CN103725661A - Special enzyme containing feed composite enzyme for piglets and preparation method thereof - Google Patents

Special enzyme containing feed composite enzyme for piglets and preparation method thereof Download PDF

Info

Publication number
CN103725661A
CN103725661A CN201310736761.9A CN201310736761A CN103725661A CN 103725661 A CN103725661 A CN 103725661A CN 201310736761 A CN201310736761 A CN 201310736761A CN 103725661 A CN103725661 A CN 103725661A
Authority
CN
China
Prior art keywords
parts
culture
enzyme
aspergillus niger
herbal medicine
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Granted
Application number
CN201310736761.9A
Other languages
Chinese (zh)
Other versions
CN103725661B (en
Inventor
张锦杰
李贵骏
刘文明
封章平
王永兰
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Hunan Hongying Biological Science & Technology Co Ltd
Original Assignee
Hunan Hongying Biological Science & Technology Co Ltd
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Hunan Hongying Biological Science & Technology Co Ltd filed Critical Hunan Hongying Biological Science & Technology Co Ltd
Priority to CN201310736761.9A priority Critical patent/CN103725661B/en
Publication of CN103725661A publication Critical patent/CN103725661A/en
Application granted granted Critical
Publication of CN103725661B publication Critical patent/CN103725661B/en
Active legal-status Critical Current
Anticipated expiration legal-status Critical

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14—Hydrolases (3)
    • C12N9/24—Hydrolases (3) acting on glycosyl compounds (3.2)
    • C12N9/2402—Hydrolases (3) acting on glycosyl compounds (3.2) hydrolysing O- and S- glycosyl compounds (3.2.1)
    • C12N9/2477—Hemicellulases not provided in a preceding group
    • C12N9/248—Xylanases
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23K—FODDER
    • A23K20/00—Accessory food factors for animal feeding-stuffs
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23K—FODDER
    • A23K20/00—Accessory food factors for animal feeding-stuffs
    • A23K20/10—Organic substances
    • A23K20/189—Enzymes
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23K—FODDER
    • A23K50/00—Feeding-stuffs specially adapted for particular animals
    • A23K50/30—Feeding-stuffs specially adapted for particular animals for swines
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/0004—Oxidoreductases (1.)
    • C12N9/0006—Oxidoreductases (1.) acting on CH-OH groups as donors (1.1)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/0004—Oxidoreductases (1.)
    • C12N9/0089—Oxidoreductases (1.) acting on superoxide as acceptor (1.15)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14—Hydrolases (3)
    • C12N9/16—Hydrolases (3) acting on ester bonds (3.1)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14—Hydrolases (3)
    • C12N9/24—Hydrolases (3) acting on glycosyl compounds (3.2)
    • C12N9/2402—Hydrolases (3) acting on glycosyl compounds (3.2) hydrolysing O- and S- glycosyl compounds (3.2.1)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14—Hydrolases (3)
    • C12N9/24—Hydrolases (3) acting on glycosyl compounds (3.2)
    • C12N9/2402—Hydrolases (3) acting on glycosyl compounds (3.2) hydrolysing O- and S- glycosyl compounds (3.2.1)
    • C12N9/2405—Glucanases
    • C12N9/2408—Glucanases acting on alpha -1,4-glucosidic bonds
    • C12N9/2411—Amylases
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14—Hydrolases (3)
    • C12N9/24—Hydrolases (3) acting on glycosyl compounds (3.2)
    • C12N9/2402—Hydrolases (3) acting on glycosyl compounds (3.2) hydrolysing O- and S- glycosyl compounds (3.2.1)
    • C12N9/2405—Glucanases
    • C12N9/2408—Glucanases acting on alpha -1,4-glucosidic bonds
    • C12N9/2411—Amylases
    • C12N9/2425—Beta-amylase (3.2.1.2)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14—Hydrolases (3)
    • C12N9/24—Hydrolases (3) acting on glycosyl compounds (3.2)
    • C12N9/2402—Hydrolases (3) acting on glycosyl compounds (3.2) hydrolysing O- and S- glycosyl compounds (3.2.1)
    • C12N9/2405—Glucanases
    • C12N9/2434—Glucanases acting on beta-1,4-glucosidic bonds
    • C12N9/2437—Cellulases (3.2.1.4; 3.2.1.74; 3.2.1.91; 3.2.1.150)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14—Hydrolases (3)
    • C12N9/24—Hydrolases (3) acting on glycosyl compounds (3.2)
    • C12N9/2402—Hydrolases (3) acting on glycosyl compounds (3.2) hydrolysing O- and S- glycosyl compounds (3.2.1)
    • C12N9/2405—Glucanases
    • C12N9/2434—Glucanases acting on beta-1,4-glucosidic bonds
    • C12N9/244—Endo-1,3(4)-beta-glucanase (3.2.1.6)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14—Hydrolases (3)
    • C12N9/24—Hydrolases (3) acting on glycosyl compounds (3.2)
    • C12N9/2402—Hydrolases (3) acting on glycosyl compounds (3.2) hydrolysing O- and S- glycosyl compounds (3.2.1)
    • C12N9/2405—Glucanases
    • C12N9/2434—Glucanases acting on beta-1,4-glucosidic bonds
    • C12N9/2445—Beta-glucosidase (3.2.1.21)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14—Hydrolases (3)
    • C12N9/24—Hydrolases (3) acting on glycosyl compounds (3.2)
    • C12N9/2402—Hydrolases (3) acting on glycosyl compounds (3.2) hydrolysing O- and S- glycosyl compounds (3.2.1)
    • C12N9/2405—Glucanases
    • C12N9/2451—Glucanases acting on alpha-1,6-glucosidic bonds
    • C12N9/2457—Pullulanase (3.2.1.41)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Y—ENZYMES
    • C12Y101/00—Oxidoreductases acting on the CH-OH group of donors (1.1)
    • C12Y101/03—Oxidoreductases acting on the CH-OH group of donors (1.1) with a oxygen as acceptor (1.1.3)
    • C12Y101/03004—Glucose oxidase (1.1.3.4)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Y—ENZYMES
    • C12Y115/00—Oxidoreductases acting on superoxide as acceptor (1.15)
    • C12Y115/01—Oxidoreductases acting on superoxide as acceptor (1.15) with NAD or NADP as acceptor (1.15.1)
    • C12Y115/01001—Superoxide dismutase (1.15.1.1)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Y—ENZYMES
    • C12Y301/00—Hydrolases acting on ester bonds (3.1)
    • C12Y301/03—Phosphoric monoester hydrolases (3.1.3)
    • C12Y301/03002—Acid phosphatase (3.1.3.2)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Y—ENZYMES
    • C12Y302/00—Hydrolases acting on glycosyl compounds, i.e. glycosylases (3.2)
    • C12Y302/01—Glycosidases, i.e. enzymes hydrolysing O- and S-glycosyl compounds (3.2.1)
    • C12Y302/01002—Beta-amylase (3.2.1.2)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Y—ENZYMES
    • C12Y302/00—Hydrolases acting on glycosyl compounds, i.e. glycosylases (3.2)
    • C12Y302/01—Glycosidases, i.e. enzymes hydrolysing O- and S-glycosyl compounds (3.2.1)
    • C12Y302/01021—Beta-glucosidase (3.2.1.21)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Y—ENZYMES
    • C12Y302/00—Hydrolases acting on glycosyl compounds, i.e. glycosylases (3.2)
    • C12Y302/01—Glycosidases, i.e. enzymes hydrolysing O- and S-glycosyl compounds (3.2.1)
    • C12Y302/01041—Pullulanase (3.2.1.41)

Landscapes

  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Organic Chemistry (AREA)
  • Engineering & Computer Science (AREA)
  • Zoology (AREA)
  • Wood Science & Technology (AREA)
  • Genetics & Genomics (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • General Health & Medical Sciences (AREA)
  • General Engineering & Computer Science (AREA)
  • Biochemistry (AREA)
  • Biotechnology (AREA)
  • Molecular Biology (AREA)
  • Biomedical Technology (AREA)
  • Medicinal Chemistry (AREA)
  • Microbiology (AREA)
  • Polymers & Plastics (AREA)
  • Animal Husbandry (AREA)
  • Food Science & Technology (AREA)
  • Birds (AREA)
  • Fodder In General (AREA)
  • Enzymes And Modification Thereof (AREA)

Abstract

The invention discloses a special enzyme containing a feed composite enzyme for piglets and a preparation method thereof, belonging to the technical field of preparation of enzyme preparations. The special enzyme for piglets is prepared by scientifically compounding Aspergillus niger culture, acidic xylanase, cellulase, beta-glucanase, amylase, phytase, Chinese herbal medicine extract, a protective agent and an activator, wherein Aspergillus niger body in the Aspergillus niger culture can inhibit the growth of pathogenic escherichia coli in processes of growth and fermentation, inhibit the growth and reproduction of Aspergillus niger, decompose aflatoxin, promote animal growth, regulate flora balance in gastrointestinal tract, and improve overall immunity; the crude enzyme liquid of fermentation liquor of the Aspergillus niger culture contains activity of multiple enzymes such as protease, mannase, alpha-galactase and pectinase, thus being especially suitable for being added into feed for piglets.

