CN113088546A - Preparation method of sargassum fusiforme polysaccharide and oligosaccharide - Google Patents
Preparation method of sargassum fusiforme polysaccharide and oligosaccharide Download PDFInfo
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Abstract
The invention relates to the technical field of sargassum fusiforme polysaccharide, and discloses a preparation method of sargassum fusiforme polysaccharide and oligosaccharide, wherein the preparation method of the sargassum fusiforme polysaccharide comprises the following steps: a1, preparing materials, namely preparing the following components in parts by weight: 100 parts of dried sargassum fusiforme, 1-3 parts of cellulase, 1-3 parts of pectinase, 1-3 parts of protease and 1-3 parts of alginic acid lyase; a2, performing enzymolysis, adding cellulase, pectinase, protease and water into dried sargassum fusiforme, wherein the ratio of feed to liquid is 1: 30-50, and performing enzymolysis at 50-60 deg.C and pH of 6.0-8.0 for 60-180 min; and A3, extracting, namely centrifuging the product obtained in the step A2, and extracting supernatant to obtain the sargassum fusiforme polysaccharide extracting solution. The preparation method of the sargassum fusiforme oligosaccharide comprises secondary enzymolysis. In conclusion, the beneficial effects of the invention are as follows: the preparation method for sargassum fusiforme polysaccharide by adopting composite enzymolysis has high preparation efficiency and low cost.
Description
Technical Field
The invention relates to the technical field of sargassum fusiforme polysaccharide, in particular to a preparation method of brown algae sulfated sargassum fusiforme polysaccharide and oligosaccharide which can be dissolved in water.
Background
The ocean is an extremely important component on the earth, and over twenty thousand organisms are bred, so that the ocean is a unique ecological system. Due to its specific ecological environment, many bioactive substances with novel structure completely different from terrestrial organisms are generated, and have important value in many fields. At present, the ocean has become one of the most promising areas. Carbohydrates are an extremely abundant active substance in the ocean, also called as saccharide compounds, and mainly comprise monosaccharide, oligosaccharide and polysaccharide; in recent years, research on the polysaccharide of the seaweed has been increasing due to the unique advantages of the seaweed polysaccharide in agricultural production, but the research on the polysaccharide of the sargassum fusiforme has been rarely reported.
Disclosure of Invention
The invention aims to provide a preparation method of sargassum fusiforme polysaccharide and oligosaccharide, which aims at solving the problems and has high preparation efficiency.
In order to achieve the aim, the invention discloses a preparation method of sargassum fusiforme polysaccharide, which comprises the following steps:
a1, preparing materials, namely preparing the following components in parts by weight: 100 parts of dried sargassum fusiforme, 1-3 parts of cellulase, 1-3 parts of pectinase, 1-3 parts of protease and 1-3 parts of alginic acid lyase.
Wherein, in the step A1, the dried sargassum fusiforme is prepared by drying sargassum fusiforme until the water content is less than or equal to 15%, grinding and sieving with a 60-mesh sieve. The sargassum fusiforme is in a powder shape, the contact area of the sargassum fusiforme and enzyme is enlarged, and the enzymolysis is facilitated.
Cyrtymenia Sparsa, Phaeophyta, Cystoseira, Fucales, Sargassaceae, Sargassum, Cyrtymenia Sparsa. Because of its unique ecological environment in the sea, many biologically active substances with novel structures, such as carbohydrates, are produced, which are completely different from terrestrial organisms.
Cellulase is one of enzymes, plays a biocatalytic role in decomposing cellulose, and is a protein capable of decomposing cellulose into polysaccharides.
Pectinase, which degrades the pectinase between cells and separates the cells from the tissue.
Proteases, proteolysis between cells to disperse cells.
Alginate lyase, which decomposes polysaccharides into oligosaccharides.
