JPH04341174A - Production of useful substance by product-inhibiting reaction or reversible reaction and its system - Google Patents

Abstract

PURPOSE:To continuously carry out production with high productivity by using reaction for producing a useful substance so as to contain the useful substance in a reaction solution containing three or more ingredients present therein, according to product-inhibitory reaction or reversible reaction. CONSTITUTION:A step for producing a reaction solution containing three or more ingredients present therein is combined with a step for chromatographically separating the ingredients in the aforementioned reaction solution into the three portions and the portion containing the raw material among the separated portions is then returned to the reaction process. Dextran and fructose are produced by enzymic reaction using sucrose as a raw material and the fructose and dextran are recovered in a pseudo-moving bed. The portion containing the sucrose and dextran is then returned to the reaction process. The aforementioned operations are continuously preformed.

Description

[Detailed description of the invention]

0001

[Industrial Application Field] The present invention relates to a method and apparatus for efficiently producing useful substances through a product inhibition reaction or a reversible reaction, typically combining an enzymatic reaction for producing dextran from sucrose with a chromatographic separation operation. The present invention relates to a method and apparatus for producing useful substances continuously and with high productivity.

0002

BACKGROUND OF THE INVENTION The technology to which the present invention is directed will be explained below, taking as a typical example the case of producing dextran by enzymatic reaction.

[0003] Dextran is a polysaccharide composed of D-glucose with a molecular weight of about 1 million, and is known as a substance synthesized from sucrose by an extracellular enzyme (dextransucrase) produced by lactic acid bacteria. It is widely used medically as a blood expander and plasma substitute.

[0004] This dextran is a viscous substance that is synthesized using sucrose as a substrate by various bacteria or dextransucrase obtained from it, as mentioned above, through the reaction of (sucrose)n → dextran + fructose. It was produced by carrying out this reaction in a medium, filtering the resulting culture solution, adding alcohol, and purifying the resulting precipitate.

[0005] Such conventional production methods are batch-type methods that require the steps of culturing, filtration, precipitation, and purification, and therefore cannot be produced continuously, and there is a limit to productivity improvement. In addition, the above enzymatic reaction is a product-inhibiting reaction in which the byproduct fructose inhibits the enzymatic reaction, so it is said that the concentration of dextran in the culture solution collected cannot exceed a certain level. There were also problems in that it was difficult to achieve high productivity.

By the way, although this does not concern a multicomponent system containing three or more components, such as the production of dextran and fructose from sucrose, it concerns the reaction of producing isomerized fructose from glucose. A report that combines a type of chromatographic separation device to perform continuous reaction and separation and improve the conversion rate of the reaction (Mr. Hashimoto, Mr. Shirai, Mr. Higuchi: Society of Chemical Engineers of Japan Yamaguchi Conference,
There is a collection of lecture abstracts p217; (1990)). This report describes a device in which enzyme reactors are installed in series and alternating with separation towers within a simulated moving bed system, which has the advantage of enabling continuous production compared to conventional batch systems. Are better.

[0007] Therefore, when producing a mixed solution containing three or more components including dextran as described above, it may be possible to apply the method of the same report to improve productivity. The same method cannot be simply applied as is. The reason is that the method in the same report is related to a method in which one reaction product is produced from one raw material component, and as mentioned above, raw material components and products are produced from a mixed liquid containing three or more components. This is because it cannot be simply applied to an operation for separating components, and there has been no known technology that can separate three or more components by combining it with a reactor.

[0008] In addition, in the case of product inhibition reactions or reversible reactions such as catalytic reactions and enzymatic reactions, there are many reactions in which equilibrium occurs without the reaction proceeding beyond a certain level due to product inhibition. This is a major constraint on improving productivity, and many studies have traditionally focused on biasing the reaction equilibrium toward the product side to increase the conversion rate and improve productivity. There is also a problem in that it is not easy to carry out such operations, such as operations for continuously separating the product from the reaction system, using industrial equipment and equipment. In addition, for the effective use of raw materials,
It has also been a common practice to separate the product from the reaction solution and reuse the remaining raw material.

However, none of the various countermeasures described above has been studied for multi-component liquids containing products and raw materials as three or more components, and conventional methods cannot be applied to such multi-component liquids. It is often extremely difficult to apply it to component systems, and at best, it is considered to improve the efficiency of raw material utilization by reusing the raw material components as raw materials after separating the product from the mixed liquid in batch-type processing. The current situation was to the extent that

[0010] Furthermore, the conversion rate from raw materials to products as described above does not proceed beyond a certain level, and moreover, there is the difficult problem that the target substance must be separated from a multicomponent liquid containing three or more components. Similar problems regarding the reaction system involved are not limited to the case of producing the above-mentioned dextran, but are exemplified in various other ways. For example, cyclodextrin (C
(abbreviated as D) is generally well known as a substance used in food additives because it clathrates hydrophobic substances and releases them under different conditions, and its production is generally carried out by C.
Although this reaction is carried out using GTase, C
Since equilibrium is reached when D is around 50%, there is a problem similar to that described above. That is, the batch-type operation that has been conventionally adopted as a manufacturing method has low raw material utilization efficiency. For this reason, saccharified starch is separated and recovered from the reaction solution using a UF membrane, etc., but this method allows the raw materials to be reused, but it is not possible to continuously produce α-CD, β-CD, and γ-CD. Since this is not possible, essential economic improvements cannot be achieved.

