CN103392911B - Feed-use high-activity lactobacillus solid preparation and preparation method thereof - Google Patents
Feed-use high-activity lactobacillus solid preparation and preparation method thereof Download PDFInfo
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Abstract
The invention discloses a feed-use high-activity lactobacillus solid preparation and a preparation method thereof. The feed-use high-activity lactobacillus solid preparation comprises growing beneficial lactobacillus bacterium mud, a protection formula and a basic formula. The invention also provides the preparation method of the feed-use high-activity lactobacillus solid preparation, lactobacillus can be protected through synergistic effects of components in the protection formula and the basic formula, and the living bacterium survival rate in the whole process can be improved. The lactobacillus in the prepared feed-use high-activity lactobacillus solid preparation has improved high temperature and high humidity resistances, has a high survival rate in a simulated intestinal fluid, and a substantial improved animal application effect.
Description
Technical field
The present invention relates to a kind of additive for microbe feedstuff, particularly a kind of feeding high-activity lactic acid bacteria solid formulation and preparation method thereof, belongs to microbial preparation field.
Background technology
Probio, as one of antibiotic substitute, is applied in feed additive field widely.In animal body, microorganism is never manufactured energy by the chyme of gastro-intestinal digestion, stimulates intestinal tract immune system, or provides essential vitamin, adjustment intestinal microecology and metabolic activity thereof.Although probiotics preparation progressively develops into the antibiotic substitute products most with application prospect; but probio especially has the lactic acid bacteria of enteron aisle adhesive attraction; in production, processing, store and very easily inactivation in use procedure; poor to the resistance of hydrochloric acid in gastric juice, cholate etc.; therefore, the probiotics preparation technology researching and developing production overall process available protecting has become focus both domestic and external.
In prior art and patent, the effect of probiotics preparation (especially lactic acid bacteria) by various factors, as the holding time etc. of the bacterium composition of animal species, physiological status, feed type, processing technology, preparation, feeding method, dosage, moisture, temperature, pH value and bacteria preparation.The stability of microorganism how is kept to be engaged in lactobacillus preparation research and produce the most real and sixty-four dollar question faced.
(1) activity problems of lactic acid bacteria.In transport, use and easy in inactivation in preservation process, thus reduce biological action, cause repeatability and the less stable of preparation result of use.
(2) stability problem in feed manufacturing process.The test of 80 DEG C of high temperature in feed manufacturing pelletization must be stood, so bacterial classification and the stability of preparation to temperature thereof seem particularly important to lactobacillus feed additive.Different bacterial classifications is comparatively large to the tolerance difference of high temperature, and general pelleting temperature is less on bacillus impact, larger on the impact such as Bacillus acidi lactici, saccharomycete.In addition, the mineral matter in holding time of feed, feed is (as heavy metal ion Cu
2+, Zn
2+, Mn
2+, Fe
2+deng), choline etc. also can affect the vigor of probio.
(3) in animal body, lactobacillus preparation must pass through gastric environment, arrives enteron aisle and be settled in its physiological function of competence exertion on intestinal mucosa with a large amount of viable bacterias.But because most of lactic acid bacteria is just dead because of hydrochloric acid in gastric juice and bile effect before entering enteron aisle, the ability of propagation of therefore surviving in host is lower, have impact on their probiotic effects to a great extent.Also an important indicator of lactic acid bacteria effect is just become in the survival rate of enteron aisle.And any product standard declaring to be added with probio is must containing at least 10
6-10
7the activated probio (FAO/WHO, 2001) of cfu/g.
(4) pharmaceutical formulation is single, and versatility is strong, thus limits the application in the lactobacillus preparation that same technique or method prepared at non-designated bacterial classification.
Summary of the invention
First technical problem that the present invention will solve is to provide a kind of feeding high-activity lactic acid bacteria solid formulation, in said preparation, Lactobacillus Survival is high, high-temp resisting high-humidity resisting ability improves, and survival rate is high in intestinal juice, alternative antibiotic, significantly can promote production performance and the health performance of Broiler chicks, effectively reduce grice diarrhoea rate.
Second technical problem that the present invention will solve is to provide a kind of preparation method of feeding high-activity lactic acid bacteria solid formulation, and this preparation method has following characteristics:
(1) there is versatility, technique is simple, to common commercial lactic acid bacterium, comprise VREF, enterococcus faecalis, lactoenterococcus, bifidobacterium bifidum, lactobacillus acidophilus, Lactobacillus casei, lactobacillus lactis, Lactobacillus plantarum, Pediococcus acidilactici, Pediococcus pentosaceus or bacillus bulgaricus, there is applicability widely, and can be used for a large amount of production, simple to operate, cost of supplementary product is cheap;
(2) adopt " two-step method ", namely add basic recipe and protection formula, prepare microorganism formulation, lactic acid bacteria is produced, store and use procedure available protecting, improve the Viable detection in whole process;
(3) unique recipe ingredient, ratio and order of addition, make lactic acid bacteria, protective agent, the various auxiliary agent of prebiotic Summing Factor interact, cooperatively interact, to reach the object effectively ensureing viable count.
