Domestication method for efficiently degrading tobacco waste to produce hydrogen by using mixed flora

Abstract

The invention discloses a domestication method for efficiently degrading tobacco waste to produce hydrogen by using mixed flora, which comprises the following steps: carrying out anaerobic enrichment domestication culture by taking manure compost as a bacterial source and cellulose prepared from tobacco waste as a core component of a domestication culture medium; the domestication culture medium contains cellulose, ammonium bicarbonate and monopotassium phosphate, and the cellulose concentration gradient is gradually increased without adjusting the pH value; stopping culturing when the hydrogen content in the biogas is reduced by 10%, transferring the biogas to a fresh domestication culture medium for passage according to an inoculum size of 10-20%, and continuously domesticating for 20-40 rounds; inoculating the domesticated mixed flora culture solution into a cellulose liquid fermentation culture medium containing microcrystalline cellulose, ammonium bicarbonate, monopotassium phosphate and a nutrient solution according to the inoculum size of 10-20%; the initial pH of the fermentation liquor is adjusted to 6.5-7.5, high-purity nitrogen is introduced to create an anaerobic environment, and hydrogen is produced through fermentation under the conditions of 35 + / -1 DEG C and 120 r / min. According to the method, flora is easy to culture, hydrogen production is efficient and stable, tobacco waste resources can be utilized, flora stability is high, adaptability is high, and strict sterile conditions are not needed.

Description

Technical Field

[0001] This invention relates to the fields of biotechnology and renewable energy technology, and in particular to a domestication method for efficiently degrading tobacco waste to produce hydrogen using mixed microbial communities. Background Technology

[0002] Hydrogen energy, as a clean, efficient, and low-carbon renewable resource, can achieve sustainable development of resources and the environment and has been widely used in many fields such as metallurgy, chemical industry, aerospace, food, and organic synthesis. Cellulose biomass is the most abundant and inexpensive renewable resource on Earth. Using cellulose biomass as a feedstock for bio-hydrogen production can achieve a high hydrogen production rate and reduce production costs, thus becoming a research hotspot in the field of bio-hydrogen production.

[0003] Traditional biohydrogen production technologies rely on pure bacterial fermentation, which suffers from a narrow substrate range, requires aseptic operation, and has low conversion efficiency. While mixed microbial fermentation can improve efficiency, the regulatory mechanisms of microbial community structure are unclear, making it difficult to optimize the synergistic effect between cellulose degradation and hydrogen production. In existing technologies, there is insufficient research on the impact of limiting substrates on microbial community structure and the synergistic optimization of hydrogen production process parameters, which restricts the application of large-scale hydrogen production from cellulosic biomass.

[0004] Therefore, this invention develops a domestication method for efficiently degrading hydrogen from tobacco waste using mixed microbial communities, and obtains cellulose-degrading microbial communities to achieve an efficient and stable cellulose hydrogen production pathway. Summary of the Invention

[0005] The technical problem to be solved by the present invention is to provide a domestication method for the efficient degradation of hydrogen from tobacco waste using mixed microbial communities, and to apply it to the degradation of cellulose for hydrogen production. This method effectively utilizes tobacco waste resources and improves the efficiency and stability of hydrogen production from cellulose degradation.

[0006] The technical problem to be solved by the present invention is achieved through the following technical solution:

[0007] A domestication method for efficiently degrading hydrogen from tobacco waste using mixed microbial communities includes the following steps:

[0008] (1) Using manure compost as the inoculum source and cellulose prepared from tobacco waste as the core component of the acclimatization culture medium, an anaerobic enrichment and acclimatization culture is carried out; the acclimatization culture medium is composed of the cellulose, ammonium bicarbonate and potassium dihydrogen phosphate, and the pH is not adjusted, and the concentration of the cellulose in the acclimatization culture medium is increased in a gradient manner.

[0009] (2) When the hydrogen content in the biogas produced by anaerobic enrichment and domestication culture decreases by 10%, stop the culture and transfer the culture medium to fresh domestication culture medium at an inoculation rate of 10-20% (V / V) for subculturing, and continue domestication for 20-40 rounds.

[0010] (3) The mixed bacterial culture obtained by domestication is inoculated into the cellulose liquid fermentation medium at an inoculation rate of 10-20% (V / V), wherein the cellulose liquid fermentation medium contains microcrystalline cellulose, ammonium bicarbonate, potassium dihydrogen phosphate and nutrient solution.

