CN108192845A - Spoil harmless treatment enzyme bacterium complexing agent and preparation method thereof - Google Patents
Spoil harmless treatment enzyme bacterium complexing agent and preparation method thereof Download PDFInfo
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- CN108192845A CN108192845A CN201810095752.9A CN201810095752A CN108192845A CN 108192845 A CN108192845 A CN 108192845A CN 201810095752 A CN201810095752 A CN 201810095752A CN 108192845 A CN108192845 A CN 108192845A
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- 241000894006 Bacteria Species 0.000 title claims abstract description 84
- 102000004190 Enzymes Human genes 0.000 title claims abstract description 50
- 108090000790 Enzymes Proteins 0.000 title claims abstract description 50
- 239000008139 complexing agent Substances 0.000 title claims abstract description 44
- 238000002360 preparation method Methods 0.000 title claims abstract description 25
- 238000000855 fermentation Methods 0.000 claims abstract description 68
- 230000004151 fermentation Effects 0.000 claims abstract description 68
- 239000000843 powder Substances 0.000 claims abstract description 37
- 241001465754 Metazoa Species 0.000 claims abstract description 36
- 241000193744 Bacillus amyloliquefaciens Species 0.000 claims abstract description 32
- 241000193755 Bacillus cereus Species 0.000 claims abstract description 28
- 240000005384 Rhizopus oryzae Species 0.000 claims abstract description 28
- 235000013752 Rhizopus oryzae Nutrition 0.000 claims abstract description 28
- 241001313536 Thermothelomyces thermophila Species 0.000 claims abstract description 27
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- 241000194108 Bacillus licheniformis Species 0.000 claims abstract description 24
- 230000000813 microbial effect Effects 0.000 claims abstract description 23
- 241000235015 Yarrowia lipolytica Species 0.000 claims abstract description 22
- 241000194107 Bacillus megaterium Species 0.000 claims abstract description 21
- 108091005804 Peptidases Proteins 0.000 claims abstract description 20
- 239000004365 Protease Substances 0.000 claims abstract description 18
- 102100037486 Reverse transcriptase/ribonuclease H Human genes 0.000 claims abstract description 18
- 102000004882 Lipase Human genes 0.000 claims abstract description 16
- 108090001060 Lipase Proteins 0.000 claims abstract description 16
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- 235000019421 lipase Nutrition 0.000 claims abstract description 16
- 238000000034 method Methods 0.000 claims abstract description 10
- 230000002269 spontaneous effect Effects 0.000 claims abstract description 8
- 239000002361 compost Substances 0.000 claims abstract description 7
- 230000000694 effects Effects 0.000 claims description 18
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- 239000002054 inoculum Substances 0.000 claims description 6
- 235000009566 rice Nutrition 0.000 claims description 6
- 241000726221 Gemma Species 0.000 claims description 5
- 239000000203 mixture Substances 0.000 claims description 5
- 238000006243 chemical reaction Methods 0.000 claims description 4
- 238000002156 mixing Methods 0.000 claims description 4
- 238000003756 stirring Methods 0.000 claims description 4
- 238000001035 drying Methods 0.000 claims description 3
- 239000002068 microbial inoculum Substances 0.000 claims description 3
- 239000002994 raw material Substances 0.000 claims description 3
- 241000223259 Trichoderma Species 0.000 claims description 2
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- 230000001580 bacterial effect Effects 0.000 abstract description 17
- 230000008569 process Effects 0.000 abstract description 3
- 229940088598 enzyme Drugs 0.000 description 32
- 235000019419 proteases Nutrition 0.000 description 14
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- 102000004169 proteins and genes Human genes 0.000 description 9
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- 238000012216 screening Methods 0.000 description 4
- 238000010563 solid-state fermentation Methods 0.000 description 4
- 108010059892 Cellulase Proteins 0.000 description 3
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- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 description 2
- 102000035195 Peptidases Human genes 0.000 description 2
- 240000004808 Saccharomyces cerevisiae Species 0.000 description 2
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- JIAARYAFYJHUJI-UHFFFAOYSA-L zinc dichloride Chemical compound [Cl-].[Cl-].[Zn+2] JIAARYAFYJHUJI-UHFFFAOYSA-L 0.000 description 2
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- 235000021240 caseins Nutrition 0.000 description 1
- 230000015556 catabolic process Effects 0.000 description 1
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- 238000000354 decomposition reaction Methods 0.000 description 1
- 238000006731 degradation reaction Methods 0.000 description 1
- BNIILDVGGAEEIG-UHFFFAOYSA-L disodium hydrogen phosphate Chemical compound [Na+].[Na+].OP([O-])([O-])=O BNIILDVGGAEEIG-UHFFFAOYSA-L 0.000 description 1
- 239000004744 fabric Substances 0.000 description 1
- 239000011790 ferrous sulphate Substances 0.000 description 1
- 235000003891 ferrous sulphate Nutrition 0.000 description 1
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- BAUYGSIQEAFULO-UHFFFAOYSA-L iron(2+) sulfate (anhydrous) Chemical compound [Fe+2].[O-]S([O-])(=O)=O BAUYGSIQEAFULO-UHFFFAOYSA-L 0.000 description 1
- 229910000359 iron(II) sulfate Inorganic materials 0.000 description 1
- 108010059345 keratinase Proteins 0.000 description 1
- 229910052943 magnesium sulfate Inorganic materials 0.000 description 1
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- 210000004681 ovum Anatomy 0.000 description 1
