CN106509468A - Composite type synbiotic microecological preparation for chickens and preparation method of composite type synbiotic microecological preparation - Google Patents

Composite type synbiotic microecological preparation for chickens and preparation method of composite type synbiotic microecological preparation Download PDF

Info

Publication number
CN106509468A
CN106509468A CN201610990924.XA CN201610990924A CN106509468A CN 106509468 A CN106509468 A CN 106509468A CN 201610990924 A CN201610990924 A CN 201610990924A CN 106509468 A CN106509468 A CN 106509468A
Authority
CN
China
Prior art keywords
chicken
probiotic bacteria
oligosaccharide
preparation
compound
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Pending
Application number
CN201610990924.XA
Other languages
Chinese (zh)
Inventor
刘君
刘华康
王晗
陈静
罗雁
刘永明
张薇
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Sichuan University of Science and Engineering
Original Assignee
Sichuan University of Science and Engineering
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Sichuan University of Science and Engineering filed Critical Sichuan University of Science and Engineering
Priority to CN201610990924.XA priority Critical patent/CN106509468A/en
Publication of CN106509468A publication Critical patent/CN106509468A/en
Pending legal-status Critical Current

Links

Classifications

    • AHUMAN NECESSITIES
    • A23FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23YINDEXING SCHEME RELATING TO LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
    • A23Y2220/00Lactobacillus
    • AHUMAN NECESSITIES
    • A23FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23YINDEXING SCHEME RELATING TO LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
    • A23Y2300/00Bifidobacterium

Abstract

The invention discloses a composite type synbiotic microecological preparation for chickens and a preparation method of the composite type synbiotic microecological preparation. The composite type synbiotic microecological preparation consists of probiotics and prebiotics, wherein the probiotics and the prebiotics are mixed according to the mass ratio of the probiotics to the prebiotics being 1 to (1-5); the probiotics consist of bacterium lacticum, bifidobacteria, bacillus and enterococcus; and the prebiotics consist of oligosaccharide and an acidulant, the oligosaccharide can be mannose oligosaccharides, fructo-oligosaccharide, isomaltooligosacharide and gynolactose, and the acidulant is a composite acidulant consisting of lactic acid, citric acid and fumaric acid. According to the composite type synbiotic microecological preparation and the preparation method thereof disclosed by the invention, the defect of separately using probiotic products for raising commodity chickens is overcome, besides, the composite type synbiotic microecological preparation is suitable for various commodity chickens, the purposes of making best use of the advantages and overcoming the disadvantages can be achieved, the field planting of exogenous probiotics in intestinal tracts of animals is promoted, the rapid growth and propagation of various probiotics in chicken bodies is selectively stimulated, the purpose of promoting the health of the chickens is achieved, the production properties are also greatly improved, the influence of conventional edible animal additives on chicken meat and egg quality is reduced, and the composite type synbiotic microecological preparation and the preparation method are economic and practical.

