CN110643593A - Preparation method of modified corn straw material immobilized agricultural microbial agent - Google Patents
Preparation method of modified corn straw material immobilized agricultural microbial agent Download PDFInfo
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Abstract
The invention belongs to the technical field of microbial agent production, and particularly relates to a preparation method of a modified corn straw material immobilized agricultural microbial agent. The microbial agent contains a modified corn straw carrier and a bacterial suspension; wherein the effective bacteria number of the bacterial suspension is more than 2 multiplied by 108one/mL. The modified corn straw of the invention has simple preparation process and H2O2As an oxidant, the composition is simple, and the oxidation effect is good; warp H2O2The activity of the oxidized straw molecules is enhanced, the adsorption capacity of the straw is promoted, and the corn straw is low in cost and easy to obtain and is suitable for large-scale production; meanwhile, the modified corn straw powder has strong adsorption capacity and rich microporous structureMicrobial adsorption provides a suitable growth environment; the added sodium carboxymethyl cellulose can be used as an adhesive to play a role in embedding and connection, the starch can meet the requirements of nutrient substances required by the growth of microorganisms, the boiling point of the glycerol is high, the freezing point is low, the microorganisms can be prevented from being stressed by the environment, and the service life and the effect of the microbial inoculum are greatly improved.
Description
Technical Field
The invention belongs to the technical field of microbial agent production, and particularly relates to a preparation method of a modified corn straw material immobilized agricultural microbial agent.
Background
China is a big agricultural country, ensures agricultural production, and causes the problem of non-point source pollution of agricultural pesticides in farmlands, and may become one of the most important factors which seriously restrict the agricultural sustainable development and the agricultural environment quality improvement of China in the future. The microbial remediation microbial inoculum can effectively degrade pesticide residues in farmlands, improve and restore soil quality, and has the characteristics of good remediation effect, low cost and no secondary pollution. The microbial agent for efficiently, stably and safely repairing pesticide residue pollution is of great significance to sustainable development of agriculture in China.
In order to prolong the shelf life of the microbial repairing microbial inoculum and enhance the adaptability of the microbial repairing microbial inoculum to the environment, an immobilization technology is a key. The immobilized microorganism technology is mainly a technology for immobilizing strains on a carrier by means of physical adsorption, an embedding method, a crosslinking method and the like, wherein the selection of a proper immobilized material is the key of the immobilized technology. In the physical adsorption immobilization method, the conventional carriers commonly used at present mainly comprise inorganic materials (such as diatomite, activated carbon, kaolin and the like), organic materials (polyvinyl alcohol, polyacrylamide and the like) and natural materials (sodium alginate, chitosan, starch and the like). However, these carriers have problems of low strength, low adsorption capacity, poor mass transfer, toxic effect on microorganisms, easy secondary pollution, etc., and thus, they have not been widely used in practical repair projects. Therefore, the microbial carrier which has strong adsorption capacity, good stability, no toxicity to microbes and environmental friendliness is obtained, and the microbial carrier has important significance for wide application of microbial remediation inocula. The modified corn straw is prepared from corn straw, and has the advantages of strong adsorption capacity, greenness, no toxicity, environmental friendliness, no secondary pollution and the like. The modified corn straw material is used as a carrier of the microbial repairing microbial inoculum, so that the survival rate of microorganisms in an application environment and the degradation capability of the microorganisms to pollutants can be improved, the problem of arrangement of agricultural wastes is solved, and the method is an economic, environment-friendly and resource-reasonably-utilizing technology.
Disclosure of Invention
The invention aims to explain a preparation method of a modified corn straw material immobilized agricultural microbial agent.
In order to achieve the purpose, the invention adopts the technical scheme that:
a modified corn stalk material immobilized agricultural microbial agent contains a modified corn stalk carrier and a bacterial suspension; wherein the effective bacteria number of the bacterial suspension is more than 2 multiplied by 108one/mL.
