WO2010098408A1

WO2010098408A1 - Method for producing sugar, method for producing ethanol, method for producing lactic acid, and method for producing starting material for enzymatic saccharification used therein

Info

Publication number: WO2010098408A1
Application number: PCT/JP2010/053007
Authority: WO
Inventors: 正浩鮫島; 圭日子五十嵐; 昌久和田; 毅上村
Original assignee: 国立大学法人東京大学; 新日本石油株式会社
Priority date: 2009-02-27
Filing date: 2010-02-25
Publication date: 2010-09-02
Also published as: CN102333882A; JP5469881B2; JP2010200624A; BRPI1013168A2

Abstract

Disclosed are a method for producing a sugar, a method for producing ethanol and a method for producing lactic acid, wherein enzymatic saccharification can be performed efficiently, thereby respectively improving the production efficiency of sugar, the production efficiency of ethanol and the production efficiency of lactic acid. Also disclosed is a method for producing a useful starting material for enzymatic saccharification which is used in the above-described production methods. Specifically disclosed is a method for producing a sugar, which is characterized by comprising: (a) a step of obtaining a modified biomass starting material by processing a biomass starting material containing a type I cellulose with a processing agent containing ammonia and/or an organic amine; (b) a step of obtaining a starting material for enzymatic saccharification by pulverizing the modified biomass starting material; and (c) a step of obtaining a sugar by enzymatically saccharifying the starting material for enzymatic saccharification.

Description

Method for producing sugar, method for producing ethanol, method for producing lactic acid, and method for producing raw material for enzyme saccharification used in these

The present invention relates to a sugar production method, an ethanol production method, and a lactic acid production method using a biomass raw material, and an enzymatic saccharification used in the sugar production method, the ethanol production method, and the lactic acid production method. The present invention relates to a method for producing raw materials.

In recent years, as part of measures against global warming, attempts have been widely made to produce ethanol from raw materials containing cellulose such as woody biomass and herbaceous biomass and use them as various fuels and chemical raw materials. Production of ethanol from biomass raw material can be performed, for example, by decomposing the collected biomass raw material into sugar in the saccharification step and then converting it to ethanol using a microorganism such as yeast in the fermentation step.

On the other hand, the use of biodegradable polymers is increasing from the viewpoint of reducing environmental impact, and lactic acid is used as one of the raw materials. This lactic acid can also be obtained by saccharifying the biomass raw material and further fermenting it.

The saccharification has been conventionally performed using concentrated sulfuric acid, but from the viewpoint of reducing the environmental load, it is desired to reduce the amount of sulfuric acid used. Therefore, in recent years, saccharification of biomass raw materials using enzymes has been widely studied as an alternative to saccharification with concentrated sulfuric acid. Enzymatic saccharification is a desirable means from the viewpoint of the influence on the environment. However, for this enzymatic saccharification, it is necessary to pre-treat biomass raw material in advance for the purpose of making the enzyme act easily. Various methods are known as a pretreatment method of this biomass raw material, and among them, steaming treatment with dilute sulfuric acid, pressurized hot water, etc. is common (for example, see Patent Documents 1 to 4 below). However, as described above, in order to obtain the desired degree of enzymatic saccharification efficiency in the case where the use of sulfuric acid is not preferable, and when these pretreatments are performed on the biomass raw material and the obtained processed product is subjected to enzymatic saccharification, There are problems that it is necessary to perform the pretreatment in multiple stages, and that the temperature must be increased to 200 ° C. or higher.

In addition, it is known that chemical and biochemical reactivity can be improved by finely pulverizing biomass raw materials by physical means, but if sufficient enzymatic saccharification efficiency is obtained by pulverization alone, The process requires a lot of energy and may lose economic rationality.

Furthermore, it is known that the chemical and biochemical reactivity of a biomass raw material is improved by pretreatment with ammonia or organic amine (see, for example, Patent Document 5 below). However, even with the pretreated biomass, the enzymatic saccharification efficiency is still not sufficient. Therefore, at present, the development of an enzyme saccharification technique that can further increase the enzyme saccharification efficiency and the development of a pretreatment technique for biomass raw materials suitable for the enzyme saccharification are still desired.

JP 2006-075007 A JP 2004-121055 A JP-T-2002-541355 JP 2002-159954 A European Patent Publication No. 77287

The object of the present invention is to solve the problems in the prior art and achieve the following objects. That is, the present invention can efficiently carry out enzymatic saccharification, and therefore can improve sugar production efficiency, ethanol production efficiency, and lactic acid production efficiency. It is an object of the present invention to provide a method and a method for producing lactic acid, and a useful method for producing a raw material for enzymatic saccharification used in the method for producing sugar, the method for producing ethanol, and the method for producing lactic acid.

In order to solve the above-mentioned problems, the present inventors have made extensive studies and obtained the following knowledge. That is, by subjecting a biomass raw material containing cellulose type I, which is a natural type cellulose, with a treatment agent containing ammonia and / or an organic amine, further pulverizing and then subjecting it to enzymatic saccharification It is a knowledge that enzymatic saccharification can be efficiently carried out, and therefore sugar production efficiency, ethanol production efficiency, and lactic acid production efficiency can be significantly improved.

In addition, the present inventors previously used enzymatic saccharification efficiently by using cellulose having a crystal density lower than that of natural cellulose (type _I cellulose) as an object of enzymatic saccharification. Patent application filed that it can be carried out, and that a biomass raw material containing cellulose type I can be treated with ammonia, particularly supercritical ammonia, to efficiently obtain cellulose for enzymatic saccharification containing cellulose III type _I. (See JP 2008-161125 A).

It has been conventionally known that the enzymatic saccharification efficiency of a biomass raw material can be remarkably improved by treating the biomass raw material with a treatment agent containing ammonia and / or an organic amine, and then further pulverizing and subjecting it to enzymatic saccharification. This is a new finding by the present inventors.

