CN1703968A - Synbiotic feed addictive of mini-peptide with beneficial microbe and its usage - Google Patents

Abstract

The invention relates to a synbiotic feed addictive of mini-peptide with beneficial microbe. which comprises small peptide dried powder and bacillus dried powder, or saccharomycete dried powder or/and lactic acid bacteria dried powder. The small peptide dried powder is prepared through the microorganism fermentation of plant protein or/and animal protein, the prepared feed addictive can be used as the substitute of antibiotics.

Description

Little peptide and beneficial microbe symphysis unit feed addictive and uses thereof

Invention field

The present invention relates to a kind of symphysis unit that is formed by beneficial microbe and little peptide or Synbiotics (Synbiotics) feed addictive and uses thereof, specifically relate to a kind of little peptide (Small Peptide that utilizes animal/vegetable protein to make, SP) and comprise symphysis unit that bacillus and/or the beneficial microbes such as saccharomycete or/and lactic acid bacteria (Benefit Microorganism, BM) form or Synbiotic feed additive and uses thereof.

Background technology

Since the mankind find antibiotic, in animal feeding and feed, extensively and for a long time use the growth promotion health-care agents such as antibiotic, chemical synthetic drug, hormone, beta-stimulants, sedative, caused following problem: livestock and poultry and aquatic products product drug residue are serious, directly are detrimental to health; The animal products quality descends, and lacks the market competitiveness and trust; Bacterial drug resistance strengthens, antibody-resistant bacterium increases, and threatens human security; Destroy ecological environment etc. Since food security in the urgent need to, every country all in accelerate development, promote the efficient feed addictive of new type of safe, with additives such as substitute antibiotics. Achievement in research in recent years shows, microecologic regulator plays an important role aspect improving in growth, growth, product formation and the product quality of animal.

Microecologic regulator comprises probio, prebiotics and symphysis unit three classes, the preparation that refers to utilize some materials of the normal useful microbial bacteria group members of animal body and metabolite and the endogenous probio growing multiplication of stimulating animal to make, the recovery of its flora that can promote to lack of proper care, keep microecological balance, improve host health level or the state that improves health. Wherein, probio or probiotic (Probiotics) refer to contain viable bacteria and (or) dead bacterium, the bacterial preparation that comprises its component and product, per os or drop into via other mucosal route, be intended to improve the balance of mucomembranous surface microorganism or enzyme, perhaps stimulate specificity or nonspecific immunity mechanism. 1989, food and drug administration (FDA) and U.S. feed supervision association (AAFCO) announced 41 kinds of safe bacterial classifications that can be used for feed. The probio bacterial classification that can be used for animal that China Ministry of Agriculture announced in 1994 has: lactic acid bacteria and bacillus, the saccharomycete etc. such as lactic acid bacteria, streptococcus fecalis, Bifidobacterium. Prebiotics (Prebiotics) refers at GI top (leading portion) that not by the COF of Host Digestion or preparation it can optionally stimulate activity or the growth and breeding of resident bacterium in one or more colons, promotes the effect of host health. It comprises oligosaccharides (such as FOS, xylo-oligosaccharide etc.), natural plant kind (comprising Chinese herbal medicine, vegetables and wild plant etc.), small peptide etc. Symphysis unit or Synbiotics (Synbiotics) are that probio and prebiotics merge the preparation that uses, and both can bring into play the physiological bacterial activity of probio, and the quantity that optionally increases again this bacterium makes prebiotic effect more lasting. Since the seventies, progressively microecologic regulator is applied to livestock and poultry and aquaculture abroad, generally be applied to the aspects such as health care, disease prevention and cure, feed addictive and water conditioner. The middle and later periods eighties 20th century, China begins little ecological goods are used for feed addictive.

Use antibiotic main purpose in a large number in livestock and poultry cultivation or in the feed, the one, in order to prevent livestock and poultry, the 2nd, for growth promotion or improve the production efficiency (being the products such as voluminous meat, egg, milk) of livestock and poultry. In order to eliminate medicament residue, overcome problems, in feed processing and livestock and poultry breeding industry, usually use at present little living conditioning agent, such as the used lactic acid bacteria of microbe additive, saccharomycete or fungi or bacillus, or the oligosaccharide of prebiotics class (such as FOS, xylo-oligosaccharide etc.), but except bacillus, other bacterial strain all is vulnerable to temperature, pressure, meta-acid, meta-alkali, metal ion (such as Cu in the used bacterial strain of microbe additive²⁺、Fe ²⁺、Mn ²⁺、Zn ²⁺) or feed in the destruction of the factor such as choline and inactivation, and the action effect of oligomerization carbohydrate additive is fainter, effectively substitute antibiotics has had a strong impact on its practical value.

Summary of the invention

The object of the invention is to: (1) overcomes in the prior art and to use micro-metal ion that microbe additive is subject to containing in the feed easily (such as Cu merely²⁺、Fe ²⁺、Mn ²⁺、Zn ²⁺), the damage of the multiple adverse factor such as Choline Chloride, antibiotic, temperature, pressure, meta-acid, meta-alkali, can not tolerate the defective of the high temperature in the feed granulating process; (2) overcome single use microbe additive and only can replenish or microbe inoculation, can not transfer and bring into play the effect of endogenous beneficial microorganism; (3) overcome and use prebiotics small peptide additive effect single, only can activate livestock and poultry endogenous profitable strain, have the beneficial microbe that produces enzymatic digestion, immune stimulatory and disease-proof functions and can not replenish. Thereby, a kind of little peptide and beneficial microbe have complementary advantages (effect of performance internal and external reasons), active height, anti-storage, stability is high, cost is low little peptide and beneficial microbe symphysis unit or Synbiotic feed additive are provided.

Another object of the present invention is to provide the purposes of described little peptide and beneficial microbe symphysis unit (element) feed addictive.

The objective of the invention is to be achieved through the following technical solutions:

Little peptide provided by the invention and beneficial microbe symphysis unit feed addictive comprise little peptide dry powder A and bacillus dry powder B; Also can comprise saccharomycete dry powder C or/and dry powder of lactic acid bacteria D. Its quality standard is: little peptide dry powder A 100mg/g, bacillus living 〉=6,000,000,000/g, saccharomycete viable bacteria 〉=400,000,000/g, viable lactic acid bacteria 〉=400,000,000/g, moisture content≤9%, ash≤8%, arsenic≤0.5mg/kg, lead≤1.0mg/kg.

Little peptide and the feed addictive S-1 of beneficial microbe symphysis unit type are comprised of little peptide dry powder A and bacillus dry powder B, and wherein, little peptide dry powder accounts for 40～60wt%; Bacillus dry powder accounts for 40～60wt%.

Little peptide and the feed addictive S-2 of beneficial microbe symphysis unit type are comprised of little peptide dry powder A, bacillus dry powder B and saccharomycete dry powder C, and wherein, little peptide dry powder accounts for 35～45wt%; Bacillus dry powder accounts for 20～30wt%; Saccharomycete dry powder accounts for 30～40wt%.

Little peptide and the feed addictive S-3 of beneficial microbe symphysis unit type are comprised of little peptide dry powder A, bacillus dry powder B and dry powder of lactic acid bacteria D, and wherein, little peptide dry powder accounts for 30～50wt%; Bacillus dry powder accounts for 20～40wt%; Dry powder of lactic acid bacteria accounts for 30～50wt%.

Little peptide and the feed addictive S-4 of beneficial microbe symphysis unit type are comprised of little peptide dry powder A, bacillus dry powder B, saccharomycete dry powder C and dry powder of lactic acid bacteria D, and wherein, little peptide dry powder accounts for 30～40wt%; Bacillus dry powder accounts for 20～30wt%; Saccharomycete dry powder accounts for 10～20wt%; Dry powder of lactic acid bacteria accounts for 20～30wt%.

Described little peptide dry powder A is with vegetable protein or/and animal protein is degraded through microbial fermentation, and its degree of hydrolysis (DH %) is controlled at and obtains after 25～40% the zymotic fluid drying, and concrete preparation method comprises the steps:

1) fermentation seed liquid preparation: will be selected from one or more the mixture in aspergillus niger, aspergillus oryzae, aspergillus terricola bacterium, bacillus subtilis, bafillus natto, the bacillus pumilus, access in 121 ℃ of fluid nutrient mediums of sterilizing 30 minutes and being cooled to after 28～37 ℃, cultivated 24～48 hours, and obtained seed liquor for 28～37 ℃; Described fluid nutrient medium is the mixed liquor in following ratio preparation: peptone 1.3～2.5g, glucose 3～5g, molasses 3～5g, potato immersion liquid 50～100ml, soluble starch 10～20g, sucrose 10～30g, beef extract 2～3g, yeast soak powder 3.5～5g, dregs of beans 1.5～2g, growth factor 1～2ml, water 1000ml; Growth factor wherein is for containing MgSO₄.7H ₂O 20.00～30.00mg/ml， CaCl ₂.2H ₂O 2.20～3.50mg/ml，ZnSO ₄.7H ₂O 0.81～1.10mg/ml，FeSO ₄.7H ₂O 0.44～0.60mg/ml， MnSO ₄.H ₂O 0.13～0.20mg/ml，CuSO ₄.5H ₂The mixture of O 0.02～0.04mg/ml;

2) fermentative degradation: one or more the mixture in vegetable protein or the animal protein is crushed to 60～120 orders, pressing protein content calculates, according to material: flowing fluid ratio=5～18: 100 ratio is put into fermentation tank, put into carbon source according to the ratio of C/N ratio=1/2～1/35 again, and adding defoamer 0.03～0.1%, mixing, the nutrient solution that obtains was in 121～125 ℃ of autoclavings 10～30 minutes, be cooled to 28～37 ℃, take the volume ratio of seed liquor and nutrient solution as 0.3～1: the seed liquor that 10 ratio access step 1) obtains, 28～42 ℃ of fermentations, throughput be 0.5～2.5vols/vol/min (hereinafter to be referred as: vvm), mixing speed 150～300rpm, use during this time conventional ninhydrin method determination of color degree of hydrolysis (Degree of Hydrolysis, DH%), degree of hydrolysis of detection in per 6 hours before 24 hours, between 24～48 hours, detected a degree of hydrolysis in per 4 hours, degree of hydrolysis of detection in per 2 hours after 48 hours, until degree of hydrolysis reaches 25～40%, then in 140～145 ℃ of sterilizations 2～8 seconds, obtain containing the protein hydrolyzate of little peptide;

3) protein hydrolyzate that contains little peptide that centrifugal slagging-off: with step 2) obtains centrifugal 10～30 minutes in 5000～7000rpm, remove slag, the supernatant that obtains is that the ultrafilter of 1000Da is removed the impurity such as high molecular weight protein by retaining molecular weight again, obtains the filtrate of molecular weight≤1000Da; This filtrate is concentrated to 50% of original volume through negative-pressure vacuum, the concentrate that obtains is added in the carrier that is pre-mixed, and presses concentrate: carrier=0.5～0.8: 1 envelope-bulk to weight ratio mixes, and it is rear 45～80 ℃ of dryings to stir, then be crushed to more than 80 orders, obtain little peptide dry powder A.

Described step 2) vegetable protein comprises soybean protein (dregs of beans, soya-bean cake), soybean protein, dregs of beans, soya-bean cake, Peas albumen, zein, red bean albumen, wheat gluten, large aleuronat, Cottonseed Meal or rapeseed dregs.

Described step 2) animal protein comprises fish meal, blood meal, feather meal, lactoprotein, muscles, casein, the discarded internal organs of animal, earthworm protein powder, squid albumen, fish-skin or other aquatic livestock albumen.

Described step 2) carbon source is for being selected from one or more the mixture in corn flour, corn syrup (honey), starch, cane molasses, beet molasses, the table sugar (white sugar or brown sugar).

Described step 3) carrier is comprised of 2～5 weight portion organic carriers and 3～5 weight portion inorganic carriers; Wherein, described organic carrier comprises bran and puffed soybean organic carrier; Described bran organic carrier is one or more the mixture in wheat bran, maize cob meal, degreasing powdered rice hulls and the defatted rice bran; Described puffed soybean organic carrier is one or more the mixture in bean cake powder, beancake powder and the corn protein powder; Described inorganic carrier is one or more the mixture in precipitated calcium carbonate, stone flour, medical stone and the zeolite powder.

Described bacillus dry powder B is made by following method:

1) dull and stereotyped cultivation rejuvenation: with method streak inoculation on the bacillus plating medium of bacillus bacterial classification employing streak inoculation, in 25～35 ℃ of cultivations 12～24 hours, make the bacillus rejuvenation, and form single bacterium colony; Described bacillus plating medium is the mixture by following w/v (W/V) preparation: peptone 0.5～1%, glucose 0.5～2%, beef extract 0.5～1.5%, potassium dihydrogen phosphate 0.3～0.6%, agar 3～4%, pH=7.5～7.6, sterilized 30 minutes for 121 ℃, for subsequent use; Described bacillus comprises: bacillus coagulans, slow bacillus, bacillus subtilis, bacillus licheniformis, bacillus pumilus, Bacillus circulans, bacillus firmus, bacillus megaterium, clostridium butyricum, bafillus natto, wax-like (sample) bacillus;

2) choose single bacterium colony the bacillus bacterium colony of making shake-flask seed: from step 1) cultivating, be inoculated in the bacillus liquid culture medium, in 28～37 ℃ of cultivations 20～30 hours, obtain the bacillus shake-flask seed liquid; Described bacillus liquid culture medium is the mixed liquor by following w/v (W/V) preparation: peptone 0.5～1%, glucose 0.5～2%, beef extract 0.5～1.5%, potassium dihydrogen phosphate 0.3～0.6%, 3.08% manganese sulfate solution 0.5～1.5%, pH=7.5～7.6, sterilized 30 minutes for 121 ℃, for subsequent use;

3) the bacillus shake-flask seed liquid that production primary seed solution: with step 2) obtains is with the inoculum concentration of 2～5v%, be inoculated in the first order seed liquid culture medium of producing used first class seed pot, cultivate 18～24h in 28～37 ℃, mixing speed 150～200rpm, throughput is 1～2vvm, obtains the primary seed solution of bacillus; Described first order seed liquid culture medium prescription is the mixed liquor by following w/v (W/V) preparation: 120 order bean cake powders 0.5～2.0%, 120 order fish meal 0.2～1.5%, 120 order corn flours 0.5～1.0%, glucose 1.0～4.5%, NaCl 0.1～0.3%, NaOH 0.05～0.10%, Na₂HPO ₄0.1～0.5%, synergist 0.05～0.2%, defoamer 0.05～0.1%, and pH=7.5～7.6 through 121 ℃ of sterilizations 30～45 minutes, are cooled to 32～37 ℃; Described synergist is by MgSO₄ 0.1～0.5g/L，CaCl ₂1.0～4.0g/L，ZnSO ₄ 0.2～1.5g/L，FeSO ₄ 0.1～0.8g/L，CuSO ₄ 0.0 1～0.05g/L，Co(NO ₃) ₂ 0.02～ 0.06g/L，MnSO ₄0.3 the aqueous solution that～1.5g/L forms;

4) the bacillus primary seed solution that second order fermentation (producing fermentation): with step 3) obtains, volume ratio with 5～10% is inoculated into and is equipped with in the fermentation tank of producing fermentation culture, prescription and the controlled condition of fermentation culture are same as step 3) first order seed liquid culture medium, ferment after 16～24 hours, by microscopic examination, when being arranged, a large amount of sporulation stops fermentation;

5) with the zymotic fluid of bacillus, be that 0.5～0.8: 1 ratio mixes according to the weight ratio of zymotic fluid and carrier, be the wet-milling shape, then on fluidizing drying bed in 80～100 ℃ of crushed after being dried to more than 80 orders, obtain bacillus dry powder B, at last according to dull and stereotyped used culture medium and the condition of culture of rejuvenation of cultivating of bacillus, the viable bacteria content of dry powder is carried out conventional plate count detect, according to count results, utilize carrier viable bacteria content to be diluted to respectively 〉=6,000,000,000/g; Described carrier (adsorbent) proportioning is: lightweight CaCO₃30～60wt%, inferior powder 20～40wt%, corn protein powder 20～30wt%.

