CN112048026B - Method for extracting and purifying licorice submicron powder polysaccharide - Google Patents
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Abstract
The invention provides a method for extracting and purifying licorice submicron powder polysaccharide, which comprises the following steps: s1, preparing liquorice superfine powder; s2, preparing a glycyrrhiza polysaccharide crude product; s3, dissolving the crude product of the glycyrrhiza polysaccharide in distilled water, and filtering the solution through a 0.30-0.50um microporous filter membrane; s4, diluting the glycyrrhiza polysaccharide solution, separating the solution by a DEAE-52 cellulose chromatographic column for 3 times, and freeze-drying to obtain the purified glycyrrhiza polysaccharide. The extraction method of the invention has high glycyrrhiza polysaccharide yield, and the glycyrrhiza polysaccharide obtained by the purification method has high purity.
Description
Technical Field
The invention relates to the field of extraction and purification of chemical components, in particular to an extraction and purification method of licorice submicron powder polysaccharide.
Background
The licorice is a perennial herb of Glycyrrhiza of Leguminosae, has high medicinal value and sweet and neutral nature, and has the effects of clearing away heat and toxic materials, invigorating spleen and qi, moistening lung and relieving cough. The main chemical component of Glycyrrhiza Polysaccharide (Glycyrrhiza Polysaccharide) in Glycyrrhiza has various biological activities such as immunoregulation, enhancement of phagocytosis function of macrophage cells, antioxidation and the like. The crude polysaccharide of licorice can be separated and extracted by different methods, such as water extraction and alcohol precipitation method, ultrasonic-assisted method, microwave, enzyme method, etc. However, in order to obtain glycyrrhiza polysaccharide with high purity, the crude extract needs to be further purified. At present, domestic research on glycyrrhiza polysaccharide mainly focuses on the extraction aspect, but few research reports on separation and purification are provided, and the biological chromatography of the crystal gel medium can effectively enrich bioactive substances and has high selectivity. Therefore, methods for isolation and purification of glycyrrhiza polysaccharides by gel chromatography are explored herein. The method takes Xinjiang Glycyrrhiza glabra (Glycyrrhiza glabra) as a raw material, and uses ultra-large pore anion exchange crystal gel as a chromatography medium to separate and purify crude polysaccharide of the licorice, so as to establish a new method for separating and purifying natural medicinal active polysaccharide by using crystal gel chromatography, and lay a foundation for the research and development and utilization of medicinal polysaccharide.
Disclosure of Invention
The technical problem to be solved is as follows: the invention aims to provide a method for extracting and purifying glycyrrhiza uralensis ultra-fine powder polysaccharide so as to obtain high-purity glycyrrhiza uralensis polysaccharide.
The technical scheme is as follows: a method for extracting and purifying licorice submicron powder polysaccharide comprises the following steps:
s1, weighing liquorice, cleaning, cutting, drying for 72 h at 50 ℃, crushing and sieving by a 80-mesh sieve to obtain liquorice coarse powder, then processing the coarse powder by a vibration type ultrafine crusher to obtain 300-mesh and 500-mesh liquorice ultrafine powder, and storing the liquorice ultrafine powder in a dryer for later use;
s2, adding licorice powder into distilled water, wherein the ratio of material to liquid is 1: 30-50, ultrasonic power of 450-: 1 ultrasonic-assisted extraction for 30min, standing for 1h after ultrasonic treatment, and then performing ultrasonic extraction at a rotation speed of 8000- -1 Centrifuging at 4 deg.C for 10min to obtain supernatant and precipitate, and rotary evaporating the supernatant to obtain crude productAdding 1/10 ethanol solution for several times, adjusting ethanol concentration, and separating to obtain crude product of Glycyrrhiza polysaccharide;
