阅读说明：本技术 褐藻胶寡糖二酸的组合物 (Algin oligosacchride diacid composition ) 是由 耿美玉 张真庆 晋迎申 肖中平 丁健 于 2018-06-29 设计创作，主要内容包括：本发明涉及一种褐藻胶寡糖二酸组合,其包含具有式(Ⅲ)的甘露糖醛二酸或其药学上可接受的盐,其中n为选自1-9的整数,m选自0,1或2,m’选自0或1,并且其中n＝1-5的褐藻胶寡糖二酸的重量总和占所述组合总重量的60％以上,古罗糖醛酸的重量总和占所述组合物重量的50％以下。<Image he="195" wi="700" file="DDA0001716975190000011.GIF" imgContent="drawing" imgFormat="GIF" orientation="portrait" inline="no"></Image>(The present invention relates to an alginate-oligosaccharaide diacid combination comprising mannuronic diacid of formula (iii) or a pharmaceutically acceptable salt thereof, wherein n is an integer selected from 1 to 9, m is selected from 0, 1 or 2, m' is selected from 0 or 1, and wherein the sum of the weights of the alginate-oligosaccharaide diacids n ═ 1 to 5 represents more than 60% of the total weight of the combination, the sum of the weights of guluronic acid represents 50% of the weight of the composition% or less.)

1. An alginate oligosacchridic diacid composition comprising mannuronic and/or guluronic acid having formula (IV):

wherein n is an integer selected from 1 to 9, m is selected from 0, 1 or 2, m' is selected from 0 or 1,

and wherein the one or more of the one,

the sum of the weights of the alginate oligosaccharide diacids n-1-5 accounts for more than 60% of the total weight of the combination;

wherein the total weight of guluronic acid is less than 50% of the weight of the composition.

2. The algin oligosacchridic diacid composition according to claim 1, wherein the sum of the weights of algin oligosacchridic diacids m + m ═ 1 or 2 is not less than 50% by total weight of said combination, preferably from 60% to 90%, more preferably from 70% to 90%.

3. The alginate oligosacchridic diacid composition according to claim 2, wherein the sum of the weights of alginate oligosacchridic diacids m + m' ═ 1 is not less than 10% of the total weight of the combination, preferably between 30 and 40%.

4. The alginate oligosacchridic diacid composition according to claim 1, wherein the sum of the weights of the alginate oligosacchridic diacids m + m' ═ 2 is not less than 10% of the total weight of the combination, preferably between 30 and 50%.

5. The alginate oligosacchridic diacid composition of claim 1, wherein the sum of the weights of alginate oligosacchridic diacids n-1-5 is 80-95% of the total weight of the combination.

6. The alginate oligosacchridic diacid composition according to claim 1, wherein the sum of the weights of alginate oligosacchridic diacids n-1-3 is comprised between 20 and 70% of the total weight of the combination.

7. The alginate oligosacchridic diacid composition of claim 1, wherein the ratio of the sum of the weights of alginate oligosacchridic diacids n-1-3 to n-4-7 is between 1.0 and 3.5.

8. The alginate oligosacchridic diacid composition of claim 7, wherein the ratio of the sum of the weights of alginate oligosacchridic diacids n-1-3 to n-4-7 is between 1.0 and 3.0.

9. The alginate oligosaccharaide diacid composition according to claim 1, wherein the sum of the weights of guluronic acid is between 0.1 and 50%, preferably between 1 and 30% of the weight of the composition.

10. The alginate oligosacchridic diacid composition of any one of claims 1 to 9, wherein the weight percentage of alginate oligosacchridic diacid of each degree of polymerization in the composition is: 5-25% of disaccharide, 15-30% of trisaccharide, 15-28% of tetrasaccharide, 10-25% of pentasaccharide, 5-15% of hexasaccharide, 3-10% of heptasaccharide, 2-5% of octasaccharide, 1-5% of nonasaccharide and 1-5% of decasaccharide.

11. The alginate oligosacchridic diacid composition of claim 10, wherein the weight percentage of alginate oligosacchridic diacid of each degree of polymerization in the composition is: 10-20% of disaccharide, 18-30% of trisaccharide, 15-28% of tetrasaccharide, 15-20% of pentasaccharide, 5-10% of hexasaccharide, 3-5% of heptasaccharide, 2-3% of octasaccharide, 1-3% of nonasaccharide and 1-3% of decasaccharide.

