CN115568532A - Preparation method and device of daily ration microbial agent - Google Patents
Preparation method and device of daily ration microbial agent Download PDFInfo
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- 238000002360 preparation method Methods 0.000 title claims abstract description 54
- 230000000813 microbial effect Effects 0.000 title claims abstract description 50
- 239000000284 extract Substances 0.000 claims abstract description 81
- 241000193749 Bacillus coagulans Species 0.000 claims abstract description 71
- 229940054340 bacillus coagulans Drugs 0.000 claims abstract description 71
- 238000010438 heat treatment Methods 0.000 claims abstract description 38
- 239000003795 chemical substances by application Substances 0.000 claims abstract description 37
- 238000012258 culturing Methods 0.000 claims abstract description 23
- 238000000855 fermentation Methods 0.000 claims abstract description 18
- 230000004151 fermentation Effects 0.000 claims abstract description 18
- 238000012216 screening Methods 0.000 claims abstract description 13
- 238000000034 method Methods 0.000 claims abstract description 12
- 239000000126 substance Substances 0.000 claims abstract description 11
- 239000002994 raw material Substances 0.000 claims abstract description 5
- 239000001963 growth medium Substances 0.000 claims description 91
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- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 30
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 claims description 22
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 20
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 20
- 239000002609 medium Substances 0.000 claims description 18
- 238000002156 mixing Methods 0.000 claims description 17
- 238000001035 drying Methods 0.000 claims description 15
- 238000001914 filtration Methods 0.000 claims description 15
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- 241000283913 Bacillus coagulans DSM 1 = ATCC 7050 Species 0.000 claims description 13
- 229920001817 Agar Polymers 0.000 claims description 11
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 claims description 11
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- 239000003242 anti bacterial agent Substances 0.000 abstract description 2
- 229940088710 antibiotic agent Drugs 0.000 abstract description 2
- 241001465754 Metazoa Species 0.000 description 6
- 239000003674 animal food additive Substances 0.000 description 3
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- A23K10/10—Animal feeding-stuffs obtained by microbiological or biochemical processes
- A23K10/16—Addition of microorganisms or extracts thereof, e.g. single-cell proteins, to feeding-stuff compositions
- A23K10/18—Addition of microorganisms or extracts thereof, e.g. single-cell proteins, to feeding-stuff compositions of live microorganisms
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Abstract
The invention relates to the technical field of daily ration microbial agents, in particular to a preparation method and a device of a daily ration microbial agent, wherein the method comprises the following steps: preparing a basic substance extract by taking radix bupleuri, rhizoma anemarrhenae and orange peel as raw materials; culturing, screening and improving Bacillus coagulans, inoculating to fermentation culture solution prepared from bupleuri radix, rhizoma anemarrhenae and pericarpium Citri Tangerinae as raw materials and used as basic extract to obtain daily ration microbial agent; the device comprises a shell, a pot body, a heating pipe, a thermal constant temperature controller and a pipe fitting, wherein the pot body, the heating pipe, the thermal constant temperature controller and the pipe fitting are arranged on the shell; the whole process is reasonable in design, and the prepared daily ration microbial agent can effectively replace the traditional antibiotics, so that the preparation method is more environment-friendly; the device has the advantages of simple integral structure, high efficiency and convenience in actual use, and is suitable for large-scale popularization.
Description
Technical Field
The invention relates to the technical field of daily ration microbial agents, in particular to a preparation method and a preparation device of a daily ration microbial agent.
Background
The animal husbandry industry is one of the major components of agriculture. Feed, which is a crucial factor in the animal husbandry, is generally referred to as plant feed. The daily ration is the total amount of various feeds which are required by various nutrient substances for one animal day and night (one day).
