CN113729109B - Fermented feed rich in antibacterial substances and preparation method thereof - Google Patents
Fermented feed rich in antibacterial substances and preparation method thereof Download PDFInfo
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Abstract
The invention discloses a fermented feed rich in antibacterial substances and a preparation method thereof. The fermented feed is obtained by fermenting raw materials comprising a carbon source, a nitrogen source, dipotassium hydrogen phosphate, magnesium sulfate, water and paenibacillus polymyxa SP 1; the Paenibacillus polymyxa SP1 is deposited in the Guangdong province microorganism strain collection center, the deposition date is 2021, 4 months and 16 days, and the deposition number is GDMCC No:61605. the feed preparation method disclosed by the invention is simple in process, and the fermented feed obtained by the method is rich in antibacterial substances and can replace the application of partial antibiotics; meanwhile, the content of available phosphorus, protein solubility and acid soluble protein is increased, the content of crude fat is obviously reduced, and the method is favorable for animals, especially young animals, to absorb and utilize and has good popularization and application values.
Description
Technical Field
The invention relates to the technical field of fermented feed, in particular to a fermented feed rich in antibacterial substances and a preparation method thereof.
Background
In the existing breeding feed technology, antibiotics are used in a large amount, a series of problems are caused, the application of antibiotics can increase the drug resistance of microorganisms and parasites of an application target, and because the drug resistance is often used again, the dosage and the variety of the antibiotics need to be further increased, the drug resistance is further increased, so that malignant circulation is formed, and the abuse of the antibiotics is caused; not only does this add significant cost and hazard to the breeding process, but the antibiotics are difficult to disappear for a short time during the food chain operation, and humans act as victims of antibiotic drug residues on top of the food chain. On the premise that the national livestock and poultry feed forbids the comprehensive implementation of the antibiotic policy, a feed formula for avoiding the problems of drug resistance and drug residue caused by antibiotics is urgently needed.
The microbial fermentation feed has the outstanding functions of improving the utilization rate and palatability of animal feed, maintaining the microecological balance of intestinal flora, enhancing the immunity of an organism, improving the disease resistance and the production performance of animals and the like, and highlights the potential effects of the microbial fermentation feed in the aspects of replacing the feed, reducing the use of antibiotics, improving the immunity of hosts, resisting diseases and the like in the animal cultivation process. If antibacterial peptide (Chinese patent CN 102150752A), terramycin, formate and other antibacterial additives are added into the feed, the antibacterial effect (Chinese patent CN 1242689C) can be achieved, but the antibacterial additive is often distributed in various food raw materials and sewage due to the strong viability of food-borne pathogenic microorganisms, has high cost after long-term use, and produces certain toxic effect on cultured animals.
In addition, the bred animals, especially young animals, have poor intestinal function, so that the high-content protein and crude fat in the food are difficult to digest and absorb, digestive disorder is generated, and the diseases such as vomiting, inappetence, gastroenteritis and the like are caused, so that the health of the animals is influenced, and death can be caused when the diseases are serious, so that the survival rate of the bred animals is influenced. The phosphorus is not easy to be absorbed by animals, the available phosphorus in the feed becomes effective phosphorus, and the higher the content is, the more beneficial to the animals to absorb the phosphorus. For this problem, the absorption of animals is often promoted by adding enzyme preparations such as lipase, protease and phytase (CN 103719596A, CN104621413 a), but the cost for adding enzyme preparations is high.
Disclosure of Invention
The invention provides a fermented feed rich in antibacterial substances and a preparation method thereof, which are used for solving the technical defects that the prior feed adopts antibiotics and antibacterial additives, which can cause drug resistance, drug residue, drug toxicity and high cost due to complex process caused by the addition of enzyme preparations.
In order to achieve the above purpose, the present invention adopts the following technical scheme:
the invention provides a fermented feed rich in antibacterial substances and a preparation method thereof, which are used for improving the utilization rate of feed raw materials, effectively acting on common food pathogenic microorganisms, being used for replacing partial antibiotics, being simple in process, low in cost, safe and nontoxic, being widely applied to livestock and poultry cultivation, having broad-spectrum antibacterial effect on bacteria used for fermentation, inhibiting common livestock and poultry pathogenic bacteria such as staphylococcus aureus, escherichia coli K12D31, escherichia coli O78, salmonella typhimurium, salmonella cholerae and the like, and having good antibacterial effect on common pathogenic bacteria such as bacillus thuringiensis and septicemia nigricans of insect cultivation objects such as silkworms and the like, and being used as a feed additive for insect cultivation, in particular silkworms. The bacteria SP1 used for fermentation can produce protease, lipase and phytase, has good fermentation effect on protein, fat and phosphorus, can ensure that the content of effective phosphorus, protein solubility and acid soluble protein of the feed is increased, the content of crude fat is obviously reduced, and the fermented feed is beneficial to the absorption of protein, fat and phosphorus of livestock and poultry, particularly young livestock and poultry, reduces the addition amount of protein and fat raw materials of the feed, and can effectively control the cultivation cost. The feed fermentation utilizes paenibacillus polymyxa which has high-efficiency fermentation and bacteriostasis functions.
