A composition, method and use thereof for alleviating ammonia inhibition in anaerobic fermentation

Abstract

The application provides a composition and method for relieving anaerobic fermentation ammonia nitrogen inhibition and application thereof, and belongs to the technical field of microbial fermentation.The composition for relieving anaerobic fermentation ammonia nitrogen inhibition comprises the following raw materials in parts by weight: 8-10 parts of saccharomyces cerevisiae, 5-8 parts of mucor circinelloides, 5-8 parts of pseudomonas putida, 3-4 parts of bacillus licheniformis and 10-15 parts of alfalfa extract.The symbiotic and intergrowth relationship among different strains in the application enhances the adaptability of microorganisms to environmental changes, the alfalfa extract provides additional nutrients and growth factors for microorganisms, helps maintain the diversity and stability of the microbial community, and ensures the continuous and stable operation of anaerobic fermentation under adversity such as ammonia nitrogen inhibition.The method for relieving anaerobic fermentation ammonia nitrogen inhibition can improve the methane production, speed up the entire anaerobic fermentation process, and improve the system efficiency and stability by using the composition.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of microbial fermentation, and particularly relates to a composition for relieving ammonia nitrogen inhibition in anaerobic fermentation, a method and application thereof. BACKGROUND

[0002] Anaerobic fermentation, as an environmentally friendly method for organic waste treatment, is attracting more and more attention. It not only realizes the resource utilization of organic waste, such as biological fuel gas, but also effectively reduces the pollution of waste to the environment. Especially in the treatment of livestock manure and other nitrogen-rich organic waste, anaerobic fermentation process has significant advantages, providing a feasible way to solve the problem of livestock waste treatment.

[0003] However, in practical application, anaerobic fermentation process faces many challenges, among which the ammonia nitrogen inhibition problem is particularly prominent. Taking livestock manure as an example, a large amount of organic nitrogen in this kind of nitrogen-rich organic waste will be decomposed and converted into ammonia nitrogen by microorganisms during anaerobic fermentation. When the ammonia nitrogen concentration exceeds a certain threshold, it will have a serious inhibitory effect on the activity of anaerobic microorganisms. Studies have shown that ammonia toxicity, especially the toxicity of free ammonia (NH3), can cause more than 30% loss of methane production, and even cause the collapse of anaerobic fermentation reactor, making the whole treatment system unable to operate normally.

[0004] To solve the problem of ammonia nitrogen inhibition in anaerobic fermentation process, many studies have been carried out and various methods have been proposed. By adding exogenous substances to remove ammonia nitrogen, a large amount of expensive chemicals is consumed, a large amount of chemical sludge needs to be disposed of, and ions that interfere with fermentation may be introduced. By adding acid (such as hydrochloric acid, sulfuric acid) to reduce the pH of the system, the toxic free ammonia (NH3) is converted into ammonium ion (NH4 + ), which needs to consume a large amount of acid, significantly increasing the operating cost, and frequent pH fluctuations will also inhibit microbial activity, making it difficult to maintain system stability. Biological reinforcement using the microorganism's own ability is considered a more environmentally friendly and sustainable potential solution, but existing microbial strategies (especially single strain addition) have core defects such as single function of strains, lack of multi-microbial population synergy, low colonization efficiency in complex and harsh fermentation environment, poor adaptability and stability, and insufficient universality. It is difficult to meet the actual needs of quickly, efficiently, stably, economically and simply operating to restore and maintain the performance of anaerobic fermentation system in a high ammonia nitrogen environment. SUMMARY

[0005] Therefore, one of the purposes of the present application is to provide a composition for relieving ammonia nitrogen inhibition in anaerobic fermentation.

[0006] The second purpose of the present application is to provide the application of the composition in relieving and / or relieving ammonia nitrogen inhibition in anaerobic fermentation.

[0007] The third object of the present application is to provide a method for relieving the inhibition of anaerobic fermentation by ammonia nitrogen.

[0008] The fourth object of the present application is to provide the use of the composition or the method in promoting the decomposition of anaerobic fermentation substrate, increasing the production of methane, and increasing the production of volatile organic acids.

[0009] In order to achieve the above-mentioned objects, the present application provides the following technical solutions.

