JP2007151433A - Method for pretreatment of lignocellulose and method for producing ethanol - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a method for pretreatment of lignocellulose for obtaining a syrup enabling efficient ethanol fermentation at low cost using a lignocellulose-containing feedstock such as waste construction materials, and to provide a method for producing ethanol. <P>SOLUTION: The method for the pretreatment of lignocellulose comprises the following process: A lignocellulose-containing feedstock is subjected to hydrolysis treatment in dilute sulfuric acid at 140-220°C for 3-20 min, the resultant hydrolyzate is subjected to solid/liquid separation into a primary syrup and a solid. The solid is incorporated with 0.5-20 mass% of quicklime on a dry basis heated at 90-200°C for 10-120 min to have calcification. Subsequently, cellulase is added to the resulting reaction product to be subjected to an enzymatic hydrolysis treatment to obtain a secondary syrup. <P>COPYRIGHT: (C)2007,JPO&INPIT

Description

  The present invention relates to a lignocellulose pretreatment method for obtaining a sugar solution that can be efficiently and efficiently fermented with ethanol using a lignocellulose-containing raw material such as waste building materials, and an ethanol production method for producing ethanol using the sugar solution.

Attempts to produce ethanol from natural lignocellulose-containing raw materials such as bagasse, rice straw, and wood chips, which are renewable resources, and to use them as energy and chemical raw materials are underway in Japan and overseas.
In woody biomass, which is one of lignocellulose-containing raw materials, hemicellulose is first hydrolyzed with acid or alkali to obtain hemicellulose-derived sugar. Saccharides constituting hemicellulose are mainly pentoses such as xylose and arabinose and hexoses such as glucose, galactose and mannose, and the ratio of these amounts varies depending on the type of woody biomass.

  After obtaining hemicellulose-derived sugar with acid or alkali, the residue is treated with acid or enzyme to obtain cellulose-derived sugar (glucose), but the method using dilute sulfuric acid has a low glucose yield of about 40%. In addition, sugar overdegradation substances such as formic acid, levulinic acid, and hydroxymethylfurfural (HMF) are likely to be produced, affecting fermentation.

One of the properties common to lignocellulose-containing raw materials is that there is a difference in the decomposition conditions of hemicellulose and cellulose. Hemicellulose is relatively easy to be decomposed by acid, alkali, and hot water treatment, and sugar can be obtained with a high recovery rate of 90% or more, whereas cellulose is decomposed under more severe conditions. Sugar overdegradation occurs at approximately the same rate, resulting in low sugar recovery.
Thus, hydrolysis by enzymes has been studied as a method to replace acids and alkalis, but it is necessary to select an appropriate method for pretreatment for making the enzyme work effectively depending on the raw materials.

As described above, since the sugar recovery rate from hemicellulose can be 90% or more by acid, alkali, and hot water treatment, the raw material is first subjected to primary hydrolysis that also serves as pretreatment for enzyme hydrolysis. Thus, it is desirable to recover the hemicellulose-derived sugar as much as possible.
A well-studied method as a pretreatment for enzyme hydrolysis is a method using acid, alkali, hydrothermal treatment, and explosion. All of the pretreatments are effective, but when saccharides are to be obtained as primary hydrolysis, the decomposition of saccharides is likely to proceed under alkaline conditions, so treatment using acid, hydrothermal treatment, or explosion is advantageous. is there. In addition, as a primary treatment for raw materials, caustic soda cooking and lime treatment under oxidizing conditions (for example, see Non-Patent Documents 1 to 4) have been reported, but the yield of hemicellulose-derived sugar is low.

