CN100457889C - Method for preparing microbial feed addictive by multiple bacteria strain mixed culture - Google Patents
Method for preparing microbial feed addictive by multiple bacteria strain mixed culture Download PDFInfo
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Abstract
The invention relates to a method of producing microbe feed stuff addition agent by multi-bacteria, among which, the bacteria is: plant lactobacillus, Lactobacillus acidophilus, fibroin-producing candida, Saccharomyces cerevisiae; make fibroin-producing candida and Saccharomyces cerevisiae into saccharomycete mixing seed liquid; make plant lactobacillus and Lactobacillus acidophilus into lactobacillus mixing seed liquid; inoculate said saccharomycete mixing seed liquid and lactobacillus mixing seed liquid to saccharomycete and lactic acid bacteria mixing seed culture medium to get saccharomycete and lactic acid bacteria mixing seed liquid; inoculate saccharomycete and lactic acid bacteria mixing seed liquid at the quantity of 0.2-0.3% to the completed product produced through grading mixing fermentation in mixing fermentation culture medium; The mixing fermentation culture of saccharomycete and lactic acid bacteria is divided into two stages, the earlier stage is aerobic culture, and the later stage is anaerobic culture. The invention is characterized on low cost of production and low energy consumption; the bacteria could cooperate with each other well which enhances the phylactic power of animals and the availability ratio of feed stuff; the method relates to the multi-bacterial strain mixing culture of microbe feed stuff addition agent which accelerates the health growth of animals
Description
Technical field
The present invention relates to a kind of production method of probiotics, particularly adopt the method for multi-strain bacteria kind mixed culture probiotic feed additive.
Background technology
Bread is the staff of life.Food-safety problem is the significant problem that is related to people ' s health and national economy, and particularly after China's WTO, the agricultural product security problem has become the key factor that influences the agricultural products in China industrial competitiveness.Pork is the main animal food that people absorb, and safe feed is the basis of animal food safety.Develop safe in utilizationly, the feed additive of favourable pig growth is the key of development " nuisanceless pig raising technology ", and the microorganism fodder preparation is the good method of the first-selection of relatively generally acknowledging at present.Along with extensively carrying out of microorganism application technology, in the application facet of active bacteria formulation, people wish that a kind of product can exercise multiple function, solve many-sided problem, and many products all are made up of multiple microorganism.At present, on the production form of this series products, the general employing separately cultivated single culture, and the culture with multiple bacterium mixes then, forms many bacterial classifications multifunctional product.This mode of production requires every kind of bacterium to use the one time fermentation jar, requires investment of production equipment bigger, and energy consumption is higher, has improved production cost greatly.Present for the bacterial strain screening with synergy relation and combination or a stochastic process, lack effective theoretical direction, and can not coordinate relation between bacterium effectively for the mixed culture system of having used, make it reach best ecological level, the performance maximum effect.This has seriously hindered development and application that mixed bacterium is cultivated.
Summary of the invention
Purpose of the present invention aims to provide and a kind ofly can reduce production costs well, production energy consumption, can act synergistically well mutually between bacterial classification, improve the immunizing power of animal, improve efficiency of feed utilization, promote the method for the multi-strain bacteria kind mixed culture production microorganism feed addictive of animal health growth.
