CN101580856A - Method utilizing agricultural wastes as raw materials to prepare microbial grease by fermentation - Google Patents
Method utilizing agricultural wastes as raw materials to prepare microbial grease by fermentation Download PDFInfo
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Abstract
The invention discloses a method utilizing agricultural wastes as raw materials to prepare microbial grease by fermentation. The method has the technical scheme that the agricultural wastes are hydrolyzed by diluted acid, a hydrolysis solution is used for preparing fermentation media after detoxifcation pretreatment, and the step of concentration is carried out until the concentration of reducing sugar ranges from 80 to 120 g/L; according to the concentration thereof in a mixture, 1.2 to 2.4 g/L of peptone and 0.25 to 0.75 g/L of dusty yeast, and the like are added; after sterilization, 5 to 15 percent of microorganism seed solution is introduced according to volume ratio; and after 7 to 9 days of culture, thalli is centrifugally collected, and the microbial grease is obtained according to the method of organic solvent extraction. The detoxifcation pretreatment is the treatment and the adsorption treatment of CaOH2. The method has the advantages of wide source of raw materials, low price and simple and easily executed process, can realize the high value utilization of hemicellulose resources in the agricultural wastes, and can reduce the preparation cost of the microbial grease.
Description
Technical field
The invention belongs to biological chemical field, particularly a kind of microorganism utilizes the method for agricultural wastes (agricultural crop straw, bagasse and corn cob) hemicellulose hydrolysate prepare microbial grease by fermentation.
Background technology
Biofuel be a kind of be the green regenerative energy sources that can substitute fossil diesel fuel of raw material production with the animal-plant oil, in petroleum-based energy substitutes strategy, have crucial status.The suitability for industrialized production of biofuel is a raw material with soybean oil, rapeseed oil and waste oil mainly at present.China has a large population and a few land, and for the raw material production biofuel does not meet national conditions, and the limited amount of waste oil can't satisfy the great demand of modern economy society to the energy with the edible oil.Microbial oil claims Unicell Oils and Fats again, is a kind of meta-bolites in the microbial life activity.Certain micro-organisms such as yeast, mould, little algae, bacterium can accumulate a large amount of greases, and fat content even reach is done mycelial more than 70% in the thalline that has.These greases are made up of similar lipid acid with general Vegetable oil lipoprotein, are a kind of new bio diesel production raw materials with great potential therefore.Compare with animal-plant oil, have following advantage with the microorganisms producing grease: growth cycle is short, and floor space is little, is not subjected to place, weather and season limit, can continuous large-scale production.But traditional oil fermentation is a carbon source with glucose etc., and cost is too high, has limited its application in production of biodiesel.Utilize various agriculture and industry wastes,, both can reduce the production cost of microbial oil greatly, help environment protection again as fermentative production greases such as agricultural crop straw, foodstuffs industry and paper industry waste water.
Lignocellulose is a renewable resources the abundantest on the earth, and its main component is Mierocrystalline cellulose, hemicellulose and xylogen.At present, Mierocrystalline cellulose and xylogen all can find the approach of effective utilization, and hemicellulose is not effectively used as yet.China is a large agricultural country, various agricultural wastes, and as enormous amount such as agricultural crop straw, bagasses, only agricultural crop straw just reaches more than 700,000,000 tons every year.In the past, this part lignocellulose resource often goes out of use or devalued utilization, be used for the farmland compost as agricultural crop straw after burning, but the black smoke that produces in the burning process pollutes the environment.In recent years, though numerous studies show that can utilize stalk or bagasse to produce alcohol fuel etc., what in fact be utilized mainly is cellulose components.Similar, greasy research also is mainly to utilize its cellulose components for raw material produces much to utilize lignocellulose, and for hemicellulose components, but often can not efficiently utilize.The main ingredient of hemicellulose is an xylan, can obtain wood sugar, glucose and pectinose etc. after the hydrolysis, and based on wood sugar, and occurring in nature seldom microorganism can utilize wood sugar.Utilizing some special microorganisms can be that the carbon source and the energy accumulate greasy characteristic with the wood sugar, as fermentable trichosporon (Trichosporon fermentans), with the agricultural wastes hemicellulose hydrolysate is that fermenting raw materials is produced grease, can reduce the production cost of microbial oil greatly, realize the higher value application of hemicellulose resource in the agricultural wastes, and help improving the situation of present energy shortage and environmental degradation.
