CN105524835A - Obtaining method and application of salt-tolerant cellulose decomposition bacteria colony - Google Patents
Obtaining method and application of salt-tolerant cellulose decomposition bacteria colony Download PDFInfo
- Publication number
- CN105524835A CN105524835A CN201610007623.0A CN201610007623A CN105524835A CN 105524835 A CN105524835 A CN 105524835A CN 201610007623 A CN201610007623 A CN 201610007623A CN 105524835 A CN105524835 A CN 105524835A
- Authority
- CN
- China
- Prior art keywords
- flora
- salt tolerant
- cellulose decomposition
- salt
- bacterial classification
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 239000001913 cellulose Substances 0.000 title claims abstract description 65
- 229920002678 cellulose Polymers 0.000 title claims abstract description 65
- 238000000354 decomposition reaction Methods 0.000 title claims abstract description 61
- 241000894006 Bacteria Species 0.000 title claims abstract description 42
- 238000000034 method Methods 0.000 title claims abstract description 24
- 235000010980 cellulose Nutrition 0.000 claims abstract description 64
- 230000001580 bacterial effect Effects 0.000 claims abstract description 47
- 238000000855 fermentation Methods 0.000 claims abstract description 33
- 230000004151 fermentation Effects 0.000 claims abstract description 33
- 229920000168 Microcrystalline cellulose Polymers 0.000 claims abstract description 10
- 235000019813 microcrystalline cellulose Nutrition 0.000 claims abstract description 10
- 239000008108 microcrystalline cellulose Substances 0.000 claims abstract description 10
- 229940016286 microcrystalline cellulose Drugs 0.000 claims abstract description 10
- 238000012216 screening Methods 0.000 claims abstract description 8
- 239000000725 suspension Substances 0.000 claims abstract description 6
- 238000011081 inoculation Methods 0.000 claims abstract description 3
- 150000003839 salts Chemical class 0.000 claims description 81
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 37
- 108010059892 Cellulase Proteins 0.000 claims description 26
- 229940106157 cellulase Drugs 0.000 claims description 26
- 239000002054 inoculum Substances 0.000 claims description 26
- 239000000203 mixture Substances 0.000 claims description 25
- 239000010815 organic waste Substances 0.000 claims description 19
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 claims description 15
- 239000010802 sludge Substances 0.000 claims description 15
- 238000002360 preparation method Methods 0.000 claims description 13
- 230000015556 catabolic process Effects 0.000 claims description 12
- 238000006731 degradation reaction Methods 0.000 claims description 12
- 235000015097 nutrients Nutrition 0.000 claims description 9
- 238000012546 transfer Methods 0.000 claims description 9
- VNWKTOKETHGBQD-UHFFFAOYSA-N methane Chemical compound C VNWKTOKETHGBQD-UHFFFAOYSA-N 0.000 claims description 8
- 108090000790 Enzymes Proteins 0.000 claims description 7
- 102000004190 Enzymes Human genes 0.000 claims description 7
- 229940088598 enzyme Drugs 0.000 claims description 7
- 239000010865 sewage Substances 0.000 claims description 7
- 210000003608 fece Anatomy 0.000 claims description 6
- 230000015784 hyperosmotic salinity response Effects 0.000 claims description 6
- 239000012531 culture fluid Substances 0.000 claims description 4
- 235000015170 shellfish Nutrition 0.000 claims description 4
- 230000006866 deterioration Effects 0.000 claims description 3
- 230000002906 microbiologic effect Effects 0.000 claims description 3
- 239000000123 paper Substances 0.000 abstract description 27
- 239000010902 straw Substances 0.000 abstract description 7
- 239000010794 food waste Substances 0.000 description 12
- 239000010813 municipal solid waste Substances 0.000 description 10
- 235000012055 fruits and vegetables Nutrition 0.000 description 6
- 239000007788 liquid Substances 0.000 description 6
- 230000000052 comparative effect Effects 0.000 description 5
- 230000000694 effects Effects 0.000 description 5
- 238000002474 experimental method Methods 0.000 description 5
- 239000000047 product Substances 0.000 description 5
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N lactic acid Chemical compound CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 description 4
- 244000005700 microbiome Species 0.000 description 4
- 235000013619 trace mineral Nutrition 0.000 description 4
- 239000011573 trace mineral Substances 0.000 description 4
- 230000009977 dual effect Effects 0.000 description 3
- 238000010335 hydrothermal treatment Methods 0.000 description 3
- 239000000758 substrate Substances 0.000 description 3
- 239000001888 Peptone Substances 0.000 description 2
- 108010080698 Peptones Proteins 0.000 description 2
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
- 230000003042 antagnostic effect Effects 0.000 description 2
- 229940041514 candida albicans extract Drugs 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 238000011033 desalting Methods 0.000 description 2
- 238000000502 dialysis Methods 0.000 description 2
- 239000003337 fertilizer Substances 0.000 description 2
- 239000004519 grease Substances 0.000 description 2
- 239000004310 lactic acid Substances 0.000 description 2
- 235000014655 lactic acid Nutrition 0.000 description 2
- 238000004519 manufacturing process Methods 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 235000012054 meals Nutrition 0.000 description 2
