CN105925645A - Preparation method of active lactobacillin and application of active lactobacillin in biological feed - Google Patents
Preparation method of active lactobacillin and application of active lactobacillin in biological feed Download PDFInfo
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- CN105925645A CN105925645A CN201610299281.4A CN201610299281A CN105925645A CN 105925645 A CN105925645 A CN 105925645A CN 201610299281 A CN201610299281 A CN 201610299281A CN 105925645 A CN105925645 A CN 105925645A
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P21/00—Preparation of peptides or proteins
- C12P21/02—Preparation of peptides or proteins having a known sequence of two or more amino acids, e.g. glutathione
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- 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
- C12N11/00—Carrier-bound or immobilised enzymes; Carrier-bound or immobilised microbial cells; Preparation thereof
- C12N11/02—Enzymes or microbial cells immobilised on or in an organic carrier
- C12N11/10—Enzymes or microbial cells immobilised on or in an organic carrier the carrier being a carbohydrate
Abstract
The invention discloses a preparation method of active lactobacillin and an application of the active lactobacillin in biological feed, wherein the preparation method comprises the following steps: skimming fresh milk so as to obtain skimmed milk, and regulating pH value to 4-5, so as to obtain a yellow green transparent whey liquid; adding 5-15g of peptone and 2-5g of a yeast extract in terms of each liter of the whey liquid, and regulating pH value so as to obtain a neutral culture solution; sterilizing the culture solution at 100-120 DEG C for 10-20min, cooling the sterilized solution to 30-40 DEG C, adding a compound Heteroclitin I, then inoculating immobilized lactobacillus acidophilus, fermenting and cultivating for 18-24h and controlling pH value at 6-7; and after fermentation, centrifuging a fermentation broth, uniformly mixing a supernatant and a composite thick slurry according to a weight ratio of (1-3) to 1, and freeze-drying at minus 30-50 DEG C so as to obtain the active lactobacillin, wherein the composite thick slurry is a composite solution of glucose and starch, the mass percentage concentration of the glucose is 20-30% and the mass percentage concentration of the starch is 5-15%, and the composite thick slurry is used after being sterilized. The active lactobacillin provided by the invention can be used for significantly improving the growth performance of weaned piglets.
Description
Technical field
The invention belongs to biological feedstuff field, relate to antibacterial peptide, be specifically related to a kind of active lactein preparation method and
Application in biological feedstuff.
Background technology
Lactein is a kind of active polypeptide or protein produced by lactic acid bacteria.Major part bacteriocin is to Characterization
Antibacterial has inhibitory action, and minority bacteriocin antimicrobial spectrum is the widest.Lactein can be divided mainly into heat-staple small-molecular peptides, thermo-responsive
High molecular weight protein and lantibiotics.
The lactein of commercial type is the preparation that a kind of pure lactic acid bacteria produces, and is through sterilizing, being concentrated in vacuo by skimmed milk, then
The lactic acid thalline being dried after inoculating lactic acid bacterium fermentation and make and the mixing powder of metabolite thereof, can make pharmaceutical or food additive
Add agent, function of dominant type milk product.
Lactein stable in properties, is not limited by processing conditions, so range of application is the most extensive, can be applicable to include fermentation system
The fields such as milk, health food, cosmetics and food fresh keeping.The antibacterial functions of lactein is mainly by its protein structure
On the difference of different loci charge number, object bacteria membrane structure part can be caused to change, there is similar cleaning agent function.One
For as, its bactericidal mechanism be first adsorb on the cell membrane of object bacteria, invade film again in and form penetrating duct, to cause born of the same parents
Interior important substance, such as ATP, K+Deng loss or biochemical reaction obstacle, and cause Bacteria Indicators dead.The effect class of lactein
Like antibiotic, but mechanism of action is different, has specificity, does not develops immunity to drugs and toxicity, and be difficult to by small intestinal Trypsin
Enzyme destroys, and can effectively suppress the growth of pathogen.
Lactein may be used for the additive of biological feedstuff, can improve the Anti-bacterium ability of domestic animal, improve the appetite of domestic animal,
And then improve the growth performance of domestic animal.
Summary of the invention
The first object of the present invention is to provide a kind of highly active lactein, for the additive of biological feedstuff, significantly changes
The growth performance of kind domestic animal；
The second object of the present invention is to provide a kind of biological feedstuff containing above-mentioned high-activity lactic acid bacteria element.
