CN111825779A - Extraction method of dendrobium officinale polysaccharide - Google Patents
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Abstract
The invention provides an extraction method of dendrobium officinale polysaccharide, which is a novel extraction method of dendrobium officinale polysaccharide, and specifically comprises the following steps: pulverizing dried herba Dendrobii, weighing appropriate amount, adding anhydrous ethanol, and sealing and soaking; after removing the ethanol, adding distilled water according to a certain feed-liquid ratio, and setting ultrasonic time, extraction temperature, extraction time, ultrasonic power and extraction times for extraction; after sufficient extraction, the polysaccharide solution is concentrated under reduced pressure, and the impurities are removed by using absolute ethyl alcohol. The results show that: the yield of the dendrobium officinale polysaccharide is 61.12%. The invention solves the comprehensive problem that the extraction rate and production are influenced because the optimal process condition cannot be determined due to the complex extraction method and excessive influence factors during the extraction of the dendrobium officinale polysaccharide.
Description
Technical Field
The invention belongs to the field of extraction of effective components in plants, and particularly relates to various factors influencing the extraction rate of dendrobium officinale, so that a dendrobium officinale polysaccharide extraction method with determined optimal experimental parameters under comprehensive influencing factors is realized. The dendrobium officinale polysaccharide can obtain higher extraction rate under a simple extraction process, has stable work and low energy consumption, and can be produced in large quantities.
Background
Dendrobium officinale (Dendrobium officinale) is an important plant used as both medicine and food, and can tonify stomach, promote fluid production, regulate immunity, delay aging and the like. Research shows that the Dendrobium officinale contains various active ingredients such as polysaccharide, alkaloid, polyphenol, flavone, amino acid and the like, wherein Dendrobium Officinale Polysaccharide (DOP) is a main active ingredient of the Dendrobium officinale polysaccharide, is also a substance with the highest content, and has the effects of resisting bacteria, enhancing immunity, reducing blood sugar, blood fat and blood pressure, resisting tumors, resisting oxidation and the like. A large number of researches report that the existing extraction method for DOP mainly comprises a hot water extraction method, an enzyme extraction method, an ultrasonic-assisted extraction method, a flash extraction method, an ultrahigh pressure extraction method and a microwave-assisted extraction method. Due to the problems of complex extraction process, low extraction rate and the like, the process exploration for selecting a simple extraction process and ensuring the extraction rate becomes a current research hotspot. The hot water extraction method is the most common method for extracting the dendrobium officinale polysaccharide, but the extraction rate is not high, the extraction efficiency can be greatly improved by the intervention of ultrasonic waves, and the size of the ultrasonic power is not explored. Therefore, the determination of the dendrobium officinale polysaccharide extraction process determined by the optimal experimental parameters under the comprehensive influence factors has a higher development prospect.
Disclosure of Invention
The technical problems to be solved by the invention are as follows: provides an optimal extraction process of dendrobium officinale polysaccharide so as to overcome the defects in the prior art.
The invention adopts the following technical scheme for solving the technical problems:
the invention provides a method for extracting dendrobium officinale polysaccharide, which specifically comprises the following steps: adding anhydrous ethanol into dried and pulverized herba Dendrobii, and sealing and soaking; removing ethanol, adding distilled water, performing ultrasonic treatment, extracting, concentrating the obtained polysaccharide solution under reduced pressure, and removing impurities with anhydrous ethanol.
The granularity of the dendrobium officinale is 0.1-0.2mm, 4.8-5.2g of the dendrobium officinale is weighed, 48-52mL of absolute ethyl alcohol is added, and the dendrobium officinale is sealed by a preservative film and soaked for 11-13 h.
The extraction method of the dendrobium officinale polysaccharide provided by the invention can comprise the following steps:
step 1, removing absolute ethyl alcohol in the soaked sample, and adding the sample and distilled water into a three-neck flask according to a proper proportion;
step 2, setting working parameters of an ultrasonic cleaner, and stirring and extracting;
step 3, filtering the extracted solution into a conical flask, putting the filtrate into an ultrasonic cleaner for ultrasonic treatment, heating by using an electric heating temperature control device, simultaneously performing vacuum pumping by using a circulating water pump for reduced pressure concentration, and centrifuging the concentrated solution;
step 4, taking the supernatant, adding absolute ethyl alcohol for alcohol precipitation, standing overnight, centrifuging to obtain a precipitate, roughly filtering the extracted solution into a 1000mL conical flask by using a 100-mesh gauze, and adding pure water for washing for several times to dissolve the precipitate (a filter cake);
step 5, adding absolute ethyl alcohol into the solution for secondary alcohol precipitation, standing in a refrigerator, centrifuging to obtain precipitate, and adding pure water for dissolving;
step 6, after full dissolution, centrifuging the solution, and taking supernatant liquid in a beaker;
and 7, carrying out freeze drying on the polysaccharide solution to obtain the target dendrobium officinale polysaccharide.