Specifically, the raw material components for preparing the fermented feed comprise: carbon source, nitrogen source, dipotassium hydrogen phosphate, magnesium sulfate, water and paenibacillus polymyxa SP1. The Paenibacillus polymyxa SP1 is deposited in the Guangdong province microorganism strain collection center, the deposition date is 2021, 4 months and 16 days, and the deposition number is GDMCC No:61605.
further, the components in parts by weight are as follows: 10-13 parts of carbon source, 1-5 parts of nitrogen source, 0.0025-0.25 part of dipotassium hydrogen phosphate, 0.000073-0.0073 part of magnesium sulfate, 15-25 parts of water and 2-4 parts of Paenibacillus polymyxa SP1.
Preferably, the carbon source is bran or corn meal, and the nitrogen source is soybean meal or soybean meal.
Further, dipotassium hydrogen phosphate adopts hydrogen phosphate hydrate: k (K) 2 HPO 4 ·2H 2 O, magnesium sulfate adopts magnesium sulfate hydrate: mgSO (MgSO) 4 ·7H 2 O。
Further, the weight portions of the components are as follows: 12 parts of bran, 3 parts of bean pulp and K 2 HPO 4 ·2H 2 O0.03 part, mgSO 4 ·7H 2 0.0015 part of O, 20 parts of water and 3 parts of Paenibacillus polymyxa SP1.
Furthermore, the water adopted in the fermented feed is sterile water.
Furthermore, the fermented feed adopts the use form of fermentation liquor of Paenibacillus polymyxa SP1, wherein the bacterial density is not less than 1 multiplied by 10 6 cfu。
Preferably, the Paenibacillus polymyxa SP1 broth has a bacterial density of (1X 10) 6 )-(1×10 10 )cfu。
The optimal initial adding density is matched with other materials, and the optimal feed product with optimal bacteriostasis and fermentation effects can be achieved under the corresponding temperature condition.
In order to achieve the purpose, the preparation method of the fermented feed is shown in figure 1, and comprises the following specific steps:
s1, pretreatment of raw materials: taking a carbon source, a nitrogen source, dipotassium hydrogen phosphate and magnesium sulfate, and adding water to uniformly mix; then sterilizing;
s2, culturing SP1 bacteria to obtain fermentation liquor;
s3, fermenting feed: inoculating SP1 bacteria fermentation liquor into the mixture sterilized in the step S1 for fermentation in a sterile environment, and intermittently turning over water materials in the fermentation process; after the fermentation is completed, the fermented feed rich in antibacterial substances is obtained.
Wherein, the sterilization temperature in the step S1 is preferably 100-150 ℃ and the sterilization time is preferably 15-30min.
Preferably, the fermentation conditions of step S3 are: the temperature is controlled at 29-31 ℃, the fermentation time is 30-72h, and the turnover frequency is as follows: 1 every 12 hours.
The beneficial effects of the invention are as follows:
according to the invention, paenibacillus polymyxa (Paenibacillus polymyxa) SP1 is added into feed raw materials for fermentation (Paenibacillus polymyxa is listed as a first-class strain which is free from safety identification by agricultural department in China), SP1 is a Paenibacillus polymyxa which is obtained by separating a subject group of the inventor and can produce various bioactive substances such as polypeptide antibiotics, antagonistic proteins, enzyme proteins and the like, and has good antibacterial effect on various strains such as staphylococcus aureus, escherichia coli K12D31, escherichia coli O78, salmonella typhimurium, salmonella cholerae, bacillus thuringiensis, black-chest and the like, and hydrogen sulfide gas is not produced by fermentation. Compared with the common fermented feed which is mainly used for improving the disease resistance of livestock and poultry by maintaining the microecological balance of animal intestinal flora through microorganisms, the fermented feed provided by the invention not only can be used for regulating the microecological balance of animal intestinal flora, is rich in antibacterial substances and can directly kill pathogenic bacteria, the fermentation and bacteriostasis can be completed through one bacterium, and the obtained fermented feed has a better inhibition effect on common intestinal pathogenic bacteria of common livestock and poultry, in particular to silkworm pathogenic bacteria of insect breeding objects, and can be used for preventing silkworm damping-off and sepsis through inhibiting bacillus thuringiensis and black thoracocentesis.
