Efficient and sustainable denitrification method for anaerobic ammonium oxidation bacteria

Abstract

The invention provides an anaerobic ammonium oxidation bacteria efficient sustainable denitrification method, which comprises the following steps: inoculating a biological hybridization system constructed by anaerobic ammonium oxidation bacteria and cadmium sulfide in an anaerobic ammonium oxidation reactor, adding nitrate-containing wastewater and a sacrificial agent, and applying light for treatment. The cadmium sulfide is prepared by reacting cadmium chloride with thioacetamide in polyethylene glycol, and the cadmium sulfide and anaerobic ammonium oxidation bacteria liquid are mixed and incubated to form a hybrid system. According to the method, photo-induced electrons of cadmium sulfide are used for driving anaerobic ammonium oxidation bacteria, nitrate generated by metabolism of the anaerobic ammonium oxidation bacteria is converted into nitrite and ammonium through a way of reducing dissimilatory nitrate into ammonium, and then the nitrite and the ammonium are converted into nitrogen through an anaerobic ammonium oxidation way, so that conversion and deep removal of the nitrate are realized in the system. The nitrogen removal rate limit of 89% of a traditional process is broken through, the total nitrogen removal rate can reach 98% or above, no organic carbon source is needed, no greenhouse gas nitrous oxide is discharged, solar driving is utilized, and the process is clean, efficient and sustainable.

Description

Technical Field

[0001] This invention relates to the field of environmental protection and wastewater treatment technology, specifically to a highly efficient and sustainable denitrification method using anaerobic ammonia-oxidizing bacteria. Background Technology

[0002] Anaerobic ammonia oxidation (Anammox) is a novel autotrophic biological nitrogen removal process that directly converts ammonium and nitrite into nitrogen gas under anaerobic conditions. Compared with traditional nitrification-denitrification nitrogen removal technologies, this process does not require an external organic carbon source, significantly reducing aeration energy consumption and excess sludge production. It offers significant economic and environmental advantages for treating wastewater with high ammonia nitrogen and low carbon-to-nitrogen ratios (such as sludge digestate, landfill leachate, and semiconductor wastewater).

[0003] Although Anammox technology can achieve highly efficient autotrophic nitrogen removal, the inherent nitrate production (approximately 11% of the total nitrogen in the influent) in its metabolic mechanism limits the upper limit of total nitrogen removal efficiency. To address this issue, existing technologies often couple the denitrification process with Anammox by adding organic carbon sources or inorganic electron donors. However, traditional denitrification pathways inevitably produce greenhouse gas emissions, such as nitrous oxide. Simultaneously, these exogenous electron donors easily induce excessive proliferation of denitrifying bacteria in the Anammox system. These denitrifying bacteria compete with Anammox bacteria (AnAOB) for nitrite substrates and ecological niches, thus severely inhibiting AnAOB activity and making it difficult to achieve stable operation of the Anammox process.

[0004] Current research indicates that AnAOB possesses not only a single denitrification function but also a highly flexible nitrogen metabolism pathway. In addition to the core Anammox pathway, the AnAOB genome encodes a complete enzyme system related to the dissimilatory nitrate reduction to ammonium (DNRA) pathway. Previous studies have shown that AnAOB can drive DNRA through small organic molecules or intracellular glycogen, reducing nitrate to nitrite and ammonium. However, the addition of small organic molecules easily induces the proliferation of heterotrophic denitrifying bacteria, and AnAOB's utilization of endogenous glycogen for nitrate reduction is limited by its own reserves, making it unsustainable. Therefore, developing strategies to effectively drive AnAOB to remove nitrate is crucial for achieving integrated, sustainable, and deep denitrification.

[0005] AnAOB is an electroactive bacterium with a complete electron transport pathway and outer membrane cytochrome, enabling it to directly take up external electrons. After taking up external electrons (such as those from electrodes and photoelectric conversion materials), microorganisms can convert them into endogenous reducing forces (such as NADH and ATP), thereby driving intracellular reduction reactions. Therefore, it is hoped that external electrons can drive nitrate reduction in AnAOB. ​​In recent years, solar energy, as an inexhaustible and clean energy source on Earth, has become an important energy source for bioavailable electrons. Therefore, using solar energy as an energy source for external electrons, combined with the diverse nitrogen metabolism pathways of AnAOB, can achieve highly efficient integrated denitrification based on AnAOB and surpass the theoretical denitrification rate (89%) of the Anammox process. Summary of the Invention

[0006] The purpose of this invention is to provide an efficient and sustainable denitrification method using anaerobic ammonia oxidizing bacteria to solve the problem of nitrate production inherent in traditional anaerobic ammonia oxidation processes.