Description

One breeder compound symphysis unit microbial ecological agent and preparation method thereof
Technical field
The invention belongs to microbial ecological agent technical field, more particularly to a breeder with compound symphysis unit microbial ecological agent and Its preparation method.
Background technology
At present, applied research of China's symphysis unit microbial ecological agent in the production of animal is also less, farm animals food Service condition with microbial ecological agent in additive is also less, and people are suitable for addition to microbial ecological agent and relatively obscure, that is, obscure Targetedly add object, cause the effect of growth of animal to be often below Expected Results.The appearance of microbial ecological agent be due to The improvement of people's living standards, people gradually recognize that antibiotic can be good for the mankind by food chain in making an addition to feedstuff for a long time Health works the mischief.The research application of microbial ecological agent overcomes a variety of drawbacks that life-time service antibiotic is caused to aquaculture, right Development green feed additive has broad application prospects, and oneself is classified as 21st century emphasis promotional technique item by the Ministry of Agriculture Mesh.This means that the research to microbial ecological agent, popularization meet the general requirement of China's sustainable development.
In existing microbial ecological agent, the stability of bacterial strain is not ensured well, and throwing must be with the shape of viable bacteria to bacterium Formula reaches gut metabolism by stomach, and colonization is trained in intestinal and grown, and this is not to be easy to do for antibacterial to general spore form Arrive;Microbial ecological agent does not targetedly add object, and domestic abundant not enough to the application of microbial ecological agent, popularization degree is not Enough, the not a height of solution abuse of antibiotics of the degree that received by raiser causes increasing pathogenic bacterium to develop immunity to drugs, medicine Thing is remained, and reduces chicken meat product quality and as food chain is accumulated to having a strong impact on that human body health is brought, and production is replaced For the microbial ecological agent of antibiotic.
The content of the invention
It is an object of the invention to provide a breeder compound symphysis unit microbial ecological agent and preparation method thereof, it is intended to solve Certainly existing abuse of antibiotics causes increasing pathogenic bacterium to develop immunity to drugs, drug residue, reduces Carnis Gallus domesticus, scrambled egg product quality And as food chain accumulates the problem having a strong impact on brought to human body health.
The present invention is achieved in that the compound symphysis unit microbial ecological agent of a breeder, the compound symphysis of the chicken First microbial ecological agent is made up of probiotic bacteria and prebioticses;
The probiotic bacteria:Prebioticses=1:1~5 quality mixes than ratio.
Further, the probiotic bacteria constitutes (lactobacilluss, bifid bar by lactobacilluss, bacillus bifiduss, bacillus cereuss and enterococcus Bacterium, bacillus cereuss and enterococcal reference culture are purchased from Beijing North Na Chuanlian Bioteknologisk Institut).
Further, the prebioticses are made up of oligosaccharide and acidulant;
The oligosaccharide is MOS, fruit oligose, oligomeric isomaltose, newborn oligosaccharide, acidulant by lactic acid, citric acid, The compound acidulant of Fumaric acid composition.
Further, the probiotic bacteria is 0.02-15g/kg, and probiotic bacteria nutrient substance is 0.05-80g/kg, and oligosaccharide is 0.01-90g/kg。
Further, the addition of oligosaccharide be MOS 0.25-2g/kg, fruit oligose 0.01-3g/kg, oligomeric Isomalt Sugared 0.01-3g/kg, newborn oligosaccharide 0.01-3g/kg;The compound acidulant that addition is made up of lactic acid, citric acid, Fumaric acid 0.1%-0.5% (accounts for microbial ecological agent ratio).
Another object of the present invention is to a kind of preparation method of chicken with compound symphysis unit microbial ecological agent is provided, The preparation method is comprised the following steps:
Separation first against property, beneficial bacteria of intestinal tract is filtered out, by the work for comparing probiotic bacteria under simulation intestinal environment Bacterium number changes, and filters out optimal bacterial strain, all kinds of oligosaccharide contents are measured, optimum addition is selected, by suitable chicken Stomach, the probiotic bacteria of intestinal and oligosaccharide are mixed and made into compound symphysis unit microbial ecological agent.
Further, the chicken is specifically included with the preparation method of compound symphysis unit microbial ecological agent:
Step one, the soil near the gastrointestinal tract of chicken, chicken farm separate lactobacilluss, bacillus bifiduss, variable X bacterium Probiotic bacteria, lactobacilluss, bacillus bifiduss, bacillus cereuss and enterococcal probiotic bacteria morphological characteristic that analytical separation is obtained are gone forward side by side The identification of row strain;
Step 2, isolated lactobacilluss, bacillus bifiduss, bacillus cereuss and enterococcal probiotic bacteria are carried out into biology The identification of characteristic, the comparison of function and safety, and external probiotic effects evaluation is carried out, preliminary screening high-quality probiotic strain;
Step 3, the high-quality probiotic bacteria in the gastrointestinal fluid of manual simulation after culture preliminary screening, determine all kinds of prebiotic The viable count of bacterium, reproductive order of generation are further screened, and select optimal bacterial strain;
Step 4:Species, the functional characteristic of all kinds of prebioticses according to probiotic bacteria and the utilization journey with corresponding probiotic bacteria Degree selects the prebioticses being made up of oligosaccharide and acidulant, optimization oligosaccharide and acidulant optimum addition;
The prebioticses that step 5, probiotic strain and step 4 to step 3 screening are selected, add probiotic bacteria nutrition Material mixing makes compound symphysis unit microbial ecological agent.
Further, the identification that strain is carried out in step one includes:
General survey:Lactobacilluss are Gram-positive sporeless bacterium, cellular morphology in chain is shaft-like or club shape, bifid bar Bacterium is gram-positive sporeless bacterium, and varying length, bacillus bifiduss have V-shape or a Y-shaped, arrangement in single, double, short chain, X, Y, V or palisade;Bacillus cereuss breeding is quick, breeds 100,000 times within four hours, vitality by force without wet condition can low temperature resistant -60 DEG C, + 280 DEG C of high temperature resistant, resistance to strong acid, resistance to highly basic, antibiotic and sterilizing, resistance to hyperoxia (thermophilic oxygen breeding), lower oxygen concentration resistance (anaerobism breeding).Volume ratio The four times greater number of general nosophyte numerator, take up space advantage, suppresses the growth and breeding of harmful bacteria.Enterococcus are circular or oval Shape, in the gram positive coccus in single paired or short catenation, without brood cell, atrichia, be aerobic or facultative anaerobic bacteria. This bacterium is higher to nutritional requirement, it is desirable to high salt (6.5%NaCl), high-alkali (pH9.6), 40% bile culture-medium and 10-45 DEG C of ring Grow under border.
Lactobacilluss, bacillus bifiduss, bacillus cereuss and enterococcus probiotic bacteria and the prebioticses selected are combined, Adding is beneficial to the nutrient substance for being conducive to growth of probiotics, makes compound symphysis unit microbial ecological agent.