For solving first technical problem, the present invention by the following technical solutions:
The invention provides a kind of feeding high-activity lactic acid bacteria solid formulation, it comprises probiotic lactobacillus bacterium mud, protection formula and basic recipe; Wherein,
Described probiotic lactobacillus bacterium mud is VREF (Enterococcus faecium), enterococcus faecalis (Enterococcusfaecalis), lactoenterococcus (Enterococcus lactis), bifidobacterium bifidum (Bifidobacterium bifidum), lactobacillus acidophilus (Lactobacillus acidophilus), Lactobacillus casei (Lactobacillus casei), lactobacillus lactis (Lactobacillus lactis), Lactobacillus plantarum (Lactobacillus plantarum), Pediococcus acidilactici (Pedicoccusacidilacticii), the bacterium mud of one or more in Pediococcus pentosaceus (Pediococcus pentosaceus) or bacillus bulgaricus (Lactobacillusbulgaria), parts by weight are 1 ~ 5 part,
The mode that these bacterial classifications above-mentioned all can be bought by business obtains.
Described protection formula comprises the component of following parts by weight: the food grade glycerin of 0.1 ~ 2 part, the sucrose of 0.4 ~ 2 part, the maltodextrin of 0.3 ~ 5 part, the plant extracts of the whey powder of 0.1 ~ 3 part and 0.06 ~ 1.5 part;
Described basic recipe comprises the component of following parts by weight: the prebiotic factor of 3 ~ 10 parts, the adhesive of 3 ~ 10 parts, the coating agent of 20.5 ~ 55 parts, the starch of 34 ~ 67 parts.
Further, plant extracts of the present invention is preferably one or more in eucommia leaf extract, dried orange peel extracts or licorice.
The described prebiotic factor is one or both in xylo-oligosaccharide or FOS.
Described adhesive is one or both in sodium carboxymethylcellulose or microcrystalline cellulose.
Described coating agent is one or more in polyacrylic resinⅡ, sodium alginate or calcium chloride; Preferably, described coating agent is polyacrylic resinⅡ or sodium alginate and two kinds, calcium chloride combines or three kinds, polyacrylic resinⅡ, sodium alginate and calcium chloride combines.
Described starch is one or more in cornstarch, wheat kind of starch or farina.
For solving second technical problem, the present invention by the following technical solutions:
A preparation method for feeding high-activity lactic acid bacteria solid formulation, the method comprises the following steps:
(1) with one or more in VREF, enterococcus faecalis, lactoenterococcus, bifidobacterium bifidum, lactobacillus acidophilus, Lactobacillus casei, lactobacillus lactis, Lactobacillus plantarum, Pediococcus acidilactici, Pediococcus pentosaceus or bacillus bulgaricus for bacterial classification ferments, then zymotic fluid is centrifugal, obtained bacterium mud;
(2) by parts by weight be 1 ~ 5 part above-mentioned bacterium mud with protection fill a prescription in various components mix, then water is added, dispersion stirs evenly, obtained bacterium liquid, wherein, described protection formula comprises the component of following parts by weight: the food grade glycerin of 0.1 ~ 2 part, the sucrose of 0.4 ~ 2 part, the maltodextrin of 0.3 ~ 5 part, the plant extracts of the whey powder of 0.1 ~ 3 part and 0.06 ~ 1.5 part; In this step, the object added water is to disperse bacterium mud, conveniently operating, as long as amount of water guarantees to make bacterium mud disperse, the weight ratio of water described in ordinary circumstance and bacterium mud is (5-7): 1;
(3) by the various component mix and blends in bacterium liquid obtained for step (2) and basic recipe, and add water in whipping process, then carry out wet granulation, drying, obtain finished product, wherein, described amount of water is 40% ~ 50% of basic recipe component weight; Described basic recipe comprises the component of following parts by weight: the prebiotic factor of 3 ~ 10 parts, the adhesive of 3 ~ 10 parts, the coating agent of 20.5 ~ 55 parts, the starch of 34 ~ 67 parts.
Further, described plant extracts is one or more in eucommia leaf extract, dried orange peel extracts or licorice, concrete preparation method is: 1 ~ 10 part of folium cortex eucommiae, dried orange peel or Radix Glycyrrhizae are dried to moisture≤10% under 40 DEG C ~ 55 DEG C conditions, be crushed to 60 ~ 80 orders, absolute ethyl alcohol extraction 1 ~ 3h, filter, rotary evaporation is extracted thing, with adding 10 ~ 200 parts of water, to obtain final product.
Further, the present invention is when carrying out strain fermentation, the culture medium being suitable for growth well known in the prior art can be used, such as, MRS culture medium, LB culture medium etc., the present invention there is provided herein a kind of formula of preferred culture medium: sucrose 1 ~ 4% (W/V), corn steep liquor 1 ~ 3% (W/V), yeast extract 0.4 ~ 2% (W/V), magnesium chloride 0.06 ~ 0.1% (W/V), manganese sulfate 0.04 ~ 0.2% (W/V), calcium carbonate 0.1 ~ 0.4% (W/V), sodium chloride 0.2 ~ 0.6% (W/V), water 100ml, pH7.0 ± 0.2.