[0011] (4) Adjust the initial pH of the fermentation broth to 6.5-7.5, introduce high-purity nitrogen into the fermentation system to create an anaerobic environment, and control the temperature at 35±1℃ and the rotation speed at 120r / min to produce hydrogen through fermentation.

[0012] Preferably, in the above technical solution, in step (1), the manure compost is at least one of cow manure compost, horse manure compost, and sheep manure compost; the tobacco waste is at least one of broken tobacco leaves, broken tobacco stems, and tobacco dust.

[0013] Preferably, in the above technical solution, the preparation process of the cellulose in step (1) includes: crushing tobacco waste through a 100-200 mesh sieve, mixing it with the degradation liquid at a mass ratio of 1:8-12, heating and refluxing at 70-95℃ for 2-4 hours, then ultrasonically treating it with 400-800W power for 1-2 hours, and finally filtering, washing and drying.

[0014] Preferably, in the above technical solution, the degradation solution is prepared by mixing H2O2 solution and NaOH solution in a mass ratio of 1:(20-40), wherein the concentration of H2O2 solution is 10-30wt% and the concentration of NaOH solution is 2-4wt%.

[0015] Preferably, in the above technical solution, in step (1), the initial concentration of cellulose in the acclimatization culture medium is 0.5-10 g / L, the concentration of ammonium bicarbonate is 0.5-1.5 g / L, and the concentration of potassium dihydrogen phosphate is 0.8-2 g / L; the cellulose concentration increases by 0.5-2 g / L each time.

[0016] Preferably, in the above technical solution, the total duration of the acclimatization process in step (2) is 10-30 days.

[0017] Preferably, in the above technical solution, in step (3), the concentration of microcrystalline cellulose in the cellulose liquid fermentation medium is 6-10 g / L, the concentration of ammonium bicarbonate is 0.5-1.5 g / L, the concentration of potassium dihydrogen phosphate is 0.8-2 g / L, and the concentration of nutrient solution is 5-20 mL / L.

[0018] Preferably, in the above technical solution, the nutrient solution is composed of ferrous chloride, sodium chloride, magnesium sulfate heptahydrate, calcium chloride dihydrate, sodium molybdate dihydrate, and manganese sulfate heptahydrate; wherein the concentration of ferrous chloride is 0.00278 g / L, the concentration of sodium chloride is 0.01-0.03 g / L, the concentration of magnesium sulfate heptahydrate is 0.1-0.2 g / L, the concentration of calcium chloride dihydrate is 0.01-0.05 g / L, the concentration of sodium molybdate dihydrate is 0.01-0.04 g / L, and the concentration of manganese sulfate heptahydrate is 0.015 g / L.

[0019] Application of a mixed microbial community obtained through the above-mentioned domestication method in the degradation of tobacco waste to produce hydrogen.

[0020] The above-described technical solution of the present invention has the following beneficial effects:

[0021] (1) Fermentation using mixed microbial communities is easy to cultivate and can efficiently and stably produce hydrogen.

[0022] (2) It can effectively utilize tobacco waste resources.

[0023] (3) The microbial community has high stability and strong adaptability, and does not require strict aseptic conditions. Attached Figure Description

[0024] The accompanying drawings, which are incorporated in and form part of this specification, illustrate embodiments of the invention and, together with their description, serve to explain the principles of the invention.

[0025] Figure 1 The hydrogen production, cellulase activity, and cellulose degradation rate of microbial communities at different domestication stages were measured.

[0026] Figure 2 The distribution of microbial communities at the phylum level before and after domestication.

[0027] Figure 3 The distribution of microbial communities at the genus level before and after domestication. Detailed Implementation

[0028] Various exemplary embodiments of the present invention will now be described in detail with reference to the accompanying drawings. It should be noted that, unless otherwise specifically stated, the relative arrangement, numerical expressions, and values ​​of the components and steps set forth in these embodiments do not limit the scope of the invention.

[0029] Unless otherwise specified, the experimental methods used in the following examples are conventional methods, and the materials and reagents used are commercially available. Unless otherwise specified, the equipment used in the experiments is well known to those skilled in the art.

[0030] A domestication method for efficiently degrading hydrogen from tobacco waste using mixed microbial communities includes the following steps:

[0031] (1) Using cow manure compost, horse manure compost or sheep manure compost as the inoculum source and cellulose as the acclimatization culture medium, anaerobic enrichment and acclimatization culture is carried out.

