JPH0342077B2 - - Google Patents

Description

[Detailed description of the invention]

[Field of Application of the Invention] The present invention provides a method for saccharifying cellulosic substances such as straw, bagasse, corn stover, and wood chips using a fibrinolytic enzyme, by adding a high concentration of cellulosic substances and continuously or semi-continuously The present invention relates to an improved saccharification method and apparatus for saccharification. [Background of the Invention] Conventionally, cellulose-based materials require delignification and destruction of the cellulose crystal structure as pretreatment in order to allow fibrinolytic enzymes to act on them. However, in order to put this pretreatment method into practical use, (1) the energy consumption required for pretreatment is low, (2) the conversion rate of pretreated raw materials to sugar is high, and (3) the saccharification reaction It is required to efficiently obtain a highly concentrated sugar solution with less energy consumption. Among these, for (1) and (2), combinations of pretreatment methods, such as a combination of alkali treatment and pulverization or ozone treatment, are being considered. On the other hand, in order to obtain the highly concentrated sugar solution (3), it is necessary to prepare the pretreated raw materials at a high concentration. In that case,
The fiber length of the raw material affects the saccharification reaction. If the raw material fiber length is finely pulverized to about 10 to 30μ,
Although there is no problem with stirring and mixing and continuous saccharification, the energy consumption required for this fine pulverization is extremely large, so it is not practical. Therefore, methods have been studied that allow pretreated raw materials that are not pulverized to be charged at a high concentration. However, the current situation is that no satisfactory method has been developed yet. For example, in a method in which raw materials are sequentially added and saccharification is performed in a slurry state, as the amount added increases, residue accumulates and becomes a sludge, so there is a problem that there is a limit to the amount added. As another method, a saccharification method using a saccharification tank that can perform sludge kneading operations requires a large capacity saccharification tank and the installation of a large-capacity stirring motor, and the mixing within the tank is incomplete. However, there are problems in that continuous saccharification is difficult due to the production of raw materials. Therefore, there is a need for the development of a saccharification method that can continuously saccharify pretreated raw materials that are not pulverized at high concentrations and at high loading rates. [Object of the Invention] The present invention was made to solve the problems of the prior art described above, and its purpose is to prepare pretreated raw materials that are not pulverized at a high concentration and perform saccharification without stirring. An object of the present invention is to provide a saccharification method and an apparatus for the same. [Summary of the Invention] To summarize the present invention, the first invention of the present invention is an invention relating to a method for saccharification of cellulose-based substances,
In a saccharification method for obtaining a sugar solution by decomposing a cellulose substance with a fibrinolytic enzyme, (1) a step of mixing the cellulose substance and a fibrinolytic enzyme via water;
(2) a step of supplying the mixture to a tower-type saccharification tank and extracting the product mixture from the bottom of the tank after residence for a predetermined time; and (3)
It is characterized in that it includes the steps of separating the product mixture into solid and liquid and recovering a sugar solution. Further, the second invention of the present invention is an invention related to a saccharification device for cellulosic substances, and in the saccharification device for obtaining a sugar solution by decomposing a cellulose material with a fibrinolytic enzyme, a cellulose-based material raw material and a fibrinolytic enzyme are used. A means for mixing with an aqueous solution containing a processing agent containing an enzyme, a means for charging the mixture into the saccharification tank, a tower-type saccharification tank with a jacket, a means for extracting the product mixture from the lower part of the saccharification tank, and a solid-liquid separation means. Features. The present invention was completed by analyzing the saccharification reaction of high-concentration raw materials as described below. Batch saccharification of unpulverized alkali-treated or ozonated bagasse (average particle size 0.7 mm) at a concentration of 10 to 15% by weight is carried out in a horizontal tank (with ribbon-shaped blades) with an inner diameter of 20 cm and a length of 40 cm. Ivy. In addition,
As cellulase, commercially available Cellulase Onozuka R-10 (manufactured by Kinki Yakult Co., Ltd., optimal temperature 50°C,
pH4.8) was used. As a result, both the alkali-treated and ozone-treated bagasse were in a sludge state before saccharification, but with the saccharification reaction, the sludge volume decreased and free aqueous liquid increased, resulting in slurry. Then, the slurry state at this time was analyzed. A reaction tube with an inner diameter of 4 cm and a height of 6 cm with a net installed at the bottom was prototyped, and free aqueous liquid was sampled over time to measure the amount of change. Figure 1 shows the measurement results. In other words, Figure 1 shows the relationship between time (hours) (horizontal axis), the volume of the packed layer (ml), the concentration of glucose and cellobiose (g/h), and the outflow rate of free aqueous liquid (ml/h).
and glucose, cellobiose concentration (g/)
(vertical axis). As is clear from FIG. 1, a large amount of free aqueous liquid is produced at the early stage of saccharification. The total amount is approximately the total water content.
