JP2006141244A - Method for saccharifying lignocellulose and the resultant saccharide - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a method for saccharifying lignin-containing lignocellulose. <P>SOLUTION: The method for saccharifying lignocellulose comprises the following process: Ozone is blown into an aqueous suspension containing the lignocellulose comprising lignin, hemicellulose and cellulose to decompose the lignin. Among holocellulose obtained after filtration of the resultant aqueous suspension, the hemicellulose is solubilized with hot water, and the hot water is filtered off to collect the cellulose. Thereafter, the cellulose is solubilized in hot water and then hydrolyzed by a relevant enzyme to obtain cellooligosaccharide and/or glucose. Another version of this method comprises the following process: Ozone is blown into an aqueous suspension containing the lignocellulose comprising lignin, hemicellulose and cellulose to decompose the lignin. Among holocellulose obtained after filtration of the resultant aqueous suspension, the hemicellulose is solubilized with hot water, and the hot water is filtered off to remove the cellulose. Thereafter, the hemicellulose is hydrolyzed by a relevant enzyme to obtain xylooligosaccharide and/or xylose. <P>COPYRIGHT: (C)2006,JPO&NCIPI

Description

  The present invention relates to a method for saccharification of lignocellulose by enzymatic treatment, and more particularly, to a method for saccharification of lignocellulose in which ozone treatment is performed prior to the decomposition of lignocellulose comprising lignin, hemicellulose and cellulose using an enzyme. The present invention relates to a sugar obtained by a saccharification method.

Technologies for effectively using so-called biomass such as wood chips, wood flour, rice straw, and fir shells have been developed.
Biomass is extremely cheap because it exists as a waste material, and since it contains lignin, hemicellulose, cellulose, etc., various substances are produced using these components as starting materials.
Among these, development of technology for producing sugar (eg, xylooligosaccharide, cellooligosaccharide, xylose, glycolose, etc.) from biomass has been attracting attention recently due to the demand for alcoholic fermentation technology.

For example, as a method for saccharification of lignocellulose, there is a chemical method using acetic anhydride or concentrated sulfuric acid, but a drying process is required, and the hydrolysis of cellulose is intense, and the equipment of the reaction system is complicated. Become.
Moreover, the entire equipment is easily corroded by acid, and extra cost is required for recovery of sulfuric acid.
Furthermore, the yield of the produced sugar is low.
On the other hand, in order to eliminate such drawbacks, an enzymatic method capable of performing hydrolysis under mild conditions has been devised.

  This method hydrolyzes using an enzyme (for example, cellulase), and does not require a reaction vessel having pressure resistance and acid resistance in terms of apparatus, and also has little environmental pollution.

Moreover, it is also advantageous that the yield is higher than that of the chemical method.
As a saccharification method using such an enzyme, for example, Patent Document 1, Patent Document 2, and the like have been proposed.
JP 2001-95594 A JP-A-8-308589

However, in general, lignocellulose using biomass as a raw material contains lignin, hemicellulose, and cellulose, and this lignin is a major obstacle to saccharification.
That is, lignocellulose is solidified in a state in which hemicellulose and cellulose are glued by lignin, and the presence of this lignin greatly reduces the efficiency of hydrolysis of hemicellulose and cellulose, resulting in a very low saccharification rate. It is.
Furthermore, since most biomass is supplied in a wet state with a moisture content of 100% or more, development of a treatment method that does not require a drying step is required.
The present invention has been made based on such technical background.
That is, the present invention provides a saccharification method capable of efficiently saccharifying lignocellulose containing lignin.
Moreover, the sugar obtained by the saccharification method is provided.

  As a result of diligent research on the above problems, the present inventors, when hemicellulose and saccharification of cellulose are performed by hydrothermal treatment, the lignin that solidifies the lignin is first decomposed with ozone, so that I also found that the problem could be solved.

  That is, the present invention is (1) a method for decomposing and saccharifying lignocellulose comprising lignin, hemicellulose and cellulose using an enzyme, and decomposing lignin by performing ozone treatment before the decomposition using the enzyme. A method for saccharification of lignocellulose, characterized in that

  The present invention also provides (2) a holo obtained by blowing ozone gas into an aqueous suspension containing lignocellulose composed of lignin, hemicellulose and cellulose to decompose lignin, and then filtering the aqueous suspension. Among cellulose, hemicellulose is solubilized with heated water, the heated water is filtered to remove the cellulose, then the cellulose is solubilized in heated water, and hydrolyzed with an enzyme to obtain cellooligosaccharide and / or glycolose. A method for saccharification of lignocellulose characterized by

  The present invention also relates to (3), a holo obtained by blowing ozone gas into an aqueous suspension containing lignocellulose composed of lignin, hemicellulose and cellulose to decompose lignin and then filtering the aqueous suspension. Lignocellulose characterized in that hemicellulose of cellulose is solubilized with heated water, the heated water is filtered to remove cellulose, and then the hemicellulose is hydrolyzed with an enzyme to obtain xylooligosaccharide and / or xylose. Saccharification method.

