JPS61271296A - Production of n-acetylchito-oligosaccharide - Google Patents

Abstract

PURPOSE:To obtain the titled compound useful as a low-cariogenic sweetener, etc., smoothly, in large quantity and high efficiency, by partially hydrolyzing chitin with an acid, neutralizing the partial hydrolyzate with an alkali, and separating and removing the by-produced salt from the neutralized liquid by an ion-exchange membrane electrodialysis, etc. CONSTITUTION:Chitin used as a starting material is partially hydrolyzed with an acid, preferably with concentrated hydrochloric acid. The partially hydrolyzed liquid is neutralized with an alkali to obtain N-acetylchito-oligosaccharide. The neutralized solution is desalted by ion-exchange membrane electrodialysis to effect the separation and removal of by-produced salt. The desalted solution is preferably treated with an ion exchange resin to effect the complete decoloration and the purification to a high purity, and usually pulverized to obtain the objective compound.

Description

DETAILED DESCRIPTION OF THE INVENTION [Industrial Application Field] The present invention relates to a method for producing N-acetyl chito-oligosaccharide. The present invention relates to a novel method for producing N-acetyl chito-oligosaccharides, which consists of a specific desalting step from a neutralized solution.

[Conventional technology and problems]

N-acetylchito-oligosaccharide is a oligosaccharide H (oligosaccharide) in which 2 to 7 N-7 cetylglucosamines are linked through β-1,4 bonds, and is a polysaccharide contained in the carapace of crabs, shrimp, krill, etc. It is obtained by partially hydrolyzing the saccharide chitin.

Furthermore, oligosaccharides are saccharides in which several monosaccharides such as glucose and 7-lactose are bonded together, and have recently been attracting attention as new substances. For example, a coupling sugar in which 1 to 8 glucose molecules are bonded to the glucose molecule of sucrose, a 7-lactooligosaccharide in which 1 to 4 7-lactose molecules are bonded to sucrose, and 3 to 7 glucose molecules in α-1,4 bonds.
Examples include malto-oligosaccharides having 10 bonds. These are used as sweeteners that take advantage of their properties such as low cariogenicity (property that does not cause cavities) or indigestibility, use as bifidus factors in dairy products, and use in reagents and medicines. The future potential is great. N-acetylchito-oligosaccharide is a similar substance, and is always an interesting substance because of the structure shown below.

(n = 0 to 5) According to the method for producing N-acetylchitooligosaccharide, chitin is partially hydrolyzed with an acid, the partially hydrolyzed solution is neutralized with an alkali, and the resulting neutralized solution is A method is known for separating and removing by-product salt from. For the separation and removal of raw salt a from the neutralized solution, an adsorption separation method using, for example, a 1:1 mixture of activated carbon and celite has been proposed. Activated charcoal generally contains the monosaccharide N-acetylglucosamine,
Since D-glucosamine and the salts produced by neutralization are not adsorbed, and only oligosaccharides are adsorbed, according to the above adsorption separation method, N-acetyl chitochloride in the neutralized solution is It is possible to adsorb only oligosaccharides and separate them from large amounts of salts. N-- adsorbed on activated carbon
Acetylchito-oligosaccharide can be obtained as a solid by breaking the adsorption and eluting it using a solvent such as alcohol, and further concentrating the eluate to drive out the alcohol and the like.

However, in the above-mentioned adsorption separation method, the adsorption amount of N-acetylchitooligosaccharide on activated carbon is small, so a large amount of activated carbon is required, or a large amount of solvent is required for elution of N-acetylchitooligosaccharide. There are problems in the use of this method, and there is also the drawback that useful N-acetylglucosamine, which is produced in a considerable amount in a chitin partial hydrolysis solution, cannot be recovered.

The purpose of the present invention is to eliminate the above-mentioned problems and difficulties, and to efficiently, smoothly and advantageously produce a large amount of N-acetylchitooligosaccharides, including N-7 cetylglucosamine, which is a sugar, using chitin as a raw material. The goal is to provide a way to obtain

[Structure of the invention]

The present invention provides an N-acetylchitooligosaccharide, which comprises partially hydrolyzing chitin with an acid, neutralizing it with an alkali to produce an N-acetylchitooligosaccharide, and separating and removing raw III salt from the neutralized solution. The present invention provides a novel method for producing 9J N-acetyl chito-oligosaccharides, which is characterized in that the neutralized solution is desalted by ion-exchange membrane electrodialysis.

In the present invention, it is important to desalinate the neutralized solution by ion exchange membrane electrodialysis.

