CN104611248A - Composite biological living bacteria preparation and preparation technology thereof - Google Patents
Composite biological living bacteria preparation and preparation technology thereof Download PDFInfo
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Abstract
The invention relates to a composite biological living bacteria preparation and a preparation method thereof. The total number of effective viable bacteria is not less than 108cfu/ml. The method includes: selecting optimal probiotics, carrying out supporting medium activation, performing enlarged culture and mixing, conducting culture by means of a mixed bacteria medium and a fermentation tank, carrying out filtration, concentration and proportioning so as to form the composite living bacteria preparation. The preparation contains the following bacteria with the total number of effective viable bacteria of: 22-25% of photosynthetic bacteria, 21-24% of yeast, 15-20% of bacillus subtilis, 15-20% of Pediococcus acidilactici, 12-16% of Streptococcus faecalis, and 12-16% of Lactobacillus acidophilus. The composite biological living bacteria preparation provided by the invention can be applied to agricultural cultivation, livestock and poultry breeding, aquaculture, feed adding, municipal sewage and domestic garbage treatment and other fields.
Description
Technical field
The invention belongs to application and the manufacture craft thereof of microbial strains, specifically a kind of NEW TYPE OF COMPOSITE biologic live bacteria preparation and preparation technology thereof that can be applicable to the fields such as agricultural planting, aquaculture, livestock and poultry cultivation, feed manufacturing, environment protection, sewage disposal.
Background technology
The end of the forties in last century, scholars found that tsiklomitsin has promoter action to growth of animals or poultry first time, thus had started the epoch of microbiotic as fodder additives.In these epoch, people widely use the microbiotic of sub-therapeutic dose and other pharmaceutical chemicalss are used as fodder additives, promote the growth of animal or prevent the generation of pathogenic bacterial infection, disease, indelible contribution has been made to the development of fodder industry and aquaculture.But along with the dark people of scientific knowledge, people, while acquisition economic benefit, have had more deep understanding to the side effect that these microbiotic, chemicals bring.As abuse of antibiotics causes animal endogenous or superinfection, produce the strain of resistance mattress, animal cell immunity, humoral immune function decline and residual etc. in animal product of medicine, for these reasons, a lot of country and raiser oppose that microbiotic is as growth stimulant, are even forbidden more and more many types of drugs by legislation.
After China joined WTO, highly beneficial to the macroeconomic development of China, but also there is the problem that can not be ignored, many outlet agricultural byproducts, as the food such as meat, fowl, egg, milk, fishery products are kept outside of the door due to problems such as pesticide residue, decrease the international market share of China, in the face of world market with keen competition, the vital task of pendulum in face of us reduces pesticide residue to greatest extent, produce green agricultural byproducts that are pollution-free, noresidue, make it to occupy a tiny space in world market.
Under this background, people one after another sight invest having multi-functions nontoxic, have no side effect, the active bacteria formulation of noresidue.For milk-acid bacteria, record the universally recognized several milk-acid bacteria of display according to pertinent data, physiological function shows as:
Lactobacterium acidophilum LA: have immuno-potentiation, is attached on animal intestinal epithelial cell, balance intestinal flora.
Lactobacterium casei Shirota: control intestinal disorder balance intestinal bacterium, reduces ight soil toxic enzymic activity.
Roy is for Bacterium lacticum: be colonizated in enteron aisle, fine to the prevention effect of Animal diseases, also has good result, can show more comprehensive more outstanding if multiple beneficial bacterium to be combined into its effect of preparation to people.Therefore, carry out that compound bio viable bacteria technical study effect is great, of far-reaching significance, space is wide.
Further research finds, has influence each other restraining effect and the active effect mutually excited between bacterial classification.Through suitable collocation can improve greatly single exist time effect and efficiency, thus range of application and using value are significantly improved.The marketable value of composite bacteria is improve agricultural quality, produces green food and promote the thinking that agricultural byproducts international status provides enlightening meaning, is also that we carry out the important evidence of this technical study.
