CN108484787B - Preparation method of moringa oleifera selenium-rich polysaccharide - Google Patents
Preparation method of moringa oleifera selenium-rich polysaccharide Download PDFInfo
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- 229910052711 selenium Inorganic materials 0.000 title claims abstract description 144
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- 238000012545 processing Methods 0.000 claims description 15
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- CIWBSHSKHKDKBQ-JLAZNSOCSA-N Ascorbic acid Chemical compound OC[C@H](O)[C@H]1OC(=O)C(O)=C1O CIWBSHSKHKDKBQ-JLAZNSOCSA-N 0.000 description 2
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- FPIPGXGPPPQFEQ-UHFFFAOYSA-N 13-cis retinol Natural products OCC=C(C)C=CC=C(C)C=CC1=C(C)CCCC1(C)C FPIPGXGPPPQFEQ-UHFFFAOYSA-N 0.000 description 1
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- ZZZCUOFIHGPKAK-UHFFFAOYSA-N D-erythro-ascorbic acid Natural products OCC1OC(=O)C(O)=C1O ZZZCUOFIHGPKAK-UHFFFAOYSA-N 0.000 description 1
- 241000196324 Embryophyta Species 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
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- FPIPGXGPPPQFEQ-OVSJKPMPSA-N all-trans-retinol Chemical compound OC\C=C(/C)\C=C\C=C(/C)\C=C\C1=C(C)CCCC1(C)C FPIPGXGPPPQFEQ-OVSJKPMPSA-N 0.000 description 1
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- WIGCFUFOHFEKBI-UHFFFAOYSA-N gamma-tocopherol Natural products CC(C)CCCC(C)CCCC(C)CCCC1CCC2C(C)C(O)C(C)C(C)C2O1 WIGCFUFOHFEKBI-UHFFFAOYSA-N 0.000 description 1
- 238000009775 high-speed stirring Methods 0.000 description 1
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Classifications
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08B—POLYSACCHARIDES; DERIVATIVES THEREOF
- C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
- C08B37/0003—General processes for their isolation or fractionation, e.g. purification or extraction from biomass
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08B—POLYSACCHARIDES; DERIVATIVES THEREOF
- C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
Abstract
The invention discloses a preparation method of moringa oleifera selenium-rich polysaccharide, which comprises the following steps: freezing and drying moringa leaves at low temperature, crushing the moringa leaves by using liquid nitrogen, sieving the crushed moringa leaves by using a 100-mesh sieve, adding absolute ethyl alcohol, stirring the mixture for 20 to 30 minutes, obtaining filter residue through an ultrafiltration membrane, recovering filtrate from the filter residue, mixing the filter residue with deionized water and yeast selenium, standing the mixture for 1 to 2 hours at the temperature of between 40 and 50 ℃, placing the mixture in a microwave, performing circulating treatment at the temperature of between 20 ℃ below zero and 10 ℃ below zero, performing centrifugal extraction on supernatant, adding the absolute ethyl alcohol into the supernatant for multiple times for precipitation, filtering the precipitate through the ultrafiltration membrane to obtain a precipitate, and performing vacuum freeze-drying on the precipitate to obtain; the added selenium yeast is placed in microwaves and treated in a circulating mode at low temperature, so that the combination of polysaccharide and selenium yeast can be obviously improved to form stable selenium-rich polysaccharide, the extraction amount of the selenium-rich polysaccharide can be obviously improved, and the activity of the selenium-rich polysaccharide is not damaged when the selenium yeast is extracted at relatively low temperature.
Description
Technical Field
The invention relates to the technical field of polysaccharide extraction, and particularly relates to a preparation method of moringa oleifera selenium-rich polysaccharide.
Background
Moringa oleifera, which is originally produced in northern India, is also planted in China at present, and has the functions of improving nutrition and dietotherapy health care as vegetables and food; also widely applied to the aspects of medicine, health care and the like, and is praised as 'living tree' and 'diamond in plants'. In 2012, the moringa oleifera is approved as a new resource food by the Ministry of health of China, and the seeds and leaves of the moringa oleifera contain rich nutritional ingredients. Sunny Sun (L.S. Ching) et al published in two, zero and one year, the vitamin E content in fresh leaves of 100 g Indian traditional moringa oleifera was about 9 mg, and the content in dried leaves was about 16.2 mg. According to calculation, only three tablespoons (about 25 g) of the moringa oleifera leaf dry powder contain 270% of vitamin A, 42% of protein, 125% of calcium, 70% of iron and 22% of vitamin C required by infants every day. Moringa leaves and pods also help oneself and fetus or baby to maintain health for pregnant and lactating women, supplying large amounts of iron, protein, copper, sulfur and vitamin B. The moringa oleifera contains abundant polysaccharides, the moringa oleifera polysaccharides are one of main effective components of the moringa oleifera and are mainly distributed in moringa oleifera leaves and moringa oleifera seeds, and the moringa oleifera polysaccharide compounds have the effects of reducing blood sugar and blood fat, reducing blood pressure, resisting oxidation and viruses, improving immunity and the like, so the moringa oleifera polysaccharides also become a hotspot of research of people.
