KR20000012173A - Method of manufacturing polysaccharites having low molecular weight and oligosaccharites - Google Patents

Abstract

PURPOSE: Low molecular polysaccharide and oligosaccharide are prepared by ultrasonic irradiation of the polysaccharide in a solvent. CONSTITUTION: The polysaccharides such as chitin, chitosan, cellulose and its derivatives, starch and its derivatives, glycogen and its derivatives, heparin hyaluronic acid, alginic acid, gum or pectin are dissolved in the solvent selected from sulfonic acid and benzoic acid etc, followed by irradiation of light, ultrasonic, ultraviolet, gamma ray or electron beam to give polysaccharide and oligosaccharide. For an example, 1.5 g of chitosan is suspended in 100 g of ion exchange water, followed by addition of 0.6 g of acetic acid, and swelled at room temperature. Obtained solution is irradiated by high pressure mercury lamp having 190- 340 nm wave length to give low molecular polysaccharide and oligosaccaharide.

Description

METHODS OF MANUFACTURING POLYSACCHARITES HAVING LOW MOLECULAR WEIGHT AND OLIGOSACCHARITES}

The present invention relates to a method for producing low molecular weight polysaccharides and oligosaccharides, and more particularly to a method for low molecular weight and oligosaccharides of polysaccharides such as chitin, chitosan, cellulose that can be widely used in food additives, natural pesticides, pharmaceuticals and the like.

In recent years, low molecular weight polysaccharides and their oligosaccharides are expected to have a wide range of applications in the food and medical fields, as well as to have a large ripple effect on life sciences. In particular, elucidating the biological functions of glycoproteins and glycolipids for oligosaccharides leads to the development of new forms of pharmaceuticals and their applications.

As for plants, the structure of oligosaccharides isolated from cell walls and bioprotective activity against microbial infections have been revealed, and thus, the discovery of a new group of plant function regulators is expected to use them in agriculture. In addition, it can be widely used in the fields of food, cosmetics, medicine, diagnostic reagents.

On the other hand, chitin, one of the polysaccharides, is present along with proteins in vivo, such as shrimp, crab, and squid bone, and is produced by alkali and acid treatment. Chitosan is industrially treated with N-deacetylation to produce agricultural, food, Used as a raw material for pesticides and other industrial products. In general, chitosan has a relatively high molecular weight and is difficult to dissolve in a solvent when used in the above-mentioned fields, so that high molecular weight chitosan is low molecular weight or oligosaccharide.

In particular, water-soluble low-molecular chitin, chitosan and chitin, chitosan oligosaccharides are useful as plant growth promoters, soil improving agents and growth regulators in agriculture, and are widely used in fields such as food, cosmetics, medicine, and reagents.

The method for producing such water-soluble low molecular chitin, chitosan and chitin, chitosan oligosaccharide is known a lot of methods of acid degradation by concentrated hydrochloric acid or the like by enzymes such as chitinase, chitosanase. However, these methods have drawbacks such as complicated pretreatment-decomposition-aftertreatment process and high production cost.

In order to solve this problem, the present inventors have previously applied for a method of preparing low-molecular chitosan and chitosan oligosaccharides by dispersing chitin or chitosan in distilled water, dissolving it in an inorganic acid or an organic acid, and irradiating light, especially ultraviolet rays. 98-32474, filed August 11, 1998).

Following this research, efforts have been made to lower the molecular weight of polysaccharides such as cellulose and starch other than chitin and chitosan, or to prepare oligosaccharides. As a result, polysaccharides other than chitin and chitosan are also dissolved in a suitable concentration and irradiated with light or ultrasonic waves. By knowing that the low molecular polysaccharides and oligosaccharides thereof can be prepared, the present invention has been completed.

An object of the present invention is to dissolve polysaccharides such as cellulose and its derivatives, starch and its derivatives, glycogen and its derivatives, alginic acid and its derivatives, heparin, hyaluronic acid, as well as chitosan or chitin in a certain concentration in water, inorganic or organic acids, The present invention provides a method for producing low molecular polysaccharides and oligosaccharides thereof by irradiation with light or ultrasonic waves.