Summary of the invention
The present invention is for the purpose of compound bio active bacteria formulation product, with multiple batches of experiment, be detected as basis, in conventional, common prebiotic bacterial classification, filter out economic and practical kind, the combination of ratio, can economical, to produce by the Secondary Fermentation of efficiency induction compound bio active bacteria formulation be more applicable for agricultural planting, livestock and poultry cultivation, aquaculture, feed add and numerous products in the fields such as municipal effluent domestic rubbish disposal.Comprehensively to solve the problems such as chemistry, hormone residues, urban environment living garbage pollution.Be of value to circular agriculture, as pollution-free vegetable, grain and oil, meat and egg production, maintaining ecological balance.Produce useful people ' s health, the many-sided effect improved the economic and social benefits.Match by adaptability substratum, NEW TYPE OF COMPOSITE biologic live bacteria preparation second product and production technique thereof can be formed.
The technical solution used in the present invention is:
One, compound bio active bacteria formulation
In reagent of the present invention, effective total viable count is no less than 10
8cfu/ml.Said preparation activates in the test tube that supporting substratum is housed by mixed strains, cultivates, generate in fermentor tank through shaking table, more after filtration, concentrated, proportioning forms active bacteria formulation, in said preparation, the per-cent of the living bacteria count of contained various bacterial classification is:
Photosynthetic bacteria 22 ~ 25%,
Yeast 21 ~ 24%,
Subtilis 15 ~ 20%
Pediococcus acidilactici 15 ~ 20%,
Streptococcus faecium 12 ~ 16%,
Lactobacterium acidophilum 12 ~ 16%.
Two, the preparation technology of compound bio active bacteria formulation
Comprise the following steps in this technique:
A, bacterial classification are chosen and activation:
In test tube, carry out inclined-plane according to above-mentioned selected species, the bacterial strain choosing health respectively and supporting activation medium and activate cultivation,
The determination of B, Shake flask medium and the cultivation of liquid spawn:
Required Shake flask medium is obtained than relation: multivitamin aqueous solution 2 parts, trace element water-soluble liquid 1 part, the activation culture based 1.5 part mixture supporting with bacterial classification shake up and disperses to implant flask by following volume parts, then add on the bacterial classification after activation, shaking table in 28-33 DEG C of sterilisable chamber and cultivate 24-48 hour produce liquid bacterial classification
The proportioning of C, seeding tank substratum and the fermentation of mixed strains are cultivated:
With reference to following component weight ratio configuration seeding tank substratum: 35% sulfuric acid 1g, 7 water magnesium sulfate 2g, sodium-chlor 1g, potassium primary phosphate 0.5g, sodium malate 2g, 2 water calcium chloride 0.5g, yeast leach liquor 0.5g, 7 aqueous ferrous sulfate 0.1g, trace element water-soluble liquid 1ml, multivitamin aqueous solution 1ml, agar 10g, sucrose 5g, peptone 5g, add pure water 1L and form liquid nutrient medium
Implant liquid nutrient medium in seeding tank after, in adjustment composite fluid bacterial classification, the effective total viable count of each bacterial classification reaches 10
6cfu/ml-10
7cfu/ml, and carry out adjustment with reference to the following per-cent composition of each bacterial classification number of viable and become mixed strains:
Photosynthetic bacteria 20 ~ 22%,
Yeast 18 ~ 22%,
Subtilis 17 ~ 20%
Pediococcus acidilactici 18 ~ 21%,
Streptococcus faecium 22 ~ 24%,
Lactobacterium acidophilum 22 ~ 18%,
Then join in liquid nutrient medium according to volume ratio 100:7 ~ 10 and stir, adjustment PH7.0, maintenance temperature 28-30 DEG C, incubation time obtains liquid spawn in 28 days.