Described saccharomycete dry powder C is made by following method:

1) dull and stereotyped cultivation rejuvenation: with method streak inoculation on the yeast plating medium of yeast inclined-plane seed employing streak inoculation, in 24～30 ℃ of cultivations 20～24 hours, make the yeast rejuvenation, and form single bacterium colony; Described yeast plating medium is the mixture by following w/v (W/V) preparation: soy peptone 0.2～0.7%, glucose 0.5～1.5%, yeast soak powder 0.1～0.5%, malt extract 0.2～1%, agar powder 2～3%, pH=5.8～6.0, sterilized 15 minutes for 121 ℃, for subsequent use; Described yeast comprises: saccharomyces cerevisiae or brewer's yeast, rhodothece rubra, red phaffia rhodozyma, Torulopsis candida, Pichia pastoris, Chinese red yeast, candida utili;

2) choose single bacterium colony the Yeast Cultivation flat board of making shake-flask seed: from step 1) cultivating, be inoculated in the triangular flask liquid medium, in 28～32 ℃ of cultivations 24～37 hours, obtain the yeast shake-flask seed liquid; Described nutrient solution is the mixed liquor by following w/v (W/V) preparation: soy peptone 0.2～0.7%, glucose 0.5～1.5%, yeast soak powder 0.1～0.5%, malt extract 0.2～1%, pH=5.8～6.0, sterilized 15 minutes for 121 ℃, for subsequent use;

3) the yeast shake-flask seed that production primary seed solution: with step 2) obtains is by the inoculum concentration of 5～12v%, be inoculated in the first order seed nutrient solution of first class seed pot, cultivated 24～37 hours in 28～37 ℃, mixing speed 160～220rpm, throughput is 1.0～3.0vvm, obtains the primary seed solution of yeast; Described yeast first order seed nutrient solution is the mixed liquor by following w/v (W/V) preparation: 120 order bean cake powders 0.5～1.5%, 120 order fish meal 0.2～0.8%, 120 order corn flours 0.5～1.0%, glucose 0.3～1.0%, NaCl 0.1～0.3%, Na₂HPO ₄0.2～0.5%, synergist 0.05～0.2%, defoamer 0.03～0.1%, and pH=5.8～6.0 through 121 ℃ of sterilizations 30～45 minutes, are cooled to 28～37 ℃; Described synergist is by MgSO₄ 0.05～0.3g/L，CaCl ₂ 2.5～4g/L，ZnSO ₄ 0.6～1.5g/L，FeSO ₄0.1～1.0g/L，MnSO ₄ 0.4～0.8g/L，CuSO ₄0.02 the aqueous solution that～0.08g/L forms;

4) the yeast first order seed that second order fermentation (producing fermentation): with step 3) obtains, be inoculated in the fermentation tank that the production fermentation culture is housed with 4～10% volume ratios, prescription and the controlled condition of fermentation culture are same as step 3) the first order seed nutrient solution, ferment after 24～60 hours, with step 1) culture medium, detect with dull and stereotyped meter method or counting method of blood cell and to stop fermentation when yeast count reaches 1,000,000,000/ml;

5) above-mentioned saccharomycetic zymotic fluid is added in the carrier that is pre-mixed, the weight ratio of zymotic fluid and carrier is 0.6～0.9: 1, fluidized bed drying, 45～60 ℃ of baking temperatures obtain saccharomycete dry powder C, then according to dull and stereotyped used culture medium and the condition of rejuvenation of cultivating of yeast, viable bacteria content to dry powder carries out conventional plate count detection, according to count results, utilize above-mentioned carrier viable bacteria content to be diluted to 〉=400,000,000/g, for subsequent use; Described carrier proportioning is: lightweight CaCO₃30～60wt%, inferior powder 20～40wt%, corn protein powder 20～30wt%.

Described dry powder of lactic acid bacteria D is made by following method:

1) dull and stereotyped cultivation rejuvenation: with method streak inoculation on the lactic acid bacteria plating medium of lactic acid bacteria inclined-plane seed employing streak inoculation, in 33～38 ℃ of cultivations 24～48 hours, make the lactic acid bacteria rejuvenation, and form single bacterium colony; Described lactic acid bacteria plating medium is the mixture by following w/v (W/V) preparation: soy peptone 0.5～1%, glucose 1～2%, yeast soak powder 0.3～0.7%, potassium dihydrogen phosphate 0.3～0.5%, beef extract 0.5～1.5%, Triammonium citrate 1.5～2.3%, Tween-80 0.5～1.4%, sodium acetate 0.2～0.6%, agar powder 2～2.5%, pH=5.5～5.8, sterilized 15 minutes for 121 ℃, for subsequent use; Described lactic acid bacteria comprises: lactobacillus acidophilus, lactobacillus bulgaricus, Lactobacillus casei, lactobacillus cellobiosas, Lactobacillus delbrueckii, lactobacillus fermenti, lactobacillus lactis, Lactobacillus plantarum, streptococcus cremoris, streptococcus fecalis, streptococcus lactis, streptococcus thermophilus, Pediococcus acidilactici, pediococcus cerevisiae, streptococcus faecalis, lactobacillus, lactobacillus thermophilus;

2) choose single bacterium colony in the lactic acid bacteria bacterium colony of making shake-flask seed: with step 1) cultivating, be inoculated in the lactic acid bacteria liquid culture medium, the employing conventional method in 35～40 ℃ of cultivations 24～48 hours, obtains the lactic acid bacteria shake-flask seed on the triangular flask fluid nutrient medium; Described culture medium is the mixed liquor by following w/v (W/V) preparation: soy peptone 0.5～1%, glucose 1～2%, yeast soak powder 0.3～0.7%, potassium dihydrogen phosphate 0.3～0.5%, beef extract 0.5～1.5%, Triammonium citrate 1.5～2.3%, Tween-80 0.5～1.4%, sodium acetate 0.2～0.6%, pH=5.5～5.8, sterilized 15 minutes for 121 ℃, for subsequent use;

3) the lactic acid bacteria shake-flask seed that production primary seed solution: with step 2) obtains is with the inoculum concentration of 6～12v%, be inoculated in the fluid nutrient medium in the first class seed pot, cultivate 17～60h in 35～38 ℃, mixing speed 150～200rpm, interval ventilation in 1 hour 5～10 minutes, throughput is 1～2.5vvm, until lactic acid bacteria number reaches 1,000,000,000, obtains the primary seed solution of lactic acid bacteria; Described first order seed liquid culture medium prescription is the mixed liquor by following w/v (W/V) preparation: 120 order bean cake powders 0.5～2.0%, 120 order fish meal 0.5～1.0%, fish peptone 0.5～2%, lactose 0.2～1.5%, 120 order corn flours 0.5～1.0%, glucose 1～2.5%, NaCl 0.1～0.3%, Na₂HPO ₄0.15～0.3%, synergist 0.05～0.3%, defoamer 0.03～0.1%, and pH=5.5～5.8 through 121 ℃ of sterilizations 30～45 minutes, are cooled to 32～37 ℃; Described synergist is by CaCl₂ 2～4g/L，ZnSO ₄ 0.5～1.5g/L，FeSO ₄ 0.2～1.0g/L，CuSO ₄0.01～0.08g/L，Co(NO ₃) ₂ 0.02～0.06g/L，MgSO ₄0.1 the aqueous solution that～0.5g/L forms;

4) the lactic acid bacteria first order seed that second order fermentation (producing fermentation): with step 3) obtains, be inoculated in the fermentation tank that the production fermentation culture is housed with 5～10% volume ratios, prescription and the controlled condition of fermentation culture are same as step 3) first order seed liquid culture medium, ferment after 17～60 hours, when detecting the pH value and dropping to 5.0 left and right sides (at this moment, the result of test of many times shows that the lactic acid bacteria amount reaches greater than 10⁹The CFU/ml zymotic fluid) stops fermentation;

5) streptococcus acidi lactici fermented solution is added in the carrier that is pre-mixed, the weight ratio of concentrate and carrier is 0.4～0.9: 1, then through 45～60 ℃ of dryings of fluid bed, is crushed to more than 80 orders, obtains dry powder of lactic acid bacteria D; At last, according to dull and stereotyped used culture medium and the condition of culture of rejuvenation of cultivating of lactic acid bacteria, the viable bacteria content of dry powder is carried out conventional plate count detect, according to count results, utilize described carrier viable bacteria content to be diluted to respectively 〉=400,000,000/g, for subsequent use; Described carrier proportioning is: lightweight CaCO₃30～60wt%, inferior powder 20～40wt%, corn protein powder 20～30wt%.

The invention provides the purposes of a kind of described little peptide and beneficial microbe symphysis unit (element) feed addictive. This additive is as Substitutes For Antibiotic, eliminate medicament residue, improve the product quality such as meat, eggs and milk, can be used for preventing and treating the various animals diarrhoea of having loose bowels, improve breeding performonce fo animals (increasing meat, egg, milk yield), improve immunity, premunition and feed efficiency, reduction animal death rate, also can be used for making ensiling, yellow storage feed and biological feedstuff, cotton dregs (cake) and the dish dregs of rice (cake) detoxification purify mew environment (ammonia falls in deodorizing) and purify fishes and shrimps pond water quality etc.

Little peptide provided by the invention and beneficial microbe symphysis unit feed addictive takes full advantage of the advantage that benefit is given birth to type bacillus, saccharomycete, lactic acid bacteria and little peptide: (1) benefit gives birth to that type bacillus is difficult for inactivation, stability is high, arrives in the animal intestinal effectively regulating intestinal canal microecological balance; (2) benefit is given birth to and can be grown rapidly, grow after the type bacillus enters alimentary canal, produces very strong protease, lipase, amylase isoreactivity, promotes the digestibility and utilization of protein, fat and starch; Simultaneously, xylan, araban, galactan, pectin etc. also there is degradation capability; The bacillus that has can also produce Cobastab and vitamin K. Therefore, gemma has well aid digestion, growth promoting function. (3) the living type bacillus of benefit this type of bacterium in metabolic process passes through to reduce oxidation-reduction potential in the alimentary canal, and produces a small amount of antibacterial material, harmful microorganism is played suppress and killing action. (4) spore of the living type bacillus of benefit has very strong tolerance to poor environments such as heat, pressure, acid, alkali, enzyme and some drugs, X ray, good stability in feed processing, storage and alimentary canal, need not the special protection processing, so use comparatively extensive; (5) lactic acid bacteria is the dominant microflora in the normal flora in the animal alimentary canal, " be throughout one's life the companion; be throughout one's life benefit " with animal, bringing into play antagonism pathogenic microorganism (such as Escherichia coli and salmonella etc.), start special and nospecific immunity system (strengthening immunity and premunition), reduce cholesterol, in and enterotoxin, synthesize nutriment, produce digestive enzymes and regulate the function such as raising poultry nutritive metabolism. (6) saccharomycete can not only help animal digestion, can also promote the endogenous beneficial microorganism vigor of alimentary canal, replenishes the B family vitamin, and its cell wall polysaccharides also has stimulating animal immunity, improves the effect of premunition. (7) little peptide of the present invention has in the remarkable promotion intestines and stomach such as the microbial growths such as bacillus, lactic acid bacteria, saccharomycete propagation and enlivens its metabolism, and active high, improve 70% such as the speed of growth that can make rumen bacteria, the small peptide of every liter of rumen fluid reaches 10mg can be enough to make the growth of beneficial bacterium to reach optimal degree, therefore, with the effect brought into play after little peptide and bacillus, lactic acid bacteria, the saccharomycete combination or the effect effect considerably beyond independent use, it is the main force of the growth promotion health-care agent such as substitute antibiotics; (8) all right stimulating animal Development of Immune Organs of product of the present invention, strengthen immunity, reduce the incidence of disease, substitute antibiotics, reduce medicament residue, can be widely used in fowl, ox, sheep, pig, aquatic products and economic animal, realize the greenization cultivation, this has great economic benefit, social benefit and ecological benefits; (9) little peptide provided by the invention and beneficial microbe symphysis unit feed addictive is a kind of green feed additive, and its promotion and application are conducive to human health and the raising of animal products quality.

The specific embodiment

Embodiment 1, utilize dregs of beans and bacillus subtilis to prepare little peptide dry powder A-1

1) with in 121 ℃ of fluid nutrient mediums of sterilizing 30 minutes and being cooled to after 28 ℃ of bacillus subtilis access, cultivated 24 hours, and obtained seed liquor for 28 ℃; Described fluid nutrient medium is the mixed liquor in following ratio preparation: peptone 1.3g, glucose 3g, molasses 3g, potato immersion liquid 50ml, soluble starch 10g, sucrose 10g, beef extract 2g, yeast soak powder 3.5g, bean cake powder (60 order) 1.5g, growth factor 1ml, water 1000ml; Growth factor wherein is for containing MgSO₄.7H ₂O20.00mg/ml，CaCl ₂.2H ₂O 2.20mg/ml，ZnSO ₄.7H ₂O 0.81mg/ml，FeSO ₄.7H ₂O 0.44mg/ml， MnSO ₄.H ₂O 0.13mg/ml，CuSO ₄.5H ₂The mixture of O 0.02mg/ml.

2) with fermentation substrate---soyabean protein powder is put into fermentation tank after being crushed to 60 orders, add successively entry and carbon source---corn flour and cane molasses (weight ratio is 1: 1), add the weight of entry and the weight ratio of the contained protein of fermentation substrate is 100: 5, add the nitrogen content of the phosphorus content of carbon source and fermentation substrate weight ratio be 1: 2, add again defoamer 0.03%, mixing, with mixture (nutrient solution) at 0.1MPa, sterilized 30 minutes for 121 ℃, be cooled to 28 ℃, adding step 1) seed liquor that obtains, the volume ratio of seed liquor and nutrient solution is 0.3: 10, mix rear at 28 ℃, speed with 0.5vvm passes into filtrated air, stir fermentation with 150rpm speed, use during this time conventional ninhydrin method determination of color degree of hydrolysis (Degree of Hydrolysis, DH%), degree of hydrolysis of detection in per 6 hours before 24 hours, between 24～48 hours, detect a degree of hydrolysis in per 4 hours, and detected a degree of hydrolysis in per 2 hours after 48 hours, until the degree of hydrolysis of protein hydrolyzate reaches 25 %, then through the ultra high temperature short time sterilization machine in 140 ℃ of sterilizations 8 seconds, obtain containing the proteolysis liquor of little peptide.

3) centrifugal 30 minutes of the protein hydrolyzate 5000rpm that contains little peptide that centrifugal slagging-off: with step 2) obtains, remove slag, the supernatant that obtains is that the ultrafilter of 1000Da is removed the impurity such as high molecular weight protein by retaining molecular weight again, obtains the filtrate of molecular weight≤1000Da; This filtrate is concentrated to 50% of original volume through negative-pressure vacuum, the concentrate that obtains is added in the carrier that is pre-mixed, press concentrate: the envelope-bulk to weight ratio of carrier=0.5: 1 mixes, described carrier is 5 weight portion organic carriers (2 weight portion wheat bran, 2 weight portion bean cake powders, 1 weight portion corn protein powder) and 5 weight portion inorganic carriers (precipitated calcium carbonate) composition, it is rear 45 ℃ of dryings to stir, then be crushed to more than 80 orders, obtain little peptide dry powder A-1.

Embodiment 2, utilize dregs of beans and black-koji mould to prepare little peptide dry powder A-2

1) with in 121 ℃ of fluid nutrient mediums of sterilizing 30 minutes and being cooled to after 35 ℃ of black-koji mould access, cultivated 24 hours, and obtained seed liquor for 35 ℃; Described fluid nutrient medium is the mixed liquor in following ratio preparation: peptone 2.5g, glucose 5g, molasses 5g, potato immersion liquid 100ml, soluble starch 20g, sucrose 30g, beef extract 3g, yeast soak powder 5g, dregs of beans 2g, growth factor-2 ml, water 1000ml; Growth factor wherein is for containing MgSO₄.7H ₂O 30.00mg/ml， CaCl ₂.2H ₂O 3.50mg/ml，ZnSO ₄.7H ₂O 1.10mg/ml，FeSO ₄.7H ₂O 0.60mg/ml，MnSO ₄.H ₂O0.20mg/ml，CuSO ₄.5H ₂The mixture of O 0.04mg/ml.