Description

A kind of suckling piglet specific enzyme containing complex enzyme for feed and preparation method thereof
Technical field
The invention belongs to enzyme preparation technical field, specifically a kind of suckling piglet specific enzyme containing complex enzyme for feed and preparation method thereof.
Background technology
In feed, add zymin mainly by following 4 reasons: 1. the antinutritional factor that degraded exists in animal-feed.These materials can not be degraded by animal endogenous enzyme, thereby disturb the eubolism of animal, cause animal digestion bad, and production performance declines.2. improve the utilization ratio of starch, protein and mineral substance.These materials or surrounded by the cell walls of fiber-enriched, or with some, can not be had by the structure formation of animal digestion that (for example, in plant feed raw material, a large amount of phosphorus exists with the form of phytate phosphorus.）。3. some specific chemical bond of degrading in raw material.These chemical bonds can not be degraded by the enzyme of animal self, add exogenous enzyme can discharge more nutrition later.4. because young animal autodigestion system is also immature, the not enough interpolation of endogenous enzyme exogenous enzyme can improve feed digestibility, prevents indigestion symptom.Except can improving the utilization ratio of daily ration, the enzyme-added difference that can also reduce between feedstuff raw material, the accuracy of raising feed formulation, the while can also be improved the regularity of growth of animal, reduces handling cost, increases economic efficiency.Use all right protection of the environment of zymin.Because the utilization ratio of feed has improved, corresponding ight soil quantity discharged has declined.In the situation that effect is obvious, the quantity discharged of ight soil can reduce by 20% left and right, the discharge of nitrogen 15% left and right that declines in pig manure, and in chicken manure, the discharge of nitrogen declines 20%.For phytate phosphorus, can significantly reduce the pollution of phosphorus to environment.
The zymin of applying in fodder industry at present mainly contains 4 large classes: be used for respectively degraded cellulose, protein, starch and phytic acid.
Fiber degradation enzyme: for monogastric animal, the maximum resistance of digestion is the enzyme that can not produce degradation of fibers, in the daily ration that contains the components such as wheat, barley, oat, fiber is araboxylan and beta-glucan greatly.Water miscible fiber can improve the viscosity of small intestine contents, hinders the absorption of nutrient, thereby reduces the growth performance of animal.Simultaneously this situation also with some because the disease that maldigestion causes is relevant.As black toe disease and the piggy of the hyposexuality of pig, fowl have loose bowels.Due to the impact of the factors such as kind, growth place gentle time condition, the altering a great deal of fibre content in barley and wheat, the nutritive value of the daily ration that causes containing these components is widely different.Fiber degradation enzyme, zytase and beta-glucan can reduce these differences, improve growth performance and the reguarity of animal.Can also reduce some dyspeptic disease simultaneously.
Proteolytic degradation enzyme: protein is from all feeds raw material in animal diets, and they finally stockpile in lean meat by the amino acid of degraded.In monogastric animal daily ration, add proteolytic enzyme (DIFFERENT FEED material protein and quality and utilizability) except fully degrading most of storage protein or Storage protein or for the available small-molecular peptides of animal, can also improve feed nutritive value by degraded anti-nutritional factors.The efficiency variance stockpiling is very large.At plant protein source, as existed some antinutritional factor in soybean cake powder, as several tannins and trypsin ihhibitor, may cause damage to intestinal absorption surface, affect nutraceutical absorption.In addition, the incomplete digestion system of young animal be the protein in vegetable-protein (as soybean cake powder) can not well be utilized.
Starch degrading enzyme: many nutritionists think corn" golden standard " of feedstuff raw material.Large absolutely number nutritionist thinks that corn does not exist lienteric, digestibility surpasses 95%, but Noy and Sklan research show (1994) in the ideal situation recently, in the daily ration of broiler of 4-12 age in days, the digestibility of starch seldom surpasses 85%, adds amylase and can make starch obtain more degradeds faster at small intestine.In the weaned piglet phase, due to nutrition, environment and immune variation, body weight can decline.In daily ration, add amylase and some other enzyme, can increase the endogenous digestive ferment secretion of animal, and then improve digesting and assimilating of nutrition, improve food conversion ratio and growth of animal rate.
Phytic acid degrading enzyme: for all animals, phosphorus is all vital for mineralising, immunity, breeding, the growth of bone.Pig and poultry monogastric animal can only in utilize the phosphorus of 30-40% in plant feed, the phytate phosphorus accident of all the other 60-70% is unserviceable.In many cases, in feed diet, must supplement the needs that inorganic phosphorus meets growth of animal.Phosphorus over half in feed is along with ight soil is discharged in environment, contaminate environment.Add the phytase phytic acid of can degrading, discharge the phosphorus in phytic acid molecule.Can produce like this 2 benefits: 1. the addition that has reduced Dietary phosphorus.2. having reduced feces of livestock and poultry pollutes the phosphorus of environment.
Apparent: as four large leading roles of feed enzyme, their mechanism of action and pattern determine or promoted animal-feed industry to use for the absorption of zymin technology to a great extent.In broiler chicken material, add at present the ratio for input and output example of enzyme over 2: 1.Comparatively speaking, in pig industry field, the service condition of zymin, with regard to more complicated, seems uncertain.Intensive degree is low, relates to link many, and the result of use of zymin is difficult to carry out business calculating.Although had the imagination of using zymin in the 1950's, until just start to understand strength how to bring into play enzyme in fodder industry the eighties in 20th century.Feed grains, as Wheat and barley all contains the unavailable fiber of higher monogastric animal.As fruit fiber can be degraded, animal just can utilize nutrition better.In Europe, barley is more cheap, and bird nutritionist and zymologist have dropped into great effort and studied and in containing the daily ration of broiler of barley, add beta-glucan enzyme to reduce the possibility of its negative impact.Its result is proved to be successfully, and has obtained a gold law: barley+beta-glucan enzyme=wheat.Be subject to above-mentioned successful inspiration, wheat is the research object of second.Theory hypothesis is: wheat+zytase=corn.The research of this step has also obtained success.In the mid-90 in 20th century, enzyme has obtained generally approval at fodder industry.Can not rant out: 1996, in the broiler chicken material (viscosity cereal is energy derive) in Europe 80%, contain fiber degradation enzyme.Strengthen thus and accelerate the application of feed industry to new technology.In the world, about 65% the poultry feed that can produce viscosity cereal that contains has added fiber degradation enzyme.And application percentage in pig feed is much lower, approaches 10%.Its major cause is the complicated structure in market, and market is diversification, even cannot calculate.From regional distribution, use the area of cellulose degrading enzyme for example mainly to concentrate on viscosity cereal, as the producing region of main energy feed: Europe, Canada, Australia and New Zealand.In addition, in the U.S., South America and the Asian-Pacific area, service condition depends on the rate of exchange between corn and wheat.In this sense, Europe is to use the core of degraded cellulose enzyme to segment market.In order to obtain global approval, feeding enzyme producer must carry out on a large scale marching take that corn---bean pulp type daily ration is main North America and the Asian-Pacific area.Corn---bean pulp type daily ration is always counted as " golden standard ", although many nutritionists think that the mobility of these raw materials is more much bigger than the mobility of original imagination.Now, increasing evidence shows that this gold daily ration also can improve its production performance by enzyme, although this class daily ration problem relevant to robust fibre or viscosity is not serious.Past 10 years was expended a lot in research and development Corn-soybean first-generation feed enzyme, and started successful Application in 1996, and initial stage application result is multifarious, but industry is just starting to become, how more and more understand could be handy, adds zymotechnic and obtain maximum economy return.It is estimated, this feed a part enzyme market share is 2,000 ten thousand dollars, and the actual broiler fodder using Corn-soybean daily ration only has 5% for enzyme-added feed.Within 1999/2000 year one, reduce feed enzyme marketable value that viscosity and robust fibre are daily ration over 100,000,000 dollars.At present, phytase has obtained admitting and applying of the whole world.The market share of phytase is approximately annual 5000 ten thousand dollars, approximately has the pig fowl feed of 8.0% left and right to add phytase in the whole world.Except the reason of economic interests, also having a factor is to have reduced the content of phytate phosphorus in excrement to be conducive to protection of the environment.
In sum, the application of suckling piglet specific enzyme has its wide market space and huge economic worth, but the thermostability of suckling piglet specific enzyme, security, composite comprehensive and giving full play to of action effect are still zymin manufacturer and the common major issue of paying close attention to of numerous raisers, prepare safer, more comprehensively, the better suckling piglet specific enzyme of enzyme action effect is industry technician corporation responsibility and pursuit.
Summary of the invention
Technical problem solved by the invention is that the aspergillus niger culture of take containing the complex enzyme for feed of heat-flash stability and pH stability is basis, and the composite Chinese herbal medicine extract of science, protective material, activator and other food grade feed enzyme, the feed complex enzyme making not only provides safety for raising livestock and poultry, comprehensive digestive ferment, alleviate digestion burden, improve raw material availability and growth rate, effective protection of the environment, appropriate activator can under equal conditions be given full play to the effect of zymin simultaneously, make the best use of everything, save zymin addition, the composite quality guaranteed period that both can extend compound enzymic preparation of science of Chinese herbal medicine extract, can improve again the immunizing power of raising livestock and poultry, thereby reach the multiplex effect of an enzyme.