The present invention is based on the above findings by the present inventors, and means for solving the above problems are as follows. That is,
<1> (a) A step of obtaining a modified biomass raw material by treating a biomass raw material containing cellulose type I with a treatment agent containing ammonia and / or an organic amine;
(B) a step of obtaining a raw material for enzyme saccharification by grinding the modified biomass raw material; and
(C) a step of saccharifying the enzyme saccharification raw material to obtain a saccharide;
It is a manufacturing method of the sugar characterized by including.
<2> The sugar production method according to <1>, wherein the treating agent used in the step (a) is ammonia.
<3> The sugar production method according to <1> or <2>, wherein the biomass material containing cellulose type I is woody biomass.
<4> Any one of the above <1> to <3>, wherein the average particle size represented by the median size of the raw material for enzymatic saccharification obtained in step (b) is 5 to 80 μm. The method for producing sugar as described in 1. above.
<5> A method for producing ethanol, wherein the sugar obtained by the method for producing sugar according to any one of <1> to <4> is fermented to obtain ethanol.
<6> A method for producing lactic acid, wherein the saccharide obtained by the method for producing saccharide according to any one of <1> to <4> is fermented to obtain lactic acid.
<7> (a) A process for obtaining a modified biomass raw material by treating a biomass raw material containing cellulose type I with a treatment agent containing ammonia and / or an organic amine; and
(B) a step of obtaining a raw material for enzymatic saccharification by pulverizing the modified biomass raw material,
It is a manufacturing method of the raw material for enzyme saccharification characterized by including this.

According to the present invention, the above-mentioned objects can be achieved, various problems in the prior art can be solved, and enzymatic saccharification can be performed efficiently. Therefore, sugar production efficiency, ethanol production efficiency, and lactic acid production efficiency Enzymatic saccharification used in sugar production method, ethanol production method, lactic acid production method, and sugar production method, ethanol production method, and lactic acid production method The raw material manufacturing method can be provided.

FIG. 1 is an X-ray diffraction pattern of coarsely crushed eucalyptus. FIG. 2 is an X-ray diffraction pattern of a sample obtained by pulverizing coarsely crushed eucalyptus. FIG. 3 is an X-ray diffraction pattern of a sample obtained by treating crushed eucalyptus with ammonia. FIG. 4 is an X-ray diffraction pattern of a sample obtained by crushing coarsely crushed eucalyptus and further treating with ammonia. FIG. 5 is an X-ray diffraction pattern of a sample obtained by subjecting coarsely crushed eucalyptus to ammonia treatment and further pulverization treatment. FIG. 6 is an X-ray diffraction diagram of coarsely ground Ezo nokinu willow. FIG. 7 is an X-ray diffraction pattern of a sample obtained by pulverizing coarsely crushed Ezo nonu willow. FIG. 8 is an X-ray diffraction pattern of a sample obtained by subjecting coarsely ground Ezo nokinu willow to ammonia treatment. FIG. 9 is an X-ray diffraction pattern of a sample obtained by pulverizing coarsely crushed Ezo nonu willow and further ammonia treatment. FIG. 10 is an X-ray diffraction pattern of a sample obtained by subjecting coarsely milled Ezo noki willow to ammonia treatment and further grinding treatment. FIG. 11 is an X-ray diffraction pattern of coarsely ground cedar. FIG. 12 is an X-ray diffraction pattern of a sample obtained by pulverizing coarsely cedar cedar. FIG. 13 is an X-ray diffraction pattern of a sample obtained by treating coarsely ground cedar with ammonia. FIG. 14 is an X-ray diffraction pattern of a sample obtained by pulverizing coarsely cedar and further treating with ammonia. FIG. 15 is an X-ray diffraction pattern of a sample obtained by subjecting coarsely ground cedar to ammonia treatment and further grinding treatment.

(Method for producing sugar)
The sugar production method of the present invention comprises (a) a step of obtaining a modified biomass raw material by treating a biomass raw material containing cellulose type I with a treating agent containing ammonia and / or an organic amine; A step of obtaining a raw material for enzyme saccharification by crushing the quality biomass raw material, and (c) a step of saccharifying the raw material for enzyme saccharification to obtain a sugar, and further further steps as necessary Including.

<Process (a)>
In the step (a), a biomass raw material containing cellulose type I is treated with a treatment agent containing ammonia and / or an organic amine to obtain a modified biomass raw material.

-Biomass raw materials containing cellulose type I-
There is no restriction | limiting in particular as a biomass raw material containing the said cellulose I type, According to the objective, it can select suitably. For example, “waste biomass” obtained as a residue from production activities such as agriculture and forestry, “resource crop biomass” obtained by intentional cultivation for the purpose of obtaining energy, etc. can be used. . Examples of the “waste-based biomass” include waste building materials, thinned wood, rice straw, wheat straw, rice husk, bagasse and the like, and examples of the “resource crop-based biomass” include sugarcane and corn. Eucalyptus, poplar, acacia, willow, cedar, switchgrass, napiergrass, Eliansus, Miscanthus, Susuki, etc. It is done. The biomass raw material containing cellulose type I is also classified into “woody biomass” using wood as a raw material, “herbaceous biomass” using grass as a raw material, and the like. In the present invention, both woody biomass and herbaceous biomass can be used, but woody biomass is preferably used from the viewpoint that the effects of the present invention can be obtained more remarkably. Moreover, as a biomass raw material containing the said cellulose I type, the cellulose I type itself obtained by refine | purifying etc. from various biomass as mentioned above may be sufficient. The biomass raw material containing the said cellulose I type may be used individually by 1 type, and may use 2 or more types together. Cellulose I type, which is a natural type cellulose, is classified into cellulose I _α type and cellulose I _β type, and any of these may be used as the cellulose I type contained in the biomass raw material. Both of these may be used.

