Micro-nano aeration technology-based efficient denitrification method of nitrification aerobic granular sludge and sulfur autotrophic denitrification coupling system

Abstract

The invention discloses an efficient denitrification method of a nitrification aerobic granular sludge and sulfur autotrophic denitrification coupling system based on a micro-nano aeration technology, and belongs to the technical field of water treatment. According to the method, a micro-nano aeration technology is adopted for culture of the nitrified aerobic granular sludge, culture and domestication of the aerobic granular sludge are facilitated, removal of pollutants can be promoted, the richness index and diversity index of the prepared nitrified aerobic granular sludge are both in a rising trend, and therefore the stability of a nitrification system is enhanced, and the nitrification effect of the nitrified aerobic granular sludge is improved. And the relative abundance of each nitration gene of the nitrated aerobic granular sludge is in a rising trend. The nitrification aerobic granular sludge system and the sulfur autotrophic denitrification system are coupled, the total nitrogen removal rate of 95% can be achieved when the HRT is equal to 1.5 h, the total nitrogen of effluent is 1.0 mg / L or below, a carbon source does not need to be added, and the denitrification performance is excellent.

Description

Technical Field

[0001] This invention relates to a highly efficient nitrogen removal method based on a coupled system of nitrifying aerobic granular sludge and sulfur autotrophic denitrification using micro-nano aeration technology, belonging to the field of water treatment technology. Background Technology

[0002] Wastewater treatment plants face immense operational pressures, as traditional wastewater treatment processes are no longer sufficient to meet the demands for high efficiency and sustainability. For issues such as poor removal of total nitrogen and total phosphorus, and low influent carbon-to-nitrogen ratios, wastewater treatment plants primarily employ methods such as adding carbon sources, adjusting process parameters, and upgrading to new technologies. These methods are technically demanding and costly, and can easily generate potential secondary hazards. Furthermore, meeting increasingly stringent effluent discharge standards has become a major challenge for wastewater treatment plants to achieve stable compliance. Therefore, continuous improvement and innovation of existing nitrogen removal technologies are necessary to explore more economical, efficient, and environmentally friendly nitrogen removal processes.

[0003] While traditional wastewater treatment technologies are constantly being improved and innovated, various emerging wastewater treatment technologies are also constantly emerging. Aerobic granular sludge technology and sulfur autotrophic denitrification technology are being gradually explored and promoted due to their significant advantages. Nitrifying aerobic granular sludge possesses characteristics such as a dense structure, good settling properties, and high toxicity resistance. Its excellent nitrification performance demonstrates good treatment effects on inorganic high-ammonia nitrogen wastewater, laying the foundation for improving the denitrification performance of reactors. Sulfur autotrophic denitrification technology, on the other hand, has advantages such as simple process facilities, zero carbon source addition, and low cost. It has been researched and applied in the deep denitrification and denitrification processes of groundwater with low carbon-to-nitrogen ratios and wastewater treatment plant effluent. Both technologies, due to their unique advantages, show broad application prospects in the field of wastewater treatment.

[0004] Currently, CN 118307135 A discloses a short-process autotrophic wastewater treatment process that achieves ultra-low carbon and nitrogen pollutant emissions by forming a combination of biological adsorption, nitrifying aerobic granular sludge, and sulfur autotrophic denitrification without the need for external carbon source addition. Although this short-process nitrification-denitrification process can achieve nitrogen removal through regulation, it is actually greatly affected by water quality and still requires a high carbon source. Furthermore, when using this process to treat wastewater with low carbon source, the nitrogen removal efficiency will decrease significantly.

[0005] Patents CN 119822501 A and CN 116589079 A both disclose technologies related to micro-nano aeration for cultivating aerobic granular sludge, but the technical solutions described are all for cultivating aerobic granular sludge, not for nitrifying aerobic granular sludge, and do not provide technical inspiration for treating low C / N wastewater. Summary of the Invention

[0006] Technical issues

[0007] Traditional nitrogen removal processes involve nitrification liquor recirculation, leading to competition among microorganisms. Low C / N ratios inhibit the activity of nitrifying and denitrifying bacteria, necessitating the addition of external carbon sources to achieve nitrogen removal. While current short-cut nitrification-denitrification processes can achieve nitrogen removal through regulation, their effectiveness is significantly influenced by water quality and still requires relatively high carbon sources. Therefore, a method is needed that treats low C / N wastewater without requiring external carbon sources.

[0008] Technical solution

[0009] To address the aforementioned technical challenges, this invention leverages the advantages of nitrifying aerobic granular sludge and sulfur autotrophic denitrification processes to construct a combined nitrifying aerobic granular sludge / sulfur autotrophic denitrification process. This combined process not only reduces costs and saves land area but also eliminates the need for carbon source addition, effectively reducing sludge production and the risk of secondary pollution. Furthermore, it demonstrates the feasibility and long-term stability of treating wastewater with low C / N ratios. In this invention, the nitrifying aerobic granular sludge is cultivated using micro-nano aeration technology. Compared to conventional aeration methods, micro-nano aeration exhibits higher oxygen mass transfer efficiency and continuously generates hydroxyl radicals, which not only promote the growth of aerobic granular sludge and enhance pollutant removal but also save costs and improve energy efficiency.