Nutrient substance:As chicken belongs to birds, dilution normal saline used slightly has area with mammalss, Amphibian Not, General N aCl concentration is 0.9%, peptone 1-100g/kg, Carnis Bovis seu Bubali cream 2-8g/kg, sucrose 1-50g/kg, yeast powder 1- 20g/kg, potassium iodate 1-3g/kg, sodium dihydrogen phosphate 0.5-2.5g/kg, magnesium sulfate 0.1-1g/kg, compound vitamin 0.01- 10g/kg, aminoacids complex 0.01-10g/kg.
Chicken compound symphysis unit microbial ecological agent and preparation method thereof that the present invention is provided, solves raising commercial chicken list Solely using the weakness of probiotic products, and be suitable for all types of commercial chickens, maximize favourable factors and minimize unfavourable ones, exogenous probiotic bacteria is promoted dynamic The field planting of thing intestinal, and the mushrooming out in the chicken body of the various probiotic bacterias of selective stimulating and breed, so as to reach promotion The purpose of chicken health, its production performance are also greatly improved, and reduce conventional animal edible additive to Carnis Gallus domesticus, egg quality Impact, it is more economical to be suitable for.It is comprehensive not enough to the mechanism of action degree of understanding of animal microecological formulation at present, lack deep layer Secondary mechanism, Mechanism Study, and the present invention more clearly elaborates the mechanism of action and courageously of the composite type microecology preparation Simulation animal intestinal environmental experiment is employed, is compared by the function and biological characteristicses of the beneficial bacteria of intestinal tract to filtering out, Compare the viable count change of probiotic bacteria under simulation intestinal environment, screening obtains optimal bacterial strain;Again all kinds of oligosaccharide contents are entered Measure is gone, has selected optimum addition, beneficial bacteria of intestinal tract and oligosaccharide are mixed and made into into chicken with compound symphysis unit Tiny ecosystem Preparation.The Substitutes For Antibiotic function of using at present is also relatively simple, and the composite type microecology preparation of present invention gained is abundant This shortcoming is overcome, by various probiotic bacterias and prebioticses are realized the micro- life of regulation animal intestinal with most suitable ratio mixing Thing is balanced, and maintains intestinal health, stimulates the growth of profitable strain, competitive inhibition harmful bacteria to be finally reached the life for promoting animal It is long, the immunity and production performance of chicken are improved, and the strain for obtaining can directly apply to fermentation and prepare microbial ecological agent, Pollution-free, noresidue.Prior art simply mixes probiotic bacteria and prebioticses simple and mechanical, not targetedly, and then must To the effect of composite type microecology preparation be not that very significantly, and present invention gained microbial ecological agent is by various probiotic bacterias and benefit Raw unit is obtained according to optimal ratio, has very much specific aim, and effect is also apparent from.
The compound symphysis unit microbial ecological agent of chicken of the present invention can adjust chicken intestinal microbial balance, maintain intestinal to be good for Health, stimulates the growth of profitable strain, competitive inhibition harmful bacteria to be finally reached the growth for promoting chicken, improve the immunity of chicken Purpose;The strain of acquisition can directly apply to fermentation and prepare microbial ecological agent, pollution-free, noresidue, be standard at this stage Green animal edible additive, meets health standards of the people to food, meets the food safety and prescription for increasingly improving; This breeder is played an important role in animal husbandry with compound symphysis unit microbial ecological agent, and chicken not only can be promoted to grow, also The conversion ratio of feedstuff can be improved, and meets the standard of Food Quality and Safety.
The present invention substitutes traditional chicken edible additive to find, for solving that livestock faces because antibiotic is indiscriminate With safety and the problem of quality of the Carnis Gallus domesticus for causing, the compound symphysis unit microbial ecological agent of a breeder is proposed;It is intended to make up Probiotic bacteria or prebioticses are added directly into into the deficiency in feedstuff and the impact to chicken meat quality, it is possible to increase Carnis Gallus domesticus produce matter Amount, and farming industry animal tradition edible additive is updated with facilitation.
The present invention relates to a kind of be beneficial to chicken growth, improve Carnis Gallus domesticus, the compound symphysis unit microbial ecological agent of eggs ' quality; Used as a kind of new, green animal edible additive, Main Function is the intestinal microbial population for optimizing chicken to symphysis unit microbial ecological agent, Chicken intestinal microbial balance is adjusted, its intestinal health is maintained, stimulates the growth of profitable strain using oligosaccharide probiotics, it is competitive Suppress harmful bacteria, be finally reached the growth for promoting chicken, improve resistance of the animal body to cause of disease material, adjust immunity of organism system System, so as to improve the immunity of animal, reaches the quality purpose for improving Carnis Gallus domesticus;Present invention simulation intestinal environment filters out probioticss Afterwards, suitable prebioticses, the such as oligosaccharide such as xylooligosaccharide, MOS, fruit oligose are selected according to all kinds of probiotic bacterias, synthesis is a kind of Compound symphysis unit microbial ecological agent, is that China's animal husbandry solves new animal edible additive to provide new selection, and solves Determined brought by traditional antibiotic drug residue, develop immunity to drugs, animal intestinal micro-ecology balance destroyed and dropped Food safety and the quality problems such as low meat productss quality.
Description of the drawings
Fig. 1 is preparation method flow chart of the chicken provided in an embodiment of the present invention with compound symphysis unit microbial ecological agent.
Specific embodiment
In order that the objects, technical solutions and advantages of the present invention become more apparent, with reference to embodiments, to the present invention It is further elaborated.It should be appreciated that specific embodiment described herein is not used to only to explain the present invention Limit the present invention.
Below in conjunction with the accompanying drawings the application principle of the present invention is explained in detail.
The chicken of the embodiment of the present invention is probiotic bacteria, prebioticses with compound symphysis unit microbial ecological agent;According to mass ratio institute State probiotic bacteria (lyophilized powder):Prebioticses=1:1~5 ratio mixing.
The probiotic bacteria is made up of lactobacilluss, bacillus bifiduss, bacillus cereuss and enterococcus;Probiotic bacteria is 0.02-15g/kg, Probiotic bacteria nutrient substance is 0.05-80g/kg, and oligosaccharide is 0.01-90g/kg;(lactobacilluss, bacillus bifiduss, bacillus cereuss and intestinal The reference culture of coccus is purchased from Beijing North Na Chuanlian Bioteknologisk Institut)
The prebioticses are made up of oligosaccharide and acidulant;
