CN102505003B - High-density fermenting method of bacillus coagulans for livestock and poultry, as well as preparation prepared by method and application thereof - Google Patents

High-density fermenting method of bacillus coagulans for livestock and poultry, as well as preparation prepared by method and application thereof Download PDF

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CN102505003B
CN102505003B CN 201110428027 CN201110428027A CN102505003B CN 102505003 B CN102505003 B CN 102505003B CN 201110428027 CN201110428027 CN 201110428027 CN 201110428027 A CN201110428027 A CN 201110428027A CN 102505003 B CN102505003 B CN 102505003B
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高书锋
刘惠知
胡新旭
周映华
吴胜莲
缪东
周小玲
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HUNAN INST OF MICROBE
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Abstract

The invention discloses a high-density fermenting method of bacillus coagulans for livestock and poultry, as well as a preparation prepared by the method and application thereof. The strain collection number of the bacillus coagulans is CICC (China Center of Industrial Culture Collection) No.20138. The method comprises the following steps: performing liquid fermentation on the bacillus coagulans; performing solid alternating temperature continuous fermentation; ventilating in later phase of solid fermentation; continuing fermenting and drying, and preparing a bacillus coagulans preparation initial product after the drying is completed; further crushing and screening to obtain a high-density bacillus coagulans preparation final product, wherein the number of strains of the final product is 115.5*10<8>cfu.g<-1>, and the spore rate is 77.2%. The product has the effects of increasing the weight and preventing diarrhea of piglet, and the application adding concentration is 0.05wt%. The number of strain preparation is far higher than that of similar strain preparations (20*10<8>cfu.g<-1>) in the market, so that the preparation has wide market competitively in the livestock and poultry breeding industry.

Description

Preparation and the application of Bacillus coagulans fermentation process in high density and preparation thereof of a kind of livestock and poultry
Technical field
The present invention relates to microbial technology field, relate in particular to a kind of method that livestock and poultry continuously ferment with the liquid-solid segmentation alternating temperature of Bacillus coagulans and microbial inoculum and the application for preparing.
Background technology
In China, microbiotic uses the history of existing decades as fodder additives, and it has promoted giving full play to of breeding performonce fo animals, has prevented the infection of germ, and major contribution has been made in the development of intensive livestock industry.But Recent study shows, microbiotic also generally uses and brought the drawback that is difficult to overcome to the mankind, as the problems such as drug residue in destruction, animal products and the environment of the generation of the autogenous infection that causes and superinfection, resistance, normal intestinal flora.
Probiotics is as a kind of novel Substitutes For Antibiotic, has to have no side effect, have no drug resistance, noresidue, significantly reduce the plurality of advantages such as livestock and poultry noxious gas emission, obtained affirming of aquaculture, especially lactic acid bacteria class and bacillus category.But lactic acid bacteria class resistance is poor, can't tolerate the high temperature in feed processing pelletization, and inactivation is very fast in normal temperature transportation and preserving process simultaneously.Though bacillus category has can withstand high temperatures, the gemma of hydrochloric acid in gastric juice, choline, but do not possess intestinal canal colonization that milk-acid bacteria has, keep the functions such as the normal eubiosis and regulating intestinal canal pH in enteron aisle.
As a kind of livestock and poultry probiotics, Bacillus coagulans is lactic acid producing but also produce gemma not only, integrates the advantage of lactic acid bacteria class and gemma mushroom probiotics, possesses the unrivaled effect of feeding of other probiotics.At first, Bacillus coagulans possesses the characteristics of Strains of lactobacillus, can grow enteron aisle is decided at the higher level but not officially announced, be supplemented with bacteria group, adjust environment and micro-flora balance in digestive tube, form biological barrier at small bowel, prevention and treatment digestive function disorder and digestive tract infection, and can secrete the acidic substance such as lactic acid, regulating intestinal canal pH, promote the wriggling of enteron aisle to improve digestion, absorptive function; Secondly, Bacillus coagulans also can form gemma, the characteristics that possess gemma mushroom probiotics, can well tolerate high temperature in the feed course of processing, enteron aisle hydrochloric acid in gastric juice, choline etc., and greatly reduce deactivation phenomenom in the transportation preserving process, can produce simultaneously multiple digestive ferment, abundant VITAMIN and UGF etc. in digestive tube, can play the effect that assist digestion, promotion are grown.Numerous results of study show: Bacillus coagulans can promote digesting and assimilating, improve the animal body immunologic function, improving the effects such as animal disease resistant ability of animal body, thereby promotes the growth of animal, reduces its mortality ratio, mortality and feedstuff-meat ratio.Outstanding feed function and good processing characteristics make Bacillus coagulans will become irreplaceable important member in feed probiotics industry.