12. The alginate oligosacchridic diacid composition of any one of claims 1 to 11, wherein the pharmaceutically acceptable salt is a sodium or potassium salt.

13. A pharmaceutical composition or health product comprising an effective amount of the alginate oligosacchride diacid composition of any one of claims 1 to 12 and, if necessary, a suitable carrier.

14. Use of the alginate-oligosacchride diacid composition of any one of claims 1 to 12 for the preparation of a medicament or health product for the treatment of diseases selected from senile dementia, parkinson's disease, inflammation, pain, diabetes or vascular dementia.

15. The algin oligosacchride diacid composition according to any one of claims 1-12 as a medicament or nutraceutical against senile dementia, parkinson's disease, inflammation, pain, diabetes or vascular dementia.

16. A method of treating a patient having a disease selected from the group consisting of: senile dementia, parkinson's disease, inflammation, pain, diabetes or vascular dementia, which comprises administering to a patient in need thereof an effective amount of an alginate oligosaccharaide diacid composition according to any one of claims 1 to 12.

Technical Field

The invention relates to an optimal composition of alginate-derived oligosaccharide diacid obtained by a biological activity screening method, which adopts an senile dementia animal model to evaluate the influence of different polymerization degrees and the matching ratio of the alginate-derived oligosaccharide on the biological activity. Finally, the composition with the optimal biological activity is screened and separated by an ultrafiltration membrane to prepare the required target substance.

Background

Alginate oligosaccharides have been widely regarded for their potential medicinal value. Alginate oligosaccharides are generally prepared from alginic acid through multiple steps.

In the algin oligose molecule, there are M segment formed by mannuronic acid (D-mannuronic acid) connected by beta-1, 4-glycosidic bond, G segment formed by guluronic acid (L-guluronic acid) connected by alpha-1, 4-glycosidic bond, and MG segment formed by the heterozygosis of these two sugars. The structural formulas of mannuronic acid and guluronic acid are shown as the following formulas (I) and (II):

the algin oligosaccharide has a structural formula shown as the following formula (III):

the M and G stages can be separated from the raw alginic acid. The general method can be described simply as: primarily degrading alginic acid to obtain mixed polysaccharide of polymannuronic acid and polyguluronic acid, and precipitating the mixed polysaccharide by an acid method to remove a certain amount of polyguluronic acid. See, for example, chinese patent application No.98806637.8 and CN02823707.2 for methods disclosed therein.

The preparation of oligomannuronic acid is as follows: heating the M-stage intermediate under acidic condition for further acidolysis to obtain small-stage mannuronic acid polymer with desired molecular weight range. In addition, there is a method of improving degradation efficiency by oxidative degradation and simultaneously oxidizing the reduced end into ring-opened glucaric acid, as described in chinese patent application 200580009396.5 (patent document 1) and US 8835403B2 (patent document 2) to gunn jasper et al. For ease of description, patent documents 1 and 2 are hereinafter collectively referred to as prior patents, and are incorporated herein by reference in their entirety.

The reaction process of mannuronic acid disclosed in the prior patent can be represented by the following reaction equation (V), that is, the C1-position aldehyde group of mannuronic acid at the reducing end of the oligomannuronic acid polysaccharide is oxidized into carboxyl group.

In the above oxidation conversion process, the commonly used oxidant is alkaline copper sulfate solution, i.e. film reagent, and the prior patent adopts the oxidation method, which specifically comprises the following steps: the reaction substrate polymannuronic acid, the above M-stage intermediate, was added to a copper sulfate solution under alkaline conditions and reacted in a boiling water bath for 15 minutes to 2 hours. The method is to use Cu²⁺Ions are oxidants to oxidize aldehyde groups, brick red cuprous oxide precipitates are generated in the reaction, and the reaction is commonly used for identifying reducing sugars.

The prior patent discloses that the manno-oligosacchride diacid has the effects of resisting Alzheimer's Disease (AD) and resisting diabetes, and the activity of the manno-oligosacchride diacid with the polymerization degree of 6 is optimal. The pathogenesis of Alzheimer's disease and type II diabetes is closely related to amyloid (beta-amyloid and amyloid). Amyloid aggregates to form protein oligomers, which further aggregate to form fibers. These aggregates are cytotoxic, induce oxidative damage to mitochondria in cells and trigger a cascade of inflammatory responses, which results in massive neuronal and beta cell damage and ultimately the development of alzheimer's disease and type II diabetes. The mannooligosaccharidic diacid targets amyloid and antagonizes cascade reaction induced by the amyloid, thereby having the effect of preventing and treating Alzheimer disease and type II diabetes.