A2, performing enzymolysis, namely adding cellulase, pectinase, protease and water into dried sargassum fusiforme, wherein the feed-liquid ratio of the dried sargassum fusiforme, the cellulase, the pectinase, the protease and the water is 1: 30-50, and performing enzymolysis at 50-60 deg.C and pH of 6.0-8.0 for 60-180 min;
during the enzymolysis process, pectinase is matched with protease to separate the cells of the sargassum fusiforme, and then cellulose in the sargassum fusiforme cells is decomposed into polysaccharide under the action of cellulase.
A21, in order to verify the influence of cellulase, pectinase and protease on the polysaccharide extraction rate of the enzymolysis product, the polysaccharide extraction rate of the enzymolysis product is determined by a single-factor test under different addition amounts of cellulase, pectinase and protease.
Specifically, 1 g of dried sargassum fusiforme is added into a 50mL centrifuge tube for testing, and different addition amounts of the third enzyme are adopted under the condition that the addition amounts, the temperature, the pH value and the enzymolysis time of the two enzymes are consistent.
When determining the influence of cellulase, 1 part, 2 parts or 3 parts of cellulase, 2 parts of pectinase and 2 parts of protease are added, and enzymolysis is carried out for 120 min at 55 ℃ and pH 6.0.
When the influence of pectinase is measured, 2 parts of cellulase, 1 part, 2 parts or 3 parts of pectinase and 2 parts of protease are added for enzymolysis for 120 min at 55 ℃ and pH 6.0.
When the influence of protease is measured, 2 parts of cellulase, 2 parts of pectinase and 1 part, 2 parts or 3 parts of protease are added, and enzymolysis is carried out for 120 min at 55 ℃ and pH 6.0.
After the enzymolysis is finished, centrifuging for 4-6min at 3000-. Preferably, the rotating speed can be 3000 revolutions per minute, 4000 revolutions per minute or 5000 revolutions per minute; the centrifugation time is selected from 4 min, 5 min or 6 min.
A22, in order to verify the influence of the enzyme addition ratio, the temperature, the pH value and the enzymolysis time on the polysaccharide extraction rate of the enzymolysis product, the polysaccharide extraction rate of the enzymolysis product is determined by a single-factor test under different enzyme addition ratios, temperatures, pH values and enzymolysis times.
Specifically, 1 g of dried sargassum fusiforme is added into a 50mL centrifuge tube, and different values are adopted for the fourth factor under the condition that three factors of enzyme addition ratio, temperature, pH value and enzymolysis time are consistent.
When the influence of the enzyme addition ratio is measured, 2 parts of cellulase, 3 parts of pectinase and 1 part of protease, or 1 part of cellulase, 2 parts of pectinase and 3 parts of protease, or 3 parts of cellulase, 1 part of pectinase and 2 parts of protease are added, and enzymolysis is carried out for 120 min at 55 ℃ and pH 6.0.
When measuring the temperature influence, adding cellulase 2 parts, pectinase 3 parts, and protease 1 parts, and performing enzymolysis at 50 or 55 or 60 deg.C and pH6.0 for 120 min.
When measuring the influence of pH, 2 parts of cellulase, 3 parts of pectinase and 1 part of protease are added, and enzymolysis is carried out at 55 ℃ and pH of 6.0 or 6.0 for 120 min.
When the influence of enzymolysis time is measured, 2 parts of cellulase, 3 parts of pectinase and 1 part of protease are added, and enzymolysis is carried out for 60 or 120 or 180min at 55 ℃ and pH 6.0.
After the enzymolysis is finished, centrifuging for 4-6min at 3000-. Preferably, the rotating speed can be 3000 revolutions per minute, 4000 revolutions per minute or 5000 revolutions per minute; the centrifugation time is selected from 4 min, 5 min or 6 min.
A23, in order to verify the influence of the enzyme addition ratio, the temperature, the pH value and the enzymolysis time on the polysaccharide extraction rate of the enzymolysis product, the polysaccharide extraction rate of the enzymolysis product is determined by orthogonal tests under different enzyme addition ratios, temperatures, pH values and enzymolysis times.