2) with fermentation substrate---dregs of beans is put into fermentation tank after being crushed to 120 orders, add successively entry and carbon source---corn molasses and white granulated sugar (weight ratio=3: 2), add the weight of entry and the weight ratio of the contained protein of fermentation substrate is 100: 18, add the nitrogen content of the phosphorus content of carbon source and fermentation substrate weight ratio be 1: 35, add again defoamer 0.1%, mixing, with mixture (being nutrient solution) at 0.1MPa, sterilized 10 minutes for 125 ℃, be cooled to 20 ℃, adding step 1) seed liquor that obtains, the volume ratio of seed liquor and nutrient solution is 1: 10, mix rear at 42 ℃, speed with 2.5vvm passes into filtrated air, stir fermentation with 300rpm speed, use during this time conventional ninhydrin method determination of color degree of hydrolysis (DH%), degree of hydrolysis of detection in per 6 hours before 24 hours, between 24～48 hours, detected a degree of hydrolysis in per 4 hours, degree of hydrolysis of detection in per 2 hours after 48 hours, until the degree of hydrolysis of protein hydrolyzate (DH%) reaches 40%, then through the ultra high temperature short time sterilization machine in 145 ℃ of sterilizations 2 seconds, obtain containing the proteolysis liquor of little peptide.

3) protein hydrolyzate that contains little peptide that centrifugal slagging-off: with step 2) obtains centrifugal 10 minutes in 7000rpm, remove slag, the supernatant that obtains is that the ultrafilter of 1000Da is removed the impurity such as high molecular weight protein by retaining molecular weight again, obtains the filtrate of molecular weight≤1000Da; This filtrate is concentrated to 50% of original volume through negative-pressure vacuum, the concentrate that obtains is added in the carrier that is pre-mixed, press concentrate: the envelope-bulk to weight ratio of carrier=0.8: 1 mixes, described carrier is that 2 weight portion organic carriers (maize cob meal) and 3 weight portion inorganic carriers (stone flour) form, it is rear 80 ℃ of dryings to stir, then be crushed to more than 80 orders, obtain little peptide dry powder A-2.

Embodiment 3, utilize beancake powder and aspergillus oryzae to prepare little peptide dry powder A-3

1) with in 121 ℃ of fluid nutrient mediums of sterilizing 30 minutes and being cooled to after 32 ℃ of aspergillus oryzae access, cultivated 36 hours, and obtained seed liquor for 32 ℃; Described fluid nutrient medium is the mixed liquor in following ratio preparation: peptone 1.8g, glucose 4g, molasses 4g, potato immersion liquid 70ml, soluble starch 15g, sucrose 20g, beef extract 2.5g, yeast soak powder 4g, bean cake powder (80 order) 1.8g, growth factor 1.5ml, water 1000ml; Growth factor wherein is for containing MgSO₄.7H ₂O25.00mg/ml，CaCl ₂.2H ₂O 3.00mg/ml，ZnSO ₄.7H ₂O 1.00mg/ml，FeSO ₄.7H ₂O 0.50mg/ml， MnSO ₄.H ₂O 0.17mg/ml，CuSO ₄.5H ₂The mixed liquor of O 0.03mg/ml.

2) with fermentation substrate---soya-bean cake is put into fermentation tank after being crushed to 80 orders, add successively entry and carbon source---starch, add the weight of entry and the weight ratio of the contained protein of fermentation substrate is 100: 10, add the nitrogen content of the phosphorus content of carbon source and fermentation substrate weight ratio be 1: 10, add again defoamer 0.06%, mixing, with mixture (being nutrient solution) at 0.11MPa, sterilized 20 minutes for 123 ℃, be cooled to 32 ℃, adding step 1) seed liquor that obtains, the volume ratio of seed liquor and nutrient solution is 0.5: 10, mix rear at 28 ℃, speed with 1.5vvm passes into filtrated air, stir fermentation with 180rpm speed, use during this time conventional ninhydrin method determination of color degree of hydrolysis (DH%), degree of hydrolysis of detection in per 6 hours before 24 hours, between 24～48 hours, detected a degree of hydrolysis in per 4 hours, degree of hydrolysis of detection in per 2 hours after 48 hours, until the degree of hydrolysis of protein hydrolyzate (DH%) reaches 30%, then through the ultra high temperature short time sterilization machine in 142 ℃ of sterilizations 6 seconds, obtain containing the proteolysis liquor of little peptide.

3) protein hydrolyzate that contains little peptide that centrifugal slagging-off: with step 2) obtains centrifugal 15 minutes in 6000rpm, remove slag, the supernatant that obtains is that the ultrafilter of 1000Da is removed the impurity such as high molecular weight protein by retaining molecular weight again, obtains the filtrate of molecular weight≤1000Da; This filtrate is concentrated to 50% of original volume through negative-pressure vacuum, the concentrate that obtains is added in the carrier that is pre-mixed, press concentrate: the envelope-bulk to weight ratio of carrier=0.7: 1 mixes, described carrier is that 3 weight portion organic carriers (degreasing powdered rice hulls) and 4 weight portion inorganic carriers (medical stone) form, it is rear 60 ℃ of dryings to stir, then be crushed to more than 80 orders, obtain little peptide dry powder A-3.

Embodiment 4, utilize fish meal and bafillus natto to prepare little peptide dry powder A-4

1) with in 121 ℃ of fluid nutrient mediums of sterilizing 30 minutes and being cooled to after 30 ℃ of bafillus natto access, cultivated 24 hours, and obtained seed liquor for 30 ℃; Described fluid nutrient medium is the mixed liquor in following ratio preparation: peptone 2.0g, glucose 4g, molasses 4g, potato immersion liquid 80ml, soluble starch 15g, sucrose 17g, beef extract 2.2g, yeast soak powder 4.5g, bean cake powder (80 order) 1.7g, growth factor 1.7ml, water 1000ml; Growth factor wherein is for containing MgSO₄.7H ₂O 25.00mg/ml，CaCl ₂.2H ₂O 3.00mg/ml，ZnSO ₄.7H ₂O 0.90mg/ml，FeSO ₄.7H ₂O0.50mg/ml，MnSO ₄.H ₂O 0.18mg/ml，CuSO ₄.5H ₂The mixed liquor of O 0.033mg/ml.

2) with fermentation substrate---fish meal is put into fermentation tank after being crushed to 80 orders, add successively entry and carbon source---beet molasses, add the weight of entry and the weight ratio of the contained protein of fermentation substrate is 100: 15, add the nitrogen content of the phosphorus content of carbon source and fermentation substrate weight ratio be 1: 20, add again defoamer 0.08%, mixing, with mixture (being nutrient solution) at 0.11MPa, sterilized 30 minutes for 121 ℃, be cooled to 36 ℃, adding step 1) seed liquor that obtains, the volume ratio of seed liquor and nutrient solution is 0.5: 10, mix rear at 32 ℃, speed with 0.8vvm passes into filtrated air, stir fermentation with 260rpm speed, use during this time conventional ninhydrin method determination of color degree of hydrolysis (DH%), degree of hydrolysis of detection in per 6 hours before 24 hours, between 24～48 hours, detected a degree of hydrolysis in per 4 hours, degree of hydrolysis of detection in per 2 hours after 48 hours, until the degree of hydrolysis of protein hydrolyzate (DH%) reaches 35%, then through the ultra high temperature short time sterilization machine in 143 ℃ of sterilizations 5 seconds, obtain containing the proteolysis liquor of little peptide.

3) protein hydrolyzate that contains little peptide that centrifugal slagging-off: with step 2) obtains centrifugal 13 minutes in 6500rpm, remove slag, the supernatant that obtains is that the ultrafilter of 1000Da is removed the impurity such as high molecular weight protein by retaining molecular weight again, obtains the filtrate of molecular weight≤1000Da; This filtrate is concentrated to 50% of original volume through negative-pressure vacuum, the concentrate that obtains is added in the carrier that is pre-mixed, press concentrate: the envelope-bulk to weight ratio of carrier=0.6: 1 mixes, described carrier is that 2 weight portion organic carriers (wheat bran) and 3 weight portion inorganic carriers (precipitated calcium carbonate) form, it is rear 70 ℃ of dryings to stir, then be crushed to more than 80 orders, obtain little peptide dry powder A-4.

Embodiment 5～24, the little peptide dry powder A-5～A-24 of preparation

Method according to embodiment 1～4 prepares little peptide dry powder A-5～A-24, and preparation condition is listed in table 1.

The preparation of table 1, little peptide dry powder A-5～A-24

Embodiment	Little peptide dry powder	Preparation method (with embodiment X)	The used bacterial classification of preparation seed liquor	Fermentation substrate	Carbon source	Protein hydrolysis degree	Organic carrier (weight portion)	Inorganic carrier (weight portion)
									5	A-5	1	The aspergillus terricola bacterium	Peas protein maize albumen	Corn flour	25	Maize cob meal 3	Stone flour 4
6	A-6	2	Aspergillus oryzae	Red bean albumen wheat gluten blood meal	Edible brown sugar	27	Degreasing powdered rice hulls 4	Medical stone 5
									7	A-7	3	Bafillus natto	Large aleuronat Cottonseed Meal fish meal	Edible sugar	34	Defatted rice bran 5	Zeolite powder 5
8	A-8	4	Aspergillus niger	Cottonseed Meal rapeseed dregs feather meal	Corn syrup	32	Bean cake powder 2	Precipitated calcium carbonate 2 stone flours 2
									9	A-9	1	Bacillus subtilis	Lactoprotein muscles casein	Cane molasses	35	Beancake powder 3	Stone flour 1 medical stone 1 zeolite powder 1
10	A-10	2	Bacillus pumilus	Animal discards internal organs earthworm protein powder	Beet molasses	40	Corn protein powder 4	Medical stone 2 zeolite powders 1
									11	A-11	3	The bacillus pumilus aspergillus niger	Squid albumen fish-skin	Corn flour brown sugar	40	Wheat bran 2 maize cob meals 3	Stone flour 2 medical stones 2
12	A-12	4	The bacillus pumilus bacillus subtilis	Zein	The brown sugar edible sugar	35	Wheat bran 1 degreasing powdered rice hulls 1	Precipitated calcium carbonate 2 stone flours 1 medical stone 2
									13	A-13	1	Aspergillus terricola bacterium bacillus subtilis	Muscles	The edible sugar corn syrup	33	Wheat bran 2 defatted rice brans 1	Precipitated calcium carbonate 2 stone flours 2
14	A-14	2	Aspergillus oryzae aspergillus terricola bacterium	The earthworm protein powder	Corn syrup starch	40	Wheat bran 1 bean cake powder 3	Stone flour 2 medical stones 1
									15	A-15	3	The bacillus subtilis bafillus natto	Squid albumen	The starch cane molasses	38	Wheat bran 2 beancake powders 3	Precipitated calcium carbonate 3
16	A-16	4	Aspergillus oryzae aspergillus terricola bacterium	Fish-skin	The starch beet molasses	33	Maize cob meal 1 degreasing powdered rice hulls 1	Stone flour 4
									17	A-17	1	Aspergillus oryzae aspergillus terricola bacterium aspergillus niger	Red bean albumen	Starch cane molasses beet molasses	36	Maize cob meal 1 bean cake powder 2	Medical stone 5
18	A-18	2	The bacillus subtilis bafillus natto	Wheat gluten	Corn flour	30	Degreasing powdered rice hulls 1 bean cake powder 2 corn protein powders 1	Zeolite powder 5
									19	A-19	3	Aspergillus oryzae aspergillus terricola bacterium	Large aleuronat	Edible brown sugar	35	Defatted rice bran 1 bean cake powder 2 beancake powders 2	Precipitated calcium carbonate 4
20	A-20	4	The aspergillus terricola bacterium	Cottonseed Meal	Edible sugar	33	Maize cob meal 1 bean cake powder 1	Stone flour 3
									21	A-21	1	Aspergillus oryzae	Rapeseed dregs	Corn syrup	26	Wheat bran 3	Medical stone 3
22	A-22	2	Bafillus natto	Feather meal	Starch	29	Maize cob meal 4	Zeolite powder 4
									23	A-23	3	Aspergillus niger	Lactoprotein	Cane molasses	33	Degreasing powdered rice hulls 5	Precipitated calcium carbonate 5
24	A-24	4	Bacillus subtilis	Casein	Beet molasses	31	Defatted rice bran 2	Stone flour 5

Embodiment 25, preparation bacillus subtilis dry powder B-1

1) dull and stereotyped cultivation rejuvenation: Bacillus subtilis strain is adopted method streak inoculation on the bacillus plating medium of streak inoculation, in 25 ℃ of cultivations 12 hours, make the bacillus subtilis rejuvenation, and form single bacterium colony; Described bacillus subtilis bacterium culture medium is the mixture by following w/v (W/V) preparation: peptone 0.5%, glucose 0.5%, beef extract 0.5%, potassium dihydrogen phosphate 0.3%, agar 3%, pH=7.5～7.6, sterilized 30 minutes for 121 ℃, for subsequent use;

2) select typical single bacterium colony the bacillus subtilis bacterium colony of making shaking flask (triangular flask) seed: from step 1) cultivating, be inoculated in the bacillus liquid culture medium, in 28 ℃ of cultivations 20 hours, obtain the bacillus subtilis shake-flask seed liquid; Described bacillus liquid culture medium is the mixed liquor by following w/v (W/V) preparation: peptone 0.5%, glucose 0.5%, beef extract 0.5%, potassium dihydrogen phosphate 0.3%, 3.08% manganese sulfate solution 0.5%, pH=7.5～7.6, sterilized 30 minutes for 121 ℃, be cooled to 28 ℃, for subsequent use;

3) production primary seed solution: first according to following weight volume ratio (W/V) preparation first order seed liquid culture medium: 120 order bean cake powders 0.5%, 120 order fish meal 0.2%, 120 order corn flours 0.5%, glucose 1.0%, NaCl 0.1%, NaOH0.05%, Na₂HPO ₄0.1%, synergist 0.05%, defoamer 0.08%; Synergist wherein is by MgSO₄ 0.1g/L， CaCl ₂ 1.0g/L，ZnSO ₄ 0.2g/L，FeSO ₄ 0.1g/L，CuSO ₄ 0.01g/L，Co(NO ₃) ₂ 0.02g/L，MnSO ₄0.3g/L the aqueous solution that forms; The pH=7.5 of this first order seed liquid culture medium～7.6, through 121 ℃ of sterilizations 45 minutes, be cooled to 32 ℃, then with step 2) the bacillus subtilis shake-flask seed liquid that obtains is with the inoculum concentration of 2v%, be inoculated in the first order seed liquid culture medium of producing used first class seed pot, cultivate 18h, mixing speed 150rpm in 32 ℃, throughput is 1vvm, obtains the primary seed solution of bacillus subtilis;

4) the bacillus subtilis primary seed solution that second order fermentation (producing fermentation): with step 3) obtains; ratio with 5% volume is inoculated in the fermentation tank that the production fermentation culture is housed; the same step 3) of the prescription of fermentation culture and controlled condition; ferment after 16 hours; by microscopic examination; when a large amount of sporulation is arranged, shut down, obtain the zymotic fluid of bacillus subtilis.

5) with the zymotic fluid of bacillus subtilis, be that 0.5: 1 ratio mixes according to the weight ratio of zymotic fluid and carrier, be the wet-milling shape, described carrier (adsorbent) proportioning is: lightweight CaCO₃60wt%, inferior powder 20wt%, corn protein powder 20wt%, then on fluidizing drying bed in 80 ℃ of crushed after being dried to more than 80 orders, obtain bacillus dry powder B-1, at last according to dull and stereotyped used culture medium and the condition of culture of rejuvenation of cultivating of bacillus, viable bacteria content to dry powder carries out conventional plate count detection, according to count results, utilize carrier viable bacteria content to be diluted to respectively 〉=6,000,000,000/g.