The above-mentioned purpose of the present invention is achieved by techniques below scheme:
A kind of active lactein, is made by the steps and forms:
Step S1, first obtains skimmed milk by sweet milk defat, and the pH value of regulation skimmed milk, to 4～5, removes isolating protein, obtains Huang
Green transparent whey；
Step S2, modulates culture fluid: every liter of whey adds peptone 5～15 grams, and yeast extract 2～5 grams, regulation pH value is to neutral；
Step S3, prepares bacillus acidophilus's immobilized cell: use MARS culture medium culturing bacillus acidophilus extremely
10^10cfu/mL, the centrifugal 0.9% normal saline mixing going supernatant, lower floor's bacterium mud to add 1:1 mass ratio, prepare addicted to yogurt bar
Bacterium diluent, every 1 volume bacillus acidophilus's diluent adds 40～the sodium alginate soln of 60 volumes 2.5%～3.5%, 38 DEG C of water-baths
Inhalation syringe after 5 minutes, is slowly dropped in 3% calcium chloride solution of 200～300 volumes, forms 2～3mm diameter beads,
Ambient-temp-stable 8～10 hours.
Step S4, by the culture fluid of preparation sterilizing 10～20 minutes at 100～120 DEG C in step S2, after being cooled to 30～40 DEG C,
First add compound Heteroclitin I so that it is the concentration in culture fluid is 20～30 μ g/g, then access by the inoculum concentration of 3～5%
In step S3 preparation immobilization lactobacillus acidophilus cells's fermentation culture 18～24 hours, control ph is 6～7；
Step S5, after fermentation ends, mixes fermentation liquid centrifuged supernatant according to weight ratio 1～3:1 with compound underflow,
After mix homogeneously, at-30～-50 DEG C, lyophilization obtains active lactein；Wherein, described compound underflow is Fructus Vitis viniferae sugar and starch
Composite solution, the mass percentage concentration of glucose is 20～30%, and the mass percentage concentration of starch is 5～15%, makes after sterilizing
With.
Further, described active lactein is made by the steps and forms:
Step S1, first obtains skimmed milk by sweet milk defat, and the pH value of regulation skimmed milk, to 4.5, removes isolating protein, obtains yellowish green
Color transparent milk clear liquid；
Step S2, modulates culture fluid: every liter of whey adds peptone 10 grams, yeast extract 3.5 grams, and regulation pH value is to neutral；
Step S3, prepares bacillus acidophilus's immobilized cell: use MARS culture medium culturing bacillus acidophilus extremely
10^10cfu/mL, the centrifugal 0.9% normal saline mixing going supernatant, lower floor's bacterium mud to add 1:1 mass ratio, prepare addicted to yogurt bar
Bacterium diluent, every 1 volume bacillus acidophilus's diluent adds the sodium alginate soln of 40 volumes 2.5%, after 38 DEG C of water-baths 5 minutes
Inhalation syringe, is slowly dropped in 3% calcium chloride solution of 200 volumes, forms 2mm diameter bead, ambient-temp-stable 8 hours.
Step S4, by above-mentioned culture fluid sterilizing 15 minutes at 110 DEG C of preparation in step S2, after being cooled to 35 DEG C, first adds
Add compound Heteroclitin I so that it is the concentration in culture fluid is 25 μ g/g, then access in step S3 by the inoculum concentration of 4%
Immobilization lactobacillus acidophilus cells's fermentation culture of preparation 21 hours, control ph is 6～7；
Step S5, after fermentation ends, mixes fermentation liquid centrifuged supernatant according to weight ratio 2:1 with compound underflow, mixed
After closing uniformly, at-40 DEG C, lyophilization obtains active lactein；Wherein, described compound underflow is the compound of Fructus Vitis viniferae sugar and starch
Solution, the mass percentage concentration of glucose is 25%, and the mass percentage concentration of starch is 10%, uses after sterilizing.