The feed industry in China is a new industry, and the growth and development process of the feed industry is basically synchronous with the reform and open process. In recent years, more and more new feeds have been designed and developed to meet the needs of animal husbandry. Wherein the microbial feed is a mainstream product in the feed industry. The microbial feed takes microbes and complex enzyme as biological feed starter strains, and raw materials of the microbial feed are converted into microbial mycoprotein, bioactive small peptide amino acid, microbial active probiotics and a complex enzyme preparation which are integrated into a biological fermentation feed. The product not only can make up amino acid which is easy to be lacked in conventional feed, but also can quickly convert the nutrient components of other raw feed materials to achieve the effect of enhancing digestion, absorption and utilization.
The microbial feed additive is prepared with normal microbe and its metabolic product or growth promoter, and has the functions of replenishing, regulating or maintaining the micro ecological balance in animal's intestinal tract, preventing and treating diseases, promoting health and raising production performance. The microbial feed additive comprises normal microbe members, especially dominant microbe groups, and also comprises a preparation prepared from substances capable of promoting the growth and reproduction of the normal microbe groups, and can generate certain biological effect or ecological effect.
Disclosure of Invention
Aiming at the defects in the prior art, the invention provides a preparation method and a device of a daily ration microbial agent.
The technical scheme of the invention is as follows: a preparation method of a daily ration microbial agent comprises the following steps:
s1, preparation of basic material extract
The Chinese thorowax root, the rhizoma anemarrhenae and the orange peel are mixed according to the mass percentage of 30-60: 12-17, and extracting the extract after mixing the raw materials in the ratio of 20-75 to obtain a basic substance extract;
s2, culture of Bacillus coagulans
Culturing the bacillus coagulans NRS-609 to obtain bacillus coagulans;
s3, screening of Bacillus coagulans
S3-1, preparation of improved LB culture medium
According to the formula: preparing an improved LB culture medium by 10g/L of tryptone, 10g/L of sodium chloride, 5g/L of yeast extract, 5g/L of glucose and X g/L of basic extract, and adjusting the pH of the culture medium to 7.0-7.4 by NaOH;
the improved LB culture medium comprises a primary improved LB culture medium, a secondary improved LB culture medium and a third improved LB culture medium; the value of the primary improved LB culture medium X, the value of the secondary improved LB culture medium X and the value of the third improved LB culture medium X are increased in sequence;
s3-2 screening of improved bacillus coagulans
Inoculating the single colony bacillus coagulans obtained in the S2 into an initial generation improved LB culture medium according to the inoculation amount of 1% of the volume ratio, culturing for 24 hours at 37 ℃, diluting and coating, and selecting the initial generation bacillus coagulans with good growth; then sequentially inoculating the bacillus coagulans into a secondary generation LB culture medium for culture and selection and a third generation LB culture medium for culture and selection to obtain the screened improved bacillus coagulans;
s4, preparation of daily ration microbial agent
S4-1, preparation of fermentation culture solution
According to the formula: tryptone 10g/L, sodium chloride 10g/L, yeast extract 5g/L, glucose 5g/L, K 2 HPO 4 2g/L of agar powder, 20g/L of agar powder and 25-30 g/L of a basic substance extract are used for preparing a fermentation culture solution, and NaOH is used for adjusting the pH value of the culture medium to 7.0-7.4;
s4-2, and preparation of microbial agent for daily ration
Inoculating the improved bacillus coagulans into a fermentation culture solution according to the inoculation amount of 2-3% of the volume ratio, culturing and fermenting for 2-3 d at 37 ℃, drying, and collecting the dried product to obtain the daily ration microbial agent.
Further, the preparation of the base extract of S1 comprises the following specific steps:
s1-1-1, mixing 30-60 mass percent of radix bupleuri, rhizoma anemarrhenae and orange peel: 12-17, washing, crushing, adding 5-8 times of purified water, heating and extracting for 3-6 hours at the temperature of 60-65 ℃, cooling the extracting solution to room temperature, and filtering to obtain an extracting solution;
s1-1-2, concentrating the extracting solution to 25-40% of the original volume, dissolving with ethanol, desolventizing with composite resin, and drying to obtain a basic extract containing saponin extract and flavanone extract; the microbial agent prepared by taking the basic extract as a reference is used for replacing a feed additive in the daily feed, and the method has the advantages of environmental protection and better performance.