[0010] This invention provides a method for cultivating nitrifying aerobic granular sludge based on micro-nano aeration technology, the method comprising the following steps:

[0011] (1) Inoculate sludge into a sequencing batch reactor and start the reactor for cultivation. In the initial stage of operation, the influent time is 3-7 min, aeration time is 200-205 min, settling time is 10-20 min, drainage time is 3-7 min, and idle time is 5-15 minutes. After the reactor is running stably, the aeration and settling time are adjusted to 210-215 min and 5-7 min, respectively, and the drainage ratio is 55-65%. After cultivation for 25-35 days, the aeration and settling time are adjusted to 150-160 min and 3-7 min, respectively, to obtain aerobic granular sludge.

[0012] (2) After the aerobic granular sludge is formed, the influent conditions are gradually changed by simultaneously reducing the C / N ratio, and the sludge is acclimatized and cultivated into nitrified aerobic granular sludge.

[0013] Furthermore, in step (1), a cylindrical sequencing batch reactor can be selected as the sequencing batch reactor; if a cylindrical sequencing batch reactor is selected, the effective volume is 4 to 8 L.

[0014] Furthermore, in step (1), an aeration device is installed at the bottom of the sequencing batch reactor; the aeration device has a fine aeration head with a pore size of 0.2 to 0.3 μm.

[0015] Furthermore, the aeration device is capable of generating air bubbles with a particle size between 1 and 100 μm.

[0016] Furthermore, in step (1), the aeration rate of the control device during the cultivation process is 1 to 5 L / min.

[0017] Furthermore, the sludge used for inoculation in step (1) is taken from a wastewater treatment plant (A). 2 The aerobic tank of the / O process.

[0018] Furthermore, in step (1), the MLSS of the inoculated sludge is 4-5 g / L.

[0019] Furthermore, in step (1), stable operation means that the SVI in the reactor is maintained at 25-30 mL / g.

[0020] Furthermore, the components of the influent in step (1) include: a mixed carbon source of CH3COONa and CH3CH2COONa, a COD of 100-1000 mg / L, and NH4+. + -N concentrations are 30-50 mg / L, 20-30 mg / L KH2PO4, 30-40 mg / L K2HPO4, 90-100 mg / L MgSO4, 70-80 mg / L CaCl2, 5-15 mg / L EDTA, 1-2 mg / L FeCl3·6H2O, 0.1-0.2 mg / L H3BO3, 0.03-0.05 mg / L CuSO4·5H2O, 0.03-0.05 mg / L KI, 0.1-0.2 mg / L MnCl2·4H2O, 0.05-0.07 mg / L ZnCl2, 0.1-0.2 mg / L CoCl2·6H2O, and 0.05-0.1 mg / L Na2MoO4·2H2O.

[0021] Furthermore, the acclimatization and cultivation process described in step (2) includes: gradually changing the influent conditions by simultaneously reducing the C / N ratio; the acclimatization and cultivation conditions are divided into two stages:

[0022] Phase I: Influent COD concentration decreases from 1000 mg / L to 100 mg / L, NH4+... + The -N concentration was decreased from 50 mg / L to 30 mg / L, and the treatment period was 35–45 days.

[0023] Stage Ⅱ: Influent COD concentration is 0, NH4+ + The -N concentration was 30 mg / L, and the treatment period was 30–35 days.

[0024] The present invention provides nitrified aerobic granular sludge prepared according to the above method.

[0025] The application of the nitrified aerobic granular sludge provided by this invention in wastewater treatment.

[0026] This invention also provides a nitrogen removal method based on a coupled system of nitrifying aerobic granular sludge and sulfur autotrophic denitrification, the method comprising:

[0027] Wastewater first enters a nitrifying aerobic granular sludge tank for treatment, then enters a storage tank to adjust the pH, and finally enters a sulfur autotrophic denitrification filter for further treatment, with the final effluent being denitrified.

[0028] In one embodiment of the present invention, the COD of the wastewater is 0-500 mg / L and the TN is 10-50 mg / L.

[0029] In one embodiment of the present invention, nitrified aerobic granular sludge obtained according to the above method is added to the nitrified aerobic granular sludge tank, and the amount added is 25-35% of the volume of the nitrified aerobic granular sludge tank.

[0030] In one embodiment of the present invention, the pH is adjusted using an alkaline agent; the alkaline agent includes sodium carbonate and sodium bicarbonate; the pH is adjusted to 8.5-9.

[0031] In one embodiment of the present invention, a gravel support layer is arranged at the bottom of the sulfur autotrophic denitrification filter, which is inoculated with sulfur autotrophic denitrification sludge and filled with elemental sulfur.

[0032] In one embodiment of the present invention, the sludge concentration of the sulfur autotrophic denitrification sludge is 1500-2500 mg / L.

[0033] In one embodiment of the present invention, the elemental sulfur has a particle size of 2-4 mm and a porosity of 40%-60%.

[0034] In one embodiment of the present invention, the amount of elemental sulfur is 70-90% of the volume of the sulfur autotrophic denitrification filter.

[0035] In one embodiment of the present invention, wastewater is treated in a nitrifying aerobic granular sludge tank for one cycle; the cycle includes 5-8 minutes of influent, 210-215 minutes of aeration, 5-10 minutes of settling, 5-10 minutes of drainage, and 5-15 minutes of idle time.

[0036] In one embodiment of the present invention, the hydraulic retention time of wastewater in the sulfur autotrophic denitrification filter is 1.5 to 10 hours.

