Method for improving denitrification performance of free anaerobic ammonium oxidation bacteria

Abstract

The invention discloses a method for improving denitrification performance of free anaerobic ammonium oxidation bacteria, which comprises the following steps: firstly, preparing hot air oxidized biochar by hot air oxidation, then adding the hot air oxidized biochar into a free anaerobic ammonium oxidation bacteria culture system, removing oxygen, and then sealing and continuously culturing in a shaking table. The hot air oxidation biochar is easy in raw material obtaining, simple in preparation and small in dosage, when the hot air oxidation biochar is added into a free anaerobic ammonium oxidation bacterium culture system, the anaerobic ammonium oxidation process starting time is shorter, the accumulated consumption of NH4 < + >-N and NO2 <->-N is higher, the system still keeps higher anaerobic ammonium oxidation activity after long-term operation, and the whole operation is simple, convenient and feasible. The prepared hot air oxidation biochar can be used as a free anaerobic ammonium oxidation bacterium growth carrier, can effectively promote growth and reproduction of free anaerobic ammonium oxidation bacteria, is rich in surface functional groups and high in electron transfer capacity, and can promote electron transfer among microorganisms, so that the anaerobic ammonium oxidation process is accelerated, and the denitrification performance of a system is improved.

Description

Technical Field

[0001] The present invention relates to the technical field of water treatment, and more particularly to a method for improving the denitrification performance of free anaerobic ammonia oxidizing bacteria. Background Art

[0002] It is difficult to utilize and treat total nitrogen in wastewater. A large amount of nitrogen is discharged into natural water bodies, which not only causes eutrophication and ecological imbalance in rivers, lakes and reservoirs, but also threatens human health. Therefore, denitrification is a key issue in the field of wastewater treatment. Since its discovery in 1995, the anaerobic ammonium oxidation mechanism has attracted widespread attention and has been widely used in various wastewater treatments. Compared with traditional biological denitrification processes, the anaerobic ammonium oxidation process uses NH4 + -N and NO2--N are used as substrates to react and generate N2 to achieve the purpose of denitrification. No external organic carbon source is required in the process. It is energy-saving and efficient, and is currently one of the most promising wastewater denitrification technologies.

[0003] At present, the anaerobic ammonium oxidation process has been successfully applied to the treatment of various high-nitrogen wastewaters such as pig farm wastewater, landfill leachate, and monosodium glutamate production wastewater, and has achieved good results and economic benefits. However, the large-scale promotion and application of this process is still limited by many factors, such as: functional microorganisms are not easy to enrich, grow slowly, and the process is poorly stable. Many studies have shown that the addition of biochar, graphene or carrier materials to the anaerobic ammonium oxidation process system can improve the stability of anaerobic ammonium oxidation granular sludge, thereby improving the system's denitrification performance and enhancing the system's stability. Most of these studies are based on anaerobic ammonium oxidation granular sludge. Anaerobic ammonium oxidation granular sludge can not only withstand higher loads, but also help accumulate and maintain high biomass concentrations. In actual engineering applications, anaerobic ammonium oxidation granular sludge is not easy to cultivate and cannot be quickly enriched. The reason is that the core functional microorganisms of the anaerobic ammonium oxidation process-anaerobic ammonium oxidizing bacteria are chemo-inorganic autotrophic bacteria, which grow slowly, have a long doubling time, and are very sensitive to environmental conditions. Therefore, providing a method for the rapid growth and reproduction of anaerobic ammonium oxidizing bacteria in a free state to quickly start the anaerobic ammonium oxidation process and improve its agglomeration and denitrification performance is of great significance for the practical application of the anaerobic ammonium oxidation process. Summary of the invention

[0004] In view of this, the present invention provides a method for improving the denitrification performance of free anaerobic ammonium oxidizing bacteria. Adding hot air oxidized biochar to the free anaerobic ammonium oxidizing bacteria culture system can accelerate the start-up of anaerobic ammonium oxidation, promote the aggregation and growth and reproduction of anaerobic ammonium oxidizing bacteria, and improve the denitrification performance of anaerobic ammonium oxidizing bacteria.

[0005] To achieve the above object, the present invention adopts the following technical solutions:

[0006] A method for improving the denitrification performance of free anaerobic ammonium oxidizing bacteria comprises the following steps:

[0007] (1) Preparation of hot air oxidized biochar: placing the biochar in a muffle furnace, heating it under air atmosphere, and then cooling it to room temperature to obtain hot air oxidized biochar;

[0008] (2) Add hot air oxidized biochar to the free anaerobic ammonia oxidizing bacteria culture system, remove oxygen and seal it in a shaker for continuous culture.

