A method and application for synchronous carbon sequestration and denitrification

Abstract

The present invention provides a method and application for synchronous carbon sequestration and denitrification, belonging to the technical field of solid waste resource utilization. The present invention provides a method for synchronous carbon sequestration and denitrification, comprising: enriching the microbial population mainly composed of hydrogen-oxidizing bacteria in soil; enriching the microbial population mainly composed of sulfur-oxidizing bacteria in soil; enriching the microbial population mainly composed of methanotrophs in soil; after mixing and inoculating the methanotroph-enriched flora, sulfur-oxidizing bacteria-enriched flora and hydrogen-oxidizing bacteria-enriched flora into a domestication medium for domestication culture, inoculating them into a domestication medium containing biogas slurry for synchronous carbon sequestration and denitrification. The method of the present invention enriches and screens a composite microbial metabolic group in which multiple microbial populations mainly composed of methanotrophic bacteria coexist to form a synergistic metabolic system. The synergistic metabolic system has H2S tolerance and can metabolize methane, carbon dioxide in biogas and N, P nutrients in biogas slurry to achieve the purpose of synchronous carbon sequestration and denitrification.

Description

Technical Field

[0001] The present invention belongs to the technical field of solid waste resource utilization, and particularly relates to a method and application for synchronous carbon sequestration and denitrification. Background Art

[0002] At present, anaerobic digestion is one of the most important methods for organic waste treatment. Anaerobic digestion is a process in which microorganisms decompose organic substances to produce biogas under anaerobic conditions. The main components of biogas are methane (CH4) and carbon dioxide (CO2), where methane accounts for about 50% - 75%, carbon dioxide accounts for about 25% - 50%, and the rest are small amounts of other gases such as hydrogen (H2), hydrogen sulfide (H2S), and trace amounts of nitrogen and oxygen. As a renewable bioenergy, the efficient utilization of biogas can replace the use of some fossil fuels and is beneficial to the reduction of greenhouse gas emissions. However, since biogas contains a large amount of carbon dioxide, trace amounts of hydrogen sulfide, and other gas impurities, it needs to be purified before meeting the transmission requirements of the gas pipeline network. Traditional biogas utilization methods, such as power generation and fuel substitution, emit a large amount of carbon dioxide again and have limited profitability. In addition, the biogas slurry generated by anaerobic digestion is usually discharged as sewage after being degraded by drugs. Some biogas slurries from the anaerobic digestion of livestock and poultry manure contain a large amount of ammonia nitrogen that has not been effectively degraded or utilized. Appropriately recovering ammonia nitrogen can effectively reduce ammonia volatilization and nitrogen oxide emissions and avoid pollution to the atmospheric environment. In addition, the ammonia nitrogen in biogas slurry can also be converted into other high-value-added chemicals, such as ammonium bicarbonate, through technical treatment for use as industrial or agricultural nitrogen fertilizers. Therefore, it is urgent to explore an efficient comprehensive utilization method for biogas and biogas slurry.

[0003] Producing microbial protein (MP) using anaerobic digestion products will be a promising approach to solve the above problems. Microbial protein refers to foods and feeds with high protein content produced by microorganisms. Since most microorganisms have strong adaptability to the environment (such as biogas and digestate), they can purify pollutants while producing MP, which provides an emerging way for waste recycling and bio-carbon capture. Compared with traditional protein production models (such as soybean cultivation), MP production requires less land resources and is more beneficial to human health and the ecosystem. In addition, the protein content of MP is usually between 30% and 70%, depending on the microbial species and environment. This will alleviate the global food crisis caused by overpopulation and meet the demand for high-protein consumption in some regions. Methane and carbon dioxide in biogas can be converted into MP by microorganisms, which is a carbon sequestration process that can effectively avoid carbon dioxide emissions. Microorganisms can effectively utilize the remaining nutrients in the digestate and purify them to meet the emission requirements. In recent years, the production of MP by bacteria has been gradually reported widely. Due to its rapid growth rate and rich protein content in its chemical structure, it has the ability to produce a large amount of MP. Methanotrophic bacteria such as hydrogen-oxidizing bacteria or methanotrophs may be suitable bacteria, which can not only fill the protein gap but also utilize methane and carbon dioxide to solve the solid waste problem caused by population growth. Both bacteria require an aerobic environment for metabolism. Hydrogen-oxidizing bacteria usually require hydrogen and carbon dioxide as the initial fermentation raw materials, while methanotrophs use methane as the main carbon source. Both of them also require a certain amount of nitrogen source to produce microbial protein. Research shows that the protein content of methanotrophic bacteria is higher than that of fungi and yeasts, and the digestibility of their cell walls is also better than that of algae.

[0004] However, reports show that H2S in biogas will significantly inhibit the growth of methanotrophic bacteria. H2S is a weak acid that can easily penetrate the membrane and inhibit various processes, such as inhibiting aerobic respiration by binding to cytochrome C oxidase. How to efficiently utilize methanotrophic bacteria to metabolize methane, carbon dioxide, and N, P nutrients in biogas or digestate to achieve the purpose of synchronous carbon sequestration and nitrogen removal still faces great challenges. Summary of the Invention

[0005] Aiming at the defects in the prior art, the purpose of the present invention is to provide a method for synchronous carbon sequestration and nitrogen removal, which can efficiently utilize methanotrophic bacteria to metabolize methane, carbon dioxide, and N, P nutrients in biogas or digestate to achieve the purpose of synchronous carbon sequestration and nitrogen removal.

[0006] The purpose of the present invention is achieved by the following technical solutions:

[0007] The present invention provides a method for synchronous carbon sequestration and nitrogen removal, comprising the following steps:

[0008] Enrich a microbial population mainly composed of hydrogen-oxidizing bacteria in the soil through a hydrogen-oxidizing bacteria mixture gas to obtain a hydrogen-oxidizing bacteria enriched flora;

[0009] Enrich a microbial population mainly composed of sulfur-oxidizing bacteria in the soil through a sulfur-oxidizing bacteria mixture gas to obtain a sulfur-oxidizing bacteria enriched flora;

[0010] Enrich a microbial population mainly composed of methane-oxidizing bacteria in the soil through a methane-oxidizing bacteria mixture gas to obtain a methane-oxidizing bacteria enriched flora;

[0011] Mix and inoculate the methane-oxidizing bacteria enriched flora, the sulfur-oxidizing bacteria enriched flora, and the hydrogen-oxidizing bacteria enriched flora into a domestication medium for domestication culture to obtain a composite microbial metabolism group;

[0012] Inoculate the composite microbial metabolism group into a domestication medium containing biogas slurry and introduce biogas for synchronous carbon fixation and denitrification;

[0013] The hydrogen-oxidizing bacteria mixture gas includes H2, O2, and CO2, and the volume ratio of H2, O2, and CO2 is (50 - 70):(30 - 20):(20 - 10);

[0014] The sulfur-oxidizing bacteria mixture gas includes H2, O2, CO2, and H2S, and the volume ratio of H2, O2, CO2, and H2S is (30 - 50):(20 - 30):(20 - 30):(20 - 30);

[0015] The methane-oxidizing bacteria mixture gas includes CH4 and O2, and the volume ratio of CH4 and O2 is (50 - 80):(50 - 20).

[0016] Preferably, the soil for enriching hydrogen-oxidizing bacteria includes the soil 2 - 10 cm below the surface layer of paddy fields storing water and / or the residue of livestock and poultry manure compost;

[0017] The soil for enriching sulfur-oxidizing bacteria includes the soil 2 - 10 cm below the surface layer of swamps and / or the sludge of oxidation ponds in sewage treatment plants;

[0018] The soil for enriching methane-oxidizing bacteria includes the soil 2 - 10 cm below the surface layer of paddy fields storing water and / or the sludge of oxidation ponds in sewage treatment plants.

[0019] Preferably, the method for enriching a microbial population mainly composed of hydrogen-oxidizing bacteria in the soil includes:

[0020] Inoculate the microorganisms in the soil into a primary hydrogen-oxidizing bacteria seed medium and introduce the hydrogen-oxidizing bacteria mixture gas for culture until more than 50% of H2 is consumed or the CO2 concentration no longer decreases to obtain a primary hydrogen-oxidizing bacteria seed liquid;

[0021] Inoculate the primary seed solution of the hydrogen-oxidizing bacteria into the secondary seed medium of the hydrogen-oxidizing bacteria, and introduce a hydrogen-oxidizing bacteria mixed gas for culturing to obtain a hydrogen-oxidizing bacteria enriched flora;

[0022] The composition of the primary seed medium of the hydrogen-oxidizing bacteria includes: Na2HPO4: 2200 - 2800 mg / L, KH2PO4: 500 - 1500 mg / L, (NH4)2SO4: 450 - 500 mg / L, MgCl2: 45 - 60 mg / L, CaCl2·2H2O: 40 - 50 mg / L, disodium EDTA: 0.2 - 0.7 mg / L, FeSO4·7H2O: 0.1 - 0.5 mg / L, ZnSO4·7H2O: 0.01 - 0.02 mg / L, MnCl2·4H2O: 0.002 - 0.003 mg / L, H3BO3: 0.01 - 0.04 mg / L, CoCl2·6H2O: 0.01 - 0.02 mg / L, CuCl2·2H2O: 0.001 - 0.01 mg / L, NiCl2·6H2O: 0.002 - 0.004 mg / L, Na2MoO4·2H2O: 0.001 - 0.003 mg / L;

[0023] The composition of the secondary seed medium of the hydrogen-oxidizing bacteria further includes, on the basis of the primary seed medium of the hydrogen-oxidizing bacteria: 1% - 3% glucose, 0.6% - 2% yeast powder, and 1.5% - 3.2% casein peptone.

[0024] Preferably, the method for enriching the microbial population mainly composed of sulfur-oxidizing bacteria in the soil includes:

[0025] Inoculate the microorganisms in the soil into the primary seed medium of the sulfur-oxidizing bacteria, and introduce a sulfur-oxidizing bacteria mixed gas for culturing until the H2S concentration no longer decreases to obtain the primary seed solution of the sulfur-oxidizing bacteria;

[0026] Inoculate the primary seed solution of the sulfur-oxidizing bacteria into the secondary seed medium of the sulfur-oxidizing bacteria, and introduce a sulfur-oxidizing bacteria mixed gas for culturing to obtain a sulfur-oxidizing bacteria enriched flora;

[0027] The composition of the primary seed medium for sulfur-oxidizing bacteria includes: phenol red: 0.001 - 0.003 g / L, 1,4-piperazinediethanesulfonic acid: 4.5 - 8.5 g / L, sodium chloride: 10 - 25.000 g / L, magnesium sulfate heptahydrate: 1.1 - 2.9 g / L, potassium chloride: 0.1 - 1 g / L, ammonium chloride: 0.015 - 0.3 g / L, sodium thiosulfate pentahydrate: 1.5 - 3.2 g / L, calcium chloride dihydrate: 0.05 - 0.2 g / L, magnesium chloride hexahydrate: 2.5 - 4.5 g / L, dipotassium hydrogen phosphate: 0.05 - 0.15 g / L, ammonium ferrous sulfate hexahydrate: 0.001 - 0.002 g / L, trace element solution: 1 mL / L, and vitamin solution: 10 mL / L;

[0028] On the basis of the primary seed medium for sulfur-oxidizing bacteria, the composition of the secondary seed medium for sulfur-oxidizing bacteria further includes: glucose 1% - 3%, yeast powder 0.6% - 2%, and casein peptone 1.5% - 3.2%.

[0029] Preferably, the method for enriching the microbial population mainly composed of methanotrophs in soil includes:

[0030] Inoculating the microorganisms in the soil into the primary seed medium for methanotrophs and culturing by introducing a methanotroph mixed gas until the CH4 concentration no longer decreases to obtain the primary seed liquid of methanotrophs;

[0031] Inoculating the primary seed liquid of methanotrophs into the secondary seed medium for methanotrophs and culturing by introducing a methanotroph mixed gas to obtain an enriched methanotroph flora;

[0032] The composition of the primary seed medium for methanotrophs includes: KNO3: 100 - 200 mg / L, KH2PO4: 100 - 200 mg / L, MgSO4·7H2O: 25 - 55 mg / L, CaCl2·2H2O: 5 - 20 mg / L, EDTA: 2 - 5 mg / L, CuCl2·5H2O: 0.05 - 0.1 mg / L, FeSO4·7H2O: 1 - 2 mg / L, ZnSO4·7H2O: 0.1 - 0.2 mg / L, NiCl2·6H2O: 0.008 - 0.02 mg / L, CoCl2·6H2O: 0.1 - 0.2 mg / L, and Na2MoO4: 0.01 - 0.03 mg / L;

[0033] On the basis of the primary seed medium for methanotrophs, the composition of the secondary seed medium for methanotrophs further includes: glucose 1% - 3%, yeast powder 0.6% - 2%, and casein peptone 1.5% - 3.2%.

