CN107916280A - A kind of lignocellulosic substep diastatic fermentation method using polyethylene glycol water as medium - Google Patents
A kind of lignocellulosic substep diastatic fermentation method using polyethylene glycol water as medium Download PDFInfo
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- 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
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- C12P19/00—Preparation of compounds containing saccharide radicals
- C12P19/14—Preparation of compounds containing saccharide radicals produced by the action of a carbohydrase (EC 3.2.x), e.g. by alpha-amylase, e.g. by cellulase, hemicellulase
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Abstract
A kind of lignocellulosic substep diastatic fermentation method using polyethylene glycol water as medium of the invention.Using the lignocellulosic of pretreatment as substrate, using polyethylene glycol/aqueous mixtures as fermentation media, using adding substrate and cellulase in batches and remove the strategy of enzymolysis liquid solid residue step by step before fermentation, the concentration of toxicity inhibition thing is reduced while solid content is improved, so as to obtain high concentration ethanol.The invention has the advantages that：Batch feeding is conducive to the heat transfer of enzymolysis process, mass transfer, can obtain the hydrolyzate of high-concentration reduced sugar；Digesting the solid residue produced can be as the adsorbent of fermentation toxicity inhibition thing, fermentation toxicant is effectively removes by centrifuging, enzymolysis liquid can be directly used for fermenting without detoxification treatment, and polyethylene glycol can effectively alleviate suppression of the residual toxic chemicals to alcohol fermentation in liquid；The acquisition of the hydrolyzate of high-concentration reduced sugar can improve later stage fermentation producing and ethanol concentration, so as to reduce cellulosic ethanol production cost.
Description
Technical field
The invention belongs to lignocellulosic hydrolysis technology field, and in particular to a kind of using polyethylene glycol-water as medium
Lignocellulosic substep diastatic fermentation method.
Background technology
With the increasingly depleted of fossil energy and increasingly sharpening for environmental pollution, renewable and clean energy resource alcohol fuel is opened
Send out and utilize the extensive concern for receiving people.For traditional alcohol fermentation using sugar or starch as raw material, the two is all the master of food
Source is wanted, producing alcohol fuel as raw material using grain constitutes safely world food threat, finds other raw materials and substitutes
Grain is imperative.The lignocellulosic renewable resource the abundantest and cheap as nature, its main component cellulose
Hemicellulose is the raw materials for production of potential alcohol fuel, is studied using lignocellulosic materials for fuel ethanol as countries in the world
Hot spot.It is the prerequisite for reducing cellulosic ethanol production cost that lignocellulosic material enzymolysis, which obtains high-concentration reduced sugar,.
And it is to obtain the direct method of hydrolyzate middle and high concentration reduced sugar to improve enzymatic hydrolysis system substrate solids content, but it is high
It is uneven that substrate solid content necessarily causes system to mix, and heat and mass is less able, as a result causes enzymolysis efficiency to reduce；It is and high
The enzymolysis substrate of solid content necessarily causes the generation of the fermentation toxicity inhibition thing of higher concentration, this seriously affects the normal of saccharomycete
Growth and subsequent alcohol fermentation.The many methods of document report：Wash detoxification, physics detoxification (is concentrated in vacuo air- extraction, UF membrane
Method), chemical detoxication (being lime neutralization, activated carbon adsorption, ion exchange, solvent extraction), biological detoxication etc. is to lignocellulosic
Hydrolyzate carries out detoxification.Such as：Bioprocess Biosyst Eng(2013)36:659-666 are begged for using active carbon adsorption
Influence of the fermentation inhibitors such as furfural, HMF, levulic acid to yeast cell growth speed is discussed；Patent CA102226204B is public
Open and soluble electrolytic salt is added in pending liquid glucose, then heating is carried out in membrane distillation removal liquid glucose by membrane module
Fermentation inhibitor.But the mode such as washing, physics, chemical detoxication consumes great lot of water resources, equipment investment cost height, enzymolysis hair
Ferment technique is complicated, adds alcohol production cycle and alcohol production cost.
The content of the invention
The present invention be directed to the heat transfer in lignocellulosic enzymolysis process, mass transfer and produce excessive toxic chemical pair
A kind of suppression problem of alcohol fermentation, there is provided lignocellulosic substep diastatic fermentation method using polyethylene glycol-water as medium.This hair
Mortifier is few in hydrolysis sugar liquid prepared by bright method, fermentable sugars concentration is high, that is, solves in lignocellulolyticenzymes hydrolytic process
The mortifier of generation produces glucose fermentation the negative effect of ethanol, and eliminates the concentration process of hydrolysis sugar liquid.
