CN103766650B - A kind of feed addictive of layer chicken and preparation thereof - Google Patents

A kind of feed addictive of layer chicken and preparation thereof Download PDF

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CN103766650B
CN103766650B CN201310736910.1A CN201310736910A CN103766650B CN 103766650 B CN103766650 B CN 103766650B CN 201310736910 A CN201310736910 A CN 201310736910A CN 103766650 B CN103766650 B CN 103766650B
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parts
feed addictive
microbial inoculum
layer chicken
bacillus subtilis
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CN201310736910.1A
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CN103766650A (en
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朱建国
王万平
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金湖县安农生态农业发展有限公司
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Abstract

The invention discloses a kind of feed addictive of layer chicken, belong to field of feed additive technology, parts by weight consist of: corn 35-65 part, rice bran 5-10 part, wheat bran 4-9 part, peanut meal 5-9 part, vegetable oil 0.5-3 part, salt 0.2-0.4 part, calcium constituent additive 0.1-0.2 part, amino acid 0.4-2.6 part, complex enzyme formulation 0.05 part, phytase 0.012 part, Choline Chloride 0.05-0.12 part, vitamin E 50-150 part, cordyceps 1-5 part, Tianmen Green bean noodle 2-6 part, licorice powder 0.5-1 part, bacillus subtilis CGMCC7926 microbial inoculum culture 5-8 part, Lactobacillus plantarum CGMCC7928 microbial inoculum 3-6 part, saccharomyces cerevisiae microbial inoculum 1-2 part.The present invention fully ensures that laying hen organ of multiplication is grown completely rapidly, and the initial stage of laying eggs lays eggs great, the height on fast on peak, and it is lasting to maintain peak of laying eggs.

Description

A kind of feed addictive of layer chicken and preparation thereof
Technical field
The present invention relates to a kind of feed addictive, belong to feed additive field, particularly a kind of feed addictive of layer chicken and preparation thereof.
Background technology
Feed is the prime cost of animal-breeding, and animal-breeding cost has 60%-70% to come from feed cost use according to statistics, therefore reduces feed waste, improves efficiency of feed utilization, is the key point carried out aquaculture cost control and improve culture benefit.Current aquaculture is common, and to there is efficiency of feed utilization not high, and not exclusively, keeping is not good at causing feed moisture absorption to be gone mouldy or the problem such as infested, and the existence of these problems not only considerably increases feeding cost, and result also in the wasting of resources in nutritional labeling digestion.How to address these problems, control aquaculture cost, reducing cultivation risk is the large bottleneck of one in aquaculture fast development.There is the much research about improving efficiency of feed utilization in recent years, but mainly concentrate on and use chemical synthesis additive agent field, chemical synthesis additive can cause animal drug resistance, the problem such as residual, harm humans is healthy, also has a large amount of excretas simultaneously, as the pernicious gas such as ammonia, hydrogen sulfide, contaminated environment.Chinese herbal feed additive, as the focus of emerging research, has the advantages such as natural, nutrition, side effect be little.Devoting Major Efforts To Developing Chinese herbal feed additive is to solution antibiotic residue problem, boost productivity, development unpolluted animal husbandry, meet the food security demand of people, reduce China's animal husbandry and developed country's gap, strengthen the competitiveness of China's livestock products in international market, there is important economic implications and social benefit.Research shows, many Chinese herbal medicines inherently have good In Vitro Bacteriostasis ability, with Western medicine unlike, Chinese herbal feed antibiotic health care agent noresidue, to have no drug resistance, its mechanism of action is not only the inhibitory or killing effect to pathogenic microorganism, the more important thing is the adjustment to body disease-resistant repair ability, namely improve the anti-stress ability of body, improve body's immunity and Defense response function, thus reach the object improving breeding performonce fo animals and food utilization efficiency.