- PJNZPQUBCPKICU-UHFFFAOYSA-N phosphoric acid;potassium Chemical compound [K].OP(O)(O)=O PJNZPQUBCPKICU-UHFFFAOYSA-N 0.000 description 1
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- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 description 1
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- 239000002689 soil Substances 0.000 description 1
- 239000007921 spray Substances 0.000 description 1
- 108010050327 trypticase-soy broth Proteins 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
- 239000011592 zinc chloride Substances 0.000 description 1
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Classifications
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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
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B09—DISPOSAL OF SOLID WASTE; RECLAMATION OF CONTAMINATED SOIL
- B09B—DISPOSAL OF SOLID WASTE NOT OTHERWISE PROVIDED FOR
- B09B3/00—Destroying solid waste or transforming solid waste into something useful or harmless
-
- 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
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/14—Fungi; Culture media therefor
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/14—Fungi; Culture media therefor
- C12N1/16—Yeasts; Culture media therefor
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- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/14—Hydrolases (3)
- C12N9/16—Hydrolases (3) acting on ester bonds (3.1)
- C12N9/18—Carboxylic ester hydrolases (3.1.1)
- C12N9/20—Triglyceride splitting, e.g. by means of lipase
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- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/14—Hydrolases (3)
- C12N9/48—Hydrolases (3) acting on peptide bonds (3.4)
- C12N9/50—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25)
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Y—ENZYMES
- C12Y301/00—Hydrolases acting on ester bonds (3.1)
- C12Y301/01—Carboxylic ester hydrolases (3.1.1)
- C12Y301/01003—Triacylglycerol lipase (3.1.1.3)
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Abstract
The present invention relates to spoil innoxious process for treating fields, provide a kind of spoil harmless treatment enzyme bacterium complexing agent, including bacillus amyloliquefaciens, bacillus cereus, bacillus licheniformis, subtropical zone streptomycete, Candida lipolytica, Rhizopus oryzae, sporotrichum thermophile part, bacillus megaterium, Trichoderma viride.Meanwhile present invention provides a kind of preparation method of enzyme bacterium complexing agent, including each strain first is made hypopus microbial dry powder；Each strain dry powder and protease and lipase are mixed and made into enzyme bacterium complexing agent in proportion again.Whereby, the present invention obtains bacillus amyloliquefaciens by being screened from farm's spoil spontaneous fermentation aerobic compost sample, and the hypopus body microbial dry powder for being prepared into the strain of the bacterial strain and other components, according to its purposes using rational proportion, it is mixed to get animals died of illness corpse harmless treatment enzyme bacterium complexing agent.
Description
Technical field
The present invention relates to spoil innoxious process for treating fields more particularly to a kind of spoil harmless treatment to use
Enzyme bacterium complexing agent and preparation method thereof.
Background technology
As China's animal husbandry is intensive, the fast development of scale, animal feeding amount increases year by year, and dying of illness for generation is dynamic
Object corpse causes huge pressure to environmental protection.The harmless treatment of animals died of illness corpse whether in place, not only concerning livestock products quality
Safety and the health of the people are measured, also concerning the sustainable development of animal husbandry and social public health security.
By taking Weifang City as an example, the whole city has more than 100,000, Scale dependency field, family, has a Animal Raising Zone, the whole city more than 4000
Year delivers ten thousand pigs, 100,000 chickens for sale, 100 cow head of livestock on hand, ten thousand laying hens feed lot up to 100 many places, have standardization raising
2.3 ten thousand are given up, generates a large amount of animals died of illness every year.If animals died of illness corpse cannot be effectively treated, environment is not only influenced
Health, contaminated soil water resource and air can also rot with the corruption of animals died of illness corpse, and the diffusion of virus and bacteria is even due to ring
The influence in border is morphed, and causes new animal epidemic disease source, and incalculable damage is brought, and can be serious to citywide animal husbandry
Human and livestock health is endangered, causes public health event.
The processing method of the spoils such as traditional burning method, landfill method, which exists, causes secondary pollution, environmental bearing capacity
The problems such as limited.Therefore trend is had become using the method for microorganism harmless treatment spoil.
At present, using the method for microorganism harmless treatment spoil also in the presence of time-consuming, complex management, degradation efficiency
Low problem.
In summary, the existing technology has inconveniences and defects in actual use, so it is necessary to be improved.
Invention content
For it is above-mentioned the defects of, it is an object of the invention to a kind of spoil harmless treatment enzyme bacterium complexing agent, packet
Include bacillus amyloliquefaciens, bacillus cereus, bacillus licheniformis, subtropical zone streptomycete, Candida lipolytica, Rhizopus oryzae,
Sporotrichum thermophile part, bacillus megaterium, Trichoderma viride.
