CN104222493A - Compound probiotic peptide as well as preparation method and application thereof - Google Patents
Compound probiotic peptide as well as preparation method and application thereof Download PDFInfo
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Abstract
The invention discloses a compound probiotic peptide as well as a preparation method and an application thereof. The compound probiotic peptide is obtained by adding a bacterium enzyme compounding fermentation agent into soya bean meal and fermenting, wherein the bacterium enzyme compounding fermentation agent is formed by mixing four types of probiotics, four types of proteases and a carbon source; the four types of probiotics comprise clostridium butyricum, bacillus pumilus S3, lactobacillus plantarum and saccharomyces boulardii; and the four types of proteases comprise an acidic protease, a neutral protease, an alkaline protease and a polypeptide protease. The invention provides a specific preparation method of the compound probiotic peptide and an application of the compound probiotic peptide to preparing a piglet feed additive. The compound probiotic peptide product is enriched with a plurality of types of probiotics and antigen proteins can be effectively degraded to generate micro-molecular substances, namely polypeptides, amino acids and the like; and in a fermentation process, a plurality of types of flavor and antagonistic substances, namely probiotic organic acids, biotins and the like are generated, so that the palatability and the safety are increased, the utilization rate of the soya bean meal and the nutritive value of a feed are remarkably improved, and the diarrhea rate of piglets can be remarkably reduced.
Description
Technical field
The invention belongs to biological enzymolysis and fermentation arts, relate to the preparation method of peptide, be specifically related to a kind of compound benefit bacterium peptide and its preparation method and application.
Background technology
Dregs of beans, as a kind of important plant protein source, has important application and research to be worth.But the existence of the anti-nutrient substances such as the protease inhibitors in soybean, phytolectin, Soybean antigen protein, urase, phytic acid, soyabean oligosaccharides, not only greatly hinder animal to the digestion of nutritional labeling, absorption and utilization, and seriously endanger the health of animal and grow.Soybean antigen protein is that in soybean processing goods, residual quantity is the highest, a kind of Soybean Anti-nutritional Factors that anti-oxidant action is the strongest, the allergic reaction of the animal generation enteron aisle such as infant and mouse, piglet, calf and other organ can be caused, and cause intestinal mucosa injury, there is damaging change in small intestine, and then it is even dead to cause diarrhoea, production performance to decline, be one of universally acknowledged eight large food allergens.Soybean antigen protein mainly comprises 11S glycinin and 7S β-soybean companion globulin, and both account for about 70% of soybean protein content.
In the bioremediation of current dregs of beans, microbe fermentation method and ferment treatment method are comparatively general, the present invention is by four kinds of bacterium, four kinds of enzymes combine with carbon source, using dregs of beans as a kind of matrix of cultivating microorganism, promote four kinds of probio amount reproductions, and in biological treatment process, effective degraded also utilizes the large molecular antigen albumen of dregs of beans, almost by its degradable be polypeptide, the small-molecule substances such as amino acid, be easy to animal absorb, and create the local flavor materials such as multiple probiotic organic acid, biotin, clostridium butyricum element waits Antagonism material, adding palatability, while security, also significantly improve protein utilization rate and the nutritive value of dregs of beans, and newborn grice diarrhoea rate can be reduced, possess certain functional.
Summary of the invention
The invention provides a kind of compound benefit bacterium peptide and its preparation method and application, described compound benefit bacterium peptide can meet the special dietary demand of newborn piglet to dregs of beans raw material.
For achieving the above object, the present invention is achieved by the following technical solutions:
A kind of compound benefit bacterium peptide, it adds fermentation in dregs of beans to by the composite leavening of bacterium enzyme and forms, the composite leavening of described bacterium enzyme is composited by four kinds of probios, four kinds of protease and carbon source, and described four kinds of probios are respectively clostridium butyricum, bacillus pumilus S3, Lactobacillus plantarum and saccharomyces boulardii; Described four kinds of protease are respectively acid protease, neutral proteinase, alkali protease and polypeptide protein enzyme; The Classification And Nomenclature of described bacillus pumilus S3 is bacillus pumilus (Bacillus pumilus), and its deposit number is: CGMCC No.9161.
Further improvement to technique scheme: the bacteria containing amount of described four kinds of probios in the composite leavening of bacterium enzyme respectively: clostridium butyricum: be not less than 1 × 10
6cFU/g; Bacillus pumilus S3: be not less than 1 × 10
6cFU/g; Lactobacillus plantarum: be not less than 1 × 10
6cFU/g; Saccharomyces boulardii: be not less than 1 × 10
5cFU/g.