Wherein the enzyme addition ratio is as follows: 2 parts of cellulase, 2 parts of pectinase and 1 part of protease, or 1 part of cellulase, 3 parts of pectinase and 2 parts of protease, or 3 parts of cellulase, 1 part of pectinase and 3 parts of protease;
wherein the temperature is: 50 or 55 or 60 ℃;
wherein the pH value is: 6.0 or 6.0;
wherein the enzymolysis time is as follows: 60 or 120 or 180 min.
The single-factor test or the orthogonal test is repeated for at least 3 times respectively, and the result is averaged to ensure the accuracy of the result.
A3, extracting, namely extracting a sargassum fusiforme polysaccharide extracting solution.
Specifically, the product obtained in the step A2 is centrifuged for 4-6min at the rotating speed of 3000-5000 r/min, and the supernatant is extracted to be used as the sargassum fusiforme polysaccharide extracting solution. Preferably, the rotating speed can be 3000 revolutions per minute, 4000 revolutions per minute or 5000 revolutions per minute; the centrifugation time is selected from 4 min, 5 min or 6 min.
A preparation method of Cyrtymenia Sparsa oligosaccharide comprises the following steps:
and B1, secondary enzymolysis, adding alginate lyase into the product of the preparation method, and carrying out enzymolysis for 60-180min at 50-60 ℃ and pH of 6.0-8.0 to obtain the sargassum fusiforme oligosaccharide degradation solution.
In the secondary enzymolysis process, under the action of alginate lyase, the polysaccharide in the sargassum fusiforme polysaccharide extract is decomposed into oligosaccharide.
And B2, freeze-drying, namely adding the product obtained in the step B1 into a freeze dryer, and drying until the water content is less than or equal to 15%. Convenient to store and transport.
In conclusion, the beneficial effects of the invention are as follows: the preparation method for sargassum fusiforme polysaccharide by adopting composite enzymolysis has high preparation efficiency and low cost.
Drawings
FIG. 1 shows the polysaccharide extraction rate of the enzyme-hydrolyzed product of step A21 of the method for preparing sargassum fusiforme polysaccharide of the present invention (55 ℃, pH6.0, enzymolysis for 120 min) with different addition amounts of cellulase, pectinase and protease;
FIG. 2 is a bar graph of polysaccharide extraction rate of the enzyme-hydrolyzed product measured by single factor test in step A21 of the preparation method of sargassum fusiforme polysaccharide of the present invention at different addition amounts of cellulase, pectinase and protease (55 deg.C, pH6.0, enzymolysis time 120 min);
FIG. 3 shows the polysaccharide extraction rate of the enzyme-hydrolyzed product of step A22 of the method for preparing sargassum fusiforme polysaccharide according to the present invention, which is determined by single factor test under different enzyme addition ratios, temperatures, pH values and enzymatic hydrolysis times;
FIG. 4 is a bar graph of polysaccharide extraction rates of enzymatic hydrolysis products at different enzyme addition ratios, temperatures, pH values and enzymatic hydrolysis times measured by single factor tests in step A22 of a method for preparing sargassum fusiforme polysaccharide according to the present invention;
FIG. 5 shows the polysaccharide extraction rate of the enzyme-hydrolyzed product of the preparation method of sargassum fusiforme polysaccharide of the present invention, which is determined by the orthogonal test in step A23, under different enzyme addition ratios, temperatures, pH values and enzymatic hydrolysis times.
Detailed Description
The invention is further described with reference to the following figures and detailed description:
a preparation method of sargassum fusiforme polysaccharide comprises the following steps:
a1, preparing materials, namely preparing the following components in parts by weight: 100 parts of dried sargassum fusiforme, 1-3 parts of cellulase, 1-3 parts of pectinase, 1-3 parts of protease and 1-3 parts of alginic acid lyase.
