KR100369517B1 - Method for preparing a low molecular weight polysaccharides and oligosaccharides - Google Patents

Abstract

The present invention relates to a method for efficiently preparing low molecular weight polysaccharides and their oligosaccharides by slightly adding a decomposition catalyst in a state in which the polysaccharide is dispersed in a solvent.These low molecular weight polysaccharides and oligosaccharides thereof are used in medicines, food additives, cosmetics, natural pesticides, and medical treatments. It is a bioactive material that can be widely applied as a material.

Description

Method for preparing a low molecular weight polysaccharides and oligosaccharides

The present invention relates to a method for producing low molecular weight polysaccharides and oligosaccharides thereof obtained by adding a small amount of a decomposition catalyst from a polysaccharide as a raw material. These low molecular weight polysaccharides and oligosaccharides are used in various fields in food, seasonings, feed, fertilizers, nutrients, cosmetics and medicines.

In particular, as the biological functions of glycoproteins and glycolipid oligosaccharides have been revealed, the development of new forms of medicines and their applications are rapidly expanding. In plants, the structure of oligosaccharides isolated from the cell wall and bioprotective activity against microbial infections have been revealed, and as a new group of plant regulatory substances are discovered, it is expected to be used in agriculture. Protein is present in squid bones and is produced by alkali and acid treatment. Chitosan is produced by N-deacetylation of alkaline chitin and is used as a raw material for agriculture, foodstuffs, pesticides and other industrial products. In general, chitosan has a relatively high molecular weight and has various problems such as difficulty in dissolving in a solvent when it is used in the above-mentioned fields, and thus high molecular weight chitosan is used for low molecular weight or oligosaccharide. In particular, low-molecular chitin, chitosan and their oligosaccharides have excellent antimicrobial activity and growth promoting effect, which is useful as a plant growth promoter, soil improving agent, and growth regulator in agriculture, and can be widely used in fields such as food, cosmetics, medicine, and reagents. have.

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, T. W et al, J. Hasmatol, 41, 573 (1979). Low molecular heparin is also used to prevent thrombosis. Furthermore, heparin of high molecular weight which is not fractionated leads to the aggregation or adhesion of platelets in plasma, while low molecular weight heparin is known to not cause the aggregation or adhesion of platelets in plasma because of its excellent compatibility with blood vessels. (Myyasaky, S., et al, Chem. Pharm. Bull., 30, 3770 (1982)

As a method of low molecular weight of heparin, a method of deamination with nitric acid solution and reducing the terminal to 2,5-anhydromannose or 2,5-anhydromannitol is generally used. Or a method of radical decomposition with hydrogen peroxide or the like.

In addition, cellulose is the most massive polysaccharide present on earth and has been used since ancient times for clothing and paper. However, cellulose has solubility problems due to intermolecular hydrogen bonding, so its processing method is very narrow, and it has been 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 or gel. Therefore, by introducing carboxymethylation of natural cellulose and introducing a certain proportion of substituents, water-soluble derivatives can be easily obtained. These are used in the fields of food, medicine, textiles, cosmetics, paper and the like. In addition, by reducing the degree of polymerization to about 200 degree of polymerization by hydrolyzing microcrystalline cellulose with acid, application methods such as formulation forming aids, polymer fillers, and filtration aids of pharmaceutical products are expanding.

On the other hand, alginic acid is a polysaccharide of marine plants that can be compared to cellulose, which is a land plant, and constitutes a material between the cell membrane and the cell membrane of brown algae. Alginic acid is widely used as a functional substance such as thickening, stabilization, and emulsification in the fields of food, medical, paper industry, and dyeing industry. Attention in recent years is water-soluble alginic acid. Water-soluble alginic acid has an excellent effect on health and medical fields such as antitumor and immune activity. This alginic acid is a substance having a low molecular weight of 30,000 to 40,000.