[0032] (2) The cellulose is prepared from tobacco resources consisting of one or more combinations of shredded tobacco leaves, shredded tobacco stems, or tobacco dust. The preparation method is as follows: the raw material, consisting of one or more combinations of shredded tobacco leaves, shredded tobacco stems, or tobacco dust, is mixed with a degradation solution and heated under reflux for 2-4 hours at a temperature of 70-95°C. After reflux, the mixture is obtained through ultrasonication, filtration, washing, and drying. The tobacco resources are obtained by pulverizing and sieving, with a sieve mesh size of 100-200 mesh. The degradation solution is prepared by mixing H₂O₂ solution and NaOH solution at a mass ratio of 1:20-40 and stirring until homogeneous. The mass ratio of tobacco resources to the degradation solution is 1:8-12, the concentration of NaOH solution is 2-4 wt%, and the concentration of H₂O₂ solution is 10-30 wt%. The ultrasonic treatment step is performed at a power of 400-800 W for 1-2 hours.

[0033] (3) The acclimatization culture medium consists of cellulose, ammonium bicarbonate, and potassium dihydrogen phosphate, wherein the concentration of cellulose is 0.5-10 g / L, the concentration of ammonium bicarbonate is 0.5-1.5 g / L, and the concentration of potassium dihydrogen phosphate is 0.8-2 g / L. The pH of the acclimatization culture medium is not adjusted. The concentration of cellulose in the acclimatization culture medium is increased gradually in increments of 0.5-2 g / L.

[0034] (4) When the hydrogen content in the biogas decreases, stop the culture. The decrease is 10%.

[0035] (5) Transfer the culture medium to fresh enrichment and acclimatization medium at an inoculation rate of 10-20% (V / V). Each subculture and transfer is one round of acclimatization. The entire enrichment and acclimatization process lasts for 20-40 rounds and the acclimatization time is 10-30 days.

[0036] (6) The culture medium of the mixed microbial community obtained by domestication is inoculated into the cellulose liquid fermentation medium at an inoculation amount of 10-20% (V / V). The cellulose fermentation medium is composed of microcrystalline cellulose, ammonium bicarbonate, potassium dihydrogen phosphate and nutrient solution, wherein the concentration of microcrystalline cellulose is 6-10 g / L, the concentration of ammonium bicarbonate is 0.5-1.5 g / L, the concentration of potassium dihydrogen phosphate is 0.8-2 g / L and the concentration of nutrient solution is 5-20 mL / L.

[0037] (7) The nutrient solution in the cellulose fermentation medium is composed of ferrous chloride, sodium chloride, magnesium sulfate heptahydrate, calcium chloride dihydrate, sodium molybdate dihydrate and manganese sulfate heptahydrate, wherein the concentration of ferrous chloride is 0.00278 g / L, the concentration of sodium chloride is 0.01-0.03 g / L, the concentration of magnesium sulfate heptahydrate is 0.1-0.2 g / L, the concentration of calcium chloride dihydrate is 0.01-0.05 g / L, the concentration of sodium molybdate dihydrate is 0.01-0.04 g / L and the concentration of manganese sulfate heptahydrate is 0.015 g / L.

[0038] (8) Adjust the initial pH of the fermentation broth to the range of 6.5-7.5, and introduce high-purity nitrogen into the fermenter to create an anaerobic environment. The fermentation conditions are: temperature controlled at 35±1℃ and rotation speed of 120r / min for hydrogen production.

[0039] Example 1

[0040] Domestication of hydrogen-producing bacteria:

[0041] (1) Anaerobic enrichment and domestication culture was carried out using cow manure compost as the inoculum source and cellulose as the domestication culture medium.

[0042] (2) Cellulose is prepared from shredded tobacco flakes. The preparation method is as follows: the shredded tobacco flakes are mixed with the degradation solution, heated and refluxed for 2 hours at a temperature of 70°C. After reflux, the cellulose is obtained by ultrasonication, filtration, washing and drying.

[0043] The tobacco resources were obtained by crushing and sieving with a 100-mesh sieve. The degradation solution was prepared by mixing H2O2 solution and NaOH solution at a mass ratio of 1:20 and stirring until homogeneous.

[0044] The mass ratio of tobacco resources to degradation liquid was 1:9, the concentration of NaOH solution was 2wt%, and the concentration of H2O2 solution was 12wt%. The ultrasonic treatment step was performed at a power of 500W for 2 hours.