Summary of the invention
The objective of the invention is in order to solve the deficiency that prior art exists, providing a kind of is raw material with agricultural wastes, efficiently utilizes the method for hemicellulose prepare microbial grease by fermentation.
The present invention is a raw material with the agricultural wastes of cheapness, and with its hydrolysis, hydrolyzed solution adds a spot of nitrogenous source and trace element after the detoxifcation pre-treatment and concentrating, obtain grease through the produce oil microbial transformation with diluted acid.Present method will significantly reduce the production cost of microbial oil, realize the higher value application of agricultural wastes hemicellulose resource.
The object of the invention is achieved through the following technical solutions:
With agricultural wastes is the method that fermenting raw materials is produced microbial oil, comprises the steps and processing condition:
(1) hydrolysis of hemicellulose: agricultural wastes are crushed to the 20-100 order, are that 1g: 10-20mL adds the hydrolysis acid solution according to the solid-to-liquid ratio of agricultural wastes and hydrolysis acid solution, obtain hemicellulose hydrolysate in 110-140 ℃ of following hydrolysis 1-3h;
(2) detoxifcation pre-treatment: comprise Ca (OH)
2Handle and adsorption treatment;
Ca (OH)
2Handle: in hydrolyzed solution, add Ca (OH)
2Powder is regulated its pH to 10.0-12.0, behind the maintenance 1-1.5h, filters, and be 5.5-6.0 with acid with the filtrate pH regulator, leave standstill 1-2h, centrifugal 10-20min removes precipitation;
Adsorption treatment is selected from a kind of in step a, b and the c absorption:
A. charcoal absorption: according to gac and Ca (OH)
2Handle gained hydrolyzed solution solid-to-liquid ratio 1g: 5-10mL adds gac, removes gac at 30-40 ℃ of following stir process 1-2h after-filtration;
B. ion exchange resin absorption: according to ion exchange resin and Ca (OH)
2Handle gained hydrolyzed solution solid-to-liquid ratio 1g: 5-10mL adds ion exchange resin, at 30-40 ℃ of following stir process 1-2h after-filtration deionizing exchange resin;
C. macroporous resin adsorption: according to macroporous resin and Ca (OH)
2Handle gained hydrolyzed solution solid-to-liquid ratio 1g: 5-10mL adds macroporous resin, removes macroporous resin at 30-40 ℃ of following stir process 1-2h after-filtration;
(3) fermention medium preparation: detoxify pretreated hydrolyzed solution under reduced pressure, being concentrated into concentration of reduced sugar under 60-70 ℃ is 80-120g/L, adds following material according to its concentration in mixture then: 1.2-2.4g/L peptone, 0.25-0.75g/L yeast powder, 0.4-0.6g/L MgSO
47H
2O, 1.5-2.5g/L KH
2PO
4, 0.003-0.005g/LMnSO
4H
2O and 0.0001-0.0002g/L CuSO
45H
2O, with acid for adjusting pH to 6.0-6.5, after sterilizing as fermention medium;
(4) 5-15 by volume: 100 insert fermentable trichosporon (Trichosporon fermentans) seed liquor in fermention medium, cultivated 7-9 days at 25-28 ℃, and centrifugal collection thalline obtains microbial oil through the organic solvent extraction method.
For further realizing the object of the invention, described organic solvent extraction method is: adding 10mL concentration by every gram dry mycelium is 3.5-4.5mol/L HCl, handles 30-60min in 80-100 ℃; Be that chloroform and the methanol mixed solvent that adds at 2: 1 extracts by volume after the cooling, collect chloroform layer, remove organic solvent, obtain microbial oil with the Rotary Evaporators decompression.
The concentration of described HCl is preferably 4mol/L.
Described agricultural wastes are preferably paddy rice stalk, wheat-straw, sorghum stalk, maize straw, bagasse or corn cob.