" W/V " used in the present invention refers to w/v, i.e. g/ml or kg/L.
Further, the prebiotic factor described in said method is one or both in xylo-oligosaccharide or FOS; Described adhesive is one or both in sodium carboxymethylcellulose or microcrystalline cellulose; Described coating agent is one or more in polyacrylic resinⅡ, sodium alginate or calcium chloride, preferably, described coating agent is polyacrylic resinⅡ a kind of component or sodium alginate and two kinds, calcium chloride combines or three kinds, polyacrylic resinⅡ, sodium alginate and calcium chloride combines.Described starch is one or more in cornstarch, wheat kind of starch or farina.
Further, in step (3), described drying is cyclone drying under 20 DEG C ~ 45 DEG C conditions.
Further, the present invention is when carrying out strain fermentation, and fermentation temperature is: 30 DEG C ~ 39 DEG C.Can also control the amount of oxygen in sweat, generally cultivate at aerobic or micro-aerobic condition bottom fermentation, when selecting the fermentation of micro-aerobic condition, the amount of oxygen is 0 ~ 0.5mg/L dissolved oxygen.
Beneficial effect of the present invention is:
1. feeding high-activity lactic acid bacteria solid formulation morphosis of the present invention is stablized, available protecting viable bacteria in production, processing, storage, use procedure.Compared with only adding the lactobacillus preparation of basic recipe, said preparation after granulation, dry after Viable detection improve; Shelf life extension, store 6 months under normal temperature, effective Viable detection is more than 80%; Bacterium survival rate in simulated intestinal fluid all reaches more than 87%; Hot and humid tolerance improves 50%.
2. animal applications Be very effective improves: feeding high-activity lactic acid bacteria solid formulation of the present invention improves meat chicken production performance, significantly improves daily gain, reduces feedstuff-meat ratio; Under 10-100g/T addition, compared with basal diet group, said preparation reduces feedstuff-meat ratio more than 9.5%; Improve broiler health performance, intestinal villus is obviously elongated, eurythmy, marshalling, and evenly, in unit are, intestinal villus quantity is many for height; Antioxygenic property strengthens, and TAC and total number born level are improved (P < 0.05), and in enteron aisle, Bifidobacterium and lactobacillus content significantly improve (P < 0.05).
In addition, can also weaned piglets be improved, improve daily gain, reduce feed-weight ratio; Under 30-100g/T addition, and only add compared with basic recipe group, said preparation improves daily gain more than 1.37%; Improve piglet health performance, significantly reduce grice diarrhoea rate (P < 0.05) more than 18%, can Some substitute zinc oxide and antibiotic.
Detailed description of the invention
Below in conjunction with specific embodiment, the present invention will be further described, but be not limit the present invention.Experimental technique in following embodiment, if no special instructions, is conventional method.Test material used in following embodiment, if no special instructions, is the purchase of routine biochemistry reagent suppliers and obtains.
Embodiment 1
(1) preparation of plant extracts
Respectively 1 ~ 10 part of folium cortex eucommiae, dried orange peel and Radix Glycyrrhizae are dried to moisture≤10% under 40 DEG C ~ 55 DEG C conditions, are crushed to 60 ~ 80 orders, absolute ethyl alcohol 10 ~ 50 parts, extraction 1 ~ 3h, filter, rotary evaporation is extracted thing (density 0.7 ~ 1.5g/cm
3), with adding 10 ~ 200 parts of water, to obtain final product.
(2) MRS culture medium and preferred culture medium is prepared
The preparation of MRS culture medium: get peptone 1% (W/V), glucose 2% (W/V), beef extract 1% (W/V), anhydrous sodium acetate 0.5% (W/V), yeast extract 0.5% (W/V), Tween 80 0.1ml, dipotassium hydrogen phosphate 0.2% (W/V), diammonium hydrogen citrate 0.2% (W/V), bitter salt 0.058% (W/V), Manganous sulfate monohydrate 0.025% (W/V), water 100ml, regulates pH to 7.0 ± 0.2; Adopt moist heat sterilization, temperature is 115 DEG C, sterilization time 30min.
The preparation of preferred culture medium: sucrose 1 ~ 4% (W/V), corn steep liquor 1 ~ 3% (W/V), yeast extract 0.4 ~ 2% (W/V), magnesium chloride 0.06 ~ 0.1% (W/V), manganese sulfate 0.04 ~ 0.2% (W/V), calcium carbonate 0.1 ~ 0.4% (W/V), sodium chloride 0.2 ~ 0.6% (W/V), water 100ml, pH7.0 ± 0.2; Adopt moist heat sterilization, temperature is 115 DEG C, sterilization time 30min.