[0045] (3) The acclimatization medium consists of cellulose, ammonium bicarbonate, and potassium dihydrogen phosphate, with a cellulose concentration of 3 g / L, an ammonium bicarbonate concentration of 0.5 g / L, and a potassium dihydrogen phosphate concentration of 1 g / L. The pH of the acclimatization medium is not adjusted. The concentration of cellulose in the acclimatization medium is increased gradually in increments of 0.5 g / L.

[0046] (4) When the hydrogen content in the biogas decreases, stop the culture. The decrease is 10%.

[0047] (5) Transfer the culture medium to fresh enrichment and acclimatization medium at an inoculation rate of 10% (V / V). Each subculture and transfer is one round of acclimatization. The entire enrichment and acclimatization process lasts for 20 rounds and the acclimatization time is 10 days.

[0048] Validation of hydrogen production by the domesticated bacterial community:

[0049] (1) The culture medium of the mixed microbial community obtained by domestication was inoculated into the cellulose liquid fermentation medium at an inoculation amount of 10% (V / V). The cellulose fermentation medium was composed of microcrystalline cellulose, ammonium bicarbonate, potassium dihydrogen phosphate and nutrient solution, wherein the concentration of microcrystalline cellulose was 3 g / L, the concentration of ammonium bicarbonate was 0.5 g / L, the concentration of potassium dihydrogen phosphate was 1 g / L and the concentration of nutrient solution was 5 mL / L.

[0050] (2) The nutrient solution in the cellulose fermentation medium is composed of ferrous chloride, sodium chloride, magnesium sulfate heptahydrate, calcium chloride dihydrate, sodium molybdate dihydrate and manganese sulfate heptahydrate, wherein the concentration of ferrous chloride is 0.00278 g / L, the concentration of sodium chloride is 0.01 g / L, the concentration of magnesium sulfate heptahydrate is 0.12 g / L, the concentration of calcium chloride dihydrate is 0.03 g / L, the concentration of sodium molybdate dihydrate is 0.01 g / L and the concentration of manganese sulfate heptahydrate is 0.015 g / L.

[0051] (3) Adjust the initial pH of the fermentation broth to 6.5, and introduce high-purity nitrogen into the fermenter to create an anaerobic environment. The fermentation conditions are: temperature controlled at 35±1℃ and rotation speed at 120r / min for hydrogen production.

[0052] Under these conditions, the maximum cumulative hydrogen production can reach 243.8 mL / g.

[0053] Example 2

[0054] Domestication of hydrogen-producing bacteria:

[0055] (1) Using horse manure compost as the microbial source and cellulose as the acclimatization culture medium, anaerobic enrichment and acclimatization culture was carried out.

[0056] (2) The cellulose is prepared from chopped tobacco stems. The preparation method is as follows: chopped tobacco stems are mixed with degradation solution, heated and refluxed for 3 hours at a temperature of 85°C. After reflux, the cellulose is obtained through ultrasonication, filtration, washing, and drying. The tobacco resources are obtained by crushing and sieving with a 150-mesh sieve. The degradation solution is prepared by mixing H₂O₂ solution and NaOH solution at a mass ratio of 1:30 and stirring until homogeneous. The mass ratio of tobacco resources to degradation solution is 1:11, the concentration of NaOH solution is 3 wt%, and the concentration of H₂O₂ solution is 20 wt%. The ultrasonic treatment step is performed at a power of 600W for 1 hour.

[0057] (3) The acclimatization culture medium consists of cellulose, ammonium bicarbonate, and potassium dihydrogen phosphate, wherein the concentration of cellulose is 7 g / L, the concentration of ammonium bicarbonate is 1.3 g / L, and the concentration of potassium dihydrogen phosphate is 1.6 g / L. The pH of the acclimatization culture medium is not adjusted. The concentration of cellulose in the acclimatization culture medium is increased gradually in increments of 1 g / L.

[0058] (4) When the hydrogen content in the biogas decreases, stop the culture. The decrease is 10%.

[0059] (5) Transfer the culture medium to fresh enrichment and acclimatization medium at an inoculation rate of 15% (V / V). Each subculture and transfer is one round of acclimatization. The entire enrichment and acclimatization process lasts for 30 rounds and the acclimatization time is 25 days.

[0060] Validation of hydrogen production by the domesticated bacterial community:

[0061] (1) The culture medium of the mixed microbial community obtained by domestication was inoculated into the cellulose liquid fermentation medium at an inoculation amount of 15% (V / V). The cellulose fermentation medium consisted of microcrystalline cellulose, ammonium bicarbonate, potassium dihydrogen phosphate and nutrient solution, wherein the concentration of microcrystalline cellulose was 7 g / L, the concentration of ammonium bicarbonate was 1.3 g / L, the concentration of potassium dihydrogen phosphate was 1.6 g / L and the concentration of nutrient solution was 10 mL / L.