[0037] Beneficial effects

[0038] (1) Compared with the prior art, the present invention adopts micro-nano aeration technology in the cultivation of nitrifying aerobic granular sludge, which is not only beneficial to the cultivation and acclimatization of aerobic granular sludge, but also promotes the removal of pollutants. The obtained nitrifying aerobic granular sludge changes from the light yellow of the initial inoculated sludge to brownish-yellow. Electron microscopy shows that the successfully cultivated nitrifying aerobic granular sludge has clearer, rounder, and more regular edges than the inoculated AGS, and the particle structure is more compact. The average particle size of the sludge increases from 189.22 μm of the inoculated aerobic granular sludge to 348.6 μm, and continues to grow steadily to 800 μm, exhibiting excellent settling performance. In addition, the richness index and diversity index of the obtained nitrifying aerobic granular sludge both show an increasing trend, thereby enhancing the stability of the nitrification system. Furthermore, the relative abundance of each nitrification gene in the nitrifying aerobic granular sludge also shows an increasing trend. Therefore, the nitrification function in the wastewater treatment process will be enhanced, achieving excellent removal efficiency of pollutants.

[0039] (2) Compared with the prior art, the wastewater treatment technology constructed in this invention has the characteristics of high nitrogen removal efficiency and no need for external carbon source. After coupling the nitrifying aerobic granular sludge system and the sulfur autotrophic denitrification system, the nitrifying aerobic granular sludge system has a high nitrogen removal efficiency and no need for external carbon source. + -N can be completely removed, and complete nitrification can be achieved even when dissolved oxygen is greater than 2 mg / L. The effluent from the nitrification stage is then fed into a sulfur autotrophic system for denitrification. At an HRT of 1.5 h, the system can achieve a total nitrogen removal rate of 95%, with total nitrogen in the effluent below 1.0 mg / L. Attached Figure Description

[0040] Figure 1 Schematic diagram of a nitrification aerobic granular sludge unit.

[0041] Figure 2 Schematic diagram of a combined process unit for nitrifying aerobic granular sludge-sulfur autotrophic denitrification.

[0042] Figure 3 : Denitrification performance of combined processes (a: nitrifying aerobic granular sludge; b: sulfur autotrophic denitrification).

[0043] Figure 4 Changes in EPS components and content at different operating stages of the combined process (a: PN; b: PS; c: EPS).

[0044] Figure 5 : Characteristics of amino acid composition in EPS hydrolysate at different operating stages (a: amino acid content in nitrifying aerobic granular sludge; b: relative abundance of amino acids in nitrifying aerobic granular sludge).

[0045] Figure 6 : Characteristics of the amino acid composition of EPS hydrolysate at different operating stages (c: content of sulfur autotrophic denitrification amino acids; d: relative abundance of sulfur autotrophic denitrification amino acids).

[0046] Figure 7 FTIR spectra (a: nitrifying aerobic granular sludge; b: sulfur autotrophic denitrification).

[0047] Figure 8 Amide I fitting spectrum of protein secondary structure (nitrifying aerobic granular sludge a: 0d; b: 40d; c: 70d; sulfur autotrophic denitrification d: 0d; e: 40d; f: 70d).

[0048] Figure 9 Correlation analysis between EPS and microbial community (a: NAGS; b: SADN).

[0049] Figure 10 COG gene functional annotation. Detailed Implementation

[0050] Example 1: Cultivation of nitrifying aerobic granular sludge

[0051] The device employs a cylindrical sequencing batch reactor with an effective volume of 4L. The aeration unit includes 0.22μm fine aeration heads, ensuring that the generated bubble particle size is approximately 100μm, with an aeration rate of 3L / min. The reaction apparatus is shown in the attached diagram. Figure 1 As shown. The inoculated sludge was taken from a wastewater treatment plant (A). 2 / The aerobic tank (O process) has an inoculation volume of 2L and an inoculated sludge MLSS of 4.5g / L. It operates for 6 cycles per day, initially with each cycle lasting 4 hours. Each cycle consists of 5 minutes of influent, 205 minutes of aeration, 15 minutes of settling, 5 minutes of effluent discharge, and a 10-minute idle period. After the reactor stabilizes (for 2 weeks), and the system SVI is maintained at 25-30 mL / g, the aeration time can be extended from 205 minutes to 215 minutes, and the settling time shortened from 15 minutes to 5 minutes. The effluent ratio is 60%. Artificial wastewater is used as the influent, with a mixed carbon source (CH3COONa and CH3CH2COONa). COD is gradually reduced from 1000 mg / L to 100 mg / L, and NH4+ is... + -N was reduced from 50 mg / L to 30 mg / L. The remaining components included 29.25 mg / L KH2PO4, 37.41 mg / L K2HPO4, 97 mg / L MgSO4, 75 mg / L CaCl2, 10 mg / L EDTA, and trace elements (1.5 mg / L FeCl3·6H2O, 0.15 mg / L H3BO3, 0.03 mg / L CuSO4·5H2O, 0.03 mg / L KI, 0.12 mg / L MnCl2·4H2O, 0.058 mg / L ZnCl2, 0.15 mg / L CoCl2·6H2O, and 0.06 mg / L Na2MoO4·2H2O).