In order to achieve the above object, the present invention is by the following technical solutions:
Containing the suckling piglet specific enzyme of complex enzyme for feed, by the raw material of following parts by weight, formed:
Aspergillus niger culture 40-60 part, acidic xylanase 15-25 part, cellulase 15-25 part, beta-glucanase 15-20 part, amylase 15-20 part, phytase 10-15 part, Chinese herbal medicine extract 10-15 part, protective material 10-15 part, activator 10-15 part.
Described acidic xylanase, cellulase, beta-glucanase, amylase, phytase are food-grade enzyme preparation;
In described aspergillus niger culture, mainly comprise feeding enzyme and aspergillus strain.
In described aspergillus niger culture fermentation broth crude enzyme liquid, contain multiple feeding enzyme, wherein, proteinase activity 6500-6700U/ml, mannosans enzyme activity 1500-1700U/ml, α-galactase vigor 1200-1300U/ml, pectinase activity 1000-1200U/ml.Fermented liquid crude enzyme liquid is carried out to heat stability test and the pH stability test of enzyme, test-results shows: at 30-75 ℃, during pH value 2-7, each enzyme all has higher enzyme activity, reaches more than 80%.
Described aspergillus strain can suppress the growth of pathogenic colon bacillus in growth and fermenting process; Suppress flavus Growth and reproduction, decompose aflatoxin; Promote growth of animal, regulate gastrointestinal bacterial flora balance, improve overall immunity.
The preparation method of described aspergillus niger culture is as follows: by the slant strains of intact aspergillus niger DM-18 through actication of culture and one-level, secondary, three grades and seeding tank liquid seeds step by step enlarged culturing obtain liquid seeds, with 6% inoculum size access fermentor tank, culture temperature 28-30 ℃, stirring velocity 200-700r/m, ventilation (V/V) 1:1-3, incubation time 10-15h; Then with 1-2 ℃/h rate of temperature fall slow cooling to 10-15 ℃, constant temperature culture 15-20h; Continuation to 2-5 ℃, now, is appended access fermentor tank, constant temperature culture 20-30h by liquid seeds with 4% inoculum size with 1-2 ℃/h rate of temperature fall slow cooling; Finally with 1-2 ℃/h temperature rise rate, be slowly warming up to 10-15 ℃, constant temperature culture 15-20h; Continuation is slowly warming up to 30-33 ℃, constant temperature culture 15-20h with 1-2 ℃/h temperature rise rate; Fermented liquid obtains aspergillus niger wet thallus and centrifugate through centrifugation, centrifugate obtains solid content 20-40% concentrated solution in the concentrated moisture of removing of 10-20 ℃ of loop ultrafiltration, concentrated solution evenly mixes with mass ratio 1-10:1 with aspergillus niger wet thallus, then through vacuum lyophilization, micronizing, obtains aspergillus niger culture.
Described slant medium consists of: casein food grade 4g, dipotassium hydrogen phosphate 2g, magnesium chloride 0.6g, Repone K 0.8g, ferrous sulfate 0.02g, glucose 20g, agar 20g, Chinese herbal medicine powder 5-20g, distilled water l000mL, 5.8,121 ℃ of sterilizing 20min of pH value.
The preparation method of described Chinese herbal medicine powder is as follows:
Take Radix Astragali 20-30 part; Radix Codonopsis 10-18 part; Radix bupleuri 10-15 part; Root of large-flowered skullcap 10-15 part; Respectively said herbal medicine being crushed to particle diameter is below 2 millimeters, then in container, evenly mix and add the water of 3-6 times of weight, control temperature 70 C～90 ℃ and keep 2～4h, add the mixture of 2-3 times of weight ethanol of mixture and propyl alcohol, control temperature to 60 ℃～78 ℃ and keep 3～4h, filter; Filtrate vacuum concentration postlyophilization obtains Chinese herbal medicine powder.
The mass ratio of described ethanol and propyl alcohol is 1:1-1.5.
Described one-level, secondary, three grades of seed culture mediums consist of: dipotassium hydrogen phosphate 2g, magnesium chloride 0.6g, Repone K 0.8g, ferrous sulfate 0.02g, glucose 20g, Chinese herbal medicine powder 15-25g, trehalose 10-30g, distilled water l000mL, 5.5,121 ℃ of sterilizing 20min of pH value.
Described seed tank culture base consists of: Semen Maydis powder 50-60g, bean powder 15-25g, wheat bran 10-15g, fish meal 10-15g, calcium chloride 6-10g, ammonium chloride 1-3g, Sodium phosphate dibasic 1-2g, Chinese herbal medicine powder 15-20g, trehalose 10-30g, pure water l000mL, pH value 5-7,121 ℃ of sterilizing 20min.
Described seeding tank fermented liquid spore concentration is 7.0x10 8-8.0x10 8individual/ml;
Described fermention medium consists of: Semen Maydis powder 50-60g, bean powder 15-25g, wheat bran 10-15g, fish meal 10-15g, calcium chloride 6-10g, ammonium chloride 1-3g, Sodium phosphate dibasic 1-2g, Chinese herbal medicine powder 30-50g, trehalose 10-30g, pure water l000mL, pH value 5-7,121 ℃ of sterilizing 20min.
Described supplemented medium weight consists of: maltodextrin 20-30%, Semen Maydis powder 10-20%, bean powder 15-25%, fish meal 1.0-1.5%, calcium chloride 0.6-1.0%, ammonium chloride 0.1-0.3%, Sodium phosphate dibasic 0.1-0.2%, Chinese herbal medicine powder 5-10%, insufficient section pure water is supplied, pH value 5-7,121-123 ℃ of sterilizing 30-40min.
The preparation method of described Chinese herbal medicine extract is: respectively Chinese herbal medicine powder being broken to particle diameter is below 2 millimeters, then in container, evenly mix and add the water of 3-6 times of weight, control temperature 70 C～90 ℃ and keep 2～4h, then being cooled to 45-60 ℃, adding mixing enzyme preparation to carry out enzymolysis, is 5.5-6.8 with newborn acid for adjusting pH value, enzymolysis 2-4h, finally add the mixture of mixture 0.5-3 times of weight ethanol and propyl alcohol, control temperature to 60 ℃～78 ℃ and keep 3～4h, filter; It is more than 20% that filtrate decompression is concentrated into solid content, and then lyophilize obtains Chinese herbal medicine extract.
The parts by weight of described herbal medicine consist of: sea-buckthorn 20-30 part, Semen Cassiae 20-30 part, matrimony vine 10-15 part, Chinese yam 10-15 part, Root of coastal Glehnia 5-10 part, Fructus Viticis Negundo 5-10 part, radix polygonati officinalis 3-5 part, seed of Job's tears 3-5 part, Fructus Hordei Germinatus 3-5 part, sweet osmanthus 3-5 part, Radix Astragali 3-5 part;
Mixed enzyme addition is the 5-10% of mixture gross weight;
The parts by weight of described mixed enzyme consist of: endo-beta-glucanase 10-20 part, outer beta-glucanase 10-20 part, beta-glucosidase 10-15 part, zytase 15-20 part, pentosanase 15-20 part, Pullulanase 20-30 part, beta-amylase 10-15 part, neutral protease 10-15 part, aspartic protease 10-15 part, superoxide-dismutase 5-10 part, glucose oxidase 5-10 part, acid phosphatase 5-10 part;
Described protective material is comprised of the raw material of following parts by weight: trehalose 20-30 part, NaCl20-30 part, (NH 4) 2sO 410-15 part, halfcystine 10-15 part.
Described activator is that the inorganic salt by following mass fraction evenly mix: zinc chloride 30-40 part, calcium chloride 10-20 part, sodium sulfate 10-20 part, magnesium chloride 5-10 part.
The preparation method of suckling piglet specific enzyme of the present invention:
By described protective material, Chinese herbal medicine extract difference micronizing; guarantee that granularity is less than described acidic xylanase and other raw material; then with aspergillus niger culture, acidic xylanase, cellulase, beta-glucanase, amylase, phytase; evenly mix; finally add activator, after mixing, pack the suckling piglet specific enzyme that gets product.
Described suckling piglet specific enzyme is applicable to feed-processing plant and plant's autogamy feed, during use, should mix with other raw material in feed, can be in advance the present invention be mixed with a small amount of feed, remix in large quantities of feeds, Direct-fed.Advise that complete diet pellet addition per ton is: 100-150g.
The Aspergillus niger strain of high yield complex enzyme for feed of the present invention is specially aspergillus niger (Aspergillus niger) DM-18, the wild mushroom HYX0022 that Jinshi City Yang You township, Shi Cong Hunan Province detritus soil, the separation of straw compound sample obtain is through through ultraviolet mutagenesis, the mutagenesis of ultraviolet nitrous acid, ultraviolet nitrosoguanidine complex mutation, then mutant strain step-sizing is eliminated, finally strain excellent is obtained producing the aspergillus niger DM-18 of complex enzyme for feed through leavening property test screen.This bacterial strain is preserved in Chinese Typical Representative culture collection center on November 3rd, 2013 and (is called for short CCTCC, address is: Wuchang District, Wuhan City, Hubei Province Luo Jia Shan Wuhan University Life Science College postcode: 430072), preserving number is CCTCC NO:M2013541, and Classification And Nomenclature is aspergillus niger (Aspergillus niger) DM-18.
The aspergillus niger DM-18 of high yield complex enzyme for feed provided by the invention has following microbial characteristic:
1, morphological feature:
Aspergillus niger DM-18, biology morphology is for comprising conidium, born of the same parents' stalk, top capsule, producing several parts such as born of the same parents' structure.Conidial head is spherical to Radiation, diameter 150-450 μ m, and conidiophore betides matrix.Born of the same parents obstruct stem 1000-3000 (length) * 12-20 (diameter) μ m, yellow or tawny, and wall is level and smooth; Spherical or the almost spherical of top capsule, diameter 35-50 μ m, surface can be educated comprehensively; Produce born of the same parents' structure double-deck, metulae 15-20 (length) * 3-4.0 (diameter) μ m, bottle stalk 6-8 (length) * 2-4 (diameter) μ m, conidium is spherical or subsphaeroidal, less, diameter 3.5-5.0 μ m, brown, wall is coarse.
2, cultivate and learn feature:
Aspergillus niger DM-18 grows rapidly on wort agar substratum, 28 ℃ of 4 days diameter 65mm; Quality velvet shape or be slightly with cotton-shaped; Conidium structure is a large amount of, and brown-black has a small amount of transudate; Bacterium colony reverse side is slightly yellow.
3, physiological and biochemical property:
Aspergillus niger DM-18 can be at potato, Semen Maydis powder, Zulkovsky starch, the upper growth such as molasses, optimum pH 4.6, optimum growth temperature 28-34 ℃, the suitableeest product enzyme temperature 28-30 ℃.
The triage techniques route of aspergillus niger DM-18 is: the preparation → mutagenic treatment → plate isolation → primary dcreening operation of original strain separation, screening and evaluation → starting strain → slant culture → spore suspension → multiple sieve → sieve → expansion experiment (leavening property mensuration) again again.
Beneficial effect:
1. in aspergillus niger culture of the present invention, aspergillus strain can suppress the growth of pathogenic colon bacillus in growth and fermenting process; Suppress flavus Growth and reproduction, decompose aflatoxin; Promote growth of animal, regulate gastrointestinal bacterial flora balance, improve overall immunity, be particularly suitable for the interpolation of the feed for piglet.