On the other hand, sugar can be recovered by acid treatment, but depending on the raw material, it may be insufficient as a pretreatment for enzyme hydrolysis.
Among wood, the residue after primary treatment of hardwood with acid or blasting is relatively easily saccharified by enzymes, but conifers have a structure containing lignin that is stronger than hardwood, so the residue after primary treatment is not changed. Enzymatic hydrolysis rate is low.
Since waste building materials that are expected to be used effectively in Japan are mainly coniferous trees such as cedar, tsuga, and pine, establishment of a pretreatment method for coniferous trees is particularly important in Japan.
In addition, a method of increasing the enzyme hydrolysis rate by conducting a secondary treatment before subjecting the first treated residue to enzyme hydrolysis has been studied. For example, ozone, hydrogen peroxide, sodium chlorite and the like have been reported.
Vincent S. Chang et al., “Oxidative lime pretreatment of high-lignin biomass, Applied Biochemistry and Biotechnology”, Applied Biochemistry and Biotechnology, Vol. 94 (1), April, 1-28 (2001) Alfredo Martinez, Maria E. Rodriguez, Sean W. York, James F. Preston, Lonnie O. Inggram, “Effects of Ca (OH) 2 Treatments (“ Overliming ”) on the Composition and Toxicity of Bagasse Hemicellulose Hydrolysates”, Biotechnology and Bioengineering , Vol.69, No.5, September 5, 526-536 (2000) Ria Millati, Claes Niklasson, Mohammad J. Taherzadeh, “Effect of pH, time and temperature of overliming on detoxification of dilute-acid hydrolyzates for fermentation by Saccharomyces cerevisiae”, Process Biochemistry, 38, 515-522 (2002) Maria de FS Barbosa et al., “Efficient Fermentation of Pinus sp. Acid Hydrolysates by and Ethanologenic Strain of Escherichia coli”, Applied and Environmental Microbiology, Vol. 58, No. 4, 1382-1384 (1992)

However, the above-described conventional pretreatment method for performing the secondary treatment is not only expensive, but there is a concern that it may act to inhibit microorganisms in the subsequent fermentation, so a step of removing the drug before fermentation is provided. There is a need.
It is desired that the chemical used in the secondary treatment is inexpensive, does not inhibit fermentation in the next step, and can be recycled.

  The present invention has been made in view of the above circumstances, and uses a lignocellulose-containing raw material such as waste building material to obtain a sugar solution that can be efficiently and efficiently fermented with ethanol, and produces ethanol using the sugar solution. An object is to provide a method for producing ethanol.

  In order to achieve the above-mentioned object, the present invention hydrolyzes a lignocellulose-containing raw material in dilute sulfuric acid at 140 to 220 ° C. for 3 to 20 minutes, and then converts the hydrolyzate into a primary saccharide liquid and a solid substance. Separated, 0.5-20 mass% of quicklime is added to the separated solid matter in terms of dry matter, heated at 90-200 ° C. for 10-120 minutes for lime treatment, and then cellulase is added to the lime-treated reaction product. In addition, the present invention provides a method for pretreatment of lignocellulose characterized in that a secondary sugar solution is obtained by enzymatic hydrolysis.

  In the lignocellulose pretreatment method of the present invention, the lime-treated reactant is preferably subjected to solid-liquid separation, and the separated liquid containing quicklime is preferably used for neutralization of the primary sugar liquid containing dilute sulfuric acid.

  In the lignocellulose pretreatment method of the present invention, the reaction product is not subjected to solid-liquid separation after lime treatment, and the reaction product can be used for neutralization of the primary sugar solution containing dilute sulfuric acid.

  In the lignocellulose pretreatment method of the present invention, the lignocellulose-containing raw material is preferably a waste building material.

  The present invention also provides an ethanol production method characterized in that ethanol fermentation is carried out using the sugar solution obtained by the above-described lignocellulose pretreatment method according to the present invention to produce ethanol.

In the method for pretreatment of lignocellulose of the present invention, a lignocellulose-containing raw material is hydrolyzed in dilute sulfuric acid to obtain a primary sugar solution, quick lime is added to the separated solid, heated to lime, and this reaction product is treated with lime. Equipped with a step to obtain secondary sugar solution by enzymatic hydrolysis, and by using quick and recyclable quick lime for secondary pretreatment before enzymatic hydrolysis, it is possible to efficiently and efficiently ferment ethanol from lignocellulose-containing raw materials A sugar solution can be obtained.
Moreover, the ethanol production method of the present invention improves ethanol yield at low cost by performing ethanol fermentation using the sugar solution obtained by the above-described lignocellulose pretreatment method according to the present invention and producing ethanol. Can be achieved.