In order to achieve the above object, a low molecular weight polysaccharide and a method for preparing the oligosaccharide thereof according to the present invention are prepared by dissolving a polysaccharide in a solvent at a predetermined concentration to form a polysaccharide solution; It characterized by consisting of the step of producing a low molecular polysaccharide and oligosaccharide by irradiating light or ultrasonic wave to the polysaccharide solution.

The present invention will be described in more detail as follows.

Specific examples of the polysaccharide that can be low molecular weight in the present invention include cellulose and its derivatives, starch and its derivatives, glycogen and its derivatives, heparin, hyaluronic acid, alginic acid, gum and pectin, including chitin, chitin and chitosan. Xanthan gum or guar gum can be used.

As mentioned above, chitin is present in the living body with proteins such as shrimp, crab, and squid bone, and is manufactured by alkali and acid treatment. Chitosan is industrially treated with N-acetylation to produce agricultural, food, pesticide, Used as raw material for other industrial products.

Heparin is a linear anionic carbohydrate, a multicomponent heterogeneous substance consisting of glucoseamine (GlcN) and uronic acid. Heparin has different physiological activities depending on the molecular weight and high molecular weight heparin has been shown to inhibit blood coagulation, while low molecular weight heparin is known to have anti-Xa activity (Barrowcliffe, TW et al, J. Hasmatol, 41, 573 (1979). )). In addition, the use of low molecular weight heparin is increasing in the prevention of thrombosis.

On the other hand, heparin of high molecular weight, which is not fractionated, causes aggregation and adhesion of platelets in plasma, while low molecular weight heparin is known to not cause aggregation or adhesion of platelets in plasma because of its excellent compatibility with blood vessels (Muuasaky, S , et al., Chem. Pharm. Bull., 30, 3770 (1982)).

As a method of low molecular weight of heparin, a method of deaminizing a nitric acid solution and reducing the end group to 2,5-anhydromannose or 2,5-anhydromannitol has been used. Or a method of radical decomposition with hydrogen peroxide or the like.

In addition, cellulose is a polysaccharide most abundantly present on earth, and has been used since ancient times for clothing, paper, and the like. However, cellulose has solubility problems due to intermolecular hydrogen bonding, and its processing method is very narrow. However, it has been able to be used in some fields since it is dissolved in a special alkaline aqueous solution. On the other hand, in the case of making and using a derivative such as carboxymethyl cellulose (CMC), it is easy to associate and gel.

Accordingly, water soluble derivatives can be easily obtained by introducing carboxymethylated natural cellulose and introducing a certain proportion of substituents, which are used in fields such as food, medicine, textiles, cosmetics and paper. In addition, microcrystalline cellulose is hydrolyzed by acid and used as a low molecular compound with a degree of polymerization of about 200, and is used in various ranges such as pharmaceutical formulation forming aids, polymer fillers, and filtration aids.

On the other hand, alginic acid is a polysaccharide of marine plants comparable to cellulose, which is a land plant, and constitutes a material between the cell membrane and the cell membrane of brown algae, and occupies 30% of its constituents, making it the most useful resource in the ocean. Alginic acid is widely used as a functional substance such as thickening, stabilization and emulsification in the fields of food manufacturing, medical materials, paper industry, and dyeing industry. Recently, attention has been paid to water-soluble alginic acid, and water-soluble alginic acid has an excellent effect on health and medical fields such as antitumor. Water-soluble alginic acid is a substance having a low molecular weight of 30,000 to 40,000.

As a method for producing such low molecular polysaccharides and oligosaccharides, many methods for acid decomposition or hydrolysis with concentrated hydrochloric acid have been known. However, this method has a disadvantage in that the pretreatment-decomposition-aftertreatment process is complicated and the production cost is expensive.

In the present invention, in order to solve the above problems, the above-mentioned polysaccharide is dissolved in a solvent at a certain concentration, and the solution is irradiated with light or ultrasonic waves to lower molecular weight and oligosaccharide in a simple manner.