- 235000019319 peptone Nutrition 0.000 description 2
- 239000007787 solid Substances 0.000 description 2
- 239000006228 supernatant Substances 0.000 description 2
- 239000012138 yeast extract Substances 0.000 description 2
- 239000002028 Biomass Substances 0.000 description 1
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 1
- 229920002488 Hemicellulose Polymers 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 229910052799 carbon Inorganic materials 0.000 description 1
- -1 chopsticks Substances 0.000 description 1
- 230000008878 coupling Effects 0.000 description 1
- 238000010168 coupling process Methods 0.000 description 1
- 238000005859 coupling reaction Methods 0.000 description 1
- 230000007812 deficiency Effects 0.000 description 1
- 230000000593 degrading effect Effects 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 238000003912 environmental pollution Methods 0.000 description 1
- 229910052739 hydrogen Inorganic materials 0.000 description 1
- 239000001257 hydrogen Substances 0.000 description 1
- 230000010354 integration Effects 0.000 description 1
- 230000003993 interaction Effects 0.000 description 1
- 239000010806 kitchen waste Substances 0.000 description 1
- 230000000813 microbial effect Effects 0.000 description 1
- 239000004033 plastic Substances 0.000 description 1
- 229920003023 plastic Polymers 0.000 description 1
- 230000001737 promoting effect Effects 0.000 description 1
- 238000004064 recycling Methods 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 239000011780 sodium chloride Substances 0.000 description 1
Classifications
-
- 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
- C12N1/02—Separating microorganisms from their culture media
-
- 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
- C12N1/36—Adaptation or attenuation of cells
-
- 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
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/14—Hydrolases (3)
- C12N9/24—Hydrolases (3) acting on glycosyl compounds (3.2)
- C12N9/2402—Hydrolases (3) acting on glycosyl compounds (3.2) hydrolysing O- and S- glycosyl compounds (3.2.1)
- C12N9/2405—Glucanases
- C12N9/2434—Glucanases acting on beta-1,4-glucosidic bonds
- C12N9/2437—Cellulases (3.2.1.4; 3.2.1.74; 3.2.1.91; 3.2.1.150)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P7/00—Preparation of oxygen-containing organic compounds
- C12P7/02—Preparation of oxygen-containing organic compounds containing a hydroxy group
- C12P7/04—Preparation of oxygen-containing organic compounds containing a hydroxy group acyclic
- C12P7/06—Ethanol, i.e. non-beverage
- C12P7/08—Ethanol, i.e. non-beverage produced as by-product or from waste or cellulosic material substrate
- C12P7/10—Ethanol, i.e. non-beverage produced as by-product or from waste or cellulosic material substrate substrate containing cellulosic material
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02E—REDUCTION OF GREENHOUSE GAS [GHG] EMISSIONS, RELATED TO ENERGY GENERATION, TRANSMISSION OR DISTRIBUTION
- Y02E50/00—Technologies for the production of fuel of non-fossil origin
- Y02E50/10—Biofuels, e.g. bio-diesel
Landscapes
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Organic Chemistry (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Genetics & Genomics (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biotechnology (AREA)
- Microbiology (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Engineering & Computer Science (AREA)
- Medicinal Chemistry (AREA)
- Biomedical Technology (AREA)
- Tropical Medicine & Parasitology (AREA)
- Virology (AREA)
- Cell Biology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Molecular Biology (AREA)
- Processing Of Solid Wastes (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention discloses an obtaining method of a salt-tolerant cellulose decomposition bacteria colony. The obtaining method comprises the following steps: S1, adding filter paper, straws and microcrystalline cellulose into a culture solution, adding a mixed bacteria source into the culture solution, carrying out screening, carrying out next-generation transfer-inoculation when the filter paper has disintegration and the straws become soft and have spots, and carrying out subculture for 20-30 generations; and S2, adding the mixed bacteria colony obtained through screening in the S1 into a culture solution with the salinity being 1% (w/v), adding filter paper and straws into the culture solution, when the filter paper has the disintegration and the straws become soft and have spots, taking the bacterial suspension, inoculating the bacterial suspension to a culture solution with the salinity being 2% (w/v), continuously increasing the salinity of the culture solution till the salinity reaches 5% (w/v), and carrying out subculture and domestication for 15-20 generations to obtain the salt-tolerant cellulose decomposition bacteria colony. The salt-tolerant cellulose decomposition bacteria colony obtained by the method provided by the invention can be applied to the fermentation of organic refuse, therefore, the purpose of improving the fermentation efficiency is improved, the yield of the product is increased by 10-50% compared with that obtained when only fermentation bacteria colony is adopted, and finally, the resource utilization ratio of the organic refuse is effectively improved.