[0009] Preferably, the biochar is wood chip charcoal.

[0010] Preferably, in step (1), the heating rate is 5°C / min, the temperature is raised to 350°C and maintained for 60 minutes.

[0011] Preferably, in step (2), the amount of hot air oxidized biochar added is 1 g / L based on the free anaerobic ammonia-oxidizing bacteria culture system.

[0012] Preferably, the preparation method of the free anaerobic ammonia-oxidizing bacteria is:

[0013] The anaerobic ammonium oxidation granular sludge was ball-milled and dispersed, and then placed in a NH4-free + -N, NO 2- -N and NO3 - -N medium and allowed to settle to obtain the supernatant, which is the free anaerobic ammonia-oxidizing bacteria.

[0014] Furthermore, the ball milling speed was 1800 rpm and the time was 10 s.

[0015] Furthermore, the composition of the culture medium is:

[0016] NaHCO3: 1g / L, KH2PO4: 10mg / L, CaCl2·H2O: 5.6mg / L, MgSO4·7H2O: 300mg / L, EDTA: 2 0mg / L, FeSO4·7H2O: 6.25g / L, MnCl2·4H2O: 0.99mg / L, CuSO4·5H2O: 0.25mg / L, ZnSO 4·7H2O: 0.43mg / L, NiCl2·6H2O: 0.19mg / L, CoCl2·6H2O: 0.24mg / L, Na2MoO4·2H2O: 0.22mg / L, Na2SeO4·10H2O: 0.21mg / L, Na2WO4·2H2O: 0.05mg / L, H3BO4: 0.014mg / L.

[0017] Preferably, the VSS concentration in the free anaerobic ammonia oxidizing bacteria culture system in step (2) is 0.07±0.01 g / L, and the NH4 + -N concentration 26.43mg / L, NO2- -N concentration 35mg / L.

[0018] Preferably, in step (2), nitrogen is filled to deoxygenate, and the filling time is 20 minutes.

[0019] Preferably, the shaking parameters in step (2) are: 35° C., 160 rpm.

[0020] Preferably, during the continuous culture in step (2), 50% of the volume of the culture medium is replaced in the anaerobic glove box every other week, and the substrate concentration is supplemented to be equal to the initial concentration.

[0021] It can be seen from the above technical solution that, compared with the prior art, the present invention discloses a method for improving the denitrification performance of free anaerobic ammonia-oxidizing bacteria, which has the following beneficial effects:

[0022] The biochar prepared by the present invention is treated with hot air, and its specific surface area and total pore volume are large, which is suitable for the attachment and growth of free anaerobic ammonium oxidizing bacteria, and can effectively promote the growth and reproduction of anaerobic ammonium oxidizing bacteria; at the same time, the hot air oxidized biochar prepared by the present invention has rich surface functional groups and strong electron transfer ability, and can act as an electron shuttle in the anaerobic ammonium oxidation process to promote electron transfer between microorganisms, thereby effectively improving the activity of anaerobic ammonium oxidation, accelerating the start-up of the anaerobic ammonium oxidation process, and enhancing the denitrification performance of the system. At the same time, the raw materials of the hot air oxidized biochar prepared by the present invention are easy to obtain and the amount used is small, which greatly saves resources and reduces costs, and has strong engineering application value. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments or the description of the prior art will be briefly introduced below. Obviously, the drawings described below are only embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on the provided drawings without paying creative work.

[0024] Figure 1 This is a picture of free anaerobic ammonium oxidizing bacteria under a fluorescence microscope;

[0025] Figure 2 This is a schematic diagram of the cumulative consumption of ammonia nitrogen and nitrite nitrogen and the accumulation of nitric nitrogen during the long-term continuous operation of Example 1;

[0026] Figure 3 This is the electron microscope image of the sludge after long-term operation of Example 1;

[0027] Figure 4 Fourier infrared image of hot air oxidized biochar prepared in Example 1;

[0028] Figure 5CV curve and electrochemical impedance spectrum of the system after long-term operation in Example 1. DETAILED DESCRIPTION

[0029] The technical solution of the present invention will be clearly and completely described below in conjunction with the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0030] Example 1

[0031] A method for improving the denitrification performance of free anaerobic ammonia oxidizing bacteria, the method comprising the following steps:

[0032] (1) Preparation of hot air oxidized biochar: Wood chip charcoal obtained from a pyrolysis gasification plant was placed in a muffle furnace, heated to 350°C at a rate of 5°C / min in an air atmosphere, maintained for 60 min, and then cooled to room temperature to obtain hot air oxidized biochar;