In step 1 of the above method, the sample is mixed with distilled water in a ratio of 1: 20- -1: 60 (g/mL), preferably 1: 50 (g/mL), was charged to a 500mL three-necked flask.
In step 2 of the method, a 240W ultrasonic cleaner is adopted, and the parameters are set as follows: heating at 50-90 deg.C for 120-.
In step 2 of the method, a 240W ultrasonic cleaner is adopted, and the parameter settings are preferably as follows: heating at 80 deg.C for 210min, ultrasonic time for 120min and ultrasonic power of 60% (140W), and stirring for extraction.
In step 3 of the above method, the extracted solution may be coarsely filtered with a 100-mesh screen into a 1000mL conical flask, and the filter cake may be washed with pure water several times, generally 3 times.
In step 3, the concentrated solution can be centrifuged at 3000-4500r/min for 10-20 min. Preferably: the concentrate was centrifuged at 4500r/min for 10 min.
In the step 4 of the method, absolute ethyl alcohol is added into the supernatant to 500mL at the temperature of 300 ℃, and the mixture is kept stand for 12 to 24 hours in a refrigerator at the temperature of 4 ℃; centrifuging at 4500r/min for 10-20min, collecting precipitate, adding pure water to 100-.
In step 4 of the above method, it is preferable that: adding anhydrous ethanol into the supernatant to 500mL, and standing in a refrigerator at 4 ℃ for 12 h. Centrifuging at 4500r/min for 10min, collecting precipitate, and adding pure water to 100mL to dissolve the precipitate.
In the step 5, absolute ethyl alcohol is added into the supernatant to 500mL, and the mixture is kept stand for 12-24h in a refrigerator at 4 ℃. Centrifuging at 4500r/min for 10-20min, collecting precipitate, adding pure water to 100-.
In step 6, the solution may be centrifuged at 3000-5000r/min for 10-20 min.
The polysaccharide extracted by the method provided by the invention can be used for cosmetic raw materials and food industry.
Compared with the prior art, the invention has the following prominent substantive characteristics:
1. the ultrasonic-assisted extraction method has the characteristics of short extraction time, low required temperature and high yield, but the ultrasonic power is not too low or too high, otherwise, the extraction rate is not high or the molecular structure and the characteristics of the polysaccharide are negatively influenced. The invention provides a novel extraction process, which fully explores the influence of ultrasonic power on the extraction rate, overcomes the defects of the traditional ultrasonic-assisted extraction method, and realizes the rapid and efficient extraction of dendrobium polysaccharides.
2. The invention balances the optimal experimental parameters under the comprehensive influence factors of the crushing degree, the feed-liquid ratio, the ultrasonic time, the extraction temperature, the extraction time, the ultrasonic power and the like on the premise of extracting for 1 time.
3. By the treatment of the method, the extraction rate of the dendrobium officinale polysaccharide is high and can reach 61.12%.
4. The method is simple to operate, and the obtained crude polysaccharide is relatively pure.
5. The method is simple to operate, easy to realize mass production of the dendrobium officinale polysaccharide, stable in work, low in energy consumption and capable of realizing industrial production, and the obtained crude polysaccharide is relatively pure.
6. The invention has no toxic substance, and the extracted polysaccharide can be used in cosmetic raw materials and food industry.