[0052] After the aerobic granular sludge is formed, the influent conditions are gradually changed by simultaneously reducing the C / N ratio. Mature aerobic granular sludge is used as inoculum sludge to acclimate and cultivate into nitrifying aerobic granular sludge. The cultivation conditions are shown in Table 1.

[0053] Table 1 Inlet Water Condition Settings

[0054]

[0055] During the acclimation process of nitrifying aerobic granular sludge, the presence of COD significantly inhibits the activity of nitrifying bacteria. Because the granular structure, initially dominated by heterotrophic bacteria, is relatively stable and the growth cycle of nitrifying bacteria is long, a certain amount of time is needed to transform it into a carrier for nitrifying bacteria growth, thus forming nitrifying aerobic granular sludge. Using aerobic granular sludge as inoculum, nitrifying aerobic granular sludge can be successfully cultivated by gradually altering external environmental conditions. This sludge exhibits a compact structure and good settling properties. Under autotrophic conditions, aerobic granular sludge may decompose to produce more small-diameter sludge particles, increasing the sludge surface area and thus enhancing nitrification.

[0056] Example 2: Nitrogen removal efficiency test and analysis of a coupled system of nitrifying aerobic granular sludge and sulfur autotrophic denitrification

[0057] The nitrogen removal efficiency of the combined process was investigated by coupling a nitrifying aerobic granular sludge system and a sulfur autotrophic denitrification system, using the methods described in the attached figure. Figure 2 The reaction apparatus shown is used for wastewater treatment. The reaction apparatus consists of a water distribution tank, a nitrifying aerobic granular sludge tank, a water storage tank, a sulfur autotrophic denitrification filter, and an aeration pump. The nitrifying aerobic granular sludge tank is cylindrical with an inner diameter of 10 cm. It contains 2 L of nitrifying aerobic granular sludge cultivated in Example 1, accounting for 30% of the tank's volume. A micro-nano aeration device is installed at the bottom to ensure that the generated bubbles have a particle size of approximately 100 μm and an aeration rate of 3 L / min. The sulfur autotrophic denitrification filter is cylindrical with an inner diameter of 10 cm. A gravel support layer is arranged at the bottom, and sulfur autotrophic denitrification sludge is inoculated with a sludge concentration of 2000 mg / L. Elemental sulfur is also added, with a particle size of 2–4 mm and a porosity of approximately 50%. The amount of elemental sulfur filling the filter is 80% of its volume.

[0058] During the treatment process, a peristaltic pump is started to pump the influent from the distribution tank into the nitrification aerobic granular sludge tank. After one reaction cycle (total time is 4 hours, including 5 minutes of influent, 215 minutes of aeration, 5 minutes of settling, 5 minutes of effluent discharge, and 10 minutes of idle time), the effluent from the nitrification aerobic granular sludge tank flows into the storage tank. Then, NaHCO3 is added to supplement the alkalinity to pH 8.5 before flowing into the sulfur autotrophic denitrification filter via a peristaltic pump. The operating cycle in the nitrification aerobic granular sludge reactor is 4 hours, and the hydraulic retention time of sulfur autotrophic denitrification is gradually shortened from 10 hours.

[0059] The influent is artificially simulated wastewater, specifically containing 30 mg / L NH4. + -N, 29.25 mg / L KH2PO4, 37.41 mg / L K2HPO4, 97 mg / L MgSO4, 75 mg / L CaCl2, 10 mg / L EDTA, 330 mg / L NaHCO3, and trace elements (1.5 mg / L FeCl3·6H2O, 0.15 mg / L H3BO3, 0.03 mg / L CuSO4·5H2O, 0.03 mg / L KI, 0.12 mg / L MnCl2·4H2O, 0.058 mg / L ZnCl2, 0.15 mg / L CoCl2·6H2O, 0.06 mg / L Na2MoO4·2H2O).

[0060] The nitrogen removal efficiency of nitrifying aerobic granular sludge is shown in the attached figure. Figure 3 As shown in (a), the nitrifying aerobic granular sludge system was found to have a high NH4+ content. + The removal rate of -N is as high as 95%, and complete nitrification can be achieved even when dissolved oxygen is greater than 2 mg / L, thus removing NH4+. + -N is converted to NO3 - -N. At this point, the microbial community structure within the system had stabilized, and the nitrifying bacteria were functioning well. However, after 40 days of operation, a large number of protozoa and metazoa appeared in the nitrifying aerobic granular sludge system, residing on the nitrifying aerobic granular sludge and causing the sludge particles to break down and disintegrate, resulting in severe sludge loss. Remedial measures such as reducing dissolved oxygen and washing the sludge were taken, and the system was still able to maintain a good nitrification effect, with the biomass remaining at 1.0 mg / L.

[0061] The effluent from the nitrification stage continues to be fed into a sulfur autotrophic system for denitrification. The hydraulic retention time (HRT) is gradually shortened from 10 h to 1.5 h, achieving a total nitrogen removal rate of 95% and effluent total nitrogen levels below 1.0 mg / L. Nitrogen in the effluent is primarily composed of NH4+. + -N is the dominant group, NH4 + When the -N concentration is below 0.5 mg / L, the effluent from the nitrification stage will have a residual 0.2 mg / L of NH4. +-N is introduced into the sulfur autotrophic system, but the denitrification system will react with NO3. - -N is reduced to generate some NH4. + -N. The sulfur autotrophic process starts with 30 mg / L NO3. - The -N operation demonstrates a rapid adaptability to the effluent from nitrifying aerobic granular sludge. This indicates that the coupling of nitrification by nitrifying aerobic granular sludge and denitrification by sulfur autotrophic sludge can achieve good nitrogen removal efficiency without the need for additional carbon sources.

