Method for improving nitration efficiency under alkaline condition by regulating extracellular polymeric substance and ion transport

Abstract

The invention discloses a method for improving nitration efficiency under alkaline conditions by regulating extracellular polymeric substances and ion transport. According to the method, through comprehensive strategies of gradually increasing the pH value of the system, controlling the number of operation days and the like, nitrobacterium extracellular polymeric substances are induced to secrete more acidic polysaccharides and acidic organic matters, and antitransporter and F0-F1 ATP enzyme activity are regulated and controlled, so that the input of intracellular H < + > is promoted to maintain the intracellular pH stable. Meanwhile, the consumption of extracellular OH <-> is accelerated by utilizing a biomineralization effect participated by carbonic anhydrase, the alkaline environment adaptability and nitrification performance of the nitrification sludge are improved, alkali-resistant nitrifying bacteria such as Nitrosomonas and Nitrospira are remarkably enriched, and the average ammonia nitrogen removal rate of the system can stably reach 96.6% under the condition that the pH is 10.0.

Description

Technical Field

[0001] The invention belongs to the field of sewage biological treatment technology and relates to a method for improving nitrification efficiency under alkaline conditions by regulating extracellular polymers and ion transport. Background Art

[0002] The effect of nitrification in biological denitrification systems is affected by various operating parameters such as temperature, hydraulic retention time (HRT), solid retention time (SRT) and pH. It is worth noting that the alkaline environment becomes one of the main challenges faced by nitrifying bacteria due to wastewater quality shock or system characteristics (such as the pH value of the bacterial-algal symbiotic system fluctuates between 6.0 and 10.0 during photosynthesis). Normally, when the pH is greater than 9.6, the biological activity of nitrifying bacteria will be inhibited or even stopped. High pH has a significant adverse effect on nitrifying bacteria, which poses a challenge to the actual stable operation of the activated sludge system. Therefore, it is necessary to develop a method to improve the adaptability and nitrification efficiency of nitrifying bacteria in an alkaline environment.

[0003] pH is an important environmental factor that affects the performance of nitrifying sludge and the structure of microbial communities, and nitrifying bacteria are very sensitive to its changes. Too high pH will not only inhibit ammonia oxidizing bacteria (AOB) and nitrite oxidizing bacteria (NOB), but also destroy NH 4 + -N and NH 3 -N, resulting in NH 3 Gas escapes and the concentration of free ammonia (FA) increases. As an inhibitor of nitrifying bacteria, FA can not only diffuse passively into the cytoplasm through the cell membrane, bind to protons in the cell and convert into ammonium, causing cell alkalization, but also inhibit the synthesis of intracellular ATP, affecting cell energy supply. Therefore, most nitrifying bacteria (such as AOB Nitrosomonas) are difficult to survive in a highly alkaline environment. NOB Nitrospira has a slightly stronger adaptability, but in the current study, only two strains of this genus can grow in an environment with a pH value higher than 8.0 (Daebeler A, Kitzinger K, Koch H, et al. Exploring the upper pH limits of nitrite oxidation: diversity, ecophysiology, and adaptive traits of haloalkalitolerant Nitrospira[J]. ISME J, 2020, 14(12): 2967-2979.).

[0004] At present, some studies have explored the domestication method of alkaline-resistant activated sludge. For example, Chinese patent CN201710835023.8 dehydrates and reactivates the activated sludge, and then adds a certain amount of ammonia solution every day to achieve alkaline-resistant domestication. In addition, Chinese patent CN201210340270.8 domesticates alkaline-resistant activated sludge by adding organic alkaline wastewater to the system to gradually increase the pH value. However, these domestication methods cannot ensure that the activated sludge is always kept in a highly alkaline environment, and none of them systematically evaluates the denitrification performance of the activated sludge after domestication, especially the research on nitrification activity is relatively lacking. Therefore, exploring how to improve the nitrification efficiency under alkaline conditions through adaptive regulation of nitrifying bacteria has important research and application value for ensuring the stable operation of the activated sludge system in a high pH environment. Summary of the invention

[0005] In view of the problems of inhibited microbial activity, increased toxicity, sludge bulking, and poor sedimentation in activated sludge systems under high pH environments, the present invention provides a method for improving nitrification efficiency under alkaline conditions by regulating extracellular polymers and ion transport. The method significantly improves the adaptability of nitrifying bacteria through a series of pathways such as gradually increasing the system pH value and controlling the number of operating days, secreting acidic organic matter in extracellular polymers, and maintaining intracellular pH stability through cation / proton antiporters, thereby enriching nitrifying sludge with strong alkaline resistance and achieving high ammonia nitrogen removal rate and nitrification activity at a constant pH of 10.0.