Further, the preparation method of the base extract of S1 comprises the following specific steps:
s1-2-1, mixing 30-60 mass percent of radix bupleuri, rhizoma anemarrhenae and orange peel: 12-17, washing, crushing, adding 5-8 times of purified water, carrying out primary heating extraction for 3-6 hours at the temperature of 60-65 ℃, and filtering after the extract is cooled to room temperature to obtain a primary extract;
s1-2-2, adding 2-3 times of purified water into the filter residue again, carrying out secondary heating extraction for 0.5-1 h at the temperature of 65-75 ℃, and filtering after the extracting solution is cooled to room temperature to obtain a secondary extracting solution;
s1-2-3, mixing the primary extract and the secondary extract, concentrating to 25-40% of the original volume, dissolving with ethanol, desolventizing with composite resin, and drying to obtain a basic extract containing saponin extract and flavanone extract;
the extraction rate can be effectively improved by using a double-extraction mode.
Further, the specific steps of the culture of the bacillus coagulans S2 are as follows: activating the strain of bacillus coagulans NRS-609, transferring the bacillus coagulans into an LB culture medium, culturing the bacillus coagulans at 37 ℃ for 24 hours, then diluting and coating the bacillus coagulans, and selecting well-grown single-colony bacillus coagulans; the bacillus coagulans is a strain lacking in toxigenic potential, and can be used as a daily ration microbial agent to be applied to daily ration feed to effectively inhibit diseases such as diarrhea of animals.
Further, the deposit number of the Bacillus coagulans NRS-609 is Bio-74771 in S2.
Further, the value of the primary modified LB medium X in S3-1 is 3, the value of the secondary modified LB medium X is 8, and the value of the third modified LB medium X is 15.
Furthermore, the value of the primary modified LB medium X in S3-1 is 5, the value of the secondary modified LB medium X is 10, and the value of the third modified LB medium X is 15.
Further, the value of the primary modified LB medium X in S3-1 is 8, the value of the secondary modified LB medium X is 12, and the value of the tertiary modified LB medium X is 20.
Further, the preparation device of the daily ration microbial inoculant comprises a shell, a pot body, a heating pipe, a thermal constant temperature controller and a pipe piece, wherein the pot body is installed on the shell, the heating pipe is installed inside the shell and used for heating the pot body, the thermal constant temperature controller is installed on the shell and used for carrying out constant temperature control on the pot body, and the pipe piece is used for carrying out water guiding/draining treatment on the pot body; the pan body comprises a first heating pan for assisting in the preparation of the base extract and a second heating pan for assisting in the culture of microorganisms; when in use, the extract of the basic substance is heated and extracted in a first heating pot; culturing and screening bacillus coagulans in a second heating pot, and heating the bacillus coagulans in a water bath for screening improved bacillus coagulans; has the advantages of high efficiency and convenience in actual use.
Compared with the prior art, the invention has the beneficial effects that: the invention has reasonable overall process design, prepares the daily ration microbial agent based on the basic material extract rich in saponin extract and flavanone extract after improving and treating the bacillus coagulans, can effectively replace the traditional antibiotics, is more environment-friendly, can effectively inhibit diseases such as diarrhea and the like, and has positive promotion effect on the breeding industry; the device has the advantages of simple integral structure, high efficiency and convenience in actual use, and suitability for mass popularization.
Reference numerals
FIG. 1 is a flow diagram of the process of the present invention;
FIG. 2 is a schematic view of the construction of a production apparatus of the present invention;
wherein, 1-shell, 2-pot body, 21-first heating pot, 22-second heating pot.