Detailed Description
The invention provides a novel extraction process of dendrobium officinale polysaccharide, which is mainly used for comparing a material-liquid ratio, ultrasonic time, extraction temperature, extraction time and ultrasonic power under the condition that the extraction frequency is 1, and further researching the extraction frequency. Crushing the dried dendrobium officinale, wherein the crushing degree is as follows: weighing appropriate amount of 0.1-1 mm, adding anhydrous alcohol, sealing and soaking; removing absolute ethyl alcohol in the soaked sample, and adding the sample and distilled water into a three-neck flask according to a proper proportion; setting working parameters of an ultrasonic cleaner, wherein the material-liquid ratio is 1: 20-1: 60 (g/mL), the ultrasonic time is 30-150min, the extraction temperature is 50-90 ℃, the extraction time is 240min and the ultrasonic power is 20-100% (240W), and stirring and extracting; filtering the extracted solution into a conical flask; putting the filtrate in an ultrasonic cleaner for ultrasonic treatment, heating by using an electric heating temperature control device, vacuumizing by using a circulating water pump for reduced pressure concentration, and centrifuging the concentrated solution; taking supernatant, adding absolute ethyl alcohol for alcohol precipitation, standing overnight, centrifuging to obtain precipitate, and adding pure water to dissolve the precipitate; adding absolute ethyl alcohol into the solution for secondary alcohol precipitation, standing in a refrigerator, centrifuging to obtain precipitate, and adding pure water for dissolving; after the solution is fully dissolved, centrifuging the solution, and taking supernatant liquid in a beaker; and (4) freeze-drying the polysaccharide solution to obtain the target dendrobium officinale polysaccharide.
The present invention will be described in detail with reference to examples, but the present invention is not limited thereto.
Example 1 extraction method of Dendrobium officinale polysaccharide
Weighing 5g of dendrobium officinale with the crushing degree of 0.8-1mm, adding about 50mL of absolute ethyl alcohol, sealing by a preservative film, and soaking for 12 hours; after removing ethanol, adding the sample into a 500mL three-necked flask, and setting experimental parameters as follows: carrying out water extraction under the conditions of feed-liquid ratio of 1: 20 (g/mL), ultrasonic time of 90 min, extraction temperature of 70 ℃, extraction time of 150min, extraction frequency of 1 time and ultrasonic power of 100% (240W); coarsely filtering the extracted solution with 100 mesh gauze into a 1000mL conical flask, and washing the filter cake with pure water for 3 times; putting the filtrate in an ultrasonic cleaner for ultrasonic treatment, heating by using an electric heating temperature control device, and simultaneously vacuumizing by using a circulating water pump for decompression and concentration; centrifuging the concentrated solution (10 min,4500 r/min); taking supernatant, adding absolute ethyl alcohol into the supernatant to 500mL for alcohol precipitation, and standing overnight in a refrigerator at 4 ℃; centrifuging (10 min,4500 r/min), collecting precipitate, adding pure water to 100mL, and dissolving the precipitate; adding anhydrous ethanol into the solution to 300mL for secondary alcohol precipitation, standing in a refrigerator at 4 ℃ for 2h, centrifuging (10 min,4500 r/min), taking precipitate, and adding pure water to 150mL for dissolution; after sufficient dissolution, the solution was centrifuged (20 min,5000 r/min) and the supernatant was taken in a beaker. The result showed an extraction rate of 13.88%.
Example 2 extraction method of Dendrobium officinale polysaccharide
Weighing 5g of dendrobium officinale with the crushing degree of 0.8-1mm, adding about 50mL of absolute ethyl alcohol, sealing by a preservative film, and soaking for 12 hours; after removing ethanol, adding the sample into a 500mL three-necked flask, and setting experimental parameters as follows: carrying out water extraction under the conditions of material-liquid ratio of 1: 50 (g/mL), ultrasonic time of 90 min, extraction temperature of 70 ℃, extraction time of 150min, extraction frequency of 1 time and ultrasonic power of 100% (240W); coarsely filtering the extracted solution with 100 mesh gauze into a 1000mL conical flask, and washing the filter cake with pure water for 3 times; putting the filtrate in an ultrasonic cleaner for ultrasonic treatment, heating by using an electric heating temperature control device, and simultaneously vacuumizing by using a circulating water pump for decompression and concentration; centrifuging the concentrated solution (10 min,4500 r/min); taking supernatant, adding absolute ethyl alcohol into the supernatant to 500mL for alcohol precipitation, and standing overnight in a refrigerator at 4 ℃; centrifuging (10 min,4500 r/min), collecting precipitate, adding pure water to 100mL, and dissolving the precipitate; adding anhydrous ethanol into the solution to 300mL for secondary alcohol precipitation, standing in a refrigerator at 4 ℃ for 2h, centrifuging (10 min,4500 r/min), taking precipitate, and adding pure water to 150mL for dissolution; after sufficient dissolution, the solution was centrifuged (20 min,5000 r/min) and the supernatant was taken in a beaker. The result showed an extraction rate of 21.21%.