Wherein, in the step A1, the dried sargassum fusiforme is prepared by drying sargassum fusiforme until the water content is less than or equal to 15%, grinding and sieving with a 60-mesh sieve. The sargassum fusiforme is in a powder shape, the contact area of the sargassum fusiforme and enzyme is enlarged, and the enzymolysis is facilitated.
Cyrtymenia Sparsa, Phaeophyta, Cystoseira, Fucales, Sargassaceae, Sargassum, Cyrtymenia Sparsa. Because of its unique ecological environment in the sea, many biologically active substances with novel structures, such as carbohydrates, are produced, which are completely different from terrestrial organisms.
Cellulase is one of enzymes, plays a biocatalytic role in decomposing cellulose, and is a protein capable of decomposing cellulose into polysaccharides.
Pectinase, which degrades the pectinase between cells and separates the cells from the tissue.
Proteases, proteolysis between cells to disperse cells.
Alginate lyase, which decomposes polysaccharides into oligosaccharides.
A2, performing enzymolysis, namely adding cellulase, pectinase, protease and water into dried sargassum fusiforme, wherein the feed-liquid ratio of the dried sargassum fusiforme, the cellulase, the pectinase, the protease and the water is 1: 30-50, and performing enzymolysis at 50-60 deg.C and pH of 6.0-8.0 for 60-180 min;
during the enzymolysis process, pectinase is matched with protease to separate the cells of the sargassum fusiforme, and then cellulose in the sargassum fusiforme cells is decomposed into polysaccharide under the action of cellulase.
A21, in order to verify the influence of cellulase, pectinase and protease on the polysaccharide extraction rate of the enzymolysis product, the polysaccharide extraction rate of the enzymolysis product is determined by a single-factor test under different addition amounts of cellulase, pectinase and protease.
Specifically, 1 g of dried sargassum fusiforme is added into a 50mL centrifuge tube for testing, and different addition amounts of the third enzyme are adopted under the condition that the addition amounts, the temperature, the pH value and the enzymolysis time of the two enzymes are consistent.
When determining the influence of cellulase, 1 part, 2 parts or 3 parts of cellulase, 2 parts of pectinase and 2 parts of protease are added, and enzymolysis is carried out for 120 min at 55 ℃ and pH 6.0.
When the influence of pectinase is measured, 2 parts of cellulase, 1 part, 2 parts or 3 parts of pectinase and 2 parts of protease are added for enzymolysis for 120 min at 55 ℃ and pH 6.0.
When the influence of protease is measured, 2 parts of cellulase, 2 parts of pectinase and 1 part, 2 parts or 3 parts of protease are added, and enzymolysis is carried out for 120 min at 55 ℃ and pH 6.0.
After the enzymolysis is finished, centrifuging for 4-6min at the rotating speed of 3000-. Preferably, the rotating speed can be 3000 revolutions per minute, 4000 revolutions per minute or 5000 revolutions per minute; the centrifugation time is selected from 4 min, 5 min or 6 min.
A22, in order to verify the influence of the enzyme addition ratio, the temperature, the pH value and the enzymolysis time on the polysaccharide extraction rate of the enzymolysis product, the polysaccharide extraction rate of the enzymolysis product is determined by a single-factor test under different enzyme addition ratios, temperatures, pH values and enzymolysis times.
Specifically, 1 g of dried sargassum fusiforme is added into a 50mL centrifuge tube, and different values are adopted for the fourth factor under the condition that three factors of enzyme addition ratio, temperature, pH value and enzymolysis time are consistent.
When the influence of the enzyme addition ratio is measured, 2 parts of cellulase, 3 parts of pectinase and 1 part of protease, or 1 part of cellulase, 2 parts of pectinase and 3 parts of protease, or 3 parts of cellulase, 1 part of pectinase and 2 parts of protease are added, and enzymolysis is carried out for 120 min at 55 ℃ and pH 6.0.
When measuring the temperature influence, adding cellulase 2 parts, pectinase 3 parts, and protease 1 parts, and performing enzymolysis at 50 or 55 or 60 deg.C and pH6.0 for 120 min.