Detailed Description
Example 1
The preparation method of the daily ration microbial agent shown in figure 1 comprises the following steps:
s1, preparation of basic material extract
S1-1-1, mixing radix bupleuri, rhizoma anemarrhenae and orange peel according to a mass percentage of 30:12, cleaning, crushing, adding 5 times of purified water, heating and extracting for 3 hours at the temperature of 60 ℃, cooling the extracting solution to room temperature, and filtering to obtain an extracting solution;
s1-1-2, concentrating the extracting solution to 25% of the original volume, dissolving with ethanol, desolventizing with composite resin, and drying to obtain a basic extract containing saponin extract and flavanone extract;
s2, culturing of Bacillus coagulans
Activating the strain of bacillus coagulans NRS-609, transferring the bacillus coagulans into an LB culture medium, culturing the bacillus coagulans at 37 ℃ for 24 hours, then diluting and coating the bacillus coagulans, and selecting well-grown single-colony bacillus coagulans; wherein the preservation number of the bacillus coagulans NRS-609 is Bio-74771;
s3, screening of Bacillus coagulans
S3-1, preparation of improved LB culture medium
According to the formula: preparing a modified LB culture medium by 10g/L tryptone, 10g/L sodium chloride, 5g/L yeast extract, 5g/L glucose and X g/L basic extract, and adjusting the pH of the culture medium to 7.0 by NaOH;
the improved LB culture medium comprises a primary improved LB culture medium, a secondary improved LB culture medium and a tertiary improved LB culture medium; the value of the primary improved LB culture medium X, the value of the secondary improved LB culture medium X and the value of the third improved LB culture medium X are increased in sequence; the value of the primary improved LB culture medium X is 3, the value of the secondary improved LB culture medium X is 8, and the value of the third improved LB culture medium X is 15;
s3-2 screening of improved bacillus coagulans
Inoculating the single colony bacillus coagulans obtained in the S2 into an initial generation improved LB culture medium according to the inoculation amount of 1% of the volume ratio, culturing for 24 hours at 37 ℃, diluting and coating, and selecting the initial generation bacillus coagulans with good growth; then sequentially inoculating the bacillus coagulans into a secondary generation LB culture medium for culture and selection and a third generation LB culture medium for culture and selection to obtain the screened improved bacillus coagulans;
s4, preparation of daily ration microbial agent
S4-1, preparation of fermentation culture solution
According to the formula: tryptone 10g/L, sodium chloride 10g/L, yeast extract 5g/L, glucose 5g/L, K 2 HPO 4 2g/L of agar powder, 20g/L of agar powder and 25g/L of a basic substance extract are used for preparing a fermentation culture solution, and the pH value of the culture medium is adjusted to 7.0 by NaOH;
s4-2, preparation of microbial agent for daily ration
Inoculating the improved bacillus coagulans into a fermentation culture solution according to the inoculation amount of 2% of the volume ratio, culturing and fermenting for 2d at the temperature of 37 ℃, drying, and collecting the dried product to obtain the daily ration microbial agent.
As shown in fig. 2, the preparation device used in the preparation process of the daily ration microbial inoculant in the embodiment comprises a shell 1, a pot body 2 mounted on the shell 1, a heating pipe mounted inside the shell 1 and used for heating the pot body 2, a thermal constant temperature controller mounted on the shell 1 and used for performing constant temperature control on the pot body 2, and a pipe fitting used for conducting water guiding/draining treatment on the pot body 2; the pot body 2 comprises a first heating pot 21 for assisting the preparation of the base extract and a second heating pot 22 for assisting the culture of the microorganism; when in use, the bupleurum, the rhizoma anemarrhenae and the orange peel are heated and extracted in the first heating pot 21; the bacillus coagulans is cultured, screened and the modified bacillus coagulans is screened in a second heating pan 22 by heating in a water bath.