2. to take the bacterial strain aspergillus niger DM-18 of high yield complex enzyme for feed be starting strain to the aspergillus niger culture in the present invention, and carry out medium optimization and zymotechnique and improve, adopt the zymotechnique of gradient cooling and gradient increased temperature, appended inoculation and feed supplement in good time simultaneously, especially the zymotechnique of gradient cooling and gradient increased temperature has significantly improved the anti-stress ability of starting strain, causes the enzymatic productivity of bacterial classification to manifest to greatest extent.And the present invention forms and implements full optimization substratum, the root of large-flowered skullcap, the radix bupleuri with the former effect of anti-heat stress have been added, added to have and adjusted and repaired body function, the immunologic function of enhancing body, the Radix Astragali with micro-Ecological regulation services, Radix Codonopsis etc., further strengthened body function, adaptation of virus and the common interoperability of microorganism under same yeasting, and then strengthened the metabolic function of microorganism, make the present invention produce that feeding composite enzyme activity is high, tolerable temperature is higher, stability is strong, be suitable for suitability for industrialized production.
3. in aspergillus niger culture fermentation broth crude enzyme liquid of the present invention, contain plurality of enzymes vigor, wherein, proteinase activity 6500-6700U/ml, mannosans enzyme activity 1500-1700U/ml, α-galactase vigor 1200-1300U/ml, pectinase activity 1000-1200U/ml.Fermented liquid crude enzyme liquid is carried out to heat stability test and the pH stability test of enzyme, test-results shows: at 30-75 ℃, during pH value 2-7, each enzyme all has higher enzyme activity, reaches more than 80%.Complex enzyme for feed thermostability of the present invention and pH stability are strong, are more applicable to fodder industry complete processing demand, and the feed enzyme loss alive after processing is low.
4. it is composite that the protective material in suckling piglet specific enzyme of the present invention adopts polysaccharide, inorganic salt and amino acid science, effectively slowed down the moisture regain of composite raw material; Can strengthen composite enzyme simultaneously resistance toly freeze, resistance toheat, keep identical enzyme activity, its heat resisting temperature can improve 20-30 ℃, resistance to freezing temp can reduce 10-15 ℃, effectively prevented the loss of composite enzyme enzyme activity in transportation, preservation and use procedure, extended the quality guaranteed period of composite enzyme, reached same enzyme activity, the like product quality guaranteed period can extend 2-3.
5. suckling piglet specific enzyme of the present invention is added inorganic salt as activator; created the top condition of enzyme catalysis; given full play to the vigor of each enzyme component of composite enzyme; the macromolecular substance such as starch in feed, protein, Mierocrystalline cellulose, phytic acid have thoroughly effectively been decomposed; greatly alleviated the digestion burden of livestock and poultry animal; improved the growth rate of raw material availability and livestock and poultry, effectively prevented the environmental pollution that feces of livestock and poultry causes, protected feeding environment simultaneously.
6. the Chinese herbal medicine extract that suckling piglet specific enzyme of the present invention is added both can extend the quality guaranteed period of composite raw material, can improve again the immunizing power of raising livestock and poultry, effectively prevented the generation of livestock and poultry epidemic disease.
7. the common synergy of protective material and zymin, activator and zymin, Chinese herbal medicine extract and zymin in suckling piglet specific enzyme of the present invention; enzyme activity and the effect of composite enzyme are brought into play to greatest extent; and the utilization ratio of feed and the growth rate of animal have been improved accordingly; strengthened appetite and the resistance against diseases of animal, extended the quality guaranteed period of composite enzyme and protected environment.
Embodiment
Below by specific embodiment narration the present invention.Unless stated otherwise, in the present invention, technique means used 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 are only limited by claims.To those skilled in the art, do not deviating under the prerequisite of essence of the present invention and scope various changes that the material component in these embodiments and consumption are carried out or change and also belong to protection scope of the present invention.
Embodiment 1
Containing the suckling piglet specific enzyme of complex enzyme for feed, by the raw material of following parts by weight, formed:
50 parts of aspergillus niger cultures, 20 parts of acidic xylanases, 20 parts of cellulases, 17 parts of beta-glucanases, 17 parts of amylase, 13 parts of phytases, 12 parts of Chinese herbal medicine extracts, 13 parts of protective materials, 12 parts of activator.
Described acidic xylanase, cellulase, beta-glucanase, amylase, phytase are food-grade enzyme preparation;
Described aspergillus niger culture preparation method comprises the steps:
(1) actication of culture
The slant strains of intact aspergillus niger DM-18 is inoculated in to slant medium, cultivates 36h for 28 ℃ and carry out actication of culture, so activate 2 times;
Described slant medium consists of: casein food grade 4g, dipotassium hydrogen phosphate 2g, magnesium chloride 0.6g, Repone K 0.8g, ferrous sulfate 0.02g, glucose 20g, agar 20g, Chinese herbal medicine powder 5g, distilled water l000mL, 5.8,121 ℃ of sterilizing 20min of pH value;
The preparation method of described Chinese herbal medicine powder is as follows:
Take 20 parts of the Radixs Astragali; 10 parts of Radix Codonopsis; 10 parts of radix bupleuri; 10 parts of the roots of large-flowered skullcap; Respectively said herbal medicine being crushed to particle diameter is below 2 millimeters, then in container, evenly mix and add the water of 3 times of weight, control temperature 70 C and keep 2h, add the mixture of 2 times of weight ethanol of mixture and propyl alcohol, control temperature to 60 ℃ and keep 3h, filter; Filtrate vacuum concentration postlyophilization obtains Chinese herbal medicine powder.
The mass ratio of described ethanol and propyl alcohol is 1:1.
(2) liquid seeds enlarged culturing
1. first order seed is cultivated: the Aspergillus usamii slant strains after step (1) activation, with spore under aseptic washing, is accessed in 500 ml shake flasks to 100 milliliters of liquid seed culture medium loading amounts, 28 ℃, 80rpm shaking table cultivation 36h;
2. secondary seed is cultivated: by first order seed, according in 500 milliliters of secondary seed shaking flasks of inoculum size access of 10%, culture condition is identical with first order seed;
3. three grades of seed culture: by secondary seed with in 5000 milliliters of three grades of seed shaking flasks of 8% inoculum size access, 1000 milliliters of liquid nutrient medium loading amounts, 28 ℃, 80rpm shaking table are cultivated 36h;
4. seed tank culture: three grades of seeds be take to the first class seed pot that 8% inoculum size access cubic capacity is 150L, seed tank culture base loading amount 100L, controlling pH value is 5,28 ℃ of culture temperature, stirring velocity 200rpm, ventilation (V/V) 1:0.8, incubation time 36h, dissolved oxygen 10%;
Described one-level, secondary, three grades of seed culture mediums consist of: dipotassium hydrogen phosphate 2g, magnesium chloride 0.6g, Repone K 0.8g, ferrous sulfate 0.02g, glucose 20g, Chinese herbal medicine powder 15g, trehalose 10g, distilled water l000mL, 5.5,121 ℃ of sterilizing 20min of pH value.
Described seed tank culture base consists of: Semen Maydis powder 50-60g, bean powder 15-25g, wheat bran 10-15g, fish meal 10g, calcium chloride 6g, ammonium chloride 1g, Sodium phosphate dibasic 1g, Chinese herbal medicine powder 15g, trehalose 10g, pure water l000mL, 5,121 ℃ of sterilizing 20min of pH value.
Described seeding tank fermented liquid spore concentration is 7.0x10 8individual/ml;
(3) ferment tank
Seeding tank liquid seeds in step (2) is accessed to fermentor tank, 28 ℃ of culture temperature, stirring velocity 200r/m, ventilation (V/V) 1:1, incubation time 10h with 6% inoculum size; Then with 1 ℃/h rate of temperature fall slow cooling to 10 ℃, constant temperature culture 15h; Continuation, with 1 ℃/h rate of temperature fall slow cooling to 2 ℃, now, is appended access fermentor tank, constant temperature culture 20h by seeding tank liquid seeds in step (2) with 4% inoculum size; Finally with 1 ℃/h temperature rise rate, be slowly warming up to 10 ℃, constant temperature culture 15h; Continuation is slowly warming up to 30 ℃ with 1 ℃/h temperature rise rate, constant temperature culture 15h;
Dissolved oxygen is controlled: by adjusting mixing speed and ventilation, control dissolved oxygen 15%;
PH controls: by mending ammoniacal liquor or dilute phosphoric acid, in controlled fermentation process, pH value remains on 5;
Control of additive raw material: when the reducing sugar content in fermented liquid is down to 3mg/ml-8mg/ml, start to add supplemented medium, feed supplement amount be take and maintained fermented liquid reducing sugar content as 2mg/ml-5mg/ml;
Put tank standard: the old and feeble self-dissolving of 60% thalline, enzyme activity increasess slowly.
Described fermention medium consists of: Semen Maydis powder 50g, bean powder 15g, wheat bran 10g, fish meal 10g, calcium chloride 6g, ammonium chloride 1g, Sodium phosphate dibasic 1g, Chinese herbal medicine powder 30g, trehalose 10g, pure water 1000mL, 5,121 ℃ of sterilizing 20min of pH value.
Described supplemented medium weight consists of: maltodextrin 20%, and Semen Maydis powder 10%, bean powder 15%, fish meal 1.0%, calcium chloride 0.6%, ammonium chloride 0.1%, Sodium phosphate dibasic 0.1%, Chinese herbal medicine powder 5%, insufficient section pure water is supplied, 5,121 ℃ of sterilizing 30min of pH value.
(4) fermented liquid obtains aspergillus niger wet thallus and centrifugate through centrifugation, centrifugate obtains solid content 30% concentrated solution in the concentrated moisture of removing of 15 ℃ of loop ultrafiltrations, concentrated solution evenly mixes with mass ratio 5:1 with aspergillus niger wet thallus, then through vacuum lyophilization, micronizing, obtains aspergillus niger culture.
In above-mentioned preparation method, in fermented liquid crude enzyme liquid, contain plurality of enzymes vigor, wherein, proteinase activity 6500U/ml, mannosans enzyme activity 1500U/ml, α-galactase vigor 1200U/ml, pectinase activity 1000U/ml.
