CN105441370B - A kind of microbial bacterial agent and its production method - Google Patents

A kind of microbial bacterial agent and its production method Download PDF

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CN105441370B
CN105441370B CN201610061792.2A CN201610061792A CN105441370B CN 105441370 B CN105441370 B CN 105441370B CN 201610061792 A CN201610061792 A CN 201610061792A CN 105441370 B CN105441370 B CN 105441370B
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CN105441370A (en
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王伟华
陈凤霞
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Shandong Hetianwang Biological Technology Co Ltd
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Abstract

The present invention provides a kind of microbial bacterial agent and its production method, including bacillusmusilaginosiengineering, bacillus subtilis, bacillus licheniformis.In the microbial bacterial agent, bacillusmusilaginosiengineering viable count is 18~2,200,000,000 cfu/g, and bacillus subtilis viable count is 17~1,900,000,000 cfu/g, and bacillus licheniformis viable count is 12~1,300,000,000 cfu/g.The production method of microbial bacterial agent, including strain selection, actication of culture and spread cultivation, strain fermentation, finished product step.Microbial bacterial agent of the present invention, bacterial content is high, and miscellaneous bacteria rate is low, can effectively improve soil property.

Description

A kind of microbial bacterial agent and its production method
Technical field
The present invention provides a kind of microbial bacterial agent and its production method, belongs to technical field of microbial fermentation.
Background technique
Microbial bacterial agent refers to a kind of particular product containing viable microbial.It is with the movable process of microbial life and Product improves crop nutritional condition, plays soil potential fertility, stimulates crop growth, germ harm is resisted, to mention High crop yield and quality.It supplies nutrients substance not as general fertilizer directly to plant, but its effect can not be ignored.
Microbial bacterial agent expands for arable farming and growing and transplanting seedling area, and continuous cropping continuous cropping increases, the soil further resulted in Earth problem is got worse, and worse and worse, quality is difficult to ensure crop yielding condition, can improve above-mentioned problem well.
Microbial bacterial agent product can be divided into liquid, pulvis, granular pattern by dosage form;It can by the microbe species or functional characteristic that include It is divided into Rhizobium Inoculant, fixed nitrogen bacteria agent, phosphorus decomposing quasi-microorganism microbial inoculum, silicate microbial bacterial agent, photosynthetic bacteria microbial inoculum, organic Material decomposing agent, growth promoting bacteria agent, bush mycorrhiza agent, biological prosthetic microbial inoculum etc..
Existing microbial bacterial agent product has many deficiencies, and especially single microbial inoculum is unable to give full play out microorganism The advantage of microbial inoculum.
The prior art has the disadvantage that
(1) bacterial content of microbial bacterial agent is lower;(2) living bacteria count of microbial bacterial agent is lower;(3) microbial bacterial agent Miscellaneous bacteria rate it is high;(4) content of beary metal of microbial bacterial agent is higher;(5) shelf-life of microbial bacterial agent is short.
Summary of the invention
The present invention be solve the shortcomings of the prior art, a kind of production method of microbial bacterial agent is provided, with realize with Lower goal of the invention:
1, the bacterial content of microbial bacterial agent is improved;
2, the living bacteria count of microbial bacterial agent is improved;
3, the miscellaneous bacteria rate of microbial bacterial agent is reduced;
4, the content of beary metal of microbial bacterial agent is reduced;
5, extend the shelf-life of microbial bacterial agent.
In order to solve the above technical problems, using following technical scheme:
A kind of microbial bacterial agent, including bacillusmusilaginosiengineering, bacillus subtilis, bacillus licheniformis.
It is further improvement to above-mentioned technical proposal below:
In the microbial bacterial agent, bacillusmusilaginosiengineering viable count is 18~2,200,000,000 cfu/g, bacillus subtilis viable bacteria Number is 17~1,900,000,000 cfu/g, and bacillus licheniformis viable count is 12~1,300,000,000 cfu/g.
A kind of production method of microbial bacterial agent, including strain selection, actication of culture and spread cultivation, strain fermentation, be made into Product step.
In the actication of culture and the step that spreads cultivation, the culture medium that spreads cultivation of the activation and the use that spreads cultivation of bacillusmusilaginosiengineering Formula are as follows: 15~18 parts by weight of sweet potato powder, 12~15 parts by weight of bean cake powder, 0.6~0.8 parts by weight of sodium alginate, yeast extract 4 ~4.5 parts by weight, 0.03~0.04 parts by weight of glutamic acid, 0.03~0.05 parts by weight of bovine bile, 0.02~0.04 weight of mannitol Measure part, 0.01~0.02 parts by weight of manganese dioxide, 0.02~0.03 parts by weight of dipotassium hydrogen phosphate, 0.05~0.07 weight of magnesium sulfate Part, 1000 parts by weight of deionized water.