(3) bacterium is clay standby
A. choose VREF (CGMCC2516), lactobacillus acidophilus (ATCC4356), respectively by bacterial classification under 35 DEG C of micro-aerobic conditions (i.e. 0.5mg/L dissolved oxygen), cultivate 8h in MRS culture medium, obtain VREF and lactobacillus acidophilus primary seed solution;
B. respectively the primary seed solution of two kinds of bacterium being cultivated by expanding in 2% inoculum concentration access seeding tank, under 35 DEG C of micro-aerobic conditions, in MRS culture medium, cultivating 6h, obtaining the secondary seed solution of two kinds of bacterium;
C. respectively the secondary seed solution of two kinds of bacterium is carried out fermented and cultured by 5% inoculum concentration access fermentation tank, use preferred culture medium to carry out fermented and cultured, under 38 DEG C of micro-aerobic conditions, cultivate 24h; By centrifugal for the zymotic fluid tube centrifuge 10000rpm bacterium mud obtaining two kinds of bacterium, according to arbitrary ratio, two kinds of bacterium mud can be mixed as required, obtained Mixed Microbes mud, such as, the mass ratio of VREF bacterium mud and lactobacillus acidophilus bacterium mud is 1:1,1:2,1:3,2:1, or 3:1 etc.;
(4) formula of solid formulation:
Mixed Microbes mud 50g;
Protection formula: 5g food grade glycerin, 20g sucrose, 30g maltodextrin, 30g whey powder, 13g eucommia leaf extract, 1g dried orange peel extracts;
Basic recipe: 300g xylo-oligosaccharide, 20g sodium carboxymethylcellulose, 450g microcrystalline cellulose, 8g polyacrylic resinⅡ, 550g sodium alginate, 600g calcium chloride, 1700g cornstarch.
(5) formulation samples preparation
Portion adds sterilized water and dilutes 20 times (sample one) for subsequent use, and portion only adds basic recipe component and granulates (sample two), and portion adds basic recipe and protection formula granulates (sample three).
A. only add basic recipe component to granulate
Get the various component mix and blends in above-mentioned bacterium mud and basic recipe, and add water in whipping process (described amount of water is 40% ~ 50% of basic recipe component weight), then wet granulation, throwing circle, cyclone drying under 20 DEG C ~ 45 DEG C conditions, obtains finished product.
B. basic recipe and the granulation of protection formula is added
Get above-mentioned bacterium mud fill a prescription with protection immediately in various component mix, then add water, dispersion stirs evenly, standing 15-60min, obtains bacterium liquid for subsequent use;
Get the various component mix and blends in bacterium liquid for subsequent use obtained above and basic recipe, and add water in whipping process (described amount of water is 40% ~ 50% of basic recipe component weight), then carry out wet granulation, throw circle, cyclone drying under 20 DEG C ~ 45 DEG C conditions, obtains finished product.Granulation, dry rear viable count are 1.0 × 10
10more than CFU/g.
(6) Performance Detection of lactobacillus preparation prepared by the present embodiment
A. the middle product of processing (after granulating, after drying) of the sample two in (5) and sample three are sampled, detect sample viable count and survival rate, in table 1; B. 3 samples in (5) are placed in simulated intestinal fluid (Chinese Pharmacopoeia formula) 2h, detect sample viable count and survival rate, in table 2; C. the sample in (5) is placed in 85 DEG C, 60% humidity environment 2min, detects sample viable count and survival rate, in table 3; D. by 3 sample sealing preservations in (5), 20 DEG C of normal temperature storage, sample viable count and survival rate is detected, in table 4.
Table 1
Note: Jun Shuo unit: CFU value/g.Lower same.
Table 2
Table 3
| Sample | 0h original value | 2min | Survival rate % after 2min |
| Sample one | 1.21×10 10 | 7.1×10 6 | -- |
| Sample two | 1.17×10 10 | 1.15×10 9 | 9.8 |
| Sample three | 1.01×10 10 | 6.16×10 9 | 61.01 |
Table 4
Can draw from above: in the middle of a. production process, product contrast shows: after granulating, after drying, improve 6.05% and 20.01% with the lactobacillus preparation Viable detection that " two-step method " is made respectively than the preparation only adding basic recipe; B. simulated intestinal fluid tolerance test shows: to utilize in " two-step method " lactobacillus preparation of making that effectively Viable detection is significantly higher than the bacterium liquid not adding the protection of any auxiliary material respectively, the preparation only adding basic recipe is 16.42% and 8.69%; C. hot and humid tolerance test shows: the preparation utilizing in " two-step method " lactobacillus preparation of making effectively Viable detection to be significantly higher than only to add basic recipe is more than 50%; D.6 store experiment the moon to show: in the lactobacillus preparation utilizing " two-step method " to make, effective Viable detection is more than 80%, far above not adding the bacterium liquid of any auxiliary material protection, only adding the preparation of basic recipe.
Embodiment 2
(1) preparation of LB culture medium and preferred culture medium is prepared
The preparation of LB culture medium: get tryptone 1% (W/V), yeast extract 0.5% (W/V), sodium chloride 1% (W/V), water 100ml, pH7.0 ± 0.2; Adopt moist heat sterilization, 121 DEG C, 20min.