[0034] Preferably, a mixed gas is introduced during the acclimation culture; the composition of the mixed gas is H2, O2, CO2, CH4, and H2S; the volume ratio of H2, O2, CO2, CH4, and H2S is (0.1~1):(10~20):(15~25):(50~60):(0.1~0.3).

[0035] Preferably, the composition of every 1 L of the acclimation culture medium includes:

[0036] 0.5~1.00 g of MgSO4·7H2O, 1.00~2.5 g of KNO3, 0.15~0.3 g of KH2PO4, 0.2~0.45 g of Na2HPO4, 0.1~0.15 g of CaCl2·2H2O, and 1 mL of trace element solution.

[0037] Preferably, the acclimation culture is continuously subcultured 3~4 times; Na2S and ammonium chloride are also included in the acclimation culture medium; during the first acclimation culture, the mass concentration of Na2S in the acclimation culture medium is 1~2 mg / L, and the mass concentration of ammonium chloride is 100~300 mg / L; during each subculture, the mass concentrations of Na2S and ammonium chloride in the acclimation culture medium are 1.5~2 times the mass concentrations of Na2S and ammonium chloride during the previous culture.

[0038] The present invention provides the application of the method according to the above technical solution in the resource utilization of biogas slurry and / or biogas.

[0039] The present invention provides the application of the method according to the above technical solution in the preparation of protein.

[0040] The beneficial effects of the present invention:

[0041] The present invention provides a method for synchronous carbon sequestration and nitrogen removal, comprising the following steps: enriching the microbial population mainly composed of hydrogen-oxidizing bacteria in soil through a hydrogen-oxidizing bacteria gas mixture to obtain a hydrogen-oxidizing bacteria enriched flora; enriching the microbial population mainly composed of sulfur-oxidizing bacteria in soil through a sulfur-oxidizing bacteria gas mixture to obtain a sulfur-oxidizing bacteria enriched flora; enriching the microbial population mainly composed of methane-oxidizing bacteria in soil through a methane-oxidizing bacteria gas mixture to obtain a methane-oxidizing bacteria enriched flora; mixing and inoculating the methane-oxidizing bacteria enriched flora, the sulfur-oxidizing bacteria enriched flora and the hydrogen-oxidizing bacteria enriched flora into a domestication medium for domestication culture to obtain a composite microbial metabolic group; inoculating the composite microbial metabolic group into a domestication medium containing biogas slurry and introducing biogas for synchronous carbon sequestration and nitrogen removal; the hydrogen-oxidizing bacteria gas mixture comprises H2, O2 and CO2, and the volume ratio of H2, O2 and CO2 is (50-70):(30-20):(20-10); the sulfur-oxidizing bacteria gas mixture comprises H2, O2, CO2 and H2S, and the volume ratio of H2, O2, CO2 and H2S is (30-50):(20-30):(20-30):(20-30); the methane-oxidizing bacteria gas mixture comprises CH4 and O2, and the volume ratio of CH4 and O2 is (50-80):(50-20). The method of the present invention enriches and screens a composite microbial metabolic group in which multiple microbial populations mainly composed of methane-oxidizing bacteria coexist to form a synergistic metabolic system. The synergistic metabolic system has H2S tolerance and can metabolize methane, carbon dioxide in biogas and N, P nutrients in biogas slurry to achieve the purpose of synchronous carbon sequestration and nitrogen removal. BRIEF DESCRIPTION OF THE DRAWINGS

[0042] In order to more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the following will briefly introduce the drawings required in the embodiments. Obviously, the drawings in the following description are only some embodiments of the present invention, and those of ordinary skill in the art can also obtain other drawings without creative efforts.

[0043] Figure 1 It is a diagram showing the growth of bacteria in the third round of culture in step S3 in Example 1;

[0044] Figure 2 It is a diagram showing the test results of the dry cell weight and protein content in step S4 in Example 1. DETAILED DESCRIPTION OF THE EMBODIMENTS

[0045] The present invention provides a method for synchronous carbon sequestration and nitrogen removal, comprising the following steps:

[0046] Enriching the microbial population mainly composed of hydrogen-oxidizing bacteria in soil through a hydrogen-oxidizing bacteria gas mixture to obtain a hydrogen-oxidizing bacteria enriched flora;

[0047] Enrich the microbial population mainly composed of sulfur-oxidizing bacteria in the soil through a sulfur-oxidizing bacteria mixed gas to obtain a sulfur-oxidizing bacteria enriched flora;

[0048] Enrich the microbial population mainly composed of methane-oxidizing bacteria in the soil through a methane-oxidizing bacteria mixed gas to obtain a methane-oxidizing bacteria enriched flora;

[0049] Mix and inoculate the methane-oxidizing bacteria enriched flora, sulfur-oxidizing bacteria enriched flora and hydrogen-oxidizing bacteria enriched flora in a domestication medium for domestication culture to obtain a composite microbial metabolism group;

[0050] Inoculate the composite microbial metabolism group into a domestication medium containing biogas slurry and introduce biogas for synchronous carbon fixation and denitrification;

[0051] The hydrogen-oxidizing bacteria mixed gas includes H2, O2 and CO2, and the volume ratio of H2, O2 and CO2 is (50 - 70):(30 - 20):(20 - 10);

[0052] The sulfur-oxidizing bacteria mixed gas includes H2, O2, CO2 and H2S, and the volume ratio of H2, O2, CO2 and H2S is (30 - 50):(20 - 30):(20 - 30):(20 - 30);

[0053] The methane-oxidizing bacteria mixed gas includes CH4 and O2, and the volume ratio of CH4 and O2 is (50 - 80):(50 - 20).

[0054] The present invention has no special limitation on the order of enriching the microbial population mainly composed of hydrogen-oxidizing bacteria in the soil, enriching the microbial population mainly composed of sulfur-oxidizing bacteria in the soil, and enriching the microbial population mainly composed of methane-oxidizing bacteria in the soil. It can first enrich the microbial population mainly composed of hydrogen-oxidizing bacteria, or first enrich the microbial population mainly composed of sulfur-oxidizing bacteria, or first enrich the microbial population mainly composed of methane-oxidizing bacteria.

[0055] The limiting terms for the mixed gas, medium or culture in the following technical solutions of the present invention do not limit the corresponding mixed gas, medium or culture, but are only used to distinguish different mixed gases, different media and different cultures.

[0056] The present invention enriches the microbial population mainly composed of hydrogen-oxidizing bacteria in soil to obtain a hydrogen-oxidizing bacteria enriched flora. In the present invention, the method for enriching the microbial population mainly composed of hydrogen-oxidizing bacteria in soil includes: inoculating the microorganisms in the soil into a primary seed culture medium and introducing a mixed gas for culturing until more than 50% of the H2 is consumed or the CO2 concentration no longer decreases, thereby obtaining a primary seed liquid of hydrogen-oxidizing bacteria. In the present invention, in order to distinguish the primary seed culture medium from other primary seed culture media in the following text, it is called the first primary seed culture medium or the primary seed culture medium of hydrogen-oxidizing bacteria; in order to distinguish the mixed gas from other mixed gases in the following text, it is called the first mixed gas of hydrogen-oxidizing bacteria; and in order to distinguish the culturing, it is called the first culturing.

[0057] In the present invention, the composition of the first primary seed culture medium includes: Na2HPO4: 2200 - 2800 mg / L, KH2PO4: 500 - 1500 mg / L, (NH4)2SO4: 450 - 500 mg / L, MgCl2: 45 - 60 mg / L, CaCl2·2H2O: 40 - 50 mg / L, disodium EDTA: 0.2 - 0.7 mg / L, FeSO4·7H2O: 0.1 - 0.5 mg / L, ZnSO4·7H2O: 0.01 - 0.02 mg / L, MnCl2·4H2O: 0.002 - 0.003 mg / L, H3BO3: 0.01 - 0.04 mg / L, CoCl2·6H2O: 0.01 - 0.02 mg / L, CuCl2·2H2O: 0.001 - 0.01 mg / L, NiCl2·6H2O: 0.002 - 0.004 mg / L, Na2MoO4·2H2O: 0.001 - 0.003 mg / L. As an optional embodiment of the present invention, the composition of the first primary seed culture medium is: Na2HPO4: 2800 mg / L, KH2PO4: 1000 mg / L, (NH4)2SO4: 500 mg / L, MgCl2: 53 mg / L, CaCl2·2H2O: 50 mg / L, disodium EDTA: 0.5 mg / L, FeSO4·7H2O: 0.2 mg / L, ZnSO4·7H2O: 0.01 mg / L, MnCl2·4H2O: 0.003 mg / L, H3BO3: 0.03 mg / L, CoCl2·6H2O: 0.02 mg / L, CuCl2·2H2O: 0.001 mg / L, NiCl2·6H2O: 0.002 mg / L, Na2MoO4·2H2O: 0.003 mg / L. In the present invention, the pH of the first primary seed culture medium can be 6.8 - 7.4. In the present invention, the pH is preferably adjusted using a phosphate buffer solution. The first primary seed culture medium of the present invention provides an optimal culture medium according to the metabolic conditions of hydrogen-oxidizing bacteria.

[0058] In the present invention, the hydrogen-oxidizing bacteria mixed gas includes H2, O2 and CO2; the volume ratio of H2, O2 and CO2 is (50-70):(30-20):(20-10). As an optional implementation mode of the present invention, the volume ratio of H2, O2 and CO2 can be 50:30:20, 50:20:10, 50:25:15, 60:30:20, 60:20:10, 60:25:15, 70:30:20, 70:20:10 or 70:25:15. In the present invention, the hydrogen-oxidizing bacteria mixed gas can also be referred to as the first hydrogen-oxidizing bacteria mixed gas. In the present invention, the first hydrogen-oxidizing bacteria mixed gas can meet the metabolic growth needs of hydrogen-oxidizing bacteria, which helps to enrich hydrogen-oxidizing bacteria. Among them, hydrogen-oxidizing bacteria use CO2 as a carbon source, H2 as an electron donor, and O2 as an electron acceptor to carry out metabolic cycles. The composition ratio setting of the first hydrogen-oxidizing bacteria mixed gas provided by the present invention can significantly shorten the enrichment time of hydrogen-oxidizing bacteria. If the gas ratio of the first mixed gas is changed, for example, increasing the volume of CO2 or increasing the proportion of H2, the enrichment time of hydrogen-oxidizing bacteria will be significantly prolonged.

[0059] After obtaining the primary hydrogen-oxidizing bacteria seed liquid, the present invention inoculates the primary hydrogen-oxidizing bacteria seed liquid into a secondary seed culture medium and introduces the first hydrogen-oxidizing bacteria mixed gas for cultivation to obtain an enriched hydrogen-oxidizing bacteria flora. In the present invention, the cultivation includes secondary cultivation and sub-cultivation. In the present invention, the secondary seed culture medium is called the first secondary seed culture medium or the hydrogen-oxidizing bacteria secondary culture medium in order to distinguish it from other secondary seed culture media in the following text.

[0060] The composition of the first secondary seed culture medium is based on the first primary seed culture medium and further includes: 1%-3% glucose, 0.6%-2% yeast powder and 1.5%-3.2% casein peptone. In the present invention, the pH of the first secondary seed culture medium can be 6.8-7.4.

[0061] In the present invention, different hydrogen-oxidizing bacteria may use only CO2 as a carbon source, or may also use organic substances in the culture medium as a carbon source. The purpose of adding is to provide different forms of carbon sources and nutrients for microorganisms as much as possible to create a suitable culture environment as much as possible.