Hereinafter, embodiments of the present invention will be described with reference to the drawings. In the following description, mass% is simply described as%.
FIG. 1 is a flow diagram illustrating a first embodiment of the method of the present invention. In this embodiment, woody biomass such as waste building materials and wood chips is used as a lignocellulose-containing raw material, which is hydrolyzed in dilute sulfuric acid to obtain a primary sugar solution, and quick lime (CaO) is added to the separated solid. It is provided with a step of adding lime treatment and enzymatic hydrolysis treatment of the reaction product to obtain a secondary sugar solution. The lignocellulose-containing raw material is not limited to the woody biomass, and various lignocellulose-containing raw materials such as rice straw, rice husk and bagasse can be used.

  In this embodiment, first, woody biomass that is a lignocellulose-containing raw material is prepared. When using waste building materials as woody biomass, the waste building materials may be crushed to a size of 1 to 20 mm, preferably 5 to 10 mm, so that hydrolysis with dilute sulfuric acid in the next step proceeds smoothly. desirable. As the chip size is as small as possible, the efficiency of saccharification increases, but the energy required for crushing also increases, so the preferred chip size is about 1 to 20 mm, more preferably about 5 to 10 mm. Moreover, when using a waste building material, it is desirable to remove extraneous contaminants, such as metal, such as a nail, and plastic as much as possible. Waste building materials are mainly generated by the demolition of wooden houses, and the percentage of conifers such as cedar, pine and tsuga is high as the tree species used.

  Next, woody biomass such as waste building materials is heated in dilute sulfuric acid and hydrolyzed. The hydrolysis conditions are as follows: sulfuric acid concentration is preferably 0.1 to 5%, more preferably about 0.5 to 3%, temperature is 140 ° C to 220 ° C, preferably about 160 ° C to 210 ° C, and reaction time is It is 3 to 20 minutes, preferably about 5 to 10 minutes. These conditions are also conditions for maximally recovering sugar from hemicellulose. By this hydrolysis treatment, hemicellulose contained in the woody biomass is hydrolyzed to produce pentose and hexose derived from hemicellulose. This hydrolysis treatment is preferably performed using an autoclave or the like.

  Next, the hydrolysis reaction product is subjected to solid-liquid separation. As the solid-liquid separation method, filtration, centrifugation, or the like can be used, but it is preferable to use filtration with low energy consumption. The filtrate obtained by solid-liquid separation includes glucose, xylose, arabinose, galactose, and mannose (hereinafter abbreviated as G, X, A, Ga, and M, respectively) that are sugars derived from hemicellulose.

  Since this filtrate (primary sugar solution) is acidic, it is neutralized and then subjected to fermentation. However, the primary sugar solution hydrolyzed under the above conditions contains furfural, HMF, levulinic acid, Formic acid and the like are included. Since these overdegraded products inhibit fermentation by microorganisms, pretreatment is necessary. However, in the method of heating by adding slaked lime or quick lime, as described above, part of furfural and HMF is also removed. (For example, refer nonpatent literature 2,3,4).

  Nitrogen and phosphorus-containing nutrient sources and ethanol-fermenting microorganisms are added to the filtrate (primary sugar solution) neutralized with slaked lime or quicklime, and the microorganisms are cultured under conditions of appropriate temperature, pH, etc. for alcoholic fermentation. And converting sugar to ethanol to produce ethanol. As the ethanol fermentation microorganism, various conventionally known ethanol fermentation microorganisms such as Saccharomyces yeast can be used.

On the other hand, the residue obtained by solid-liquid separation is added with 0.5 to 20%, preferably 1 to 15%, more preferably 4 to 12% (both in terms of residue dry matter) of quick lime, and the temperature is 90 to 90%. The lime treatment is performed by heating at 200 ° C., preferably 100 to 180 ° C., more preferably 100 to 150 ° C., for 10 to 120 minutes, preferably 10 to 60 minutes, more preferably 20 to 40 minutes. This heating is preferably performed by blowing steam into the residue to which quicklime has been added.
In the lime treatment, the solid content concentration (slurry concentration) of the residue is preferably 5 to 30%, more preferably 10 to 20%.

Since slaked lime or quicklime is inexpensive as an alkali, it is generally used as an alkali for neutralizing a primary sugar solution obtained by hydrolysis using dilute sulfuric acid.
In addition, slaked lime or quick lime causes precipitation with sulfuric acid, resulting in poorly soluble gypsum, which not only has the effect of reducing fermentation inhibition by dissolved salts, but also adds slaked lime or quick lime slightly in excess, at 60 ° C for 30 minutes Holding for about several hours has the effect of decomposing and removing most of the furans produced by excessive decomposition of sugars such as furfural and HMF contained in the hydrolyzate.
Slaked lime or quicklime used for neutralization tends to form salts and precipitates, and has little toxicity to cells.
The quick lime after being used in the lime treatment is recycled to the neutralization step so that the quick lime other than the amount consumed in the reaction can be effectively used without waste.