Further, described active lactein is made by the steps and forms:
Step S1, first obtains skimmed milk by sweet milk defat, and the pH value of regulation skimmed milk, to 4, removes isolating protein, obtains yellowish green
Color transparent milk clear liquid；
Step S2, modulates culture fluid: every liter of whey adds peptone 5 grams, yeast extract 2 grams, and regulation pH value is to neutral；
Step S3, prepares bacillus acidophilus's immobilized cell: use MARS culture medium culturing bacillus acidophilus extremely
10^10cfu/mL, the centrifugal 0.9% normal saline mixing going supernatant, lower floor's bacterium mud to add 1:1 mass ratio, prepare addicted to yogurt bar
Bacterium diluent, every 1 volume bacillus acidophilus's diluent adds the sodium alginate soln of 50 volumes 3%, and 38 DEG C of water-baths were inhaled after 5 minutes
Enter syringe, be slowly dropped in 3% calcium chloride solution of 250 volumes, form 2.5mm diameter bead, ambient-temp-stable 9 hours.
Step S4, by above-mentioned culture fluid sterilizing 20 minutes at 100 DEG C of preparation in step S2, after being cooled to 30 DEG C, first adds
Add compound Heteroclitin I so that it is the concentration in culture fluid is 20 μ g/g, then access in step S3 by the inoculum concentration of 3%
Immobilization lactobacillus acidophilus cells's fermentation culture of preparation 24 hours, control ph is 6～7；
Step S5, after fermentation ends, mixes fermentation liquid according to weight ratio 3:1 with compound underflow, at-30 DEG C after mix homogeneously
Lower lyophilization obtains active lactein；Wherein, described compound underflow is the composite solution of Fructus Vitis viniferae sugar and starch, glucose
Mass percentage concentration is 20%, and the mass percentage concentration of starch is 5%, uses after sterilizing.
Further, described active lactein is made by the steps and forms:
Step S1, first obtains skimmed milk by sweet milk defat, and the pH value of regulation skimmed milk, to 5, removes isolating protein, obtains yellowish green
Color transparent milk clear liquid；
Step S2, modulates culture fluid: every liter of whey adds peptone 15 grams, yeast extract 5 grams, and regulation pH value is to neutral；
Step S3, prepares bacillus acidophilus's immobilized cell: use MARS culture medium culturing bacillus acidophilus extremely
10^10cfu/mL, the centrifugal 0.9% normal saline mixing going supernatant, lower floor's bacterium mud to add 1:1 mass ratio, prepare addicted to yogurt bar
Bacterium diluent, every 1 volume bacillus acidophilus's diluent adds the sodium alginate soln of 60 volumes 3.5%, after 38 DEG C of water-baths 5 minutes
Inhalation syringe, is slowly dropped in 3% calcium chloride solution of 300 volumes, forms 3mm diameter bead, ambient-temp-stable 10 hours.
Step S4, by above-mentioned culture fluid sterilizing 10 minutes at 120 DEG C of preparation in step S2, after being cooled to 40 DEG C, first adds
Add compound Heteroclitin I so that it is the concentration in culture fluid is 30 μ g/g, then access in step S3 by the inoculum concentration of 5%
Immobilization lactobacillus acidophilus cells's fermentation culture of preparation 18 hours, control ph is 6～7；
Step S5, after fermentation ends, mixes fermentation liquid centrifuged supernatant according to weight ratio 1:1 with compound underflow, mixed
After closing uniformly, at-50 DEG C, lyophilization obtains active lactein；Wherein, described compound underflow is the compound of Fructus Vitis viniferae sugar and starch
Solution, the mass percentage concentration of glucose is 30%, and the mass percentage concentration of starch is 15%, uses after sterilizing.
A kind of biological feedstuff, containing above-mentioned active lactein, adds 60～80g active lacteins in every 100kg biological feedstuff.
The application in terms of improving domestic animal growth performance of the above-mentioned active lactein.
The application in terms of improving domestic animal growth performance of the above-mentioned biological feedstuff.
Advantages of the present invention:
The active lactein biological activity that the present invention provides is high, after adding in feedstuff, can significantly improve ablactational baby pig growth
Performance.The present invention compared with prior art, has prominent substantive distinguishing features and significantly progress.
Detailed description of the invention
Further illustrate the essentiality content of the present invention below in conjunction with embodiment, but do not limit scope with this.To the greatest extent
The present invention is explained in detail by pipe with reference to preferred embodiment, it will be understood by those within the art that, can be to the present invention
Technical scheme modify or equivalent, without deviating from the spirit and scope of technical solution of the present invention.