The oligosaccharide is MOS, fruit oligose, oligomeric isomaltose, newborn oligosaccharide etc., and acidulant is using by lactic acid, lemon The compound acidulant of the material compositions such as lemon acid, Fumaric acid.
The addition of all kinds of oligosaccharide be MOS 0.25-2g/kg, fruit oligose 0.01-3g/kg, oligomeric isomaltose 0.01-3g/kg, newborn oligosaccharide 0.01-3g/kg.Add by the compound acidulant of the material compositions such as lactic acid, citric acid, Fumaric acid, 0.1%-0.5% of the ratio for microbial ecological agent;
The chicken of the embodiment of the present invention with the preparation method of compound symphysis unit microbial ecological agent first against property separation, sieve Beneficial bacteria of intestinal tract is selected, by the viable count change for comparing probiotic bacteria under simulation intestinal environment, optimal bacterial strain is filtered out, will All kinds of oligosaccharide contents are measured, and select optimum addition, and the mixing of suitable chicken stomach, the probiotic bacteria of intestinal and oligosaccharide is made Into compound symphysis unit microbial ecological agent.
As shown in figure 1, chicken provided in an embodiment of the present invention is included with the preparation method of compound symphysis unit microbial ecological agent Following steps:
S101:The soil near gastrointestinal tract, chicken farm from chicken separates lactobacilluss, bacillus bifiduss, variable X bacterium Probiotic bacteria, lactobacilluss, bacillus bifiduss, bacillus cereuss and enterococcal probiotic bacteria morphological characteristic that analytical separation is obtained, and carry out The identification of strain;
S102:Isolated lactobacilluss, bacillus bifiduss, bacillus cereuss and enterococcal probiotic bacteria are carried out into biological characteristics The identification of property, the comparison of function and safety, and external probiotic effects evaluation is carried out, preliminary screening high-quality probiotic strain;
S103:The high-quality probiotic bacteria after preliminary screening is cultivated in the gastrointestinal fluid of manual simulation, all kinds of probiotic bacterias are determined Viable count, reproductive order of generation further screened, and selects optimal bacterial strain;
S104:Species, the functional characteristic of all kinds of prebioticses and the producing level with corresponding probiotic bacteria according to probiotic bacteria The prebioticses that selection is made up of oligosaccharide and acidulant, optimization oligosaccharide and acidulant optimum addition;
S105:The prebioticses selected with step S104 to the probiotic strain of step S103 screening, add probiotic bacteria battalion Foster material mixing makes compound symphysis unit microbial ecological agent.
Further, the identification that strain is carried out in step S101 includes:
General survey:Lactobacilluss are Gram-positive sporeless bacterium, cellular morphology in chain is shaft-like or club shape, bifid bar Bacterium is gram-positive sporeless bacterium, and varying length, bacillus bifiduss have V-shape or a Y-shaped, arrangement in single, double, short chain, X, Y, V or palisade;Bacillus cereuss breeding is quick, breeds 100,000 times within four hours, vitality by force without wet condition can low temperature resistant -60 DEG C, + 280 DEG C of high temperature resistant, resistance to strong acid, resistance to highly basic, antibiotic and sterilizing, resistance to hyperoxia (thermophilic oxygen breeding), lower oxygen concentration resistance (anaerobism breeding).Volume ratio The four times greater number of general nosophyte numerator, take up space advantage, suppresses the growth and breeding of harmful bacteria.Enterococcus are circular or oval Shape, the gram positive coccus in single paired or short catenation, without brood cell, atrichia, are aerobic or facultative anaerobic bacteria.This Bacterium is higher to nutritional requirement, it is desirable to high salt (6.5%NaCl), high-alkali (pH9.6), 40% bile culture-medium and 10-45 DEG C of environment Lower growth.
Lactobacilluss, bacillus bifiduss, bacillus cereuss and enterococcus probiotic bacteria and the prebioticses selected are combined, Adding is beneficial to the nutrient substance of growth of probiotics, makes compound symphysis unit microbial ecological agent.
Nutrient substance:As chicken belongs to birds, dilution normal saline used slightly has area with mammalss, Amphibian Not, General N aCl concentration is 0.9%, peptone 1-100g/kg, Carnis Bovis seu Bubali cream 2-8g/kg, sucrose 1-50g/kg, yeast powder 1- 20g/kg, potassium iodate 1-3g/kg, sodium dihydrogen phosphate 0.5-2.5g/kg, magnesium sulfate 0.1-1g/kg, compound vitamin 0.01- 10g/kg, aminoacids complex 0.01-10g/kg.
The application principle of the present invention is further described with reference to specific embodiment.
Embodiment 1:Soil preliminary screening enterococcus class probiotic bacteria from chicken plant.
The first step:Collection containing enterococcus pedotheque.The soil for selecting chicken cultivation ground in large-scale plant that raises is collection mesh Ground, randomly select 3-5 zonule, each region takes 20-40g initial soils, is divided into 2 and is assembled into sterilized valve bag In, take back rapidly laboratory and be placed in 4 DEG C of refrigerators and save backup;
Second step:Enterococcal separation.Aseptically, 2 groups of pedotheques are added separately in enrichment culture liquid, Bacterium colony is formed, then strain is inoculated into into bacteria suspension made by physiological saline solution respectively, then be diluted to not with 10 sequence diminishing methods Same Concentraton gradient, different solubility gradient sample plate streaks is seeded in culture medium, and each dilution factor does 3 and puts down OK, thus made by flat board be put into couveuse in cultivate 24-72h;
3rd step:It is enterococcal to isolate and purify.Aseptically, in 2 groups of picking, single purebred bacterial strain is identified, Probiotic bacteria inoculating loop after discriminating is inoculated in physiological saline solution, bacteria suspension is made, is inoculated on flat board, trained Observe after supporting, circulate operation, until obtaining pure bacterium colony, inclined plane inoculating is saved backup;
4th step:Enterococcal discriminating.Enterococcus after isolating and purifying are carried out into morphologic observation, physio-biochemical characteristics are determined With 16S rDNA sequencings etc. method, respectively different bacterium colonies is identified, the purpose for knowing that to which class distinguishes name is reached.
Embodiment 2
From preliminary screening bacillus subtilises in chicken feed grain.
The first step:The collection of sample in trough.It is sample collecting destination with selecting each chicken cultivation, randomly chooses several Place's trough, everywhere sampling 20-30g, is respectively charged in sterilized valve bag, takes back laboratory and is placed in guarantor in 4 DEG C of refrigerators Deposit standby;
Second step:The enrichment of bacillus subtilises, initial gross separation.Aseptically, 2 groups of trough samples are added respectively Enter in enrichment culture liquid, form bacterium colony, then strain is inoculated in physiological saline solution respectively makes bacteria suspension, then use 10 sequences Row diminishing method is diluted to different Concentraton gradient, and different solubility gradient sample plate streaks is seeded in culture medium, Each dilution factor do 3 it is parallel, thus made by flat board be put into couveuse in cultivate 24-72h;
3rd step:Bacillus subtilises isolate and purify.Aseptically, in 2 groups of picking, single purebred bacterial strain enters Row identification, the bacillus subtilises after discriminating is inoculated in physiological saline solution with inoculating loop, bacteria suspension is made, then is connect Plant on flat board, observe after culture, circulate operation, until obtaining the bacterium colony that purebred bacillus subtilises are formed, inclined plane inoculating is protected Deposit standby;
