invention; Figure 8 The results of bacterial cell analysis at the genus level in SRB culture medium, cathode biofilm, and cathode electrolyte in Example 3 of this invention; Figure 9 The embodiment of the present invention containing VB 12 The results of the degradation and defluorination of fluorochlorooctanoic acid using a bioelectrochemical method with sulfate-reducing bacteria culture medium as electrolyte; Figure 10 Different Fe in Example 5 of the present invention 2+ Concentration containing VB 12 The degradation results of fluorochlorooctanoic acid by a bioelectrochemical method using sulfate-reducing bacteria culture medium as electrolyte; Figure 11 Different Fe in Example 5 of the present invention 2+ Concentration containing VB 12 The defluorination results of fluorochlorooctanoic acid were obtained by using a sulfate-reducing bacteria culture medium as the electrolyte in a bioelectrochemical method for 168 h. Figure 12 Different Fe in Example 5 of the present invention 2+ Concentration containing VB 12 SEM images of the cathode biofilm after 168 h of bioelectrochemical reaction using sulfate-reducing bacteria culture medium as electrolyte, where a represents 5 mg / L, b represents 10 mg / L, c represents 20 mg / L, and d represents 50 mg / L. Figure 13 The results of the electrochemical method for the degradation and defluorination of fluorochlorooctanoic acid in the comparative examples of this invention are shown. Detailed Implementation

[0036] To better illustrate the purpose, technical solution, and advantages of the present invention, the present invention will be further described below in conjunction with specific embodiments.

[0037] Unless otherwise specified, all other materials and reagents used in the examples are commercially available.

[0038] The culture medium for sulfate-reducing bacteria consisted of: water, 0.5 g / L KH₂PO₄, 1.0 g / L NH₄Cl, 1.0 g / L Na₂SO₄, 0.05 g / L CaCl₂, 2.0 g / L MgCl₆H₂O, 0.1 g / L ascorbic acid, 0.1 g / L sodium thioglycolate, and 0.5 g / L FeSO₄·7H₂O (Fe2+ (Concentration of 100 mg / L) and 1.0 g / L yeast extract.

[0039] Includes VB 12 The sulfate-reducing bacteria culture medium consisted of: water, 0.5 g / L KH₂PO₄, 1.0 g / L NH₄Cl, 1.0 g / L Na₂SO₄, 0.05 g / L CaCl₂, 2.0 g / L MgCl₆H₂O, 0.1 g / L ascorbic acid, 0.1 g / L sodium thioglycolate, and 0.5 g / L FeSO₄·7H₂O (Fe 2+ The concentrations were 100 mg / L, 1.0 g / L yeast extract, and 0.0001 g / L vitamin B12 (VB). 12 (Concentration: 100 μg / L).

[0040] In this invention, the Latin names of the strains Clostridium_sensu_stricto, Desulfotomaculum, Sporomusa, Tissierella, Anaerocolumna, Caproiciproducens, desulfovibro, Sulfurospirillum, Geobacter, Desulfuromonas, Lachnoclostridium, and Sedimentibacter are all in italics. In unclassified_Lutispora, Lutispora is in italics, and in Bacillaceae_bacterium_mt6, Bacillaceae_bacterium is in italics.

[0041] Example 1 Preparation of sulfate-reducing bacterial culture medium. This invention utilizes soil obtained from petroleum hydrocarbon contaminated sites and enriches the soil microbial community under anaerobic conditions using a sulfate-reducing bacterial culture medium to obtain sulfate-reducing bacteria capable of biosynthesizing ferrous sulfide.

