CN104161172B - A kind of method utilizing Semen arachidis hypogaeae dregs two step solid fermentation to produce probiotic peptide - Google Patents
A kind of method utilizing Semen arachidis hypogaeae dregs two step solid fermentation to produce probiotic peptide Download PDFInfo
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Abstract
The invention belongs to animal feed and bionic technical field, be specifically related to a kind of method utilizing Semen arachidis hypogaeae dregs two step solid fermentation to produce probiotic peptide.With Semen arachidis hypogaeae dregs as raw material, sterilizing after being mixed and stirred for uniformly with water, press Semen arachidis hypogaeae dregs weight after sterilizing and access 2%(v/w) aspergillus niger (CGMCC9183) spore liquid, at 28 DEG C ~ 32 DEG C, cultivate 24 ~ 28h, every 4h stirring is once；Press Semen arachidis hypogaeae dregs weight after cultivation and access 10%(v/w) Pediococcus pentosaceus (CGMCC9182), mix homogeneously, 37 DEG C ~ 46 DEG C anaerobic fermentation 36 ~ 42h, after fermentation ends, 60 DEG C of dry, pulverizing, prepare probiotic peptide.The content of peptides of probiotic peptide reaches more than 25%, and lactic acid content reaches more than 8.0%, and viable count of lactobacillus reaches 109 More than CFU/g, acid protease reaches more than 200U/g.Production cost of the present invention is low, sweat is easily controllable, constant product quality.
Description
Technical field
The invention belongs to animal feed and bionic technical field, be specifically related to a kind of method utilizing Semen arachidis hypogaeae dregs two step solid fermentation to produce probiotic peptide.
Background technology
Semen arachidis hypogaeae dregs is Semen arachidis hypogaeae side-product after oil plant is refined in squeezing, containing abundant vegetable protein.The nutritive value of Semen arachidis hypogaeae dregs is higher, and its metabolizable energy is that dregs of rice apoplexy due to endogenous wind is the highest, and gross protein value reaches more than 48%, is the important plant origin albumen feedstuff of China, and China's annual Semen arachidis hypogaeae dregs resource is about at about more than 300 ten thousand tons.But, owing to the protein quality of Semen arachidis hypogaeae dregs is the same with other plant albumen, all existing with macromolecular structure, and there is antinutritional factor, hinder the digestion of albumen, absorption, the utilization rate of albumen is the most on the low side.
But; the most relatively fewer to the exploitation of Semen arachidis hypogaeae dregs; the polypeptide of high-load to be obtained (absolute magnitude is higher than 25%) is all that the method by liquid enzymolysis is carried out; product yield is relatively low; drying cost is too high; and owing to products application face is narrower, alternative materials is enriched, it is difficult to realize the large-scale production on Semen arachidis hypogaeae dregs.Recently, solid fermentation is few due to water content, does not the most produce garbage, low in the pollution of the environment, the advantage such as is easily processed, studied personnel application and Semen arachidis hypogaeae dregs high-valued in.At present, use bacillus cereus and lactic acid bacteria co-fermentation to produce fermented peanut meal, obtained bigger concern as feedstuff.But, owing to bacillus cereus produces neutral protease, under lactic acid bacteria co-fermentation, fermentation system quickly becomes acid, bacillus cereus and neutral protease are difficult to play a role, and bacillus cereus and lactic acid bacteria are respectively aerobic microbiological and anaerobe, both are inconsistent to the demand of oxygen, limited (the product lactic acid bacteria 10 of growth of microorganism8CFU/g), the beneficial metabolic product assay also causing product is limited, if content of peptides is about 8%, and lactic acid content about 3%.