[0007] To achieve the above objectives, the present invention adopts the following technical solution: A highly efficient and sustainable nitrogen removal method using anaerobic ammonia-oxidizing bacteria includes the following steps: Step S1: Inoculate the anaerobic ammonia oxidation reactor with an anaerobic ammonia oxidizing bacteria-cadmium sulfide biohybrid system; Step S2: Pass nitrate-containing wastewater into the anaerobic ammonia oxidation reactor and add a sacrificial agent; Step S3: Apply light to the anaerobic ammonia oxidation reactor to treat the nitrate-containing wastewater.

[0008] Preferably, in step S1, the preparation method of the bio-hybrid system is as follows: Step S11: Add cadmium sulfide powder to the anaerobic ammonia-oxidizing bacteria solution and mix thoroughly to obtain the first mixture; Step S12: Incubate the first mixture obtained in step S11 in a constant temperature shaker to obtain an anaerobic ammonia oxidizing bacteria-cadmium sulfide biohybrid system.

[0009] Preferably, the cadmium sulfide powder is prepared by the following method: Step S111: Add cadmium chloride to polyethylene glycol and stir to dissolve, forming the first colloid; Step S112: Add thioacetamide powder to the first colloid and stir continuously to form a second colloid; Step S113: Let the second colloid stand at 25°C for 12 hours to obtain the third colloid; Step S114: The third colloid is washed and centrifuged 6 times with deionized water, then washed and centrifuged 6 times with anhydrous ethanol to thoroughly remove the residue. After drying at 25°C for 24 hours, it is ground and sieved to obtain cadmium sulfide powder.

[0010] Preferably, in step S111, the addition ratio of cadmium chloride to polyethylene glycol is 1 mol: 175 mL, and the stirring time is 15 min; in step S112, the molar ratio of cadmium chloride to thioacetamide is 1:1, and the stirring time is 30 min; in step S113, the centrifugation conditions must at least meet the following requirements: rotation speed of 10000 rpm, time of 10 minutes, and temperature of 10 °C.

[0011] Preferably, the amount of cadmium sulfide powder added is controlled at 0.1~5 g / L.

[0012] Preferably, the anaerobic ammonia-oxidizing bacteria solution includes Broccoli spp. ( Candidatus Brocadia, Jeterella spp. Candidatus Jettenia, Cullella spp. Candidatus Kuenenia, Stephania ( ) Candidatus Scalindua, genus of anaerobic ammonia-oxidizing cocci ( Candidatus At least one of the anaerobic ammonia-oxidizing bacteria genus such as Anammoxoglobus.

[0013] Preferably, in step S11, the concentration of the anaerobic ammonia-oxidizing bacteria solution is controlled at OD. 600 The dissolved oxygen concentration is 0.5~2.0; in step S12, the incubation is strictly anaerobic, with a dissolved oxygen concentration of less than 0.05 mg / L, an incubation time of 24~96 h, a temperature of 35~37 ℃, and a shaking speed of 100~300 rpm.

[0014] Preferably, in step S2, the concentration of the nitrate wastewater is in the range of 0.1~100 mmol N / L; the sacrificial agent is at least one of sodium lactate, formic acid, acetic acid, oxalic acid, triethanolamine, cysteine, ascorbic acid, disodium ethylenediaminetetraacetate, sodium sulfite and sodium sulfide, and its dosage is 0.001~1 wt%.

[0015] Preferably, in step S3, the illumination wavelength can be 200~1000 nm, and can be monochromatic light or multi-wavelength mixed light; the illumination intensity can be 0.1~1000 mW / cm². 2 The lighting mode can be either continuous or intermittent.

[0016] Preferably, in step S1, the anaerobic ammonia oxidation reactor is an anaerobic serum bottle, an anaerobic membrane bioreactor, an upflow anaerobic sludge bed reactor, or an anaerobic expanded granular sludge bed reactor; the inoculum amount of the anaerobic ammonia oxidizing bacteria-cadmium sulfide biohybrid system is 5-95% of the volume of the anaerobic ammonia oxidation reactor.