At present, the application report of Bacillus coagulans aspect additive of livestock and poultry is increasing, but the report of relevant its industrialization aspect is relatively less.Analyze its reason, mainly that Bacillus coagulans liquid fermenting level in industrialization process is low, fermented liquid is after the aftertreatment technologys such as absorption, drying and pulverizing, the biomass rate of loss is higher, cause the finished product biomass lower, directly affected the result of use of Bacillus coagulans microbial inoculum, therefore, the production method that works out a kind of high-density Bacillus coagulans microbial inoculum will be brought boundless vital force for livestock and poultry breeding industry and the green ecological agricultural of China.
Summary of the invention
The purpose of this invention is to provide a kind of livestock and poultry methods and applications of high-density Bacillus coagulans microbial inoculum and preparation thereof.
A kind of livestock and poultry are numbered with the method for Bacillus coagulans high density fermentation, described Bacillus coagulans (Bacillus coagulan) culture presevation: CICC No.20138(is available from Chinese industrial microbial strains preservation center), specifically comprise the following steps:
1) liquid fermenting
The inclined-plane seed of described bacterial strain is linked into the fermentor tank liquid fermentation and culture after cultivating through shake-flask seed, liquid fermentation medium preparation: starch 9.0-12.0g, fish meal 18.0-22.0g(is crushed to 60 orders), dipotassium hydrogen phosphate 2.0-3.2g, manganese sulfate monohydrate 0.10-0.30g, magnesium sulfate heptahydrate 0.60-0.90g, calcium carbonate 0.10-0.30g, sodium-chlor 0.15-0.40g and SODIUMNITRATE 0.10-0.30g, water adds to 1000mL, initial pH=6.5-7.2;
In fermentor tank, the loading amount of substratum accounts for the 40-60% of fermentor tank volume, and seed liquor is according to the 3-5% inoculation of culture volume, and the ferment tank parameter is temperature 35-40 ℃, rotating speed 220-280r.min -1, air flow 12.0-14.0L.min -1, fermentation time is 33-39h;
Inclined-plane used of the present invention seed activation substratum, all pass through before shake-flask seed substratum and fermentation tank culture medium are used warm sterilization (121 ℃, 30mins).
2) solid fermentation
Press the weight access liquid fermenting seed liquor of substratum 25-33% to the solid fermentation substratum, leavening temperature 34-36 ℃, the fermentation indoor relative humidity is controlled and is not less than 75%, 45-51h continuously ferments, regularly turn over even once (generally every 12 hours), wherein ferment after 34-38h, regulating and controlling temperature is 38-42 ℃, and drying aid simultaneously ventilates;
The solid fermentation substratum is made of wheat bran, moisture content in medium 38-42%, and this water content comprises the moisture in the liquid fermenting seed liquor of access.
After the step 1) liquid fermenting is completed, biomass reaches 54.5-62.0 * 10 8Cfu.mL -1
Step 2) after solid fermentation is completed, biomass reaches 108.5-127.0 * 10 8Cfu.g -1, the gemma rate reaches 71.3-74.4%.
Step 2) in, wheat bran is made to pulverize before substratum and is the 30-50 order, dry heat treatment 1-3h intermittently under 80 ℃ of conditions.
A kind of livestock and poultry are to be pulverized by the product that above-mentioned method fermentation is prepared to obtain with high-density Bacillus coagulans preparation.
Bacteria preparation finished product biomass reaches 102.4-115.5 * 10 8Cfu.g -1, the gemma rate reaches 72.7-77.2%.
Pulverizer is adopted in described pulverizing, and rotating speed is 20000-30000r.min -1, grinding time is 7-13s, pulverizes the order number and is not less than 60 orders.
The preparation that described livestock and poultry are suffered from diarrhoea for the preparation of Animal liveweight gain and prevention with high-density Bacillus coagulans preparation.Specifically described bacteria preparation can be added in feed, feed and raise domestic animal (generally using object to be pig, especially weanling pig).Adding concentration in feed is 0.05wt%.