In the treatment with the treatment agent containing ammonia and / or organic amine, as the biomass raw material containing cellulose type I, the collected material may be used as it is, but it is reduced to a certain size by cutting, pulverizing, or the like. It is desirable to use it afterwards. The size of the biomass raw material is not particularly limited and may be appropriately selected according to the purpose. For example, the mesh opening size is preferably 5 mm or less, more preferably 3 mm or less, and further preferably 2 mm or less. preferable. When the mesh size of the mesh exceeds 5 mm, the treatment with the treatment agent may be insufficient. On the other hand, when the size is within the further preferable range, it is advantageous in that the treatment time can be shortened and the amount of the treatment agent to be used can be reduced. Hereinafter, the process of cutting and pulverizing the collected biomass material may be referred to as “coarse pulverization”.

By performing the coarse pulverization in advance, the treatment with the treatment agent containing ammonia and / or organic amine proceeds efficiently, and when pulverizing the modified biomass raw material, finer fine powder with excellent enzymatic saccharification efficiency The raw material for enzyme saccharification can be obtained efficiently. There is no restriction | limiting in particular as a grinder used for the said rough crushing, According to the objective, it can select suitably, For example, a Willet mill, a cutter mill, a hammer mill, a pin mill etc. can be used.

-Treatment with ammonia and / or organic amine-
When the biomass raw material containing the cellulose type I is treated with ammonia as a treating agent, the method is not particularly limited and can be appropriately selected depending on the purpose. For example, it can be performed by introducing a biomass raw material containing cellulose type I and ammonia into a pressure vessel, setting the inside of the pressure vessel to a desired pressure and temperature, and processing for a desired time. The ammonia may be in the liquid phase, in the gas phase, or in a supercritical state. By the treatment with ammonia, at least a part of cellulose type I in the biomass raw material is transformed into cellulose III type _I having higher enzymatic saccharification efficiency. From the viewpoint of improving the transformation efficiency, liquid ammonia or supercritical state Ammonia is suitable. However, in consideration of the target saccharification rate, energy consumption, etc., it is possible to select treatments under conditions suitable for each. The conditions for the treatment with ammonia are not limited, but generally preferred conditions are a temperature of −35 to 140 ° C. and a pressure of 0 to 12.5 MPa.

When the biomass containing cellulose type I is treated with an organic amine as a treating agent, the organic amine to be used is not particularly limited and can be appropriately selected according to the purpose. For example, ethylenedian, monomethylamine, monoethylamine and the like are preferably used, and ethylenediamine is preferred. When the biomass containing cellulose type I is treated with these organic amines, the treatment temperature and pressure can be the same as those in the treatment with ammonia.

In the present invention, it is preferable to use ammonia as the treatment agent from the viewpoint of the efficiency of transformation from cellulose type I to cellulose III type _I , ease of removal of the treatment agent after treatment, and the like.

The time for the treatment with the treatment agent containing ammonia and / or organic amine is not particularly limited, and may be a desired level depending on the amount of biomass raw material containing the cellulose type I used, the treatment pressure, the treatment temperature, and the like. Can be appropriately selected within the range in which the transformation from cellulose I type to cellulose III _I type proceeds, preferably 10 minutes to 10 hours, more preferably 30 minutes to 8 hours, particularly 30 minutes to 5 hours. preferable. If the treatment time is less than 10 minutes, the desired degree of transformation from cellulose type I to cellulose III type _I may not proceed, and if it exceeds 10 hours, more than cellulose I type to cellulose III type _I The transformation to may not proceed and may be inefficient as a whole. On the other hand, when the treatment time is within the further preferable range, it is advantageous in that the transformation from cellulose I type to cellulose III _I type can proceed efficiently.

The amount of the ammonia and / or organic amine used in the treatment with the treatment agent containing ammonia and / or organic amine is not particularly limited and may be appropriately selected depending on the purpose. 10 mg to 300 g is preferable, 100 mg to 150 g is more preferable, and 1 g to 50 g is particularly preferable with respect to 1 g of biomass raw material containing type I. When the amount of ammonia and / or organic amine used is less than 10 mg relative to 1 g of biomass raw material containing cellulose type I, the treatment may be insufficient. May be worse. On the other hand, when the amount used is within the particularly preferable range, it is advantageous in that the treatment time can be shortened and the amount of the treatment agent to be used can be reduced.

In addition, at the time of the treatment with the treatment agent containing ammonia and / or organic amine, as long as at least the treatment agent contains ammonia and / or organic amine, other compounds may be used in combination. Examples of the compound include carbon dioxide, nitrogen, ethylene, methane, ethane, propane, butane, pentane, hexane, toluene, benzene, phenol, dioxane, xylene, acetone, chloroform, carbon tetrachloride, ethanol, methanol, propanol, and butanol. Etc. In addition, it is preferable not to use water as said other compound. With the water, cellulose III type _I obtained was, in some cases back to cellulose I type.

-Reformed biomass feedstock-
A modified biomass raw material is obtained by the treatment with the treatment agent containing ammonia and / or organic amine. By the treatment, at least a part of cellulose type I contained in the biomass raw material can be transformed into cellulose III type _{I having} a lower crystal density. Cellulose III type _I is advantageous in that the enzyme is likely to act because of its low crystal density. Furthermore, by the said process, most hemicellulose contained in the said biomass raw material is decomposed | disassembled to an oligosaccharide grade, and becomes soluble in water. Therefore, by processing the biomass material, the cellulose type I and hemicellulose contained in the biomass material are changed to a state in which the enzyme is more likely to act, such as cellulose III type _I and oligosaccharide derived from hemicellulose, respectively. Therefore, the enzymatic saccharification efficiency can be improved. Incidentally, by the process, at least part of the cellulose I type is converted into cellulose III _I type, for example, can be confirmed by X-ray diffraction, FT-IR, solid-state NMR and the like. The “modified biomass raw material” in the present application means a material obtained by treating a biomass raw material containing cellulose type I with a treating agent containing ammonia and / or an organic amine, but the cellulose type I contained in the biomass raw material. It is preferable that at least a part is transformed into cellulose III type _I.