Description
Technical field
The invention belongs to microorganism field and field of environment protection, particularly, relate to a kind of preparation method and application thereof of salt tolerant cellulose decomposition flora.
Background technology
At present, the process of organic waste is disposed and has been become must needing of social development.Organic waste mainly comprises the agriculture and forestry organic waste material such as changing food waste and fruits and vegetables rubbish, straw.These rubbish contain a large amount of lignocelluloses, and lignocellulose, is difficult to be decomposed by the microorganisms by the interaction such as hydrogen bond and covalent linkage weave in by xylogen, Mierocrystalline cellulose and hemicellulose, thus reduces the resource utilization efficiency of organic waste.Therefore, how effectively the Mierocrystalline cellulose of degrading in organic waste is the key improving organic waste resource utilization rate, and the effect giving full play to the microorganism with ligocellulose degradation's function is one of important channel realizing above-mentioned purpose.
Summary of the invention
For the deficiencies in the prior art, the invention provides a kind of preparation method of salt tolerant cellulose decomposition flora.Described method using ocean, salt lake, saltern, hot spring, paper mill sludge, sewage plant sludge and cow dung carry out screening and the salt tolerance domestication of cellulose decomposition flora as bacterium source, obtain the mixed bacterial that lignocellulose degradation ability is stable.
Another object of the present invention is to provide the salt tolerant cellulose decomposition flora and salt tolerant cellulase thereof that obtain according to aforesaid method.
Another object of the present invention is to the application that above-mentioned salt tolerant cellulose decomposition flora and salt tolerant cellulase thereof are provided.
Above-mentioned purpose of the present invention is achieved by the following technical programs.
A preparation method for salt tolerant cellulose decomposition flora, comprises the steps:
S1. filter paper, stalk and Microcrystalline Cellulose are added in sterile medium, add mixed bacterium source again to screen, there is disintegration at filter paper, stalk carry out when softening and occur spot the next generation switching, in succeeding transfer culture 20 ~ 30 generation, obtain the mixed bacterial with lignocellulose degradation ability;
S2. salt tolerance domestication: it is in the sterile medium of 1% (w/v) that the mixed bacterial screened by S1 joins salinity, and adds filter paper and stalk in this sterile medium; When being immersed in that disintegration appears in the filter paper in nutrient solution, stalk softens and occur spot, get bacteria suspension to receive in the aseptic culture fluid that fresh salinity is 2% (w/v) with 10% (v/v), so improve constantly the salinity to 5% (w/v) in sterile medium; In succeeding transfer culture 15 ~ 20 generations of domestication, obtain salt tolerant cellulose decomposition flora;
Wherein, mixed bacterium source described in S1 is the blend sample from one or more in ocean, salt lake, saltern, hot spring, paper mill sludge, sewage plant sludge or cow dung.
Contriver's random selecting is in a large number from the blend sample of one or more in ocean, salt lake, saltern, hot spring, paper mill sludge, sewage plant sludge or cow dung, and the probability obtaining salt tolerant cellulose decomposition flora according to described method reaches 50 ~ 90%.Wherein, when sample be ocean, salt lake, saltern bed mud and paper mill sludge mixture time, the probability obtaining salt tolerant cellulose decomposition flora reaches 80 ~ 90%; When sample is ocean, salt lake, saltern, hot spring bed mud mixture, the probability obtaining salt tolerant cellulose decomposition flora reaches 70 ~ 80%; When sample is paper mill sludge, sewage plant sludge and cow dung mixture, the probability obtaining salt tolerant cellulose decomposition flora reaches 50 ~ 70%.The present invention proves the function-stable of the mixed bacterial lignocellulose degradation obtained by the experiment of degrade filter paper and stalk, illustrates that the method can be reused in production.
In above-mentioned screening and domestication process, the present invention can enrich substrate component as carbon source using filter paper, straw and Microcrystalline Cellulose simultaneously, increases cellulose decomposition flora and obtains probability.In addition, screening index, except filter paper disintegration, also add straw spot, and the flora obtained more effectively can utilize various lignocellulose in organic waste.When salt tolerance is tamed, salt content is set to 1-5%, and the microorganism obtained can meet the requirement of changing food waste salinity.Too high salt branch causes the bacterial classification that effectively can utilize changing food waste substrate in flora in a large number dead, and the bacterial classification of surviving may produce antagonistic action with functional flora, and the product yield that changing food waste is fermented reduces.