[0042] Fresh petroleum hydrocarbon-contaminated soil was collected from near an oil refinery in Dingbian County, Yulin City, Shaanxi Province, as the source soil for inoculum. After drying at room temperature, the soil was ground and passed through a 2 mm sieve to remove impurities, yielding soil powder. 1 g of soil powder was weighed and added to 20 mL of physiological saline. The mixture was shaken at 30°C and 180 rpm for 1 hour, then allowed to stand for 1 hour. 1 mL of the supernatant was then added to 9 mL of physiological saline to prepare a soil suspension. 1 mL of the soil suspension was added to 1 L of anaerobic sulfate-reducing bacteria culture medium and cultured anaerobically at 30°C for 5 days until the medium turned completely black. The culture was then transferred to 1 L of fresh anaerobic sulfate-reducing bacteria culture medium and cultured anaerobically at 30°C for 5 days to obtain a sulfate-reducing bacteria culture broth (SRB culture broth).

[0043] The appearance of the sulfate-reducing bacterial culture medium is as follows: Figure 1 As shown, the sulfate-reducing bacterial culture medium of the present invention is a black suspension containing a black precipitate.

[0044] The sulfate-reducing bacterial culture was centrifuged (5000 rpm), the precipitate was collected and washed with oxygen-free water, and the washed precipitate was freeze-dried for 24 h to obtain a black solid.

[0045] The black solid was characterized using a combined multifunctional horizontal X-ray diffractometer. The X-ray diffraction pattern of the black solid is shown below. Figure 2 As shown in the results, compared with the FeS standard card, the black precipitate in the SRB culture medium is FeS precipitate. This indicates that the sulfate-reducing bacterial culture medium obtained in this invention contains sulfate-reducing bacteria capable of synthesizing ferrous sulfide.

[0046] The obtained sulfate-reducing bacterial culture can be directly used for the degradation and defluorination of fluorochlorooctanoic acid in bioelectrochemical processes.

[0047] After one generation of subculturing, the sulfate-reducing bacterial culture was mixed with 30% glycerol at a 1:1 ratio and stored at -80℃. Before use, the stored sulfate-reducing bacterial culture needs to be activated using LB liquid medium containing 1 g / L Na₂SO₄. The sulfate-reducing bacteria were cultured to the logarithmic growth phase (OD₆₀₀ = 0.8). The activated solution was then used for the bioelectrochemical degradation and defluorination of fluorochlorooctanoic acid.

[0048] DNA was extracted from sulfate-reducing bacterial culture medium (SRB medium), 16S RNA was amplified, and the composition of the bacterial community was analyzed.

[0049] 16SRNA sequencing was performed on the V3-V4 region using the Illumina NovaSeq sequencing platform. OTU sequences were obtained through similarity clustering (using a similarity score of 0.97). Primer information: (F: ACTCCTACGGGAGGCAGCA; R: GGACTACHVGGGTWTCTAAT). Sequencing was performed by Beijing Biomarker Biotechnology Co., Ltd.

[0050] A pie chart showing the classification of bacteria at the genus level in SRB culture medium is shown below. Figure 3 As shown. The dominant sulfate-reducing bacteria at the genus level in the sulfate-reducing bacterial culture medium included Clostridium sensu stricto, Desulfotomaculum, Sporomausa, unclassified Lutispora, Tissierella, Anaerocolumna, Caproiciproducens, and uncultured Firmicutes bacterium, with relative abundances of 46.06%, 15.27%, 10.99%, 6.88%, 2.55%, 3.08%, 2.07%, and 1.17%, respectively.

