CN110653238A - Method for treating garden waste by using aerobic-anaerobic two-stage process - Google Patents

Method for treating garden waste by using aerobic-anaerobic two-stage process Download PDF

Info

Publication number
CN110653238A
CN110653238A CN201910909408.3A CN201910909408A CN110653238A CN 110653238 A CN110653238 A CN 110653238A CN 201910909408 A CN201910909408 A CN 201910909408A CN 110653238 A CN110653238 A CN 110653238A
Authority
CN
China
Prior art keywords
garden waste
culture
substrate
aerobic
saccharomyces cerevisiae
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
Application number
CN201910909408.3A
Other languages
Chinese (zh)
Inventor
林云琴
郑浩旻
房秀秀
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
South China Agricultural University
Original Assignee
South China Agricultural University
Priority date (The priority date 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 date listed.)
Filing date
Publication date
Application filed by South China Agricultural University filed Critical South China Agricultural University
Priority to CN201910909408.3A priority Critical patent/CN110653238A/en
Publication of CN110653238A publication Critical patent/CN110653238A/en
Pending legal-status Critical Current

Links

Images

Classifications

    • 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
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
    • A01G18/00—Cultivation of mushrooms
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
    • A01G18/00—Cultivation of mushrooms
    • A01G18/20—Culture media, e.g. compost
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B09—DISPOSAL OF SOLID WASTE; RECLAMATION OF CONTAMINATED SOIL
    • B09B—DISPOSAL OF SOLID WASTE NOT OTHERWISE PROVIDED FOR
    • B09B5/00—Operations not covered by a single other subclass or by a single other group in this subclass
    • 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
    • Y02W—CLIMATE CHANGE MITIGATION TECHNOLOGIES RELATED TO WASTEWATER TREATMENT OR WASTE MANAGEMENT
    • Y02W30/00—Technologies for solid waste management
    • Y02W30/50—Reuse, recycling or recovery technologies
    • Y02W30/78—Recycling of wood or furniture waste
    • 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
    • Y02W—CLIMATE CHANGE MITIGATION TECHNOLOGIES RELATED TO WASTEWATER TREATMENT OR WASTE MANAGEMENT
    • Y02W30/00—Technologies for solid waste management
    • Y02W30/50—Reuse, recycling or recovery technologies
    • Y02W30/82—Recycling of waste of electrical or electronic equipment [WEEE]

Landscapes

  • Life Sciences & Earth Sciences (AREA)
  • Mycology (AREA)
  • Environmental Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Environmental & Geological Engineering (AREA)
  • Preparation Of Compounds By Using Micro-Organisms (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)

Abstract

The invention provides a method for treating garden waste by using an aerobic-anaerobic two-stage process. According to the method, the auricularia polytricha is inoculated in a substrate containing wheat grains for culture to obtain cultivated species, the cultivated species is inoculated in garden waste for aerobic culture, efficient selective degradation of lignin is realized, the stubborn structure of lignocellulose is destroyed, cellulose is reserved as much as possible, the selective degradation coefficient of the lignin can reach 2-4, and the selectivity is high; the auricularia polytricha is used as a pretreatment microbial inoculum, so that the microbial source is reliable and easy to obtain, the strains are easy to store, the activation time is short, and the application field of the auricularia polytricha is widened. The method for treating garden waste by using the aerobic-anaerobic two-stage process, disclosed by the invention, has the advantages that the garden waste subjected to aerobic culture treatment is taken as a substrate, cellulase and saccharomyces cerevisiae are added for carrying out anaerobic fermentation to produce ethanol, the ethanol yield can be effectively improved, meanwhile, the conditions are mild, the energy consumption is low, the process is simple, the operability is strong, and the industrial prospect is good.