Preferably, mixed bacterium source, 5g ocean is added in every 100mL sterile medium in S1,1g filter paper, 1g stalk and 1g Microcrystalline Cellulose.
Preferably, in S1, each succeeding transfer culture microbe inoculation amount is 10% (v/v) of nutrient solution.
Preferably, the composition of sterile medium described in S1, S2 is: peptone 5g/L, yeast extract paste 5g/L, filter paper 5g/L, NaCl5g/L, CaCO
32g/L, K
2hPO
40.5g/L, MgSO
40.5g/L, trace element solution 0.5mL/L.
Preferably, described trace element solution composition is: ZnSO
40.3g/L; CaCl
20.25g/L; CuSO
40.25g/L; FeSO
40.2g/L.
Preferably, the culture condition of the cellulose decomposition of salt tolerant described in S2 flora in procurement process in sterile medium is 28 ~ 50 DEG C, quiescent culture.
Preferably, when described in S1, mixed bacterium source is Marine Bacteria source, be ocean bed mud, shellfish and marine plant mixture; When described mixed bacterium source is bacterium source, salt lake, be salt lake bed mud and salt lake plant mixture; When described mixed bacterium source is bacterium source, saltern, be saltern mud and periphery growing plants mixture.
The present invention also provides the salt tolerant cellulose decomposition flora and salt tolerant cellulase thereof that obtain according to aforesaid method.
The present invention also provides above-mentioned salt tolerant cellulose decomposition flora and salt tolerant cellulase thereof promoting the application in organic refuse fermentation.
Preferably, adopt the coculture of function bacterial classification and described salt tolerant cellulose decomposition flora, or the fermented mixture organic waste of the salt tolerant cellulase of function bacterial classification and the generation of described salt tolerant cellulose decomposition flora; Described function bacterial classification is microbiological deterioration flora, ethanol fermentation flora, lactic fermentation flora, acetic acid fermentation flora or methane fermentation flora.Described function bacterial classification is preferably mixed bacterial, different according to production object, selects corresponding functional flora; As during to produce for the purpose of fertilizer, functional flora selects microbiological deterioration flora; During to produce for the purpose of ethanol, functional flora selects ethanol fermentation flora; During to produce for the purpose of lactic acid, functional flora selects lactic fermentation flora; During to produce for the purpose of acetic acid, functional flora selects acetic acid fermentation flora; During to produce for the purpose of biogas, functional flora selects methane fermentation flora.
The preparation method of described salt tolerant cellulase is ordinary method, is preferably: get Efficient salt-tolerant cellulose-decomposing bacterium group fermentation liquid in 4 DEG C, the centrifugal 10min of 6000rpm, gained supernatant liquor is crude enzyme liquid; Salt tolerant cellulase is obtained to saltout concentrated, dialysis desalting of crude enzyme liquid.
When the fermented bacterium in organic refuse fermentation process is the coculture of function bacterial classification and described salt tolerant cellulose decomposition flora, described coculture adopts simultaneously or the mode of feeding function bacterial classification and salt tolerant cellulose decomposition flora successively.Preferably, when adding, the inoculum size of described function bacterial classification is 1 ~ 20%(v/v simultaneously), the inoculum size of described salt tolerant cellulose decomposition flora is 1 ~ 15%(v/v); When adding successively, the inoculum size of described function bacterial classification is 1 ~ 15%(v/v), the inoculum size of described salt tolerant cellulose decomposition flora is 1 ~ 10%(v/v).More preferably, when adding, the inoculum size of described function bacterial classification is 10%(v/v simultaneously), the inoculum size of described salt tolerant cellulose decomposition flora is 3%(v/v); When adding successively, the inoculum size 8%(v/v of described function bacterial classification), the inoculum size of described salt tolerant cellulose decomposition flora is 5%(v/v).
When the fermented mixture organic waste of the salt tolerant cellulase adopting function bacterial classification and described salt tolerant cellulose decomposition flora to produce, described mixture adopts simultaneously or the mode of feeding function bacterial classification and salt tolerant cellulase successively.Preferably, when adding, the inoculum size of described function bacterial classification is 1 ~ 25%(v/v simultaneously), the inoculum size of described salt tolerant cellulase is 0.1 ~ 5%(v/v); When adding successively, the inoculum size of described function bacterial classification is 1 ~ 15%(v/v), the inoculum size of described salt tolerant cellulase is 0.1 ~ 10%(v/v).More preferably, when adding, the inoculum size of described function bacterial classification is 12%(v/v simultaneously), the inoculum size of described salt tolerant cellulase is 0.2%(v/v); When adding successively, the inoculum size of described function bacterial classification is 7%(v/v), the inoculum size of described salt tolerant cellulase is 0.5%(v/v).