[0062] In the present invention, the soil includes the soil 2 - 10 cm below the surface layer of the paddy field for water storage and / or the residue of livestock and poultry manure compost. Through research, it is found in the present invention that the soil 2 - 10 cm below the surface layer of the paddy field for water storage and the residue of livestock and poultry manure compost are rich in hydrogen-oxidizing bacteria. After obtaining the corresponding soil, it is preferred to mix the soil with water to obtain a soil-water mixture in the present invention. In the present invention, the mass-volume ratio of the soil to water can be (5 - 10) g : (100 - 120) mL. As an optional implementation manner of the present invention, the mass-volume ratio of the soil to water can be 5 g:100 mL, 10 g:100 mL, 5 g:120 mL, 10 g:100 mL, 8 g:100 mL, 8 g:120 mL, 5 g:110 mL, 8 g:110 mL or 8 g:120 mL. After obtaining the soil-water mixture, it is preferred to oscillate the soil-water mixture on a shaker at 30 °C and 120 rpm for 1 - 2 h in the present invention. After the oscillation is completed, it is preferred to mix the oscillated soil-water mixture with the first primary seed medium to complete inoculation; the volume ratio of the soil-water mixture to the first primary seed medium is 1 : (1 - 2). After the inoculation is completed, a soil-medium mixed system is obtained in the present invention. After obtaining the soil-medium mixed system, it is preferred to stir the soil-medium mixed system with a magnetic stirrer at 250 - 450 rpm for 1 - 2 h in the present invention. After the stirring is completed, the mixed soil-medium mixed system is placed in a culture container; the volume ratio of the soil-medium mixed system to the culture container is 1 : (5 - 10), and it can also be 1:5, 1.1:5, 1:6, 1:7, 1:8, 1:9 or 1:10. After placing the soil-medium mixed system in the culture container, a first mixed gas is introduced in the present invention. The first mixed gas of hydrogen-oxidizing bacteria is introduced until the culture container is filled in the present invention. Before introducing the first mixed gas of hydrogen-oxidizing bacteria, it is preferred to first introduce nitrogen to evacuate the original air. After evacuating the original air, the first mixed gas of hydrogen-oxidizing bacteria is then introduced. After introducing the first mixed gas, the first cultivation is carried out in the present invention. In the present invention, the temperature of the first cultivation is 25 - 35 °C. As an optional implementation manner of the present invention, the temperature of the first cultivation can be 25, 26, 27, 28, 29, 30, 31, 32, 33, 34 or 35 °C. In the present invention, the rotation speed of the first cultivation can be 150 - 210 rpm. As an optional implementation manner of the present invention, the rotation speed can be 150, 160, 170, 180, 190, 200 or 210 rpm. The first cultivation in the present invention is preferably carried out until more than 50% of H2 is consumed or the CO2 concentration no longer decreases to obtain a primary seed solution of hydrogen-oxidizing bacteria.

[0063] After obtaining the primary seed solution of the hydrogen-oxidizing bacterium, the present invention inoculates the primary seed solution of the hydrogen-oxidizing bacterium into a first secondary seed culture medium and introduces a first mixed gas of hydrogen-oxidizing bacterium for cultivation to obtain an enriched flora of hydrogen-oxidizing bacterium. The cultivation of the present invention includes secondary cultivation and sub-cultivation.

[0064] When the present invention inoculates the primary seed solution of the hydrogen-oxidizing bacterium into the first secondary seed culture medium, it is preferred to inoculate 5-10 mL of the primary seed solution of the hydrogen-oxidizing bacterium into every 100-200 mL of the first secondary seed culture medium to obtain a secondary culture system. As an optional implementation mode of the present invention, the volume ratio of the secondary culture system to its culture container can be 1:(5-10), or can also be 1:5, 1.1:5, 1:6, 1:7, 1:8, 1:9 or 1:10. After the present invention inoculates the primary seed solution of the hydrogen-oxidizing bacterium into the first secondary seed culture medium, it is preferred to first introduce nitrogen to evacuate the original air. After evacuating the original air, then introduce the first mixed gas of hydrogen-oxidizing bacterium until the culture container is full. In the present invention, the temperature of the secondary cultivation is 25-35 °C. As an optional implementation mode of the present invention, the temperature of the secondary cultivation can be 25, 26, 27, 28, 29, 30, 31, 32, 33, 34 or 35 °C. In the present invention, the rotation speed of the secondary cultivation can be 150-210 rpm. As an optional implementation mode of the present invention, the rotation speed can be 150, 160, 170, 180, 190, 200 or 210 rpm. During the secondary cultivation process of the present invention, it is preferred to cultivate until more than 50% of H2 is consumed or the CO2 concentration no longer decreases to obtain a secondary culture seed solution. As an optional implementation mode of the present invention, the time of the secondary cultivation is shorter than the time of the primary cultivation.

[0065] After the second cultivation is completed, the present invention preferably further includes inoculating the second cultivation seed liquid into a first secondary seed culture medium for sub-cultivation. The main purpose of the sub-cultivation in the present invention is to enrich the microbial community, and at the same time enhance and stabilize its carbon fixation ability. When performing sub-cultivation in the present invention, preferably 5-10 mL of the second cultivation seed liquid is inoculated into every 100-200 mL of the first secondary seed culture medium to obtain a sub-cultivation system. As an alternative embodiment of the present invention, the volume ratio of the sub-cultivation system to its culture container can be 1:(5-10), or can also be 1:5, 1.1:5, 1:6, 1:7, 1:8, 1:9 or 1:10. During the sub-cultivation process of the present invention, preferably the secondary seed culture medium and the first mixed gas of hydrogen-oxidizing bacteria are replaced once every 2-10 days, and the interval time can be 2, 3, 4, 5, 6, 7, 8, 9 or 10 days. When performing sub-cultivation in the present invention, preferably the culture medium and the mixed gas are continuously replaced 5-10 times, which can be 5, 6, 7, 8, 9 or 10 times; the present invention preferably replaces the mixed gas while replacing the culture medium, and when replacing the culture medium each time, the volume ratio of the culture system to the culture container remains unchanged. During the sub-cultivation process of the present invention, after continuously replacing the first mixed gas of hydrogen-oxidizing bacteria 5-10 times, in the last cultivation, it is preferably cultivated until more than 50% of the H2 is consumed or the CO2 concentration no longer decreases, to obtain a hydrogen-oxidizing bacteria enriched microbial community.

[0066] The hydrogen-oxidizing bacteria enriched microbial community obtained by the present invention through the above method can efficiently utilize hydrogen and carbon dioxide: Hydrogen-oxidizing bacteria can use hydrogen (H2) as an electron donor and carbon dioxide (CO2) as a carbon source to generate organic substances through photosynthesis or chemosynthesis. The hydrogen-oxidizing bacteria enriched by the specific culture medium and gas conditions in the present invention have high hydrogen and carbon dioxide utilization abilities and can effectively convert these gases into biomass. The hydrogen-oxidizing bacteria enriched microbial community can improve the carbon fixation efficiency: During the process of hydrogen-oxidizing bacteria metabolizing using hydrogen and carbon dioxide, a large amount of carbon can be fixed as biomass to generate useful substances such as microbial protein. This helps to improve the carbon fixation efficiency of the entire system, reduce carbon dioxide emissions, and reduce the greenhouse effect. The hydrogen-oxidizing bacteria enriched microbial community can enhance the system stability: Hydrogen-oxidizing bacteria play an important role in the complex microbial metabolic community and can cooperate with other microbial populations to enhance the stability and anti-interference ability of the entire system. For example, during the synchronous carbon fixation and denitrification process, hydrogen-oxidizing bacteria can act together with methane-oxidizing bacteria and sulfur-oxidizing bacteria to improve the tolerance and adaptability of the system. The hydrogen-oxidizing bacteria enriched microbial community can promote the recycling of resources: Hydrogen-oxidizing bacteria can utilize hydrogen and carbon dioxide in biogas to achieve the recycling of resources, which not only reduces the dependence on fossil energy but also improves the resource utilization efficiency, in line with the development concept of circular economy.

[0067] The present invention enriches the microbial population mainly composed of sulfur-oxidizing bacteria in soil. The method for obtaining the sulfur-oxidizing bacteria enriched flora includes: inoculating the microorganisms in the soil into the primary seed medium for sulfur-oxidizing bacteria and introducing the first mixed gas for sulfur-oxidizing bacteria for the first culture until the concentration of H2S no longer decreases, thereby obtaining the primary seed liquid of sulfur-oxidizing bacteria. In the present invention, the primary seed medium for sulfur-oxidizing bacteria can also be referred to as the second primary seed medium.

[0068] In the present invention, the composition of the second primary seed medium includes: phenol red: 0.001 - 0.003 g / L, 1,4-piperazinediethanesulfonic acid: 4.5 - 8.5 g / L, sodium chloride: 10 - 25.000 g / L, magnesium sulfate heptahydrate: 1.1 - 2.9 g / L, potassium chloride: 0.1 - 1 g / L, ammonium chloride: 0.015 - 0.3 g / L, sodium thiosulfate pentahydrate: 1.5 - 3.2 g / L, calcium chloride dihydrate: 0.05 - 0.2 g / L, magnesium chloride hexahydrate: 2.5 - 4.5 g / L, dipotassium hydrogen phosphate: 0.05 - 0.15 g / L, ammonium iron(II) sulfate hexahydrate: 0.001 - 0.002 g / L, trace element solution: 1 mL / L, and vitamin solution: 10 mL / L. The pH of the second primary seed medium can be 6.8 - 7.4. In the present invention, the pH is preferably adjusted using a phosphate buffer solution. As an optional implementation manner of the present invention, the composition of the second primary seed medium is: phenol red: 0.003 g / L, 1,4-piperazinediethanesulfonic acid: 6.500 g / L, sodium chloride: 25.000 g / L, magnesium sulfate heptahydrate: 2.700 g / L, potassium chloride: 0.500 g / L, ammonium chloride: 0.250 g / L, sodium thiosulfate pentahydrate: 2.480 g / L, calcium chloride dihydrate: 0.140 g / L, magnesium chloride hexahydrate: 4.300 g / L, dipotassium hydrogen phosphate: 0.140 g / L, ammonium iron(II) sulfate hexahydrate: 0.002 g / L, trace element solution: 1 mL / L, and vitamin solution: 10 mL / L.

[0069] In the present invention, each liter of the trace element solution comprises: 1.00 - 3.00 g of MgSO4·7H2O, 0.1 - 1 g of MnSO4·H2O, 0.5 - 1.00 g of NaCl, 0.10 - 0.20 g of FeSO4·7H2O, 0.15 - 0.2 g of CoSO4·7H2O, 0.10 - 0.2 g of CaCl2·2H2O, 0.15 - 0.18 g of ZnSO4·7H2O, 0.01 - 0.02 g of CuSO4·5H2O, 0.01 - 0.02 g of AlK(SO4)2·12H2O, 0.01 - 0.02 g of H3BO3, 0.01 - 0.05 g of Na2MoO4·2H2O, 0.01 - 0.05 g of NiCl2·6H2O, 0.01 - 0.50 mg of Na2SeO3·5H2O, and 0.02 - 0.40 mg of Na2WO4·2H2O. As an alternative embodiment of the present invention, the composition of each liter of the trace element solution is: 2.00 g of MgSO4·7H2O, 0.5 g of MnSO4·H2O, 0.8 g of NaCl, 0.15 g of FeSO4·7H2O, 0.18 g of CoSO4·7H2O, 0.15 g of CaCl2·2H2O, 0.16 g of ZnSO4·7H2O, 0.015 g of CuSO4·5H2O, 0.015 g of AlK(SO4)2·12H2O, 0.015 g of H3BO3, 0.03 g of Na2MoO4·2H2O, 0.03 g of NiCl2·6H2O, 0.2 mg of Na2SeO3·5H2O, and 0.2 mg of Na2WO4·2H2O.

[0070] In the present invention, the vitamin solution comprises: 1 - 2 mg / L of vitamin B7, 1 - 2 mg / L of folic acid, 5 - 10 mg / L of pyridoxine hydrochloride, 2.5 - 5.5 mg / L of vitamin B12, 2 - 5 mg / L of riboflavin, 1.5 - 6 mg / L of vitamin B3, 2.5 - 5.5 mg / L of vitamin B5, vitamin B 12 0.1 - 0.2 mg / L, 2 - 5 mg / L of p-aminobenzoic acid, and 2 - 5 mg / L of (DL)-α-lipoic acid. As an alternative embodiment of the present invention, the composition of the vitamin solution is: 1.5 mg / L of vitamin B7, 1.5 mg / L of folic acid, 8 mg / L of pyridoxine hydrochloride, 4 mg / L of vitamin B1, 4 mg / L of riboflavin, 4 mg / L of vitamin B3, 4 mg / L of vitamin B5, vitamin B 12 0.15 mg / L, 4 mg / L of p-aminobenzoic acid, and 4 mg / L of (DL)-α-lipoic acid. In the present invention, the second primary seed culture medium provides the optimal culture medium according to the metabolic conditions of sulfur-oxidizing bacteria.