In the actication of culture and the step that spreads cultivation, activation and the culture medium that spreads cultivation for the use that spreads cultivation of bacillus subtilis Formula are as follows: 12~14 parts by weight of potato starch, 9~10 parts by weight of trehalose, 6~8 parts by weight of sorbitol, 4~6 weight of beef extract Measure part, 0.06~0.08 parts by weight of aspartic acid, 0.07~0.08 parts by weight of glucose, 0.02~0.04 weight of ironic citrate Part, 0.01~0.02 parts by weight of vitamin B1,0.03~0.04 parts by weight of 2,6- inositol, 0.01~0.02 weight of manganese dioxide Part, 0.03~0.05 parts by weight of dipotassium hydrogen phosphate, seven brochanites, 0.04~0.06 parts by weight, 1000 parts by weight of deionized water.
In the actication of culture and the step that spreads cultivation, activation and the culture medium that spreads cultivation for the use that spreads cultivation of bacillus licheniformis Formula are as follows: 18~20 parts by weight of lactose, 7~9 parts by weight of fish peptone, 4~5 parts by weight of sorbitol, 8~10 weight of modified starch Measure part, 0.06~0.08 parts by weight of asparagine, 0.03~0.04 parts by weight of olive oil, 0.02~0.03 weight of ironic citrate Part, 0.02~0.03 parts by weight of niacin, 0.06~0.09 parts by weight of calcium monohydrogen phosphate, 0.04~0.06 parts by weight of magnesium sulfate, boric acid 1~1.4 parts by weight, 1000 parts by weight of deionized water.
In the strain fermentation step, the temperature of bacillusmusilaginosiengineering fermentation is 34 DEG C, and inoculum concentration is fermented and cultured The 6% of matrix amount, fermentation time are 30~32 hours.
In the strain fermentation step, the fermentation temperature of fermentation of bacillus subtilis is 36 DEG C, and inoculum concentration is fermentation training The 6% of matrix amount is supported, fermentation time is 36~40 hours.
In the strain fermentation step, the fermentation temperature of bacillus licheniformis fermentation is 35 DEG C, and inoculum concentration is fermentation training The 6% of matrix amount is supported, fermentation time is 22~24 hours.
In the finished product step, the component of carrier includes medical stone, calcium carbonate, vermiculite power, and mass ratio is wheat Meal stone: calcium carbonate: vermiculite power=5:2:1.
Compared with prior art, the beneficial effects of the present invention are:
1, microbial bacterial agent of the present invention, bacterial content is high, and total bacterial content is 85~8,800,000,000 cfu/g;
2, microbial bacterial agent of the present invention, living bacteria count is high, and bacillusmusilaginosiengineering viable count is 18~2,200,000,000 cfu/g, Bacillus subtilis viable count is 17~1,900,000,000 cfu/g, and bacillus licheniformis viable count is 12~1,300,000,000 cfu/g;
3, microbial bacterial agent of the present invention, miscellaneous bacteria rate is low, and miscellaneous bacteria rate is 4.6~5.2%;
4, microbial bacterial agent of the present invention, content of beary metal is low, and arsenic and its compounds content are 6~9 mg/kg, mercury and its change Conjunction object content is 0.7~0.8 mg/kg, 9~11 mg/kg of lead and its compounds content;
5, microbial bacterial agent of the present invention, long shelf-life, shelf-life are 20~24 months.
Specific embodiment
A kind of production method of the microbial bacterial agent of embodiment 1
Step 1, strain selection
The strain of use includes:
Bacillusmusilaginosiengineering (Bacillus mucilaginosus) deposit number is ACCC10013;
Bacillus subtilis (Bacillus subtilis) deposit number be ACCC01175;
Bacillus licheniformis (Bacillus licheniformis) deposit number is ACCC11091;
Above-mentioned strain is all from commercially available.
It step 2, actication of culture and spreads cultivation
(1) activation of bacillusmusilaginosiengineering with spread cultivation
Bacillusmusilaginosiengineering strain is inoculated on plating medium, 30 DEG C are cultivated 24 hours, and activated spawn is obtained;Institute Stating plating medium is nutrient agar;
The activated spawn of bacillusmusilaginosiengineering is inoculated in the fluid nutrient medium in shaking flask, inoculum concentration is culture substrate The 5% of amount, is placed in shaking table culture case, and revolving speed is 120 revs/min, and ventilatory capacity 1:0.4,34 DEG C are cultivated 20 hours, are spread cultivation Seed liquor;
The formula of the fluid nutrient medium are as follows: sweet potato powder 15g, bean cake powder 12g, sodium alginate 0.6g, yeast extract 4g, paddy ammonia Sour 0.03g, it bovine bile 0.03g, mannitol 0.02g, manganese dioxide 0.01g, dipotassium hydrogen phosphate 0.02g, magnesium sulfate 0.05g, goes Ionized water 1kg;Adjusting pH is 7.