A kind of lignocellulosic substep diastatic fermentation method using polyethylene glycol-water as medium, concretely comprises the following steps：
(1) lignocellulosic of pretreatment is mixed according to solid substrate with fermentation media by certain solid-to-liquid ratio,
Then cellulase is added, just stage enzymatic hydrolysis is carried out under the conditions of pH is 4.0-5.5, temperature is 40-60 DEG C；
(2) the enzymolysis slurries obtained in step (1) are subjected to separation of solid and liquid, the wooden of pretreatment is added into gained liquid
Cellulose, cellulase, carry out enzyme hydrolysis again under the conditions of 40-60 DEG C；
(3) the enzymolysis slurries obtained in step (2) are subjected to separation of solid and liquid again, can obtain can ferment containing higher concentration
The hydrolyzate of sugar.
(4) saccharomyces cerevisiae is added in the hydrolyzate containing fermentable sugars obtained into step (3) step, is 4.0- in pH
5.5th, temperature carries out fermentation producing and ethanol under the conditions of being 28-40 DEG C.
The lignocellulosic material main component of the pretreatment is the agriculture containing cellulose, hemicellulose, lignin
Woods discarded object or fast-growing energy-source plant.The branch that raw material main source is wheat stalk, maize straw or/and straw, lumbering produces
One or more in leaf, discarded wood or/and sawdust, sugar grass or/and withy journey.
The preprocess method that the lignocellulosic of heretofore described pretreatment is applicable in includes chemical method, Physical, change
- Physical and bioanalysis etc..
Enzymolysis medium described in step (1) of the present invention is the matter of polyethylene glycol and the mixture of water, polyethylene glycol and water
It is 2%-50% to measure ratio.The molecular weight polyethylene glycol is 200~8000, is preferably 200-2000；
The cellulose of pretreatment described in step (1) of the present invention, (2) and the solid-to-liquid ratio (grams per milliliter) of enzymolysis medium are equal
For 5%-50%；The additive amount of cellulase is 10-80FPU/g lignocellulosic materials；
The lignocellulosic enzymolysis time of pretreatment described in step (1) of the present invention, (2) for 2-120 it is small when；The wine
The OD of the cell density of brewer yeast600It is worth and is：0.2~20, it is preferably 8.0~10；
Compared with the prior art, a kind of lignocellulosic substep diastatic fermentation using polyethylene glycol-water as medium of the present invention produces
The method of ethanol has the following advantages that：The method of the present invention batch feeding enzymolysis alleviates heat transfer, effect of mass transmitting, can obtain high concentration also
The hydrolyzate of raw sugar；The solid residue centrifuged after enzymolysis can be as the adsorbent of fermentation toxicity inhibition thing, the hydrolysis of preparation
Mortifier is few in liquid glucose；Polyethylene glycol can effectively alleviate suppression of the residual toxic chemicals to alcohol fermentation；Enzymolysis liquid is without de-
Poison processing can be directly used for fermenting, and can simplify production technology, reduce equipment investment.
(1) the method for the present invention batch feeding is advantageously used for the heat transfer of enzymolysis process, mass transfer, can obtain high-concentration reduced sugar
Hydrolyzate；
(2) solid residue centrifuged after digesting can be removed in hydrolyzate as the adsorbent of fermentation toxicity inhibition thing
Most of fermentation inhibitor；
(3) polyethylene glycol can effectively alleviate suppression of the remaining toxic chemical to alcohol fermentation
(4) enzymolysis liquid can be directly used for alcohol fermentation without detoxification treatment, can effectively simplify production technology, reduce equipment throwing
Money.
The method of the present invention solves the toxicity inhibition thing produced in lignocellulolyticenzymes hydrolytic process to glucose fermentation
The negative effect of ethanol is produced, and eliminates the concentration process of hydrolysis sugar liquid, reduces ethanol distillation energy consumption, so as to effectively reduce fine
The totle drilling cost of the plain alcohol production of dimension.
Brief description of the drawings
Fig. 1 is a kind of technique stream using polyethylene glycol-water as the lignocellulosic substep diastatic fermentation method of medium of the invention
Cheng Tu.
Embodiment
With reference to specific embodiment, the present invention is described further, but protection scope of the present invention is from embodiment
Limitation, the content described in following embodiments and specification merely illustrates the principles of the invention, and is not departing from spirit of the invention
On the premise of scope, various changes and modifications of the present invention are possible, these changes and improvements both fall within this claimed hair
In bright scope.The scope of the present invention is defined by the appended claims and its equivalents.
A kind of process flow chart such as Fig. 1 institutes using polyethylene glycol-water as the lignocellulosic substep diastatic fermentation method of medium
Show.