Embodiment 26, preparation bacillus subtilis dry powder B-2

1) dull and stereotyped cultivation rejuvenation: Bacillus subtilis strain is adopted method streak inoculation on the bacillus plating medium of streak inoculation, in 30 ℃ of cultivations 18 hours, make the bacillus subtilis rejuvenation, and form single bacterium colony; Described bacillus subtilis bacterium culture medium is the mixture by following w/v (W/V) preparation: peptone 0.8%, glucose 1.3%, beef extract 1.0%, potassium dihydrogen phosphate 0.5%, agar 3.5%, pH=7.5～7.6, sterilized 30 minutes for 121 ℃, for subsequent use;

2) select typical single bacterium colony the bacillus subtilis bacterium colony of making shaking flask (triangular flask) seed: from step 1) cultivating, be inoculated in the bacillus liquid culture medium, in 32 ℃ of cultivations 24 hours, obtain the bacillus subtilis shake-flask seed liquid; Described bacillus liquid culture medium is the mixed liquor by following w/v (W/V) preparation: peptone 0.8%, glucose 1.3%, beef extract 1.0%, potassium dihydrogen phosphate 0.5%, agar 3.5%, 3.08% manganese sulfate solution 1.0%, pH=7.5～7.6, sterilized 30 minutes for 121 ℃, be cooled to 32 ℃, for subsequent use;

3) production primary seed solution: first according to following weight volume ratio (W/V) preparation first order seed liquid culture medium: 120 order bean cake powders 1.0%, 120 order fish meal 1.0%, 120 order corn flours 0.7%, glucose 2.5%, NaCl 0.2%, NaOH0.08%, Na₂HPO ₄0.3%, synergist 0.1%, defoamer 0.08%; Synergist wherein is by MgSO₄ 0.5g/L， CaCl ₂ 4.0g/L，ZnSO ₄ 1.5g/L，FeSO ₄ 0.8g/L，CuSO ₄ 0.05g/L，Co(NO ₃) ₂ 0.06g/L，MnSO ₄1.5g/L the aqueous solution that forms; The pH=7.5 of this first order seed liquid culture medium～7.6, through 121 ℃ of sterilizations 35 minutes, be cooled to 35 ℃, then with step 2) the bacillus subtilis shake-flask seed liquid that obtains is with the inoculum concentration of 3v%, be inoculated in the first order seed liquid culture medium of producing used first class seed pot, cultivate 20h, mixing speed 180rpm in 35 ℃, throughput is 1.5vvm, obtains the primary seed solution of bacillus subtilis;

4) the bacillus subtilis primary seed solution that second order fermentation (producing fermentation): with step 3) obtains; ratio with 10% volume is inoculated in the fermentation tank that the production fermentation culture is housed; the same step 3) of the prescription of fermentation culture and controlled condition; ferment after 20 hours; by microscopic examination; shut down when a large amount of sporulation is arranged, obtain the zymotic fluid of bacillus subtilis.

5) with the zymotic fluid of bacillus subtilis, be that 0.7: 1 ratio mixes according to the weight ratio of zymotic fluid and carrier, be the wet-milling shape, described carrier (adsorbent) proportioning is: lightweight CaCO₃45wt%, inferior powder 30wt%, corn protein powder 25wt%, then on fluidizing drying bed in 90 ℃ of crushed after being dried to more than 80 orders, obtain bacillus dry powder B-2, at last according to dull and stereotyped used culture medium and the condition of culture of rejuvenation of cultivating of bacillus, viable bacteria content to dry powder carries out conventional plate count detection, according to count results, utilize carrier viable bacteria content to be diluted to respectively 〉=6,000,000,000/g.

Embodiment 27, preparation bacillus subtilis dry powder B-3

1) dull and stereotyped cultivation rejuvenation: Bacillus subtilis strain is adopted method streak inoculation on the bacillus plating medium of streak inoculation, in 35 ℃ of cultivations 24 hours, make the bacillus subtilis rejuvenation, and form single bacterium colony; Described bacillus subtilis bacterium culture medium is the mixture by following w/v preparation: peptone 1%, glucose 2%, beef extract 1.5%, potassium dihydrogen phosphate 0.6%, agar 4%, pH=7.5～7.6, sterilized 30 minutes for 121 ℃, be cooled to 35 ℃, for subsequent use;

2) select typical single bacterium colony the bacillus subtilis bacterium colony of making shaking flask (triangular flask) seed: from step 1) cultivating, be inoculated in the bacillus liquid culture medium, in 37 ℃ of cultivations 30 hours, obtain the bacillus subtilis shake-flask seed liquid; Described bacillus liquid culture medium is the mixed liquor by following w/v (W/V) preparation: peptone 1.0%, glucose 2%, beef extract 1.5%, potassium dihydrogen phosphate 0.6%, agar 4%, 3.08% manganese sulfate solution 1.5%, pH=7.5～7.6, sterilized 30 minutes for 121 ℃, be cooled to 37 ℃, for subsequent use;

3) production primary seed solution: first according to following weight volume ratio (W/V) preparation first order seed liquid culture medium: 120 order bean cake powders 2.0%, 120 order fish meal 1.5%, 120 order corn flours 1.0%, glucose 4.5%, NaCl 0.3%, NaOH0.1%, Na₂HPO ₄0.5%, synergist 0.2%, defoamer 0.1%; Synergist wherein is by MgSO₄ 0.3g/L，CaCl ₂3.0g/L，ZnSO ₄ 0.8g/L，FeSO ₄ 0.4g/L，CuSO ₄ 0.03g/L，Co(NO ₃) ₂ 0.04g/L，MnSO ₄1.0g/L the aqueous solution that forms; The pH=7.5 of this first order seed liquid culture medium～7.6, through 121 ℃ of sterilizations 30 minutes, be cooled to 37 ℃, then with step 2) the bacillus subtilis shake-flask seed liquid that obtains is with the inoculum concentration of 5v%, be inoculated in the first order seed liquid culture medium of producing used first class seed pot, cultivate 24h, mixing speed 200rpm in 37 ℃, throughput is 2.0vvm, obtains the primary seed solution of bacillus subtilis;

4) the bacillus subtilis primary seed solution that second order fermentation (producing fermentation): with step 3) obtains; ratio with 7% volume is inoculated in the fermentation tank that the production fermentation culture is housed; the same step 3) of the prescription of fermentation culture and controlled condition; ferment after 24 hours; by microscopic examination; when a large amount of sporulation is arranged, shut down, obtain the zymotic fluid of bacillus subtilis.

5) with the zymotic fluid of bacillus subtilis, be that 0.8: 1 ratio mixes according to the weight ratio of zymotic fluid and carrier, be the wet-milling shape, described carrier (adsorbent) proportioning is: lightweight CaCO₃30wt%, inferior powder 40wt%, corn protein powder 30wt%, then on fluidizing drying bed in 100 ℃ of crushed after being dried to more than 80 orders, obtain bacillus dry powder B-3, at last according to dull and stereotyped used culture medium and the condition of culture of rejuvenation of cultivating of bacillus, viable bacteria content to dry powder carries out conventional plate count detection, according to count results, utilize carrier viable bacteria content to be diluted to respectively 〉=6,000,000,000/g.

Embodiment 28～40, preparation bacillus dry powder B-4～B-16

Method according to embodiment 25～27 prepares bacillus dry powder B-4～B-16, and preparation condition is listed in table 2.

The preparation of table 2, bacillus dry powder B-4～B-16

Embodiment	Bacillus dry powder	Preparation method (with embodiment X)	The used bacterial classification of preparation seed liquor	Carrier and post-treatment condition (with embodiment X)
					28	B-4	25	Bacillus coagulans	27
29	B-5	26	Slow bacillus	26
					30	B-6	27	Bacillus licheniformis	25
31	B-7	25	Bacillus pumilus	27
					32	B-8	26	Bacillus circulans	26
33	B-9	27	Bacillus firmus	25
					34	B-10	25	Bacillus megaterium	27
35	B-11	26	Clostridium butyricum	26
					36	B-12	27	Bafillus natto	25
37	B-13	26	Bacillus cercus	27
					38	B-14	26	The bafillus natto bacillus subtilis	26
39	B-15	25	The clostridium butyricum bacillus coagulans	25
					40	B-16	27	The bacillus licheniformis Bacillus circulans	26

Embodiment 41, preparation deep red saccarom ycete dry powder C-1

1) dull and stereotyped cultivation rejuvenation: with method streak inoculation on the yeast plating medium of rhodothece rubra inclined-plane seed employing streak inoculation, in 24 ℃ of cultivations 20 hours, make the yeast rejuvenation, and form single bacterium colony; Described yeast plating medium is the mixture by following w/v (W/V) preparation: soy peptone 0.2%, glucose 0.5%, yeast soak powder 0.1%, malt extract 0.2%, agar powder 2%, pH=5.8～6.0,121 ℃ the sterilization 15 minutes, be cooled to 24 ℃ for subsequent use;

2) choose single bacterium colony the rhodothece rubra culture plate of making shake-flask seed: from step 1) cultivating, be inoculated in the triangular flask liquid medium, in 28 ℃ of cultivations 24 hours, obtain the yeast shake-flask seed liquid; Described nutrient solution is the mixed liquor by following w/v (W/V) preparation: soy peptone 0.2%, glucose 0.5%, yeast soak powder 0.1%, malt extract 0.2%, and pH=5.8～6.0,121 ℃ sterilization 15 minutes is cooled to 28 ℃, and is for subsequent use;

3) the yeast shake-flask seed that production primary seed solution: with step 2) obtains is by the inoculum concentration of 5v%, be inoculated in the first order seed nutrient solution of first class seed pot, cultivated mixing speed 160rpm 24 hours in 28 ℃, throughput is 1.0vvm, obtains the primary seed solution of yeast; Described yeast first order seed nutrient solution is the mixed liquor by following w/v (W/V) preparation: 120 order bean cake powders 0.5%, 120 order fish meal 0.2%, 120 order corn flours 0.5%, glucose 0.3%, NaCl 0.1%, Na₂HPO ₄0.2%, synergist 0.05%, defoamer 0.03%, and pH=5.8～6.0 through 121 ℃ of sterilizations 30 minutes, are cooled to 28 ℃; Described synergist is by MgSO₄ 0.05g/L，CaCl ₂ 2.5g/L，ZnSO ₄ 0.6g/L，FeSO ₄ 0.1g/L， MnSO ₄ 0.4g/L，CuSO ₄0.02g/L the aqueous solution that forms;

4) the yeast first order seed that second order fermentation (producing fermentation): with step 3) obtains, be inoculated in the fermentation tank that the production fermentation culture is housed with 4% volume ratio, prescription and the controlled condition of fermentation culture are same as step 3) the first order seed nutrient solution, ferment after 24 hours, with step 1) culture medium, detect with colony counting method or counting method of blood cell and to stop fermentation when yeast count reaches 1,000,000,000/ml;

5) above-mentioned saccharomycetic zymotic fluid is added in the carrier that is pre-mixed, described carrier comprises: lightweight CaCO₃60wt%, inferior powder 20wt%, corn protein powder 20wt%, the weight ratio of zymotic fluid and carrier is 0.6: 1, fluidized bed drying, 45 ℃ of baking temperatures obtain saccharomycete dry powder C-1, then according to dull and stereotyped used culture medium and the condition of rejuvenation of cultivating of yeast, viable bacteria content to dry powder carries out conventional plate count detection, according to count results, utilize above-mentioned carrier viable bacteria content to be diluted to 〉=400,000,000/g, for subsequent use.

Embodiment 42, preparation deep red saccarom ycete dry powder C-2

1) dull and stereotyped cultivation rejuvenation: with method streak inoculation on the yeast plating medium of rhodothece rubra inclined-plane seed employing streak inoculation, in 28 ℃ of cultivations 22 hours, make the yeast rejuvenation, and form single bacterium colony; Described yeast plating medium is the mixture by following w/v (W/V) preparation: soy peptone 0.5%, glucose 1.0%, yeast soak powder 0.3%, malt extract 0.6%, agar powder 2.5%, pH=5.8～6.0,121 ℃ the sterilization 15 minutes, be cooled to 28 ℃ for subsequent use;

2) choose single bacterium colony the Yeast Cultivation flat board of making shake-flask seed: from step 1) cultivating, be inoculated in the triangular flask liquid medium, in 30 ℃ of cultivations 30 hours, obtain the yeast shake-flask seed liquid; Described nutrient solution is the mixed liquor by following w/v (W/V) preparation: soy peptone 0.5%, glucose 1.0%, yeast soak powder 0.3%, malt extract 0.6%, and pH=5.8～6.0,121 ℃ sterilization 15 minutes is cooled to 30 ℃, and is for subsequent use;

3) the yeast shake-flask seed that production primary seed solution: with step 2) obtains is by the inoculum concentration of 9v%, be inoculated in the first order seed nutrient solution of first class seed pot, cultivated mixing speed 200rpm 30 hours in 33 ℃, throughput is 2.0vvm, obtains the primary seed solution of yeast; Described yeast first order seed nutrient solution is the mixed liquor by following w/v (W/V) preparation: 120 order bean cake powders 1.0%, 120 order fish meal 0.7%, 120 order corn flours 0.7%, glucose 0.6%, NaCl 0.2%, Na₂HPO ₄0.35%, synergist 0.13～0.2%, defoamer 0.06%, and pH=5.8～6.0 through 121 ℃ of sterilizations 40 minutes, are cooled to 33 ℃; Described synergist is by MgSO₄ 0.1g/L，CaCl ₂ 3.0g/L，ZnSO ₄ 0.9g/L，FeSO ₄ 0.5g/L， MnSO ₄ 0.6g/L，CuSO ₄0.05g/L the aqueous solution that forms;

4) the yeast first order seed that second order fermentation (producing fermentation): with step 3) obtains, be inoculated in the fermentation tank that the production fermentation culture is housed with 7% volume ratio, prescription and the controlled condition of fermentation culture are same as step 3) the first order seed nutrient solution, ferment after 48 hours, with step 1) culture medium, detect with colony counting method or counting method of blood cell and to stop fermentation when yeast count reaches 1,000,000,000/ml;

5) above-mentioned saccharomycetic zymotic fluid is added in the carrier that is pre-mixed, described carrier comprises: lightweight CaCO₃45wt%, inferior powder 30wt%, corn protein powder 25wt%, the weight ratio of zymotic fluid and carrier is 0.8: 1, fluidized bed drying, 50 ℃ of baking temperatures obtain saccharomycete dry powder C-2, then according to dull and stereotyped used culture medium and the condition of rejuvenation of cultivating of yeast, viable bacteria content to dry powder carries out conventional plate count detection, according to count results, utilize above-mentioned carrier viable bacteria content to be diluted to 〉=400,000,000/g, for subsequent use.

Embodiment 43, preparation deep red saccarom ycete dry powder C-3

1) dull and stereotyped cultivation rejuvenation: with method streak inoculation on the yeast plating medium of rhodothece rubra inclined-plane seed employing streak inoculation, in 30 ℃ of cultivations 24 hours, make the yeast rejuvenation, and form single bacterium colony; Described yeast plating medium is the mixture by following w/v (W/V) preparation: soy peptone 0.7%, glucose 1.5%, yeast soak powder 0.5%, malt extract 1%, agar powder 3%, pH=5.8～6.0,121 ℃ the sterilization 15 minutes, be cooled to 30 ℃ for subsequent use;

2) choose single bacterium colony the Yeast Cultivation flat board of making shake-flask seed: from step 1) cultivating, be inoculated in the triangular flask liquid medium, in 32 ℃ of cultivations 37 hours, obtain the yeast shake-flask seed liquid; Described nutrient solution is the mixed liquor by following w/v (W/V) preparation: soy peptone 0.7%, glucose 1.5%, yeast soak powder 0.5%, malt extract 1%, and pH=5.8～6.0,121 ℃ sterilization 15 minutes is for subsequent use;

3) the yeast shake-flask seed that production primary seed solution: with step 2) obtains is by the inoculum concentration of 12v%, be inoculated in the first order seed nutrient solution of first class seed pot, cultivated mixing speed 220rpm 37 hours in 37 ℃, throughput is 3vvm, obtains the primary seed solution of yeast; Described yeast first order seed nutrient solution is the mixed liquor by following w/v (W/V) preparation: 120 order bean cake powders 1.5%, 120 order fish meal 0.8%, 120 order corn flours 1.0%, glucose 1.0%, NaCl 0.3%, Na₂HPO ₄0.5%, synergist 0.2%, defoamer 0.1%, pH=5.8～6.0, through 121 ℃ the sterilization 45 minutes, be cooled to 37 ℃ for subsequent use; Described synergist is by MgSO₄ 0.3g/L，CaCl ₂ 4g/L，ZnSO ₄ 1.5g/L，FeSO ₄ 1.0g/L， MnSO ₄ 0.8g/L，CuSO ₄0.08g/L the aqueous solution that forms;

4) the yeast first order seed that second order fermentation (producing fermentation): with step 3) obtains, be inoculated in the fermentation tank that the production fermentation culture is housed with 10% volume ratio, prescription and the controlled condition of fermentation culture are same as step 3) the first order seed nutrient solution, ferment after 60 hours, with step 1) culture medium, detect with colony counting method or counting method of blood cell and to stop fermentation when yeast count reaches 1,000,000,000/ml;

5) above-mentioned saccharomycetic zymotic fluid is added in the mixed in advance carrier, described carrier comprises: lightweight CaCO₃30wt%, inferior powder 40wt%, corn protein powder 30wt%, the weight ratio of zymotic fluid and carrier is 0.9: 1, fluidized bed drying, 60 ℃ of baking temperatures obtain saccharomycete dry powder C-3, then according to dull and stereotyped used culture medium and the condition of rejuvenation of cultivating of yeast, viable bacteria content to dry powder carries out conventional plate count detection, according to count results, utilize above-mentioned carrier viable bacteria content to be diluted to 〉=400,000,000/g, for subsequent use.

