CN108084293B - Method for extracting artemisia vulgaris polysaccharide with high selectivity - Google Patents
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Abstract
The invention discloses a method for extracting Artemisia polysaccharide with high selectivity, which comprises the following steps of firstly), drying the Artemisia raw material, then crushing the mixture into folium artemisiae argyi powder, putting the folium artemisiae argyi powder in the step two) into a closed extraction tank, vacuumizing the extraction tank to 100 kPa, injecting distilled water as an extraction solvent, adding ionic liquid entrainer [ C4mim ] Cl for extraction, extracting, controlling the extraction temperature by an automatic controller, taking heat from a far infrared generator arranged at the bottom of the extraction tank, step three) pumping the liquid solvent into an evaporation tank, subjecting the liquid solvent in the evaporation tank to constant temperature and reduced pressure desolventizing treatment to obtain crude extract, dissolving with 95% ethanol solution, placing in a refrigerator at 4 deg.C to completely precipitate polysaccharide, filtering, freeze drying, the obtained Artemisia polysaccharide is an extraction method which has simple process, low investment and high efficiency and can be used for industrialized production of high-quality Artemisia polysaccharide.
Description
Technical Field
The invention relates to the field of natural product extraction, in particular to a method for extracting artemisia vulgaris polysaccharide, which has the advantages of simple process, high selectivity, high extraction rate and no damage to the activity of the polysaccharide.
Background
Northern Asia drug (B)Artemisia vulgarisL.) is perennial of Artemisia of CompositaeOne variety of the herbal plant ai, tomorrow li shizhen (compendium of materia Medica), year: "(folium Artemisiae Argyi) herbal medicine is not indigenous, but is clouded with the field. In Song Dynasty, the best … … is the best one for the syndrome of recovering Tangyin (the one in the town of the Tangyin county at present) and the syndrome of only Tangyin in the recent times is called Bei ai. North moxa, Hai moxa, Qi moxa and Qi moxa are called "four famous moxa" in history in China. The whole herb of the northern wormwood is used for medicine and has the effects of warming qi and blood, expelling cold and dampness, stopping bleeding, warming channels, preventing miscarriage and the like. The northern wormwood has high yield, rich nutrient components and higher content of crude protein, crude fat and nitrogen-free extract; the folium Artemisiae Argyi is rich in polysaccharide, essential oil, flavone, mineral, microelement, tannic acid and vitamins A, B1, B2, C, etc. Is a natural plant resource with great development value.
The functions of the polysaccharide of the artemisia vulgaris are mostly expressed in the aspects of immunoregulation, tumor resistance, virus resistance, cancer resistance, oxidation resistance, aging resistance, beauty maintenance, face nourishing, liver and kidney disease symptom treatment and relief, blood sugar reduction, bacteria resistance and the like. The polysaccharide is easy to dissolve in water, so the most common method for extracting the polysaccharide is a water extraction and alcohol precipitation method, the method has good polysaccharide extraction effect and low cost, but the extract has more impurities, especially protein, the subsequent purification process is complex, the time is long, and the polysaccharide loss is serious. Shenxia et al adopts response surface method to optimize the extraction process of folium Artemisiae Argyi crude polysaccharide, and determines the optimal process conditions for extracting folium Artemisiae Argyi polysaccharide by water: the extraction temperature is 99 ℃, the extraction time is 2.3 h, and the water-material ratio is 20 (Shenxia, Zhang Yanhong, Yuan Hui, and the like. research on optimizing the extraction process of the crude polysaccharide of folium artemisiae argyi by a response surface analysis method [ J ]. Chinese patent medicine, 2010, 32(1):48-51 ]. The method for extracting and purifying artemisia argyi polysaccharide (application number: 201110075571.8) discloses a method for extracting polysaccharide artemisia argyi polysaccharide by using an ethanol solution percolation method, and a finished product is obtained after the steps of dissolving by normal saline, repeatedly removing protein and desalting, wherein the process is complicated.