(2) activation of bacillus subtilis with spread cultivation
Bacillus subtilis strain is inoculated on plating medium, 32 DEG C are cultivated 36 hours, and activated spawn is obtained;It is described Plating medium is cornstarch culture medium;
The activated spawn of bacillus subtilis is inoculated in the fluid nutrient medium in shaking flask, inoculum concentration is culture medium quality 4%, be placed in shaking table culture case, revolving speed is 180 revs/min, ventilatory capacity 1:0.6,33 DEG C and cultivates 30 hours, obtains the kind that spreads cultivation Sub- liquid;
The formula of the fluid nutrient medium are as follows: potato starch 12g, trehalose 9g, sorbitol 6g, beef extract 4g, asparagus fern Propylhomoserin 0.06g, glucose 0.07g, ironic citrate 0.02g, vitamin B1 0.01g, 2,6- inositol 0.03g, manganese dioxide 0.01g, dipotassium hydrogen phosphate 0.03g, seven brochanite 0.04g, deionized water 1kg;Adjusting pH is 7.2.
(3) activation of bacillus licheniformis with spread cultivation
Bacillus licheniformis strain is inoculated on plating medium, 35 DEG C are cultivated 16 hours, and activated spawn is obtained;It is described Plating medium is beef extract-peptone agar medium;
The activated spawn of bacillus licheniformis is inoculated in the fluid nutrient medium in shaking flask, inoculum concentration is culture medium quality 4%, be placed in shaking table culture case, revolving speed is 200 revs/min, ventilatory capacity 1:0.6,33 DEG C and cultivates 22 hours, obtains the kind that spreads cultivation Sub- liquid;
The formula of the fluid nutrient medium are as follows: lactose 18g, fish peptone 7g, sorbitol 4g, modified starch 8g, Tianmen Winter amide 0.06g, olive oil 0.03g, ironic citrate 0.02g, niacin 0.02g, calcium monohydrogen phosphate 0.06g, magnesium sulfate 0.04g, boron Sour 1g, deionized water 1kg;Adjusting pH is 6.8.
Step 3, strain fermentation
(1) bacillusmusilaginosiengineering ferments
Fermentation medium sterilizing: fermentation medium each component is weighed, and is added in fermentor, is started stirring, 120rpm Stir 10min;Heating, pressurization, fermentation pressure inside the tank reach 0.105MPa, and temperature reaches 121 DEG C, and sterilize 25~30min;
The fermentation medium comprises the following components in parts by weight:
20 parts of wheat bran, 18 parts of cottonseed meal, 4 parts of maltose, 4 parts of soy meal, 1 part of beef extract, 0.5 part of bovine bile, wood 0.2 part of sugar alcohol, 0.1 part of manganese dioxide, 0.06 part of dipotassium hydrogen phosphate, 0.1 part of magnesium sulfate, 2000 parts of deionized water;Adjusting pH is 7.
Inoculation: fermentation jar temperature is down to 30 DEG C, under aseptic condition, and the kind that spreads cultivation of bacillusmusilaginosiengineering is added from inoculation mouth Sub- liquid, inoculum concentration are the 6% of fermentation medium quality;
Fermentation: after inoculation, controlling 34 DEG C of fermentation jar temperature, and 80rpm stirring is continually fed into filtrated air, avoids miscellaneous bacteria dirty Dye;Fermentation 30 hours.
(2) fermentation of bacillus subtilis
Fermentation medium sterilizing: fermentation medium each component is weighed, and is added in fermentor, is started stirring, 120rpm Stir 10min;Heating, pressurization, fermentation pressure inside the tank reach 0.105MPa, and temperature reaches 121 DEG C, and sterilize 25~30min;
The fermentation medium comprises the following components in parts by weight:
18 parts of tapioca starch, 4 parts of sucrose, 1 part of sorbitol, 2 parts of beef extract, 0.06 part of aspartic acid, 8 parts of corn flour, lemon 0.05 part of lemon acid iron, 0.03 part of vitamin B1,0.05 part of 2,6- inositol, 0.06 part of manganese dioxide, 0.1 part of dipotassium hydrogen phosphate, Seven 0.05 part of brochanites, 2000 parts of deionized water;Adjusting pH is 7.2.
Inoculation: fermentation jar temperature is down to 30 DEG C, under aseptic condition, and the seed that spreads cultivation of bacillus subtilis is added from inoculation mouth Liquid, inoculum concentration are the 6% of fermentation medium quality;
Fermentation: after inoculation, controlling 36 DEG C of fermentation jar temperature, and 90rpm stirring is continually fed into filtrated air, avoids miscellaneous bacteria dirty Dye;Fermentation 36 hours.
