KR0139615B1 - Production method of 0-carboxymethyl chitin - Google Patents

Abstract

The present invention relates to a process for the preparation of low insoluble content of 0-carboxymethyl chitin.

The method of the present invention specifically comprises (1) mixing chitin with 35 to 60% by weight of NaOH solution at 4 ° C. to room temperature and then freeze aged at −20 ° C. for about 15 to 20 hours to obtain alkaline chitin, (2) The obtained alkaline chitin was dispersed and swelled in isopropanol at 25 to 60 ° C. for 30 minutes to 1.5 hours to obtain a slurry, and (3) in the obtained slurry, chloroacetic acid was dissolved in isopropanol and added in portions over 0.5 to 2 hours. After carboxymethylation for 2 to 10 hours while maintaining at 30 to 60 ℃, (4) deionized water was added to the slurry, the pH of the slurry was adjusted to 7 with glacial acetic acid and then filtered (5) the residue obtained 0-carboxy prepared by the method of the present invention, which comprises washing with an ethanol / deionized water mixture, washing with ethanol and then filtration and drying under reduced pressure. Methyl chitin has a viscosity of having an insoluble matter content of less than 10% eseo 0.5% aqueous solution of 500 to 1500 centipoise.

Description

Method for preparing 0-carboxymethyl chitin

The present invention relates to a process for the preparation of 0-carboxymethyl chitin with low insoluble content. The present invention also relates to a process for obtaining 0-carboxymethyl chitin with low insoluble content from high molecular weight chitin.

Recently, with the large-scale catch of crabs and shrimps, interest in the disposal of shellfish, aquatic waste, has led to the availability of extractable chitin from all over the world. Chitin can be extracted not only from crustaceans, but also from insect shells, mushrooms, and fungi, and it is known that there are approximately 10 ¹¹ tons of chitin on Earth.

Chitin and its derivable chitosan were initially used for wastewater treatment and flocculant, but recently, with the establishment of high-quality chitin / chitosan manufacturing technology, their wide range of application has become known, especially in medicine, food and cosmetics. The direction is shifting to applications in high value-added fields. As the necessity of high quality and high purity chitin / chitosan in this field is increasing, several countries, such as Canada, Japan and Korea, are focusing on the development of high quality chitin / chitosan products.

However, chitin, defined as [(1 → 4) -acetamido-2-deoxy-β-D-glucose], is a strong acid sulfuric acid because it is highly crystalline due to the presence of intramolecular and intermolecular hydrogen bonds in its internal structure. It has the disadvantage that it is hardly dissolved except formic acid and methanesulfonic acid.

Hexafluoroisopropanol, hexafluoroacetone, 1,2-dichloro alcohol, formic acid, etc. are mentioned as organic solvents capable of dissolving chitin, but the solubility varies greatly depending on the source from which chitin is extracted. For example, chitin extracted from shrimp or cuttlefish is relatively easy to dissolve, but only partial dissolution is possible for chitin-derived high crystallinity. In order to improve the poor solubility of chitin, various measures have been taken. The most representative method is the following two methods:

The first method is to prepare water soluble chitin. The process is to treat the chitin with a concentrated NaOH solution to convert it to alkaline chitin and then partially deacetylate it, or N-acetylate by reacting chitosan with at least 90% deacetylation with acetic acid. In this case, it is known that the solubility in water is expressed when the degree of partial deacetylation is 50 to 55%. In the partial deacetylation method, the solubility expression in water is not constant due to the source of chitin, so that the solubility is not expressed in many cases. In addition, both processes are complicated and the manufacturing process takes a long time, so there are many problems in actual production.

The second method is to prepare a chitin derivative having a hydrophilic functional group inside the chitin molecular chain through a polymer reaction. Examples of hydrophilic chitin derivatives include O-carboxymethyl chitin, N-carboxymethyl chitin, hydroxypropyl chitin, phosphorylated chitin and sulfonated chitin, among which 0-carboxymethyl chitin (O-CM chitin) in industrial applications. And hydroxypropyl chitin have received positive reviews. In particular, carboxymethyl chitin can be applied to various fields because of the relatively simple manufacturing method and excellent solubility in water. For example, 70% carboxymethylated chitin is effective for maintaining long-term immunoadjuvant activity, and 80% carboxymethylated chitin is effective for maintaining short-term immunoadjuvant activity with low substitution degree of carboxymethylated chitin. In addition, it can be used as a substrate for evaluation of activity, such as lysozyme, a chitin degrading enzyme, has excellent adsorption ability to Ca ions, can be used as an artificial red blood cell adjuvant, and also has a tumor growth inhibitory effect.