That is, when the neutralized solution is subjected to ion exchange electrodialysis, the by-product salts are efficiently separated and removed, and all of the generated N-acetylchito-oligosaccharide including the useful N-7 cetylglucosamine is recovered. Can be done. Not only can N-7 cetylglucosamine, which could not be recovered by conventional adsorption separation methods, be recovered smoothly and advantageously, but it can also be produced without the disadvantages such as losses that are seen in the methods of adsorption with activated carbon and elution with alcohol, etc. Almost all of the N-acetylchito-oligosaccharides obtained can be obtained.

The present invention will be explained in detail below.

The chitin used in the present invention is prepared r) by treating the carapace of crustaceans such as shrimp, crabs, and krill with hydrochloric acid to remove calcium, and further treating with sodium hydroxide to remove protein. Of course, chitin obtained by other acquisition routes, preparation methods, etc. may also be used.

Partial hydrolysis of chitin with an acid is carried out using hydrochloric acid, formic acid, acetic acid, sulfuric acid, etc., but it is preferably carried out using concentrated hydrochloric acid and stirring at 30 to 50''C for 2 to 3 hours. Next, in order to complete the hydrolysis reaction, it is diluted with water of about the same volume as the partial hydrolysis solution, and further hydroxylated by 25 to 50% to prevent the temperature from rising.)IJ
Neutralize with an alkali using an aqueous solution of aluminum.

The resulting neutralized aqueous solution has a slightly brownish color due to the decomposition of the sugar, and some undecomposed insoluble chitin remains, but these can be removed by suction filtration using a small amount of activated carbon and filter paper, leaving it colorless. A clear neutralized solution is obtained. In the present invention, the partial hydrolysis of fJ+ with an acid and the neutralization with a 7-alkali are not particularly limited, and may be carried out by appropriately adopting various conventionally known and well-known means and conditions. Can be done.

The neutralized solution obtained in this way differs slightly depending on the type and amount of acid used for partial hydrolysis of chitin or the amount of dilution water, the type and amount of alkali used for neutralization, and the conditions used in each process, but it is suitable. In a preferred embodiment, the solid content may be 15 to 20% and the salt content may be 10-15%.

This neutralized solution is subjected to a desalination process according to the invention via ion exchange membrane electrodialysis. An ion exchange membrane electrodialysis device generally has a large number of anion exchange membranes and cation exchange membranes arranged one after another, and a pair of electrodes arranged at both ends.

When a direct current is passed between the electrodes, the cations in the neutralizing solution move toward the cathode, and the anions move toward the anode. However, cations pass through the cation exchange membrane but not the anion exchange membrane, and on the other hand, anions pass through the anion exchange membrane but not the cation exchange membrane. As a result, a chamber where ions are concentrated (concentration chamber),
A chamber is created where ions are removed (desalination chamber).

Salts in the neutralization solution, such as common salt and D-glucosamine hydrochloride, which is an acetylated product of N-acetylglucosamine;
Sodium acetate and other substances produced by the deacetylation reaction move to a chamber where ions are concentrated and become uncharged N-7.
Cetylglucosamine and N-acetylchitooligosaccharide remain in the desalting chamber, and the neutralized solution is desalted.

The ion exchange membrane that can be used in the present invention is not particularly limited, and various conventionally known or well-known membranes may be used. For example, Neocepta CL-257,
CM-1~2. Examples include AM-1 to AM-3 (manufactured by Tokuyama Soda Co., Ltd.), Selemion CMV, and AMV (manufactured by Asahi Glass Co., Ltd.). In addition, each manufacturer sells ion exchange membrane electrodialysis devices that are systematized, so they can be easily handled. According to the present invention, it is possible to hydrolyze chitin with an acid and remove almost 100% of the salt in the neutralized solution. The desalted solution thus obtained may be pulverized as it is using a freeze dryer or spray dryer, but it is preferably pretreated with an ion exchange resin for complete decolorization and high purity purification. 1.
1.

The N-acetylchito-oligosaccharide obtained by the present invention is unique in that it contains the monosaccharide N-acetylglufusamine and N-7 cetylchitobio to N-acetylchitoheptaose, and can be easily produced in large quantities. It can be manufactured in

〔Example〕

Next, embodiments of the present invention will be described in more detail, but it goes without saying that the present invention is not limited by such explanations. In addition, in the following examples, the proportions are based on weight unless otherwise specified.

[Example 11 Chitin 400F (20 mesh pulverized product) was mixed with concentrated hydrochloric acid for 1.
In addition to 29, hydrolysis was carried out with stirring at 40"C for 3 hours. After completion of hydrolysis, dilution with the same volume of water 1..
It was neutralized to pH 7.0 with a 25% hydroxide solution. 100 g of activated carbon was added to this neutralized solution, and after suction filtration with Toyo Roshi No. 2, the residue was washed with a small amount of water, and this washing liquid was combined to form a colorless and transparent washing liquid 4.
! -I got it.