Embodiment 1
The Fast growth poplar of steam explosion method pretreatment, polyethylene glycol (PEG-1000), deionized water are mixed according to solid-to-liquid ratio 10%
Close, polyethylene glycol and deionized water quality are than 20%, then addition cellulase 15FPU/g substrates, pH be 4.85, temperature be
When carrying out that just stage enzymatic hydrolysis 24 is small under the conditions of 50 DEG C, the enzymolysis slurries of acquisition are subjected to separation of solid and liquid, grape in gained hydrolyzate
Sugared concentration is 40g/L；Fast growth poplar, the cellulase 15FPU/ of steam explosion method pretreatment are added by solid-to-liquid ratio 10% into hydrolyzed liquid
G substrates carry out enzyme hydrolysis 24h again, the enzymolysis slurries of acquisition are carried out separation of solid and liquid again, the glucose for obtaining hydrolyzate is dense
Spend for 70g/L；Saccharomyces cerevisiae OD is added into the hydrolyzate of acquisition600For 2.0, temperature carries out fermentation production second under the conditions of being 33 DEG C
Alcohol, when fermentation 96 is small after the concentration of alcohol that obtains be 30g/L.
Embodiment 2
The Fast growth poplar of steam explosion method pretreatment, polyethylene glycol (PEG-200), deionized water are mixed according to solid-to-liquid ratio 10%
Close, polyethylene glycol and deionized water quality are than 20%, then addition cellulase 15FPU/g substrates, pH be 4.85, temperature be
When carrying out that just stage enzymatic hydrolysis 24 is small under the conditions of 50 DEG C, the enzymolysis slurries of acquisition are subjected to separation of solid and liquid, grape in gained hydrolyzate
Sugared concentration is 41g/L；Fast growth poplar, the cellulase 15FPU/ of steam explosion method pretreatment are added by solid-to-liquid ratio 10% into hydrolyzed liquid
G substrates are carried out when enzyme hydrolysis 24 is small again, and the enzymolysis slurries of acquisition are carried out separation of solid and liquid again, obtain the glucose of hydrolyzate
Concentration is 72g/L；Saccharomyces cerevisiae OD is added into the hydrolyzate of acquisition600For 2.0, temperature carries out fermentation production under the conditions of being 33 DEG C
Ethanol, when fermentation 96 is small after the concentration of alcohol that obtains be 26g/L.
Embodiment 3
The stalk of steam explosion method pretreatment, polyethylene glycol (PEG-2000), deionized water are mixed according to solid-to-liquid ratio 10%,
Polyethylene glycol and deionized water quality are than 20%, then addition cellulase 15FPU/g substrates, pH be 4.85, temperature 50
When carrying out that just stage enzymatic hydrolysis 24 is small under the conditions of DEG C, the enzymolysis slurries of acquisition are subjected to separation of solid and liquid, glucose in gained hydrolyzate
Concentration is 45g/L；Fast growth poplar, the cellulase 15FPU/g of steam explosion method pretreatment are added by solid-to-liquid ratio 10% into hydrolyzed liquid
Substrate is carried out when enzyme hydrolysis 24 is small again, and the enzymolysis slurries of acquisition are carried out separation of solid and liquid again, obtain the glucose of hydrolyzate
Concentration is 86g/L；Saccharomyces cerevisiae OD is added into the hydrolyzate of acquisition600For 2.0, temperature carries out fermentation production under the conditions of being 33 DEG C
Ethanol, when fermentation 96 is small after the concentration of alcohol that obtains be 35g/L.
Embodiment 4
The stalk of steam explosion method pretreatment, polyethylene glycol (PEG-8000), deionized water are mixed according to solid-to-liquid ratio 10%,
Polyethylene glycol and deionized water quality are than 20%, then addition cellulase 15FPU/g substrates, pH be 4.85, temperature 50
When carrying out that just stage enzymatic hydrolysis 24 is small under the conditions of DEG C, the enzymolysis slurries of acquisition are subjected to separation of solid and liquid, glucose in gained hydrolyzate
Concentration is 43g/L；Fast growth poplar, the cellulase 15FPU/g of steam explosion method pretreatment are added by solid-to-liquid ratio 10% into hydrolyzed liquid
Substrate carries out enzyme hydrolysis 24h again, and the enzymolysis slurries of acquisition are carried out separation of solid and liquid again, obtain the concentration of glucose of hydrolyzate
For 78g/L；Saccharomyces cerevisiae OD is added into the hydrolyzate of acquisition600For 2.0, temperature carries out fermentation producing and ethanol under the conditions of being 33 DEG C,
Ferment 96 it is small when after the concentration of alcohol that obtains be 32g/L.