As for the method for producing such low molecular polysaccharides and oligosaccharides, there are many known methods of acid degradation using concentrated hydrochloric acid or the like, or enzymes such as chitinase, chitosanase, and cellulase. However, the enzymatic digestion method has a disadvantage in that the location of cleavage and degradation of polysaccharides is limited due to the specificity of the enzyme, thereby limiting the types of degradation products that can be obtained. In addition, in the case of acid decomposition, since polysaccharides are decomposed in a strong acid solution such as concentrated hydrochloric acid, a large amount of monosaccharides are generated, and the conversion efficiency is low. Finally, a neutralization process is required and a desalting process is essential. There are also methods using nitrite salts (Japanese Patent Laid-Open No. 62-184002), methods using peracetic acid (Japanese Laid-Open Patent Publication No. 2-41301), methods using hydrogen peroxide (Japanese Laid-Open Patent Publication No. 2-22301), and enzymes. They are all difficult to obtain in a solid state because they are prepared in a dissolved state, and there is a problem that spray drying or lyophilization is required to obtain a solid. In addition, in the case of polysaccharide having a large molecular weight, since the concentration of dissolving is limited (1 to 5% by weight), production efficiency is not good, and thus a problem occurs in terms of production price. The present inventors have also provided a method for preparing low molecular weight polysaccharides and oligosaccharides thereof in a state in which a polysaccharide is dissolved in a solvent (10-1999-010444, 10-2000-0005039, 10-2000-0021021). .

The present invention is a method for producing low molecular weight polysaccharides and oligosaccharides thereof, in which ammonium persulfate and persulfate are used as a decomposition catalyst in a dispersion solution in a state in which the polysaccharide is dispersed in a mixed solvent such as distilled water, an organic solvent or an organic solvent / distilled water, depending on the type of polysaccharide. Potassium, acetyl peroxide, peracetic acid, hydrogen peroxide, hydrochloric acid, nitrous acid, hydrogen peroxide, etc. are added or heated, or a photolysis catalyst is added and irradiated with light to precipitate in solids dispersed in a solvent, precipitated by filtration, or spray dried Or it relates to a method for efficiently producing a low molecular weight polysaccharide and its oligosaccharides without a process such as lyophilization.

The present invention relates to a process for preparing low molecular polysaccharides and their oligosaccharides.

That is, dissolving the decomposition catalyst at a constant concentration in a mixed solvent such as distilled water, organic solvent, organic solvent / distilled water, and dispersing the polysaccharide at a constant concentration in the above solution to irradiate a constant temperature or light to disperse the solid in the solvent. The present invention relates to a method for efficiently preparing low molecular polysaccharides and oligosaccharides thereof made by reacting and filtering in a mixture.

Polysaccharides used in the present invention include chitin, chitosan, alginic acid, pectin, inulin, cellulose, gum, starch, glycogen, heparin, hyaluronic acid, pullulan, glucomannan, polygalactoseamine, and derivatives thereof. The concentration of the polysaccharide solution that can be used in preparation is the concentration at which the polysaccharide can be immersed, preferably 20 to 60% by weight.

The decomposition catalyst used in the present invention may be divided into a pyrolysis catalyst and a photolysis catalyst, and the pyrolysis catalyst may be 4,4'-azobis-4-cyanopentanoic acid or 2,2'-azobis-isobutylamidine. And salts thereof, 2,2'-azobis-2- (methylcarboxy) propane, ammonium persulfate, potassium persulfate, hydrogen peroxide-Fe ²⁺ , potassium persulfate or ammonium persulfate-sodium bisulfite or sodium bisulfite, Cumene hydroperoxide-amine, peroxyacetic acid, acetyl peroxide, azo-nitriles (such as N, N'-azobisisobutylonitrile), alkyl peroxides (such as t-butylperoxide), aryl per Oxides (such as dicumyl peroxide), hydropuck oxides (such as t-butyl hydroperoxide), ketone peroxides (such as diisopropyl ketone), peresters, percarbonates, hydrochloric acid, nitric acid, sulfuric acid, Inorganic acids such as nitrous acid, chloric acid, sodium chlorate, chlorite, hydrogen peroxide, ozone, nitrous acid A catalyst such as a mountain cove is characterized in that selected according to the solvent. Examples of photolysis catalysts include halogen compounds (iodine, bromine, iodine chloride), halogenated organic compounds (phenyliodo dichloride, etc.), hydrogen peroxide, alkyl hydroperoxides (t-butyl hydroperoxide, etc.). , Cumene hydroperoxide, peroxides (diisopropyl peroxide, dicumyl peroxide, benzoyl peroxide, etc.), ketones (acetone, 3-pentanone, 2-heptanone, methoxyacetone, 1-phenylacetone, aceto Phenone, benzoin, benzophenone, etc.), quinones (hydroquinone, anthraquinone, etc.), azo compounds, hydrazones, titanium oxide, zinc oxide, iron oxide, cadmium sulfide, zinc sulfide, hydrogen peroxide, ozone, and the like. It is characterized in that it is selected according to the solvent and the metal oxide is used to coat or disperse in the lamp. In addition, the amount of the catalyst may be 0.01 to 60% by weight of the weight of the polysaccharide, but 0.05 to 30% by weight is suitable.