Description

Method for treating garden waste by using aerobic-anaerobic two-stage process
Technical Field
The invention belongs to the field of renewable energy sources and environmental protection, and particularly relates to a method for treating garden waste by using an aerobic-anaerobic two-stage process.
Background
With the acceleration of the urbanization process and the continuous increase of the green land area of urban gardens, landscaping garbage including dead branches and fallen leaves and landscaping trimmings becomes an important component of urban solid waste, and great pressure is brought to the environment. The treatment of garden waste is mainly concentrated in the modes of landfill, incineration and the like for a long time, the treatment cost is high, great influence is generated on the atmosphere and soil, and the precious biomass resource of garden waste is wasted. The garden waste is a natural lignocellulose raw material which is rich renewable energy substances on the earth, and if the garden waste can be developed and utilized to replace non-renewable resources to a certain extent, the pressure caused by the increasingly reduced non-renewable resources can be weakened, and a larger development space is provided for the cyclic utilization of the resources. How to reasonably and efficiently treat and utilize garden waste becomes a problem to be solved urgently.
At present, garden waste is utilized in the following ways: paper making, garden organic covering preparation, biofuel preparation, mixed composting and the like. But the utilization mode has the problems of large occupied area, long period, secondary pollution, low added value of regenerated products and the like. In addition, the two or more methods are combined for treatment, so that the treatment efficiency can be improved, and the economic benefit can be increased. How to apply the combined treatment method to garden waste becomes a key for realizing the reutilization of the lignocellulose raw material in the garden waste to the maximum extent.
The lack of energy is a serious problem in the development process of human beings, and the reduction of fossil fuels such as petroleum, coal and the like makes people have to step on the long and diffuse way of searching new energy. After the oil crisis in the 70 s of the 20 th century, the biofuel ethanol becomes an ideal product for replacing petrochemical fuels such as gasoline and the like due to the advantages of reproducibility, environmental friendliness, mature technology, convenience in use, easiness in popularization and the like, is concerned by various countries, and is a core of the second-generation bioethanol technology for producing the fuel ethanol by taking crop straws, wood processing residues, garden wastes and the like as main raw materials. However, lignocellulose, which is a main component in garden waste, has a complex structure and is difficult to degrade, so that the biological conversion rate is not high, and resource utilization of the garden waste is restricted to a certain extent.
In order to improve the energy utilization efficiency of the garden waste, the raw materials usually need to be pretreated to destroy the lignocellulose structure and improve the subsequent biological conversion rate. However, the traditional physical and chemical pretreatment methods are high in cost and easy to generate secondary pollution. The biological pretreatment method has mild conditions, easy operation and no secondary pollution, but when the structure of lignocellulose is destroyed by the existing biological pretreatment method, the degradation rate of the cellulose is often higher than that of lignin, which is extremely unfavorable for preparing bioethanol, and the degradation of the cellulose directly causes the reduction of the subsequent bioethanol yield.
With the growing world economy and the increasing population, the problem of energy shortage is becoming more serious, and the development of renewable energy is called a global concern. Ethanol has received considerable attention from governments and many researchers as a clean and renewable energy source. At present, in the production of fuel ethanol in China, saccharides or grains are used as raw materials, and the further expansion of yield is limited by resources, so that the production of ethanol by using cheap and easily available cellulosic raw materials is a necessary trend for future development, and how to break the lignocellulose structure in the cellulosic raw materials and improve the yield of bioethanol is a key research point in the future.
Disclosure of Invention
The invention aims to overcome the defects of the prior art and provide a method for selectively degrading garden waste, which can realize selective degradation of lignin and retain as much cellulose as possible.
The invention also aims to provide application of the method for selectively degrading garden waste.
The invention further aims to provide a method for treating garden waste by using an aerobic-anaerobic two-stage process.
The purpose of the invention is realized by the following technical scheme:
a method for selectively degrading garden waste comprises the following steps:
(1) preparation of cultivars: au781 is inoculated into a sterilized substrate containing wheat grains for culture to obtain a cultivated species;
(2) aerobic culture: adding water into garden waste, mixing to obtain culture substrate, sterilizing, inoculating the said cultivated species, and aerobic culturing.
The auricularia polytricha described in the step (1) is preferably activated before inoculation.
The activating culture medium is preferably a PDA culture medium: 200g of potato, 20.0g of glucose and KH2PO4 3.0g，MgSO4·7H2O1.5 g, vitamin B18mg, agar 25g, distilled water 1000mL, pH natural.
The specific operation of the activation is preferably to culture the mycelia at 28 ℃ until the mycelia are full of the culture medium, transfer the thalli to another culture medium, and continue the culture until the mycelia are full of the culture medium.
The substrate containing wheat grains in the step (1) preferably comprises wheat grains, wood chips and calcium carbonate.
The wheat grains, the wood chips and the calcium carbonate are preferably mixed according to the mass ratio of 94:5: 1.