In the modified biomass raw material, cellulose may have other compounds between its molecular structures. For example, as described above, the modified biomass raw material can be obtained by treating a biomass raw material containing cellulose type I, which is a natural type cellulose, with a treatment agent containing ammonia and / or an organic amine. It may be in a state of a complex of cellulose and ammonia and / or an organic amine (hereinafter sometimes referred to as “complex such as cellulose / ammonia”) produced in the process. However, the complex of cellulose / ammonia is difficult to adjust pH during enzymatic saccharification, and has the property of returning to cellulose type I by the action of water. It is preferable to use a modified biomass raw material from which ammonia and / or organic amine has been removed from the composite of cellulose and ammonia.

Therefore, after the treatment with ammonia and / or organic amine, it is preferable to provide a removal step of removing ammonia and / or organic amine from the composite of cellulose and ammonia. The method for removing the ammonia and / or organic amine is not particularly limited and may be appropriately selected depending on the purpose. For example, the cellulose / ammonia obtained after the treatment with the ammonia and / or organic amine, etc. Examples include a method of washing the modified biomass raw material containing the composite with methanol, ethanol, acetone or the like, a method of drying under reduced pressure, and a method of drying at a temperature equal to or higher than the boiling point of the treating agent. In the case of using ammonia as the treating agent, the removal method is not using an organic solvent, and is excellent in safety, at a temperature equal to or higher than the boiling point of ammonia (for example, room temperature to 50 ° C.) at normal pressure or A method of drying under reduced pressure is preferred.

In addition, it is preferable that the said modified biomass raw material contains cellulose III type _I, and there is no restriction | limiting in the ratio, However, The more is preferable at the point from which the outstanding enzyme saccharification efficiency is obtained. In addition to the cellulose III type _I , the modified biomass material includes, for example, cellulose type I (cellulose I _α type, cellulose I _β type) and other components such as hemicellulose and lignin. May be. However, from the viewpoint of improving the efficiency of enzymatic saccharification, it is preferable that lignin is not contained or the content thereof is small.

<Step (b)>
In the step (b), the raw material for enzyme saccharification is obtained by pulverizing the modified biomass raw material obtained in the step (a).

-Grinding-
In the step (b), the modified biomass raw material obtained in the step (a) is pulverized.

A method for pulverizing the modified biomass raw material is not particularly limited and may be appropriately selected depending on the intended purpose. For example, a pulverizer such as a flat mill, a planetary ball mill, a vibration ball mill, a bead mill, a jet mill or the like may be used. Can be used. Among these, a flat mill is preferable as the pulverizer because a fine powdery raw material for enzymatic saccharification excellent in enzymatic saccharification efficiency can be obtained with relatively low energy. By the pulverization, the modified biomass raw material can be used as the raw material for enzymatic saccharification of the present invention. By pulverizing the modified biomass raw material, it is possible to further improve the enzymatic saccharification efficiency.

The conditions for the pulverization are not particularly limited, and can be appropriately selected depending on the type of pulverizer, the type of biomass raw material, the average particle size of the pulverized product to be obtained, and the like. In addition, when using a flat mill as a pulverizer, since the pulverized biomass raw material is discharged from the mortar, when further pulverizing, the discharged pulverized material will be collected and supplied to the mortar again. Further, this may be repeated a plurality of times. In this case, the number of times of pulverization is not particularly limited, and can be appropriately selected according to the type of pulverizer used, the time per pulverization, the energy applied, and the like. This is advantageous in that a finer powdery raw material for enzyme saccharification with excellent enzyme saccharification efficiency can be obtained. The pulverization is advantageous in that a finer powdery raw material for enzyme saccharification with excellent enzymatic saccharification efficiency can be obtained each time the pulverization is repeated. 4 times or less is preferable from the standpoint of inefficiency as a whole.

-Raw material for enzymatic saccharification-
The “enzymatic saccharification raw material” according to the method of the present invention refers to a material obtained by pulverizing the modified biomass raw material. By pulverizing the modified biomass raw material, it is possible to further improve the enzymatic saccharification efficiency. The particle size of the enzyme saccharification raw material particles obtained by the pulverization is not particularly limited, and the preferred size varies depending on the type of biomass raw material to be used. The average particle size is preferably 5 to 80 μm, more preferably 5 to 50 μm, and even more preferably 5 to 30 μm. If the average particle size of the raw material for enzyme saccharification is to be less than 5 μm, it will take a great deal of energy and time for pulverization and lose economic rationality, while if it exceeds 80 μm, the enzymatic saccharification efficiency May not be sufficiently improved. On the other hand, if the average particle size of the enzyme saccharification raw material is within the further preferable range, it is advantageous in terms of the balance of energy and time required for pulverization and enzyme saccharification efficiency. In addition, as an average particle diameter of the raw material for enzyme saccharification in the present application, a median diameter obtained by measurement by a laser diffraction confusion method is adopted. Here, the median diameter refers to a particle diameter in which the cumulative volume of particles having a particle diameter equal to or larger than the particle diameter is equal to particles having a particle diameter equal to or smaller than the particle diameter.