Embodiment 5
The stalk of steam explosion method pretreatment, polyethylene glycol (PEG-1000), deionized water are mixed according to solid-to-liquid ratio 10%,
Polyethylene glycol and deionized water quality are than 20%, then addition cellulase 10FPU/g substrates, pH be 4.85, temperature 50
When carrying out that just stage enzymatic hydrolysis 24 is small under the conditions of DEG C, the enzymolysis slurries of acquisition are subjected to separation of solid and liquid, glucose in gained hydrolyzate
Concentration is 30g/L；Fast growth poplar, the cellulase 10FPU/g of steam explosion method pretreatment are added by solid-to-liquid ratio 10% into hydrolyzed liquid
Substrate carries out enzyme hydrolysis 24h again, and the enzymolysis slurries of acquisition are carried out separation of solid and liquid again, obtain the concentration of glucose of hydrolyzate
For 48g/L；Saccharomyces cerevisiae OD is added into the hydrolyzate of acquisition600For 2.0, temperature carries out fermentation producing and ethanol under the conditions of being 33 DEG C,
Ferment 96 it is small when after the concentration of alcohol that obtains be 19g/L.
Embodiment 6
The stalk of steam explosion method pretreatment, polyethylene glycol (PEG-1000), deionized water are mixed according to solid-to-liquid ratio 10%,
Polyethylene glycol and deionized water quality are than 20%, then addition cellulase 15FPU/g substrates, pH be 4.85, temperature 50
When carrying out that just stage enzymatic hydrolysis 24 is small under the conditions of DEG C, the enzymolysis slurries of acquisition are subjected to separation of solid and liquid, glucose in gained hydrolyzate
Concentration is 50g/L；Fast growth poplar, the cellulase 15FPU/g of steam explosion method pretreatment are added by solid-to-liquid ratio 10% into hydrolyzed liquid
Substrate is carried out when enzyme hydrolysis 24 is small again, and the enzymolysis slurries of acquisition are carried out separation of solid and liquid again, obtain the glucose of hydrolyzate
Concentration is 90g/L；Saccharomyces cerevisiae OD is added into the hydrolyzate of acquisition600For 2.0, temperature carries out fermentation production under the conditions of being 33 DEG C
Ethanol, when fermentation 96 is small after the concentration of alcohol that obtains be 42g/L.
Embodiment 7
The stalk of steam explosion method pretreatment, polyethylene glycol (PEG-1000), deionized water are mixed according to solid-to-liquid ratio 10%,
Polyethylene glycol and deionized water quality are than 20%, then addition cellulase 30FPU/g substrates, pH be 4.85, temperature 50
When carrying out that just stage enzymatic hydrolysis 24 is small under the conditions of DEG C, the enzymolysis slurries of acquisition are subjected to separation of solid and liquid, glucose in gained hydrolyzate
Concentration is 60g/L；Fast growth poplar, the cellulase 30FPU/g of steam explosion method pretreatment are added by solid-to-liquid ratio 10% into hydrolyzed liquid
Substrate carries out enzyme hydrolysis 24h again, and the enzymolysis slurries of acquisition are carried out separation of solid and liquid again, obtain the concentration of glucose of hydrolyzate
For 110g/L；Saccharomyces cerevisiae OD is added into the hydrolyzate of acquisition600For 2.0, temperature carries out fermentation production second under the conditions of being 33 DEG C
Alcohol, when fermentation 96 is small after the concentration of alcohol that obtains be 50g/L.
Embodiment 8
Fast growth poplar furfural dregs, polyethylene glycol (PEG-2000), deionized water are mixed according to solid-to-liquid ratio 15%, poly- second two
Alcohol and deionized water quality are than 20%, then addition cellulase 50FPU/g substrates, pH be 4.85, temperature be 50 DEG C of conditions
When the first stage enzymatic hydrolysis 8 of lower progress is small, the enzymolysis slurries of acquisition are subjected to separation of solid and liquid, concentration of glucose is in gained hydrolyzate
65g/L；Into hydrolyzed liquid by solid-to-liquid ratio 15% add steam explosion method pretreatment Fast growth poplar, cellulase 50FPU/g substrates into
The enzymolysis slurries of acquisition are carried out separation of solid and liquid, the concentration of glucose for obtaining hydrolyzate is 116g/ by capable enzyme hydrolysis 8h again again
L；Saccharomyces cerevisiae OD is added into the hydrolyzate of acquisition600For 2.0, temperature carries out fermentation producing and ethanol, fermentation 96 under the conditions of being 33 DEG C
The concentration of alcohol obtained after hour is 55g/L.