The objective of the invention is to realize by following manner:
Employed bacterial classification is: plant lactobacillus (Lactobacillus plantarum), Lactobacterium acidophilum (Lactobacillus acidophilus), Candida utilis (Candida tails), yeast saccharomyces cerevisiae (Saccharomyces cerevisiae); Its technology is: yeast saccharomyces cerevisiae and Candida utilis are made yeast mixed seeds liquid; Plant lactobacillus, Lactobacterium acidophilum are made milk-acid bacteria mixed seeds liquid; Above-mentioned resulting yeast mixed seeds liquid, Lactobacterium acidophilum mixed seeds liquid are inoculated in yeast and the milk-acid bacteria mixed seeds substratum, obtain yeast and milk-acid bacteria mixed seeds liquid; Yeast and milk-acid bacteria mixed seeds liquid is inoculated in the mixed fermentive culture medium by the inoculum size of 0.2-0.3% obtains finished product after the segmentation mixed fermentation; Described mixed fermentive culture medium is: 1-3% brown sugar, 1.5% glucose, 0.1-0.3% peptone, 0.5-1.5% dregs of beans, 0.1-0.6% radish juice, 0.1%K
2HPO
4, 0.1%MgSO
4.7H
2O, water 1000mL, pH 6.4-6.6; The segmentation mixing fermentation culture of described yeast and milk-acid bacteria is carried out in two stages, aerobic cultivation: culture temperature 28-30 ℃, air flow 300L/min whenever led to 1h and stopped 10min, mixing speed 40-50r/min early stage, every 3h 30min stops 30min, cultivates 20-30h; Later stage anaerobically fermenting: culture temperature 32-34 ℃, air flow 0L/min, mixing speed 40-50r/min, every 30min stops 30min, cultivates 20-30h.
Also add the 0.0-0.3% vegetables oil in the described mixed fermentive culture medium.
Best results when mixed fermentive culture medium is following component: 2% brown sugar, 1.5% glucose, 0.1% peptone, 1% dregs of beans, 0.5% radish juice, 0.1%K
2HPO
4, 0.1%MgSO
4.7H
2O, 0.15% vegetables oil, water 1000mL transfer pH 6.4-6.6.
Yeast mixed seeds liquid, Lactobacterium acidophilum mixed seeds liquid are inoculated in yeast and the milk-acid bacteria mixed seeds substratum by the inoculum size of 1-2%, 0.5-1.5% respectively.
Yeast mixed seeds liquid, Lactobacterium acidophilum mixed seeds liquid are inoculated in yeast and the milk-acid bacteria mixed seeds substratum by 1.5%, 1% inoculum size respectively; Yeast and milk-acid bacteria mixed seeds liquid is inoculated in by 0.2% inoculum size carries out mixed fermentation, best results in the mixed fermentive culture medium.
The present invention selects with bacterial classification by repeatedly studying and test for producing, and, plant lactobacillus (Lactobacillus plantarum), Lactobacterium acidophilum (Lactobacillus acidophilus), Candida utilis (Candida tails), yeast saccharomyces cerevisiae (Saccharomyces cerevisiae) carried out research, test by being produced with characteristic and the performance of bacterium for 4 kinds, test, the contriver finds that effective combination of above-mentioned 4 kinds of bacterial classifications has effect well for product of the present invention.And bacterial classification used in the present invention is the feed grade probiotic bacterium bacterial classification that the Ministry of Agriculture allows use.The present invention utilizes yeast to be facultative aerobe, milk-acid bacteria is microaerophilic characteristics, advanced person's gas cultivation of acting charitably, make yeast produce a large amount of thalline and meta-bolites, this culture condition does not damage the survival of milk-acid bacteria again simultaneously, after cultivating early stage, stop aerated culture, make yeast can carry out zymamsis, produce a certain amount of alcohol and carbonic acid gas, for the generation of ester perfumery matter and little aerobic growth of milk-acid bacteria provide condition, milk-acid bacteria can utilize saccharomycetic meta-bolites to grow simultaneously, further reduces the pH value, for the pollution that reduces the assorted bacterium of product creates conditions, and reduced production energy consumption.
The present invention utilizes saccharomycetic medium component simple, meta-bolites is nutritious, and the medium component complexity of milk-acid bacteria, and price is more expensive, saccharomycetic meta-bolites or dead cell autolyzate can provide the nutritional requirement of lactobacter growth, and research optimization has obtained plant lactobacillus (Lactobacillus plantarum), Lactobacterium acidophilum (Lactobacillus acidophilus), Candida utilis (Candida tails), the culture medium prescription of 4 kinds of bacterium mixed culture of yeast saccharomyces cerevisiae (Saccharomyces cerevisiae) (i.e. 2% brown sugar, 1.5% glucose, 0.1% peptone, 1% dregs of beans, 0.5% radish juice, 0.1%K
2HPO
4, 0.1%MgSO
4.7H
2O, 0.15% vegetables oil, water 1000mL, pH 6.4-6.6.Greatly reduce the peptone content in the general milk-acid bacteria substratum, saved yeast powder (cream) composition, and adopted low price, abundant bean cake powder to do nitrogenous source, and adopt the red pool to replace part sucrose or glucose to do carbon source, reduce about 1510.9 yuan/ton of raw materials for production costs.
