CN102851325A - Fermentation method for producing ethanol by using enzymatic saccharification of corn cob - Google Patents
Fermentation method for producing ethanol by using enzymatic saccharification of corn cob Download PDFInfo
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- CN102851325A CN102851325A CN2012103043584A CN201210304358A CN102851325A CN 102851325 A CN102851325 A CN 102851325A CN 2012103043584 A CN2012103043584 A CN 2012103043584A CN 201210304358 A CN201210304358 A CN 201210304358A CN 102851325 A CN102851325 A CN 102851325A
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- enzymolysis
- cellulase
- cellobiase
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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
- Y02E—REDUCTION OF GREENHOUSE GAS [GHG] EMISSIONS, RELATED TO ENERGY GENERATION, TRANSMISSION OR DISTRIBUTION
- Y02E50/00—Technologies for the production of fuel of non-fossil origin
- Y02E50/10—Biofuels, e.g. bio-diesel
Abstract
The invention provides a fermentation method for producing ethanol by using enzymatic saccharification of corn cob. Corn cob is used as a raw material, and subjected to enzymatic saccharification by cellulase. Main factors influencing enzymolysis efficiency in the saccharification process are studied, and feasibility of using hydrolysate to conduct ethanol fermentation is further explored.
Description
Technical field
The invention belongs to fermentation technical field, be specifically related to a kind of method of utilizing Enzymatic Hydrolysis of Corncob for Ethanol Production.
Background technology
Ethanol is the most promising liquid fuel from renewable resources, has cleaning, safety and the clean-burning characteristics of support gasoline.The production of present domestic alcohol fuel is mainly take foodstuff starch such as corn, wheats as raw material, and the present situation that China has a large population in a small area to have limited based on grain be the ethanol scale operation of raw material.China has abundant and cheap vegetable fibre resource, especially agriculture and forestry organic waste material, adopts biotechnology that fibrous material is degraded into fermentable glucose, further fermentative production of ethanol, to tap a new source of energy and the aspect such as environment protection significant.Utilize cellulase natural cellulose to be degraded in the process of glucose, must rely on the synergy of 3 kinds of components of cellulase to finish, namely cellulose macromolecule is at first at endo-type-beta-glucanase (C
xEnzyme) and circumscribed-type-beta-glucanase (C
1Enzyme) degraded fibroblast dimension disaccharides further generates glucose again under the effect under cellobiase (also claiming beta-glucosidase) effect.In the now widely used trichoderma reesei cellulase preparation, endo-type and circumscribed-type-activity of beta-glucanase are higher, but because the cellobiose enzyme activity is very low, in the cellulase hydrolysis process, easily cause the accumulation of cellobiose, thereby the katalysis of endo-type and circumscribed-type-beta-glucanase is formed strong feedback inhibition.
Corn cob is the large and concentrated agricultural fibre waste of a class amount, utilizes this fiber waste-material to replace the grain fermentative production of ethanol to have important economic benefit and social benefit.This patent utilizes cellulase that corn cob is carried out the enzyme process saccharification take corn cob as raw material, the Main Factors that affects enzymolysis efficiency in the saccharifying is studied, and has further probed into and utilized enzymolysis solution to carry out the feasibility of ethanol fermentation.
Summary of the invention
The invention provides a kind of method of utilizing Enzymatic Hydrolysis of Corncob for Ethanol Production.
1. a method of utilizing Enzymatic Hydrolysis of Corncob for Ethanol Production is characterized in that, the ethanol fermentation process of enzymolysis solution: the enzymolysis solution that the batch feeding enzymolysis obtains, add yeast extract paste to 3 g/L, (NH
4)
2HPO
4To 0.25 g/L, 50 mL liquid measures are packed in the 100 mL triangular flasks, access 0.5 mL cultivates the brewing yeast cell concentrated solution (cell precipitation of centrifugal rear collection is suspended in the sterilized water) of 24 h after the sterilization, and in 38 ℃ of anaerobically fermenting 30 h, timing sampling detects.