[0071] The sulfur-oxidizing bacteria mixed gas includes H2, O2, CO2, and H2S; the volume ratio of H2, O2, CO2, and H2S is (30-50):(20-30):(20-30):(20-30). As an alternative embodiment of the present invention, the volume ratio of H2, O2, CO2, and H2S can be 30:20:20:20, 30:30:30:30, 50:20:20:20, 50:30:30:30, 30:20:30:20, 30:20:30:30, 50:20:30:20, or 50:20:30:30. In the present invention, the sulfur-oxidizing bacteria mixed gas can also be referred to as the first sulfur-oxidizing bacteria mixed gas. The composition ratio setting of the first sulfur-oxidizing bacteria mixed gas provided by the present invention can significantly shorten the enrichment time of sulfur-oxidizing bacteria. If the gas ratio of the first mixed gas is changed, the enrichment time of sulfur-oxidizing bacteria will be significantly prolonged.

[0072] After obtaining the primary sulfur-oxidizing bacteria seed liquid, the present invention inoculates the primary sulfur-oxidizing bacteria seed liquid into a secondary sulfur-oxidizing bacteria seed medium and introduces the first sulfur-oxidizing bacteria mixed gas for cultivation to obtain an enriched sulfur-oxidizing bacteria flora. In the present invention, the secondary sulfur-oxidizing bacteria seed medium can be referred to as the second secondary seed medium. In the present invention, the cultivation includes secondary cultivation and sub-cultivation.

[0073] In the present invention, the composition of the second secondary seed medium is based on the second primary seed medium and further includes: 1%-3% glucose, 0.6%-2% yeast powder, and 1.5%-3.2% casein peptone. The pH of the second secondary seed medium can be 6.8-7.4.

[0074] In the process of enriching the microbial population mainly composed of sulfur-oxidizing bacteria in the present invention, the sulfur-oxidizing bacteria use H2S as an electron donor, O2 as an electron acceptor, and CO2 as a carbon source for metabolic cycling. The components in the medium provide necessary nutrients and a buffer system to maintain a stable pH, which is conducive to the growth of sulfur-oxidizing bacteria. During the enrichment process of sulfur-oxidizing bacteria, a relatively high H2S ratio (20%-30%) ensures that sulfur-oxidizing bacteria have sufficient electron donors for metabolism, and the ratios of H2, O2, and CO2 meet their respiration and carbon fixation requirements. When enriching the microbial population mainly composed of sulfur-oxidizing bacteria in the present invention, the pH value of the medium is adjusted between 6.8 and 7.4, which is suitable for the growth of sulfur-oxidizing bacteria. The present invention creates a micro-oxic environment suitable for the growth of sulfur-oxidizing bacteria by introducing the mixed gas and exhausting the original air, which is conducive to enriching the microbial population mainly composed of sulfur-oxidizing bacteria.

[0075] In the present invention, the soil includes the soil 2 - 10 cm below the surface layer of the swamp and / or the sludge in the oxidation pond of the sewage treatment plant. Through research, it is found that the soil 2 - 10 cm below the surface layer of the swamp and / or the sludge in the oxidation pond of the sewage treatment plant is rich in sulfur-oxidizing bacteria. After obtaining the corresponding soil, preferably, the soil is mixed with water to obtain a soil-water mixture. In the present invention, the mass-volume ratio of the soil to water can be (5 - 10) g : (100 - 120) mL. As an optional implementation mode of the present invention, the mass-volume ratio of the soil to water can be 5 g:100 mL, 10 g:100 mL, 5 g:120 mL, 10 g:100 mL, 8 g:100 mL, 8 g:120 mL, 5 g:110 mL, 8 g:110 mL or 8 g:120 mL. After obtaining the soil-water mixture, preferably, the soil-water mixture is oscillated on a shaker at 30 °C and 120 rpm for 1 - 2 h. After the oscillation is completed, preferably, the oscillated soil-water mixture is mixed with the second primary seed medium to complete inoculation; the volume ratio of the soil-water mixture to the second primary seed medium is 1 : (1 - 2). After the inoculation is completed, a soil-medium mixed system is obtained. After obtaining the soil-medium mixed system, preferably, the soil-medium mixed system is stirred with a magnetic stirrer at 250 - 450 rpm for 1 - 2 h. After the stirring is completed, the mixed soil-medium mixed system is placed in a culture container; the volume ratio of the soil-medium mixed system to the culture container is 1 : (5 - 10), and can also be 1:5, 1.1:5, 1:6, 1:7, 1:8, 1:9 or 1:10. After placing the soil-medium mixed system in the culture container, a first mixed gas of sulfur-oxidizing bacteria is introduced. The first mixed gas of sulfur-oxidizing bacteria is introduced until the culture container is full. Before introducing the first mixed gas of sulfur-oxidizing bacteria, preferably, nitrogen is first introduced to evacuate the original air, and after evacuating the original air, the first mixed gas of sulfur-oxidizing bacteria is introduced. After introducing the first mixed gas, the first cultivation is carried out. In the present invention, the temperature of the first cultivation is 25 - 35 °C. As an optional implementation mode of the present invention, the temperature of the first cultivation can be 25, 26, 27, 28, 29, 30, 31, 32, 33, 34 or 35 °C. In the present invention, the rotation speed of the first cultivation can be 150 - 210 rpm. As an optional implementation mode of the present invention, the rotation speed can be 150, 160, 170, 180, 190, 200 or 210 rpm. The first cultivation in the present invention is preferably carried out until the concentration of H2S no longer decreases, and a primary seed solution of sulfur-oxidizing bacteria is obtained.

[0076] After obtaining the primary seed solution of sulfur-oxidizing bacteria, the present invention inoculates the primary seed solution of sulfur-oxidizing bacteria into a secondary seed culture medium and introduces a first mixed gas of sulfur-oxidizing bacteria for secondary culture and subculture to obtain an enriched flora of sulfur-oxidizing bacteria.

[0077] When the present invention inoculates the primary seed solution of sulfur-oxidizing bacteria into the secondary seed culture medium, it is preferred to inoculate 5-10 mL of the primary seed solution of sulfur-oxidizing bacteria into 100-200 mL of the secondary seed culture medium to obtain a secondary culture system. As an optional implementation manner of the present invention, the volume ratio of the secondary culture system to its culture container can be 1:(5-10), or can also be 1:5, 1.1:5, 1:6, 1:7, 1:8, 1:9 or 1:10. After the present invention inoculates the primary seed solution of sulfur-oxidizing bacteria into the secondary seed culture medium, it is preferred to first introduce nitrogen to exhaust the original air. After exhausting the original air, then introduce the first mixed gas of sulfur-oxidizing bacteria until the culture container is full. In the present invention, the temperature of the secondary culture is 25-35 °C. As an optional implementation manner of the present invention, the temperature of the secondary culture can be 25, 26, 27, 28, 29, 30, 31, 32, 33, 34 or 35 °C. In the present invention, the rotation speed of the secondary culture can be 150-210 rpm. As an optional implementation manner of the present invention, the rotation speed can be 150, 160, 170, 180, 190, 200 or 210 rpm. During the secondary culture process of the present invention, it is preferred to culture until the concentration of H2S no longer decreases to obtain a secondary culture seed solution. As an optional implementation manner of the present invention, the time of the secondary culture is shorter than the time of the first culture.

[0078] After the second cultivation is completed, the present invention preferably further includes inoculating the second cultivation seed liquid into a second-stage seed culture medium for sub-cultivation. The sub-cultivation of the present invention is mainly to enrich bacteria, and at the same time enhance and stabilize their sulfur oxidation ability. When the present invention conducts sub-cultivation, preferably 5-10 mL of the second cultivation seed liquid is inoculated into 100-200 mL of the second-stage seed culture medium to obtain a sub-cultivation system. As an alternative embodiment of the present invention, the volume ratio of the sub-cultivation system to its culture container can be 1:(5-10), or can be 1:5, 1.1:5, 1:6, 1:7, 1:8, 1:9 or 1:10. During the sub-cultivation process of the present invention, preferably, the first mixed gas of sulfur-oxidizing bacteria is replaced every 2-10 days; the interval time can be 2, 3, 4, 5, 6, 7, 8, 9 or 10 days. When the present invention conducts sub-cultivation, it is preferably continuously replaced 5-10 times for the culture medium and the mixed gas, which can be 5, 6, 7, 8, 9 or 10 times; the present invention preferably replaces the mixed gas while replacing the culture medium, and when replacing the culture medium each time, the volume ratio of the culture system to the culture container remains unchanged. During the sub-cultivation process of the present invention, after continuously replacing the first mixed gas of sulfur-oxidizing bacteria 5-10 times, it is preferably cultured until the concentration of H2S no longer decreases during the last cultivation to obtain an enriched flora of sulfur-oxidizing bacteria.

[0079] The sulfur-oxidizing bacteria enriched by the present invention through specific culture medium and gas conditions have a high hydrogen sulfide degradation ability, can use hydrogen sulfide (H2S) as an electron donor, and generate substances such as sulfates by oxidizing hydrogen sulfide, thereby effectively reducing the hydrogen sulfide concentration in the environment and being able to treat a large amount of hydrogen sulfide gas in a short time. The enriched flora of sulfur-oxidizing bacteria has strong tolerance: the enriched sulfur-oxidizing bacteria can grow well under specific culture conditions and have strong tolerance. The enriched flora of sulfur-oxidizing bacteria can reduce environmental pollution: hydrogen sulfide is a toxic and harmful gas with a strong odor, which harms the environment and human health. By degrading hydrogen sulfide, sulfur-oxidizing bacteria can effectively reduce its concentration in the environment, reduce pollution to the atmosphere and water bodies, and protect the ecological environment. The synergistic effect of the enriched flora of sulfur-oxidizing bacteria: sulfur-oxidizing bacteria and other microbial populations (such as methane-oxidizing bacteria and hydrogen-oxidizing bacteria) together form a composite microbial metabolic group, which can play a synergistic role. For example, during the synchronous carbon fixation and denitrification process, sulfur-oxidizing bacteria can act together with methane-oxidizing bacteria and hydrogen-oxidizing bacteria to improve the metabolic efficiency and stability of the whole system and achieve the synchronous removal of multiple pollutants.

[0080] The method for enriching the microbial population mainly composed of methanotrophs in soil of the present invention includes: inoculating the microorganisms in the soil into the primary seed medium for methanotrophs, introducing the first mixed gas for methanotrophs for the first cultivation, and culturing until the CH4 concentration no longer decreases to obtain the primary seed liquid for methanotrophs. In the present invention, the primary seed medium for methanotrophs can also be referred to as the third primary seed medium.

[0081] In the present invention, the composition of the third primary seed medium includes: KNO3: 100 - 200 mg / L, KH2PO4: 100 - 200 mg / L, MgSO4·7H2O: 25 - 55 mg / L, CaCl2·2H2O: 5 - 20 mg / L, EDTA: 2 - 5 mg / L, CuCl2·5H2O: 0.05 - 0.1 mg / L, FeSO4·7H2O: 1 - 2 mg / L, ZnSO4·7H2O: 0.1 - 0.2 mg / L, NiCl2·6H2O: 0.008 - 0.02 mg / L, CoCl2·6H2O: 0.1 - 0.2 mg / L, and Na2MoO4: 0.01 - 0.03 mg / L. The pH of the third primary seed medium can be 6.8 - 7.4. In the present invention, the pH is preferably adjusted using a phosphate buffer solution. As an optional implementation manner of the present invention, the composition of the third primary seed medium is: KNO3: 100 mg / L, KH2PO4: 100 mg / L, MgSO4·7H2O: 50 mg / L, CaCl2·2H2O: 10 mg / L, EDTA: 5 mg / L, CuCl2·5H2O: 0.1 mg / L, FeSO4·7H2O: 2 mg / L, ZnSO4·7H2O: 0.1 mg / L, NiCl2·6H2O: 0.02 mg / L, CoCl2·6H2O: 0.2 mg / L, and Na2MoO4: 0.03 mg / L. In the present invention, the third primary seed medium provides the optimal medium according to the metabolic conditions of methanotrophs.

[0082] In the present invention, the methane-oxidizing bacteria mixed gas includes CH4 and O2; the volume ratio of CH4 to O2 is (50-80):(50-20), which can be (50-75):(50-25), or can be 50:50, 75:25, 80:20 or 60:40. In the present invention, the methane-oxidizing bacteria mixed gas can also be referred to as the first methane-oxidizing bacteria mixed gas. In the present invention, the first methane-oxidizing bacteria mixed gas can meet the metabolic growth needs of methane-oxidizing bacteria. Among them, the metabolism of methane-oxidizing bacteria carries out metabolic cycles with methane as the carbon source. The composition ratio setting of the first methane-oxidizing bacteria mixed gas provided by the present invention can significantly shorten the enrichment time of methane-oxidizing bacteria. If the gas ratio of the first mixed gas is changed, for example, increasing the volume of CH4 or increasing the proportion of O2, the enrichment time of methane-oxidizing bacteria will be significantly prolonged.

