JPS585197A - Saccharifying method of cellulose - Google Patents

Abstract

PURPOSE:To saccharify the titled substance, by ozonizing a pulverized fibrous or granular material containing cellulose with a specific moisture content, and reacting the treated cellulose with an enzyme. CONSTITUTION:A pulverized fibrous or granular material, containing cellulose and having a granular size <=5mm. is adjusted to a moisture content of 30- 90wt%, and then treated with 1-30wt%, based on the dried material containing the cellulose, ozone in the gaseous form. The tissue is destroyed by the ozone treatment, and the treated material is then reacted with an enzyme containig cellulose derived from a microorganism belonging to the genus Aspergillus or Trichoderma and decomposed enzymically. The material containing the cellulose is saccharified and converted into a monosaccharide.

Description

[Detailed description of the invention] The present invention relates to a method for enzymatic saccharification of cellulose.

Specifically, the present invention relates to a wood saccharification method in which crushed cellulose is treated with ozone and then enzymatically decomposed.

In the past, methods for enzymatically decomposing and saccharifying cellulose-containing materials such as wood have been studied in various fields, and in particular, wood saccharification methods that utilize cellulase-containing enzymes are said to require destroying the wood tissue. ing. Methods for destroying the structure of such cellulose-containing materials include, for example, pulverizing them into particles with a particle size of 50 μm or less using a ball mill, freeze-pulverization methods, and radiation irradiation methods. However, these methods take a long time to destroy the structure of cellulose.
It usually takes 15 to 16 hours and requires a large amount of electrical energy. In particular, the freeze-grinding method and the radiation irradiation method require a large amount of electrical power as well as the creation of special processing equipment and operating environments. on the other hand.

Chemical tissue destruction methods such as the chlorite method and peracetic acid method are known, but not only do they require large amounts of these chemicals, but they also require large amounts of chemicals and lignin-containing treatments. There is a drawback that a large amount of expense is required for processing the waste liquid for disposal.

The present inventors have overcame the deficiencies of conventional methods for chemically destroying the tissue of cellulose-containing substances, and have conducted repeated research on a method for effectively chemically destroying the tissue and saccharifying cellulose through enzymatic decomposition. result.

We have found a very effective method.

The present invention involves treating a cellulose-containing material that has been ground into fibers or granules with ozone under conditions of a moisture content of 30 to 90% by weight to destroy its structure, and then enzymatically decomposing it by acting on an enzyme containing cellulase. The gist of this paper is a method for saccharification of cellulose, which is characterized by the following.

For example, it is composed of three typical cellulose-containing materials: wood cellulose, hemicellulose, and phosphorus. The proportions vary depending on the species of wood, but usually cellulose is 40 to 50% by weight.

Hemicellulose 20-30% by weight and lignin 20-3%
It consists of about 0 weight factor. As described above, the wood tissue of such constituent components is destroyed as much as possible by various means, and then saccharified by cellulase-containing enzymes.

In the present invention, the cellulose-containing material is ground into fibrous or granular forms. In this case, 1, for example, wood can be easily ground into fibers or granules using a conventional mechanical bulging method such as a Uille set grinder or a disc refiner.

The tissue of the thus obtained pulverized product is destroyed by the action of ozone in the presence of water, and is effectively decomposed and saccharified by saccharifying enzymes. However, if the diameter of the pulverized product is 5 mm or more, it is inconvenient because little improvement in the enzymatic decomposition rate of the ozonated product can be expected.

Therefore, in the present invention, it is desirable that the cellulase-containing enzyme be pulverized into fibers or particles smaller than 5 groups.

The ground cellulose-containing material is then treated with ozonation in the presence of water to chemically disrupt its structure. In this ozone treatment, it has been found that the amount of admixture water, mainly adhering water and impregnated water, which is in contact with the cellulose-containing material and the cellulose-containing material, has a very large influence on the enzymatic decomposition rate. The amount of ozone used to treat the loin-containing material varies depending on the pulverized state of the cellulose-containing material and the amount of water present, but is approximately 1 to 30 ozone per dry cellulose-containing material, preferably 10 to 25 ozone.
The weight factor may be conveniently employed and the suitable amount selected depending on other conditions.

In the present invention, ozone is preferably applied in gaseous form to the pulverized hydrated cellulose.

Ozone only needs to destroy the structure of the cellulose-containing material, and a sufficient amount for that purpose is advantageously used, but during the destruction process, the crushed cellulose-containing material must be adjusted to have a moisture content in the range of 30 to 90% by weight. It is important to.