[0006] The technical solution of the present invention is as follows:

[0007] The method of improving the nitrification efficiency under alkaline conditions by regulating extracellular polymers and ion transport is as follows:

[0008] A nitrifying membrane bioreactor was constructed, which adopted a continuous flow operation mode, with continuous aeration at the bottom of the reactor, and the effluent was filtered using an immersed microfiltration membrane module to separate biomass and wastewater;

[0009] During the operation of the nitrifying membrane bioreactor, the environmental pH value in the reactor is gradually increased through three periods, including the stabilization period, the alkaline stimulation period and the alkaline adaptation period, and the number of operating days is controlled by monitoring the level of active oxygen in the system. These comprehensive control strategies are used to promote the secretion of acidic substances in the extracellular polymers of nitrifying bacteria, regulate the antiporter and F 0 -F 1 ATPase, and biomineralization using carbonic anhydrase to accelerate OH - The consumption and H + The input of nitrifying sludge can further improve its alkaline adaptability and nitrification performance;

[0010] After entering the alkaline adaptation period, the nitrifying membrane bioreactor continues to operate in a constant high pH environment. When the activity of nitrifying bacteria recovers to the same level as that in the stable period and the content of extracellular polymers and the activity of related functional proteins change significantly, an alkaline-resistant and efficient nitrifying microbial community is obtained.

[0011] Preferably, the stable period is a neutral environment; the alkaline stimulation period is a slightly alkaline environment; and the alkaline adaptation period is a highly alkaline environment.

[0012] Preferably, the method for controlling the number of operations is to gradually increase the pH value of the environment and enter the next stage when the active oxygen level in the system drops to less than 50% of that in the middle stage.

[0013] Preferably, an automated pH control system is used to control the pH value in the reactor in real time. When the pH value is lower than the set value, the dosing pump is turned on to deliver NaOH solution to the reactor to increase the pH value. When the pH value is higher than the set value, the dosing pump is turned off. Due to the ammonia oxidation effect of microorganisms, the pH value will be further reduced. The automated pH control system consists of a pH probe, a pH controller, a dosing pump, and a NaOH solution storage tank.

[0014] Preferably, when the active oxygen level drops below 50% of that in the middle period during each pH gradient, the sludge microbial samples are collected to measure the content of extracellular polymers and the activity of related functional proteins.

[0015] Preferably, the initial sludge concentration in the nitrifying membrane bioreactor is 2000-3000 mg / L, the dissolved oxygen concentration in the system is maintained at 3.0-4.0 mg / L, and the hydraulic retention time is 6-9 h.

[0016] Compared with the prior art, the present invention has the following advantages:

[0017] (1) The present invention utilizes comprehensive control strategies such as pH gradient enhancement and operation days control. At pH 10.0, the alkali-resistant nitrifying bacteria are controlled to secrete acidic polysaccharides in the extracellular polymer (EPS), and glutamate dehydrogenase (GDH) is used to increase acidic organic matter to neutralize the external alkaline environment. In addition, by regulating the antiporters (such as NhaA, TrkA, and KefB) and F 0 -F 1 ATPase promotes H + The input of OH is used to maintain the intracellular pH stability, and a series of biomineralization processes involving carbonic anhydrase (CA) are used to accelerate the OH - The consumption of Nitrosomonas and Nitrospira increased the abundance to 1.3% and 5.4%, respectively, significantly improving the adaptability of nitrifying bacteria in a high alkaline environment.