  Next, the reaction product subjected to the quicklime treatment is solid-liquid separated into a lime treatment solution and a residue. As the solid-liquid separation method, filtration, centrifugation, or the like can be used, but it is preferable to use filtration with low energy consumption. The filtrate obtained by solid-liquid separation is used for neutralization of the primary sugar solution. The residue has a high cellulose content and is subjected to enzymatic hydrolysis using cellulase.

  Enzymatic hydrolysis is carried out by adding cellulase to a liquid in which the residue is suspended and reacting, for example, with stirring at a pH of about 4-6, a temperature of about 35-60 ° C., for about 10-100 hours. The secondary sugar solution obtained by this enzymatic hydrolysis contains glucose (G), which is a sugar derived from cellulose.

  This secondary sugar solution is mixed with the above-described neutralized primary sugar solution or separately, and a nutrient source containing nitrogen and phosphorus and an ethanol-fermenting microorganism such as yeast are added, and an appropriate temperature, pH, etc. are added. Microorganisms are cultured under conditions to perform alcoholic fermentation, sugar is converted to ethanol, and ethanol is produced. As the ethanol fermentation microorganism, various conventionally known ethanol fermentation microorganisms such as Saccharomyces yeast can be used.

In the pretreatment method of lignocellulose of the present embodiment, a lignocellulose-containing raw material is hydrolyzed in dilute sulfuric acid to obtain a primary sugar solution, and quick lime is added to the separated solid, heated to lime, and this reaction product. It is possible to efficiently ferment ethanol from lignocellulose-containing raw materials by using low-cost and recyclable quick lime for secondary pretreatment before enzymatic hydrolysis. Can be obtained.
In addition, the ethanol production method of the present embodiment achieves an improvement in ethanol yield at low cost by performing ethanol fermentation using the sugar solution obtained by the lignocellulose pretreatment method and producing ethanol. Is possible.

  FIG. 2 is a flow diagram illustrating a second embodiment of the method of the present invention. This embodiment uses woody biomass, hydrolyzes it in dilute sulfuric acid to obtain a primary sugar solution, and adds the quick lime to the separated solid to perform lime treatment with the above-described first embodiment. Although it is performed in the same manner, the lime-treated reactant is added to the primary sugar solution as a lime-treated slurry without solid-liquid separation, and the mixture is neutralized and then subjected to enzyme hydrolysis using cellulase to obtain a secondary sugar solution. It is characterized by obtaining.

  In this embodiment, since all the lime processing slurries obtained by lime treatment are used for neutralization of the primary sugar solution, among the quick lime added for lime treatment, quick lime excluding the portion consumed in the lime treatment is used as the primary sugar. It can be used for neutralization of the liquid. The alkali which is insufficient for neutralization is added separately. It may be quick lime, slaked lime, or a mixture of quick lime and slaked lime. After neutralizing the mixture of the primary sugar solution and the lime processing slurry, cellulase is added, and the reaction is performed, for example, by reacting for about 4 to 6 hours at a temperature of about 35 to 60 ° C. for about 10 to 100 hours with stirring. The secondary sugar solution obtained by this enzyme hydrolysis includes glucose, xylose, arabinose, galactose, mannose (G, X, A, Ga, M), which are sugars derived from hemicellulose contained in the primary sugar solution, and enzyme hydrolysis. The glucose (G) produced | generated by decomposition | disassembly is contained.

  After the enzymatic hydrolysis is completed, ethanol fermentation microorganisms such as yeast and other nutrient sources containing nitrogen and phosphorus are added to the resulting secondary sugar solution, and the microorganisms are cultured under conditions of appropriate temperature, pH, etc. To convert the sugar to ethanol to produce ethanol. As the ethanol fermentation microorganism, various conventionally known ethanol fermentation microorganisms such as Saccharomyces yeast can be used.