In the above application, in the method of lowering the molecular weight and oligosaccharide of chitin or chitosan, α-chitosan obtained by deacetylating chitin obtained by treating shrimp shell or crab shell with hydrochloric acid as chitin or chitosan, or cuttlefish bone Β-chitosan prepared by treatment with caustic soda is used. Preferably, the chitosan used in the present invention has a degree of deacetylation of 0 to 100% and a degree of polymerization of about 200 to 30,000. Alternatively, chitosan may be used on the market.

Inorganic acids used to dissolve chitosan in the present invention include sulfuric acid, hydrochloric acid, phosphoric acid, nitric acid, and the like, with hydrochloric acid being preferred. The organic acid may include acetic acid, tartaric acid, salicylic acid, citric acid, hydroxyacetic acid, ringoic acid, propionic acid, formic acid, lactic acid, glycerin acid, ascorbic acid, xenoic acid, sulfamic acid, benzoic acid, sorbic acid, glutamic acid, and glutamic acid. Among them, acetic acid, salicylic acid, benzoic acid and sorbic acid are preferable. In addition, 0.5-10 times mole can be used with respect to the amine group of a chitosan, and, as for the density | concentration of the acid used for dissolving chitosan, it is preferable to use the same mole with respect to mole of glucose amine units.

The concentration of chitosan solution used in the production is preferably 0.5 to 10% by weight, and may be used in excess of 10% by weight.

On the other hand, the production may be carried out in an inert gas atmosphere such as nitrogen gas, carbon dioxide gas, helium gas, argon gas, etc., but may be performed in an inert gas atmosphere. However, although it can carry out in an inert gas atmosphere according to the temperature at the time of irradiating an acid or ultraviolet-ray which melt | dissolves chitosan, it is not necessary to carry out in inert gas atmosphere. However, discoloration of the manufactured article can be prevented by using an inert gas depending on the acid at which the chitosan is dissolved or the temperature when irradiated with ultraviolet rays.

In the low molecular weight and oligosaccharide of the polysaccharide, when the polysaccharide is dissolved in a solvent, the solvent to be dissolved may vary depending on the type of polysaccharide.

Specifically, in the case of a water-soluble polysaccharide, distilled water can be used without using the above organic acid or inorganic acid as a solvent.

Preferred solvents for dissolving cellulose and its derivatives are benzyltrimethylammonium, N-methylmorphyrine-N-oxide, dimethylsulfoxide (DMSO), SO ₂ -amine, N ₂ O ₄ -DMSO, paraformaldehyde- DMSO, distilled water and the like can be used.

In the case of chitin, LiCl-DMSO, dimethylacetamide, N-methylpyrrolidone, hexafluoro-2-propanol, hexafluoroacetone, trichloroacetic acid-dichloroethane and the like are used. Uses distilled water.

In the case of alginic acid, distilled water and alkaline aqueous solution can be used.

For heparin, distilled water or formic acid is used.

As described above, polysaccharides including chitosan are dissolved in a predetermined solvent to prepare a polysaccharide solution, and then light or ultrasonic irradiation is performed to produce low molecular chitosan and oligosaccharides thereof. The light used in the preparation is gamma rays, ultraviolet rays, electron beams, and the like. It may be used, but is preferably ultraviolet.

In the case of ultraviolet rays, it is preferable to use 190-340 nm, and it is preferable that it is 50-580 Kev for gamma rays, and it is preferable to carry out using 10 Kev-1 Mev as an electron beam. And the irradiation temperature of light is 4-80 degreeC, It is preferable to carry out at room temperature.

The light irradiation time varies depending on the intensity of light, and in the case of ultraviolet rays, 1 to 240 hours is preferable. If the irradiation time of gamma rays or electron beams of high energy becomes long, problems such as discoloration and deterioration may occur.

On the other hand, the irradiation of light may be irradiated by placing the ultraviolet, gamma rays or electron beam generating device closer to the sample solution or immersed in the solution.

On the other hand, the ultrasonic irradiation is preferably performed for 1 to 72 hours so that the frequency of 15 to 100KHz output 100 to 4800W.