Embodiment 9
Stalk furfural dregs, polyethylene glycol (PEG-400), deionized water are mixed according to solid-to-liquid ratio 15%, polyethylene glycol
With deionized water quality than 20%, cellulase 80FPU/g substrates are then added, under the conditions of pH is 4.85, temperature is 50 DEG C
When the first stage enzymatic hydrolysis 8 of progress is small, the enzymolysis slurries of acquisition are subjected to separation of solid and liquid, concentration of glucose is 63g/ in gained hydrolyzate
L；The Fast growth poplar of steam explosion method pretreatment is added by solid-to-liquid ratio 15%, cellulase 80FPU/g substrates carry out again into hydrolyzed liquid
The enzymolysis slurries of acquisition are carried out separation of solid and liquid by secondary enzyme hydrolysis 8h again, and the concentration of glucose for obtaining hydrolyzate is 114g/L；To
Saccharomyces cerevisiae OD is added in the hydrolyzate of acquisition600For 2.0, temperature carries out fermentation producing and ethanol under the conditions of being 33 DEG C, when fermentation 96 is small
The concentration of alcohol obtained afterwards is 52g/L.
Embodiment 10
Stalk furfural dregs, polyethylene glycol (PEG-4000), deionized water are mixed according to solid-to-liquid ratio 20%, polyethylene glycol
With deionized water quality than 20%, cellulase 60FPU/g substrates are then added, under the conditions of pH is 4.85, temperature is 40 DEG C
When the first stage enzymatic hydrolysis 72 of progress is small, the enzymolysis slurries of acquisition are subjected to separation of solid and liquid, concentration of glucose is in gained hydrolyzate
55g/L；Into hydrolyzed liquid by solid-to-liquid ratio 20% add steam explosion method pretreatment Fast growth poplar, cellulase 60FPU/g substrates into
The enzymolysis slurries of acquisition are carried out separation of solid and liquid, the concentration of glucose for obtaining hydrolyzate is by capable enzyme hydrolysis 72h again again
100g/L；Saccharomyces cerevisiae OD is added into the hydrolyzate of acquisition600For 8.0, temperature carries out fermentation producing and ethanol under the conditions of being 38 DEG C,
Ferment 96 it is small when after the concentration of alcohol that obtains be 42g/L.
Embodiment 11
Stalk furfural dregs, polyethylene glycol (PEG-4000), deionized water are mixed according to solid-to-liquid ratio 20%, polyethylene glycol
With deionized water quality than 20%, cellulase 60FPU/g substrates are then added, under the conditions of pH is 4.85, temperature is 50 DEG C
When the first stage enzymatic hydrolysis 72 of progress is small, the enzymolysis slurries of acquisition are subjected to separation of solid and liquid, concentration of glucose is in gained hydrolyzate
85g/L；Into hydrolyzed liquid by solid-to-liquid ratio 20% add steam explosion method pretreatment Fast growth poplar, cellulase 60FPU/g substrates into
The enzymolysis slurries of acquisition are carried out separation of solid and liquid, the concentration of glucose for obtaining hydrolyzate is by capable enzyme hydrolysis 72h again again
170g/L；Saccharomyces cerevisiae OD is added into the hydrolyzate of acquisition600For 8.0, temperature carries out fermentation producing and ethanol under the conditions of being 38 DEG C,
Ferment 96 it is small when after the concentration of alcohol that obtains be 73g/L.
Embodiment 12
Stalk furfural dregs, polyethylene glycol (PEG-4000), deionized water are mixed according to solid-to-liquid ratio 20%, polyethylene glycol
With deionized water quality than 20%, cellulase 60FPU/g substrates are then added, under the conditions of pH is 4.85, temperature is 60 DEG C
When the first stage enzymatic hydrolysis 72 of progress is small, the enzymolysis slurries of acquisition are subjected to separation of solid and liquid, concentration of glucose is in gained hydrolyzate
88g/L；Into hydrolyzed liquid by solid-to-liquid ratio 20% add steam explosion method pretreatment Fast growth poplar, cellulase 60FPU/g substrates into
The enzymolysis slurries of acquisition are carried out separation of solid and liquid, the concentration of glucose for obtaining hydrolyzate is by capable enzyme hydrolysis 72h again again
174g/L；Saccharomyces cerevisiae OD is added into the hydrolyzate of acquisition600For 8.0, temperature carries out fermentation producing and ethanol under the conditions of being 38 DEG C,
Ferment 96 it is small when after the concentration of alcohol that obtains be 75g/L.