[0083] After obtaining the primary methane-oxidizing bacteria seed solution, the present invention inoculates the primary methane-oxidizing bacteria seed solution into the secondary methane-oxidizing bacteria seed culture medium and introduces the first methane-oxidizing bacteria mixed gas for cultivation to obtain an enriched methane-oxidizing bacteria flora. In the present invention, the secondary methane-oxidizing bacteria seed culture medium can also be referred to as the third secondary seed culture medium. The cultivation includes secondary cultivation and sub-cultivation.

[0084] In the present invention, the third secondary seed culture medium is based on the third primary seed culture medium and further includes: 1%-3% glucose, 0.6%-2% yeast powder, and 1.5%-3.2% casein peptone. The pH of the third secondary seed culture medium can be 6.8-7.4.

[0085] In the process of enriching the microbial population mainly composed of methane-oxidizing bacteria in the soil in the present invention, the methane-oxidizing bacteria use CH4 as the carbon source and electron donor, and O2 as the electron acceptor to carry out metabolic cycles. The relatively high CH4 ratio (50%-85%) in the mixed gas ensures that the methane-oxidizing bacteria have sufficient carbon sources and electron donors for metabolism, while the proportion of O2 meets its respiratory needs. By introducing the mixed gas and exhausting the original air, a micro-oxic environment suitable for the growth of methane-oxidizing bacteria is created. The culture medium for enriching the microbial population mainly composed of methane-oxidizing bacteria in the soil in the present invention contains components such as KNO3, KH2PO4, MgSO4·7H2O, CaCl2·2H2O, EDTA, CuCl2·5H2O, FeSO4·7H2O, etc. These components provide the nutrients and buffer systems required by methane-oxidizing bacteria. The pH value of the culture medium is adjusted between 6.8 and 7.4, which is suitable for the growth of methane-oxidizing bacteria.

[0086] In the present invention, the soil includes: the soil 2 - 10 cm below the surface of the paddy field for water storage and / or the sludge in the oxidation pond of the sewage treatment plant. Through research, it is found in the present invention that the soil collected from the soil 2 - 10 cm below the surface of the paddy field for water storage and / or the sludge in the oxidation pond of the sewage treatment plant is rich in methane - oxidizing bacteria. After obtaining the corresponding soil, it is preferred to mix the soil with water to obtain a soil - water mixture. In the present invention, the mass - volume ratio of the soil to water can be (5 - 10) g:(100 - 120) mL. As an optional embodiment of the present invention, the mass - volume ratio of the soil to water can be 5 g:100 mL, 10 g:100 mL, 5 g:120 mL, 10 g:100 mL, 8 g:100 mL, 8 g:120 mL, 5 g:110 mL, 8 g:110 mL or 8 g:120 mL. After obtaining the soil - water mixture, it is preferred to oscillate the soil - water mixture on a shaker at 30 °C and 120 rpm for 1 - 2 h. After the oscillation is completed, it is preferred to mix the oscillated soil - water mixture with the third primary seed medium; the volume ratio of the soil - water mixture to the third primary seed medium is 1:(1 - 2). After the mixing is completed, a soil - medium mixing system is obtained. After obtaining the soil - medium mixing system, it is preferred to stir the soil - medium mixing system with a magnetic stirrer at 250 - 450 rpm for 1 - 2 h. After the stirring is completed, the mixed soil - medium mixing system is placed in a culture container; the volume ratio of the soil - medium mixing system to the culture container is 1:(5 - 10), and it can also be 1:5, 1.1:5, 1:6, 1:7, 1:8, 1:9 or 1:10. After placing the soil - medium mixing system in the culture container, a first mixed gas is introduced. Before introducing the first mixed gas for methane - oxidizing bacteria, it is preferred to first introduce nitrogen to evacuate the original air. After evacuating the original air, then introduce the first mixed gas for methane - oxidizing bacteria. In the present invention, the temperature of the first culture is 25 - 35 °C. As an optional embodiment of the present invention, the temperature of the first culture can be 25, 26, 27, 28, 29, 30, 31, 32, 33, 34 or 35 °C. In the present invention, the rotation speed of the first culture can be 150 - 210 rpm. As an optional embodiment of the present invention, the rotation speed can be 150, 160, 170, 180, 190, 200 or 210 rpm. The first culture in the present invention is preferably cultured until the CH4 concentration no longer decreases to obtain a primary seed liquid of methane - oxidizing bacteria.

[0087] After obtaining the primary seed liquid of methane - oxidizing bacteria, the present invention inoculates the primary seed liquid of methane - oxidizing bacteria into the third secondary seed medium, introduces the first mixed gas for methane - oxidizing bacteria for the second culture and sub - culture to obtain an enriched flora of methane - oxidizing bacteria.

[0088] When inoculating the primary seed solution of the methanotrophic bacteria into the third secondary seed culture medium in the present invention, it is preferably to inoculate 5 - 10 mL of the primary seed solution of the methanotrophic bacteria into 100 - 200 mL of the third secondary seed culture medium to obtain a secondary culture system. As an optional implementation mode of the present invention, the volume ratio of the secondary culture system to its culture container can be 1:(5 - 10), or can also be 1:5, 1.1:5, 1:6, 1:7, 1:8, 1:9 or 1:10. After inoculating the primary seed solution of the methanotrophic bacteria into the third secondary seed culture medium in the present invention, it is preferably to first introduce nitrogen to exhaust the original air. After exhausting the original air, then introduce the first mixed gas of the methanotrophic bacteria until the culture container is filled. In the present invention, the temperature of the second culture is 25 - 35 °C. As an optional implementation mode of the present invention, the temperature of the second culture can be 25, 26, 27, 28, 29, 30, 31, 32, 33, 34 or 35 °C. In the present invention, the rotation speed of the second culture can be 150 - 210 rpm. As an optional implementation mode of the present invention, the rotation speed can be 150, 160, 170, 180, 190, 200 or 210 rpm. During the second culture process of the present invention, it is preferably to culture until the CH4 concentration no longer decreases to obtain the second culture seed solution. As an optional implementation mode of the present invention, the time of the second culture is shorter than that of the lower culture.

[0089] After the second culture is completed, the present invention preferably further includes inoculating the second culture seed solution into the third secondary seed culture medium for subculture. The main purpose of the subculture in the present invention is to enrich bacteria, and at the same time enhance and stabilize their methane oxidation ability. When conducting the subculture in the present invention, it is preferably to inoculate 5 - 10 mL of the second culture seed solution into 100 - 200 mL of the third secondary seed culture medium to obtain a subculture system. As an optional implementation mode of the present invention, the volume ratio of the subculture system to its culture container can be 1:(5 - 10), or can also be 1:5, 1.1:5, 1:6, 1:7, 1:8, 1:9 or 1:10. During the subculture process of the present invention, it is preferably to replace the secondary seed culture medium and the first mixed gas of the methanotrophic bacteria every 2 - 10 days; the interval time can be 2, 3, 4, 5, 6, 7, 8, 9 or 10 days. When conducting the subculture in the present invention, it is preferably to continuously replace the culture medium and the mixed gas 5 - 10 times, which can be 5, 6, 7, 8, 9 or 10 times; the present invention preferably replaces the mixed gas while replacing the culture medium, and when replacing the culture medium each time, the volume ratio of the culture system to the culture container remains unchanged. During the subculture process of the present invention, after continuously replacing the first mixed gas of the methanotrophic bacteria 5 - 10 times, it is preferably to culture until the CH4 concentration no longer decreases during the last culture to obtain a methanotrophic bacteria enriched flora.

[0090] In the present invention, the methane-oxidizing bacteria enriched under specific culture medium and gas conditions have a high methane degradation ability, can process a large amount of methane gas in a short time, and generate carbon dioxide and water by oxidizing methane. The enriched methane-oxidizing bacteria flora has strong adaptability: the enriched methane-oxidizing bacteria can grow well under specific culture conditions and have strong adaptability. The enriched methane-oxidizing bacteria flora can promote carbon fixation: during the process of degrading methane, the methane-oxidizing bacteria can fix part of the carbon as biomass and generate useful substances such as microbial protein, which not only helps to reduce greenhouse gas emissions, but also enables the effective utilization of carbon resources, meeting the requirements of sustainable development. The enriched methane-oxidizing bacteria flora has a synergistic effect: the methane-oxidizing bacteria and other microbial populations (such as hydrogen-oxidizing bacteria and sulfur-oxidizing bacteria) together form a complex microbial metabolic group and can play a synergistic role. For example, in the process of synchronous carbon fixation and denitrification, the methane-oxidizing bacteria can act together with sulfur-oxidizing bacteria and hydrogen-oxidizing bacteria to improve the metabolic efficiency and stability of the whole system.

[0091] Through the optimized design of the above-mentioned culture medium, gas composition, culture conditions, etc., the present invention can effectively enrich hydrogen-oxidizing bacteria, sulfur-oxidizing bacteria and methane-oxidizing bacteria. The enrichment process of each microbial population is based on its specific metabolic requirements and environmental conditions. By precisely controlling the culture medium components and gas ratios, the efficient enrichment and activity maintenance of the target microbial population are ensured. These optimized conditions not only improve the enrichment efficiency of the microorganisms, but also provide a stable microbial metabolic group for the subsequent synchronous carbon fixation and denitrification process.

[0092] After obtaining the enriched methane-oxidizing bacteria flora, the enriched sulfur-oxidizing bacteria flora and the enriched hydrogen-oxidizing bacteria flora respectively, the present invention mixes and inoculates the enriched methane-oxidizing bacteria flora, the enriched sulfur-oxidizing bacteria flora and the enriched hydrogen-oxidizing bacteria flora into a domestication culture medium for domestication culture to obtain a complex microbial metabolic group.

[0093] In the present invention, a third mixed gas is introduced during domestication culture; the composition of the third mixed gas is H2, O2, CO2, CH4 and H2S; the volume ratio of H2, O2, CO2, CH4 and H2S is (0.1~1):(10~20):(15~25):(50~60):(0.1~0.3). The third mixed gas of the present invention is closest to the true composition of biogas, in order to further domesticate the survival and digestion ability of the mixed bacterial system under true biogas.

[0094] In the present invention, the composition of each 1 L of the acclimation medium includes: 0.5 - 1.00 g of MgSO4·7H2O, 1.00 - 2.5 g of KNO3, 0.15 - 0.3 g of KH2PO4, 0.2 - 0.45 g of Na2HPO4, 0.1 - 0.15 g of CaCl2·2H2O, and 1 mL of trace element solution. As an optional embodiment of the present invention, the composition of each 1 L of the acclimation medium is: 1.00 g of MgSO4·7H2O, 1.00 g of KNO3, 0.27 g of KH2PO4, 0.28 g of Na2HPO4, 0.13 g of CaCl2·2H2O, and 1 mL of trace element solution. In the present invention, the composition of each 1 L of the trace element solution includes 1.00 g of EDTA-2Na, 0.50 g of FeSO4·7H2O, 0.07 g of ZnSO4·7H2O, 0.03 g of MnCl2·4H2O, 0.30 g of H3BO3, 0.60 g of CoCl2·6H2O, 0.02 g of NiCl2·6H2O, 0.03 g of Na2MoO4·2H2O, and 0.01 g of CuCl2·H2O. In the present invention, the acclimation medium is a basal nitrate mineral medium, and adding nutrient elements can simultaneously meet the metabolic requirements of the enriched methanotrophic bacteria population, the enriched sulfur-oxidizing bacteria population, and the enriched hydrogen-oxidizing bacteria population.