[0062] Comparative Example 1

[0063] Referring to Examples 1-4 of CN 118307135 A, wastewater treatment was performed on the artificially simulated wastewater (COD = 0 mg / L) described in Example 2 of this invention. The results showed that the nitrogen removal efficiency of the combined process of biological adsorption + nitrifying aerobic granular sludge + sulfur autotrophic denitrification in CN 118307135 A was approximately 73.33%, significantly lower than the nitrogen removal efficiency (95%) of the combined process of micro-nano aerated nitrifying aerobic granular sludge + sulfur autotrophic denitrification in this patent.

[0064] Table 2 Comparison of the performance of different processes in cultivating nitrifying aerobic granular sludge under COD reduction conditions

[0065]

[0066] In the cultivation process of nitrifying aerobic granular sludge under ordinary aeration, the aeration device is a common aeration head, and the generated bubble particle size is about 2mm. Under the premise of controlling the COD of the influent connected to the system and without other external conditions, the comparison indicators with those of cultivating nitrifying aerobic granular sludge under ordinary aeration are shown in Table 2. The successful cultivation time of nitrifying aerobic granular sludge in Example 1 of this invention ranged from 50 to 70 days, while the successful cultivation time of nitrifying aerobic granular sludge under ordinary aeration ranged from 80 to 145 days. It is evident that the formation of nitrifying aerobic granular sludge cultivated under micro-nano aeration takes significantly less time than the granulation process of nitrifying aerobic granular sludge under ordinary aeration, greatly reducing the cultivation time and accelerating the formation of nitrifying aerobic granular sludge. Therefore, the aeration time required for cultivating nitrifying aerobic granular sludge using micro-nano aeration is shorter. Furthermore, the oxygen volumetric mass transfer coefficient of micro-nano aeration is 4.11 times that of conventional aeration, and its oxygenation power efficiency is 3.30 times higher. This means that micro-nano aeration can transfer oxygen to the water more efficiently, thus significantly reducing energy consumption during the cultivation process. For example, using electromagnetic air pumps, micro-nano aeration for cultivating nitrifying aerobic granular sludge can achieve approximately 30% energy savings in aeration.

[0067] In addition, compared with nitrifying aerobic granules grown by ordinary aeration, the nitrifying aerobic granules grown by micro-nano aeration under COD reduction conditions have an average particle size of about 800μm, an SVI of 21.56mL / g, a smooth surface, a dense structure, and excellent settling performance.

[0068] Example 3: Changes in the extracellular polymeric properties of sludge in a coupled system

[0069] By classifying TB-EPS into Ionic TB-EPS and Hydrophobic TB-EPS, the effects of different forces on the sludge aggregation process were investigated. The changes in proteoglycan content in nitrifying aerobic granular sludge and sulfur autotrophic denitrifying sludge are shown in the attached figure. Figure 4 As shown in the figure, the hydrophobic TB-EPS content of nitrifying aerobic granular sludge was significantly higher than that of sulfur autotrophic denitrifying sludge, indicating that the sludge aggregation performance in the sulfur autotrophic denitrifying sludge system was weaker than that of nitrifying aerobic granular sludge. The ionic TB-PN content of nitrifying aerobic granular sludge surged at day 40, increasing from 24.83 mg / g VSS to 73.35 mg / g VSS. This increase may be due to the loose granular structure exposing more cation binding sites, making it easier for the sludge to bind with metal ions. The SMP-PN content of sulfur autotrophic sludge showed a trend of first decreasing and then increasing, indicating that changes in the external water quality environment inevitably cause fluctuations in the extracellular polymers secreted by microorganisms. The LB-PN content of sulfur autotrophic sludge increased significantly, reaching a maximum of 37.71 mg / g VSS from the initial stage of 5.58 mg / g VSS. LB-PN possesses large polymer chains and a steric hindrance structure, indicating that under the stimulation of changes in the external environment, LB-PN can improve the adhesion of microbial sulfur packing through greater hydrophobic interactions. In addition, the hydrophobic TB-PN of the sulfur autotrophic sludge increased from 57.07 mg / g VSS to 73.82 mg / g VSS on day 70, and the enhanced hydrophobicity is beneficial to the adhesion of sludge to the packing material.