Chinese herbal medicine is some plants with given efficacy, these plants are each powerful, the effectiveness of such as Activities of Some Plants is as follows: lucid asparagus: its medicinal part is block root mainly, there is replenishing the vital essence and removing heat, moisturize the effect of promoting the production of body fluid, modern pharmacology research shows that it has good curative effect to symptoms such as pulmonary tuberculosis, bronchitis, diphtheria, pertussis, dry mouth and throats, also has antibacterial immunity and anti-oxidation function simultaneously.Fructus Corni: its taste is sour, puckery, and tepor, returns liver kidney channel.Containing various active composition, comprise volatile ingredient, glucosides class and aglycon, tannin and organic acid etc.Fructus Corni water decoction significantly can raise mice serum hemolytic antibody and serum antibody IgG content.The effect that carbohydrate in Fructus Corni has obvious Promote immunity to react.Cattail pollen: be the dry pollen of Typhaceae plant raupo cattail, typha orientalis, theory of traditional Chinese medical science thinks that this property of medicine taste is sweet flat, returns liver, the heart, the spleen channel, effect of tool analgesia stagnation resolvation.Echinacea: the class Echinacea plant originating in North America and southern Canada, its main active is polysaccharide, alkylamide compound and Caffeic acids derivative.Research shows that different xylan in Echinacea and arabinose by stimulating the activity of the lymphocytic propagation of monokaryon and macrophage, thus can have significant immunostimulation.Echinacea also has antibacterial and anti-inflammation functions, its active component Cichoric acid, has and strengthens immune anti-inflammatory properties, can suppress hyaluronic acid, and protection collagen III, from degraded, has obvious inhibitory action to Escherichia coli.
How effectively to utilize existing herbal raw material to prepare and effectively can improve animal immunizing power and the feed addictive improving feed complicated utilization efficiency, it is a project being conducive to feed industry and developing in a healthy way that the high value realizing feed utilizes.
Summary of the invention:
The object of the invention is to develop a kind of feed addictive of layer chicken and preparation method thereof.
Technical scheme of the present invention is as follows:
A kind of feed addictive of layer chicken, parts by weight consist of: corn 35-65 part, rice bran 5-10 part, wheat bran 4-9 part, peanut meal 5-9 part, vegetable oil 0.5-3 part, salt 0.2-0.4 part, calcium constituent additive 0.1-0.2 part, amino acid 0.4-2.6 part, complex enzyme formulation 0.05 part, phytase 0.012 part, Choline Chloride 0.05-0.12 part, vitamin E 50-150 part, cordyceps 1-5 part, Tianmen Green bean noodle 2-6 part, licorice powder 0.5-1 part, bacillus subtilis CGMCC7926 microbial inoculum culture 5-8 part, Lactobacillus plantarum CGMCC7928 microbial inoculum 3-6 part, saccharomyces cerevisiae microbial inoculum 1-2 part.
Described lucid asparagus powder, preparation method thereof is as follows: it is less than 1 millimeter that lucid asparagus is crushed to particle diameter through pulverizer.
Described licorice powder preparation method is as follows: it is less than 1 millimeter that Radix Glycyrrhizae is crushed to particle diameter through pulverizer.
Described cordyceps preparation method: cordyceps species obtains seed liquor through inoculation fermentation, cultivate step by step and obtain zymotic fluid, zymotic fluid centrifugation obtains mycelia, and mycelia drying and crushing obtains mycelium powder of sinensis.
Described Cordyceps Militaris fermentation medium percentage by weight consists of: sucrose 4%, glucose 3%, Tianmen Green bean noodle 08%, Fructus Corni 0.7%, peptone 0.4%, soyabean protein powder 0.6%, potassium dihydrogen phosphate 0.2%, and magnesium sulfate 0.1%, insufficient section pure water is supplied, pH6.0.
Described calcium constituent additive is one in calcium carbonate, calcium monohydrogen phosphate or calcium dihydrogen phosphate or two kinds.
Described amino acid is two or more the mixing in lysine, methionine, threonine or tryptophan.
In described complex enzyme formulation, the parts by weight content of each component is as follows: zytase 3-6 part, mannase 1-2 part, protease 1-3 part;
The unit of above-mentioned enzyme is as follows: zytase 20000-30000U/g, mannase 1000-40000U/g, protease 3 000-5000U/g.