The above trace element water-soluble liquid is mixed by following composition:
EDTA-2Na:190 ~ 210mg, 2 water Sodium orthomolybdate: 18 ~ 22mg, 7 aqueous ferrous sulfates: 80 ~ 120mg, 6 water nickel oxide 8 ~ 12mg: boric acid 80 ~ 120mg, 2 water cupric chloride: 80 ~ 120mg, zinc chloride: 80 ~ 120mg, 4 water Manganous chloride tetrahydrate: 80 ~ 120mg, distilled water: 0.8 ~ 1.2L.
Above-described multivitamin aqueous solution is mixed by following composition:
VITMAIN B1: 480 ~ 520g, para-amino benzoic acid: 280 ~ 320g, vitamins B
3: 480 ~ 520mg, vitamins B
1240 ~ 60mg, vitamins B
6: 80 ~ 120mg, vitamins B
7: 40 ~ 60mg, distilled water 0.8 ~ 1.2L.
Three, compound bio viable bacteria pulvis.
Four, after step C completes in above processing step, compound bio viable bacteria liquid agent is obtained.
Again liquid preparation drying is stayed thing to consolidating of moisture < 15%, then through pulverizing, cross 60 mesh sieves, packaging, forms effective total viable count and is no less than 10
7the powderous preparations of cfu/ gram.
Above-described composite bacteria liquid or powder product, directly or by means of Secondary Fermentation will be applied to the fields such as agricultural planting, livestock and poultry cultivation, aquaculture, feed interpolation and municipal effluent domestic rubbish disposal respectively, to solve the problems such as chemistry, hormone residues and city environmental pollution, produce green, pollution-free vegetable, grain and oil, meat and egg production, maintaining ecological balance, useful people ' s health, improves the economic and social benefits.
First it is considered that the selection of probiotics in the present invention's design.First be the production technique environment according to it, the applied environment of Secondary Fermentation product, the technical feature feature of leavened prod set out, the biological characteristics determined from their respective Track characters goes understanding, compares, to find out excellent healthy bacterial classification.Again the every class bacterial classification selected, by the conditional request needed for each bacterial classification, select the substratum at different levels of applicable growth and breeding.Need through initial option, carry out mixed culture experiment to a classifying type enlarged culturing class active bacteria.Select symbiosis feature according in their symbiosis propagation, interdependent required condition, then they are combined carry out compound criteria conditional filtering out.By symbiosis culture based formulas required in the mixed fermentation tank determined after repeatedly culture experiment, repeated screening, could finally implement mixing fermentation culture, generate the composite bacteria reagent wanted required for the present invention.
The beneficial effect that the present invention produces is: directly application or Secondary Fermentation product can be applied to agriculture and animal husbandry aquatic products field and produce the economic benefit of low cost high added value energetically and make full use of biomass waste material and realize Agricultural recycling economy and environmental protect benefit:
1, compound bio active bacteria formulation of the present invention can directly be applied in aquaculture aspect: be administered to water body or add in bait, can make to set up profitable strain in the digestive tube of animal and in water body environment, suppress harmful microbe Growth and reproduction.Normal microflora can be the nutrient substance that breed variety provides a large amount of, strengthens its resistance against diseases, and its thalline effect can also be killed or suppress pathogenic micro-organism, eliminates objectionable impurities, promotes the growth of beneficial algae, for aquatic animal provides good growing environment.
2, compound bio active bacteria formulation of the present invention is applied in livestock and poultry cultivation aspect: adjustable animal microecological balance, prevent and overcome microbial state treatment, improve immunologic function of livestock and birds, strengthen its resistance against diseases, reduce M & M, and then few with or without veterinary drug, improve food conversion ratio, suppress, eliminate colony house obnoxious flavour, promote animal growth and development.
3, compound bio active bacteria formulation of the present invention is applied in plant husbandry aspect: the decomposition that can promote the soil organism, improves fertilizer efficiency; Can improve the physico-chemical property of soil, make it soft, permeable, ventilative, thus promote the metabolism of long-term cropping, the generation of prevention and minimizing disease and pest, improves crop quality.