The wheat grains are preferably pretreated as follows: after being cleaned, the wheat grains are soaked in cold water for 8-10 h, boiled in boiling water for 20-30 min, and drained until the water content is 85-92%.
The wood dust refers to sawdust powder left after wood processing, does not contain wood shavings, and has the water content of 40-50%.
The specific method for preparing the cultivar is preferably as follows: preparing the activated auricularia polytricha and a culture medium into round slices to obtain fungus plugs; the plugs were placed in a container with substrate and cultured until the container was filled with hyphae.
The filling level of the substrate in the container is preferably 70% and the material is not compacted.
The container is preferably a tissue culture bottle.
The circular sheet preferably has a diameter of 0.5cm and can be made by a punch.
The adding amount of the bacterial plugs is preferably 5-7 bacterial plugs per 250mL of substrate.
The culture conditions are that the temperature is 28 ℃ and the humidity is 60%; shaking the mixture once every 24h to make the hyphae grow uniformly.
Preferably, the garden waste in the step (2) is firstly crushed to below 3 cm; the water content of the garden waste is preferably 40-50%.
Calcium carbonate can also be added into the culture substrate in the step (2).
Waste yeast can be added into the culture substrate in the step (2) to serve as a nitrogen source supplement in an aerobic pretreatment stage, so that the activity of the auricularia polytricha can be further improved, the substrate property can be optimized, the waste yeast is generated after main fermentation and after-fermentation brewing processes in the beer production process, and contains nutrients such as proteins, vitamins and trace elements, and the water content is 5-10% after drying treatment.
The garden waste, the waste yeast and the calcium carbonate are preferably mixed according to the mass ratio of (90-95) to (4-9) to 1.
The water is preferably added in the step (2) according to the mass ratio of the materials to the water of 1: 1.
Preferably, the culture substrate in the step (2) is kept still for 8-10 hours before sterilization, so that the culture substrate fully absorbs moisture and the materials are uniform.
The specific operation of the post-sterilization inoculation described in the step (2) is preferably: and bagging the culture substrate according to 250-300 g per bag, sterilizing, cooling, and inoculating cultivated species at two ends of the fungus bag.
The bagging is preferably performed using thick polypropylene high pressure plastic bags.
The amount of inoculum of the cultivar described in step (2) is preferably inoculated at 10% of the wet weight of the culture substrate before sterilization.
The temperature of the aerobic culture condition in the step (2) is preferably 28 ℃; the humidity is preferably 60%; the time for aerobic culture is preferably 20 days.
The sterilization in the steps (1) and (2) is preferably performed at 121 ℃ for 30 min.
By the method for selectively degrading garden waste, the selective degradation coefficient of lignin can reach 2-4, and the selectivity is high; the selective degradation coefficient SV of the lignin is the degradation rate of the lignin/the degradation rate of the cellulose.
The method for selectively degrading garden waste is applied to the preparation of ethanol; because the method can selectively degrade lignin and retain as much cellulose as possible, the garden waste treated by the method can be used for preparing ethanol, and the yield of the ethanol can be effectively improved.
A method for treating garden waste by using an aerobic-anaerobic two-stage process comprises the following steps: the garden waste treated by the method for selectively degrading garden waste is used as a substrate, and cellulase and saccharomyces cerevisiae are added for anaerobic fermentation to produce ethanol (the flow chart of the method is shown in figure 1).
The substrate preferably comprises the garden waste and a nutrient solution, the nutrient solution is a malt juice liquid culture medium, and the formula of the nutrient solution is 5.0g of peptone, 10.0g of glucose, 3.0g of yeast extract, 3.0g of malt extract and 1000mL of distilled water, and the pH is natural.
The mass fraction of dry matters of the substrate in the fermentation container is preferably 5-10%.
Sterilizing the substrate before adding cellulase and saccharomyces cerevisiae; the sterilization is preferably carried out at 121 ℃ for 30 min.
The addition amount of the cellulase is preferably 20-40U/gVS, and further preferably 25-30U/gVS; wherein VS refers to the mass of volatile substances contained in the raw mix.
The saccharomyces cerevisiae is preferably saccharomyces cerevisiae GIM 2.188; the concentration of the saccharomyces cerevisiae bacterial liquid is preferably 108～109cfu/mL。
The saccharomyces cerevisiae liquid is preferably obtained by inoculating saccharomyces cerevisiae into a wort solid culture medium for culture.
The formula of the wort solid culture medium is as follows: 5.0g of peptone, 10.0g of glucose, 3.0g of yeast extract, 3.0g of malt extract, 25.0g of agar powder and 1000mL of distilled water, and the pH is natural.
The culture is preferably carried out for 12-18 h at the temperature of 28 +/-1 ℃ and 150 rev/min.
The inoculation amount of the saccharomyces cerevisiae bacterial liquid is preferably 5-10% of the total volume of the substrate.
The anaerobic fermentation condition is preferably 28 +/-1 ℃ and 150 rev/min; the time of anaerobic fermentation is preferably 7-10 days, and the concentration of ethanol in the fermentation container is basically unchanged.
The invention utilizes the defects that garden wastes are difficult to be efficiently utilized in the prior art, and the agaric fungi are utilized for aerobic pretreatment to selectively degrade lignin, destroy the stubborn structure of lignocellulose, reduce the degradation of the cellulose as far as possible, and realize high yield of ethanol while reducing the harm of waste treatment.
Compared with the prior art, the invention has the following advantages and effects:
1. the method utilizes fungus to perform aerobic pretreatment on garden waste under the condition of medium temperature, the pretreatment process does not need to be performed under the aseptic condition, the condition is mild, the process does not need to be turned over, the energy consumption is low, the process is simple, and the operability is strong.
2. The invention uses the fungus as the pretreatment microbial inoculum, has reliable and easily obtained bacterial source, easy preservation of the strain and short activation time, and widens the application field of the fungus.
3. The method realizes the degradation of lignocellulose in garden waste in the pretreatment process, wherein the lignin degradation rate is higher than that of the cellulose, and the purposes of selectively degrading lignin, reserving as much cellulose as possible for subsequent ethanol fermentation and improving the ethanol yield are achieved.
4. According to the invention, a small amount of waste yeast is used as a nitrogen source supplement in an aerobic pretreatment stage, so that the activity of fungus agaric is improved, the high-efficiency degradation of lignocellulose is realized, the environmental pollution of the waste yeast is reduced, and the resource utilization of the waste yeast is realized.
5. The invention realizes the 'harmlessness, reduction and recycling' of garden wastes, has potential environmental, social and economic benefits, and has huge market and application potentials in the aspects of renewable energy production, waste resource conversion and utilization and solid waste biological treatment technology.
Drawings
FIG. 1 is a schematic view of the process for treating garden waste by the aerobic-anaerobic two-stage process of the present invention.
Detailed Description
The present invention will be described in further detail with reference to examples and drawings, but the present invention is not limited thereto.
Example 1
(1) The garden waste for the test is taken from southern China agricultural university, the taken fresh dry branches trimmed in the campus are uniformly air-dried, the moisture content is 40-50% after the air-drying, the fresh dry branches are crushed into wood sections of 30-40 cm by using saws, and finally the wood sections are crushed to 0-3 cm by using a hammer mill.
(2) The waste yeast for the test is taken from Zhujiang beer GmbH of Guangzhou city, and the waste yeast is produced by main fermentation and after-fermentation brewing processes in the production process of the beer, contains nutrients such as protein, vitamins and trace elements, and is dried, so that the water content is 5-10%. Auricularia polytricha (Mont.) Sacc. Auricularia (Auricularia polytricha ) with the number of Au781 is purchased from Guangdong province microorganism strain collection center, and the Auricularia polytricha with the number of GIMCC5.174 is selected from the microorganism strain library.
(3) Preparation of PDA Medium (Potato 200g, glucose 20.0g, KH)2PO4 3.0g，MgSO4·7H2O1.5 g, vitamin B18mg, 25g of agar, 1000mL of distilled water and natural pH), transferring the agaric hyphae in the frozen tube to a PDA culture medium by using a plate-scribing method, culturing the solid plate with the strain at 28 ℃ for about 7 days (taking hypha overgrowing the culture medium as a standard), repeating the steps, transferring the thalli to the other solid plate again, and continuously culturing for 7 days to realize thalli activation.
(4) Selecting commercially available wheat grains, removing shrunken grains and impurities, cleaning with clear water, soaking in cold water for 8 hr, boiling in boiling water for 25min, and draining to water content of 88.9%. Measuring the water content of the sawdust according to the following steps: wood chip: mixing calcium carbonate 94:5:1 (by mass), placing into 250mL tissue culture bottle (with no material compaction, 70% filling amount), sterilizing at 121 deg.C for 30min, scattering wheat grains, and cooling to obtain culture substrate.
The wood dust refers to sawdust powder left after wood processing, does not contain wood shavings, and has a water content of 40-50%; calcium carbonate is commercially available technical grade.
(5) Cutting the activated agaric and the culture medium into round sheets by using a puncher with the diameter of 0.5cm to prepare fungus plugs, inoculating 5 fungus plugs into each tissue culture bottle, and uniformly distributing the fungus plugs in the tissue culture bottles filled with the culture seed substrates obtained in the step (4). And (3) transferring the inoculated tissue culture bottle into an artificial climate box, shaking up the culture material in the tissue culture bottle once every 24 hours at the temperature of 28 ℃ and the humidity of 60% to ensure that hyphae uniformly grow in the bottle, and obtaining the cultivated species after the hyphae overgrow the tissue culture bottle.
(6) Uniformly mixing garden waste, waste yeast and calcium carbonate according to the mass ratio of 95:4:1, adding water until the material-water ratio is 1:1, placing in a 20L plastic barrel for 8 hours, then filling the materials into thick polypropylene high-pressure plastic bags according to the dosage of 250g per bag, compacting, and sterilizing for 30min at 121 ℃ by using an autoclave. And (3) cooling the sterilized fungus bags on an ultraclean workbench, and inoculating cultivated species at two ends of the fungus bags, wherein the inoculated cultivated species account for 10% of the mass of the fungus bag mixture before sterilization (calculated on a wet basis). The inoculated fungus bags are placed in an artificial climate box regularly, the temperature is 28 ℃, the humidity is 60 percent, and the standing culture is carried out for 20 days.
(7) After the cultivation, the content of lignin and cellulose in the garden waste before and after the treatment was measured by the american NREL method (slave, et al.2008). The degradation rate of lignin in the material is 16.7%, the degradation rate of cellulose is 7.3%, and the selective degradation coefficient of lignocellulose is 2.29.