In the present invention, do not do the test method emphasized especially and reagent is this area conventional methods and reagent.Addicted to yogurt bar
Bacterium selects Lactobacillus acidophilus ATCC 4356, and other also may be used.The preparation method of compound Heteroclitin I sees document:
Compounds from Kadsura heteroclita and related anti-HIV activity,Phytochemistry,69(2008)
1266-1272。
Embodiment 1: the preparation of active lactein
Step S1, first obtains skimmed milk by sweet milk defat, and the pH value of regulation skimmed milk, to 4.5, removes isolating protein, obtains yellowish green
Color transparent milk clear liquid；
Step S2, modulates culture fluid: every liter of whey adds peptone 10 grams, yeast extract 3.5 grams, and regulation pH value is to neutral；
Step S3, prepares bacillus acidophilus's immobilized cell: use MARS culture medium culturing bacillus acidophilus extremely
10^10cfu/mL, the centrifugal 0.9% normal saline mixing going supernatant, lower floor's bacterium mud to add 1:1 mass ratio, prepare addicted to yogurt bar
Bacterium diluent, every 1 volume bacillus acidophilus's diluent adds the sodium alginate soln of 40 volumes 2.5%, after 38 DEG C of water-baths 5 minutes
Inhalation syringe, is slowly dropped in 3% calcium chloride solution of 200 volumes, forms 2cm diameter bead, ambient-temp-stable 8 hours.
Step S4, by above-mentioned culture fluid sterilizing 15 minutes at 110 DEG C of preparation in step S2, after being cooled to 35 DEG C, first adds
Add compound Heteroclitin I so that it is the concentration in culture fluid is 25 μ g/g, then access in step S3 by the inoculum concentration of 4%
Immobilization lactobacillus acidophilus cells's fermentation culture of preparation 21 hours, control ph is 6～7；
Step S5, after fermentation ends, mixes fermentation liquid centrifuged supernatant according to weight ratio 2:1 with compound underflow, mixed
After closing uniformly, at-40 DEG C, lyophilization obtains active lactein；Wherein, described compound underflow is the compound of Fructus Vitis viniferae sugar and starch
Solution, the mass percentage concentration of glucose is 25%, and the mass percentage concentration of starch is 10%, uses after sterilizing.
Embodiment 2: the preparation of active lactein
Step S1, first obtains skimmed milk by sweet milk defat, and the pH value of regulation skimmed milk, to 4, removes isolating protein, obtains yellowish green
Color transparent milk clear liquid；
Step S2, modulates culture fluid: every liter of whey adds peptone 5 grams, yeast extract 2 grams, and regulation pH value is to neutral；
Step S3, prepares bacillus acidophilus's immobilized cell: use MARS culture medium culturing bacillus acidophilus extremely
10^10cfu/mL, the centrifugal 0.9% normal saline mixing going supernatant, lower floor's bacterium mud to add 1:1 mass ratio, prepare addicted to yogurt bar
Bacterium diluent, every 1 volume bacillus acidophilus's diluent adds the sodium alginate soln of 50 volumes 3%, and 38 DEG C of water-baths were inhaled after 5 minutes
Enter syringe, be slowly dropped in 3% calcium chloride solution of 250 volumes, form 2.5mm diameter bead, ambient-temp-stable 9 hours.
Step S4, by above-mentioned culture fluid sterilizing 20 minutes at 100 DEG C of preparation in step S2, after being cooled to 30 DEG C, first adds
Add compound Heteroclitin I so that it is the concentration in culture fluid is 20 μ g/g, then access in step S3 by the inoculum concentration of 3%
Immobilization lactobacillus acidophilus cells's fermentation culture of preparation 24 hours, control ph is 6～7；
Step S5, after fermentation ends, mixes fermentation liquid centrifuged supernatant according to weight ratio 3:1 with compound underflow, mixed
After closing uniformly, at-30 DEG C, lyophilization obtains active lactein；Wherein, described compound underflow is the compound of Fructus Vitis viniferae sugar and starch
Solution, the mass percentage concentration of glucose is 20%, and the mass percentage concentration of starch is 5%, uses after sterilizing.