It was 30%. At this time, the raw material sludge remained in a sludge state and its volume decreased as a result of the saccharification reaction. This volume reduction rate was approximately 30-40%. Based on this, it was thought that if a saccharification tank having a structure similar to the reaction tube described above was used to carry out the saccharification reaction using the sequential addition method, a large amount of raw material could be charged to cover the reduced volume of the raw material sludge. Therefore, we used a tower-type saccharification tank with an internal diameter of 5 cm and a height of 20 cm, which had a mesh installed at the bottom of the tank, and added 100 g of the ozonated bagasse and cellulase aqueous solution (water content approximately 85% by weight) every 24 hours (filling height: Sa7
cm) was added sequentially to carry out the saccharification reaction. Figure 2 shows the results. In other words, Figure 2 shows the relationship between the saccharification time (hours) (horizontal axis), the amount of free aqueous liquid produced (cumulative flow rate) (ml) extracted from the bottom of the saccharification tank, and the glucose concentration (mg/ml) (vertical axis). It is a graph showing the relationship between. As is clear from Fig. 2, the sugar concentration of the free aqueous liquid increases as the amount of addition increases, and 3
- After the 4th time, it became almost constant. Each addition also produced 50-60 ml of free aqueous liquid. The total volume of raw materials charged at this time was about 1/3 compared to a batch reaction system in which free aqueous liquid was not removed. but,
The sugar concentration of the liquid separated from the total residual sludge in the saccharification tank was lower than in the batch reaction system in which free aqueous liquid was not removed. This is because the saccharification reaction is distributed in the filling height direction. For this reason, in the above-mentioned saccharification method, in addition to sequential addition,
It has been found that it is better to extract the sludge from the bottom of the saccharification tank, that is, to operate continuously or semi-continuously. Therefore, the saccharification reaction rate is higher in the early stages of the saccharification reaction, and the rate of increase gradually decreases and becomes constant. In this experiment, it became constant after 30 to 48 hours. Therefore, it is efficient to extract the sludge from the tank when the residence time of the raw material at the bottom of the saccharification tank reaches a time at which the saccharification rate becomes constant. Then, by separating the liquid from the extracted raw material sludge, a highly concentrated sugar solution can be obtained. However, there were concerns about dilution due to free water. Therefore, we investigated the flow of water in the sludge-filled layer. As a result, it was found that water was released from the packed layer only when the amount of water retained in the packed layer reached 80% by weight or more, and the water moved downward due to the extrusion flow. Furthermore, in the above experimental data, the maximum amount of free water produced per unit time after the sixth addition is as small as about 2.6% of the total amount of retained water. It takes about 50 hours or more for the free water in the upper stage to reach the lower stage. Therefore, it was estimated that the free aqueous liquid obtained from the bottom of the saccharification tank was extruded from the sludge located near the bottom of the tank. In fact, in the experiment described above, the sugar concentration of the free aqueous liquid in the sixth experiment almost matched the concentration of the sugar solution contained in the sludge near the bottom of the tank. From this, it was determined that there was no dilution due to free water. Therefore, semi-continuous saccharification was performed using the saccharification tank described above. After adding 100 g of the mixture of pretreated bagasse and cellulase solution to 5 portions every 12 hours, the residence time was
Saccharification was performed for 60 hours. In addition, a net was installed between each sludge so that the raw material sludge in each category could be determined. As a result, the sugar concentrations of the liquid separated from the sampled sludge and the free aqueous liquid were almost the same (glucose 54 g/g), which was the same as the value in the batch system saccharification reaction. Next, conditions for effectively implementing the present invention,
That is, the conditions under which the volume reduction rate of the sludge-like raw material becomes large were investigated. We focused on the retained moisture content of the sludge-like raw material and the fiber length of the cellulose material as influencing factors. First, saccharification raw materials with various mixing ratios of bagasse and cellulase aqueous solution were prepared, and
The material was charged into a saccharification tube with a height of 6 cm and a screen installed at the bottom of the tank, and the volume reduction rate of the raw material, that is, (volume after saccharification/volume before saccharification) x 100 (%), was determined. The average fiber length of bagasse is 0.7
mm. The measurement results are shown in Figure 3. That is, FIG. 3 is a graph showing the relationship between solid content concentration (weight %) (horizontal axis) and volume reduction rate (%) (vertical axis). As is clear from FIG. 3, at a solid content concentration of 5% by weight, the prepared raw material was in the form of a slurry, the liquid flowed out through the net, and bagasse remained in the form of sludge in the saccharification tube. At this time, the retained moisture content of the residual bagasse, that is, [(retained water amount/(retained water amount + solid material amount)] × 100 (%)) was 80 to 90% by weight. progressed, and the sludge volume decreased.On the other hand, when the solid content concentration was 10 to 20% by weight, the raw material became sludge-like.