Further improvement to technique scheme: the content of described four kinds of protease in the composite leavening of bacterium enzyme respectively: acid protease: be not less than 500U/g; Neutral proteinase: be not less than 1000U/g; Alkali protease: be not less than 4000U/g; Polypeptide protein enzyme: be not less than 4500U/g.
Further improvement to technique scheme: described carbon source is the glucose accounting for the composite leavening quality 9%-10% of bacterium enzyme.
Present invention also offers the preparation method of described compound benefit bacterium peptide, it comprises the steps:
(1) in dregs of beans, add the water accounting for Soybean Meal 50-60% and form fermentation bed material, inoculation accounts for the composite leavening of described bacterium enzyme of Soybean Meal 10%, mixes formation fermentation materials;
(2) described fermentation materials is stored in tinfoil paper sealing bag, in 200 DEG C of sealings, is placed in 37 DEG C of constant incubators, fermentation 72h;
(3) after having fermented, low temperature drying, pulverizes and sieves, and forms described compound benefit bacterium peptide.
Further improvement to technique scheme: in described step (3), fermentation materials is in 40 DEG C of low temperature dryings, pulverizes 60 mesh sieves.
Invention further provides described compound benefit bacterium peptide for the preparation of the application of feeding in the feed addictive of newborn piglet.
Described compound benefit bacterium peptide compares for the ratio of 5-15% is added in feed to account for feeding quality, described compound benefit bacterium peptide antagonism pig manure coliform.
Compared with prior art, the advantage of the inventive method and technique effect are:
1, the composite leavening of bacterium enzyme that the present invention is used is in biological enzymolysis fermentation theoretical foundation, in conjunction with years'experiences, for the conventional dregs of beans material characteristic of production, and the selectivity leavening prepared voluntarily; The composite leavening of selectivity bacterium enzyme described in employing, biological treatment dregs of beans prepares compound benefit bacterium peptide product.
2, the compound benefit bacterium peptide product prepared by the present invention, is rich in multiple probio, comprises clostridium butyricum, bacillus pumilus S3, Lactobacillus plantarum, saccharomyces boulardii.In its enzymatic hydrolysis and fermentation process, antigen protein in dregs of beans of can effectively degrading produces the small-molecule substance such as polypeptide, amino acid, and produce local flavor, the Antagonism materials such as multiple probiotic organic acid, biotin, not only increase palatability, security, and gemma class probio has good storage stability.
3, the compound benefit bacterium peptide product prepared by the present invention, can significantly improve dregs of beans utilization rate and feed nutritive value, have pig manure coliform resistance, also can significantly reduce grice diarrhoea rate.
4, bacterium involved in the present invention, enzyme etc., all belong to feed addictive category, wide material sources, and safety is easy to get, with low cost.
5, the method for the invention simple and reasonable steps, swift to operate, has good industrializing implementation prospect.
After reading the specific embodiment of the present invention by reference to the accompanying drawings, other advantages of the present invention will become clearly.
Accompanying drawing explanation
Fig. 1 shows compound benefit bacterium peptide-probiotics bacterial amount composition and stability half a year of the preparation after biological treatment of No. 1 dregs of beans.
Fig. 2 shows compound benefit bacterium peptide-probiotics bacterial amount composition and stability half a year of the preparation after biological treatment of No. 2 dregs of beans.
Fig. 3 shows that dregs of beans raw material is prepared as the antigen protein SDS-PAGE electrophoretogram of compound benefit bacterium peptide after biological treatment.
Fig. 4 shows that dregs of beans raw material is prepared as compound benefit bacterium peptide and is rich in probiotic organic acid after biological treatment.
Fig. 5 shows that dregs of beans raw material is prepared as compound benefit bacterium peptide after biological treatment can antagonism pig manure coliform.
Fig. 6 shows that dregs of beans raw material is prepared as compound benefit bacterium peptide and significantly can reduces newborn grice diarrhoea rate after biological treatment.
Fig. 7 is the production technological process of bacillus pumilus S3.
detailed description of the invention
For understanding the function of this compound benefit bacterium peptide product better, below in conjunction with the drawings and specific embodiments, feature being quoted to product of the present invention and being further described in detail.