[0018] (2) The present invention successfully obtained alkali-resistant nitrification sludge that maintained efficient nitrification performance under highly alkaline conditions by using a nitrification bioreactor, and its average ammonia removal rate at pH 10.0 was as high as 96.6±2.2%. At the same time, the alkali-resistant nitrification bacteria group was applicable to a wide pH range and had high activity at pH 7.5-10.0, with NOB activity reaching 14.6 mg N·g -1 MLVSS·h -1 , which is equivalent to the activity level in a neutral environment, shows high stability and strong adaptability, and has potential application prospects in the field of biological sewage treatment. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1 Schematic diagram of the nitrifying membrane bioreactor.

[0020] Figure 2 (a) Protein, (b) Polysaccharide and (c) Total EPS content changes in EPS, (df) Three components in EPS obtained by three-dimensional fluorescence parallel factor analysis and their (gi)F max Changes in value.

[0021] Figure 3 The changes in the content of relevant functional proteins obtained by PICRUSt2 based on the KEGG database.

[0022] Figure 4 This is the daily changes in the inlet and outlet water quality during the operation of the nitrifying membrane bioreactor.

[0023] Figure 5 (a) AOB and NOB activities of nitrifying sludge at the end of each stage, (b) AOB and NOB activities of alkaline-tolerant nitrifying sludge at pH values ​​of 7.5, 8.0, 9.0 and 10.0 after acclimation, and (c) changes in reactive oxygen species (ROS) levels in nitrifying sludge in the middle and end of each stage.

[0024] Figure 6 The abundance changes of some functional bacterial genera in the reactor at different stages.

[0025] Figure 7 (a) Changes in activated sludge concentration during sludge acclimation, (b) changes in SVI, (c) comparison of XRD spectra of seed sludge and alkali-resistant nitrification sludge, (d) comparison of photos after calcination, (e) comparison of infrared spectra, (f) scanning electron microscope images of seed sludge and (g) alkali-resistant nitrification sludge. DETAILED DESCRIPTION

[0026] The present invention is further described in detail below in conjunction with specific embodiments and drawings, but the embodiments of the present invention are not limited thereto.

[0027] The reagents used in the following examples are all available from commercial sources.

[0028] Example 1

[0029] Construction of nitrifying membrane bioreactor:

[0030] A continuous flow nitrification membrane bioreactor with an effective volume of about 2 L was established. Figure 1 As shown, the reactor is made of plexiglass. The effluent uses a filtration submerged microfiltration membrane assembly (made of PVDF membrane, membrane fiber inner diameter 1.6mm, outer diameter 2.8mm, micropore diameter 0.1μm) to separate biomass and wastewater to ensure that all biomass is retained in the bioreactor. The initial sludge concentration is 2560mg / L. Microporous aeration is set at the bottom of the reactor, and continuous aeration is performed by an air pump. A pH probe and a dissolved oxygen (DO) probe are set in the reactor to monitor the pH and DO values ​​in the reactor in real time, so that the DO concentration is maintained at 3.0-4.0mg / L, and the pH value is regulated by an automated pH control system. The reactor inlet uses synthetic domestic sewage without a carbon source, and the inlet and outlet flow rates are controlled by a peristaltic pump. The HRT is 8.94h, and the HRT is shortened to 6.19h after the 75th day to increase the nitrification rate.

[0031] The operation of the nitrifying membrane bioreactor gradually increases the environmental pH value in the reactor through three periods: first, the stabilization period (Phase I, 1-28d), the environmental pH is 7.5. Then there is the alkaline stimulation period, during which the environmental pH value gradually increases to 8.0±0.1 (Phase II, 29-58d), 8.5±0.1 (Phase III, 59-90d), 9.0±0.1 (Phase IV, 91-120d), and finally 9.5±0.1 (Phase V, 121-150d). Finally, there is the alkaline adaptation period, when the pH value rises to 10.0±0.1 (Phase VI, 151-320d). In addition, the control strategy of the number of operating days is used to further domesticate the nitrifying sludge. Whenever the active oxygen level in the system drops to less than 50% of the mid-stage, the pH value is increased by gradient, which is considered to enter the next stage.