Described hydrolysis acid is preferably sulfuric acid, hydrochloric acid or phosphoric acid.
Described step (2) Ca (OH)
2Acid is preferably sulfuric acid, hydrochloric acid or phosphoric acid in the processing.
Regulate the used acid of pH in the preparation of described step (3) fermention medium and be preferably sulfuric acid, hydrochloric acid or phosphoric acid.
With respect to prior art, of the present invention is the method that fermenting raw materials is produced microbial oil with the agricultural wastes hemicellulose hydrolysate, has following advantage:
(1) the present invention's hemicellulose components that can effectively utilize in the agricultural wastes is come prepare microbial grease by fermentation, provides novel method and new way for solving the difficult problem that hemicellulose is difficult to utilize in the lignocellulose resource;
(2) agricultural waste material that is utilized is renewable resources, and wide material sources, cheap, is that the raw material production grease not only can be turned waste into wealth with it, reduces the production cost of microbial oil, and helps environment protection.
Embodiment
Below in conjunction with embodiment the present invention is described further, embodiments of the invention are not limit so.
Embodiment 1
1) rice straw hemicellulose hydrolysate preparation
Rice straw being crushed to 20 orders, taking by weighing 40g, is that 1g: 20mL adds dilution heat of sulfuric acid according to the solid-to-liquid ratio of rice straw (solid) and 2% (weight) dilute sulphuric acid (liquid), in 121 ℃ of following hydrolysis 1h, after the filtration hydrolyzed solution.
2) detoxifcation of hydrolyzed solution is handled
A.Ca (OH)
2Handle: in hydrolyzed solution, add Ca (OH)
2Powder is regulated its pH to 10.0, behind the maintenance 1h, filters.Regulate filtrate pH to 5.5 with HCl, leave standstill 1h, centrifugal again 10min removes precipitation.
B. charcoal absorption: by solid-to-liquid ratio 1g: 10mL, gac (particle diameter is 2.5mm, and German Merck company produces) is added in the hydrolyzed solution of step a gained, remove gac at 40 ℃ of following stir process 1h after-filtration.
3) preparation of fermention medium
Hydrolyzed solution after the detoxifcation is under reduced pressure, and being concentrated into concentration of reduced sugar under 60 ℃ is 100g/L, adds the peptone of following material: 1.8g/L, the yeast powder of 0.5g/L, the MgSO of 0.4g/L according to its concentration in this step final mixture then
47H
2The KH of O, 2.0g/L
2PO
4, 0.003g/L MnSO
4H
2O and 0.0001g/LCuSO
45H
2O uses H
2SO
4Regulate pH to 6.5,121 ℃ of sterilization 20min make fermention medium.
4) liquid seeds is cultivated
Fermentable trichosporon (Trichosporon fermentans CICC 1368, purchase in Chinese industrial microbial strains preservation center) at liquid seed culture medium (glucose 20g/L, peptone 10g/L, yeast powder 10g/L, pH is 6.0) in cultivate 24h in 28 ℃, 160r/min, make fermentation seed liquid.
5) microbe oil fermentation
With 10% (volume) inoculum size seed liquor is inserted in the 50mL fermention medium, cultivated 8 days, stop fermentation in 25 ℃, 160r/min; Fermented liquid in the centrifugal 10min of 8000r/min, is collected thalline, washes the back with water and be dried to constant weight in 105 ℃, dry mycelium 18.2g/L.
6) microbial oil extracts
Add 10mL 4mol/L HCl by every gram dry mycelium, handle 60min in 80 ℃; Adding volume ratio in cooling back is that 2: 1 chloroform and methanol mixed solvent extracts, and collects chloroform layer, removes organic solvent with the Rotary Evaporators decompression, obtains microbial oil, and the thalline fat content is 35.6%.
Embodiment 2
1) corn cob hemicellulose hydrolysate preparation
Corn cob meal being broken to 40 orders, taking by weighing 40g, is that 1g: 10mL adds dilute hydrochloric acid solution according to the solid-to-liquid ratio of corn cob (solid) and 3% (weight) dilute hydrochloric acid solution (liquid), in 140 ℃ of following hydrolysis 3h, filters the back and obtains hydrolyzed solution.
