CN105132470A - Method for enzyme recycling in enzymolysis process of lignocellulose hydrolase - Google Patents
Method for enzyme recycling in enzymolysis process of lignocellulose hydrolase Download PDFInfo
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- CN105132470A CN105132470A CN201510636139.XA CN201510636139A CN105132470A CN 105132470 A CN105132470 A CN 105132470A CN 201510636139 A CN201510636139 A CN 201510636139A CN 105132470 A CN105132470 A CN 105132470A
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- 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
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- Y02E—REDUCTION OF GREENHOUSE GAS [GHG] EMISSIONS, RELATED TO ENERGY GENERATION, TRANSMISSION OR DISTRIBUTION
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Abstract
The invention belongs to the technical field of cellulosic ethanol production, and discloses a method for enzyme recycling in enzymolysis process of lignocellulose hydrolase. The method specifically comprises the following steps that an enzymolysis tank is started, materials and water are continuously and evenly put into the enzymolysis tank in batches according to the set substrate concentration, and meanwhile the lignocellulose hydrolase is continuously and evenly put into the enzymmolysis tank in batches for enzymolysis; after enzymolysis is completed, the temperature of enzymatic hydrolysate is lowered to 30-32 DEG C, saccaromyces cerevisiae is inoculated, and fermentation is performed; obtained fermentation liquor is guided into a vacuum distillation tower, and negative-pressure distillation is performed; residual distillation liquor obtained through negative-pressure distillation is guided into the enzymolysis tank, enzyme recycling is performed for 1-2 times in a cycling mode. According to the method, through negative-pressure distillation, the activity of the residual lignocellulose hydrolase in the fermentation liquor is kept; through recycling, the residual distillation liquor is added in a new enzymolysis system, therefore, the usage amount of the ignocellulose hydrolase can be lowered by 20-25%, the utilization rate of the ignocellulose hydrolase is greatly increased, and then the production cost is reduced.
Description
Technical field
The invention belongs to the technical field that cellulose ethanol is produced, be specifically related to the method for enzyme recycle in a kind of lignocellulose enzymolysis process.
Background technology
Lignocellulose raw material is renewable resources abundant, the most cheap on the earth.The wood fiber biomass that the whole world is produced by photosynthesis is every year up to 1,000 hundred million tons.China is large agricultural country, has abundant biomass resource, and only agricultural crop straw one just has nearly 8 × 10 every year
8ton (being roughly equal to 400,000,000 tons of mark coals), account for 10 ~ 15% of national primary energy consumption total amount, add forestry fiber waste material and the industrial fiber waste residue of enormous amount, annual available lignocellulose raw material total amount can reach more than 2,000,000,000 tons.In lignocellulose raw material, Mierocrystalline cellulose accounts for 35 ~ 45% of dry weight, hemicellulose accounts for 20 ~ 40%, adopt appropriate technology that they are hydrolyzed into fermentable sugar, further fermentative production of ethanol, make full use of the pressure that these biomass resources can alleviate china natural resources shortage, alleviate environmental pollution, promote Agro-ecology benign cycle, to construction, energy-saving low-carbon society is significant.
The production process of usual cellulose ethanol comprises the pre-treatment of raw material, enzymic hydrolysis, fermentation and distilation, and wherein enzymolysis is the whole cellulose ethanol major technique of producing and economical one.At present, the topmost problem of cellulose hydrolysis saccharification is that enzymolysis efficiency is low, and the enzymolysis cycle is long, and enzyme dosage is large, and the not high aspect of transformation efficiency, these problems seriously constrain bio-transformation and the utilization of lignocellulose raw material.Cellulase preparation be costly one of principal element affecting bio-ethanol price, the technology reducing cellulase usage quantity or expense becomes bio-ethanol and produces emphasis.