[0095] When the present invention mixes and inoculates the methane-oxidizing bacteria enriched flora, the sulfur-oxidizing bacteria enriched flora, and the hydrogen-oxidizing bacteria enriched flora into the acclimation medium, it is preferably to mix the corresponding enriched floras into the acclimation medium at an inoculation ratio of 5% - 25% (v) respectively; the volume of the acclimation medium in the culture flask is 10% - 25% (v). After the inoculation of the present invention is completed, an enriched flora medium mixed system is obtained. After obtaining the enriched flora medium mixed system, the present invention introduces a third mixed gas into the enriched flora medium mixed system until the culture flask is full. Before introducing the third mixed gas, the present invention preferably first introduces nitrogen to evacuate the original air. After evacuating the original air, the third mixed gas is introduced. The temperature of the acclimation culture of the present invention can be 28 - 35°C; intermittent oscillation is preferably carried out during the acclimation culture process; the rotation speed of the oscillation can be 120 - 280 rpm; during the intermittent oscillation, it is stationary for 15 - 30 min and oscillates for 10 - 20 min. The acclimation culture of the present invention is preferably carried out in a shaker. During the acclimation culture process of the present invention, it is preferably to replace the fresh acclimation medium and the fresh third mixed gas every 1 - 2 days. Each time the fresh acclimation medium is replaced, it is preferably to make the acclimation medium in the culture flask be 10% - 25% (v). When the present invention replaces the fresh acclimation medium for subculture, it is preferably to inoculate 40% - 60% (v), or 50% of the bacterial liquid in the original culture system into the fresh acclimation medium. In the present invention, the acclimation culture is continuously subcultured 3 - 4 times; the acclimation medium also includes Na2S and ammonium chloride; the mass concentration of Na2S in the acclimation medium during the first acclimation culture is 1 - 2 mg / L, and the mass concentration of ammonium chloride is 100 - 300 mg / L; each time during subculture, the mass concentration of Na2S and ammonium chloride in the acclimation medium is 1.5 - 2 times that of the previous culture. The present invention preferably measures the OD 600 value every 2 - 4 h during each subculture to monitor the growth of the mixed bacteria. The present invention adds Na2S and ammonium chloride to the acclimation medium mainly to improve the H2S tolerance and ammonia nitrogen degradation ability of the acclimation culture mixed flora.

[0096] After the acclimation culture is completed, the present invention obtains a composite microbial metabolic group.

[0097] After obtaining the composite microbial metabolic group, the present invention preferably inoculates the composite microbial metabolic group into an acclimation medium containing biogas slurry and introduces biogas for synchronous carbon fixation and denitrification.

[0098] The present invention has no special limitation on the composition and source of the biogas slurry, and conventional biogas slurry in the art can be used. For verifying the effect, the biogas slurry used in the present invention is the biogas slurry simulated in the laboratory; the composition of the biogas slurry includes: acetic acid: 1.5 - 2.5 g / L, propionic acid: 0.5 - 1.0 g / L, butyric acid: 0.2 - 0.5 g / L, glucose: 0.1 - 0.3 g / L, ammonium salt (NH4Cl): 0.3 - 0.5 g / L, potassium dihydrogen phosphate (KH2PO4): 0.05 - 0.1 g / L, potassium chloride (KCl): 0.05 - 0.1 g / L, sodium sulfate (Na2SO4): 0.02 - 0.05 g / L, ferric chloride (FeCl3): 0.001 - 0.005 g / L, magnesium sulfate (MgSO4·7H2O): 0.01 - 0.02 g / L, calcium chloride (CaCl2): 0.01 - 0.02 / L, with water as the solvent. In the present invention, the addition amount of the biogas slurry is 10% - 20% (v / v) of the culture container; the addition amount of the acclimation medium is 10% - 25% of the culture container. In the present invention, when inoculating the composite microbial metabolic group into the acclimation medium containing biogas slurry, the inoculation amount of the composite microbial metabolic group is 20% - 50% of the volume of the acclimation medium containing biogas slurry. After inoculation, the present invention obtains a microbial metabolic group - biogas slurry mixed system. The present invention introduces biogas into the microbial metabolic group - biogas slurry mixed system. The present invention has no special limitation on the composition and source of the biogas, and conventional biogas in the art can be used. For verifying the effect, the third mixed gas described in the above technical solution is used as the biogas in the present invention. Before introducing the third mixed gas, the present invention preferably first introduces nitrogen to evacuate the original gas, and then introduces the third mixed gas until the culture container is filled with the third mixed gas. The temperature for synchronous carbon fixation and denitrification in the present invention can be 28 - 35°C, or can be 28, 29, 30, 31, 32, 33, 34 or 35°C; intermittent oscillation is preferably carried out during the synchronous carbon fixation and denitrification process; the rotation speed of the oscillation can be 120 - 280 rpm; when intermittently oscillating, it is static for 15 - 30 min and oscillates for 10 - 20 min. The synchronous carbon fixation and denitrification in the present invention is preferably carried out in a shaker; the time for synchronous carbon fixation and denitrification can be 12 - 35 d. During the synchronous carbon fixation and denitrification process in the present invention, the third mixed gas is preferably replaced every 1 - 2 days. The purpose of replacing the third mixed gas every 1 - 2 days during this process is that the composite microbial metabolic group can efficiently utilize gases such as H2, CO2, CH4 and H2S. If not replaced in time, it will reduce the growth rate of the composite microbial metabolic period, and thus reduce the protein yield during the synchronous carbon fixation and denitrification process.

[0099] Through the synchronous carbon fixation and denitrification process, the present invention can enable the enriched composite microbial metabolic group to efficiently utilize methane and carbon dioxide in biogas and nutrients such as nitrogen and phosphorus in biogas slurry to produce microbial protein.

[0100] After the synchronous carbon fixation and denitrification of the present invention are completed, it is preferred to centrifuge the fermentation product at 10,000 - 12,000 rpm for 10 - 30 minutes, and pour off the supernatant; wash the centrifuged product with phosphate buffer and then centrifuge again, repeating 1 - 3 times; the fermentation product after multiple centrifugation washings is baked at 103 - 110 °C for 20 - 30 hours to obtain microbial protein.

[0101] The present invention provides the application of the method described in the above technical solution in the resource utilization of biogas slurry and / or biogas.

[0102] In order to further illustrate the present invention, the technical solution provided by the present invention will be described in detail below in conjunction with the drawings and embodiments, but they cannot be understood as limiting the protection scope of the present invention.

[0103] Example 1

[0104] A method for synchronous carbon fixation and denitrification, the steps are as follows:

[0105] S1: Collect different soils at different sampling sites respectively, and introduce a first mixed gas into different seed media for primary enrichment culture:

[0106] The soil samples are taken from the soil 2 - 10 cm below the surface of a paddy field for storing water, livestock and poultry manure compost residues, the soil 2 - 10 cm below the surface of a marsh, and the sludge in the oxidation pond of a sewage treatment plant. Among them, the soil 2 - 10 cm below the surface of the paddy field for storing water is taken from a paddy field in Shunyi District, Beijing; the livestock and poultry manure compost residues are taken from a composting site on a farm; the soil 2 - 10 cm below the surface of the marsh is taken from a riverside marsh; the sludge in the oxidation pond of the sewage treatment plant is taken from the Xiaohongmen Sewage Treatment Plant in Beijing.

[0107] The reason for selecting these four kinds of soils is that for sulfide-oxidizing bacteria (abbreviated as sulfur-oxidizing bacteria), they are easily enriched in the soil 2 - 10 cm below the surface of a marsh and / or the sludge in the oxidation pond of a sewage treatment plant. Similarly, hydrogen-oxidizing bacteria are easily enriched in the soil 2 - 10 cm below the surface of a paddy field for storing water and / or livestock and poultry manure compost residues, and methane-oxidizing bacteria are easily enriched in the soil 2 - 10 cm below the surface of a paddy field for storing water and / or the sludge in the oxidation pond of a sewage treatment plant.

[0108] The corresponding relationship of the primary seed liquid medium:

[0109] For the soil corresponding to the enrichment of hydrogen-oxidizing bacteria: the soil is 2-10 cm below the surface of the paddy field and the livestock manure compost residue. The corresponding culture medium for this soil is the first primary seed culture medium, and the composition of the first primary seed culture medium is: Na2HPO4: 2800 mg / L, KH2PO4: 1000 mg / L, (NH4)2SO4: 500 mg / L, MgCl2: 53 mg / L, CaCl2·2H2O: 50 mg / L, disodium EDTA: 0.5 mg / L, FeSO4·7H2O: 0.2 mg / L, ZnSO4·7H2O: 0.01 mg / L, MnCl2·4H2O: 0.003 mg / L, H3BO3: 0.03 mg / L, CoCl2·6H2O: 0.02 mg / L, CuCl2·2H2O: 0.001 mg / L, NiCl2·6H2O: 0.002 mg / L, Na2MoO4·2H2O: 0.003 mg / L.

[0110] For the soil corresponding to the enrichment of ② sulfur-oxidizing bacteria: the soil 2 - 10 cm below the surface layer of the swamp and the sludge in the oxidation pond of the sewage treatment plant. The culture medium corresponding to this soil is the second primary seed culture medium, and the composition of the second primary seed culture medium is: phenol red: 0.003 g / L, 1,4-piperazinediethanesulfonic acid: 6.500 g / L, sodium chloride: 25.000 g / L, magnesium sulfate heptahydrate: 2.700 g / L, potassium chloride: 0.500 g / L, ammonium chloride: 0.250 g / L, sodium thiosulfate pentahydrate: 2.480 g / L, calcium chloride dihydrate: 0.140 g / L, magnesium chloride hexahydrate: 4.300 g / L, dipotassium hydrogen phosphate: 0.140 g / L, ammonium ferrous sulfate hexahydrate: 0.002 g / L, trace element solution: 1 mL / L, vitamin solution: 10 mL / L. The composition of the trace element solution is: MgSO4·7H2O 2.00 g / L, MnSO4·H2O 0.5 g / L, NaCl 0.8 g / L, FeSO4·7H2O 0.15 g / L, CoSO4·7H2O 0.18 g / L, CaCl2·2H2O 0.15 g / L, ZnSO4·7H2O 0.16 g / L, CuSO4·5H2O 0.015 g / L, AlK(SO4)2·12H2O 0.015 g / L, H3BO3 0.015 g / L, Na2MoO4·2H2O 0.03 g / L, NiCl2·6H2O 0.03 g / L, Na2SeO3·5H2O 0.2 mg / L and Na2WO4·2H2O 0.2 mg / L. The composition of the vitamin solution is: vitamin B7 1.5 mg / L, folic acid 1.5 mg / L, pyridoxine hydrochloride 8 mg / L, vitamin B1 4 mg / L, riboflavin 4 mg / L, vitamin B3 4 mg / L, vitamin B5 4 mg / L, vitamin B 12 0.15 mg / L, p-aminobenzoic acid 4 mg / L and (DL)-α-lipoic acid 4 mg / L.

[0111] For the soil corresponding to the enrichment of ③ methane-oxidizing bacteria: the soil 2 - 10 cm below the surface layer of the paddy field with water storage and the sludge in the oxidation pond of the sewage treatment plant. The culture medium corresponding to this soil is the third primary seed culture medium, and the composition of the third primary seed culture medium is: KNO3: 100 mg / L, KH2PO4: 100 mg / L, MgSO4·7H2O: 50 mg / L, CaCl2·2H2O: 10 mg / L, EDTA: 5 mg / L, CuCl2·5H2O: 0.1 mg / L, FeSO4·7H2O: 2 mg / L, ZnSO4·7H2O: 0.1 mg / L, NiCl2·6H2O: 0.02 mg / L, CoCl2·6H2O: 0.2 mg / L, Na2MoO4: 0.03 mg / L.

[0112] The above culture media were all dissolved in 1 L of distilled water, and the pH was adjusted to 6.8 - 7.4 with phosphate buffer, and then autoclaved at 121 °C for 30 min.

[0113] Enrichment process of hydrogen-oxidizing bacteria, sulfur-oxidizing bacteria and methanotrophic bacteria: Take the corresponding soil samples respectively, and then take 10 g of the corresponding soil and add it to a conical flask containing 120 mL of sterilized distilled water, and shake it on a shaker at 30 °C and 120 rpm for 1.5 h to obtain the soil-water mixture enriched with each bacterium.

[0114] Mix 100 mL of the well-mixed soil sample (i.e., the soil-water mixture) in the conical flask with 200 mL of the above culture medium correspondingly, and stir it with a magnetic stirrer at 250 rpm for 1 h.

[0115] Put 100 mL of the mixed solution into a 500 mL sterile serum bottle, pass nitrogen for 5 min to empty the original air, and then pass the first mixed gas until the sterile serum bottle is filled with the first mixed gas.

[0116] The first mixed gas:

[0117] For the enrichment of the above-mentioned ① hydrogen-oxidizing bacteria, the corresponding first mixed gas I (referred to as the hydrogen-oxidizing bacteria first mixed gas for easy distinction), the composition is H2, O2 and CO2, and the volume ratio of H2:O2:CO2 = 7:2:1;

[0118] For the enrichment of the above-mentioned ② sulfur-oxidizing bacteria, the corresponding first mixed gas II (referred to as the sulfur-oxidizing bacteria first mixed gas for easy distinction), the composition is H2, O2, CO2, H2S, and the volume ratio of H2:O2:CO2:H2S = 3:3:2:2;

[0119] For the enrichment of ③ methanotrophic bacteria, the corresponding first mixed gas III (referred to as the methanotrophic bacteria first mixed gas for easy distinction), the composition is CH4 and O2, and the volume ratio of CH4:O2 = 3:1.