(8) Preparing a wort solid culture medium, wherein the formula is as follows: 5.0g of peptone, 10.0g of glucose, 3.0g of yeast extract, 3.0g of malt extract, 25.0g of agar powder and 1000mL of distilled water, and the pH is natural. Inoculating Saccharomyces cerevisiae (purchased from Guangdong province microorganism culture Collection, and numbered as GIM2.188 in microorganism culture library) in a freezing tube to a wort solid culture medium by streaking, inoculating yeast mycelium to the wort liquid culture medium (except not containing agar, the other formula is the same as the wort solid culture medium), and culturing at 28 + -1 deg.C and 150rev/min for 16h to obtain Saccharomyces cerevisiae liquid.
(9) Putting the materials cultured in the step (6) into a 250mL fermentation bottle, adding a malt wort liquid culture medium as a nutrient solution, ensuring that the mass fraction of dry matters of the materials in the fermentation bottle is 5%, and sterilizing the fermentation bottle at 121 ℃ for 30min under high pressure; adding cellulase (green xylanase, enzyme activity greater than 15U/mg, purchased from Shanghai Bo and ao Biotech Co., Ltd.) into the sterilized fermentation bottle at an amount of 30U/gVS, wherein VS refers to the mass of volatile substances contained in the raw material mixture; then adding the mixture with the concentration of 108cfu/mL of saccharomyces cerevisiae bacterial liquid, wherein the inoculation amount of the saccharomyces cerevisiae bacterial liquid is 10% (V/V in volume) of the total volume of the fermentation bottle material.
(10) And (3) placing the fermentation bottle at the temperature of 28 +/-1 ℃ and under the condition of 150rev/min, and culturing for 9 days until the ethanol concentration in the fermentation bottle is basically unchanged. The final ethanol concentration in the fermentation bottle was 181.4 g/L. Wherein, the ethanol concentration is determined by potassium dichromate oxidation spectrophotometry (see sandai, research on tubular membrane integration technology for preparing bioethanol by cellulose fermentation, Master graduate thesis of Tianjin university, 2015, "determination of ethanol content of 3.3.2").
Example 2
(1) The garden waste for the test is taken from southern China agricultural university, fresh dry branches trimmed in a campus are taken back to be uniformly air-dried, then the dry branches are crushed into wood segments of 30-40 cm by using saws, and finally the wood segments are crushed to 0-3 cm by using a hammer mill.
(2) The waste yeast for test was obtained from Zhujiang beer GmbH, Guangzhou city, and Auricularia polytricha (Mont.) was Auricularia polytricha (Auricularia polytricha ) and numbered Au 781.
(3) Preparation of PDA Medium (Potato 200g, glucose 20.0g, KH)2PO4 3.0g，MgSO4·7H2O1.5 g, vitamin B18mg, agar 25g, distilled water 1000mL, pH Natural), plate-streakingTransferring the fungus hypha in the freezing tube to a PDA culture medium, culturing the solid plate with the fungus at 28 ℃ for about 7 days (based on the fact that the fungus hypha grows over the culture medium), repeating the steps, transferring the fungus to the other solid plate again, and continuously culturing for 7 days to realize the activation of the fungus.
(4) Selecting commercially available wheat grains, removing shrunken grains and impurities, cleaning with clear water, soaking in cold water for 8 hr, boiling in boiling water for 30min, and draining to water content of 91.7%. Measuring the water content of the sawdust according to the following steps: wood chip: mixing calcium carbonate 94:5:1 (by mass), placing into 250mL tissue culture bottle (with no material compaction, 70% filling amount), sterilizing at 121 deg.C for 30min, scattering wheat grains, and cooling to obtain culture substrate.
(5) Cutting the activated agaric and the culture medium into round sheets by using a puncher with the diameter of 0.5cm to prepare fungus plugs, inoculating 7 fungus plugs into each tissue culture bottle, and uniformly distributing the fungus plugs in the tissue culture bottles filled with the culture seed substrates obtained in the step (4). And (3) transferring the inoculated tissue culture bottle into an artificial climate box, shaking up the culture material in the tissue culture bottle once every 24 hours at the temperature of 28 ℃ and the humidity of 60% to ensure that hyphae uniformly grow in the bottle, and obtaining the cultivated species after the hyphae overgrow the tissue culture bottle.
(6) Uniformly mixing garden waste, waste yeast and calcium carbonate according to a mass ratio of 90:9:1, adding water until the material-water ratio is 1:1, placing the mixture in a 20L plastic barrel for 8-10 hours, then filling the materials into thick polypropylene high-pressure plastic bags according to the amount of 250g per bag, compacting, and sterilizing for 30min at 121 ℃ by using an autoclave. And (3) cooling the sterilized fungus bags on an ultraclean workbench, and inoculating cultivated species at two ends of the fungus bags, wherein the inoculated cultivated species account for 10% of the mass of the fungus bag mixture before sterilization (calculated on a wet basis). The inoculated fungus bags are placed in an artificial climate box regularly, the temperature is 28 ℃, the humidity is 60 percent, and the standing culture is carried out for 20 days.
(7) After the culture is finished, the degradation rate of lignin in the material is 22.3%, the degradation rate of cellulose is 7.1%, and the selective degradation coefficient of lignocellulose is 3.14.
(8) Preparing a wort solid culture medium, wherein the formula is as follows: 5.0g of peptone, 10.0g of glucose, 3.0g of yeast extract, 3.0g of malt extract, 25.0g of agar powder and 1000mL of distilled water, and the pH is natural. And (3) carrying out streak transfer on the saccharomyces cerevisiae GIM2.188 in the frozen tube to a wort solid culture medium, after 3 times of transfer, selecting yeast hyphae to inoculate to the wort liquid culture medium, and culturing for 16h at the temperature of 28 +/-1 ℃ and 150rev/min to obtain saccharomyces cerevisiae bacterial liquid.
(9) Putting the materials cultured in the step (6) into a 250mL fermentation bottle, adding a malt wort liquid culture medium as a nutrient solution, ensuring that the mass fraction of dry matters of the materials in the fermentation bottle is 8%, and sterilizing the fermentation bottle at 121 ℃ for 30min under high pressure; adding cellulase (same as example 1) into the sterilized fermentation bottle in an amount of 25U/gVS, wherein VS refers to the mass of volatile substances contained in the raw material mixture; then adding the mixture with the concentration of 108cfu/mL of saccharomyces cerevisiae bacterial liquid, wherein the inoculation amount of the saccharomyces cerevisiae bacterial liquid is 10% (V/V in volume) of the total volume of the fermentation bottle material.