When measuring the influence of pH, 2 parts of cellulase, 3 parts of pectinase and 1 part of protease are added, and enzymolysis is carried out at 55 ℃ and pH of 6.0 or 6.0 for 120 min.
When the influence of enzymolysis time is measured, 2 parts of cellulase, 3 parts of pectinase and 1 part of protease are added, and enzymolysis is carried out for 60 or 120 or 180min at 55 ℃ and pH 6.0.
After the enzymolysis is finished, centrifuging for 4-6min at 3000-. Preferably, the rotating speed can be 3000 revolutions per minute, 4000 revolutions per minute or 5000 revolutions per minute; the centrifugation time is selected from 4 min, 5 min or 6 min.
A23, in order to verify the influence of the enzyme addition ratio, the temperature, the pH value and the enzymolysis time on the polysaccharide extraction rate of the enzymolysis product, the polysaccharide extraction rate of the enzymolysis product is determined by orthogonal tests under different enzyme addition ratios, temperatures, pH values and enzymolysis times.
Wherein the enzyme addition ratio is as follows: 2 parts of cellulase, 2 parts of pectinase and 1 part of protease, or 1 part of cellulase, 3 parts of pectinase and 2 parts of protease, or 3 parts of cellulase, 1 part of pectinase and 3 parts of protease;
wherein the temperature is: 50 or 55 or 60 ℃;
wherein the pH value is: 6.0 or 6.0;
wherein the enzymolysis time is as follows: 60 or 120 or 180 min.
The single-factor test or the orthogonal test is repeated for at least 3 times respectively, and the result is averaged to ensure the accuracy of the result.
A3, extracting, namely extracting a sargassum fusiforme polysaccharide extracting solution.
Specifically, the product obtained in the step A2 is centrifuged for 4-6min at 3000-5000 rpm, and the supernatant is extracted to be used as the sargassum fusiforme polysaccharide extract. Preferably, the rotating speed can be 3000 revolutions per minute, 4000 revolutions per minute or 5000 revolutions per minute; the centrifugation time is selected from 4 min, 5 min or 6 min.
A preparation method of Cyrtymenia Sparsa oligosaccharide comprises the following steps:
and B1, secondary enzymolysis, adding alginate lyase into the product of the preparation method, and carrying out enzymolysis for 60-180min at 50-60 ℃ and pH of 6.0-8.0 to obtain the sargassum fusiforme oligosaccharide degradation solution.
In the secondary enzymolysis process, under the action of alginate lyase, the polysaccharide in the sargassum fusiforme polysaccharide extract is decomposed into oligosaccharide.
And B2, freeze-drying, namely adding the product obtained in the step B1 into a freeze dryer, and drying until the water content is less than or equal to 15%. Convenient to store and transport.
Specifically, example 1: when determining the influence of cellulase, 1 part of cellulase, 2 parts of pectinase and 2 parts of protease are added, and enzymolysis is carried out for 120 min at 55 ℃ and pH 6.0.
Specifically, example 2: when determining the influence of cellulase, adding 2 parts of cellulase, 2 parts of pectinase and 2 parts of protease, and performing enzymolysis at 55 ℃ and pH6.0 for 120 min.
Specifically, example 3: when determining the influence of cellulase, adding 3 parts of cellulase, 2 parts of pectinase and 2 parts of protease, and performing enzymolysis at 55 ℃ and pH6.0 for 120 min.
Specifically, example 4: when the influence of pectinase is measured, 2 parts of cellulase, 1 part of pectinase and 2 parts of protease are added, and enzymolysis is carried out for 120 min at 55 ℃ and pH 6.0.
Specifically, example 5: when the influence of pectinase is measured, 2 parts of cellulase, 2 parts of pectinase and 2 parts of protease are added, and enzymolysis is carried out for 120 min at 55 ℃ and pH 6.0.