Beneficial effect of the present invention is, has greatly promoted biomass (127.0 * 10 by the liquid-solid two double dry technical schemes of continuous temperature-variable fermentations of step 8Cfu.g -1), biomass is liquid fermenting biomass (62.0 * 10 8Cfu.mL -1) 2.05 times, by the solid fermentation later stage alternating temperature regulation and control dry technical scheme of holding concurrently, make the gemma rate bring up to 74.4% from 63.7% of contrast simultaneously, the solid fermentation dry whole process of holding concurrently is that low temperature carries out in addition, energy consumption is little, has strengthened the competitive power of the present invention in the livestock and poultry cultivation industry.
Embodiment
Be intended to further illustrate the present invention below in conjunction with embodiment, and unrestricted the present invention.
Embodiment 1
(1) liquid fermenting
A) inclined-plane seed activation: primordial seed is seeded to test tube ordinary nutrient agar slant substratum, 37 ℃ of incubated overnight 24h.The ordinary nutrient agar slant substratum (peptone 10g, extractum carnis 3g, sodium-chlor 5g, agar 20g, water adds to 1L, pH=6.5-7.2);
B) shake-flask seed substratum: inclined-plane seed inoculation one ring to three rings are arrived common nutrient broth medium, 37 ℃, 200r.min -1Incubated overnight 24h; Common nutrient broth medium (peptone 10g, extractum carnis 3g, sodium-chlor 5g, water adds to 1L, pH=6.5-7.2);
C) above-mentioned seed liquor is inoculated into the 20L fermentor tank according to 4% of culture volume and carries out liquid culture, culture volume is 50% of fermentor tank volume;
Fermention medium preparation: starch 10.6g.L -1, fish meal 20.2g.L -1, dipotassium hydrogen phosphate 2.72g.L -1, manganese sulfate monohydrate 0.169g.L -1, magnesium sulfate heptahydrate 0.739g.L -1, calcium carbonate 0.20g.L -1, sodium-chlor 0.234g.L -1With SODIUMNITRATE 0.17g.L -1, solvent is water, initial pH=7.0;
Fermentation: press 4% inoculation of culture volume, be seeded to the ferment tank substratum, regulation and control ferment tank parameter is 37 ℃ of temperature, rotating speed 250r.min -1, air flow 13.3L.min -1, treat that fermentation time is 36h, fermentation ends is taken a sample to fermented liquid, carries out gradient dilution with sterilized water, and tilt-pour process detects bacteria containing amount, triplicate, biomass reaches 62.0 * 10 8Cfu.mL -1Detect substratum: glucose 20g, Tryptones 20g, yeast extract paste 10g, agar 18g, salts solution 10mL (containing in the 1mL salts solution: magnesium sulfate heptahydrate 5mg, manganese sulfate monohydrate 1mg, calcium carbonate 2mg, sodium-chlor 5mg)
The described inclined-plane used of the present embodiment seed activation substratum, shake-flask seed substratum and fermentation tank culture medium are all passed through high-temperature sterilization before using, and embodiment 2 and 3 is identical.
(2) solid fermentation
A) substratum configuration: wheat bran (pulverizing is 40 orders left and right, 80 ℃ intermittently dry heat treatment 2h), water content 40%, this water content comprise the moisture in the liquid fermenting seed liquor of access.
B) fermentation: connect the liquid fermenting seed liquor to the solid fermentation substratum by 30% of substratum weight, temperature maintains 35 ℃, the fermentation indoor relative humidity is controlled at more than 75%, the 48h that continuously ferments, rear 12h carries out air seasoning, end to be dried, sampling 10g, detect biomass and gemma amount, triplicate, biomass and gemma amount are respectively 115.5 * 10 8Cfu.g -1With 73.5 * 10 8Cfu.g -1Biomass detection method and detection substratum are with embodiment 1(1).The gemma quantity measuring method: get the 10g sample, put in the 90mL sterilized water, 80 ℃ of water-bath thermal treatment 10mins carry out gradient dilution with sterilized water, and tilt-pour process detects the gemma amount, and the gemma amount detects substratum with embodiment 1(1).