s3, combining the glycyrrhiza polysaccharide crude products extracted for multiple times in the steps, dissolving the glycyrrhiza polysaccharide crude products into distilled water with the concentration of 15-20mg/mL, uniformly stirring the glycyrrhiza polysaccharide crude products at 8000-10000r/min, centrifuging the glycyrrhiza polysaccharide crude products, and passing supernate through a 0.30-0.50um microporous filter membrane;
s4, diluting the glycyrrhiza polysaccharide solution into a solution of 10mg/mL, carrying out DEAE-52 cellulose chromatographic column, loading the sample with 2mL, carrying out elution at the flow rate of 2.5mL/min by using a 0.1-0.4mol/L NaCl solution, collecting the eluent, concentrating, dialyzing and freeze-drying the eluent;
s5, preparing freeze-dried components collected by the DEAE-52 cellulose chromatographic column into 5mg/mL polysaccharide solution, loading the polysaccharide solution with the sample amount of 2mL, purifying by using the DEAE-52 cellulose chromatographic column, wherein the eluent is distilled water, the elution flow rate is set to be 0.5mL/min, collecting the eluent, concentrating and dialyzing the eluent, and freeze-drying;
s6, preparing the freeze-dried component obtained in the step S5 into 5mg/mL polysaccharide solution, loading the polysaccharide solution with the sample volume of 2mL, purifying the polysaccharide solution by using a DEAE-52 cellulose chromatographic column, setting the elution flow rate to be 0.5mL/min, collecting the elution solution, concentrating and dialyzing the elution solution, and freeze-drying the elution solution to obtain the purified glycyrrhiza polysaccharide.
Preferably, the licorice root ultra-fine powder in the step S1 is 300-400 meshes.
Preferably, in the step S2, 95% ethanol solution is first added into the supernatant to make the final ethanol concentration of the sample be 30%, and the sample is left for 12 hours and centrifuged to obtain a crude glycyrrhiza polysaccharide product; and continuously dropwise adding 95% ethanol solution into the supernatant gradually to make the final concentration of the supernatant reach 60%, standing, centrifuging to obtain crude glycyrrhiza polysaccharide, adding ethanol into the supernatant until the concentration of ethanol in the supernatant reaches 80%, standing, centrifuging to obtain crude glycyrrhiza polysaccharide, and mixing the crude glycyrrhiza polysaccharide for several times.
Preferably, the pore size of the microporous filter membrane in the step S3 is 0.35-0.40 um.
Preferably, the cellulose of DEAE-52 in step S4 is OH - DEAE-52 cellulose form.
Preferably, DE in the step S5AE-52 cellulose is Cl - DEAE-52 cellulose form.
Preferably, the DEAE-52 cellulose in the step S5 is HCO 3 - DEAE-52 cellulose form.
Has the advantages that: the extraction and purification of the invention has the following advantages:
1. after the liquorice is subjected to superfine grinding treatment, the granularity is uniform, the dispersity is good, intracellular components can be fully exposed, the dissolution and the release of effective components are promoted, the activity of the effective components is improved, and the loss of polysaccharide components is reduced to the maximum extent;
2. in the purification process of polysaccharide, 3 DEAE-52 celluloses are selected and used according to OH - Type, Cl - Type, HCO 3 - Sequentially setting the type, wherein the polysaccharide belongs to weak electrolyte and has different interaction force with DEAE-52 cellulose, and eluting the glycyrrhiza polysaccharide in different order according to OH - Type, Cl - Type, HCO 3 - The elution effect of the type is sequentially improved, and highly purified glycyrrhiza polysaccharide is obtained.