Embodiment 3: the preparation of active lactein
Step S1, first obtains skimmed milk by sweet milk defat, and the pH value of regulation skimmed milk, to 5, removes isolating protein, obtains yellowish green
Color transparent milk clear liquid；
Step S2, modulates culture fluid: every liter of whey adds peptone 15 grams, yeast extract 5 grams, and regulation pH value is to neutral；
Step S3, prepares bacillus acidophilus's immobilized cell: use MARS culture medium culturing bacillus acidophilus extremely
10^10cfu/mL, the centrifugal 0.9% normal saline mixing going supernatant, lower floor's bacterium mud to add 1:1 mass ratio, prepare addicted to yogurt bar
Bacterium diluent, every 1 volume bacillus acidophilus's diluent adds the sodium alginate soln of 60 volumes 3.5%, after 38 DEG C of water-baths 5 minutes
Inhalation syringe, is slowly dropped in 3% calcium chloride solution of 300 volumes, forms 3mm diameter bead, ambient-temp-stable 10 hours.
Step S4, by above-mentioned culture fluid sterilizing 10 minutes at 120 DEG C of preparation in step S2, after being cooled to 40 DEG C, first adds
Add compound Heteroclitin I so that it is the concentration in culture fluid is 30 μ g/g, then access in step S3 by the inoculum concentration of 5%
Immobilization lactobacillus acidophilus cells's fermentation culture of preparation 18 hours, control ph is 6～7；
Step S5, after fermentation ends, mixes fermentation liquid centrifuged supernatant according to weight ratio 1:1 with compound underflow, mixed
After closing uniformly, at-50 DEG C, lyophilization obtains active lactein；Wherein, described compound underflow is the compound of Fructus Vitis viniferae sugar and starch
Solution, the mass percentage concentration of glucose is 30%, and the mass percentage concentration of starch is 15%, uses after sterilizing.
Embodiment 4: the contrast of embodiment 1, without compound Heteroclitin I
Step S1, first obtains skimmed milk by sweet milk defat, and the pH value of regulation skimmed milk, to 4.5, removes isolating protein, obtains yellowish green
Color transparent milk clear liquid；
Step S2, modulates culture fluid: every liter of whey adds peptone 10 grams, yeast extract 3.5 grams, and regulation pH value is to neutral；
Step S3, prepares bacillus acidophilus's immobilized cell: use MARS culture medium culturing bacillus acidophilus extremely
10^10cfu/mL, the centrifugal 0.9% normal saline mixing going supernatant, lower floor's bacterium mud to add 1:1 mass ratio, prepare addicted to yogurt bar
Bacterium diluent, every 1 volume bacillus acidophilus's diluent adds the sodium alginate soln of 40 volumes 2.5%, after 38 DEG C of water-baths 5 minutes
Inhalation syringe, is slowly dropped in 3% calcium chloride solution of 200 volumes, forms 2cm diameter bead, ambient-temp-stable 8 hours.
Step S4, by above-mentioned culture fluid sterilizing 15 minutes at 110 DEG C of preparation in step S2, after being cooled to 35 DEG C, by 4%
Inoculum concentration access immobilization lactobacillus acidophilus cells's fermentation culture of preparation 21 hours in step S3, control ph is 6～7；
Step S5, after fermentation ends, mixes fermentation liquid centrifuged supernatant according to weight ratio 2:1 with compound underflow, mixed
After closing uniformly, at-40 DEG C, lyophilization obtains active lactein；Wherein, described compound underflow is the compound of Fructus Vitis viniferae sugar and starch
Solution, the mass percentage concentration of glucose is 25%, and the mass percentage concentration of starch is 10%, uses after sterilizing.
Embodiment 5: effect example, the impact on Growth Performance of Weaning Piglets
One, materials and methods
1, material: prepare active lactein 1～4 by embodiment 1～4 method respectively.
2, method
2.1 experimental animals and packet
With reference to NRC (1998) preparation Basic drawing.Select 100 healthy (21 ± 1) age in days wean DLY ternarys miscellaneous
Hand over piglet, be randomly divided into 5 groups, often group 20, male and female half and half.Matched group uses Basic drawing to feed.Test group 1～4 is respectively
On the basis of Basic drawing, add active lactein 1～4 respectively, every 100kg feedstuff adds 70g active lactein.
2.2 method for breeding
30 days test periods.Use routine to raise and immunity during whole test, free choice feeding, freely drink water.
2.3 test indexs and method
Growth performance: statistics initial weight, end weight, Average weight increasing a day and feed-weight ratio.
3, statistical analysis
Test data uses SPSS20.0 software to carry out variance analysis, and the significance of difference carries out Duncans multiple comparisons, number in table
Value is expressed as Mean ± SD.
Two, result
The results are shown in Table 1.