The preparation of preferred culture medium: sucrose 1 ~ 4% (W/V), corn steep liquor 1 ~ 3% (W/V), yeast extract 0.4 ~ 2% (W/V), magnesium chloride 0.06 ~ 0.1% (W/V), manganese sulfate 0.04 ~ 0.2% (W/V), calcium carbonate 0.1 ~ 0.4% (W/V), sodium chloride 0.2 ~ 0.6% (W/V), water 100ml, pH7.0 ± 0.2; Adopt moist heat sterilization, temperature is 115 DEG C, sterilization time 30min.
(2) bacterium is clay standby
A. choose enterococcus faecalis (ATCC19433), by bacterial classification under 35 DEG C of conditions, cultivate 8h in LB culture medium, obtain primary seed solution;
B. primary seed solution being cultivated by expanding in 2% inoculum concentration access seeding tank, under 37 DEG C of micro-aerobic conditions, cultivating 6h in LB culture medium, obtaining secondary seed solution;
C. secondary seed solution is carried out fermented and cultured by 5% inoculum concentration access fermentation tank, use preferred culture medium to carry out fermented and cultured, under 38 DEG C of micro-aerobic conditions, cultivate 24h; By centrifugal for zymotic fluid desk centrifuge 6000rpm obtained bacterium mud.
(3) formula of solid formulation
Bacterium mud 100g;
Protection formula is: 25g food grade glycerin, 45g sucrose, 30g maltodextrin, 28g whey powder, 8g eucommia leaf extract, 1.5g dried orange peel extracts, 4.5g licorice; The preparation method of described plant extracts is with embodiment 1;
Basic recipe is: 325g xylo-oligosaccharide, 1.0g sodium carboxymethylcellulose, 334g microcrystalline cellulose, 18g polyacrylic resinⅡ, 1050g sodium alginate, 1200g sodium chloride, 6700g cornstarch;
(4) formulation samples preparation
Portion only adds the granulation of basic recipe component, and portion adds basic recipe, and (after granulation, drying, viable count is 1.0 × 10 with the granulation of protection formula
10more than CFU/g).Concrete preparation method is with embodiment 1.
(5) Performance Detection of the lactobacillus preparation prepared by the present embodiment: to the action effect of broiler growth and health.
Animal used as test: 120 public broiler chicken of AA;
Feeding and management: whole feeding period amounts to 42d, be divided into two stage: 1-21d for early stage, 22-42d is the later stage.Employing is raised in cages (each cage is exactly a repetition), and whole experimental period carries out immunity, sterilization routinely, whole day 24h illumination, full phase free choice feeding and drinking-water, and temperature automatically controlled control is wet.Before brood time, 3d room temperature controls at 33 DEG C, declines weekly 3 DEG C until 24 DEG C of constant temperature later; Humidity 1-21 day remains on 65-70%, and 22-42 keeps 60-65%; 1-42d continues 24h illumination and ventilation.
Packet design:
120 Broiler chicks are divided into 4 groups at random, often organize 6 repetitions, often repeat 5.Raise in Pilot Base SPF room, conputer controlled humiture.
A group: the basal diet of the not added with antibiotic of feeding+only add basic recipe lactobacillus preparation
B group: the lactobacillus preparation 1.00 × 10 that basal diet+" two-step method " of the not added with antibiotic of feeding is protected
10cFU/kg feed.
Experimental result is as table 5-6.
The lactobacillus preparation that table 5 " two-step method " is made is on the impact of broiler growth performance
The different lowercase alphabet of colleague's shoulder mark shows significant difference (P<0.05), identical lowercase or without letter representation difference not significantly (P>0.05).Following table is same.
Table 6 TAC (T-AOC)
| A group | B group | |
| 21 ages in days | 11.11±0.84 a | 15.54±1.19 c |
| 42 ages in days | 9.49±0.93 bc | 10.89±1.28 c |
The lactobacillus preparation that table 7 " two-step method " is made is on the impact of Broiler chicks Morphological Indices of Duodenum Villus structure
The lactobacillus preparation that table 8 " two-step method " is made is on the impact of Broiler chicks gut flora
Table 5-table 8 shows, compared with only adding the group of basic recipe preparation, the lactobacillus preparation group broiler chicken feedstuff-meat ratio that " two-step method " is made significantly reduces by 9.66%; TAC significantly improves 14.75%-39.87%; Unit are intestinal villus quantity is many, and intestines suede height, Crypt depth all have significant change; In ileum, Bifidobacterium, lactobacillus quantity significantly increase.
Embodiment 3
(1) culture medium is prepared
Preparation LB culture medium: get tryptone 1% (W/V), yeast extract 0.5% (W/V), sodium chloride 1% (W/V), water 100ml, pH7.0 ± 0.2; Adopt moist heat sterilization, 121 DEG C, 20min.
Preparation preferred culture medium: sucrose 1 ~ 4% (W/V), corn steep liquor 1 ~ 3% (W/V), yeast extract 0.4 ~ 2% (W/V), magnesium chloride 0.06 ~ 0.1% (W/V), manganese sulfate 0.04 ~ 0.2% (W/V), calcium carbonate 0.1 ~ 0.4% (W/V), sodium chloride 0.2 ~ 0.6% (W/V), water 100ml, pH7.0 ± 0.2; Sterilizing methods: moist heat sterilization, 115 DEG C, 30min.