Above 20% by weight, the material became lumpy. The volume of these sludge and nodular raw materials decreased due to the saccharification reaction. As a result, it was found that the volume reduction rate becomes large when the solid content concentration is 10 to 20% by weight, and becomes maximum when the solid content concentration is 15% by weight. This value coincides with the limit value for the raw material bagasse to absorb water (limit water retention amount of approximately 85% by weight). These results were similar when rice straw was used as the sample. Next, the effect of fiber length (average diameter) on the volume reduction rate was investigated under the above-mentioned limit water retention conditions. The measurement results are shown in FIG. In other words, Figure 4 shows the fiber length (mm) (horizontal axis) and volume reduction rate (%) (vertical axis).
It is a graph showing the relationship between As is clear from Figure 4, the fiber length is 0.1 mm.
The above is effective. Incidentally, when the fiber length is 2 mm or more, the volume reduction rate improves, but since the volume at the time of preparation increases, this is not necessarily effective. Further, when the fiber length is 0.06 mm or less, there is almost no decrease in volume. This is because although free water was formed, it was not eluted through the screen due to clogging. In view of the above, the fiber length is preferably about 0.1 mm to 2.0 mm. The above explanation was based on the case where a screen was installed at the bottom of the saccharification tank in order to study free aqueous liquid, but in reality, as will be explained later, the presence or absence of a screen will affect the saccharification reaction itself. I confirmed that there is no. As described above, according to the saccharification method of the present invention, continuous saccharification could be performed at a high concentration and at a high charging rate even with a cellulosic material that is not pulverized. Embodiments of the apparatus of the present invention will be specifically described below based on the accompanying drawings. FIG. 5 is a partially cross-sectional schematic diagram showing an example of the device of the present invention. In Fig. 5, numeral 1 is a cellulose-based material, 2 is an enzyme aqueous solution, 3 is a mixing tank, 3a is a hopper, 3b is a stirring motor, 3c is a ribbon blade, 3d is an alkali or acid aqueous solution, 3e is a pump, and 3f is a pump , 4 is screw feeder,
4a is a screw blade, 5 is a saccharification tank, 5a is a hot water jacket, 6 is an excluder, 6a is a screw blade, 6b is a drain port, 7 is a screen, 7a is a free aqueous liquid, 8 is a centrifugal separator, 8a is a residue Cellulose-based material, 8b means separation liquid. The apparatus shown in FIG. 5 consists of four steps: mixing, transportation, saccharification, and solid-liquid separation. (1) Mixing process Cellulose-based material 1, which is a raw material, is supplied to the mixing tank 3 from the hopper 3a. Then, an aqueous enzyme solution 2 such as cellulase or hemicellulase is supplied by the pump 3f. The mixture is then mixed by the rotation of the ribbon blade 3c installed on the rotating shaft of the stirring motor 3b. The amount of water at this time is set to the above-mentioned limit retention amount of water, and this value may be determined in advance and the calculated amount may be added. The pH of the mixture is adjusted by supplying an aqueous alkali or acid solution 3d via a pump 3e. The amount added at this time is measured in advance.
Note that the structure of the mixing tank is not particularly limited. (2) Transportation process The mixture was transported to the saccharification tank 5 using a screw feeder 4. The supply amount of the mixture was controlled by the rotation speed and operating time of the screw blade 4a. (3) Saccharification process The saccharification tank 5 is a tower-shaped packed tank, and a hot water jacket 5a is installed on the tank wall. The bottom of the tank is connected to a cross-feeding excluder 6.
A net 7 is installed below the excluder 6 on the downward extension of the saccharification tank 5. The diameter of the net 7 may be any diameter as long as it can hold the cellulose material 1. The mixture supplied from the screw feeder 4 forms a packed layer in the saccharification tank 5. Then, the water is heated to the optimum temperature for the fibrinolytic enzyme by the hot water jacket 5a.
When the saccharification tank is enlarged, heat transfer tubes may be installed in the packed layer so as not to interfere with the descent of the mixture. The free aqueous liquid produced by the saccharification reaction was extracted from the mixture using a net 7, and the cellulose-based material 1 near the bottom of the tank was extracted using an excluder 6. The extraction amount at this time was controlled by the rotational speed of the screw blade 6a of the excluder 6 and the operating time. The operations of the screw feeder 4 and excluder 6 are related, and after the excluder 6 finishes operating, the screw feeder 4 starts operating. The operating time for each can be determined based on the transport capacity of each by checking in advance the volume reduction of the mixture during the set residence time in the saccharification tank. That is, the operating times of the screw feeder 4 and excluder 6 are calculated using the following formulas: Δt ₆ =V _F /Q _F …(1) Δt ₇ =V _EX /Q _EX …(2) V _EX /V _F = α ... (3) α = f (T) ... (4) [In each of the above formulas, Δt ₆ : Operating time of the screw feeder (minutes), Δt ₇ : Operating time of the excluder (min), Q _F : Transport speed of screw feeder (m ³ / min), Q _EX : Transport speed of excluder (m ³ / min), V _F : Volume of feed mixture (m ³ ), V _EX : Volume of extracted material (m ³ ), α:
Reduction rate of mixture volume due to saccharification, T: Residence time of mixture (hours)] (4) Solid-liquid separation step The mixture extracted by excluder 6 (consisting of residual cellulose substance 8a and aqueous solution containing sugar and enzymes) is , solid-liquid separation is performed using a centrifuge 8. The separated liquid 8b and the free aqueous liquid 7a are combined to form a sugar solution. Note that the solid-liquid separation method is not particularly limited to the above centrifugation method. Next, FIG. 6 is a partially cross-sectional schematic diagram showing another example of the apparatus of the present invention. Each symbol in FIG. 6 has the same meaning as in FIG. 5, and the difference from FIG. 5 is that the net 7 is not installed at the bottom of the saccharification tank 5. The process in the apparatus shown in FIG. 6 consists of four steps as in FIG. 5, and the mixing step and transport step are the same as in FIG. The steps after the saccharification step are performed as follows. so that the mixture of the cellulose material 1 and the fibrinolytic enzyme aqueous solution supplied from the screw feeder 4 can form a packed layer in the saccharification tank 5.
Adjust the operation of screw feeder 4 and excluder 6. An aqueous solution containing sugar is formed from this packed layer through a saccharification reaction. This aqueous solution is
It descends inside the filling tank and reaches the bottom of the tank. and,
It passes through the excluder 6 and is extracted out of the tank system from its outlet 6b. This aqueous solution and excluder 6
The cellulose-based substance 1 extracted by the method is subjected to a centrifuge 8 to separate the residual cellulose-based substance 8a and a separated liquid 8b containing sugar. At this time, depending on the fiber length of the cellulose-based material 1, the aqueous solution produced by the saccharification reaction may not descend within the filled layer and may stay in the upper part. If a new raw material is supplied and saccharification is performed in this state, the amount of aqueous solution will increase, so it is sufficient to provide an outlet in the upper part of the saccharification tank 5 and draw out the aqueous solution to keep the liquid level constant. Note that since the extracted liquid contains sugar, it is collected together with the above-mentioned separated liquid 8b. [Examples of the Invention] Hereinafter, the present invention will be explained in more detail with reference to Examples and Comparative Examples, but the present invention is not limited to these Examples. Example 1 Using ozone-treated bagasse as a raw material, continuous saccharification was performed using the apparatus shown in FIG. Note that the ozone treatment method was the method described in Japanese Patent Application Laid-Open No. 57-29293. The saccharification tank has an inner diameter of 20 cm and a height of 65 cm. The retained water percentage of the mixture of ozonated bagasse (average fiber length 0.7 mm, cellulose content 40%) and Cellulase Onozuka R-10 containing aqueous solution was 85% by weight. This mixture was supplied at a rate of 1 kg/hour, the residence time of this mixture section in the tank was about 24 hours, and the temperature in the tank was controlled at 50°C. As a result, 48 hours after the start of the reaction, the glucose concentration in the free aqueous liquid and the aqueous solution contained in the extracted bagasse was 44 g/hour, and the combined amount of sugar solution was 0.65 per unit time. Comparative Example 1 Using the same bagasse as in Example 1, a continuous saccharification reaction was carried out in a mechanical stirring tank (with ribbon type blades) having an internal capacity of 50 ml. A mixture of ozonated bagasse and cellulase aqueous solution whose pH was adjusted to 4.8 in advance was added to
Kg/hour was fed into the mechanically stirred tank, and the residence time was 48 hours. The temperature inside the tank was 50°C. As a result, the glucose concentration in the effluent is approximately
It was 42g/. Furthermore, when the residence time was 24 hours, the sugar concentration of the effluent was approximately 24 g/g/. Therefore, at a feed rate of 1 kg/hour of the mixture, approximately
In order to obtain a sugar solution with a glucose concentration of about 42 g/g, a residence time of 48 hours and a sugar volume of about 50 g were required. As is clear from Comparative Example 1 above, it was found that according to the present invention, the tank volume could be reduced to about 2/5 of that of the conventional method. Example 2 An experiment similar to Example 1 was conducted using alkali-treated bagasse. Alkali-treated bagasse was prepared as follows. A 0.5% by weight aqueous solution of caustic soda and bagasse (average fiber length 0.7mm) were mixed at a mixing ratio of 20:
1 and heated at 100°C for 30 minutes.
After collecting the bagasse, it was washed and dried. This alkali-treated bagasse and an aqueous cellulase solution were mixed. The mixing ratio was 3:17. Other saccharification conditions were the same as in Example 1. After the reaction stabilizes, the glucose concentration of the free aqueous liquid and the aqueous solution contained in the withdrawal mixture is approximately 68 g/g/g.
It was hot. Note that the sugar concentration is higher here than in Example 1 because the alkali-treated bagasse has a higher cellulose content. On the other hand, as in Comparative Example 1, a saccharification experiment was conducted in a mechanical stirring tank (with ribbon type blades). As a result, in order to obtain a sugar solution having the sugar concentration described in Example 2, the tank volume was required to be about 50, as in Example 1. As is clear from this, it has been found that the tank volume can be reduced by the present invention even when alkali-treated bagasse is used. Example 3 An experiment similar to Example 1 was conducted using rice straw, corn cob, and wood chips as cellulosic materials. The pretreatment method was alkaline treatment, and the treatment conditions were the same as in Example 2. The length of each fiber is
The average diameter was 0.8 mm. The results are shown in Table 1 below.