Example 2
The preparation method of the daily ration microbial agent shown in figure 1 comprises the following steps:
s1, preparation of basic material extract
1-1-1, mixing radix bupleuri, rhizoma anemarrhenae and orange peel according to a mass percentage of 40:13, cleaning, crushing, adding 6 times of purified water, heating and extracting for 5 hours at the temperature of 60 ℃, cooling the extracting solution to room temperature, and filtering to obtain an extracting solution;
s1-1-2, concentrating the extracting solution to 30% of the original volume, dissolving with ethanol, desolventizing with composite resin, and drying to obtain a basic extract containing saponin extract and flavanone extract;
s2, culture of Bacillus coagulans
Activating the strain of bacillus coagulans NRS-609, transferring the bacillus coagulans into an LB culture medium, culturing the bacillus coagulans at 37 ℃ for 24 hours, then diluting and coating the bacillus coagulans, and selecting well-grown single-colony bacillus coagulans; wherein the preservation number of the bacillus coagulans NRS-609 is Bio-74771;
s3, screening of Bacillus coagulans
S3-1, preparation of improved LB culture medium
According to the formula: preparing a modified LB culture medium by 10g/L tryptone, 10g/L sodium chloride, 5g/L yeast extract, 5g/L glucose and X g/L basic extract, and adjusting the pH of the culture medium to 7.2 by NaOH;
the improved LB culture medium comprises a primary improved LB culture medium, a secondary improved LB culture medium and a third improved LB culture medium; the value of the primary improved LB culture medium X, the value of the secondary improved LB culture medium X and the value of the third improved LB culture medium X are increased in sequence; the value of the primary modified LB culture medium X is 3, the value of the secondary modified LB culture medium X is 8, and the value of the third modified LB culture medium X is 15;
s3-2 screening of modified Bacillus coagulans
Inoculating the single colony bacillus coagulans obtained in the S2 into an initial generation improved LB culture medium according to the inoculation amount of 1% of the volume ratio, culturing for 24 hours at 37 ℃, diluting and coating, and selecting the initial generation bacillus coagulans with good growth; then sequentially inoculating the bacillus coagulans to a secondary generation LB culture medium for culture and selection and a third generation LB culture medium for culture and selection to obtain the screened improved bacillus coagulans;
s4, preparation of daily ration microbial agent
S4-1, preparation of fermentation culture solution
According to the formula: tryptone 10g/L, sodium chloride 10g/L, yeast extract 5g/L, glucose 5g/L, K 2 HPO 4 2g/L of agar powder, 20g/L of agar powder and 28g/L of a basic substance extract are used for preparing a fermentation culture solution, and the pH value of the culture medium is adjusted to 7.2 by NaOH;
s4-2, preparation of microbial agent for daily ration
Inoculating the improved bacillus coagulans into a fermentation culture solution according to the inoculation amount of 2% of the volume ratio, culturing and fermenting for 2.5 days at the temperature of 37 ℃, drying, and collecting the dried product to obtain the daily ration microbial agent.
Example 3
The preparation method of the daily ration microbial agent shown in figure 1 comprises the following steps:
s1, preparation of basic material extract
S1-1-1, mixing the following components in percentage by mass: 17, cleaning, crushing, adding 8 times of purified water, heating and extracting for 6 hours at the temperature of 65 ℃, cooling the extracting solution to room temperature, and filtering to obtain an extracting solution;
s1-1-2, concentrating the extracting solution to 40% of the original volume, dissolving with ethanol, desolventizing with composite resin, and drying to obtain a basic extract containing saponin extract and flavanone extract;
s2, culturing of Bacillus coagulans
Activating the strain of bacillus coagulans NRS-609, transferring the bacillus coagulans into an LB culture medium, culturing the bacillus coagulans at 37 ℃ for 24 hours, then diluting and coating the bacillus coagulans, and selecting well-grown single-colony bacillus coagulans; wherein the preservation number of the bacillus coagulans NRS-609 is Bio-74771;
s3, screening of Bacillus coagulans
S3-1, preparation of improved LB culture medium
According to the formula: preparing a modified LB culture medium by 10g/L tryptone, 10g/L sodium chloride, 5g/L yeast extract, 5g/L glucose and X g/L basic extract, and adjusting the pH of the culture medium to 7.4 by NaOH;