Disclosure of Invention
The invention aims to overcome the defects in the prior art and provide the extraction method which has simple process, low investment and high efficiency and can be used for industrially producing the high-quality Artemisia polysaccharide.
The technical scheme of the invention is realized as follows:
a method for extracting Artemisia polysaccharide with high selectivity is characterized in that: the method comprises the following steps:
step one), pretreatment: before extraction, drying the raw material of the artemisia vulgaris L, and then crushing the dried material of the artemisia vulgaris L into artemisia vulgaris L powder with the granularity of 40-60 meshes;
step two), extraction: placing the pretreated folium Artemisiae Argyi powder in a sealed extraction tank, vacuumizing until the vacuum degree in the extraction tank reaches-100 kPa, injecting distilled water into the extraction tank as extraction solvent, completely immersing folium Artemisiae Argyi sample, and adding ionic liquid entrainer [ C ] for extraction4mim]Extracting after Cl, wherein the amount of the entrainer is 0.1-1% of the volume of the extraction solvent;
the extraction temperature is controlled by an automatic controller, the heat source is a far infrared generating device arranged at the bottom of the extraction tank, a glass window is arranged at the bottom of the extraction tank, so that far infrared rays penetrate through the extraction tank, the power is 500-1000W, and the far infrared wavelength range is 25-200 mu m;
step three), desolventizing: after extraction, pumping the liquid solvent into an evaporating pot, and respectively carrying out constant-temperature reduced-pressure desolventizing treatment on the material residues in the extracting pot and the liquid solvent in the evaporating pot to obtain a crude extract after desolventizing the liquid solvent;
step four), alcohol precipitation: dissolving the above extract with 95% ethanol solution, placing in refrigerator at 4 deg.C to precipitate polysaccharide completely, filtering, and freeze drying to obtain Artemisia princeps Pampanini polysaccharide.
The water content of the dried folium artemisiae argyi raw material in the step one) is lower than 8%.
The mass-volume ratio of the pretreated folium artemisiae argyi powder to water in the step two) is 1: 5-15 g/ml, the extraction time is 5-10 minutes, the temperature is 105-115 ℃, the pressure is 0.1-0.2 Mpa, and the cyclic extraction is performed for 1-3 times.
The entrainer for extraction in the second step can be used [ C ]4C11OHim]CF3SO3And [ C4mim]CF3SO3The entrainer is 0.1-1% of the volume of the extraction solvent.
The temperature of the decompression desolventizing in the step three) is 50-65 ℃.
And the water vapor evaporated from the material slag and the liquid solvent in the step three) flows back to the solvent storage tank after being condensed and liquefied for recycling.
The mass volume ratio of the extract to the 95% ethanol solution in the step four) is 1: 3-4, and the standing time of the extract in a refrigerator at 4 ℃ is 2-4 h.
The structure that the water vapor evaporated from the material slag and the liquid solvent flows back to the solvent storage tank after being condensed and liquefied is as follows: the extraction tank and the evaporating tank are communicated with the inlet of the compressor through pipelines, the outlet of the compressor is communicated with the inlet of the condenser through a pipeline, and the outlet of the condenser is communicated with the top end of the solvent storage tank.
The invention has the positive effects that: (1) the method comprises the steps of taking water in a subcritical state as an extraction solvent, transferring polysaccharide components in materials into the liquid extraction solvent in a closed, oxygen-free and low-pressure extraction tank according to the principle that organic matters are similar and soluble mutually, separating the extraction solvent from polysaccharide through a reduced-pressure evaporation process, and removing impurities to finally obtain pure polysaccharide of Artemisia princeps. The polysaccharide has high dissolution rate and high dissolution speed. (2) Photons in the far infrared region have lower energy than photons in the visible and ultraviolet regions and therefore excite the polysaccharide molecules only in the form of stretching, bending, shaking and twisting, and these vibrations and molecular friction generate heat that raises the temperature of the system, thus allowing the polysaccharide to be dissolved out with high selectivity. The far infrared three-dimensional heating mode and the enhancement of the interaction between the sample and the solvent caused by molecular vibration greatly improve the extraction efficiency and obviously shorten the extraction time. In addition, far infrared radiation is compatible with both polar and non-polar solvents and is not harmful to the human body. (3) The ionic liquid is selectively combined with polysaccharide components in a sample by taking hydrophilic interaction, hydrogen bonds and steric hindrance effect as main acting force, so that the dissolving capacity of the extraction system for the polysaccharide is improved, and the selective extraction of the system for the polysaccharide is realized.