For solving the defect of Semen arachidis hypogaeae dregs Forage quality, in conjunction with polypeptide products, probiotic bacteria, probiotics and enzyme preparation that current field of fodder is conventional, by biotechnology processing procedure, the function of these benefit-additives is rolled into one, is prepared for probiotic peptide.First, sweat is considered as nutrient substance predigestion process in vitro, in Semen arachidis hypogaeae dregs, antinutritional factor is decomposed removal, macro-molecular protein is broken down into polypeptide and the little peptide of part, the absorption of peptide has that transport speed is fast, it is low to consume energy, carrier is difficult between the advantage such as saturated and various peptide transport uncompetitive and inhibition, animal tissue can also directly utilize the Amino acid synthesis histone in peptide, reduces energy expenditure；Secondly, young animal gastrointestinal physiology and microbiota are grown and are not also perfected, stomachial secretion hydrochloric acid ability is poor, pH value in stomach is caused to raise and suppress pepsin activity and along with the interpolation in feedstuff after achlorhydria, the digestibility making nutrient substance is relatively low, and the external source digestive enzyme of sweat accumulation and a large amount of of beneficial metabolites supplement, and contributes to improving intestinal microflora balance, strengthen animal body immunity, significantly improve nutrient substance digestibility and utilization efficiency in animal body.Simultaneously as food safety increasingly receives publicity, European Union has completely forbidden in 2006 and has added antibiotic in feedstuff, finds the work of Substitutes For Antibiotic also achieved with certain progress.Research finds, in feedstuff, interpolation probiotic bacteria (lactic acid bacteria) or its metabolite lactic acid can be effectively improved the microbial environment of animal the intestines and stomach, effectively prevent digestive tract disease, excitating organism immunologic function, improve body disease-resistant, anti-stress ability, improve utilization rate and the breeding performonce fo animals of feedstuff.
Therefore, on the basis of current Animal nutrition theoretical system, for presently, there are the deficiency of technology, by aerobic microbiological and anaerobe two-step fermentation, it is achieved the external degradation of Semen arachidis hypogaeae dregs albumen, accumulate useful metabolite polypeptide and digestive enzyme, the substantial amounts of probiotic bacteria of Sync enrichment, prepares probiotic peptide, to the utilization rate and the breeding performonce fo animals that improve feedstuff, alleviate abuse of antibiotics and there is important effect, the healthy and sustainable development of feed industry and aquaculture is had the meaning of the property changed.
Summary of the invention
The technical problem to be solved is to utilize Semen arachidis hypogaeae dregs two step solid fermentation to produce probiotic peptide, and the content of peptides of probiotic peptide reaches more than 25%, and lactic acid content reaches more than 8.0%, and viable count of lactobacillus reaches 109More than CFU/g, acid protease reaches more than 200U/g.Two step solid fermentation are respectively aerobic fermentation and anaerobic fermentation, used microorganism meets the aspergillus niger (CGMCCNo.9183) of " feed additive kind catalogue (2013) ", respectively high yield acid protease and high temperature resistant, the Pediococcus pentosaceus (CGMCCNo.9182) of high-yield quick lactic acid.
Pediococcus pentosaceus, circular colonies, milky, smooth, protruding, neat in edge, opaque, Gram-positive, coccus.Utilizing glucose, xylose, arabinose, galactose, mannose, the multiple sugar such as glucosamine and cellobiose produces acid, acidproof, bile tolerance, has good intestinal colonisation ability.
Aspergillus niger, pitchy, top capsule is big, spherical, and stigma is double-deck, and conidium is spherical, darkly, pitchy, smooth or coarse.Mycelia prosperity multi-branched, has the many cells fungus every multinuclear.Produce acid protease, phytase, cellulase and xylanase etc..
The incubation step of Pediococcus pentosaceus is:
The activated inclined-plane of Pediococcus pentosaceus accesses MRS culture medium 37 DEG C and cultivates 36h, then spreads cultivation step by step 12h with 5% inoculum concentration, until enough solid fermentation consumptions.
MRS culture medium (g L-1): glucose 20, Carnis Bovis seu Bubali cream 10, tryptone 10, yeast extract 5, K2HPO42, Triammonium citrate 2, sodium acetate 5, MgSO40.1, MnSO4·H2O0.05, tween 80 1, pH is natural, 121 DEG C of sterilizing 20min.
The incubation step of aspergillus niger is:
Behind the activated inclined-plane of aspergillus niger, 5d cultivated by line potato culture flat board 30 DEG C, uses aseptic water washing flat board, makes spore suspension.
Potato culture: Rhizoma Solani tuber osi 20g, sucrose 2g, tap water 100mL, natural (about 6.0) the 121 DEG C of 20min that go out of agar 2g, pH.