Embodiment 1
One, the separation screening of bacillus pumilus S3
In year September in May, 2013 to 2013, from Jiangzhou, coastal many places gather water sample, bed mud, the gut of shrimp ight soil in shrimp aquaculture region.Bed mud, gut of shrimp fecal specimens, respectively at 10 times of dilutions in SPSS, fully grind, and getting supernatant is sample liquid; Water sample is directly as sample liquid; Sample liquid is inoculated in 2216E fluid nutrient medium respectively dilutes after 30 DEG C of enrichment culture 48h, coat carry out bacterial strain in 2216E flat board separation, purifying, filter out the pure bacterium of 26 strain altogether.
Purifying single strain adopts dull and stereotyped dibbling method to screen the bacterium that flys up and down.Take vibrio parahaemolytious as indicator bacteria, be inoculated in by vibrio parahaemolytious in 2216E fluid nutrient medium and cultivate 18h, dilution is prepared into 10
5-10
6the bacteria suspension of CFU/mL, gets 0.1mL and is coated with 2216E flat board, for subsequent use.With the bacterial strain that sterile toothpick picking is to be screened, point is connected to containing in bacterium flat board, and 25 DEG C of constant temperature culture 24h, point of observation connects periphery of bacterial colonies and whether occurs obvious inhibition zone or the area of coverage.
To the 26 strain bacteriums be separated to adopt dibbling methods to screen 3 strains to have antagonistic action bacterial strain to vibrio parahaemolytious, wherein S3 has stronger antagonistic action to vibrio parahaemolytious, obviously can suppress the growth of pathogen, around pathogen, produce obviously inhibition zone clearly.
Through Bacterial Physiological biochemical method Preliminary Identification, bacterial strain S3 is bacillus, its biological characteristics: Gram-positive, and the colony diameter that nutrient agar panel is formed is about 2mm, and neat in edge is smooth, middle fold, milky.Thalli morphology is shaft-like, and gemma is oval, middle life.
Two, the molecular genetics taxonomic identification of bacillus pumilus
First extract the DNA of bacterial strain S3, with this DNA for template, 16S rRNA universal primer is primer, increases to 16SrRNA fragment, and amplified fragments carries out sequencing.Sequencing result Blast software the belong to 16S rRNA sequence of planting relevant to GenBank is compared, the 16s rRNA sequence homology degree that result shows the Bacillus pumilus in this bacterial strain 16S rRNA sequence and GenBank gene pool in bacillus is the highest, and homology reaches 99%.Phylogenetic analysis is carried out by the 16S rRNA sequence of DNAMAN6.0 to bacillus existing in Genbank, result shows, bacterial strain S316S rRNA and Bacillus pumilus homology the highest, judge the bacillus pumilus of this bacterial strain as bacillus.
The bacterial strain S3 screened is carried out culture presevation, depositary institution: China Committee for Culture Collection of Microorganisms's common micro-organisms center (CGMCC); Address: No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City, Institute of Microorganism, Academia Sinica; Preservation date: on May 13rd, 2014; The deposit number of bacillus pumilus Bacillus pumilus is CGMCC No.9161.
Three, the composition of compound benefit bacterium peptide
Compound benefit bacterium peptide of the present invention adds fermentation in dregs of beans to by the composite leavening of bacterium enzyme and forms, the composite leavening of described bacterium enzyme is composited by four kinds of probios, four kinds of protease and carbon source, described four kinds of probios are respectively clostridium butyricum, bacillus pumilus S3, Lactobacillus plantarum, saccharomyces boulardii; Described four kinds of protease are respectively acid protease, neutral proteinase, alkali protease, polypeptide protein enzyme; The Classification And Nomenclature of described bacillus pumilus S3 is bacillus pumilus (Bacillus pumilus), and its deposit number is: CGMCC No.9161.Described carbon source is the glucose accounting for the composite leavening quality 9%-10% of bacterium enzyme, the glucose crop carrier of interpolation and carbon source, to meet the growth demand of various bacterium, and promotes that the probiotic organic acid such as lactic acid, acetic acid, butyric acid produces.
The bacteria containing amount of described four kinds of probios in the composite leavening of bacterium enzyme be respectively: clostridium butyricum: be not less than 1 × 10
6cFU/g; Bacillus pumilus S3: be not less than 1 × 10
6cFU/g; Lactobacillus plantarum: be not less than 1 × 10
6cFU/g; Saccharomyces boulardii: be not less than 1 × 10
5cFU/g.The content of described four kinds of protease in the composite leavening of bacterium enzyme is respectively: acid protease: be not less than 500U/g; Neutral proteinase: be not less than 1000U/g; Alkali protease: be not less than 4000U/g; Polypeptide protein enzyme: be not less than 4500U/g.