Moreover, the strain can secrete various enzyme proteins such as lipase, phytase and the like, can increase the content of available phosphorus, protein solubility and acid soluble protein in raw materials, remarkably reduces the content of crude fat, and is beneficial to absorption and utilization of animals, especially young animals.
The fermented feed is safe, harmless to cultured organisms, short in fermentation time, simple in preparation process and suitable for popularization and application.
Drawings
FIG. 1 is a flow chart of a preparation method of the fermented feed
FIG. 2 is a graph showing the effect of different carbon sources on the fermentation effect of SP1 bacteria
FIG. 3 comparison of the effects of different nitrogen sources on the number of SP1 colonies
FIG. 4 is a graph showing the effect of mass ratio of bran to soybean meal on the colony count of SP1
FIG. 5 addition of different concentrations of K 2 HPO 4 Comparison of effects on SP1 colony count
FIG. 6 addition of different concentrations of MgSO 4 Effect on SP1 colony count comparison
FIG. 7 addition of different concentrations of MnSO 4 Effect on SP1 colony count comparison
FIG. 8 is a graph showing the effect of different feed water ratios on the number of viable bacteria in fermentation
FIG. 9 is a graph showing the effect of different inoculum sizes on the number of viable bacteria in fermentation
FIG. 10 is a graph showing the effect of different fermentation periods on the number of viable bacteria
FIG. 11 antibacterial activity of fermentation supernatant and fermented feed extract
Detailed Description
The invention is further illustrated in the following drawings and specific examples, which are not intended to limit the invention in any way. Unless specifically stated otherwise, the reagents, methods and apparatus employed in the present invention are those conventional in the art.
Reagents and materials used in the following examples are commercially available unless otherwise specified.
Examples 1-2 are examples of feeds and examples 3-11 are examples of the present fermented feed in combination with the feed preparation process.
EXAMPLE 1 fermentation broth screening
The applicant obtains a strain with highest antibacterial activity from soil in mulberry field of agricultural university of south China by adopting an agar hole diffusion method and using escherichia coli and staphylococcus aureus as indicator bacteria through comparison and screening, and the strain is named as a strain SP1.
The integrated result of morphological identification and molecular identification shows that the identified strain SP1 belongs to Paenibacillus polymyxa (Paenibacillus polymyxa) and is preserved in the microorganism strain collection of Guangdong province at the year 2021, month 4 and day 16, and the preservation unit address is the microbiological institute of the Guangdong province (building 5 of the national institute of sciences of Guangzhou, mitsui, 100 th university, and building 59) with the preservation number of GDMCC No:61605.
example 2 preparation of fermented feed
1. Weighing the following components in parts by weight:
12 parts of carbon source bran,
3 parts of nitrogen source soybean meal,
Dipotassium hydrogen phosphate hydrate (K) 2 HPO 4 ·2H 2 O) 0.03 part,
Magnesium sulfate hydrate (MgSO) 4 ·7H 2 O) 0.0015 part,
20 parts of water, and the water content is equal to 20 parts,
3 parts of Paenibacillus polymyxa, wherein Paenibacillus polymyxa is SP1 bacteria, and the density of the SP1 bacteria is 1 multiplied by 10 6 cfu per serving.
2. The preparation method of the fermented feed comprises the following steps:
s1, pretreatment of raw materials: taking a carbon source, a nitrogen source, dipotassium hydrogen phosphate hydrate and magnesium sulfate hydrate, and adding water to uniformly mix; sterilizing at 120deg.C for 20min;
s2, culturing SP1 bacteria to obtain fermentation liquor;
s3, fermenting feed: inoculating 3mL of the SP1 strain fermentation liquor (1×10) per 30g of the mixture after sterilization in step S1 under aseptic conditions 6 cfu/mL), fermenting at constant temperature of 30+/-1 ℃ for 48 hours, intermittently stirring the water material, wherein the stirring frequency is 1 time every 12 hours; after the fermentation is completed, the fermented feed rich in antibacterial substances is obtained.
