Method for treating blue-green algae through fermentation of saccharomonospora viridis and high-temperature bacillus subtilis

Abstract

The invention discloses a method for treating blue-green algae through fermentation of saccharomonospora viridis and high-temperature bacillus subtilis, the method comprises preparation of a compound microbial agent and fermentation of the blue-green algae, and the compound microbial agent is formed by compounding solid-state fermentation products of the high-temperature bacillus subtilis and the saccharomonospora viridis. Wherein the effective viable count of the high-temperature bacillus subtilis is greater than or equal to 2.5 * 10 < 8 > cfu / g, and the effective viable count of the saccharomonospora viricola is greater than or equal to 1.0 * 10 < 8 > cfu / g. According to the method, the blue-green algae can be subjected to safe and non-toxic fermentation treatment, meanwhile, due to the heat production effect of the fermentation process, electric energy can be saved, the aim of saving and utilizing resources is achieved, and products obtained after blue-green algae fermentation can also be used as feed and fertilizer raw materials for resource utilization.

Description

Technical Field

[0001] The invention belongs to the technical field of blue algae treatment, and specifically relates to a method for treating blue algae by fermentation with green Saccharomonas viridis and thermophilic Bacillus subtilis. Background Art

[0002] Cyanobacteria, also known as blue-green algae, have cell walls and are single-celled organisms. They are larger than bacteria, with a diameter of about 3 to 15 microns. Usually, cyanobacteria tend to aggregate to form filamentous or single-cell aggregates.

[0003] The most representative feature of the large-scale "water bloom" phenomenon that occurs in lakes in my country in summer is the massive reproduction of cyanobacteria, which forms a blue-green floating algae layer on the water surface. The large-scale reproduction of cyanobacteria consumes the dissolved oxygen in the water, causing the death of a large number of fish and shrimp, and produces highly toxic algae toxins. Therefore, the control of cyanobacteria in lakes has become a key and difficult issue in the current water pollution control.

[0004] With the deterioration of water environment quality and the high attention of society, the technology of collecting and treating cyanobacteria has received more and more attention from researchers and has been put into practical application in many lake management projects. The use of reasonable, economical and efficient technical means to achieve the rapid separation and resource utilization of cyanobacteria in water is a new development direction for the research on eutrophication control of water bodies. It is worth noting that in the process of control and resource utilization, in addition to economical and efficient technical methods, how to ensure safe technical methods without secondary pollution should also be a key research direction. Summary of the invention

[0005] The purpose of the present invention is to overcome the deficiencies in the prior art and provide a composite microbial agent for safely and non-toxically fermenting cyanobacteria, thereby achieving the purpose of treating cyanobacteria and utilizing them as resources.

[0006] In order to solve the above technical problems, the technical solution adopted by the present invention is: A method for treating cyanobacteria by fermentation of green saccharomonas and thermophilic Bacillus subtilis, comprising the preparation of a composite microbial agent and cyanobacteria fermentation, wherein the composite microbial agent is compounded from solid-state fermentation products of thermophilic Bacillus subtilis and green saccharomonas, wherein the effective viable count of thermophilic Bacillus subtilis is ≥2.5×10 8 cfu / g, effective viable count of Saccharomonas viridis ≥ 1.0×10 8 cfu / g.

[0007] The preparation method of the composite microbial agent comprises the following steps: (1) Solid-state fermentation: Thermobaric Bacillus subtilis and Saccharomonas viridis are subjected to solid-state fermentation to obtain a large number of microbial cells; (2) Boiling drying: Dry the fermented product at 70°C to obtain a microbial agent product with a moisture content of less than 10%; (3) Crushing: crushing the dried solid-state fermentation product; (4) Mixing: The solid-state fermentation products of thermophilic Bacillus subtilis and green Saccharomonas viridis are mixed and compounded according to the proportion of bacterial content, and auxiliary materials are added.