The preparation method of described Chinese herbal medicine extract is: respectively Chinese herbal medicine powder being broken to particle diameter is below 2 millimeters, then in container, evenly mix and add the water of 5 times of weight, control 80 ℃ of temperature and keep 3h, then being cooled to 53 ℃, adding mixing enzyme preparation to carry out enzymolysis, is 6.2 with newborn acid for adjusting pH value, enzymolysis 3h, finally add the mixture of 2 times of weight ethanol of mixture and propyl alcohol, control temperature to 69 ℃ and keep 4h, filter; It is more than 20% that filtrate decompression is concentrated into solid content, and then lyophilize obtains Chinese herbal medicine extract.
The parts by weight of described herbal medicine consist of: 25 parts of sea-buckthorns, 25 parts of Semen Cassiaes, 17 parts of matrimony vines, 13 parts of Chinese yams, 8 parts of Root of coastal Glehnia, 7 parts of Fructus Viticis Negundo, 4 parts of radix polygonati officinalis, 4 parts of the seeds of Job's tears, 4 parts of Fructus Hordei Germinatus, 4 parts of sweet osmanthus, 4 parts of the Radixs Astragali;
Mixed enzyme addition is the 5-10% of mixture gross weight;
The parts by weight of described mixed enzyme consist of: 15 parts of endo-beta-glucanases, 15 parts of outer beta-glucanases, 13 parts of beta-glucosidases, 13 parts of zytases, 13 parts of pentosanases, 25 parts of Pullulanases, 12 parts of beta-amylases, 13 parts of neutral proteases, 13 parts of aspartic proteases, 7 parts of superoxide-dismutases, 7 parts of glucose oxidases, 7 parts of acid phosphatases;
Described protective material is comprised of the raw material of following parts by weight: 25 parts of trehaloses, NaCl25 part, (NH 4) 2sO 413 parts, 12 parts of halfcystines.
Described activator is that the inorganic salt by following quality component evenly mix: 35 parts of zinc chloride, 15 parts, calcium chloride, 15 parts, sodium sulfate, 7 parts, magnesium chloride.
The preparation method of suckling piglet specific enzyme of the present invention:
By described protective material, Chinese herbal medicine extract difference micronizing; guarantee that granularity is less than described acidic xylanase and other raw material; then with aspergillus niger culture, acidic xylanase, cellulase, beta-glucanase, amylase, phytase; evenly mix; finally add activator, after mixing, pack the suckling piglet specific enzyme that gets product.
Embodiment 2
Containing the suckling piglet specific enzyme of complex enzyme for feed, by the raw material of following parts by weight, formed:
40 parts of complex enzyme for feed, 15 parts of acidic xylanases, 15 parts of cellulases, 15 parts of beta-glucanases, 15 parts of amylase, 10 parts of phytases, 10 parts of Chinese herbal medicine extracts, 10 parts of protective materials, 10 parts of activator.
Described acidic xylanase, cellulase, beta-glucanase, amylase, phytase are food-grade enzyme preparation;
Described aspergillus niger culture preparation method comprises the steps:
(1) actication of culture
The slant strains of intact aspergillus niger DM-18 is inoculated in to slant medium, cultivates 48h for 34 ℃ and carry out actication of culture, so activate 4 times;
Described slant medium consists of: casein food grade 4g, dipotassium hydrogen phosphate 2g, magnesium chloride 0.6g, Repone K 0.8g, ferrous sulfate 0.02g, glucose 20g, agar 20g, Chinese herbal medicine powder 20g, distilled water 1000mL, 5.8,121 ℃ of sterilizing 20min of pH value.
The preparation method of described Chinese herbal medicine powder is as follows:
Take 30 parts of the Radixs Astragali; 18 parts of Radix Codonopsis; 15 parts of radix bupleuri; 15 parts of the roots of large-flowered skullcap; Respectively said herbal medicine being crushed to particle diameter is below 2 millimeters, then in container, evenly mix and add the water of 6 times of weight, control 90 ℃ of temperature and keep 4h, add the mixture of 3 times of weight ethanol of mixture and propyl alcohol, control temperature to 78 ℃ and keep 4h, filter; Filtrate vacuum concentration postlyophilization obtains Chinese herbal medicine powder.
The mass ratio of described ethanol and propyl alcohol is 1:1.5.
(2) liquid seeds enlarged culturing
1. first order seed is cultivated: the slant strains after step (1) activation, with spore under aseptic washing, is accessed in 500 ml shake flasks to 100 milliliters of liquid seed culture medium loading amounts, 34 ℃, 120rpm shaking table cultivation 48h;
2. secondary seed is cultivated: by first order seed, according in 500 milliliters of secondary seed shaking flasks of inoculum size access of 10%, culture condition is identical with first order seed;
3. three grades of seed culture: by secondary seed with in 5000 milliliters of three grades of seed shaking flasks of 8% inoculum size access, 1000 milliliters of liquid nutrient medium loading amounts, 34 ℃, 120rpm shaking table are cultivated 48h;
4. seed tank culture: three grades of seeds be take to the first class seed pot that 8% inoculum size access cubic capacity is 150L, seed tank culture base loading amount 100L, controlling pH value is 5-7,30 ℃ of culture temperature, stirring velocity 400rpm, ventilation (V/V) 1:1.2, incubation time 48h, dissolved oxygen 20%;
Described one-level, secondary, three grades of seed culture mediums consist of: dipotassium hydrogen phosphate 2g, magnesium chloride 0.6g, Repone K 0.8g, ferrous sulfate 0.02g, glucose 20g, Chinese herbal medicine powder 25g, trehalose 30g, distilled water 1000mL, 5.5,121 ℃ of sterilizing 20min of pH value.
Described seed tank culture base consists of: Semen Maydis powder 60g, bean powder 25g, wheat bran 15g, fish meal 15g, calcium chloride 10g, ammonium chloride 3g, Sodium phosphate dibasic 2g, Chinese herbal medicine powder 20g, trehalose 30g, pure water l000mL, 7,121 ℃ of sterilizing 20min of pH value.
Described seeding tank fermented liquid spore concentration is 8.0x10 8individual/ml;
(3) ferment tank
Seeding tank liquid seeds in step (2) is accessed to fermentor tank, 30 ℃ of culture temperature, stirring velocity 700r/m, ventilation (V/V) 1:3, incubation time 15h with 6% inoculum size; Then with 2 ℃/h rate of temperature fall slow cooling to 15 ℃, constant temperature culture 20h; Continuation, with 2 ℃/h rate of temperature fall slow cooling to 5 ℃, now, is appended access fermentor tank, constant temperature culture 30h by seeding tank liquid seeds in step (2) with 4% inoculum size; Finally with 2 ℃/h temperature rise rate, be slowly warming up to 15 ℃, constant temperature culture 20h; Continuation is slowly warming up to 34 ℃ with 2 ℃/h temperature rise rate, constant temperature culture 20h;
Dissolved oxygen is controlled: by adjusting mixing speed and ventilation, control dissolved oxygen 30%;
PH controls: by mending ammoniacal liquor or dilute phosphoric acid, in controlled fermentation process, pH value remains on 6;
Control of additive raw material: when the reducing sugar content in fermented liquid is down to 3mg/ml-8mg/ml, start to add supplemented medium, feed supplement amount be take and maintained fermented liquid reducing sugar content as 2mg/ml-5mg/ml;
Put tank standard: the old and feeble self-dissolving of 80% thalline, enzyme activity increasess slowly.
Described fermention medium consists of: Semen Maydis powder 60g, bean powder 25g, wheat bran 15g, fish meal 15g, calcium chloride 10g, ammonium chloride 3g, Sodium phosphate dibasic 2g, Chinese herbal medicine powder 50g, trehalose 30g, pure water 1000mL, 7,121 ℃ of sterilizing 20min of pH value.
Described supplemented medium weight consists of: maltodextrin 30%, and Semen Maydis powder 20%, bean powder 25%, fish meal 1.5%, calcium chloride 1.0%, ammonium chloride 0.3%, Sodium phosphate dibasic 0.2%, Chinese herbal medicine powder 10%, insufficient section pure water is supplied, 7,123 ℃ of sterilizing 40min of pH value.
(4) fermented liquid obtains aspergillus niger wet thallus and centrifugate through centrifugation, centrifugate obtains solid content 20% concentrated solution in the concentrated moisture of removing of 10 ℃ of loop ultrafiltrations, concentrated solution evenly mixes with mass ratio 1:1 with aspergillus niger wet thallus, then through vacuum lyophilization, micronizing, obtains aspergillus niger culture.
In above-mentioned preparation method, in fermented liquid crude enzyme liquid, contain plurality of enzymes vigor, wherein, proteinase activity 6600U/ml, mannosans enzyme activity 1600U/ml, α-galactase vigor 1240U/ml, pectinase activity 1080U/ml.
The preparation method of described Chinese herbal medicine extract is: respectively Chinese herbal medicine powder being broken to particle diameter is below 2 millimeters, then in container, evenly mix and add the water of 3 times of weight, control temperature 70 C and keep 2h, then being cooled to 45 ℃, adding mixing enzyme preparation to carry out enzymolysis, is 5.5 with newborn acid for adjusting pH value, enzymolysis 2h, finally add the mixture of 0.5 times of weight ethanol of mixture and propyl alcohol, control temperature to 60 ℃ and keep 3h, filter; It is more than 20% that filtrate decompression is concentrated into solid content, and then lyophilize obtains Chinese herbal medicine extract.
The parts by weight of described herbal medicine consist of: 20 parts of sea-buckthorns, 20 parts of Semen Cassiaes, 10 parts of matrimony vines, 10 parts of Chinese yams, 5 parts of Root of coastal Glehnia, 5 parts of Fructus Viticis Negundo, 3 parts of radix polygonati officinalis, 3 parts of the seeds of Job's tears, 3 parts of Fructus Hordei Germinatus, 3 parts of sweet osmanthus, 3 parts of the Radixs Astragali;
Mixed enzyme addition is 5% of mixture gross weight;
The parts by weight of described mixed enzyme consist of: 10 parts of endo-beta-glucanases, 10 parts of outer beta-glucanases, 10 parts of beta-glucosidases, 15 parts of zytases, 15 parts of pentosanases, 20 parts of Pullulanases, 10 parts of beta-amylases, 10 parts of neutral proteases, 10 parts of aspartic proteases, 5 parts of superoxide-dismutases, 5 parts of glucose oxidases, 5 parts of acid phosphatases;
Described protective material is comprised of the raw material of following parts by weight: 20 parts of trehaloses, NaCl20 part, (NH 4) 2sO 410 parts, 10 parts of halfcystines.
Described activator is that the inorganic salt by following quality component evenly mix: 30 parts of zinc chloride, 10 parts, calcium chloride, 10 parts, sodium sulfate, 5 parts, magnesium chloride.
The preparation method of suckling piglet specific enzyme of the present invention is as embodiment 1.
Embodiment 3
Containing the suckling piglet specific enzyme of complex enzyme for feed, by the raw material of following parts by weight, formed:
60 parts of complex enzyme for feed, 25 parts of acidic xylanases, 25 parts of cellulases, 20 parts of beta-glucanases, 0 part of amylase 2,15 parts of phytases, 15 parts of Chinese herbal medicine extracts, 15 parts of protective materials, 15 parts of activator.
Described acidic xylanase, cellulase, beta-glucanase, amylase, phytase are food-grade enzyme preparation;
Described aspergillus niger culture preparation method comprises the steps:
(1) actication of culture
The slant strains of intact aspergillus niger DM-18 is inoculated in to slant medium, cultivates 42h for 30 ℃ and carry out actication of culture, so activate 3 times;