If the frequency, power, or irradiation time of the ultrasonic irradiation exceeds the above range, there may be a problem of discoloration and deterioration.

Hereinafter, the present invention will be described in detail with reference to Examples, but the present invention is not limited by the Examples.

Example 1

1.5 g (9.3 mol amino group) of chitosan was suspended in 100 g of ion-exchanged water, followed by adding 0.6 g (9.3 mol) of acetic acid to swell at room temperature. The solution thus prepared was irradiated for 2 hours using an ultraviolet (high pressure mercury lamp) having a wavelength of 190 ~ 340nm to prepare a low molecular chitosan and oligosaccharides. Then, the low molecular weight chitosan and oligosaccharide were dissolved, lyophilized or spray dried to obtain a powder.

EXAMPLES 2-7

Prepared in the same manner as in Example 1, except that the UV irradiation time was changed to 4, 12, 18, 24, 36 and 72 hours, respectively.

Example 8

The low molecular weight chitosan and oligosaccharide obtained by the method of Example 1 were dispersed in distilled water-methanol mixed solution to which triethylamine was added, and acetic anhydride was added to N-acetylate to prepare low molecular weight chitin and oligosaccharides thereof.

Example 9

2 g of chitosan was suspended in 96 g of ion-exchanged water, and 2 g of acetic acid was added to swell at room temperature for 12 hours to irradiate ultraviolet light having a wavelength of 190 to 340 nm to prepare a chitosan decomposition product. The decomposed product was used as a solution as it was decomposed or lyophilized or spray dried.

[Examples 10-12]

Prepared in the same manner as in Example 9, except that the UV irradiation time was changed to 24, 36 and 72 hours, respectively.

Example 13

Prepared in the same manner as in Example 9 except that 2g of carboxymethyl cellulose was dissolved in 98g of ion-exchanged water.

Example 14

Manufactured in the same manner as in Example 9, except that 2 g of alginic acid was dissolved in 98 g of ion-exchanged water.

Example 15

Prepared in the same manner as in Example 9, except that heparin 2g was used.

Example 16

2 g of carboxymethyl cellulose was dissolved in 100 g of ion-exchanged water, and the solution was irradiated with ultraviolet light (high pressure mercury lamp) having a wavelength of 190 to 340 nm for 12 hours to prepare a carboxymethyl cellulose decomposition product. The decomposed product was used as a solution as it was decomposed or lyophilized or spray dried.

[Examples 17-19]

Prepared in the same manner as in Example 16, except that UV irradiation time was changed to 12, 24, 36, 72 hours, respectively.

Example 20

Prepared in the same manner as in Example 16, but irradiated with an ultraviolet wavelength of 190 ~ 340nm.

Example 21

Prepared in the same manner as in Example 16, except that starch was used as a sample.

Example 22

Prepared in the same manner as in Example 16, but instead of ultraviolet light was irradiated with ultrasonic waves to a temperature of 25 ℃ for 2 hours at a frequency of 20kHz, 1000W capacity.

[Examples 23 to 26]

Prepared in the same manner as in Example 22, except that the ultrasonic irradiation time was changed to 4, 12, 18, 24 hours, respectively.

Comparative Example 1

Prepared in the same manner as in Example 9, but did not irradiate ultraviolet rays.

Comparative Example 2

Prepared in the same manner as in Example 13, but did not irradiate ultraviolet rays.

Comparative Example 3

Prepared in the same manner as in Example 14, but did not irradiate ultraviolet rays.

[Comparative Example 4]

Prepared in the same manner as in Example 15, but did not irradiate ultraviolet rays.

[Comparative Example 5]

Prepared in the same manner as in Example 16, but did not irradiate ultraviolet light.

Comparative Example 6

Prepared in the same manner as in Example 21, but did not irradiate ultraviolet rays.