[0032] The composition of synthetic domestic sewage is as follows: NH 4 Cl(229.28mg / L), K 2 HPO 4 (21.95mg / L), MgSO 4 7H 2 O (91.2 mg / L), CaCl 2 (11.1mg / L), trace element solution (1mL / L). Trace element solution composition: 1.5g / LFeCl 3 6H2 O, 0.18 g / L KI, 0.15 g / L H 3 BO 3 , 0.12g / L MnCl 2 ·4H 2 O, 0.12g / L ZnSO 4 7H 2 O, 0.06g / LNaMoO 4 ·2H 2 O, 0.03 g / L CuSO 4 ·5H 2 O, 10 g / L EDTA-2Na.

[0033] Example 2

[0034] Methods for improving the alkali resistance of nitrifying bacteria and the effect of improving nitrification efficiency:

[0035] By regulating the various physiological characteristics of nitrifying bacteria, the alkali resistance of nitrifying bacteria can be improved. Figure 2 As shown in Figure 2, under a high alkaline environment, by promoting bacteria to secrete a large amount of acidic organic matter (such as acidic polysaccharides, humic acid, etc.) in EPS, these substances can act as buffers to neutralize the external alkalinity, thereby inhibiting the excessive OH - Damage to cells. Figure 3 As shown, it catalyzes Na + / K + The output and H + The input of 200 μL of dissolved oxygen can maintain the intracellular pH at around 7-8.5. This pathway is the core of bacteria maintaining intracellular pH homeostasis. Generally, a high pH environment can provide a higher transmembrane potential under high aeration conditions. Therefore, a continuous high dissolved oxygen environment in the reactor can increase F 0 -F 1 ATPase transcription, H + At the same time, by increasing the GDH content, the production of organic acids (i.e., glutamate) is increased to ensure the normal energy metabolism of the cells. In addition, by increasing the content of carbonic anhydrase, CO 2 To HCO 3 - and convert HCO 3 - Converted to CO 3 2- To further consume OH - , and then with Ca in the water 2+ The reaction forms calcite crystals CaCO 3The biomineralization accelerates the formation of OH - consumption, preventing cell damage.

[0036] Figure 4 The NH in the inlet and outlet water during the operation of the nitrifying membrane bioreactor is shown. 4 + -N, NO 2 - -N and NO 3 - -N concentration. Obviously, when the pH is below 9.0, the ammonia nitrogen removal rate is always 100%, so the HRT is shortened to 6.19h after 75d to improve the nitrification efficiency. Although the stability of the effluent decreased slightly when the pH increased to 9.5 and 10.0, the average ammonia nitrogen removal rate remained at 96.6±2.2%, significantly improving the nitrification efficiency in a high pH environment. Figure 5 As shown in (a), the activity of AOB decreased significantly by 78.5% at high pH, ​​while NOB was inhibited at pH 8.5 and 9.0, but its activity gradually recovered to 14.6 mg N g -1 MLVSS·h -1 , has been significantly improved, achieving efficient and stable nitrification. At the same time, the applicable pH range of the alkaline nitrification sludge is wide, and it maintains high activity in the pH range of 7.5 to 10.0 ( Figure 5 (b)), and reached the highest activity at pH 10.0, indicating the effective enrichment of alkali-tolerant nitrifying bacteria. In addition, the reduction of ROS levels at the end of each stage ( Figure 5 (c)) The nitrifying sludge gradually adapted to the stress stimulation of the external alkaline environment and improved its adaptability. In addition, the bacteria significantly improved their alkaline adaptability through the above-mentioned multiple pathways, so nitrifying bacteria such as AOB Nitrosomonas (1.3%), norank_f__JG30-KF-CM45 (2.0%) and NOB Nitrospira (5.4%) were significantly enriched ( Figure 6 ), which means the successful domestication of alkali-resistant and efficient nitrifying sludge.

[0037] according to Figure 7 (f) and (g) show that there are large pieces of flaky sediments and some granular aggregates on the surface of the alkali-resistant nitrification sludge, mainly CaCO 3 ( Figure 7 (c)-(e)), but its structure is relatively loose, while the seed sludge has stronger agglomeration. Despite this, the alkali-resistant nitrification sludge still has good sedimentation performance, with an SVI value of only 121.4±3.6mL / g, and no sludge bulking phenomenon occurs ( Figure 7(a)-(b)), which is attributed to the increase of carbonic anhydrase, which leads to a large amount of CaCO attached to the sludge surface. 3 Improved the settling of sludge flocs.

