JPS6013678B2 - Method for producing cyclodextrin - Google Patents

Description

[Detailed description of the invention] The present invention relates to a method for producing cyclodextrin.

More specifically, a cyclodextrin synthase is allowed to act on at least one substance selected from starch, a starch composition fraction, and a starch separation reaction product, and the produced cyclodextrin is separated by ultrafiltration. In particular, it relates to a method for producing cyclodextrins. Cyclodextrin has 6 or more glucose molecules Q-1
・It is a 4-linked cyclic polysaccharide, and so far, Q-cyclodextrin with 6 glucose units, 8-cyclodextrin with 7 glucose units, y-cyclodextrin with 8 units, 9
Branched cyclodextrins in which 61 cyclodextrins and glucose or more Q-1.4 glucans are attached to the ring are known. 'These cyclodextrins have various uses; for example, in the food industry, pharmaceutical industry, and other fields, they are used to stabilize unstable substances and solubilize insoluble substances.

However, in the production of cyclodextrin using the secondary method, separation has always been a problem.

That is, the organic solvent precipitation method uses a toxic secondary starch agent and requires complicated steps such as steam distillation. Furthermore, it is difficult to separate cyclodextrin even with recently developed methods using glucoamylase and methods using surfactants. Cyclodextrin Synthesis The fact that superhydrogens have a wide variety of actions is one of the reasons why it has been difficult to produce cyclodextrins by conventional methods. In addition to the cyclic sugar production reaction, this cyclodextrin synthase also uses linear sugar as an acceptor to open the cyclic sugar ring and transfer it to it. Therefore, as the reaction progresses, the yield of cyclodextrin decreases due to decomposition of the produced cyclic sugar. Therefore, the present inventors believe that it is essential to remove the produced cyclic sugar from the reaction system as quickly as possible in order to increase its yield, and as a means to do so, the method is limited to the separation of cyclodextrin. As a result of intensive study on the application of the outer furnace filtration method, they found that very good results could be obtained and completed the present invention. In the present invention, the starch that can be used is one obtained from any raw material such as horse radish, shiso, sweet potato, corn, waxy corn, barley, wheat, tapioca, or the like.

Examples of starch composition fractions include amylose and amylobectin. Furthermore, starch decomposition reaction products include, for example, white dextrin, yellow dextrin, and calcined dextrin such as British gum; oxidized starch, and modified starch with low viscosity (by treatment with enzymes, acids, mechanical high-pelagic agitation, etc.) and other modified starches. ; Starch ethers represented by starch phosphate, starch acetate, etc.
Examples include starch derivatives such as starch esters; physically treated starches such as starches irradiated with radiation or neutron beams, high frequency treatment, or moist heat treatment; Q-starch; These starches may be used alone or in combination of two or more. Any known cyclodextrin synthase can be used, such as the enzyme produced by Bacillus macerans (macerans enzyme).
is known as a typical example.

When cyclodextrin synthase is added to starch, it liquefies and produces cyclodextrin.

In addition to cyclodextrin synthase, the liquefaction of starches requires Q-amylase (
(liquefied type), and also those solubilized by physical methods such as acids and alkalis can also be used. The biomass of cyclodextrin is affected by enzyme concentration, reaction time, etc.
Appropriate reaction conditions should be selected depending on the purpose. In the present invention, ultrafiltration is usually performed while reacting at a temperature of 40 to 50°C. The produced cyclodextrin is separated as a permeate through ultrafiltration. This permeate is a colorless and transparent liquid containing 90% or more of cyclodextrin, and can be concentrated by appropriate concentration measures if desired. General methods such as thermal evaporation can be used as the concentration means, but reverse osmosis is also a suitable method and can be extremely easily concentrated. For example, the permeate can be concentrated to 30 to 40% by reverse osmosis, and by leaving it at a low temperature, 6-cyclodextrin will first crystallize. When the mother liquor from which 3-cyclodextrin has been removed is further concentrated, a small amount of ethanol is added, and the solution is left at low temperature, Q-
Cyclodextrin is obtained. Next, as much as possible
- Cyclodextrin is obtained by removing the cyclodextrin, concentrating the mother liquor again by reverse osmosis, and leaving it at a low temperature. As described above, according to the method of the present invention, a clear liquid containing cyclodextrin as a main component is obtained, so crystallization is easy, and Q-, 8-, y-cyclodextrin, etc. can be obtained from a water or ethanol solution. Can be crystallized.

On the other hand, branched dextrin and a part of cyclodextrin remain in the part that is not subjected to extraocular heating, that is, the non-permeated liquid, and the branched dextrin is useful for producing branched cyclodextrin.