[0051] Example 2 The sulfate-reducing bacterial culture broth obtained in Example 1 was used to degrade fluorochlorooctanoic acid (CTFE4). Fluorochlorooctanoic acid was added to the sulfate-reducing bacterial culture medium, and the effect of the sulfate-reducing bacteria on fluorochlorooctanoic acid was tested. Specifically, 60 mL of sulfate-reducing bacterial culture medium (FeSO4·7H2O concentration of 0.5 g / L) was placed in a 100 mL anaerobic serum bottle. Three parallel controls and a blank control were performed for each experiment. Nitrogen gas was passed through the anaerobic serum bottle and culture medium at a flow rate of 0.2 L / min for 30 min to remove oxygen, followed by sterilization. Then, CTFE4 methanol solution (CTFE4 methanol solution prepared with methanol, concentration of 10 mM) was added to the anaerobic environment until the CTFE4 concentration in the culture medium reached 24 mg / L. A 2% sulfate-reducing bacterial culture medium was added as an inoculum, and the culture was incubated statically at 30℃ in the dark for 90 days. Samples were taken at 0, 4, 8, 12, 16, 20, 35, 50, 70, and 90 days (2 mL each time). Immediately after sampling, the samples were centrifuged at 16000 g at 4℃ for 15 min. The bacterial cells were stored at -80℃ for subsequent biological analysis, and the supernatant was stored at 4℃ for the determination of CTFE4 and fluoride ions.

[0052] The concentration of CTFE4 was determined using ultra-high performance liquid chromatography (UHPLC) coupled to an API 5500 triple quadrupole mass spectrometer. UHPLC separation was performed using a C18 column.

[0053] Fluoride ion concentration was determined using an ion-selective electrode. Initially, concentrations of 0, 0.1, 0.25, 0.5, 1, 5, and 10 ppm F were measured. － A standard solution was used to plot a calibration curve. 1 mL of the supernatant sample after centrifugation was diluted with 2 mL of deionized water, followed by 3 mL of TISAB II solution. The ISE electrode was then immersed in the mixed solution to determine the fluoride ion concentration. The defluorination rate (def.) was calculated using the standard curve. - Divide the molar concentration by the molar concentration of fluorine atoms in the sample on day 0, and then multiply by 100%.

[0054] The results of pure culture of sulfate-reducing bacteria on the degradation and defluorination of fluorochlorooctanoic acid are as follows: Figure 4 As shown, the results indicated that after culturing sulfate-reducing bacterial culture medium with fluorochlorooctanoic acid for 90 days, the concentration of CTFE4 in the medium significantly decreased, while the concentration of fluoride ions significantly increased, indicating that CTFE4 underwent degradation and defluorination to produce fluoride ions. After 90 days of culture, 90.2% of CTFE4 was degraded, and 254.84 μM of fluoride ions were produced. － The defluorination rate reached 46.3%. This indicates that the sulfate-reducing bacterial culture medium can effectively degrade and defluorinate fluorochlorooctanoic acid (CTFE4), demonstrating that not only carbon-carbon and carbon-chlorine bonds are broken during the degradation of CTFE4, but also carbon-fluorine bonds. This proves that the sulfate-reducing bacterial culture medium of this invention contains bacteria with the functions of degrading CTFE4 and defluorinating it.

[0055] Example 3 A two-chamber bioelectrochemical reactor was used as the reactor, and the sulfate-reducing bacterial culture broth from Example 1 was used as the microorganism to degrade fluorochlorooctanoic acid by a bioelectrochemical method.

[0056] A schematic diagram of the structure of a two-chamber bioelectrochemical reactor is shown below. Figure 5 As shown, the dual-chamber bioelectrochemical reactor is divided into a cathode chamber and an anode chamber by a cation exchange membrane. The cathode chamber is equipped with a working electrode and a reference electrode, and the anode chamber is equipped with a counter electrode. The working electrode is raw carbon paper (30 mm × 25 mm × 0.19 mm). Before use, the raw carbon paper is soaked in acetone solution for 2 hours, then washed with ethanol and air-dried. The reference electrode is an Ag / AgCl electrode. The counter electrode is a platinum sheet electrode (10 mm × 10 mm × 0.1 mm). The cathode chamber and the anode chamber are also equipped with one or more sealable sampling ports. The effective volume of a single electrode chamber is approximately 180 mL, and the solution volume is 160 mL.