(2) bacterium is clay standby
A. lactobacillus lactis (ATCC12315), bacillus bulgaricus (ATCC11842) and lactoenterococcus (ATCC19435) is chosen, respectively by bacterial classification under 35 DEG C of micro-aerobic conditions (i.e. 0.3mg/L dissolved oxygen), cultivate 8h in LB culture medium, obtain the primary seed solution of three kinds of bacterium;
B. respectively the primary seed solution of three kinds of bacterium being cultivated by expanding in 2% inoculum concentration access seeding tank, under 37 DEG C of micro-aerobic conditions, in LB culture medium, cultivating 6h, obtaining the secondary seed solution of three kinds of bacterium;
C. respectively the secondary seed solution of three kinds of bacterium is carried out fermented and cultured by 5% inoculum concentration access fermentation tank, use preferred culture medium to carry out fermented and cultured, under 38 DEG C of micro-aerobic conditions, cultivate 24h; Bacterium mud is obtained by centrifugal for zymotic fluid desk centrifuge 6000rpm.Can be mixed by three kinds of bacterium mud according to arbitrary ratio as required, obtained Mixed Microbes mud, such as, the mass ratio of lactobacillus lactis bacterium mud, bacillus bulgaricus bacterium mud and lactoenterococcus bacterium mud is 1:1:1,1:2:1,1:1:3,2:1:1 etc.;
(3) formula of solid formulation
Mixed Microbes mud 50g
Protection formula: 10g food grade glycerin, 23g sucrose, 33g maltodextrin, 26g whey powder, 8g eucommia leaf extract, 1g dried orange peel extracts, 4g licorice; The preparation method of plant extracts is see embodiment 1;
Basic recipe: 450g xylo-oligosaccharide, 18g sodium carboxymethylcellulose, 320g microcrystalline cellulose, 10g polyacrylic resinⅡ, 1250g sodium alginate, 1550g calcium chloride, 3250g cornstarch.
(4) formulation samples preparation
Portion only adds basic recipe component and granulates (sample one), and portion adds basic recipe and protection formula granulates (sample two) concrete preparation method with embodiment 1.
(5) Performance Detection of lactobacillus preparation prepared of the present embodiment
Experiment purpose: contrast only adds lactobacillus preparation that basic recipe and " two-step method " make to grice diarrhoea, daily gain.
Experimental animal: weanling pig 102;
Duration of trial: test is from weaning beginning, 35 days experimental periods;
Packet design:
Weanling pig is divided at random 2 groups (one group, sample and two groups, samples), often organizes 3 repetitions, often repeat 17.
Diarrhea rate=diarrhoea piglet head/total head of test piglet
Diarrhoea piglet head=diarrhoea piglet head time (the 35th age in days)+diarrhoea piglet head time (the 36th age in days)+... + diarrhoea piglet head time (the 63rd age in days); The total head of piglet=piglet head number (the 35th age in days)+piglet head number (the 36th age in days)+... + piglet head number (the 63rd age in days).
Scours index: Scours index reflects the order of severity of diarrhoea, this index is higher, and representative diarrhoea is more serious.
Scours index=stools scored sum/total head number of confession examination pig.Standards of grading see the following form.
Diarrhoea status standards of grading
| Diarrhoea degree | Ight soil outward appearance | Scoring |
| Normally | Bar shaped or granular | 0 |
| Slightly | Soft fecal energy is shaped | 1 |
| Moderate | Thick shape, shapeless, liquid dung is without segregation phenomenon | 2 |
| Seriously | Aqueous, shapeless, liquid dung has segregation phenomenon, and mucus just or just dense | 3 |
| Pollute | The anus of pig and surrounding thereof have excrement thing to hang sample | 4 |
| Red and swollen | The anus of pig is red and swollen | 5 |
To the testing result of two groups of samples as shown in table 9, table 10:
Table 9 piglet daily gain compares
| Process | Average daily gain, g/ days |
| One group, sample | 464.13±30.1 |
| Two groups, sample | 470.5±15.3 |
Table 10 grice diarrhoea rate compares
| Process | Diarrhea rate (%) | Scours index (%) | Death rate (%) |
| One group, sample | 2.48 b | 0.865 b | 1.6 |
| Two groups, sample | 2.01 a | 0.801 a | 0 |
As from the foregoing, compared with only adding the preparation of basic recipe, the piglet daily gain of the lactobacillus preparation group utilizing " two-step method " to make improves 1.37%; Diarrhea rate significantly reduces (P < 0.05), and compared with control group, diarrhea rate declines 18.95%.
In addition, the ruddy degree of test group piglet skin and hair color brightness, in disorder degree are greatly improved.