2. the process of the described batch feeding enzymolysis of step 1: in 2 L self-control enzymolysis reactor, carry out, the initial substrate mass concentration is 100 g/L, add a certain amount of cellulase and cellobiase, the Tween-80 addition is 1% (v/v), pH value 4.8,50 ℃ of Water Under solutions.12 and 24 at twice feed supplements of h after enzymolysis begins make substrate final quality concentration reach 200 g/L, add corresponding cellulase and cellobiase in the time of feed supplement, ask during enzymolysis to be controlled to be 60 h.
3. the process of described batch of formula enzymolysis of step 2: carry out in 2 L self-control enzymolysis reactor, the substrate mass concentration is 100 g/L, adds a certain amount of zymin, in pH value 4.8,50 ℃ of Water Under solutions.
4. the optimum amount of the described cellulase of step 3 is 25 FPIU.
5. the Best Times of described batch of formula enzymolysis of step 3 is 54 h.
6. the described cellobiase of adding of step 3 plays synergy.
7. the optimum amount of the described nonionic surface active agent Tween-80 of step 3 is 1.5%.
8. the described batch feeding enzymolysis process of step 2 has shortened the reaction times greatly, has improved production efficiency.
9. the alcohol concn behind ethanol fermentation 18 h of the described batch feeding enzymolysis solution of step 1 reaches 45.8 g/L.
Beneficial effect of the present invention is: the present invention is take corn cob as raw material, utilize cellulase that corn cob is carried out the enzyme process saccharification, the Main Factors that affects enzymolysis efficiency in the saccharifying is studied, and has further probed into and utilized enzymolysis solution to carry out the feasibility of ethanol fermentation.
Description of drawings
Enzymolysis time process under the effect of Fig. 1 cellulase.
Fig. 2 adds the coordinated enzymatic hydrolysis behind the cellobiase.
Fig. 3 batch feeding coordinated enzymatic hydrolysis time course.
The ethanol fermentation of Fig. 4 enzymolysis solution.
Embodiment
The following examples elaborate to the present invention, but to the present invention without limits.
The used bacterial strain of this patent be yeast saccharomyces cerevisiae (
Saccharomyces cerevisiae) CICC 1027, be preserved in Chinese industrial microbial strains preservation administrative center.
Embodiment 1
The implementation case illustrates different cellulase consumptions to the impact of batch formula enzymolysis, and the cost that effectively reduces cellulase is one of key link in the Mierocrystalline cellulose enzyme process Mashing process.The implementation case has been studied different cellulase consumptions to the impact of corn cob enzymolysis efficiency, the results are shown in Table l.
As can be seen from Table 1, different cellulase consumptions have considerable influence to enzymolysis efficiency, and when the cellulase consumption was increased to 25 FPIU from 10 FPIU, the enzymolysis yield brought up to 74.3% from 48.7%, further improve the cellulase consumption, the ascensional range of enzymolysis yield tends towards stability.Consider that from the economic angle that reduces as far as possible the cellulase cost Optimum of cellulase is 25 FPIU.
Table 1 cellulase consumption is on the impact of enzymolysis efficiency
| Cellulase consumption/FPIU | Reducing sugar (g/L) | Enzymolysis yield/% |
| 10 | 35.4 | 48.7 |
| 15 | 43.2 | 61.2 |
| 20 | 50.4 | 68.9 |
| 25 | 56.3 | 74.3 |
| 30 | 56.6 | 74.5 |
Embodiment 2
The implementation case illustrates time that different corn cobs criticize the formula enzymolysis to the impact of batch formula enzymolysis, and the result as shown in Figure 1.
As can be seen from Figure 1, along with the carrying out of enzyme digestion reaction, the reducing sugar mass concentration increases sharply, 54 h reducing sugar mass concentrations reach 49.6 g/L (wherein glucose is 26.7 g/L), the enzymolysis yield is 68.6%, continues to prolong enzymolysis time, and the enzymolysis yield does not significantly improve.The content of finding in addition cellobiose in the enzymolysis solution carries out and builds up with reaction, 54 h reach 11.8 g/L, this is owing to cellobiose enzyme activity in the trichoderma reesei cellulase system is not enough, the cellobiose that causes producing can not further be degraded to glucose and accumulate, thereby the katalysis of endo-type in the cellulase system and circumscribed-type-beta-glucanase is formed strong feedback inhibition.