As shown in Table 1, the Average weight increasing a day of test group 1～3 is above matched group, and feed-weight ratio is below matched group, with matched group phase
Than significant difference (P ＜ 0.05)；Test group 4 Average weight increasing a day and matched group there was no significant difference (P ＞ 0.05), and feed-weight ratio is with right
(P ＞ 0.05) is there was no significant difference according to group.
The table 1 impact on Growth Performance of Weaning Piglets
Result shows, the feedstuff being added with the active lactein that the present invention provides can significantly improve the growth performance of ablactational baby pig.
The effect of above-described embodiment indicates that the essentiality content of the present invention, but does not limit protection scope of the present invention with this.
It will be understood by those within the art that, technical scheme can be modified or equivalent, and not take off
Essence and protection domain from technical solution of the present invention.
Claims (7)
1. an active lactein, it is characterised in that be made by the steps and form:
Step S1, first obtains skimmed milk by sweet milk defat, and the pH value of regulation skimmed milk, to 4～5, removes isolating protein, obtains Huang
Green transparent whey；
Step S2, modulates culture fluid: every liter of whey adds peptone 5～15 grams, and yeast extract 2～5 grams, regulation pH value is to neutral；
Step S3, prepares bacillus acidophilus's immobilized cell: use MARS culture medium culturing bacillus acidophilus extremely
10^10cfu/mL, the centrifugal 0.9% normal saline mixing going supernatant, lower floor's bacterium mud to add 1:1 mass ratio, prepare addicted to yogurt bar
Bacterium diluent, every 1 volume bacillus acidophilus's diluent adds 40～the sodium alginate soln of 60 volumes 2.5%～3.5%, 38 DEG C of water-baths
Inhalation syringe after 5 minutes, is slowly dropped in 3% calcium chloride solution of 200～300 volumes, forms 2～3mm diameter beads,
Ambient-temp-stable 8～10 hours.
Step S4, by the culture fluid of preparation sterilizing 10～20 minutes at 100～120 DEG C in step S2, after being cooled to 30～40 DEG C,
First add compound Heteroclitin I so that it is the concentration in culture fluid is 20～30 μ g/g, then access by the inoculum concentration of 3～5%
In step S3 preparation immobilization lactobacillus acidophilus cells's fermentation culture 18～24 hours, control ph is 6～7；
Step S5, after fermentation ends, mixes fermentation liquid centrifuged supernatant according to weight ratio 1～3:1 with compound underflow,
After mix homogeneously, at-30～-50 DEG C, lyophilization obtains active lactein；Wherein, described compound underflow is Fructus Vitis viniferae sugar and starch
Composite solution, the mass percentage concentration of glucose is 20～30%, and the mass percentage concentration of starch is 5～15%, makes after sterilizing
With.
Active lactein the most according to claim 1, it is characterised in that be made by the steps and form:
Step S1, first obtains skimmed milk by sweet milk defat, and the pH value of regulation skimmed milk, to 4.5, removes isolating protein, obtains yellowish green
Color transparent milk clear liquid；
Step S2, modulates culture fluid: every liter of whey adds peptone 10 grams, yeast extract 3.5 grams, and regulation pH value is to neutral；
Step S3, prepares bacillus acidophilus's immobilized cell: use MARS culture medium culturing bacillus acidophilus extremely
10^10cfu/mL, the centrifugal 0.9% normal saline mixing going supernatant, lower floor's bacterium mud to add 1:1 mass ratio, prepare addicted to yogurt bar
Bacterium diluent, every 1 volume bacillus acidophilus's diluent adds the sodium alginate soln of 40 volumes 2.5%, after 38 DEG C of water-baths 5 minutes
Inhalation syringe, is slowly dropped in 3% calcium chloride solution of 200 volumes, forms 2mm diameter bead, ambient-temp-stable 8 hours.
Step S4, by above-mentioned culture fluid sterilizing 15 minutes at 110 DEG C of preparation in step S2, after being cooled to 35 DEG C, first adds
Add compound Heteroclitin I so that it is the concentration in culture fluid is 25 μ g/g, then access in step S3 by the inoculum concentration of 4%
Immobilization lactobacillus acidophilus cells's fermentation culture of preparation 21 hours, control ph is 6～7；
Step S5, after fermentation ends, mixes fermentation liquid centrifuged supernatant according to weight ratio 2:1 with compound underflow, mixed
After closing uniformly, at-40 DEG C, lyophilization obtains active lactein；Wherein, described compound underflow is the compound of Fructus Vitis viniferae sugar and starch
Solution, the mass percentage concentration of glucose is 25%, and the mass percentage concentration of starch is 10%, uses after sterilizing.