[0011] In addition, a method of producing fructose and glucose by acid decomposition by contacting sucrose with mineral acids such as hydrochloric acid or sulfuric acid or a strongly acidic cation exchange resin is also commonly used, and the resulting product is an invert sugar. It has traditionally been used as a raw material for producing oligosaccharides by separating fructose and glucose, or for medical purposes. However, this acid decomposition reaction generally does not proceed to more than about 60%.

0012

[Problems to be Solved by the Invention] As described above, when producing useful substances by product inhibition reactions and reversible reactions such as enzymatic reactions and catalytic reactions, in a system where the reaction in which raw materials are converted to products reaches equilibrium, It has been desired to solve the above problems and realize efficient and economical manufacturing. In particular, when the reaction solution contains three or more components, a new method has been sought since there have been no previous proposals.

[0013] From the various viewpoints mentioned above, the present inventor has conducted various studies and found that when a useful substance is produced by a product inhibition reaction or a reversible reaction in which the reaction solution contains three or more components including raw materials. Therefore, the present invention was developed with the aim of providing a method that can realize continuous and highly economical manufacturing.

Another object of the present invention is to provide a method for efficiently producing a target substance using a compact device.

Other more specific objects of the present invention are as follows. In other words, for example, when dextran is produced in a batch process using the conventional method, alcohol is added to a mixed solution containing the raw material sucrose and the products dextran and fructose to precipitate and recover the dextran, so the remaining solution is It contains alcohol along with sucrose and fructose. Therefore, alcohol separation is required to recover each component from the remaining liquid, and recovery of the raw material sucrose is time-consuming. For this reason, it is generally used as a sweetener separately or discarded without performing a recovery operation, but the present invention further improves the economic efficiency of producing the target substance by efficiently separating these components. The purpose of the present invention is to provide a method for producing a new useful substance that can be produced using the following methods.

[0016]

[Means and Actions for Solving the Problems] In order to achieve the above-mentioned various objects, the present inventor has proposed a method in which three or more components of raw materials and reaction products are mixed together by a product inhibition reaction or a reversible reaction. a first step of producing a mixed solution, a second step of transferring this mixed solution to a chromatographic separation device, and a third step of chromatographically separating the transferred mixed solution into three or more sections.
Production of a useful substance by a product inhibition reaction or a reversible reaction, characterized in that the step of step 1 and the fourth step of returning the separated section containing the raw material components to the first step are carried out in succession. completed the method.

[0017] The apparatus used to carry out such a method includes a reactor which receives a stock solution and causes a reaction using a catalyst or an enzyme supported on an internal immobilized carrier, and a reactor which receives a stock solution and performs a reaction using a catalyst or an enzyme supported on an internal immobilized carrier. A simulated moving bed device consisting of a plurality of columns to which a mixed liquid of raw material components and products is sent and chromatographically separates each component, a liquid transfer system that sends a reaction liquid from a reactor to a simulated moving bed device, and a simulated moving bed. A typical example is a device that combines a liquid transfer system that returns components including the stock solution separated in the device to the reactor, and more specifically, a device that combines three or more raw materials and reaction products by a product inhibition reaction or a reversible reaction. It consists of a reactor that produces a mixed solution containing a mixture of components, and a plurality of adsorbent-packed towers forming an endless circulation system, and contains the components contained in the mixed solution that is continuously supplied to any of the adsorbent-packed towers. A group of adsorbent packed towers that sequentially separate and form zones enriched with each of the above components along the flow direction of the liquid, and from the adsorbent packed tower upstream of the mixed liquid supply position, a first extracting means for extracting the first section of the mixed liquid, a second extracting means for extracting the separated second section from the adsorbent packed tower downstream of the mixed liquid supply position; A separated third
a simulated moving bed device equipped with a third extraction means for extracting the segment; and a mixed liquid transfer means for selecting any adsorbent packed tower of the simulated moving bed device from the reactor and supplying the mixed liquid. , means for returning to the reactor the liquid of the enriched fraction of the raw material extracted from the first to third extraction means of the simulated moving bed device, and The position at which the mixed liquid is transferred and the position at which each section of liquid is extracted by the first to third extraction means are synchronized with respect to the group of a plurality of adsorbent packed towers formed as the endless circulation system. An example of an apparatus for producing a useful substance by a product inhibition reaction or a reversible reaction is characterized in that it has a control means for moving the mixed liquid to the downstream side of the flow over time.