The effect of the coordinated enzymatic hydrolysis of cellobiase is added in the implementation case explanation, and the feedback inhibition that causes for weakening the cellobiose accumulation improves the enzymolysis yield, adds cellobiase in enzymatic hydrolysis system.The result is shown in Figure 2.
By shown in Figure 2, ask process during coordinated enzymatic hydrolysis when enzyme dosage is 25 FPIU and 6.5 CBIU under the substrate mass concentration of 100 g/L.Can find out, add behind the cellobiase that cellobiose content all maintains lower level, not significantly accumulation in the whole enzymolysis process.Add an amount of cellobiase in the enzymatic hydrolysis system, the cellobiose that generates can be hydrolyzed into rapidly glucose, greatly weaken the feedback inhibition to endo-type in the cellulase system and circumscribed-type-beta-glucanase, thereby improved glucose quality concentration and enzymolysis yield.Behind enzymolysis 54 h, the reducing sugar mass concentration reaches 62.4 g/L (wherein glucose is 50.4 g/L), and the enzymolysis yield is increased to 84.3%.
Embodiment 4
The implementation case explanation nonionic surface active agent Tween-80 is on the impact of batch formula enzymolysis, carry out in the process of enzymolysis as substrate take the pretreated dragon spruce of steam explosion, add Tween-80 and can promote enzymolysis, it promotes that mechanism may be that nonionic surface active agent Tween-80 can be adsorbed on the lignin structure surface in the substrate, thereby reduced the absorption of cellulase on xylogen, cellulase just can fully act on Mierocrystalline cellulose like this, and then improves enzymolysis efficiency.Study under the coordinated enzymatic hydrolysis condition (enzyme dosage is 25 FPIU and 6.5 CBIU) Tween-80 addition in the test to the impact of enzymolysis efficiency, the results are shown in Table 2 after being hydrolyzed 54 h.
Add Tween-80 and can effectively promote enzyme digestion reaction, when the Tween-80 addition is 1.5% (volume fraction), the reducing sugar mass concentration is 68.2 g/L, the enzymolysis yield reaches 92.4%, further improve the addition of Tween-80, reducing sugar mass concentration and enzymolysis yield almost no longer change.
The Tween-80 addition is on the impact of enzymolysis efficiency under the table 2 coordinated enzymatic hydrolysis condition
| Tween-80 addition/% | Reducing sugar (g/L) | Enzymolysis yield/% |
| 0.5 | 60.3 | 84.2 |
| 1.0 | 62.3 | 86.3 |
| 1.5 | 68.2 | 92.4 |
| 2.0 | 69.1 | 92.7 |
Embodiment 5
The effect of the implementation case explanation batch feeding enzymolysis, in the enzyme process Mashing process of fibrous material, improve concentration of substrate and help to obtain higher sugared concentration, if but initial substrate concentration is too high, then feed liquid is too sticky, stir and the heat transmission resistance increase, be unfavorable for that enzyme digestion reaction carries out, can this difficult problem of fine solution by the batch feeding enzymolysis process.The result as shown in Figure 3.
As seen from Figure 3, adopt the batch feeding enzymolysis process, respectively 18 and feed supplement during 30 h, make substrate final quality concentration be increased to 200 g/L, the reducing sugar mass concentration reaches 116.7 g/L behind enzymolysis 66 h, wherein glucose quality concentration is 95.5 g/L, and the enzymolysis yield reaches 80.3%, a little less than the level of batch formula enzymolysis; For the substrate of degraded equivalent, the batch feeding enzymolysis process has shortened the reaction times greatly, has improved production efficiency.
The ethanol fermentation of the implementation case explanation batch feeding enzymolysis solution utilizes an Accharomyces cerevisiae under 38 ℃ the corn cob enzymolysis solution to be carried out ethanol fermentation, and experimental result is seen Fig. 4.
As seen from Figure 4, yeast saccharomyces cerevisiae can ferment rapidly, and glucose generates ethanol in the enzymolysis solution, after the inoculation in 18 h glucose utilized fully.The ethanol mass concentration reaches 45.8 g/L in the fermented liquid, and every gram glucose makes ethanol 0.48 g, reaches 94% (theoretical yield is 0.51 g ethanol (with every gram glucose meter)) of theoretical yield.