Rhizoma Solani tuber osi processing method: peeling potatoes, is cut into block and adds water, boil 30min, with filtered through gauze, filtrate sugaring, supplies water to 100ml, loading triangular flask.
The technical scheme is that
A kind of method utilizing Semen arachidis hypogaeae dregs two step solid fermentation to produce probiotic peptide, step is as follows:
With Semen arachidis hypogaeae dregs as raw material, 115-121 DEG C of sterilizing 10-20min after mixing homogeneously with tap water by material water weight ratio 1:0.8-1:1.1, the 2%(v/w of Semen arachidis hypogaeae dregs weight is pressed after sterilizing) access Aspergillus niger spores liquid, at 28 DEG C ~ 32 DEG C, cultivate 24 ~ 28h after mix homogeneously, every 4h stirring is once；After cultivation by 3 batches mixing, by Semen arachidis hypogaeae dregs weight access 10%(v/w) Pediococcus pentosaceus, after mix homogeneously at 37 DEG C ~ 46 DEG C anaerobic fermentation 36 ~ 42h, after fermentation ends 60 DEG C be dried, pulverize, prepare probiotic peptide；Described aspergillus niger (Aspergillusniger) preservation is to being positioned at the China General Microbiological culture presevation administrative center of Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, and on May 20 2014 preservation time, its deposit number is CGMCCNo.9183；Pediococcus pentosaceus (Pediococcuspentosaceus) preservation is to being positioned at the China General Microbiological culture presevation administrative center of Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, and on May 20 2014 preservation time, its deposit number is CGMCCNo.9182.
The invention have the benefit that
The present invention is by the flora of screening effect excellence, and by optimizing preparation technology and parameters, a kind of method providing new production probiotic peptide, Semen arachidis hypogaeae dregs two step solid fermentation is specially utilized to produce probiotic peptide, the content of peptides of the probiotic peptide obtained reaches more than 25%, lactic acid content reaches more than 8.0%, and viable count of lactobacillus reaches 109More than CFU/g, acid protease reaches more than 200U/g.This product realizes the external predigestion of high molecular weight protein in Semen arachidis hypogaeae dregs, and accumulates useful metabolite lactic acid, the polypeptide of absorption easy to digest and acid protease, is enriched with useful lactic acid bacteria.
The probiotic peptide product that the present invention obtains, adds about 5% in feed formula, can improve the abilities of digestive and absorption of animal, regulates animal alimentary canal microecological balance, the suppression pathogenic microorganism such as escherichia coli, Salmonella growth, strengthens the disease resistance ability of animal.
In a word, the preparation method of the present invention is easily controllable, the probiotic peptide constant product quality obtained, and performance is good, and the method possesses prominent substantive distinguishing features and improves significantly.
Contrast effect:
Using fermented peanut meal prepared by one-step fermentation, content of peptides is 8%, and lactic acid content is 3.0%, and probiotics number of viable is 108CFU/g, far below the probiotic peptide prepared by this method.
Test example:
With add 6% fish flour normal feedstuff as test control group, the fish flour substituting in feedstuff 2%, 4%, 6% by probiotic peptide respectively (is equivalent to the 1/3 of fish flour total amount, 2/3,3/3), be made into etc. nitrogen, etc. can feedstuff, supplement crystal lysine, methionine and threonine in each fish flour alternate sets so that it is reach the level consistent with matched group simultaneously.Feedstuff is all pulverized with pulverizer, crosses 40 mesh sieves, is crushed to sinking pellet diet (pelleting temperature 85 DEG C), dries, plastic bag sealing at ventilation lucifugal after mix homogeneously.Carry out 8 weeks raising experiments:
Probiotic peptide prepared by table 1 present invention replaces the fish flour impact on Growth Op Tilapia
Note: the different letter person of colleague's data tool, represents significant difference (P < 0.05)
Table 2 probiotic peptide replaces the fish flour impact on tilapia body index
Note: the different letter person of colleague's data tool, represents significant difference (P < 0.05)
By table 1, table 2 understands, in the tilapia feed that fish meal content is 6%, under conditions of supplementary lysine, methionine and threonine, probiotic peptide prepared by the present invention can replace the 2/3 (4%) of fish flour consumption, test group Growth Op Tilapia index and body index and matched group to be quite even better than matched group.In the feeding process of product probiotic peptide Substitute For Partial fish flour of the present invention, improve acid-base balance in tilapia body, improve the rate of digesting and absorbing, improve immunity, decrease the water pollution brought in breeding process.