Detailed Description
Example 1
A method for extracting and purifying liquorice submicron powder polysaccharide comprises the following steps:
s1, weighing liquorice, cleaning, cutting, drying for 72 h at 50 ℃, crushing, sieving by a 80-mesh sieve to obtain liquorice coarse powder, then processing the coarse powder by a vibration type ultrafine grinder to obtain 500-mesh liquorice ultrafine powder, and storing in a dryer for later use;
s2, adding licorice powder into distilled water, wherein the ratio of material to liquid is 1: 30, at an ultrasonic power of 600W, duty cycle 1: 1 ultrasonic-assisted extraction for 30min, standing for 1h after ultrasonic treatment, and rotating at 8000r/min -1 Centrifuging at 4 deg.C for 10min to obtain supernatant and precipitate, rotary evaporating the supernatant to 1/10 of original volume, adding 95% ethanol solution into the supernatant to make the final ethanol concentration of the sample be 30%, standing for 12h, and centrifuging to obtain crude product of Glycyrrhiza polysaccharide; continuously dropwise adding 95% ethanol solution into the supernatant to make the final concentration of the supernatant reach 60%, standing, centrifuging, and separating to obtain crude Glycyrrhiza polysaccharideAdding ethanol into the supernatant until the concentration of ethanol in the supernatant reaches 80%, standing, centrifuging to obtain crude product of Glycyrrhiza polysaccharide, and mixing the crude products of Glycyrrhiza polysaccharide for several times;
s3, combining the crude glycyrrhiza polysaccharide products extracted for multiple times in the steps, dissolving the crude glycyrrhiza polysaccharide products into distilled water, wherein the concentration of the crude glycyrrhiza polysaccharide products is 15-20mg/mL, uniformly stirring the crude glycyrrhiza polysaccharide products at 8000r/min, centrifuging the mixture, and filtering supernate through a 0.50-micrometer microporous filter membrane;
s4, diluting the glycyrrhiza polysaccharide solution into 10mg/mL solution, OH - Performing DEAE-52 cellulose chromatography column with sample loading capacity of 2mL and flow rate of 2.5mL/min, eluting with 0.4mol/L NaCl solution, collecting eluate, concentrating, dialyzing, and lyophilizing;
s5, adding OH - Lyophilized fractions collected from DEAE-52 cellulose chromatography column were prepared into 5mg/mL polysaccharide solution, loaded in 2mL, and concentrated with Cl - Purifying with DEAE-52 cellulose chromatographic column, eluting with distilled water at 0.5mL/min, collecting eluate, concentrating, dialyzing, and lyophilizing;
s6, reconstituting the lyophilized fraction obtained in step S5 into a polysaccharide solution of 5mg/mL, loading 2mL, and adding HCO 3 - Purifying with DEAE-52 cellulose chromatographic column, eluting with distilled water at flow rate of 0.5mL/min, collecting eluate, concentrating, dialyzing, and lyophilizing to obtain purified Glycyrrhiza polysaccharide.
Example 2
A method for extracting and purifying liquorice submicron powder polysaccharide comprises the following steps:
s1, weighing liquorice, cleaning, cutting, drying for 72 h at 50 ℃, crushing, sieving by a 80-mesh sieve to obtain liquorice coarse powder, then processing the coarse powder by a vibration type ultrafine grinder to obtain 300-mesh liquorice ultrafine powder, and storing in a dryer for later use;
s2, adding liquorice powder into distilled water, wherein the ratio of material to liquid is 1: 50, at an ultrasonic power of 450W, duty cycle 1: 1 ultrasonic-assisted extraction for 30min, standing for 1h after ultrasonic treatment, and rotating at a speed of 10000 r.min -1 Centrifuging at 4 deg.C for 10min to obtain supernatant and precipitate, rotary evaporating the supernatant to 1/10, collecting supernatantFirstly, adding 95% ethanol solution to ensure that the final ethanol concentration of the sample is 30%, standing for 12h, and centrifuging to obtain a crude product of the glycyrrhiza polysaccharide; continuously dropwise adding 95% ethanol solution into the supernatant gradually to make the final concentration of the supernatant reach 60%, standing, centrifuging to obtain crude product of Glycyrrhiza polysaccharide, adding ethanol into the supernatant until the ethanol concentration of the supernatant reaches 80%, standing, centrifuging to obtain crude product of Glycyrrhiza polysaccharide, and mixing the crude product of Glycyrrhiza polysaccharide several times;
s3, combining the crude glycyrrhiza polysaccharide products extracted for multiple times in the steps, dissolving the crude glycyrrhiza polysaccharide products into distilled water at the concentration of 15-20mg/mL, uniformly stirring the mixture at 10000r/min, centrifuging the mixture, and filtering the supernatant through a 0.30-micrometer microporous filter membrane;
s4, diluting the glycyrrhiza polysaccharide solution into 10mg/mL solution, OH - Performing DEAE-52 cellulose chromatography column with sample loading capacity of 2mL and flow rate of 2.5mL/min, eluting with 0.1mol/L NaCl solution, collecting eluate, concentrating, dialyzing, and lyophilizing;
s5, adding OH - Lyophilized fractions collected from DEAE-52 cellulose chromatography column were prepared into 5mg/mL polysaccharide solution, loaded in 2mL, and concentrated with Cl - Purifying with DEAE-52 cellulose chromatographic column, eluting with distilled water at 0.5mL/min, collecting eluate, concentrating, dialyzing, and lyophilizing;
s6, reconstituting the lyophilized fraction obtained in step S5 into a polysaccharide solution of 5mg/mL, loading 2mL, and adding HCO 3 - Purifying with DEAE-52 cellulose chromatographic column, eluting with distilled water at 0.5mL/min, collecting eluate, concentrating, dialyzing, and lyophilizing to obtain purified Glycyrrhiza polysaccharide.