  The present invention also resides in (4) the lignocellulose saccharification method according to 2 or 3 above, wherein the heating temperature for solubilizing hemicellulose is 140 to 200 ° C.

  The present invention also resides in (5) the method for saccharification of lignocellulose as described in 2 above, wherein the temperature of the heated water for solubilizing the cellulose is 190 ° C to 250 ° C.

  The present invention also resides in (6) the lignocellulose saccharification method according to the above 2, characterized in that xylanase is used as an enzyme for hydrolyzing hemicellulose.

  The present invention also resides in (7) the method for saccharification of lignocellulose as described in 2 above, wherein cellulase is used as an enzyme for hydrolyzing cellulose.

  The present invention also relates to (8) a sugar produced by the saccharification method according to any one of 1 to 7 above.

  In addition, as long as the objective of this invention is followed, the structure which combined said (1) to (8) suitably is also employable.

The present invention is a stage before decomposing lignocellulose composed of lignin, hemicellulose and cellulose using an enzyme, and in order to decompose lignin by performing ozone treatment, hemicellulose and cellulose are efficiently hydrolyzed. The yield is also greatly improved.
Specifically, by performing ozone treatment, the heat treatment temperature for solubilizing hemicellulose and cellulose is lowered and heat efficiency is improved as compared with the case where ozone treatment is not performed.

The present invention is characterized in that in a method for decomposing and saccharifying lignocellulose comprising lignin, hemicellulose, and cellulose using an enzyme, the lignin is decomposed by performing ozone treatment before the decomposition using the enzyme. .
FIG. 1 is a process diagram showing ozone treatment, and FIG. 2 is a process diagram showing hot water treatment and saccharification.

[Ozone treatment]
Lignocellulose is one in which hemicellulose and cellulose are solidified by lignin, and in this state, the efficiency is low even if it is attempted to solubilize hemicellulose or cellulose by hydrothermal treatment (heat treatment with water).
Therefore, before heat treatment with water, thermal heat, this lignin is decomposed by ozone treatment.
Specifically, first, when water is added to a powdered lignocellulose and stirred, an aqueous suspension containing a solid content is formed.

The aqueous suspension is subjected to ozone treatment. Specifically, for example, ozone is blown into the aqueous suspension in the container to decompose lignin contained in the solid lignocellulose.
Lignin is oxidatively decomposed by performing ozone treatment at room temperature for a certain period of time and decomposed into organic acids such as gallic acid, oxalic acid, acetic acid, formic acid, etc., and this organic acid is dissolved in water.

This aqueous suspension contains organic acid and solid content of holocellulose (hemicellulose and cellulose).
In order to separate the organic acid and the holocellulose from such an aqueous suspension, the aqueous suspension is filtered to take out the holocellulose as a solid content.
The remaining liquid becomes an organic acid solution and can be used effectively for others.
The holocellulose taken out by filtration may be kept in a wet state or dried after washing with water.

[Method of saccharification of cellulose]
(Solubilization of hemicellulose)
The holocellulose from which the lignin has been removed through the ozone treatment step is then treated with heated water to solubilize the hemicellulose in the holocellulose (hot water treatment).
Usually, the heating temperature of this heated water is preferably 140 ° C. to 200 ° C. from the viewpoint of mainly solubilizing hemicellulose.
Since cellulose does not dissolve and remains as a solid, it is filtered and separated from hemicellulose and taken out as a filtrate.

(Solubilization of cellulose)
The taken-out cellulose is solubilized with heated water (hot water treatment).
In this case, the heating temperature of the heated water is preferably 190 ° C. to 250 ° C. from the viewpoint of solubilizing cellulose.

(Saccharification of cellulose)
Cellulose solubilized in heated water is hydrolyzed by enzymes to obtain cellooligosaccharides and / or glycolose.
Here, cellulase, glucoamylase, tannase, phytase, protease, chitinase, etc. are appropriately selected and used as the enzyme that hydrolyzes cellulose.
The enzymatic hydrolysis is preferably performed at a temperature slightly higher than room temperature, for example, 40 to 60 ° C.