Meanwhile present invention provides a kind of preparation method of enzyme bacterium complexing agent, including each strain first is made hypopus
Microbial dry powder；Each strain dry powder and protease and lipase are mixed and made into enzyme bacterium complexing agent in proportion again.
To achieve these goals, the present invention provides a kind of spoil harmless treatment enzyme bacterium complexing agent, including such as
The component of lower parts by weight：
The Rate activity of spoil harmless treatment according to the present invention enzyme bacterium complexing agent, the protease and lipase
For 100000U/g.
Spoil harmless treatment according to the present invention enzyme bacterium complexing agent, the bacillus amyloliquefaciens are from cultivation
The dominant bacteria that can generate protease filtered out in the spoil spontaneous fermentation aerobic compost of field.
A kind of preparation method of spoil harmless treatment enzyme bacterium complexing agent according to the present invention, including walking as follows
Suddenly：
Step 1：Respectively by the bacillus amyloliquefaciens, Bacillus cereus, bacillus licheniformis, solution fat vacation silk ferment
Mother, bacillus megaterium, subtropical zone streptomycete, Rhizopus oryzae, sporotrichum thermophile and Trichoderma viride strain that hypopus is made is micro-
Biological dry powder；
Step 2：By hypopus microbial dry powder made of each strain of the step 1 and protease and lipase in proportion
It is mixed and made into the enzyme bacterium complexing agent.
The preparation method of spoil harmless treatment enzyme bacterium complexing agent according to the present invention, the hypopus microorganism
The preparation method of dry powder includes the following steps：
A. activation culture
Each strain is seeded to equipped with volume ratio to live in the conical flask of 15~30% activation mediums respectively
Change culture；
B. the strain after the step A culture is inoculated into seeding tank and is enlarged culture；Inoculum concentration is：Volume ratio 1
~3%；
C. the cultured strain of the step B is inoculated into fermentation tank and carries out fermentation expansion culture；Inoculum concentration is：Volume
Than 5~10%；
D. each single bacterium that the step C obtains is dehydrated, the hypopus microbial dry powder is made；
Same fermentation medium is housed in the seeding tank and fermentation tank；Equipped with fermentation medium in the seeding tank
It measures and is：Volume ratio 50~70%；
Bacillus amyloliquefaciens, Bacillus cereus, bacillus licheniformis, solution fat are false as described in A~D steps culture
The parameter of silk yeast and the hypopus microbial dry powder of bacillus megaterium is shown in Table one, table two；
Subtropical zone streptomycete, Rhizopus oryzae, sporotrichum thermophile and Trichoderma viride are stopped as described in A~D steps culture
The parameter of dormancy body microbial dry powder is shown in Table four, table five；
Bacillus amyloliquefaciens, Bacillus cereus, bacillus licheniformis, solution fat vacation silk ferment described in the step C culture
During female and bacillus megaterium, the amount equipped with fermentation medium is in the fermentation tank：Volume ratio 50~70%；
Described in the step C culture when subtropical zone streptomycete, Rhizopus oryzae, sporotrichum thermophile and Trichoderma viride, the hair
The amount equipped with fermentation medium is in fermentation tank：Volume ratio 30~50%.
The preparation method of spoil harmless treatment enzyme bacterium complexing agent according to the present invention solves starch described in step C
Bacillus, Bacillus cereus, bacillus licheniformis, Candida lipolytica and bacillus megaterium fermentation tank in convert
Rate is more than 95%；Bacterium number content is respectively in fermentation tank：Bacillus amyloliquefaciens >=1010A/ml, Bacillus cereus >=7*
109A/ml, bacillus licheniformis >=8*109A/ml, Candida lipolytica >=4*109A/ml, bacillus megaterium >=1010
A/ml；
Subtropical zone streptomycete described in step C, Rhizopus oryzae, sporotrichum thermophile and Trichoderma viride fermentation tank in conversion ratio it is equal
More than 90%, in fermentation tank bacterium number content >=109A/ml.
The preparation method of spoil harmless treatment enzyme bacterium complexing agent according to the present invention, the solution starch gemma bar
Bacterium, Bacillus cereus, bacillus licheniformis, Candida lipolytica and bacillus megaterium carry out the D steps using spray
Mist is dried.
The preparation method of spoil harmless treatment enzyme bacterium complexing agent according to the present invention, the subtropical zone strepto-
Bacterium, Rhizopus oryzae, sporotrichum thermophile and Trichoderma viride carry out the D steps and use fluidized drying.
The preparation method of spoil harmless treatment enzyme bacterium complexing agent according to the present invention, the solution starch gemma bar
Bacterium, Bacillus cereus, bacillus licheniformis, Candida lipolytica and bacillus megaterium hypopus microbial dry powder in
Bacterium number content is >=109A/gram；
The subtropical zone streptomycete, Rhizopus oryzae, sporotrichum thermophile and Trichoderma viride hypopus microbial dry powder in bacterium
Counting content is respectively：Subtropical zone streptomycete >=3*109It is a/gram, Rhizopus oryzae >=4*109It is a/gram, sporotrichum thermophile >=6*109A/
Gram, Trichoderma viride >=5*109A/gram.