[0010] A composition for relieving the inhibition of anaerobic fermentation by ammonia nitrogen, comprising the following raw materials in parts by weight: 8-10 parts of Saccharomyces cerevisiae, 5-8 parts of Mucor circinelloides, 5-8 parts of Pseudomonas putida, 3-4 parts of Bacillus licheniformis, and 10-15 parts of Medicago sativa extract.

[0011] Preferably, the preparation method of the Medicago sativa extract comprises the following steps: taking fresh Medicago sativa, drying and crushing, mixing the powder with water, and then extracting at 40-50℃, collecting the extract liquid by filtration, and drying the concentrated extract liquid to obtain the Medicago sativa extract.

[0012] The present application also provides the use of the composition in relieving and / or alleviating the inhibition of anaerobic fermentation by ammonia nitrogen.

[0013] The present application also provides a method for relieving the inhibition of anaerobic fermentation by ammonia nitrogen, comprising the following steps: activating the composition, inoculating the activated composition into a fermentation substrate, and performing anaerobic fermentation.

[0014] Preferably, the mass ratio of the composition to the total solid of the fermentation substrate is (0.01-1.5):1.

[0015] Preferably, during the anaerobic fermentation, the total solid concentration in the fermentation substrate is 4%-30%, the temperature of the anaerobic fermentation is 30-55℃, and the pH of the anaerobic fermentation system is 6.5-7.5.

[0016] Preferably, the activation method of the composition comprises the following steps: weighing the raw materials in proportion, respectively inoculating the microorganisms into an activation culture medium, performing a first shock culture, obtaining four preliminary activated bacterial liquids after the culture, mixing the four preliminary activated bacterial liquids in a volume ratio of 1:1:1:1, adding the activation culture medium, performing a second shock culture, adding the Medicago sativa extract after the culture, and obtaining the activated composition.

[0017] Preferably, the first shock culture is performed under the following conditions: the culture temperature is 28-32℃, the culture time is 24-48h, and the rotation speed is 150-200rpm.

[0018] Preferably, the second shock culture conditions are as follows: the culture temperature is 28-32 DEG C, the culture time is 6-12h, and the rotation speed is 100-120rpm.

[0019] The application further provides application of the composition or the method in promoting anaerobic fermentation substrate decomposition, increasing methane production and increasing volatile organic acid production.

[0020] Compared with the prior art, the application has the following beneficial effects:

[0021] The application provides a composition for relieving ammonia nitrogen inhibition in anaerobic fermentation, and the symbiotic and intergrowth relationship among different strains in the application enhances the adaptability of microorganisms to environmental changes, the alfalfa extract provides additional nutrients and growth factors for the microorganisms, helps maintain the diversity and stability of the microbial community, and guarantees the continuous and stable operation of the anaerobic fermentation under adversity such as ammonia nitrogen inhibition. The complex microorganisms and the alfalfa extract can reduce the amount of ammonia nitrogen in the anaerobic fermentation system by synergistic action and self metabolism, and relieve the toxicity of ammonia nitrogen to the microorganisms. The alfalfa extract is rich in various active ingredients, can regulate the permeability of the microbial cell membrane, enhances the ability of the microorganisms to resist ammonia nitrogen stress, and further strengthens the relieving effect on ammonia nitrogen inhibition in synergistic action with the complex microorganisms.

[0022] The method for relieving ammonia nitrogen inhibition in anaerobic fermentation utilizes the composition of the application to perform anaerobic fermentation, the complex microorganisms can secrete various extracellular enzymes, accelerate the hydrolysis and acidification process of organic matter, convert complex macromolecular organic matter into small-molecule volatile fatty acids (VFAs), provide sufficient substrate for the subsequent methane production stage, and promote the material conversion of the anaerobic fermentation. The bioactive substances in the alfalfa extract can also stimulate the activity of methanogens, increase methane production, speed up the entire anaerobic fermentation process, and improve the system efficiency and stability.

[0023] The raw materials of the composition of the application are easy to obtain and cultivate, are environment-friendly, can effectively relieve ammonia nitrogen inhibition and reduce treatment cost when applied in the field of anaerobic fermentation on a large scale, and have extremely high popularization and application value. DETAILED DESCRIPTION

[0024] The application provides a composition for relieving ammonia nitrogen inhibition in anaerobic fermentation, and the composition comprises the following raw materials in parts by weight: 8-10 parts of saccharomyces cerevisiae, 5-8 parts of mucor circinelloides, 5-8 parts of pseudomonas putida, 3-4 parts of bacillus licheniformis and 10-15 parts of alfalfa extract.