[0017] Compared with the prior art, the present invention has the following beneficial effects: (1) This invention promotes the removal of nitrate by AnAOB through the construction of AnAOB-CdS biohybrid system. When the nitrate concentration in the influent is 4 mM, the removal rate can reach 100%.

[0018] (2) The AnAOB-CdS biohybrid system constructed in this invention can be used in conjunction with the Anammox process, and the overall denitrification rate can be increased to more than 98%, effectively breaking through the upper limit of the denitrification rate of the Anammox process (89%).

[0019] (3) This invention utilizes the high photoelectric conversion capability of CdS to promote the activity of DNRA and Anammox functional enzymes in AnAOB, thereby enhancing the denitrification efficiency of the Anammox process while avoiding the emission of greenhouse gas nitrous oxide during denitrification.

[0020] (4) The method of the present invention is green and environmentally friendly, makes full use of renewable energy, meets the global development needs for pollution reduction and carbon reduction, and is suitable for improving the denitrification efficiency of Anammox process.

[0021] (5) Constructing an AnAOB-CdS biohybrid system using AnAOB and CdS. This helps AnAOB enhance the activity of DNRA functional enzymes, thereby promoting Anammox activity and improving the system's denitrification performance, which has guiding significance for the engineering application of Anammox. Attached Figure Description

[0022] Figure 1 This is the X-ray diffraction spectrum of the CdS material prepared in Example 1 of this invention; Figure 2 This is a scanning electron microscope (SEM) image of the CdS material obtained in Example 1 of this invention. Figure 3 This is an elemental distribution diagram of the CdS material obtained in Example 1 of the present invention; Figure 4 This is a high-resolution transmission electron microscope (TEM) image of the CdS material obtained in Example 1 of this invention. Figure 5 This is a genus-level composition diagram of the microorganisms that formed the AnAOB-CdS biohybrid system in Embodiment 2 of the present invention; Figure 6This is a scanning electron microscope image showing the results of the AnAOB-CdS biohybrid system obtained in Example 2 of this invention. Figure 7 This is an elemental distribution diagram of the AnAOB-CdS biohybrid system obtained in Example 2 of the present invention; Figure 8 These are the photocurrent test results of the AnAOB-CdS biohybrid system obtained in Example 2 of this invention and Comparative Example 1; Figure 9 This is a graph showing the nitrate removal performance test results of the AnAOB-CdS biohybrid system in Embodiment 3 and Comparative Examples 1 to 4 of the present invention; Figure 10 The AnAOB-CdS biohybrid system in Embodiment 3 of this invention 15 Test diagram of N-labeled nitrate products; Figure 11 This is a graph showing the abundance of nitrogen conversion functional genes of AnAOB in the AnAOB-CdS biohybrid system during nitrate removal in Example 3 and Comparative Example 2 of this invention. Figure 12 This is a comparison chart of the effluent water quality of Example 4 and Comparative Example 5 of the present invention in enhancing the denitrification efficiency of the existing Anammox process; Figure 13 This is a schematic diagram of a highly efficient and sustainable denitrification method using anaerobic ammonia-oxidizing bacteria provided by the present invention. Detailed Implementation

[0023] The technical solutions of the present invention will be clearly and completely described below with reference to the accompanying drawings of the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments.

[0024] The endpoints and any values ​​of the ranges disclosed in this embodiment are not limited to the precise ranges or values, and these ranges or values ​​should be understood to include values ​​close to these ranges or values. For numerical ranges, the endpoint values ​​of the various ranges, the endpoint values ​​of the various ranges and individual point values, and individual point values ​​can be combined with each other to obtain one or more new numerical ranges, which should be considered as specifically disclosed herein.

[0025] like Figure 13 As shown, the first aspect of this embodiment provides a highly efficient and sustainable denitrification method using anaerobic ammonia-oxidizing bacteria, comprising the following steps: Step S1: Inoculate the anaerobic ammonia oxidation (Anammox) reactor with an anaerobic ammonia oxidizing bacteria-cadmium sulfide (AnAOB-CdS) biohybrid system (hereinafter referred to as the AnAOB-CdS biohybrid system). Furthermore, in this embodiment, the anaerobic ammonia oxidation reactor is an anaerobic serum bottle, an anaerobic membrane bioreactor, an upflow anaerobic sludge bed reactor, or an anaerobic expanded granular sludge bed reactor. The inoculum size of the anaerobic ammonia oxidizing bacteria-cadmium sulfide hybrid system is 5-95% of the volume of the anaerobic ammonia oxidation reactor.