Selenium is a trace nutrient element necessary for human body, has various functions of resisting cancer, improving immunity, delaying senility and the like, and is known as 'vital fire', 'king of resisting cancer' and the like. In 1973, the WHO and the international nutrition organization regulated selenium to be a micronutrient essential to humans and animals. At present, it is proved that selenium is an important component for forming more than 30 kinds of selenium-containing proteins and selenium-containing enzymes in mammals, such as glutathione peroxidase, thioredoxin, iodothyronine, deiodinase and the like, and has various biological functions of resisting oxidation, resisting cancer, improving the immunity of organisms and the like. There are over 40 countries and regions around the world, with about 5-10 million people lacking selenium. Two thirds of areas in China belong to selenium deficiency areas, and 30 percent of the areas are serious selenium deficiency areas. The selenium intake of human bodies is recommended to be 50-200 mug/d by the Chinese society of nutrition, while the selenium intake of urban residents in China is only 26-32 mug/d, and is seriously low. The biological conversion method is used for converting inorganic selenium into organic selenium, so that the selenium content in food is increased, and the method is an important way for improving selenium nutrition of human bodies. The selenium source is supplemented through diet, the immunity of the human body is improved, the health and the long life of people are realized, and the selenium-enriched health-care food is a national problem which is focused on by all countries in the world. The selenium-rich agricultural product is a high-value agricultural product with a health care function, and can be used for scientifically supplementing selenium. The selenium is scientifically supplemented by selenium-rich agricultural products, and the per-capita selenium intake of residents in selenium-deficient countries such as British, Finland, Australia and the like is effectively improved.
At present, selenium-rich moringa tea and selenium-rich moringa related food gradually appear, the research on moringa selenium-rich polysaccharide is very few, a patent application with the patent application number of 201610279274.8 discloses a biotransformation high-selenium polysaccharide moringa health tea, which mainly utilizes strain inoculation culture to improve the medium selenium polysaccharide of the moringa health tea, the raw materials of the moringa health tea comprise tea components and organic selenium, the tea components comprise 10-30% of moringa, 5-15% of moringa branch leaf powder, 2-8% of ganoderma spore powder and 50-70% of tea, and 100-organic selenium and 150 micrograms of tea are added into each gram of the tea components, and the production process comprises the following steps: weighing the raw materials in the proportion, mixing and stirring, and adjusting the water content to 55-60%; bottling or bagging, and sterilizing; inoculating strains and culturing; the method is simple and uncontrollable, the conversion rate of the selenium-rich polysaccharide is uncontrollable, and the extraction and purification of the selenium-rich polysaccharide compound in the moringa oleifera are not mentioned.
Disclosure of Invention
The invention aims to provide a preparation method of moringa oleifera selenium-rich polysaccharide, which is characterized in that moringa oleifera leaves are used as a raw material to extract the selenium-rich polysaccharide, absolute ethyl alcohol can quickly and effectively remove impurities dissolved in the ethanol in moringa oleifera leaf powder in a high-pressure quick stirring process, and added yeast selenium is placed in microwaves and is treated in a circulating mode at the temperature of-20 to-10 ℃ to remarkably improve the combination of the polysaccharide and the yeast selenium to form stable selenium-rich polysaccharide, so that the extraction amount of the selenium-rich polysaccharide can be remarkably improved finally, and the activity of the selenium-rich polysaccharide is not damaged during extraction at a relatively low temperature.
The technical scheme provided by the invention is as follows:
a preparation method of moringa oleifera selenium-rich polysaccharide comprises the following steps:
firstly, selecting moringa leaves as a raw material, wherein the moringa leaves can comprise old leaves and tender leaves, the proportion can be selected at will, freezing and drying the moringa leaves at a low temperature, crushing the moringa leaves by using liquid nitrogen, sieving the crushed moringa leaves by using a 100-mesh sieve, adding absolute ethyl alcohol, stirring for 20-30 minutes, obtaining filter residue by using an ultrafiltration membrane, recovering filtrate, and removing impurities dissolved in the absolute ethyl alcohol from moringa leaf powder;
step two, mixing the filter residue with deionized water and yeast selenium according to the weight ratio of 10:10-50:0.01-0.02, placing the mixture at the temperature of 40-50 ℃ for 1-2 hours, placing the mixture in microwaves and performing circulating treatment at the temperature of-20 to-10 ℃, centrifuging and extracting supernatant, placing the supernatant in microwaves and performing circulating treatment at the temperature of-20 to-10 ℃ to obviously improve the selenium content in the polysaccharide, thereby finally obviously improving the extraction amount of the selenium-rich polysaccharide and extracting the selenium-rich polysaccharide at a relatively low temperature without damaging the activity of the selenium-rich polysaccharide;
and step three, adding absolute ethyl alcohol into the supernatant obtained in the step two for multiple times for precipitation, filtering through an ultrafiltration membrane to obtain a precipitate, and performing vacuum freeze-drying on the precipitate to obtain the moringa oleifera selenium-rich polysaccharide.