The raw material for enzymatic saccharification obtained by pulverization of the modified biomass may be used, for example, as it is in the enzymatic saccharification of step (c) described later, or after appropriately passing through other steps, step (c) described later. It may be subjected to enzymatic saccharification. There is no restriction | limiting in particular as said other process, According to the objective, it can select suitably, For example, the raw material for enzyme saccharification of the fine powder obtained by the said grinding | pulverization is suitable for the enzyme saccharification mentioned later. The pH adjustment process etc. which adjust to appropriate pH are mentioned.

<Step (c)>
In the step (c), the enzyme saccharification raw material obtained in the step (b) is enzymatically saccharified to obtain a saccharide.

-Enzymatic saccharification-
There is no restriction | limiting in particular as the method of performing the said enzyme saccharification, For example, it can carry out on the conditions as shown below.
There is no restriction | limiting in particular as an enzyme used for the said enzyme saccharification, According to the objective, it can select suitably, For example, a cellulase, cellobiase ((beta) -glucosidase), etc. are mentioned.

The amount of the enzyme used in the enzymatic saccharification is not particularly limited and may be appropriately selected according to the purpose. For example, 0.001 to 100 mg is preferable with respect to 1 g of the enzyme saccharification raw material. 0.01 to 10 mg is more preferable, and 0.1 to 1 mg is still more preferable. If the amount of the enzyme used is less than 0.001 mg with respect to 1 g of the enzyme saccharification raw material, enzyme saccharification may be insufficient, and if it exceeds 100 mg, saccharification inhibition may occur. On the other hand, when the amount of the enzyme used is within the more preferable range, it is advantageous in that the amount of sugar obtained is larger than the amount of enzyme added.

The temperature for the enzymatic saccharification is not particularly limited and may be appropriately selected depending on the intended purpose. For example, it is preferably 10 to 70 ° C, more preferably 20 to 60 ° C, and further preferably 30 to 50 ° C. preferable. If the temperature is less than 10 ° C, enzyme saccharification may not proceed sufficiently, and if it exceeds 70 ° C, the enzyme may be deactivated. On the other hand, when the temperature is within the further preferable range, it is advantageous in that a large amount of sugar is obtained with respect to the amount of enzyme added.

The pH at the time of enzymatic saccharification is not particularly limited and may be appropriately selected depending on the intended purpose. For example, it is preferably 3.0 to 8.0, more preferably 3.5 to 7.0, 4.0 to 6.0 is more preferable. If the pH is less than 3.0 or more than 8.0, the enzyme may be deactivated. On the other hand, when the pH is within the more preferable range, it is advantageous in that a large amount of sugar is obtained with respect to the amount of enzyme added.

-sugar-
By the enzyme saccharification, for example, a sugar solution containing glucose which is a saccharide derived from cellulose III type _I contained in the raw material for enzyme saccharification obtained in the step (b) can be obtained. In addition, the sugar solution obtained by the enzymatic saccharification may contain, for example, glucose derived from the cellulose type I or sugar derived from hemicellulose. Examples of sugars derived from hemicellulose include pentose sugars such as xylose and arabinose, and hexose sugars such as glucose, galactose, and mannose.
The sugar solution may be used, for example, as it is in the ethanol production method or lactic acid production method of the present invention, which will be described later, or after the other steps described below, the ethanol production method or lactic acid of the present invention, which will be described later. You may use for the manufacturing method of.
There is no restriction | limiting in particular as said other process, According to the objective, it can select suitably, For example, the pH adjustment process etc. which adjust the said sugar liquid to pH suitable for each fermentation process mentioned later, etc. Is mentioned.

(Ethanol production method)
The method for producing ethanol of the present invention includes a step (fermentation step) of fermenting the sugar obtained by the above-described method for producing sugar of the present invention to obtain ethanol, and further includes other steps as necessary. .

<Fermentation process (alcohol fermentation process)>
In the method for producing ethanol, the method for fermenting the sugar is not particularly limited and may be appropriately selected depending on the intended purpose. For example, an alcohol-fermenting microorganism such as yeast is added to the solution containing the sugar. In particular, a method of performing alcoholic fermentation is particularly preferable. There is no restriction | limiting in particular as said yeast, According to the objective, it can select suitably, For example, Saccharomyces genus yeast etc. are mentioned. The yeast may be natural yeast or genetically modified yeast. Specific examples of the ethanol-fermenting microorganism include Saccharomyces cerevisiae, Kluyveromyces fragilis, Kluyveromyces lactis (K. lactis), Kluyveromyces marxianus Recombinant yeasts such as K. marxianus, Pichia stipitis, P. pastoris, Pachisolen tannofilus, Candida glabrata, or Candida glabrata Zymomonas mobilis, Symobacter pa Main (Zymobacter palmae), Clostridium thermocellum (Clostridium thermocellum), Clostridium Rujungudari (C.ljungdahlii) like bacterial or can be used these genetic recombinant.

In the fermentation, the amount of yeast used, fermentation temperature, pH, fermentation time, etc. are not particularly limited, and are appropriately selected according to, for example, the amount of sugar to be used for alcohol fermentation, the type of yeast to be used, etc. can do.

<Other processes>
There is no restriction | limiting in particular as said other process, According to the objective, it can select suitably, For example, the process etc. which isolate | separate and refine | purify the ethanol obtained by the said fermentation process are mentioned. The method for separation and purification is not particularly limited and may be appropriately selected depending on the intended purpose. Examples thereof include distillation.

The ethanol obtained by the ethanol production method can be suitably used as, for example, fuel ethanol, industrial ethanol, and the like. Since the ethanol can be obtained from a biomass raw material, it can be reproduced as long as the biomass raw material can be produced, and the plant absorbs carbon dioxide in the atmosphere at the time of cultivation. Even if carbon dioxide is generated by combustion, it does not increase the carbon dioxide concentration in the atmosphere. Therefore, it can be said that ethanol is a desirable energy source for preventing global warming. In recent years, such ethanol is particularly expected to be mixed with gasoline and used as an environmentally friendly automobile fuel.