Example 3
A method for extracting and purifying liquorice submicron powder polysaccharide comprises the following steps:
s1, weighing liquorice, cleaning, cutting, drying at 50 ℃ for 72 h, crushing, sieving with a 80-mesh sieve to obtain coarse liquorice powder, processing the coarse powder with a vibration type ultrafine grinder to obtain 300-mesh liquorice ultrafine powder, and storing in a dryer for later use;
s2, adding liquorice powder into distilled water, wherein the ratio of material to liquid is 1: 40, by ultrasonic workRate 600W, duty cycle 1: 1 ultrasonic-assisted extraction for 30min, standing for 1h after ultrasonic treatment, and rotating at 8500 r.min -1 Centrifuging at 4 deg.C for 10min to obtain supernatant and precipitate, rotary evaporating the supernatant to 1/10 of original volume, adding 95% ethanol solution into the supernatant to make the final ethanol concentration of the sample be 30%, standing for 12h, and centrifuging to obtain crude product of Glycyrrhiza polysaccharide; continuously dropwise adding 95% ethanol solution into the supernatant gradually to make the final concentration of the supernatant reach 60%, standing, centrifuging to obtain crude product of Glycyrrhiza polysaccharide, adding ethanol into the supernatant until the ethanol concentration of the supernatant reaches 80%, standing, centrifuging to obtain crude product of Glycyrrhiza polysaccharide, and mixing the crude product of Glycyrrhiza polysaccharide several times;
s3, combining the crude glycyrrhiza polysaccharide products extracted for multiple times in the steps, dissolving the crude glycyrrhiza polysaccharide products into distilled water at the concentration of 15-20mg/mL, uniformly stirring the mixture at 9000r/min, centrifuging the mixture, and filtering the supernatant with a 0.40-micron microporous filter membrane;
s4, diluting the glycyrrhiza polysaccharide solution into 10mg/mL solution, OH - Performing DEAE-52 cellulose chromatography column with sample loading amount of 2mL and flow rate of 2.5mL/min, eluting with 0.2-0.3mol/L NaCl solution, collecting eluate, concentrating, dialyzing, and lyophilizing;
s5, adding OH - Lyophilized fractions collected from DEAE-52 cellulose chromatography column were prepared into 5mg/mL polysaccharide solution, loaded in 2mL, and concentrated with Cl - Purifying with DEAE-52 cellulose chromatographic column, eluting with distilled water at 0.5mL/min, collecting eluate, concentrating, dialyzing, and lyophilizing;
s6, reconstituting the lyophilized fraction obtained in step S5 into a 5mg/mL polysaccharide solution, loading 2mL, and applying HCO 3 - Purifying with DEAE-52 cellulose chromatographic column, eluting with distilled water at flow rate of 0.5mL/min, collecting eluate, concentrating, dialyzing, and lyophilizing to obtain purified Glycyrrhiza polysaccharide.