the improved LB culture medium comprises a primary improved LB culture medium, a secondary improved LB culture medium and a tertiary improved LB culture medium; the value of the primary improved LB culture medium X, the value of the secondary improved LB culture medium X and the value of the third improved LB culture medium X are increased in sequence; the value of the primary modified LB culture medium X is 3, the value of the secondary modified LB culture medium X is 8, and the value of the third modified LB culture medium X is 15;
s3-2 screening of improved bacillus coagulans
Inoculating the single colony bacillus coagulans obtained in the S2 into an initial generation improved LB culture medium according to the inoculation amount of 1% of the volume ratio, culturing for 24 hours at 37 ℃, diluting and coating, and selecting the initial generation bacillus coagulans with good growth; then sequentially inoculating the bacillus coagulans into a secondary generation LB culture medium for culture and selection and a third generation LB culture medium for culture and selection to obtain the screened improved bacillus coagulans;
s4, preparation of daily ration microbial agent
S4-1, preparation of fermentation culture solution
According to the formula: tryptone 10g/L, sodium chloride 10g/L, yeast extract 5g/L, glucose 5g/L, K 2 HPO 4 2g/L of agar powder, 20g/L of agar powder and 25-30 g/L of a basic substance extract are used for preparing a fermentation culture solution, and the pH value of the culture medium is adjusted to 7.4 by NaOH;
s4-2, preparation of microbial agent for daily ration
Inoculating the improved bacillus coagulans into a fermentation culture solution according to the inoculation amount of 3% of the volume ratio, culturing and fermenting for 3d at the temperature of 37 ℃, drying, and collecting the dried product to obtain the daily ration microbial agent.
Example 4
The difference from the embodiment 1 is:
the preparation method of the basic extract comprises the following specific steps:
1-2-1, mixing radix bupleuri, rhizoma anemarrhenae and orange peel according to a mass percentage of 30:12, cleaning, crushing, adding 5 times of purified water, carrying out primary heating extraction for 3 hours at the temperature of 60 ℃, cooling the extracting solution to room temperature, and filtering to obtain a primary extracting solution;
s1-2-2, adding 2 times of purified water into the filter residue again, carrying out secondary heating extraction for 0.5h at the temperature of 65 ℃, and filtering after the extracting solution is cooled to room temperature to obtain a secondary extracting solution;
s1-2-3, mixing the primary extract and the secondary extract, concentrating to 25% of the original volume, dissolving with ethanol, and desolventizing and drying with composite resin to obtain a basic extract containing saponin extract and flavanone extract.
Example 5
In contrast to example 1:
the preparation method of the basic extract comprises the following specific steps:
s1-2-1, mixing the following components in percentage by mass: 15, cleaning, crushing, adding 6 times of purified water, heating and extracting for the first time at 60 ℃ for 4 hours, cooling the extracting solution to room temperature, and filtering to obtain a first extracting solution;
s1-2-2, adding 2 times of purified water into the filter residue again, carrying out secondary heating extraction for 1 hour at 70 ℃, cooling the extracting solution to room temperature, and filtering to obtain a secondary extracting solution;
s1-2-3, mixing the primary extract and the secondary extract, concentrating to 30% of the original volume, dissolving with ethanol, and desolventizing and drying with composite resin to obtain a basic extract containing saponin extract and flavanone extract.
Example 6
In contrast to example 1:
the preparation method of the basic extract comprises the following specific steps:
1-2-1, mixing radix bupleuri, rhizoma anemarrhenae and orange peel according to a mass percentage of 60:17, cleaning, crushing, adding 8 times of purified water, heating and extracting for the first time for 6 hours at the temperature of 65 ℃, cooling the extracting solution to room temperature, and filtering to obtain a first extracting solution;
s1-2-2, adding purified water of which the amount is 3 times that of the filter residue again, carrying out secondary heating extraction for 1 hour at the temperature of 75 ℃, and filtering after the extracting solution is cooled to room temperature to obtain a secondary extracting solution;
s1-2-3, mixing the primary extract and the secondary extract, concentrating to 40% of the original volume, dissolving with ethanol, and desolventizing and drying with composite resin to obtain a basic extract containing saponin extract and flavanone extract.
Example 7
The difference from example 1 is: s3-1, the value of the X of the primary modified LB culture medium is 5, the value of the X of the secondary modified LB culture medium is 10, and the value of the X of the third modified LB culture medium is 15.