Embodiment
Each probiotic bacterium in the present invention all selects the bacterial classification preserved by China Committee for Culture Collection of Microorganisms's common micro-organisms center, wherein, photosynthetic bacteria (Inoculant of photosynthetic bacteria) be numbered 1.2352, yeast (Saccharomyces sp) be numbered 20122, subtilis (Bacillus subtilissubsp.subtilis) be numbered 1.0015, pediococcus acidilactici (Pediococcus acidilactici) be numbered 1.2696, streptococcus faecium (Lactobacillus acidophilus) be numbered 10180, Lactobacterium acidophilum (Lactobacillus acidophilus) be numbered 1.2686.
When buying collection bacterial classification, the planning standard activation medium model that bacterium Tibetan keeping unit can provide on the application direction of different strain is for reference, and the substratum as activated yeast is YPD ... activation medium LB ... activation medium MRS.The above substratum being matched in actication of culture is referred to as supporting substratum.
A, bacterial classification are chosen and activation:
In test tube, carry out inclined-plane according to selected species, the bacterial strain choosing health respectively and supporting activation medium and activate cultivation,
The determination of B, Shake flask medium and the cultivation of liquid spawn:
Required Shake flask medium is obtained than relation: multivitamin aqueous solution 2 parts, trace element water-soluble liquid 1 part, the activation culture based 1.5 part mixture supporting with bacterial classification shake up and disperses to implant flask by following volume parts, then add on the bacterial classification after activation, shaking table in 28-33 DEG C of sterilisable chamber and cultivate 24-48 hour produce liquid bacterial classification
The proportioning of C, seeding tank substratum and the fermentation of mixed strains are cultivated:
With reference to following component weight ratio configuration seeding tank substratum: 35% sulfuric acid 1g, 7 water magnesium sulfate 2g, sodium-chlor 1g, potassium primary phosphate 0.5g, sodium malate 2g, 2 water calcium chloride 0.5g, yeast leach liquor 0.5g, 7 aqueous ferrous sulfate 0.1g, trace element water-soluble liquid 1ml, multivitamin aqueous solution 1ml, agar 10g, sucrose 5g, peptone 5g, add the formation liquid nutrient medium that pure water forms 1L
Inject liquid nutrient medium in seeding tank after, the effective total viable count of each bacterial classification in composite fluid bacterial classification must be adjusted and reach 10
6cfu/ml-10
7cfu/ml, and carry out adjustment with reference to the following per-cent composition of each bacterial classification number of viable and become mixed strains:
Photosynthetic bacteria 20 ~ 22%,
Yeast 18 ~ 22%,
Subtilis 17 ~ 20%
Pediococcus acidilactici 18 ~ 21%,
Streptococcus faecium 22 ~ 24%,
Lactobacterium acidophilum 22 ~ 18%,
Then stir with long-pending to join in liquid nutrient medium than 100:7 ~ 10 according to the fermentor tank size of enlarged culturing, adjustment PH7.0, keep temperature 28-30 DEG C, incubation time obtains liquid spawn in 28 days.
The above trace element water-soluble liquid is the experience proportioning being adapted to each uniform growth demand in mixed strains and drawing through repetition test and formula rate adjustment.
Embodiment is as follows:
Embodiment (mg) 1234
EDTA-2Na 190 195 205 210
2 water Sodium orthomolybdates 18 19.5 20.5 22
7 aqueous ferrous sulfates 80 90 105 120
6 water nickel oxides 12 11 10 8
Boric acid 80 90 105 120
2 water cupric chlorides 80 90 105 120
Zinc chloride 80 90 105 120
4 water Manganous chloride tetrahydrates 80 90 105 120
Add distilled water 1 liter
Above-described multivitamin aqueous solution is also the experience proportioning being adapted to each uniform growth demand in mixed strains and drawing through repetition test and formula rate adjustment.Specific embodiment is as follows:
Embodiment is as follows:
Embodiment (g) A B C D
VITMAIN B1 480 490 505 520
Para-amino benzoic acid 320 310 300 180
Vitamins B
3480 490 505 520
Vitamins B
1240 50 55 60
Vitamins B
6120 110 95 80
Vitamins B
40 48 52 60
Distilled water (L) 1.2 1.1 9.5 8.