[Method for saccharification of hemicellulose]
(Solubilization of hemicellulose)
The holocellulose from which the lignin has been removed through the ozone treatment step is then treated with heated water to solubilize the hemicellulose in the holocellulose (hot water treatment).
The heating temperature of this heated water is preferably 140 ° C. to 200 ° C. from the viewpoint of solubilizing only hemicellulose as described above.
Since cellulose is not dissolved and remains a solid, it is separated by filtration and taken out.

(Saccharification of hemicellulose)
The hemicellulose solubilized solution from which the cellulose has been removed is hydrolyzed by an enzyme to obtain xylooligosaccharide and / or xylose.
Here, as the enzyme that hydrolyzes hemicellulose, xylanase, hemicellulase, cellulase, or the like is appropriately selected and used.
The enzymatic hydrolysis is preferably performed at a temperature slightly higher than room temperature, for example, 40 to 60 ° C., as in the case of cellulose described above.
EXAMPLES Hereinafter, although this invention is demonstrated based on an Example, this invention is not limited only to an Example described below.

[Example 1]
Here, the effectiveness of ozone treatment was tested.
Sugi sawdust (2.5 g, 4 lumps) was prepared and stirred in 1 liter of water in a beaker to create individual water suspensions.
Ozone (specifically, oxygen containing ozone) was blown into and supplied to this water suspension.
The supply amount of ozone was 0.5 liter / min.
The main component of cedar sawdust is lignocellulose composed of lignin, hemicellulose, and cellulose, among which lignin was oxidized by ozone and decomposed.
This ozone treatment was performed in four cases: 0 hours, 2 hours, 4 hours (twice), and 8 hours.
At that time, it can be seen that the residual solids (lignin, hemicellulose, cellulose) are decomposed in this order by the total amount of carbon solubilized in the aqueous suspension.

FIG. 3 shows the result.
The reaction time (ozone treatment time), the ratio of the solubilized carbon amount (TC) to the total carbon amount (TCo) of the aqueous suspension, the pH of the aqueous suspension, and the residual solid content (% by weight) are shown.

[Example 2]
(Hot water treatment)
The aqueous suspension that had been subjected to ozone treatment for 4 hours in the experiment as in Example 1 was filtered and dried to prepare 0.5 g of a solid content.
This solid content was decomposed by a hot water flow type reactor (the temperature of the reaction vessel in the device can be increased with time).
The pressure in the reaction vessel was 10 MPa.
The decomposition solution was collected at 10-minute intervals to obtain 20 mL of reaction solution.
About the reaction liquid, pH and total carbon amount were measured.

FIG. 4 shows the result.
The vertical axis represents the heating temperature (reaction temperature), the total amount of carbon solubilized in water (mg / 20 mL), and the horizontal axis represents the reaction time (minutes).

(Glycation treatment with enzymes)
3 mg / 5 mL of cellulase (Cellulosin T2 manufactured by HBI) was added as an enzyme to the reaction solution after measuring the pH and the total amount of carbon, and reacted at 50 ° C. for 20 hours.
The amount of glucose in the reaction solution was measured with a glucose test Wako reagent (manufactured by Wako Pure Chemical Industries).

FIG. 5 shows the result.
The vertical axis shows the amount of glucose produced (mg / 20 mL) and pH, and the horizontal axis shows the reaction time (minutes).

[Comparative Example 1]
Here, it experimented about the case where ozone treatment is not performed.

(Hot water treatment)
A solid content of 0.5 g of cedar sawdust was prepared.
This solid content was decomposed by a hot water flow type reactor (the temperature of the reaction vessel in the device can be increased with time).
The pressure in the reaction vessel was 10 MPa.
The decomposition solution was collected at 10-minute intervals to obtain 20 mL of reaction solution.
About the reaction liquid, pH and total carbon amount were measured.
FIG. 6 shows the result.

(Glycation treatment with enzymes)
3 mg / 5 mL of cellulase (Cellulosin T2 manufactured by HBI) was added as an enzyme to the reaction solution after measuring the pH and the total amount of carbon, and reacted at 50 ° C. for 20 hours.
The amount of glucose in the reaction solution was measured with a glucose test Wako device (manufactured by Wako Pure Chemical Industries).
FIG. 7 shows the result.

Example 3
Individual water suspensions that had been subjected to ozone treatment by experiments as in Example 1 were filtered and dried to produce 1 g of a solid content.
This was left in 40 mL of heated water (150 ° C.) for 4 hours to solubilize hemicellulose, and 10 mL of the supernatant was collected.
3 mg / 5 mL of xylanase (Helcer Cellulose HC100) was added to 10 mL of the supernatant and reacted at 50 ° C. for 24 hours.
An ion chromatograph (manufactured by DIONEX) was used for analysis of saccharides in the reaction solution.
The sugar concentration was substituted by the peak area.
FIG. 8 shows the result.