Mixing the modified corn stalk carrier with the bacterial suspension, adding 0.5-0.6g of the modified corn stalk carrier into every 20mL of the bacterial suspension, mixing, adding the mixed solution, uniformly mixing, standing for adsorption for 3-4h, and performing vacuum freeze drying to obtain the immobilized agricultural microbial agent; wherein the volume ratio of the mixed solution to the bacterial suspension is 1: 1; the final concentration of sodium carboxymethylcellulose in the mixed solution is 1-1.2 wt%, the final concentration of starch is 2-2.5 wt%, and the final concentration of glycerol is 1-1.2 wt%.
The modified corn stalk carrier is alkaline H2O2And (4) treating the corn straws by using the solution.
The modified corn stalk carrier is prepared by mixing H2O2Mixing with deionized water to obtain 1-1.2% H2O2Then adjusting the pH of the solution to 11-12; mixing with dried corn stalk powder, stirring at 22-25 deg.C for 14-16h, adjusting pH to 6-7, filtering, and drying to obtain modified corn stalk carrier.
Further, the reaction product of H2O2Mixing with deionized water to obtain 1-1.2% H2O2Adding flake NaOH to adjust the pH value of the solution to 11-12; mixing with dried corn stalk powder passing through 1mm sieve, stirring at 22-25 deg.C for 14-16h, adjusting pH to 6-7 with HCl, filtering, and drying to obtain modified corn stalk carrier.
The basicity H2O2The mass ratio of the solution to the corn stalk powder is 5-5.5: mixing at a ratio of 1-1.1.
The bacterial suspension is prepared by resuspending the thalli after fermentation culture and centrifugation by using skimmed milk powder as a protective agent, wherein the addition amount of the skimmed milk powder is 2.5-3 wt%; wherein, the thalli is pesticide degradation bacteria and other biocontrol and bacterial manure microorganisms, and specifically comprises the following components: arthrobacter (Arthrobacter), Hericium (Chengganzhangella), Pseudomonas (Pseudomonas), Burkholderia (Burkholderia), and the like.
A method for preparing modified corn stalk material immobilized agricultural microbial agent comprises soaking dried corn stalk powder in alkaline H2O2Obtaining a modified corn straw carrier in the solution; centrifugally collecting bacteria after fermentation culture, and resuspending the bacteria by using protective agent skimmed milk powder; and uniformly mixing the re-suspension and the modified corn stalk carrier, adding the mixed solution after uniformly mixing, standing for adsorption for 3-4h, and freeze-drying to obtain the modified corn stalk material immobilized agricultural microbial agent.
The invention has the advantages that:
1) the carrier is simple to manufacture: the modified corn straw has simple preparation process H2O2As an oxidant, the composition is simple, and the oxidation effect is good; warp H2O2The activity of the oxidized straw molecules is enhanced, the adsorption capacity of the straw is promoted, and the corn straw is low in cost and easy to obtain and is suitable for large-scale production;
2) the immobilization efficiency is high: the modified corn straw powder has strong adsorption capacity, and the abundant microporous structure provides a proper growth environment for microbial adsorption; the added sodium carboxymethylcellulose can be used as an adhesive to play roles of embedding and connection, the starch can meet nutrient substances required by the growth of microorganisms, the boiling point of the glycerol is higher, the freezing point is lower, the microorganisms can be prevented from being stressed by the environment, and the service life and the effect of the microbial inoculum are greatly improved.
3) The immobilized carrier is green and environment-friendly: the corn straw is a natural degradable substance, is nontoxic and pollution-free to the environment, solves the problem of arrangement of agricultural wastes, and is an economic and environment-friendly carrier.
Detailed Description
The principles and features of this invention are described below in conjunction with examples, which are set forth to illustrate, but are not to be construed to limit the scope of the invention.
Example 1
A modified corn stalk material immobilized agricultural microbial agent comprises the following steps:
1) 485mL of deionized water and 15mL of H are measured2O2Mixing well to obtain 500mL of 1% H2O2Adding flake NaOH, adjusting pH to 11-12 to obtain alkaline 1% H2O2A solution;
2) weighing 100g of dried corn straw powder, sieving the powder by a 1mm sieve, and adding the powder to the alkaline 1% H obtained in the step 1)2O2Stirring the solution for 14 hours by using a constant-temperature magnetic stirrer;
3) adjusting the pH of the mixture in the step 2) to 6-7 by using 6mol/L HCl, and filtering by using a nylon net;
4) washing the filtrate with deionized water, and drying in an electric heating constant temperature blast drying oven at 60 deg.C to constant weight to obtain modified corn stalk carrier.