Example 8
The difference from example 1 is: s3-1, the value of the primary modified LB culture medium X is 8, the value of the secondary modified LB culture medium X is 12, and the value of the third modified LB culture medium X is 20.
Claims (10)
1. A preparation method of a daily ration microbial agent is characterized by comprising the following steps:
s1, preparation of basic material extract
The Chinese thorowax root, the rhizoma anemarrhenae and the orange peel are mixed according to the mass percentage of 30-60: 12-17, and extracting the extract after mixing the raw materials in the ratio of 20-75 to obtain a basic substance extract;
s2, culturing of Bacillus coagulans
Culturing the bacillus coagulans NRS-609 to obtain bacillus coagulans;
s3, screening of Bacillus coagulans
S3-1, preparation of improved LB culture medium
According to the formula: preparing an improved LB culture medium by 10g/L of tryptone, 10g/L of sodium chloride, 5g/L of yeast extract, 5g/L of glucose and X g/L of basic extract, and adjusting the pH of the culture medium to 7.0-7.4 by NaOH;
the improved LB culture medium comprises a primary improved LB culture medium, a secondary improved LB culture medium and a third improved LB culture medium; the value of the primary improved LB culture medium X, the value of the secondary improved LB culture medium X and the value of the third improved LB culture medium X are increased in sequence;
s3-2 screening of modified Bacillus coagulans
Inoculating the single colony bacillus coagulans obtained in the S2 into an initial generation improved LB culture medium according to the inoculation amount of 1% of the volume ratio, culturing for 24 hours at 37 ℃, diluting and coating, and selecting the initial generation bacillus coagulans with good growth; then sequentially inoculating the bacillus coagulans to a secondary generation LB culture medium for culture and selection and a third generation LB culture medium for culture and selection to obtain the screened improved bacillus coagulans;
s4, preparation of daily ration microbial agent
S4-1, preparation of fermentation culture solution
According to the formula: tryptone 10g/L, sodium chloride 10g/L, yeast extract 5g/L, glucose 5g/L, K 2 HPO 4 2g/L of agar powder, 20g/L of agar powder and 25-30 g/L of a basic substance extract are used for preparing a fermentation culture solution, and NaOH is used for adjusting the pH value of the culture medium to 7.0-7.4;
s4-2, preparation of microbial agent for daily ration
Inoculating the improved bacillus coagulans into a fermentation culture solution according to the inoculation amount of 2-3% of the volume ratio, culturing and fermenting for 2-3 d at 37 ℃, drying, and collecting the dried product to obtain the daily ration microbial agent.
2. The method for preparing a microbial agent for ration according to claim 1, wherein the base extract of S1 is prepared by the following specific steps:
s1-1-1, mixing 30-60 mass percent of radix bupleuri, rhizoma anemarrhenae and orange peel: 12-17, washing, crushing, adding 5-8 times of purified water, heating and extracting for 3-6 hours at the temperature of 60-65 ℃, cooling the extracting solution to room temperature, and filtering to obtain an extracting solution;
s1-1-2, concentrating the extracting solution to 25-40% of the original volume, dissolving with ethanol, desolventizing with composite resin, and drying to obtain a basic extract containing saponin extract and flavanone extract.
3. The method for preparing a microbial agent for ration according to claim 1, wherein the base extract of S1 is prepared by the following specific steps:
s1-2-1, mixing 30-60 mass percent of radix bupleuri, rhizoma anemarrhenae and orange peel: 12-17, 20-75, cleaning, crushing, adding 5-8 times of purified water, carrying out primary heating extraction for 3-6 hours at the temperature of 60-65 ℃, and filtering after the extract is cooled to room temperature to obtain a primary extract;
s1-2-2, adding 2-3 times of purified water into the filter residue again, carrying out secondary heating extraction for 0.5-1 h at the temperature of 65-75 ℃, and filtering after the extracting solution is cooled to room temperature to obtain a secondary extracting solution;
s1-2-3, mixing the primary extract and the secondary extract, concentrating to 25-40% of the original volume, dissolving with ethanol, desolventizing with composite resin, and drying to obtain a basic extract containing saponin extract and flavanone extract.
