CN100497647C - Method for adsorbing and decomposing stalk cellulose using cellulase - Google Patents
Method for adsorbing and decomposing stalk cellulose using cellulase Download PDFInfo
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- CN100497647C CN100497647C CNB2006100112163A CN200610011216A CN100497647C CN 100497647 C CN100497647 C CN 100497647C CN B2006100112163 A CNB2006100112163 A CN B2006100112163A CN 200610011216 A CN200610011216 A CN 200610011216A CN 100497647 C CN100497647 C CN 100497647C
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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
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
- Y02P20/00—Technologies relating to chemical industry
- Y02P20/50—Improvements relating to the production of bulk chemicals
- Y02P20/59—Biological synthesis; Biological purification
Abstract
This invention relates to a method for preparing straw cellulose by adsorption-enzymolysis with cellulose. The method comprises: (1) steam-exploding straw, drying, mixing with cellulose solution, performing adsorption-desorption at -4-0 deg.C, and performing solid-liquid separation to obtain supernatant and cellulose-adsorbed steam-exploded straw; (2) performing enzymolysis on the straw at 40-50 deg.C for 48-72 h; (3) repeating adsorption-desorption and enzymolysis for 1-4 times. The cellulase consumption is 6 IU/g substrate, and the productivity of reducing sugar is 17.70-27.07 mg/IU cellulase. The method also has such advantages as short enzymolysis time, low energy consumption and recoverable cellulose.
Description
Technical field
The invention belongs to the cellulase hydrolysis field, particularly a kind of method of using the cellulase adsorbing stalk cellulose of enzymolysisby, the method that cellulase is efficiently utilized.
Background technology
The available plant cellulose aboundresources of China, only the agricultural crop straw annual output can reach 700,000,000 tons more than.The research of saccharification of cellulose has just begun before nearly 200 years.Early stage research mainly adopts the concentrated acid method to become glucose with diluted acid method hydrolysis and saccharification Mierocrystalline cellulose.Acid-hydrolysis method has fast this great advantage of speed of response; but owing to consuming a large amount of acid, conversion unit being existed corrodibility, energy consumption height, the sugar that acid hydrolysis produces is easy to degrade and can produces uronic acid or furfural etc. usually to follow-up fermentation or the totally unfavorable by product of the course of processing.Though there are the problems referred to above in the acid saccharification cellulosic material, because cognitive level and scientific research condition at that time, the acid system degraded was also once for the moment brilliant; Up to twentieth century four, the fifties, enzymic degradation is subjected to investigator's favor gradually and is entering coming into one's own further instantly of 21st century, becomes various countries correlative study person's focus.Enzymolysis process mild condition, efficiency of pcr product height, equipment requirements is low, energy consumption is low, but its maximum shortcoming is exactly that speed of reaction is slow.For improving speed of response, will increase the enzyme dosage in the conversion process, this faces the problem of enzyme application cost far above the commercial operations level in the time of certainly will making its commercial operations.
In enzymolysis process, the recycle enzyme is a kind of very effective enzyme of economizing on, and then reduces the mode of enzymolysis cost.At present, most researchers adopts form direct mode that repeats adsorptive enzyme on fresh substrate in batches, promptly behind batch enzymolysis, carries out system and separates, and supernatant can adsorb fresh substrate and continue enzymolysis; The substrate that contains residual cellulase adds in the new substrate and continues enzymolysis.So repeatedly, can carry out several cycles.Many substrates such as wheat straw, aspen carry out system and separate behind the enzymolysis certain hour, and the residue of adsorptive enzyme can obtain higher sugar amount mix the continuation enzymolysis with fresh substrate after.Though this mode can make full use of enzyme, seek out desirable hydrolysis efficiency, then will by prolong each reaction times of taking turns hydrolysis (take turns hydrolysis as the 1st and take 3 days, and the 5th take turns will be up to 16 days about).This just needs to consume extra energy, has increased cost of labor simultaneously, makes to save to such an extent that the cost of enzyme is offset by this extra " burden ".As seen, it is imperative to seek the mode of other recycle enzymes.Through a large amount of investigation and research, we have invented a kind of method of cellulase absorption enzymolysis steam puffed stalk.
Summary of the invention
The objective of the invention is at the low problem of cellulase utilization ratio in the present cellulase hydrolysis process, propose a kind of method of utilizing stalk pre-treatment and cellulase to combine to make the cellulase absorption enzymolysis steam puffed stalk that every international unit cellulase produces more reducing sugars.