4th step:The discriminating of bacillus subtilises.Bacillus subtilises after isolating and purifying are carried out into morphologic observation, physiology Biochemical characteristic determine and 16S rDNA sequencings etc. method, respectively different bacterium colonies is identified, is reached and class is known to which Distinguish the purpose of name.
Example 3
Preliminary screening lactobacilluss, bacillus bifiduss probiotic bacteria from animal intestinal.
The first step:Collection containing probiotic bacteria sample.Select each large-scale slaughterhouse to be sample collecting destination, cut at random The stomach intestinal tissue of different healthy chickens, is divided into 2 and is assembled in sterilized valve bag, and quickly takes back laboratory and be placed in 4 DEG C of ice Save backup in case;
Second step:The enrichment of probiotic bacteria, screening.Aseptically, by 2 Zu Ji stomach intestinal tissues sample inner surface difference The enrichment culture in corresponding culture fluid, forms bacterium colony, then strain is inoculated in bacteria suspension made by addition physiological saline solution, Different Concentraton gradient are diluted to 10 sequence diminishing methods again, different solubility gradient sample plate streaks is seeded to into phase Answer in culture medium, each dilution factor do 3 it is parallel, thus made by flat board be put into couveuse in cultivate 24-72h;
3rd step:Probiotic bacteria is isolated and purified.Aseptically, in 2 groups of picking, single purebred bacterial strain is identified, Probiotic bacteria inoculating loop after discriminating is inoculated in physiological saline solution, bacteria suspension is made, then is connect with plate streak Plant on flat board, observe after culture, circulate operation, until obtaining pure bacterium colony, inclined plane inoculating is saved backup;
4th step:The discriminating of probiotic bacteria.Lactobacilluss after isolating and purifying are carried out into morphologic observation, physio-biochemical characteristics are determined With 16S rDNA sequencings etc. method, respectively different bacterium colonies is identified, the purpose for knowing that to which class distinguishes name is reached.
Embodiment 4
The further screening of probiotics strain.
The first step:The probiotic bacteria of preliminary screening is carried out into function test measure respectively, for example, the probiotic bacteria of new culture is connect Plant in the culture fluid of different temperatures, determine the tolerance to temperature;Probiotic bacteria is cultivated 3-5 days in the environment of different pH, so The degree of probiotic bacteria acid and alkali-resistance change is judged afterwards;
Second step:The probiotic bacteria after preliminary screening is cultivated in manual simulation chicken gastrointestinal microecological environment, to all kinds of The survival rate of probiotic bacteria, reproductive order of generation are measured, and filter out the preferable probiotic bacteria of upgrowth situation;
3rd step:The above is every to determine the probiotic bacteria that qualified probiotic bacteria is final screening, finally give 1 plant of enterococcus, 1 Bacillus subtilis, 2 plants of lactobacilluss and 2 plants of bacillus bifiduss;.
Embodiment 5
The preparation of symphysis unit microbial ecological agent.
The first step:Species, the functional characteristic of all kinds of prebioticses according to probiotic bacteria and the utilization journey with corresponding probiotic bacteria Degree selects the prebioticses that are made up of oligosaccharide, and selectable oligosaccharide is xylooligosaccharide, MOS, fruit oligose, oligomeric Isomalt Sugar, newborn oligosaccharide,.
Second step:The addition of all kinds of oligosaccharide is tested, the addition of all kinds of oligosaccharide is MOS 0.25-2g/kg, Fruit oligose 0.01-3g/kg, oligomeric isomaltose 0.01-3g/kg, newborn oligosaccharide 0.01-3g/kg.
3rd step:4 kinds of probiotic bacterias and the prebioticses selected are combined, add be beneficial to be conducive to it is prebiotic The nutrient substance of bacteria growing, makes compound symphysis unit microbial ecological agent;
4th step:The product being eventually fabricated is tested in being added to chicken food materials, the gain in weight, feedstuff according to experimental chicken Consumption and feedstuff-meat ratio etc. carry out the production performance situation of judgment experiment chicken.
The application principle of the present invention is further described with reference to test.
Step one, the soil from chicken farm, feed the trough of chicken, gastrointestinal region and separate lactobacilluss, double Isolated probiotic bacteria is carried out biological characteristicses, function and safety by discrimination bacillus, bacillus cereuss and enterococcus probiotic bacteria Identification, and carry out external probiotic effects evaluation, preliminary screening probiotic bacteria;
Step 2, using chicken intestinal microorganism and carbohydrate metabolism, the relation of vitamin, filter out raising chicken obtain from daily ration The high-quality probiotic bacteria of energy, the acquisition and storage that improve energy, raising chicken using high-quality probiotic bacteria in chicken intestinal is from daily ration The middle ability for obtaining energy, further according to microbial technique index (prior art:1. probiotic bacteria is separated on selective medium, Then carry out the physiological property detection of acidproof, bile tolerance, digestion resistant enzyme etc. to every plant of selected antibacterial one by one, finally carry out prebiotic Bacterium other characteristic tests.2. a small amount of content is taken in animal alimentary canal first, and be diluted, under anaerobic state, trained in MRS Enrichment culture is carried out in foster base, then obtain can gradually increase culture medium by gradient culture again as the antibacterial of prebiotic strain Acidity (pH be 3.0 or so) and bile salt concentration, select the bacterial strain with acidproof and resistance to bile characteristic, further according to probiotic bacteria spy Property requirement, add other inhibitor or pathogen in the medium, filter out and grow fine and the bacterium for killing pathogen can be pressed down Strain, finally carries out the forms such as Gram’s staining, catalase and biochemical identification again, and determines its kind to the bacterial strain.) screening high-quality chicken Beneficial bacteria of intestinal tract;3. screening and determining carries out biological characteristicses, the comparison of function and safety by isolated probiotic bacteria Identification, and carry out external probiotic effects evaluation, preliminary screening high-quality probiotic bacteria;1) lactobacilluss are (to screen from animal gastrointestinal tract As a example by):Take a certain amount of stirring in 50mL anaerobism diluents of gastroduodenal large intestines jejunum cecal content respectively to make 10 times are serially diluted, and are taken each 0.1mL of 3 dilution factors respectively and are coated with LBS flat boards, 37-38 DEG C, anaerobic jar culture.With inoculating loop from On flat board, picking all kinds bacterium colony carries out gram stain microscopy, and the bacterium colony with typical lactobacilluss morphological characteristic is entered to advance One screening.The diameters observed of 48-72h are cultivated on LBS flat boards in 0.5mm-2mm, the smooth moistening side of white circular surface The neat raised bacterium colony of edge is lactobacilluss.2) bacillus