2) detoxifcation of hydrolyzed solution is handled
A.Ca (OH)
2Handle: in hydrolyzed solution, add Ca (OH)
2Powder is regulated its pH to 10.0, behind the maintenance 1.5h, filters.Use H
2SO
4Regulate filtrate pH to 6.0, leave standstill 2h, centrifugal again 15min removes precipitation.
B. ion exchange resin absorption: ion exchange resin Bio-Rad AG 1-X8 is added in the hydrolyzed solution, at 30 ℃ of following stir process 2h after-filtration deionizing exchange resins by solid-to-liquid ratio 1g: 5mL.
3) preparation of fermention medium
Hydrolyzed solution after the detoxifcation is under reduced pressure, and being concentrated into concentration of reduced sugar under 60 ℃ is 80g/L, adds following material according to its concentration in this step final mixture then: peptone 1.2g/L; Yeast powder 0.75g/L; MgSO
47H
2O 0.6g/L; KH
2PO
42.5g/L; MnSO
4H
2O 0.004g/L; CuSO
45H
2O 0.0001g/L regulates pH to 6.0 with HCl, and 121 ℃ of sterilization 20min make fermention medium.
4) liquid seeds is cultivated
Fermentable trichosporon (Trichosporon fermentans CICC 1368, purchase in Chinese industrial microbial strains preservation center) at liquid seed culture medium (glucose 20g/L, peptone 10g/L, yeast powder 10g/L, pH is 6.0) in cultivate 24h in 28 ℃, 160r/min, make fermentation seed liquid.
5) microbe oil fermentation
With 5% inoculum size (volume) seed liquor is inserted in the 50mL fermention medium, cultivated 9 days, stop fermentation in 28 ℃, 160r/min; Fermented liquid in the centrifugal 10min of 8000r/min, is collected thalline, washes the back with water and be dried to constant weight in 105 ℃, dry mycelium 21.6g/L.
6) microbial oil extracts
Add 10mL 4.5mol/L HCl by every gram dry mycelium, in 90 ℃ of water bath processing 40min; Adding volume ratio in cooling back is that 2: 1 chloroform and methanol mixed solvent extracts, and collects chloroform layer, removes organic solvent with the Rotary Evaporators decompression, obtains microbial oil, and the thalline fat content is 34.3%.
Embodiment 3
1) bagasse hemicellulose hydrolyzed solution preparation
Bagasse being crushed to 100 orders, taking by weighing 40g, is that 1g: 15mL adds dilution heat of sulfuric acid according to the solid-to-liquid ratio of bagasse (solid) and 4% (weight) dilute sulphuric acid (liquid), in 140 ℃ of following hydrolysis 1.5h, filters the back and obtains hydrolyzed solution.
2) detoxifcation of hydrolyzed solution is handled
A.Ca (OH)
2Handle: in hydrolyzed solution, add Ca (OH)
2Powder is regulated its pH to 11.0, behind the maintenance 1h, filters.Use H
3PO
4Regulate filtrate pH to 5.5, leave standstill 1.5h, centrifugal again 20min removes precipitation.
B. macroporous resin adsorption: macroporous resin Amberlite XAD-4 is added in the hydrolyzed solution by solid-to-liquid ratio 1g: 5mL, remove macroporous resin at 40 ℃ of following stir process 1h after-filtration.
3) preparation of fermention medium
Hydrolyzed solution after the detoxifcation is under reduced pressure, and being concentrated into concentration of reduced sugar under 70 ℃ is 120g/L, adds following material according to its concentration in this step final mixture then: peptone 2.4g/L; Yeast powder 0.25g/L; MgSO
47H
2O 0.5g/L; KH
2PO
41.5g/L; MnSO
4H
2O 0.005g/L; CuSO
45H
2O 0.0002g/L uses H
3PO
4Regulate pH to 6.0,121 ℃ of sterilization 20min make fermention medium.
4) liquid seeds is cultivated
Fermentable trichosporon (Trichosporon fermentans CICC 1368, purchase in Chinese industrial microbial strains preservation center) at liquid seed culture medium (glucose 20g/L, peptone 10g/L, yeast powder 10g/L, pH is 6.0) in cultivate 24h in 28 ℃, 160r/min, make fermentation seed liquid.