Specifically, example 6: when the influence of pectinase is measured, 2 parts of cellulase, 3 parts of pectinase and 2 parts of protease are added, and enzymolysis is carried out for 120 min at 55 ℃ and pH 6.0.
Specifically, example 7: when the influence of protease is measured, 2 parts of cellulase, 2 parts of pectinase and 1 part of protease are added, and enzymolysis is carried out for 120 min at 55 ℃ and pH 6.0.
Specifically, example 8: when the influence of the protease is measured, 2 parts of cellulase, 2 parts of pectinase and 2 parts of protease are added, and enzymolysis is carried out for 120 min at 55 ℃ and pH 6.0.
Specifically, example 9: when the influence of protease is measured, 2 parts of cellulase, 2 parts of pectinase and 3 parts of protease are added, and enzymolysis is carried out for 120 min at 55 ℃ and pH 6.0.
The test result is shown in figure 1, under the single-factor test condition, when 2 parts of cellulase is added, the extraction rate is highest; when 3 parts of pectinase is added, the extraction rate is highest; when 1 part of protease is added, the extraction rate is highest.
Specifically, example 10: adding 2 parts of cellulase, 3 parts of pectinase and 1 part of protease, and performing enzymolysis at 55 deg.C and pH of 6.0 for 120 min.
Specifically, example 11: adding 1 part of cellulase, 2 parts of pectinase and 3 parts of protease, and performing enzymolysis at 55 deg.C and pH of 6.0 for 120 min.
Specifically, example 12: adding cellulase 3 parts, pectase 1 part, and protease 2 parts, and performing enzymolysis at 55 deg.C and pH6.0 for 120 min.
Specifically, example 13: when measuring the temperature influence, 2 parts of cellulase, 3 parts of pectinase and 1 part of protease are added, and enzymolysis is carried out at 50 ℃ and pH6.0 for 120 min.
Specifically, example 14: when measuring the temperature influence, 2 parts of cellulase, 3 parts of pectinase and 1 part of protease are added, and enzymolysis is carried out for 120 min at 55 ℃ and pH 6.0.
Specifically, example 15: when measuring the temperature influence, 2 parts of cellulase, 3 parts of pectinase and 1 part of protease are added, and enzymolysis is carried out at 60 ℃ and pH6.0 for 120 min.
Specifically, example 16: when the influence of the pH value is measured, 2 parts of cellulase, 3 parts of pectinase and 1 part of protease are added, and enzymolysis is carried out for 120 min at 55 ℃ and pH 6.0.
Specifically, example 17: when the influence of the pH value is measured, 2 parts of cellulase, 3 parts of pectinase and 1 part of protease are added, and enzymolysis is carried out for 120 min at 55 ℃ and pH 6.0.
Specifically, example 18: when the influence of the pH value is measured, 2 parts of cellulase, 3 parts of pectinase and 1 part of protease are added, and enzymolysis is carried out for 120 min at 55 ℃ and pH 6.0.
Specifically, example 19: when the influence of enzymolysis time is measured, 2 parts of cellulase, 3 parts of pectinase and 1 part of protease are added, and enzymolysis is carried out for 60 min at 55 ℃ and pH 6.0.
Specifically, example 20: when the influence of enzymolysis time is measured, 2 parts of cellulase, 3 parts of pectinase and 1 part of protease are added, and enzymolysis is carried out for 120 min at 55 ℃ and pH 6.0.
Specifically, example 21: when the influence of enzymolysis time is measured, 2 parts of cellulase, 3 parts of pectinase and 1 part of protease are added, and enzymolysis is carried out for 180min at 55 ℃ and pH 6.0.
The test result is shown in fig. 2, under the single-factor test condition, when the enzyme addition ratio is 2:3:1, the extraction rate is highest; when the temperature is 55 ℃, the extraction rate is highest; when the pH value is 6.0, the extraction rate is highest; the extraction rate is highest when the enzymolysis time is 120 min.
The level of orthogonality and the factors are explained.