Example 4
A method for extracting and purifying licorice submicron powder polysaccharide comprises the following steps:
s1, weighing liquorice, cleaning, cutting, drying at 50 ℃ for 72 h, crushing, sieving with a 80-mesh sieve to obtain coarse liquorice powder, processing the coarse powder with a vibration type ultrafine grinder to obtain 400-mesh ultrafine liquorice powder, and storing in a dryer for later use;
s2, adding liquorice powder into distilled water, wherein the ratio of material to liquid is 1: 45, at an ultrasonic power of 500W, duty cycle 1: 1 ultrasonic-assisted extraction for 30min, standing for 1h after ultrasonic treatment, and rotating at 9500 r.min -1 Centrifuging at 4 deg.C for 10min to obtain supernatant and precipitate, rotary evaporating the supernatant to 1/10 of original volume, adding 95% ethanol solution into the supernatant to make the final ethanol concentration of the sample be 30%, standing for 12h, and centrifuging to obtain crude product of Glycyrrhiza polysaccharide; continuously dropwise adding 95% ethanol solution into the supernatant gradually to make the final concentration of the supernatant reach 60%, standing, centrifuging to obtain crude product of Glycyrrhiza polysaccharide, adding ethanol into the supernatant until the ethanol concentration of the supernatant reaches 80%, standing, centrifuging to obtain crude product of Glycyrrhiza polysaccharide, and mixing the crude product of Glycyrrhiza polysaccharide several times;
s3, combining the glycyrrhiza polysaccharide crude products extracted for multiple times in the steps, dissolving the glycyrrhiza polysaccharide crude products into distilled water at the concentration of 15-20mg/mL, uniformly stirring the glycyrrhiza polysaccharide crude products at 8500r/min, centrifuging the glycyrrhiza polysaccharide crude products, and passing supernate through a 0.35um microporous filter membrane;
s4, diluting the glycyrrhiza polysaccharide solution into 10mg/mL solution, OH - Performing DEAE-52 cellulose chromatography column with sample loading capacity of 2mL and flow rate of 2.5mL/min, eluting with 0.2-0.3mol/L NaCl solution, collecting eluate, concentrating, dialyzing, and lyophilizing;
s5, adding OH - Lyophilized fractions collected from DEAE-52 cellulose chromatography column were prepared into 5mg/mL polysaccharide solution, loaded in 2mL, and concentrated with Cl - Purifying with DEAE-52 cellulose chromatographic column, eluting with distilled water at 0.5mL/min, collecting eluate, concentrating, dialyzing, and lyophilizing;
s6, reconstituting the lyophilized fraction obtained in step S5 into a polysaccharide solution of 5mg/mL, loading 2mL, and adding HCO 3 - Purifying with DEAE-52 cellulose chromatographic column, eluting with distilled water at 0.5mL/min, collecting eluate, concentrating, dialyzing, and lyophilizing to obtain purified Glycyrrhiza polysaccharide.