Selected 6 kinds of probiotic bacteriums to be activated respectively and further after shaking table enlarged culturing becomes liquid spawn, live body bacterium total quantity reaches 10
6cfu/ml-10
7cfu/ml.Further, per-cent composition regulation when the ratio of each bacterial classification number of viable should meet inoculation in mixed strains.To ensure in spawn fermentation tank, product final in specific substratum can reach the standard of mixed strains product.
The liquid spawn drying of generation is stayed thing admittedly to moisture < 15%, then through pulverizing, crosses 60 mesh sieves, packaging, forms effective total viable count and is no less than 10
7the powderous preparations of cfu/ gram.
Compound bio active bacteria formulation, as raw material mixed strains based on probiotic bacterium, itself possesses the base attribute that decomposing biomass material converting is biomass fertilizers, has again the ability suppressing harmful fungus grown Sum decomposition dirt.According to different objects, the auxiliary composition zymophyte system having developed at present component by biomass substratum and supporting aerobic, anaerobism fungus strain, generate microbial nutrition complementation, Various Functions grows synchronous effects.The compound bio active bacteria formulation Secondary Fermentation product being made as basis with this compound bio viable bacteria has extensively been tried out at aquatic products, livestock and poultry, agricultural planting and field of environment protection.More than ten plant compound bio active bacteria formulation directly and non-resultant product shown powerful vitality.
Claims (5)
1. compound bio active bacteria formulation, effective total viable count is no less than 10
8cfu/ml, is characterized in that: this preparation generates in the fermentor tank that supporting substratum is housed by mixed strains, and after filtration, concentrated, proportioning forms active bacteria formulation, and in said preparation, the per-cent of the living bacteria count of contained each bacterial classification is:
Photosynthetic bacteria 22 ~ 25%,
Yeast 21 ~ 24%,
Subtilis 15 ~ 20%
Pediococcus acidilactici 15 ~ 20%,
Streptococcus faecium 12 ~ 16%,
Lactobacterium acidophilum 12 ~ 16%.
2. the preparation technology of compound bio active bacteria formulation according to claim 1, is characterized in that: comprise the following steps in this technique:
A, bacterial classification are chosen and activation:
In test tube, carry out inclined-plane according to selected species, the bacterial strain choosing health respectively and supporting activation medium and activate cultivation,
The determination of B, Shake flask medium and the cultivation of liquid spawn:
Required Shake flask medium is obtained than relation: multivitamin aqueous solution 2 parts, trace element water-soluble liquid 1 part, the activation culture based 1.5 part mixture supporting with bacterial classification shake up and disperses to inject flask by following volume parts, then add on the bacterial classification after activation, shaking table in 28-33 DEG C of sterilisable chamber and cultivate 24-48 hour produce liquid bacterial classification
The proportioning of C, seeding tank substratum and the fermentation of mixed strains are cultivated:
With reference to following component weight ratio configuration seeding tank substratum: 35% dilute sulphuric acid 1ml, 7 water magnesium sulfate 2g, sodium-chlor 1g, potassium primary phosphate 0.5g, sodium malate 2g, 2 water calcium chloride 0.5g, yeast leach liquor 0.5g, 7 aqueous ferrous sulfate 0.1g, trace element water-soluble liquid 1ml, multivitamin aqueous solution 1ml, agar 10g, sucrose 5g, peptone 5g, add pure water 1L and form liquid nutrient medium
Inject aforesaid liquid substratum in spawn fermentation tank after, in adjustment composite fluid bacterial classification, the effective total viable count of each bacterial classification reaches 10
6cfu/ml-10
7cfu/ml, and the mixed strains reaching following per-cent composition with reference to each bacterial classification number of viable:
Photosynthetic bacteria 20 ~ 22%,
Yeast 18 ~ 22%,
Subtilis 17 ~ 20%
Pediococcus acidilactici 18 ~ 21%,
Streptococcus faecium 22 ~ 24%,
Lactobacterium acidophilum 22 ~ 18%,
Then join in liquid nutrient medium according to volume ratio 100:7 ~ 10 and stir, adjustment PH7.0, maintenance temperature 28-30 DEG C, incubation time obtains liquid spawn in 28 days.