Organic solvents usable in the present invention include alcohols such as methyl alcohol, ethyl alcohol, (iso) propyl alcohol, (t-, iso) butyl alcohol, and the like as acetone, ethyl methyl ketone, diethyl ketone, methyl propyl ketone, Ethyl propyl ketone and the like dimethyl permamide (DMF), dimethyl acetamide (DMAc), tetrahydrofuran (THF), dioxane, benzene, toluene, acetonitrile, xylene, hexane, ethylene glycol, etc. can be used. Water, aqueous acetic acid solution, aqueous hydrochloric acid solution, caustic soda solution, etc. may be used, but these solvents may be used singly or in combination of two or more solvents depending on the type of polysaccharide and the type of decomposition catalyst.

In the present invention, the reaction temperature depends on the solvent and the kind of catalyst to which the catalyst may act, and is 0 to 150 ° C, preferably 25 to 100 ° C.

In addition, the light used in the present invention uses ultraviolet light, visible light, and radiation depending on the type of catalyst, and ultraviolet light is suitable to use in the range of 200 to 400 nm, and radiation may use α-ray, β-ray, r-ray, or electron beam. r line and electron beam are convenient. The dose of the electron beam is used up to 0.01 to 2,000 kGy, it is preferable to use 100 to 2,000 kGy when used in a dry state and 0.01 to 20 kGy in a solution state.

In the present invention, in order to increase decomposition efficiency or prevent discoloration, one type of air, oxygen, nitrogen, helium, etc. may be selected and used according to the type of decomposition catalyst.

Hereinafter, the present invention will be described in detail with reference to Examples.

Example 1

5 g of potassium persulfate was dissolved in 200 g of distilled water, and 50 g of chitosan (molecular weight 150,000) was dispersed in this solution, and then reacted at 60 ° C. for 3 hours to prepare a chitosan decomposition product. The digested product was filtered, washed several times and dried.

Example 2

To prepare a chitosan decomposition product by reacting an ozone / oxygen mixed gas at a rate of 5.4 L / h at 60 ° C. for 3 hours in a solution obtained by dispersing chitosan (molecular weight 200,000) in distilled water in 200 g of ethyl alcohol / distilled water (3: 1). Except that was carried out in the same manner as in Example 1.

Example 3

5 g of potassium persulfate was dissolved in 200 g of ethyl alcohol / distilled water (3: 1), 50 g of starch (molecular weight 340,000) was dispersed in this solution, and reacted at 80 ° C. for 3 hours to prepare a starch decomposition product. The digested product was filtered and washed several times with ethyl alcohol / distilled water (1: 1) solution and dried.

Example 4

The same procedure as in Example 3 was carried out except that 50 g of chitosan (molecular weight: 250,000) was dispersed.

Example 5

The same procedure as in Example 3 was carried out except that 50 g of sodium alginate (molecular weight: 450,000) was dispersed.

Example 6

The same procedure as in Example 3 was carried out except that 50 g of carboxymethyl cellulose (molecular weight: 340,000) was dispersed.

Example 7

The same procedure as in Example 3 was carried out except that 50 g of heparin (molecular weight: 17,000) was dispersed.

Example 8

10 g of hydrogen peroxide (30%) was dissolved in 200 g of ethyl alcohol / distilled water (3: 1), 50 g of chitosan (molecular weight: 250,000) was dispersed in this solution, and then reacted at 80 ° C. for 3 hours to prepare a chitosan decomposition product. The digested product was filtered and washed several times with ethyl alcohol / distilled water (1: 1) solution and dried.

Example 9

10 g of hydrogen peroxide (30%) was dissolved in 200 g of ethyl alcohol / distilled water (3: 1), and 50 g of chitosan (molecular weight: 250,000) was dispersed in this solution, and then irradiated with ultraviolet (high pressure mercury lamp) at 254 nm for 6 hours to decompose the chitosan. Was prepared. The digested product was filtered and washed several times with ethyl alcohol / distilled water (1: 1) solution and dried.