The advantage of maximum of the present invention is, the present invention proposes plant lactobacillus (Lactobacillusplantarum), Lactobacterium acidophilum (Lactobacillus acidophilus), Candida utilis (Candida tails), yeast saccharomyces cerevisiae (Saccharomyces cerevisiae) are carried out mixed culture, the stepwise fermentation technology has creatively been proposed, mixed fermentation main aerobic culturing yeast bacterium in early stage of the present invention obtains thalline and meta-bolites, and main anaerobism of later stage is cultivated milk-acid bacteria and obtained thalline and make saccharomycetes to make fermentation produce wine flavour.And worked out well at four kinds of bacterium mixed fermentive culture mediums of the present invention simultaneously, four kinds of bacterium mixing fermentation culture conditions, these all do not have the pertinent literature report.And the crux that product of the present invention can successfully prepare also is determining of above-mentioned condition.
Product of the present invention is used for feeding of pig and chicken to be had promotes growth, improve feedstuff-meat ratio, improves immunizing power, reduces functions such as the environmental pollution and the generation that wards off disease animal; Can improve the vigor of boar sperm, reduce rate of teratosperm; There are not hormone and antibiotic remains in the meat product, resistance and drug residue have been avoided, of the present inventionly applying the competitiveness in the international market that can improve China's pork product, guarantee the quality and the hygienic safety of pork product, is the new generation product after microbiotic.Thereby the present invention be a kind of effectively utilize 4 kinds of bacterium well mixed fermentation produce the production method of this superior prod of microorganism feed addictive, the present invention can reduce production costs greatly, promotes applying of this series products.
Product of the present invention in the ChangSha, Hunan Province, part districts and cities such as Liuyang, Ningxiang, Lanshan County, Zhuzhou and large-scale pig farm promote, and delivers 260,000 bull pigs for sale, the raiser on average increases income more than 50 yuan/head, direct economic benefit reaches more than 1,500 ten thousand yuan.
Embodiment
Embodiment 1
For example the present invention is done more detailed description below:
Bacterial classification: Candida utilis [ACC20060], yeast saccharomyces cerevisiae [ACC20065], plant lactobacillus [ACC10644], Lactobacterium acidophilum [ACC10637] (being the purchase of Chinese agriculture microbial strains preservation center obtains).
Respectively above-mentioned four kinds of bacterial classifications are carried out actication of culture, Candida utilis, yeast saccharomyces cerevisiae are inoculated in respectively on the PDA slant medium, 20-30 ℃, cultivate 30-40h; After the activation, cultivate respectively and produce Candida utilis, yeast saccharomyces cerevisiae liquid seeds continuously; And plant lactobacillus, Lactobacterium acidophilum are inoculated in the MRS slant medium respectively, 30-40 ℃, cultivate 30-40h; After the activation, cultivate respectively and produce plant lactobacillus, Lactobacterium acidophilum liquid seeds continuously.
With yeast saccharomyces cerevisiae and Candida utilis combined inoculation in the YPD seed culture medium, under 26-30 ℃, 160-180r/min, cultivate 20-30h, obtain the yeast mixed seeds, and then be inoculated in the yeast mixed seeds substratum in 26-30 ℃ 160-180r/min, cultivate 20-30h, get yeast mixed seeds liquid.Being prepared as of yeast mixed seeds substratum wherein: peptone 10.0g, glucose 20.0g, yeast extract paste 5.0g, distilled water 1000mL, adjust pH are 6.8～7.0, sterilising treatment is stand-by.The contriver finds also in the yeast mixed seeds liquid that it is 1 that yeast saccharomyces cerevisiae is counted ratio with the bacterium of Candida utilis: during 1-3, can optimize product of the present invention.