Preferably, the weight ratio of the absolute ethyl alcohol to the moringa oleifera leaf powder is 1-5: 1.
Preferably, the absolute ethyl alcohol added in the step one is placed in a reaction kettle, stirred for 20-30 minutes at the temperature of 0-4 ℃ and the pressure of 10-20Mpa at the stirring speed of 2000-5000 rpm, wherein the weight ratio of the absolute ethyl alcohol to the moringa oleifera leaf powder is 1:1, and the impurities dissolved in the ethanol in the moringa oleifera leaf powder can be quickly dissolved out by utilizing the conditions of low temperature, high pressure and quick stirring, so that the impurity removal efficiency is improved, and the consumption of the absolute ethyl alcohol can be reduced.
Preferably, the filter residue in the first step is further added with anhydrous ethanol and anhydrous ethyl ether for at least 3 times of cleaning and filtering, wherein the weight ratio of the anhydrous ethanol to the anhydrous ethyl ether is 1:1-2, so that the extracted selenium-enriched polysaccharide can reduce impurities, and the polysaccharide and the yeast selenium can be combined to form stable selenium-enriched polysaccharide.
Preferably, the cyclic treatment in microwave and at-20 to-10 ℃ in the second step specifically comprises: treating in 500-2000MHz microwave for 10-30 sec, and then at-20 deg.c to-10 deg.c for 10-30 min for 3-5 times.
Preferably, the method further comprises the following steps: and (2) placing the mixture in microwave and carrying out cyclic treatment at the temperature of between 20 ℃ below zero and 10 ℃ below zero, adding 3-6% by weight of trichloroacetic acid-acetone, treating the mixture for 0.5-1 hour at the temperature of between 0 ℃ and 4 ℃ and under the pressure of between 1 and 5Mpa, centrifuging and filtering the mixture at 500 r/min, extracting the trichloroacetic acid-acetone to remove the trichloroacetic acid-acetone to obtain a polysaccharide extracting solution, deproteinizing the polysaccharide extracting solution by adopting low-concentration trichloroacetic acid-acetone under the condition of low temperature and relative high pressure, so that the removal efficiency of the protein can be improved, the removal time of the protein can be shortened, and the damage of an organic solvent to the activity of the selenium-enriched.
Preferably, after 5.5 weight percent of trichloroacetic acid-acetone is added and uniformly stirred, the mixture is placed in a reaction kettle, the temperature is adjusted to be 0-4 ℃ and the pressure is 1-5Mpa for stirring treatment for 5-10 minutes, the temperature is not changed and the pressure is kept constant, the mixture is placed for 10-20 minutes, circulation treatment is carried out, the treatment is finished after the cumulative time of stirring treatment is 40-60 minutes under the temperature of 0-4 ℃ and the pressure is 1-5Mpa, the mixture is placed in a centrifuge for 500 r/min for centrifugation for 5-10 minutes, the normal pressure circulation treatment is recovered after high pressure treatment, various proteins in moringa leaves can be rapidly separated and removed, and the.
The invention has the following beneficial effects:
firstly, the polysaccharide is insoluble in absolute ethyl alcohol, absolute ethyl alcohol is adopted to remove substances which are easy to be dissolved in the ethanol in the moringa leaves, preferably, the substances in the moringa leaves can be quickly and effectively dissolved in the ethanol under the condition of high-pressure and high-speed stirring at 0-4 ℃, the impurity removal time is obviously shortened, the impurity removal efficiency is improved, and the activity of the polysaccharide is not damaged;
secondly, the invention mixes the absolute ethyl alcohol preliminary impurity removal extract (namely filter residue) with deionized water and selenium yeast, and then utilizes microwave and ultralow temperature (-20 to-10 ℃) to circularly process, thus not only dissolving the polysaccharide, but also effectively combining with the selenium yeast to form stable selenium-rich polysaccharide, and improving the extraction amount of the selenium-rich polysaccharide and the content of organic selenium in the polysaccharide;
thirdly, the invention uses trichloroacetic acid-acetone with low concentration to deproteinize the moringa oleifera polysaccharide, and adopts trichloroacetic acid-acetone with low concentration to deproteinize under the condition of low temperature and relative high pressure, thereby improving the removal efficiency of the protein, shortening the removal time of the protein, and reducing the damage of organic solvent to the activity of the selenium-enriched polysaccharide under the condition of low temperature and high pressure.