Detailed description of the invention
Below by specific embodiment, technical scheme is described in detail.
Embodiment 1
With Semen arachidis hypogaeae dregs as raw material, 121 DEG C of sterilizing 20min after mixing homogeneously with tap water by material-water ratio 1:0.8, the 2%(v/w of Semen arachidis hypogaeae dregs weight is pressed after sterilizing) access aspergillus niger (CGMCCNo.9183) spore liquid, at 28 DEG C ~ 32 DEG C, cultivate 28h after mix homogeneously, every 4h stirring is once；By 3 batch mixing after cultivation, access 10%(v/w by Semen arachidis hypogaeae dregs weight) Pediococcus pentosaceus (CGMCCNo.9182), after mix homogeneously, anaerobic fermentation 42h, dry after fermentation ends 60 DEG C, pulverizing at 37 DEG C ~ 46 DEG C, prepares probiotic peptide.The content of peptides of probiotic peptide reaches 28.4%, and lactic acid content reaches 8.8%, and viable count of lactobacillus reaches 1.2 × 109CFU/g, acid protease reaches 228U/g.
Embodiment 2
With Semen arachidis hypogaeae dregs as raw material, 121 DEG C of sterilizing 20min after mixing homogeneously with tap water by material-water ratio 1:0.8, the 2%(v/w of Semen arachidis hypogaeae dregs weight is pressed after sterilizing) access aspergillus niger (CGMCCNo.9183) spore liquid, at 28 DEG C ~ 32 DEG C, cultivate 26h after mix homogeneously, every 4h stirring is once；By 3 batch mixing after cultivation, access 10%(v/w by Semen arachidis hypogaeae dregs weight) Pediococcus pentosaceus (CGMCCNo.9182), after mix homogeneously, anaerobic fermentation 36h, dry after fermentation ends 60 DEG C, pulverizing at 37 DEG C ~ 46 DEG C, prepares probiotic peptide.The content of peptides of probiotic peptide reaches 26.2%, and lactic acid content reaches 8.3%, and viable count of lactobacillus reaches 1.0 × 109CFU/g, acid protease reaches 212U/g.
Embodiment 3
With Semen arachidis hypogaeae dregs as raw material, 120 DEG C of sterilizing 15min after mixing homogeneously with tap water by material-water ratio 1:1.1, the 2%(v/w of Semen arachidis hypogaeae dregs weight is pressed after sterilizing) access aspergillus niger (CGMCCNo.9183) spore liquid, at 28 DEG C ~ 32 DEG C, cultivate 28h after mix homogeneously, every 4h stirring is once；By 3 batch mixing after cultivation, access 10%(v/w by Semen arachidis hypogaeae dregs weight) Pediococcus pentosaceus (CGMCCNo.9182), after mix homogeneously, anaerobic fermentation 36h, dry after fermentation ends 60 DEG C, pulverizing at 37 DEG C ~ 46 DEG C, prepares probiotic peptide.The content of peptides of probiotic peptide reaches 25.4%, and lactic acid content reaches 8.2%, and viable count of lactobacillus reaches 1.0 × 109CFU/g, acid protease reaches 201U/g.
More than show and describe the ultimate principle of the present invention and principal character and advantages of the present invention.Skilled person will appreciate that of the industry; the present invention is not restricted to the described embodiments; the principle that the present invention is simply described of the description in above-described embodiment and description; without departing from the spirit and scope of the present invention; the present invention also has various changes and modifications; these changes and improvements both fall within scope of the claimed invention, and in claimed scope, appending claims and equivalent thereof define.