Cellulase is adsorbed on above substrate at the hydrolysis of lignocellulose initial stage, after reaction terminates, due to the change of concentration of substrate, enzyme spins off gradually from substrate, become the state of resolvase, be scattered in enzymolysis solution, another part is then adsorbed in the enzymolysis residue of high lignin content.Resolvase in enzymolysis solution, by recycling, gives full play to it and is worth.
Realize efficient enzymolysis, reduce enzymolysis cost, can set about from following three: the pre-treatment 1) expected; 2) reduce the industrial production cost of lytic enzyme as far as possible, enzyme cost is reduced to certain limit, and the whole cost of investment of cellulose ethanol just can have competitive edge with oil; 3) effective fiber element the recycling and reusing of enzyme, even if the enzyme in enzymic hydrolysis process participates in reaction fully, increases enzyme and cellulosic touch opportunity, and raising enzymolysis efficiency, realizes enzyme and recycle and reuse.
Patent CN101974570A discloses a kind of method of recycling cellulose complete components in fuel ethanol production, refer to after lignocellulose simultaneous saccharification and fermentation workshop section, utilize coupling fresh substrate heavily to adsorb the method with immobilized cellobiase technique recycling cellulase total composition, this invention relates to immobilization process, owing to there is very large mass transfer obstacle between immobilized enzyme and cellulose solids particle, in lignocellulose enzymatic hydrolysis system, be therefore only limitted to cellobiose enzyme immobilizatio recycling at present.
Summary of the invention
Technical problem to be solved by this invention is, underuses for traditional enzymolysis process free cellulose enzyme, provides the method for enzyme recycle in a kind of lignocellulose enzymolysis process, improves the utilization ratio of free cellulose enzyme, reduces production cost.
For solving the problems of the technologies described above, the technical solution adopted in the present invention is: the method for enzyme recycle in a kind of lignocellulose enzymolysis process, the series connection of enzymatic vessel, fermentor tank and distillation tower is adopted to carry out, first carry out enzymolysis by the feed ratio of setting, after enzymolysis terminates, cooling inoculation is fermented, again fermented liquid is imported vacuum still and carry out vacuum distillation, removing ethanol, live yeast inactivation, leave free cellulase, finally imported in enzymatic hydrolysis system by vinasse and carry out reuse, concrete steps are as follows:
Step 1): start enzymatic vessel, according to the concentration of substrate of setting, criticizes continuous uniform be added in enzymatic vessel by material, moisture, and continuous uniform adds lignocellulose lytic enzyme in batches simultaneously, carries out enzymolysis;
Step 2): after enzymolysis terminates, enzymolysis solution is cooled to 30 ~ 32 DEG C, inoculation yeast saccharomyces cerevisiae, ferments;
Step 3): by step 2) fermentation gained fermented liquid imports in vacuum still and carries out vacuum distillation;
Step 4): imported in enzymatic vessel by step 3) vacuum distillation gained vinasse and carry out enzyme reuse, cycle index is 1 ~ 2 time.
Further, the material described in step 1) is through pretreated lignocellulosic material, and described lignocellulosic material can be the one in maize straw, wheat stalk, rice straw, or is lignocellulosic material known in those skilled in the art; Described pre-treatment comprises the steps: first to be adopted by lignocellulosic material mechanical means to carry out pulverization process, particle diameter is made to reach 0.14 ~ 32 millimeter, then be conveyed into steam explosion tank by rotary conveyor to process, then make the water content of material be 50 ~ 65% through plate-and-frame filter press press filtration.
Further, the concentration of substrate set described in step 1) is 20 ~ 25%, described material and moisture are criticized continuous uniform be added in enzymatic vessel, the concrete grammar that simultaneously continuous uniform adds lignocellulose lytic enzyme is in batches: the weight adding material for the first time accounts for and always adds 1/8 ~ 1/6 of inventory, the amount adding lignocellulose lytic enzyme is for the first time always add enzyme amount 5/9 ~ 7/9, when enzymatic vessel completes always add inventory 1/2 ~ 3/4, once add remaining 2/9 ~ 4/9 enzyme amount, feed intake end;
Every gram of over dry material adds lignocellulose lytic enzyme 30 ~ 60FPU.