Embodiment 13
Stalk furfural dregs, polyethylene glycol (PEG-4000), deionized water are mixed according to solid-to-liquid ratio 20%, polyethylene glycol
With deionized water quality than 5%, then add cellulase 60FPU/g substrates, under the conditions of pH is 4.85, temperature is 50 DEG C into
When the first stage enzymatic hydrolysis 72 of row is small, the enzymolysis slurries of acquisition are subjected to separation of solid and liquid, concentration of glucose is 84g/ in gained hydrolyzate
L；The Fast growth poplar of steam explosion method pretreatment is added by solid-to-liquid ratio 20%, cellulase 60FPU/g substrates carry out again into hydrolyzed liquid
The enzymolysis slurries of acquisition are carried out separation of solid and liquid by secondary enzyme hydrolysis 72h again, and the concentration of glucose for obtaining hydrolyzate is 172g/L；
Saccharomyces cerevisiae OD is added into the hydrolyzate of acquisition600For 8.0, temperature carries out fermentation producing and ethanol under the conditions of being 38 DEG C, and fermentation 96 is small
When after the concentration of alcohol that obtains be 18g/L.
Embodiment 14
Stalk furfural dregs, polyethylene glycol (PEG-4000), deionized water are mixed according to solid-to-liquid ratio 30%, polyethylene glycol
With deionized water quality than 20%, cellulase 80FPU/g substrates are then added, under the conditions of pH is 4.85, temperature is 50 DEG C
When the first stage enzymatic hydrolysis 72 of progress is small, the enzymolysis slurries of acquisition are subjected to separation of solid and liquid, concentration of glucose is in gained hydrolyzate
100g/L；Into hydrolyzed liquid by solid-to-liquid ratio 30% add steam explosion method pretreatment Fast growth poplar, cellulase 80FPU/g substrates into
The enzymolysis slurries of acquisition are carried out separation of solid and liquid, the concentration of glucose for obtaining hydrolyzate is by capable enzyme hydrolysis 72h again again
200g/L；Saccharomyces cerevisiae OD is added into the hydrolyzate of acquisition600For 10.0, temperature carries out fermentation producing and ethanol under the conditions of being 33 DEG C,
Ferment 96 it is small when after the concentration of alcohol that obtains be 92g/L.
Embodiment 15
Stalk furfural dregs, polyethylene glycol (PEG-4000), deionized water are mixed according to solid-to-liquid ratio 30%, polyethylene glycol
With deionized water quality than 20%, cellulase 80FPU/g substrates are then added, under the conditions of pH is 4.85, temperature is 50 DEG C
When the first stage enzymatic hydrolysis 72 of progress is small, the enzymolysis slurries of acquisition are subjected to separation of solid and liquid, concentration of glucose is in gained hydrolyzate
105g/L；Into hydrolyzed liquid by solid-to-liquid ratio 30% add steam explosion method pretreatment Fast growth poplar, cellulase 80FPU/g substrates into
The enzymolysis slurries of acquisition are carried out separation of solid and liquid, the concentration of glucose for obtaining hydrolyzate is by capable enzyme hydrolysis 72h again again
204g/L；Saccharomyces cerevisiae OD is added into the hydrolyzate of acquisition600For 8.0, temperature carries out fermentation producing and ethanol under the conditions of being 38 DEG C,
Ferment 96 it is small when after the concentration of alcohol that obtains be 94g/L.
Embodiment 16
Stalk furfural dregs, polyethylene glycol (PEG-4000), deionized water are mixed according to solid-to-liquid ratio 30%, polyethylene glycol
With deionized water quality than 20%, cellulase 80FPU/g substrates are then added, under the conditions of pH is 4.85, temperature is 50 DEG C
When the first stage enzymatic hydrolysis 8 of progress is small, the enzymolysis slurries of acquisition are subjected to separation of solid and liquid, concentration of glucose is 78g/ in gained hydrolyzate
L；The Fast growth poplar of steam explosion method pretreatment is added by solid-to-liquid ratio 30%, cellulase 80FPU/g substrates carry out again into hydrolyzed liquid
The enzymolysis slurries of acquisition are carried out separation of solid and liquid by secondary enzyme hydrolysis 8h again, and the concentration of glucose for obtaining hydrolyzate is 135g/L；To
Saccharomyces cerevisiae OD is added in the hydrolyzate of acquisition600For 8.0, temperature carries out fermentation producing and ethanol under the conditions of being 38 DEG C, when fermentation 96 is small
The concentration of alcohol obtained afterwards is 58g/L.