(10) And (3) culturing the fermentation bottle for 10 days at the temperature of 28 +/-1 ℃ and 150rev/min, wherein the final ethanol concentration in the fermentation bottle is 207.5 g/L.
Example 3
(1) The garden waste for the test is taken from southern China agricultural university, fresh dry branches trimmed in a campus are taken back to be uniformly air-dried, then the dry branches are crushed into wood segments of 30-40 cm by using saws, and finally the wood segments are crushed to 0-3 cm by using a hammer mill.
(2) The waste yeast for test is obtained from Zhujiang beer GmbH of Guangzhou city, Pleurotus ostreatus with the number of P40 is selected from Pleurotus ostreatus, and purchased from Guangdong province microorganism strain collection center, and the number of the microorganism strain library is GDMCC 5.539.
(3) Preparation of PDA Medium (Potato 200g, glucose 20.0g, KH)2PO4 3.0g，MgSO4·7H2O1.5 g, vitamin B18mg, 25g of agar and 1000mL of distilled water, and natural pH), transferring the oyster mushroom hyphae in the frozen tube to a PDA culture medium by using a plate-scribing method, culturing the solid plate with the strain at 28 ℃ for about 7 days (taking the hyphae to overgrow the culture medium as a standard), repeating the steps, transferring the thallus to another solid plate again, and continuously culturing for 7 days to realize the living thallusAnd (4) transforming.
(4) Selecting commercially available wheat grains, removing shrunken grains and impurities, cleaning with clear water, soaking in cold water for 8 hr, boiling in boiling water for 30min, and draining to water content of 91.7%. Measuring the water content of the sawdust according to the following steps: wood chip: mixing calcium carbonate 94:5:1 (by mass), placing into 250mL tissue culture bottle (with no material compaction, 70% filling amount), sterilizing at 121 deg.C for 30min, scattering wheat grains, and cooling to obtain culture substrate. The water content of the sawdust is 40-50%.
(5) Cutting the activated oyster mushroom and the culture medium into round slices by using a puncher with the diameter of 0.5cm to prepare the mushroom plugs, inoculating 5 mushroom plugs into each tissue culture bottle, and uniformly distributing the mushroom plugs in the tissue culture bottles filled with the culture seed substrates obtained in the step (4). And (3) transferring the inoculated tissue culture bottle into an artificial climate box, shaking up the culture material in the tissue culture bottle once every 24 hours at the temperature of 28 ℃ and the humidity of 60% to ensure that hyphae uniformly grow in the bottle, and obtaining the cultivated species after the hyphae overgrow the tissue culture bottle.
(6) Uniformly mixing garden waste, waste yeast and calcium carbonate according to the mass ratio of 95:4:1, adding water until the material-water ratio is 1:1 (by mass), placing the mixture in a 20L plastic barrel for 8-10 hours, then filling the materials into thick polypropylene high-pressure plastic bags according to the dosage of 250g per bag, compacting, and sterilizing for 30min at 121 ℃ by using an autoclave. And (3) cooling the sterilized fungus bags on an ultraclean workbench, and inoculating cultivated species at two ends of the fungus bags, wherein the inoculated cultivated species account for 10% of the mass of the fungus bag mixture before sterilization (calculated on a wet basis). The inoculated fungus bags are placed in an artificial climate box regularly, the temperature is 28 ℃, the humidity is 60 percent, and the standing culture is carried out for 20 days.
(7) After the culture is finished, the degradation rate of lignin in the material is 15.2%, the degradation rate of cellulose is 32.4%, and the selective degradation coefficient of lignocellulose is 0.47.
(8) Preparing a wort solid culture medium, wherein the formula is as follows: 5.0g of peptone, 10.0g of glucose, 3.0g of yeast extract, 3.0g of malt extract, 25.0g of agar powder and 1000mL of distilled water, and the pH is natural. And (3) carrying out streak transfer on the saccharomyces cerevisiae GIM2.188 in the frozen tube to a wort solid culture medium, after 3 times of transfer, selecting yeast hyphae to inoculate to the wort liquid culture medium, and culturing for 16h at the temperature of 28 +/-1 ℃ and 150rev/min to obtain saccharomyces cerevisiae bacterial liquid.
(9) Putting the materials cultured in the step (6) into a 250mL fermentation bottle, adding a malt wort liquid culture medium as a nutrient solution, ensuring that the mass fraction of dry matters of the materials in the fermentation bottle is 5%, and sterilizing the fermentation bottle at 121 ℃ for 30min under high pressure; adding cellulase (same as example 1) into the sterilized fermentation bottle in an amount of 25U/gVS, wherein VS refers to the mass of volatile substances contained in the raw material mixture; then adding the mixture with the concentration of 108cfu/mL of saccharomyces cerevisiae bacterial liquid, wherein the inoculation amount of the saccharomyces cerevisiae bacterial liquid is 10% (V/V in volume) of the total volume of the fermentation bottle material.
(10) And (3) placing the fermentation bottle at the temperature of 28 +/-1 ℃ and under the condition of 150rev/min, culturing for 8 days until the ethanol concentration in the fermentation bottle is basically unchanged, wherein the final ethanol concentration in the fermentation bottle is 128.3 g/L.
The above embodiments are preferred embodiments of the present invention, but the present invention is not limited to the above embodiments, and any other changes, modifications, substitutions, combinations, and simplifications which do not depart from the spirit and principle of the present invention should be construed as equivalents thereof, and all such changes, modifications, substitutions, combinations, and simplifications are intended to be included in the scope of the present invention.