Claims (7)
1. a feeding high-activity lactic acid bacteria solid formulation, is characterized in that, comprises probiotic lactobacillus bacterium mud, protection formula and basic recipe; Wherein,
Described probiotic lactobacillus bacterium mud is the bacterium mud of one or more in VREF, enterococcus faecalis, lactoenterococcus, bifidobacterium bifidum, lactobacillus acidophilus, Lactobacillus casei, lactobacillus lactis, Lactobacillus plantarum, Pediococcus acidilactici, Pediococcus pentosaceus or bacillus bulgaricus, and parts by weight are 1 ~ 5 part;
Described protection formula comprises the component of following parts by weight: the food grade glycerin of 0.1 ~ 2 part, the sucrose of 0.4 ~ 2 part, the maltodextrin of 0.3 ~ 5 part, the plant extracts of the whey powder of 0.1 ~ 3 part and 0.06 ~ 1.5 part; Described plant extracts is eucommia leaf extract and dried orange peel extracts, or eucommia leaf extract, dried orange peel extracts and licorice, wherein, described eucommia leaf extract, dried orange peel extracts or licorice are: 1 ~ 10 part of folium cortex eucommiae, dried orange peel or Radix Glycyrrhizae are dried to moisture≤10% under 40 DEG C ~ 55 DEG C conditions, are crushed to 60 ~ 80 orders, absolute ethyl alcohol extraction 1 ~ 3h, filter, rotary evaporation is extracted thing, adds 10 ~ 200 parts of water, to obtain final product;
Described basic recipe comprises the component of following parts by weight: the prebiotic factor of 3 ~ 10 parts, the adhesive of 3 ~ 10 parts, the coating agent of 20.5 ~ 55 parts, the starch of 34 ~ 67 parts;
Described feeding high-activity lactic acid bacteria solid formulation is obtained by following method:
(1) with one or more in VREF, enterococcus faecalis, lactoenterococcus, bifidobacterium bifidum, lactobacillus acidophilus, Lactobacillus casei, lactobacillus lactis, Lactobacillus plantarum, Pediococcus acidilactici, Pediococcus pentosaceus or bacillus bulgaricus for bacterial classification ferments, then zymotic fluid is centrifugal, obtained bacterium mud;
(2) by parts by weight be 1 ~ 5 part above-mentioned bacterium mud with protection fill a prescription in various components mix, then water is added, dispersion stirs evenly, obtained bacterium liquid, wherein, described protection formula comprises the component of following parts by weight: the food grade glycerin of 0.1 ~ 2 part, the sucrose of 0.4 ~ 2 part, the maltodextrin of 0.3 ~ 5 part, the plant extracts of the whey powder of 0.1 ~ 3 part and 0.06 ~ 1.5 part;
(3) by the various component mix and blends in bacterium liquid obtained for step (2) and basic recipe, and add water in whipping process, then carry out wet granulation, drying, obtain finished product, wherein, amount of water is 40% ~ 50% of basic recipe component weight; Described basic recipe comprises the component of following parts by weight: the prebiotic factor of 3 ~ 10 parts, the adhesive of 3 ~ 10 parts, the coating agent of 20.5 ~ 55 parts, the starch of 34 ~ 67 parts.
2. feeding high-activity lactic acid bacteria solid formulation according to claim 1, is characterized in that, the described prebiotic factor is one or both in xylo-oligosaccharide or FOS; Described adhesive is one or both in sodium carboxymethylcellulose or microcrystalline cellulose; Described coating agent is one or more in polyacrylic resinⅡ, sodium alginate or calcium chloride; Described starch is one or more in cornstarch, wheat kind of starch or farina.
3. a preparation method for feeding high-activity lactic acid bacteria solid formulation, it is characterized in that, the method comprises the following steps:
(1) with one or more in VREF, enterococcus faecalis, lactoenterococcus, bifidobacterium bifidum, lactobacillus acidophilus, Lactobacillus casei, lactobacillus lactis, Lactobacillus plantarum, Pediococcus acidilactici, Pediococcus pentosaceus or bacillus bulgaricus for bacterial classification ferments, then zymotic fluid is centrifugal, obtained bacterium mud;
(2) by parts by weight be 1 ~ 5 part above-mentioned bacterium mud with protection fill a prescription in various components mix, then water is added, dispersion stirs evenly, obtained bacterium liquid, wherein, described protection formula comprises the component of following parts by weight: the food grade glycerin of 0.1 ~ 2 part, the sucrose of 0.4 ~ 2 part, the maltodextrin of 0.3 ~ 5 part, the plant extracts of the whey powder of 0.1 ~ 3 part and 0.06 ~ 1.5 part; Described plant extracts is eucommia leaf extract and dried orange peel extracts, or eucommia leaf extract, dried orange peel extracts and licorice, and the preparation method of described eucommia leaf extract, dried orange peel extracts or licorice is: 1 ~ 10 part of folium cortex eucommiae, dried orange peel or Radix Glycyrrhizae are dried to moisture≤10% under 40 DEG C ~ 55 DEG C conditions, be crushed to 60 ~ 80 orders, absolute ethyl alcohol extraction 1 ~ 3h, filter, rotary evaporation is extracted thing, add 10 ~ 200 parts of water, to obtain final product;
(3) by the various component mix and blends in bacterium liquid obtained for step (2) and basic recipe, and add water in whipping process, then carry out wet granulation, drying, obtain finished product, wherein, amount of water is 40% ~ 50% of basic recipe component weight; Described basic recipe comprises the component of following parts by weight: the prebiotic factor of 3 ~ 10 parts, the adhesive of 3 ~ 10 parts, the coating agent of 20.5 ~ 55 parts, the starch of 34 ~ 67 parts.