5) microbe oil fermentation
With 15% inoculum size (volume) seed liquor is inserted in the 50mL fermention medium, cultivated 7 days, stop fermentation in 25 ℃, 160r/min; Fermented liquid in the centrifugal 10min of 8000r/min, is collected thalline, washes the back with water and be dried to constant weight in 105 ℃, dry mycelium 25.2g/L.
6) microbial oil extracts
Add 10mL 3.5mol/L HCl by every gram dry mycelium, in 100 ℃ of water bath processing 30min; Adding volume ratio in cooling back is that 2: 1 chloroform and methanol mixed solvent extracts, and collects chloroform layer, removes organic solvent with the Rotary Evaporators decompression, obtains microbial oil, and the thalline fat content is 38.4%.
Embodiment 4
1) maize straw hemicellulose hydrolysate preparation
Maize straw being crushed to 60 orders, taking by weighing 40g, is that 1g: 10mL adds dilute phosphoric acid solution according to the solid-to-liquid ratio of maize straw (solid) and 2% (weight) dilute phosphoric acid (liquid), in 110 ℃ of following hydrolysis 3h, filters the back and obtains hydrolyzed solution.
2) detoxifcation of hydrolyzed solution is handled
A.Ca (OH)
2Handle: in hydrolyzed solution, add Ca (OH)
2Powder is regulated its pH to 12.0, behind the maintenance 1h, filters.Use H
2SO
4Regulate filtrate pH to 5.5, leave standstill 1h, centrifugal again 20min removes precipitation.
B. charcoal absorption: gac (particle diameter is 2.5mm, and German Merck company produces) is added in the hydrolyzed solution by solid-to-liquid ratio 1g: 5mL, remove gac at 30 ℃ of following stir process 2h after-filtration.
3) preparation of fermention medium
Hydrolyzed solution after the detoxifcation is under reduced pressure, and being concentrated into concentration of reduced sugar under 65 ℃ is 110g/L, adds following material according to its concentration in this step final mixture then: peptone 1.6g/L; Yeast powder 0.75g/L; MgSO
47H
2O, 0.4g/L; KH
2PO
42.0g/L; MnSO
4H
2O0.004g/L; CuSO
45H
2O0.0002g/L uses H
2SO
4Regulate pH to 6.5,121 ℃ of sterilization 20min make fermention medium.
4) liquid seeds is cultivated
Fermentable trichosporon (Trichosporon fermentans CICC 1368, purchase in Chinese industrial microbial strains preservation center) at liquid seed culture medium (glucose 20g/L, peptone 10g/L, yeast powder 10g/L, pH is 6.0) in cultivate 24h in 28 ℃, 160r/min, make fermentation seed liquid.
5) microbe oil fermentation
With 12.5% inoculum size (volume) seed liquor is inserted in the 50mL fermention medium, cultivated 7 days, stop fermentation in 25 ℃, 160r/min; Fermented liquid in the centrifugal 10min of 8000r/min, is collected thalline, washes the back with water and be dried to constant weight in 105 ℃, dry mycelium 19.2g/L.
6) microbial oil extracts
Add 10mL 4mol/L HCl by every gram dry mycelium, in 80 ℃ of water bath processing 60min; Adding volume ratio in cooling back is that 2: 1 chloroform and methanol mixed solvent extracts, and collects chloroform layer, removes organic solvent with the Rotary Evaporators decompression, obtains microbial oil, and the thalline fat content is 28.0%.
Embodiment 5
1) broomcorn straw hemicellulose hydrolysate preparation
Broomcorn straw being crushed to 80 orders, taking by weighing 40g, is that 1g: 10mL adds dilute hydrochloric acid solution according to the solid-to-liquid ratio of broomcorn straw (solid) and 6% (weight) dilute hydrochloric acid (liquid), in 130 ℃ of following hydrolysis 2.5h, filters the back and obtains hydrolyzed solution.