[0026] Step S2: Flow wastewater containing nitrates into the anaerobic ammonia oxidation reactor and add a sacrificial agent; Furthermore, the concentration of nitrate in the nitrate-containing wastewater ranges from 0.1 to 100 mmol N / L.

[0027] Furthermore, the sacrificial agent is at least one selected from sodium lactate, formic acid, acetic acid, oxalic acid, triethanolamine, cysteine, ascorbic acid, disodium ethylenediaminetetraacetate, sodium sulfite, and sodium sulfide. The dosage of the sacrificial agent is 0.001~1 wt%.

[0028] Step S3: Apply light to the anaerobic ammonia oxidation reactor to treat the nitrate-containing wastewater.

[0029] Furthermore, in this embodiment, the wavelength of the illumination is 200~1000 nm, and it can be monochromatic light or multi-wavelength mixed light, with an illumination intensity of 0.1~1000 mW / cm². 2 The lighting mode is either continuous or intermittent.

[0030] The second aspect of this embodiment provides a method for preparing an anaerobic ammonia-oxidizing bacteria-cadmium sulfide biohybrid system as follows: Step S11: Add cadmium sulfide (CdS) powder (hereinafter referred to as CdS powder) to anaerobic ammonia oxidizing bacteria (AnAOB) solution (hereinafter referred to as AnAOB solution) and mix thoroughly to obtain the first mixture; Furthermore, in this embodiment, the CdS powder dosage is controlled at 0.1~5 g / L; the concentration of the anaerobic ammonia-oxidizing bacteria solution is controlled at OD0.05. 600 The value is 0.5~2.0.

[0031] Step S12: Incubate the first mixture obtained in step S11 in a constant temperature shaker to obtain the anaerobic ammonia oxidizing bacteria-cadmium sulfide biohybrid system.

[0032] Furthermore, in this embodiment, the incubation is strictly anaerobic, with a dissolved oxygen concentration of less than 0.05 mg / L, an incubation time of 24~96 h, a temperature of 35~37 ℃, and a shaker speed of 100~300 rpm.

[0033] The third aspect of this embodiment provides that the CdS powder used in step S1 is prepared by the following method: Step S111: Add cadmium chloride (CdCl2·2.5H2O) to polyethylene glycol (PEG 400) and stir continuously until fully mixed and dissolved to form the first colloid; Furthermore, in this embodiment, the addition ratio of cadmium chloride to polyethylene glycol is 1 mol: 175 mL, and the stirring time is 15 min.

[0034] Step S112: Add thioacetamide (C2H5NS) powder to the first colloid and stir continuously to form a second colloid; Furthermore, in this embodiment, the molar ratio of cadmium chloride to the thioacetamide is 1:1, and the stirring time is 30 min.

[0035] Step S113: The second colloid is left to stand at 25°C for 12 hours to obtain the third colloid; Step S114: The third colloid is washed and centrifuged 6 times with deionized water, then washed and centrifuged 6 times with anhydrous ethanol to thoroughly remove the residue. After drying at 25°C for 24 hours, it is ground and sieved to obtain the CdS powder.

[0036] Furthermore, in this embodiment, the centrifugation conditions must at least meet the following requirements: a rotation speed of 10,000 rpm, a time of 10 minutes, and a temperature of 10 °C.

[0037] The present invention will be described in detail below through specific data examples. In the following examples, unless otherwise specified, all raw materials are commercially available products.

[0038] Example 1

[0039] This example provides a method for preparing cadmium sulfide (CdS), including the following steps: (1) Add 40 mmol CdCl2·2.5H2O to 7 mL PEG 400 and stir continuously for 15 min until fully mixed and dissolved to form the first colloid; (2) Add 40 mmol of thioacetamide to the first colloid and stir continuously for 30 min to form the second colloid; (3) The second colloid is placed in an environment of 25 °C and left to stand for 12 h to obtain the third colloid; (4) The third colloid was washed and centrifuged with deionized water 6 times, then washed and centrifuged with anhydrous ethanol 6 times to remove the residue. After drying at 25 °C for 24 h, it was ground and sieved to obtain CdS photoelectric conversion material. The centrifugation conditions are: 10,000 rpm, 10 ℃, and 10 min.