During to produce for the purpose of fertilizer, preferably, described organic refuse fermentation condition is 28 ~ 40 DEG C, aerobic cultivation of ventilating.
During to produce for the purpose of the anaerobism products such as ethanol, lactic acid, acetic acid and methane, preferably, described organic refuse fermentation condition is 28 ~ 40 DEG C, leaves standstill Anaerobic culturel.
Preferably, before organic refuse fermentation, adopt microwave, ultrasonic wave and hydrothermal technique combined pretreatment organic waste.Research finds, during by combined pretreatment and fermented bacterium of the present invention and enzyme coupling, can further improve the productive rate of fermentation products obtained therefrom afterwards.More preferably, the power of described microwave is 100w ~ 2500kw, and the time is 2 ~ 120min, and temperature is 90 ~ 250 DEG C; Described hyperacoustic power is 15 ~ 1500w/L, and the treatment time is 5 ~ 60min; The temperature of described hydrothermal treatment consists is 90 ~ 250 DEG C, and the treatment time is 5 ~ 150min.
Preferably, described organic waste is one or more in organic moiety, agricultural rubbish or municipal sludge in changing food waste, domestic refuse.Preferably, described agricultural rubbish is fruits and vegetables rubbish or stalk.
If described organic waste is changing food waste, preferably, before its fermentation, meal kitchen grease is separated.
Compared with prior art, the present invention has following beneficial effect:
(1) the present invention carries out the screening of salt tolerant cellulose decomposition flora with ocean, salt lake, saltern, hot spring, paper mill sludge, sewage plant sludge and cow dung bacterium source and carries out salt tolerance domestication, prove that this mixed bacterial has the function of stable lignocellulose degradation by filter paper degradation experiment, the lignocellulose in degradable organic waste.
(2) the present invention is directed to the feature being difficult to utilization in organic waste containing a large amount of lignocellulose, after obtaining salt tolerant cellulose decomposition flora, by itself and functional flora Dual culture, or utilize its salt tolerant cellulase and functional flora acting in conjunction, reach the object improving fermentation efficiency, after fermentation, products obtained therefrom productivity ratio only adopts functional flora to increase by 10 ~ 50%, effectively improves resource utilization rate.The invention provides the novel method solving organic waste environmental pollution and realize its recycling.Prove that the method effectively can improve product yield through fruits and vegetables rubbish and changing food waste degradation experiment, and step is simple, workable.
Accompanying drawing explanation
Fig. 1 is the effect inoculating salt tolerant cellulose decomposition flora Dual culture fermented garbage from kitchen after embodiment 1 first inoculates ethanol bacterium.
Fig. 2 is that embodiment 1 mixed bacterial of resistance to salt cellulose is to the degradation effect of filter paper.
Fig. 3 is the effect that embodiment 2 adds salt tolerant cellulase and ethanol bacterium fermentation fruits and vegetables rubbish simultaneously.
Embodiment
Below in conjunction with specific embodiment, the present invention is described in further details, but embodiment does not limit in any form the present invention.Unless stated otherwise, the present invention adopts reagent, method and apparatus are the art conventional reagent, method and apparatus.
Embodiment 1
A preparation method for salt tolerant cellulose decomposition flora, comprises the steps:
S1. filter paper, stalk and Microcrystalline Cellulose are added in sterile medium, add ocean bed mud, shellfish and marine plant mixture again and carry out first stage screening, carry out the next generation's switching when softening and occur spot at filter paper just disintegration (dissolving), stalk, obtain the mixed bacterial with lignocellulose degradation ability; 5g ocean bed mud, shellfish and marine plant mixture is added, 1g filter paper, 1g stalk and 1g Microcrystalline Cellulose in every 100mL sterile medium; Each subinoculation microbial biomass is 10% (v/v) of sterile medium, in succeeding transfer culture 20 ~ 30 generation, obtains the mixed bacterial with lignocellulose degradation ability;
S2. the flora with cellulose-decomposing ability obtained in step S1 is carried out salt tolerance domestication: it is in the sterile medium of 1% (w/v) that the cellulose decomposition flora that experiment obtains is added to salinity; When being immersed in that disintegration (dissolving) appears in the filter paper in nutrient solution, stalk softens and occurs spot, get bacteria suspension to receive in the aseptic culture fluid that fresh salinity is 2% (w/v) with 10% (v/v), so improve constantly salinity to 5% in nutrient solution (w/v); In succeeding transfer culture domestication 15-20 generation, obtains salt tolerant cellulose decomposition flora.