Prepared by bacillus subtilis culture: cultivate bacillus subtilis from inclined-plane switching, the seed liquor after spreading cultivation step by step is transferred in fermentation tank, and control temperature is 28-30 DEG C, aerlbic culture 19-24 hour, and throughput is 2.0m 3/ minute; Zymotic fluid plate-frame filtering, the dry culture obtained containing bacillus subtilis.
Prepared by saccharomyces cerevisiae microbial inoculum: slant strains obtains yeast starter liquid through the conventional multistage technique that spreads cultivation, and transfer in fermentation tank, control temperature is 28 DEG C, early stage aerlbic culture 14 hours, throughput controls as 2.0m 3/ minute, later stage Anaerobic culturel 15 hours; Zymotic fluid low temperature concentrates, and after mixing with carrier, through fluidized bed drying preparation, vehicle group becomes CaCO 340 parts, 20 parts, dextrin, corn protein powder 20 parts.
Prepared by Lactobacillus plantarum: inclined-plane is cultivated Lactobacillus plantarum seed liquor and transferred in fermentation tank, and control temperature is 28-32 DEG C, Anaerobic culturel 22 hours, and throughput is 2.0m 3/ minute; Complete centrifugation of fermenting obtains wet thallus, and add the protective agent consisting of 10% defatted milk, 5% trehalose, 5% glycerine, obtain microbial inoculum by freeze drying, moisture is lower than 10%.
The preparation method of product of the present invention is as follows: the Tianmen Green bean noodle after pulverizing and licorice powder are mixed with other raw materials according to formula rate.
The invention has the beneficial effects as follows:
Bacillus subtilis culture is with the addition of in feed, the composite bacillus subtilis microbial agent of science of the present invention, Lactobacillus plantarum microbial inoculum and saccharomyces cerevisiae microbial inoculum, bacillus subtilis, saccharomyces cerevisiae microbial inoculum and Lactobacillus plantarum microbial inoculum are achieved organic assembling, in chicken stomach, under effect due to body temperature factor, enzyme preparation starts to have an effect, and decomposes wherein nutriment; Lactobacillus plantarum body fluid and around after digestion trophic factors start growth and breeding and produce lactic acid, the distinctive strong galactopoiesis acidity of Lactobacillus plantarum is played, and the generation of lactic acid effectively promotes and formed the digestive environments of stomach, promotes the digesting and assimilating of nutriment; The factors such as product bacillus subtilis of the present invention enter enteron aisle and effectively can reduce the usage quantity of antibiotics in letting animals feed, improve the immunity of letting animals feed, improve the security of animal meat product, improve the food utilization efficiency of animal, reduce the use of antibiotic and medicine in conventional raising, strengthen the resistivity of animal, improve raise benefit.
Hen secondary sex characters cockscomb is grown and red is looked fast soon, and wattle is also red looks fast soon, and whole chicken group cockscomb reddens rapidly greatly, grows have medium above correlation soon and well with organ of multiplication.One of target that the present invention reaches fully ensures that laying hen organ of multiplication is grown completely rapidly, and body weight increases fast, guarantees that physique is strong, for stable high yield is taken a firm foundation; Be embodied in the weightening finish of laying hen largest body, maximum organ of multiplication is grown, and ensures physical efficiency deposit (energy and calcium phosphorus), is getting rid of hat phase lay heavily!
The present invention adopts science compound feed, coordinates the organic assembling of accurate scale of feeding, fully ensures that laying hen organ of multiplication is grown completely rapidly, body weight increases fast, guarantee that physique is strong, body maturation and sexal maturity are synchronously grown, thus reach the initial stage of laying eggs and lay eggs great, fast on peak, on height, and it is lasting to maintain peak of laying eggs, and reaches peak of laying eggs for more than 7 months, lay eggs 72 week age and reach 311 pieces of eggs, egg production more than 19.6 kilograms.28 weekend average egg weight 61.3 grams, improve 3.03% than average level; Laying rate reaches more than 95%, improves 1-3.6% than average level; Egg production, from 48.2 grams/only/day, brings up to 50.1 grams/only/day; 16 week age-28, all age grade sections death rate was reduced to 0.75%, every chicken fecund 3 pieces, egg.