2) detoxifcation of hydrolyzed solution is handled
A.Ca (OH)
2Handle: in hydrolyzed solution, add Ca (OH)
2Powder is regulated its pH to 10.0, behind the maintenance 1.5h, filters.Use H
2SO
4Regulate filtrate pH to 5.5, leave standstill 1h, centrifugal again 10min removes precipitation.
B. ion exchange resin absorption: ion exchange resin Purolite MN-150 is added in the hydrolyzed solution, at 40 ℃ of following stir process 1h after-filtration deionizing exchange resins by solid-to-liquid ratio 1g: 10mL.
3) preparation of fermention medium
Hydrolyzed solution after the detoxifcation is under reduced pressure, and being concentrated into concentration of reduced sugar under 60 ℃ is 90, adds following material according to its concentration in this step final mixture then: peptone 1.8g/L; Yeast powder 0.5g/L; MgSO
47H
2O 0.4g/L; KH
2PO
42.0g/L; MnSO
4H
2O 0.003g/L; CuSO
45H
2O 0.0001g/L uses H
3PO
4Regulate pH to 6.5,121 ℃ of sterilization 20min make fermention medium.
4) liquid seeds is cultivated
Fermentable trichosporon (Trichosporon fermentans CICC 1368, purchase in Chinese industrial microbial strains preservation center) at liquid seed culture medium (glucose 20g/L, peptone 10g/L, yeast powder 10g/L, pH is 6.0) in cultivate 24h in 28 ℃, 160r/min, make fermentation seed liquid.
5) microbe oil fermentation
With 10% inoculum size (volume) seed liquor is inserted in the 50mL fermention medium, cultivated 9 days, stop fermentation in 28 ℃, 160r/min; Fermented liquid in the centrifugal 10min of 8000r/min, is collected thalline, washes the back with water and be dried to constant weight in 105 ℃, dry mycelium 21.8g/L.
6) microbial oil extracts
Add 10mL 4mol/L HCl by every gram dry mycelium, in 100 ℃ of water bath processing 30min; Adding volume ratio in cooling back is that 2: 1 chloroform and methanol mixed solvent extracts, and collects chloroform layer, removes organic solvent with the Rotary Evaporators decompression, obtains microbial oil, and the thalline fat content is 33.1%.
Embodiment 6
1) wheat stalk hemicellulose hydrolysate preparation
Wheat stalk being crushed to 100 orders, taking by weighing 40g, is that 1g: 10mL adds dilution heat of sulfuric acid according to the solid-to-liquid ratio of wheat stalk (solid) and 4% (weight) dilute sulphuric acid (liquid), in 140 ℃ of following hydrolysis 1h, filters the back and obtains hydrolyzed solution.
2) detoxifcation of hydrolyzed solution is handled
A.Ca (OH)
2Handle: in hydrolyzed solution, add Ca (OH)
2Powder is regulated its pH to 12.0, behind the maintenance 1h, filters.Regulate filtrate pH to 5.5 with HCl, leave standstill 1h, centrifugal again 15min removes precipitation.
B. ion exchange resin absorption: ion exchange resin Bio-Rad AG50W-X8 is added in the hydrolyzed solution, at 40 ℃ of following stir process 1h after-filtration deionizing exchange resins by solid-to-liquid ratio 1g: 10mL.
3) preparation of fermention medium
Hydrolyzed solution after the detoxifcation is under reduced pressure, and being concentrated into concentration of reduced sugar under 60 ℃ is 90g/L, adds following material according to its concentration in this step final mixture then: peptone 1.8g/L; Yeast powder 0.25g/L; MgSO
47H
2O 0.5g/L; KH
2PO
42.0g/L; MnSO
4H
2O 0.004g/L; CuSO
45H
2O 0.0002g/L regulates pH to 6.5 with HCl, and 121 ℃ of sterilization 20min make fermention medium.
4) liquid seeds is cultivated
Fermentable trichosporon (Trichosporon fermentans CICC 1368, purchase in Chinese industrial microbial strains preservation center) at liquid seed culture medium (glucose 20g/L, peptone 10g/L, yeast powder 10g/L, pH is 6.0) in cultivate 24h in 28 ℃, 160r/min, make fermentation seed liquid.