  According to this embodiment, the same effect as the first embodiment described above can be obtained, and further, the lime treatment slurry after lime treatment is mixed with the primary sugar solution and then the enzyme hydrolysis is performed, whereby the processing step is performed. It can be simplified. Moreover, since the lime processing slurry is mixed with the primary sugar solution and used for neutralization of the primary sugar solution, there is no waste of quick lime added for lime treatment.

  In this embodiment, after neutralizing the mixture of the primary sugar solution and the lime treatment slurry, it is also possible to add both cellulase and ethanol-fermenting microorganisms to this mixture and simultaneously perform enzyme hydrolysis and alcohol fermentation. It is.

The lignocellulose-containing raw material was pretreated under the following experimental conditions.
[Common conditions] <Raw materials>
Waste building materials for boards (mainly cedar, tsuga and pine).
<Primary hydrolysis conditions>
Sulfuric acid concentration 1%, 170 ° C., 10 minutes.
<Filtration>
After primary hydrolysis, solid-liquid separation was performed by suction filtration, and the residue was washed well with water to obtain a raw material for secondary treatment.
<Secondary processing>
Lime treatment was performed under the conditions described in each Example and each Comparative Example described below.
<Enzymatic hydrolysis>
Cellulase: 0.4 g of SPEZYME CE, manufactured by Genencor International Japan Limited, was added to 1 g of solids in the residue separated after lime treatment.
Reactor: To the 200 mL Erlenmeyer flask, 2.5 g of the residue was added as a solid, and a buffer solution and ion-exchanged water were added together with the adhering moisture of the residue to 50 mL.
pH: 0.2N sodium acetate was added as a buffer to adjust to pH 4.5.
Temperature: 45 ° C.
Agitation: Shake at 100 rpm.

[Experiment 1: Comparison of lime treatment temperature]
The temperature of the lime treatment was set as in the following Examples 1 to 4 and Comparative Example 1, pretreatment was performed, and further enzymatic hydrolysis was performed, and the glucose yield of the resulting secondary sugar solution was examined and compared. In addition, quick lime addition amount is a ratio with respect to a residue dry matter.
Example 1: The lime treatment conditions were a quick lime addition amount of 4%, a temperature of 100 ° C., and a treatment time of 30 minutes.
Example 2: The lime treatment conditions were a quick lime addition amount of 4%, a temperature of 121 ° C., and a treatment time of 30 minutes.
Example 3: The lime treatment conditions were a quick lime addition amount of 4%, a temperature of 150 ° C., and a treatment time of 30 minutes.
Example 4: The lime treatment conditions were a quick lime addition amount of 4%, a temperature of 180 ° C., and a treatment time of 30 minutes.
Comparative Example 1: The lime treatment was not performed, and the residue was directly subjected to enzymatic hydrolysis.
Table 1 shows the amount of glucose recovered. In Table 1, “glucose yield (g / 100 g—pretreated product)” refers to the glucose yield relative to the solid content (dry weight) of the residue after lime treatment, and “glucose yield (g / 100 g—pre-treatment). “Solid content before treatment” means the glucose yield relative to the solid content (dry weight) before lime treatment.

From these results, the mass loss due to the secondary pretreatment varied depending on the temperature of the lime treatment.
The glucose yield (based on the solid content subjected to the primary treatment) taking into account the mass loss of the solid content is 33 g due to the lime treatment for 4 minutes at 121 ° C. for 30 minutes with respect to 20 g in the case of only the primary treatment (Comparative Example 1). Increased by 65%. In addition, an increase in yield was observed even at a temperature of 180 ° C.

[Experiment 2: Comparison of lime addition amount]
The amount of quicklime added to the lime treatment is set as in the following Examples 1 to 3 and Comparative Examples 1 and 2, pretreatment is performed, and further, enzymatic hydrolysis is performed, and the glucose yield of the resulting secondary sugar solution is examined. Compared.
Example 1: The lime treatment conditions were a quick lime addition amount of 4%, a temperature of 121 ° C., and a treatment time of 30 minutes.
Example 2: The lime treatment conditions were 8% quick lime addition, 121 ° C. temperature, and 30 minutes treatment time.
Example 3: The lime treatment conditions were a quick lime addition amount of 12%, a temperature of 121 ° C., and a treatment time of 30 minutes.
Comparative Example 1: The lime treatment conditions were 0% quick lime addition, a temperature of 121 ° C., and a treatment time of 30 minutes.
Comparative Example 2: The lime treatment was not performed, and the residue was directly subjected to enzymatic hydrolysis.
Table 2 shows the amount of glucose recovered.