(3) solid fermentation later stage differing temps regulation and control
Embodiment 1(2) the solid fermentation later stage (after fermentation 36h), carrying out hold concurrently dry alternating temperature of solid fermentation by 30 ℃, 40 ℃ and 45 ℃ respectively processes, take 35 ℃ of fermentations of temperature not as contrast, after the drying of holding concurrently wait fermenting finishes, get respectively the sample 10g that different alternating temperatures are processed, detect biomass and gemma amount, calculate the gemma rate, triplicate, detection method is with embodiment 1(2); Result shows: in the solid fermentation later stage (fermentation 36h after), when alternating temperature was regulated to 40 ℃ and 45 ℃, the gemma rate was respectively 74.4% and 75.9%, is better than contrasting gemma rate 63.7%; The solid fermentation later stage (after fermentation 36h), when alternating temperature was regulated to 40 ℃, biomass and gemma amount were all the highest, are respectively 127.0 * 10 8Cfu.g -1And 94.5.0 * 10 8Cfu.g -1Therefore, the solid fermentation later stage (fermentation 36h after) selects 40 ℃ to carry out the alternating temperature regulation and control.
The impact of table 1 solid fermentation later stage alternating temperature regulation and control on microbial inoculum biomass and gemma rate
Figure GDA00002462312300041
(4) impact of different grinding times on the microbial inoculum biomass
From pressing embodiment 1(1), embodiment 1(2) and embodiment 1(3) the product of most preferred technique scheme preparation, get respectively the 100g sample and pulverize, the pulverizing rotating speed is 24000r.min -1Grinding time is respectively 0s, 3s, 7s, 10s, 13s and 17s.After pulverizing end, get respectively the product 10g of different grinding times, detect biomass, triplicate.Biomass detection method and detection substratum are with embodiment 1(1).Detect at last finished product gemma number, calculate the gemma rate.Result shows: from smashing fineness, and grinding time 10-17s, smashing fineness satisfies production requirement greater than 60 orders; When grinding time was 17s, biomass was 102.5 * 10 8Cfu.g -1, rate of loss is 19.3%, loses greatlyr, and therefore, grinding time is strict controlled in 17s, and the optimal selection grinding time is 10s.Namely obtain finished product after pulverizing 10s, finished product thalline number is 115.5 * 10 8Cfu.g -1, the gemma number is 89.2 * 10 8Cfu.g -1, the gemma rate is 77.2%.
The impact of the different grinding times of table 2 on the microbial inoculum biomass
Figure GDA00002462312300051
Embodiment 2
(1) liquid fermenting
A) inclined-plane seed activation: primordial seed is seeded to test tube ordinary nutrient agar slant substratum, 37 ℃ of incubated overnight 24h.The ordinary nutrient agar slant substratum (peptone 10g, extractum carnis 3g, sodium-chlor 5g, agar 20g adds water to 1L, pH=6.5-7.2);
B) shake-flask seed substratum: inclined-plane seed inoculation one ring to three rings are arrived common nutrient broth medium, 37 ℃, 200r.min -1Incubated overnight 24h; Common nutrient broth medium (sodium-chlor 5g adds water to 1L, pH=6.5-7.2 for peptone 10g, extractum carnis 3g);
C) above-mentioned seed liquor is inoculated into the 20L fermentor tank according to 3% of culture volume and carries out liquid culture, culture volume is 40% of fermentor tank volume;
Fermention medium preparation: starch 9.0g.L -1, fish meal (the conventional purchase is crushed to 60 orders) 18.0g.L -1, dipotassium hydrogen phosphate 2.0g.L -1, manganese sulfate monohydrate 0.10g.L -1, magnesium sulfate heptahydrate 0.60g.L -1, calcium carbonate 0.10g.L -1, sodium-chlor 0.15g.L -1With SODIUMNITRATE 0.10g.L -1, solvent is water, initial pH=6.5;
Fermentation: press 3% inoculation of culture volume, be seeded to the ferment tank substratum, regulation and control ferment tank parameter is 35 ℃ of temperature, rotating speed 220r.min -1, air flow 12.0L.min -1, treat that fermentation time is 33h, fermentation ends is taken a sample to fermented liquid, carries out gradient dilution with sterilized water, and tilt-pour process detects bacteria containing amount, triplicate, biomass reaches 54.5 * 10 8Cfu.mL -1Detect substratum with embodiment 1(1).