The method of cellulase absorption enzymolysis steam puffed stalk provided by the invention is at first carried out the steam explosion pre-treatment to stalk; Get a certain amount of cellulose enzyme liquid then, mix at a certain temperature with steam puffed stalk, wait to reach the absorption-desorption balance after, carry out system and separate, the residue of adsorptive enzyme recovers solid-to-liquid ratio and carries out enzymolysis at a certain temperature; Supernatant continues to mix with fresh steam puffed stalk, reach the absorption-desorption balance after, carry out the system separation second time ... repeat above step, carry out 2-4 times successively.
The method of use cellulase adsorbing stalk cellulose of enzymolysisby provided by the invention, its step comprises:
(1) stalk steam explosion under the condition of vapor pressure 1.0～1.5Mpa of water content 10～35% is handled, again the steam stalk after steam-explosion is handled is carried out drying treatment afterwards;
(2) with the damping fluid mixed preparing cellulose enzyme liquid of cellulase with pH4.8;
(3) the exsiccant steam puffed stalk is mixed by the proportioning of the weight part proportioning 1:6～1:10 of solid-liquid with cellulose enzyme liquid, its cellulase consumption is the quick-fried wheat straw of 30～150IU/g vapour, carries out absorption-desorption under-4 ℃～0 ℃; After treating that absorption-desorption reaches balance, carry out solid-liquid separation, must go up the cleer and peaceful steam puffed stalk that is adsorbed with cellulase;
Described damping fluid is selected from acetic acid, sodium-acetate, citric acid or sodium citrate buffer solution;
Described cellulase is selected from the cellulase that Trichodermareesei, viride, aspergillus niger or koning trichoderma produce;
(4) steam puffed stalk that will be adsorbed with cellulase was 40 ℃～50 ℃ following enzymolysis 48～72 hours.
The method of use cellulase adsorbing stalk cellulose of enzymolysisby provided by the invention, described step also is included in and repeats following step 1-4 time after the step 3): add fresh steam puffed stalk in the separating obtained supernatant in step (3), the weight part proportioning that makes the mixed solution solid-liquid is 1:6～1:10, under-4 ℃～0 ℃, carry out absorption-desorption, after treating that absorption-desorption reaches balance, carry out solid-liquid separation, must go up the cleer and peaceful steam puffed stalk that is adsorbed with cellulase, with the steam puffed stalk that is adsorbed with cellulase 40 ℃～50 ℃ following enzymolysis 48～72 hours.
The time of described absorption-desorption is 2～4 hours.
Described stalk is selected from wheat straw, maize straw, straw, sorghum straw, reed, peanut seedling or Pericarppium arachidis hypogaeae.
After enzymolysis finishes, suction filtration, the gained solution reducing sugar amount in the DNS method measurement solution.When the cellulase consumption was the 50IU/g substrate, the reducing sugar amount (27.07mg) that the unit enzyme produces contrast (9.88mg) had improved nearly 2 times.
After enzymolysis finished, the cellulase in the system can reclaim: adopt the ammonium sulfate precipitation method in the supernatant, the enzyme that is adsorbed in solid residue can adopt Ca (OH)
2-Tween20 reclaims.
Method of the present invention has the following advantages:
(1) many batches of substrates of a collection of enzyme enzymolysis: after a collection of enzyme can adsorb 5 times with fresh substrate repeatedly and obtain good hydrolysis result, make the average consumption of cellulase be low to moderate the 6IU/g substrate most, be lower than the minimum amount standard 10IU/g substrate of the enzyme that can implement the industrialization estimation.
(2) shorten enzymolysis time than recycling technology behind the conventional batch enzymolysis, saved big energy;
(3) behind the enzymolysis, cellulase is recyclable.
Embodiment
The invention will be further described below by embodiment.
Embodiment 1
1, with water content be 10% wheat straw 15g, drop in the steam-explosion jar and under vapor pressure 1.0Mpa, handled 5 minutes that instant decompression discharges afterwards, obtain the wheat straw after steam explosion is handled, and with its 60 ℃ dry 48 hours down;
2, the quick-fried wheat straw of dried vapour and cellulase solution (being the cellulase solution of 4.8 hac buffer preparation with pH) the weight part proportioning 1:7 by solid-liquid is mixed, its cellulase consumption is the quick-fried wheat straw of 30IU/g vapour, carries out system after 2 hours in absorption under-1 ℃ and separates;
3, the substrate of adsorptive enzyme recovers solid-to-liquid ratio (the weight part proportioning 1:7 of solid-liquid) under 50 ℃, enzymolysis 72 hours.