The above-mentioned simulated moving bed device is not particularly limited as long as it can be combined with a reactor and three or more components can be separated and recovered. A layer device or a device based on the pseudo moving bed device (Japanese Patent Application Laid-open No. 1982-91205) applied to the “Specific Component Separation Method” proposed by the present applicant, as well as “ A suitable example is a pseudo moving bed device based on the Japanese Patent Application No. 2-402826 entitled "Multicomponent Separation Method and Apparatus". Of these, the latter "multi-component separation device" uses a large number of unit packed beds filled with adsorbents to form an endless series circulation channel, and this circulation channel is used for circulating and blocking. This is a system in which a raw material fluid containing three or more components having different affinities for the adsorbent is passed through the plurality of unit packed beds, so that components with a weaker affinity for the adsorbent are replaced by a component with a stronger affinity for the adsorbent. For a system that forms adsorption zones that are sequentially divided into components, the circulation of the system is blocked at a position upstream of the adsorption zone where components with weak affinity are formed, and the circulation of the system is blocked at a position downstream of this blockage. a step of supplying the raw material fluid and extracting from the system a predetermined component among the components forming the adsorption zone upstream of the cutoff position; and releasing the cutoff and supplying the raw material fluid. While circulating the above system, each component remaining in the above process is removed.
The steps of supplying the desorbent fluid and withdrawing each separately according to a two-component simulated moving bed method are repeated as one cycle, or if necessary, the latter step can be followed by removing the fluid in the system. While circulating, the desorbent fluid is supplied to each adsorption zone of each component, which is divided into adsorption zones from weak to strong affinity in the circulation direction, and each component is extracted. This device is used in a method in which the supply and extraction positions of fractions are sequentially shifted to the downstream side of the circulating flow in accordance with the movement of the adsorption zone. for example,
An endless serial circulation flow path formed using a large number of unit packed tanks filled with adsorbent, one cutoff valve that can be opened and closed provided in the middle of this circulation flow, and the downstream position of this cutoff valve. a raw material fluid supply means connected to the circulation flow path at a position upstream of the cutoff valve, a fluid extraction means connected to the circulation flow path at a position upstream of the cutoff valve, and a desorbent fluid supply means connected to each adjacent space of the unit packed bed. An example of this is a device having a configuration including a device and a fluid extraction means, and the device of the present invention can be constructed by connecting a reactor to the device to realize the method of the present invention. Further, in the method of the present invention, it is also possible to use, for example, a chromatographic separation apparatus based on JP-A-63-158105.

[0019] In carrying out the method of the present invention, it is particularly preferable that in the first step, before the reaction in which the product is produced from the raw material components by an enzymatic reaction or the like reaches equilibrium, the conversion of the raw material components to the product is preferably carried out. The conversion rate is 1/10 to 8/10, optimally 2/10 to 6/1 of the conversion rate when equilibrium is reached.
It is preferable to move the reaction solution to the second step at a stage of about 0. Such operations can be performed as appropriate by adjusting the residence time in the reactor depending on the flow rate setting of the raw material fluid, the shape and size of the reactor, etc. By doing this, high reaction rates can be achieved. In addition to allowing the reaction to occur and increasing the amount of product produced per unit time, it is also possible to effectively utilize the enzyme activity that deactivates over time in enzymatic reactions. There is an advantage that the reactor can be made small.

[0020] In the fourth step, it is also preferable that the liquid containing the raw material components is concentrated using, for example, an ultrafiltration membrane device and then returned to the first step.

[0021]

Effects of the Invention According to the method of the present invention, useful substances produced by a product inhibition reaction or reversible reaction system in which three or more components are contained in a mixed solution can be produced by combining the production reaction and separation operation. Since it is possible to do this continuously, the target substance can be efficiently produced and separated and recovered.

[0022] Furthermore, by enriching the raw material components in the reaction solution and returning them to the reactor, the raw material components can be used extremely effectively without waste, and continuous production as described above is possible. In combination with this, highly economical production can be carried out. Especially when it is difficult to separate the target substance and raw material components,
In a general simulated moving bed, it is necessary to increase the length of the column filled with adsorbent in order to separate these mixed parts, but in the device of the present invention, this mixed part is separated into a reactor. This has the advantage that even if the column length is shortened, the yield will not be affected.

In addition, in the operation of the method of the present invention, by transferring the raw material to the chromatographic separation device before the reaction of converting the raw material to the product reaches equilibrium, especially at a stage where the conversion rate is sufficiently low, generally a high reaction rate can be achieved. In addition to making it possible to carry out the reaction at a high speed and maintaining a high amount of product produced per unit time, for example, in the case of enzymatic reactions, it is possible to effectively utilize the activity of enzymes that deactivate over time. There are excellent effects such as the ability to downsize the equipment, especially the reactor, required to ensure a fixed production amount.