In step S1, described filter paper is commercially available qualitative filter paper; Described stalk drying is to constant weight; Described Microcrystalline Cellulose is commercially available Microcrystalline Cellulose.Described in step S1, S2, the composition of sterile medium is (L
-1): peptone 5g; Yeast extract paste 5g; Filter paper 5g; NaCl5g; CaCO
32g; K
2hPO
40.5g; MgSO
40.5g; Trace element solution 0.5mL.Described trace element solution composition is (L
-1): ZnSO
40.3g; CaCl
20.25g; CuSO
40.25g; FeSO
40.2g.
Described salt tolerant cellulose decomposition flora culture condition is 30 DEG C, quiescent culture.
The cellulose decomposition flora that S1 obtains to the degradation effect of filter paper as shown in Figure 2.
Salt tolerant cellulose decomposition flora obtained above is applied to the method improving organic refuse fermentation efficiency, specifically comprises the steps:
(1) described organic waste is changing food waste, removes the foreign material such as chopsticks, plastics bag in changing food waste, changing food waste is carried out successively microwave, ultrasonic wave and hydrothermal treatment consists; The power of described microwave is 900kw, and the treatment time is 15min; Described hyperacoustic power is 1000w/L, and the treatment time is 10min; Hydrothermal treatment consists temperature is 160 DEG C, and the treatment time is 50min; Above-mentioned process is carried out in the integration apparatus that microwave generator, ultrasonic generator and hydrothermal device are housed;
(2) meal kitchen grease is separated;
(3) step (2) remaining solids in add 10%(v/v) ethanol mixed bacterial anaerobically fermenting at 30 DEG C; After 24 hours, add 3%(v/v) salt tolerant cellulose decomposition flora; Can reach the maximum value of alcohol yied when the 4th day, than Zao 2 days of the time only inoculating ethanol bacterium and reach alcohol yied maximum value, and Dual culture gained alcohol yied is than only inoculating ethanol bacterium high (as shown in Figure 1).
Embodiment 2
The present embodiment key step is identical with embodiment 1, difference is that the bacterium source added in the present embodiment S1 is salt lake bed mud, salt lake plant and sewage plant sludge mixture, is applied to by salt tolerant cellulose decomposition flora obtained above and improves in organic refuse fermentation efficiency.In the present embodiment step (1), organic waste is fruits and vegetables rubbish; Without step (2), replace salt tolerant cellulose decomposition flora to add salt tolerant cellulase in step (3), the dosing method of ethanol mixed bacterial and salt tolerant cellulase is for add simultaneously.Ethanol bacterium inoculum size is 7%(v/v), salt tolerant cellulase inoculum size is 0.5%(v/v); 30 DEG C of Anaerobic culturel.Wherein, the preparation method of salt tolerant cellulase is as follows: get the fermentation liquid of the salt tolerant cellulose decomposition flora of acquisition in 4 DEG C, the centrifugal 10min of 6000rpm, gained supernatant liquor is crude enzyme liquid; Salt tolerant cellulase is obtained to saltout concentrated, dialysis desalting of crude enzyme liquid.Fermentation results as shown in Figure 3.Result shows, the present embodiment gained alcohol yied is higher than only inoculating ethanol bacterium, illustrates and adds the alcohol yied that salt tolerant cellulase can improve fruits and vegetables rubbish fermentation.
Comparative example 1
The acquisition of this comparative example salt tolerant cellulose decomposition flora and application method thereof are substantially identical with embodiment 1, difference is, in the method for this comparative example salt tolerant cellulose decomposition flora, S2 is: it is in the sterile medium of 6% (w/v) that the cellulose decomposition flora that experiment obtains is added to salinity; When being immersed in that disintegration (dissolving) appears in the filter paper in nutrient solution, stalk softens and occurs spot, get bacteria suspension to receive in the aseptic culture fluid that fresh salinity is 7% (w/v) with 10% (v/v), so improve constantly salinity to 10% in nutrient solution (w/v); In succeeding transfer culture domestication 15-20 generation, obtains salt tolerant cellulose decomposition flora.
This comparative example is obtained the salt tolerant cellulose decomposition flora that salt tolerant cellulose decomposition flora alternate embodiment 1 obtains, be added in changing food waste solids and ferment.Ferment the 4th day alcohol yied 6.3%(v/v fewer than embodiment 1), though its reason may be the salt tolerant Mierocrystalline cellulose flora that this comparative example obtains can tolerate 10% (w/v) high density NaCl, but because salinity increase in nutrient solution makes the bacterial classification that effectively can utilize changing food waste substrate in flora in a large number dead, the bacterial classification with cellulose decomposition function of simultaneously surviving and ethanol mixed bacterial produce antagonistic action, thus decrease the productive rate of kitchen waste ethanol fermentation.