The enzyme that present embodiment uses is the cellulase that Trichodermareesei produced, and the reducing sugar amount that its unit enzyme produces is 22.62mg.
Embodiment 2
1, with water content be 10% jowar stalk 15g, drop in the steam-explosion jar and under vapor pressure 1.4Mpa, handled 6 minutes that instant decompression discharges afterwards, obtain the jowar stalk after steam explosion is handled, and with its 60 ℃ dry 48 hours down;
2, the quick-fried jowar stalk of dried vapour and cellulase (being the enzyme liquid of 4.8 citric acid solution preparation with pH) the weight part proportioning 1:6 by solid-liquid is mixed, the cellulase consumption is the 50IU/g substrate, carries out the system separation at-3 ℃ after adsorbing 2 hours down;
3, add the quick-fried jowar stalk of fresh vapour in the separating obtained supernatant in step 2, the weight part proportioning that makes the mixed solution solid-liquid is 1:6, carries out absorption-desorption under-3 ℃ again;
After treating that absorption-desorption reaches balance, carry out solid-liquid separation again, must go up the cleer and peaceful steam puffed stalk that is adsorbed with cellulase; With the steam puffed stalk that is adsorbed with cellulase 50 ℃ of following enzymolysis 60 hours.
The enzyme that present embodiment uses is the cellulase that viride produced, and the reducing sugar amount that the unit enzyme produces is 27.07mg.
Embodiment 3
1, with water content be 35% straw 15g, drop in the steam-explosion jar and under vapor pressure 1.1Mpa, handled 7 minutes that instant decompression discharges afterwards, obtain the straw after steam explosion is handled, and with its 60 ℃ dry 48 hours down;
2, the quick-fried straw of dried vapour and cellulase (being the enzyme liquid of 4.8 sodium acetate buffer solution preparation with pH) are mixed by the weight part proportioning 1:8 by solid-liquid, the cellulase consumption is the 100IU/g substrate, carries out the system separation at 0 ℃ after adsorbing 3 hours down;
3, present embodiment also comprises and repeats following step 2 time: add the quick-fried jowar stalk of fresh vapour in the separating obtained supernatant in step 2, the weight part proportioning that makes the mixed solution solid-liquid is 1:8, carrying out absorption-desorption under 0 ℃ after 4 hours, carry out solid-liquid separation, must go up the cleer and peaceful steam puffed stalk that is adsorbed with cellulase; With the steam puffed stalk that is adsorbed with cellulase 40 ℃ of following enzymolysis 52 hours.
The enzyme that present embodiment uses is the cellulase that aspergillus niger produced, and the reducing sugar amount that the unit enzyme produces is 19.90mg.
Embodiment 4
1, with water content be 20% maize straw 15g, drop in the steam-explosion jar and under vapor pressure 1.5Mpa, handled 6 minutes that instant decompression discharges afterwards, obtain the maize straw after steam explosion is handled, and with its 60 ℃ dry 48 hours down;
2, the quick-fried maize straw of dried vapour and cellulase (being the enzyme liquid of 4.8 sodium citrate buffer preparation with pH) the weight part proportioning 1:10 by solid-liquid is mixed, the cellulase consumption is the 150IU/g substrate, carries out system after 4 hours in absorption under-4 ℃ and separates;
3, present embodiment also comprises and repeats following step 3 or 4 times: add the quick-fried jowar stalk of fresh vapour in the separating obtained supernatant in step 2, the weight part proportioning that makes the mixed solution solid-liquid is 1:10, carrying out absorption-desorption under-4 ℃ after 2 hours, carry out solid-liquid separation, must go up the cleer and peaceful steam puffed stalk that is adsorbed with cellulase; With the steam puffed stalk that is adsorbed with cellulase 45 ℃ of following enzymolysis 48 hours.
The enzyme that present embodiment uses is the cellulase that koning trichoderma produced, and the reducing sugar amount that the unit enzyme produces is 17.70mg.
Claims (4)
1, a kind of method of using the cellulase adsorbing stalk cellulose of enzymolysisby, its step comprises:
(1) stalk steam explosion under the condition of vapor pressure 1.0～1.5Mpa of water content 10～35% is handled, again the steam stalk after steam-explosion is handled is carried out drying treatment afterwards;
(2) with the damping fluid mixed preparing cellulose enzyme liquid of cellulase with pH4.8;
(3) the exsiccant steam puffed stalk is mixed by the proportioning of the weight part proportioning 1:6～1:10 of solid-liquid with cellulose enzyme liquid, its cellulase consumption is the quick-fried wheat straw of 30～150IU/g vapour, carries out absorption-desorption under-4 ℃～0 ℃; After treating that absorption-desorption reaches balance, carry out solid-liquid separation, must go up the cleer and peaceful steam puffed stalk that is adsorbed with cellulase;
Described damping fluid is selected from acetic acid, sodium-acetate, citric acid or sodium citrate buffer solution;
Described cellulase is selected from the cellulase that Trichodermareesei, viride, aspergillus niger or koning trichoderma produce;
(4) steam puffed stalk that will be adsorbed with cellulase was 40 ℃～50 ℃ following enzymolysis 48～72 hours.