[0070] Changes in extracellular polymeric amino acids can reflect changes in protein composition. During reactor operation, the amino acid content changes of two types of sludge were continuously monitored (see attached diagram). Figure 5To further analyze the changes in the functional structure of sludge in the system, this study aimed to investigate the effects of alanine (Ala) on ATP production and its role in catabolism. Ala plays a crucial role in the catabolism of ATP and serves as an important indicator of microbial stress response. On day 40 of the nitrifying aerobic granular sludge process, the alanine content significantly increased from 12.58 mg / g VSS initially to 38.55 mg / g VSS. After the sulfur autotrophic process received the effluent from the nitrifying aerobic granular sludge process, the alanine content in the sulfur autotrophic sludge increased more than fourfold compared to the initial value, indicating that changes in the aquatic environment have a certain impact on the sulfur autotrophic sludge, enhancing the catabolism capacity of the microorganisms. Glutamic acid (Glu), as one of the most abundant metabolites in bacterial cells, consistently exhibits a high content and relative proportion. The relative proportion of glutamate in the nitrifying aerobic granular sludge ranges from 9.44% to 20.32%, while that in the sulfur autotrophic denitrifying sludge ranges from 11.68% to 30.03%. Glutamic acid, a key amino acid in nitrogen metabolism, indicates enhanced reduction of nitrogen pollutants in systems primarily undergoing denitrification. The concentration of the hydrophobic amino acid valine (Val) in the nitrifying aerobic granular sludge system increased from 6.10 mg / g VSS to 17.73 mg / g VSS after sludge particle breakage, indicating that the microbial community can secrete more hydrophobic substances to promote sludge aggregation. The valine concentration in the sulfur-autotrophic denitrifying sludge also increased from 6.92 mg / g VSS to 34.80 mg / g VSS on day 70; the increase in hydrophobic substances can enhance intercellular aggregation, thereby accelerating biofilm formation. Proline (Pro) has the function of regulating mitochondrial electron transport complexes, maintaining protein stability, and enhancing the activity of various enzymes. The proline concentration in the nitrifying aerobic granular sludge increased from 12.17 mg / g VSS on day 0 to 61.97 mg / g VSS on day 70; the increased proline content plays a role in maintaining sludge protein stability and enhancing related enzyme activity. The proline concentration in the sulfur-autotrophic sludge increased from 6.23 mg / g VSS on day 0 to 57.48 mg / g VSS on day 70. The increase in this amino acid concentration in the sulfur-autotrophic system may be related to the enhanced electron transfer capacity caused by the shortened hydraulic retention time.

[0071] Furthermore, histidine (His) plays a crucial role in cell communication and signal transduction among Gram-negative bacteria. Its changes and the genes encoding histidine and serine enzymes are closely related. The relative proportion of histidine in nitrifying aerobic granular sludge decreased from 14.50% to 2.33% on day 70, while the relative proportion of histidine in sulfur-autotrophic sludge increased from 4.87% to 12.18% on day 30. This suggests that signal transduction may be weakened in nitrifying aerobic granular sludge, while intercellular communication is enhanced with increasing nitrogen removal load in sulfur-autotrophic denitrification systems, a finding also confirmed by metagenomic annotation analysis.

[0072] Example 4: Infrared spectroscopy and protein secondary structure analysis of sludge in a coupled system

[0073] Fourier transform infrared spectroscopy (FTIR) was used to analyze the functional group structures that play an important role in the cultivation of nitrifying aerobic granular sludge and sulfur autotrophic denitrifying sludge. Freeze-dried sludge samples from day 0 (inoculated), day 40, and finally day 70 were selected for characterization. The FTIR spectra are attached. Figure 6 As shown, the FTIR peak positions and numbers of the two types of sludge are very similar, indicating that the chemical group types in the nitrifying aerobic granular sludge and the sulfur autotrophic denitrifying sludge are similar, with differences only in peak intensity at different cultivation stages. Vibrations in activated sludge are generally associated with proteins and polysaccharides, such as those at 3500-3100 cm⁻¹. -1 The tensile vibrations at 1635 cm⁻¹ are NH₄⁺ and OH⁻. -1 Tensile vibration near C=O, 1384 cm -1 The bending vibration of CH2, and 1153-1072 cm -1 COC stretching vibration of polysaccharides.

[0074] Protein secondary structure influences microbial aggregation, adsorption, and biofilm formation. The stability of a protein's secondary structure is maintained through hydrogen bonds formed between carbonyl and amide groups on its backbone. Further processing of infrared spectroscopy data reveals the amide I band (1700-1600 cm⁻¹), which accurately and sensitively reflects protein conformation and changes. -1 Quantitative analysis of protein secondary structure was performed (Table 3). The analysis was conducted using methods such as Fourier deconvolution and peak fitting. The secondary structure peak fitting results are attached. Figure 7 As shown.

[0075] Table 3. Relative percentage content of protein secondary structure at different stages

[0076]

[0077]

[0078] The results showed that both nitrifying aerobic granular sludge and sulfur autotrophic denitrification sludge possessed six protein secondary structures. Random coils and reverse β-sheets were unfavorable for aggregation, while the rest were favorable. Aggregation-favorable structures dominated in both groups of sludge, accounting for more than 60% of the total, indicating strong aggregation properties of the microbial communities in both nitrifying aerobic granular sludge and sulfur autotrophic denitrification sludge. α-helices are the main form of protein secondary structure; their aggregation enhances the strength and elasticity of the sludge. In nitrifying aerobic granular sludge, the relative proportion of α-helices decreased by 0.58% at day 40, indicating instability in the system, which subsequently stabilized. The relative proportion of α-helices in sulfur autotrophic denitrification sludge was lower than that in nitrifying aerobic granular sludge, suggesting weaker hydrogen bonding forces within the denitrification system. β-sheets are special sheet-like structures with wrinkles, providing numerous hydrophobic sites and thus promoting sludge aggregation. At day 40, the β-sheet structure of the nitrifying aerobic granular sludge had the highest relative proportion among the six structures, reaching 22.42%, indicating that the nitrification system may need to expose more hydrophobic sites at this time to maintain the stability of the nitrifying aerobic granular sludge. In the sulfur autotrophic denitrification system, the β-sheet structure increased compared to day 0. This may be due to the stimulating effect of changes in the external environment on the denitrification system, enhancing hydrophobicity and promoting sludge aggregation and attachment. Polymer chains play an important role in connecting α-helices and β-sheets; the increase in polymer chains in the nitrifying aerobic granular sludge is beneficial for enhancing the binding force between sludge particles.