Claims (7)
1. utilizing the method that Semen arachidis hypogaeae dregs two step solid fermentation produces probiotic peptide, step is as follows:
With Semen arachidis hypogaeae dregs as raw material, 115-121 DEG C of sterilizing 10-20min after mixing homogeneously with tap water by material water weight ratio 1:0.8-1:1.1, pressing 2% access Aspergillus niger spores liquid of Semen arachidis hypogaeae dregs weight after sterilizing, cultivate 24 ~ 28h after mix homogeneously at 28 DEG C ~ 32 DEG C, every 4h stirring is once；After cultivation by 3 batches mixing, access 10% Pediococcus pentosaceus by Semen arachidis hypogaeae dregs weight, after mix homogeneously at 37 DEG C ~ 46 DEG C anaerobic fermentation 36 ~ 42h, after fermentation ends 60 DEG C be dried, pulverize, prepare probiotic peptide；
Described Pediococcus pentosaceus is the Pediococcus pentosaceus of lactic acid producing high temperature resistant, quick, and preservation is to China General Microbiological culture presevation administrative center, and its deposit number is CGMCC9182.
The method utilizing Semen arachidis hypogaeae dregs two step solid fermentation to produce probiotic peptide the most according to claim 1, it is characterized in that, described aspergillus niger is the aspergillus niger of high yield acid protease, and preservation is to China General Microbiological culture presevation administrative center, and its deposit number is CGMCC9183.
The method utilizing Semen arachidis hypogaeae dregs two step solid fermentation to produce probiotic peptide the most according to claim 1, it is characterised in that the incubation step of described Pediococcus pentosaceus is:
The activated inclined-plane of Pediococcus pentosaceus accesses MRS culture medium 37 DEG C and cultivates 36h, then spreads cultivation step by step 12h with 5% inoculum concentration, until enough solid fermentation consumptions；
MRS culture medium: glucose 20g/L, Carnis Bovis seu Bubali cream 10g/L, tryptone 10g/L, yeast extract 5g/L, K2HPO42g/L, Triammonium citrate 2g/L, sodium acetate 5g/L, MgSO40.1g/L, MnSO4·H2O0.05g/L, tween 80 1g/L, pH are natural, 121 DEG C of sterilizing 20min.
The method utilizing Semen arachidis hypogaeae dregs two step solid fermentation to produce probiotic peptide the most according to claim 2, it is characterised in that the incubation step of described aspergillus niger is:
Behind the activated inclined-plane of aspergillus niger, 5d cultivated by line potato culture flat board 30 DEG C, uses aseptic water washing flat board, makes spore suspension；
Potato culture: Rhizoma Solani tuber osi 20g, sucrose 2g, tap water 100mL, agar 2g, pH are natural, 121 DEG C of sterilizing 20min；
Rhizoma Solani tuber osi processing method: peeling potatoes, is cut into block and adds water, boil 30min, with filtered through gauze, filtrate sugaring, supplies water to 100ml, loading triangular flask.
5. the probiotic peptide that prepared by the method as described in any one of claim 1-4.
6. the probiotic peptide that prepared by the method as described in any one of claim 1-4, content of peptides reaches more than 25%, and lactic acid content reaches more than 8.0%, and viable count of lactobacillus reaches 109More than CFU/g, acid protease reaches more than 200U/g.
7. the application in preparing feedstuff of the probiotic peptide as described in claim 5 or 6.
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| CN104543374A (en) * | 2015-01-20 | 2015-04-29 | 青岛天祥食品集团有限公司 | High-value utilization method for high-nutrition peanut hot-pressed cakes |
| CN105146050B (en) * | 2015-09-25 | 2019-05-14 | 华南理工大学 | A kind of nutrient base material and its production method and purposes rich in essential amino acid |
| CN110818459B (en) * | 2018-08-10 | 2021-10-22 | 中科绿帅生物科技(广州)有限公司 | Liquid peanut meal organic fertilizer and preparation method and application thereof |
| CN109258921A (en) * | 2018-11-01 | 2019-01-25 | 青岛农业大学 | A kind of fermentation material of the Maize-Peanut dregs of rice, its fermentation process and application |
| CN110583964A (en) * | 2019-09-23 | 2019-12-20 | 江南大学 | Biological removal method for efficiently removing four aflatoxins in peanut meal |
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| CN103361395B (en) * | 2013-08-07 | 2015-10-14 | 山东省花生研究所 | A kind of solid state fermentation prepares the method for peanut non-starch polysaccharide |
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