The preparation method of spoil harmless treatment enzyme bacterium complexing agent according to the present invention, it is compound using the enzyme bacterium
It is as follows that agent carries out the step of animals died of illness corpse harmless treatment：
First, animals died of illness corpse in fermentation machine is crushed, then adds powdered rice hulls, allow the spoil of crushing
With the abundant mixing of powdered rice hulls, and moisture is adjusted to 38~42%；Then fermentation machine internal temperature is risen to 120~125 DEG C, maintained
25~35 minutes；
Secondly, when fermentation machines internal temperature is down to 36~38 DEG C, the enzyme of weight ratio 1-3% is added into fermentation machine
Bacterium complexing agent simultaneously opens stirring, is sufficiently mixed microbial inoculum and raw material；It is persistently stirred during fermentation；
Fermentation time 24~48 hours；37~50 DEG C of fermentation temperature.
The purpose of the present invention is to provide a kind of spoil harmless treatment enzyme bacterium complexing agents and preparation method thereof, lead to
It crosses from farm's spoil spontaneous fermentation aerobic compost sample screening and obtains a bacillus amyloliquefaciens, and by the bacterial strain
With Bacillus cereus, bacillus licheniformis, Candida lipolytica, subtropical zone streptomycete, Rhizopus oryzae, sporotrichum thermophile, green
The strain of trichoderma and bacillus megaterium carries out the hypopus body microbial dry powder being prepared into after High Density Cultivation, according to its purposes
Using rational proportion, it is mixed to get animals died of illness corpse harmless treatment enzyme bacterium compound formulation.Animals died of illness corpse is handled with it,
Protein, starch, fat, cellulose, hair in corpse and its auxiliary material etc. can thoroughly be decomposed, be converted to plant and easily inhale
Receive the small-molecule substance utilized.Corpse decrement can be more than 90% in 48 hours, and surplus material can do organic fertilizer.These have simultaneously
Beneficial microorganism and its metabolite can also inhibit the growth and breeding of pathogenic bacteria in spoil, reduce the diffusion of pathogenic bacteria.This
Invention is for animals died of illness corpse harmless treatment, and compared with prior art, production cost is low, easy to use, environmentally protective, processing
Effect is high, turns waste into wealth.
Specific embodiment
In order to make the purpose , technical scheme and advantage of the present invention be clearer, with reference to embodiments, to the present invention
It is further elaborated.It should be appreciated that the specific embodiments described herein are merely illustrative of the present invention, it is not used to
Limit the present invention.
The present invention provides a kind of spoil harmless treatment enzyme bacterium complexing agent, the component mixing including following parts by weight
It is prepared：
Preferably, the Rate activity of the protease of the invention and lipase is 100000U/g.
The preparation method of the spoil harmless treatment enzyme bacterium complexing agent includes the following steps：
Step 1: respectively by the bacillus amyloliquefaciens, Bacillus cereus, bacillus licheniformis, solution fat vacation silk ferment
Mother, bacillus megaterium, subtropical zone streptomycete, Rhizopus oryzae, sporotrichum thermophile and Trichoderma viride strain that hypopus is made is micro-
Biological dry powder；
Step 2: by hypopus microbial dry powder made of each strain of the step 1 and protease and lipase in proportion
It is mixed and made into the enzyme bacterium complexing agent；
Preferably, the preparation method of the hypopus microbial dry powder of the invention includes the following steps：
A. activation culture
Each strain is seeded to equipped with volume ratio to live in the conical flask of 15~30% activation mediums respectively
Change culture；
B. the strain after the step A culture is inoculated into seeding tank and is enlarged culture；Inoculum concentration is：Volume ratio 1
~3%；
C. the cultured strain of the step B is inoculated into fermentation tank and carries out fermentation expansion culture；Inoculum concentration is：Volume
Than 5~10%；
D. each single bacterium that the step C obtains is dehydrated, the hypopus microbial dry powder is made；
Same fermentation medium is housed in the seeding tank and fermentation tank；Equipped with fermentation medium in the seeding tank
It measures and is：Volume ratio 50~70%；
The bacillus amyloliquefaciens, Bacillus cereus, bacillus licheniformis, Candida lipolytica and huge gemma
Bacillus uses High Density Cultivation；It is shown in Table by the parameter of the hypopus microbial dry powder of each strain of A~D steps culture
First, table two；Bacterium number content in ferment effect and dry powder is shown in Table three；
Preferably, the bacillus amyloliquefaciens of the invention, Bacillus cereus, bacillus licheniformis, solution fat are false
Silk yeast and bacillus megaterium carry out the D steps using spray drying；
Preferably, bacillus amyloliquefaciens, Bacillus cereus, lichens bud described in the step C culture of the invention
When spore bacillus, Candida lipolytica and bacillus megaterium, the amount equipped with fermentation medium is in the fermentation tank：Volume ratio
50~70%；