[0040] Example 2

[0041] This embodiment provides a method for constructing an AnAOB-CdS biohybrid system, and the method includes the following steps: (1) Add the CdS powder described in Example 1 to 300 mL of AnAOB bacterial solution and mix thoroughly. The amount of CdS powder added is controlled at 1 g / L, and the concentration of AnAOB bacterial solution is OD. 600 = 1.0, thus obtaining the first mixture; (2) The first mixture obtained in step (1) was incubated in a constant temperature shaker. The temperature of the constant temperature shaker was controlled at 37 °C and the rotation speed was controlled at 180 rpm. The dissolved oxygen concentration was controlled to be less than 0.05 mg / L. After incubation for 48 h, the AnAOB-CdS bio-hybrid system was obtained.

[0042] AnAOB bacterial suspensions were cultured for more than one year. The composition and content of the culture medium used included, based on the total amount of the culture medium, ammonium chloride concentration of 2.0 g / L, sodium nitrite concentration of 3.1 g / L, potassium bicarbonate concentration of 1 g / L, potassium dihydrogen phosphate concentration of 0.02 g / L, ferrous sulfate heptahydrate concentration of 0.02 g / L, magnesium sulfate heptahydrate concentration of 0.02 g / L, calcium chloride concentration of 0.05 g / L, disodium ethylenediaminetetraacetate concentration of 0.005-2 g / L, and mineral mixture concentration of 0.02 g / L.

[0043] AnAOB bacterial culture mainly consists of the genus Candidatus Brocadia.

[0044] Example 3

[0045] This example demonstrates the application of the AnAOB-CdS biohybrid system in the field of photo-enhanced nitrate removal, and the method includes the following steps: (1) The AnAOB-CdS biohybrid system described in Example 2 was inoculated into a 50 mL anaerobic serum bottle. The hydraulic retention time was set to 3 days, and the inoculation amount of the AnAOB-CdS biohybrid system was 50% of the volume of the anaerobic serum bottle. (2) Add nitrate wastewater to the anaerobic serum bottle in step (1), the nitrate concentration of the wastewater is 4 mmol N / L, and add sodium sulfite as a sacrificial agent, the content of the sacrificial agent is 0.1 wt%; (3) Apply visible light to the anaerobic serum bottle with a wavelength range of 400-800 nm, using continuous irradiation mode, and set the light intensity to 0.8 mW / cm². 2 The nitrate-containing wastewater was then treated.

[0046] Example 4

[0047] This example follows the method of Example 3, except that actual Anammox process effluent is added in step (2), wherein the nitrate concentration is 5 mmol N / L and the sacrificial agent content is 0.05 wt%, and the remaining steps and parameters are the same as in Example 3.

[0048] Comparative Example 1 The procedure was carried out according to Example 3, except that AnAOB bacterial culture was directly inoculated instead of the AnAOB-CdS biohybrid system, and no light was applied. The remaining steps and parameters were the same as in Example 3.

[0049] Comparative Example 2 The procedure was carried out according to Example 3, except that AnAOB bacterial culture was directly inoculated instead of the AnAOB-CdS biohybrid system, and the remaining steps and parameters were the same as in Example 3.

[0050] Comparative Example 3 The procedure was carried out according to Example 3, except that no light was applied, while the remaining steps and parameters were the same as in Example 3.

[0051] Comparative Example 4 The procedure was carried out according to Example 3, except that the AnAOB-CdS biohybrid system was subjected to high-pressure steam sterilization beforehand at a temperature of 121 °C for 20 min. The remaining steps and parameters were the same as in Example 3.

[0052] Comparative Example 5 The procedure was carried out according to Example 4, except that AnAOB bacterial culture was directly inoculated instead of the AnAOB-CdS biohybrid system, and the remaining steps and parameters were the same as in Example 4.