[0038] In summary, the method of the present invention promotes the secretion of acidic polysaccharides, regulates cation / proton antiporters and F 0 -F 1 ATPase and increase the activity of carbonic anhydrase to accelerate OH - The adaptability of nitrifying bacteria to highly alkaline environments has been significantly improved through multiple channels such as the consumption of nitrifying bacteria. At the same time, the nitrification efficiency has been dynamically improved, and the alkali-resistant nitrifying bacteria have been successfully enriched, showing significant environmental benefits, with good practical value, and of great significance for promoting the development of sewage treatment technology.

[0039] The above embodiments are preferred implementation modes of the present invention, but the implementation modes of the present invention are not specifically limited by the above embodiments. Within the technical concept of the present invention, a variety of simple modifications can be made to the technical scheme of the present invention, and these simple modifications all belong to the protection scope of the present invention.

[0040] It should also be noted that the various specific technical features described in the above specific embodiments can be combined in any suitable manner without contradiction. In order to avoid unnecessary repetition, the present invention will not further describe various possible combinations.

[0041] In addition, various embodiments of the present invention may be arbitrarily combined, and as long as they do not violate the concept of the present invention, they should also be regarded as the contents disclosed by the present invention.

Claims

1. A method for improving nitrification efficiency under alkaline conditions by regulating extracellular polymers and ion transport, characterized in that: The details are as follows: A nitrifying membrane bioreactor was constructed, which adopted a continuous flow operation mode, with continuous aeration at the bottom of the reactor, and the effluent was filtered using an immersed microfiltration membrane module to separate biomass and wastewater; During the operation of the nitrifying membrane bioreactor, the environmental pH value in the reactor is gradually increased through three periods, including the stabilization period, the alkaline stimulation period and the alkaline adaptation period, and the number of operating days is controlled by monitoring the level of active oxygen in the system. These comprehensive control strategies are used to accelerate OH by promoting the secretion of acidic substances in the extracellular polymers of nitrifying bacteria, regulating antiporters and F0-F1 ATPase, and utilizing the biomineralization process involving carbonic anhydrase. - The consumption and H + The input of nitrifying sludge can further improve its alkaline adaptability and nitrification performance; After entering the alkaline adaptation period, the nitrifying membrane bioreactor continues to operate in a constant high pH environment. When the activity of nitrifying bacteria recovers to the same level as that in the stable period and the content of extracellular polymers and the activity of related functional proteins change significantly, an alkaline-resistant and efficient nitrifying microbial community is obtained.

2. The method according to claim 1, characterized in that The stable period is a neutral environment; the alkaline stimulation period is a slightly alkaline environment; and the alkaline adaptation period is a highly alkaline environment.

3. The method according to claim 1, characterized in that The method for controlling the number of operating days is to gradually increase the environmental pH value and enter the next stage when the active oxygen level in the system drops to less than 50% of the mid-stage.

4. The method according to claim 1, characterized in that: An automated pH control system is used to control the pH value in the reactor in real time. When the pH value is lower than the set value, the dosing pump is turned on to deliver NaOH solution to the reactor to increase the pH value. When the pH value is higher than the set value, the dosing pump is turned off, and the pH value will be further reduced due to the ammonia oxidation effect of microorganisms.

5. The method according to claim 4, characterized in that The automatic pH control system is composed of a pH probe, a pH controller, a dosing pump and a NaOH solution storage tank.

6. The method according to claim 1, characterized in that When the reactive oxygen level dropped below 50% of that at the middle stage during each pH gradient, sludge microbial samples were collected to measure the content of extracellular polymers and the activity of related functional proteins.

7. The method according to claim 1, characterized in that The initial sludge concentration in the nitrifying membrane bioreactor was 2000~3000 mg / L, the dissolved oxygen concentration in the system was maintained at 3.0~4.0 mg / L, and the hydraulic retention time was 6~9 h.

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