Example 3 preparation of fermented feed
1. Weighing the following components in parts by weight:
12 parts of carbon source corn flour,
3 parts of nitrogen source bean flour,
Dipotassium hydrogen phosphate hydrate (K) 2 HPO 4 ·2H 2 O) 0.03 part,
Magnesium sulfate hydrate (MgSO) 4 ·7H 2 O) 0.0015 part,
20 parts of water, and the water content is equal to 20 parts,
3 parts of Paenibacillus polymyxa, wherein Paenibacillus polymyxa is SP1 bacteria, and the density of the SP1 bacteria is 1 multiplied by 10 6 cfu per serving.
2. The preparation method of the fermented feed is the same as in example 2.
Example 4 preparation of fermented feed
1. Weighing the following components in parts by weight:
10 parts of carbon source bran,
1 part of nitrogen source soybean meal,
Dipotassium hydrogen phosphate hydrate (K) 2 HPO 4 ·2H 2 O) 0.0025 part,
Magnesium sulfate hydrate (MgSO) 4 ·7H 2 0.000073 parts of O),
15 parts of water, and the water is mixed with the water,
2 parts of Paenibacillus polymyxa, wherein the Paenibacillus polymyxa is SP1 bacteria, and the density of the SP1 bacteria is 1 multiplied by 10 6 cfu per serving.
2. The preparation method of the fermented feed is the same as in example 2.
Example 5 preparation of fermented feed
1. Weighing the following components in parts by weight:
13 parts of carbon corn powder,
5 parts of nitrogen source bean flour,
Dipotassium hydrogen phosphate hydrate (K) 2 HPO 4 ·2H 2 O) 0.25 part,
Magnesium sulfate hydrate (MgSO) 4 ·7H 2 O) 0.0073 part,
25 parts of water, and the water is mixed with the water,
4 parts of Paenibacillus polymyxa, wherein Paenibacillus polymyxa is SP1 bacteria, and the density of the SP1 bacteria is 1 multiplied by 10 6 cfu per serving.
2. The preparation method of the fermented feed is the same as in example 2.
Example 6 preparation of fermented feed
1. Weighing the following components in parts by weight:
10 parts of carbon source bran,
5 parts of nitrogen source soybean meal,
Dipotassium hydrogen phosphate hydrate (K) 2 HPO 4 ·2H 2 O) 0.0025 part,
Magnesium sulfate hydrate (MgSO) 4 ·7H 2 O) 0.0073 part,
20 parts of water, and the water content is equal to 20 parts,
3 parts of Paenibacillus polymyxa, wherein Paenibacillus polymyxa is SP1 bacteria, and the density of the SP1 bacteria is 1 multiplied by 10 6 cfu per serving.
2. The preparation method of the fermented feed is the same as in example 2.
Example 7 index test of fermented feed
1. Physicochemical Properties
The physicochemical properties of the feeds obtained in examples 2 to 6 before and after fermentation were observed and the relevant index was measured, and the results showed that: the fermented feed prepared by the invention is yellow brown, has bean fragrance and has viable count as high as 6 multiplied by 10 9 cfu/g or more, wherein the viable count of the fermented feed prepared in example 2 was 6.33X10 9 cfu/g, has good antibacterial effect.
2. Fermentation effect
The fermented feed prepared in example 2 was used as an example to test the change of the related index before and after fermentation.
The results show that the fermented feed has good fermentation effect, compared with the prior fermentation, the effective phosphorus content is improved by 48.44 percent, the nitrogen content in the protein solubility is improved by 11.11 percent, the nitrogen content in the acid soluble protein is improved by 35.29 percent, the crude fat content is reduced by 43.79 percent, and the fermented feed is more beneficial to the absorption and utilization of fat and protein by animals, particularly by children with gastrointestinal dysfunction, and the vomiting and dyspepsia caused by excessive fat are reduced (table 1).
TABLE 1 variation of various indexes before and after fermentation
Example 8 selection of carbon sources in fermented feed Material
Experiments were performed by changing the carbon source based on the method of example 2. The method specifically comprises the following steps: respectively taking 12g of rice hulls, 12g of bran and 12g of corn meal as carbon sources, adding 3g of soybean meal as nitrogen sources into each group, and adding K with a certain mass 2 HPO 4 ·2H 2 O and MgSO 4 ·7H 2 O, after being stirred and mixed uniformly, each of the components is added with 15mL of sterile water, then sterilized at a high temperature of 120 ℃ for 20min, and after the temperature is reduced to about 30 ℃, 3mL (about 10) 6 cfu/mL) SP1 seed liquid, stirring and mixing uniformly, placing in an incubator for fermentation culture, and sampling after stopping fermentation to determine the bacterial content of the solid fermentation product and the antibacterial activity of the product.