Embodiment 44～51, preparation saccharomycete dry powder C-4～C-11

Method according to embodiment 41～43 prepares saccharomycete dry powder C-4～C-11, and preparation condition is listed in table 3.

The preparation of table 3, saccharomycete dry powder C-4～C-11

Embodiment	Saccharomycete dry powder	Preparation method (with embodiment X)	The used bacterial classification of preparation seed liquor	Carrier and post-treatment condition (with embodiment X)
					44	C-4	41	Brewer's yeast	43
45	C-5	42	Red phaffia rhodozyma	42
					46	C-6	43	Torulopsis candida	41
47	C-7	41	Pichia pastoris	43
					48	C-8	42	The Chinese red yeast	41
49	C-9	43	Candida utili	42
					50	C-10	41	The brewer's yeast Pichia pastoris	43
51	C-11	42	The Torulopsis candida candida utili	41

Embodiment 52, preparation lactobacillus acidophilus dry powder D-1

1) dull and stereotyped cultivation rejuvenation: with method streak inoculation on the lactic acid bacteria plating medium of lactobacterium acidophilus inclined-plane seed employing streak inoculation, in 33 ℃ of cultivations 24 hours, make the lactic acid bacteria rejuvenation, and form single bacterium colony; Described lactic acid bacteria plating medium is the mixture by following w/v (W/V) preparation: soy peptone 0.5%, glucose 1%, yeast soak powder 0.3%, potassium dihydrogen phosphate 0.3%, beef extract 0.5%, Triammonium citrate 1.5%, Tween-80 0.5%, sodium acetate 0.2%, agar powder 2%, pH=5.5～5.8,121 ℃ the sterilization 15 minutes, be cooled to 33 ℃ for subsequent use;

2) choose single bacterium colony in lactobacterium acidophilus's bacterium colony of making shake-flask seed: with step 1) cultivating, be inoculated in the lactic acid bacteria liquid culture medium, the employing conventional method in 35 ℃ of cultivations 24 hours, obtains the lactic acid bacteria shake-flask seed on the triangular flask fluid nutrient medium; Described culture medium is the mixed liquor by following w/v (W/V) preparation: soy peptone 0.5%, glucose 1%, yeast soak powder 0.3%, potassium dihydrogen phosphate 0.3%, beef extract 0.5%, Triammonium citrate 1.5%, Tween-80 0.5%, sodium acetate 0.2%, pH=5.5～5.8,121 ℃ the sterilization 15 minutes, be cooled to 35 ℃ for subsequent use;

3) the lactic acid bacteria shake-flask seed that production primary seed solution: with step 2) obtains is with the inoculum concentration of 6v%, be inoculated in the first order seed liquid culture medium of producing used first class seed pot, cultivate 17h in 35 ℃, mixing speed 150rpm, interval ventilation in 1 hour 5 minutes, throughput is 1vvm, until lactic acid bacteria number reaches 1,000,000,000, obtains the primary seed solution of lactic acid bacteria; Described first order seed liquid culture medium prescription is the mixed liquor by following w/v (W/V) preparation: 120 order bean cake powders 0.5%, 120 order fish meal 0.5%, fish peptone 0.5%, lactose 0.2%, 120 order corn flour 0.5%, glucose 1%, NaCl 0.1%, Na₂HPO ₄0.15%, synergist 0.05%, defoamer 0.03%, and pH=5.5～5.8 through 121 ℃ of sterilizations 30 minutes, are cooled to 35 ℃; Described synergist is by CaCl₂ 2g/L，ZnSO ₄ 0.5g/L，FeSO ₄ 0.2g/L， CuSO ₄ 0.01g/L，Co(NO ₃) ₂ 0.02g/L，MgSO ₄0.1g/L the aqueous solution that forms;

4) the lactic acid bacteria first order seed that second order fermentation (producing fermentation): with step 3) obtains, be inoculated in the fermentation tank that the production fermentation culture is housed with 5% volume ratio, prescription and the controlled condition of fermentation culture are same as step 3) first order seed liquid culture medium, ferment after 17 hours, when detecting the pH value and dropping to 5.0 left and right sides (at this moment, the result of test of many times shows that the lactic acid bacteria amount reaches greater than 10⁹The CFU/ml zymotic fluid) stops fermentation;

5) streptococcus acidi lactici fermented solution is added in the carrier that is pre-mixed, the weight ratio of zymotic fluid and carrier is 0.4: 1, and described carrier comprises: lightweight CaCO₃30wt%, inferior powder 40wt%, corn protein powder 30wt%, then through 45 ℃ of dryings of fluid bed, be crushed to more than 80 orders, obtain dry powder of lactic acid bacteria D-1, last, according to dull and stereotyped used culture medium and the condition of culture of rejuvenation of cultivating of lactic acid bacteria, viable bacteria content to dry powder carries out conventional plate count detection, according to count results, utilize described carrier viable bacteria content to be diluted to respectively 〉=400,000,000/g, for subsequent use.

Embodiment 53, preparation lactobacillus acidophilus dry powder D-2

1) dull and stereotyped cultivation rejuvenation: with method streak inoculation on the lactic acid bacteria plating medium of lactobacterium acidophilus inclined-plane seed employing streak inoculation, in 35 ℃ of cultivations 36 hours, make the lactic acid bacteria rejuvenation, and form single bacterium colony; Described lactic acid bacteria plating medium is the mixture by following w/v (W/V) preparation: soy peptone 0.8%, glucose 1.5%, yeast soak powder 0.5%, potassium dihydrogen phosphate 0.4%, beef extract 1%, Triammonium citrate 2%, Tween-80 1%, sodium acetate 0.4%, agar powder 2.2%, pH=5.5～5.8,121 ℃ the sterilization 15 minutes, be cooled to 35 ℃ for subsequent use;

2) choose single bacterium colony in lactobacterium acidophilus's bacterium colony of making shake-flask seed: with step 1) cultivating, be inoculated in the lactic acid bacteria liquid culture medium, the employing conventional method in 37 ℃ of cultivations 36 hours, obtains the lactic acid bacteria shake-flask seed on the triangular flask fluid nutrient medium; Described culture medium is the mixed liquor by following w/v (W/V) preparation: soy peptone 0.8%, glucose 1.5%, yeast soak powder 0.5%, potassium dihydrogen phosphate 0.4%, beef extract 1%, Triammonium citrate 2%, Tween-80 1%, sodium acetate 0.4%, pH=5.5～5.8, sterilized 15 minutes for 121 ℃, for subsequent use;

3) the lactobacterium acidophilus's shake-flask seed that production primary seed solution: with step 2) obtains is with the inoculum concentration of 10v%, be inoculated in the first order seed liquid culture medium of producing used first class seed pot, cultivate 36h in 37 ℃, mixing speed 180rpm, interval ventilation in 1 hour 8 minutes, throughput is 2vvm, until lactic acid bacteria number reaches 1,000,000,000, obtains lactobacterium acidophilus's primary seed solution; Described first order seed liquid culture medium prescription is the mixed liquor by following w/v (W/V) preparation: 120 order bean cake powders 1.5%, 120 order fish meal 0.8%, fish peptone 1.5%, lactose 1%, 120 order corn flour 0.8%, glucose 2%, NaCl 0.2%, Na₂HPO ₄0.25%, synergist 0.2%, defoamer 0.06%, and pH=5.5～5.8 through 121 ℃ of sterilizations 40 minutes, are cooled to 35 ℃; Described synergist is by CaCl₂ 4g/L，ZnSO ₄ 1.5g/L，FeSO ₄1.0g/L，CuSO ₄ 0.08g/L，Co(NO ₃) ₂ 0.06g/L，MgSO ₄0.5g/L the aqueous solution that forms;

4) the lactic acid bacteria first order seed that second order fermentation (producing fermentation): with step 3) obtains, be inoculated in the fermentation tank that the production fermentation culture is housed with 8% volume ratio, prescription and the controlled condition of fermentation culture are same as step 3) first order seed liquid culture medium, ferment after 48 hours, when detecting the pH value and dropping to 5.0 left and right sides (at this moment, the result of test of many times shows that the lactic acid bacteria amount reaches greater than 10⁹The CFU/ml zymotic fluid) stops fermentation;

5) streptococcus acidi lactici fermented solution is added in the carrier that is pre-mixed, the weight ratio of zymotic fluid and carrier is 0.7: 1, and described carrier comprises: lightweight CaCO₃60wt%, inferior powder 20wt%, corn protein powder 20wt%, then through 50 ℃ of dryings of fluid bed, be crushed to more than 80 orders, obtain dry powder of lactic acid bacteria D-2, last, according to dull and stereotyped used culture medium and the condition of culture of rejuvenation of cultivating of lactic acid bacteria, viable bacteria content to dry powder carries out conventional plate count detection, according to count results, utilize described carrier viable bacteria content to be diluted to respectively 〉=400,000,000/g, for subsequent use.

Embodiment 54, preparation lactobacillus acidophilus dry powder D-3

1) dull and stereotyped cultivation rejuvenation: with method streak inoculation on the lactic acid bacteria plating medium of lactobacterium acidophilus inclined-plane seed employing streak inoculation, in 38 ℃ of cultivations 48 hours, make the lactic acid bacteria rejuvenation, and form single bacterium colony; Described lactic acid bacteria plating medium is the mixture by following w/v (W/V) preparation: soy peptone 1%, glucose 2%, yeast soak powder 0.7%, potassium dihydrogen phosphate 0.5%, beef extract 1.5%, Triammonium citrate 2.3%, Tween-80 1.4%, sodium acetate 0.6%, agar powder 2.5%, pH=5.5～5.8,121 ℃ the sterilization 15 minutes, be cooled to 38 ℃ for subsequent use;

2) choose single bacterium colony in the lactic acid bacteria bacterium colony of making shake-flask seed: with step 1) cultivating, be inoculated in the lactic acid bacteria liquid culture medium, the employing conventional method in 40 ℃ of cultivations 48 hours, obtains the lactic acid bacteria shake-flask seed on the triangular flask fluid nutrient medium; Described culture medium is the mixed liquor by following w/v (W/V) preparation: soy peptone 1%, glucose 2%, yeast soak powder 0.7%, potassium dihydrogen phosphate 0.5%, beef extract 1.5%, Triammonium citrate 2.3%, Tween-80 1.4%, sodium acetate 0.6%, pH=5.5～5.8,121 ℃ the sterilization 15 minutes, be cooled to 40 ℃ for subsequent use;

3) the lactic acid bacteria shake-flask seed that production primary seed solution: with step 2) obtains is with the inoculum concentration of 12v%, be inoculated in the first order seed liquid culture medium of producing used first class seed pot, cultivate 60h in 38 ℃, mixing speed 200rpm, interval ventilation in 1 hour 10 minutes, throughput is 2.5vvm, until lactic acid bacteria number reaches 1,000,000,000, obtains the primary seed solution of lactic acid bacteria; Described first order seed liquid culture medium prescription is the mixed liquor by following w/v (W/V) preparation: 120 order bean cake powders 2.0%, 120 order fish meal 1.0%, fish peptone 2%, lactose 1.5%, 120 order corn flour 1.0%, glucose 2.5%, NaCl 0.3%, Na₂HPO ₄0.3%, synergist 0.3%, defoamer 0.1%, and pH=5.5～5.8 through 121 ℃ of sterilizations 45 minutes, are cooled to 38 ℃; Described synergist is by CaCl₂ 3g/L，ZnSO ₄ 1.0g/L，FeSO ₄ 0.7g/L， CuSO ₄ 0.04g/L，Co(NO ₃) ₂ 0.04g/L，MgSO ₄0.4g/L the aqueous solution that forms;

4) the lactic acid bacteria first order seed that second order fermentation (producing fermentation): with step 3) obtains, be inoculated in the fermentation tank that the production fermentation culture is housed with 10% volume ratio, prescription and the controlled condition of fermentation culture are same as step 3) first order seed liquid culture medium, ferment after 60 hours, when detecting the pH value and dropping to 5.0 left and right sides (at this moment, the result of test of many times shows that the lactic acid bacteria amount reaches greater than 10⁹The CFU/ml zymotic fluid) stops fermentation;

5) streptococcus acidi lactici fermented solution is added in the carrier that is pre-mixed, the weight ratio of zymotic fluid and carrier is 0.9: 1, and described carrier comprises: lightweight CaCO₃45wt%, inferior powder 30wt%, corn protein powder 25wt%, then through 60 ℃ of dryings of fluid bed, be crushed to more than 80 orders, obtain dry powder of lactic acid bacteria D-3, last, according to dull and stereotyped used culture medium and the condition of culture of rejuvenation of cultivating of lactic acid bacteria, viable bacteria content to dry powder carries out conventional plate count detection, according to count results, utilize described carrier viable bacteria content to be diluted to respectively 〉=400,000,000/g, for subsequent use.

Embodiment 55～75, preparation lactobacillus acidophilus dry powder D-4～D-24

Method according to embodiment 52～54 prepares dry powder of lactic acid bacteria D-4～D-24, and preparation condition is listed in table 4.

The preparation of table 4, dry powder of lactic acid bacteria D-4～D-24

Embodiment number	Dry powder of lactic acid bacteria	Preparation method (with embodiment X)	The used bacterial classification of preparation seed liquor	Carrier and post-treatment condition (with embodiment X)
					55	D-4	53	Lactobacillus bulgaricus	55
56	D-5	54	Lactobacillus casei	53
					57	D-6	55	Lactobacillus cellobiosas	54

58	D-7	53	Lactobacillus delbrueckii	55
					59	D-8	54	Lactobacillus fermenti	53
60	D-9	55	Lactobacillus lactis	54
					61	D-10	53	Lactobacillus plantarum	55
62	D-11	54	Streptococcus cremoris	53
					63	D-12	55	Streptococcus fecalis	54
64	D-13	53	Streptococcus lactis	55
					65	D-14	54	Streptococcus thermophilus	53
66	D-15	55	Pediococcus acidilactici	54
					67	D-16	53	Pediococcus cerevisiae	55
68	D-17	54	Streptococcus faecalis	53
					69	D-18	55	Lactobacillus	54
70	D-19	53	Lactobacillus thermophilus	55
					71	D-20	54	The lactobacillus Pediococcus acidilactici	53
72	D-21	55	The lactobacillus thermophilus streptococcus thermophilus	54
					73	D-22	53	The lactobacillus streptococcus fecalis	55
74	D-23	54	The lactobacillus thermophilus streptococcus cremoris	53
					75	D-24	55	The Lactobacillus casei pediococcus cerevisiae	54

Embodiment 76～91, the preparation little peptide of S-1 type of the present invention and the feed addictive S-1-1～S-1-16 of beneficial microbe symphysis unit

The bacillus dry powder blend of the little peptide dry powder of 40～60wt% and 40～60wt% is even, obtain the feed addictive S-1-1～S-1-16 of the little peptide of S-1 type of the present invention and beneficial microbe symphysis unit, its composition is as shown in table 5.

The quality standard of the little peptide that obtains and beneficial microbe symphysis unit feed addictive is: little peptide 100mg/g, bacillus living 〉=6,000,000,000/g is yellowish white powder, moisture content≤9%, ash≤8%, arsenic≤0.5mg/kg, lead≤1.0mg/kg.