(2) hold concurrently alternating temperature regulation and control of solid fermentation
A) substratum configuration: wheat bran (pulverizing is 30 orders left and right, 80 ℃ intermittently dry heat treatment 1h), water content 38%, this water content comprise the moisture in the liquid fermenting seed liquor of access.
B) fermentation: connect the liquid fermenting seed liquor to the solid fermentation substratum by 25% of substratum weight, temperature maintains 34 ℃, the fermentation indoor relative humidity is controlled at more than 75%, the 45h that continuously ferments, after fermentation 34h, regulating and controlling temperature is 38 ℃ and carries out solid fermentation and hold concurrently dryly, drying aid simultaneously ventilates, end to be dried, sample thief 10g detects biomass and gemma amount, triplicate.Biomass and gemma quantity measuring method and detection substratum are with embodiment 1(2).Result shows: the sample biomass is 122.5 * 10 8Cfu.g -1, the gemma amount is 87.3 * 10 8Cfu.g -1, the gemma rate is 71.3%.
(3) impact of different grinding times on the microbial inoculum biomass
From by embodiment 2(1) and embodiment 2(2) the product of technical scheme preparation get respectively the 100g sample and pulverize, the pulverizing rotating speed is 20000r.min -1Grinding time is respectively 0s, 3s, 7s, 10s, 13s, 17s and 20s.After pulverizing end, detect fineness, the product that fineness is processed greater than the 60 minimum grinding times of purpose is finished product, gets finished product 10g, detects biomass and gemma amount, calculates the gemma rate, triplicate.Biomass, gemma quantity measuring method and detection substratum are with embodiment 1(2).Result shows: from smashing fineness, grinding time is 13s or during greater than 13s, smashing fineness satisfies production requirement greater than 60 orders, therefore, namely obtains finished product after pulverizing 13s, and finished product biomass and gemma amount are respectively 112.0 * 10 8Cfu.g -1With 81.5 * 10 8Cfu.g -1, the gemma rate is 72.7%.
Embodiment 3
(1) liquid fermenting
A) inclined-plane seed activation: primordial seed is seeded to test tube ordinary nutrient agar slant substratum, 37 ℃ of incubated overnight 24h.The ordinary nutrient agar slant substratum (peptone 10g, extractum carnis 3g, sodium-chlor 5g, agar 20g adds water to 1L, pH=6.5-7.2);
B) shake-flask seed substratum: inclined-plane seed inoculation one ring to three rings are arrived common nutrient broth medium, 37 ℃, 200r.min -1Incubated overnight 24h; Common nutrient broth medium (sodium-chlor 5g adds water to 1L, pH=6.5-7.2 for peptone 10g, extractum carnis 3g);
C) above-mentioned seed liquor is inoculated into the 20L fermentor tank according to 5% of culture volume and carries out liquid culture, culture volume is 60% of fermentor tank volume;
Fermention medium preparation: starch 12.0g.L -1, fish meal (the conventional purchase is crushed to 60 orders) 22.0g.L -1, dipotassium hydrogen phosphate 3.2g.L -1, manganese sulfate monohydrate 0.30g.L -1, magnesium sulfate heptahydrate 0.90g.L -1, calcium carbonate 0.30g.L -1, sodium-chlor 0.40g.L -1With SODIUMNITRATE 0.30g.L -1, water is solvent, initial pH=7.2;
Fermentation: press 5% inoculation of culture volume, be seeded to the ferment tank substratum, regulation and control ferment tank parameter is 40 ℃ of temperature, rotating speed 280r.min -1, air flow 14.0L.min -1, treat that fermentation time is 39h, fermentation ends is taken a sample to fermented liquid, carries out gradient dilution with sterilized water, and tilt-pour process detects bacteria containing amount, triplicate, biomass reaches 57.2 * 10 8Cfu.mL -1Detect substratum and detect substratum with embodiment 1(1);
(2) hold concurrently alternating temperature regulation and control of solid fermentation
A) substratum configuration: wheat bran (pulverizing is 50 orders left and right, 80 ℃ intermittently dry heat treatment 3h), water content 42%, this water content comprise the moisture in the liquid fermenting seed liquor of access.