As a result, as shown in FIG. 2, the numbers of viable bacteria produced in the two groups of the culture media containing bran and corn flour were detected to be 2.0X10, respectively 8 cfu/g、1.55×10 8 cfu/g, it can be seen that bran is suitable for solid fermentation of strain SP1 compared with corn meal. Because the effect of the solid fermentation of the bran is better than that of corn flour, the bran is better than that of rice husk.
Example 9 selection of Nitrogen Source in fermented feed Material
Experiments were performed by varying the nitrogen source based on the method of example 2. The method specifically comprises the following steps: taking 12g of bran as a carbon source, respectively taking 3g of yeast powder, 3g of soybean powder and 3g of soybean meal as nitrogen sources, and adding K with a certain mass 2 HPO 4 ·2H 2 O and MgSO 4 ·7H 2 O, after being stirred and mixed uniformly, each of the components is added with 15mL of sterile water, then sterilized at a high temperature of 120 ℃ for 20min, and after the temperature is reduced to about 30 ℃, 3mL (about 10) 6 cfu/mL) SP1 seed liquid, stirring and mixing uniformly, placing in an incubator for fermentation culture, and sampling after stopping fermentation to determine the bacterial content of the solid fermentation product and the antibacterial activity of the product.
The results are shown in FIG. 3, in which SP1 bacteria did not grow in the yeast-containing bran. The generated viable bacteria are detected by bran containing soybean meal and soybean mealThe numbers were 2.53X 10 respectively 8 cfu/g、3.67×10 8 cfu/g, so the soybean meal is superior to soybean meal, and the Paenibacillus polymyxa SP1 cannot utilize yeast powder.
Example 10 optimization of the carbon Source to Nitrogen Source ratio in fermented feed Material
Preparing 6 groups of bran and bean pulp with mass ratio of 1:2, 1:1, 2:1, 4:1, 6:1 and 8:1 respectively, and adding K with certain mass respectively 2 HPO 4 ·2H 2 O and MgSO 4 ·7H 2 O, mixing, adding 15mL of sterile water, sterilizing at 120deg.C for 20min, cooling to about 30deg.C, and inoculating 3mL (about 10 6 cfu/mL) SP1 seed solution, stirring and mixing uniformly, placing in an incubator for fermentation culture, and sampling after stopping fermentation to determine the bacterial content of the solid fermentation product. The control feed water ratio is 1:1, wherein the mass ratio of the bran to the bean pulp is 1:2, namely 5g of the bran and 10g of the bean pulp; the mass ratio of the bran to the bean pulp is 1:1, namely 7.5g of bran and 7.5g of bean pulp; the mass ratio of the bran to the bean pulp is 2:1 respectively, namely 10g of bran and 5g of bean pulp; the mass ratio of the bran to the bean pulp is 4:1, namely 12g of bran and 3g of bean pulp; the mass ratio of the bran to the bean pulp is 6:1, namely 12.9g of bran and 2.1g of bean pulp; the mass ratio of the bran to the bean pulp is 8:1, 13.3g of bran and 1.7g of bean pulp.
The result is shown in fig. 4, the optimal mass ratio is 12g of bran and 3g of bean pulp, namely bran: the mass ratio of the bean pulp is 4:1.
EXAMPLE 11 fermentation of K in feed stock 2 HPO 4 Content selection
Weighing 5 parts of bran (12 g) and soybean meal (3 g) raw materials, and respectively adding 2%, 0.2%, 0.02%, 0.002% and 0% (w/w) K 2 HPO 4 ·2H 2 O, mixing, adding 15mL of sterile water, sterilizing at 120deg.C for 20min, cooling to about 30deg.C, and inoculating 3mL (about 10 6 cfu/mL) SP1 seed solution, stirring and mixing uniformly, placing in an incubator for fermentation culture, and sampling after stopping fermentation to determine the bacterial content of the solid fermentation product.
The result is shown in FIG. 5, wherein K is optimally added 2 HPO 4 The hydrate concentration was 0.2%.