4. The method for preparing a microbial inoculant for ration according to claim 1, wherein the step of culturing the bacillus coagulans S2 comprises the following steps: activating the strain of the bacillus coagulans NRS-609, transferring the bacillus coagulans NRS-609 into an LB culture medium, culturing the bacillus coagulans in the LB culture medium at 37 ℃ for 24 hours, then diluting and coating the bacillus coagulans, and selecting well-grown single-colony bacillus coagulans.
5. The method for preparing microbial inoculant for daily ration according to claim 4, wherein the Bacillus coagulans NRS-609 with the preservation number Bio-74771 is S2.
6. The preparation method of a ration microbial agent according to claim 1, wherein the value of the primary modified LB medium X in S3-1 is 3, the value of the secondary modified LB medium X is 8, and the value of the third modified LB medium X is 15.
7. The preparation method of a ration microbial agent according to claim 1, wherein the value of the primary modified LB medium X in S3-1 is 5, the value of the secondary modified LB medium X is 10, and the value of the third modified LB medium X is 15.
8. The preparation method of a microbial agent for ration according to claim 1, wherein the value of the primary modified LB medium X in S3-1 is 8, the value of the secondary modified LB medium X is 12, and the value of the tertiary modified LB medium X is 20.
9. The method for preparing microbial inoculant for daily ration of claim 1, wherein the fermentation broth S4-1 is prepared from tryptone 10g/L, sodium chloride 10g/L, yeast extract 5g/L, glucose 5g/L, K 2 HPO 4 2g/L, 20g/L agar powder and 25-30 g/L base extract.
10. The preparation device of a microbial agent for ration according to claim 1, which comprises a shell (1), a pot body (2) installed on the shell (1), a heating pipe installed inside the shell (1) and used for heating the pot body (2), a thermal thermostat controller installed on the shell (1) and used for controlling the temperature of the pot body (2), and a pipe fitting used for conducting water guiding/draining treatment on the pot body (2); the pan body (2) comprises a first heating pan (21) for assisting in the preparation of the base extract and a second heating pan (22) for assisting in the cultivation of microorganisms.
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Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103289910A (en) * | 2012-10-23 | 2013-09-11 | 广州格拉姆生物科技有限公司 | Solid fermentation production method of bacillus coagulans |
| CN104988075A (en) * | 2015-06-11 | 2015-10-21 | 天津中天精科科技有限公司 | Improved fermentation culture medium of pleurotus eryngii liquid strain |
| CN105104712A (en) * | 2015-10-09 | 2015-12-02 | 沈阳丰美生物技术有限公司 | Microbiological feed additive and preparation method thereof |
| CN205850902U (en) * | 2016-08-11 | 2017-01-04 | 谢贤英 | A kind of adjustable integrated water-bath of temperature |
| CN210683764U (en) * | 2019-10-09 | 2020-06-05 | 广州千江生物科技有限公司 | Water bath for microbial cultivation |
-
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- 2022-09-16 CN CN202211130322.9A patent/CN115568532A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103289910A (en) * | 2012-10-23 | 2013-09-11 | 广州格拉姆生物科技有限公司 | Solid fermentation production method of bacillus coagulans |
| CN104988075A (en) * | 2015-06-11 | 2015-10-21 | 天津中天精科科技有限公司 | Improved fermentation culture medium of pleurotus eryngii liquid strain |
| CN105104712A (en) * | 2015-10-09 | 2015-12-02 | 沈阳丰美生物技术有限公司 | Microbiological feed additive and preparation method thereof |
| CN205850902U (en) * | 2016-08-11 | 2017-01-04 | 谢贤英 | A kind of adjustable integrated water-bath of temperature |
| CN210683764U (en) * | 2019-10-09 | 2020-06-05 | 广州千江生物科技有限公司 | Water bath for microbial cultivation |
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