Clostridium butyricum used in the present invention, Lactobacillus plantarum and saccharomyces boulardii are the bacterium powder of commercial goods, the acid protease used, neutral proteinase, alkali protease and polypeptide protein enzyme are the enzyme preparation of commercial goods, and described bacillus pumilus S3 also preserves from row filter, its technological process of production as shown in Figure 7.
Four, the preparation method of compound benefit bacterium peptide of the present invention comprises the steps:
(1) in dregs of beans, add the water accounting for Soybean Meal 50-60% and form fermentation bed material, inoculation accounts for the composite leavening of described bacterium enzyme of Soybean Meal 10%, mixes formation fermentation materials;
(2) described fermentation materials is stored in tinfoil paper sealing bag, in 200 DEG C of sealings, is placed in 37 DEG C of constant incubators, fermentation 72h.
In described step (2), fermentation materials is in complete sealed environment, and in front 24h, still remaining in material have oxygen, aerobic bacillus pumilus S3 and facultative aerobic saccharomyces boulardii ramp; During 24h-48h, along with the continuous consumption of oxygen and the generation gradually of other gases, amphimicrobian Lactobacillus plantarum occupies advantage gradually, and saccharomycete bacterium also progresses into fermentation pattern, can produce the multiple probiotic organic acid such as lactic acid, acetic acid around here; During 48h-72h, after oxygen exhausts completely, complete anaerobic bacteria clostridium butyricum occupies advantage gradually, and secretes butyric acid, is conducive to improving animal intestinal.Under totally sealed environment, achieve the relay growth of four kinds of characteristic probios in 72h, multiple probiotic material can be produced.
In described step (2), fermentation materials is in complete sealed environment, and bacterium dominant growths different in different time sections, causes environment pH also to gradually change, the basic trend for reducing.Early stage pH higher than 5 time, alkali protease and polypeptide protein enzyme can play certain enzymolysis, and later stage pH, lower than after 5, plays biological degradation function primarily of neutral proteinase and acid protease.The restriction enzyme site of four kinds of enzymes is different, can effective degradation antigen albumen, significantly improves acid-soluble albumen, alkaline hydrolysis protein ratio, increases free aminoacid content, improve the external protein digestibility of product.
In described step (2), fermentation materials is in complete sealed environment, pH reduces gradually, four kinds of enzymes alternately work, and restriction enzyme site is different, can effective degradation antigen albumen, significantly improve acid-soluble albumen, alkaline hydrolysis protein ratio, increase free aminoacid content, improve the external protein digestibility of product.
(3) after having fermented, in 40 DEG C of low temperature dryings, pulverize 60 mesh sieves, form compound benefit bacterium peptide product;
In described step (3), before and after low temperature drying process, final compound benefit bacterium peptide product is compared with fermentation wet feed, clostridium butyricum, bacillus pumilus S3, Lactobacillus plantarum, saccharomyces boulardii etc. have a certain proportion of survival, Bacillus survival rate higher than 89%, and in half a year under room temperature condition Viable detection higher than 90%.
Described compound benefit bacterium peptide product is rich in probio, organic acid etc., and has miscellaneous bacteria resistance, for the feed for piglet preparation, adds, can significantly reduce grice diarrhoea rate with 5-15% ratio.
Described compound benefit bacterium peptide product, possesses certain pig manure coliform resistance.
With the method for the invention, No. 1 and No. 2 two kinds of dregs of beans raw materials (being commercially available common dregs of beans raw material) are carried out a biological disposal upon, obtain No. 1 bacterium peptides beneficial to No. 2 two kinds of compounds.
Embodiment 2
Described compound benefit bacterium peptide product respectively during mensuration fermentation 72h after fermentation termination wet feed, low temperature drying, room temperature deposit the content of four kinds of bacterium in the benefit of the compound after June bacterium peptide product.Utilize tryptone-sulphite-seromycin culture medium, adopt layer plating method, measure clostridium butyricum bacterium amount; Utilize nutrient agar, adopt method of dilution butteron on plate, measure bacillus pumilus S3 bacterium amount; Utilize MRS culture medium, adopt pour plate method, measure Lactobacillus plantarum bacterium amount; Utilize YPD culture medium, adopt method of dilution butteron on plate, measure saccharomyces boulardii bacterium amount.In view of four kinds of bacterium bacterium amounts differ greatly, represent bacterium amount with log CFU/g.