EXAMPLE 12 MgSO in fermented feed Material 4 Content selection
Weighing 5 parts of bran (12 g) and soybean meal (3 g) raw materials, adding 30mg K 2 HPO 4 ·2H 2 O was added to 1%, 0.1%, 0.01%, 0.001%, 0% (w/w) MgSO, respectively 4 ·7H 2 O, mixing, adding 15mL of sterile water, sterilizing at 120deg.C for 20min, cooling to about 30deg.C, and inoculating 3mL (about 10 6 cfu/mL) SP1 seed liquid, stirring and mixing uniformly, placing in an incubator for fermentation culture, and sampling after stopping fermentation to determine the bacterial content of the solid fermentation product and the antibacterial activity of the product.
The results are shown in FIG. 6, best MgSO 4 The hydrate addition concentration was 0.01%, i.e., 1.5mg.
Example 13 MnSO in fermented feed Material 4 Screening of (C)
MnSO 4 Is selected from the group consisting of a screening of the above. Weighing 5 parts of bran (12 g) and soybean meal (3 g) raw materials, adding 30mg K 2 HPO 4 ·2H 2 O，1.5mg MgSO 4 ·7H 2 O, after mixing uniformly, mnSO of 1%, 0.1%, 0.01%, 0.001%, 0% (w/w) is added respectively 4 Adding 15mL of sterile water, sterilizing at 120deg.C for 20min, cooling to about 30deg.C, and inoculating 3mL (about 10 6 cfu/mL) SP1 seed solution, stirring and mixing uniformly, placing in an incubator for fermentation culture, and sampling after stopping fermentation to determine the bacterial content of the solid fermentation product.
As a result, FIG. 7 shows that unlike other Paenibacillus polymyxa, mn ions have an inhibitory effect on the growth of Paenibacillus polymyxa SP1, and that growth without addition is preferable.
Example 14 selection of feed Water ratio in fermented feed Material
3 parts of bran (12 g) and soybean meal (3 g) are weighed and added with 30mg K 2 HPO 4 ·2H 2 O，1.5mg MgSO 4 ·7H 2 Adding distilled water 10mL,15mL, 20mL and 25mL according to the ratio of 3:2, 1:1, 3:4 and 3:4 (g/mL), sterilizing at 120deg.C for 20min, cooling to about 30deg.C, and introducing 3mL (about 10) 6 cfu/mL) SP1 seed solution, stirring and mixing uniformly, and placing in an incubatorAnd (3) fermenting and culturing, sampling after stopping fermentation, and measuring the bacterial content of the solid fermentation product.
As a result, see FIG. 8, the optimal feed water ratio was 3 g/4 mL, i.e., 20mL distilled water.
EXAMPLE 15 fermentation initial inoculum size selection
Weighing 4 parts of bran (12 g) and soybean meal (3 g) raw materials, and adding 30mg K 2 HPO 4 ·2H 2 O、1.5mg MgSO 4 ·7H 2 O, 20mL of sterile water, sterilizing at 120deg.C for 20min, and respectively introducing 1mL, 2mL, 3mL, and 4mL (about 10) 6 cfu/mL) SP1 seed solution, stirring and mixing uniformly, placing in an incubator for fermentation culture, and sampling after stopping fermentation to determine the bacterial content of the solid fermentation product.
The results are shown in FIG. 9, and the optimal initial inoculum size is 3mL.
EXAMPLE 16 fermentation time selection
3 parts of bran (12 g) and soybean meal (3 g) are weighed and added with 30mg K 2 HPO 4 ·2H 2 O、1.5mg MgSO 4 ·7H 2 O, 20mL of sterile water, sterilizing at 120deg.C for 20min, cooling to about 30deg.C, and inoculating 3mL (about 10 6 cfu/mL) SP1 seed solution, stirring and mixing uniformly, placing in an incubator for fermentation culture for 24 hours, 48 hours and 72 hours, sampling and measuring the bacterial content of the solid fermentation product.
The results are shown in FIG. 10, and the optimal fermentation time is 48h.
Example 17 antibacterial Effect verification
1. Experimental methods and procedures
The size of the bacteriostasis zone is detected by an agar pore diffusion method to judge the bacteriostasis effect of the fermentation liquid and the fermented feed.
LB medium includes LB liquid medium and LB solid medium.
LB liquid medium (1000 mL): 10g/L peptone, 5g/L yeast extract and 10g/L sodium chloride, and sterilizing at 121deg.C for 20min under high pressure with distilled water to 1L pH of 6.0-7.0.