Further, in step 1), described material and moisture are criticized continuous uniform be added in enzymatic vessel, the concrete grammar that simultaneously continuous uniform adds lignocellulose lytic enzyme is in batches: the weight adding material for the first time accounts for and always adds 1/6 of inventory, the weight adding lignocellulose lytic enzyme is for the first time always add enzyme amount 2/3, when enzymatic vessel completes always add inventory 3/4, once add remaining 1/3 enzyme amount, feed intake end; Start to from enzymatic vessel the end that feeds intake, complete within 16h, enzymolysis proceeds to step 2 to during 48h).
Further, step 2) described in technological condition for fermentation be: leavening temperature is 30 ~ 32 DEG C, and inoculum size is 0.5 ~ 0.8 ‰, and rotating speed is 60 ~ 150r/min.
Further, the processing condition of vacuum distillation described in step 3) are: distillation temperature is 50 ~ 70 DEG C, and vacuum tightness is-1.2 ~-0.90Mpa, and distillation time is 0.4 ~ 1.0h, fully by complete for the ethanolic extraction in fermented liquid.
Further, the vinasse volume importing enzymatic vessel described in step 4) is 25% of the actual tinning volume of enzymatic vessel.
Compared with prior art, its beneficial effect is in the present invention: the live yeast in fermented liquid of the present invention is inactivation under the hot environment of vacuum distillation, the cellulase containing non-inactivation in the fermented liquid in vacuum still.The present invention passes through underpressure distillation, remain cellulase activity remaining in fermented liquid, by reuse, vinasse is added in new enzymatic hydrolysis system, the lignocellulose lytic enzyme consumption of 20 ~ 25% can be reduced, greatly improve the utilization ratio of cellulase, thus reduce production cost.
Embodiment
In order to understand the present invention better, set forth content of the present invention below in conjunction with embodiment is clear further, but protection content of the present invention is not only confined to the following examples.In the following description, a large amount of concrete details is given to provide more thorough understanding of the invention.But, it will be apparent to one skilled in the art that the present invention can be implemented without the need to these details one or more.In other example, in order to avoid obscuring with the present invention, technical characteristics more well known in the art are not described.
In following embodiment, lignocellulose lytic enzyme, also can be called cellulose complex enzyme or complex cellulase, and compound method is: cellulase, zytase, cellobiase are mixed in proportion, and adds 1 ‰ PEG4000, obtained.In the present invention, the filter paper enzyme activity (FPU) of described lignocellulose lytic enzyme is 800FPU/mL.Enzyme is lived and is defined as international unit (IU), and namely 1min produces enzyme amount needed for 1 μm of ol glucose is 1 Mei Huo unit.Cellulase, zytase, cellobiase, PEG4000 are common agents, by commercially available acquisition.
In following embodiment, yeast saccharomyces cerevisiae is provided by Angel Yeast Co., Ltd, its activation method is the ordinary skill in the art, method is in the present invention: take active dry yeast, adding the 2%(w/v through sterilising treatment) D/W is under 30 DEG C of conditions, aerlbic culture 30 ~ 45min, foaming is as the criterion, and can do inoculation and use.
In following embodiment 1 ~ 3, in enzymatic vessel, the density of enzymolysis solution is pressed 1.1kg/L and is calculated (in situ measurement numerical value), and the density of water is pressed 1.0kg/L and calculated.
Below only for wheat stalk, set forth technical scheme of the present invention and implement to enable those skilled in the art.Same, those skilled in the art can use other material, comprise in maize straw, rice straw and other known lignocellulosic material one or more, under the record or enlightenment of the technology of the present invention content, realize the recycle problem of enzyme in its enzymolysis process; Must drop within protection scope of the present invention.