5) 20mL of bacterial liquid obtained by fermenting the herbicide Arthrobacter keyseri ATR1 obtained by the room sieve in a conventional mode is centrifuged for 10min at 4000r/min, the thalli are collected, 10-13mL of 3 wt% skimmed milk powder aqueous solution is used for resuspending the thalli, and bacterial suspension is prepared. The effective viable count is controlled to reach the technical index of agricultural microbial agent products, which is more than 2 multiplied by 108Adding 0.5-0.6g of the modified corn straw carrier obtained in the step 4) into the bacterial suspension, and uniformly mixing.
6) And (3) adding 10-13mL of mixed solution containing 1 wt% of sodium carboxymethylcellulose, 2% (v/v) of starch and 1 wt% of glycerol into the mixture obtained in the step 5), standing for adsorption for 3-4h, and carrying out vacuum freeze drying to obtain a finished product of the fixed microbial inoculum.
7) Respectively diluting and coating the fermentation liquor obtained in the step 5) and the fixed microbial inoculum finished product obtained in the step 6) by deionized water according to a 10-time gradient dilution method, wherein the effective viable count of the fermentation liquor is 1.075 multiplied by 1012Freeze drying the fermented liquid to obtain 5.8 × 10 viable count9Per mL; the effective viable count of the immobilized bacteria agent prepared in the step 6) is 8.129 multiplied by 1011one/mL.
The experimental results show that the effective viable count survival rate is high and the immobilization efficiency is high after the immobilization is carried out by the immobilization method.
8) Taking 20mL of fermentation liquor obtained in the step 5), centrifugally collecting thalli and the immobilized microbial inoculum prepared in the step 6), and respectively adding a basic inorganic salt solution (NaCl 00.5g/L, NH) containing 500mg/L atrazine4Cl 2g/L，MgSO4·7H2O 0.4g/L，K2HPO4 1.5g/L，KH2PO40.5g/L of distilled water, 1000mL of distilled water, pH6.8-7.2), 150r/min, carrying out shake culture for 8h, detecting the residue of atrazine, calculating to obtain that the degradation rate of free thalli to atrazine is 62.66%, and the degradation rate of modified corn straw immobilized microbial agent to atrazine is 98.05%, which indicates that the modified corn straw immobilized microbial agent has high-efficiency degradation rate.
Example 2
A preparation method of a modified corn straw material immobilized agricultural microbial agent comprises the following steps:
1) 485mL of deionized water and 15mL of H are measured2O2Mixing well to obtain 500mL of 1% H2O2Adding flake NaOH, adjusting pH to 11-12 to obtain alkaline 1% H2O2A solution;
2) weighing 100g of dried corn straw powder, sieving the powder by a 1mm sieve, and adding the powder to the alkaline 1% H obtained in the step 1)2O2Stirring the solution for 14 hours by using a constant-temperature magnetic stirrer;
3) adjusting the pH of the mixture in the step 2) to 6-7 by using 6mol/L HCl, and filtering by using a nylon net;
4) washing the filtrate with deionized water, and drying in an electric heating constant temperature blast drying oven at 60 deg.C to constant weight to obtain modified corn stalk carrier.
5) 20mL of bacterial liquid obtained by fermenting the herbicide methanolivorans CHL1 obtained by the room sieve according to a conventional mode is centrifuged for 10min at 4000r/min, and thalli are collected and resuspended by 10-13mL of 3 wt% of skimmed milk powder aqueous solution to prepare bacterial suspension. The effective viable count is controlled to reach the technical index of agricultural microbial agent products, which is more than 2 multiplied by 108Adding 0.5-0.6g of the modified corn straw carrier obtained in the step 4) into the bacterial suspension, and uniformly mixing.
6) And (3) adding 10-13mL of a mixed solution containing 1 wt% of sodium carboxymethylcellulose, 2% (v/v) of starch and 1 wt% of glycerol into the mixture obtained in the step 5), standing for adsorption for 3-4h, and carrying out vacuum freeze drying to obtain a finished product of the fixed microbial inoculum.