The subtropical zone streptomycete, Rhizopus oryzae, sporotrichum thermophile and Trichoderma viride use solid state fermentation culture；By described
The parameter of the hypopus microbial dry powder of each strain of A~D step cultures is shown in Table four, table five；In ferment effect and dry powder
Bacterium number content is shown in Table six；
Preferably, the subtropical zone streptomycete of the invention, Rhizopus oryzae, sporotrichum thermophile and Trichoderma viride carry out institute
D steps are stated using fluidized drying；
Preferably, subtropical zone streptomycete described in the step C culture of the invention, Rhizopus oryzae, sporotrichum thermophile and
During Trichoderma viride, the amount equipped with fermentation medium is in the fermentation tank：Volume ratio 30~50%；
Preferably, the bacillus amyloliquefaciens of the invention are from farm's spoil spontaneous fermentation aerobic compost
Middle screening generates the dominant bacteria of protease.Then Jin Hang Oscillating bottles of experiment, pilot experiment finally produce qualified single bacterium system
Agent.Concrete operation step is as described below：
Farm's spoil spontaneous fermentation aerobic compost temperature raising period sample is taken using four point samplings, then with dilution
Physiological saline after rubbing method sterilizes sample does gradient dilution, takes 10-5、10-6、10-7Three each 100ul of gradient are applied respectively
In cloth and following tablet.Bacteria culture media uses beef-protein medium, and unwrapping wire bacterium culture medium is using No. 1 culture of Gao Shi
Base, fungi culture medium use PDA culture medium.Each gradient does four repetitions, and bacteria culture media tablet is placed in 37 degree of incubators
Culture 1-2 days, actinomyces culture medium flat plate and fungi culture medium tablet are placed in 30 degree of incubators and cultivate 3-4 days.
Cultured tablet is taken out, dominant strain is observed, tri- dominant bacterias of 1#, 2#, 3# is found that on Bacterial Plate
Strain, actinomyces tablet and fungi tablet do not find dominant strain, then respectively draw 1#, 2# and 3# bacterial strain on Bacterial Plate
Line detaches single bacterium colony, and is positioned in 37 degree of incubators and cultivates 1-2 days.
The single bacterium colony of cultured three bacterial strains is inoculated in respectively in protease strain screening and culturing medium；The culture medium
It is formulated and is：Potassium dihydrogen phosphate 0.36g/L, magnesium sulfate 0.5g/L, zinc chloride 0.014g/L, disodium hydrogen phosphate 1.07g/L, sodium chloride
0.16g/L, calcium chloride 0.002g/L, ferrous sulfate 0.002g/L, casein 4g/L, trypticase 0.05g/L, agar powder 20g/L,
PH6.8-7.0 is then placed into cultivating 1-2 days in 37 degree of incubators.The screening flat board of cultured three bacterial strains is observed, as a result
The periphery of bacterial colonies of only 1# bacterial strains has transparent circle, and the periphery of bacterial colonies of 2# and 3# bacterial strains does not have transparent circle, illustrates that 1# bacterial strains have
Generate the ability of protease.1# bacterial strains are subjected to culture presevation, and send the inclined-plane of the bacterial strain to associated mechanisms and carry out strain mirror
Fixed, qualification result shows that the bacterial strain is bacillus amyloliquefaciens.
Bacillus amyloliquefaciens hypopus microorganism is made by abovementioned steps in the bacillus amyloliquefaciens of gained again later
Dry powder.
Since the bacillus amyloliquefaciens in the present invention are screened in farm's spoil spontaneous fermentation sample
The dominant strain with very strong protease production capacity, the protein substance in animals died of illness corpse can thoroughly be divided
Solution.
Bacillus cereus in the present invention has the activity of higher protease, amylase, can decompose animals died of illness corpse
Starch, protein substance in body.
Bacillus licheniformis in the present invention has higher keratinase activity, can be by the hair in animals died of illness corpse
Hair is decomposed, which can also generate resistant activity material, can inhibit the growth and breeding of pathogenic bacteria.
Subtropical zone streptomycete in the present invention has stronger starch and protein hydrolysis ability, especially with stronger
Keratin hydrolysis ability can decompose starch, protein, the hair class substance in animals died of illness corpse.The generation of the bacterium
The growth and breeding of pathogenic bacteria can also directly be inhibited by thanking to product.It is easily absorbed by plants by the substance that streptomycete is decomposed, so as to increase
Strong plant is to the repellence and immunity of various diseases.
Candida lipolytica in the present invention has the very strong ability to reduce fat with protein, can be by animals died of illness
Fat, protein substance in corpse are thoroughly decomposed, and have higher lipase and proteinase activity.
Rhizopus oryzae in the present invention has very high lipase and amylase activity, can be by the fat in animals died of illness corpse
Fat, starchy material are thoroughly decomposed.
Sporotrichum thermophile in the present invention belongs to Thermophilic Bacteria, and optimum growth temperature is 45~50 degree, can be in hot environment
The lower cellulose substances more thoroughly decomposed in animals died of illness corpse auxiliary material.