[0025] The amount of the Saccharomyces cerevisiae in the composition is preferably 9 parts; the amount of the Mucor circinelloides is preferably 6-7 parts; the amount of the Pseudomonas putida is preferably 6-7 parts; the amount of the Bacillus licheniformis is preferably 3.5 parts; and the amount of the Medicago sativa extract is preferably 12-14 parts, and more preferably 13 parts. 7 ~10 8 cfu / g.

[0026] In the present application, the preparation method of the Medicago sativa extract preferably comprises the following steps: taking fresh Medicago sativa, drying and crushing, mixing the powder with water, extracting at 40-50℃, collecting the extract liquid by filtration, drying after concentration, and obtaining the Medicago sativa extract.

[0027] In the preparation method of the Medicago sativa extract, the drying is preferably performed at 40-50℃, and the drying time is preferably 3-5h; after drying, crushing is performed, preferably to 40-60 mesh; the powder is mixed with water at a solid-liquid ratio of 1g: (10-15)mL; the extraction temperature is preferably 43-48℃, and more preferably 45℃; and the extraction time is preferably 2-4h, and more preferably 2.5-3.5h. The concentration method and the drying method are not limited in the present application, and the conventional methods in the art can be adopted.

[0028] The Medicago sativa extract in the present application provides additional nutrients and growth factors for microorganisms, helps to maintain the diversity and stability of the microbial community, ensures the continuous and stable operation of anaerobic fermentation under adversity such as ammonia nitrogen inhibition, stimulates the activity of methanogenic bacteria, improves the methane production, speeds up the entire anaerobic fermentation process, and improves the system efficiency and stability.

[0029] The present application also provides the use of the composition in relieving and / or alleviating the ammonia nitrogen inhibition in anaerobic fermentation.

[0030] The composition of the present application can reduce the amount of ammonia nitrogen in the anaerobic fermentation system through synergistic effect and its own metabolism, reduce its toxicity to microorganisms, provide sufficient substrate for the subsequent methanogenesis stage, and promote the material conversion of anaerobic fermentation.

[0031] The present application also provides a method for relieving the ammonia nitrogen inhibition in anaerobic fermentation, comprising the following steps: activating the composition, inoculating the activated composition into the fermentation substrate, and performing anaerobic fermentation.

[0032] In the method for alleviating ammonia nitrogen inhibition in anaerobic fermentation, the mass ratio of the composition to the total solids of the fermentation substrate is preferably (0.01-1.5):1, more preferably (0.05-1):1, and further preferably (0.06-0.08):1, based on the mass of the unactivated composition. In the anaerobic fermentation, the total solids concentration in the fermentation substrate is preferably 4%-30% (w / w), and more preferably 15%-20%; the temperature of the anaerobic fermentation is preferably 30-55°C, more preferably 35-45°C, and further preferably 38-42°C; and the pH of the anaerobic fermentation system is preferably 6.5-7.5.

[0033] In the method for alleviating ammonia nitrogen inhibition in anaerobic fermentation, the activation method of the composition preferably comprises: weighing each raw material in proportion; introducing each microorganism into an activation culture medium, respectively, for first shock culture, and obtaining four preliminary activated bacterial liquids after the culture; mixing the four preliminary activated bacterial liquids in a volume ratio of 1:1:1:1, adding the activation culture medium, and performing second shock culture; and adding alfalfa extract after the culture to obtain the activated composition. As an implementable manner, the composition of the activation culture medium is: glucose 10 g / L, peptone 5 g / L, yeast extract 3 g / L, potassium dihydrogen phosphate 1 g / L, and magnesium sulfate 0.5 g / L; after dissolving each component with distilled water, the pH is adjusted to 6.5-7.0, and sterilization is performed at 121°C for 20 min.

[0034] In the activation method of the composition, the inoculation amount of each microorganism is 3%-5% (w / v); the volume ratio of the mixed liquid of the four preliminary activated bacterial liquids to the added activation culture medium is preferably 1:(4-6), and more preferably 1:5; the temperature of the first shock culture is preferably 28-32°C, and more preferably 30°C; the culture time is preferably 24-48 h, and more preferably 30-40 h; and the rotation speed during the culture is preferably 150-200 rpm, and more preferably 180 rpm. The temperature of the second shock culture is preferably 28-32°C, and more preferably 30°C; the culture time is preferably 6-12 h, and more preferably 8-10 h; and the rotation speed during the culture is preferably 100-120 rpm, and more preferably 110 rpm.