Claims (10)

1. A method for selectively degrading garden waste is characterized by comprising the following steps:
(1) preparation of cultivars: au781 is inoculated into a sterilized substrate containing wheat grains for culture to obtain a cultivated species;
(2) aerobic culture: adding water into garden waste, mixing to obtain culture substrate, sterilizing, inoculating the said cultivated species, and aerobic culturing.
2. The method for selectively degrading garden waste according to claim 1, wherein:
the substrate containing the wheat grains in the step (1) comprises the wheat grains, wood chips and calcium carbonate;
calcium carbonate and waste yeast are also added into the culture substrate in the step (2);
the specific operation of the inoculation after the sterilization in the step (2) is as follows: and bagging the culture substrate according to 250-300 g per bag, sterilizing, cooling, and inoculating cultivated species at two ends of the fungus bag.
3. The method for selectively degrading garden waste according to claim 2, wherein:
activating the auricularia polytricha in the step (1) before inoculation;
inoculating the inoculum size of the cultivar in the step (2) according to the 10 percent of the wet weight of the culture substrate before sterilization;
the substrate containing the wheat grains is prepared by mixing the wheat grains, the wood chips and calcium carbonate according to the mass ratio of 94:5: 1;
the water content of the sawdust is 40-50%;
the garden waste, the waste yeast and the calcium carbonate in the culture substrate are mixed according to the mass ratio of (90-95) to (4-9) to 1;
the water content of the waste yeast is 5-10%.
4. The method for selectively degrading garden waste according to claim 1, wherein:
the wheat grains are pretreated as follows: after being cleaned, the wheat grains are soaked in cold water for 8-10 h, boiled in boiling water for 20-30 min, and drained until the water content is 85-92%;
firstly crushing the garden waste in the step (2) to be less than 3 cm; the water content is 40-50%;
the specific method for preparing the cultivar comprises the following steps: preparing the activated auricularia polytricha and a culture medium into round slices to obtain fungus plugs; placing the fungus plugs in a container filled with a substrate for culturing until hyphae grow over the container;
the temperature of the aerobic culture condition in the step (2) is 28 ℃; the humidity is 60%; the aerobic culture time is 20 days.
5. The method for selectively degrading garden waste according to claim 4, wherein:
in the preparation of said cultivar:
the filling amount of the substrate in the container is 70%, and the material is not compacted;
the diameter of the round slice is 0.5 cm;
adding 5-7 bacterial plugs into each 250mL of substrate;
the culture conditions are that the temperature is 28 ℃ and the humidity is 60%;
adding water in the step (2) according to the mass ratio of the materials to the water of 1: 1;
and (3) standing the culture substrate in the step (2) for 8-10 hours before sterilization.
6. The method for selectively degrading garden waste according to any one of claims 1 to 5, wherein the method is applied to ethanol preparation.
7. A method for treating garden waste by using an aerobic-anaerobic two-stage process is characterized by comprising the following steps:
the garden waste treated by the method for selectively degrading garden waste according to any one of claims 1 to 5 is used as a substrate, and cellulase and saccharomyces cerevisiae are added for anaerobic fermentation to produce ethanol.
8. The method for treating garden waste by using the aerobic-anaerobic two-stage process according to claim 7, wherein the method comprises the following steps:
the substrate comprises the garden waste and a nutrient solution, the nutrient solution is a malt wort liquid culture medium, and the formula of the nutrient solution is 5.0g of peptone, 10.0g of glucose, 3.0g of yeast extract, 3.0g of malt extract and 1000mL of distilled water;
inoculating the strain of the saccharomyces cerevisiae liquid according to 5-10% of the total volume of the substrate;
the saccharomyces cerevisiae is saccharomyces cerevisiae GIM 2.188;
the saccharomyces cerevisiae bacterial liquid is obtained by inoculating saccharomyces cerevisiae to a wort solid culture medium for culture.
9. The method for treating garden waste by using the aerobic-anaerobic two-stage process according to claim 8, wherein the method comprises the following steps:
the mass fraction of dry matters of the substrate in the fermentation container is 5-10%;
the formula of the wort solid culture medium is as follows: 5.0g of peptone, 10.0g of glucose, 3.0g of yeast extract, 3.0g of malt extract, 25.0g of agar powder and 1000mL of distilled water;
the saccharomyces cerevisiae bacterial liquid is obtained by culturing for 12-18 h at the temperature of 28 +/-1 ℃ and the speed of 150 rev/min.
10. The method for treating garden waste by using the aerobic-anaerobic two-stage process according to claim 7, wherein the method comprises the following steps:
the concentration of the saccharomyces cerevisiae bacterial liquid is 108～109cfu/mL；
The addition amount of the cellulase is 20-40U/gVS, wherein VS refers to the mass of volatile substances contained in the raw material mixture;
the anaerobic fermentation condition is 28 +/-1 ℃ and 150 rev/min;
the anaerobic fermentation is subject to the condition that the ethanol concentration in the fermentation container is basically unchanged.
CN201910909408.3A 2019-09-25 2019-09-25 Method for treating garden waste by using aerobic-anaerobic two-stage process Pending CN110653238A (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN201910909408.3A CN110653238A (en) 2019-09-25 2019-09-25 Method for treating garden waste by using aerobic-anaerobic two-stage process