The bulky grains such as the protein in animals died of illness corpse can be resolved into amino by the bacillus megaterium in the present invention
The small-molecule substances such as acid have very high proteinase activity.
Trichoderma viride in the present invention can generate a variety of biologically active enzyme systems, such as：Cellulase, chitinase,
Zytase etc..Trichoderma viride is one of highest bacterial strain of cellulase-producing activity, and generated cellulase can degrade disease
Fiber substance in dead spoil auxiliary material, effect is very good, and is a kind of resourceful antagonistic microbe, can effectively press down
Pathogenic bacteria in animals died of illness corpse processed.
Protease and lipase in the present invention are mainly used for decomposing protein and fats object in animals died of illness corpse
Matter accelerates the decomposition of spoil, and quickly and effectively food is provided for the microorganism in compound formulation, makes these beneficial micro- lifes
Object obtains fast-growth breeding, becomes dominant strain, inhibits the diffusion of pathogenic bacteria.
Preferably, each strain in the present invention did antagonistic experiment before compounding, as a result showed between them without short of money
Resistance.
It is as follows that animals died of illness corpse harmless treatment step is carried out using the present invention：
First, animals died of illness corpse in fermentation machine is crushed, then adds powdered rice hulls according to a certain percentage, allow powder
The broken abundant mixing of spoil and powdered rice hulls, 38~42% are adjusted to by moisture, fermentation machine internal temperature is then risen to 120~
125 DEG C, maintain 25~35 minutes, by spoil germ, virus thoroughly kill.
Secondly, when fermentation machines internal temperature is down to 36~38 DEG C, the enzyme bacterium that weight ratio 1-3% is added into fermentation machine is answered
Mixture opens stirring, microbial inoculum and raw material is allowed to be sufficiently mixed, and ferment, wants always on stirring during fermentation, and by temperature
It maintains between 37~50 DEG C.
The weight ratio of each component sets embodiment and carries out animals died of illness corpse in enzyme bacterium complexing agent by changing the present invention
Body harmless treatment；As a result it is as follows：
Embodiment one：The weight proportion of each component be 30 parts of bacillus amyloliquefaciens, 5 parts of bacillus cereus, lichens bud
8 parts of spore bacillus, 15 parts of subtropical zone streptomycete, 10 parts of Candida lipolytica, 10 parts of Rhizopus oryzae, 10 parts of sporotrichum thermophile, huge bud
3 parts of spore bacillus, 4 parts of Trichoderma viride, 2 parts of protease, the enzyme bacterium complexing agent that 3 parts of lipase is mixed carry out animals died of illness corpse
Harmless treatment, ferments 24 hours, and acquired achievement is：Spoil decrement 92%, material organic matter after fermentation
Content 53%, total nutrient 6%, moisture 28%, pH 6.9, induced worm egg death rate 100%, and excrement colibacillus group is not detected.
Embodiment two：The weight proportion of each component be 15 parts of bacillus amyloliquefaciens, 10 parts of bacillus cereus, lichens bud
4 parts of spore bacillus, 8 parts of subtropical zone streptomycete, 20 parts of Candida lipolytica, 20 parts of Rhizopus oryzae, 20 parts of sporotrichum thermophile, huge gemma
1 part of bacillus, 2 parts of Trichoderma viride .5 parts of proteinase-10, the enzyme bacterium complexing agent that 5 parts of lipase is mixed carry out animals died of illness corpse
Harmless treatment, ferments 36 hours, and acquired achievement is：Spoil decrement 96%, material organic matter after fermentation
Content 55%, total nutrient 10%, moisture 20%, pH 7.2, induced worm egg death rate 100%, and fecal coliform is not detected
Group.
Embodiment three：The weight proportion of each component be 20 parts of bacillus amyloliquefaciens, 7 parts of bacillus cereus, lichens bud
6 parts of spore bacillus, 18 parts of subtropical zone streptomycete, 15 parts of Candida lipolytica, 14 parts of Rhizopus oryzae, 13 parts of sporotrichum thermophile, huge bud
5 parts of spore bacillus, 1 part of Trichoderma viride, protease 3 part, the enzyme bacterium complexing agent that 0.8 part of lipase is mixed carry out animals died of illness corpse
Body harmless treatment, ferments 48 hours, and acquired achievement is：Spoil decrement 97%, material after fermentation is organic
Matter content 60%, total nutrient 9%, moisture 23%, pH 7.5, induced worm egg death rate 100%, and fecal coliform is not detected
Group.