LB solid medium (1000 mL): 10g/L peptone, 5g/L yeast extract, 10g/L sodium chloride, 1.5% agar powder, and distilled water to 1L, pH 6.0-7.0, and sterilizing at 121deg.C for 20min.
The method comprises the following steps: and (3) placing the sterilized LB culture medium at room temperature, cooling to 48-50 ℃, absorbing 100 mu L of indicator bacteria staphylococcus aureus diluent, adding the indicator bacteria diluent into 10mL of LB solid culture medium which is dissolved and cooled to 40-50 ℃, immediately pouring the mixture into a culture dish with the diameter of 90mm, rapidly shaking and uniformly shaking to uniformly disperse the thalli, and punching 3 thalli by a puncher with the diameter of 2.7mm for later use after the thalli is cooled and solidified. 5uL of sterile water, 5uL of an SP1 fermentation liquid and 5uL of a fermented feed extracting solution are respectively added into each hole, after standing for 30min, the mixture is cultured for 24 hours at 37 ℃, and the size of a bacteriostasis zone is observed and measured and recorded.
2. Test results
The bacteriostatic effect on staphylococcus aureus is shown in fig. 11, where CK1 is the sterile water control (2.7 mm, i.e. diameter of spotted wells); CK2 is a bacteriostasis zone (12.78 mm) of the SP1 fermentation broth; 1 is a bacteriostasis zone (18.86 mm) of the fermented feed extract.
The result shows that the Paenibacillus polymyxa SP1 fermentation liquor has good antibacterial effect, and the antibacterial effect of the fermented feed prepared by the method is obviously improved.
The above examples are preferred embodiments of the present invention, but the embodiments of the present invention are not limited to the above examples, and any other changes, modifications, substitutions, combinations, and simplifications that do not depart from the spirit and principle of the present invention should be made in the equivalent manner, and the embodiments are included in the protection scope of the present invention.
Claims (10)
1. The fermented feed rich in antibacterial substances is characterized by being obtained by fermenting raw materials comprising a carbon source, a nitrogen source, dipotassium hydrogen phosphate, magnesium sulfate, water and paenibacillus polymyxa SP 1; the Paenibacillus polymyxa SP1 is deposited in the Guangdong province microorganism strain collection center, the deposition date is 2021, 4 months and 16 days, and the deposition number is GDMCC No:61605.
2. the fermented feed of claim 1, wherein the components are in parts by weight: 10-13 parts of carbon source, 1-5 parts of nitrogen source, 0.0025-0.25 part of dipotassium hydrogen phosphate, 0.000073-0.0073 part of magnesium sulfate, 15-25 parts of water and 2-4 parts of Paenibacillus polymyxa SP1.
3. The fermented feed of claim 2, wherein the carbon source is bran or corn meal and the nitrogen source is soybean meal or soy flour.
4. The fermented feed according to claim 2, characterized in that dipotassium hydrogen phosphate is used as hydrogen phosphate hydrate: k (K) 2 HPO 4 ·2H 2 O, magnesium sulfate adopts magnesium sulfate hydrate: mgSO (MgSO) 4 ·7H 2 O。
5. The fermented feed of claim 2, wherein the weight parts of the components are 12 parts of bran, 3 parts of soybean meal and K 2 HPO 4 ·2H 2 O0.03 part, mgSO 4 ·7H 2 0.0015 part of O, 20 parts of water and 3 parts of Paenibacillus polymyxa SP1.
6. The fermented feed according to claim 4, wherein Paenibacillus polymyxa SP1 is used in the form of a fermentation broth, wherein the bacterial density is not less than 1X 10 6 cfu/mL。
7. The fermented feed according to claim 4, wherein Paenibacillus polymyxa SP1 is used in the form of a fermentation broth, wherein the bacterial density is (1×10 6 )-(1×10 10 )cfu/mL。
8. The fermented feed according to any one of claims 1 to 7, characterized in that it is prepared by a process comprising the steps of:
s1, pretreatment of raw materials: taking a carbon source, a nitrogen source, dipotassium hydrogen phosphate and magnesium sulfate, and adding water to uniformly mix; then sterilizing;
s2, culturing SP1 bacteria to obtain fermentation liquor;
s3, fermenting feed: inoculating SP1 bacteria fermentation liquor into the mixture sterilized in the step S1 for fermentation in a sterile environment, and intermittently turning over water materials in the fermentation process; after the fermentation is completed, the fermented feed rich in antibacterial substances is obtained.