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN201910909408.3A CN110653238A (en) 2019-09-25 2019-09-25 Method for treating garden waste by using aerobic-anaerobic two-stage process

Publications (1)

Publication Number Publication Date
CN110653238A true CN110653238A (en) 2020-01-07

Family

ID=69039022

Family Applications (1)

Application Number Title Priority Date Filing Date
CN201910909408.3A Pending CN110653238A (en) 2019-09-25 2019-09-25 Method for treating garden waste by using aerobic-anaerobic two-stage process

Country Status (1)

Country Link
CN (1) CN110653238A (en)

Citations (10)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20070227063A1 (en) * 2006-03-30 2007-10-04 Board Of Trustees Of Michigan State University Process for conversion of mushroom lignocellulosic waste to useful byproducts
JP2007319860A (en) * 2007-09-03 2007-12-13 Sadatsune Fukuda System for treating organic waste
CN101128595A (en) * 2005-02-28 2008-02-20 株式会社雪国舞茸 Pretreatment of waste mushroom bed and method of converting the same into sugars and ethanol
CN101235392A (en) * 2008-01-16 2008-08-06 柏晓东 Cellulose fuel ethanol and preparation method thereof
CN102449156A (en) * 2009-03-17 2012-05-09 全技术公司 Compositions and methods for conversion of lignocellulosic material to fermentable sugars and products produced therefrom
CN102757904A (en) * 2011-04-25 2012-10-31 广州珠江啤酒股份有限公司 Method for preparing yeast culture media by using waste brewer's yeast
CN103409383A (en) * 2013-07-25 2013-11-27 江苏大学 Method used for accelerating lignin degradation in Aspergillus oryzae solid state fermentation
CN107360856A (en) * 2017-07-24 2017-11-21 北京中林智艺技术有限公司 Gardens dry branches and fallen leaves weeds cultivation flat mushroom method
CN109168977A (en) * 2018-10-30 2019-01-11 湖南奇硒健康产业有限公司 A kind of selenium-rich wheat food and its breeding method
CN109588208A (en) * 2018-12-22 2019-04-09 甘肃省科学院生物研究所 A kind of biological stephanoporate material and preparation method

Patent Citations (10)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101128595A (en) * 2005-02-28 2008-02-20 株式会社雪国舞茸 Pretreatment of waste mushroom bed and method of converting the same into sugars and ethanol
US20070227063A1 (en) * 2006-03-30 2007-10-04 Board Of Trustees Of Michigan State University Process for conversion of mushroom lignocellulosic waste to useful byproducts
JP2007319860A (en) * 2007-09-03 2007-12-13 Sadatsune Fukuda System for treating organic waste
CN101235392A (en) * 2008-01-16 2008-08-06 柏晓东 Cellulose fuel ethanol and preparation method thereof
CN102449156A (en) * 2009-03-17 2012-05-09 全技术公司 Compositions and methods for conversion of lignocellulosic material to fermentable sugars and products produced therefrom
CN102757904A (en) * 2011-04-25 2012-10-31 广州珠江啤酒股份有限公司 Method for preparing yeast culture media by using waste brewer's yeast
CN103409383A (en) * 2013-07-25 2013-11-27 江苏大学 Method used for accelerating lignin degradation in Aspergillus oryzae solid state fermentation
CN107360856A (en) * 2017-07-24 2017-11-21 北京中林智艺技术有限公司 Gardens dry branches and fallen leaves weeds cultivation flat mushroom method
CN109168977A (en) * 2018-10-30 2019-01-11 湖南奇硒健康产业有限公司 A kind of selenium-rich wheat food and its breeding method
CN109588208A (en) * 2018-12-22 2019-04-09 甘肃省科学院生物研究所 A kind of biological stephanoporate material and preparation method

Non-Patent Citations (3)

* Cited by examiner, † Cited by third party
Title
史雅静等: "毛木耳降解木质纤维素的研究", 《微生物学杂志》 *
宫志远等: "《食用菌保护地栽培技术》", 31 January 2002, 山东科学技术出版社 *
杨玉红等: "《生命科学综合实验指导》", 31 July 2016, 沈阳：东北大学出版社 *

Similar Documents

Publication Publication Date Title
CN105567612B (en) A kind of degradation composite bacteria agent preparation of garden waste and application
CN101974436B (en) Lignocellulose degrading bacteria and application thereof
CN102765972B (en) Organic garbage decomposing bacterium agent, as well as preparation method and application of organic garbage composting bacterium agent
CN103444436B (en) Method for cultivating lucid ganoderma and phoenix mushroom by waste mango branches
CN101037639A (en) Method for producing biologic grease and diesel oil
CN103993042A (en) Method for combined production of bioethanol and pullulan from lignocellulose substances
CN102220380B (en) Method for preparing fuel ethanol efficiently from large-algae biomass as raw material
CN105803004A (en) Method for low-temperature fermentation of biogas from agricultural waste
CN1858213A (en) Method for co-producing hydrogen and methane by biomass and solid organic waste fermenting method
CN101532038A (en) Method for preparing fuel ethanol by plant straws
CN103992958B (en) One strain rice straw degradative fungi intends healthy and free from worry Trichoderma spp. ZJC-1 and microbial inoculum thereof
CN109089732B (en) Edible fungus culture medium and preparation method and application thereof
CN101914576B (en) Method for producing ethanol and methane by mixed fermentation of paper-making sludge and monosodium glutamate waste liquid
CN110653238A (en) Method for treating garden waste by using aerobic-anaerobic two-stage process
CN102492634B (en) High-temperature resistant yeast and application thereof
CN107893043A (en) A kind of zymomonas mobilis mutant strain of enduring high-concentration acetic acid and its application
CN103881948B (en) A kind of detoxification hydrolysis rapeseed cake produces microbial inoculum and the technique of agricultural amino acid fertilizer
CN110592047B (en) Novel method for producing feruloyl esterase by fermenting straws with Verbena pyricularis and application
CN107630047A (en) A kind of method of biophysics Combined Treatment straw production ethanol and volatile fatty acid
CN108330087B (en) Solid leaven for fermenting peanut straw
CN105524835A (en) Obtaining method and application of salt-tolerant cellulose decomposition bacteria colony
CN102161971B (en) Fermentation inoculum for producing cellulase and hemicellulase
CN105779506B (en) Method for co-producing methane and ethanol
CN110583867A (en) Method for producing protein feed from potato residue and method for utilizing potato starch processing wastewater residue
CN111662113A (en) Method for producing organic fertilizer by fermenting garden pruning matters

Legal Events

Date Code Title Description
PB01 Publication
PB01 Publication
SE01 Entry into force of request for substantive examination
SE01 Entry into force of request for substantive examination