Claims (9)
1. a method of utilizing Enzymatic Hydrolysis of Corncob for Ethanol Production is characterized in that, the ethanol fermentation process of enzymolysis solution: the enzymolysis solution that the batch feeding enzymolysis obtains, add yeast extract paste to 3 g/L, (NH
4)
2HPO
4To 0.25 g/L, 50 mL liquid measures to be packed in the 100 mL triangular flasks, access 0.5 mL cultivates the brewing yeast cell concentrated solution of 24 h after the sterilization, and in 38 ℃ of anaerobically fermenting 30 h, timing sampling detects.
2. method according to claim 1, it is characterized in that, the process of batch feeding enzymolysis: in 2 L self-control enzymolysis reactor, carry out, the initial substrate mass concentration is 100 g/L, add a certain amount of cellulase and cellobiase, the Tween-80 addition is 1%, pH value 4.8,50 ℃ of Water Under solutions; 12 and 24 at twice feed supplements of h after enzymolysis begins make substrate final quality concentration reach 200 g/L, add corresponding cellulase and cellobiase in the time of feed supplement, ask during enzymolysis to be controlled to be 60 h.
3. method according to claim 2 is characterized in that, the process of batch formula enzymolysis: carry out in 2 L self-control enzymolysis reactor, the substrate mass concentration is 100 g/L, adds a certain amount of zymin, in pH value 4.8,50 ℃ of Water Under solutions.
4. method according to claim 3 is characterized in that, the optimum amount of cellulase is 25 FPIU.
5. method according to claim 3 is characterized in that, the Best Times of batch formula enzymolysis is 54 h.
6. method according to claim 3 is characterized in that, adds cellobiase and plays synergy.
7. method according to claim 3 is characterized in that, the optimum amount of nonionic surface active agent Tween-80 is 1.5%.
8. method according to claim 2 is characterized in that, the batch feeding enzymolysis process has shortened the reaction times greatly, has improved production efficiency.
9. method according to claim 1 is characterized in that, the alcohol concn behind ethanol fermentation 18 h of batch feeding enzymolysis solution reaches 45.8 g/L.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104846033A (en) * | 2015-05-14 | 2015-08-19 | 天津大学 | Method for preparing bioethanol by recovering and reusing resistant cellulase of coupling surface active agents of recombinant bacteria |
| CN110982848A (en) * | 2019-11-18 | 2020-04-10 | 福建农林大学 | Method for producing ethanol by semi-synchronous enzymolysis and fermentation of defatted peanut powder prehydrolysis liquid and pretreated solid matrix |
| CN113462504A (en) * | 2021-07-05 | 2021-10-01 | 西北农林科技大学 | Apple wine preparation process |
-
2012
- 2012-08-24 CN CN2012103043584A patent/CN102851325A/en active Pending
Non-Patent Citations (3)
| Title |
|---|
| 刘钟骏: "纤维素物质同时糖化法制备燃料乙醇的研究", 《中国优秀硕士论文全文数据库》 * |
| 赵晶, 等: "酶法糖化玉米芯发酵生产乙醇的研究", 《林产化学与工业》 * |
| 赵晶: "玉米芯酶法糖化发酵生产乙醇的研究", 《中国优秀硕士学位论文全文数据库》 * |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104846033A (en) * | 2015-05-14 | 2015-08-19 | 天津大学 | Method for preparing bioethanol by recovering and reusing resistant cellulase of coupling surface active agents of recombinant bacteria |
| CN110982848A (en) * | 2019-11-18 | 2020-04-10 | 福建农林大学 | Method for producing ethanol by semi-synchronous enzymolysis and fermentation of defatted peanut powder prehydrolysis liquid and pretreated solid matrix |
| CN113462504A (en) * | 2021-07-05 | 2021-10-01 | 西北农林科技大学 | Apple wine preparation process |
| CN113462504B (en) * | 2021-07-05 | 2022-09-13 | 西北农林科技大学 | Apple wine preparation process |
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Application publication date: 20130102 |