Detailed Description
The present invention is further described in detail with reference to specific examples, so that those skilled in the art can implement the invention with reference to the description.
Example 1
The preparation method of the moringa oleifera selenium-rich polysaccharide comprises the following steps:
firstly, selecting moringa leaves as a raw material, freeze-drying the moringa leaves at a low temperature of-20 ℃, then crushing the dried moringa leaves by liquid nitrogen, screening the crushed moringa leaves by a 100-mesh screen, adding absolute ethyl alcohol into the screened moringa leaves, stirring the mixture for 20 minutes, obtaining filter residues by an ultrafiltration membrane, recovering the filtrate, and removing impurities dissolved in the absolute ethyl alcohol from moringa leaf powder;
step two, mixing the filter residue with deionized water and yeast selenium according to the weight ratio of 10:10:0.01, placing the mixture at the temperature of 40 ℃ for 1 hour, placing the mixture in microwaves and performing circulating treatment at the temperature of-20 ℃, centrifuging and extracting supernatant, placing the mixture in microwaves and performing circulating treatment at the temperature of-20 ℃, so that the selenium content in the polysaccharide can be obviously improved, the extraction amount of the selenium-rich polysaccharide can be obviously improved finally, and the activity of the selenium-rich polysaccharide is not damaged during extraction at relatively low temperature;
and step three, adding absolute ethyl alcohol into the supernatant obtained in the step two for multiple times for precipitation, filtering through an ultrafiltration membrane to obtain a precipitate, and performing vacuum freeze-drying on the precipitate to obtain the moringa oleifera selenium-rich polysaccharide.
Example 2
The preparation method of the moringa oleifera selenium-rich polysaccharide comprises the following steps:
firstly, selecting moringa leaves as a raw material, freeze-drying the moringa leaves at a low temperature of-20 ℃, crushing the moringa leaves by using liquid nitrogen, sieving the crushed moringa leaves by using a 100-mesh sieve, adding absolute ethyl alcohol, stirring the mixture for 30 minutes, passing the mixture through an ultrafiltration membrane to obtain filter residue, recovering a filtrate, wherein the weight ratio of the absolute ethyl alcohol to the moringa leaf powder is 5:1, and removing impurities dissolved in the absolute ethyl alcohol from the moringa leaf powder;
step two, mixing the filter residue with deionized water and yeast selenium according to the weight ratio of 10:50:0.02, placing the mixture at the temperature of 50 ℃ for 2 hours, placing the mixture in microwaves and performing circulating treatment at the temperature of-10 ℃, centrifuging and extracting supernatant, placing the mixture in microwaves and performing circulating treatment at the temperature of-10 ℃ to obviously improve the selenium content in the polysaccharide, thereby finally obviously improving the extraction amount of the selenium-rich polysaccharide and extracting the selenium-rich polysaccharide at relatively low temperature without damaging the activity of the selenium-rich polysaccharide,
and step three, adding absolute ethyl alcohol into the supernatant obtained in the step two for multiple times for precipitation, filtering through an ultrafiltration membrane to obtain a precipitate, and performing vacuum freeze-drying on the precipitate to obtain the moringa oleifera selenium-rich polysaccharide.
Example 3
The preparation method of the moringa oleifera selenium-rich polysaccharide comprises the following steps:
firstly, selecting moringa leaves as a raw material, freeze-drying the moringa leaves at a low temperature, crushing the moringa leaves by using liquid nitrogen, sieving the crushed moringa leaves with a 100-mesh sieve, adding absolute ethyl alcohol, placing the absolute ethyl alcohol in a reaction kettle, setting the temperature at 0 ℃, stirring the absolute ethyl alcohol for 25 minutes under the pressure of 15Mpa, and stirring the absolute ethyl alcohol and the moringa leaves at a stirring speed of 4000 revolutions per minute, wherein the weight ratio of the absolute ethyl alcohol to the moringa leaves powder is 1:1, impurities dissolved in the ethanol in the moringa leaves powder can be quickly dissolved out by using the conditions of low temperature, high pressure and quick stirring, the impurity removal efficiency is improved, the consumption of the absolute ethyl alcohol can be reduced, filter residue is obtained by using an ultrafiltration;
step two, mixing the filter residue with deionized water and yeast selenium according to the weight ratio of 10:20:0.01, placing the mixture at the temperature of 40-50 ℃ for 1-2 hours, placing the mixture in microwaves and circularly processing the mixture at the temperature of-15 ℃, centrifugally extracting supernatant, placing the mixture in microwaves and circularly processing the mixture at the temperature of-15 ℃ to obviously improve the selenium content in the polysaccharide, thereby finally obviously improving the extraction amount of the selenium-rich polysaccharide and extracting the selenium-rich polysaccharide at relatively low temperature without damaging the activity of the selenium-rich polysaccharide,
and step three, adding absolute ethyl alcohol into the supernatant obtained in the step two for multiple times for precipitation, filtering through an ultrafiltration membrane to obtain a precipitate, and performing vacuum freeze-drying on the precipitate to obtain the moringa oleifera selenium-rich polysaccharide.