Comparative example 1
A method for extracting and purifying liquorice submicron powder polysaccharide comprises the following steps:
s1, weighing liquorice, cleaning, cutting, drying for 72 hours at 50 ℃, crushing, and sieving with a 80-mesh sieve to obtain liquorice coarse powder;
s2, adding liquorice powder into distilled water, wherein the ratio of material to liquid is 1: 40, at an ultrasonic power of 600W, duty cycle 1: 1 ultrasonic-assisted extraction for 30min, standing for 1h after ultrasonic treatment, and rotating at 8500 r.min -1 Centrifuging at 4 deg.C for 10min to obtain supernatant and precipitate, rotary evaporating the supernatant to 1/10 of original volume, adding 95% ethanol solution into the supernatant to make the final ethanol concentration of the sample be 30%, standing for 12h, and centrifuging to obtain crude product of Glycyrrhiza polysaccharide; continuously dropwise adding 95% ethanol solution into the supernatant gradually to make the final concentration of the supernatant reach 60%, standing, centrifuging to obtain crude product of Glycyrrhiza polysaccharide, adding ethanol into the supernatant until the ethanol concentration of the supernatant reaches 80%, standing, centrifuging to obtain crude product of Glycyrrhiza polysaccharide, and mixing the crude product of Glycyrrhiza polysaccharide several times;
s3, combining the glycyrrhiza polysaccharide crude products extracted for multiple times in the steps, dissolving the glycyrrhiza polysaccharide crude products into distilled water at the concentration of 15-20mg/mL, uniformly stirring the glycyrrhiza polysaccharide crude products at 9000r/min, centrifuging the glycyrrhiza polysaccharide crude products, and passing supernate through a 0.40um microporous filter membrane;
s4, diluting the glycyrrhiza polysaccharide solution into 10mg/mL solution, OH - Performing DEAE-52 cellulose chromatography column with sample loading amount of 2mL and flow rate of 2.5mL/min, eluting with 0.2-0.3mol/L NaCl solution, collecting eluate, concentrating, dialyzing, and lyophilizing;
s5, adding OH - The lyophilized fraction collected by DEAE-52 cellulose chromatography column was prepared into 5mg/mL polysaccharide solution, loaded in 2mL, and treated with Cl - Purifying with DEAE-52 cellulose chromatographic column, eluting with distilled water at 0.5mL/min, collecting eluate, concentrating, dialyzing, and lyophilizing;
s6, reconstituting the lyophilized fraction obtained in step S5 into a polysaccharide solution of 5mg/mL, loading 2mL, and adding HCO 3 - Purifying with DEAE-52 cellulose chromatographic column, and evaporating the eluateDistilling with water, setting the flow rate of elution to 0.5mL/min, collecting eluate, concentrating, dialyzing, and lyophilizing to obtain purified Glycyrrhiza polysaccharide.
Comparative example 2
A method for extracting and purifying liquorice submicron powder polysaccharide comprises the following steps:
s1, weighing liquorice, cleaning, cutting, drying at 50 ℃ for 72 h, crushing, sieving with a 80-mesh sieve to obtain coarse liquorice powder, processing the coarse powder with a vibration type ultrafine grinder to obtain 300-mesh liquorice ultrafine powder, and storing in a dryer for later use;
s2, adding liquorice powder into distilled water, wherein the ratio of material to liquid is 1: 40, at an ultrasonic power of 600W, duty cycle 1: 1 ultrasonic-assisted extraction for 30min, standing for 1h after ultrasonic treatment, and rotating at 8500 r.min -1 Centrifuging at 4 deg.C for 10min to obtain supernatant and precipitate, rotary evaporating the supernatant to 1/10 of original volume, adding 95% ethanol solution into the supernatant to make the final ethanol concentration of the sample be 30%, standing for 12h, and centrifuging to obtain crude product of Glycyrrhiza polysaccharide; continuously dropwise adding 95% ethanol solution into the supernatant gradually to make the final concentration of the supernatant reach 60%, standing, centrifuging to obtain crude glycyrrhiza polysaccharide product, adding ethanol into the supernatant until the ethanol concentration of the supernatant reaches 80%, standing, centrifuging to obtain crude glycyrrhiza polysaccharide product, and mixing the crude glycyrrhiza polysaccharide products for several times;
s3, combining the crude glycyrrhiza polysaccharide products extracted for multiple times in the steps, dissolving the crude glycyrrhiza polysaccharide products into distilled water at the concentration of 15-20mg/mL, uniformly stirring the mixture at 9000r/min, centrifuging the mixture, and filtering the supernatant with a 0.40-micron microporous filter membrane;
s4, diluting the glycyrrhiza polysaccharide solution into 10mg/mL solution, OH - And (2) carrying out DEAE-52 cellulose chromatographic column with the sample loading amount of 2mL and the flow rate of 2.5mL/min, eluting with 0.2-0.3mol/L NaCl solution, collecting eluent, concentrating, dialyzing and freeze-drying the eluent to obtain the purified glycyrrhiza polysaccharide.