B) fermentation: connect the liquid fermenting seed liquor to the solid fermentation substratum by 33% of substratum weight, temperature maintains 36 ℃, the fermentation indoor relative humidity is controlled at more than 75%, the 51h that continuously ferments, after fermentation 38h, regulating and controlling temperature is 42 ℃ and carries out solid fermentation and hold concurrently dryly, drying aid simultaneously ventilates, end to be dried, sample thief 10g detects biomass and gemma amount, triplicate.Biomass and gemma quantity measuring method and detection substratum are with embodiment 1(2).Result shows: the sample biomass is 108.5 * 10 8Cfu.g -1, the gemma amount is 78.4 * 10 8Cfu.g -1, the gemma rate is 72.2%;
(3) impact of different grinding times on the microbial inoculum biomass
From by embodiment 3(1) and embodiment 3(2) the product of technical scheme preparation get respectively the 100g sample and pulverize, the pulverizing rotating speed is 30000r.min -1, grinding time is respectively 0s, 3s, 7s, 10s, 13s, 17s and 20s.After pulverizing end, detect fineness, the product that fineness is processed greater than the 60 minimum grinding times of purpose is finished product, gets finished product 10g, detects biomass and gemma amount, calculates the gemma rate, triplicate.Biomass, gemma quantity measuring method and detection substratum are with embodiment 1(2).Result shows: from smashing fineness, grinding time is 7s or during greater than 7s, smashing fineness satisfies production requirement greater than 60 orders, therefore, namely obtains finished product after pulverizing 7s, and finished product biomass and gemma amount are respectively 102.4 * 10 8Cfu.g -1With 77.5 * 10 8Cfu.g -1, the gemma rate is 75.7%.
Embodiment 4, the Bacillus coagulans preparation impact on child care pig production performance
Finished product by most preferred embodiment 1 technical scheme preparation is carried out clinical trial, and concrete scheme is: test site: there is greatly the back of the body kind of pig farm, cultivation Development Co., Ltd's Changsha County river in the Hunan; Experimental animal: Du Luoke * length is white * Da Bai original seed weanling pig; Test grouping: with the Du Luoke of 120 7kg left and right * length in vain * Da Bai original seed weanling pig is divided into 4 groups, 30 every group at random.
The conventional daily ration of feeding take the pig farm is basal diet (seeing Table 3), control group and the test group scheme (seeing Table 4) of feeding; Preliminary trial period 5d, 28 days trial periods,, the duration of test free choice feeding is freely drunk water.Calculate the initial mean body weight of test, the off-test mean body weight, average daily gain (ADG), average daily ingestion amount (ADFI), feed conversion rate (FCR), every day, observed and recorded diarrhoea situation, calculated diarrhea rate.
Result shows: the FCR that B, C and D process is respectively 1.397,1.413 and 1.427, all processes FCR1.55 less than CK; The diarrhea rate that B, C and D process is respectively 3.2%, 3.8% and 4.3%, all process diarrhea rate 13.2% less than CK, therefore product of the present invention is by the 0.05wt%(B processing) and the 0.1%wt%(C processing) concentration is added the daily ration of feeding to and weanling pig had the effect of weightening finish and prevention diarrhoea, and action effect is better than the common Bacillus coagulans preparation (D processing) through the direct preparation of liquid fermenting by the interpolation of 0.20wt% concentration.
The basic diet raw material of table 3 forms and trophic level
Figure GDA00002462312300081
Table 4 test design
The impact of table 5 Bacillus coagulans preparation on the child care weaned piglets
The present invention has worked out a kind of liquid-solid segmentation alternating temperature and has continuously fermented and prepare method and the using method of high-density Bacillus coagulans microbial inoculum.The finished product biomass can reach 115.5 * 10 8Cfu.g -1, the gemma rate can reach 77.2%, and product of the present invention has the effect of weightening finish and prevention diarrhoea to piglet.This microbial inoculum biomass is far above similar microbial inoculum biomass (20 * 10 on market 8Cfu.g -1), have the wide market competitiveness in the livestock and poultry cultivation industry.
Should be understood that, for those of ordinary skills, can be improved according to the above description or conversion, and all these improvement or conversion all belong to the protection domain of claims of the present invention.