The water-soluble carboxymethyl chitin can generally be prepared by reacting chitin and chloroacetic acid in a concentrated alkaline colorant, which is accompanied by partial hydrolysis in the -NHCOCH ₃ group present in the chitin. In addition to the COO ^- Na ⁺ group, a small amount of -NH ₂ can also be produced. Carboxymethylation is most active in the OH group at the C ₆ position.

In connection with the above, Trujillo's article Carbohydr. Res. Vol. 7, p483-485,1968, describes a method for preparing carboxymethyl chitin, in which 4 g of chitin is immersed in 20 ml of dimethyl sulfoxide (DMSO) for 24 hours, pulverized and filtered, and obtained in a cake form. The solid was washed with ethanol, regrind and dried, then treated with 50 ml of 65% NaOH solution for 1 hour, the resulting slurry was filtered and compressed to a wet weight of 14 g, to which 50 ml of isopropanol and 6 g of chloroacetic acid were simultaneously added. After stirring and filtration for 1 hour, 200 ml of water and concentrated hydrochloric acid were added to neutralize the liquid to neutrality, and the obtained high-viscosity solution was filtered through a glass filter, followed by 1 l of acetone to obtain sodium salt of carboxymethylchitin, This was washed and dried with anhydrous ethanol to obtain 3.5 g of carboxymethyl chitin.

It is described that a more refined fibrous carboxymethylchitin can be prepared by dissolving the final carboxymethyl chitin in 120 ml of water, followed by centrifugation to remove the insoluble matter, and adding 1 liter of acetone.

In Kosugi, European Patent Publication No. 0013512 of Zuiichi, 20 g of chitin and 65 g of 11N NaOH solution were mixed with each other at 15 ° C., and then frozen at −20 ° C. for 24 hours, and the frozen chitin was extracted with 400 ml of ethanol and chloroacetic acid. After dispersing in 30 g of a mixed solution, reacting with carboxymethylated at 3 ° C. for 2 hours at 20 ° C. for 45 hours, neutralizing the reaction product, dialyzing through a Visking tube and precipitating in ethanol to give carboxymethyl having a degree of substitution of 0.6. A method for producing carboxymethyl chitin is described which captures 28 g of chitin.

In addition, Tokura's paper, Polymer j. Vol. 15, No. 6, p485-489, 1983], 10 g of chitin was dispersed in 40 ml of 60% NaOH solution containing 0.2% sodium dodecyl sulfate while maintaining at 4 DEG C., and the resulting slurry was left at -20 DEG C for 10 to 10 hours. Alkaline chitin was prepared by freezing for 20 hours, which was then dispersed in 200 ml of isopropanol at room temperature, chloroacetic acid was slowly added dropwise to the dispersion slurry to neutralize the liquidity of the slurry, and then the slurry was filtered and washed with ethanol and then desorbed. A method of dissolving in 2 l of ionized water and slowly dropping into 5 l of acetone to precipitate solidify and obtaining the solidified carboxymethyl chitin by centrifugation followed by washing with acetone to obtain the final carboxymethyl chitin is described.

In the methods described in the preceding documents, the insoluble content of the prepared carboxymethyl chitin is accompanied by a separate process for removing it, which makes the manufacturing process complicated and difficult to apply to high molecular weight chitin raw materials. The above documents do not mention the whiteness, insoluble content and viscosity of the obtained carboxymethyl chitin, and there are many inadequate points for obtaining a high viscosity carboxymethyl chitin with low insoluble content.

Accordingly, the present inventors have diligently studied to solve the above problems, and as a result, the timing and amount of chloroacetic acid added to chitin, carboxymethylation reaction conditions, swelling time and temperature of alkaline chitin in an organic solvent, and the amount of alkaline solution used It was found that carboxymethylchitin having a minimum insoluble content, high molecular weight and high purity can be obtained by adjusting the determinative conditions such as the insoluble content and eliminating centrifugation for removing insoluble matter and reprecipitation by acetone.

It is an object of the present invention to provide a method for preparing O-carboxymethyl chitin with low insoluble content.