Example 4
The preparation method of the moringa oleifera selenium-rich polysaccharide comprises the following steps:
firstly, selecting moringa leaves as a raw material, freeze-drying the moringa leaves at a low temperature, crushing the moringa leaves by using liquid nitrogen, sieving the crushed moringa leaves by using a 100-mesh sieve, adding absolute ethyl alcohol, placing the crushed moringa leaves into a reaction kettle, stirring the mixture for 30 minutes at the temperature of 4 ℃ and the pressure of 20Mpa, wherein the stirring speed is 5000 r/min, wherein the weight ratio of the absolute ethyl alcohol to the moringa oleifera leaf powder is 1:1, the ethanol-soluble impurities in the moringa oleifera leaf powder can be quickly dissolved out by using the conditions of low temperature, high pressure and quick stirring, the impurity removal efficiency is improved, and can reduce the consumption of absolute ethyl alcohol, filter residue is obtained through an ultrafiltration membrane and the filtrate is recovered, the filter residue is also added with absolute ethyl alcohol and absolute ethyl ether for at least 3 times of cleaning and filtering, wherein the weight ratio of the absolute ethyl alcohol to the absolute ethyl ether is 1:1, so that the extracted selenium-rich polysaccharide can reduce impurities, and the polysaccharide and the selenium yeast can be combined to form stable selenium-rich polysaccharide;
step two, mixing the filter residue with deionized water and yeast selenium according to the weight ratio of 10:30:0.01, placing the mixture at the temperature of 45 ℃ for 1 hour, placing the mixture in microwave for cyclic treatment at the temperature of between 20 ℃ below zero and 10 ℃ below zero, centrifugally extracting supernatant, placing the mixture in microwave for cyclic treatment at the temperature of between 20 ℃ below zero and 10 ℃ below zero, so that the selenium content in polysaccharide can be obviously improved, the extraction amount of the selenium-enriched polysaccharide can be obviously improved finally, the activity of the selenium-enriched polysaccharide is not damaged during extraction at relatively low temperature,
and step three, adding absolute ethyl alcohol into the supernatant obtained in the step two for multiple times for precipitation, filtering through an ultrafiltration membrane to obtain a precipitate, and performing vacuum freeze-drying on the precipitate to obtain the moringa oleifera selenium-rich polysaccharide.
Example 5
The preparation method of the moringa oleifera selenium-rich polysaccharide comprises the following steps:
firstly, selecting moringa leaves as a raw material, freeze-drying the moringa leaves at a low temperature, crushing the moringa leaves by using liquid nitrogen, sieving the crushed moringa leaves by using a 100-mesh sieve, adding absolute ethyl alcohol, placing the mixture into a reaction kettle, stirring the mixture for 25 minutes at the temperature of 1 ℃ and the pressure of 15Mpa, wherein the stirring speed is 3500 revolutions per minute, wherein the weight ratio of the absolute ethyl alcohol to the moringa oleifera leaf powder is 1:1, the ethanol-soluble impurities in the moringa oleifera leaf powder can be quickly dissolved out by using the conditions of low temperature, high pressure and quick stirring, the impurity removal efficiency is improved, and can reduce the consumption of absolute ethyl alcohol, filter residue is obtained through an ultrafiltration membrane and is recovered into filtrate, the filter residue is also added with the absolute ethyl alcohol and the absolute ethyl ether for 3 times of cleaning and filtering, wherein the weight ratio of the absolute ethyl alcohol to the absolute ethyl ether is 1:2, so that the extracted selenium-rich polysaccharide can reduce impurities, and the polysaccharide and the selenium yeast can be combined to form stable selenium-rich polysaccharide;
step two, mixing the filter residue with deionized water and yeast selenium according to the weight ratio of 10:35:0.01, placing the mixture at 45 ℃ for 1 hour, placing the mixture in 500MHZ microwave for processing for 25 seconds, then placing the mixture at-20 to-10 ℃ for 15 minutes, circulating the mixture for 3 times, centrifugally extracting supernatant, placing the supernatant in microwave and processing the supernatant at-20 to-10 ℃ in a circulating way, so that the selenium content in polysaccharide can be obviously improved, the extraction amount of the selenium-rich polysaccharide can be obviously improved finally, and the activity of the selenium-rich polysaccharide is not damaged when the selenium-rich polysaccharide is extracted at relatively low temperature,
and step three, adding absolute ethyl alcohol into the supernatant obtained in the step two for multiple times for precipitation, filtering through an ultrafiltration membrane to obtain a precipitate, and performing vacuum freeze-drying on the precipitate to obtain the moringa oleifera selenium-rich polysaccharide.