4. preparation method according to claim 3, it is characterized in that, the culture medium prescription used during fermentation is: sucrose 1 ~ 4% (W/V), corn steep liquor 1 ~ 3% (W/V), yeast extract 0.4 ~ 2% (W/V), magnesium chloride 0.06 ~ 0.1% (W/V), manganese sulfate 0.04 ~ 0.2% (W/V), calcium carbonate 0.1 ~ 0.4% (W/V), sodium chloride 0.2 ~ 0.6% (W/V), water 100ml, pH7.0 ± 0.2.
5. preparation method according to claim 3, is characterized in that, the described prebiotic factor is one or both in xylo-oligosaccharide or FOS; Described adhesive is one or both in sodium carboxymethylcellulose or microcrystalline cellulose; Described coating agent is one or more in polyacrylic resinⅡ, sodium alginate or calcium chloride; Described starch is one or more in cornstarch, wheat kind of starch or farina.
6. preparation method according to claim 3, is characterized in that, in step (3), described drying is cyclone drying under 20 DEG C ~ 45 DEG C conditions.
7. preparation method according to claim 3, is characterized in that, in step (1), temperature during fermentation is: 30 DEG C ~ 39 DEG C.
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Families Citing this family (21)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| AU2015259359A1 (en) * | 2014-05-12 | 2016-11-17 | BiOWiSH Technologies, Inc. | Compositions and methods for improving human health and nutrition |
| CN104431560B (en) * | 2014-12-15 | 2017-08-08 | 江苏傲农生物科技有限公司 | A kind of pig feed additive with enteron aisle health care and raising luring function |
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| CN112262917A (en) * | 2020-10-26 | 2021-01-26 | 合肥五粮泰生物科技有限公司 | Preparation method of high-viable bacteria fermented feed |
| CN114304688B (en) * | 2021-11-25 | 2023-08-01 | 中粮集团有限公司 | Zein coating agent, probiotics preparation coated by coating agent and preparation method thereof |
| CN117256730B (en) * | 2023-11-23 | 2024-04-02 | 山东广元药业科技有限公司 | Probiotic-containing feed additive for conditioning animal intestinal tracts and preparation method thereof |
Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1843385A (en) * | 2006-02-17 | 2006-10-11 | 哈尔滨美华生物技术股份有限公司 | Enteral microecological formulation and its preparation process |
| CN101297820A (en) * | 2007-04-30 | 2008-11-05 | 大连森佰澳科技有限公司 | Streptococcus faecium function signal molecule formulation and product thereof for reducing fat and slimming |
| CN101496822A (en) * | 2009-02-18 | 2009-08-05 | 上海谱莱生物技术有限公司 | Composite probiotics micro-ecological formulation and preparation method |
| CN101696406A (en) * | 2009-10-19 | 2010-04-21 | 李军训 | Double-layer embedment technique of semidry method and probiotics orifice-powder bed method |
| CN102018216A (en) * | 2009-09-19 | 2011-04-20 | 菲伯纳生物医药有限责任公司 | Composition containing bifidobacteria for adjusting intestinal flora and enhancing immunity |
| CN102599345A (en) * | 2012-01-13 | 2012-07-25 | 山东华牧天元动物保健品有限公司 | Multicomponent biological compound agent for livestock |
Family Cites Families (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101461453B (en) * | 2009-01-15 | 2012-01-04 | 润盈生物工程（上海）有限公司 | Composite microbial feed additive for beasts and birds |
| CN102021162A (en) * | 2009-09-19 | 2011-04-20 | 菲伯纳生物医药有限责任公司 | High-activity bifidobacterium powder and preparation method thereof |
| CN101773207B (en) * | 2010-02-08 | 2012-12-12 | 鲁东梁 | Hybrid probiotic desiccate and preparation method thereof |
-
2013
- 2013-08-06 CN CN201310339372.2A patent/CN103392911B/en active Active
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1843385A (en) * | 2006-02-17 | 2006-10-11 | 哈尔滨美华生物技术股份有限公司 | Enteral microecological formulation and its preparation process |
| CN101297820A (en) * | 2007-04-30 | 2008-11-05 | 大连森佰澳科技有限公司 | Streptococcus faecium function signal molecule formulation and product thereof for reducing fat and slimming |
| CN101496822A (en) * | 2009-02-18 | 2009-08-05 | 上海谱莱生物技术有限公司 | Composite probiotics micro-ecological formulation and preparation method |
| CN102018216A (en) * | 2009-09-19 | 2011-04-20 | 菲伯纳生物医药有限责任公司 | Composition containing bifidobacteria for adjusting intestinal flora and enhancing immunity |
| CN101696406A (en) * | 2009-10-19 | 2010-04-21 | 李军训 | Double-layer embedment technique of semidry method and probiotics orifice-powder bed method |
| CN102599345A (en) * | 2012-01-13 | 2012-07-25 | 山东华牧天元动物保健品有限公司 | Multicomponent biological compound agent for livestock |
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