In order to obtain the manno-oligosacchride diacid with anti-Alzheimer disease and anti-diabetes disclosed in the previous patent, guluronic acid in raw alginic acid is removed, and the content of guluronic acid in alginic acid is usually above 30% and up to about 70%, so the actual production cost is very high in order to obtain high-purity manno-oligosacchride diacid.

Disclosure of Invention

The first aspect of the present invention relates to a composition of alginate oligosaccharaide diacid comprising mannuronic and/or guluronic acid having formula (IV) or a pharmaceutically acceptable salt thereof:

wherein n is an integer selected from 1 to 9, m is selected from 0, 1 or 2, m' is selected from 0 or 1,

and wherein the one or more of the one,

the sum of the weights of the alginate oligosaccharide diacids n-1-5 accounts for more than 60% of the total weight of the combination; the total weight of guluronic acid diacid is less than 50% of the weight of the composition.

Another aspect of the present invention relates to a pharmaceutical composition or nutraceutical comprising the composition of alginate oligosaccharaide diacid as described above. Still other aspects of the invention relate to the use of compositions of alginate-derived oligosaccharaides for the treatment of a disease selected from the group consisting of senile dementia, parkinson's disease, inflammation, pain, diabetes and vascular dementia.

In particular, the algin oligosacchride diacid composition of the invention is a mixture of mannuronic acid and guluronic acid with different degrees of polymerization, the main components of which are M segment formed by mannuronic acid connected by beta-1, 4-glycosidic bond, G segment formed by guluronic acid connected by alpha-1, 4-glycosidic bond, and MG segment formed by the heterozygosis of the two sugars, the oligosaccharide has degree of polymerization of 2 to 10. Mannuronic acid is known to have some pharmacological activity against alzheimer's disease and diabetes. The most active sugars are the 5-8 sugars, especially the 6 sugars. However, the inventors have found that the oligogalactaric acid mixture of mannuronic acid and guluronic acid having a degree of polymerization of 2 to 10 also has pharmacological activity against alzheimer's disease and diabetes, provided that the guluronic acid content needs to be controlled within a certain range. That is, the alginate oligosaccharide diacid composition can be prepared by greatly reducing the production cost, is easier to realize in actual production and is easier to realize industrial mass production.

Drawings

FIG. 1 intermediate nuclear magnetic spectrum.

FIG. 2 is a mass spectrum of disaccharide, trisaccharide and tetrasaccharide in product A.

FIG. 3 is a mass spectrum of pentasaccharide, hexasaccharide and heptasaccharide in product A.

FIG. 4 is a mass spectrum of octasaccharides, nonasaccharides and decasaccharides in product A.

FIG. 5 shows the NMR spectrum of product A.

FIG. 6 shows the NMR spectrum of product B.

FIG. 7 shows the NMR spectrum of product C.

FIG. 8 shows the NMR spectrum of product D.

FIG. 9 shows the effect of different oligosaccharide compositions and hexasaccharide mannosyluronate on the number of platform crossings in AD animals; the numbers on the abscissa in the figure correspond to samples: i: a control group; ii: a model group; iii: product A; iv: product B; v: product C; vi: product D; vii: mannose aldehydic acid hexasaccharide.

FIG. 10 shows the effect of different oligosaccharide compositions and hexasaccharides of mannosylaldehydic acid on the swimming distance of AD animals; wherein the abscissa reference numerals are the same as those of fig. 9.

FIG. 11 shows the effect of different oligosaccharide compositions and hexasaccharides of mannosylaldehydic acid on the day 11 creep time of PD animals; wherein the abscissa reference numerals are the same as those of fig. 9.

FIG. 12 shows the effect of different oligosaccharide compositions and hexasaccharide mannosyluronate on day 11 latency in PD animals; wherein the abscissa reference numerals are the same as those of fig. 9.

FIGS. 13a and 13b show the therapeutic effect of different oligosaccharide compositions and hexa-mannose aldehydic acid on inflammatory bowel disease in mice; the abscissa of the figure is denoted by the same reference numerals as in fig. 9.

FIG. 14 shows the effect of different oligosaccharide compositions, hexasaccharide mannosyluronate on postprandial blood glucose in diabetic mice; the abscissa of the figure is denoted by the same reference numerals as in fig. 9.