Remaining embodiment process repeats no more, and the results are shown in Table seven.By each embodiment it is known that the weight of each component is matched
Than for 15~30 parts of bacillus amyloliquefaciens, 5~10 parts of bacillus cereus, 4~8 parts of bacillus licheniformis, subtropical zone strepto-
8~18 parts of bacterium, 10~20 parts of Candida lipolytica, 10~25 parts of Rhizopus oryzae, 10~20 parts of sporotrichum thermophile, bacillus megaterium
1~5 part, 1~4 part of Trichoderma viride, proteinase-10 .5~3 part, the enzyme bacterium complexing agent that 0.8~5 part of lipase is mixed carries out sick
Dead spoil harmless treatment, fermentation time 24~48 hours, acquired achievement are：Spoil is reduced more than 90%,
The material content of organic matter 45~60% after fermentation, total nutrient 5~10%, moisture 25~30%, pH 6.8-7.5, ascarid
The worm's ovum death rate is 100%, and does not detect excrement colibacillus group；Material after fermentation meets the relevant criterion of organic fertilizer, can be with
It is used as organic fertilizer.
Table one：Activation culture parameter (High Density Cultivation)
Table two：Expand culture parameters (High Density Cultivation)
Table three：Bacterium number content (High Density Cultivation) in ferment effect and dry powder
Table four：Activation culture parameter (solid state fermentation culture)
Table five：Expand culture parameters (solid state fermentation culture)
Table six：Bacterium number content (solid state fermentation culture) in ferment effect and dry powder
Table seven：The weight and application effect of each embodiment
Note：A is bacillus amyloliquefaciens in table；B is bacillus cereus；C is bacillus licheniformis；D is subtropical zone chain
Mould；E is Candida lipolytica；F is Rhizopus oryzae；G is sporotrichum thermophile；H is bacillus megaterium；I is Trichoderma viride；J is
Protease；K is lipase
In conclusion the purpose of the present invention is to provide a kind of spoil harmless treatment enzyme bacterium complexing agent and its systems
Preparation Method obtains a bacillus amyloliquefaciens by being screened from farm's spoil spontaneous fermentation aerobic compost sample,
And by the bacterial strain and Bacillus cereus, bacillus licheniformis, Candida lipolytica, subtropical zone streptomycete, Rhizopus oryzae, thermophilic side
Spore is mould, the strain of Trichoderma viride and bacillus megaterium carries out the hypopus body microbial dry powder being prepared into after High Density Cultivation,
According to its purposes using rational proportion, it is mixed to get animals died of illness corpse harmless treatment enzyme bacterium compound formulation.With its processing disease
Dead spoil can thoroughly be decomposed protein, starch, fat, cellulose, hair in corpse and its auxiliary material etc., be turned
The small-molecule substance that chemical conversion plant is easily absorbed and utilized.Corpse decrement can be more than 90% in 48 hours, and surplus material can do organic
Fertilizer.These beneficial microbes and its metabolite can also inhibit the growth and breeding of pathogenic bacteria in spoil simultaneously, reduce and cause
The diffusion of germ.The present invention is used for animals died of illness corpse harmless treatment, and compared with prior art, production cost is low, easy to use,
Environmentally protective, processing effect is high, turns waste into wealth.
Certainly, the present invention can also have other various embodiments, without deviating from the spirit and substance of the present invention, ripe
It knows those skilled in the art and makes various corresponding changes and deformation, but these corresponding changes and change in accordance with the present invention
Shape should all belong to the protection domain of appended claims of the invention.
Claims (10)
1. a kind of spoil harmless treatment enzyme bacterium complexing agent, which is characterized in that include the component of following parts by weight：
2. spoil harmless treatment according to claim 1 enzyme bacterium complexing agent, which is characterized in that the protease
Rate activity with lipase is 100000U/g.
3. spoil harmless treatment according to claim 1 or 2 enzyme bacterium complexing agent, which is characterized in that the solution
Bacillus amyloliquefaciens are the dominant bacterias that can generate protease filtered out from farm's spoil spontaneous fermentation aerobic compost
Kind.
4. a kind of preparation of spoil harmless treatment enzyme bacterium complexing agent according to claims 1 to 3 any one
Method, which is characterized in that include the following steps：
Step 1：Respectively by bacillus amyloliquefaciens, Bacillus cereus, bacillus licheniformis, Candida lipolytica, huge
Bacterium anthracoides, subtropical zone streptomycete, Rhizopus oryzae, sporotrichum thermophile and Trichoderma viride strain hypopus microorganism be made do
Powder；
Step 2：Hypopus microbial dry powder made of each strain of the step 1 is mixed in proportion with protease and lipase
The enzyme bacterium complexing agent is made.