Example 10

The average molecular weights of the degradation products obtained by the method of Example 1-9 and their oligosaccharides were analyzed by GPC. Shodex OHpak SB-801 + SB-802 + SB-803 (7.5mm ID × 300mm L) was used, and the eluate was used for 0.1M acetic acid buffer solution and the flow rate was adjusted to 0.8ml / min. The results are shown in Table 1.

Item Example 1 Example 2 Example 3 Example 4 Example 5 Example 6 Example 7 Example 8 Example 9 Molecular Weight (Mn) 49,000 55,000 45,000 57,000 43,000 41,000 6,000 52,000 48,000 Molecular weight distribution 1.9 2.0 2.1 2.2 1.8 2.1 1.5 2.4 2.3 yield(%) 98 97 98 96 97 95 94 98 97

As can be seen in the above examples, the polysaccharide was dispersed in a solvent, and then reacted by adding a decomposition catalyst to be dispersed at a high concentration, and there was no process such as precipitation or spray drying or lyophilization. It was not necessary and the discoloration which appeared at the time of decomposition did not appear, and the quality was also favorable. The present invention can efficiently prepare low molecular weight polysaccharides and oligosaccharides in the solid state, process cost is low, and can produce polysaccharides of desired molecular weight in a short time and can be produced in high concentrations, which is much more economical.

Claims (6)

20 to 60% of the polysaccharide solution is dispersed in a solvent, and in the dispersed state, 0.01 to 60% by weight of a decomposition catalyst is added thereto, and the reaction is heated to 0 to 150 ° C. or 200 to 400 nm ultraviolet, α, β A low molecular polysaccharide and a method for producing oligosaccharide thereof, characterized in that any one selected from?,?, and 0.01 to 2000 kGy electron beams is reacted and filtered. The catalyst according to claim 1, wherein the decomposition catalyst is 4,4'-azobis-4-cyanopentanoic acid, 2,2'-azobis-isobutylamidine and salts thereof, 2,2'-azobis- 2- (methylcarboxy) propane, ammonium persulfate, potassium persulfate, hydrogen peroxide-Fe ²⁺ , potassium persulfate or ammonium persulfate-sodium bisulfite or sodium bisulfite, cumene hydroperoxide-amine, peroxyacetic acid, acetyl Peroxide, N, N'-azobisisobutylonitrile, t-butylperoxide, dicumylperoxide, t-butylhydroperoxide, diisopropylketone, hydrochloric acid, nitric acid, sulfuric acid, chloric acid, sodium chlorate, azo Chloric acid, hydrogen peroxide, ozone, ascorbic acid, iodine, bromine, iodine chloride, phenyliodo dichloride, hydrogen peroxide, alkyl hydroperoxide, t-butyl hydroperoxide, cumene hydroperoxide, di Isopropyl peroxide, Dicumyl peroxide, Ben Coyl peroxide, acetone, 3-pentanone, 2-heptanone, methoxyacetone, 1-phenylacetone, acetophenone, benzoin, benzophenone, hydroquinone, anthraquinone, azo compound, hyrazone, titanium oxide, A method for producing low molecular weight polysaccharides and oligosaccharides thereof, characterized by using any one or two or more of zinc oxide, iron oxide, cadmium sulfide, zinc sulfide, hydrogen peroxide, and ozone. delete The method of claim 1, wherein the polysaccharide is selected from chitin, chitosan, alginic acid, pectin, inulin, cellulose, gum, starch, glycogen, heparin, hyaluronic acid, pullulan, glucomannan and polygalactoseamine. A low molecular polysaccharide and a method for producing the oligosaccharide thereof. The organic solvent according to claim 1, wherein the organic solvent is methyl alcohol, ethyl alcohol, (iso) propyl alcohol, (t-, iso) butyl alcohol, acetone, ethyl methyl ketone, diethyl ketone, methyl propyl ketone, ethyl propyl ketone, dimethyl permamide (DMF), dimethylacetamide (DMAc), tetrahydrofuran (THF), dioxane, benzene, toluene, acetonitrile, xylene, hexane, ethylene glycol, aqueous acetic acid solution, aqueous hydrochloric acid solution, caustic soda solution A method for producing a low molecular polysaccharide and an oligosaccharide thereof characterized by mixing and using the above. The method of claim 1, wherein any one of air, oxygen, nitrogen, helium is selected and used to increase the decomposition efficiency of the polysaccharide solution or to prevent discoloration.