An alcohol other than ethanol is produced by fermenting the sugar obtained by the sugar production method of the present invention with a microorganism that produces the desired alcohol, instead of the yeast that produces ethanol. You can also For example, butanol can be produced by fermentation using acetone / butanol bacteria.

(Production method of lactic acid)
The method for producing lactic acid of the present invention includes a step (fermentation step) of fermenting the saccharide obtained by the above-described method for producing saccharide of the present invention to obtain lactic acid, and further includes other steps as necessary. .

<Fermentation process (lactic acid fermentation process)>
In the lactic acid production method, the method for fermenting the sugar is not particularly limited and may be appropriately selected depending on the intended purpose. For example, a lactic acid-fermenting microorganism such as lactic acid bacteria is added to the solution containing the sugar. In particular, a method of performing lactic acid fermentation is particularly preferable. The lactic acid bacterium is not particularly limited and may be appropriately selected depending on the intended purpose. Streptococcus thermophilus) and Lactobacillus bulgaricus. The lactic acid bacterium may be a natural lactic acid bacterium or a genetically modified lactic acid bacterium.

In the fermentation, the amount of lactic acid bacteria used, fermentation temperature, pH, fermentation time, etc. are not particularly limited, and are appropriately selected according to, for example, the amount of sugar to be used for lactic acid fermentation, the type of lactic acid bacteria used, etc. can do.

<Other processes>
There is no restriction | limiting in particular as said other process, According to the objective, it can select suitably, For example, the process etc. which isolate | separate and refine | purify the lactic acid obtained by the said fermentation process are mentioned. The separation / purification method is not particularly limited and may be appropriately selected depending on the intended purpose.

The lactic acid obtained by the lactic acid production method can be suitably used for producing polylactic acid by chemical polymerization, for example. Currently, it is desirable to be able to produce lactic acid, which is often produced from starch such as corn, from biomass raw materials containing cellulose that cannot be used for food. According to the method for producing lactic acid, It is possible to efficiently produce polylactic acid from a biomass raw material containing

The saccharide obtained by the method for producing saccharides of the present invention is fermented by using microorganisms that produce the desired organic acid in place of the lactic acid bacteria, so that organic acids other than lactic acid, such as citric acid and succinic acid, can be obtained. Acid, malic acid, oxalic acid and the like can also be produced.

(Method for producing raw materials for enzymatic saccharification)
The method for producing a raw material for enzyme saccharification according to the present invention comprises (a) a step of obtaining a modified biomass raw material by treating a biomass raw material containing cellulose type I with a treating agent containing ammonia and / or an organic amine; and (B) A step of obtaining the enzyme saccharification raw material by pulverizing the modified biomass raw material, and further including other steps as necessary.

In the method for producing the enzyme saccharification raw material, the step (a) and the step (b) are as described in the item of the method for producing a sugar of the present invention.

It is generally known that chemical and biological reactivity is improved by pulverizing a biomass raw material containing cellulose. In addition, the present inventors have already disclosed that by treating a biomass raw material containing cellulose type I with ammonia or the like, cellulose type _I is transformed into cellulose type III and type _I to improve enzymatic saccharification efficiency.

In the sugar production method of the present invention, when enzymatic saccharification is performed after the biomass raw material is only treated with a treatment agent containing ammonia and / or organic amine, and when the biomass raw material is only pulverized, enzymatic saccharification is performed. Furthermore, it is possible to carry out enzymatic saccharification with much superior efficiency compared to the case where the biomass raw material is first pulverized and then treated with a treatment agent containing ammonia and / or organic amine to carry out enzymatic saccharification. is there. Although this mechanism of action is not clear, the present inventors presume as follows.

In the sugar production method according to the present invention, the enzyme saccharification raw material of the present invention obtained in the step (b) has its X-ray diffraction pattern in comparison with the modified biomass raw material obtained in the step (a). Has been shown to change significantly. That is, there is a possibility that some change has occurred in the cellulose structure by pulverizing the modified biomass raw material. On the other hand, when the biomass raw material is first pulverized by the same operation as in step (b) according to the method of the present invention and then treated with a treatment agent containing ammonia and / or organic amine, the pulverization is not performed. There is no significant difference in the X-ray diffraction pattern in comparison with the case of treatment with a treatment agent containing ammonia and / or organic amine. From this, the structural change reflected in the change in the X-ray diffraction pattern generated when the modified biomass raw material is pulverized is related to the fact that the raw material for enzyme saccharification according to the present invention has high enzyme saccharification efficiency. It is possible that

Examples of the present invention will be described below, but the present invention is not limited to these examples.

Example 1
-Biomass raw material-
Eucalyptus was used as a biomass raw material containing cellulose type I.

-Coarse grinding-
The prepared eucalyptus was coarsely pulverized using a Willet mill with a target average particle size of 200 μm.

-Ammonia treatment-
The coarsely crushed eucalyptus was subjected to treatment with supercritical ammonia by the following operation.
A 4 g sample of coarsely crushed eucalyptus dried in an oven at 60 ° C. for 24 hours is placed in a portable reactor TVS-N2 type (produced by TAIATSU, hereinafter referred to as “container”) having an internal volume of 120 ml, and is sealed in a cooling device. Then, while cooling the container to −13 ° C., ammonia was allowed to flow in at a pressure of 0.5 MPa for 30 minutes. Thereafter, a PC-V type heater (manufactured by TAIATSU) was attached to the container, and a heating / pressurizing treatment was performed at 140 ° C. for 1 hour. At this time, it was confirmed that the pressure in the container was 11 MPa or more at which ammonia became a supercritical state. After the treatment, the inside of the container was brought to atmospheric pressure to remove ammonia, the temperature was cooled to room temperature, and a solid sample in the container was collected. The sample was allowed to evaporate well overnight without sealing.