[Evaluation]
(Knowledge in Example 1)
When the reaction time is 2 hours, the residual solid content is reduced to 70% by weight, indicating that lignin (usually occupying 30% by weight of lignocellulose) is decomposed by the reaction for 2 hours.
Since the ozone durability gradually increases in the order of lignin, hemicellulose, and cellulose, it was confirmed that the decomposition of hemicellulose (cellulose is also decomposed into an organic acid) started at a reaction time of 4 hours.
This shows that a reaction time of about 2 hours is sufficient.

(Knowledge in Example 2)
Hemicellulose started to melt from 140 ° C. and was almost completely solubilized at 190 ° C., while cellulose began to melt at 190 ° C. and almost all was solubilized at 220 ° C.
Moreover, it turned out that hemicellulose or a cellulose will decompose excessively when it exceeds 250 degreeC.
In addition, after overdecomposition, it becomes, for example, hydroxymethylfurfural.

(Knowledge in Comparative Example 1)
Hemicellulose began to melt at 150 ° C and solubilized almost at 180 ° C, while cellulose began to melt at 225 ° C and solubilized almost at 250 ° C.

(Knowledge in Example 3)
It can be seen that the yield of xylo-oligosaccharide is greater when ozone treatment is performed than when ozone treatment is not performed.
In particular, when ozone treatment is performed for 2 hours, xylo-oligosaccharide can be obtained efficiently.

(Comparison between Example 2 and Comparative Example 1)
From the comparison between FIG. 4 and FIG. 6, when both hemicellulose and cellulose are subjected to ozone treatment, the heat treatment temperature for solubilization is lower than when ozone treatment is not performed.
Therefore, it can be understood that it can be efficiently solubilized also thermally, and as a result, saccharification is efficiently performed.
From the comparison between FIG. 5 and FIG. 7, it was found that the yield of glucose was several times higher when the ozone treatment was performed than when the ozone treatment was not performed.

FIG. 1 is a process diagram showing ozone treatment. FIG. 2 is a process diagram showing heat treatment and saccharification. FIG. 3 shows the result of the hot water treatment when the ozone treatment time is changed. FIG. 4 shows the result of the hot water treatment when the ozone treatment is performed for 4 hours. FIG. 5 shows the result of saccharification treatment when ozone treatment is performed for 4 hours. FIG. 6 shows the result of the hot water treatment when no ozone treatment is performed. FIG. 7 shows the result of saccharification treatment when ozone treatment is not performed. FIG. 8 shows the result of saccharification in the hot water treatment when the ozone treatment time is changed.

Claims (8)

  A method of decomposing and saccharifying lignocellulose consisting of lignin, hemicellulose and cellulose using an enzyme, wherein lignin is decomposed by performing ozone treatment before the decomposition using the enzyme. Saccharification method.   Lignin is decomposed by blowing ozone gas into an aqueous suspension containing lignocellulose consisting of lignin, hemicellulose, and cellulose, and then hemicellulose is solubilized with heated water from the holocellulose obtained by filtering the aqueous suspension. A method for saccharification of lignocellulose, wherein cellulose is removed by filtering the heated water, and then the cellulose is solubilized in heated water and hydrolyzed by an enzyme to obtain cellooligosaccharides and / or glycolose.   Lignin is decomposed by blowing ozone gas into an aqueous suspension containing lignocellulose consisting of lignin, hemicellulose, and cellulose, and then hemicellulose is solubilized with heated water from the holocellulose obtained by filtering the aqueous suspension. A method for saccharification of lignocellulose, wherein the heated water is filtered to remove cellulose, and then the hemicellulose is hydrolyzed with an enzyme to obtain xylooligosaccharide and / or xylose.   The method for saccharification of lignocellulose according to claim 2 or 3, wherein the heating temperature for solubilizing hemicellulose is 140 ° C to 200 ° C.   The method for saccharification of lignocellulose according to claim 2, wherein the temperature of the heated water for solubilizing the cellulose is 190 ° C to 250 ° C or higher.   The method for saccharification of lignocellulose according to claim 2, wherein xylanase is used as an enzyme for hydrolyzing hemicellulose.   The method for saccharification of lignocellulose according to claim 2, wherein cellulase is used as an enzyme for hydrolyzing cellulose. A sugar produced by the saccharification method according to any one of claims 1 to 7.

Images