Therefore, the invention has the following advantages under the conditions of far infrared strengthening and ionic liquid assistance: the method has the advantages of no toxicity, environmental protection, non-thermal processing, oxidation prevention, no damage to the activity of the product, high extraction selectivity, high product purity, high yield, industrial large-scale production, energy conservation, low operation cost and the like, and has very wide application prospect.
Drawings
Fig. 1 is a schematic diagram of an extraction apparatus of the present application.
FIG. 2 is a glucose curve of example 1.
Detailed Description
Reference numerals: 1-solvent storage tank, 2-extraction tank, 3-evaporation tank, 4-collection bottle, 5-metering pump, 6-compressor, 7-condenser, 8-hot water tank, 9-hot water pump, 101-pressure gauge I, 102-pressure gauge II, 103-pressure gauge III, 111-temperature gauge I, 112-temperature gauge II, 113-temperature gauge III, 13-vacuum pump, 14-far infrared generator, 15-controller
A method for extracting Artemisia polysaccharide with high selectivity comprises the following steps:
step one), pretreatment: carrying out hot air drying on the artemisia leaf to enable the water content of the artemisia leaf raw material to be lower than 8%, and then crushing the artemisia leaf raw material into powder with the granularity of 40-60 meshes to obtain a pretreated artemisia leaf sample;
step two), extraction: placing the crushed folium artemisiae argyi powder into a closed extraction tank, and adding ionic liquid [ C ] according to 0.1-1% of the volume of an extraction solvent4mim]Cl、[C4C11OHim]CF3SO3、[C4mim]CF3SO3One of the folium artemisiae argyi and the folium artemisiae argyi is used as an entrainer, the extraction is carried out in a vacuum manner until the vacuum degree in an extraction tank reaches-100 kPa, distilled water is injected into the extraction tank to be used as an extraction solvent, the folium artemisiae argyi sample is completely immersed, the extraction conditions are that the mass-volume ratio of pretreated folium artemisiae argyi powder to water is 1: 5-15 g/ml, the extraction time is 5-10 minutes, the temperature is 105-115 ℃, the pressure is 0.1-0.2 Mpa, the extraction is carried out for 1-3 times in a circulating manner, wherein the extraction temperature is controlled by an automatic controller, the heat source is a far infrared generator arranged at the bottom of the extraction tank, the power is 500-1000;
step three), desolventizing: and after extraction is finished, pumping the liquid solvent into an evaporating pot, and respectively carrying out constant-temperature reduced-pressure desolventizing treatment on the folium artemisiae argyi material residues and the liquid solvent at the desolventizing temperature of 50-65 ℃ to obtain the extract after the liquid solvent is desolventized. The water vapor evaporated from the material slag and the liquid solvent flows back to the solvent storage tank after condensation and liquefaction for recycling.
Step four), alcohol precipitation: dissolving the extract and a 95% ethanol solution according to the mass volume ratio of 1: 3-4, standing in a refrigerator at 4 ℃ for 2-4 h, filtering, and freeze-drying to obtain the blumea polysaccharide.