Embodiment 17
Stalk furfural dregs, polyethylene glycol (PEG-4000), deionized water are mixed according to solid-to-liquid ratio 30%, polyethylene glycol
With deionized water quality than 2%, then add cellulase 80FPU/g substrates, under the conditions of pH is 4.85, temperature is 50 DEG C into
When the first stage enzymatic hydrolysis 8 of row is small, the enzymolysis slurries of acquisition are subjected to separation of solid and liquid, concentration of glucose is 77g/L in gained hydrolyzate；
The Fast growth poplar of steam explosion method pretreatment is added by solid-to-liquid ratio 30%, cellulase 80FPU/g substrates carry out again into hydrolyzed liquid
The enzymolysis slurries of acquisition are carried out separation of solid and liquid by enzyme hydrolysis 8h again, and the concentration of glucose for obtaining hydrolyzate is 132g/L；To obtaining
Saccharomyces cerevisiae OD is added in the hydrolyzate obtained600For 8.0, temperature carries out fermentation producing and ethanol under the conditions of being 38 DEG C, when fermentation 96 is small after
The concentration of alcohol of acquisition is 15g/L.
Embodiment 18
Stalk furfural dregs, polyethylene glycol (PEG-4000), deionized water are mixed according to solid-to-liquid ratio 30%, polyethylene glycol
With deionized water quality than 20%, cellulase 80FPU/g substrates are then added, under the conditions of pH is 4.85, temperature is 50 DEG C
When the first stage enzymatic hydrolysis 8 of progress is small, the enzymolysis slurries of acquisition are subjected to separation of solid and liquid, concentration of glucose is 78g/ in gained hydrolyzate
L；The Fast growth poplar of steam explosion method pretreatment is added by solid-to-liquid ratio 30%, cellulase 80FPU/g substrates carry out again into hydrolyzed liquid
The enzymolysis slurries of acquisition are carried out separation of solid and liquid by secondary enzyme hydrolysis 8h again, and the concentration of glucose for obtaining hydrolyzate is 135g/L；To
Saccharomyces cerevisiae OD is added in the hydrolyzate of acquisition600For 8.0, temperature carries out fermentation producing and ethanol under the conditions of being 38 DEG C, when fermentation 96 is small
The concentration of alcohol obtained afterwards is 58g/L.
Embodiment 19
Stalk furfural dregs, polyethylene glycol (PEG-400), deionized water are mixed according to solid-to-liquid ratio 10%, polyethylene glycol
With deionized water quality than 50%, cellulase 15FPU/g substrates are then added, under the conditions of pH is 4.85, temperature is 50 DEG C
When the first stage enzymatic hydrolysis 8 of progress is small, the enzymolysis slurries of acquisition are subjected to separation of solid and liquid, concentration of glucose is 42g/ in gained hydrolyzate
L；The Fast growth poplar of steam explosion method pretreatment is added by solid-to-liquid ratio 10%, cellulase 15FPU/g substrates carry out again into hydrolyzed liquid
The enzymolysis slurries of acquisition are carried out separation of solid and liquid by secondary enzyme hydrolysis 8h again, and the concentration of glucose for obtaining hydrolyzate is 68g/L；To
Saccharomyces cerevisiae OD is added in the hydrolyzate of acquisition600For 8.0, temperature carries out fermentation producing and ethanol under the conditions of being 38 DEG C, when fermentation 96 is small
The concentration of alcohol obtained afterwards is 29g/L.
Comparative example 1
Contrasted with embodiment 1, the Fast growth poplar of steam explosion method pretreatment is mixed with deionized water according to solid-to-liquid ratio 10%
Close, then add cellulase 15FPU/g substrates, it is small that just stage enzymatic hydrolysis 24 is carried out under the conditions of pH is 4.85, temperature is 50 DEG C
When, the enzymolysis slurries of acquisition are subjected to separation of solid and liquid, concentration of glucose is 38g/L in gained hydrolyzate；Pressed into hydrolyzed liquid
Solid-to-liquid ratio 10% adds the Fast growth poplar of steam explosion method pretreatment, cellulase 15FPU/g substrates carry out enzyme hydrolysis 24h again, will obtain
The enzymolysis slurries obtained carry out separation of solid and liquid again, and the concentration of glucose for obtaining hydrolyzate is 65g/L；Add into the hydrolyzate of acquisition
Add saccharomyces cerevisiae OD600For 2.0, temperature carries out fermentation producing and ethanol under the conditions of being 33 DEG C, when fermentation 96 is small after the concentration of alcohol that obtains
For 10g/L.