Plant lactobacillus, Lactobacterium acidophilum combined inoculation in the MRS seed culture medium, under 30-34 ℃, are cultivated 20-30h, obtain the milk-acid bacteria mixed seeds, and then be inoculated in the milk-acid bacteria mixed seeds substratum under 30-34 ℃, cultivate 20-30h, milk-acid bacteria mixed seeds liquid.Wherein milk-acid bacteria mixed seeds medium preparation is: peptone 10g, extractum carnis 10g, yeast extract paste 5g, dipotassium hydrogen phosphate 2g, dibasic ammonium citrate 2g, sodium acetate 5g, glucose 20g, tween 80 1mL, magnesium sulfate heptahydrate 0.58g, four water manganous sulfate 0.25g, water 1000mL, adjust pH 6.2-6.4, sterilising treatment, stand-by.The contriver finds that also it is 1 that the bacterium of plant lactobacillus, Lactobacterium acidophilum in the milk-acid bacteria mixed seeds liquid is counted ratio: can optimize product of the present invention during 1-2.
Above-mentioned resulting yeast mixed seeds liquid, Lactobacterium acidophilum mixed seeds liquid are inoculated in yeast and the milk-acid bacteria mixed seeds substratum by the inoculum size of 1-2%, 0.5-1.5% respectively, under 30-34 ℃, 160-180r/min, cultivate 30-40h, obtain yeast and milk-acid bacteria mixed seeds liquid.Described yeast and milk-acid bacteria mixed seeds medium preparation are: 2% brown sugar, 1.5% glucose, 0.5% peptone, 0.5% radish juice, 0.1%K
2HPO
4, 0.1%MgSO
4.7H
2O, 0.15% vegetables oil, water 1000mL, adjust pH 6.4-6.6; Sterilising treatment, stand-by.
Yeast and milk-acid bacteria mixed seeds liquid is inoculated in by the inoculum size of 0.2-0.3% carries out mixed fermentation in the mixed fermentive culture medium, mixed fermentive culture medium is by the preparation of following component: 1-3% brown sugar, 1.5% glucose, 0.1-0.3% peptone, 0.5-1.5% dregs of beans, 0.1-0.6% radish juice, 0.1%K
2HPO
4, 0.1%MgSO
4.7H
2O, 0.0-0.3% vegetables oil, water 1000mL, pH 6.4-6.6; Sterilising treatment, stand-by.
The segmentation mixing fermentation culture condition of yeast and milk-acid bacteria is: early stage aerobic cultivation: culture temperature 28-30 ℃, air flow 300L/min, whenever logical 1h stops 10min, mixing speed 40-50r/min, every 3h 30min stops 30min, cultivates 20-30h; Later stage anaerobically fermenting: culture temperature 32-34 ℃, air flow 0L/min, mixing speed 40-50r/min, every 30min stops 30min, cultivates 20-30h.
Last multiparity product quality detects, and can, packing are checked and accepted.
With the prepared product of the present invention, in feed, to add product and directly feed by 0.3% of daily ration, its effect is obvious, and is simple, is worthy to be popularized.But feed after adding 0.3% microorganism fodder preparation fermentation feed, also may can play feeding effect preferably, save cost.
The product of the application of the invention proves that the resulting complex microorganism preparations of the present invention makes the pig colour of skin ruddy, and growth potential is good, and the physique of pig strengthens; Day weight gain improves 15.4%; Efficiency of feed utilization improves 11.7%; Coliform quantity in the ight soil reduces, and obnoxious flavoures such as ammonia reduce; The excrement apparent digestibility significant difference of protein and fat; Total phosphorous reduces in the ight soil.
The product the key technical indexes that the present invention produces:
Living bacteria count: 〉=2.5 * 10
8CFU/mL
Wherein: milk-acid bacteria 〉=2.0 * 10
8CFU/mL
Yeast 〉=0.5 * 10
8CFU/mL
Coliform group count :≤3/100mL.
Salmonellas: must not detect.
PH value: 3.0-5.0.
Quality guaranteed period: must not be lower than 6 months (〉=6 months).