5) microbe oil fermentation
With 10% inoculum size (volume ratio) seed liquor is inserted in the 50mL fermention medium, cultivated 9 days, stop fermentation in 28 ℃, 160r/min; Fermented liquid in the centrifugal 10min of 8000r/min, is collected thalline, washes the back with water and be dried to constant weight in 105 ℃, dry mycelium 24.8g/L.
6) microbial oil extracts
Add 10mL 4mol/L HCl by every gram dry mycelium, in 80 ℃ of water bath processing 60min; Adding volume ratio in cooling back is that 2: 1 chloroform and methanol mixed solvent extracts, and collects chloroform layer, removes organic solvent with the Rotary Evaporators decompression, obtains microbial oil, and the thalline fat content is 35.5%.
Embodiment 7
Be that with the difference of embodiment 3 raw material is a rice straw in the step 1), be crushed to 40 orders, hydrolysis temperature and time are respectively 130 ℃ and 2h; Step 2) the big pore resin is Amberlite XAD-8 in, and solid-to-liquid ratio is 1g: 10mL, and the stir process temperature and time is respectively 30 ℃ and 2h; The microbe oil fermentation time is 8 days in the step 5).Result of implementation obtains dry mycelium 28.0g/L, and the thalline fat content is 40.1%.
Embodiment 8
Be that with the difference of embodiment 2 raw material is a maize straw in the step 1), used diluted acid is 3% dilute sulphuric acid, step 2) in the institute spent ion exchange resin be Purolite MN-150.Result of implementation obtains dry mycelium 21.2g/L, and the thalline fat content is 29.6%.
Embodiment 9
Be that with the difference of embodiment 2 raw material is a broomcorn straw in the step 1), used diluted acid is 2% dilute sulphuric acid, step 2) in the institute spent ion exchange resin be Purolite A-860S.Result of implementation obtains dry mycelium 20.1g/L, and the thalline fat content is 34.4%.
Embodiment 10
Be that with the difference of embodiment 3 raw material is a rice straw in the step 1), be crushed to 60 orders, used diluted acid is 2% dilute sulphuric acid, step 2) in the big pore resin be Amberlite XAD-8.Result of implementation obtains dry mycelium 25.8g/L, and the thalline fat content is 38.5%.
Claims (7)
1, be the method that fermenting raw materials is produced microbial oil with agricultural wastes, it is characterized in that comprising the steps and processing condition:
(1) hydrolysis of hemicellulose: agricultural wastes are crushed to the 20-100 order, are that 1g: 10-20mL adds the hydrolysis acid solution according to the solid-to-liquid ratio of agricultural wastes and hydrolysis acid solution, obtain hemicellulose hydrolysate in 110-140 ℃ of following hydrolysis 1-3h;
(2) detoxifcation pre-treatment: comprise Ca (OH)
2Handle and adsorption treatment;
Ca (OH)
2Handle: in hydrolyzed solution, add Ca (OH)
2Powder is regulated its pH to 10.0-12.0, behind the maintenance 1-1.5h, filters, and be 5.5-6.0 with acid with the filtrate pH regulator, leave standstill 1-2h, centrifugal 10-20min removes precipitation;
Adsorption treatment is selected from a kind of in step a, b and the c absorption:
A. charcoal absorption: according to gac and Ca (OH)
2Handle gained hydrolyzed solution solid-to-liquid ratio 1g: 5-10mL adds gac, removes gac at 30-40 ℃ of following stir process 1-2h after-filtration;
B. ion exchange resin absorption: according to ion exchange resin and Ca (OH)
2Handle gained hydrolyzed solution solid-to-liquid ratio 1g: 5-10mL adds ion exchange resin, at 30-40 ℃ of following stir process 1-2h after-filtration deionizing exchange resin;
C. macroporous resin adsorption: according to macroporous resin and Ca (OH)
2Handle gained hydrolyzed solution solid-to-liquid ratio 1g: 5-10mL adds macroporous resin, removes macroporous resin at 30-40 ℃ of following stir process 1-2h after-filtration;
(3) fermention medium preparation: detoxify pretreated hydrolyzed solution under reduced pressure, being concentrated into concentration of reduced sugar under 60-70 ℃ is 80-120g/L, adds following material according to its concentration in mixture then: 1.2-2.4g/L peptone, 0.25-0.75g/L yeast powder, 0.4-0.6g/L MgSO
47H
2O, 1.5-2.5g/L KH
2PO
4, 0.003-0.005g/LMnSO
4H
2O and 0.0001-0.0002g/L CuSO
45H
2O, with acid for adjusting pH to 6.0-6.5, after sterilizing as fermention medium;
(4) 5-15 by volume: 100 insert fermentable trichosporon seed liquor in fermention medium, cultivated 7-9 days at 25-28 ℃, and centrifugal collection thalline obtains microbial oil through the organic solvent extraction method.