[0033] (2) Continuous culture was performed in 150 ml serum bottles. Two experimental groups were set up and named: CK and mod_C. 20 ml of free anaerobic ammonia oxidizing bacteria were inoculated in serum bottles and culture medium was added to supplement the culture system to 100 ml. VSS was 0.07 ± 0.01 g / L, NH4 + -N(NH4Cl) and NO2 - -N(NaNO2) initial concentrations were 26.43 mg / L and 35 mg / L, in which 0.1 g of hot air oxidized biochar was additionally added to the mod_C group, and then nitrogen was immediately filled for deoxygenation for 20 minutes. After the aeration, the rubber stopper was quickly plugged in, and the reaction device was placed in a shaker at 35°C for continuous culture at a speed of 160 rpm. The culture medium in the 50% serum bottle was replaced in the anaerobic glove box every other week, and the substrate concentration was supplemented to be equal to the initial concentration.

[0034] The preparation method of free anaerobic ammonium oxidizing bacteria is as follows: a number of anaerobic ammonium oxidizing granular sludge is placed in a ball mill and ground at a rate of 1800 rpm for 10 seconds, and then 5 ml of the ground free sludge is placed in 150 ml of NH4-free + -N, NO2 - -N and NO3 - -N medium was precipitated and allowed to stand for 30 min, and the supernatant obtained was the free anaerobic ammonia-oxidizing bacteria prepared, and the microscopic image thereof showed that the cells were in a free state, as shown in the attached Figure 1 shown.

[0035] The composition of the culture medium is: NaHCO3: 1g / L, KH2PO4: 10mg / L, CaCl2·H2O: 5.6mg / L, MgSO4·7H2O: 300mg / L, EDTA: 20mg / L, FeSO4·7H2O: 6.25g / L, MnCl2·4H2O: 0.99mg / L, CuSO4·5H2O: 0.25mg / L, ZnSO4·7H2O: 0.43mg / L, NiCl2·6H2O: 0.19mg / L, CoCl2·6H2O: 0.24mg / L, Na2MoO4·2H2O: 0.22mg / L, Na2SeO4·10H2O: 0.21mg / L, Na2WO4·2H2O: 0.05mg / L, H3BO4: 0.014mg / L.

[0036] The cumulative consumption of ammonia nitrogen and nitrite nitrogen in CK and mod_C treatments during long-term continuous operation is as follows Figure 2 As shown in the figure, since the inoculated bacteria were free anaerobic ammonium oxidizing bacteria, their ammonia nitrogen metabolism capacity was inhibited. At the beginning of the experiment, only nitrite nitrogen was consumed in both treatments, but no ammonia nitrogen was consumed. It was not until the 21st day that the mod_C group began to consume ammonia nitrogen, and the ammonia nitrogen consumption continued to increase during the subsequent culture process, while the CK group did not consume ammonia nitrogen until the 49th day, and the subsequent ammonia nitrogen consumption was always at a low level. On the 84th day, the cumulative consumption of ammonia nitrogen (517.3 mg / L) and nitrite nitrogen (854.3 mg / L) in the mod_C group were 27.4 times and 3.2 times that of the CK group (18.9 mg / L and 263.5 mg / L), respectively. It can be seen that the addition of heated air oxidized biochar started the anaerobic ammonia oxidation process faster and the denitrification performance was also better.

[0037] The specific surface area of ​​the hot air oxidized biochar prepared is 238.2 m 2 / g, total pore volume is 0.15cm 3 / g, SEM results of sludge after long-term operation ( Figure 3 ) showed that anaerobic ammonium oxidizing bacteria can attach to the surface of hot air oxidized biochar, and hot air oxidized biochar can provide a fulcrum for the growth of free anaerobic ammonium oxidizing bacteria, facilitating their agglomeration and growth. The surface of hot air oxidized biochar is rich in functional groups such as CO, COC (1060), and OH (1434). Cyclic voltammetry (CV) and electrochemical impedance spectroscopy can be used to reflect the electron transfer capacity of the system, such as Figure 5 As shown in A, under the same applied voltage, the response current of the mond_C group is much higher than that of the CK group; at the same time, the electrochemical impedance spectroscopy of the mod_C group ( Figure 5 B) The semicircle diameter of the fitting curve is smaller, which means that its electron transfer resistance is lower than that of the CK group. The above results show that the system with hot air oxidized biochar has a stronger electron transfer capacity.