Example 3 extraction method of Dendrobium officinale polysaccharide
Weighing 5g of dendrobium officinale with the crushing degree of 0.4-0.6mm, adding about 50mL of absolute ethyl alcohol, sealing by a preservative film, and soaking for 12 h; after removing ethanol, adding the sample into a 500mL three-necked flask, and setting experimental parameters as follows: carrying out water extraction under the conditions of material-liquid ratio of 1: 50 (g/mL), ultrasonic time of 90 min, extraction temperature of 70 ℃, extraction time of 150min, extraction frequency of 1 time and ultrasonic power of 100% (240W); coarsely filtering the extracted solution with 100 mesh gauze into a 1000mL conical flask, and washing the filter cake with pure water for 3 times; putting the filtrate in an ultrasonic cleaner for ultrasonic treatment, heating by using an electric heating temperature control device, and simultaneously vacuumizing by using a circulating water pump for decompression and concentration; centrifuging the concentrated solution (10 min,4500 r/min); taking supernatant, adding absolute ethyl alcohol into the supernatant to 500mL for alcohol precipitation, and standing overnight in a refrigerator at 4 ℃; centrifuging (10 min,4500 r/min), collecting precipitate, adding pure water to 100mL, and dissolving the precipitate; adding anhydrous ethanol into the solution to 300mL for secondary alcohol precipitation, standing in a refrigerator at 4 ℃ for 2h, centrifuging (10 min,4500 r/min), taking precipitate, and adding pure water to 150mL for dissolution; after sufficient dissolution, the solution was centrifuged (20 min,5000 r/min) and the supernatant was taken in a beaker. The result showed an extraction rate of 25.35%.
Example 4 extraction method of Dendrobium officinale polysaccharide
Weighing 5g of dendrobium officinale with the crushing degree of 0.1-0.2mm, adding about 50mL of absolute ethyl alcohol, sealing with a preservative film, and soaking for 12 h; after removing ethanol, adding the sample into a 500mL three-necked flask, and setting experimental parameters as follows: carrying out water extraction under the conditions of material-liquid ratio of 1: 50 (g/mL), ultrasonic time of 90 min, extraction temperature of 70 ℃, extraction time of 150min, extraction frequency of 1 time and ultrasonic power of 100% (240W); coarsely filtering the extracted solution with 100 mesh gauze into a 1000mL conical flask, and washing the filter cake with pure water for 3 times; putting the filtrate in an ultrasonic cleaner for ultrasonic treatment, heating by using an electric heating temperature control device, and simultaneously vacuumizing by using a circulating water pump for decompression and concentration; centrifuging the concentrated solution (10 min,4500 r/min); taking supernatant, adding absolute ethyl alcohol into the supernatant to 500mL for alcohol precipitation, and standing overnight in a refrigerator at 4 ℃; centrifuging (10 min,4500 r/min), collecting precipitate, adding pure water to 100mL, and dissolving the precipitate; adding anhydrous ethanol into the solution to 300mL for secondary alcohol precipitation, standing in a refrigerator at 4 ℃ for 2h, centrifuging (10 min,4500 r/min), taking precipitate, and adding pure water to 150mL for dissolution; after sufficient dissolution, the solution was centrifuged (20 min,5000 r/min) and the supernatant was taken in a beaker. The result showed an extraction rate of 34.43%.
Example 5 extraction method of Dendrobium officinale polysaccharide
Weighing 5g of dendrobium officinale with the crushing degree of 0.1-0.2mm, adding about 50mL of absolute ethyl alcohol, sealing with a preservative film, and soaking for 12 h; after removing ethanol, adding the sample into a 500mL three-necked flask, and setting experimental parameters as follows: carrying out water extraction under the conditions of material-liquid ratio of 1: 50 (g/mL), ultrasonic time of 30 min, extraction temperature of 70 ℃, extraction time of 150min, extraction frequency of 1 time and ultrasonic power of 100% (240W); coarsely filtering the extracted solution with 100 mesh gauze into a 1000mL conical flask, and washing the filter cake with pure water for 3 times; putting the filtrate in an ultrasonic cleaner for ultrasonic treatment, heating by using an electric heating temperature control device, and simultaneously vacuumizing by using a circulating water pump for decompression and concentration; centrifuging the concentrated solution (10 min,4500 r/min); taking supernatant, adding absolute ethyl alcohol into the supernatant to 500mL for alcohol precipitation, and standing overnight in a refrigerator at 4 ℃; centrifuging (10 min,4500 r/min), collecting precipitate, adding pure water to 100mL, and dissolving the precipitate; adding anhydrous ethanol into the solution to 300mL for secondary alcohol precipitation, standing in a refrigerator at 4 ℃ for 2h, centrifuging (10 min,4500 r/min), taking precipitate, and adding pure water to 150mL for dissolution; after sufficient dissolution, the solution was centrifuged (20 min,5000 r/min) and the supernatant was taken in a beaker. The result showed an extraction rate of 29.56%.