So as to obtain high ethano concentration.It can be seen from the above that polyethylene glycol is ligno-cellulose hydrolysate original position detoxification fermentation system
The key factor of ethanol.Certainly, the dosage of cellulase, solid-to-liquid ratio, hydrolysis temperature, enzymolysis time, fermentation temperature, fermentation time
It is also interrelated etc. factor, really need to adjust parameters as the case may be during the technique is implemented, to up to
To preferable ferment effect.
Claims (13)
- A kind of 1. lignocellulosic substep diastatic fermentation method using polyethylene glycol-water as medium, it is characterised in that the step of this method Suddenly it is：(1) by the lignocellulosic solid substrate of pretreatment with digesting being mixed according to certain solid-to-liquid ratio for medium, Ran Houtian Add cellulase, tentatively digested under the conditions of pH is 4.0-5.5, temperature is 40-60 DEG C；(2) the enzymolysis slurries obtained in step (1) are subjected to separation of solid and liquid, the wood fibre of pretreatment is added into gained liquid Element, cellulase, are digested again under the conditions of 40-60 DEG C；(3) the enzymolysis slurries obtained in step (2) are subjected to separation of solid and liquid again, can obtained containing higher concentration fermentable sugars Hydrolyzate；(4) saccharomyces cerevisiae is added in the hydrolyzate containing fermentable sugars obtained into step (3) step, is 4.0-5.5, temperature in pH Spend for 28-40 DEG C, fermentation time be 4-120h under the conditions of carry out fermentation producing and ethanol.
- 2. a kind of lignocellulosic substep diastatic fermentation method using polyethylene glycol-water as medium described in accordance with the claim 1, its It is characterized in that：Medium is digested as polyethylene glycol and the mixture of water described in step (1), and the mass ratio of polyethylene glycol and water is 2%-50%.
- 3. a kind of lignocellulosic substep diastatic fermentation method using polyethylene glycol-water as medium described in accordance with the claim 3, its It is characterized in that：The molecular weight polyethylene glycol is 200~8000, is preferably 200-2000.
- 4. a kind of lignocellulosic substep diastatic fermentation method using polyethylene glycol-water as medium described in accordance with the claim 1, its It is characterized in that：The cellulose of pretreatment described in step (1), (2) and the solid-to-liquid ratio of enzymolysis medium are 5-50%g/ml；It is fine The additive amount of the plain enzyme of dimension is 10-80FPU/g lignocellulosic materials.
- 5. a kind of lignocellulosic substep diastatic fermentation method using polyethylene glycol-water as medium described in accordance with the claim 1, its It is characterized in that：The lignocellulosic enzymolysis time of pretreatment described in step (1) of the present invention, (2) for 2-120 it is small when.
- 6. a kind of lignocellulosic substep diastatic fermentation method using polyethylene glycol-water as medium described in accordance with the claim 1, its It is characterized in that：The OD of the cell density of saccharomyces cerevisiae described in step (4)600It is worth and is：0.2~20, it is preferably 8~10.
- 7. a kind of lignocellulosic substep diastatic fermentation method using polyethylene glycol-water as medium described in accordance with the claim 1, its It is characterized in that：The preprocess method that the lignocellulosic of the pretreatment is applicable in includes chemical method, Physical, chemical-physical Method and bioanalysis.
- 8. a kind of lignocellulosic substep diastatic fermentation method using polyethylene glycol-water as medium described in accordance with the claim 1, its It is characterized in that：The lignocellulosic material main component of the pretreatment is the agriculture containing cellulose, hemicellulose, lignin Woods discarded object or fast-growing energy-source plant.
- 9. a kind of lignocellulosic substep diastatic fermentation method using polyethylene glycol-water as medium described in accordance with the claim 1, its It is characterized in that：The lignocellulosic of the pretreatment from agricultural wastes, forestry waste, special energy crop or/ With the discarded object of various containing celluloses.
- 10. according to a kind of lignocellulosic substep diastatic fermentation method using polyethylene glycol-water as medium described in claim 9, It is characterized in that：The agricultural wastes are wheat stalk, maize straw or/and straw.
- 11. according to a kind of lignocellulosic substep diastatic fermentation method using polyethylene glycol-water as medium described in claim 9, It is characterized in that：Branches and leaves, discarded wood or/and the sawdust that the forestry waste produces for lumbering.
- 12. according to a kind of lignocellulosic substep diastatic fermentation method using polyethylene glycol-water as medium described in claim 9, It is characterized in that：The special energy crop is sugar grass or/and withy journey.
- 13. according to a kind of lignocellulosic substep diastatic fermentation method using polyethylene glycol-water as medium described in claim 9, It is characterized in that：The discarded object of the various containing celluloses is urban solid garbage, waste paper or/and bagasse.
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