Furthermore, if a concentration means such as an ultrafiltration membrane device is provided in the process of returning the raw material-enriched liquid from the chromatographic separation device to the reactor, the recovery rate of the fraction containing the raw material will be reduced. Even if the concentration is increased, the concentration of the liquid returned to the reactor can be appropriately controlled, so the amount of raw material liquid fed to the reactor is not limited, and the reaction can be carried out with high efficiency.

[0025]

EXAMPLES The present invention will be explained in detail below based on Examples, but it is obvious that the present invention is not limited to these Examples.

Example 1 Preparation of immobilized enzyme Leuconostoc mesenteroid
s in a sucrose-based medium at 25°C for 24 hours.
After culturing for an hour, the medium supernatant was dialyzed using an ultrafiltration membrane with a molecular weight cutoff of 10,000 and concentrated under reduced pressure. This solution was added to a 2.5% hydroxylpropyl methyl cellulose acetate succinate solution, carbodiimide as a crosslinking agent was gradually added, the pH was adjusted to 5.5, and the mixture was reacted for 6 hours. Thereafter, the pH was lowered to 4.0, and the immobilized enzyme was precipitated and collected. When this immobilized enzyme was dissolved in a sucrose solution adjusted to pH 5.5 and the activity was measured from the amount of fructose produced, it was confirmed that the enzyme had an enzyme activity of 0.5 u/g of carrier.

Dextran Production Apparatus The above-mentioned immobilized enzyme used in this example dissolves and precipitates depending on the pH value, and exhibits enzyme activity in the dissolved state. For this reason, the production apparatus of this example is configured by a combination of a reactor whose basic configuration is a main reactor 1, a pH adjuster 2, and a separator 3, and a pseudo moving bed device.

The reactor of this example using immobilized enzyme is shown in FIG.
As shown in the figure, it consists of a main reactor 1, a pH regulator 2, and a separator 3.
The regulator and separator each have a capacity of 50 ml.

Reference numeral 4 denotes a stock solution tank filled with sucrose, and the stock solution is supplied to the main reactor 1 by a stock solution feed pump 5. 6 is an acid tank,
7 is an alkali tank, in which acid or alkali is sent to the main reactor 1 through a pump 9 from a feed line 8 having an on-off valve in each of the branch routes, so as to adjust the pH inside the main reactor 1. It has become. 10 is a temperature sensor, 11
is a pH sensor, 12 is a stirring device, 13 is a heater,
Each reactor is attached to the main reactor 1 so that the reaction conditions within the main reactor 1 can be maintained at optimum conditions.

The stock solution supplied to the main reactor 1 produces dextran and fructose from sucrose by enzyme activity, and the reaction solution (produced mixed solution) containing these is sent to the pH regulator 2 by the peristaltic pump 14. Acid tank 1
The immobilized enzyme precipitated in the lower separator 3 returns to the main reactor 1, and the reaction solution is sent to the reaction solution storage tank 16 in an overflow manner. Note that 17 is a pH sensor.

The reaction solution obtained as described above is sent from the reaction solution storage tank 16 to the simulated moving bed device via the feed line 19 by the feed pump 18.

Next, the simulated moving bed separation device of this example will be explained. The simulated moving bed separation device of this example has a diameter of 12 mmφ.
It consists of a column group in which 12 columns 31 to 42 of 00 mmH are directly connected in an endless series, and each column is coated with Amberlite CG-6000, a strongly acidic cation exchange resin.
(Product name: D.V.B. 6%, average diameter 0.25 mm) is filled in 270 ml each. 50 is a circulation pump for this endless series of columns.

This simulated moving bed device is connected so that the reaction liquid is transferred from the reaction liquid storage tank 16 to a predetermined column by a feed pump 18, and is also connected to a line for extracting fructose from the column on the upstream side. 51, a line 52 for extracting a mixed section of dextran and sucrose from a downstream column, a line 53 for extracting dextran from a further downstream column, and a line 54 for introducing an eluent are connected as shown in the figure, and these positions While maintaining the relationship, the connection position is shifted to the downstream side over time by a control device (not shown). Therefore, at the upstream and downstream ends of each column, the lines 19,
Opening/closing valves and the like are provided for switching connections to 51 to 54, but these are the same as in the case of a normal pseudo moving bed device, and are not shown because they would complicate the diagram.

A line 52 for extracting the liquid from the dextran and sucrose mixing section is connected to return the liquid to the stock solution tank 4. Note that a concentrating means 55 such as a membrane device may be provided in the middle of this line 52 as shown in the figure.

Production of Dextran The following production of dextran was carried out using the apparatus shown in FIG.

Dissolve 72 g of sucrose in pure water to make 1 liter, and add 0.1N hydrochloric acid to this solution to adjust the pH to 5.
．． It was adjusted to 3-5.4. Add 100 g of the above immobilized enzyme to about 100 ml of this solution, and add it to the main reactor 1 equipped with a stirrer.
The temperature was maintained at 45°C, and the pH was adjusted to 5.3-5.
The reaction was carried out for 5 hours while stirring at a temperature of 4.