1: adding enzyme at 50 deg.C and pH6.0 at a ratio of 2:3:1, and performing enzymolysis for 60 min;
2: performing enzymolysis at 55 deg.C and pH6.0 for 120 min at a ratio of 1:2: 3;
3: the enzyme addition ratio is 3:1:2, 60 deg.C, pH6.0, and enzymolysis for 180 min.
The test results are shown in fig. 3.
According to the results of orthogonal experiments, the four factors influence the extraction efficiency of the sargassum fusiforme polysaccharide in the sequence of extraction time, PH, temperature and enzyme quantity ratio. Wherein k3 is the largest among the three average values of the enzyme quantity ratio, and the average values of the extraction time k3, the temperature k1 and the pH k2 are the largest in the same way. Therefore, the optimal extraction process of the sargassum fusiforme polysaccharide comprises the following steps: the enzyme amount ratio is cellulase: and (3) pectinase: papain 2:3:1, extracting at 50 deg.C and pH6.0 for 180 min.
The above description is only a preferred embodiment of the present invention, and it should be noted that, for those skilled in the art, various modifications and substitutions can be made without departing from the technical principle of the present invention, and these modifications and substitutions should also be regarded as the protection scope of the present invention.
Claims (10)
1. A preparation method of sargassum fusiforme polysaccharide is characterized by comprising the following steps:
a1, preparing materials, namely preparing the following components in parts by weight: 100 parts of dried sargassum fusiforme, 1-3 parts of cellulase, 1-3 parts of pectinase, 1-3 parts of protease and 1-3 parts of alginic acid lyase;
a2, performing enzymolysis, namely adding cellulase, pectinase, protease and water into dried sargassum fusiforme, wherein the feed-liquid ratio of the dried sargassum fusiforme, the cellulase, the pectinase, the protease and the water is 1: 30-50, and performing enzymolysis at 50-60 deg.C and pH of 6.0-8.0 for 60-180 min;
and A3, extracting, namely centrifuging the product obtained in the step A2, and extracting supernatant to obtain the sargassum fusiforme polysaccharide extracting solution.
2. The method of claim 1, wherein the dried sargassum fusiforme is prepared by drying sargassum fusiforme until the water content is less than or equal to 15%, grinding, and sieving with a 60-mesh sieve in step A1.
3. The method of claim 1, wherein the polysaccharide extraction rate of the enzyme-hydrolyzed product is determined by a one-way test in step A2 at different addition levels of cellulase, pectinase and protease.
4. The method of claim 3, wherein the polysaccharide extraction rate of the enzyme-hydrolyzed product is determined by a one-way test in step A2 at different enzyme addition ratio, temperature, pH and enzymolysis time.
5. The method of claim 4, wherein the polysaccharide extraction rate of the enzyme-hydrolyzed product is determined by orthogonal test at step A2 under different enzyme ratio, temperature, pH value and enzymolysis time.
6. The method of preparing sargassum fusiforme polysaccharide as claimed in any one of claims 3 to 5, wherein 1 g of dried sargassum fusiforme is added into a 50mL centrifuge tube for testing in a one-way test or an orthogonal test.
7. The method of preparing sargassum fusiforme polysaccharide as claimed in any one of claims 3 to 5, wherein the single factor test or the orthogonal test is repeated at least 3 times each, and the results are averaged.
8. The method as claimed in claim 1, wherein the product of step A2 is centrifuged at 3000-5000 rpm/min for 4-6min in step A3.
9. A preparation method of sargassum fusiforme oligosaccharide is characterized by comprising the following steps:
and B1, secondary enzymolysis, adding alginate lyase into the product of the preparation method of claim 1, and carrying out enzymolysis for 60-180min at 50-60 ℃ and pH of 6.0-8.0 to obtain the sargassum fusiforme oligosaccharide degradation solution.
10. The method of claim 9, further comprising the steps of:
and B2, freeze-drying, namely adding the product obtained in the step B1 into a freeze dryer, and drying until the water content is less than or equal to 15%.
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