Embodiment 1
For 50L enzymatic vessel, the present invention will be further described.
The method of enzyme recycle in a kind of lignocellulose enzymolysis process, the series connection of enzymatic vessel, fermentor tank and distillation tower is adopted to carry out, first feed intake by the concentration of substrate of setting and carry out enzymolysis, after enzymolysis terminates, cooling inoculation is fermented, again fermented liquid is imported in vacuum still and carry out vacuum distillation, removing ethanol, live yeast inactivation, leaves free cellulase, finally imported in enzymatic hydrolysis system by vinasse and carry out reuse, concrete steps are as follows:
1) tap water 19.5L is added in the 50L enzymatic vessel M1 after disappearing process to sky, open and stir, setting concentration of substrate is 25%(w/v), drop into M1 the wheat stalk 3.33kg(1/6 that water content is 50% (w/w)) by dog-house, to be mixed evenly after, add lignocellulose lytic enzyme 0.33L (2/3), control temperature of reaction 50 DEG C, start to enter enzymolysis process; Then continuous uniform charging, when inlet amount reaches 15kg (3/4), adds remaining lignocellulose lytic enzyme 0.17L, continues continuous uniform and feeds intake, and after starting to feed intake, in 16h, feed supplement reaches 20kg to total amount; Every gram of over dry material adds lignocellulose lytic enzyme 40FPU;
2), when enzymolysis is to 48h, be cooled to 30 DEG C of inoculation yeast saccharomyces cerevisiaes and ferment, controlling fermentation condition is: leavening temperature is 30 DEG C, and inoculum size is 0.5 ‰ (v/v), and rotating speed is 60r/min;
3) after fermentation ends, in 55 ± 2 DEG C, under the vacuum tightness of-0.95Mpa, underpressure distillation 0.8h is carried out to fermented liquid, fully by complete for ethanolic extraction wherein;
4) be 20%(w/v by 10L concentration of substrate) vinasse import in M1, add water 9.5L, lignocellulose lytic enzyme 0.33L, material 3.4kg successively, control temperature of reaction 50 ± 2 DEG C, start to enter second batch of enzymolysis process, the same step 1) of specific implementation method, circulate 1 end.
In the present embodiment, wheat stalk is through pretreated material, its pretreated method is: first adopted by wheat stalk mechanical means to carry out pulverization process, particle diameter is made to reach 0.14 ~ 32 millimeter, then be conveyed into steam explosion tank by rotary conveyor to process, then make the water content of material be 50% through plate-and-frame filter press press filtration.Wheat stalk described in following content is identical with the present embodiment, repeats no more.
As calculated, in loop cycle compared with intermittent type enzymatic saccharification zymotechnique, lignocellulose lytic enzyme consumption reduce 20%.
Embodiment 2
With 150m
3enzymatic vessel is that the present invention will be further described for example.