Example 6 extraction method of Dendrobium officinale polysaccharide
Weighing 5g of dendrobium officinale with the crushing degree of 0.1-0.2mm, adding about 50mL of absolute ethyl alcohol, sealing with a preservative film, and soaking for 12 h; after removing ethanol, adding the sample into a 500mL three-necked flask, and setting experimental parameters as follows: carrying out water extraction under the conditions of material-liquid ratio of 1: 50 (g/mL), ultrasonic time of 120min, extraction temperature of 70 ℃, extraction time of 150min, extraction frequency of 1 time and ultrasonic power of 100% (240W); coarsely filtering the extracted solution with 100 mesh gauze into a 1000mL conical flask, and washing the filter cake with pure water for 3 times; putting the filtrate in an ultrasonic cleaner for ultrasonic treatment, heating by using an electric heating temperature control device, and simultaneously vacuumizing by using a circulating water pump for decompression and concentration; centrifuging the concentrated solution (10 min,4500 r/min); taking supernatant, adding absolute ethyl alcohol into the supernatant to 500mL for alcohol precipitation, and standing overnight in a refrigerator at 4 ℃; centrifuging (10 min,4500 r/min), collecting precipitate, adding pure water to 100mL, and dissolving the precipitate; adding anhydrous ethanol into the solution to 300mL for secondary alcohol precipitation, standing in a refrigerator at 4 ℃ for 2h, centrifuging (10 min,4500 r/min), taking precipitate, and adding pure water to 150mL for dissolution; after sufficient dissolution, the solution was centrifuged (20 min,5000 r/min) and the supernatant was taken in a beaker. The result showed 36.70% extraction yield.
Example 7 extraction method of Dendrobium officinale polysaccharide
Weighing 5g of dendrobium officinale with the crushing degree of 0.1-0.2mm, adding about 50mL of absolute ethyl alcohol, sealing with a preservative film, and soaking for 12 h; after removing ethanol, adding the sample into a 500mL three-necked flask, and setting experimental parameters as follows: carrying out water extraction under the conditions of material-liquid ratio of 1: 50 (g/mL), ultrasonic time of 120min, extraction temperature of 70 ℃, extraction time of 150min, extraction frequency of 1 time and ultrasonic power of 20% (240W); coarsely filtering the extracted solution with 100 mesh gauze into a 1000mL conical flask, and washing the filter cake with pure water for 3 times; putting the filtrate in an ultrasonic cleaner for ultrasonic treatment, heating by using an electric heating temperature control device, and simultaneously vacuumizing by using a circulating water pump for decompression and concentration; centrifuging the concentrated solution (10 min,4500 r/min); taking supernatant, adding absolute ethyl alcohol into the supernatant to 500mL for alcohol precipitation, and standing overnight in a refrigerator at 4 ℃; centrifuging (10 min,4500 r/min), collecting precipitate, adding pure water to 100mL, and dissolving the precipitate; adding anhydrous ethanol into the solution to 300mL for secondary alcohol precipitation, standing in a refrigerator at 4 ℃ for 2h, centrifuging (10 min,4500 r/min), taking precipitate, and adding pure water to 150mL for dissolution; after sufficient dissolution, the solution was centrifuged (20 min,5000 r/min) and the supernatant was taken in a beaker. The result showed 37.63% extraction yield.