Example 1 preparation of glucose Standard Curve and detection method of Artemisia polysaccharide
Detecting polysaccharide by adopting a phenol-sulfuric acid method, firstly drawing a glucose standard curve: preparing a mother solution with the concentration of 0.1 mg/mL from a glucose standard substance dried to constant weight at 105 ℃, performing step dilution, and respectively preparing standard solutions with the final concentrations of 0.01, 0.02, 0.03, 0.04 and 0.05 mg/mL; sequentially adding 1 mL of 5% phenol solution and 5 mL of concentrated sulfuric acid into 2 mL of standard solutions with different concentrations, uniformly mixing, carrying out boiling water bath for 10 min, taking out, cooling to room temperature, using distilled water as a blank, measuring a light absorption value at 490 nm, and recording data; taking the concentration of the glucose standard solution as an abscissa and the absorbance OD490 as an ordinate to prepare a standard curve (FIG. 2), and fitting to obtain a regression equation:
Y=13.205x+0.236
correlation coefficient:R 2= 0.98977
linear range: 0.01-0.05 mg/mL.
Detection of Artemisia polysaccharide: preparing 0.001 g of polysaccharide extract into 50.0 mug/mL aqueous solution, placing 2.0 mL into a 15 mL test tube with a plug, sequentially adding 1 mL of 5% phenol solution and 5 mL of concentrated sulfuric acid, uniformly mixing, carrying out boiling water bath for 10 min, taking out, cooling to room temperature, using distilled water as a blank, measuring the light absorption value at 490 nm, recording data, and substituting into a regression equation to obtain the concentration of the polysaccharide.
Example 2 extraction of polysaccharides by conventional Water extraction and alcohol precipitation
The method comprises the following steps of (A) carrying out the extraction of the artemisia argyi crude polysaccharide by using a literature report (Shenxia, Zhang Yanhong, Yuan Hui, and the like; research on optimizing the extraction process of the artemisia argyi crude polysaccharide by a response surface analysis method [ J ]. Chinese patent medicine, 2010, 32(1): 48-51.) and optimizing the obtained extraction optimal conditions of the artemisia argyi crude polysaccharide by the response surface analysis method: the leaching temperature is 99 ℃, the leaching time is 2.3 h, and the water-material ratio is 20. Centrifuging the extractive solution at 5000 rpm for 15 min; concentrating the centrifuged supernatant under reduced pressure, adding 4 times volume of 95% ethanol solution, placing in a refrigerator at 4 deg.C for 4 hr to completely precipitate polysaccharide, filtering, and freeze drying to obtain Artemisia princeps Pampanini polysaccharide.
The purity of the polysaccharide obtained was 284.61 mg/g, calculated as used in example 1.
Example 3 extraction of Artemisia polysaccharide by the method of the invention
Pulverizing folium Artemisiae Argyi with water content less than 8%, sieving with 40 mesh sieve, placing 100 g into extraction tank, vacuumizing until vacuum degree in the extraction tank reaches-100 kPa, injecting distilled water into the extraction tank as extraction solvent, completely immersing folium Artemisiae Argyi sample at a mass-volume ratio of folium Artemisiae Argyi and distilled water of 1:8 g/ml, and extracting with entrainer [ C ]4mim]8 g of Cl ionic liquid, controlling the extraction temperature to be 110 ℃ by the cooperation of an automatic controller and a far infrared generating device at the bottom of the extraction tank, controlling the far infrared power to be 1000W, controlling the wavelength range to be 25-200 mu m, controlling the extraction pressure to be 0.15 Mpa, controlling the extraction time to be 6 minutes, and circularly extracting for 2 times; after extraction is finished, pumping the liquid solvent into an evaporating pot, and respectively carrying out constant-temperature decompression desolventizing treatment on the folium artemisiae argyi material residues and the liquid solvent, wherein the desolventizing temperature is 65 ℃; dissolving the desolventized extract with 95% ethanol solution at a mass-to-volume ratio of 1:3, standing in a refrigerator at 4 deg.C for 2 h, filtering, and freeze-drying to obtain Artemisia princeps Pampanini polysaccharide.
The purity of the polysaccharide obtained was 761.48mg/g, calculated as used in example 1. The result of comparative example 2 can show that, the extraction efficiency is significantly improved by performing the present embodiment under the subcritical condition and combining far infrared irradiation and ionic liquid entrainment, and the extraction time is shortened from the original 2.3 h to 6 minutes for 2 times of circulation. Meanwhile, the selectivity of the extraction process to the polysaccharide of the Artemisia princeps is obviously enhanced, and the content of the polysaccharide in the extract is improved from 284.61 mg/g to 761.48 mg/g.