〔Effect of the invention〕

As explained in detail above, according to the present invention,
Since saccharification can be performed without pulverization, energy consumption for pretreatment can be significantly reduced. In addition, the process is practical as it is continuous saccharification, and the saccharification tank requires only 1/3 to 2/5 of the volume of a conventional mechanical stirring tank, and since there is no stirring, the stirring power is zero. A remarkable effect was achieved.

[Brief explanation of drawings]

Figures 1 to 4 are graphs showing the results of various experiments conducted to determine the operating conditions of the present invention, and Figures 5 and 6 are partial cross-sectional schematic diagrams showing an example of the apparatus of the present invention. It is a diagram. 1... Cellulose-based material, 2... Enzyme aqueous solution, 3...
Mixing tank, 4... Screw feeder, 5... Saccharification tank, 6... Excluder, 7... Net, 8... Centrifugal separator.

Claims (1)

[Scope of Claims] 1. A saccharification method for obtaining a sugar solution by decomposing a cellulose substance with a fibrinolytic enzyme, which comprises: (1) mixing a cellulose substance and a fibrinolytic enzyme via water; (2) ) supplying the mixture to a tower-type saccharification tank and extracting the product mixture from the bottom of the tank after residence for a predetermined time; and (3) separating the product mixture into solid and liquid and recovering the sugar solution. A method for saccharification of cellulosic substances characterized by encapsulation. 2. The method for saccharifying a cellulosic material according to claim 1, wherein the water content in the mixture containing the cellulosic material, fibrinolytic enzyme, and water is the maximum amount of water that can retain the cellulosic material. 3. The method for saccharifying a cellulosic material according to claim 1, wherein the mixture containing the cellulosic material, fibrinolytic enzyme, and water has a water content of 80 to 90% by weight. 4. In a saccharification device that obtains a sugar solution by decomposing a cellulose material with a fibrinolytic enzyme, means for mixing the cellulosic material raw material and an aqueous solution containing a treatment agent containing a fibrinolytic enzyme, and charging the mixture into the saccharification tank. 1. An apparatus for saccharification of cellulosic substances, comprising: a jacketed tower-type saccharification tank; a means for extracting a product mixture at the bottom of the saccharification tank; and a solid-liquid separation means. 5. A patent claim in which the solid-liquid separation means comprises a means for separating a free aqueous liquid from a product mixture, a means for separating solid-liquid from a residue, and a means for mixing the separated liquid with the free aqueous liquid and recovering it as a sugar solution. The apparatus for saccharification of cellulosic substances according to item 4.