Claims (10)
1. a preparation method for salt tolerant cellulose decomposition flora, is characterized in that, comprises the steps:
S1. filter paper, stalk and Microcrystalline Cellulose are added in sterile medium, add mixed bacterium source again to screen, there is disintegration at filter paper, stalk carry out when softening and occur spot the next generation switching, in succeeding transfer culture 20 ~ 30 generation, obtain the mixed bacterial with lignocellulose degradation ability;
S2. salt tolerance domestication: it is in the sterile medium of 1% (w/v) that the mixed bacterial screened by S1 joins salinity, and adds filter paper and stalk in this sterile medium; When being immersed in that disintegration appears in the filter paper in nutrient solution, stalk softens and occur spot, get bacteria suspension to receive in the aseptic culture fluid that fresh salinity is 2% (w/v) with 10% (v/v), so improve constantly the salinity to 5% (w/v) in sterile medium; In succeeding transfer culture 15 ~ 20 generations of domestication, obtain salt tolerant cellulose decomposition flora;
Wherein, mixed bacterium source described in S1 is the blend sample from one or more in ocean, salt lake, saltern, hot spring, paper mill sludge, sewage plant sludge or cow dung.
2. preparation method according to claim 1, is characterized in that, adds 5g mixed bacterium source, 1g filter paper, 1g stalk and 1g Microcrystalline Cellulose in S1 in every 100mL sterile medium.
3. preparation method according to claim 1, is characterized in that, in S1, each succeeding transfer culture microbe inoculation amount is 10% (v/v) of nutrient solution.
4. preparation method according to claim 1, is characterized in that, the culture condition of the cellulose decomposition of salt tolerant described in S2 flora in screening process in sterile medium is 28 ~ 50 DEG C, quiescent culture.
5. preparation method according to claim 1, is characterized in that, when described in S1, mixed bacterium source is Marine Bacteria source, is ocean bed mud, shellfish and marine plant mixture; When described mixed bacterium source is bacterium source, salt lake, be salt lake bed mud and salt lake plant mixture; When described mixed bacterium source is bacterium source, saltern, be saltern mud and periphery growing plants mixture.
6. the salt tolerant cellulose decomposition flora that obtains of preparation method described in any one of claim 1 ~ 5 and enzyme thereof.
7. salt tolerant cellulose decomposition flora and enzyme thereof the application in organic refuse fermentation described in claim 6.
8. application according to claim 7, it is characterized in that, adopt the coculture of function bacterial classification and described salt tolerant cellulose decomposition flora, or the fermented mixture organic waste of the salt tolerant cellulase of function bacterial classification and the generation of described salt tolerant cellulose decomposition flora; Described function bacterial classification is microbiological deterioration flora, ethanol fermentation flora, lactic fermentation flora, acetic acid fermentation flora or methane fermentation flora.
9. application according to claim 8, it is characterized in that, when fermented bacterium is the coculture of function bacterial classification and described salt tolerant cellulose decomposition flora, described coculture adopts simultaneously or the mode of feeding function bacterial classification and salt tolerant cellulose decomposition flora successively; When adding, the inoculum size of described function bacterial classification is 1 ~ 20%(v/v simultaneously), the inoculum size of described salt tolerant cellulose decomposition flora is 1 ~ 15%(v/v); When adding successively, the inoculum size of described function bacterial classification is 1 ~ 15%(v/v), the inoculum size of described salt tolerant cellulose decomposition flora is 1 ~ 10%(v/v).
10. application according to claim 8, it is characterized in that, when the fermented mixture organic waste of the salt tolerant cellulase adopting function bacterial classification and described salt tolerant cellulose decomposition flora to produce, described mixture adopts simultaneously or the mode of feeding function bacterial classification and salt tolerant cellulase successively; When adding, the inoculum size of described function bacterial classification is 1 ~ 25%(v/v simultaneously), the inoculum size of described salt tolerant cellulase is 0.1 ~ 5%(v/v); When adding successively, the inoculum size of described function bacterial classification is 1 ~ 15%(v/v), the inoculum size of described salt tolerant cellulase is 0.1 ~ 10%(v/v).