The composition of table 5, the little peptide of S-1 type and beneficial microbe symphysis unit feed addictive

Embodiment	Little peptide dry powder A		Bacillus dry powder B			Feed addictive S-1
							Kind	Shared percentage by weight (%)	Kind	Used bacterial classification	Shared percentage by weight (%)
	76	A-1	40	B-1	Bacillus subtilis							60	S-1-1
77						A-2	50	B-2	Bacillus subtilis	50	S-1-2
	78	A-3	60	B-3	Bacillus subtilis							40	S-1-3
79						A-4	40	B-4	Bacillus coagulans	60	S-1-4
	80	A-5 A-24	50	B-5	Slow bacillus							50	S-1-5
81						A-6	60	B-6	Bacillus licheniformis	40	S-1-6
	82	A-7	40	B-7	Bacillus pumilus							60	S-1-7
83						A-8	50	B-8	Bacillus circulans	50	S-1-8
	84	A-9 A-23	60	B-9	Bacillus firmus							40	S-1-9
85						A-10  A-22	40	B-10	Bacillus megaterium	60	S-1-10
	86	A-11  A-19	50	B-11	Clostridium butyricum							50	S-1-11
87						A-12  A-18	60	B-12	Bafillus natto	40	S-1-12

88	A-13  A-17	40	B-13	Bacillus cercus	60	S-1-13
							89	A-14  A-15   A-16	50	B-14	The bafillus natto bacillus subtilis	50	S-1-14
90	A-13  A-17   A-21	60	B-15	The clostridium butyricum bacillus coagulans	40	S-1-15
							91	A-14  A-15   A-16  A-20	50	B-16	The bacillus licheniformis Bacillus circulans	50	S-1-16

Embodiment 92～107, the preparation little peptide of S-2 type of the present invention and the feed addictive S-2-1～S-2-16 of beneficial microbe symphysis unit

The saccharomycete dry powder blend of the bacillus dry powder of the little peptide dry powder of 35～45wt%, 20～30wt% and 30～40wt% is even, obtain the feed addictive S-2-1～S-2-16 of the little peptide of S-2 type of the present invention and beneficial microbe symphysis unit, its composition is as shown in table 6.

The quality standard of the little peptide that obtains and beneficial microbe symphysis unit feed addictive is: little peptide 100mg/g, bacillus living 〉=6,000,000,000/g, saccharomycete viable bacteria 〉=400,000,000/g, be yellowish white powder, moisture content≤9%, ash≤8%, arsenic≤0.5mg/kg, lead≤1.0mg/kg.

The composition of table 6, the little peptide of S-2 type and beneficial microbe symphysis unit feed addictive

Embodiment	Little peptide dry powder A	Bacillus dry powder B and used bacterial classification		Saccharomycete dry powder C	A: B: C weight ratio	Feed addictive S-2
							92	A-1	B-2	Bacillus coagulans	C-1	35∶30∶35	S-2-1
93	A-2	B-3	Slow bacillus	C-2	40∶20∶40	S-2-2
							94	A-3	B-1	Bacillus licheniformis	C-3	45∶25∶30	S-2-3
95	A-4	B-4	Bacillus coagulans	C-1	35∶30∶35	S-2-4
							96	A-5 A-24	B-5	Slow bacillus	C-2	40∶20∶40	S-2-5
97	A-6	B-6	Bacillus licheniformis	C-3	45∶25∶30	S-2-6
							98	A-7	B-7	Bacillus pumilus	C-4	35∶30∶35	S-2-7
99	A-8	B-8	Bacillus circulans	C-5	40∶20∶40	S-2-8
							100	A-9  A-23	B-9	Bacillus firmus	C-6	45∶25∶30	S-2-9
101	A-10  A-22	B-10	Bacillus megaterium	C-7	35∶30∶35	S-2-10
							102	A-11  A-19	B-11	Clostridium butyricum	C-8	40∶20∶40	S-2-11
103	A-12  A-18	B-12	Bafillus natto	C-9	45∶25∶30	S-2-12
							104	A-13  A-17	B-13	Bacillus cercus	C-10	35∶30∶35	S-2-13
105	A-14  A-15     A-16	B-14	The bafillus natto bacillus subtilis	C-11	40∶20∶40	S-2-14
							106	A-13  A-17     A-21	B-15	The clostridium butyricum bacillus coagulans	C-3	45∶25∶30	S-2-15
107	A-14  A-15     A-16  A-20	B-16	The bacillus licheniformis Bacillus circulans	C-4	35∶30∶35	S-2-16

Embodiment 108～123, the preparation little peptide of S-3 type of the present invention and the feed addictive S-3-1～S-3-16 of beneficial microbe symphysis unit

The little peptide dry powder of 30～50wt%, the bacillus dry powder of 20～40wt% and the dry powder of lactic acid bacteria of 30～50wt% are mixed, obtain the feed addictive S-3-1～S-3-16 of the little peptide of S-3 type of the present invention and beneficial microbe symphysis unit, its composition is as shown in table 7.

The quality standard of the little peptide that obtains and beneficial microbe symphysis unit feed addictive is: little peptide 100mg/g, bacillus living 〉=6,000,000,000/g, viable lactic acid bacteria 〉=400,000,000/g, be yellowish white powder, moisture content≤9%, ash≤8%, arsenic≤0.5mg/kg, lead≤1.0mg/kg.

The composition of table 7, the little peptide of S-3 type and beneficial microbe symphysis unit feed addictive

Embodiment	Little peptide dry powder A	Bacillus dry powder B	Dry powder of lactic acid bacteria D	A: B: D weight ratio	Feed addictive S-3
						108	A-1	B-1	D-1	30∶20∶50	S-3-1
109	A-2	B-2	D-3	30∶40∶30	S-3-2
						110	A-3	B-3	D-2	50∶20∶30	S-3-3
111	A-4	B-4	D-4	30∶20∶50	S-3-4
						112	A-5 A-24 equal portions	B-5	D-5	30∶40∶30	S-3-5
113	A-6	B-6	D-6	50∶20∶30	S-3-6
						114	A-7	B-7	D-7	30∶20∶50	S-3-7
115	A-8	B-8	D-8	30∶40∶30	S-3-8
						116	A-9 A-23 equal portions	B-9	D-9	50∶20∶30	S-3-9
117	A-10 A-22 equal portions	B-10	D-10	30∶20∶50	S-3-10
						118	A-11 A-19 equal portions	B-11	D-11	30∶40∶30	S-3-11
119	A-12 A-18 equal portions	B-12	D-12	50∶20∶30	S-3-12
						120	A-13 A-17 equal portions	B-13	D-13	30∶20∶50	S-3-13
121	A-14 A-15 A-16 equal portions	B-14	D-14	30∶40∶30	S-3-14
						122	A-13 A-17 A-21 equal portions	B-15	D-15	50∶20∶30	S-3-15
123	A-14 A-15 A-16 A-20 equal portions	B-16	D-16	30∶40∶30	S-3-16

Embodiment 124～139, the preparation little peptide of S-4 type of the present invention and the feed addictive S-4-1～S-4-16 of beneficial microbe symphysis unit

The little peptide dry powder of 30～40wt%, the bacillus dry powder of 20～30wt%, the saccharomycete dry powder of 10～20wt% and the dry powder of lactic acid bacteria of 20～30wt% are mixed, obtain the feed addictive S-4-1～S-4-16 of the little peptide of S-4 type of the present invention and beneficial microbe symphysis unit, its composition is as shown in table 8.

The quality standard of the little peptide that obtains and beneficial microbe symphysis unit feed addictive is: little peptide 100mg/g, bacillus living 〉=6,000,000,000/g, viable lactic acid bacteria 〉=400,000,000/g, saccharomycete viable bacteria 〉=400,000,000/g, be yellowish white powder, moisture content≤9%, ash≤8%, arsenic≤0.5mg/kg, lead≤1.0mg/kg.

The composition of table 8, the little peptide of S-4 type and beneficial microbe symphysis unit feed addictive

Embodiment	Little peptide dry powder A	Bacillus dry powder B	Saccharomycete dry powder C	Dry powder of lactic acid bacteria D	A: B: C: D weight ratio	Feed addictive S-4
							124	A-1	B-1	C-1	D-1	30∶30∶10∶30	S-4-1
125	A-2	B-2	C-3	D-3	35∶25∶20∶20	S-4-2
							126	A-3	B-3	C-2	D-2	40∶20∶15∶25	S-4-3
127	A-4	B-4	C-1	D-4	30∶30∶10∶30	S-4-4
							128	A-5  A-24	B-5	C-2	D-5	35∶25∶20∶20	S-4-5
129	A-6	B-6	C-3	D-6	40∶20∶15∶25	S-4-6
							130	A-7	B-7	C-4	D-7	30∶30∶10∶30	S-4-7
131	A-8	B-8	C-5	D-8	35∶25∶20∶20	S-4-8
							132	A-9 A-23	B-9	C-6	D-9	40∶20∶15∶25	S-4-9
133	A-10  A-22	B-10	C-7	D-10	30∶30∶10∶30	S-4-10

134	A-11  A-19	B-11	C-8	D-11	35∶25∶20∶20	S-4-11
							135	A-12  A-18	B-12	C-9	D-12 D-24	40∶20∶15∶25	S-4-12
136	A-13  A-17	B-13	C-10	D-13 D-23	30∶30∶10∶30	S-4-13
							137	A-14  A-15  A-16	B-14	C-11	D-14 D-20  D-21	35∶25∶20∶20	S-4-14
138	A-13  A-17  A-21	B-15	C-3	D-15  D-18  D-22	40∶20∶15∶25	S-4-15
							139	A-14  A-15  A-16  A-20	B-16	C-4	D-16 D-17  D-19	40∶20∶15∶25	S-4-16

The impact that embodiment 140, little peptide provided by the invention and beneficial microbe symphysis unit feed addictive are produced laying hen

Select extra large blue grey laying hen 900 plumages of 349～377 ages in days, be divided into 3 processed group: the blank group is not added any composition; Test 1 group and add 0.2% the symphysis feed addictive S-3-1 of unit of the present invention; Test 2 groups and add 0.2% the symphysis feed addictive S-4-1 of unit of the present invention; Each processes 6 repetitions, and each repeats 50 chickens. Butcher the situation of change of different intestinal segment lactic acid bacterias, Escherichia coli and total aerobic bacteria in the observation analysis laying hen enteron aisle after the raising. Result of the test is listed in table 9.

Table 9, laying hen gut flora separate counting and decide result [log₁₀(bacterium is counted the CFU/g intestinal contents)]

Grouping	Measure bacterium	Blank	The S-3-1 (0.2%) of symphysis unit	The S-4-1 (0.2%) of symphysis unit
					Crop	Lactic acid bacteria	8.3406±0.0560 Bb	9.5118±0.0095 Aa	9.5966±0.0078 Aa
Escherichia coli	8.3277±0.0685 Aa	7.2246±0.0276 Bb	7.3116±0.0150 Bb
				Total aerobic bacteria		9.0396±0.0560 A	7.9243±0.0000 C	8.0183±0.0187 B
Jejunum	Lactic acid bacteria	7.4512±0.0429 Bc	8.5450±0.0440 Ab						8.8663±0.0042 Aa
				Escherichia coli	8.6325±0.0086 Aa	7.5269±0.0036 Bb	7.4284±0.0057 Bc
	Total aerobic bacteria	8.7323±0.0113 B	8.0470±0.0035 C					7.8370±0.0035 A
				Caecum	Lactic acid bacteria	7.6430±0.0280 Bc	8.7522±0.0053 Ab		8.9319±0.0170 Aa
Escherichia coli	8.5050±0.0190 Aa	7.6284±0.0013 Bb	7.5246±0.0276 Bc
					Total aerobic bacteria	8.9085±0.0000 Aa	7.8892±0.0119 Bb	7.8834±0.0201 Bb
Rectum	Lactic acid bacteria	8.0050±0.0192 Bc	8.9370±0.0035 Ab						8.9823±0.0000 Aa
				Escherichia coli	8.1300±0.0228 Aa	7.6578±0.0202 Bb	7.3710±0.0131 Bc
	Total aerobic bacteria	8.8892±0.0119 Aa	7.9846±0.0032 Bb					7.3314±0.0429 Cc

Annotate: colleague's sample room is all to be indicated different capitalization persons and reaches the utmost point level of signifiance (P＜0.01) for difference;

Colleague's sample room is all to be indicated different lowercase persons and reaches the level of signifiance (P＜0.05) for difference; There is not difference without being labeled as in statistics.

As can be seen from Table 9: add after little peptide provided by the invention and the beneficial microbe symphysis unit feed addictive, lactic acid bacteria number in the crop has increased by 10～12 times than the obvious of blank group, Escherichia coli and total aerobic bacteria digital display work descend, and difference is (p＜0.01) extremely significantly; 10～15 times of the obvious increases of the lactic acid bacteria number contrast group in the jejunum, Escherichia coli and total aerobic bacteria digital display work descend, and difference is (p＜0.01) extremely significantly; Lactic acid bacteria number in the caecum obviously increases than blank group, and Escherichia coli and total aerobic bacteria digital display work descend more than 10 times (p＜0.01); Nearly 10 times of the obvious increase of the lactic acid bacteria number contrast group in the rectum, Escherichia coli and total aerobic bacteria digital display work descend more than 10 times, and difference is (p＜0.01) extremely significantly.

This result of the test explanation, when little peptide provided by the invention and beneficial microbe symphysis unit feed addictive is applied to laying hen and produces, can promote significantly and stimulate the growth of residing beneficial microbe-lactic acid bacteria in the laying hen intestines and stomach (comprising crop, jejunum, caecum and rectum) by the utmost point, quantity improves 10～15 times, and can extremely significantly suppress the growth and breeding of Escherichia coli and some other aerobic bacteria (such as salmonella, clostridium etc.), significantly descend more than 10 times, can regulate the animal and bird intestines colony balance, improve the laying hen premunition, be conducive to safeguard livestock birds health. In addition, in livestock and poultry cultivation, use the first feed addictive of symphysis provided by the invention can reduce antibiotic consumption, to reducing medicament residue, it also is favourable improving the animal products quality.

Embodiment 141, symphysis of the present invention unit feed addictive is on the impact of broiler chicken premunition

Select 500 of the Avain chickens that 1-28 age in days male and female raises together with, be divided at random 5 processing, be blank group, antibiotic control group (aureomycin 100g/t), add symphysis unit's S-1-1 group of the present invention (0.5kg/t), add S-2-1 group of the present invention (1kg/t), add S-4-2 group of the present invention (1kg/t), each processes 100. Brood after 1 week, each processing is divided into 5 repetitions, and each repeats 20. Butcher the situation of change (being the bacterium phase change) of different intestinal segment lactic acid bacterias, Escherichia coli, salmonella and total aerobic bacteria in the observation analysis Intestine of Broiler after the raising. Result of the test is listed in table 10.

Table 10, Intestine of Broiler flora separate counting and decide result [log₁₀(bacterium is counted the CFU/g intestinal contents]

Group	Measure microorganism	Ileum	Caecum	Rectum	Diarrhea rate %
						Blank group (not adding any material)	Escherichia coli	8.639±0.123 Aab	12.719±0.039 Aa	10.221±0.111 Aa	15.23±0.13 A
Salmonella	8.794±0.101 Aa	12.100±0.062 Aa	10.049±0.028 Aa
				Lactic acid bacteria	8.427±0.243 Cc		9.046±0.038 Cd	9.087±0.057 Cd
Total aerobic bacteria	10.430±0.016 Bb	12.712±0.019 Aa	11.095±0.082 Aa
				Antibiotic control group (adding aureomycin 100g/t feed)	Escherichia coli		8.599±0.087 Ab	11.350±0.042 Cc	9.732±0.064 Bb	5.87±0.11 B
Salmonella	8.747±0.103 Aa	11.812±0.093 Bb	9.453±0.068 Cc
					Lactic acid bacteria	8.462±0.039 Cc	9.129±0.021 Ccd	9.548±0.074 Bc
Total aerobic bacteria	10.787±0.010 Aa	11.753±0.090 Bb	10.809±0.075 Bb
					S-1-1 (adding the 0.5kg/t feed)	Escherichia coli	8.079±0.074 Bc	12.137±0.054 Bb	9.568±0.098 Bb		4.91±0.09 B
Salmonella	8.142±0.085 Bb	11.671±0.050 Bb	9.775±0.113 Bb
				Lactic acid bacteria		9.207±0.063 Bb	9.266±0.101 Cc	9.778±0.103 Bb
Total aerobic bacteria	9.697±0.055 Cc	11.417±0.102 Cc	8.688±0.081 Cc
				S-1-1 (adding the 1kg/t feed)		Escherichia coli	8.902±0.116 Aa	11.198±0.068 CDd	8.647±0.061 Cc	2.88±0.07 C
Salmonella	8.099±0.043 Bb	11.355±0.048 Cc	8.340±0.051 Dd
					Lactic acid bacteria	9.321±0.077 Bb	9.758±0.1051 Bb	10.312±0.021 Aa
Total aerobic bacteria	9.531±0.061 CDd	11.417±0.053 Cc	8.324±0.066 Dd
					S-4-2 (adding the 1kg/t feed)	Escherichia coli	7.590±0.072 Cd	11.117±0.028 Dd	7.551±0.037 Dd		2.13±0.7 C
Salmonella	7.767±0.027 Cc	10.545±0.022 Dd	8.328±0.043 Dd
				Lactic acid bacteria		10.055±0.058 Aa	10.333±0.048 Aa	10.378±0.033 Aa
Total aerobic bacteria	9.493±0.072 Dd	10.388±0.077 Dd	8.044±0.035 Ee

Annotate: colleague's sample room is all to be indicated different capitalization persons and reaches the utmost point level of signifiance (P＜0.01) for difference;

Colleague's sample room is all to be indicated different lowercase persons and reaches the level of signifiance (P＜0.05) for difference; There is not difference without being labeled as in statistics.