Detailed description of the invention
The present invention is described below by specific embodiment.Unless stated otherwise, technological means used in the present invention is method known in those skilled in the art.In addition, embodiment is interpreted as illustrative, but not limits the scope of the invention, and the spirit and scope of the invention only limited by claims.To those skilled in the art, under the prerequisite not deviating from essence of the present invention and scope, the various change carry out the material component in these embodiments and consumption or change also belong to protection scope of the present invention.
Saccharomyces cerevisiae saccharomyces cerevisiae CICC31481 provided by the present invention is purchased from Chinese industrial Microbiological Culture Collection administrative center.
Bacillus subtilis (Bacillussubtilis) Li-2013-02, this bacterial strain is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center on July 15th, 2013 and (is called for short CGMCC, address is: No. 3, No. 1, North Star West Road, Chaoyang District, city of BeiJing, China institute), preserving number is CGMCCNo.7926.
Described strain characteristic is that the enzyme activity of product Thermostable α-Amylase is high, heat-resisting, acid resistance is strong.
Thermostable α-Amylase enzyme activity prepared by described bacterial strain is 30000-35000u/ml; Applicable temperature scope is 105-115 DEG C, optimal reactive temperature 110 DEG C, at 110 DEG C of enzymes complete stability alive; Being suitable for pH value in reaction scope is 3.0-7.0, and the enzyme complete stability alive when pH value is 3.0, optimal reaction pH value is 4.2.
Described bacterial strain feature is as follows:
Described bacterial strain colony colour on solid plate is milky, and dry tack free is opaque, neat in edge, for having the aerobic bacteria of motility.Microscopy is elongated rod shape, and Gram's staining is positive.This bacterium can utilize citrate, and nitrate reductase, V-P test into the positive.
Described bacillus subtilis (Bacillussubtilis) Li-2013-02 is produced Thermostable α-Amylase bacillus subtilis Li-2013 by a strain obtains through UV-LiCl-dithyl sulfate Mutation screening, and concrete screening step is as follows:
(1) preparation of bacteria suspension
The mono-bacterium colony of Li-2013 grown after plate streaking is separated is accessed in seed culture medium, 100r/min, after 40 DEG C of cultivation 12h, after getting 1mL medium centrifugal, use brine twice, and resuspended with 9mL physiological saline.
(2) UV-LiCl-dithyl sulfate complex mutation
Bacteria suspension is placed in aseptic flat board, is 30cm in distance, stirs and irradiate 100s under the uviol lamp of power 15w.Bacterium liquid through irradiating is coated lithium chloride flat board after gradient dilution, and contrasts to be coated with flat board without the bacterium liquid dilution of ultraviolet irradiation.Above-mentioned coating is dull and stereotyped uniformly, wrap with the cloth of black or newspaper, put 40 DEG C and cultivate 48h, the flat board growing bacterium colony filters out hydrolysis circle choose preserve to inclined-plane with colony diameter ratio the maximum, bacteria suspension is mixed with after purifying, fully mix with dithyl sulfate stoste after gradient dilution, and in 40 DEG C of concussion process 40min, the bacterium liquid processed is coated lithium chloride flat board after gradient dilution.
(3) primary dcreening operation of high-yield strains
Above-mentioned coating is dull and stereotyped uniformly, and put 40 DEG C and cultivate 48h, on the flat board growing bacterium colony, primary dcreening operation goes out hydrolysis circle and chooses preserve to inclined-plane with colony diameter ratio the greater, obtains three strain bacterium Li-2013-01, Li-2013-02, Li-2013-03 after purifying
(4) shake flask fermentation sieves again
By the three strain bacterium Li-2013-01 obtained, Li-2013-02, Li-2013-03 carry out shake flask fermentation in the 250mL shaking flask containing 30mL fermentation medium, seed inoculum concentration 10% (V/V), 40 DEG C, 100r/min cultivates 72h, centrifuging and taking fermented supernatant fluid obtains crude enzyme liquid.