[0057] 1. Bioelectrochemical methods for the degradation of fluorochlorooctanoic acid Sulfate-reducing bacteria culture medium was added to a two-chamber bioelectrochemical reactor. Then, 2% sulfate-reducing bacteria culture medium was added to the cathode. The reactor was turned on and reacted at -0.85 V for 3 days to form a cathode biofilm. Then, 24 mg / L of fluorochlorooctanoic acid (CTFE4) was added to the cathode and reacted at -0.85 V for 192 h. Samples were taken at 0, 24, 48, 96, 144, and 192 h, and the bacteria were isolated and collected according to the method in Example 2 for later use. The supernatant was collected to test the CTFE4 and fluoride ion concentrations.

[0058] The bioelectrochemical method using sulfate-reducing bacteria culture medium as the electrolyte for the degradation and defluorination of fluorochlorooctanoic acid yields the following results: Figure 6 As shown, the results indicate that sulfate-reducing bacteria culture medium, as a bioelectrochemical electrolyte, can rapidly degrade and defluorinate CTFE4. Within 192 hours, 97.7% of CTFE4 can be degraded and the defluorination rate reaches 42.3%.

[0059] 2. Effects of bioelectrochemical methods on the degradation of fluorochlorooctanoic acid on sulfate-reducing bacteria in sulfate-reducing bacterial cultures. The sulfate-reducing bacterial culture medium (SRB culture medium) of Example 1, the cathode biofilm (cathode biofilm) after 192 h of bioelectrochemical degradation of fluorochlorooctanoic acid, and the electrolyte (cathode electrolyte) after 192 h of bioelectrochemical degradation of fluorochlorooctanoic acid were used as samples to test the bacterial species classification of the samples.

[0060] DNA was extracted from bacteria in SRB culture medium, cathode biofilm, and cathode electrolyte, and 16S RNA was amplified and the composition of the bacterial community was analyzed.

[0061] 16SRNA sequencing was performed on the V3-V4 region using the Illumina NovaSeq sequencing platform. OTU sequences were obtained through similarity clustering (using a similarity score of 0.97). Primer information: (F: ACTCCTACGGGAGGCAGCA; R: GGACTACHVGGGTWTCTAAT). Sequencing was performed by Beijing Biomarker Biotechnology Co., Ltd.

[0062] The phylum-level analysis results of bacteria in SRB culture medium, cathode biofilm, and cathode electrolyte are as follows: Figure 7As shown, in the SRB culture medium, cathode biofilm, and cathode electrolyte, Firmicutes were the dominant phylum among sulfate-reducing bacteria, with relative abundances of 97.06%, 93.70%, and 95.94%, respectively. Actinobacteria were the second most abundant, with relative abundances of 1.55%, 3.83%, and 1.05%, respectively. Proteobacteria were the third most abundant, with relative abundances of 0. The relative abundances of Bacteroides were 0.32%, 2.34%, and 2.15%, respectively. It also contained small amounts of Bacteroides, with relative abundances of 0.21%, 0.12%, and 0.57% in SRB culture medium, cathode biofilm, and cathode electrolyte, respectively. This is consistent with the literature reports that Firmicutes contains various facultative and obligate dehalogenating bacteria, and that Proteobacteria such as desulfovibro, Sulfurospirillum, Geobacter, and Desulfuromonas all possess facultative dehalogenating abilities.

[0063] The relative abundance of Actinobacteria increased from 1.55% (SRB culture medium) to 3.83% (cathode biofilm), achieving enrichment on the biofilm. The relative abundance of Proteobacteria increased from 0.32% (SRB culture medium) to 2.34% (cathode biofilm) and 2.15% (cathode electrolyte), also achieving enrichment on the biofilm.

[0064] The results of bacterial cell analysis at the genus level in SRB culture medium, cathode biofilm, and cathode electrolyte are as follows: Figure 8 As shown.