As can be seen from Table 10: add the increase that the first feed addictive of symphysis of the present invention can make the lactic acid bacteria number contrast group in the ileum, coliform count and salmonella are counted 4～10 times of the reductions of contrast group, total aerobic bacteria number descends 5～8 times than control group, and difference is (p＜0.01) extremely significantly; 2～19 times of the increases of the lactic acid bacteria number contrast group in the caecum, coliform count, salmonella number and total aerobic bacteria number reduce than control group, and difference is (p＜0.01) extremely significantly; The remarkable increase of the lactic acid bacteria number contrast group in the rectum, coliform count, salmonella number and total aerobic bacteria number reduce than control group, and difference is (p＜0.01) extremely significantly. Simultaneously, in raising, the meat chick use symphysis of the present invention first, its diarrhea rate extremely significantly is lower than blank group and antibiotic control group, difference is analysed significantly (p＜0.01), illustrate that little peptide of the present invention and beneficial microbe symphysis unit feed addictive can effectively prevent and treat the diarrhoea of meat chick, can replace antibiotic fully.

Result of the test shows, can promote significantly and stimulate the growth of residing beneficial microbe-lactic acid bacteria in the broiler chicken intestines and stomach (comprising ileum, caecum and rectum) by the utmost point when little peptide provided by the invention and beneficial microbe symphysis unit feed addictive are applied to broiler production, quantity improves 20～40 times; And can extremely significantly suppress the growth and breeding of Escherichia coli and some other aerobic bacteria (such as salmonella, clostridium etc.), make harmful bacteria reduce more than 10 times, regulate the Intestine of Broiler colony balance, improve the broiler chicken premunition, diarrhea rate extremely significantly descends, be conducive to safeguard livestock birds health, fully substitute antibiotics.

Embodiment 142, symphysis provided by the invention unit feed addictive is on the impact of performance in layers, Egg Quality, death rate and antibody horizontal

With identical 18000 of the blue laying hens in later stage sea of laying eggs of age in days, minutes 6 groups, one group 3000 of every buildings, minute 3 repetitions, each repeats 1000. The grouping situation is as follows:

The A group, blank group, the laying hens essential ration of feeding; The B group is added terramycin in laying hens essential ration, 80g/t is the antibiotic blank; The C group is added little peptide of the present invention and the feed addictive S-1-10 of beneficial microbe symphysis unit, 1kg/t in laying hens essential ration; The D group is added little peptide of the present invention and the feed addictive S-2-16 of beneficial microbe symphysis unit, 1kg/t in laying hens essential ration; The E group is added little peptide of the present invention and the feed addictive S-3-11 of beneficial microbe symphysis unit, 1kg/t in laying hens essential ration; The F group is added little peptide of the present invention and the feed addictive S-4-8 of beneficial microbe symphysis unit, 1kg/t in laying hens essential ration.

Employed laying hens essential ration prescription (percentage by weight) is: corn 57.74%, dregs of beans 25.8%, wheat bran 3.0%, stone flour 8.42%, hydrogen calcium 1.53%, salt 0.33%, vegetable oil 2.0%, 1% premix 1.0%.

Front 7 days is preliminary trial period, and rear 28 days is experimental period, and presses above-mentioned grouping repeated test 3 times, has measured NH in the house₃、 H ₂The indexs such as S concentration, broken rate of eggshell, albumen quality (Hough unit), laying rate, egg size, death rate, egg-shell gloss cleanliness, eggshell color and luster, antibody horizontal. The results are shown in table 11.

Table 11, on the impact of performance in layers, Egg Quality, death rate

Later stage laying rate of chicken (%)	80.2	83.5	84.6	84.7	84.9	85.7
							Hough unit	78.0	82.2	85.3	84.5	85.9	86.6
Egg size (g)	65.0	66.0	67.3	67.1	66.9	67.7
							The egg-shell gloss cleanliness*	++	+++	++++	++++	++++	++++
Death rate %	3.9	1.9	1.5	1.7	1.6	1.2
							**Ewcastle disease virus antibody (ELISA, OD value)	0.77	0.75	1.31	1.36	1.48	1.43

^*The egg-shell gloss cleanliness with+how much represent ,+more multilist shows the egg-shell gloss cleanliness better.^**BSA is bovine serum albumin(BSA).

As can be seen from Table 11, the laying rate of the laying hen of each group of the little peptide of the difference provided by the invention of feeding and beneficial microbe symphysis unit's feed addictive feed addictive (S-1-10, S-2-16, S-3-11, S-4-8), egg size, Hough unit, egg-shell gloss cleanliness, antibody horizontal are all significantly better than the indices of control group and antibiotic control group, use simultaneously death rate, the broken rate of eggshell of the laying hen of little peptide and beneficial microbe symphysis unit each group of feed addictive, ammonia and the concentration of hydrogen sulfide of hen house all to be starkly lower than control group and antibiotic group, and can to replace antibiotic fully.

The above results explanation, little peptide provided by the invention and beneficial microbe symphysis unit feed addictive can not only significantly improve production performance, Egg Quality, the environmental quality of laying hen, and can obviously improve antibody horizontal and reduce death rate, and significantly better than blank and antibody element control group. To be little peptide contain little peptide and different beneficial microbe from beneficial microbe symphysis unit feed addictive to its reason, can not only replenish or inoculate beneficial microbe and bring into play exogenous beneficial microorganism effect (as producing the benefit materials such as digestive ferment, antibacterial material, multivitamin to animal like this, the functions such as performance digestion, immune stimulatory and diseases prevention), but also contain little peptide (being a kind of prebiotics) because of little peptide and beneficial microbe symphysis unit feed addictive, can activate endogenic profitable strain in the animal gastrointestinal tract, and then the functions such as enzymatic digestion, immune stimulatory and diseases prevention are produced in reinforcement, thereby, little peptide and beneficial microbe are had complementary advantages, promote secretory volume, the raising digestive enzyme activity of microorganism digestive ferment, improve absorption conversion and the utilization ratio of feed nutrition element, and the ammonia in the reduction pouity dwelling place and hydrogen sulfide content, produce good economic benefit, ecological benefits and, social benefit.

Embodiment 143～146, little peptide provided by the invention and beneficial microbe symphysis unit feed addictive are on the impact on piglet, growing and fattening pigs, broiler chicken and grass carp, and the effect of substitute antibiotics

Table 12, to the effect in piglet, growing-finishing pig, meat chick and the grass carp cultivation

Instance number	Symphysis unit	Test method	Experimental scale	Result of the test
					Embodiment 143	A group: contrast B group: S-1-2 C group: S-1-3 D group: S-1-14 E group: S-1-15 F group: S-1-16 G group: S-1-12	With embodiment 144, A group adds flavomycoin 50g/t, aureomycin 100g/t contrast, and the additive of B, C, D, E, F, G group is as described in the left hurdle, each 1kg/t, every group of 30 pigs, 3 repetitions.	210 of piglets	Contrast with adding the antibiosis group, the indices that adds the symphysis tuple improves as follows: daily ingestion amount raising 9.2%～11.7% daily gain improves 11.2%～12.3% feed efficiency improvement, 11.9%～13.8% diarrhea rate and reduces by 59.1%～60.5%

Embodiment 144	A group: contrast B group: S-2-1 C group: S-2-2 D group: S-2-3 E group: S-2-4 F group: S-2-5	With example 144, the A group adds aureomycin 100g/t, sulfanilamide (SN) 100g/t contrast, and B, C, D, E, F group are added benefit-8,10,14,15,16 each 1.5kg/t, every group of 60 pigs, 6 repetitions.	Breed 420 of growing and fattening pigs (30-90kg)	Each growth phase feed intake of fattening pig, daily gain; Feed efficiency improves, and diarrhea rate and death rate reduce, and every production performance is not less than adds antibiotic colony; The pig house stink reduces, and dressing percentage is similar or higher to interpolation antibiotic colony with lean meat percentage, and pork color is ruddy, and is Fresh ﹠ Tender in Texture.
					Embodiment 145	Chicken: A group: contrast B group: S-3-2 C group: S-3-3 D group: S-3-4 E group: S-3-5 F group: chicken among the S-3-6: A group: contrast B group: S-2-6 C group: S-2-7 D group: S-2-8 E group: S-2-9 F group: S-2-10	Large chicken: A group: contrast B group: S-4-3 C group: S-4-4 D group: S-4-5 E group: S-4-6 F group: each symphysis unit addition of S-4-7 control group terramycin 150g/t is: the large chicken 1.0kg/t of chicken 1.0kg/t feed granulating among the chicken 1.5kg/t, every group of 4000 broiler chicken, 4 repetitions, each repeats 1000, totally 24000	24000 of broiler chicken	Increase the deposition of chicken body internal protein and reduce abdomen fat rate (P＜0.05), obviously reduce pH value (p＜0.05 of duodenum, jejunum and ileum,), significantly suppress the E.Coli breeding in (minimizing) jejunum and the rectum, and can improve the content (p＜0.05) of Bacillus acidi lactici in the enteron aisle, extremely be conducive to the health of chicken and reduce stink.
Embodiment 146	A group: contrast B group: S-4-14 C group: S-4-9 D group: S-4-10 E group: S-4-11 F group: S-4-12 G group: S-4-13	Control group feed per ton adds terramycin and each 100g test group interpolation symphysis unit amount of sulfanilamide (SN) is feed per ton: the 1.0kg/t feed granulating, and every group of 3000 grass carps, 3 repetitions, each repeats 1000, totally 21000	21000 finishing period grass carps	Feed coefficient contrast group improves 6.9%, and daily gain contrast group improves 12.7%, and survival rate improves 22.6%, and the regularity of test group is good, and resistance against diseases is strong. Water quality control aspect, 3.0 meters, 3 mu inputs of the depth of water symphysis 1kg of unit. Ammoniacal nitrogen (NH as a result3-N) in 24 hours, be reduced to the 0.21mg/L of test group by the 0.72mg/L of control group, reduce by 3 times, the white skin disease of Tilapia mossambica reduces 66.2% simultaneously.

As can be seen from Table 12, feed little peptide provided by the invention and beneficial microbe symphysis the unit feed addictive, the indices of piglet, the difference raising stage of breeding pig, meat chick and grass carp is all significantly better than different antibiotic control groups, illustrate that little peptide provided by the invention and beneficial microbe symphysis unit feed addictive can effectively be prevented and treated livestock and poultry and aquatic animal disease, the effects such as (weightening finish, feed efficiency etc.), the water quality of improving animal house environment and aquatic livestock, raising animal product quality that significantly improve production performance, further proof little peptide provided by the invention and beneficial microbe symphysis unit feed addictive can be used as Substitutes For Antibiotic, and has good economy and ecological benefits.

Embodiment 147, little peptide provided by the invention and beneficial microbe symphysis unit feed addictive are on the immunity (comprising cellular immunity and humoral immunity) of meat chick or the impact of premunition

For checking little peptide provided by the invention and beneficial microbe symphysis unit feed addictive in the immunoenhancement result on the broiler chicken and the application prospect in animal husbandry, Avain chicken 1 age in days 900 plumages are selected in test, divide 9 groups, every group of 100 broiler chicken, divide 5 repetitions, each repeats 20 chickens, feed with 9 kinds of different daily rations, be that the A group is the blank group, the B group is the antibiotic group, and C, D, E, F, G, H, I group are added respectively different symphysis each 1kg of (as shown in table 13) feed addictive of unit of the present invention. Respectively at 7 ages in days and 14 age in days injection of bovine serum albumin antigens, in order to detecting amynologic index. In 7 weeks of examination phase, 3 weeks were detected lymphocyte proliferation vigor, bovine serum albumin(BSA) antibody horizontal during age, the results are shown in Table 13, table 14.

Table 13, symphysis unit are on the impact of chicken blood lymphocyte proliferation activity (cellular immunity)

Additive	Blank (not adding)	The antibiotic contrast (enramycin, 5mg/kg)	S-1-4	S-1-5	S-1-6	S-2-11	S-2-13	S-3-11	S-3-16
										T cell viability (OD value)	2.362	2.412	2.320	2.654	2.872	2.911	2.865	2.798	3.011

Table 14, symphysis unit are on the impact of chicken blood bovine serum albumin(BSA) antibody titer (humoral immunity)

Additive	Blank (not adding)	Antibiotic (peace rice mycin, 5mg/kg)	S-1-4	S-1-5	S-1-6	S-2-11	S-2-13	S-3-11	S-3-16
										Antibody titer (OD value)	1.112	1.235	2.725	3.546	3.595	4.563	4.342	3.329	3.679

As shown in Table 13, little peptide provided by the invention and the interpolation of beneficial microbe symphysis unit's feed addictive in feed can obviously strengthen the T cell viability, illustrate that little peptide provided by the invention and beneficial microbe symphysis unit feed addictive can obviously improve the cellular immune function of animal.

As shown in Table 14, interpolation in feed little peptide provided by the invention and beneficial microbe symphysis unit feed addictive can obviously strengthen the antibody titer level, illustrate that little peptide provided by the invention and beneficial microbe symphysis unit feed addictive can obviously improve the humoral immune function of animal.

The result of the test of consolidated statement 13, table 14, little peptide provided by the invention is added in explanation in animal feed and beneficial microbe symphysis unit feed addictive can obviously improve the immunity (comprising cellular immunity and humoral immunity) of animal, and then strengthens the premunition of animal.

Comprehensive above all examples, illustrate that little peptide provided by the invention and beneficial microbe symphysis unit feed addictive can be used as Substitutes For Antibiotic, eliminate medicament residue, improve the product quality such as meat, eggs and milk, can be used for preventing and treating the various animals diarrhoea of having loose bowels, improve breeding performonce fo animals, raising immunity or premunition and feed efficiency, reduction animal death rate, also can be used for making ensiling, yellow storage feed and biological feedstuff, cotton dregs (cake) and the dish dregs of rice (cake) detoxification purify mew environment (ammonia falls in deodorizing) and purify fishes and shrimps pond water quality etc.

Claims (11)

1, a kind of little peptide and beneficial microbe symphysis unit feed addictive comprises little peptide dry powder and bacillus dry powder.

2, little peptide as claimed in claim 1 and beneficial microbe symphysis unit feed addictive also comprises saccharomycete dry powder or/and dry powder of lactic acid bacteria.

3, little peptide as claimed in claim 1 and beneficial microbe symphysis unit feed addictive is comprised of little peptide dry powder and bacillus dry powder, and wherein, little peptide dry powder accounts for 40～60wt%; Bacillus dry powder accounts for 40～60wt%.

4, little peptide as claimed in claim 2 and beneficial microbe symphysis unit feed addictive is comprised of little peptide dry powder, bacillus dry powder and saccharomycete dry powder, and wherein, little peptide dry powder accounts for 35～45wt%; Bacillus dry powder accounts for 20～30wt%; Saccharomycete dry powder accounts for 30～40wt%.

5, little peptide as claimed in claim 2 and beneficial microbe symphysis unit feed addictive is comprised of little peptide dry powder, bacillus dry powder and dry powder of lactic acid bacteria, and wherein, little peptide dry powder accounts for 30～50wt%; Bacillus dry powder accounts for 20～40wt%; Dry powder of lactic acid bacteria accounts for 30～50wt%.

6, little peptide as claimed in claim 2 and beneficial microbe symphysis unit feed addictive is comprised of little peptide dry powder, bacillus dry powder, saccharomycete dry powder and dry powder of lactic acid bacteria, and wherein, little peptide dry powder accounts for 30～40wt%; Bacillus dry powder accounts for 20～30wt%; Saccharomycete dry powder accounts for 10～20wt%; Dry powder of lactic acid bacteria accounts for 20～30wt%.