[0053] Test Example 1 X-ray diffraction spectroscopy was performed on the CdS photoelectric conversion material obtained in Example 1 above, and the results are as follows: Figure 1As shown in the figure. By comparing and analyzing with the standard card (PDF#97-002-9278), it can be seen that the diffraction peak positions of the sample are in high agreement with the standard spectrum. The characteristic diffraction peaks located at approximately 2θ of 26.5°, 44.0°, 52.1°, and 70.4° in the figure correspond to the (111), (220), (311), and (331) crystal planes of cubic CdS, respectively. No obvious impurity peaks were detected in the spectrum, indicating that the prepared CdS photoelectric conversion material has high purity. In addition, the diffraction peaks showed obvious broadening characteristics, which is usually attributed to the small grain size, indicating that the sample has nanocrystalline structure characteristics.

[0054] Test Example 2 The CdS photoelectric conversion material obtained in Example 1 was subjected to morphology scanning, elemental distribution, and structural characterization tests. The results are as follows: Figures 2-4 As shown.

[0055] in, Figure 2 This is a scanning electron microscope (SEM) image showing the CdS photoelectric conversion material obtained in Example 1 of this invention. Figure 3 This is an elemental distribution diagram of the CdS photoelectric conversion material obtained in Example 1 of the present invention. Figure 4 The image shows the high-resolution transmission electron microscope test results of CdS obtained in Example 1 of this invention.

[0056] Depend on Figure 2 As can be seen, the CdS obtained in Example 1 of this invention exhibits an irregular blocky agglomeration morphology with a relatively rough surface. These are micron-sized secondary particles formed by the self-assembly or stacking of primary nanocrystals. Figure 3 It can be seen that Cd (green) and S (red) elements are distributed very uniformly throughout the entire particle area, confirming the uniformity of the chemical composition of the CdS obtained in Example 1 of this invention, and that there are no obvious impurity elements. Figure 4 It can be seen that the CdS obtained in Example 1 of the present invention has clearly visible lattice fringes, indicating that the sample has good crystallinity. By measuring the lattice fringes spacing, the interplanar spacings of 0.338 nm, 0.208 nm, 0.170 nm and 0.134 nm were obtained, which correspond to the (111), (220), (311) and (331) crystal planes of cubic CdS, respectively.

[0057] Test Example 3 The results of the genus-level test on the AnAOB bacterial solution used in Example 2 above are as follows: Figure 5 As shown. By Figure 5 It can be seen that Candidatus Brocadia is the dominant genus of bacteria in the AnAOB bacterial solution used in Example 2, with a relative abundance of about 50%.

[0058] Test Example 4 The AnAOB-CdS biohybrid system obtained in Example 2 was subjected to morphological scanning and elemental distribution testing. The results are as follows: Figure 6 and Figure 7 As shown. By Figure 6 As can be seen, the AnAOB-CdS biohybrid system obtained in Example 2 of this invention exhibits a typical bio-inorganic hybrid structure. It was observed that a large number of particulate clusters were tightly adhered to the AnAOB surface of the AnAOB-CdS biohybrid system obtained in Example 2 of this invention. These clusters are formed by the accumulation of nanoscale particles and are relatively concentrated, indicating that the inorganic nanomaterials have achieved assembly on the cell surface, forming a stable interfacial contact. Figure 7 It can be seen that the characteristic signals of cadmium (Cd, red) and sulfur (S, green) elements in the AnAOB-CdS biohybrid system obtained in Example 2 of the present invention highly overlap in spatial distribution, and their distribution areas are similar to those of the components in the biohybrid system. Figure 6 The consistent positions of the particle clusters prove that the substance attached to the surface of AnAOB is indeed CdS.

[0059] Test Example 5 The photocurrent of the CdS photoelectric conversion material obtained in Example 1, the AnAOB bacterial solution used in Example 2, and the AnAOB-CdS biohybrid system obtained in Example 2 were tested, and the results are as follows: Figure 8 As shown.

[0060] Specifically, a standard three-electrode testing system was used, with 0.1M phosphate buffer as the electrolyte, an Ag / AgCl electrode as the reference electrode, and a platinum wire electrode as the counter electrode. The CdS photoelectric conversion material obtained in Example 1 of this invention, the AnAOB bacterial solution used in Example 2 above, and the AnAOB-CdS biohybrid system obtained in Example 2 above were used as the working electrodes. A xenon lamp was used as the light source, and photocurrent testing was performed under a light-dark cycle of 20 s:20 s.