[0065] The results showed that, before degradation, the dominant sulfate-reducing bacteria in the sulfate-reducing bacterial culture medium (SRB medium) at the genus level included Clostridium sensu stricto, Desulfotomaculum, Sporomausa, unclassified Lutispora, Tissierella, Anaerocolumna, Caproiciproducens, and uncultured Firmicutes bacterium, with relative abundances of 46.06%, 15.27%, 10.99%, 6.88%, 2.55%, 3.08%, 2.07%, and 1.17%, respectively. The relative abundances of Clostridium sensu stricto_10, Clostridium sensu stricto_13, Clostridium sensu stricto_1, Clostridium sensu stricto_12, and Clostridium sensu stricto_8 in Clostridium sensu stricto were 13.02%, 11.26%, 10.54%, 8.57%, and 2.67%, respectively.

[0066] The dominant sulfate-reducing bacteria on the cathode biofilm at the genus level included *Tissierella*, *Clostridium sensu stricto*, *Bacillaceae bacterium mt6*, *uncultured Firmicutes bacterium*, *Lachnoclostridium*, *Sedimentibacter*, *Anaerocolumna*, and *Caproiciproducens*, with relative abundances of 41.32%, 15.67%, 9.83%, 8.44%, 6.70%, 3.24%, 1.70%, and 1.45%, respectively. Within the *Clostridium sensu stricto* group, *Clostridium sensu stricto* 13, 10, 11, 12, and 8 accounted for approximately 5.21%, 4.94%, 3.23%, 1.16%, and 1.12%, respectively.

[0067] The dominant sulfate-reducing bacteria in the cathode electrolyte at the genus level included *Tissierella*, *Clostridium sensu stricto*, *Bacillaceae bacterium mt6*, *uncultured Firmicutes bacterium*, *Lachnoclostridium*, *Sedimentibacter*, *Anaerocolumna*, and *Caproiciproducens*, with relative abundances of 29.50%, 29.28%, 10.88%, 7.00%, 5.06%, 4.86%, 1.83%, and 1.29%, respectively. Among *Clostridium* species, *Clostridium sensu stricto* 13, 1, 10, 12, and 8 accounted for 11.88%, 10.41%, 3.25%, 2.84%, and 0.9%, respectively.

[0068] Tissierella was significantly enriched in the bioelectrochemical system, becoming the dominant genus on the cathode biofilm, with its abundance increasing from 2.55% to 41.32%, and also the main dominant bacterium in the electrolyte, with an abundance of 29.50%. This indicates that Tissierella has a strong extracellular electron transfer capacity and may be a potential electroactive defluorinating bacterium. This is consistent with literature reports that Tissierella can promote the dechlorination of trichloroethylene by promoting electron transfer. The relative abundance of Bacillaceae_bacterium_mt6 increased from 0.28% to 9.83% (cathode biofilm) and 10.88% (cathode electrolyte), the relative abundance of Lachnoclostridium increased from 0.37% to 6.70% (cathode biofilm) and 5.06% (cathode electrolyte), and the relative abundance of Sepimentibacter increased from 0.43% to 3.24% (cathode biofilm) and 4.86% (cathode electrolyte). The relative abundance of Clostridium sensu stricto decreased from 46.06% to 15.67% (cathode biofilm) and 29.28% (cathode electrolyte), the relative abundance of Sporomorusa decreased from 10.99% to 0.78%, and the relative abundance of Desulfotomaculum decreased from 15.27% to 0.35% (cathode biofilm) and 1.52% (cathode electrolyte).