[0008] Furthermore, the effective viable count of thermophilic Bacillus subtilis after compounding is ≥2.5×10 8 cfu / g, effective viable count of Saccharomonas viridis ≥ 1.0×10 8 cfu / g.

[0009] The preparation of the thermophilic Bacillus subtilis solid-state fermentation adopts the following steps: The thermophilic Bacillus subtilis strain was inoculated from the slant into the primary seed culture medium and cultured for 2-3 days. The spores were washed off to prepare a spore suspension. The spore suspension was spread on the secondary seed culture medium for 2-3 days. The spores were washed off to prepare a spore suspension. The spore suspension was then heated to 10 3 -10 4 The solid-state fermentation medium is inoculated with an inoculum amount of cfu / g substrate and cultured for 5-6 days to obtain a solid-state fermentation product of thermophilic Bacillus subtilis.

[0010] Furthermore, the primary and secondary seed culture media are the same, with a formula of 0.5% peptone, 0.3% beef extract, trace element solution (0.5 g / L CaCl2•2H2O, 0.2 g / L K2HPO4, 3.0 g / L (NH4)2HPO4, 0.1 g / L MgSO4•7H2O, 0.1g / L MnSO4•H2O), 1.5%-2% agar, and a pH of 7.0-8.0.

[0011] Furthermore, the solid-state fermentation medium formula is corn flour and soybean flour as solid-state fermentation matrix, and an inorganic salt nutrient solution composed of CaCl2, KH2PO4, (NH4)2HPO4, MgSO4, and MnSO4 with a final concentration of 1.0% is added.

[0012] Furthermore, the solid-state fermentation conditions are a temperature of 55-65° C. and an initial pH value of 7.0-8.0.

[0013] Further, the deposit information of the Bacillus subtilis strain used is: deposit classification name :Bacillus subtilis , accession number ：5550 , deposit date : December 8, 2011 , preservation unit :China Microbiological Culture Collection Administration Microbiology Center , address of depository :No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing .

[0014] The preparation of the green Saccharomonas solid-state fermentation adopts the following steps: (1) Shake flask culture: Inoculate the Saccharomonas viridis strain from the stored slant into the shake flask medium, and let it stand and activate the culture at 60℃±1℃ for 12-14 h; (2) Seed culture: Inoculate the shake flask culture into the seed tank at an inoculum rate of 2.0-3.0%, and culture at a speed of 30-40 r / min and 60℃±1℃ for 10-12 h to obtain the seed solution, and count the green Saccharomonas in the seed solution; (3) Solid-state fermentation: The seed liquid is inoculated into the solid-state fermentation medium for fermentation. The fermentation product can be obtained by culturing at 55-65°C for 4-5 days.

[0015] Furthermore, the formula of the shake flask culture medium is 25 g of peptone, 15 g of glucose, 1000 mL of water, and pH 7.0-7.5.

[0016] Furthermore, the formula of the seed culture medium is 10-15 g of peptone, 10-15 g of beef extract, 1000 mL of water, and pH 7.0-7.5.

[0017] Furthermore, the solid-state fermentation medium is formulated with wheat bran as the sole carbon source and fermentation substrate, with 2-3% peptone, 1.5-2.0% NaCl, and 0.4%-0.5% nutrient solution (composed of KNO3, K2HPO4, MgSO4, and FeSO4) added, and an initial pH of 7.0-7.5.

[0018] The cyanobacteria fermentation is performed by placing the cyanobacteria into a fermentation tank or a fermentation tank, adding the composite microbial agent, stirring at a speed of 300 rpm for 5 minutes, and standing to ferment for 24 hours.

[0019] Furthermore, the dosage of the composite microbial agent is 0.5-1.5kg / m³, and the water content of the blue algae is ≤90%.