[0038] In summary, the addition of hot air oxidized biochar enables the free anaerobic ammonium oxidizing bacteria culture system to start the anaerobic ammonium oxidation process faster and have better denitrification performance. The reason is that the hot air oxidized biochar has a large specific surface area and total pore volume, which can provide a growth fulcrum for the free anaerobic ammonium oxidizing bacteria and facilitate their agglomeration growth; at the same time, the hot air oxidized biochar has rich surface functional groups, and its addition to the system enhances the system's electron transfer capacity, thereby enhancing the system's anaerobic ammonium oxidation activity and improving the denitrification performance of free anaerobic ammonium oxidizing bacteria.

[0039] In this specification, each embodiment is described in a progressive manner, and each embodiment focuses on the differences from other embodiments, and the same and similar parts between the embodiments can be referred to each other. The above description of the disclosed embodiments enables professionals and technicians in this field to implement or use the present invention. Various modifications to these embodiments will be obvious to professionals and technicians in this field, and the general principles defined herein can be implemented in other embodiments without departing from the spirit or scope of the present invention. Therefore, the present invention will not be limited to these embodiments shown in this article, but will comply with the widest range consistent with the principles and novel features disclosed herein.

Claims

1. A method for improving the denitrification performance of free anaerobic ammonia-oxidizing bacteria, characterized in that: The following steps are involved: (1) Preparation of hot air oxidized biochar: placing the biochar in a muffle furnace, heating it under air atmosphere, and then cooling it to room temperature to obtain hot air oxidized biochar; (2) Add hot air oxidized biochar to the free anaerobic ammonia oxidizing bacteria culture system, remove oxygen and seal it in a shaker for continuous culture.

2. A method for improving the denitrification performance of free anaerobic ammonium oxidizing bacteria according to claim 1, characterized in that: The biochar is wood chip charcoal.

3. A method for improving the denitrification performance of free anaerobic ammonium oxidizing bacteria according to claim 1, characterized in that: In step (1), the heating rate is 5°C / min, the temperature is raised to 350°C and maintained for 60 minutes.

4. A method for improving the denitrification performance of free anaerobic ammonium oxidizing bacteria according to claim 1, characterized in that: In step (2), based on the free anaerobic ammonia-oxidizing bacteria culture system, the amount of hot air oxidized biochar added is 1 g / L.

5. A method for improving the denitrification performance of free anaerobic ammonium oxidizing bacteria according to claim 1, characterized in that: The preparation method of the free anaerobic ammonia oxidizing bacteria is as follows: The anaerobic ammonium oxidation granular sludge was ball-milled and dispersed, and then placed in a NH4-free + -N, NO 2- -N and NO3 - -N medium and allowed to settle to obtain the supernatant, which is the free anaerobic ammonia-oxidizing bacteria.

6. A method for improving the denitrification performance of free anaerobic ammonium oxidizing bacteria according to claim 5, characterized in that: The ball milling speed was 1800 rpm and the time was 10 s.

7. A method for improving the denitrification performance of free anaerobic ammonium oxidizing bacteria according to claim 5, characterized in that: The composition of the culture medium is: NaHCO3: 1g / L, KH2PO4: 10mg / L, CaCl2·H2O: 5.6mg / L, MgSO4·7H2O: 300mg / L, EDTA: 2 0mg / L, FeSO4·7H2O: 6.25g / L, MnCl2·4H2O: 0.99mg / L, CuSO4·5H2O: 0.25mg / L, ZnSO 4·7H2O: 0.43mg / L, NiCl2·6H2O: 0.19mg / L, CoCl2·6H2O: 0.24mg / L, Na2MoO4·2H2O: 0.22mg / L, Na2SeO4·10H2O: 0.21mg / L, Na2WO4·2H2O: 0.05mg / L, H3BO4: 0.014mg / L.

8. A method for improving the denitrification performance of free anaerobic ammonium oxidizing bacteria according to claim 1, characterized in that: The VSS concentration in the free anaerobic ammonia oxidizing bacteria culture system in step (2) is 0.07±0.01 g / L, and the NH4 + -N concentration 26.43mg / L, NO2 - -N concentration 35mg / L.

9. A method for improving the denitrification performance of free anaerobic ammonium oxidizing bacteria according to claim 1, characterized in that: In step (2), nitrogen is filled to deoxygenate, and the filling time is 20 minutes.

10. A method for improving the denitrification performance of free anaerobic ammonium oxidizing bacteria according to claim 1, characterized in that: The shaking parameters in step (2) are: 35°C, 160 rpm.

11. A method for improving the denitrification performance of free anaerobic ammonium oxidizing bacteria according to claim 1, characterized in that: During the continuous culture process in step (2), 50% of the volume of the culture medium was replaced every other week in the anaerobic glove box, and the substrate concentration was supplemented to be equal to the initial concentration.

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