[0069] According to literature reports, Clostridium can become a dominant genus on the cathode biofilm in bioelectrochemistry, possessing electron transfer and hydrogen production capabilities. However, its role in promoting the degradation and defluorination of perfluorinated compounds is very limited. Although this study enriched a sulfate-reducing bacterial community with Clostridium sensu stricto as the first dominant genus (46.06%), its abundance in the cathode biofilm decreased to 15.67%, becoming the second dominant genus, replaced by Tissierella, which reached a high abundance of 41.32%. However, Clostridium sensu stricto maintained an abundance of 29.28% in the electrolyte, essentially matching the abundance of Tissierella (29.5%). In bioelectrochemistry, the cathode is the interface that directly gains electrons, indicating that Tissierella has a stronger electron transfer capacity than Clostridium sensu stricto, and also exhibits stronger degradative and defluorinating abilities for perfluorinated and polyfluoroalkyl compounds.

[0070] Example 4 The bioelectrochemical method for degrading fluorochlorooctanoic acid is the same as in Example 3, except that the electrolyte is replaced with a culture medium containing vitamin B12 (VB12) instead of the sulfate-reducing bacteria culture medium used in Example 3. 12 Sulfate-reducing bacteria culture medium, the culture medium contains VB 12 The concentration is 100 μg / L. Specifically, Will contain VB 12 Sulfate-reducing bacteria culture medium was added to a two-chamber bioelectrochemical reactor. Then, 2% sulfate-reducing bacteria culture medium was added to the cathode. The reactor was turned on and reacted at -0.85 V for 3 days to form a cathode biofilm. Then, 24 mg / L of fluorochlorooctanoic acid (CTFE4) was added to the cathode and reacted at -0.85 V for 96 h. Samples were taken at 0, 12, 24, 48, 72 and 96 h, and the bacteria were isolated and collected according to the method in Example 2 for later use. The supernatant was collected to test the CTFE4 and fluoride ion concentrations.

[0071] Includes VB 12 The bioelectrochemical method using sulfate-reducing bacteria culture medium as electrolyte for the degradation and defluorination of fluorochlorooctanoic acid yielded the following results: Figure 9 As shown, the results indicate that VB is included. 12 The sulfate-reducing bacteria culture medium can improve the efficiency of sulfate-reducing bacteria in the degradation and defluorination of CTFE4 in bioelectrochemical methods, achieving complete degradation of CTFE4 and a defluorination rate of 45% within 96 hours; the degradation rate and defluorination rate are higher than those without VB. 12The culture medium concentrations were increased by more than two times. Compared to degradation using sulfate-reducing bacterial cultures alone, the degradation time of CTFE4 was shortened from 90 days (incomplete degradation) to 4 days (complete degradation), a reduction of 22.5 times in reaction time while achieving almost the same defluorination rate. This indicates that VB-containing... 12 The sulfate-reducing bacteria culture medium under bioelectrochemical culture conditions can promote the degradation and defluorination of fluorochlorooctanoic acid by sulfate-reducing bacteria, significantly shortening the degradation time.

[0072] Example 5 Different Fe 2+ The effect of a bioelectrochemical method using sulfate-reducing bacteria culture medium of a certain concentration as the electrolyte on the degradation and defluorination of fluorochlorooctanoic acid.

[0073] 1. Different Fe 2+ Concentration containing VB 12 Sulfate-reducing bacteria culture medium was used as the electrolyte, containing VB. 12 The sulfate-reducing bacteria culture medium consisted of: water, 0.5 g / L KH₂PO₄, 1.0 g / L NH₄Cl, 1.0 g / L Na₂SO₄, 0.05 g / L CaCl₂, 2.0 g / L MgCl₆H₂O, 0.1 g / L ascorbic acid, 0.1 g / L sodium thioglycolate, FeSO₄·7H₂O, 1.0 g / L yeast extract, and 0.0001 g / L vitamin B12 (VB₂). 12 ); The concentration of FeSO4·7H2O in the culture medium was 0.025 g / L (Fe 2+ Concentration of 5 mg / L), 0.05 g / L (Fe 2+ Concentration of 10 mg / L), 0.1 g / L (Fe 2+ Concentration of 20 mg / L), 0.25 g / L (Fe 2+ (Concentration is 50 mg / L).