-Crushing process-
Using a ceramic flat mortar and home mortar-type tea powder device “Marugoto Green Tea EU6820” (trade name, manufactured by Panasonic Corporation), 2 g of the above ammonia-treated sample was set for 10 minutes each time with the setting of “fine” on the scale. Milled repeatedly 3 times.

-X-ray diffraction analysis-
100 mg of the crushed sample was pressure-molded at a pressure of 200 kg / cm ² and subjected to X-ray diffraction analysis. X-ray diffraction was performed by a diffractometry method using a tube-type X-ray generator RINT2200 (trade name, manufactured by Rigaku Corporation). Step scan method with a voltage of 38 kV, a current of 50 mA, a monochromator CuKα ray (wavelength 0.15418 nm), operating range 2θ = 5 to 30 °, step width 0.1 °, integration time 20 seconds Measured with Moreover, the X-ray diffraction analysis was performed by the same operation also about the sample after coarse grinding | pulverization and the sample after ammonia treatment. The diffraction pattern of the sample after coarse pulverization is shown in FIG. 1, the diffraction pattern of the sample after ammonia treatment is shown in FIG. 3, and the diffraction pattern of the sample after ammonia treatment and pulverization treatment is shown in FIG.

-Enzymatic saccharification reaction-
About the sample which performed the ammonia process and the grinding | pulverization process by the above, enzymatic saccharification reaction was performed by the following operation.
Take 10 mg of precisely weighed sample in a microtube with an internal volume of 1.5 ml, sample concentration 1% (wt / vol), Celluclast@1.5L and Novozyme @ 188 (both trade names, manufactured by Novozyme) as enzymes An enzyme saccharification reaction solution was prepared so that the enzyme concentration was 0.01% (wt / vol), the enzyme concentration was 0.02% (wt / vol) in total, and the pH was 4.5 (acetate buffer). This was subjected to an enzymatic saccharification reaction by shaking for 24 hours using a rotary shaker (15 rpm) in a constant temperature room at 37 ° C. The glucose concentration in the supernatant obtained by centrifugation after the reaction was measured using Glucose CII Test Wako (trade name, manufactured by Wako Pure Chemical Industries, Ltd.), the glucose yield was calculated, and the results are shown in Table 1.
The glucose yield is defined by the following formula.

Glucose yield (%) = [amount of glucose in enzyme saccharification reaction solution / (amount of enzyme saccharification raw material × total glycation rate / 100)] × 100

Total glucosylation rate (%): (amount of glucose obtained when a biomass raw material is completely hydrolyzed separately / amount of biomass raw material) × 100 (corresponding to a theoretical yield based on biomass raw material)

The total glycation rate of the used eucalyptus was 43.3%.

(Comparative Example 1-1: Enzymatic saccharification of untreated sample)
The roughly crushed eucalyptus used in Example 1 was subjected to an enzymatic saccharification reaction as it was in the same manner as the enzymatic saccharification reaction in Example 1 without performing ammonia treatment and pulverization treatment. The results are shown in Table 1.

(Comparative Example 1-2: No ammonia treatment, enzymatic saccharification of pulverized sample)
-Crushing process-
The coarsely crushed eucalyptus used in Example 1 was pulverized by the same operation as the operation of pulverizing the sample after ammonia treatment in Example 1 without being ammonia-treated.

-X-ray diffraction analysis-
X-ray diffraction analysis of the crushed sample was performed in the same manner as in Example 1. The diffraction pattern is shown in FIG.

-Enzymatic saccharification reaction-
The smashed sample was subjected to an enzymatic saccharification reaction by the same operation as the enzymatic saccharification reaction operation in Example 1. The glucose yield was calculated in the same manner as in Example 1, and the results are shown in Table 1.

(Comparative Example 1-3: Enzymatic saccharification of ammonia-treated sample)
About the sample which processed the coarsely ground eucalyptus obtained in Example 1 with ammonia, enzyme saccharification reaction was performed by operation similar to the enzyme saccharification reaction operation in Example 1 without pulverizing after that. The glucose yield was calculated in the same manner as in Example 1, and the results are shown in Table 1.

(Comparative Example 1-4: Enzymatic saccharification of a sample treated with ammonia after grinding)
-Crushing process-
The coarsely crushed eucalyptus used in Example 1 was pulverized by the same operation as the operation of pulverizing the sample after ammonia treatment in Example 1.

The pulverized sample was treated with ammonia in the same manner as the ammonia treatment in Example 1.

-X-ray diffraction analysis-
X-ray diffraction analysis of the sample treated with ammonia after the pulverization was performed in the same manner as in Example 1. The diffraction pattern is shown in FIG.

-Enzymatic saccharification reaction-
The sample subjected to the ammonia treatment after the pulverization treatment was subjected to the enzyme saccharification reaction by the same operation as the enzyme saccharification reaction operation in Example 1. The glucose yield was calculated in the same manner as in Example 1, and the results are shown in Table 1.

(Example 2)
Rough pulverization, ammonia treatment, pulverization, and enzymatic saccharification reaction were carried out in this order in the same manner as in Example 1 except that Ezo no Kinu willow was used instead of eucalyptus used in Example 1. The results are shown in Table 1. Further, the X-ray diffraction analysis was performed on the sample at each stage by the same operation as in Example 1. The X-ray diffraction pattern of each sample was similar to the corresponding eucalyptus sample in Example 1. The diffraction pattern after coarse pulverization is shown in FIG. 6, the diffraction pattern of the sample after ammonia treatment is shown in FIG. 8, and the diffraction pattern of the sample after ammonia treatment and pulverization treatment is shown in FIG.
In addition, the total glucoseation rate of the used Ezo nokinu willow was 45.1%, and the average particle diameter of the coarsely pulverized Ezo noki willow was 252 μm.