[0035] The activated composition has strong stability, can quickly colonize when inoculated into the fermentation substrate, develop into a dominant flora, reduce the ammonia nitrogen content in the fermentation system through synergistic action, convert complex macromolecular organic matter into small-molecule volatile fatty acids (VFAs), and provide sufficient substrate for the subsequent methanogenesis stage, thereby promoting the material conversion in the anaerobic fermentation.

[0036] The application also provides application of the composition or the method in promoting anaerobic fermentation substrate decomposition, increasing methane production and increasing volatile organic acid production. The application can be used for anaerobic fermentation treatment of high-nitrogen organic matter such as livestock and poultry breeding manure, kitchen waste, municipal sludge and the like, promotes fermentation substrate decomposition, and also provides methane and volatile organic acid, thereby providing a feasible solution for waste treatment problems.

[0037] The technical solutions provided by the application are described in detail below in combination with examples, but they should not be understood as limitations to the protection scope of the application.

[0038] Example 1

[0039] A composition for relieving ammonia nitrogen inhibition in anaerobic fermentation, raw materials are as follows in terms of weight fraction:

[0040] Saccharomyces cerevisiae 9 parts, Mucor circinelloides 6 parts, Pseudomonas putida 6 parts, Bacillus licheniformis 3.5 parts and alfalfa extract 13 parts. The viable cell count of each microorganism is 10 7 ~10 8 cfu / g.

[0041] Preparation method of the alfalfa extract:

[0042] Fresh alfalfa is dried at 45℃ for 5h, crushed to pass through a 60-mesh sieve, mixed with water at a solid-liquid ratio of 1g:12mL, extracted at 45℃ for 3.5h, filtered to collect the extract, and the filter residue is extracted again once, the extract is combined, concentrated and dried to obtain the alfalfa extract.

[0043] Example 2

[0044] A composition for relieving ammonia nitrogen inhibition in anaerobic fermentation, raw materials are as follows in terms of weight fraction:

[0045] Saccharomyces cerevisiae 8 parts, Mucor circinelloides 7 parts, Pseudomonas putida 7 parts, Bacillus licheniformis 4 parts and alfalfa extract 15 parts. The viable cell count of each microorganism is 10 7 ~10 8 cfu / g.

[0046] The preparation method of the alfalfa extract is the same as that in Example 1.

[0047] Example 3

[0048] A composition for relieving ammonia nitrogen inhibition in anaerobic fermentation, raw materials are as follows in terms of weight fraction:

[0049] Saccharomyces cerevisiae 10 parts, Mucor circinelloides 5 parts, Pseudomonas putida 5 parts, Bacillus licheniformis 3 parts and alfalfa extract 10 parts. The viable cell count of each microorganism is 107 10 8 cfu / g.

[0050] The preparation method of the alfalfa extract is the same as that in Example 1.

[0051] Example 4

[0052] A method for relieving the inhibition of anaerobic fermentation by ammonia nitrogen, comprising the following steps:

[0053] The raw materials in the composition provided in Example 1 are weighed according to the proportion;

[0054] Saccharomyces cerevisiae, Mucor circinelloides, Pseudomonas putida and Bacillus licheniformis are inoculated into the activated culture medium respectively, and the inoculation amount is 5% (w / v), and the culture is carried out at 30°C and a rotation speed of 150 rpm for 30 h, to obtain four kinds of preliminary activated bacterial liquids;

[0055] The four kinds of preliminary activated bacterial liquids are mixed according to a volume ratio of 1:1:1:1, 5 times the volume of the mixed liquid of the activated culture medium is added, and the culture is carried out at 30°C and a rotation speed of 110 rpm for 10 h, to obtain the activated composition.

[0056] The activated composition is inoculated into the fermentation substrate, and the mass ratio of the composition to the total solid of the fermentation substrate is 0.08:1 based on the mass of the unactivated composition, wherein the total solid concentration in the fermentation substrate is 20% (w / w); the fermentation system has a pH of 7.0.