The method of enzyme recycle in a kind of lignocellulose enzymolysis process, the series connection of enzymatic vessel, fermentor tank and distillation tower is adopted to carry out, first feed intake by the concentration of substrate of setting and carry out enzymolysis, after enzymolysis terminates, cooling inoculation is fermented, again fermented liquid is imported vacuum still and carry out vacuum distillation, removing ethanol, live yeast inactivation, leaves free cellulase, finally imported in enzymatic hydrolysis system by vinasse and carry out reuse, concrete steps are as follows:
1) 150m after disappearing process to sky
3enzymatic vessel M1 adds tap water 58.5m
3, open and stir, setting concentration of substrate is 25%(w/v), dropping into water content by dog-house to M1 is the wheat stalk 10 tons (1/6) of 50% (w/w), to be mixed evenly after, add lignocellulose lytic enzyme 1.0m
3(2/3), control temperature of reaction 50 ± 1 DEG C, start to enter enzymolysis process, then continuous uniform charging, when inlet amount reaches 45 tons (3/4), add remaining lignocellulose lytic enzyme 0.5m
3, continue continuous uniform and feed intake, after starting to feed intake, in 16h, feed supplement to total amount reaches 60 tons; Every gram of over dry material adds lignocellulose lytic enzyme 40FPU;
2), when enzymolysis is to 48h, be cooled to 32 DEG C of inoculation yeast saccharomyces cerevisiaes and ferment, controlling fermentation condition is: leavening temperature is 32 DEG C, and inoculum size is 0.8 ‰ (v/v), and rotating speed is 150r/min;
3) after fermentation ends, in 60 ± 2 DEG C, under the vacuum tightness of-1.1Mpa, underpressure distillation 0.5h is carried out to fermented liquid, fully by complete for cut ethanolic extraction wherein;
4) by 30m
3concentration of substrate is 20%(w/v) vinasse import in M1, add water 28.5m successively
3, lignocellulose lytic enzyme 1.0m
3, material 50 tons, control temperature of reaction 50 DEG C, start to enter second batch of enzymolysis process, the same step 1) of specific implementation method, circulate 1 end.
As calculated, in loop cycle compared with intermittent type enzymatic saccharification zymotechnique, lignocellulose lytic enzyme consumption reduce 25%.
Embodiment 3
With 150m
3enzymatic vessel is that the present invention will be further described for example.
The method of enzyme recycle in a kind of lignocellulose enzymolysis process, the series connection of enzymatic vessel, fermentor tank and distillation tower is adopted to carry out, first feed intake by the concentration of substrate of setting and carry out enzymolysis, after enzymolysis terminates, cooling inoculation is fermented, again fermented liquid is imported vacuum still and carry out vacuum distillation, removing ethanol, live yeast inactivation, leaves free cellulase, finally imported in enzymatic hydrolysis system by vinasse and carry out reuse, concrete steps are as follows:
1) 150m after disappearing process to sky
3enzymatic vessel M1 adds 70.8 tons, tap water, open and stir, setting concentration of substrate is 20%(w/v), drop into M1 the wheat stalk 8 tons (1/6) that water content is 50% (w/w) by dog-house, to be mixed evenly after, add lignocellulose lytic enzyme 0.8m
3(2/3), control temperature of reaction 50 ± 1 DEG C, start to enter enzymolysis process, then continuous uniform charging, when inlet amount reaches 36 tons (3/4), add remaining lignocellulose lytic enzyme 0.4m
3, continue continuous uniform and feed intake, after starting to feed intake, in 16h, feed supplement to total amount reaches 48 tons; Every gram of over dry material adds lignocellulose lytic enzyme 40FPU;
2), when enzymolysis is to 48h, be cooled to 32 DEG C of inoculation yeast saccharomyces cerevisiaes and ferment, controlling fermentation condition is: leavening temperature is 32 DEG C, and inoculum size is 0.6 ‰ (v/v), and rotating speed is 120r/min;
2) after fermentation ends, in 65 ± 2 DEG C, under the vacuum tightness of-1.2Mpa, underpressure distillation 0.4h is carried out to fermented liquid, fully by complete for cut ethanolic extraction wherein;
3) by 30m
3concentration of substrate 20%(w/v) vinasse imports in M1, adds water 40.8m successively
3, lignocellulose lytic enzyme 0.8m
3, material 8 tons, control temperature of reaction 50 DEG C, start to enter second batch of enzymolysis process, the same step 1) of specific implementation method, circulate 1 end.
As calculated, in loop cycle compared with intermittent type enzymatic saccharification zymotechnique, lignocellulose lytic enzyme consumption reduce 20%.