5. the preparation method of spoil harmless treatment enzyme bacterium complexing agent according to claim 4, which is characterized in that
The preparation method of the hypopus microbial dry powder includes the following steps：
A. activation culture
Each strain is seeded to equipped with volume ratio to carry out activation training in the conical flask of 15~30% activation mediums respectively
It supports；
B. the strain after the step A culture is inoculated into seeding tank and is enlarged culture；Inoculum concentration is：Volume ratio 1~
3%；
C. the cultured strain of the step B is inoculated into fermentation tank and carries out fermentation expansion culture；Inoculum concentration is：Volume ratio 5
~10%；
D. each single bacterium that the step C obtains is dehydrated, the hypopus microbial dry powder is made；
Same fermentation medium is housed in the seeding tank and fermentation tank；The amount of fermentation medium is housed in the seeding tank
For：Volume ratio 50~70%；
Bacillus amyloliquefaciens, Bacillus cereus, bacillus licheniformis, solution fat vacation silk ferment as described in A~D steps culture
The parameter of female and bacillus megaterium hypopus microbial dry powder is shown in Table one, table two；
As described in A~D steps culture subtropical zone streptomycete, Rhizopus oryzae, sporotrichum thermophile and Trichoderma viride hypopus
The parameter of microbial dry powder is shown in Table four, table five；
Bacillus amyloliquefaciens, Bacillus cereus, bacillus licheniformis, Candida lipolytica described in the step C culture with
And during bacillus megaterium, the amount equipped with fermentation medium is in the fermentation tank：Volume ratio 50~70%；
Described in the step C culture when subtropical zone streptomycete, Rhizopus oryzae, sporotrichum thermophile and Trichoderma viride, the fermentation tank
The interior amount equipped with fermentation medium is：Volume ratio 30~50%.
Table one
Table two
Table four
Table five
6. the preparation method of spoil harmless treatment enzyme bacterium complexing agent according to claim 5, which is characterized in that
Bacillus amyloliquefaciens, Bacillus cereus, bacillus licheniformis, Candida lipolytica and huge gemma bar described in step C
Conversion ratio is more than 95% in the fermentation tank of bacterium；Bacterium number content is respectively in fermentation tank：Bacillus amyloliquefaciens >=1010A/ml,
Bacillus cereus >=7*109A/ml, bacillus licheniformis >=8*109A/ml, Candida lipolytica >=4*109It is a/ml, huge
Bacterium anthracoides >=1010A/ml；
Subtropical zone streptomycete described in step C, Rhizopus oryzae, sporotrichum thermophile and Trichoderma viride fermentation tank in conversion ratio be more than
90%, bacterium number content is >=10 in fermentation tank9A/ml.
7. the preparation method of spoil harmless treatment enzyme bacterium complexing agent according to claim 5, which is characterized in that
The bacillus amyloliquefaciens, Bacillus cereus, bacillus licheniformis, Candida lipolytica and bacillus megaterium carry out
The D steps are using spray drying.
8. the preparation method of spoil harmless treatment enzyme bacterium complexing agent according to claim 5, which is characterized in that
The subtropical zone streptomycete, Rhizopus oryzae, sporotrichum thermophile and Trichoderma viride carry out the D steps and use fluidized drying.
9. the preparation method of spoil harmless treatment enzyme bacterium complexing agent according to claim 5, which is characterized in that
The bacillus amyloliquefaciens, Bacillus cereus, bacillus licheniformis, Candida lipolytica and bacillus megaterium stop
In dormancy body microbial dry powder bacterium number content >=109A/gram；
The subtropical zone streptomycete, Rhizopus oryzae, sporotrichum thermophile and Trichoderma viride hypopus microbial dry powder in bacterium number contain
Amount is respectively：Subtropical zone streptomycete >=3*109It is a/gram, Rhizopus oryzae >=4*109It is a/gram, sporotrichum thermophile >=6*109It is a/gram, it is green
Color trichoderma >=5*109A/gram.
10. spoil harmless treatment enzyme bacterium complexing agent and its preparation side according to claim 4~9 any one
Method, which is characterized in that the step of carrying out animals died of illness corpse harmless treatment using the enzyme bacterium complexing agent is as follows：
First, animals died of illness corpse in fermentation machine is crushed, then adds powdered rice hulls, allow the spoil and rice of crushing
The abundant mixing of shell powder, and moisture is adjusted to 38~42%；Then fermentation machine internal temperature is risen to 120~125 DEG C, maintain 25~
35 minutes；
Secondly, when fermentation machines internal temperature is down to 36~38 DEG C, the enzyme bacterium that weight ratio 1-3% is added into fermentation machine is answered
Mixture simultaneously opens stirring, is sufficiently mixed microbial inoculum and raw material；It is persistently stirred during fermentation；
Fermentation time 24~48 hours；37~50 DEG C of fermentation temperature.
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Denomination of invention: Enzyme bacteria compound agent for harmless treatment of animal corpse and its preparation method Effective date of registration: 20220412 Granted publication date: 20210820 Pledgee: Weifang rural commercial bank Limited by Share Ltd. hi tech sub branch Pledgor: SHANDONG SUKAHAN BIO-TECHNOLOGY Co.,Ltd. Registration number: Y2022980004100 |
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Date of cancellation: 20230804 Granted publication date: 20210820 Pledgee: Weifang rural commercial bank Limited by Share Ltd. hi tech sub branch Pledgor: SHANDONG SUKAHAN BIO-TECHNOLOGY Co.,Ltd. Registration number: Y2022980004100 |
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Denomination of invention: Enzyme and bacterial complex agents for harmless treatment of animal carcasses and their preparation methods Granted publication date: 20210820 Pledgee: Weifang rural commercial bank Limited by Share Ltd. hi tech sub branch Pledgor: SHANDONG SUKAHAN BIO-TECHNOLOGY Co.,Ltd. Registration number: Y2024980006749 |