Claims (10)

1. livestock and poultry are with the method for Bacillus coagulans high density fermentation, described Bacillus coagulans (Bacillus coagulan) culture presevation numbering: CICC No.20138, it is characterized in that, and specifically comprise the following steps:
1) liquid fermenting
The inclined-plane seed of described bacterial strain is linked into the fermentor tank liquid fermentation and culture after cultivating through shake-flask seed, liquid fermentation medium preparation: starch 9.0-12.0g, fish meal 18.0-22.0g, dipotassium hydrogen phosphate 2.0-3.2g, manganese sulfate monohydrate 0.10-0.30g, magnesium sulfate heptahydrate 0.60-0.90g, calcium carbonate 0.10-0.30g, sodium-chlor 0.15-0.40g and SODIUMNITRATE 0.10-0.30g, water adds to 1000mL, initial pH=6.5-7.2;
In fermentor tank, the loading amount of substratum accounts for the 40-60% of fermentor tank volume, and seed liquor is according to the 3-5% inoculation of culture volume, and the ferment tank parameter is temperature 35-40 ℃, rotating speed 220-280r.min -1, air flow 12.0-14.0L.min -1, fermentation time is 33-39h;
2) solid fermentation
Press the obtained liquid fermentation liquid of weight access step 1) of substratum 25-33% to the solid fermentation substratum, leavening temperature 34-36 ℃, the fermentation indoor relative humidity is controlled and is not less than 75%, 45-51h continuously ferments, regularly turn over once even, wherein ferment after 34-38h, regulating and controlling temperature is 38-42 ℃, and drying aid simultaneously ventilates;
The solid fermentation substratum is made of wheat bran, moisture content in medium 38-42%, and this water content comprises the moisture in the liquid fermentation liquid of access.
2. preparation method according to claim 1, is characterized in that, after the step 1) liquid fermenting is completed, biomass reaches 54.5-62.0 * 10 8Cfu.mL -1
3. preparation method according to claim 1, is characterized in that step 2) solid fermentation complete after biomass reach 108.5-127.0 * 10 8Cfu.g -1, the gemma rate reaches 71.3-74.4%.
4. preparation method according to claim 1, is characterized in that step 2) in wheat bran make to pulverize before substratum and be the 30-50 order, dry heat treatment 1-3h intermittently under 80 ℃ of conditions.
5. livestock and poultry with high-density Bacillus coagulans preparation, is characterized in that, described bacteria preparation is to be pulverized by the product that the described method fermentation of claim 1-4 any one is prepared to obtain.
6. bacteria preparation according to claim 5, is characterized in that, bacteria preparation finished product biomass reaches 102.4-115.5 * 10 8Cfu.g -1, the gemma rate reaches 72.7-77.2%.
7. bacteria preparation according to claim 6, is characterized in that, pulverizer is adopted in described pulverizing, and rotating speed is 20000-30000r.min -1, grinding time is 7-13s, pulverizes the order number and is not less than 60 orders.
8. claim 5 or 6 or 7 described livestock and poultry with the purposes of high-density Bacillus coagulans preparation, is characterized in that, for the preparation of the preparation that can promote Animal liveweight gain and prevention diarrhoea.
9. livestock and poultry according to claim 8 with the purposes of high-density Bacillus coagulans preparation, is characterized in that, described bacteria preparation is added in feed, feed and raise domestic animal.
10. livestock and poultry according to claim 9 with the purposes of high-density Bacillus coagulans preparation, is characterized in that, adding concentration in feed is 0.05wt%.
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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
CN102943057B (en) * 2012-11-16 2014-06-25 南京工业大学 Bacillus coagulans and process for high-density fermentation of bacillus coagulans
CN103255080B (en) * 2013-03-22 2015-05-27 内蒙古和美科盛生物技术有限公司 Excellent bacillus coagulans for breeding piglets and application thereof
CN103255082B (en) * 2013-03-22 2018-09-21 内蒙古和美科盛生物技术有限公司 One plant of excellent bacillus coagulans and its application for growing and fattening pigs cultivation
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CN104762231A (en) * 2015-03-26 2015-07-08 浙江省农业科学院 Method for preparing feed living bacteria agent in combination of bacillus coagulans and saccharomyces cerevisiae
CN104974966B (en) * 2015-07-22 2017-12-12 江南大学 A kind of bacillus coagulans and its fermentation process in high density and dry bacterium powder preparation method
CN104974967B (en) * 2015-07-22 2018-04-17 江南大学 One bacillus pumilus and its child care piglet compound micro-ecological preparation
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