Embodiment 2
Yeast mixed seeds liquid, Lactobacterium acidophilum mixed seeds liquid are inoculated in yeast and the milk-acid bacteria mixed seeds substratum by 1.5%, 1% inoculum size respectively; Yeast and milk-acid bacteria mixed seeds liquid inoculated by 0.2% inoculum size carry out mixed fermentation in the sub-mixed fermentive culture medium.
Mixed fermentive culture medium can reach best by the preparation of following component: 2% brown sugar, 1.5% glucose, 0.1% peptone, 1% dregs of beans, 0.5% radish juice, 0.1%K
2HPO
4, 0.1%MgSO
4.7H
2O, 0.15% vegetables oil, water 1000mL transfer pH 6.4-6.6; Sterilising treatment, stand-by.Other step is with embodiment 1.
Claims (8)
1, the method for preparing microbial feed addictive by multiple bacteria strain mixed culture, employed bacterial classification is: plant lactobacillus, Lactobacterium acidophilum, Candida utilis, yeast saccharomyces cerevisiae; Its technology is: yeast saccharomyces cerevisiae and Candida utilis are made yeast mixed seeds liquid; Plant lactobacillus, Lactobacterium acidophilum are made milk-acid bacteria mixed seeds liquid; Above-mentioned resulting yeast mixed seeds liquid, milk-acid bacteria mixed seeds liquid are inoculated in yeast and the milk-acid bacteria mixed seeds substratum, obtain yeast and milk-acid bacteria mixed seeds liquid; Yeast and milk-acid bacteria mixed seeds liquid is inoculated in the mixed fermentive culture medium by the inoculum size of 0.2-0.3% obtains finished product after the segmentation mixed fermentation;
In 26-30 ℃, 160-180r/min cultivates 20-30h to described yeast mixed seeds, gets yeast mixed seeds liquid in substratum; Wherein yeast mixed seeds substratum is: peptone 10.0g, and glucose 20.0g, yeast extract paste 5.0g, distilled water 1000mL, adjust pH are 6.8～7.0;
The milk-acid bacteria mixed seeds under 30-34 ℃, is cultivated 20-30h in substratum, get milk-acid bacteria mixed seeds liquid; Wherein milk-acid bacteria mixed seeds medium preparation is: peptone 10g, extractum carnis 10g, yeast extract paste 5g, dipotassium hydrogen phosphate 2g, dibasic ammonium citrate 2g, sodium acetate 5g, glucose 20g, tween 80 1mL, magnesium sulfate heptahydrate 0.58g, four water manganous sulfate 0.25g, water 1000mL, adjust pH 6.2-6.4;
Yeast mixed seeds liquid, the milk-acid bacteria mixed seeds liquid that obtains is inoculated in yeast and the milk-acid bacteria mixed seeds liquid culture medium, and under 30-34 ℃, 160-180r/min cultivates 30-40h, obtains yeast and milk-acid bacteria mixed seeds liquid; Described yeast and milk-acid bacteria mixed seeds substratum are: 2% brown sugar, 1.5% glucose, 0.5% peptone, 0.5% radish juice, 0.1%K
2HPO
4, 0.1%MgSO
4.7H
2O, 0.15% vegetables oil, water 1000mL, adjust pH 6.4-6.6;
Described mixed fermentive culture medium is: 1-3% brown sugar, 1.5% glucose, 0.1-0.3% peptone, 0.5-1.5% dregs of beans, 0.1-0.6% radish juice, 0.1%K
2HPO
4, 0.1%MgSO
4.7H
2O, water 1000mL, pH 6.4-6.6;
Described yeast and milk-acid bacteria segmentation mixing fermentation culture are then carried out in two stages, aerobic cultivation: culture temperature 28-30 ℃, air flow 300L/min whenever led to 1h and stopped 10min, mixing speed 40-50r/min early stage, every 3h 30min stops 30min, cultivates 20-30h; Later stage anaerobically fermenting: culture temperature 32-34 ℃, air flow 0L/min, mixing speed 40-50r/min, every 30min stops 30min, cultivates 20-30h.
2, the method for preparing microbial feed addictive by multiple bacteria strain mixed culture according to claim 1 adds the 0.0-0.3% vegetables oil in the mixed fermentive culture medium.