[0120] After passing the first mixed gas into each of the above culture systems, the bacteria were cultured in a constant temperature shaker at 28 °C and 180 rpm.

[0121] For the enrichment of ① hydrogen-oxidizing bacteria, when more than 50% of H2 is consumed or the CO2 concentration no longer decreases, the culture solution is obtained;

[0122] For the enrichment of ② sulfur-oxidizing bacteria, when the H2S concentration decreases to no longer decrease, the culture solution is obtained;

[0123] For the enrichment of ③ methanotrophic bacteria, when the CH4 concentration decreases to no longer decrease, the culture solution is obtained.

[0124] S2: Take 10 mL of the culture medium from S1 and re-add it to serum bottles (500 mL) respectively, and then add 100 mL of the corresponding secondary seed medium respectively.

[0125] The secondary seed medium is prepared by adding 1.5 wt.% glucose, 0.6 wt.% yeast powder, and 1.5 wt.% casein peptone to the corresponding primary seed medium, and is respectively called the first secondary seed medium (for enrichment of hydrogen-oxidizing bacteria), the second secondary seed medium (for enrichment of sulfur-oxidizing bacteria), and the third secondary seed medium (for enrichment of methanotrophic bacteria). All are dissolved in 1 L of distilled water, and the pH is adjusted to 6.8 - 7.4 with phosphate buffer, and autoclaved at 121 °C for 30 min.

[0126] After adding the culture medium to the secondary seed medium, replace the gas with nitrogen and introduce the first mixed gas, and the culture conditions are the same as in S1.

[0127] Similarly, for ① enrichment of hydrogen-oxidizing bacteria, when more than 50% of H2 is consumed or the CO2 concentration no longer decreases;

[0128] For ② enrichment of sulfur-oxidizing bacteria, the H2S concentration decreases to no longer decrease;

[0129] For ③ enrichment of methanotrophic bacteria, the CH4 concentration decreases and then no longer decreases;

[0130] After ①, ②, and ③ proceed until the corresponding gas no longer changes, sub-cultures are carried out respectively: Take 10 mL of the culture medium and re-add it to the new corresponding secondary seed medium (100 mL) to obtain a culture system, and then culture the 110 mL of the culture system in a 500 mL serum bottle. During this period, replace the secondary seed medium and the first mixed gas every 2 days, and replace them continuously for 5 times. After the last replacement of the first mixed gas, the corresponding gas volume remains unchanged, and the hydrogen-oxidizing bacteria enrichment flora, sulfur-oxidizing bacteria enrichment flora, and methanotrophic bacteria enrichment flora are obtained respectively.

[0131] S3: Inoculate different secondary enrichment floras into the same reactor containing the acclimation medium, that is, inoculate the hydrogen-oxidizing bacteria enrichment flora, sulfur-oxidizing bacteria enrichment flora, and methanotrophic bacteria enrichment flora obtained in S2, add the acclimation culture medium in stages, and carry out intermittent culture by introducing the third mixed gas:

[0132] Among them, the composition of each 1 L of acclimation medium is: 1.00 g of MgSO4·7H2O, 1.00 g of KNO3, 0.27 g of KH2PO4, 0.28 g of Na2HPO4, 0.13 g of CaCl2·2H2O, and 1 mL of trace elements. The composition of each 1 L of trace elements is 1.00 g of EDTA-2Na, 0.50 g of FeSO4·7H2O, 0.07 g of ZnSO4·7H2O, 0.03 g of MnCl2·4H2O, 0.30 g of H3BO3, 0.60 g of CoCl2·6H2O, 0.02 g of NiCl2·6H2O, 0.03 g of Na2MoO4·2H2O, and 0.01 g of CuCl2·H2O. The acclimation medium also contains 1 mg / L of Na2S and 100 mg / L of ammonium chloride. The medium and trace elements are dissolved in 1 L of distilled water, the pH is adjusted to 6.8 to 7.2, and autoclaved at 121 °C for 30 min.

[0133] The bacterial liquid cultured separately in S2 (i.e., the enriched flora of hydrogen-oxidizing bacteria, the enriched flora of sulfur-oxidizing bacteria, and the enriched flora of methanotrophic bacteria) was mixed in the acclimation medium at an inoculation ratio of 20% (v), and the acclimation medium in the culture bottle was 25% (v). The gas was replaced with nitrogen, the original gas in the culture bottle was emptied, and the headspace was filled with the third mixed gas (i.e., the entire culture bottle was filled). It was placed in a constant temperature incubator at a temperature of 28 °C, shaken at 180 rpm in an intermittent mode, that is, static for 15 min and then shaken for 20 min. Every 2 days, the fresh acclimation medium and fresh gas (fresh third mixed gas) were replaced. Each time the fresh acclimation medium was replaced, the acclimation medium in the culture bottle was 25% (v), and the bacteria in the original bottle were inoculated into the new bottle, and the inoculation amount was 50% of the bacterial liquid in the original bottle inoculated into the fresh acclimation medium. The continuous passage was carried out for 3 rounds. Each time during the passage, the OD value was measured every 4 h to monitor the growth of the mixed bacteria. The growth of the bacteria in the third round of this process is as shown in 600 the following. Figure 1 shown.

[0134] 1 mg / L of Na2S and 100 mg / L of ammonium chloride were added to the first acclimation medium, and the addition amounts of Na2S and ammonium chloride in each subsequent passage were 1.5 times that of the previous time.

[0135] The third mixed gas contains H2, O2, CO2, CH4, and H2S, and the composition is that the volume ratio of H2:O2:CO2:CH4:H2S is 1:10:25:60:0.2.

[0136] S4: The third mixed gas was introduced into the container containing the acclimation medium (the volume of the acclimation medium in the container was 25%), and 10% of biogas slurry was added to culture the composite microbial metabolism group.

[0137] Inoculate the bacterial liquid (complex microbial metabolic group) in S3 medium into a culture flask containing acclimation medium with biogas slurry, and the inoculation ratio is 40% (v). The culture time is 20 days. During the culture process, the culture temperature is 28 °C, the shaker speed is 180 rpm, and it shakes in an intermittent mode, that is, it stands still for 15 minutes and then swings for 20 minutes. Replace the new gas (the third mixed gas) every 2 days.

[0138] The above biogas slurry is a laboratory-simulated biogas slurry, and its composition is: acetic acid (vinegar): 1.5 g / L, propionic acid: 0.5 g / L, butyric acid: 0.2 g / L, glucose: 0.2 g / L, ammonium salt (NH4Cl): 0.3 g / L, potassium dihydrogen phosphate (KH2PO4): 0.05 g / L, potassium chloride (KCl): 0.05 g / L, sodium sulfate (Na2SO4): 0.02 g / L, ferric chloride (FeCl3): 0.002 g / L, magnesium sulfate (MgSO4·7H2O): 0.01 g / L, calcium chloride (CaCl2): 0.01 g / L, with water as the solvent.

[0139] After the culture is completed, separate the microbial protein. Among them, during the culture process, take the culture solution to separate the bacteria every 4 days, and detect the cell dry weight and protein content.

[0140] The steps for separating the obtained protein include: centrifuge the fermentation product at 10000 rpm for 10 minutes, pour off the supernatant; wash the centrifuged product with phosphate buffer and then centrifuge again, repeat 3 times; bake the fermentation product after multiple centrifugation and washing at 105 °C for 24 hours to obtain microbial protein.

[0141] During the culture process, the detection results of cell dry weight and protein content are as Figure 2 shown. The final biomass dry weight (i.e., cell dry weight) is 495 mg / L, and the protein content in the biomass is 59.4%.

[0142] Example 2

[0143] A method for synchronous carbon fixation and nitrogen removal, the steps are the same as in Example 1, and the only differences are: ① For the soil corresponding to the enrichment of hydrogen-oxidizing bacteria, it is the soil 2 - 10 cm below the surface of the paddy field for storing water. ② For the soil corresponding to the enrichment of sulfur-oxidizing bacteria, it is the soil 2 - 10 cm below the surface of the swamp. ③ For the soil corresponding to the enrichment of methane-oxidizing bacteria, it is the soil 2 - 10 cm below the surface of the paddy field for storing water.

[0144] The biomass dry weight is 494 mg / L, and the protein content in the biomass is 51.8%.

[0145] Example 3

[0146] A method for synchronous carbon sequestration and nitrogen removal, the steps are the same as those in Example 1, and the only difference is that: for ① the soil corresponding to the enrichment of hydrogen-oxidizing bacteria is the soil collected from the residue of livestock and poultry manure compost. ② the soil corresponding to the enrichment of sulfur-oxidizing bacteria is the soil collected from the sludge in the oxidation pond of a sewage treatment plant. ③ the soil corresponding to the enrichment of methanotrophs is the soil collected from the sludge in the oxidation pond of a sewage treatment plant.

[0147] The dry weight of biomass is 505 mg / L, and the protein content in the biomass is 50.2%.

[0148] Example 4

[0149] A method for synchronous carbon sequestration and nitrogen removal, the steps are the same as those in Example 1, and the only difference is that: for ① the soil corresponding to the enrichment of hydrogen-oxidizing bacteria is the soil 2 - 10 cm below the surface layer of a paddy field for storing water. ② the soil corresponding to the enrichment of sulfur-oxidizing bacteria is the soil collected from the sludge in the oxidation pond of a sewage treatment plant. ③ the soil corresponding to the enrichment of methanotrophs is the soil collected from the sludge in the oxidation pond of a sewage treatment plant.

[0150] The dry weight of biomass is 478 mg / L, and the protein content in the biomass is 43.5%.

[0151] Example 5

[0152] A method for synchronous carbon sequestration and nitrogen removal, the steps are the same as those in Example 1, and the only difference is that: for ① the soil corresponding to the enrichment of hydrogen-oxidizing bacteria is the soil 2 - 10 cm below the surface layer of a paddy field for storing water. ② the soil corresponding to the enrichment of sulfur-oxidizing bacteria is the soil 2 - 10 cm below the surface layer of a marsh. ③ the soil corresponding to the enrichment of methanotrophs is the soil collected from the sludge in the oxidation pond of a sewage treatment plant.

[0153] The dry weight of biomass is 483 mg / L, and the protein content in the biomass is 46.8%.

[0154] Comparative Example 1

[0155] A method for synchronous carbon sequestration and nitrogen removal, the steps are the same as those in Example 1, and the only difference is that the enrichment and use of hydrogen-oxidizing bacteria are not carried out.

[0156] The final dry weight of biomass (i.e., cell dry weight) is 358 mg / L, and the protein content in the biomass is 42.7%. And during the cultivation process, CO2 will be produced due to the metabolism of methanotrophs, resulting in a reduction in the carbon sequestration effect.

[0157] Comparative Example 2

[0158] A method for synchronous carbon sequestration and nitrogen removal, the steps are the same as those in Example 1, and the only difference is that the enrichment and use of sulfur-oxidizing bacteria are not carried out. The cultivation process is inhibited by H2S in biogas, and the growth of microorganisms is severely affected.

[0159] The final biomass dry weight (i.e., cell dry weight) was 215 mg / L, and the protein content in the biomass was 15.2%.

[0160] Comparative Example 3

[0161] A method for synchronous carbon fixation and nitrogen removal, with the steps the same as in Example 1, and the only difference being that: the enrichment and use of methanotrophic bacteria were not carried out. Methane in the biogas was not fully utilized.

[0162] The final biomass dry weight (i.e., cell dry weight) was 362 mg / L, and the protein content in the biomass was 39.5%.

[0163] Example 6

[0164] A method for synchronous carbon fixation and nitrogen removal, with the steps the same as in Example 1, and the only difference being that: the composition of the first mixed gas of hydrogen-oxidizing bacteria was H2:O2:CO2 volume ratio = 50:30:20.

[0165] The final biomass dry weight (i.e., cell dry weight) was 462 mg / L, and the protein content in the biomass was 50.6%.

[0166] Example 7

[0167] A method for synchronous carbon fixation and nitrogen removal, with the steps the same as in Example 1, and the only difference being that: the composition of the first mixed gas of hydrogen-oxidizing bacteria was H2:O2:CO2 volume ratio = 60:25:15.

[0168] The final biomass dry weight (i.e., cell dry weight) was 471 mg / L, and the protein content in the biomass was 51.5%.

[0169] Example 8

[0170] A method for synchronous carbon fixation and nitrogen removal, with the steps the same as in Example 1, and the only difference being that: the composition of the first mixed gas of sulfur-oxidizing bacteria was H2S, H2:O2:CO2:H2S volume ratio: 30:20:20:20.

[0171] The final biomass dry weight (i.e., cell dry weight) was 427 mg / L, and the protein content in the biomass was 51.6%.