Example 4 extraction of Artemisia polysaccharide by the method of the invention
Pulverizing folium Artemisiae Argyi with water content less than 8%, sieving with 40 mesh sieve, placing 100 g into extraction tank, vacuumizing until vacuum degree in the extraction tank reaches-100 kPa, injecting distilled water into the extraction tank as extraction solvent, completely immersing folium Artemisiae Argyi sample at a mass-volume ratio of folium Artemisiae Argyi and distilled water of 1:10 g/ml, and extracting with entrainer [ C ]4C11OHim]CF3SO310g of ionic liquid, the extraction temperature is controlled at 105 ℃ by the cooperation of an automatic controller and a far infrared generating device at the bottom of the extraction tank, the far infrared power is 800W, the wavelength range is 25-200 mu m, the extraction pressure is 0.10 Mpa, the extraction time is 8 minutes, and the extraction is circulated for 2 times; after extraction is finished, pumping the liquid solvent into an evaporating pot, and respectively carrying out constant-temperature decompression desolventizing treatment on the folium artemisiae argyi material residues and the liquid solvent, wherein the desolventizing temperature is 65 ℃; dissolving the desolventized extract with 95% ethanol solution at a mass-to-volume ratio of 1:4, standing in a refrigerator at 4 deg.C for 3 h, filtering, and freeze-drying to obtain Artemisia princeps Pampanini polysaccharide.
The purity of the polysaccharide obtained was 865.26 mg/g, calculated as used in example 1. The result of the second comparative example shows that the extraction efficiency is remarkably improved by performing the second embodiment under the subcritical condition and combining far infrared irradiation and ionic liquid entrainment, and the extraction time is shortened from the original 2.3 h to 8 min for 2 times of circulation. Meanwhile, the selectivity of the extraction process to the polysaccharide of the Artemisia princeps is obviously enhanced, and the content of the polysaccharide in the extract is improved from 284.61 mg/g to 865.26 mg/g.
The results of the above examples show that compared with the traditional water extraction and alcohol precipitation method, the method of the invention has simple process, the extracted material residue can not be denatured, and the further processing and utilization can not be influenced; molecular vibration excited by far infrared and intermolecular interaction between ionic liquid and a target product enable polysaccharide in a sample to be dissolved out in a high selectivity manner, and the content of polysaccharide in an extract is increased; in addition, the interaction between the sample and the solvent caused by a far infrared three-dimensional heating mode and molecular vibration is enhanced, so that the extraction efficiency is greatly improved, and the extraction time is obviously shortened.
Although the present invention has been described with reference to the preferred embodiments, it is not intended to be limited to the embodiments shown, and the invention is applicable to other types of samples. Various changes and modifications can be made by one skilled in the art without departing from the spirit and scope of the invention, and the scope of the invention should be determined by the appended claims.
Claims (7)
1. A method for extracting Artemisia polysaccharide with high selectivity is characterized in that: the method comprises the following steps:
step one), pretreatment: before extraction, drying the raw material of the artemisia vulgaris L, and then crushing the dried material of the artemisia vulgaris L into artemisia vulgaris L powder with the granularity of 40-60 meshes;
step two), extraction: placing the pretreated folium Artemisiae Argyi powder in a sealed extraction tank, vacuumizing until the vacuum degree in the extraction tank reaches-100 kPa, injecting distilled water into the extraction tank as extraction solvent, completely immersing folium Artemisiae Argyi sample, and adding ionic liquid entrainer [ C ] for extraction4mim]Extracting after Cl, wherein the amount of the entrainer is 0.1-1% of the volume of the extraction solvent;
the extraction temperature is controlled by an automatic controller, the heat source is a far infrared generating device arranged at the bottom of the extraction tank, a glass window is arranged at the bottom of the extraction tank, so that far infrared rays penetrate through the extraction tank, the power is 500-1000W, and the far infrared wavelength range is 25-200 mu m;
step three), desolventizing: after extraction, pumping the liquid solvent into an evaporating pot, and respectively carrying out constant-temperature reduced-pressure desolventizing treatment on the material residues in the extracting pot and the liquid solvent in the evaporating pot to obtain a crude extract after desolventizing the liquid solvent;
step four), alcohol precipitation: dissolving the above extract with 95% ethanol solution, placing in refrigerator at 4 deg.C to precipitate polysaccharide completely, filtering, and freeze drying to obtain Artemisia princeps Pampanini polysaccharide;
the mass-volume ratio of the pretreated folium artemisiae argyi powder to water in the step two) is 1: 5-15 g/ml, the extraction time is 5-10 minutes, the temperature is 105-115 ℃, the pressure is 0.1-0.2 Mpa, and the cyclic extraction is performed for 1-3 times.