7) Respectively diluting and coating the fermentation liquor obtained in the step 5) and the fixed microbial inoculum finished product obtained in the step 6) by deionized water according to a 10-time gradient dilution method, wherein the effective viable count of the fermentation liquor is 8.5 multiplied by 1010Freeze drying the fermented liquid to obtain effective viable count of 3.07 × 108Per mL; the effective viable count of the immobilized bacteria agent prepared in the step 6) is 1.7 multiplied by 109one/mL.
The experimental results show that the effective viable count survival rate is high and the immobilization efficiency is high after the immobilization is carried out by the immobilization method.
8) And (3) centrifugally collecting 20mL of fermentation liquor in the step 5) and the immobilized microbial agent prepared in the step 6) are respectively added into a basic inorganic salt solution containing 100mg/L of metsulfuron-methyl, the solution is subjected to shaking culture for 5 days, the residue of the metsulfuron-methyl is detected, the degradation rate of free bacteria to the metsulfuron-methyl is calculated to be 78.13%, the degradation rate of the modified corn straw immobilized microbial agent to the metsulfuron-methyl is 99.44%, and the modified corn straw immobilized microbial agent has a high-efficiency degradation rate.
The above description is only for the purpose of illustrating the preferred embodiments of the present invention and is not to be construed as limiting the scope of the present invention, as any modifications, equivalent replacements, improvements, etc., which are within the spirit and principle of the present invention, should be included in the present invention.
Claims (8)
1. A modified corn stalk material immobilized agricultural microbial agent is characterized in that the microbial agent contains a modified corn stalk carrier and a bacterial suspension; wherein the effective bacteria number of the bacterial suspension is more than 2 multiplied by 108one/mL.
2. The immobilized agricultural microbial agent of modified corn stalk material according to claim 1, wherein the modified corn stalk carrier is mixed with bacterial suspension, 0.5-0.6g of modified corn stalk carrier is added into every 20mL of bacterial suspension, mixed solution is added after mixing, and mixed uniformly, and the mixture is kept stand for adsorption for 3-4h and is subjected to vacuum freeze drying to prepare the immobilized agricultural microbial agent; wherein the volume ratio of the mixed solution to the bacterial suspension is 1: 1; the final concentration of sodium carboxymethylcellulose in the mixed solution is 1-1.2 wt%, the final concentration of starch is 2-2.5 wt%, and the final concentration of glycerol is 1-1.2 wt%.
3. The modified corn stalk material immobilized agricultural microbial agent as claimed in claim 1, wherein the modified corn stalk carrier is alkaline H2O2And (4) treating the corn straws by using the solution.
4. The modified corn stalk material immobilized agricultural microbial agent as claimed in claim 3, wherein: h is to be2O2Mixing with deionized water to obtain 1-1.2% H2O2Then adjusting the pH of the solution to 11-12; mixing with dried corn stalk powder, stirring at 22-25 deg.C for 14-16h, adjusting pH to 6-7, filtering, and drying to obtain modified corn stalk carrier.
5. The modified corn stalk material immobilized agricultural microbial agent as claimed in claim 4, wherein: h is to be2O2Mixing with deionized water to obtain 1-1.2% H2O2Adding flake NaOH to adjust the pH value of the solution to 11-12; mixing with dried corn stalk powder passing through 1mm sieve, stirring at 22-25 deg.C for 14-16h, adjusting pH to 6-7 with HCl, filtering, and drying to obtain modified corn stalk carrier.
6. The modified corn stover material-immobilized agricultural microbial inoculant of claim 3, wherein the alkaline H is2O2The mass ratio of the solution to the corn stalk powder is 5-5.5: mixing at a ratio of 1-1.1.
7. The modified corn stalk material immobilized agricultural microbial agent as claimed in claim 1 or 2, wherein the bacterial suspension is prepared by resuspending thallus after fermentation culture and centrifugation with skimmed milk powder as a protective agent, and the addition amount of the skimmed milk powder is 2.5-3 wt%.