(3) bacillus licheniformis ferments
Fermentation medium sterilizing: fermentation medium each component is weighed, and is added in fermentor, is started stirring, 120rpm Stir 10min;Heating, pressurization, fermentation pressure inside the tank reach 0.105MPa, and temperature reaches 121 DEG C, and sterilize 25min;
The fermentation medium comprises the following components in parts by weight:
30 parts of potato starch, 1 part of yeast powder, 10 parts of cottonseed meal, 4 parts of glucose, 0.06 part of asparagine, olive oil 0.1 part, 0.1 part of ironic citrate, 0.05 part of niacin, 0.1 part of calcium monohydrogen phosphate, 0.05 part of magnesium sulfate, 1 part of boric acid, deionized water 2000 parts;Adjusting pH is 6.8.
Inoculation: fermentation jar temperature is down to 30 DEG C, under aseptic condition, and the seed that spreads cultivation of bacillus subtilis is added from inoculation mouth Liquid, inoculum concentration are the 6% of fermentation medium quality;
Fermentation: after inoculation, controlling 35 DEG C of fermentation jar temperature, and 80rpm stirring is continually fed into filtrated air, avoids miscellaneous bacteria dirty Dye;Fermentation 22 hours.
Step 4, finished product
Medical stone, calcium carbonate, vermiculite power are mixed by the weight ratio of 5:2:1, after sterilizing, as carrier;
Step 3 is fermented 3 kinds of fermentation liquids obtained, filtering obtains bacillusmusilaginosiengineering bacterium solution, bacillus subtilis bacterium Liquid, bacillus licheniformis bacterium solution;
After bacterium solution in above-mentioned 3 is mixed by the mass ratio of 1:1:1, it is sprayed on carrier, makes biodiversity percentage composition 28%。
A kind of production method of the microbial bacterial agent of embodiment 2
Step 1, strain selection
The strain of use is same as Example 1.
It step 2, actication of culture and spreads cultivation
(1) activation of bacillusmusilaginosiengineering with spread cultivation
Bacillusmusilaginosiengineering strain is inoculated on plating medium, 32 DEG C are cultivated 24 hours, and activated spawn is obtained;Institute Stating plating medium is nutrient agar;
The activated spawn of bacillusmusilaginosiengineering is inoculated in the fluid nutrient medium in shaking flask, inoculum concentration is culture substrate The 5% of amount, is placed in shaking table culture case, and revolving speed is 120 revs/min, and ventilatory capacity 1:0.4,35 DEG C are cultivated 21 hours, are spread cultivation Seed liquor;
The formula of the fluid nutrient medium are as follows: sweet potato powder 17g, bean cake powder 14g, sodium alginate 0.7g, yeast extract 4.2g, paddy Propylhomoserin 0.03g, bovine bile 0.04g, mannitol 0.04g, manganese dioxide 0.01g, dipotassium hydrogen phosphate 0.03g, magnesium sulfate 0.06g, Deionized water 1kg;Adjusting pH is 7.
(2) activation of bacillus subtilis with spread cultivation
Bacillus subtilis strain is inoculated on plating medium, 33 DEG C are cultivated 36 hours, and activated spawn is obtained;It is described Plating medium is cornstarch culture medium;
The activated spawn of bacillus subtilis is inoculated in the fluid nutrient medium in shaking flask, inoculum concentration is culture medium quality 4%, be placed in shaking table culture case, revolving speed is 180 revs/min, ventilatory capacity 1:0.6,33 DEG C and cultivates 30 hours, obtains the kind that spreads cultivation Sub- liquid;
The formula of the fluid nutrient medium are as follows: potato starch 13g, trehalose 9.5g, sorbitol 7g, beef extract 4.5g, Aspartic acid 0.07g, glucose 0.07g, ironic citrate 0.03g, vitamin B1 0.01g, 2,6- inositol 0.03g, titanium dioxide Manganese 0.02g, dipotassium hydrogen phosphate 0.04g, seven brochanite 0.05g, deionized water 1kg;Adjusting pH is 7.2.