  Although the mass loss due to the secondary pretreatment changed depending on the amount of lime added, the amount of change was smaller than that in the case of temperature. The glucose yield (based on the solid material subjected to the primary treatment) taking into account the mass loss of the solid content is 33 g due to the lime treatment for 4 minutes at 121 ° C. for 30 minutes with respect to 20 g in the case of only the primary treatment (Comparative Example 2). Increased by 65%. The yield did not increase even when the amount of quicklime was increased to 8% or 12%. In addition, when only heating was performed without adding quicklime, the total yield was lower than when no treatment was performed.

[Experiment 3: Comparison between lime treatment and caustic soda treatment]
A secondary sugar solution obtained by performing pretreatment of each of cases where quick lime is added during lime treatment (Examples 1 and 2) and when caustic soda is added (Comparative Examples 1 and 2), and further enzymatic hydrolysis is performed. The glucose yields were examined and compared.
Example 1: The lime treatment conditions were a quick lime addition amount of 4%, a temperature of 121 ° C., and a treatment time of 30 minutes.
Example 2: The lime treatment conditions were 8% quick lime addition, 121 ° C. temperature, and 30 minutes treatment time.
Comparative Example 1: Caustic soda was used in place of quick lime, and the treatment conditions were 8% caustic soda addition, temperature 121 ° C., and treatment time 30 minutes.
Comparative Example 2: Caustic soda was used in place of quick lime, and the treatment conditions were a caustic soda addition amount of 12%, a temperature of 121 ° C., and a treatment time of 30 minutes.
Comparative Example 3: The lime treatment was not performed, and the residue was directly subjected to enzymatic hydrolysis.
Table 3 shows the amount of glucose recovered.

  When the secondary pretreatment was performed with caustic soda, the glucose yield was lower than in the case of only the primary treatment (Comparative Example 3).

[Experiment 4: Recycling lime after reaction to neutralization process]
In Experiment 3, the secondary treatment product treated with lime under the conditions of Example 1 (calcified lime addition amount 4%, 121 ° C., 30 minutes) was used for neutralization of the primary sugar solution. This neutralization was performed using slaked lime until reaching a pH of 10 at which the effect of removing the fermentation inhibitor was obtained (Example 1).
Table 4 shows the amount of slaked lime used per 1000 g of raw material. In addition, the primary sugar liquid obtained by primary hydrolysis with respect to 1000g of raw materials was 3200g, and the primary hydrolysis residue was 750g as a solid substance.

  By using the processed product of the secondary pretreatment with quick lime for neutralization of the primary sugar solution, the slaked lime newly used for neutralization of the primary sugar solution is 12 g than when the lime treatment was not performed (Comparative Example 1). Reduced.

It is a flowchart which shows 1st Embodiment of the method of this invention. It is a flowchart which shows 2nd Embodiment of the method of this invention.

Claims (5)

  The lignocellulose-containing raw material is hydrolyzed in dilute sulfuric acid at 140 to 220 ° C. for 3 to 20 minutes, and then the hydrolyzate is solid-liquid separated into a primary sugar solution and a solid, and the separated solid is dried. 0.5-20 mass% of quicklime is added in conversion, heated at 90-200 ° C. for 10-120 minutes for lime treatment, and then cellulase is added to the lime-treated reaction product to carry out enzyme hydrolysis treatment, secondary sugar A method for pretreating lignocellulose, comprising obtaining a liquid.   2. The lignocellulose pretreatment method according to claim 1, wherein the lime-treated reactant is subjected to solid-liquid separation, and the liquid containing the separated quick lime is used for neutralization of the primary sugar solution containing dilute sulfuric acid.   The method for pretreatment of lignocellulose according to claim 1, wherein the reaction product is used for neutralization of the primary sugar solution containing dilute sulfuric acid without solid-liquid separation after the lime treatment.   The pretreatment method for lignocellulose according to any one of claims 1 to 3, wherein the lignocellulose-containing raw material is a waste building material. An ethanol production method, wherein ethanol is fermented using the sugar solution obtained by the lignocellulose pretreatment method according to any one of claims 1 to 4 to produce ethanol.

Images