10 ml of this reaction solution was pumped into peristaltic pump 1.
4 and introduced into pH adjuster 2. In the pH adjuster 2, a 0.1N hydrochloric acid solution was added while stirring with N2 gas to adjust the pH to 3.8 to 3.9. By lowering the pH in this manner, the immobilized enzyme in the solution was coagulated and precipitated. This solution was then introduced into separator 3 and the precipitated immobilized enzyme was returned to main reactor 1 for continuous processing. On the other hand, the reaction liquid flows into the reaction liquid storage tank 16 due to overflow.
Moved to. The above operation was performed by placing 50 ml of sucrose solution in the pH adjuster 2 in advance.

The solution (reaction liquid) thus obtained contains 0.01 mol of fructose (therefore, 0.01 mol of dextran).
01 mol), and unreacted sucrose to about 0.01 mol).
It contained 19 moles. This reaction solution was supplied to a simulated moving bed apparatus at a flow rate of 8.4 ml/hr.

In the simulated moving bed device, the standard flow rate after the extraction position (downstream) of the dextran section is adjusted to 50°C while moving the liquid supply and extraction ports to the downstream side every 5 minutes.
ml/hr, reaction solution supply flow rate 8.4 ml/hr, eluent supply flow rate 42 ml/hr, fructose fraction withdrawal flow rate 20 ml/hr, dextran division withdrawal flow rate 16 ml/hr.
．． The operation was carried out at a rate of 1 ml/hr and a discharge flow rate of 14.3 ml/hr from the sucrose and dextran mixing section.

The liquid in the sucrose and dextran mixed section extracted above was concentrated to about 3 times (11 ml) using a flat membrane RO membrane device, and then added to the sucrose solution (about 9 ml).
The solution was supplied to the main reactor 1 in a mixed form at a rate of 0.2 ml/hr (0 g/liter), and the difference in the reaction state from when the stock solution was supplied was investigated. Note that the liquid after the reaction was continuously introduced into the pH adjuster 2 and the separator 3.

As a result, the fructose concentration of the reaction solution flowing out from the separator 3 gradually increased to 0.01 mol after about 5 hours, and the concentration of unreacted sucrose to 0.24 mol.

The results of simulating the above material balance relationship are shown in Table 1 below, and the actual results also agreed well with this as described above.

[0043]

[Table 1]

Next, the reaction solution obtained in the above reaction was supplied to a simulated moving bed apparatus at a flow rate of 14.1 ml/hr.
ml/hr, the withdrawal flow rate of the fructose section is 25 ml/hr, the withdrawal flow rate of the dextran and sucrose mixed section is 25 ml/hr.
．． 4 ml/hr, the withdrawal flow rate of the dextran section was 28.
Separation was possible by changing the conditions to 7 ml/hr. In addition, the liquid in the dextran and sucrose mixing section is 1
It was concentrated using an RO membrane with a diameter of 0 cm.

Next, the liquid in the dextran and sucrose mixed section and 0.32 ml of the sucrose stock solution were mixed and supplied to the main reactor 1, but the fructose concentration produced was 0.
．． At 0.025 mol, there was almost no change from the above case. Therefore, it was confirmed that it is possible to produce dextran while performing the above-mentioned return and to perform the operations of separating fructose and dextran continuously.

[0046] When the results of the above operation are compared with those of the operation without separation, the dextran production capacity is about 1.7 times, which confirms the excellent effect of the present invention.

The results of simulating the above material balance relationship are shown in Table 2 below, and as mentioned above, the actual results also match well.

[0048]

[Table 2]

Example 2 Preparation of immobilized enzyme Bacillus macerans (IFO 3)
490) was cultured with shaking at 40° C. for 2 days in a potato-calcium carbonate medium supplemented with ammonium sulfate, and the supernatant after centrifugal precipitation was used as the crude enzyme.

After adding 1 liter of ethyl alcohol to 2 liters of this enzyme solution, next 50 g of cornstarch,
50g of Celite was added and stirred to adsorb starch. 3
After washing with 3% ethyl alcohol solution, 20 ml of 40°C
It was desorbed with deionized water heated to . The resulting colorless and transparent liquid was dissolved at pH 6.20 and 1 containing 5 x 10-4M Ca++.
Dialyzed in 0mM acetate buffer by collodion bag, concentrated, and 0-0.5M by DEAE-cellulose column.
Gradient elution was performed with NaCl solution. It was further concentrated using a collodion bag to obtain purified enzyme. Specific activity was 10 IU/mg.

[0051] This enzyme is CGTase, and FE-4
611 (manufactured by Organo) was adsorbed as a carrier. Note that FE-4611 is a carrier in which an amine group is introduced by reacting glycidyl methacrylate with dimethylaminoethanol, the epoxy ring is opened, an amino group is introduced, and an alcohol hydroxyl group is generated.