2, method by the described use cellulase of claim 1 adsorbing stalk cellulose of enzymolysisby, it is characterized in that, described step also is included in and repeats following step 1-4 time after the step 3): add fresh steam puffed stalk in the separating obtained supernatant in step (3), the weight part proportioning that makes the mixed solution solid-liquid is 1:6～1:10, under-4 ℃～0 ℃, carry out absorption-desorption, after treating that absorption-desorption reaches balance, carry out solid-liquid separation, must go up the cleer and peaceful steam puffed stalk that is adsorbed with cellulase, with the steam puffed stalk that is adsorbed with cellulase 40 ℃～50 ℃ following enzymolysis 48～72 hours.
3, by the method for claim 1 or 2 described use cellulase adsorbing stalk cellulose of enzymolysisby, it is characterized in that the time of described absorption-desorption is 2～4 hours.
4, by the method for claim 1 or 2 described use cellulase adsorbing stalk cellulose of enzymolysisby, it is characterized in that described stalk is selected from wheat straw, maize straw, straw, sorghum straw, reed, peanut seedling or Pericarppium arachidis hypogaeae.
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| CNB2006100112163A CN100497647C (en) | 2006-01-18 | 2006-01-18 | Method for adsorbing and decomposing stalk cellulose using cellulase |
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| CNB2006100112163A CN100497647C (en) | 2006-01-18 | 2006-01-18 | Method for adsorbing and decomposing stalk cellulose using cellulase |
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| CN100497647C true CN100497647C (en) | 2009-06-10 |
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Families Citing this family (11)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| FR2925521B1 (en) * | 2007-12-20 | 2009-12-18 | Inst Francais Du Petrole | COMPLEMENTATION OF THE TRICHODERMA REESEI SECRETOME LIMITING MICROBIOLOGICAL CONTAMINATION IN INDUSTRIAL PROCESSES. |
| CN101418289B (en) * | 2008-11-28 | 2011-05-04 | 天津科技大学 | Trichoderma reesei liquid submerged fermentation cellulase and enzymatic beating process thereof |
| EP2505657A1 (en) * | 2009-11-27 | 2012-10-03 | Mitsui Chemicals, Inc. | Process for production of monosaccharide |
| CN102154243B (en) * | 2011-01-10 | 2012-09-26 | 南阳天冠生物发酵有限公司 | Method for producing liquid cellulose by mixed fermentation of microbe |
| CN102864192B (en) * | 2011-07-07 | 2014-09-17 | 中国石油化工股份有限公司 | Method for extracting sugar from wood fiber substances |
| CN103255185B (en) * | 2012-02-21 | 2015-04-22 | 华东理工大学 | Method for producing microbial oil through lignocellulose simultaneous saccharification and fermentation, and for recycling cellulase |
| CN102824897B (en) * | 2012-08-28 | 2015-09-16 | 华南理工大学 | A kind of polluted-water petroleum adsorbent and preparation method thereof |
| CN103243137A (en) * | 2013-05-08 | 2013-08-14 | 天津大学 | Method for improving straw enzymolysis saccharification efficiency by two-step crushing combined with steam explosion |
| CN105544237B (en) * | 2016-02-25 | 2018-05-11 | 东莞市联洲知识产权运营管理有限公司 | A kind of manufacture method of the combined insoles material of ventilation wear-resistance |
| CN110282953A (en) * | 2019-06-12 | 2019-09-27 | 魏炎梅 | A kind of preparation method of high temperature resistant anti-thermal shock electrothermal dark-red pottery |
| CN115786423B (en) * | 2023-02-03 | 2023-04-28 | 中国科学院过程工程研究所 | Method for reducing enzyme amount for enzymolysis of cellulose by steam explosion grading utilization of straw |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN1451755A (en) * | 2002-04-12 | 2003-10-29 | 中国农业大学 | Method for producing alcohol by solid fermentation of stalks |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN1451755A (en) * | 2002-04-12 | 2003-10-29 | 中国农业大学 | Method for producing alcohol by solid fermentation of stalks |
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