FIG. 15 shows the effect of oligosaccharide compositions, hexamannose-aldehydic acid on latency of acetic acid-induced writhing response in mice; the numbers on the abscissa in the figure correspond to samples: i: a model group; ii: product A; iii: product B; iv: product C; v: product D; vi: mannose aldehydic acid hexasaccharide.

FIG. 16 shows the effect of different oligosaccharide compositions, hexa-mannose glucuronate on the number of writhing responses in mice to acetic acid; wherein the abscissa reference numerals are the same as those of fig. 15.

FIG. 17 shows the effect of different oligosaccharide compositions, hexa-mannose alduronate on the number of head curls in nitroglycerin migraine rats; wherein the abscissa reference numerals are the same as those of fig. 9.

FIG. 18 shows the effect of different oligosaccharide compositions, hexasaccharide mannosyluronate on the number of caudate c-fos positive cells in the trigeminal nerve bundle of rats electrically stimulated to migraine in trigeminal ganglia; wherein the abscissa reference numerals are the same as those of fig. 9.

FIG. 19 shows the effect of different oligosaccharide compositions, hexa-mannose-aldehydic acid on the latency of the darkness-avoidance experiment in bilateral common carotid artery ligation mice with vascular dementia; wherein the abscissa reference numerals are the same as those of fig. 9.

FIG. 20 shows the effect of different oligosaccharide compositions, hexa-mannose-aldehydic acid on the number of errors in the darkening-prevention experiment in bilateral common carotid artery ligated mice with vascular dementia; wherein the abscissa reference numerals are the same as those of fig. 9.

FIG. 21 shows the effect of different oligosaccharide compositions, hexa-mannose-aldaric acid on the escape latency of the water maze experiment in bilateral common carotid artery ligation mice with vascular dementia; wherein the abscissa reference numerals are the same as those of fig. 9.

FIG. 22 shows the effect of different oligosaccharide compositions, hexasaccharide mannosyluronate on the number of platform crossings in bilateral common carotid artery ligated vascular dementia mice; wherein the abscissa reference numerals are the same as those of fig. 9.

Detailed Description

Various aspects of the present invention will be described in detail below, but the present invention is not limited to these specific embodiments. Modifications and adaptations of the present invention that come within the spirit of the following disclosure may be made by those skilled in the art and are within the scope of the present invention.

Algin oligosaccharidic diacid composition

The first aspect of the present invention relates to a composition of alginate oligosaccharaide diacid comprising mannuronic and/or guluronic acid having formula (IV) or a pharmaceutically acceptable salt thereof:

wherein n is an integer selected from 1 to 9, m is selected from 0, 1 or 2, m' is selected from 0 or 1,

and wherein the one or more of the one,

the sum of the weights of the alginate oligosaccharide diacids n-1-5 accounts for more than 60% of the total weight of the combination;

wherein the total weight of guluronic acid diacid is less than 50% of the weight of the composition.

The algin oligosacchride diacid composition is a mixture of mannuronic acid and guluronic acid with different polymerization degrees, and the main components of the algin oligosacchride diacid composition are an oligosaccharide with the polymerization degree of 2-10, wherein the oligosaccharide mainly comprises an M segment formed by connecting mannuronic acid through beta-1, 4-glycosidic bonds, a G segment formed by connecting guluronic acid through alpha-1, 4-glycosidic bonds, and an MG segment formed by hybridizing the two saccharides. According to the prior application, it is known that mannuronic acid has pharmacological activity against Alzheimer's disease and diabetes, the most active sugar being the 5-8 sugar, in particular the 6 sugar. However, unlike the known prior art, the inventors found that the oligodiacid mixture of mannuronic and guluronic acids with a degree of polymerization of 2 to 10, also has pharmacological activity against alzheimer's disease and diabetes, but requires control of the guluronic acid content within a certain range.

In the actual preparation process, the content of guluronic acid in the original product after primary degradation of alginic acid is usually above 30%, and can reach about 70% at most, if guluronic acid should be separated and removed as much as possible in order to obtain high-activity manno-oligosacchride diacid according to the previous application. Based on the above findings of the inventors, it is not necessary to separate and remove guluronic acid from the degradation product. Further, the inventors have found that by controlling the conditions of the acid precipitation reaction to control the ratio of guluronic acid within a certain range, the activity of the resulting composition is achieved or even superior to that of the manno-oligosaccharidic diacid 6 saccharide disclosed in the prior application. And as guluronic acid is not needed to be removed as an impurity, the product yield is also obviously higher than that disclosed in the prior application theoretically, the production cost is greatly reduced, the emission of wastes is reduced, the method is easier to realize in actual production and is easier to realize industrialized mass production.