(5) enzyme activity determination
The definition of Mei Huo unit: 1mL crude enzyme liquid, in 105 DEG C, under pH4.2 condition, 1min liquefies 1mg soluble starch,
Be 1 enzyme activity unit, represent with U/mL.
After measured, bacterial strain Li-2013-02 is stable most superior strain, and enzyme is lived and reached 30000U/mL.
Described lithium chloride is dull and stereotyped: starch 1%, peptone 1%, (NH) 2sO 40.4%, K 2hPO 40.8%, CaCl 20.2%, lithium chloride 0.9%, agar 2%.
Described seed culture medium: dusty yeast 0.5%, peptone 1%, soluble starch 1%, NaCl1%.
Described fermentation medium: corn flour 5%-15%, beancake powder 4%-10%, (NH) 2sO 40.4%, K 2hPO 40.8%, CaCl 20.2%.
Described shake flask culture conditions: this bacterium in the 250mL shaking flask containing 30mL fermentation medium, inoculum concentration 10% (V/V), 100r/min, 40 DEG C of fermented and cultured 72h.
Described high-temperature resistant alpha-amylase, its zymologic property is as follows:
(1) this enzyme Acclimation temperature wider range, optimum temperature between 100-110 DEG C, and to be preserved below 110 DEG C, and temperature stability is better, and more than 110 DEG C to preserve long-time temperature stability poor.
(2) this enzyme optimal reaction pH value is 4.2.High enzyme vigor is all had, the enzyme complete stability alive when pH value is 3.0 between pH value 3.0-7.0.
(3) enzymatic activity: by mutant strain Li-2013-02 provided by the present invention, the Thermostable α-Amylase enzyme activity of preparation is 30000-35000U/ml.
Lactobacillus provided by the present invention is Lactobacillus plantarum (Lactobacillusplantarum) Li-2013-01, this bacterial strain is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center, deposit number is CGMCCNo.7928, preservation address: No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City, Institute of Microorganism, Academia Sinica, postcode 100101.Preservation date on July 15th, 2013.This bacterial strain feature is as follows: examine under a microscope, and this bacterial strain is rod-short, and Gram's staining is positive, and boundless hair, does not produce gemma; On solid medium, this bacterium bacterium colony is white, and smooth surface is fine and close, and form is circular, and edge is more neat.Physicochemical characteristics is: catalase (-), gelatin liquefaction (-), indoles experiment (+), motility (-), fermentation gas (-), nitrate reductase (-), and fermentation gas (-) produces hydrogen sulfide gas (-), grows (+) in pH4.5MRS culture medium.
Lactobacillus plantarum of the present invention adopts following flow process to carry out seed selection:
The original bacterial classification that sets out → test tube activation → dithyl sulfate (DES) mutagenesis → dull and stereotyped primary dcreening operation → nitrosoguanidine (NTG) mutagenesis → dull and stereotyped primary dcreening operation → shaking flask sieves → mitotic stability test again.
The original bacterial classification that sets out is CICC20242, is purchased from Chinese industrial Microbiological Culture Collection administrative center.
Original strain of the present invention is in xylan culture medium, and the output of lactic acid is 12.5g/L.In order to improve its lactic acid production, DES and NTG is adopted to carry out mutagenesis to this bacterial classification successively, mutagenesis adopts MRS calcium carbonate flat board to carry out primary dcreening operation, then 500mL shake flask fermentation is adopted, biosensor analysis instrument carries out multiple sieve to Producing Strain, the lactobacillus plantarum strain that seed selection is excellent, then does passage assays, evaluates its genetic stability.
Bacterial strain CGMCCNo.7928 genetic stability result shows: through continuous passage ten times, property indices is all more stable, and heredity is better, and proterties is not replied, therefore using the object bacterial strain that bacterial strain CGMCCNo.7928 obtains as seed selection.