Example 8 extraction method of Dendrobium officinale polysaccharide
Weighing 5g of dendrobium officinale with the crushing degree of 0.1-0.2mm, adding about 50mL of absolute ethyl alcohol, sealing with a preservative film, and soaking for 12 h; after removing ethanol, adding the sample into a 500mL three-necked flask, and setting experimental parameters as follows: carrying out water extraction under the conditions of material-liquid ratio of 1: 50 (g/mL), ultrasonic time of 120min, extraction temperature of 70 ℃, extraction time of 150min, extraction frequency of 1 time and ultrasonic power of 60% (240W); coarsely filtering the extracted solution with 100 mesh gauze into a 1000mL conical flask, and washing the filter cake with pure water for 3 times; putting the filtrate in an ultrasonic cleaner for ultrasonic treatment, heating by using an electric heating temperature control device, and simultaneously vacuumizing by using a circulating water pump for decompression and concentration; centrifuging the concentrated solution (10 min,4500 r/min); taking supernatant, adding absolute ethyl alcohol into the supernatant to 500mL for alcohol precipitation, and standing overnight in a refrigerator at 4 ℃; centrifuging (10 min,4500 r/min), collecting precipitate, adding pure water to 100mL, and dissolving the precipitate; adding anhydrous ethanol into the solution to 300mL for secondary alcohol precipitation, standing in a refrigerator at 4 ℃ for 2h, centrifuging (10 min,4500 r/min), taking precipitate, and adding pure water to 150mL for dissolution; after sufficient dissolution, the solution was centrifuged (20 min,5000 r/min) and the supernatant was taken in a beaker. The result showed 41.47% extraction yield.
Example 9 extraction method of Dendrobium officinale polysaccharide
Weighing 5g of dendrobium officinale with the crushing degree of 0.1-0.2mm, adding about 50mL of absolute ethyl alcohol, sealing with a preservative film, and soaking for 12 h; after removing ethanol, adding the sample into a 500mL three-necked flask, and setting experimental parameters as follows: carrying out water extraction under the conditions of feed-liquid ratio of 1: 50 (g/mL), ultrasonic time of 120min, extraction temperature of 50 ℃, extraction time of 150min, extraction frequency of 1 time and ultrasonic power of 60% (240W); coarsely filtering the extracted solution with 100 mesh gauze into a 1000mL conical flask, and washing the filter cake with pure water for 3 times; putting the filtrate in an ultrasonic cleaner for ultrasonic treatment, heating by using an electric heating temperature control device, and simultaneously vacuumizing by using a circulating water pump for decompression and concentration; centrifuging the concentrated solution (10 min,4500 r/min); taking supernatant, adding absolute ethyl alcohol into the supernatant to 500mL for alcohol precipitation, and standing overnight in a refrigerator at 4 ℃; centrifuging (10 min,4500 r/min), collecting precipitate, adding pure water to 100mL, and dissolving the precipitate; adding anhydrous ethanol into the solution to 300mL for secondary alcohol precipitation, standing in a refrigerator at 4 ℃ for 2h, centrifuging (10 min,4500 r/min), taking precipitate, and adding pure water to 150mL for dissolution; after sufficient dissolution, the solution was centrifuged (20 min,5000 r/min) and the supernatant was taken in a beaker. The results showed an extraction rate of 38.48%.
Example 10 extraction method of Dendrobium officinale polysaccharide
Weighing 5g of dendrobium officinale with the crushing degree of 0.1-0.2mm, adding about 50mL of absolute ethyl alcohol, sealing with a preservative film, and soaking for 12 h; after removing ethanol, adding the sample into a 500mL three-necked flask, and setting experimental parameters as follows: carrying out water extraction under the conditions of feed-liquid ratio of 1: 50 (g/mL), ultrasonic time of 120min, extraction temperature of 80 ℃, extraction time of 150min, extraction frequency of 1 time and ultrasonic power of 60% (240W); coarsely filtering the extracted solution with 100 mesh gauze into a 1000mL conical flask, and washing the filter cake with pure water for 3 times; putting the filtrate in an ultrasonic cleaner for ultrasonic treatment, heating by using an electric heating temperature control device, and simultaneously vacuumizing by using a circulating water pump for decompression and concentration; centrifuging the concentrated solution (10 min,4500 r/min); taking supernatant, adding absolute ethyl alcohol into the supernatant to 500mL for alcohol precipitation, and standing overnight in a refrigerator at 4 ℃; centrifuging (10 min,4500 r/min), collecting precipitate, adding pure water to 100mL, and dissolving the precipitate; adding anhydrous ethanol into the solution to 300mL for secondary alcohol precipitation, standing in a refrigerator at 4 ℃ for 2h, centrifuging (10 min,4500 r/min), taking precipitate, and adding pure water to 150mL for dissolution; after sufficient dissolution, the solution was centrifuged (20 min,5000 r/min) and the supernatant was taken in a beaker. The results showed an extraction of 44.64%.