Example 6
The preparation method of the moringa oleifera selenium-rich polysaccharide comprises the following steps:
firstly, selecting moringa leaves as a raw material, freeze-drying the moringa leaves at a low temperature, crushing the moringa leaves by using liquid nitrogen, sieving the crushed moringa leaves by using a 100-mesh sieve, adding absolute ethyl alcohol, placing the crushed moringa leaves into a reaction kettle, stirring the mixture for 30 minutes at the temperature of 4 ℃ and the pressure of 20Mpa, wherein the stirring speed is 5000 r/min, wherein the weight ratio of the absolute ethyl alcohol to the moringa oleifera leaf powder is 1:1, the ethanol-soluble impurities in the moringa oleifera leaf powder can be quickly dissolved out by using the conditions of low temperature, high pressure and quick stirring, the impurity removal efficiency is improved, and can reduce the consumption of absolute ethyl alcohol, filter residue is obtained through an ultrafiltration membrane and recovered to obtain filter liquor, the filter residue is also added with absolute ethyl alcohol and absolute ethyl ether to be cleaned and filtered for 4 times, wherein the weight ratio of the absolute ethyl alcohol to the absolute ethyl ether is 1:2, so that the extracted selenium-rich polysaccharide can reduce impurities, and the polysaccharide and the selenium yeast can be combined to form stable selenium-rich polysaccharide;
step two, mixing the filter residue with deionized water and yeast selenium according to the weight ratio of 10:10:0.01, placing the mixture at the temperature of 40-50 ℃ for 1 hour, placing the mixture in 2000MHZ microwave for processing for 15 seconds, then placing the mixture at the temperature of-20 to-10 ℃ for 20 minutes, circulating the mixture for 5 times, adding 3 percent by weight of trichloroacetic acid-acetone, processing the mixture at the temperature of 2 ℃ and the pressure of 4Mpa for 0.5 hour, performing centrifugal filtration at 500 r/min, removing the trichloroacetic acid-acetone through extraction to obtain polysaccharide extract, performing deproteinization by adopting low-concentration trichloroacetic acid-acetone under the condition of low temperature and high pressure, improving the removal efficiency of protein, shortening the removal time of protein, reducing the damage of organic solvent to the activity of polysaccharide under the condition of low temperature and high pressure, performing centrifugal extraction on supernatant fluid by placing the mixture in microwave and processing the mixture at the temperature of-20 to-10 ℃ to obviously improve the selenium content in the polysaccharide, thereby finally remarkably improving the extraction quantity of the selenium-rich polysaccharide and extracting the selenium-rich polysaccharide at relatively low temperature without damaging the activity of the selenium-rich polysaccharide,
and step three, adding absolute ethyl alcohol into the supernatant obtained in the step two for multiple times for precipitation, filtering through an ultrafiltration membrane to obtain a precipitate, and performing vacuum freeze-drying on the precipitate to obtain the moringa oleifera selenium-rich polysaccharide.
Example 7
The preparation method of the moringa oleifera selenium-rich polysaccharide comprises the following steps:
firstly, selecting moringa leaves as a raw material, freeze-drying the moringa leaves at a low temperature, crushing the moringa leaves by using liquid nitrogen, sieving the crushed moringa leaves by using a 100-mesh sieve, adding absolute ethyl alcohol, placing the crushed moringa leaves into a reaction kettle, stirring the mixture for 30 minutes at the temperature of 4 ℃ and the pressure of 10Mpa, wherein the stirring speed is 5000 r/min, wherein the weight ratio of the absolute ethyl alcohol to the moringa oleifera leaf powder is 1:1, the ethanol-soluble impurities in the moringa oleifera leaf powder can be quickly dissolved out by using the conditions of low temperature, high pressure and quick stirring, the impurity removal efficiency is improved, and can reduce the consumption of absolute ethyl alcohol, filter residue is obtained through an ultrafiltration membrane and the filtrate is recovered, the filter residue is also added with absolute ethyl alcohol and absolute ethyl ether for at least 3 times of cleaning and filtering, wherein the weight ratio of the absolute ethyl alcohol to the absolute ethyl ether is 1:2, so that the extracted selenium-rich polysaccharide can reduce impurities, and the polysaccharide and the selenium yeast can be combined to form stable selenium-rich polysaccharide;