Object bacterial strain CGMCCNo.7928 is done the experiment of 10L fermentation tank, result shows: after fermentation 72h, take xylan as carbon source, the lactic acid concn of Lactobacillus plantarum CGMCCNo.7928 can reach 57g/L, improves 356% compared with starting strain.
Object bacterial strain CGMCCNo.7928 is done the experiment of 10L fermentation tank, result shows: after fermentation 72h, take glucose as carbon source, the lactic acid concn of Lactobacillus plantarum CGMCCNo.7928 can reach 68g/L.
Detailed process is as follows:
Culture medium:
Liquid MRS xylan culture medium (beef extract 2g, peptone 10g, yeast extract 5g, xylan 20g, sodium acetate 5g, ammonium citrate 2g, dipotassium hydrogen phosphate 2g, epsom salt 0.2g, seven water manganese sulfate 0.05g, after dissolving one by one, running water constant volume 1000mL, regulates pH7.0-7.2); MRS xylan screening solid medium (beef extract 2g, peptone 10g, yeast extract 5g, xylan 90g, sodium acetate 5g, ammonium citrate 2g, dipotassium hydrogen phosphate 2g, epsom salt 0.2g, seven water manganese sulfate 0.05g, after dissolving one by one, running water constant volume 1000mL, regulate pH7.0-7.2, add 20g agar).
Dithyl sulfate (DES) mutagenic and breeding:
Super-clean bench is got Lactobacillus plantarum one ring on test tube slant, and access is equipped with in the 250mL triangular flask of 50mL liquid MRS xylan culture medium, 200rpm, cultivates about 12h for 40 DEG C, makes thalline be in logarithmic growth in earlier stage.
Get 5mL bacterium liquid, the centrifugal 10min of 5000rpm collects thalline, with brine 2 times.
107/mL bacteria suspension is diluted to pH7.0 phosphate buffer.
Get the kaliumphosphate buffer of 32mLpH7.0,8mL bacteria suspension, 150mL triangular flask that 0.4mLDES to put into rotor in advance fully mix, make DES ultimate density be 1%(v/v).
In 30 DEG C of shaking tables, 150rpm reacts 30min, gets 1mL mixed liquor, adds 0.5mL25%Na 2s2O 3solution stopped reaction.
Dilution spread screens in solid medium plate in the MRS xylan containing 90g/L xylan.The bacterial strain that after cultivating 2 ~ 3 days at 40 DEG C, picking transparent circle/colony diameter is maximum, label is DES bacterium.
Nitrosoguanidine mutagenesis:
Super-clean bench is got Lactobacillus plantarum DES mono-ring on test tube slant, and access is equipped with in the 250mL triangular flask of 50mL liquid MRS xylan culture medium, 200rpm, cultivates about 12h for 40 DEG C, makes thalline be in logarithmic growth in earlier stage.
Get the centrifugal 10min of 5mL bacterium liquid 5000rpm and collect thalline, with brine 2 times.
107/mL bacteria suspension is diluted to pH6.0 phosphate buffer.
Get 10mL bacteria suspension to be transferred in 100mL triangular flask, add the NTG of 10mg, be mixed with the NTG solution that final concentration is 10mg/mL, and add 4-5 and drip acetone, be beneficial to NTG and dissolve.
At 30 DEG C, the centrifugal 10min of 200rpm oscillating reactions 30min, 5000rpm collects thalline, with SPSS washing several, and stopped reaction.
Suitable dilution is coated with, and gets last dilution bacterium liquid 0.2mL, and dilution spread screens in solid medium plate in the MRS xylan containing 90g/L xylan.The bacterial strain 150 that after cultivating 2 ~ 3 days at 40 DEG C, picking transparent circle/colony diameter is larger.
Shaking flask is sieved again:
Super-clean bench is got Lactobacillus plantarum one ring on each test tube slant respectively, and access is equipped with in the 250mL triangular flask of 50mL liquid MRS xylan culture medium, 200rpm, cultivates 3-4 days, detects concentration of glucose and Pfansteihl change in concentration every day for 40 DEG C.After fermentation ends, compare the xylan wear rate of 150 strain bacterial classifications and lactic acid and produce speed, the co