When the water content deviates from the above range, the decomposition rate of the ozone-treated cellulose by enzymes decreases, which is not preferable. This will be further clarified in the specific examples below.

Such water content can be easily adjusted by immersing the ground cellulose-containing material in water and then squeezing it, for example.

The ozone-treated cellulose-containing material is decomposed by a commonly known saccharifying enzyme, such as a cellulase-containing enzyme produced by various microorganisms such as Aspergillus, Irpex, and Trichoderma. In this way, cellulose-containing substances are saccharified and converted into monosaccharides.

The method of the present invention is extremely simple to operate compared to conventional methods for saccharification of cellulose-containing substances.

There is also no need to worry about environmental pollution such as water pollution.

In the following, the present invention will be explained in more detail with reference to Examples.

Example 1 A thermomechanical valve (hereinafter referred to as TMP) was obtained by treating red pine tree with a pressurized disc refiner. This undried TMP (moisture content 69, 61)16.
45. ! i' (Collect 5.1 as dry TMP in a 500-shaped eggplant flask, attach it to a rotary evaporator, and rotate it while aerating 3 volumes of ozone from one end at a flow rate of 200 m4/min, and The tissue was destroyed by discharging it from the end. Various treated samples were prepared with different aeration times, or in other words, different amounts of ozone action.
They were washed with water 1-air dried.

To each 2 g of the ozonated air-dried sample obtained in this way,
0 containing commercially available cellulase (meicelase) 50
, add 1M acetate buffer (pH 5,0) 1.0-,
It was incubated at a temperature of 40°C.

After 48 hours, the decomposition liquid was filtered through a 304 glass filter, the undecomposed residue was thoroughly dried at a temperature of 105°C, and its weight was measured. In addition, the reducing sugar content of the obtained F solution was quantified, and the relationship between the amount of ozone action and the pulp ≠ decomposition rate is summarized in Table 1 below.

For comparison, an experiment was conducted on TMP that was not treated with ozone under the same enzymatic decomposition conditions as above, and the results are also shown in the table below.

As can be seen from the table above, the reducing sugar production rate of un-ozonated TMP is 19%, while that of ozonated TMP is 5.0% by weight, and as the amount of ozone action increases, the reduction increases. Sugar production rate, that is, saccharification and decomposition 4 shows the maximum enzyme decomposition rate when 24 weight percent ozone is applied,
At an ozone action amount of 36% by weight, there is a tendency for the decomposition rate to decrease somewhat.

Example 2 The TMP obtained in Example 1 was air-dried, 5 g of the completely dried net amount was collected, immersed in water for 1 hour, and then degassed for 50 minutes to obtain a sample that had sufficiently permeated with water. Samples with various moisture contents were prepared by dehydration on a Buchner funnel. Ozone was aerated through each of these for 6 cycles, and then the same procedure as in Example 1 was carried out for 1 cycle.

The sample thus obtained was subjected to enzymatic decomposition and treated in the same manner to determine the relationship between the water content and saccharide degradation rate of the sample. The results are shown in Table 2.

Table 2 TMP It can be seen that when red pine ° is treated with ozone having a hexagonal relationship, the enzymatic decomposition rate peaks at a water content of around 46 wt.

Example 3 Red pine wood flour (60-8o mesh, alcohol and benzene extracted) and air-dried TMP of Example 2.

After adjusting the moisture content to 50 to 54% by weight using the method shown in Example 2 and aerating with ozone for various times, the relationship between the amount of ozone action and the saccharide decomposition rate was determined in the same manner as in Example 1. The results are shown in Table 3.

Table 3

Claims (1)

[Claims] 1. A cellulose-containing material ground into fibrous or granular form,
A method for saccharification of cellulose, which comprises treating the cellulose with ozone under conditions of a moisture content of 30 to 90% by weight to destroy its tissue, and then enzymatically decomposing the cellulose by applying an enzyme containing cellulase. 2. The method according to claim 1, wherein the cellulose-containing material is wood. 3. Claim 1 in which ozone is applied to the dry absolute amount of the pulverized cellulose-containing material by 1 to 30% by weight.
2. The method according to any one of paragraphs 1 and 2. 4. The method according to any one of claims 1 to 3, wherein the cellulose-containing material is pulverized into fibers or particles of 5 sm or less.