step two, mixing filter residue with deionized water and yeast selenium according to a weight ratio of 10:25:0.01, placing the mixture at 45 ℃ for 1 hour, placing the mixture in 1500MHZ microwave for processing for 30 seconds, then placing the mixture at-20 to-10 ℃ for 10 minutes, circulating the mixture for 4 times, adding 5.5 percent by weight of trichloroacetic acid-acetone, uniformly stirring the mixture, placing the mixture in a reaction kettle, regulating the temperature to 4 ℃ and the pressure to 1MPa for stirring processing for 5 minutes, regulating the temperature to 0 ℃ and the pressure to 1MPa for stirring processing for 5 minutes after the temperature is unchanged and the pressure is 1MPa is placed for 10 minutes, circulating the mixture after the temperature is unchanged and the pressure is 1MPa is placed for 10 minutes, finishing the processing after the cumulative time of stirring processing at 4 ℃ and the pressure is 40 minutes, placing the mixture in a centrifuge for 500 rpm for centrifuging for 5 minutes, recovering the normal pressure circulation processing after high pressure processing, can quickly separate out and remove various proteins in the moringa leaves and improve the impurity removal efficiency, adopts low-concentration trichloroacetic acid-acetone to remove proteins under the conditions of low temperature and relative high pressure, can improve the protein removal efficiency and shorten the protein removal time, can reduce the damage of organic solvents to the activity of the selenium-enriched polysaccharide under the conditions of low temperature and high pressure, centrifugally extracts supernatant, places the supernatant in microwaves and processes the supernatant in a circulating mode at the temperature of between 20 ℃ below zero and 10 ℃ below zero to obviously improve the selenium content in the polysaccharide, thereby finally obviously improving the extraction amount of the selenium-enriched polysaccharide and extracting the selenium-enriched polysaccharide at relative low temperature without damaging the activity of the selenium-enriched polysaccharide,
and step three, adding absolute ethyl alcohol into the supernatant obtained in the step two for multiple times for precipitation, filtering through an ultrafiltration membrane to obtain a precipitate, and performing vacuum freeze-drying on the precipitate to obtain the moringa oleifera selenium-rich polysaccharide.
Comparative example 1
As in example 1, step one was replaced with: firstly, selecting moringa leaves as a raw material, freeze-drying the moringa leaves at a low temperature of-20 ℃, crushing the dried moringa leaves by using liquid nitrogen, and screening the crushed moringa leaves through a 100-mesh screen to obtain moringa leaf powder, wherein the moringa leaf powder replaces filter residues to carry out the following steps.
Comparative example 2
Step two was replaced with, as in example 1: and step two, mixing the filter residue and deionized water according to the weight ratio of 10:10, standing at the temperature of 40 ℃ for 1 hour, then placing in a microwave, performing circulating treatment at the temperature of-20 ℃, and centrifuging to extract supernatant.
Comparative example 3
Step two was replaced with, as in example 1: and step two, mixing the filter residue with deionized water and selenium yeast according to the weight ratio of 10:10:0.01, standing for 1 hour at the temperature of 40 ℃, and centrifuging to extract supernatant.
Example 4
Step two was replaced with, as in example 1: and step two, mixing the filter residue and deionized water according to the weight ratio of 10:10, standing for 1 hour at the temperature of 40 ℃, and centrifuging to extract supernatant.
Testing the content of the moringa oleifera selenium-enriched polysaccharide by using a spectrophotometer, calculating the extraction rate and the purity of the moringa oleifera selenium-enriched polysaccharide, and analyzing the total selenium content and the organic selenium content in the extracted moringa oleifera selenium-enriched polysaccharide by using a high performance liquid phase-hydride generation-atomic fluorescence spectrometry method, wherein the results are as follows:
| extraction rate of moringa oleifera selenium-rich polysaccharide | Purity of moringa oleifera selenium-rich polysaccharide | Organic selenium content | Total selenium content | |
| Example 1 | 25.6％ | 90.2％ | 2.32mg/kg | 2.45mg/kg |
| Example 2 | 26.5％ | 90.5％ | 2.35mg/kg | 2.42mg/kg |
| Example 3 | 26.7％ | 91.5％ | 2.38mg/kg | 2.47mg/kg |
| Example 4 | 27.8％ | 93.6％ | 2.33mg/kg | 2.43mg/kg |
| Example 5 | 28.5％ | 95.8％ | 2.42mg/kg | 2.50mg/kg |
| Example 6 | 28.9％ | 97.5％ | 2.47mg/kg | 2.53mg/kg |
| Example 7 | 29.2％ | 98.6％ | 2.46mg/kg | 2.50mg/kg |
| Comparative example 1 | 20.2％ | 80.2％ | 1.92mg/kg | 2.15mg/kg |
| Comparative example 2 | 23.5％ | 90.1％ | 0.89mg/kg | 1.23mg/kg |
| Comparative example 3 | 15.2％ | 88.2％ | 1.11mg/kg | 1.35mg/kg |
| Comparative example 4 | 11.6％ | 85.2％ | 0.88mg/kg | 1.28mg/kg |
As can be seen from the data in the table, the method provided by the invention can obviously improve the extraction rate and purity of the moringa oleifera selenium-rich polysaccharide and improve the content of organic selenium in the moringa oleifera selenium-rich polysaccharide.