The purity of the glycyrrhiza polysaccharide obtained in examples 1-4 and comparative examples 1-2 of the present invention was 96.5%, 97.5%, 96.8%, 97.7%, 96.4%, 91.5%, and the extraction rate of glycyrrhiza polysaccharide was 13.2%, 14.1%, 13.6%, 13.7%, 12.5%, and 13.8%, as determined by phenol-sulfuric acid colorimetry.
Claims (4)
1. The method for extracting and purifying the licorice submicron powder polysaccharide is characterized by comprising the following steps:
s1, weighing liquorice, cleaning, cutting, drying for 72 h at 50 ℃, crushing and sieving by a 80-mesh sieve to obtain liquorice coarse powder, then processing the coarse powder by a vibration type ultrafine crusher to obtain 300-mesh and 500-mesh liquorice ultrafine powder, and storing the liquorice ultrafine powder in a dryer for later use;
s2, adding liquorice powder into distilled water, wherein the ratio of material to liquid is 1: 30-50, ultrasonic power of 450-: 1 ultrasonic-assisted extraction for 30min, standing for 1h after ultrasonic treatment, and then performing ultrasonic extraction at a rotation speed of 8000- -1 Centrifuging at 4 deg.C for 10min to obtain supernatant and precipitate, rotary evaporating the supernatant to 1/10 of original volume, adding ethanol solution for several times, adjusting ethanol concentration, and separating to obtain crude product of Glycyrrhiza polysaccharide;
s3, combining the crude glycyrrhiza polysaccharide products extracted for multiple times in the steps, dissolving the crude glycyrrhiza polysaccharide products into distilled water, wherein the concentration of the crude glycyrrhiza polysaccharide products is 15-20mg/mL, uniformly stirring the crude glycyrrhiza polysaccharide products at 8000-10000r/min, centrifuging the mixture, and filtering supernate through a 0.30-0.50um microporous filter membrane;
s4, diluting the glycyrrhiza polysaccharide solution into 10mg/mL solution, OH - Performing DEAE-52 cellulose chromatography column with sample loading amount of 2mL and flow rate of 2.5mL/min, eluting with 0.1-0.4mol/L NaCl solution, collecting eluate, concentrating, dialyzing, and lyophilizing;
s5, adding OH - Lyophilized fractions collected from DEAE-52 cellulose chromatography column were prepared into 5mg/mL polysaccharide solution, loaded in 2mL, and concentrated with Cl - Purifying with DEAE-52 cellulose chromatographic column with distilled water as eluent at flow rate of 0.5mL/min, collecting eluate, concentrating, dialyzing, and lyophilizing;
s6, reconstituting the lyophilized fraction obtained in step S5 into a polysaccharide solution of 5mg/mL, loading 2mL, and adding HCO 3 - Purifying with DEAE-52 cellulose chromatographic column, eluting with distilled water at flow rate of 0.5mL/min, collecting eluate, concentrating, dialyzing, and lyophilizing to obtain purified Glycyrrhiza polysaccharide.
2. The method for extracting and purifying licorice submicron powder polysaccharide as claimed in claim 1, characterized in that: the licorice root ultra-fine powder in the step S1 is 300-400 meshes.
3. The method for extracting and purifying licorice submicron powder polysaccharide as claimed in claim 1, characterized in that: in the step S2, firstly adding 95% ethanol solution into the supernatant to make the final ethanol concentration of the sample be 30%, standing for 12h, and centrifuging to obtain a crude product of the glycyrrhiza polysaccharide; and continuously dropwise adding 95% ethanol solution into the supernatant gradually to make the final concentration of the supernatant reach 60%, standing, centrifuging to obtain crude glycyrrhiza polysaccharide, adding ethanol into the supernatant until the concentration of ethanol in the supernatant reaches 80%, standing, centrifuging to obtain crude glycyrrhiza polysaccharide, and mixing the crude glycyrrhiza polysaccharide for several times.
4. The method for extracting and purifying licorice submicron powder polysaccharide as claimed in claim 1, characterized in that: the pore diameter of the microporous filter membrane in the step S3 is 0.35-0.40 um.
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