bifiduss (as a example by screening from animal gastrointestinal tract):Stomach 12 is taken respectively Duodenum 12 large intestines jejunum cecal content is a certain amount of to be dissolved in 100mL modified MRS culture mediums, and enrichment culture 24h takes 1mL gradients dilute Release, coat on flat board, cultivate 48h under anaerobic, purified 3 times with inoculating loop picking blue colonies, obtain bacillus bifiduss, The bacterial strain of picking strictly anaerobic carries out whether Gram’s staining, microscopy observation thalline color size shape etc. meet bacillus bifiduss The basic feature of category.3) bacillus cereuss (as a example by screening from animal gastrointestinal tract) take gastroduodenal large intestines Blind jejunum respectively The a certain amount of stirring in 50mL anaerobism diluents of intestinal contentses is made 10 times and is serially diluted, and takes each 0.1mL of 3 dilution factors respectively and applies Cloth LBS flat boards, 37 DEG C of culture 48h-72h, are suspected to be bacterium colony with inoculating loop picking, first carry out azaleine and simply dye observation, then carry out Gram’s staining and spore staining, picking Bacillus colonies carry out pure culture, and preserve in inclined-plane, then carry out catalase Determine, will each separation strains be inoculated in LB culture medium slants, 37 DEG C culture 18h-24h, take a clean slide, upper Drip 10% H in face2O2, the cultured lawn of one ring of picking, in H2O2In smear, if there is bubble, for the positive, otherwise be It is negative.4) enterococcus (by taking the soil near plant as an example):
1st, the acquisition process of soil
Sample in soil using five point sampling near plant.As the microorganism in soil is typically in 5-25cm Soil layer in, so first rooting out the epidermis of soil with sterilizing, then take soil 200-300g, as in aseptic plastic bag, And indicate and take place, date.Preserve as in refrigerator after being transported back laboratory, it is standby.
2nd, in soil strain separation
The sample adopted back is obtained into sample 20-30g with quartering, grinding is uniform in being placed on mortar.1g soil is weighed, It is placed in fluid medium sterilizing Sucus Vitis viniferae, 28 DEG C of Jing, 12h, 180rpm shaking table enrichment culture.1mL enrichment culture liquid is taken, is entered respectively Five gradient dilutions (1 of row:1000、1:5000、1:10000、1:50000、1:100000), then with liquid-transfering gun respectively from each dilute Degree of releasing respectively takes 0.1mL diluents, and on injection flat board, sample is uniformly coated in media surface using spreading rod, and (attention can not Rupture media surface), culture dish is covered, then culture dish is inverted, in preference temperature (generally 25 DEG C or 28 DEG C) Cultivate in calorstat, each gradient does three repetitions.Observation culture medium, till strain forms the bacterium colony clearly easily debated.For The interference of antibacterial is excluded in screening process, in the medium plus 30ug/mL streptomycin or 100mg/L chloromycetin. Colony colour and form are observed and record, and then cell size, form are recorded in microexamination, are analyzed and return with preliminary Class.
Step 3, the Intestinum Gallus domesticus in the chicken gastrointestinal fluid of manual simulation after culture preliminary screening are determined all kinds of to probiotic bacteria The viable count of probiotic bacteria, reproductive order of generation are further screened, and screen optimal bacterial strain;1. identify:Lactobacilluss are blue for leather Family name positive sporeless bacterium, cellular morphology is typically shaft-like in chain or club shape.Bacillus bifiduss are gram-positive without spore bar Bacterium, varying length, typical bacillus bifiduss have V-shape or Y-shaped, and arrangement is in single, double, short chain, X, Y, V or palisade.Spore bar There are two kinds of forms in bacterium:Spore and trophosome.The cell grown form of trophosome has shaft-like and ellipticity.There is stock in shaft-like Shape, rod-short, ellipticity have the oval and short ellipse of length.Enterococcus are single celled eukaryotic microorganisms, and the form of enterococcus cell is led to Often there are spherical, oval, sausage shape, ellipse, Fructus Citri Limoniae shape or Nodus Nelumbinis Rhizomatis shape etc..It is more much greater than the unicellular individuality of antibacterial, one As be 1-5 microns × 5-30 microns, enterococcus atrichia, it is impossible to travelling.2. compare:It is special according to the form of the different probiotic bacterias of comparison Levy, so as to identify each probiotic bacteria.
Step 4:Species, the functional characteristic of all kinds of prebioticses according to probiotic bacteria and the utilization journey with corresponding probiotic bacteria Degree is selected by all kinds of oligosaccharide and the prebioticses for constituting, and selects all kinds of oligosaccharide optimum additions;
Step 5, the prebioticses that probiotics and step 4 are selected are screened to step 3, chicken is mixed and made into compound Symphysis unit microbial ecological agent.Add by lactic acid, citric acid, Fumaric acid material composition compound acidulant 0.1%-0.5%. Under optimal acidulant addition:1. the digestive tract environment of animal can be improved, so as to improve health status and the growth of animal Performance.2. gastrointestinal acidity is improved, so as to improve pepsic activity.3. improve microbiota in gastrointestinal tract, suppress Harmful microbe breeding 4. promotes absorption 5. of the animal body to mineral element and vitamin to promote the propagation of probioticss, can Substitute the function of some antibiotic)
Further, in the optimal bacterial strain of step 3 screening preferably:1 plant of enterococcus, 1 bacillus subtilis, 2 plants of lactic acid bars Bacterium and 2 plants of bacillus bifiduss.1. chicken feed conversion ratio is improved, promotes meat, laying hen growth;2. nutrient substance is produced, it is possible to adjusted The activity of enzyme in animal body;3. strengthen commercial chicken immunologic function 4. improve chicken survival rate 5. improve broiler daily gain and Laying hen egg laying performance, including egg color and egg size etc. 6, strengthen the non-specific immunity of animal.
Further, in step 4, all kinds of oligosaccharide are:Xylooligosaccharide, MOS, fruit oligose, oligomeric isomaltose, breast are few Sugar;
The preferred xylooligosaccharide 0.05-5g/kg of all kinds of oligosaccharide optimum additions, MOS 0.25-2g/kg, fruit oligose 0.02-2g/kg, oligomeric isomaltose 0.5-3g/kg, newborn oligosaccharide 0.5-3g/kg.1. body can be promoted to generate various nutrients Matter, including vitamin B1, B2, B6, B12, nicotinic acid and Folic Acid;2. bifidus bacilluss, lactobacilluss, bacillus acidophilus etc. in pair intestinal The selective proliferation function of difference, makes profitable strain take advantage in intestinal, suppresses the growth of harmful bacteria, reduces noxious substance The formation of (such as endotoxin, Ammonia etc.), and there is protective effect to intestinal mucosa cells regulating liver-QI, so as to prevent sending out for pathological changes intestinal cancer It is raw, enhancing human body immunity power;3. the inhibitory action of pair harmful bacteria, especially has obvious inhibitory action to clostridium, such as low IMO not only has inhibitory action to clostridium, also significantly inhibits effect to escherichia coli.
Presently preferred embodiments of the present invention is the foregoing is only, not to limit the present invention, all essences in the present invention Any modification, equivalent and improvement made within god and principle etc., should be included within the scope of the present invention.