[0061] Depend on Figure 8 As can be seen, the CdS photoelectric conversion material obtained in Example 1 of this invention exhibits a rapid and strong current response, with a steady-state photocurrent density of approximately 6.2 mA / m. 2 This confirms its excellent photoelectric conversion capability. The AnAOB bacterial solution used in Example 2 of this invention did not show significant current changes under light irradiation, indicating that the original microorganism itself does not possess photoelectric response characteristics. For the AnAOB-CdS biohybrid system obtained in Example 2 of this invention, a significant photocurrent signal was also observed at the moment of light activation, and it exhibited good repeatability. Its photocurrent density was approximately 5.0 mA / m². 2This indicates that the AnAOB-CdS biohybrid system obtained in Example 2 of this invention can effectively capture light energy and generate electrons.

[0062] Test Example 6 During reactor operation, the denitrification efficiency of Example 3 and Comparative Examples 1-4 was tested, and the results are as follows: Figure 9 As shown.

[0063] Depend on Figure 9 As can be seen, compared with Comparative Examples 1 to 4, the AnAOB-CdS biohybrid system in Example 3 of the present invention exhibited excellent nitrate removal performance under light conditions, achieving complete removal within 72 hours. This indicates that the external electrons generated by CdS can be used as electron donors by AnAOB, thereby enhancing the nitrate removal performance of AnAOB.

[0064] Test Example 7 During reactor operation, the above-described Example 3 was tested. 15 N isotope products, results as follows Figure 10 As shown.

[0065] Depend on Figure 10 It can be seen that in Embodiment 3 of the present invention 15 The N isotope product is only nitrogen gas, indicating that the denitrification pathway of the AnAOB-CdS biohybrid system is the Anammox pathway rather than the denitrification pathway, thus avoiding the emission of the greenhouse gas nitrous oxide.

[0066] Test Example 8 During reactor operation, the abundance maps of nitrogen transformation functional genes in Example 3 and Comparative Example 1 were analyzed, and the results are as follows: Figure 11 As shown.

[0067] Depend on Figure 11 As can be seen, compared with pure AnAOB bacterial culture, in the AnAOB-CdS biohybrid system of Example 3 of this invention, genes related to the core Anammox reaction pathway showed a significant upregulation trend. Specifically, the expression abundance of the hzsABC gene encoding hydrazine synthase and the hdh gene encoding hydrazine dehydrogenase increased significantly, by 1.79 and 2.59 times, respectively. This indicates that external electrons enhance the core Anammox denitrification by entering the cellular metabolic network. Simultaneously, the AnAOB-CdS biohybrid system of Example 3 of this invention activated the expression of genes related to periplasmic nitrate reduction and nitrite reduction to ammonium (napA and nrfA). This suggests that AnAOB utilizes external electrons to enhance the conversion pathway from nitrate to nitrite and ammonium, thereby providing sufficient reaction substrates for the Anammox reaction and achieving efficient nitrate removal and biological denitrification.

[0068] Test Example 9 During reactor operation, the denitrification efficiency of Examples 4 and 5 was tested, and the results are as follows: Figure 12 As shown.

[0069] Depend on Figure 12 It can be seen that the AnAOB-CdS bio-hybrid system of Example 3 of the present invention can reduce the nitrate in the actual Anammox process effluent from 74.5 mg / L to 12.7 mg / L and increase the nitrogen removal rate from 89.2% to 98.2%, significantly improving the total nitrogen removal efficiency of the Anammox system.

[0070] The preferred embodiments of the present invention have been described in detail above; however, the present invention is not limited thereto. Within the scope of the inventive concept, various simple modifications can be made to the technical solutions of the present invention, including combinations of various technical features in any other suitable manner. These simple modifications and combinations should also be considered as the content disclosed in the present invention and are all within the protection scope of the present invention.

Claims

1. A highly efficient and sustainable nitrogen removal method using anaerobic ammonia-oxidizing bacteria, characterized in that, Includes the following steps: Step S1: Inoculate the anaerobic ammonia oxidation reactor with an anaerobic ammonia oxidizing bacteria-cadmium sulfide biohybrid system; Step S2: Pass nitrate-containing wastewater into the anaerobic ammonia oxidation reactor and add a sacrificial agent; Step S3: Apply light to the anaerobic ammonia oxidation reactor to treat the nitrate-containing wastewater.