(Comparative Examples 2-1 to 2-4)
In each of Comparative Examples 1-1 to 1-4, Comparative Example 1 corresponding to Ex. 1 was used except that the coarsely ground Ezo nocilia used in Example 2 was used instead of the coarsely ground Eucalyptus used as the starting material. Pretreatment was performed in the same manner as in -1 to 1-4, and enzymatic saccharification reaction was performed in the same manner as in Example 2 for each sample obtained. For each comparative example, the glucose yield was calculated and the results are shown in Table 1. Further, for Comparative Examples 2-2 to 2-4, X-ray diffraction analysis was performed on each pretreated sample. The X-ray diffraction pattern of each sample was similar to each eucalyptus sample in Comparative Examples 1-2 to 1-4 corresponding to each sample. FIG. 7 shows the diffraction pattern after the pulverization treatment, and FIG. 9 shows the diffraction pattern of the sample treated with ammonia after the pulverization treatment.

(Example 3)
Rough pulverization, ammonia treatment, pulverization, and enzymatic saccharification reaction were performed in this order in the same manner as in Example 1 except that cedar was used instead of eucalyptus used in Example 1. The results are shown in Table 1. Further, the X-ray diffraction analysis was performed on the sample at each stage by the same operation as in Example 1. The X-ray diffraction pattern of each sample was similar to the corresponding eucalyptus sample in Example 1. The diffraction pattern after coarse pulverization is shown in FIG. 11, the diffraction pattern of the sample after ammonia treatment is shown in FIG. 13, and the diffraction pattern of the sample after ammonia treatment and pulverization treatment is shown in FIG.
The total glycation rate of the used cedar was 42.7%, and the average particle size of the coarsely pulverized cedar was 207 μm.

(Comparative Examples 3-1 to 3-4)
In each of Comparative Examples 1-1 to 1-4, Comparative Example 1 corresponding to Example 1 was used except that the coarsely ground cedar used in Example 3 was used instead of the coarsely ground Eucalyptus used as the starting material. Pretreatment was performed in the same manner as in -1 to 1-4, and enzymatic saccharification reaction was performed in the same manner as in Example 3 for the samples obtained in each. For each comparative example, the glucose yield was calculated and the results are shown in Table 1. Further, for Comparative Examples 3-2 to 3-4, X-ray diffraction analysis was performed on each pretreated sample. The X-ray diffraction pattern of each sample was similar to each eucalyptus sample in Comparative Examples 1-2 to 1-4 corresponding to each sample. FIG. 12 shows the diffraction pattern after the grinding treatment, and FIG. 14 shows the diffraction pattern of the sample treated with ammonia after the grinding treatment.

From the results in Table 1, the biomass saccharification efficiency is improved with respect to the untreated biomass raw material, the biomass raw material subjected to the ammonia treatment or the pulverization treatment by further pulverizing the biomass raw material and subjecting it to enzymatic saccharification In addition, it was shown that the enzyme saccharification efficiency can be improved even when compared with the case where the ammonia treatment is performed after the pulverization treatment. Further, from the results shown in FIGS. 1 to 15, by treating the biomass raw material with ammonia, X-ray diffraction peaks attributed to cellulose III type _{I of} 2θ = 12 °, 17 ° and 21 ° appear, and this is pulverized. As a result, it was revealed that the diffraction pattern changes remarkably, and that the diffraction pattern when pulverized after the ammonia treatment is different from that when the biomass raw material is ammonia-treated after pulverization. This suggests that the biomass raw material has a specific structure by pulverization after ammonia treatment.

The sugar production method, ethanol production method, and lactic acid production method of the present invention can significantly improve sugar production efficiency, ethanol production efficiency, and lactic acid production efficiency. In addition, according to the method for producing a raw material for enzyme saccharification of the present invention, it is possible to efficiently obtain a material for enzyme saccharification suitable for the above-described sugar production method, ethanol production method, and lactic acid production method of the present invention. it can. Therefore, the sugar production method, ethanol production method, lactic acid production method, and enzyme saccharification raw material production method of the present invention aim to produce an environmentally friendly fuel that has been attracting attention in recent years, for example. It can be suitably used for the production of ethanol from biomass raw materials, and the production of environmentally friendly biodegradable plastics.

Claims

(A) a step of obtaining a modified biomass raw material by treating a biomass raw material containing cellulose type I with a treatment agent containing ammonia and / or an organic amine;
(B) a step of obtaining a raw material for enzyme saccharification by grinding the modified biomass raw material; and
(C) a step of saccharifying the enzyme saccharification raw material to obtain a saccharide;
A method for producing sugar, comprising:
The method for producing sugar according to claim 1, wherein the treating agent used in step (a) is ammonia.
The method for producing sugar according to claim 1 or 2, wherein the biomass raw material containing cellulose type I is woody biomass.
The average particle diameter represented by the median diameter of the enzyme saccharification raw material obtained in step (b) is 5 to 80 µm, according to any one of claims 1 to 3, A method for producing the sugar as described.
A method for producing ethanol, wherein the saccharide obtained by the method for producing saccharide according to any one of claims 1 to 4 is fermented to obtain ethanol.
A method for producing lactic acid, wherein the saccharide obtained by the method for producing saccharide according to any one of claims 1 to 4 is fermented to obtain lactic acid.
(A) a step of obtaining a modified biomass raw material by treating a biomass raw material containing cellulose type I with a treatment agent containing ammonia and / or an organic amine; and
(B) a step of obtaining a raw material for enzymatic saccharification by pulverizing the modified biomass raw material,
The manufacturing method of the raw material for enzyme saccharification characterized by including this.