Fig. 1 illustrates compound benefit bacterium peptide-probiotics bacterial amount composition and stability half a year of the preparation after biological treatment of No. 1 dregs of beans, and Fig. 2 illustrates compound benefit bacterium peptide-probiotics bacterial amount composition and stability half a year of the preparation after biological treatment of No. 2 dregs of beans.Before and after low temperature drying process, final compound benefit bacterium peptide product has a certain proportion of survival with each bacterium in fermentation wet feed, and wherein Bacillus dries survival rate higher than 89%, and after depositing half a year under room temperature condition, Viable detection is higher than 90%.In the fermentation wet feed of No. 1 compound benefit bacterium peptide, clostridium butyricum bacterium amount reaches 1.08 × 10
6cFU/g, after low temperature drying, containing clostridium butyricum 9.7 × 10
5cFU/g, dry survival rate 89.81%, and after half a year, bacterium amount still keeps 8.8 × 10
5cFU/g, survival rate 90.72%, illustrate that in this product, clostridium butyricum heat resistance is strong, good stability, which greatly improves the possibility that this bacterium plays a role in animal body; Bacillus pumilus S3 and clostridium butyricum similar, fermentation wet feed in bacterium amount reach 4.5 × 10
7cFU/g, after low temperature drying, bacterium amount reaches 4.2 × 10
7cFU/g, survival rate 93.34% after drying, after room temperature deposits half a year, bacterium amount still keeps 4 × 10
7cFU/g, Viable detection reaches 95.24%, illustrates that its spore is allowed a choice through drying course, and tolerance is stronger, has good stability, can strengthen the application potential of this product; In the fermentation wet feed of No. 1 compound benefit bacterium peptide, Lactobacillus plantarum bacterium amount reaches 3.3 × 10
7cFU/g, decline 1 order of magnitude in drying course, and bacterium amount is 3.83 × 10
6cFU/g, after room temperature deposits half a year, still keeps 1.96 × 10
6cFU/g, the viable bacteria bacterium amount order of magnitude does not become, survival rate 51.17%, illustrates containing more Lactobacillus plantarum in compound benefit bacterium peptide product, though there is loss can tolerate the storage of certain hour; In the fermentation wet feed of No. 1 compound benefit bacterium peptide, saccharomyces boulardii amount can reach 7.5 × 10
5cFU/g, but its tolerance is poor, and in low temperature drying milling product, bacterium amount reaches 3.5 × 10
4cFU/g, after room temperature deposits half a year, saccharomycete only reaches 4.1 × 10
3cFU/g.In No. 2 compounds benefit bacterium peptides each bacterium content and ratio, dries front and back survival rate, the index such as room temperature to deposit before and after half a year Viable detection, with similar in the beneficial bacterium peptide of No. 1 compound, but its Lactobacillus plantarum bacterium amount is slightly high, and other bacterium bacterium amounts are lower slightly.
Embodiment 3
Utilize the qualitative degraded situation determining antigen protein of SDS-PAGE (polyacrylamide gel electrophoresis), utilize ELISA (enzyme-linked immunosorbent assay) kit to measure the clearance of antigen protein.TCA (trichloroacetic acid extraction) is utilized to measure acid-soluble albumen (accounting for crude protein, %); With reference to GB/T19541-2004 feed Soybean Meal, utilize 0.2%KOH to measure KOH solubility, with reference to the total quantitative determination of GB GBT-8314-2002 free amino acid, measure free aminoacid content; Utilize SDS-II Bionic digestive system for monogastric animals that Beijing animal and veterinary research institute of the Chinese Academy of Agricultural Sciences researches and develops, the external protein digestibility of simulated determination.
As shown in Figure 3, ELISA quantified results is as shown in table 1 for the SDS-PAGE qualitative determination result of antigen protein.No. 1 from the band depth slightly difference of No. 2 different protein protomers of dregs of beans raw material, glycinin, β-soybean companion globulin, the concrete measured value of antigen protein is also slightly different, but the contrast trend between dregs of beans raw material and the benefit of the compound after carrying out a biological disposal upon bacterium peptide product is similar: after carrying out a biological disposal upon, the large molecular antigen albumen of compound benefit bacterium peptide is almost degradable is small molecular protein (molecular weight of albumen is lower than 14KDa), from No. 1 with No. 2 dregs of beans raw materials to compound benefit bacterium peptide product, the antigen protein clearance that ELISA kit measures also reaches 91.98% and 93.21% respectively.