(3) activation of bacillus licheniformis with spread cultivation
Bacillus licheniformis strain is inoculated on plating medium, 35 DEG C are cultivated 16 hours, and activated spawn is obtained;It is described Plating medium is beef extract-peptone agar medium;
The activated spawn of bacillus licheniformis is inoculated in the fluid nutrient medium in shaking flask, inoculum concentration is culture medium quality 4%, be placed in shaking table culture case, revolving speed is 200 revs/min, ventilatory capacity 1:0.6,33 DEG C and cultivates 22 hours, obtains the kind that spreads cultivation Sub- liquid;
The formula of the fluid nutrient medium are as follows: lactose 19g, fish peptone 8g, sorbitol 4.5g, modified starch 9g, day Asparagine 0.07g, olive oil 0.03g, ironic citrate 0.03g, niacin 0.02g, calcium monohydrogen phosphate 0.08g, magnesium sulfate 0.05g, Boric acid 1.2g, deionized water 1kg;Adjusting pH is 6.8.
Step 3, strain fermentation
(1) bacillusmusilaginosiengineering ferments
Fermentation medium sterilizing: fermentation medium each component is weighed, and is added in fermentor, is started stirring, 120rpm Stir 10min;Heating, pressurization, fermentation pressure inside the tank reach 0.105MPa, and temperature reaches 121 DEG C, and sterilize 25~30min;
The fermentation medium comprises the following components in parts by weight:
22 parts of wheat bran, 18 parts of cottonseed meal, 5 parts of maltose, 4.2 parts of soy meal, 1.5 parts of beef extract, bovine bile 0.5 Part, 0.3 part of xylitol, 0.1 part of manganese dioxide, 0.07 part of dipotassium hydrogen phosphate, 0.1 part of magnesium sulfate, 2000 parts of deionized water;Adjust pH It is 7.
Inoculation: fermentation jar temperature is down to 33 DEG C, under aseptic condition, and the kind that spreads cultivation of bacillusmusilaginosiengineering is added from inoculation mouth Sub- liquid, inoculum concentration are the 6% of fermentation medium quality;
Fermentation: after inoculation, controlling 34 DEG C of fermentation jar temperature, and 80rpm stirring is continually fed into filtrated air, avoids miscellaneous bacteria dirty Dye;Fermentation 31 hours.
(2) fermentation of bacillus subtilis
Fermentation medium sterilizing: fermentation medium each component is weighed, and is added in fermentor, is started stirring, 120rpm Stir 10min;Heating, pressurization, fermentation pressure inside the tank reach 0.105MPa, and temperature reaches 121 DEG C, and sterilize 25~30min;
The fermentation medium comprises the following components in parts by weight:
19 parts of tapioca starch, 5 parts of sucrose, 1.5 parts of sorbitol, 2 parts of beef extract, 0.07 part of aspartic acid, 10 parts of corn flour, 0.06 part of ironic citrate, 0.04 part of vitamin B1,0.06 part of 2,6- inositol, 0.07 part of manganese dioxide, dipotassium hydrogen phosphate 0.15 Part, 0.06 part of seven brochanite, 2000 parts of deionized water;Adjusting pH is 7.2.
Inoculation: fermentation jar temperature is down to 32 DEG C, under aseptic condition, and the seed that spreads cultivation of bacillus subtilis is added from inoculation mouth Liquid, inoculum concentration are the 6% of fermentation medium quality;
Fermentation: after inoculation, controlling 36 DEG C of fermentation jar temperature, and 90rpm stirring is continually fed into filtrated air, avoids miscellaneous bacteria dirty Dye;Fermentation 38 hours.
(3) bacillus licheniformis ferments
Fermentation medium sterilizing: fermentation medium each component is weighed, and is added in fermentor, is started stirring, 120rpm Stir 10min;Heating, pressurization, fermentation pressure inside the tank reach 0.105MPa, and temperature reaches 121 DEG C, and sterilize 25~30min;
The fermentation medium comprises the following components in parts by weight:
32 parts of potato starch, 1.5 parts of yeast powder, 13 parts of cottonseed meal, 5 parts of glucose, 0.07 part of asparagine, olive Oily 0.1 part, 0.2 part of ironic citrate, 0.05 part of niacin, 0.2 part of calcium monohydrogen phosphate, 0.07 part of magnesium sulfate, 1.2 parts of boric acid, deionization 2000 parts of water;Adjusting pH is 6.8.
Inoculation: fermentation jar temperature is down to 33 DEG C, under aseptic condition, and the seed that spreads cultivation of bacillus subtilis is added from inoculation mouth Liquid, inoculum concentration are the 6% of fermentation medium quality;
Fermentation: after inoculation, controlling 35 DEG C of fermentation jar temperature, and 80rpm stirring is continually fed into filtrated air, avoids miscellaneous bacteria dirty Dye;Fermentation 23 hours.
Step 4, finished product
It is identical as the operating method of embodiment 1.
A kind of production method of the microbial bacterial agent of embodiment 3
Step 1, strain selection
The strain of use is same as Example 1.