According to a preferred embodiment, the total weight of the alginate oligosacchridic diacid with n ═ 1 to 5 in the composition of the invention is 80 to 95% of the total weight of the combination, and the total weight of guluronic acid is less than 50% of the weight of the composition.

According to a preferred embodiment, the ratio of the sum of the weights of the oligoalginate-oligosacchridic diacids n-1 to 3 to the sum of the weights of the oligoalginate-oligosacchridic diacids n-4 to 7 in the alginate-oligosacchridic diacid composition of the invention is between 1.0 and 3.5.

According to a preferred embodiment, the sum of the weights of the alginate oligosacchride diacids m + m ═ 1 or 2 in the alginate oligosacchride diacid combination of the invention is not less than 50% of the total weight of the combination, preferably from 60% to 90%, more preferably from 70% to 90%. In particular, the sum of the weights of the alginate oligosacchride diacids m + m' ═ 1 in the alginate oligosacchride composition of the invention is not less than 10% of the total weight of the combination, preferably 30-40%. In another preferred embodiment, the sum of the weights of the alginate oligosacchridic diacids m + m' ═ 2 in the alginate oligosacchridic diacid composition of the invention is not less than 10% of the total weight of the combination, preferably between 30 and 50%.

According to a preferred embodiment, the sum of the weights of the alginate oligosacchride diacids in the alginate oligosacchride composition of the invention, n-1-5, is comprised between 80 and 95% of the total weight of the combination.

According to a preferred embodiment, the sum of the weights of the alginate oligosacchride diacids in the alginate oligosacchride composition of the invention n-1-3 is 20-70% of the total weight of the combination.

According to a preferred embodiment, the ratio of the sum of the weights of the alginate oligosacchride diacids n-1-3 to the sum of the weights of the alginate oligosacchride diacids n-4-7 in the alginate oligosacchride diacid composition of the invention is between 1.0 and 3.5, preferably between 1.0 and 3.0.

According to a preferred embodiment, the weight percentage of the alginate oligosaccharide diacid in each degree of polymerization in the alginate oligosaccharide diacid composition is as follows: 5-25% of disaccharide, 15-30% of trisaccharide, 15-28% of tetrasaccharide, 10-25% of pentasaccharide, 5-15% of hexasaccharide, 3-10% of heptasaccharide, 2-5% of octasaccharide, 1-5% of nonasaccharide and 1-5% of decasaccharide. In particular, the weight percentage of oligosaccharides in the combination is: 10-20% of disaccharide, 18-30% of trisaccharide, 15-28% of tetrasaccharide, 15-20% of pentasaccharide, 5-10% of hexasaccharide, 3-5% of heptasaccharide, 2-3% of octasaccharide, 1-3% of nonasaccharide and 1-3% of decasaccharide.

According to a preferred embodiment, the sum of the weights of guluronic acid in the alginate oligogalactaric acid composition of the invention is between 0.1 and 50%, preferably between 1 and 30% of the weight of the composition.

In the alginate oligosaccharide diacid composition, the pharmaceutically acceptable salt is sodium salt or potassium salt.

Preparation method of alginate oligosaccharide diacid composition

The preparation process of the algin oligose diacid of the invention is summarized as follows:

primarily degrading alginic acid to obtain mixed polysaccharide of polymannuronic acid and polyguluronic acid, and precipitating the mixed polysaccharide by an acid method to remove a certain amount of polyguluronic acid; the higher the pH control during the acid precipitation process, the higher the polyguluronic acid content of the resulting mixed polysaccharide. See, for example, chinese patent application No.98806637.8 and CN02823707.2 for methods disclosed therein. The mixed polysaccharide is subjected to oxidative degradation in the presence of an oxidant to obtain oxidized oligosaccharide with different polymerization degrees, and the oxidized oligosaccharide is characterized in that mannuronic acid or guluronic acid at the reducing end of the oligosaccharide is oxidized into saccharic acid with 3-6 carbons.