[0074] Different Fe 2+ Concentration containing VB 12 Sulfate-reducing bacteria culture medium was added to a two-chamber bioelectrochemical reactor. Then, 2% sulfate-reducing bacteria culture medium was added to the cathode. The reactor was turned on and reacted at -0.85 V for 3 days to form a biofilm at the cathode. Then, 24 mg / L of fluorochlorooctanoic acid (CTFE4) was added to the cathode and reacted at -0.85 V for 168 h. Samples were taken at 0, 12, 24, 36, 48, 72, 96, 120, 144, and 168 h, and the bacterial cells were isolated and collected according to the method in Example 2 for later use. The supernatant was collected to test the CTFE4 and fluoride ion concentrations.

[0075] Different Fe 2+Concentration containing VB 12 The degradation results of fluorochlorooctanoic acid by a bioelectrochemical method using sulfate-reducing bacteria culture medium as the electrolyte are as follows: Figure 10 As shown, different Fe 2+ Concentration containing VB 12 The defluorination results of fluorochlorooctanoic acid were obtained by using a sulfate-reducing bacteria culture medium as the electrolyte in a bioelectrochemical method for 168 hours. Figure 11 As shown, the results indicate that with Fe 2+ With increasing concentration (5-50 mg / L), the degradation effect of CTFE4 initially increases and then decreases. 2+ At a concentration of 5 mg / L, complete degradation of CTFE4 requires 96 hours, with a defluorination rate of 75.59%. 2+ At concentrations of 10 and 20 mg / L, CTFE4 degradation was more effective, with near-complete degradation achieved at 36 and 48 h, respectively, and defluorination rates of 91.86% and 90.38% at 168 h, respectively. 2+ When the concentration increased to 50 mg / L, the degradation effect of CTFE4 weakened, requiring 72 hours to completely degrade, and the defluorination rate decreased to 74.41%.

[0076] 2. After 168 hours of reaction, the cathode biofilm was removed from the anaerobic chamber and fixed with 2.5% glutaraldehyde solution at 4°C for 8 hours (overnight). It was then washed three times with PBS buffer and subsequently dehydrated in a gradient of 25%, 50%, 75%, 90%, and 100% ethanol, with each concentration dehydrating for 20 minutes. Afterward, it was replaced three times with tert-butanol and finally freeze-dried under vacuum. The biofilm was observed using a scanning electron microscope.

[0077] Different Fe 2+ Concentration containing VB 12 The SEM image of the cathode biofilm after 168 h of reaction using sulfate-reducing bacteria culture medium as the electrolyte is shown below. Figure 12 As shown, a represents 5 mg / L, b represents 10 mg / L, c represents 20 mg / L, and d represents 50 mg / L. The results show that with Fe... 2+ With increasing Fe concentration (5-50 mg / L), the attachment of sulfate-reducing bacteria on the cathode initially increased and then decreased, indicating that at 10-20 mg / L Fe... 2+ At higher concentrations, this facilitates microbial attachment to the cathode. Direct cell-cathode contact improves electron transfer efficiency, promotes CTFE4 degradation, and increases defluorination rate. 2+ At concentrations of 10 and 20 mg / L, the sulfate-reducing bacteria showed the highest attachment amount on the cathode, while Fe... 2+ As the concentration continued to increase, almost no microbial cell attachment was observed on the biofilm. Fe2+ It is toxic to cells; at excessively high concentrations, it is detrimental to cell growth, reduces cell activity, or even leads to cell death. In this invention, high concentrations of Fe... 2+ The absence of microbial cells may be due to cell death, leading to a rapid decrease in CTFE4 degradation and defluorination rate.

[0078] Comparative Example The effects of electrochemical methods on fluorochlorooctanoic acid were tested using an electrochemical workstation.