[0020] Compared with the prior art, the present invention has the following advantages: The composite microbial agent of the present invention is composed of thermophilic Bacillus subtilis ( Bacillus subtilis ) and Saccharomonas viridis ( Saccharomonospora viridis ) is compounded with the solid fermentation products of cyanobacteria. The present invention can ferment cyanobacteria safely and non-toxicly. At the same time, the heat generation of the fermentation process itself can save electric energy and achieve the purpose of resource conservation and utilization. The products after cyanobacteria fermentation can also be utilized as feed and fertilizer raw materials for resource utilization. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] Figure 1The present invention is a flow chart of the preparation method of the composite microbial agent. Implementation

[0022] In order to make the objects and advantages of the present invention more clearly understood, the present invention is further described below in conjunction with embodiments; it should be understood that the specific embodiments described herein are only used to explain the present invention and are not used to limit the present invention.

[0023] The preferred embodiments of the present invention are described below with reference to the accompanying drawings. The accompanying drawings are simplified schematic diagrams, which only illustrate the basic process of the present invention in a schematic manner, and therefore only show processes related to the present invention. It should be understood by those skilled in the art that these embodiments are only used to explain the technical principles of the present invention and are not intended to limit the scope of protection of the present invention. Example

[0024] The following is combined with Figure 1 The process flow chart for implementing the first technical solution of the present invention is described in detail. The preparation of the composite microbial agent in this embodiment and the method of fermenting cyanobacteria using the composite microbial agent are implemented as follows: A method for treating cyanobacteria by fermentation of green saccharomonas and thermophilic Bacillus subtilis, comprising the preparation of a composite microbial agent and cyanobacteria fermentation, wherein the composite microbial agent is compounded from solid-state fermentation products of thermophilic Bacillus subtilis and green saccharomonas, wherein the effective viable count of thermophilic Bacillus subtilis is ≥2.5×10 8 cfu / g, effective viable count of Saccharomonas viridis ≥ 1.0×10 8 cfu / g.

[0025] The preparation method of the composite microbial agent comprises the following steps: (1) Solid-state fermentation: Thermomyces subtilis and Saccharomonas viridis are subjected to solid-state fermentation to obtain a large number of microbial cells; (2) Boiling drying: Dry the fermented product at 70°C to obtain a microbial agent product with a moisture content of less than 10%; (3) Crushing: crushing the dried solid-state fermentation product; (4) Mixing: The solid-state fermentation products of thermophilic Bacillus subtilis and green Saccharomonas viridis are mixed and compounded according to the proportion of bacterial content, and auxiliary materials are added.

[0026] Furthermore, the effective viable count of thermophilic Bacillus subtilis after compounding is ≥2.5×10 8 cfu / g, effective viable count of Saccharomonas viridis ≥ 1.0×10 8 cfu / g.

[0027] The preparation of the thermophilic Bacillus subtilis solid-state fermentation adopts the following steps: The thermophilic Bacillus subtilis strain was inoculated from the slant into the primary seed culture medium and cultured for 2-3 days. The spores were washed off to prepare a spore suspension. The spore suspension was spread on the secondary seed culture medium for 2-3 days. The spores were washed off to prepare a spore suspension. The spore suspension was then heated to 10 3 -10 4 The solid-state fermentation medium is inoculated with an inoculum amount of cfu / g substrate and cultured for 5-6 days to obtain a solid-state fermentation product of thermophilic Bacillus subtilis.

[0028] Furthermore, the primary and secondary seed culture media are the same, with a formula of 0.5% peptone, 0.3% beef extract, trace element solution (0.5 g / L CaCl2•2H2O, 0.2 g / L K2HPO4, 3.0 g / L (NH4)2HPO4, 0.1 g / L MgSO4•7H2O, 0.1g / L MnSO4•H2O), 1.5%-2% agar, and a pH of 7.0-8.0.

[0029] Furthermore, the solid-state fermentation medium formula is corn flour and soybean flour as solid-state fermentation matrix, and an inorganic salt nutrient solution composed of CaCl2, KH2PO4, (NH4)2HPO4, MgSO4, and MnSO4 with a final concentration of 1.0% is added.