The oxidant particularly useful for the reaction of the present invention is ozone. In the reaction process, ozone is introduced into the solution containing the mixed polysaccharide, so that the oxidation degradation reaction of the sugar chain can be generated. The temperature at which the oxidative degradation step is carried out is preferably 0 to 70 deg.C, more preferably 10 to 45 deg.C. The above oxidative degradation step is carried out at a pH of 3 to 13, preferably 4 to 10, more preferably 6 to 8.

The oxidative degradation reaction using ozone in the present invention is the same as the oxidative degradation using basic copper sulfate (prior patent) or acid hydrolysis in the presence of hydrogen peroxide and sodium hypochlorite (chinese patent application 01107952.5) used in the prior art in that three methods can degrade sugar chains, except that the sugar chain reducing end structures of the degradation products are different, and the reducing end of the oxidative degradation product mannuronic acid or guluronic acid obtained in the present invention contains a diacid structure of 3 to 6 carbons. In addition, the process employed in the oxidative degradation step of the present invention also has other advantages: 1. the reaction condition is mild, and no special reaction condition is needed; 2. the used ozone can be prepared on the reaction site, and the transportation pressure is reduced in industrial production; 3. after the reaction, the ozone is automatically decomposed into oxygen, so that the harm of reaction reagent residue is avoided, and the environment is not polluted. The reaction process is shown in the following equation (VI):

the above reaction can be simplified as follows:

in the above reaction formula (VI) and the schematic diagram of the compound of the general formula (IV),

the oligosaccharide with m ═ 2 and m' ═ 1 is a sugar diacid with 6 terminal carbons;

oligosaccharides with m ═ 1 and m '1 or m ═ 2 and m' 0 are sugar diacids with a terminal 5 carbons;

oligosaccharides with m ═ 1 and m 'or with m ═ 0 and m' 1 are saccharic acids with a terminal 4 carbon number;

the oligosaccharide with m ═ 0 and m' ═ 0 is a sugar diacid with a terminal 3 carbons.

The total weight of the algin oligosacchride diacid with n-1-5 accounts for 80-95% of the total weight of the combination, and the total weight of the algin oligosacchride diacid with n-1-3 accounts for 20-70% of the total weight of the combination. Wherein the ratio of the sum of the weights of the alginate oligosaccharide diacids n-1-3 to the sum of the weights of the alginate oligosaccharide diacids n-4-7 is between 1.0 and 3.5, preferably between 1.0 and 3.0. The total weight of guluronic acid is less than 50% of the weight of the composition, preferably 0.1-50%, most preferably 1-30%.

In an exemplary embodiment, the method of the present invention comprises the steps of:

(1) preparing an alginate oligosaccharide diacid product:

preparation of mixed polysaccharide of polymannuronic acid and polyguluronic acid. As described above, the mixed polysaccharide of polymannuronic acid and polyguluronic acid, which are the starting materials used in the present invention, can be prepared by methods known in the art. Such as the methods disclosed in chinese patent application No.98806637.8 and CN 02823707.2. The general method can be described simply as: the polymannuronic acid and polyguluronic acid mixed polysaccharide can be obtained after alginic acid is primarily degraded, and after the mixed polysaccharide is precipitated by an acid method, the content of partial polyguluronic acid in the mixed polysaccharide can be adjusted to obtain the polymannuronic acid and polyguluronic acid mixed polysaccharide.

And (4) ozone oxidation degradation. Dissolving the mixed polysaccharide in a proper amount of water at room temperature or under a heating condition, stirring, continuously introducing ozone, and starting the reaction. The pH of the reaction can be adjusted to between 3 and 13, preferably between 4 and 10, more preferably between 6 and 8, by dropwise addition of dilute hydrochloric acid or dilute NaOH solution. The temperature is preferably from 0 to 70 ℃ and more preferably from 10 to 45 ℃. After the reaction is completed, the introduction of ozone is stopped, and the pH is adjusted to be neutral.

And (5) membrane separation and purification. The reaction product obtained above was prepared into a solution of about 10% concentration, and the solution was separated by a molecular interception membrane to remove degradation products below monosaccharide, and the non-permeated liquid was collected. The MWCO specification of the molecular cut-off membrane used is between 1000Da and 3000Da, preferably 2000 Da. Concentrating the collected liquid by a rotary evaporator, and drying in vacuum to obtain the mixture of the oligomeric alginate-derived oligosaccharides. The analysis shows that the products are all compositions which are disaccharide-decasaccharide oligosaccharide and the content of the oligosaccharide is in a certain proportion range.