Experimental Example

The average molecular weight and yield of the degradation products and oligosaccharides obtained according to the above Examples and Comparative Examples were analyzed by gel permeation chromatography (GPC). At this time, Shodex OHpakSB-801 + SB-802 + SB-803 (7.5mm ID × 300mm L) was used as the column, and the eluate was 0.2% acetic acid (including 0.1M NaCl) for chitosan. 0.1M phosphate buffer solution was used. Flow rate was analyzed by adjusting to 0.8ml / min.

The results are shown in Table 1 below.

division yield(%) Weight average molecular weight (Mw) Example One 100 120,000 2 100 85,000 3 100 34,000 4 100 20,000 5 100 12,000 6 100 8,000 7 100 2,000 8 100 9 100 34,000 10 100 20,000 11 100 12,000 12 100 8,000 13 100 42,000 14 100 52,000 15 100 4,200 16 100 34,000 17 100 20,000 18 100 12,000 19 100 8,000 20 100 120,000 21 100 42,000 22 100 52,000 23 100 31,000 24 100 12,400 25 100 10,000 26 100 7,400 Comparative example One 620,000 2 540,000 3 670,000 4 15,000 5 620,000 6 540,000

From the results of Table 1, it can be seen that according to the present invention, even a simple process using light or ultrasonic waves can effectively produce low molecular weight polysaccharides and oligosaccharides thereof.

As described in detail above, according to the present invention, low-molecular polysaccharides can be easily used by using light, especially ultraviolet or ultrasonic waves, without going through complicated decomposition and post-treatment processes such as conventional acid decomposition with concentrated hydrochloric acid or enzymes. And since the oligosaccharides thereof can be produced, it is possible to obtain the effect that the manufacturing process is simplified and the manufacturing cost is reduced.

Although the preferred embodiments of the present invention have been described above, those skilled in the art can variously modify and change the present invention without departing from the spirit and scope of the present invention as set forth in the claims below. I can understand.

Claims (7)

Dissolving the polysaccharide in a solvent at a predetermined concentration to form a polysaccharide solution; A method of producing low molecular polysaccharides and oligosaccharides comprising the steps of preparing low molecular polysaccharides and oligosaccharides by irradiating the polysaccharide solution with light or ultrasonic waves. The method of claim 1, Polysaccharides include chitin, chitosan, cellulose, starch, glycogen, heparin, hyaluronic acid, alginic acid, xanthan gum, guar gum, pectin and derivatives thereof. The method of claim 1, As light, a method for producing low molecular polysaccharides and oligosaccharides, characterized in that it is selected from ultraviolet rays, gamma rays or electron beams. The method of claim 1, Light irradiation is a method for producing low molecular polysaccharides and oligosaccharides, characterized in that carried out for 1 to 240 hours at 4 ~ 80 ℃ using light selected from ultraviolet rays of 190 to 340 nm, gamma rays of 50 to 580 Kev or electron beam of 20 Kev to 1 Mev. The method of claim 1, Ultrasonic irradiation is a method for producing low molecular polysaccharides and oligosaccharides, characterized in that performed for 1 to 72 hours at a frequency of 15-100KHz, output 100-4800W. The method of claim 1, The polysaccharide solution is a method for producing low molecular weight polysaccharides and oligosaccharides, characterized in that the concentration is 0.5 to 10%. The method of claim 1, Examples of the solvent include distilled water, sulfuric acid, hydrochloric acid, phosphoric acid, acetic acid, tartaric acid, salicylic acid, citric acid, hydroxy acetic acid, ringoic acid, propionic acid, lactic acid, glycerin acid, ascorbic acid, xenoic acid, sulfamic acid, benzoic acid, sorbic acid, glutamic acid, Glutamic acid, benzyltrimethylammonium, N-methylmorphyrine-N-oxide, dimethyl sulfoxide, SO ₂ -amine, N ₂ O ₄ -dimethylsulfoxide, paraformaldehyde-dimethylsulfoxide, LiCl-dimethylsulfoxide, dimethyl Method for producing low molecular polysaccharides and oligosaccharides, characterized in that it is selected from the group consisting of acetamide, N-methylpyrrolidone, hexafluoro-2-propanol, hexafluoroacetone, trichloroacetic acid-dichloroethane and formic acid .