Claims (7)

1. a breeder is with compound symphysis unit microbial ecological agent, it is characterised in that the chicken is with compound symphysis unit Tiny ecosystem system Agent is made up of probiotic bacteria and prebioticses;
According to probiotic bacteria described in mass ratio:Prebioticses=1:1~5 ratio mixing.
2. chicken as claimed in claim 1 is with compound symphysis unit microbial ecological agent, it is characterised in that the probiotic bacteria is by newborn bar Bacterium, bacillus bifiduss, bacillus cereuss and enterococcus composition.
3. chicken as claimed in claim 1 is with compound symphysis unit microbial ecological agent, it is characterised in that the prebioticses are by oligomeric Sugar and acidulant are constituted;
The oligosaccharide is MOS, fruit oligose, oligomeric isomaltose, newborn oligosaccharide, and acidulant is by lactic acid, citric acid, Rhizoma Corydalis The compound acidulant of rope acid composition.
4. the chicken as described in claim 1 or 3 is with compound symphysis unit microbial ecological agent, it is characterised in that the probiotic bacteria is 0.02-15g/kg, probiotic bacteria nutrient substance are 0.05-80g/kg, and oligosaccharide is 0.01-90g/kg.
5. chicken as claimed in claim 3 is with compound symphysis unit microbial ecological agent, it is characterised in that the addition of oligosaccharide is MOS 0.25-2g/kg, fruit oligose 0.01-3g/kg, oligomeric isomaltose 0.01-3g/kg, newborn oligosaccharide 0.01-3g/kg; The compound acidulant that addition is made up of lactic acid, citric acid, Fumaric acid, compound acidulant account for the 0.1%- of microbial ecological agent ratio 0.5%.
6. preparation method of a kind of chicken as claimed in claim 1 with compound symphysis unit microbial ecological agent, it is characterised in that described Preparation method is comprised the following steps:
Step one, the soil near gastrointestinal tract, chicken farm from chicken separate lactobacilluss, bacillus bifiduss, the benefit of variable X bacterium Raw bacterium, lactobacilluss, bacillus bifiduss, bacillus cereuss and enterococcal probiotic bacteria morphological characteristic that analytical separation is obtained, and carry out bacterium The identification planted;
Isolated lactobacilluss, bacillus bifiduss, bacillus cereuss and enterococcal probiotic bacteria are carried out biological characteristics by step 2 The identification of property, the comparison of function and safety, and external probiotic effects evaluation is carried out, preliminary screening high-quality probiotic strain;
Step 3, cultivates the high-quality probiotic bacteria after preliminary screening in the gastrointestinal fluid of manual simulation, determines all kinds of probiotic bacterias Viable count, reproductive order of generation are further screened, and select optimal bacterial strain;
Step 4, the species, the functional characteristic of all kinds of prebioticses according to probiotic bacteria and the producing level choosing with corresponding probiotic bacteria Select the prebioticses being made up of oligosaccharide and acidulant, optimization oligosaccharide and acidulant optimum addition;
Step 5, the prebioticses selected with step 4 to the probiotic strain of step 3 screening, adds probiotic bacteria and nutrient Matter is mixed and made into compound symphysis unit microbial ecological agent.
7. preparation method of the chicken as claimed in claim 6 with compound symphysis unit microbial ecological agent, it is characterised in that nutrient Matter includes:NaCl concentration is 0.9%, peptone 1-100g/kg, Carnis Bovis seu Bubali cream 2-8g/kg, sucrose 1-50g/kg, yeast powder 1- 20g/kg, potassium iodate 1-3g/kg, sodium dihydrogen phosphate 0.5-2.5g/kg, magnesium sulfate 0.1-1g/kg, compound vitamin 0.01- 10g/kg, aminoacids complex 0.01-10g/kg.
CN201610990924.XA 2016-11-10 2016-11-10 Composite type synbiotic microecological preparation for chickens and preparation method of composite type synbiotic microecological preparation Pending CN106509468A (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN201610990924.XA CN106509468A (en) 2016-11-10 2016-11-10 Composite type synbiotic microecological preparation for chickens and preparation method of composite type synbiotic microecological preparation