[0052] The carrier is packed into a column, regenerated with an alkaline solution, washed with an acetic acid/sodium acetate buffer solution having a pH of around 6, and then the enzyme solution obtained in the above operation is passed downflow for a contact time of 0.5 to 1. Contacted in time. This carrier is 1.0
mg of protein was adsorbed.

Cyclodextrin Production Apparatus The simulated moving bed apparatus of this example was constructed as shown in FIG. 2 in accordance with the description in Japanese Patent Application No. 2-402826.

That is, the symbols 101 to 112 in FIG.
Strong acidic cation exchange resin Amberlite IR-1016
(Product name: D.V.B. 4%; manufactured by Organo)
It is a unit packed column (column) filled with 50 ml, and each of these first to twelfth columns are connected in an endless series by piping so that liquid can flow therethrough, and a circulation pump 120 in the system It is designed to allow fluid to flow through it. B is a cutoff valve interposed between the twelfth column 112 and the first column 101 in this circulation path, and is controlled to open and close by a control means (not shown) to put the path into a circulation state or into a cutoff state. It is provided so that it can be switched.

[0055] A reaction liquid feed line 122 is connected to the first column 101 located on the downstream side of this shutoff valve B, through which a reaction liquid is introduced from its upstream end via an on-off valve A.
Further, a pure water feed line 124 that introduces an eluent (pure water) from the upper end of the seventh column 107 via an on-off valve 123 is connected to the twelfth column 112 located upstream of the shutoff valve B. A withdrawal line 126 for withdrawing the liquid of the section forming the enriched zone in the twelfth column is connected from the end via an on-off valve C. In addition, the above-mentioned first tower to first tower
Each of the two columns 101 to 112 has a line for introducing the eluent through an on-off valve (not shown), and a section forming an enriched zone within each column (which may also be a return liquid section). )
The extraction line for extracting the liquid through the on-off valve (also not shown) is connected in the same way as above, but since it would make the diagram complicated if all of these lines were shown, the lines are shown with broken lines so that the others are This is omitted, and the specific flow relationships are specifically described in the following production examples.

On the other hand, the reactor is a main reactor (column) 128 filled with 10 ml of the above-mentioned immobilized enzyme carrier, after which the stock solution (saccharified starch) is supplied from the stock solution tank 129 with a pump 130 and retained therein for a predetermined time. , and was configured to be stored in a reaction liquid storage tank 131. The liquid in this reaction liquid storage tank 131 was transferred to a simulated moving bed device by a feed pump 132. In this main reactor 128, 10 ml of the above-mentioned immobilized enzyme carrier was packed into a column, and 4% liquefied starch was added at a temperature of 50°C.
When the liquid is passed at a flow rate of SV 0.1 and reacted for a long time, cyclodextrin (CD) is produced, and the total CD is 45 to 50%, of which α-CD is about 16% and β-
Approximately 26% of CD and 6% of γ-CD were generated (when the liquid flow rate was set to SV 0.5, the total CD was 30~30%).
35%, of which α-CD is about 12% and β-CD is about 1
8%, and γ-CD was produced at 2%).

[0057] Each tower 101 to 112 of the pseudo moving bed device
In this example, a return line 133 is provided to return the liquid in the section containing the raw material components to the stock solution tank 129, and a membrane device may be installed in the middle of this return line 133 as necessary. It is also preferable to provide a concentrating means 134 such as the like.

Production of Cyclodextrin In the main reactor 128 of the production apparatus constructed as above, four
% liquefied starch at a temperature of 50°C and a liquid flow rate of SV 0.1 to react for a long time, and the reaction solution was transferred to the above-mentioned simulated moving bed apparatus for 18 hours.
The liquid was passed through at a flow rate of 8 ml/hr.

At this time, first, the simulated moving bed apparatus closes the shutoff valve B, while leaving the valves A and C open, and injects the reaction liquid into the first column at a rate of 188 ml/hr, and then pours pure water into the first column. 7th tower 1
No. 17 was injected at a rate of 136 ml/hr, and the liquid in the β and γ-CD sections was extracted from the bottom of the twelfth column at a rate of 324 ml/hr, and this operation was run for 2.4 minutes (see Figure 3).

After that, the above-mentioned shutoff valve B is opened, and valves A,
C is closed and the eluent (pure water) is injected at 124 ml/hr into the eighth column 108 while circulating the liquid in the system.
The operation was carried out for 2.4 minutes by withdrawing the starch fraction liquid from the lower part of No. 04 at a rate of 76 ml/hr and at the same time withdrawing the α-CD fraction liquid from the lower part of the 10th column 110 at a rate of 48 ml/hr. Note that the circulation flow is 2 in the part from the 11th column to the 4th column.
It was set at 83 ml/hr (see Figure 4).

Next, while keeping the flow rates of each injection, withdrawal, and circulation constant, each inlet and withdrawal port are moved one column in the flow direction (that is, pure water is injected into the ninth column, and the starch division is The liquid was taken out from the fifth column, and the liquid in the α-CD section was taken out from the 11th column), and the operation was continued for 2.4 minutes. in this way,
Each inlet and outlet were operated and moved sequentially in the flow direction for 2.4 minutes each until the pure water inlet reached the seventh column.