Example 11 extraction method of Dendrobium officinale polysaccharide
Weighing 5g of dendrobium officinale with the crushing degree of 0.1-0.2mm, adding about 50mL of absolute ethyl alcohol, sealing with a preservative film, and soaking for 12 h; after removing ethanol, adding the sample into a 500mL three-necked flask, and setting experimental parameters as follows: carrying out water extraction under the conditions of feed-liquid ratio of 1: 50 (g/mL), ultrasonic time of 120min, extraction temperature of 80 ℃, extraction time of 120min, extraction frequency of 1 time and ultrasonic power of 60% (240W); coarsely filtering the extracted solution with 100 mesh gauze into a 1000mL conical flask, and washing the filter cake with pure water for 3 times; putting the filtrate in an ultrasonic cleaner for ultrasonic treatment, heating by using an electric heating temperature control device, and simultaneously vacuumizing by using a circulating water pump for decompression and concentration; centrifuging the concentrated solution (10 min,4500 r/min); taking supernatant, adding absolute ethyl alcohol into the supernatant to 500mL for alcohol precipitation, and standing overnight in a refrigerator at 4 ℃; centrifuging (10 min,4500 r/min), collecting precipitate, adding pure water to 100mL, and dissolving the precipitate; adding anhydrous ethanol into the solution to 300mL for secondary alcohol precipitation, standing in a refrigerator at 4 ℃ for 2h, centrifuging (10 min,4500 r/min), taking precipitate, and adding pure water to 150mL for dissolution; after sufficient dissolution, the solution was centrifuged (20 min,5000 r/min) and the supernatant was taken in a beaker. The results showed an extraction rate of 39.50%.
Example 12 extraction method of Dendrobium officinale polysaccharide
Weighing 5g of dendrobium officinale with the crushing degree of 0.1-0.2mm, adding about 50mL of absolute ethyl alcohol, sealing with a preservative film, and soaking for 12 h; after removing ethanol, adding the sample into a 500mL three-necked flask, and setting experimental parameters as follows: carrying out water extraction under the conditions of feed-liquid ratio of 1: 50 (g/mL), ultrasonic time of 120min, extraction temperature of 80 ℃, extraction time of 210min, extraction frequency of 1 time and ultrasonic power of 60% (240W); coarsely filtering the extracted solution with 100 mesh gauze into a 1000mL conical flask, and washing the filter cake with pure water for 3 times; putting the filtrate in an ultrasonic cleaner for ultrasonic treatment, heating by using an electric heating temperature control device, and simultaneously vacuumizing by using a circulating water pump for decompression and concentration; centrifuging the concentrated solution (10 min,4500 r/min); taking supernatant, adding absolute ethyl alcohol into the supernatant to 500mL for alcohol precipitation, and standing overnight in a refrigerator at 4 ℃; centrifuging (10 min,4500 r/min), collecting precipitate, adding pure water to 100mL, and dissolving the precipitate; adding anhydrous ethanol into the solution to 300mL for secondary alcohol precipitation, standing in a refrigerator at 4 ℃ for 2h, centrifuging (10 min,4500 r/min), taking precipitate, and adding pure water to 150mL for dissolution; after sufficient dissolution, the solution was centrifuged (20 min,5000 r/min) and the supernatant was taken in a beaker. The result showed 51.48% extraction yield.
Example 13 extraction method of Dendrobium officinale polysaccharide
Weighing 5g of dendrobium officinale with the crushing degree of 0.1-0.2mm, adding about 50mL of absolute ethyl alcohol, sealing with a preservative film, and soaking for 12 h; after removing ethanol, adding the sample into a 500mL three-necked flask, and setting experimental parameters as follows: carrying out water extraction under the conditions of feed-liquid ratio of 1: 50 (g/mL), ultrasonic time of 120min, extraction temperature of 80 ℃, extraction time of 210min, extraction times of 3 times and ultrasonic power of 60% (240W); coarsely filtering the extracted solution with 100 mesh gauze into a 1000mL conical flask, and washing the filter cake with pure water for 3 times; putting the filtrate in an ultrasonic cleaner for ultrasonic treatment, heating by using an electric heating temperature control device, and simultaneously vacuumizing by using a circulating water pump for decompression and concentration; centrifuging the concentrated solution (10 min,4500 r/min); taking supernatant, adding absolute ethyl alcohol into the supernatant to 500mL for alcohol precipitation, and standing overnight in a refrigerator at 4 ℃; centrifuging (10 min,4500 r/min), collecting precipitate, adding pure water to 100mL, and dissolving the precipitate; adding anhydrous ethanol into the solution to 300mL for secondary alcohol precipitation, standing in a refrigerator at 4 ℃ for 2h, centrifuging (10 min,4500 r/min), taking precipitate, and adding pure water to 150mL for dissolution; after sufficient dissolution, the solution was centrifuged (20 min,5000 r/min) and the supernatant was taken in a beaker. The result showed an extraction rate of 61.12%.