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN201610990924.XA CN106509468A (en) 2016-11-10 2016-11-10 Composite type synbiotic microecological preparation for chickens and preparation method of composite type synbiotic microecological preparation

Publications (1)

Publication Number Publication Date
CN106509468A true CN106509468A (en) 2017-03-22

Family

ID=58350935

Family Applications (1)

Application Number Title Priority Date Filing Date
CN201610990924.XA Pending CN106509468A (en) 2016-11-10 2016-11-10 Composite type synbiotic microecological preparation for chickens and preparation method of composite type synbiotic microecological preparation

Country Status (1)

Country Link
CN (1) CN106509468A (en)

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN108391758A (en) * 2018-04-25 2018-08-14 广西康佳龙农牧集团有限公司 Milking sow compound Synbiotics
CN113261629A (en) * 2021-06-28 2021-08-17 深圳绿养时代新技术有限公司 Feed additive for improving DHA content stability in eggs and application thereof

Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN103315139A (en) * 2013-07-01 2013-09-25 乌鲁木齐思科康达科技有限责任公司 Compound microbial ecological agent for monogastric animals and preparation method thereof
CN103750027A (en) * 2013-12-30 2014-04-30 何贻峰 Chicken feed
CN104206828A (en) * 2014-08-12 2014-12-17 成都巨星农牧科技有限公司 Fermentation method for preparation of pig feed
CN104758318A (en) * 2015-03-06 2015-07-08 山东宝来利来生物工程股份有限公司 A composite preparation for reducing a diarrhea rate of weaned piglets and a preparing method thereof
CN105296550A (en) * 2015-11-19 2016-02-03 润盈生物工程(上海)有限公司 Production technology for high-density fermentation of lactics and separation coupling of lactic acid by physical method

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN103315139A (en) * 2013-07-01 2013-09-25 乌鲁木齐思科康达科技有限责任公司 Compound microbial ecological agent for monogastric animals and preparation method thereof
CN103750027A (en) * 2013-12-30 2014-04-30 何贻峰 Chicken feed
CN104206828A (en) * 2014-08-12 2014-12-17 成都巨星农牧科技有限公司 Fermentation method for preparation of pig feed
CN104758318A (en) * 2015-03-06 2015-07-08 山东宝来利来生物工程股份有限公司 A composite preparation for reducing a diarrhea rate of weaned piglets and a preparing method thereof
CN105296550A (en) * 2015-11-19 2016-02-03 润盈生物工程(上海)有限公司 Production technology for high-density fermentation of lactics and separation coupling of lactic acid by physical method

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN108391758A (en) * 2018-04-25 2018-08-14 广西康佳龙农牧集团有限公司 Milking sow compound Synbiotics
CN113261629A (en) * 2021-06-28 2021-08-17 深圳绿养时代新技术有限公司 Feed additive for improving DHA content stability in eggs and application thereof

Similar Documents

Publication Publication Date Title
CN103614327B (en) A kind of subtilis and uses thereof
CN102409015B (en) Composite micro-ecological preparation as well as premixed material and application of preparation in feed additive
CN102093967B (en) Mink source enterococcus faecium and application thereof
CN106566791A (en) Porcine compound synbiotics micro-ecological preparation and preparation method thereof
CN106260540B (en) Biological feed for creep feed and creep feed
CN104293696B (en) One strain enterococcus faecalis HEW-A131 and application thereof
CN101638627B (en) Bacillus subtilis and application thereof in biological feed additives
CN103275907B (en) Bacillus amyloliquefacien and preparation method and application thereof
CN103981118B (en) A kind of bacillus subtilis feed addictive and its preparation method and application
CN101463329B (en) Saccharomyces cerevisiae, yeast preparation including the same, feed, and preparation and use thereof
CN102599348B (en) Lactic acid bacteria semi-solid fermentation product for increasing growth performance of animals and process
CN101671638B (en) New strain of bifidobacterium and fermentative preparation method and application thereof
CN106720938A (en) A kind of compound symphysis unit probiotics and preparation method thereof
CN103045498B (en) Bacillus amyloliquefaciens and application thereof
CN102517238B (en) Acid-producing bacillus cereus and application thereof
CN105420150A (en) Lactobacillus acidophilus and application thereof
CN106509455A (en) Composite type synbiotic microecological preparation for rabbits and preparation method of composite type synbiotic microecological preparation
CN105199978A (en) Bacillus coagulans preparation for livestock breeding and preparation method thereof
CN109874920A (en) A kind of composite microbial feed additive and preparation method thereof
CN102907585A (en) Probiotics and vitamin premix and mixture for middle pig
CN102517227A (en) Enterococcus faecalis and applications and feed additive and leavening agent thereof
CN102329749A (en) Bacillus subtilis bred by space mutation breeding technology and application of Bacillus subtilis
CN106509468A (en) Composite type synbiotic microecological preparation for chickens and preparation method of composite type synbiotic microecological preparation
CN103266074B (en) B.subtilis spores strain and application thereof
CN102787083B (en) Bacillus subtilis and application

Legal Events

Date Code Title Description
C06 Publication
PB01 Publication
SE01 Entry into force of request for substantive examination
SE01 Entry into force of request for substantive examination
RJ01 Rejection of invention patent application after publication
RJ01 Rejection of invention patent application after publication

Application publication date: 20170322