It is also an object of the present invention to prepare O-carboxymethyl chitin with low insoluble content from high molecular weight chitin without the need for a separate step to remove insoluble content.

That is, the present invention

(1) the chitin raw material was mixed with 35 to 60% by weight NaOH solution at 4 ° C. to room temperature, and then aged at about −20 ° C. for about 15 to 20 hours to obtain an alkaline chitin,

(2) dispersing and swelling the obtained alkaline chitin in isopropanol at 25 to 60 ° C. for 0.5 to 1.5 hours to obtain a slurry,

(3) chloroacetic acid was added to the slurry obtained over 0.5 to 2 hours, and then carboxymethylated for 2 to 10 hours while maintaining at 30 to 60 ° C,

(4) After the deionized water was added to the slurry, the pH of the slurry was adjusted to 7 with glacial acetic acid and then filtered.

(5) The obtained residue was washed with an ethanol / deionized water mixture, washed with ethanol, filtered and dried under reduced pressure to obtain carboxymethyl chitin, thereby preparing a low insoluble content of O-carboxymethyl chitin. It is about.

Hereinafter, the present invention will be described in more detail.

The starting material chitin has a viscosity measured using a Brookfield viscometer (No. 4 spindle, 30 rpm) in a 0.1% solution range of 200 to 1000 centipoise, and can be used for both low molecular weight chitin and high molecular weight chitin. have. Therefore, it can select and use according to the desired viscosity of the carboxymethyl chitin finally obtained.

First, the chitin is mixed with 35 to 60% by weight NaOH solution at 4 ° C. to room temperature and then freeze aged at −20 ° C. for about 15 to 20 hours to obtain alkaline chitin (step 1). It is preferable to add the NaOH solution of the above concentration in the range of 2 to 10 ml per 1 g of chitin. Use of NaOH solution concentrations and amounts outside the above ranges rapidly increases the insoluble content in the obtained carboxymethyl chitin.

The alkaline chitin is then dispersed and swelled in isopropanol at 25 to 60 ° C. for 30 minutes to 1.5 periods (step 2). Isopropanol is preferably added in a range of 16 to 30 ml per 1 g of chitin. At this time, the swelling time and swelling temperature has the most important influence on the viscosity and insoluble content of the carboxymethyl chitin obtained, so as not to deviate from the above conditions.

Next, croroacetic acid is added to the obtained slurry over 0.5 to 2 hours, and then carboxymethylated for 2 to 10 hours while maintaining at 30 to 60 ° C (step 3). At this time, the chloroacetic acid is preferably dissolved in isopropanol and added so as to be in the range of 1.7 to 4 g per 1 g of chitin, and it is preferable to add it several times over 30 minutes to 2 hours rather than adding it all at once. This split addition was found to reduce the insoluble content of the final carboxymethyl chitin and raise the viscosity.

When the temperature is lowered below 30 ° C. during the carboxymethylation reaction, the reaction hardly progresses or the reaction rate is very slow. Above 60 ° C., the yield and reaction rate are increased but the insoluble content is significantly increased and the cleavage of chitin molecular chain is promoted. This lowers the viscosity of the carboxymethyl chitin and can yellow it. In addition, when the reaction time is 6 hours or more, the insoluble content of the carboxymethyl chitin is no longer reduced, and the cleavage of the chitin molecular chain is promoted, so that the molecular weight of the final carboxymethyl chitin is accompanied by yellowing.

The final product is then collected from the carboxymethylated reaction product, for which deionized water is added, adjusted to pH 7 with glacial acetic acid and filtered (step 4). The obtained residue was then washed with an ethanol / deionized water mixture to remove residual salts, washed again with ethanol 2-3 times, filtered and dried under reduced pressure to give the final carboxymethyl chitin (step 5).

When the low molecular weight chitin is used as a starting material, the insoluble content of the obtained final product is close to 0%, but the high molecular weight chitin contains a much higher insoluble content (however, it is significantly reduced than the conventional method. When carboxymethyl chitin with an insoluble content of 0% is desired, the obtained carboxymethyl chitin can be preferably dissolved again in water and then purified by reprecipitation with acetone.

O-carboxymethyl chitin prepared by the method of the present invention as described above has an insoluble content of less than 10%, preferably less than 5%, most preferably 0%, in a 0.5% aqueous solution state Viscosity measurements using a Brookfield viscometer (No. 4 spindle, 30 rpm) have viscosity values in the range of 500-1500 centipoise.