9. The fermented feed according to claim 8, wherein the sterilization temperature in step S1 is 100-150 ℃ and the sterilization time is 15-30min.
10. The fermented feed of claim 8, wherein the fermentation conditions of step S3 are: the temperature is controlled at 29-31 ℃, the fermentation time is 30-72h, and the turnover frequency is as follows: 1 every 12 hours.
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Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0281091A2 (en) * | 1987-03-06 | 1988-09-07 | Hoechst Aktiengesellschaft | Agents containing Bacillus polymyxa to deliver to animals |
| CN101104637A (en) * | 2007-06-15 | 2008-01-16 | 华南农业大学 | Cecropin and its generation bacterial strain and use |
| CN101869181A (en) * | 2010-04-09 | 2010-10-27 | 武汉科诺生物科技股份有限公司 | Preparation method of bacillus polymyxa raw powder with 100 billion live spores per gram |
| CN105166385A (en) * | 2015-09-23 | 2015-12-23 | 安徽省兴源草业有限公司 | Fermented feed with ragimillet seeds and fermenting agent |
| WO2018047338A1 (en) * | 2016-09-12 | 2018-03-15 | 有限会社マヤインダストリー | Process for producing immunostimulant for animal, immunostimulant for animal, and feed for animal |
| CN110484467A (en) * | 2019-08-19 | 2019-11-22 | 山东宝来利来生物工程股份有限公司 | Antibacterial peptide and the application of one plant of bacillus polymyxa and its generation |
| CN112280721A (en) * | 2020-11-19 | 2021-01-29 | 山东龙昌动物保健品有限公司 | Paenibacillus polymyxa LCP-01, compound microbial inoculum compounded with bile acid and application thereof |
Family Cites Families (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US6989370B2 (en) * | 2003-05-01 | 2006-01-24 | The United States Of America As Represented By The Secretary Of Agriculture | Bacteriocins and novel bacterial strains |
| US9809865B2 (en) * | 2009-12-09 | 2017-11-07 | Best Environmental Technologies, Inc. | Anti-microbial agent from Paenibacillus sp. and methods and uses thereof |
-
2021
- 2021-07-20 CN CN202110821644.7A patent/CN113729109B/en active Active
Patent Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0281091A2 (en) * | 1987-03-06 | 1988-09-07 | Hoechst Aktiengesellschaft | Agents containing Bacillus polymyxa to deliver to animals |
| CN101104637A (en) * | 2007-06-15 | 2008-01-16 | 华南农业大学 | Cecropin and its generation bacterial strain and use |
| CN101869181A (en) * | 2010-04-09 | 2010-10-27 | 武汉科诺生物科技股份有限公司 | Preparation method of bacillus polymyxa raw powder with 100 billion live spores per gram |
| CN105166385A (en) * | 2015-09-23 | 2015-12-23 | 安徽省兴源草业有限公司 | Fermented feed with ragimillet seeds and fermenting agent |
| WO2018047338A1 (en) * | 2016-09-12 | 2018-03-15 | 有限会社マヤインダストリー | Process for producing immunostimulant for animal, immunostimulant for animal, and feed for animal |
| CN110484467A (en) * | 2019-08-19 | 2019-11-22 | 山东宝来利来生物工程股份有限公司 | Antibacterial peptide and the application of one plant of bacillus polymyxa and its generation |
| CN112280721A (en) * | 2020-11-19 | 2021-01-29 | 山东龙昌动物保健品有限公司 | Paenibacillus polymyxa LCP-01, compound microbial inoculum compounded with bile acid and application thereof |
Non-Patent Citations (5)
| Title |
|---|
| 慈晓通 等.多黏类芽孢杆菌发酵液抑菌效果的研究.中国家禽.2019,第41卷(第8期),25-28. * |
| 李玉洋 等.菌糠固态发酵法培养多粘类芽孢杆菌SH15的工艺.山东农业科学.2017,第49卷(第12期),119-125. * |
| 李福彬 等.多粘芽孢杆菌抗逆性与益生性的体外评价.河北农业大学学报.2010,第33卷(第3期),78-82. * |
| 程爱芳 等.多粘类芽孢杆菌HD-1产纤维素酶的条件优化.食品工业科技.2014,第36卷(第10期),173-177. * |
| 赵国群 等.梨渣固态发酵培养多粘类芽孢杆菌的工艺.农业工程学报.2016,第32卷(第7期),303-308. * |
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