It step 2, actication of culture and spreads cultivation
(1) activation of bacillusmusilaginosiengineering with spread cultivation
Bacillusmusilaginosiengineering strain is inoculated on plating medium, 34 DEG C are cultivated 24 hours, and activated spawn is obtained;Institute Stating plating medium is nutrient agar;
The activated spawn of bacillusmusilaginosiengineering is inoculated in the fluid nutrient medium in shaking flask, inoculum concentration is culture substrate The 5% of amount, is placed in shaking table culture case, and revolving speed is 120 revs/min, and ventilatory capacity 1:0.4,36 DEG C are cultivated 22 hours, are spread cultivation Seed liquor;
The formula of the fluid nutrient medium are as follows: sweet potato powder 18g, bean cake powder 15g, sodium alginate 0.8g, yeast extract 4.5g, paddy Propylhomoserin 0.04g, bovine bile 0.05g, mannitol 0.04g, manganese dioxide 0.02g, dipotassium hydrogen phosphate 0.03g, magnesium sulfate 0.07g, Deionized water 1kg;Adjusting pH is 7.
(2) activation of bacillus subtilis with spread cultivation
Bacillus subtilis strain is inoculated on plating medium, 33 DEG C are cultivated 36 hours, and activated spawn is obtained;It is described Plating medium is cornstarch culture medium;
The activated spawn of bacillus subtilis is inoculated in the fluid nutrient medium in shaking flask, inoculum concentration is culture medium quality 4%, be placed in shaking table culture case, revolving speed is 180 revs/min, ventilatory capacity 1:0.6,33 DEG C and cultivates 30 hours, obtains the kind that spreads cultivation Sub- liquid;
The formula of the fluid nutrient medium are as follows: potato starch 14g, trehalose 10g, sorbitol 8g, beef extract 6g, asparagus fern Propylhomoserin 0.08g, glucose 0.08g, ironic citrate 0.04g, vitamin B1 0.02g, 2,6- inositol 0.04g, manganese dioxide 0.02g, dipotassium hydrogen phosphate 0.05g, seven brochanite 0.06g, deionized water 1kg;Adjusting pH is 7.2.
(3) activation of bacillus licheniformis with spread cultivation
Bacillus licheniformis strain is inoculated on plating medium, 35 DEG C are cultivated 16 hours, and activated spawn is obtained;It is described Plating medium is beef extract-peptone agar medium;
The activated spawn of bacillus licheniformis is inoculated in the fluid nutrient medium in shaking flask, inoculum concentration is culture medium quality 4%, be placed in shaking table culture case, revolving speed is 200 revs/min, ventilatory capacity 1:0.6,33 DEG C and cultivates 22 hours, obtains the kind that spreads cultivation Sub- liquid;
The formula of the fluid nutrient medium are as follows: lactose 20g, fish peptone 9g, sorbitol 5g, modified starch 10g, Tianmen Winter amide 0.08g, olive oil 0.04g, ironic citrate 0.03g, niacin 0.03g, calcium monohydrogen phosphate 0.09g, magnesium sulfate 0.06g, boron Sour 1.4g, deionized water 1kg;Adjusting pH is 6.8.
Step 3, strain fermentation
(1) bacillusmusilaginosiengineering ferments
Fermentation medium sterilizing: fermentation medium each component is weighed, and is added in fermentor, is started stirring, 120rpm Stir 10min;Heating, pressurization, fermentation pressure inside the tank reach 0.105MPa, and temperature reaches 121 DEG C, and sterilize 25~30min;
The fermentation medium comprises the following components in parts by weight:
23 parts of wheat bran, 19 parts of cottonseed meal, 6 parts of maltose, 4.5 parts of soy meal, 2 parts of beef extract, 0.6 part of bovine bile, 0.4 part of xylitol, 0.2 part of manganese dioxide, 0.09 part of dipotassium hydrogen phosphate, 0.2 part of magnesium sulfate, 2000 parts of deionized water;Adjust pH be 7。
Inoculation: fermentation jar temperature is down to 35 DEG C, under aseptic condition, and the kind that spreads cultivation of bacillusmusilaginosiengineering is added from inoculation mouth Sub- liquid, inoculum concentration are the 6% of fermentation medium quality;
Fermentation: after inoculation, controlling 34 DEG C of fermentation jar temperature, and 80rpm stirring is continually fed into filtrated air, avoids miscellaneous bacteria dirty Dye;Fermentation 32 hours.