2. The efficient and sustainable denitrification method using anaerobic ammonia-oxidizing bacteria according to claim 1, characterized in that, In step S1, the preparation method of the bio-hybrid system is as follows: Step S11: Add cadmium sulfide powder to the anaerobic ammonia-oxidizing bacteria solution and mix thoroughly to obtain the first mixture; Step S12: Incubate the first mixture obtained in step S11 in a constant temperature shaker to obtain an anaerobic ammonia oxidizing bacteria-cadmium sulfide biohybrid system.

3. The efficient and sustainable denitrification method using anaerobic ammonia-oxidizing bacteria according to claim 2, characterized in that, The cadmium sulfide powder is prepared by the following method: Step S111: Add cadmium chloride to polyethylene glycol and stir to dissolve, forming the first colloid; Step S112: Add thioacetamide powder to the first colloid and stir continuously to form a second colloid; Step S113: Let the second colloid stand at 25°C for 12 hours to obtain the third colloid; The third colloid was washed and centrifuged 6 times with deionized water, then washed and centrifuged 6 times with anhydrous ethanol to thoroughly remove the residue. After drying at 25°C for 24 hours, it was ground and sieved to obtain cadmium sulfide powder.

4. The efficient and sustainable denitrification method using anaerobic ammonia-oxidizing bacteria according to claim 3, characterized in that, In step S111, the addition ratio of cadmium chloride to polyethylene glycol is 1 mol: 175 mL, and the stirring time is 15 min; in step S112, the molar ratio of cadmium chloride to thioacetamide is 1:1, and the stirring time is 30 min; in step S113, the centrifugation conditions must at least meet the following requirements: rotation speed of 10000 rpm, time of 10 minutes, and temperature of 10 ℃.

5. The efficient and sustainable denitrification method using anaerobic ammonia-oxidizing bacteria according to claim 2, characterized in that, The dosage of cadmium sulfide powder is controlled at 0.1~5 g / L.

6. The efficient and sustainable denitrification method using anaerobic ammonia-oxidizing bacteria according to claim 2, characterized in that, The anaerobic ammonia-oxidizing bacterial solution includes at least one of the genera of anaerobic ammonia oxidizing bacteria, such as Brocardia, Jeterus, Cuscuta, Stepanes, and Anaerobic ammonia-oxidizing cocci.

7. The efficient and sustainable denitrification method using anaerobic ammonia-oxidizing bacteria according to claim 2, characterized in that, In step S11, the concentration of the anaerobic ammonia-oxidizing bacteria solution is controlled at OD. 600 The dissolved oxygen concentration is 0.5~2.0; in step S12, the incubation is strictly anaerobic, with a dissolved oxygen concentration of less than 0.05 mg / L, an incubation time of 24~96 h, a temperature of 35~37 ℃, and a shaking speed of 100~300 rpm.

8. The efficient and sustainable denitrification method using anaerobic ammonia-oxidizing bacteria according to claim 1, characterized in that, In step S2, the concentration of the nitrate wastewater is in the range of 0.1~100 mmol N / L; the sacrificial agent is at least one of sodium lactate, formic acid, acetic acid, oxalic acid, triethanolamine, cysteine, ascorbic acid, disodium ethylenediaminetetraacetate, sodium sulfite and sodium sulfide, and its dosage is 0.001~1 wt%.

9. The efficient and sustainable denitrification method using anaerobic ammonia-oxidizing bacteria according to claim 1, characterized in that, In step S3, the illumination wavelength can be 200~1000 nm, and can be monochromatic light or multi-wavelength mixed light; the illumination intensity can be 0.1~1000 mW / cm². 2 The lighting mode can be either continuous or intermittent.

10. The efficient and sustainable denitrification method using anaerobic ammonia-oxidizing bacteria according to claim 1, characterized in that, In step S1, the anaerobic ammonia oxidation reactor is an anaerobic serum bottle, an anaerobic membrane bioreactor, an upflow anaerobic sludge bed reactor, or an anaerobic expanded granular sludge bed reactor; the inoculum amount of the anaerobic ammonia oxidizing bacteria-cadmium sulfide biohybrid system is 5-95% of the volume of the anaerobic ammonia oxidation reactor.

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