[0057] After the fermentation substrate is inoculated with the activated composition, anaerobic fermentation is carried out at 35°C.

[0058] In this embodiment, the composition of the activated culture medium is: glucose 10 g / L, peptone 5 g / L, yeast extract 3 g / L, potassium dihydrogen phosphate 1 g / L, magnesium sulfate 0.5 g / L; after each component is dissolved with distilled water, the pH is adjusted to 7.0, and sterilized at 121°C for 20 min.

[0059] Example 5

[0060] A method for relieving the inhibition of anaerobic fermentation by ammonia nitrogen, comprising the following steps:

[0061] The raw materials in the composition provided in Example 2 are weighed according to the proportion;

[0062] Saccharomyces cerevisiae, Mucor circinelloides, Pseudomonas putida and Bacillus licheniformis are inoculated into the activated culture medium respectively, and the inoculation amount is 5% (w / v), and the culture is carried out at 30°C and a rotation speed of 150 rpm for 30 h, to obtain four kinds of preliminary activated bacterial liquids;

[0063] The four preliminary activated bacteria liquids were mixed in a volume ratio of 1:1:1:1, 4 times the volume of the mixed liquid was added to the activated culture medium, and the mixture was cultured at 28°C and 120 rpm for 12 hours to obtain the activated composition.

[0064] The activated composition was inoculated into the fermentation substrate, and the mass ratio of the composition to the total solids in the fermentation substrate was 0.05:1 based on the mass of the unactivated composition, wherein the total solid concentration in the fermentation substrate was 30% (w / w); the pH of the fermentation system was 6.5.

[0065] After the fermentation substrate was inoculated with the activated composition, anaerobic fermentation was carried out at 45°C.

[0066] In this embodiment, the composition of the activated culture medium is: glucose 10 g / L, peptone 5 g / L, yeast extract 3 g / L, potassium dihydrogen phosphate 1 g / L, magnesium sulfate 0.5 g / L; after dissolving each component with distilled water, adjust the pH to 7.0, sterilize at 121°C for 20 min.

[0067] Example 6

[0068] A method for alleviating the inhibition of anaerobic fermentation by ammonia nitrogen, the steps are as follows:

[0069] The raw materials in the composition provided in Example 3 were weighed according to the proportion;

[0070] Saccharomyces cerevisiae, Mucor circinelloides, Pseudomonas putida and Bacillus licheniformis were inoculated into the activated culture medium, the inoculation amount was 5% (w / v), and the mixture was cultured at 32°C and 200 rpm for 26 hours to obtain four preliminary activated bacteria liquids;

[0071] The four preliminary activated bacteria liquids were mixed in a volume ratio of 1:1:1:1, 6 times the volume of the mixed liquid was added to the activated culture medium, and the mixture was cultured at 32°C and 100 rpm for 8 hours to obtain the activated composition.

[0072] The activated composition was inoculated into the fermentation substrate, and the mass ratio of the composition to the total solids in the fermentation substrate was 0.1:1 based on the mass of the unactivated composition, wherein the total solid concentration in the fermentation substrate was 10% (w / w); the pH of the fermentation system was 6.5.

[0073] After the fermentation substrate was inoculated with the activated composition, anaerobic fermentation was carried out at 50°C.

[0074] In this embodiment, the composition of the activated culture medium is: glucose 10 g / L, peptone 5 g / L, yeast extract 3 g / L, potassium dihydrogen phosphate 1 g / L, magnesium sulfate 0.5 g / L; after dissolving each component with distilled water, adjust the pH to 7.0, sterilize at 121°C for 20 min.

[0075] Comparative Example 1

[0076] The present comparative example differs from Example 1 in that no Medicago officinalis extract is added.

[0077] Comparative Example 2

[0078] The present comparative example differs from Example 1 in that no Pseudomonas putida and Bacillus licheniformis are added.

[0079] Comparative Example 3

[0080] The present comparative example differs from Example 1 in that no Mucor circinelloides is added.

[0081] Comparative Example 4

[0082] The present comparative example differs from Example 1 in that the Pseudomonas putida is replaced with an equal weight portion of Alcaligenes faecalis.