[0079] Example 5: Microbial Community Analysis in Coupled Systems

[0080] Extracellular polymers (EPS) are primarily secreted by the microbial community. Therefore, Pearson correlation analysis can identify microbial populations significantly correlated with EPS secretion at each layer. Pearson correlation analysis was performed on the PN and PS of each EPS layer with genus-level microorganisms (20 species) to explore the correlation between EPS and the microbial community. The results are shown in the appendix. Figure 8As shown in the figure, in the nitrifying aerobic granular sludge system, the changes in bacterial genera in the nitrifying aerobic granular sludge were strongly correlated with polysaccharides, especially SMP-PS and Ionic TB-PS. SMP-PS showed a significant positive correlation with genera such as Haliangium, Ahniella, norank_f_SC-I-84, Chryseobacterium, and SM1A02, indicating that these genera maintain the stability of the nitrifying aerobic granular sludge system by secreting SMP-PS. In the sulfur autotrophic denitrifying sludge, many microbial genera showed a negative correlation with extracellular polymers, and the abundance changes of these genera were significantly correlated with Hydropobic TB-PN, LB-PS, and Ionic TB-PS. Genera such as Ferritrophicum, Sulfurimonas, and Simplicispira showed a significant positive correlation with LB-PS and Ionic TB-PS, indicating that these denitrifying bacteria may enhance their ability to attach to sulfur packing by promoting polysaccharide secretion, thereby achieving efficient nitrogen removal.

[0081] Example 6: Metagene Annotation Functional Analysis in Coupled Systems

[0082] COG functional annotation was performed on genes from the nitrifying aerobic granular sludge and the sulfur autotrophic denitrifying sludge (day 0 and day 80) in the coupled system. The results of the functional gene annotation are attached. Figure 9As shown, 25 major COG functions were annotated. The COG gene function results among the samples were basically similar, with only minor differences, indicating that the main COG functions of the microorganisms in nitrifying aerobic granular sludge and sulfur autotrophic denitrifying sludge were essentially the same. The top three major functions of nitrifying aerobic granular sludge were: cell wall / membrane / envelope biogenesis (M), general function prediction only (R), and amino acid transport and metabolism (E). In contrast, the top three major functions of sulfur autotrophic denitrifying sludge were: cell wall / membrane / envelope biogenesis, general function prediction only, and signal transduction mechanisms (T). This indicates that nitrifying aerobic granular sludge has stronger amino acid transport and metabolism, while sulfur autotrophic denitrification has stronger signal transduction mechanisms. Translation, ribosomal structure, and biogenesis (J) were all elevated in both systems, indicating that the microorganisms in the reactor continuously adapted to the environment to enhance their growth and reproduction capabilities. At day 80, the relative proportion of energy production and conversion (C) of nitrifying aerobic granular sludge was 6.14%, while the relative proportion of this function of sulfur autotrophic denitrifying sludge was 6.57%, indicating that the energy production functions such as ATP synthesis of sulfur autotrophic denitrifying sludge were stronger than those of nitrifying aerobic granular sludge.

[0083] The main nitrogen metabolism pathways in the metagenomic database were screened, and genes containing nitrogen metabolism pathways were retained to construct a nitrogen metabolism gene set. Genes mainly involved in nitrification were analyzed, and nitrification genes in nitrifying aerobic granular sludge were analyzed. The main genes for nitrification were nxrA, nxrB, pmoA-amoA, pmoB-amoB, pmoC-amoC, and hao. Compared with day 0, the relative abundance of each nitrification gene showed an increasing trend, indicating that the nitrification function of the reactor gradually increased. In the NH3→NH2OH process, the ammonia monooxygenase pmoC-amoC was the main enzyme, and its relative abundance increased from 2.40% to 2.46%. The relative abundance of pmoA-amoA increased by 0.13%, and the relative abundance of pmoB-amoB increased the most, by 1.19%. The relative abundance of the hydroxylamine dehydrogenase gene hao also increased significantly, from 1.62% to 2.39%, indicating that an ammonia nitrogen environment of 30 mg / L is conducive to the enrichment of the hao gene. The relative abundance of nitrite oxidases nxrA and nxrB was the highest and showed an increasing trend, with nxrA reaching a maximum relative abundance of 3.68%. Analysis of denitrification functional genes in sulfur-autotrophic denitrifying sludge revealed that the main functional genes involved in denitrification included narH, narG, narI, napA, napB, nirS, nirK, norB, norC, and nosZ. Figure 3-1 9) During the reduction of nitrate to nitrite, the relative abundance of narH and narG decreased, but the relative abundance of narI increased from 2.96% to 5.32%, indicating that the addition of nitrifying aerobic granular sludge effluent is beneficial for the enrichment of narI gene abundance. Secondly, the relative abundance of napA decreased, while the relative abundance of napB increased, indicating that the abundance of the nitrate reductase subunit napA decreased and the abundance of the nitrate reductase electron transfer subunit increased as HRT gradually decreased. In the nitrite to nitric oxide pathway, the relative abundance of both nirS and nirK genes increased, and the abundance of nirS gene was twice that of nirK, indicating that increased denitrification load is beneficial for the enrichment of denitrification genes. During the nitric oxide to nitrous oxide reduction process, the relative abundance of norB and norC increased, with norB gene being dominant, but the relative abundance of norC gene increased significantly, from 1.44% to 3.31%. Finally, during the reduction of nitrous oxide to nitrogen gas, which is the final denitrification process, the abundance of the nosZ functional gene also increases. Overall, with the increase of the denitrification load, all genes in the denitrification pathway are effectively enriched.