The method provided by the embodiment has the following characteristics:
1. the extraction method of the dendrobium candidum polysaccharide provided by the invention is a novel ultrasonic-assisted extraction process, fully researches the factors influencing the extraction rate, overcomes the defects of the traditional ultrasonic-assisted extraction method, and realizes the rapid and efficient extraction of the dendrobium candidum polysaccharide.
2. The invention balances the optimal experimental parameters under the comprehensive influence factors of the crushing degree, the feed-liquid ratio, the ultrasonic time, the extraction temperature, the extraction time, the ultrasonic power and the like on the premise of extracting for 1 time.
3. By the treatment of the method, the yield of the dendrobium officinale polysaccharide is high and can reach 61.12%.
4. The method is simple to operate, easy to realize mass production of the dendrobium officinale polysaccharide, stable in work, low in energy consumption and capable of realizing industrial production, and the obtained crude polysaccharide is relatively pure.
5. The invention has no toxic substance, and the extracted polysaccharide can be used in cosmetic raw materials and food industry.
Claims (10)
1. The extraction method of the dendrobium candidum polysaccharide is characterized in that absolute ethyl alcohol is added into dried and crushed dendrobium candidum, and the mixture is sealed and soaked; removing ethanol, adding distilled water, performing ultrasonic treatment, extracting, concentrating the obtained polysaccharide solution under reduced pressure, and removing impurities with anhydrous ethanol.
2. The extraction method of the dendrobium officinale polysaccharide as claimed in claim 1, wherein the granularity of the dendrobium officinale is 0.1-0.2mm, 4.8-5.2g of the dendrobium officinale is weighed, 48-52mL of absolute ethyl alcohol is added, and the mixture is sealed by a preservative film and soaked for 11-13 h.
3. The extraction method of dendrobium officinale polysaccharide as claimed in claim 1, which is characterized by comprising the following steps:
step 1, removing absolute ethyl alcohol in the soaked sample, and adding the sample and distilled water into a three-neck flask according to a proper proportion;
step 2, setting working parameters of an ultrasonic cleaner, and stirring and extracting;
step 3, filtering the extracted solution into a conical flask, putting the filtrate into an ultrasonic cleaner for ultrasonic treatment, heating by using an electric heating temperature control device, simultaneously performing vacuum pumping by using a circulating water pump for reduced pressure concentration, and centrifuging the concentrated solution;
step 4, adding absolute ethyl alcohol into the supernatant for alcohol precipitation, standing overnight, centrifuging to obtain a precipitate, and adding pure water to dissolve the precipitate;
step 5, adding absolute ethyl alcohol into the solution for secondary alcohol precipitation, standing in a refrigerator, centrifuging to obtain precipitate, and adding pure water for dissolving;
step 6, after full dissolution, centrifuging the solution, and taking supernatant liquid in a beaker;
and 7, carrying out freeze drying on the polysaccharide solution to obtain the target dendrobium officinale polysaccharide.
4. The method according to claim 3, wherein the ratio of the sample to distilled water in step 1 is 1: 20-1: 60g/mL, and charged into a 500mL three-necked flask.
5. The method of claim 3, wherein the 240W ultrasonic cleaner parameters are set in step 2 to: heating at 50-90 deg.C for 120-.
6. The method according to claim 3, wherein the extracted solution is coarsely filtered with a 100 mesh screen into a 1000mL conical flask in step 3, and the filter cake is washed several times with pure water.
7. The method as claimed in claim 3, wherein the concentrate is centrifuged at 3000-4500r/min for 10-20min in step 3.
8. The method as claimed in claim 3, wherein the absolute ethanol is added to the supernatant in step 4 to 500mL, and the mixture is left standing in a refrigerator at 4 ℃ for 12-24 h; centrifuging at 4500r/min for 10-20min, collecting precipitate, adding pure water to 100-.
9. The method according to claim 3, wherein in step 5, absolute ethanol is added for secondary alcohol precipitation, and the mixture is kept still for 2 to 8 hours at 4 ℃ in a refrigerator; centrifuging at 3000-4500r/min for 10-20min, collecting the precipitate, adding pure water to 150-200mL for dissolution.
10. The method according to claim 3, wherein the solution is centrifuged at 3000-5000r/min for 10-20min in step 6.
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