(2) fermentation of bacillus subtilis
Fermentation medium sterilizing: fermentation medium each component is weighed, and is added in fermentor, is started stirring, 120rpm Stir 10min;Heating, pressurization, fermentation pressure inside the tank reach 0.105MPa, and temperature reaches 121 DEG C, and sterilize 25~30min;
The fermentation medium comprises the following components in parts by weight:
20 parts of tapioca starch, 6 parts of sucrose, 2 parts of sorbitol, 3 parts of beef extract, 0.08 part of aspartic acid, 11 parts of corn flour, lemon 0.07 part of lemon acid iron, 0.04 part of vitamin B1,0.07 part of 2,6- inositol, 0.08 part of manganese dioxide, 0.2 part of dipotassium hydrogen phosphate, Seven 0.07 part of brochanites, 2000 parts of deionized water;Adjusting pH is 7.2.
Inoculation: fermentation jar temperature is down to 35 DEG C, under aseptic condition, and the seed that spreads cultivation of bacillus subtilis is added from inoculation mouth Liquid, inoculum concentration are the 6% of fermentation medium quality;
Fermentation: after inoculation, controlling 36 DEG C of fermentation jar temperature, and 90rpm stirring is continually fed into filtrated air, avoids miscellaneous bacteria dirty Dye;Fermentation 40 hours.
(3) bacillus licheniformis ferments
Fermentation medium sterilizing: fermentation medium each component is weighed, and is added in fermentor, is started stirring, 120rpm Stir 10min;Heating, pressurization, fermentation pressure inside the tank reach 0.105MPa, and temperature reaches 121 DEG C, and sterilize 25~30min;
The fermentation medium comprises the following components in parts by weight:
34 parts of potato starch, 2 parts of yeast powder, 15 parts of cottonseed meal, 6 parts of glucose, 0.08 part of asparagine, olive oil 0.2 part, 0.2 part of ironic citrate, 0.06 part of niacin, 0.2 part of calcium monohydrogen phosphate, 0.08 part of magnesium sulfate, 1.4 parts of boric acid, deionized water 2000 parts;Adjusting pH is 6.8.
Inoculation: fermentation jar temperature is down to 35 DEG C, under aseptic condition, and the seed that spreads cultivation of bacillus subtilis is added from inoculation mouth Liquid, inoculum concentration are the 6% of fermentation medium quality;
Fermentation: after inoculation, controlling 35 DEG C of fermentation jar temperature, and 80rpm stirring is continually fed into filtrated air, avoids miscellaneous bacteria dirty Dye;Fermentation 24 hours.
Step 4, finished product
It is identical as the operating method of embodiment 1.
The microbial bacterial agent of embodiment 1-3 preparation is detected;Testing result is shown in Table 1;
Table 1
Seen from table 1, microbial bacterial agent of the present invention, total bacterial content are 85~8,800,000,000 cfu/g, bacillusmusilaginosiengineering viable bacteria Number be 18~2,200,000,000 cfu/g, bacillus subtilis viable count be 17~1,900,000,000 cfu/g, bacillus licheniformis viable count be 12~ 1300000000 cfu/g, miscellaneous bacteria rate are 4.6~5.2%, and fineness is 120 mesh, and arsenic and its compounds content are 6~9 mg/kg, mercury and its change Conjunction object content is 0.7~0.8 mg/kg, and 9~11 mg/kg of lead and its compounds content, the shelf-life is 20~24 months.
Effect experiment:
The microbial bacterial agent of embodiment 1-3 is applied in eggplant plantation plot, applies 40~50kg of microbial bacterial agent per acre, With topsoil mix, plant eggplant 1 year, soil property be improved significantly, specific targets are shown in Table 2;
Performance indicator after 2 soil improvement of table
As can be seen from Table 2, after using microbial bacterial agent, soil property be improved significantly, soil pH is 6.8~7.2, soil Bulk density is 1.36~1.41g/cm3, ventilation voidage is 20~23%;Heavy metal in soil content is substantially reduced, content of microorganisms It significantly improves.
After microbial bacterial agent, eggplant yield improves 15~18%, and commodity fruit rate improves 23~25%, malformed fruit rate Reduce 9~13%.
Finally, it should be noted that the foregoing is only a preferred embodiment of the present invention, it is not intended to restrict the invention, It, for those skilled in the art, still can be with although describing the invention in detail with reference to the foregoing embodiments It modifies the technical solutions described in the foregoing embodiments or equivalent replacement of some of the technical features.It is all Within the spirit and principles in the present invention, any modification, equivalent replacement, improvement and so on should be included in guarantor of the invention Within the scope of shield.The above is the citing of best mode for carrying out the invention, wherein the part that do not address in detail is this field The common knowledge of those of ordinary skill;Protection scope of the present invention is based on the contents of the claims, any based on of the invention Technical inspiration and the equivalent transformation carried out, also within protection scope of the present invention.