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610007623.0A CN105524835A (en) | 2016-01-07 | 2016-01-07 | Obtaining method and application of salt-tolerant cellulose decomposition bacteria colony |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610007623.0A CN105524835A (en) | 2016-01-07 | 2016-01-07 | Obtaining method and application of salt-tolerant cellulose decomposition bacteria colony |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN105524835A true CN105524835A (en) | 2016-04-27 |
Family
ID=55767366
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201610007623.0A Pending CN105524835A (en) | 2016-01-07 | 2016-01-07 | Obtaining method and application of salt-tolerant cellulose decomposition bacteria colony |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN105524835A (en) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107779424A (en) * | 2017-11-10 | 2018-03-09 | 中国科学院沈阳应用生态研究所 | A kind of method and its application that straw degradative fungus strain is screened using stalk as bacterium source |
| WO2021190585A1 (en) * | 2020-03-26 | 2021-09-30 | 中国科学院南京土壤研究所 | Quick isolation method for salt-tolerant cellulose decomposing bacteria |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102952765A (en) * | 2011-08-31 | 2013-03-06 | 中国石油化工股份有限公司 | Enrichment culture method of salt-tolerant efficient nitrosobacteria |
| CN104651293A (en) * | 2015-01-30 | 2015-05-27 | 华南农业大学 | Compound bacteria with function of synergistically degrading lignocelluloses and organic chlorophenol |
-
2016
- 2016-01-07 CN CN201610007623.0A patent/CN105524835A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102952765A (en) * | 2011-08-31 | 2013-03-06 | 中国石油化工股份有限公司 | Enrichment culture method of salt-tolerant efficient nitrosobacteria |
| CN104651293A (en) * | 2015-01-30 | 2015-05-27 | 华南农业大学 | Compound bacteria with function of synergistically degrading lignocelluloses and organic chlorophenol |
Non-Patent Citations (1)
| Title |
|---|
| 杨军 等: "高盐高浓度CMC废水处理工程调试运行", 《环境工程》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107779424A (en) * | 2017-11-10 | 2018-03-09 | 中国科学院沈阳应用生态研究所 | A kind of method and its application that straw degradative fungus strain is screened using stalk as bacterium source |
| WO2021190585A1 (en) * | 2020-03-26 | 2021-09-30 | 中国科学院南京土壤研究所 | Quick isolation method for salt-tolerant cellulose decomposing bacteria |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Wang et al. | Study on the comprehensive utilization of city kitchen waste as a resource in China | |
| Sawyerr et al. | An overview of biogas production: Fundamentals, applications and future research | |
| Liu et al. | Fermentative hydrogen production from macro-algae Laminaria japonica using anaerobic mixed bacteria | |
| Xu et al. | Bioconversion of biowaste into renewable energy and resources: A sustainable strategy | |
| Zhu et al. | Bioenergy from dairy manure: technologies, challenges and opportunities | |
| CN105948853B (en) | Organic fertilizer stack type fermentation method taking mushroom dregs as substrate | |
| CN105002221A (en) | Method for producing biogas through high-efficiency anaerobic fermentation of organic waste resource in intensive vegetable area | |
| CN104630292A (en) | Method for preparing butyric acid by fermenting lignocellulose by using mixed flora | |
| CN105948841B (en) | Organic fertilizer tank type fermentation method taking mushroom dregs as substrate | |
| CN106811438B (en) | Straw degradation acidification microbial inoculum and preparation method thereof | |
| CN102533609A (en) | Methane dry fermentation compound bacteria | |
| Chatellard et al. | Trends and challenges in biohydrogen production from agricultural waste | |
| Mehariya et al. | Bio-based and agriculture resources for production of bioproducts | |
| CN108676821A (en) | A method of utilizing kitchen garbage fermentation and acid | |
| CN105803004A (en) | Method for low-temperature fermentation of biogas from agricultural waste | |
| CN114561327A (en) | Cellulose degradation composite microbial inoculum and preparation method and application thereof | |
| CN114276176A (en) | Harmless treatment method suitable for mixed composting of kitchen waste and breeding waste | |
| CN103602715A (en) | Method for preparing hydrogen from straws | |
| Kumar et al. | Optimization of dry anaerobic fermentation of solid organic wastes | |
| CN110396483B (en) | High-temperature straw degradation bacterium B-8, and microbial inoculum and application thereof | |
| CN101063152A (en) | Kitchen residual garbage normal temperature anaerobic fermentation method | |
| CN105524835A (en) | Obtaining method and application of salt-tolerant cellulose decomposition bacteria colony | |
| CN102363794A (en) | Method for producing hydrogen through kitchen waste enzymolysis and reinforced dark fermentation | |
| Rose et al. | Food industry waste: potential pollutants and their bioremediation strategies | |
| CN103058478B (en) | Method for improving anaerobic digestion performance of sludge and synchronously enlarging cultivation by functional bacteria |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20160427 |
|
| RJ01 | Rejection of invention patent application after publication |