[0084] The embodiments provided above are not intended to limit the scope of the invention, nor are the described steps intended to limit the order of execution. Any obvious modifications made to the invention by those skilled in the art based on existing common knowledge also fall within the scope of protection defined by the claims.

Claims

1. A method for cultivating nitrifying aerobic granular sludge based on micro-nano aeration technology, characterized in that, The method includes the following steps: (1) Inoculate sludge into a sequencing batch reactor and start the reactor for cultivation. In the initial stage of operation, the influent time is 3-7 min, aeration time is 200-205 min, settling time is 10-20 min, drainage time is 3-7 min, and idle time is 5-15 minutes. After the reactor is running stably, the aeration and settling time are adjusted to 210-215 min and 5-7 min, respectively, and the drainage ratio is 55-65%. After cultivation for 25-35 days, the aeration and settling time are adjusted to 150-160 min and 3-7 min, respectively, to obtain aerobic granular sludge. An aeration device is installed at the bottom of the sequencing batch reactor, and the aeration device is equipped with a fine aeration head with an pore size of 0.2 to 0.3 μm; the aeration rate of the control device is 1 to 5 L / min during the cultivation process; (2) After the aerobic granular sludge is formed, the influent conditions are gradually changed by simultaneously reducing the C / N ratio, and the sludge is acclimatized and cultivated into nitrified aerobic granular sludge.

2. The method according to claim 1, characterized in that, The sludge used for inoculation in step (1) was taken from the aerobic tank of the wastewater treatment plant; the MLSS of the inoculated sludge was 4-5 g / L.

3. The method according to claim 1, characterized in that, The components of the influent mentioned in step (1) include: a mixed carbon source of CH3COONa and CH3CH2COONa, a COD of 100-1000 mg / L, and NH4+. + -N concentrations are 30-50 mg / L, 20-30 mg / L KH2PO4, 30-40 mg / L K2HPO4, 90-100 mg / L MgSO4, 70-80 mg / L CaCl2, 5-15 mg / L EDTA, 1-2 mg / L FeCl3·6H2O, 0.1-0.2 mg / L H3BO3, 0.03-0.05 mg / L CuSO4·5H2O, 0.03-0.05 mg / L KI, 0.1-0.2 mg / L MnCl2·4H2O, 0.05-0.07 mg / L ZnCl2, 0.1-0.2 mg / L CoCl2·6H2O, and 0.05-0.1 mg / L Na2MoO4·2H2O.

4. The method according to claim 1, characterized in that, The acclimatization and cultivation process described in step (2) includes: gradually changing the influent conditions by simultaneously reducing the C / N ratio; the acclimatization and cultivation conditions are divided into two stages: Phase I: Influent COD concentration decreases from 1000 mg / L to 100 mg / L, NH4+... + The -N concentration was decreased from 50 mg / L to 30 mg / L, and the treatment period was 35–45 days. Stage Ⅱ: Influent COD concentration is 0, NH4+ + The -N concentration was 30 mg / L, and the treatment period was 30–35 days.

5. Nitrified aerobic granular sludge prepared by the method according to any one of claims 1 to 4.

6. The application of the nitrified aerobic granular sludge as described in claim 5 in wastewater treatment.

7. A nitrogen removal method based on a coupled system of nitrifying aerobic granular sludge and sulfur autotrophic denitrification, characterized in that, The method includes: Wastewater first enters the nitrification aerobic granular sludge tank for treatment, then enters the storage tank to adjust the pH, and finally enters the sulfur autotrophic denitrification filter for treatment, and finally the effluent completes denitrification. The nitrifying aerobic granular sludge described in claim 5 is added to the nitrifying aerobic granular sludge tank, with the addition amount being 25-35% of the volume of the nitrifying aerobic granular sludge tank; the sulfur autotrophic denitrification filter is inoculated with sulfur autotrophic denitrification sludge and filled with elemental sulfur; the sludge concentration of the sulfur autotrophic denitrification sludge is 1500-2500 mg / L; the filling amount of the elemental sulfur is 70-90% of the volume of the sulfur autotrophic denitrification filter.

8. The denitrification method according to claim 7, characterized in that, The wastewater has a COD of 0–500 mg / L and a TN of 10–50 mg / L.

9. The denitrification method according to claim 7, characterized in that, Wastewater is treated in a nitrifying aerobic granular sludge tank for one cycle; in the cycle, water inflow is 5-8 minutes, aeration is 210-215 minutes, settling is 5-10 minutes, drainage is 5-10 minutes, and idle time is 5-15 minutes.

10. The denitrification method according to claim 7, characterized in that, The hydraulic retention time of wastewater in a sulfur autotrophic denitrification filter is 1.5 to 10 hours.

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