3. the preparation technology of compound bio active bacteria formulation according to claim 2, is characterized in that: described trace element water-soluble liquid is mixed by following composition:
EDTA-2Na:190 ~ 210mg, 2 water Sodium orthomolybdate: 18 ~ 22mg, 7 aqueous ferrous sulfates: 80 ~ 120mg, 6 nickel oxide 8 ~ 12mg: boric acid 80 ~ 120mg, 2 water cupric chloride: 80 ~ 120mg, zinc chloride: 80 ~ 120mg, 4 water Manganous chloride tetrahydrate: 80 ~ 120mg, distilled water: 0.8 ~ 1.2L.
4. the preparation technology of compound bio active bacteria formulation according to claim 2, is characterized in that: described multivitamin aqueous solution is mixed by following composition:
VITMAIN B1: 480 ~ 520g, para-amino benzoic acid: 280 ~ 320g, vitamins B
3: 480 ~ 520mg, vitamins B
1240 ~ 60mg, vitamins B
6: 80 ~ 120mg, vitamins B
7: 40 ~ 60mg, distilled water 0.8 ~ 1.2L.
5. a compound bio viable bacteria pulvis, the processing step said according to claim 2 makes, and it is characterized in that: after step C completes, by liquid preparation drying to moisture < 15%, then through pulverizing, cross 60 mesh sieves, packaging, forms effective total viable count and is no less than 10
7the powderous preparations of cfu/ gram.
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Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105145632A (en) * | 2015-07-20 | 2015-12-16 | 潘灿平 | Composite bacterial enzyme system with multiple effects and application thereof |
| CN105540874A (en) * | 2016-02-29 | 2016-05-04 | 苏州市相城区阳澄湖镇剑成水产生态养殖专业合作社 | Water quality modifying agent for aquaculture |
| CN106942355A (en) * | 2017-04-09 | 2017-07-14 | 湖南易科生物工程有限公司 | A kind of compound method of fruit vegetables antistaling agent |
| CN107079983A (en) * | 2017-04-09 | 2017-08-22 | 湖南易科生物工程有限公司 | A kind of compound method of fruit vegetables antistaling agent |
| CN107079984A (en) * | 2017-04-09 | 2017-08-22 | 湖南易科生物工程有限公司 | A kind of compound method of fruit vegetables antistaling agent |
| CN110125149A (en) * | 2019-05-31 | 2019-08-16 | 上海天微生物科技有限公司 | A kind of processing method of life wet refuse |
| CN111018133A (en) * | 2019-12-20 | 2020-04-17 | 深圳清谷环境科技有限公司 | Environment-friendly microbial sewage treatment agent and preparation method and application thereof |
| CN113430150A (en) * | 2021-08-11 | 2021-09-24 | 真爱生命生物技术(重庆)有限公司 | Probiotics and application thereof |
| CN115918777A (en) * | 2022-12-23 | 2023-04-07 | 福建师范大学 | Method for treating fruit and vegetable garbage by using microbial agent |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN105145632A (en) * | 2015-07-20 | 2015-12-16 | 潘灿平 | Composite bacterial enzyme system with multiple effects and application thereof |
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| CN111018133B (en) * | 2019-12-20 | 2022-03-29 | 深圳清谷环境科技有限公司 | Environment-friendly microbial sewage treatment agent and preparation method and application thereof |
| CN113430150A (en) * | 2021-08-11 | 2021-09-24 | 真爱生命生物技术(重庆)有限公司 | Probiotics and application thereof |
| CN115918777A (en) * | 2022-12-23 | 2023-04-07 | 福建师范大学 | Method for treating fruit and vegetable garbage by using microbial agent |
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Application publication date: 20150513 |