After table 2 shows that two kinds of dregs of beans are treated and is prepared as compound benefit bacterium peptide product, the change of the indexs such as acid-soluble albumen (accounting for crude protein, %), the external Bionic digestion rate of KOH solubility, free amino acid, nonruminant.From No. 1 with No. 2 dregs of beans raw materials to compound benefit bacterium peptide product, the amplification of acid-soluble albumen/cp is respectively 27.64% and 29.15%, the amplification of KOH solubility is respectively 14.13% and 16.81%, the amplification of free amino acid is respectively 9.34% and 9.26%, and the amplification of external protein digestibility is respectively 13.82% and 13.51%.The change of these indexs, mainly has benefited from the composite leavening of selectivity bacterium enzyme to the biological enzymolysis of dregs of beans and fermentation process, and also almost degradable with high molecular weight protein is before that small molecular protein result of the test is corresponding.This compound benefit bacterium peptide product is used for the preparation of newborn baby pig feedstuff, advantageously in absorption and the digestion of protide nutrition, and the irritability can effectively avoiding large molecular antigen albumen to cause reaction.
Table 1 shows that dregs of beans raw material is prepared as the antigen protein ELISA kit measurement result of compound benefit bacterium peptide after biological treatment
Table 2 shows that dregs of beans raw material is prepared as the indexs such as the acid-soluble albumen of compound benefit bacterium peptide, KOH solubility, free amino acid and external protein digestibility and changes after biological treatment
Embodiment 4
With the method for the invention, No. 1 and No. 2 two kinds of dregs of beans raw materials are carried out a biological disposal upon, obtain No. 1 bacterium peptides beneficial to No. 2 two kinds of compounds.Adopt HPLC high performance liquid chromatography, the lactic acid in product, acetic acid, butyric acid content are analyzed.
Fig. 4 shows that dregs of beans raw material is prepared as compound benefit bacterium peptide and is rich in probiotic organic acid after biological treatment.Lactic acid is a kind of important antimicrobial compound, plays an important role to the antagonism of pathogenic bacteria, also has the effect promoting growth of animal, extend the feed shelf-life, prevent feed mold.The lactic acid content of No. 2 compound benefit bacterium peptides is a little more than No. 1 compound benefit bacterium peptide, and this result and its Lactobacillus plantarum bacterium amount are slightly high corresponding.In two kinds of compound benefit bacterium peptide products, certain density acetic acid is had to produce, adjustable environment pH, and the extraordinary local flavor forming product.Butyric acid is produced by clostridium butyricum specificity, can reduce hindgut pH, simultaneously the growth of harmful clostridium such as Competitive assays clostridieum welchii.In addition, butyric acid can also promote large intestine and intestinal epithelial cell breeding, has the function of repairing intestinal mucosa, Promote immunity.Relative to No. 2 compound benefit bacterium peptides, containing more clostridium butyricum in No. 1 compound benefit bacterium peptide product, thus also create more butyric acid.These probiotic organic acids, can promote that benefit materials absorbs, and make product be provided with the complex functions such as germ antagonism, intestinal regeneration, Immune-enhancing effect.
Embodiment 5
With the method for the invention, No. 1 and No. 2 two kinds of dregs of beans raw materials are carried out a biological disposal upon, obtain No. 1 bacterium peptides beneficial to No. 2 two kinds of compounds.On the methylene blue culture medium of Yihong, with the pig manure coliform of even spread for target bacterium, digest simulated solution for antagonism comparative group with the monogastric animal bionic of dregs of beans raw material and compound benefit bacterium peptide product, investigate before and after biological treatment, whether have antibacterial material and produce.
As shown in Figure 5, showing that dregs of beans raw material is prepared as compound benefit bacterium peptide after biological treatment can antagonism pig manure coliform for the result of Antagonism contrast test.Left figure shows that No. 1 compound benefit bacterium peptide is compared with No. 1 dregs of beans raw material, and inhibition zone, by not having, increases to 29 ± 0.4mm, and amplification is remarkable; Right figure shows No. 2 compound benefit bacterium peptides compared with No. 2 dregs of beans raw materials, and inhibition zone, by not having, increases to 27 ± 0.3mm, and amplification is also more remarkable.These antibacterial material, the materials such as the lactic acid that the bacteriocin that the butyric acid and the clostridium butyricum that mainly comprise clostridium butyricum secretion are plain, bacillus pumilus S3 secretes, Lactobacillus plantarum are secreted.Initial guess, this compound benefit bacterium peptide product is applied in the feed for piglet, can reach the object suppressing the pathogenic microorganism growth and breedings such as coliform.