2, according to claim 1 is the method that fermenting raw materials is produced microbial oil with agricultural wastes, it is characterized in that described organic solvent extraction method is: adding 10mL concentration by every gram dry mycelium is 3.5-4.5mol/L HCl, handles 30-60min in 80-100 ℃; Be that chloroform and the methanol mixed solvent that adds at 2: 1 extracts by volume after the cooling, collect chloroform layer, remove organic solvent, obtain microbial oil with the Rotary Evaporators decompression.
3, according to claim 2 is the method that fermenting raw materials is produced microbial oil with agricultural wastes, and the concentration that it is characterized in that described HCl is 4mol/L.
4, according to claim 1 is the method that fermenting raw materials is produced microbial oil with agricultural wastes, it is characterized in that described agricultural wastes are paddy rice stalk, wheat-straw, sorghum stalk, maize straw, bagasse or corn cob.
5, according to claim 1 is the method that fermenting raw materials is produced microbial oil with agricultural wastes, it is characterized in that described hydrolysis acid is sulfuric acid, hydrochloric acid or phosphoric acid.
6, according to claim 1 is the method that fermenting raw materials is produced microbial oil with agricultural wastes, it is characterized in that described step (2) Ca (OH)
2Acid is sulfuric acid, hydrochloric acid or phosphoric acid in the processing.
7, according to claim 1 is the method that fermenting raw materials is produced microbial oil with agricultural wastes, it is characterized in that regulating the used acid of pH in the preparation of described step (3) fermention medium is sulfuric acid, hydrochloric acid or phosphoric acid.
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| CN101948883A (en) * | 2010-09-06 | 2011-01-19 | 长春工业大学 | Method for fermentatively producing microbial oil by utilizing corn husk dregs as raw materials |
| CN102061319A (en) * | 2010-11-09 | 2011-05-18 | 佛山市正合生物能源有限公司 | Method for preparing biological fatty oil from Dioscorea camposita |
| CN102061317A (en) * | 2010-11-09 | 2011-05-18 | 佛山市正合生物能源有限公司 | Method for producing biolipid by cassava fermentation |
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| CN102618591A (en) * | 2012-04-11 | 2012-08-01 | 安徽安生生物化工科技有限责任公司 | Method for directly fermenting starchy materials and producing oil by aid of mortierella elongata |
| CN102618447A (en) * | 2012-01-10 | 2012-08-01 | 安徽安生生物化工科技有限责任公司 | Mortierella elongata PFY capable of utilizing lignocellulose for fermentation production of microbial oil and method of producing microbial oil by same |
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| CN102618591B (en) * | 2012-04-11 | 2013-08-21 | 安徽安生生物化工科技有限责任公司 | Method for directly fermenting starchy materials and producing oil by aid of mortierella elongata |
| CN102634549A (en) * | 2012-04-28 | 2012-08-15 | 中国科学院广州能源研究所 | Production method for microbial oil |
| CN102634549B (en) * | 2012-04-28 | 2014-06-04 | 中国科学院广州能源研究所 | Production method for microbial oil |
| CN104140884A (en) * | 2014-08-08 | 2014-11-12 | 融安恒裕香油科技有限公司 | Method for improving edible plant oilseed squeezing process |
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| CN110495520A (en) * | 2019-08-23 | 2019-11-26 | 福建师范大学 | Tea grounds fermentation liquid and its preparation method and application |
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