[0172] Example 9

[0173] A method for synchronous carbon fixation and nitrogen removal, with the steps the same as in Example 1, and the only difference being that: the composition of the first mixed gas of sulfur-oxidizing bacteria was H2S, H2:O2:CO2:H2S volume ratio: 50:30:30:30. The too high initial proportion of hydrogen sulfide may have a counterproductive effect and instead reduce the tolerance of sulfur-oxidizing bacteria.

[0174] The final biomass dry weight (i.e., cell dry weight) was 401.5 mg / L, and the protein content in the biomass was 49.5%.

[0175] Example 10

[0176] A method for synchronous carbon fixation and nitrogen removal, with the steps the same as in Example 1, and the only difference being that the composition of the first mixed gas of sulfur-oxidizing bacteria is H2S, and the volume ratio of H2:O2:CO2:H2S is 40:25:25:25.

[0177] The final biomass dry weight (i.e., cell dry weight) was 435 mg / L, and the protein content in the biomass was 50.4%.

[0178] Example 11

[0179] A method for synchronous carbon fixation and nitrogen removal, with the steps the same as in Example 1, and the only difference being that the first mixed gas of methanotrophic bacteria has a composition with a volume ratio of CH4:O2 of 50:50.

[0180] The final biomass dry weight (i.e., cell dry weight) was 436 mg / L, and the protein content in the biomass was 52.8%.

[0181] Example 12

[0182] A method for synchronous carbon fixation and nitrogen removal, with the steps the same as in Example 1, and the only difference being that the first mixed gas of methanotrophic bacteria has a composition with a volume ratio of CH4:O2 of 60:40.

[0183] The final biomass dry weight (i.e., cell dry weight) was 485 mg / L, and the protein content in the biomass was 55.4%.

[0184] Example 13

[0185] A method for enriching methanotrophic bacteria, with the steps the same as in Example 1, and the only difference being that the volume ratio of CH4 and O2 is 80:20.

[0186] Comparative Example 4

[0187] A method for enriching hydrogen-oxidizing bacteria, with the steps the same as in Example 1, and the only difference being that the composition of the first mixed gas of hydrogen-oxidizing bacteria is H2:O2:CO2 volume ratio = 50:20:30.

[0188] Comparative Example 5

[0189] A method for enriching hydrogen-oxidizing bacteria, with the steps the same as in Example 1, and the only difference being that the composition of the first mixed gas of hydrogen-oxidizing bacteria is H2:O2:CO2 volume ratio = 75:5:20.

[0190] Comparative Example 6

[0191] A method for enriching sulfur-oxidizing bacteria, the steps are the same as in Example 1, and the only difference is that the composition of the first mixed gas of sulfur-oxidizing bacteria is H2S, and the volume ratio of H2:O2:CO2:H2S is 30:20:20:40.

[0192] Comparative Example 7

[0193] A method for enriching sulfur-oxidizing bacteria, the steps are the same as in Example 1, and the only difference is that the composition of the first mixed gas of sulfur-oxidizing bacteria is H2S, and the volume ratio of H2:O2:CO2:H2S is 50:30:30:10.

[0194] Comparative Example 8

[0195] A method for enriching methane-oxidizing bacteria, the steps are the same as in Example 1, and the only difference is that the volume ratio of CH4 and O2 is 40:60.

[0196] Application Example 1

[0197] The time for enriching functional bacteria in Example 1 and Example 13 and Comparative Examples 4-8 is shown in Table 1. The time here is the time to obtain the corresponding enriched functional bacteria flora after subculture.

[0198] Table 1 Time for enriching functional bacteria in Example 1 and Example 13 and Comparative Examples 4-8

[0199]

[0200] In summary, the method for synchronous carbon fixation and nitrogen removal provided by the present invention enriches and screens a composite microbial metabolic group in which multiple microbial populations mainly composed of methane-oxidizing bacteria coexist to form a synergistic metabolic system. The synergistic metabolic system has H2S tolerance and can metabolize methane, carbon dioxide in biogas and N and P nutrients in biogas slurry to achieve the purpose of synchronous carbon fixation and nitrogen removal.

[0201] Although the above embodiments have made a detailed description of the present invention, they are only a part of the embodiments of the present invention, not all embodiments. People can also obtain other embodiments without creative efforts based on these embodiments, and these embodiments all belong to the protection scope of the present invention.

Claims

1. A method for simultaneous carbon fixation and nitrogen removal, characterized in that: The following steps are involved: The hydrogen-oxidizing bacteria mixed gas is used to enrich the microbial population dominated by hydrogen-oxidizing bacteria in the soil to obtain a hydrogen-oxidizing bacteria-enriched bacterial population; The sulfur-oxidizing bacteria mixed gas is used to enrich the microbial population dominated by sulfur-oxidizing bacteria in the soil to obtain a sulfur-oxidizing bacteria enriched bacterial population; The methane-oxidizing bacteria mixed gas is used to enrich the microbial population dominated by methane-oxidizing bacteria in the soil to obtain a methane-oxidizing bacteria-enriched bacterial population; The enriched bacterial groups of methane oxidizing bacteria, the enriched bacterial groups of sulfur oxidizing bacteria and the enriched bacterial groups of hydrogen oxidizing bacteria are mixed and inoculated into an acclimation medium for acclimation and culture to obtain a composite microbial metabolic group; The composite microbial metabolites are inoculated into an acclimated culture medium containing biogas and biogas is introduced to perform simultaneous carbon fixation and nitrogen removal; The hydrogen oxidizing bacteria mixed gas comprises H2, O2 and CO2, and the volume ratio of H2, O2 and CO2 is (50-70): (30-20): (20-10); The sulfur oxidizing bacteria mixed gas includes H2, O2, CO2 and H2S, and the volume ratio of H2, O2, CO2 and H2S is (30-50): (20-30): (20-30): (20-30); The methane oxidizing bacteria mixed gas includes CH4 and O2, and the volume ratio of CH4 to O2 is (50-80): (50-20); Methods for enriching the soil microbial population dominated by sulfur-oxidizing bacteria include: Inoculating soil microorganisms into a primary seed culture medium for sulfur-oxidizing bacteria and introducing a sulfur-oxidizing bacteria mixed gas for culturing until the H2S concentration no longer decreases, thereby obtaining a primary seed solution for sulfur-oxidizing bacteria; The primary seed liquid of sulfur-oxidizing bacteria is inoculated into the secondary seed culture medium of sulfur-oxidizing bacteria, and a sulfur-oxidizing bacteria mixed gas is introduced for culturing to obtain a sulfur-oxidizing bacteria enriched bacterial population; The composition of the primary seed culture medium for sulfur oxidizing bacteria includes: phenol red: 0.001-0.003 g / L, 1,4-piperazine diethanesulfonic acid: 4.5-8.5 g / L, sodium chloride: 10-25.000 g / L, magnesium sulfate heptahydrate: 1.1-2.9 g / L, potassium chloride: 0.1-1 g / L, ammonium chloride: 0.015-0.3 g / L, sodium thiosulfate pentahydrate: 1.5-3.2 g / L, calcium chloride dihydrate: 0.05-0.2 g / L, magnesium chloride hexahydrate: 2.5-4.5 g / L, potassium hydrogen phosphate: 0.05-0.15 g / L, ammonium ferrous sulfate hexahydrate: 0.001-0.002 g / L, trace element solution: 1 mL / L and vitamin solution: 10 mL / L; The composition of the secondary seed culture medium for sulfur-oxidizing bacteria is based on the primary seed culture medium for sulfur-oxidizing bacteria and further includes: 1% to 3% glucose, 0.6% to 2% yeast powder and 1.5% to 3.2% casein peptone; A mixed gas is introduced during the acclimation culture; the mixed gas is composed of H2, O2, CO2, CH4 and H2S; the volume ratio of H2, O2, CO2, CH4 and H2S is (0.1-1): (10-20): (15-25): (50-60): (0.1-0.3); The acclimation culture is continuously subcultured 3 to 4 times; the acclimation culture medium also includes Na2S and ammonium chloride; during the first acclimation culture, the mass concentration of Na2S in the acclimation culture medium is 1 to 2 mg / L, and the mass concentration of ammonium chloride is 100 to 300 mg / L; during each subculture, the mass concentration of Na2S and ammonium chloride in the acclimation culture medium is 1.5 to 2 times the mass concentration of Na2S and ammonium chloride in the previous culture.

2. The method according to claim 1, characterized in that: Soils rich in hydrogen oxidizing bacteria include soil 2 to 10 cm below the surface of waterlogged rice fields and / or compost residues of livestock and poultry manure; Soils rich in sulfur-oxidizing bacteria include 2-10 cm soil below the surface of marshes and / or sludge from oxidation ponds in sewage treatment plants; Soils enriched with methane-oxidizing bacteria include soil 2 to 10 cm below the surface of impounded rice fields and / or sludge from oxidation ponds in sewage treatment plants.

3. The method according to claim 1, characterized in that: Methods for enriching soil microbial populations dominated by hydrogen oxidizing bacteria include: Inoculate the microorganisms in the soil into the primary seed culture medium of hydrogen oxidizing bacteria and introduce the hydrogen oxidizing bacteria mixed gas for cultivation until the H2 consumption is more than 50% or the CO2 concentration no longer decreases, thereby obtaining the primary seed solution of hydrogen oxidizing bacteria; The hydrogen oxidizing bacteria primary seed liquid is inoculated into the hydrogen oxidizing bacteria secondary seed culture medium, and a hydrogen oxidizing bacteria mixed gas is introduced for culturing to obtain a hydrogen oxidizing bacteria enriched bacterial population; The composition of the primary seed culture medium for hydrogen oxidizing bacteria includes: Na2HPO4: 2200~2800 mg / L, KH2PO4: 500~1500mg / L, (NH4)2SO4: 450~500 mg / L, MgCl2: 45~60mg / L, CaCl2·2H2O: 40~50 mg / L, disodium EDTA: 0.2~0.7 mg / L, FeSO4·7H2O: 0.1~0.5 mg / L, ZnSO4·7H2O: 0.01~0.02 mg / L, MnCl2·4H2O: 0.002~0.003 mg / L, H3BO3: 0.01~0.04 mg / L, CoCl2·6H2O: 0.01~0.02 mg / L, CuCl2·2H2O: 0.001~0.01 mg / L, NiCl2·6H2O: 0.002~0.004 mg / L, Na2MoO4·2H2O: 0.001~0.003 mg / L; The composition of the hydrogen oxidizing bacteria secondary seed culture medium is based on the hydrogen oxidizing bacteria primary seed culture medium and further includes: 1%~3% glucose, 0.6%~2% yeast powder and 1.5%~3.2% casein peptone.

4. The method according to claim 1, characterized in that: Methods for enriching soil microbial populations dominated by methanotrophs include: Inoculating soil microorganisms into a primary seed culture medium for methane oxidizing bacteria and introducing a mixed gas of methane oxidizing bacteria for cultivation until the CH4 concentration no longer decreases, thereby obtaining a primary seed solution for methane oxidizing bacteria; The primary seed liquid of methanotrophic bacteria is inoculated into the secondary seed culture medium of methanotrophic bacteria, and a mixed gas of methanotrophic bacteria is introduced for cultivation to obtain an enriched bacterial population of methanotrophic bacteria; The composition of the primary seed culture medium for methanotrophic bacteria includes: KNO3: 100~200 mg / L, KH2PO4: 100~200 mg / L, MgSO4·7H2O: 25~55 mg / L, CaCl2·2H2O: 5~20 mg / L, EDTA: 2~5 mg / L, CuCl2·5H2O: 0.05~0.1 mg / L, FeSO4·7H2O: 1~2 mg / L, ZnSO4·7H2O: 0.1~0.2 mg / L, NiCl2·6H2O: 0.008~0.02 mg / L, CoCl2·6H2O: 0.1~0.2 mg / L and Na2MoO4: 0.01~0.03 mg / L; The composition of the secondary seed culture medium for methanotrophic bacteria is based on the primary seed culture medium for methanotrophic bacteria and further includes: 1% to 3% glucose, 0.6% to 2% yeast powder and 1.5% to 3.2% casein peptone.

5. The method according to claim 1, characterized in that: The composition of each 1L acclimatization medium includes: 0.5~1.00 g MgSO4·7H2O, 1.00~2.5 g KNO3, 0.15~0.3 g KH2PO4, 0.2~0.45 g Na2HPO4, 0.1~0.15 g CaCl2·2H2O and 1 mL trace element solution.

6. Application of the method according to any one of claims 1 to 5 in the resource utilization of biogas slurry and / or biogas.

7. Use of the method according to any one of claims 1 to 5 in preparing protein.

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