While embodiments of the invention have been described above, it is not limited to the applications set forth in the description and the embodiments, which are fully applicable to various fields of endeavor for which the invention may be embodied with additional modifications as would be readily apparent to those skilled in the art, and the invention is therefore not limited to the details given herein and to the embodiments shown and described without departing from the generic concept as defined by the claims and their equivalents.
Claims (5)
1. A preparation method of moringa oleifera selenium-rich polysaccharide is characterized by comprising the following steps:
step one, selecting moringa leaves as a raw material, freezing and drying the moringa leaves at a low temperature, crushing the moringa leaves by using liquid nitrogen, sieving the crushed moringa leaves by using a 100-mesh sieve, adding absolute ethyl alcohol, stirring for 20-30 minutes, passing through an ultrafiltration membrane to obtain filter residue, recovering filtrate,
step two, mixing the filter residue with deionized water and yeast selenium according to the weight ratio of 10:10-50:0.01-0.02, placing for 1-2 hours at the temperature of 40-50 ℃, placing in microwave, circularly processing at the temperature of-20 to-10 ℃, centrifugally extracting supernatant, adding 3-6% trichloroacetic acid-acetone, processing for 0.5-1 hour at the temperature of 0-4 ℃ and under the pressure of 1-5Mpa, centrifugally filtering at 500 r/min, extracting to remove trichloroacetic acid-acetone, adding 5.5% trichloroacetic acid-acetone, stirring uniformly, placing in a reaction kettle, adjusting the temperature to 0-4 ℃ and the pressure of 1-5Mpa, stirring for 5-10 minutes, placing for 10-20 minutes under the constant temperature and the constant pressure, adjusting the temperature to 0-4 ℃ and the pressure of 1-5Mpa, stirring for 5-10 minutes, standing at constant temperature and constant pressure for 10-20 min, circulating, stirring at 0-4 deg.C and 1-5Mpa for 40-60 min, centrifuging at 500 rpm for 5-10 min to obtain polysaccharide extractive solution,
and step three, adding the polysaccharide extracting solution obtained in the step two into absolute ethyl alcohol for multiple times for precipitation, filtering through an ultrafiltration membrane to obtain a precipitate, and performing vacuum freeze-drying on the precipitate to obtain the moringa oleifera selenium-rich polysaccharide.
2. The method for preparing moringa oleifera selenium-rich polysaccharide as claimed in claim 1, wherein the weight ratio of the absolute ethyl alcohol to the moringa oleifera leaf powder is 1-5: 1.
3. The method for preparing the moringa oleifera selenium-rich polysaccharide as claimed in claim 2, wherein in the step one, absolute ethyl alcohol is added, the absolute ethyl alcohol is placed in a reaction kettle, the temperature is set to be 0-4 ℃, the stirring speed is 2000-5000 revolutions per minute, and the pressure is 10-20Mpa, wherein the weight ratio of the absolute ethyl alcohol to the moringa oleifera leaf powder is 1: 1.
4. The method for preparing moringa oleifera selenium-rich polysaccharide as claimed in claim 1, wherein in step one, absolute ethyl alcohol and absolute ethyl ether are further added into the filter residue for at least 3 times of cleaning and filtering, wherein the weight ratio of the absolute ethyl alcohol to the absolute ethyl ether is 1: 1-2.
5. The method for preparing moringa oleifera selenium-rich polysaccharide as claimed in claim 1, wherein the second step of placing in microwave and circulating treatment at-20 to-10 ℃ specifically comprises: treating in 500-2000MHz microwave for 10-30 sec, and then at-20 deg.c to-10 deg.c for 10-30 min for 3-5 times.
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| WO1992015314A1 (en) * | 1991-02-28 | 1992-09-17 | Phytopharm Ltd. | Pharmaceutical compositions for the treatment of skin disorders |
| CN1069495A (en) * | 1992-07-31 | 1993-03-03 | 黄祥贤 | Selenium polysaccharide and manufacture method thereof |
| CN102241786A (en) * | 2011-06-07 | 2011-11-16 | 江苏大学 | Preparation method and application of selenium enriched puerarin polysaccharide |
| CN104829743A (en) * | 2015-05-25 | 2015-08-12 | 萧丽雅 | Preparation method and use of moringa oleifera leaf polysaccharides |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| WO1992015314A1 (en) * | 1991-02-28 | 1992-09-17 | Phytopharm Ltd. | Pharmaceutical compositions for the treatment of skin disorders |
| CN1069495A (en) * | 1992-07-31 | 1993-03-03 | 黄祥贤 | Selenium polysaccharide and manufacture method thereof |
| CN102241786A (en) * | 2011-06-07 | 2011-11-16 | 江苏大学 | Preparation method and application of selenium enriched puerarin polysaccharide |
| CN104829743A (en) * | 2015-05-25 | 2015-08-12 | 萧丽雅 | Preparation method and use of moringa oleifera leaf polysaccharides |
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