[0083] Comparative Example 5

[0084] The present comparative example differs from Example 1 in that the raw materials are as follows, in parts by weight:

[0085] 5 parts of Saccharomyces cerevisiae, 8 parts of Mucor circinelloides, 8 parts of Pseudomonas putida, 3.5 parts of Bacillus licheniformis and 13 parts of Medicago officinalis extract.

[0086] Comparative Example 6

[0087] The present comparative example differs from Example 4 in that the composition of Example 1 is replaced with the composition of Comparative Example 1.

[0088] Comparative Example 7

[0089] The present comparative example differs from Example 4 in that the composition of Example 1 is replaced with the composition of Comparative Example 2.

[0090] Comparative Example 8

[0091] The present comparative example differs from Example 4 in that the composition of Example 1 is replaced with the composition of Comparative Example 3.

[0092] Comparative Example 9

[0093] The present comparative example differs from Example 4 in that the composition of Example 1 is replaced with the composition of Comparative Example 4.

[0094] Comparative Example 10

[0095] The present comparative example differs from Example 4 in that the composition of Example 1 is replaced with the composition of Comparative Example 5.

[0096] Example 7

[0097] Effect of different compositions in relieving ammonia nitrogen inhibition.

[0098] The compositions of Examples 1-3 and Comparative Examples 1-5 were activated by the method of Example 4 to obtain activated compositions.

[0099] A simulated fermentation broth was prepared: glucose 20 g / L, ammonium chloride 2500 mg / L, potassium dihydrogen phosphate 1.5 g / L, magnesium sulfate 0.5 g / L, calcium chloride 0.1 g / L, FeSO4·7H2O 0.1 g / L, CoCl2·6H2O 0.01 g / L, NiCl2·6H2O 0.01 g / L, pH adjusted to 7.0, and autoclaved at 121°C for 20 min.

[0100] The simulated fermentation broth was divided into 8 groups, and the activated compositions of Examples 1-3 and Comparative Examples 1-5 were inoculated into each group at an inoculation amount of 10% (v / v), and cultured at 35°C and 120 rpm. At 0h, 24h and 48h after inoculation, the ammonia nitrogen concentration was detected by the Nessler's reagent method, and the ammonia nitrogen removal rate was calculated: ammonia nitrogen removal rate (%) = (initial ammonia nitrogen amount - ammonia nitrogen amount after fermentation) / initial ammonia nitrogen amount x 100. The results are shown in Table 1.

[0101] Table 1 Ammonia nitrogen removal rate

[0102]

[0103] According to the results in Table 1, the composition provided by the present application can quickly and effectively remove ammonia nitrogen and relieve ammonia nitrogen inhibition.

[0104] Example 8

[0105] Effect of the method for relieving ammonia nitrogen inhibition in anaerobic fermentation.

[0106] 1. Add exogenous ammonia nitrogen.

[0107] The test materials were cow dung and waste activated sludge provided by the Animal Science Experiment Station of Northwest A&F University. Before fermentation, the total solid concentration and pH of each group of fermentation substrates were adjusted. The ammonia nitrogen concentration was adjusted to 6000 mg / L by adding ammonium chloride. Anaerobic fermentation was carried out by the methods of Examples 4-6 and Comparative Examples 6-10. At the same time, a control group was set up: the fermentation substrate was directly subjected to anaerobic fermentation at 35°C without inoculation of exogenous microorganisms.

[0108] The organic matter (COD), volatile fatty acids (VFAs) and cumulative methane production in the fermentation system were detected before and after fermentation, and the improvement multiples of different methods were calculated: improvement multiple = experimental group quantitative index / control group quantitative index. The results are shown in Table 2.

[0109] Table 2 Effect of different methods on alleviating ammonia inhibition in anaerobic fermentation (6000 mg / L ammonia nitrogen)

[0110]

[0111] According to the results in Table 2, the method provided by the present application effectively alleviates ammonia inhibition and improves the production of organic matter, volatile organic acid and cumulative methane.

[0112] 2. No exogenous ammonia nitrogen was added.

[0113] The test raw materials were cow dung and waste activated sludge provided by the Animal Science Experiment Station of Northwest A & F University. Before fermentation, the total solid concentration and pH of the fermentation substrate of each group were adjusted. The fermentation substrate did not add ammonium chloride, and the initial ammonia nitrogen concentration was 720 mg / L.