[0030] Furthermore, the solid-state fermentation conditions are a temperature of 55-65° C. and an initial pH value of 7.0-8.0.

[0031] Further, the deposit information of the Bacillus subtilis strain used is: deposit classification name: Bacillus subtilis, deposit number: CGMCC No.5550, deposit date: December 8, 2011, deposit unit: General Microbiology Center of China Microorganism Culture Collection Administration, deposit unit address: No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing.

[0032] The preparation of the green Saccharomonas solid-state fermentation adopts the following steps: (1) Shake flask culture: Inoculate the Saccharomonas viridis strain from the stored slant into the shake flask medium, and let it stand and activate the culture at 60℃±1℃ for 12-14 h; (2) Seed culture: Inoculate the shake flask culture into the seed tank at an inoculum rate of 2.0-3.0%, and culture at a speed of 30-40 r / min and 60℃±1℃ for 10-12 h to obtain the seed solution, and count the green Saccharomonas in the seed solution; (3) Solid-state fermentation: The seed liquid is inoculated into the solid-state fermentation medium for fermentation. The fermentation product can be obtained by culturing at 55-65°C for 4-5 days.

[0033] Furthermore, the formula of the shake flask culture medium is 25 g of peptone, 15 g of glucose, 1000 mL of water, and pH 7.0-7.5.

[0034] Furthermore, the formula of the seed culture medium is 10-15 g of peptone, 10-15 g of beef extract, 1000 mL of water, and pH 7.0-7.5.

[0035] Furthermore, the solid-state fermentation medium is formulated with wheat bran as the sole carbon source and fermentation substrate, with 2-3% peptone, 1.5-2.0% NaCl, and 0.4%-0.5% nutrient solution (composed of KNO3, K2HPO4, MgSO4, and FeSO4) added, and an initial pH of 7.0-7.5.

[0036] The cyanobacteria fermentation is performed by placing the cyanobacteria into a fermentation tank or a fermentation tank, adding the composite microbial agent, stirring at a speed of 300 rpm for 5 minutes, and standing to ferment for 24 hours.

[0037] Furthermore, the dosage of the composite microbial agent is 0.5-1.5kg / m³, and the water content of the blue algae is ≤90%.

[0038] So far, the technical solutions of the present invention have been described in conjunction with the preferred embodiments shown in the accompanying drawings. However, it is easy for those skilled in the art to understand that the protection scope of the present invention is obviously not limited to these specific embodiments. Without departing from the principle of the present invention, those skilled in the art can make equivalent changes or substitutions to the relevant technical features, and the technical solutions after these changes or substitutions will fall within the protection scope of the present invention.

Claims

1. A method for treating cyanobacteria by fermentation of green Saccharomonas and thermophilic Bacillus subtilis, comprising the preparation of a composite microbial agent and cyanobacterial fermentation, wherein the composite microbial agent is compounded from solid-state fermentation products of thermophilic Bacillus subtilis and green Saccharomonas, wherein the effective viable count of thermophilic Bacillus subtilis is ≥ 2.5×10 8 cfu / g, effective viable count of Saccharomonas viridis ≥ 1.0×10 8 cfu / g.

2. The method for treating cyanobacteria by fermentation with Saccharomonas viridis and Bacillus subtilis according to claim 1, characterized in that: The preparation method of the composite microbial agent comprises the following steps: (1) Solid-state fermentation: Thermomyces subtilis and Saccharomonas viridis are subjected to solid-state fermentation to obtain solid-state fermentation products respectively; (2) Boiling drying: drying the solid-state fermentation product at 70° C. to obtain a microbial inoculant product with a water content of less than 10%; (3) Crushing: crushing the dried solid-state fermentation product; (4) Mixing: The solid-state fermentation products of thermophilic Bacillus subtilis and green Saccharomonas viridis are mixed and compounded according to the proportion of bacterial content, and auxiliary materials are added.