7, little peptide as claimed in claim 1 and beneficial microbe symphysis unit feed addictive, described little peptide dry powder, with vegetable protein or/and animal protein is degraded through microbial fermentation, and its degree of hydrolysis is controlled at and obtains after 25～40% the zymotic fluid drying, and concrete preparation method comprises the steps:

1) fermentation seed liquid preparation: will be selected from one or more the mixture in aspergillus niger, aspergillus oryzae, aspergillus terricola bacterium, bacillus subtilis, bafillus natto, the bacillus pumilus, access was through 121 ℃ of sterilizations 30 minutes and be cooled in 28～37 ℃ the fluid nutrient medium, cultivated 24～48 hours, and obtained seed liquor for 28～37 ℃; Described fluid nutrient medium is the mixed liquor in following ratio preparation: peptone 1.3～2.5g, glucose 3～5g, molasses 3～5g, potato immersion liquid 50～100ml, soluble starch 10～20g, sucrose 10～30g, beef extract 2～3g, yeast soak powder 3.5～5g, dregs of beans 1.5～2g, growth factor 1～2ml, water 1000ml; Growth factor wherein is for containing MgSO₄·7H ₂O 20.00～30.00mg/ml， CaCl ₂.2H ₂O 2.20～3.50mg/ml，ZnSO ₄.7H ₂O 0.81～1.10mg/ml，FeSO ₄.7H ₂O 0.44～0.60mg/ml， MnSO ₄.H ₂O 0.13～0.20mg/ml，CuSO ₄.5H ₂The mixture of O 0.02～0.04mg/ml;

2) fermentative degradation: one or more the mixture in vegetable protein or the animal protein is crushed to 60～120 orders, pressing protein content calculates, according to material: flowing fluid ratio=5～18: 100 ratio is put into fermentation tank, put into carbon source according to the ratio of C/N ratio=1/2～1/35 again, and adding defoamer 0.03～0.1%, mixing, the nutrient solution that obtains was in 121～125 ℃ of autoclavings 10～30 minutes, be cooled to 28～37 ℃, take the volume ratio of seed liquor and nutrient solution as 0.3～1: the seed liquor that 10 ratio access step 1) obtains, 28～42 ℃ of fermentations, throughput is 0.5～2.5vvm, mixing speed 150～300rpm, use during this time conventional ninhydrin method determination of color degree of hydrolysis, detected a degree of hydrolysis in per 6 hours before 24 hours, between 24～48 hours, degree of hydrolysis of detection in per 4 hours, degree of hydrolysis of detection in per 2 hours after 48 hours, until degree of hydrolysis reaches 25～40%, then in 140～145 ℃ of sterilizations 2～8 seconds, obtain containing the protein hydrolyzate of little peptide;

3) protein hydrolyzate that contains little peptide that centrifugal slagging-off: with step 2) obtains centrifugal 10～30 minutes in 5000～7000rpm, remove slag, the supernatant that obtains is that the ultrafilter of 1000Da is removed the impurity such as high molecular weight protein by retaining molecular weight again, obtains the filtrate of molecular weight≤1000Da; This filtrate is concentrated to 50% of original volume through negative-pressure vacuum, the concentrate that obtains is added in the carrier that is pre-mixed, and presses concentrate: carrier=0.5～0.8: 1 envelope-bulk to weight ratio mixes, and it is rear 45～80 ℃ of dryings to stir, then be crushed to more than 80 orders, obtain little peptide dry powder; Described step 3) carrier is comprised of 2～5 weight portion organic carriers and 3～5 weight portion inorganic carriers; Wherein, described organic carrier comprises bran and puffed soybean organic carrier; Described inorganic carrier is one or more the mixture in precipitated calcium carbonate, stone flour, medical stone and the zeolite powder.

8, little peptide as claimed in claim 1 and beneficial microbe symphysis unit feed addictive, described bacillus dry powder is to be made by following method:

1) dull and stereotyped cultivation rejuvenation: with method streak inoculation on the bacillus plating medium of bacillus bacterial classification employing streak inoculation, in 25～35 ℃ of cultivations 12～24 hours, make the bacillus rejuvenation, and form single bacterium colony; Described bacillus plating medium is the mixture by following w/v preparation: peptone 0.5～1%, glucose 0.5～2%, beef extract 0.5～1.5%, potassium dihydrogen phosphate 0.3～0.6%, agar 3～4%, pH=7.5～7.6, sterilized 30 minutes for 121 ℃, for subsequent use; Described bacillus comprises: bacillus coagulans, slow bacillus, bacillus subtilis, bacillus licheniformis, bacillus pumilus, Bacillus circulans, bacillus firmus, bacillus megaterium, clostridium butyricum, bafillus natto, Bacillus cercus;

2) choose single bacterium colony the bacillus bacterium colony of making shake-flask seed: from step 1) cultivating, be inoculated in the bacillus liquid culture medium, in 28～37 ℃ of cultivations 20～30 hours, obtain the bacillus shake-flask seed liquid; Described bacillus liquid culture medium is the mixed liquor by following w/v preparation: peptone 0.5～1%, glucose 0.5～2%, beef extract 0.5～1.5%, potassium dihydrogen phosphate 0.3～0.6%, 3.08% manganese sulfate solution 0.5～1.5%, pH=7.5～7.6, sterilized 30 minutes for 121 ℃, for subsequent use;

3) the bacillus shake-flask seed liquid that production primary seed solution: with step 2) obtains is with the inoculum concentration of 2～5v%, be inoculated in the first order seed liquid culture medium of producing used first class seed pot, cultivate 18～24h in 28～37 ℃, mixing speed 150～200rpm, throughput is 1～2vvm, obtains the primary seed solution of bacillus; Described first order seed liquid culture medium prescription is the mixed liquor by following w/v preparation: 120 order bean cake powders 0.5～2.0%, 120 order fish meal 0.2～1.5%, 120 order corn flours 0.5～1.0%, glucose 1.0～4.5%, NaCl 0.1～0.3%, NaOH 0.05～0.10%, Na₂HPO ₄0.1～0.5%, synergist 0.05～0.2%, defoamer 0.05～0.1%, and pH=7.5～7.6 through 121 ℃ of sterilizations 30～45 minutes, are cooled to 32～37 ℃; Described synergist is by MgSO₄ 0.1～0.5g/L，CaCl ₂ 1.0～ 4.0g/L，ZnSO ₄ 0.2～1.5g/L，FeSO ₄ 0.1～0.8g/L，CuSO ₄ 0.01～0.05g/L，Co(NO ₃) ₂ 0.02～ 0.06g/L，MnSO ₄0.3 the aqueous solution that～1.5g/L forms;

4) the bacillus primary seed solution that second order fermentation: with step 3) obtains, volume ratio with 5～10% is inoculated into and is equipped with in the fermentation tank of producing fermentation culture, prescription and the controlled condition of fermentation culture are same as step 3) first order seed liquid culture medium, ferment after 16～24 hours, by microscopic examination, when being arranged, a large amount of sporulation stops fermentation;

5) with the zymotic fluid of bacillus, be that 0.5～0.8: 1 ratio mixes according to the weight ratio of zymotic fluid and carrier, be the wet-milling shape, then on fluidizing drying bed in 80～100 ℃ of crushed after being dried to more than 80 orders, obtain bacillus dry powder, at last according to dull and stereotyped used culture medium and the condition of culture of rejuvenation of cultivating of bacillus, the viable bacteria content of dry powder is carried out conventional plate count detect, according to count results, utilize carrier viable bacteria content to be diluted to respectively 〉=6,000,000,000/g; Described carrier proportioning is: lightweight CaCO₃30～60wt%, inferior powder 20～40wt%, corn protein powder 20～30wt%.

9, little peptide as claimed in claim 2 and beneficial microbe symphysis unit feed addictive, described saccharomycete dry powder is to be made by following method:

1) dull and stereotyped cultivation rejuvenation: with method streak inoculation on the yeast plating medium of yeast inclined-plane seed employing streak inoculation, in 24～30 ℃ of cultivations 20～24 hours, make the yeast rejuvenation, and form single bacterium colony; Described yeast plating medium is the mixture by following w/v preparation: soy peptone 0.2～0.7%, glucose 0.5～1.5%, yeast soak powder 0.1～0.5%, malt extract 0.2～1%, agar powder 2～3%, pH=5.8～6.0,121 ℃ the sterilization 15 minutes, be cooled to 24～30 ℃ for subsequent use; Described yeast comprises: saccharomyces cerevisiae or brewer's yeast, rhodothece rubra, red phaffia rhodozyma, Torulopsis candida, Pichia pastoris, Chinese red yeast, candida utili;

2) choose single bacterium colony the Yeast Cultivation flat board of making shake-flask seed: from step 1) cultivating, be inoculated in the triangular flask liquid medium, in 28～32 ℃ of cultivations 24～37 hours, obtain the yeast shake-flask seed liquid; Described nutrient solution is the mixed liquor by following w/v preparation: soy peptone 0.2～0.7%, glucose 0.5～1.5%, yeast soak powder 0.1～0.5%, malt extract 0.2～1%, pH=5.8～6.0,121 ℃ the sterilization 15 minutes, be cooled to 28～32 ℃ for subsequent use;

3) the yeast shake-flask seed that production primary seed solution: with step 2) obtains is by the inoculum concentration of 5～12v%, be inoculated in the first order seed nutrient solution of first class seed pot, cultivated 24～37 hours in 28～37 ℃, mixing speed 160～220rpm, throughput is 1.0～3.0vvm, obtains the primary seed solution of yeast; Described yeast first order seed nutrient solution is the mixed liquor that connects the preparation of following weight volume ratio: 120 order bean cake powders 0.5～1.5%, 120 order fish meal 0.2～0.8%, 120 order corn flours 0.5～1.0%, glucose 0.3～1.0%, NaCl 0.1～0.3%, Na₂HPO ₄0.2～0.5%, synergist 0.05～0.2%, defoamer 0.03～0.1%, and pH=5.8～6.0 through 121 ℃ of sterilizations 30～45 minutes, are cooled to 28～37 ℃; Described synergist is by MgSO₄ 0.05～0.3g/L，CaCl ₂ 2.5～4g/L，ZnSO ₄ 0.6～1.5g/L，FeSO ₄ 0.1～1.0g/L， MnSO ₄ 0.4～0.8g/L，CuSO ₄0.02 the aqueous solution that～0.08g/L forms;

4) the yeast first order seed that second order fermentation: with step 3) obtains, be inoculated in the fermentation tank that the production fermentation culture is housed with 4～10% volume ratios, prescription and the controlled condition of fermentation culture are same as step 3) the first order seed nutrient solution, ferment after 24～60 hours, with step 1) culture medium, detect with dull and stereotyped meter method or counting method of blood cell and to stop fermentation when yeast count reaches 1,000,000,000/ml;

5) above-mentioned saccharomycetic zymotic fluid is added in the carrier that is pre-mixed, the weight ratio of zymotic fluid and carrier is 0.6～0.9: 1, fluidized bed drying, 45～60 ℃ of baking temperatures obtain saccharomycete dry powder, then according to dull and stereotyped used culture medium and the condition of rejuvenation of cultivating of yeast, viable bacteria content to dry powder carries out conventional plate count detection, according to count results, utilize above-mentioned carrier viable bacteria content to be diluted to 〉=400,000,000/g, for subsequent use; Described carrier proportioning is: lightweight CaCO₃30～60wt%, inferior powder 20～40wt%, corn protein powder 20～30wt%.

10, little peptide as claimed in claim 2 and beneficial microbe symphysis unit feed addictive, described dry powder of lactic acid bacteria is to be made by following method:

1) dull and stereotyped cultivation rejuvenation: with method streak inoculation on the lactic acid bacteria plating medium of lactic acid bacteria inclined-plane seed employing streak inoculation, in 33～38 ℃ of cultivations 24～48 hours, make the lactic acid bacteria rejuvenation, and form single bacterium colony; Described lactic acid bacteria plating medium is the mixture by following w/v preparation: soy peptone 0.5～1%, glucose 1～2%, yeast soak powder 0.3～0.7%, potassium dihydrogen phosphate 0.3～0.5%, beef extract 0.5～1.5%, Triammonium citrate 1.5～2.3%, Tween-80 0.5～1.4%, sodium acetate 0.2～0.6%, agar powder 2～2.5%, pH=5.5～5.8,121 ℃ the sterilization 15 minutes, be cooled to 33～38 ℃ for subsequent use; Described lactic acid bacteria comprises: lactobacillus acidophilus, lactobacillus bulgaricus, Lactobacillus casei, lactobacillus cellobiosas, Lactobacillus delbrueckii, lactobacillus fermenti, lactobacillus lactis, Lactobacillus plantarum, streptococcus cremoris, streptococcus fecalis, streptococcus lactis, streptococcus thermophilus, Pediococcus acidilactici, pediococcus cerevisiae, streptococcus faecalis, lactobacillus, lactobacillus thermophilus;

2) choose single bacterium colony in the lactic acid bacteria bacterium colony of making shake-flask seed: with step 1) cultivating, be inoculated in the lactic acid bacteria liquid culture medium, the employing conventional method in 35～40 ℃ of cultivations 24～48 hours, obtains the lactic acid bacteria shake-flask seed on the triangular flask fluid nutrient medium; Described culture medium is the mixed liquor by following w/v preparation: soy peptone 0.5～1%, glucose 1～2%, yeast soak powder 0.3～0.7%, potassium dihydrogen phosphate 0.3～0.5%, beef extract 0.5～1.5%, Triammonium citrate 1.5～2.3%, Tween-80 0.5～1.4%, sodium acetate 0.2～0.6%, pH=5.5～5.8, sterilized 15 minutes for 121 ℃, for subsequent use;

3) the lactic acid bacteria shake-flask seed that production primary seed solution: with step 2) obtains is with the inoculum concentration of 6～12v%, be inoculated in the first order seed liquid culture medium of producing used first class seed pot, cultivate 17～60h in 35～38 ℃, mixing speed 150～200rpm, interval ventilation in 1 hour 5～10 minutes, throughput is 1～2.5vvm, until lactic acid bacteria number reaches 1,000,000,000, obtains the primary seed solution of lactic acid bacteria; Described first order seed liquid culture medium prescription is the mixed liquor by following w/v preparation: 120 order bean cake powders 0.5～2.0%, 120 order fish meal 0.5～1.0%, fish peptone 0.5～2%, lactose 0.2～1.5%, 120 order corn flours 0.5～1.0%, glucose 1～2.5%, NaCl 0.1～0.3%, Na₂HPO ₄0.15～0.3%, synergist 0.05～0.3%, defoamer 0.03～0.1%, and pH=5.5～5.8 through 121 ℃ of sterilizations 30～45 minutes, are cooled to 32～37 ℃; Described synergist is by CaCl₂ 2～4g/L，ZnSO ₄ 0.5～1.5g/L，FeSO ₄ 0.2～1.0g/L， CuSO ₄ 0.01～0.08g/L，Co(NO ₃) ₂ 0.02～0.06g/L，MgSO ₄0.1 the aqueous solution that～0.5g/L forms;

4) the lactic acid bacteria first order seed that second order fermentation: with step 3) obtains, be inoculated in the fermentation tank that the production fermentation culture is housed with 5～10% volume ratios, prescription and the controlled condition of fermentation culture are same as step 3) first order seed liquid culture medium, ferment after 17～60 hours, detection pH value stops fermentation when dropping to 5.0 left and right sides;

5) streptococcus acidi lactici fermented solution is added in the carrier that is pre-mixed, the weight ratio of concentrate and carrier is 0.4～0.9: 1, then through 45～60 ℃ of dryings of fluid bed, is crushed to more than 80 orders, obtains dry powder of lactic acid bacteria; At last, according to dull and stereotyped used culture medium and the condition of culture of rejuvenation of cultivating of lactic acid bacteria, the viable bacteria content of dry powder is carried out conventional plate count detect, according to count results, utilize described carrier viable bacteria content to be diluted to respectively 〉=400,000,000/g, for subsequent use; Described carrier proportioning is: lightweight CaCO₃30～60wt%, inferior powder 20～40wt%, corn protein powder 20～30wt%.

11, the purposes of a kind of claim 1 or 2 described little peptides and beneficial microbe symphysis unit feed addictive.