Embodiment 6
With the method for the invention, No. 1 and No. 2 two kinds of dregs of beans raw materials are carried out a biological disposal upon, obtain No. 1 bacterium peptides beneficial to No. 2 two kinds of compounds.On May 10th, 2014, to June 13, in Qingdao Pingdu City south villages and small towns plant of Hong Lan village, to the weanling pig of 30 ages in days, carries out piglet culture efficiency contrast test.The piglet dregs of beans corn type basal diet that control group adopts base feed factory in Weifang to provide; Two test group respectively with 10% No. 1 with No. 2 compounds benefit bacterium peptide product Substitution for Soybean Meal corn type basal diets; Take diarrhea rate as index, investigate before and after biological treatment, whether effectively can alleviate grice diarrhoea.
Zooperal result as shown in Figure 6, shows that dregs of beans raw material is prepared as compound benefit bacterium peptide after biological treatment, significantly can reduce newborn piglet diarrhea of pigs rate.The dregs of beans corn type basal diet that control group piglet is fed common, containing ANFs such as more antigen protein, trypsin inhibitors, easily cause the stress reactions such as newborn grice diarrhoea, in this test, control group diarrhea rate reaches 19.06% ± 0.25%.In two test group, basal diet is substituted respectively with 10% No. 1 and No. 2 compounds benefit bacterium peptide products, significantly can reduce the ratio of normal meal, reduce the consumption of the ANFs such as large molecular antigen, and be rich in the probio such as clostridium butyricum, bacillus pumilus, the probiotic organic acid such as butyric acid, lactic acid, can antagonism germ, reduce the generation of the stress reactions such as diarrhoea, result also shows, and two test group all grice diarrhoea phenomenon do not occur.The method of the invention, can carry out effective biodegradation to ANFs such as antigen proteins in dregs of beans raw material, and be rich in multiple prebiotic substance, is prepared as feed product and feeds after piglet, can significantly reduce grice diarrhoea rate.
Above embodiment only in order to technical scheme of the present invention to be described, but not is limited; Although with reference to previous embodiment to invention has been detailed description, for the person of ordinary skill of the art, still can modify to the technical scheme described in previous embodiment, or equivalent replacement is carried out to wherein portion of techniques feature; And these amendments or replacement, do not make the essence of appropriate technical solution depart from the spirit and scope of the present invention's technical scheme required for protection.
Claims (9)
1. a compound benefit bacterium peptide, it is characterized in that it adds fermentation in dregs of beans to by the composite leavening of bacterium enzyme and forms, the composite leavening of described bacterium enzyme is composited by four kinds of probios, four kinds of protease and carbon source, and described four kinds of probios are respectively clostridium butyricum, bacillus pumilus S3, Lactobacillus plantarum and saccharomyces boulardii; Described four kinds of protease are respectively acid protease, neutral proteinase, alkali protease and polypeptide protein enzyme; The Classification And Nomenclature of described bacillus pumilus S3 be bacillus pumilus (
bacillus pumilus), its deposit number is: CGMCC No. 9161.
2. compound according to claim 1 benefit bacterium peptide, is characterized in that: the bacteria containing amount of described four kinds of probios in the composite leavening of bacterium enzyme respectively: clostridium butyricum: be not less than 1 × 10
6cFU/g; Bacillus pumilus S3: be not less than 1 × 10
6cFU/g; Lactobacillus plantarum: be not less than 1 × 10
6cFU/g; Saccharomyces boulardii: be not less than 1 × 10
5cFU/g.
3. compound according to claim 1 benefit bacterium peptide, is characterized in that: the content of described four kinds of protease in the composite leavening of bacterium enzyme respectively: acid protease: be not less than 500U/g; Neutral proteinase: be not less than 1000U/g; Alkali protease: be not less than 4000U/g; Polypeptide protein enzyme: be not less than 4500U/g.
4. compound benefit bacterium peptide according to claim 1, is characterized in that: described carbon source is the glucose accounting for the composite leavening quality 9%-10% of bacterium enzyme.