Claims (1)

1. a kind of production method of microbial bacterial agent, it is characterised in that: including strain selection, actication of culture and spread cultivation, strain hair Ferment, finished product step;
In the actication of culture and the step that spreads cultivation, the activation of bacillusmusilaginosiengineering and matching for the culture medium that spreads cultivation for the use that spreads cultivation Side are as follows: 15~18 parts by weight of sweet potato powder, 12~15 parts by weight of bean cake powder, 0.6~0.8 parts by weight of sodium alginate, yeast extract 4~ 4.5 parts by weight, 0.03~0.04 parts by weight of glutamic acid, 0.03~0.05 parts by weight of bovine bile, 0.02~0.04 weight of mannitol Part, 0.01~0.02 parts by weight of manganese dioxide, 0.02~0.03 parts by weight of dipotassium hydrogen phosphate, 0.05~0.07 weight of magnesium sulfate Part, 1000 parts by weight of deionized water;
In the actication of culture and the step that spreads cultivation, the activation of bacillus subtilis and the formula of the culture medium that spreads cultivation for the use that spreads cultivation Are as follows: 12~14 parts by weight of potato starch, 9~10 parts by weight of trehalose, 6~8 parts by weight of sorbitol, 4~6 weight of beef extract Part, 0.06~0.08 parts by weight of aspartic acid, 0.07~0.08 parts by weight of glucose, 0.02~0.04 parts by weight of ironic citrate, 0.01~0.02 parts by weight of vitamin B1,0.03~0.04 parts by weight of 2,6- inositol, 0.01~0.02 parts by weight of manganese dioxide, 0.03~0.05 parts by weight of dipotassium hydrogen phosphate, seven brochanites, 0.04~0.06 parts by weight, 1000 parts by weight of deionized water;
In the actication of culture and the step that spreads cultivation, the activation of bacillus licheniformis and the formula of the culture medium that spreads cultivation for the use that spreads cultivation Are as follows: 18~20 parts by weight of lactose, 7~9 parts by weight of fish peptone, 4~5 parts by weight of sorbitol, 8~10 weight of modified starch Part, 0.06~0.08 parts by weight of asparagine, 0.03~0.04 parts by weight of olive oil, 0.02~0.03 weight of ironic citrate Part, 0.02~0.03 parts by weight of niacin, 0.06~0.09 parts by weight of calcium monohydrogen phosphate, 0.04~0.06 parts by weight of magnesium sulfate, boric acid 1~1.4 parts by weight, 1000 parts by weight of deionized water;
In the strain fermentation step, the temperature of bacillusmusilaginosiengineering fermentation is 34 DEG C, and inoculum concentration is fermented and cultured matrix The 6% of amount, fermentation time are 30~32 hours;
In the strain fermentation step, the fermentation temperature of fermentation of bacillus subtilis is 36 DEG C, and inoculum concentration is fermentation medium The 6% of quality, fermentation time are 36~40 hours;
In the strain fermentation step, the fermentation temperature of bacillus licheniformis fermentation is 35 DEG C, and inoculum concentration is fermentation medium The 6% of quality, fermentation time are 22~24 hours;
In the finished product step, the component of carrier includes medical stone, calcium carbonate, vermiculite power, and mass ratio is medical stone: Calcium carbonate: vermiculite power=5:2:1;
In microbial bacterial agent obtained, bacillusmusilaginosiengineering viable count is 18~2,200,000,000 cfu/g, bacillus subtilis viable count For 17~1,900,000,000 cfu/g, bacillus licheniformis viable count is 12~1,300,000,000 cfu/g;Miscellaneous bacteria rate is 4.6~5.2%;Arsenic and its change Conjunction object content is 6~9 mg/kg, and mercury and mercuric compounds content is 0.7~0.8 mg/kg, lead and its compounds content 9~11 mg/kg;Shelf-life is 20~24 months;
The bacillusmusilaginosiengineering (Bacillus mucilaginosus) deposit number be ACCC10013;
The bacillus subtilis (Bacillus subtilis) deposit number be ACCC01175;
The bacillus licheniformis (Bacillus licheniformis) deposit number be ACCC11091.
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Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN102229900A (en) * 2011-05-20 2011-11-02 周剑平 Compound probiotics preparation for biological organic fertilizers and preparation method thereof
CN102816726A (en) * 2012-09-10 2012-12-12 临沂中磷生物科技有限公司 Novel compound microbial agent and applications thereof

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN102229900A (en) * 2011-05-20 2011-11-02 周剑平 Compound probiotics preparation for biological organic fertilizers and preparation method thereof
CN102816726A (en) * 2012-09-10 2012-12-12 临沂中磷生物科技有限公司 Novel compound microbial agent and applications thereof

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
微生物肥料菌种应用与效果分析;陈慧君;《中国优秀硕士学位论文数据库 农业科技辑》;20140215;全文,特别是摘要
微生物肥料菌种的应用及效果分析;李晓娇;《农业与技术》;20160108;34

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