Described slant medium consists of: casein food grade 4g, dipotassium hydrogen phosphate 2g, magnesium chloride 0.6g, Repone K 0.8g, ferrous sulfate 0.02g, glucose 20g, agar 20g, Chinese herbal medicine powder 12g, distilled water 1000mL, 5.8,121 ℃ of sterilizing 20min of pH value.
The preparation method of described Chinese herbal medicine powder is as follows:
Take 25 parts of the Radixs Astragali; 15 parts of Radix Codonopsis; 12 parts of radix bupleuri; 12 parts of the roots of large-flowered skullcap; Respectively said herbal medicine being crushed to particle diameter is below 2 millimeters, then in container, evenly mix and add the water of 5 times of weight, control 80 ℃ of temperature and keep 3h, add the mixture of 3 times of weight ethanol of mixture and propyl alcohol, control temperature to 70 ℃ and keep 4h, filter; Filtrate vacuum concentration postlyophilization obtains Chinese herbal medicine powder.
The mass ratio of described ethanol and propyl alcohol is 1:1.2.
(2) liquid seeds enlarged culturing
1. first order seed is cultivated: the slant strains after step (1) activation, with spore under aseptic washing, is accessed in 500 ml shake flasks to 100 milliliters of liquid seed culture medium loading amounts, 30 ℃, 100rpm shaking table cultivation 42h;
2. secondary seed is cultivated: by first order seed, according in 500 milliliters of secondary seed shaking flasks of inoculum size access of 10%, culture condition is identical with first order seed;
3. three grades of seed culture: by secondary seed with in 5000 milliliters of three grades of seed shaking flasks of 8% inoculum size access, 1000 milliliters of liquid nutrient medium loading amounts, 30 ℃, 100rpm shaking table are cultivated 42h;
4. seed tank culture: three grades of seeds be take to the first class seed pot that 8% inoculum size access cubic capacity is 150L, seed tank culture base loading amount 100L, controlling pH value is 6,30 ℃ of culture temperature, stirring velocity 300rpm, ventilation (V/V) 1:1, incubation time 42h, dissolved oxygen 20%;
Described one-level, secondary, three grades of seed culture mediums consist of: dipotassium hydrogen phosphate 2g, magnesium chloride 0.6g, Repone K 0.8g, ferrous sulfate 0.02g, glucose 20g, Chinese herbal medicine powder 20g, trehalose 20g, distilled water 1000mL, 5.5,121 ℃ of sterilizing 20min of pH value.
Described seed tank culture base consists of: Semen Maydis powder 55g, bean powder 20g, wheat bran 12g, fish meal 12g, calcium chloride 8g, ammonium chloride 2g, Sodium phosphate dibasic 2g, Chinese herbal medicine powder 18g, trehalose 20g, pure water 1000mL, 6,121 ℃ of sterilizing 20min of pH value.
Described seeding tank fermented liquid spore concentration is 7.5x10 8individual/ml;
(3) ferment tank
Seeding tank liquid seeds in step (2) is accessed to fermentor tank, 30 ℃ of culture temperature, stirring velocity 350r/m, ventilation (V/V) 1:2, incubation time 12h with 6% inoculum size; Then with 2 ℃/h rate of temperature fall slow cooling to 12 ℃, constant temperature culture 18h; Continuation, with 2 ℃/h rate of temperature fall slow cooling to 4 ℃, now, is appended access fermentor tank, constant temperature culture 25h by seeding tank liquid seeds in step (2) with 4% inoculum size; Finally with 2 ℃/h temperature rise rate, be slowly warming up to 12 ℃, constant temperature culture 18h; Continuation is slowly warming up to 30 ℃ with 2 ℃/h temperature rise rate, constant temperature culture 18h;
Dissolved oxygen is controlled: by adjusting mixing speed and ventilation, control dissolved oxygen 20%;
PH controls: by mending ammoniacal liquor or dilute phosphoric acid, in controlled fermentation process, pH value remains on 5.2;
Control of additive raw material: when the reducing sugar content in fermented liquid is down to 3mg/ml-8mg/ml, start to add supplemented medium, feed supplement amount be take and maintained fermented liquid reducing sugar content as 2mg/ml-5mg/ml;
Put tank standard: the old and feeble self-dissolving of 70% thalline, enzyme activity increasess slowly.
Described fermention medium consists of: Semen Maydis powder 55g, bean powder 20g, wheat bran 12g, fish meal 12g, calcium chloride 8g, ammonium chloride 2g, Sodium phosphate dibasic 2g, Chinese herbal medicine powder 40g, trehalose 20g, pure water 1000mL, 6,121 ℃ of sterilizing 20min of pH value.
Described supplemented medium weight consists of: maltodextrin 25%, and Semen Maydis powder 15%, bean powder 18%, fish meal 1.2%, calcium chloride 0.8%, ammonium chloride 0.2%, Sodium phosphate dibasic 0.2%, Chinese herbal medicine powder 8%, insufficient section pure water is supplied, 6,123 ℃ of sterilizing 40min of pH value.
(4) fermented liquid obtains aspergillus niger wet thallus and centrifugate through centrifugation, centrifugate obtains solid content 40% concentrated solution in the concentrated moisture of removing of 20 ℃ of loop ultrafiltrations, concentrated solution evenly mixes with mass ratio 10:1 with aspergillus niger wet thallus, then through vacuum lyophilization, micronizing, obtains aspergillus niger culture.
In above-mentioned preparation method, in fermented liquid crude enzyme liquid, contain plurality of enzymes vigor, wherein, proteinase activity 6700U/ml, mannosans enzyme activity 1700U/ml, α-galactase vigor 1300U/ml, pectinase activity 1200U/ml.
The preparation method of described Chinese herbal medicine extract is: respectively Chinese herbal medicine powder being broken to particle diameter is below 2 millimeters, then in container, evenly mix and add the water of 6 times of weight, control 90 ℃ of temperature and keep 4h, then being cooled to 60 ℃, adding mixing enzyme preparation to carry out enzymolysis, is 6.8 with newborn acid for adjusting pH value, enzymolysis 4h, finally add the mixture of 3 times of weight ethanol of mixture and propyl alcohol, control temperature to 78 ℃ and keep 4h, filter; It is more than 20% that filtrate decompression is concentrated into solid content, and then lyophilize obtains Chinese herbal medicine extract.
The parts by weight of described herbal medicine consist of: 30 parts of sea-buckthorns, 30 parts of Semen Cassiaes, 15 parts of matrimony vines, 15 parts of Chinese yams, 10 parts of Root of coastal Glehnia, 10 parts of Fructus Viticis Negundo, 5 parts of radix polygonati officinalis, 5 parts of the seeds of Job's tears, 5 parts of Fructus Hordei Germinatus, 5 parts of sweet osmanthus, 5 parts of the Radixs Astragali;
Mixed enzyme addition is 10% of mixture gross weight;
The parts by weight of described mixed enzyme consist of: 20 parts of endo-beta-glucanases, 20 parts of outer beta-glucanases, 15 parts of beta-glucosidases, 20 parts of zytases, 20 parts of pentosanases, 30 parts of Pullulanases, 15 parts of beta-amylases, 15 parts of neutral proteases, 15 parts of aspartic proteases, 10 parts of superoxide-dismutases, 10 parts of glucose oxidases, 10 parts of acid phosphatases;
Described protective material is comprised of the raw material of following parts by weight: 30 parts of trehaloses, NaCl30 part, (NH 4) 2sO 415 parts, 15 parts of halfcystines.
Described activator is that the inorganic salt by following quality component evenly mix: 40 parts of zinc chloride, 20 parts, calcium chloride, 20 parts, sodium sulfate, 10 parts, magnesium chloride.
The preparation method of suckling piglet specific enzyme of the present invention is as embodiment 1.
The thermal stability analysis of embodiment 4 complex enzyme for feed
Thermostability to complex enzyme for feed is analyzed, embodiment 3 fermented liquid crude enzyme liquids are placed in respectively at 30 ℃, 40 ℃, 45 ℃, 50 ℃, 55 ℃, 60 ℃, 65 ℃, 70 ℃, 75 ℃, and in 10 minutes sampling and measuring complex enzyme for feed, the enzyme of each constitutive enzyme is lived.At 30 ℃, 40 ℃, 45 ℃, 50 ℃, 60 minutes each constitutive enzyme enzymes are lived and are not declined.At 55 ℃, 60 ℃ and 65 ℃, each constitutive enzyme enzyme work in 30 minutes drops to constitutive enzyme each constitutive enzyme enzyme work in 95%, 60 minute alive and drops to 85%.At 70 ℃, each constitutive enzyme enzyme work in 30 minutes drops to constitutive enzyme each constitutive enzyme enzyme work in 90%, 60 minute alive and drops to 85%.At 75 ℃, each constitutive enzyme enzyme work in 85%, 60 minute that each constitutive enzyme enzyme work in 30 minutes drops to constitutive enzyme work drops to 80% of constitutive enzyme work, and compared with prior art, similarity condition is issued to same enzyme tolerable temperature alive and has on average improved 5-10 ℃.
The pH stability analysis of embodiment 5 complex enzyme for feed
PH stability to complex enzyme for feed enzyme is analyzed, and embodiment 3 fermented liquid crude enzyme liquids are placed in respectively to pH value 2.0,2.5,3.5,4.5,5.5,6.0,6.5,7.0 times, and in 10 minutes sampling and measuring complex enzyme for feed, the enzyme of each constitutive enzyme is lived.Crude enzyme liquid pH value 5.5,6.0,6.5,7.0 times, 60 minutes each constitutive enzyme enzymes are lived and are not declined.PH value 4.5,3.5 times, each constitutive enzyme enzyme work in 95%, 60 minute that each constitutive enzyme enzyme work in 30 minutes drops to constitutive enzyme work drops to 85% of constitutive enzyme work.PH value 2.5 times, each constitutive enzyme enzyme work in 30 minutes drops to that constitutive enzyme lives within 90%, 60 minute, drop to that constitutive enzyme lives 85%.PH value 2.0 times, each constitutive enzyme enzyme work in 30 minutes drops to that constitutive enzyme lives within 85%, 60 minute, drop to that constitutive enzyme lives 80%, compared with prior art, it is more wide in range that similarity condition is issued to the same enzyme resistance to pH value of living.
The result of use test of embodiment 6 embodiment of the present invention 1 suckling piglet specific enzyme in weanling pig
Test method:
Subjects is a certain large-scale regular plant in Hunan mean body weight is 180 of the healthy weanling pigs of (7.16 ± 0.15) kg, and statistical analysis body weight difference is not remarkable.Adopt single-factor randomized design, by 180 healthy weanling pigs, by male and female half and half, be divided into 3 groups (control group 1, control group 2 and test group), every group of 6 repetitions, each repeats 10 pigs.The product 100g of the present invention that in test group feed, interpolation embodiment 1 per ton makes, interpolation market milk piglet specific enzyme 100g per ton in control group 1 feed, control group 2 does not add product of the present invention.Feed continuously 30 days, growth indexes is as table 1
Table 1
Project Test group Control group 1 Deviation Control group 2 Deviation
Starting weight (kg) 7.23 7.18 -- 7.09 --
End heavy (kg) 21.58 19.62 -- 17.34 --
Day weight gain (g/d ﹒ head) 478 415 15.18% 342 39.77%
Daily ingestion amount (g/d ﹒ head) 598.5 605.8 -1.20% 612.5 -22.86%
Feedstuff-meat ratio 1.25 1.46 -14.38% 1.79 -30.17%
Diarrhea rate (%) 1 11 -90.90% 13 -92.31%
Test-results shows, in weanling pig daily ration, adds this product, and the day weight gain of piglet has improved respectively 15.18% and 39.77% than control group 1 and control group 2; Feedstuff-meat ratio has reduced respectively 14.38% and 30.17%; Diarrhea rate has reduced respectively 90.90% and 92.31%, and product of the present invention can significantly improve piglet body immunity, increases cultivation quality and benefits.