Claims (6)
1. a method for enzyme recycle in lignocellulose enzymolysis process, it is characterized in that, concrete steps are as follows:
Step 1): start enzymatic vessel, according to the concentration of substrate of setting, criticizes continuous uniform be added in enzymatic vessel by material, moisture, and continuous uniform adds lignocellulose lytic enzyme in batches simultaneously, carries out enzymolysis;
Step 2): after enzymolysis terminates, enzymolysis solution is cooled to 30 ~ 32 DEG C, inoculation yeast saccharomyces cerevisiae, ferments;
Step 3): by step 2) fermentation gained fermented liquid imports in vacuum still and carries out vacuum distillation;
Step 4): imported in enzymatic vessel by step 3) vacuum distillation gained vinasse and carry out enzyme reuse, cycle index is 1 ~ 2 time.
2. the method for enzyme recycle in lignocellulose enzymolysis process as claimed in claim 1, it is characterized in that: the material described in step 1) is through pretreated lignocellulosic material, described pre-treatment comprises the steps: first to be adopted by lignocellulosic material mechanical means to carry out pulverization process, particle diameter is made to reach 0.14 ~ 32 millimeter, then be conveyed into steam explosion tank by rotary conveyor to process, then make the water content of material be 50 ~ 65% through plate-and-frame filter press press filtration.
3. the method for enzyme recycle in lignocellulose enzymolysis process as claimed in claim 1, it is characterized in that: the concentration of substrate set described in step 1) is 20 ~ 25%, described material and moisture are criticized continuous uniform be added in enzymatic vessel, the concrete grammar that simultaneously continuous uniform adds lignocellulose lytic enzyme is in batches: the weight adding material for the first time accounts for and always adds 1/8 ~ 1/6 of inventory, the amount adding lignocellulose lytic enzyme is for the first time always add enzyme amount 5/9 ~ 7/9, when enzymatic vessel completes always add inventory 1/2 ~ 3/4, once add remaining 2/9 ~ 4/9 enzyme amount, feed intake end, every gram of over dry material adds lignocellulose lytic enzyme 30 ~ 60FPU.
4. the method for enzyme recycle in lignocellulose enzymolysis process as claimed in claim 1, is characterized in that: step 2) described in technological condition for fermentation be: leavening temperature is 30 ~ 32 DEG C, and inoculum size is 0.5 ~ 0.8 ‰, and rotating speed is 60 ~ 150r/min.
5. the method for enzyme recycle in lignocellulose enzymolysis process as claimed in claim 1, it is characterized in that: the processing condition of vacuum distillation described in step 3) are: distillation temperature is 50 ~ 70 DEG C, vacuum tightness is-1.2 ~-0.90Mpa, distillation time is 0.4 ~ 1.0h, fully by complete for the ethanolic extraction in fermented liquid.
6. the method for enzyme recycle in lignocellulose enzymolysis process as claimed in claim 1, is characterized in that: the vinasse volume importing enzymatic vessel described in step 4) is 25% of the actual tinning volume of enzymatic vessel.
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101974570A (en) * | 2010-09-09 | 2011-02-16 | 天津大学 | Method for recycling cellulose complete components in fuel ethanol production |
| CN104611381A (en) * | 2013-11-05 | 2015-05-13 | 中国石油化工股份有限公司 | Method for producing ethanol by continuous enzymolysis and fermentation of lignocellulose |
| CN104774876A (en) * | 2015-04-10 | 2015-07-15 | 山东龙力生物科技股份有限公司 | Method for comprehensive utilization of lignocellulosic biomasses |
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Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101974570A (en) * | 2010-09-09 | 2011-02-16 | 天津大学 | Method for recycling cellulose complete components in fuel ethanol production |
| CN104611381A (en) * | 2013-11-05 | 2015-05-13 | 中国石油化工股份有限公司 | Method for producing ethanol by continuous enzymolysis and fermentation of lignocellulose |
| CN104774876A (en) * | 2015-04-10 | 2015-07-15 | 山东龙力生物科技股份有限公司 | Method for comprehensive utilization of lignocellulosic biomasses |
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