This branched dextrin is resistant to the action of cyclodextrin synthase, but by adding a surfactant such as sodium dodecyl sulfate to it, a large amount of branched cyclodextrin can be produced. Note that if a small amount of a debranching enzyme such as pullulanase is added to branched dextrin, branched cyclodextrin is not produced, but a large amount of Q1, 8-, y-cyclodextrin, etc. is produced. The action of pullulanase is strongly inhibited by cyclodextrin, so in normal batch reactions a large amount of pullulanase is required to increase the amount of Q-,8-y-cyclodextrin produced by debranching. As mentioned above, the method of the present invention is characterized in that a small amount of burulanase is sufficient because cyclodextrin exits the reaction system. Furthermore, the method of the present invention can be applied to a method for producing cyclodextrin using glucoamylase, a method for producing branched cyclodextrin, etc., and is very effective in removing glucose and oligosaccharides.

In carrying out the method of the present invention, conditions for the ultra-furnace method and reverse osmosis method may be appropriately determined. The cyclodextrin obtained by the method of the present invention has higher purity and safety than those obtained by conventional methods,
It is expected that it will find applications in many fields.

Next, the present invention will be explained in detail with reference to examples.

In addition, in the examples, a DDS Hirakoshi type ultra module, membranes used: heat-resistant ultra-furnace (manufactured by DDS, GR-61P) with nominal molecular weight cutoffs of 20,000 and 25,000, sucrose cu
RO membrane with t20-40% (manufactured by Nippon Denko, NTRIOOO
(No. series) was used. Example 1 Add 350% macerans enzyme to Mareshi, Shiyo starch lk9
001HU was added to the starch, and the starch was liquefied in tap water at a temperature of 65 to 6,800 degrees Celsius, allowed to cool at room temperature for 3 minutes, and then heated in an ultra-furnace for 2 hours while maintaining the temperature at 48 to 50 qo.

The total cyclodextrin yield was 16%, and its composition was as shown in FIG.

Example 2 Liquefied Q-amylase 5 in horse starch and perilla starch lk9
The mixture was liquefied at 60 to 70 QC, then autoclaved, and insoluble matter was removed by heating in an oven.Next, 500% of Macerans enzyme solution (20,000 THU) was added and ultrafiltration was carried out at 4,000 QC for 6 hours.

As a result, the amount of Q-cyclodextrin in the produced cyclodextrin was higher than in Example 1, but oligosaccharides such as glucose to maltoheptaose were contained as impurities.

Total cyclodextrin yield was 38%. Example 3 1kg of mareshi, perilla starch and 1000 macerans enzymes
0 THU was added to the mixture, and the mixture was liquefied at 60 to 70 OO in tap water, allowed to cool for 3 minutes at room temperature, and then autoclaved.

In addition, a new version of Macerans Enzyme Solution 500' (20,000
THU) was added and ultrafiltered at 4000 for 6 hours. In addition, pullulanase 50
0 m was added, and ultrafiltration was performed for 6 hours.

The permeate was concentrated using a reverse osmosis membrane to obtain a sugar solution containing 34% Bnx. The total cyclodextrin yield was 62%, and its composition was Q1: 42%, 8-: 36%, Y-: 12%, 6-: 4%, and other brans: 6%. Example 4 The oak crepe liquid IQ liquid 2.5Z shown in Example 3 was added to the part of Example 3 that was not subjected to ultrafurnace treatment, that is, the non-permeate liquid, and ultrafiltering was carried out at 40°C for 6 hours. Ta.

The total cyclodextrin yield was 34% of the bran feed.

Example 5 Waxy corn starch lk9 and macerans enzyme 10
00 Chunichi U was added, and the following treatment was carried out in the same manner as in Example 3.

Total cyclodextrin yield was 65%.

Example 6 Corn starch lk 9 was added to 20% hydrochloric acid 1
After suspending the solution in 40 qo solution for 6 hours, the hydrochloric acid was removed and the solution was neutralized with an alkali to pH 6-7.

Then, 10,000 THU of macerans enzyme was added and liquefied. After autoclaving, 20,000 THU of macerans enzyme was added and ultrafiltered at 50°C for 2 hours.

Note that when performing the reverse osmosis method, salts can be removed by using a membrane for removing salts. Total cyclodextrin yield was 30%.

[Brief explanation of the drawing]

FIG. 1 shows the cyclodextrin composition in the ultrafurnace permeate prepared by the method of the present invention. (High performance liquid chromatography, 65% acetonitrile) Q: is monocyclodextrin, 8: 8-cyclodextrin, y: y-cyclodextrin, 6
: 6-Cyclodextrin, G, ~G3: Glucose ~ Maltotriose. Figure 1

Claims (1)

[Scope of Claims] 1. A cyclodextrin synthase is allowed to act on at least one substance selected from starch, a compositional fraction of starch, and a starch decomposition reaction product, and the produced cyclodextrin is separated by ultrafiltration. A method for producing cyclodextrin, characterized by: 2. After allowing cyclodextrin synthase to act on at least one substance among starch, a compositional fraction of starch, and a starch decomposition reaction product, a permeate is obtained by ultrafiltration, and then the permeate is subjected to reverse osmosis. A method for producing cyclodextrin, which comprises concentrating it by a method.