Claims (10)

1. the suckling piglet specific enzyme containing complex enzyme for feed, raw material by following parts by weight forms: aspergillus niger culture 40-60 part, acidic xylanase 15-25 part, cellulase 15-25 part, beta-glucanase 15-20 part, amylase 15-20 part, phytase 10-15 part, Chinese herbal medicine extract 10-15 part, protective material 10-15 part, activator 10-15 part;
The preparation method of described aspergillus niger culture is as follows: by the slant strains of intact aspergillus niger DM-18 through actication of culture and one-level, secondary, three grades and seeding tank liquid seeds step by step enlarged culturing obtain liquid seeds, with 6% inoculum size access fermentor tank, culture temperature 28-30 ℃, stirring velocity 200-700r/m, ventilation (V/V) 1:1-3, incubation time 10-15h; Then with 1-2 ℃/h rate of temperature fall slow cooling to 10-15 ℃, constant temperature culture 15-20h; Continuation to 2-5 ℃, now, is appended access fermentor tank, constant temperature culture 20-30h by liquid seeds with 4% inoculum size with 1-2 ℃/h rate of temperature fall slow cooling; Finally with 1-2 ℃/h temperature rise rate, be slowly warming up to 10-15 ℃, constant temperature culture 15-20h; Continuation is slowly warming up to 30-33 ℃, constant temperature culture 15-20h with 1-2 ℃/h temperature rise rate; Fermented liquid obtains aspergillus niger wet thallus and centrifugate through centrifugation, centrifugate obtains solid content 20-40% concentrated solution in the concentrated moisture of removing of 10-20 ℃ of loop ultrafiltration, concentrated solution evenly mixes with mass ratio 1-10:1 with aspergillus niger wet thallus, then through vacuum lyophilization, micronizing, obtains aspergillus niger culture;
Described aspergillus niger DM-18 is preserved in Chinese Typical Representative culture collection center on November 3rd, 2013, and preserving number is CCTCC NO:M2013541;
Described protective material is comprised of the raw material of following parts by weight: trehalose 20-30 part, NaCl20-30 part, (NH 4) 2sO 410-15 part, halfcystine 10-15 part;
Described activator is that the inorganic salt by following mass fraction evenly mix: zinc chloride 30-40 part, calcium chloride 10-20 part, sodium sulfate 10-20 part, magnesium chloride 5-10 part.
2. a kind of suckling piglet specific enzyme containing complex enzyme for feed according to claim 1, it is characterized in that, the preparation method of described Chinese herbal medicine extract is: respectively Chinese herbal medicine powder being broken to particle diameter is below 2 millimeters, then in container, evenly mix and add the water of 3-6 times of weight, control temperature 70 C～90 ℃ and keep 2～4h, then be cooled to 45-60 ℃, add the mixing enzyme preparation of mixture gross weight 5-10% to carry out enzymolysis, with newborn acid for adjusting pH value, be 5.5-6.8, enzymolysis 2-4h, finally add the mixture of mixture 0.5-3 times of weight ethanol and propyl alcohol, control temperature to 60 ℃～78 ℃ and keep 3～4h, filter, it is more than 20% that filtrate decompression is concentrated into solid content, and then lyophilize obtains Chinese herbal medicine extract,
The parts by weight of described herbal medicine consist of: sea-buckthorn 20-30 part, Semen Cassiae 20-30 part, matrimony vine 10-15 part, Chinese yam 10-15 part, Root of coastal Glehnia 5-10 part, Fructus Viticis Negundo 5-10 part, radix polygonati officinalis 3-5 part, seed of Job's tears 3-5 part, Fructus Hordei Germinatus 3-5 part, sweet osmanthus 3-5 part, Radix Astragali 3-5 part;
The parts by weight of described mixed enzyme consist of: endo-beta-glucanase 10-20 part, outer beta-glucanase 10-20 part, beta-glucosidase 10-15 part, zytase 15-20 part, pentosanase 15-20 part, Pullulanase 20-30 part, beta-amylase 10-15 part, neutral protease 10-15 part, aspartic protease 10-15 part, superoxide-dismutase 5-10 part, glucose oxidase 5-10 part, acid phosphatase 5-10 part.
3. a kind of suckling piglet specific enzyme containing complex enzyme for feed according to claim 1, is characterized in that, the slant medium of preparing aspergillus niger culture consists of: casein food grade 4g, dipotassium hydrogen phosphate 2g, magnesium chloride 0.6g, Repone K 0.8g, ferrous sulfate 0.02g, glucose 20g, agar 20g, Chinese herbal medicine powder 5-20g, distilled water l000mL, 5.8,121 ℃ of sterilizing 20min of pH value.
4. a kind of suckling piglet specific enzyme containing complex enzyme for feed according to claim 1, is characterized in that, the seed tank culture base of preparing aspergillus niger culture consists of: Semen Maydis powder 50-60g, bean powder 15-25g, wheat bran 10-15g, fish meal 10-15g, calcium chloride 6-10g, ammonium chloride 1-3g, Sodium phosphate dibasic 1-2g, Chinese herbal medicine powder 15-20g, trehalose 10-30g, pure water l000mL, pH value 5-7,121 ℃ of sterilizing 20min.
5. a kind of suckling piglet specific enzyme containing complex enzyme for feed according to claim 1, is characterized in that, while preparing aspergillus niger culture, seeding tank fermented liquid spore concentration is 7.0x10 8-8.0x10 8individual/ml.
6. a kind of suckling piglet specific enzyme containing complex enzyme for feed according to claim 1, is characterized in that, the fermention medium of preparing aspergillus niger culture consists of: Semen Maydis powder 50-60g, bean powder 15-25g, wheat bran 10-15g, fish meal 10-15g, calcium chloride 6-10g, ammonium chloride 1-3g, Sodium phosphate dibasic 1-2g, Chinese herbal medicine powder 30-50g, trehalose 10-30g, pure water l000mL, pH value 5-7,121 ℃ of sterilizing 20min.
7. according to the arbitrary described a kind of suckling piglet specific enzyme containing complex enzyme for feed of claim 3-6, it is characterized in that, the preparation method of described Chinese herbal medicine powder is as follows: take Radix Astragali 20-30 part; Radix Codonopsis 10-18 part; Radix bupleuri 10-15 part; Root of large-flowered skullcap 10-15 part; Respectively said herbal medicine being crushed to particle diameter is below 2 millimeters, then in container, evenly mix and add the water of 3-6 times of weight, control temperature 70 C～90 ℃ and keep 2～4h, add the mixture of 2-3 times of weight ethanol of mixture and propyl alcohol, the mass ratio of ethanol and propyl alcohol is 1:1-1.5, control temperature to 60 ℃～78 ℃ and keep 3～4h, filter; Filtrate vacuum concentration postlyophilization obtains Chinese herbal medicine powder.
8. the preparation method containing the suckling piglet specific enzyme of complex enzyme for feed; it is characterized in that; by protective material, Chinese herbal medicine extract difference micronizing; guarantee that granularity is less than described acidic xylanase and other raw material; then evenly mix with aspergillus niger culture, acidic xylanase, cellulase, beta-glucanase, amylase, phytase; finally add activator, after mixing, pack the suckling piglet specific enzyme that gets product.
9. the preparation method of a kind of suckling piglet specific enzyme containing complex enzyme for feed according to claim 8, it is characterized in that, described suckling piglet specific enzyme is comprised of the raw material of following parts by weight: aspergillus niger culture 40-60 part, acidic xylanase 15-25 part, cellulase 15-25 part, beta-glucanase 15-20 part, amylase 15-20 part, phytase 10-15 part, Chinese herbal medicine extract 10-15 part, protective material 10-15 part, activator 10-15 part;
The preparation method of described aspergillus niger culture is as follows: by the slant strains of intact aspergillus niger DM-18 through actication of culture and one-level, secondary, three grades and seeding tank liquid seeds step by step enlarged culturing obtain liquid seeds, with 6% inoculum size access fermentor tank, culture temperature 28-30 ℃, stirring velocity 200-700r/m, ventilation (V/V) 1:1-3, incubation time 10-15h; Then with 1-2 ℃/h rate of temperature fall slow cooling to 10-15 ℃, constant temperature culture 15-20h; Continuation to 2-5 ℃, now, is appended access fermentor tank, constant temperature culture 20-30h by liquid seeds with 4% inoculum size with 1-2 ℃/h rate of temperature fall slow cooling; Finally with 1-2 ℃/h temperature rise rate, be slowly warming up to 10-15 ℃, constant temperature culture 15-20h; Continuation is slowly warming up to 30-33 ℃, constant temperature culture 15-20h with 1-2 ℃/h temperature rise rate; Fermented liquid obtains aspergillus niger wet thallus and centrifugate through centrifugation, centrifugate obtains solid content 20-40% concentrated solution in the concentrated moisture of removing of 10-20 ℃ of loop ultrafiltration, concentrated solution evenly mixes with mass ratio 1-10:1 with aspergillus niger wet thallus, then through vacuum lyophilization, micronizing, obtains aspergillus niger culture;
Described protective material is comprised of the raw material of following parts by weight: trehalose 20-30 part, NaCl20-30 part, (NH 4) 2sO 410-15 part, halfcystine 10-15 part;
Described activator is that the inorganic salt by following quality component evenly mix: zinc chloride 30-40 part, calcium chloride 10-20 part, sodium sulfate 10-20 part, magnesium chloride 5-10 part.
10. the preparation method of a kind of suckling piglet specific enzyme containing complex enzyme for feed according to claim 9, it is characterized in that, the described suckling piglet specific enzyme containing complex enzyme for feed is comprised of the raw material of following parts by weight: 50 parts of aspergillus niger cultures, 20 parts of acidic xylanases, 20 parts of cellulases, 17 parts of beta-glucanases, 17 parts of amylase, 13 parts of phytases, 12 parts of Chinese herbal medicine extracts, 13 parts of protective materials, 12 parts of activator;
Described aspergillus niger culture preparation method comprises the steps:
(1) actication of culture
The slant strains of intact aspergillus niger DM-18 is inoculated in to slant medium, cultivates 36h for 28 ℃ and carry out actication of culture, so activate 2 times;
Described slant medium consists of: casein food grade 4g, dipotassium hydrogen phosphate 2g, magnesium chloride 0.6g, Repone K 0.8g, ferrous sulfate 0.02g, glucose 20g, agar 20g, Chinese herbal medicine powder 5g, distilled water 1000mL, 5.8,121 ℃ of sterilizing 20min of pH value;
The preparation method of described Chinese herbal medicine powder is as follows:
Take 20 parts of the Radixs Astragali; 10 parts of Radix Codonopsis; 10 parts of radix bupleuri; 10 parts of the roots of large-flowered skullcap; Respectively said herbal medicine being crushed to particle diameter is below 2 millimeters, then in container, evenly mix and add the water of 3 times of weight, control temperature 70 C and keep 2h, add the mixture of 2 times of weight ethanol of mixture and propyl alcohol, control temperature to 60 ℃ and keep 3h, filter; Filtrate vacuum concentration postlyophilization obtains Chinese herbal medicine powder;
The mass ratio of described ethanol and propyl alcohol is 1:1;
(2) liquid seeds enlarged culturing
1. first order seed is cultivated: the Aspergillus usamii slant strains after step (1) activation, with spore under aseptic washing, is accessed in 500 ml shake flasks to 100 milliliters of liquid seed culture medium loading amounts, 28 ℃, 80rpm shaking table cultivation 36h;
2. secondary seed is cultivated: by first order seed, according in 500 milliliters of secondary seed shaking flasks of inoculum size access of 10%, culture condition is identical with first order seed;
3. three grades of seed culture: by secondary seed with in 5000 milliliters of three grades of seed shaking flasks of 8% inoculum size access, 1000 milliliters of liquid nutrient medium loading amounts, 28 ℃, 80rpm shaking table are cultivated 36h;
4. seed tank culture: three grades of seeds be take to the first class seed pot that 8% inoculum size access cubic capacity is 150L, seed tank culture base loading amount 100L, controlling pH value is 5,28 ℃ of culture temperature, stirring velocity 200rpm, ventilation (V/V) 1:0.8, incubation time 36h, dissolved oxygen 10%;
Described one-level, secondary, three grades of seed culture mediums consist of: dipotassium hydrogen phosphate 2g, magnesium chloride 0.6g, Repone K 0.8g, ferrous sulfate 0.02g, glucose 20g, Chinese herbal medicine powder 15g, trehalose 10g, distilled water 1000mL, 5.5,121 ℃ of sterilizing 20min of pH value;
Described seed tank culture base consists of: Semen Maydis powder 50-60g, bean powder 15-25g, wheat bran 10-15g, fish meal 10g, calcium chloride 6g, ammonium chloride 1g, Sodium phosphate dibasic 1g, Chinese herbal medicine powder 15g, trehalose 10g, pure water l000mL, 5,121 ℃ of sterilizing 20min of pH value;
Described seeding tank fermented liquid spore concentration is 7.0x10 8individual/ml;
(3) ferment tank
Seeding tank liquid seeds in step (2) is accessed to fermentor tank, 28 ℃ of culture temperature, stirring velocity 200r/m, ventilation (V/V) 1:1, incubation time 10h with 6% inoculum size; Then with 1 ℃/h rate of temperature fall slow cooling to 10 ℃, constant temperature culture 15h; Continuation, with 1 ℃/h rate of temperature fall slow cooling to 2 ℃, now, is appended access fermentor tank, constant temperature culture 20h by seeding tank liquid seeds in step (2) with 4% inoculum size; Finally with 1 ℃/h temperature rise rate, be slowly warming up to 10 ℃, constant temperature culture 15h; Continuation is slowly warming up to 30 ℃ with 1 ℃/h temperature rise rate, constant temperature culture 15h;
Dissolved oxygen is controlled: by adjusting mixing speed and ventilation, control dissolved oxygen 15%;
PH controls: by mending ammoniacal liquor or dilute phosphoric acid, in controlled fermentation process, pH value remains on 5;
Control of additive raw material: when the reducing sugar content in fermented liquid is down to 3mg/ml-8mg/ml, start to add supplemented medium, feed supplement amount be take and maintained fermented liquid reducing sugar content as 2mg/ml-5mg/ml;
Put tank standard: the old and feeble self-dissolving of 60% thalline, enzyme activity increasess slowly;
Described fermention medium consists of: Semen Maydis powder 50g, bean powder 15g, wheat bran 10g, fish meal 10g, calcium chloride 6g, ammonium chloride 1g, Sodium phosphate dibasic 1g, Chinese herbal medicine powder 30g, trehalose 10g, pure water 1000mL, 5,121 ℃ of sterilizing 20min of pH value;
Described supplemented medium weight consists of: maltodextrin 20%, and Semen Maydis powder 10%, bean powder 15%, fish meal 1.0%, calcium chloride 0.6%, ammonium chloride 0.1%, Sodium phosphate dibasic 0.1%, Chinese herbal medicine powder 5%, insufficient section pure water is supplied, 5,121 ℃ of sterilizing 30min of pH value;
(4) fermented liquid obtains aspergillus niger wet thallus and centrifugate through centrifugation, centrifugate obtains solid content 30% concentrated solution in the concentrated moisture of removing of 15 ℃ of loop ultrafiltrations, concentrated solution evenly mixes with mass ratio 5:1 with aspergillus niger wet thallus, then through vacuum lyophilization, micronizing, obtains aspergillus niger culture;
The preparation method of described Chinese herbal medicine extract is: respectively Chinese herbal medicine powder being broken to particle diameter is below 2 millimeters, then in container, evenly mix and add the water of 5 times of weight, control 80 ℃ of temperature and keep 3h, then being cooled to 53 ℃, adding mixing enzyme preparation to carry out enzymolysis, is 6.2 with newborn acid for adjusting pH value, enzymolysis 3h, finally add the mixture of 2 times of weight ethanol of mixture and propyl alcohol, control temperature to 69 ℃ and keep 4h, filter; It is more than 20% that filtrate decompression is concentrated into solid content, and then lyophilize obtains Chinese herbal medicine extract;
The parts by weight of described herbal medicine consist of: 25 parts of sea-buckthorns, 25 parts of Semen Cassiaes, 17 parts of matrimony vines, 13 parts of Chinese yams, 8 parts of Root of coastal Glehnia, 7 parts of Fructus Viticis Negundo, 4 parts of radix polygonati officinalis, 4 parts of the seeds of Job's tears, 4 parts of Fructus Hordei Germinatus, 4 parts of sweet osmanthus, 4 parts of the Radixs Astragali;
Mixed enzyme addition is the 5-10% of mixture gross weight;
The parts by weight of described mixed enzyme consist of: 15 parts of endo-beta-glucanases, 15 parts of outer beta-glucanases, 13 parts of beta-glucosidases, 13 parts of zytases, 13 parts of pentosanases, 25 parts of Pullulanases, 12 parts of beta-amylases, 13 parts of neutral proteases, 13 parts of aspartic proteases, 7 parts of superoxide-dismutases, 7 parts of glucose oxidases, 7 parts of acid phosphatases;
Described protective material is comprised of the raw material of following parts by weight: 25 parts of trehaloses, NaCl25 part, (NH 4) 2sO 413 parts, 12 parts of halfcystines;
Described activator is that the inorganic salt by following quality component evenly mix: 35 parts of zinc chloride, 15 parts, calcium chloride, 15 parts, sodium sulfate, 7 parts, magnesium chloride;
By described protective material, Chinese herbal medicine extract difference micronizing; guarantee that granularity is less than described acidic xylanase and other raw material; then with aspergillus niger culture, acidic xylanase, cellulase, beta-glucanase, amylase, phytase; evenly mix; finally add activator, after mixing, pack the suckling piglet specific enzyme that gets product.
CN201310736761.9A 2013-12-27 2013-12-27 A kind of suckling piglet specific enzyme containing complex enzyme for feed and preparation method thereof Active CN103725661B (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN201310736761.9A CN103725661B (en) 2013-12-27 2013-12-27 A kind of suckling piglet specific enzyme containing complex enzyme for feed and preparation method thereof