After that, shut off valve B is closed again, and valves A and C are closed.
The reaction solution was injected at 188 ml/hr and pure water at 136 ml/hr into the 7th column, and β, γ-C was injected from the bottom of the 12th column.
2.4 while drawing out the liquid in section D at a rate of 324 ml/hr.
I drove for a minute.

[0063] Thereafter, shutoff valve B was closed and valves A and C were opened, and as in the previous operation, each inlet and outlet were sequentially switched and moved while circulating the liquid in the column.
This was repeated.

[0064] Through the operation as described above, the liquid separated into zones enriched with each component could be extracted from the column in a substantially continuous operation. However, β, γ
-CD sections were extracted every cycle.

[0065] The starch fraction liquid obtained by the above operation of the simulated moving bed device was concentrated in the UF membrane device 134 and returned to the raw material tank 129 to be mixed with the raw material.

After 10 cycles, the α-CD fraction was purified to a purity of about 85-90%, the β- and γ-CD fractions to a purity of about 80-85%, and the starch fraction to a purity of about 80%.

The obtained CD fraction was recovered as a product, and the starch fraction was returned as described above, mixed with the stock solution in the stock solution tank 129, and reused for CD production. The starch fraction mixed with this stock solution contains a small amount of CD, but when reacted with the immobilized enzyme, there is almost no difference from the treatment of the pure stock solution, and the results of separation using a simulated moving bed apparatus are also similar. It was the same.

[0068] The productivity with the above manufacturing apparatus was approximately three times as high as the production capacity when separation was not performed using the pseudo moving bed apparatus. Moreover, the separation results of the pseudo moving bed device were very stable.

Example 3 Example 2 was carried out using an H-type column (filling amount: 10 ml) packed with Amberlite IR-120 (trade name) as the main reactor.
A manufacturing apparatus was constructed, and 50% sucrose as a stock solution was passed through it at 50°C and SV2. This hydrolyzes sucrose, producing approximately 28% of each of glucose and fructose.
%, and a reaction solution containing about 44% unreacted sucrose was obtained in the reaction solution storage tank 131. Next, when the liquid was passed through SV5, about 20% each of glucose and fructose was produced.

[0070] This reaction solution was passed through a simulated moving bed apparatus and operated in substantially the same manner as in Example 2.

That is, first, the shutoff valve B of the pseudo moving bed device
is closed, while valves A and C are left open. The reaction solution is injected into the first column at a rate of 188 ml/hr, pure water is injected into the seventh column 117 at a rate of 178 ml/hr, and the reaction solution is injected into the seventh column 117 at a rate of 178 ml/hr. The glucose compartment liquid was drawn out from the bottom of the tank at a rate of 366 ml/hr, and this operation was run for 3 minutes (see Figure 5).

After that, the above-mentioned shutoff valve B is opened, and valves A,
C is closed, and the eluent (pure water) is injected at 162 ml/hr into the eighth column 108 while circulating the liquid in the system.
57ml/hr of liquid from the sucrose section from the bottom of 04
At the same time, the fructose fraction was extracted from the lower part of the 10th column 110 at a rate of 105 ml/hr, and the operation was continued for 3 minutes. The circulating flow was 283 ml/hr from the 11th column to the 4th column (see Figure 6).

Next, while keeping the flow rates of each injection, withdrawal, and circulation constant, each inlet and withdrawal port are moved one column at a time in the flow direction (that is, pure water is injected into the ninth column, and the sucrose is The liquid in the section was drawn out from the 5th column, and the liquid in the fructose section was drawn out from the 11th column.) The operation was carried out for 3 minutes. in this way,
Run each inlet and outlet in the flow direction for 3 minutes each.
The movement was repeated until the pure water inlet reached the seventh column.

After that, shut off valve B is closed again and valves A and C are closed.
The reactor was opened, the reaction solution was injected at 188 ml/hr, pure water was injected into the 7th column at 178 ml/hr, and the operation was continued for 3 minutes while extracting the glucose fraction liquid from the bottom of the 12th column at 366 ml/hr.

[0075] Thereafter, shutoff valve B was closed and valves A and C were opened, and as in the previous operation, each inlet and outlet were sequentially switched and moved while circulating the liquid in the tower.
This was repeated.

[0076] Through the operation as described above, the liquid separated into zones enriched with each component could be extracted from the column in a substantially continuous operation. However, the glucose classification was extracted every cycle.

The sucrose fraction liquid obtained by the above operation of the simulated moving bed device was concentrated in the UF membrane device 134 and returned to the raw material tank 129 to be mixed with the raw material.

[0078] After 10 cycles, the purity of the fructose category is about 80-85%, and the purity of the glucose category is about 85-9%.
0%, the sucrose fraction was purified to approximately 80% purity.