Active lactein the most according to claim 1, it is characterised in that be made by the steps and form:
Step S1, first obtains skimmed milk by sweet milk defat, and the pH value of regulation skimmed milk, to 4, removes isolating protein, obtains yellowish green
Color transparent milk clear liquid；
Step S2, modulates culture fluid: every liter of whey adds peptone 5 grams, yeast extract 2 grams, and regulation pH value is to neutral；
Step S3, prepares bacillus acidophilus's immobilized cell: use MARS culture medium culturing bacillus acidophilus extremely
10^10cfu/mL, the centrifugal 0.9% normal saline mixing going supernatant, lower floor's bacterium mud to add 1:1 mass ratio, prepare addicted to yogurt bar
Bacterium diluent, every 1 volume bacillus acidophilus's diluent adds the sodium alginate soln of 50 volumes 3%, and 38 DEG C of water-baths were inhaled after 5 minutes
Enter syringe, be slowly dropped in 3% calcium chloride solution of 250 volumes, form 2.5mm diameter bead, ambient-temp-stable 9 hours.
Step S4, by above-mentioned culture fluid sterilizing 20 minutes at 100 DEG C of preparation in step S2, after being cooled to 30 DEG C, first adds
Add compound Heteroclitin I so that it is the concentration in culture fluid is 20 μ g/g, then access in step S3 by the inoculum concentration of 3%
Immobilization lactobacillus acidophilus cells's fermentation culture of preparation 24 hours, control ph is 6～7；
Step S5, after fermentation ends, mixes fermentation liquid centrifuged supernatant according to weight ratio 3:1 with compound underflow, mixed
After closing uniformly, at-30 DEG C, lyophilization obtains active lactein；Wherein, described compound underflow is the compound of Fructus Vitis viniferae sugar and starch
Solution, the mass percentage concentration of glucose is 20%, and the mass percentage concentration of starch is 5%, uses after sterilizing.
Active lactein the most according to claim 1, it is characterised in that be made by the steps and form:
Step S1, first obtains skimmed milk by sweet milk defat, and the pH value of regulation skimmed milk, to 5, removes isolating protein, obtains yellowish green
Color transparent milk clear liquid；
Step S2, modulates culture fluid: every liter of whey adds peptone 15 grams, yeast extract 5 grams, and regulation pH value is to neutral；
Step S3, prepares bacillus acidophilus's immobilized cell: use MARS culture medium culturing bacillus acidophilus extremely
10^10cfu/mL, the centrifugal 0.9% normal saline mixing going supernatant, lower floor's bacterium mud to add 1:1 mass ratio, prepare addicted to yogurt bar
Bacterium diluent, every 1 volume bacillus acidophilus's diluent adds the sodium alginate soln of 60 volumes 3.5%, after 38 DEG C of water-baths 5 minutes
Inhalation syringe, is slowly dropped in 3% calcium chloride solution of 300 volumes, forms 3mm diameter bead, ambient-temp-stable 10 hours.
Step S4, by above-mentioned culture fluid sterilizing 10 minutes at 120 DEG C of preparation in step S2, after being cooled to 40 DEG C, first adds
Add compound Heteroclitin I so that it is the concentration in culture fluid is 30 μ g/g, then access in step S3 by the inoculum concentration of 5%
Immobilization lactobacillus acidophilus cells's fermentation culture of preparation 18 hours, control ph is 6～7；
Step S5, after fermentation ends, mixes fermentation liquid centrifuged supernatant according to weight ratio 1:1 with compound underflow, mixed
After closing uniformly, at-50 DEG C, lyophilization obtains active lactein；Wherein, described compound underflow is the compound of Fructus Vitis viniferae sugar and starch
Solution, the mass percentage concentration of glucose is 30%, and the mass percentage concentration of starch is 15%, uses after sterilizing.
5. a biological feedstuff, it is characterised in that: containing the arbitrary described active lactein of Claims 1 to 4, every 100kg
Biological feedstuff adds 60～80g active lacteins.
6. Claims 1 to 4 arbitrary described active lactein application in terms of improving domestic animal growth performance.
7. the application in terms of improving domestic animal growth performance of the biological feedstuff described in claim 5.
Priority Applications (1)
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