[0079] Add sulfate-reducing bacteria culture medium (without VB) to the reactor connected to the electrochemical workstation. 12 As an electrolyte, 24 mg / L of fluorochlorooctanoic acid (CTFE4) was added to the cathode, and the reaction was carried out at a voltage of -0.85 V. Samples were taken at different times (0, 6, 12, 24, 36, 48, 72, 96, 120, 144 h) and the concentrations of CTFE4 and fluoride ions were separated and tested according to the method in Example 2.

[0080] The electrochemical methods for the degradation and defluorination of fluorochlorooctanoic acid are as follows: Figure 13 As shown, the results indicate that at -0.85 V, the electrochemical degradation of CTFE4 without the addition of VB... 12 Under these conditions, less than 20% of CTFE4 was degraded after 144 hours, with a defluorination rate of only 2.24%, indicating that electrochemical conditions cannot effectively degrade and defluorinate fluorochlorooctanoic acid.

[0081] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit the scope of protection of the present invention. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can be made to the technical solutions of the present invention without departing from the essence and scope of the technical solutions of the present invention.

Claims

1. A highly efficient bioelectrochemical method for defluorinating and degrading perfluorinated compounds, characterized in that, Includes the following steps: S1. Construct a bioelectrochemical reactor, add sulfate-reducing bacteria culture medium and electrolyte to the bioelectrochemical reactor, pass electricity to react, and form a cathode biofilm of sulfate-reducing bacteria at the cathode to complete the start-up of the bioelectrochemical reactor. S2. Adding perfluorinated compounds to the cathode of the bioelectrochemical reactor after startup and passing electricity through it can achieve the degradation and defluorination of perfluorinated compounds. The dominant sulfate-reducing bacteria in the sulfate-reducing bacterial culture medium at the genus level included Clostridium sensu stricto, Desulfotomaculum, Sporomausa, unclassified Lutispora, Tissierella, Anaerocolumna, Caproiciproducens, and uncultured Firmicutes bacterium, with relative abundances of 46.06%, 15.27%, 10.99%, 6.88%, 2.55%, 3.08%, 2.07%, and 1.17%, respectively.

2. The bioelectrochemical method as described in claim 1, characterized in that, In step S1, the electrolyte is a sulfate-reducing bacteria culture medium; the sulfate-reducing bacteria culture medium contains ferrous ions.

3. The bioelectrochemical method as described in claim 2, characterized in that, The concentration of ferrous ions in the sulfate-reducing bacteria culture medium is 1~100 mg / L.

4. The bioelectrochemical method as described in claim 2 or 3, characterized in that, The sulfate-reducing bacteria culture medium also contains vitamin B12.

5. The bioelectrochemical method as described in claim 1, characterized in that, The method for preparing the sulfate-reducing bacterial culture medium is as follows: the source soil is cultured using a sulfate-reducing bacterial culture medium. When the culture medium turns completely black, it is replaced with a fresh sulfate-reducing bacterial culture medium and cultured for 5 days to obtain the sulfate-reducing bacterial culture medium.

6. The bioelectrochemical method as described in claim 5, characterized in that, The soil from which the bacteria originated was contaminated with petroleum hydrocarbons.

7. The bioelectrochemical method as described in claim 5, characterized in that, In the preparation method of the sulfate-reducing bacterial culture medium, the culture temperature is 28~37℃ and the culture time is 3~7 days.

8. The bioelectrochemical method as described in claim 1, characterized in that, In step S1, the volume ratio of the sulfate-reducing bacterial culture medium to the electrolyte is 1~5:100; And / or, in steps S1 and S2, the bioelectrochemical reactor is a two-chamber reactor.

9. The bioelectrochemical method as described in claim 1, characterized in that, In step S2, the voltage applied is -0.6 to -1.0V; And / or, in step S2, the perfluorinated compound is fluorochlorooctanoic acid.

10. The application of the bioelectrochemical method according to any one of claims 1 to 9 in the defluorination and degradation of perfluorinated compounds in wastewater and / or contaminated soil.

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