[0062] (2) The nutrient solution in the cellulose fermentation medium is composed of ferrous chloride, sodium chloride, magnesium sulfate heptahydrate, calcium chloride dihydrate, sodium molybdate dihydrate and manganese sulfate heptahydrate, wherein the concentration of ferrous chloride is 0.00278 g / L, the concentration of sodium chloride is 0.02 g / L, the concentration of magnesium sulfate heptahydrate is 0.16 g / L, the concentration of calcium chloride dihydrate is 0.04 g / L, the concentration of sodium molybdate dihydrate is 0.03 g / L and the concentration of manganese sulfate heptahydrate is 0.015 g / L.

[0063] (3) Adjust the initial pH of the fermentation broth to 7.0, and introduce high-purity nitrogen into the fermenter to create an anaerobic environment. The fermentation conditions are: temperature controlled at 35±1℃ and rotation speed at 120r / min for hydrogen production.

[0064] Under these conditions, the maximum cumulative hydrogen production can reach 253.9 mL / g.

[0065] Example 3

[0066] Domestication of hydrogen-producing bacteria:

[0067] (1) Anaerobic enrichment and domestication culture was carried out using cow manure compost and horse manure compost as inoculum sources and cellulose as domestication culture medium.

[0068] (2) The cellulose is prepared from tobacco dust. The preparation method is as follows: tobacco dust is mixed with degradation solution, heated and refluxed for 4 hours at a temperature of 95°C. After reflux, it is obtained through ultrasonication, filtration, washing, and drying. The tobacco resources are obtained by crushing and sieving with a 200-mesh sieve. The degradation solution is prepared by mixing H2O2 solution and NaOH solution at a mass ratio of 1:40 and stirring until homogeneous. The mass ratio of tobacco resources to degradation solution is 1:12, the concentration of NaOH solution is 4wt%, and the concentration of H2O2 solution is 30wt%. The ultrasonic treatment step is performed at 800W for 1 hour.

[0069] (3) The acclimatization culture medium consists of cellulose, ammonium bicarbonate, and potassium dihydrogen phosphate, wherein the concentration of cellulose is 8 g / L, the concentration of ammonium bicarbonate is 1.5 g / L, and the concentration of potassium dihydrogen phosphate is 2 g / L. The pH of the acclimatization culture medium is not adjusted. The concentration of cellulose in the acclimatization culture medium is increased gradually in increments of 2 g / L.

[0070] (4) When the hydrogen content in the biogas decreases, stop the culture. The decrease is 10%.

[0071] (5) Transfer the culture medium to fresh enrichment and acclimatization medium at an inoculation rate of 20% (V / V). Each subculture and transfer is one round of acclimatization. The entire enrichment and acclimatization process lasts for 40 rounds and the acclimatization time is 30 days.

[0072] Validation of hydrogen production by the domesticated bacterial community:

[0073] (1) The culture medium of the mixed microbial community obtained by domestication was inoculated into the cellulose liquid fermentation medium at an inoculation amount of 20% (V / V). The cellulose fermentation medium consisted of microcrystalline cellulose, ammonium bicarbonate, potassium dihydrogen phosphate and nutrient solution, wherein the concentration of microcrystalline cellulose was 8 g / L, the concentration of ammonium bicarbonate was 1.5 g / L, the concentration of potassium dihydrogen phosphate was 2 g / L and the concentration of nutrient solution was 20 mL / L.

[0074] (2) The nutrient solution in the cellulose fermentation medium is composed of ferrous chloride, sodium chloride, magnesium sulfate heptahydrate, calcium chloride dihydrate, sodium molybdate dihydrate and manganese sulfate heptahydrate, wherein the concentration of ferrous chloride is 0.00278 g / L, the concentration of sodium chloride is 0.03 g / L, the concentration of magnesium sulfate heptahydrate is 0.2 g / L, the concentration of calcium chloride dihydrate is 0.05 g / L, the concentration of sodium molybdate dihydrate is 0.04 g / L and the concentration of manganese sulfate heptahydrate is 0.015 g / L.

[0075] (3) Adjust the initial pH of the fermentation broth to 7.5, and introduce high-purity nitrogen into the fermenter to create an anaerobic environment. The fermentation conditions are: temperature controlled at 35±1℃ and rotation speed at 120r / min for hydrogen production.