The invention is further illustrated by the following examples.

In the following Examples, about 0.1% chitin solution, in which 0.01 g of chitin was dissolved in a mixed solution of 5 g of dimethylacetamide, 5 g of 1-methyl-2-pyrrolidone and 0.5 g of LiC1, was left at room temperature for 7 days, and then a Brookfield viscometer Although only chitin in the range of 200 to 1000 centipoise was used as the starting chitin material when measuring viscosity with (4 spindles, 30 rpm), chitin in other viscosity ranges may also be used.

Example 1

At room temperature, 5 g of chitin was mixed with 20 ml of an aqueous 45% NaOH solution (containing 0.2% sodium dodecyl sulfate), and left at 4 ° C. for 1 hour and then freeze aged at -20 ° C. for 15 to 20 hours. The freeze-aged alkaline chitin was finely pulverized, placed in a 500 ml three-necked flask, and 120 ml of isopropanol was added thereto, followed by slowly stirring for 40 minutes while maintaining 45 ° C. to disperse and swell the alkaline chitin. The reaction was proceeded while adding a solution of 14.75 g of chloroacetic acid in 15 ml of isopropanol in 6 to 10 portions over 1 hour and 15 minutes to the dispersion slurry. When the addition of chloroacetic acid was completed, the carboxymethylation reaction was performed for 4 hours and 45 minutes while maintaining 40 degreeC (reaction time after completion of addition of chloroacetic acid was defined as the carboxymethylation reaction time). Upon completion of the reaction, 8.2 ml of deionized water was added to the reaction system, glacial acetic acid was added dropwise to adjust the liquidity of the reaction system to pH7, and the solids were filtered and washed several times with 400 ml of a 80/20% by volume solution of ethanol / deionized water. After removal, the mixture was washed again with 400 ml of ethanol. The washed solid was collected by filtration and dried for 24 hours at 30 ° C. and reduced pressure to obtain 9.7 g of O-carboxymethylchitin. The insoluble content of the obtained product was less than 1% and the viscosity was 972 centipoise.

[Examples 2 to 5]

Example 1 was carried out in the same manner as in Example 1 except that various process conditions were changed as shown in Table 1, and the results are also shown in Table 1 Example.

[Comparative Examples 1 and 2]

These comparisons show the effect of NaOH solution concentration and chloroacetic acid addition time outside the scope of the present invention on insoluble content. Several process conditions were performed in the same manner as in Example 1 except that Table 1 was carried out, and the results are shown in Table 1.

[Comparative Examples 3 and 4]

It was carried out as Table 1 according to the method described in the articles of European Patent Publication No. 0013512 and Tokura mentioned as prior art, the results are also shown in Table 1.

Viscosity is the viscosity for the soluble part.

Claims (6)

(1) a chitin having a viscosity in the range of 200 to 1000 centipoise is mixed with a 40 to 60 wt% NaOH solution at 4 ° C. to room temperature, and (2) the mixture obtained in step (1) is about 15 to about 20 ° C. Freeze aging for 20 hours to obtain alkaline chitin, (3) the obtained alkaline chitin was dispersed and swelled in isopropanol at 25 to 60 ° C. for 30 minutes to 1.5 hours to obtain a slurry, and (4) chloro to the obtained slurry. After acetic acid was added in portions over 0.5 to 2 hours, carboxymethylation was carried out at 35 to 60 ° C. for 2 to 10 hours, (5) deionized water was added to the slurry, and the pH of the slurry was adjusted to 7 with glacial acetic acid. And then (6) the obtained residue was washed with a mixture of ethanol / deionized water, washed with ethanol, filtered and dried under reduced pressure to obtain O-carboxymethylchitin, having an insoluble content of 15% or less. Preparation of carboxymethyl chitin. The crime prevention according to claim 1, wherein in step (1), a 40 to 60 wt% NaOH solution is added in an amount ranging from 2 to 10 ml per 1 g of chitin. The method of claim 1 wherein said chitin is a high molecular weight chitin having a viscosity of at least 300 centipoise. The method of claim 2, further comprising the step of dissolving the obtained 0-carboxymethylchitin again in water and reprecipitation with acetone to purify it. The method of claim 1 wherein chloroacetic acid is dissolved in isopropanol and added. The method according to claim 5, wherein chloroacetic acid is added at a ratio of 1.7 to 4 g with respect to 1 g of chitin.