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN201310736761.9A CN103725661B (en) 2013-12-27 2013-12-27 A kind of suckling piglet specific enzyme containing complex enzyme for feed and preparation method thereof

Publications (2)

Publication Number Publication Date
CN103725661A true CN103725661A (en) 2014-04-16
CN103725661B CN103725661B (en) 2015-08-12

Family

ID=50449954

Family Applications (1)

Application Number Title Priority Date Filing Date
CN201310736761.9A Active CN103725661B (en) 2013-12-27 2013-12-27 A kind of suckling piglet specific enzyme containing complex enzyme for feed and preparation method thereof

Country Status (1)

Country Link
CN (1) CN103725661B (en)

Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN105309754A (en) * 2014-08-04 2016-02-10 湖南新鸿鹰生物工程有限公司 Mold-culture-containing enzyme special for anthony pigs and preparation method thereof
CN108576402A (en) * 2018-03-22 2018-09-28 昆明三正生物科技（集团）有限公司 A kind of suckling piglet specific enzyme production method
CN109370915A (en) * 2018-11-28 2019-02-22 河北京安瑞能环境科技有限公司 One kind being used for the pretreated bacteria agent production method of straw methane fermentation and application
CN114601058A (en) * 2022-04-09 2022-06-10 河南宏展生物科技有限公司 Piglet concentrated feed, complete ration and preparation method thereof

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101423826A (en) * 2008-12-15 2009-05-06 宁夏大学 Method for producing complex enzyme preparation for feeding by using tylosin herb residue
CN101591619A (en) * 2009-07-03 2009-12-02 中国农业科学院饲料研究所 Aspergillus niger strain and application thereof
CN102119768A (en) * 2011-01-25 2011-07-13 北京挑战农业科技有限公司 Complex enzyme preparation for feeding piglets

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101423826A (en) * 2008-12-15 2009-05-06 宁夏大学 Method for producing complex enzyme preparation for feeding by using tylosin herb residue
CN101591619A (en) * 2009-07-03 2009-12-02 中国农业科学院饲料研究所 Aspergillus niger strain and application thereof
CN102119768A (en) * 2011-01-25 2011-07-13 北京挑战农业科技有限公司 Complex enzyme preparation for feeding piglets

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
秦涛 等: ""复合型饲料添加因素研究评述"", 《安徽农业科学》, vol. 36, no. 4, 31 December 2008 (2008-12-31) *
费笛波 等: ""黑曲霉6042 液体发酵生产饲用复合酶的工艺研究"", 《浙江农业学报》, vol. 8, no. 6, 31 December 1996 (1996-12-31) *

Cited By (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN105309754A (en) * 2014-08-04 2016-02-10 湖南新鸿鹰生物工程有限公司 Mold-culture-containing enzyme special for anthony pigs and preparation method thereof
CN105309754B (en) * 2014-08-04 2019-03-29 湖南新鸿鹰生物工程有限公司 A kind of suckling piglet specific enzyme of the object containing mycotic culture and preparation method thereof
CN108576402A (en) * 2018-03-22 2018-09-28 昆明三正生物科技（集团）有限公司 A kind of suckling piglet specific enzyme production method
CN109370915A (en) * 2018-11-28 2019-02-22 河北京安瑞能环境科技有限公司 One kind being used for the pretreated bacteria agent production method of straw methane fermentation and application
CN109370915B (en) * 2018-11-28 2021-07-20 河北京安瑞能环境科技有限公司 Application method of biological agent for straw biogas fermentation pretreatment
CN114601058A (en) * 2022-04-09 2022-06-10 河南宏展生物科技有限公司 Piglet concentrated feed, complete ration and preparation method thereof

Also Published As

Publication number Publication date
CN103725661B (en) 2015-08-12

Similar Documents

Publication Publication Date Title
CN103667222B (en) Feed compound enzyme-containing dedicated enzyme for growing pigs and preparation method of feed compound enzyme-containing dedicated enzyme
CN103734481B (en) Feed compound enzyme containing feeding complex enzyme and preparation method of feed compound enzyme
CN103667221B (en) Alkaline xylanase-containing dedicated enzyme for piglets and preparation method of alkaline xylanase-containing dedicated enzyme
CN110934238A (en) Fermented soft pellet feed for prawn culture and preparation method thereof
CN103667220B (en) Neutral protease-containing growing pig dedicated enzyme and preparation method thereof
CN103734480B (en) Feed compound enzyme capable of improving animal appetite and preparation method thereof
CN103704486B (en) Poultry enzyme including neutral protease and preparation method thereof
CN103766611B (en) Neutral protease-containing coarse grain daily ration enzyme and preparation method thereof
CN103082136A (en) Pig feed additive
CN101120719B (en) Microorganism ferment purple sweet potato and preparation method thereof
CN107853480A (en) The fermentation process of one boar food and application
CN110214871A (en) A kind of layer breeding probiotics and preparation method thereof
CN103725661B (en) A kind of suckling piglet specific enzyme containing complex enzyme for feed and preparation method thereof
CN103859169A (en) Feed compound enzyme containing neutral protease and preparation method thereof
CN108432979A (en) A kind of Australia freshwater lobster ecological feed
CN103750023B (en) Special beta-dextranase-containing complex enzyme for piglet and preparation method thereof
CN103749987A (en) Coarse cereal, mixed meal and daily ration enzyme preparation and preparation method thereof
CN103875935B (en) A kind of suckling piglet special composite enzyme containing profitable probliotics and preparation method thereof
CN104757279A (en) Suckling piglet special-purpose compound enzyme and preparation method thereof
CN103695396B (en) A kind of high vigor complex enzyme for feed
CN103740683B (en) A kind of Wheat ration enzyme containing neutral protease and preparation method thereof
CN103750011B (en) Special compound enzyme containing cellulase for piglets and preparation method thereof
CN103865900B (en) A kind of feed complex enzyme containing alkalescent xylanase and preparation method thereof
CN103689241B (en) Preparation method of high-activity composite enzyme for feed
CN103725663B (en) A kind of growing swine specific enzyme containing alkalescent xylanase and preparation method thereof

Legal Events

Date Code Title Description
C06 Publication
PB01 Publication
C10 Entry into substantive examination
SE01 Entry into force of request for substantive examination
C14 Grant of patent or utility model
GR01 Patent grant