2. The method for extracting polysaccharide of Artemisia princeps according to claim 1, wherein: the water content of the dried folium artemisiae argyi raw material in the step one) is lower than 8%.
3. The method for extracting polysaccharide of Artemisia princeps according to claim 1, wherein: the entrainer for extraction in the step two) adopts [ C ]4C11OHim]CF3SO3And [ C4mim]CF3SO3The entrainer is 0.1-1% of the volume of the extraction solvent.
4. The method for extracting polysaccharide of Artemisia princeps according to claim 1, wherein: the temperature of the decompression desolventizing in the step three) is 50-65 ℃.
5. The method for extracting Artemisia polysaccharide with high selectivity according to any one of claims 1-4, wherein: and thirdly), condensing and liquefying the water vapor evaporated from the material slag and the liquid solvent, and then returning the condensed and liquefied water vapor to the solvent storage tank for recycling.
6. The method for extracting Artemisia polysaccharide with high selectivity according to any one of claims 1-4, wherein: the mass volume ratio of the extract to the 95% ethanol solution in the step four) is 1: 3-4, and the standing time of the extract in a refrigerator at 4 ℃ is 2-4 h.
7. The method for extracting Artemisia polysaccharide with high selectivity according to claim 5, wherein: the structure that the water vapor evaporated from the material slag and the liquid solvent flows back to the solvent storage tank after being condensed and liquefied is as follows: the extraction tank and the evaporating tank are communicated with the inlet of the compressor through pipelines, the outlet of the compressor is communicated with the inlet of the condenser through a pipeline, and the outlet of the condenser is communicated with the top end of the solvent storage tank.
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Address after: 455000 west section of the Yellow River Road, Anyang, Henan Province, Anyang Institute of Technology Applicant after: Anyang Institute of Technology Applicant after: Tangyin County Medical Industry Management Office Applicant after: Henan subcritical extraction technology Research Institute Co., Ltd. Address before: 455000 the Yellow River, Anyang, Henan reaches the western part of Anyang Institute of Technology. Applicant before: Anyang Institute of Technology Applicant before: Tangyin County Medical Industry Management Office Applicant before: Henan subcritical extraction technology Research Institute Co., Ltd. |
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| CP01 | Change in the name or title of a patent holder | ||
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Address after: 455000 Anyang Institute of technology, west section of Huanghe Avenue, Anyang City, Henan Province Patentee after: ANYANG INSTITUTE OF TECHNOLOGY Patentee after: Tangyin Pharmaceutical Industry Development Service Center Patentee after: HENAN YALINJIE EXTRACTION TECHNOLOGY RESEARCH INSTITUTE Co.,Ltd. Address before: 455000 Anyang Institute of technology, west section of Huanghe Avenue, Anyang City, Henan Province Patentee before: ANYANG INSTITUTE OF TECHNOLOGY Patentee before: TANGYIN COUNTY PHARMACEUTICAL INDUSTRY MANAGEMENT OFFICE Patentee before: HENAN YALINJIE EXTRACTION TECHNOLOGY RESEARCH INSTITUTE Co.,Ltd. |