8. The preparation method of the modified corn stalk material immobilized agricultural microbial agent of claim 1, wherein the dried corn stalk powder is soaked in alkaline H2O2Obtaining a modified corn straw carrier in the solution; centrifugally collecting bacteria after fermentation culture, and resuspending the bacteria by using protective agent skimmed milk powder; and uniformly mixing the re-suspension and the modified corn stalk carrier, adding the mixed solution after uniformly mixing, standing for adsorption for 3-4h, and freeze-drying to obtain the modified corn stalk material immobilized agricultural microbial agent.
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Cited By (7)
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| CN111235056A (en) * | 2020-01-19 | 2020-06-05 | 南阳师范学院 | Grown-up bacterium BI-1 capable of degrading tribenuron-methyl and culture method and application thereof |
| CN111517477A (en) * | 2020-05-14 | 2020-08-11 | 江西省科学院 | Sulfate reducing bacteria embedded particles for treating ionic rare earth mine wastewater, and preparation method and application thereof |
| CN111545181A (en) * | 2020-05-12 | 2020-08-18 | 山东交通学院 | Preparation and oil removal method of biologically-modified corn straw oil absorption material |
| CN111662897A (en) * | 2020-06-19 | 2020-09-15 | 中农新科(苏州)有机循环研究院有限公司 | Preparation method of modified wheat bran material immobilized microbial agent |
| CN112813059A (en) * | 2021-02-08 | 2021-05-18 | 杭州楠大环保科技有限公司 | Immobilization method of microbial agent |
| CN112852666A (en) * | 2021-01-19 | 2021-05-28 | 新疆河润水业有限责任公司 | Preparation method of microbial agent and microbial fertilizer prepared by adopting microbial agent |
| CN113512425A (en) * | 2021-06-16 | 2021-10-19 | 广西博世科环保科技股份有限公司 | Preparation method of wood fiber-based microorganism-loaded material for repairing organic contaminated soil |
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Cited By (11)
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| CN111235056A (en) * | 2020-01-19 | 2020-06-05 | 南阳师范学院 | Grown-up bacterium BI-1 capable of degrading tribenuron-methyl and culture method and application thereof |
| CN111235056B (en) * | 2020-01-19 | 2023-07-04 | 南阳师范学院 | Rigid fungus BI-1 capable of degrading tribenuron-methyl, and culture method and application thereof |
| CN111545181A (en) * | 2020-05-12 | 2020-08-18 | 山东交通学院 | Preparation and oil removal method of biologically-modified corn straw oil absorption material |
| CN111517477A (en) * | 2020-05-14 | 2020-08-11 | 江西省科学院 | Sulfate reducing bacteria embedded particles for treating ionic rare earth mine wastewater, and preparation method and application thereof |
| CN111517477B (en) * | 2020-05-14 | 2021-01-22 | 江西省科学院 | Sulfate reducing bacteria embedded particles for treating ionic rare earth mine wastewater, and preparation method and application thereof |
| CN111662897A (en) * | 2020-06-19 | 2020-09-15 | 中农新科(苏州)有机循环研究院有限公司 | Preparation method of modified wheat bran material immobilized microbial agent |
| CN111662897B (en) * | 2020-06-19 | 2023-03-21 | 中农新科(苏州)有机循环研究院有限公司 | Preparation method of modified wheat bran material immobilized microbial agent |
| CN112852666A (en) * | 2021-01-19 | 2021-05-28 | 新疆河润水业有限责任公司 | Preparation method of microbial agent and microbial fertilizer prepared by adopting microbial agent |
| CN112813059A (en) * | 2021-02-08 | 2021-05-18 | 杭州楠大环保科技有限公司 | Immobilization method of microbial agent |
| CN113512425A (en) * | 2021-06-16 | 2021-10-19 | 广西博世科环保科技股份有限公司 | Preparation method of wood fiber-based microorganism-loaded material for repairing organic contaminated soil |
| CN113512425B (en) * | 2021-06-16 | 2023-05-05 | 广西博世科环保科技股份有限公司 | Preparation method of lignocellulose-based loaded microbial material for repairing organic contaminated soil |
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