3. A method for fermenting cyanobacteria with Saccharomonas viridis and Bacillus subtilis as claimed in claim 2, characterized in that: The bacterial content in the thermophilic Bacillus subtilis fermentation product is 2.5×10 8 cfu / g, the bacterial content in the fermentation product of Saccharomonas viridis was 1.0×10 8 cfu / g.

4. The method for fermenting cyanobacteria with Saccharomonas viridis and Bacillus subtilis as claimed in claim 2, characterized in that: The preparation of the thermophilic Bacillus subtilis solid-state fermentation adopts the following steps: The thermophilic Bacillus subtilis strain was inoculated from the slant into the primary seed culture medium and cultured for 2-3 days. The spores were washed off to prepare a spore suspension. The spore suspension was spread on the secondary seed culture medium for 2-3 days. The spores were washed off to prepare a spore suspension. The spore suspension was then heated to 10 3 -10 4 cfu / g inoculated into the solid fermentation medium, cultured for 5-6 days, and obtained the solid fermentation product of thermophilic Bacillus subtilis; The first-level and second-level seed culture media are the same, with a formula of 0.5% peptone, 0.3% beef extract, trace element solution (0.5 g / L CaCl2•2H2O, 0.2 g / L K2HPO4, 3.0 g / L (NH4)2HPO4, 0.1 g / L MgSO4•7H2O, 0.1 g / L MnSO4•H2O), 1.5%-2% agar, and a pH of 7.0-8.0; The solid-state fermentation medium is formulated by using corn flour and soybean flour as solid-state fermentation matrix, and adding an inorganic salt nutrient solution composed of CaCl2, KH2PO4, (NH4)2HPO4, MgSO4, and MnSO4 with a final concentration of 1.0%; The solid-state fermentation conditions are a temperature of 55-65° C. and an initial pH value of 7.0-8.

0.

5. The method for treating cyanobacteria by fermentation with Saccharomonas viridis and Bacillus subtilis as claimed in claim 2, characterized in that: The preparation of the green Saccharomonas solid-state fermentation adopts the following steps: (1) Shake flask culture: Inoculate the Saccharomonas viridis strain from the stored slant into the shake flask medium, and let it stand and activate the culture at 60℃±1℃ for 12-14 h; (2) Seed culture: Inoculate the shake flask culture into the seed tank at an inoculum rate of 2.0-3.0%, and culture at a speed of 30-40 r / min and 60℃±1℃ for 10-12 h to obtain the seed solution, and count the green Saccharomonas in the seed solution; (3) Solid-state fermentation: The seed solution is inoculated into the solid-state fermentation medium for fermentation. The fermentation product can be obtained by culturing at 55-65°C for 4-5 days. The formula of the shake flask medium is 25 g of peptone, 15 g of glucose, 1000 mL of water, pH 7.0-7.5; The formula of the seed culture medium is 10-15 g of peptone, 10-15 g of beef extract, 1000 mL of water, pH 7.0-7.5; The solid fermentation medium is formulated with wheat bran as the sole carbon source and fermentation matrix, with 2-3% peptone, 1.5-2.0% NaCl, and 0.4%-0.5% nutrient solution (composed of KNO3, K2HPO4, MgSO4, and FeSO4) added, and an initial pH of 7.0-7.

5.

6. The method for treating cyanobacteria by fermentation with Saccharomonas viridis and Bacillus subtilis according to claim 1, characterized in that: The cyanobacteria fermentation is performed by placing the cyanobacteria into a fermentation tank or a fermentation tank, adding the composite microbial agent, stirring at a speed of 300 rpm for 5 minutes, and standing to ferment for 24 hours.

7. A method for treating cyanobacteria by fermentation with Saccharomonas viridis and Bacillus subtilis as claimed in claim 6, characterized in that: The dosage of the composite microbial agent is 0.5-1.5kg / m³, and the water content of the blue algae is ≤90%.

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