3, the method for preparing microbial feed addictive by multiple bacteria strain mixed culture according to claim 1 is inoculated in yeast and milk-acid bacteria mixed seeds liquid and carries out mixed fermentation in the mixed fermentive culture medium by 0.2% inoculum size.
4, the method for preparing microbial feed addictive by multiple bacteria strain mixed culture according to claim 2, described mixed fermentive culture medium is: 2% brown sugar, 1.5% glucose, 0.1% peptone, 1% dregs of beans, 0.5% radish juice, 0.1%K
2HPO
4, 0.1%MgSO
4.7H
2O, 0.15% vegetables oil, water 1000mL transfer pH 6.4-6.6.
5, the method for preparing microbial feed addictive by multiple bacteria strain mixed culture according to claim 1, yeast mixed seeds liquid, milk-acid bacteria mixed seeds liquid are inoculated in yeast and the milk-acid bacteria mixed seeds substratum by the inoculum size of 1-2%, 0.5-1.5% respectively.
6, the method for preparing microbial feed addictive by multiple bacteria strain mixed culture according to claim 5 is inoculated in yeast mixed seeds liquid, milk-acid bacteria mixed seeds liquid in yeast and the milk-acid bacteria mixed seeds substratum by 1.5%, 1% inoculum size respectively.
7, the method for preparing microbial feed addictive by multiple bacteria strain mixed culture according to claim 1, to count ratio with the bacterium of Candida utilis be 1 to yeast saccharomyces cerevisiae in the yeast mixed seeds liquid: 1-3.
8, the method for preparing microbial feed addictive by multiple bacteria strain mixed culture according to claim 1, in the milk-acid bacteria mixed seeds liquid bacterium of plant lactobacillus, Lactobacterium acidophilum to count ratio be 1: 1-2.
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| CN101703169B (en) * | 2009-11-02 | 2011-12-07 | 许民强 | Method for preparing live lactobacillus planterum preparation and application of live lactobacillus planterum preparation in preventing and treating piglet diarrhoea |
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| CN101946853B (en) * | 2010-08-25 | 2012-11-28 | 天津农学院 | Method for preparing silage feed by fermenting straw through composite microbes |
| CN105010731A (en) * | 2015-05-12 | 2015-11-04 | 河南科技大学 | Plant protein mixture preparation method |
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| CN107712266A (en) * | 2016-08-10 | 2018-02-23 | 湖南山河美生物环保科技股份有限公司 | Secondary fermentation grain slag produces the method and application method of high activity high nutrition feed |
| CN109593666B (en) * | 2017-09-30 | 2020-11-10 | 河南牧一动物药业有限公司 | Composite microecological preparation and preparation method and application thereof |
| CN107647086A (en) * | 2017-11-05 | 2018-02-02 | 长沙秋点兵信息科技有限公司 | Method for producing microbial feed additive by adopting multi-strain mixed culture |
| CN107853452A (en) * | 2017-11-13 | 2018-03-30 | 广西塔锡科技有限公司 | A kind of production method of additive for microbe feedstuff |
| CN111808786B (en) * | 2020-08-12 | 2024-03-29 | 沈阳市皇姑区乐伯欧生物技术信息咨询服务部 | Preparation method and application of plant yeast lactic acid bacteria |
| CN113180149B (en) * | 2021-05-11 | 2023-10-13 | 江南大学 | Method for producing probiotics feed by continuously fermenting vinasse serving as raw material |
| CN116058432B (en) * | 2023-03-02 | 2023-06-16 | 莱芜泰禾生化有限公司 | Citric acid mycelium residue microbial feed additive and preparation method and application thereof |
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Assignee: Hunan Taigu Biological Technology Co.,Ltd. Assignor: Li Jianwen Contract fulfillment period: 2008.10.10 to 2018.10.9 contract change Contract record no.: 2009430000196 Denomination of invention: Method for preparing microbial feed addictive by multiple bacteria strain mixed culture Granted publication date: 20090204 License type: Exclusive license Record date: 20091117 |
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