The obtained grape sugar and fructose fractions were recovered as products, and the sucrose fraction was returned, mixed with raw materials, and reused for the reaction. Sucrose, which is a mixed raw material, contains a small amount of monosaccharide, but when reacted with an ion exchange resin, the results are almost the same as when reacted with a sucrose stock solution, and the separation results with a simulated moving bed device are also the same. Met. Comparing the productivity of this example with the case without separation using a simulated moving bed device, the production capacity with immobilized enzyme is approximately 1.8
The separation result with the simulated moving bed apparatus was very stable.

[Brief explanation of the drawing]

FIG. 1 is a diagram showing an outline of the configuration of a dextran production apparatus of Example 1.

FIG. 2 is a diagram showing an outline of the configuration of a dextrin manufacturing apparatus of Example 2.

FIG. 3 is a diagram illustrating a part of the operating status of the simulated moving bed device in the manufacturing apparatus of Example 2.

FIG. 4 is a diagram illustrating another operating situation of the simulated moving bed device in the manufacturing apparatus of Example 2.

FIG. 5 is a diagram illustrating a part of the operating status of the simulated moving bed device in the manufacturing apparatus of Example 3.

FIG. 6 is a diagram illustrating another operating situation of the simulated moving bed device in the manufacturing apparatus of Example 3.

[Explanation of symbols]

1: Main reactor, 2: pH adjuster, 3: Separator, 4: Stock solution tank, 16: Reaction solution storage tank, 18: Feed pump, 1
9: Feed line, 31-42: Column, 50: Circulation pump, 51-53: Extraction line, 54: Eluent introduction line, B: Shutoff valve.

Claims (6)

[Claims] Claim 1: By a product inhibition reaction or a reversible reaction,
A first step of producing a mixed solution containing three or more components of raw materials and reaction products, a second step of transferring this mixed solution to a chromatographic separation device, and a second step of producing a mixed solution containing three or more components of raw materials and reaction products; A product inhibition reaction characterized in that a third step of chromatographic separation into fractions and a fourth step of returning the fraction containing raw material components among the separated fractions to the first step are performed continuously; A method for producing useful substances through reversible reactions. 2. In claim 1, the raw material supplied to the first step passes through the first step and moves to the second step before the reaction in which the raw material components are converted into products reaches equilibrium. A method for producing a useful substance by a product inhibition reaction or a reversible reaction. 3. A method for producing a useful substance by a product inhibition reaction or a reversible reaction according to claim 1 or 2, wherein the chromatographic separation in the third step is performed using a simulated moving bed device. [Claim 4] In any one of claims 1 to 3,
A product inhibition reaction or a reversible reaction characterized by having a step of concentrating the returned liquid during the fourth step of returning the fraction containing the raw material components chromatographically separated in the third step to the first step. A method for producing useful substances. [Claim 5] In any one of claims 1 to 4,
In the first step, dextran is produced by an enzymatic reaction using sucrose as a raw material, and the mixed solution containing the dextran produced in the first step, fructose, and unreacted sucrose is subjected to pseudo-transfer in the second step. The mixture is transferred to a third step in which chromatographic separation is carried out using a layer device, and in the third step, the first fraction enriched with fructose is separated by a pseudo moving bed operation and is separated upstream from the supply position of the mixed liquid. A second section in which dextran and sucrose are mixed is separated by a simulated moving bed operation, extracted from the downstream side of the supply position, and returned to the first step through a fourth step. By an enzymatic reaction characterized in that the third section enriched with is separated by the pseudo moving bed operation and extracted from a further downstream side than the extraction position of the second section. A method of producing dextran. 6. A reactor for producing a mixture of three or more components of raw materials and reaction products through a product inhibition reaction or a reversible reaction, comprising a plurality of adsorbent-packed columns forming an endless circulation system, and A group of adsorbent-packed towers that sequentially separate and form zones enriched with each of the above components contained in the mixed liquid that is continuously supplied to one of the adsorbent-packed towers along the flow direction of the liquid. and a first extraction means for extracting the first section of the separated portion from the adsorbent packed tower upstream of the mixed liquid supply position, and from the adsorbent packed tower downstream of the mixed liquid supply position, A second extracting means for extracting the separated second section, and a third extracting means for extracting the separated third section from the adsorbent packed tower located further downstream than the second extracting means. a simulated moving bed device comprising a discharge means, a mixed liquid transfer means for selecting one of the adsorbent packed towers of the simulated moving bed device from the reactor and supplying the mixed liquid, a first of the simulated moving bed device; ~ means for returning the liquid of the raw material-enriched fraction extracted from the third extraction means to the reactor;
A plurality of adsorbent packings formed as the endless circulation system are arranged at positions where the mixed liquid is transferred from the reactor to the simulated moving bed device and where liquids in each section are extracted by the first to third extracting means. 1. An apparatus for producing a useful substance by a product inhibition reaction or a reversible reaction, comprising a control means for synchronously moving the flow of a mixed liquid to a downstream side of a group of columns over time.