[0114] The methods in Examples 4-6 and Comparative Examples 6-10 were used for anaerobic fermentation. At the same time, a control group was set up: the fermentation substrate was directly subjected to anaerobic fermentation at 35℃ without inoculating exogenous microorganisms.

[0115] The organic matter (COD), volatile organic acid (VFAs) and cumulative methane production in the fermentation system were detected before and after fermentation, and the improvement multiples of different methods were calculated: improvement multiple = experimental group quantitative index / control group quantitative index. The results are shown in Table 3.

[0116] Table 3 Effect of different methods on alleviating ammonia inhibition in anaerobic fermentation (no exogenous ammonia nitrogen was added)

[0117]

[0118] According to the results in Table 3, under the condition of no ammonia inhibition, the method provided by the present application can also promote the anaerobic fermentation, improve the production of organic matter, volatile organic acid and cumulative methane.

[0119] Therefore, the composition of the present application has the advantages of less dosage, cleanliness, easy availability and effective cost reduction without secondary pollution. The combined action of the composite microorganism and alfalfa extract in the composition of the present application can effectively alleviate ammonia inhibition, and the microorganism can better utilize the substrate for metabolic activity, reducing the waste of substrate caused by ammonia toxicity. The present application promotes the decomposition and transformation of organic matter, so that more substrate can be converted into useful products such as methane, improves the utilization rate of substrate, realizes the efficient resource utilization of organic waste, produces more clean energy while treating organic waste, and has good environmental and economic benefits.

[0120] The above merely describes the preferred embodiments of the present application, and it should be pointed out that, for those skilled in the art, several improvements and refinements can be made without departing from the principles of the present application, and these improvements and refinements should also be considered as falling within the protection scope of the present application.

Claims

1. A composition for alleviating ammonia nitrogen inhibition in anaerobic fermentation, characterized in that: The invention is composed of the following raw materials in parts by weight: 8-10 parts of saccharomyces cerevisiae, 5-8 parts of Mucor circinelloides, 5-8 parts of Pseudomonas putida, 3-4 parts of Bacillus licheniformis and 10-15 parts of alfalfa extract; The viable counts of the Saccharomyces cerevisiae, Mucor circinelloides, Pseudomonas putida and Bacillus licheniformis were all 10 7 ~10 8 cfu / g; The preparation method of the alfalfa extract comprises the following steps: taking fresh alfalfa, drying and then crushing it, mixing the powder with water and then extracting it at 40-50° C., filtering and collecting the extract, concentrating the extract and then drying it to obtain the alfalfa extract.

2. Use of the composition according to claim 1 in relieving and / or alleviating ammonia nitrogen inhibition in anaerobic fermentation.

3. A method for alleviating ammonia nitrogen inhibition in anaerobic fermentation, characterized in that: The method comprises the following steps: activating the composition according to claim 1, inoculating the activated composition into a fermentation substrate, and performing anaerobic fermentation.

4. The method according to claim 3, characterized in that The mass ratio of the composition to the total solids of the fermentation substrate is 0.01-1.5:

1.

5. The method according to claim 3, characterized in that During anaerobic fermentation, the total solid concentration in the fermentation substrate is 4%~30%; the temperature of anaerobic fermentation is 30~55℃; and the pH of the anaerobic fermentation system is 6.5~7.

5.

6. The method according to claim 3, characterized in that The activation method of the composition comprises: weighing various raw materials in proportion; inoculating various microorganisms into activation culture medium respectively, performing a first shaking culture, and obtaining four preliminary activated bacterial liquids after the culture is completed; mixing the four preliminary activated bacterial liquids in a volume ratio of 1:1:1:1, adding the activation culture medium, performing a second shaking culture, and adding alfalfa extract after the culture is completed to obtain an activated composition.

7. The method according to claim 6, characterized in that The conditions for the first shaking culture are: culture temperature of 28~32°C, culture time of 24~48h, and rotation speed of 150~200rpm.

8. The method according to claim 6, characterized in that The conditions for the second shaking culture are: culture temperature of 28~32°C, culture time of 6~12h, and rotation speed of 100~120rpm.

9. Use of the composition according to claim 1 or the method according to any one of claims 3 to 8 in any one or more of the following (1) to (3); (1) Promote the decomposition of anaerobic fermentation substrates; (2) Increase methane production from anaerobic fermentation; (3) Increase the yield of volatile organic acids in anaerobic fermentation.

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