5. the preparation method of the compound benefit bacterium peptide described in any one of claim 1-4, is characterized in that it comprises the steps:
(1) in dregs of beans, add the water accounting for Soybean Meal 50-60% and form fermentation bed material, inoculation accounts for the composite leavening of described bacterium enzyme of Soybean Meal 10%, mixes formation fermentation materials;
(2) described fermentation materials is stored in tinfoil paper sealing bag, in 200 DEG C of sealings, is placed in 37 DEG C of constant incubators, fermentation 72h;
(3) after having fermented, low temperature drying, pulverizes and sieves, and forms described compound benefit bacterium peptide.
6. the preparation method of compound benefit bacterium peptide according to claim 5, is characterized in that: in described step (3), fermentation materials is in 40 DEG C of low temperature dryings, pulverizes 60 mesh sieves.
7. the compound benefit bacterium peptide described in any one of claim 1-4 is for the preparation of the application of feeding in the feed addictive of newborn piglet.
8. compound benefit bacterium peptide according to claim 7 is for the preparation of the application of feeding in the feed addictive of newborn piglet, it is characterized in that: described compound benefit bacterium peptide adds in feed than for the ratio of 5-15% to account for feeding quality.
9. compound benefit bacterium peptide according to claim 7 is for the preparation of the application of feeding in the feed addictive of newborn piglet, it is characterized in that: described compound benefit bacterium peptide antagonism pig manure coliform.
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| CN106472932A (en) * | 2016-10-18 | 2017-03-08 | 李德田 | A kind of complex microorganism beverage and preparation method thereof |
| CN106922948A (en) * | 2015-12-29 | 2017-07-07 | 丰益(上海)生物技术研发中心有限公司 | The production technology of fermented bean dregs is carried out suitable for edible fat production factory environment |
| CN106962609A (en) * | 2017-04-01 | 2017-07-21 | 福建龙岩闽雄生物科技股份有限公司 | A kind of direct putting type biological fermentation feed additive and its production and use |
| CN110804564A (en) * | 2019-11-18 | 2020-02-18 | 江苏三仪生物工程有限公司 | Preparation of clostridium butyricum powder and detection culture medium thereof |
| CN114617191A (en) * | 2022-03-15 | 2022-06-14 | 广东海洋大学 | Functional feed prepared from euphausia superba and method thereof |
| CN114617190A (en) * | 2022-03-15 | 2022-06-14 | 广东海洋大学 | Functional feed protein prepared from euphausia superba and method thereof |
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| CN104872379A (en) * | 2015-05-18 | 2015-09-02 | 青岛尚德生物技术有限公司 | Fermentation substance prepared by adding fish soluble in corn-soybean meal diet and fermentation method of fermentation substance |
| CN104872379B (en) * | 2015-05-18 | 2018-07-24 | 青岛尚德生物技术有限公司 | A kind of fermentate and its fermentation process adding the molten slurry of fish in corn-soybean meal diet |
| CN106922948A (en) * | 2015-12-29 | 2017-07-07 | 丰益(上海)生物技术研发中心有限公司 | The production technology of fermented bean dregs is carried out suitable for edible fat production factory environment |
| CN105876154A (en) * | 2016-04-22 | 2016-08-24 | 宜昌市昌伟饲料科技有限公司 | Antibiotic-free feed for livestock and poultry and production method thereof |
| CN106472932A (en) * | 2016-10-18 | 2017-03-08 | 李德田 | A kind of complex microorganism beverage and preparation method thereof |
| CN106962609A (en) * | 2017-04-01 | 2017-07-21 | 福建龙岩闽雄生物科技股份有限公司 | A kind of direct putting type biological fermentation feed additive and its production and use |
| CN110804564A (en) * | 2019-11-18 | 2020-02-18 | 江苏三仪生物工程有限公司 | Preparation of clostridium butyricum powder and detection culture medium thereof |
| CN110804564B (en) * | 2019-11-18 | 2020-10-09 | 江苏三仪生物工程有限公司 | Preparation of clostridium butyricum powder and detection culture medium thereof |
| CN114617191A (en) * | 2022-03-15 | 2022-06-14 | 广东海洋大学 | Functional feed prepared from euphausia superba and method thereof |
| CN114617190A (en) * | 2022-03-15 | 2022-06-14 | 广东海洋大学 | Functional feed protein prepared from euphausia superba and method thereof |
| WO2023175153A1 (en) * | 2022-03-18 | 2023-09-21 | Fumi Ingredients B.V. | A microbial cell extract, method for obtaining said microbial cell extract and use of said microbial cell extract |
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