[0076] Under these conditions, the maximum cumulative hydrogen production can reach 265.7 mL / g.

[0077] Although the present invention has been disclosed above with reference to embodiments, it is not intended to limit the present invention. Any person skilled in the art can make various different choices and modifications without departing from the spirit and scope of the present invention. Therefore, the scope of protection of the present invention is defined by the claims and their equivalents.

Claims

1. A domestication method for efficiently degrading hydrogen from tobacco waste using mixed microbial communities, characterized in that, Includes the following steps: (1) Using manure compost as the inoculum source and cellulose prepared from tobacco waste as the core component of the acclimatization culture medium, an anaerobic enrichment and acclimatization culture is carried out; the acclimatization culture medium is composed of the cellulose, ammonium bicarbonate and potassium dihydrogen phosphate, and the pH is not adjusted, and the concentration of the cellulose in the acclimatization culture medium is increased in a gradient manner. (2) When the hydrogen content in the biogas produced by anaerobic enrichment and domestication culture decreases by 10%, stop the culture and transfer the culture medium to fresh domestication culture medium at an inoculation rate of 10-20% (V / V) for subculturing, and continue domestication for 20-40 rounds. (3) The mixed bacterial culture obtained by domestication is inoculated into the cellulose liquid fermentation medium at an inoculation rate of 10-20% (V / V), wherein the cellulose liquid fermentation medium contains microcrystalline cellulose, ammonium bicarbonate, potassium dihydrogen phosphate and nutrient solution. (4) Adjust the initial pH of the fermentation broth to 6.5-7.5, introduce high-purity nitrogen into the fermentation system to create an anaerobic environment, and control the temperature at 35±1℃ and the rotation speed at 120r / min to produce hydrogen through fermentation.

2. The domestication method according to claim 1, characterized in that, In step (1), the manure compost is at least one of cow manure compost, horse manure compost, and sheep manure compost; the tobacco waste is at least one of broken tobacco leaves, broken tobacco stems, and tobacco dust.

3. The domestication method according to claim 1, characterized in that, In step (1), the preparation process of the cellulose includes: crushing tobacco waste through a 100-200 mesh sieve, mixing it with the degradation liquid at a mass ratio of 1:8-12, heating and refluxing at 70-95℃ for 2-4 hours, then ultrasonically treating it with 400-800W power for 1-2 hours, and finally filtering, washing and drying.

4. The domestication method according to claim 3, characterized in that, The degradation solution is prepared by mixing H2O2 solution and NaOH solution in a mass ratio of 1:(20-40), wherein the concentration of H2O2 solution is 10-30wt% and the concentration of NaOH solution is 2-4wt%.

5. The domestication method according to claim 1, characterized in that, In step (1), the initial concentration of cellulose in the acclimatization culture medium is 0.5-10 g / L, the concentration of ammonium bicarbonate is 0.5-1.5 g / L, and the concentration of potassium dihydrogen phosphate is 0.8-2 g / L; the cellulose concentration increases in increments of 0.5-2 g / L each time.

6. The domestication method according to claim 1, characterized in that, In step (2), the total duration of the acclimatization process is 10-30 days.

7. The domestication method according to claim 1, characterized in that, In step (3), the concentration of microcrystalline cellulose in the cellulose liquid fermentation medium is 6-10 g / L, the concentration of ammonium bicarbonate is 0.5-1.5 g / L, the concentration of potassium dihydrogen phosphate is 0.8-2 g / L, and the concentration of nutrient solution is 5-20 mL / L.

8. The domestication method according to claim 7, characterized in that, The nutrient solution is composed of ferrous chloride, sodium chloride, magnesium sulfate heptahydrate, calcium chloride dihydrate, sodium molybdate dihydrate, and manganese sulfate heptahydrate; wherein the concentration of ferrous chloride is 0.00278 g / L, the concentration of sodium chloride is 0.01-0.03 g / L, the concentration of magnesium sulfate heptahydrate is 0.1-0.2 g / L, the concentration of calcium chloride dihydrate is 0.01-0.05 g / L, the concentration of sodium molybdate dihydrate is 0.01-0.04 g / L, and the concentration of manganese sulfate heptahydrate is 0.015 g / L.

9. The application of a mixed microbial community obtained by any one of the domestication methods described in claims 1-8 in the degradation of tobacco waste to produce hydrogen.

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