This washing solution (solid content: 19.6%, salt content: 13.4%) was prepared using 10 pairs of cation exchange membranes 'Neosepta CM-1'' and anion exchange membrane 1 'Neosepta A M-1''. TS-2-10 with 10 desalination chambers and effective membrane area of 2d1/sheet
Using a type electrodialysis device (manufactured by Koe 111 Soda Co., Ltd.), add 4Q of the above washing liquid to the desalination chamber and 4Q of 13% saline to the concentration chamber.
4Q of 2% saline was used in the electrode chamber, and the liquid temperature was 1.
Electrodialysis was performed for 210 minutes under the conditions of 4 to 35°C, current of 8 to 1.5 A, and voltage of 10 to 24 V. The volume of the desalted solution after electrodialysis is 1.7%, the Cj solid content is 5.6%, the salt concentration is 0.002%, and the desalination rate is 99.98%.

Next, the desalination solution described above was transferred to a column (φ4゜5X30cm) packed with cation exchange resin ``Amberlite I R120B'' (Oru/Company Mine) and an anion exchange resin 1.
Complete decolorization and purification were performed by passing it through a column (φ4, 5 x 30 cm) packed with Amberlite IRA400'' (manufactured by Olga Zensha).Furthermore, this liquid was concentrated under reduced pressure to obtain 3001, and a freeze dryer was used. 176 g of white powder was obtained.

The yield was 44.0% based on 400 g of raw chitin.

The composition of the white powder described in 1. is 31% N-acetylglucosamine, 19% N-acetylchitobiose, 19% N-acetylchitotriose, 15% N-7 cetylchitotetraose, Cetyl chitobentaose 10%,
The contents were 5% N-7 cetylchitohexaose and 1% N-acetylchitohexaose.

[Example 21 Salt production from 1 kg of chitin (20 mesh pulverized product)] ! 3
．． 7. After completion of the hydrolysis, the solution was diluted with the same volume of water and neutralized to pH 7.0 with 25% sodium hydroxide solution.

Add 250g of activated carbon to this neutralized solution and use Toyo Roshi No.
After suction filtration using No. 2, the residue was further washed with a small amount of water, and this washing liquid was combined to obtain a colorless and transparent washing liquid No. 12 Summer.

This washing solution (solid content 15.4%, salt content 11.0%) was desalinated using the same TS-2-10 type electrodialyzer (S! Use the above washing liquid 4Q in the chamber, 10% saline 4Q in the concentration chamber, and 2% saline 4Q as the electrode solution, liquid temperature 19-28°C, current 8-0.5A,
Electrodialysis was performed for 180 minutes under voltage conditions of 9 to 23V. Desalinated solution 1 after electrodialysis has a solid content of 2.1 summer and a solid content of 8.
4% and a salt concentration of 0.001%, the desalination rate is 99.99%. The following desalting operation was performed three times, and a total of 8.59 desalting solutions were obtained including the washing solution in the desalting chamber.

This desalted solution was completely decolorized and purified by passing through ion exchange 8 (fat) in the same manner as in Example 1. This solution was dried using a spray dryer to obtain 448 g of white powder. Its composition was 26% N-acetylglucosamine, 16% N-acetylchitobiose, 17% N-7 cetylchitotriose, and 15% N-acetylchitotetraose. %, N-acetylchitopentaose 13%, N-7 cetylchitohexaose 9%, and N-acetylchitotetraose 4%.

〔Effect of the invention〕

According to the present invention, it is possible to smoothly and advantageously recover N-acetylchitooligosaccharide produced by partial hydrolysis and neutralization of chitin, and it is possible to achieve the excellent effect of efficiently desalting. Furthermore, the effect that useful N-acetylglucosamine, which could not be recovered by conventional adsorption separation methods, can also be recovered is also achieved. According to the present invention, the drawbacks of conventional adsorption separation methods such as the use of a large amount of adsorbent and the use of a large amount of elution solvent are solved, and a large amount of N- including N-7 cetylglufusamine is produced using chitin as a raw material.
Acetylchito-oligosaccharides can be produced efficiently.

Claims (1)

[Claims] In a method for producing N-acetylchitooligosaccharides, which comprises partially hydrolyzing chitin with an acid, neutralizing with an alkali to produce N-acetylchitooligosaccharides, and separating and removing by-product salts from the neutralized solution. , a method for producing N-acetylchito-oligosaccharides, characterized in that desalting from the neutralized solution is carried out by ion exchange membrane electrodialysis.