CN112545926A - Comprehensive extraction method of folium artemisiae argyi volatile oil and folium artemisiae argyi total flavonoids and oxidation resistance evaluation of folium artemisiae argyi volatile oil - Google Patents
Comprehensive extraction method of folium artemisiae argyi volatile oil and folium artemisiae argyi total flavonoids and oxidation resistance evaluation of folium artemisiae argyi volatile oil Download PDFInfo
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Abstract
The invention discloses a comprehensive extraction method of folium artemisiae argyi volatile oil and folium artemisiae argyi total flavonoids, and the method is used for evaluating the oxidation resistance of the folium artemisiae argyi volatile oil, belongs to the technical field of comprehensive utilization of plant resources, and comprises the following steps: drying and pulverizing folium Artemisiae Argyi, extracting volatile oil by steam distillation over water, and purifying with GPerforming qualitative and quantitative analysis on folium Artemisiae Argyi volatile oil by C-MS to obtain folium Artemisiae Argyi volatile oil components, comparing with Vc, and performing antioxidant evaluation on folium Artemisiae Argyi volatile oil to obtain folium Artemisiae Argyi volatile oil with good antioxidant property and IC capable of eliminating DPPH50It was 5.7 mg/mL. Performing orthogonal optimization ethanol heating reflux extraction on the folium artemisiae argyi residues subjected to steam distillation, concentrating and drying by rotary evaporation to obtain folium artemisiae argyi total flavonoids, wherein under the optimal extraction condition, the purity of the crude folium artemisiae argyi total flavonoids extracted once reaches 44.1%, and the yield reaches 9.21%.
Description
Technical Field
The invention belongs to the technical field of comprehensive utilization of plant resources, and particularly relates to a comprehensive extraction method of folium artemisiae argyi volatile oil and folium artemisiae argyi total flavonoids and an evaluation method of the oxidation resistance of the folium artemisiae argyi volatile oil.
Background
The wormwood has a close relationship with the life of people in China, is recorded in compendium of materia Medica and new materia Medica, is a traditional Chinese medicine in China, and is usually picked and dried in the sun when flowers are not bloomed and leaves are luxuriant in summer. The folium artemisiae argyi is pungent, bitter and warm in nature, has little toxicity, enters liver, spleen and kidney channels, has the effects of warming the channels to stop bleeding, dispelling cold to stop pain, regulating menstruation and preventing miscarriage, removing dampness to relieve itching, clearing and activating the channels and collaterals and the like, is a medicinal and edible traditional Chinese medicinal material, is high in safety, low in price and easy to obtain, and has wide development and utilization values. The folium Artemisiae Argyi contains volatile oil, 1, 8-cineole, folium Artemisiae Argyi flavone, etc. Modern experiments prove that the folium artemisiae argyi has the effects of resisting bacteria and viruses, relieving asthma and cough, eliminating phlegm, stopping bleeding and resisting coagulation, calming and resisting allergy, protecting liver and benefiting gallbladder and the like. The volatile oil of the folium artemisiae argyi is the basis of main drug effect substances of the folium artemisiae argyi and is regarded as the evaluation standard of the quality of the folium artemisiae argyi medicinal material; the flavonoids compounds in the folium artemisiae argyi have strong effects of resisting oxidation and removing free radicals, and have good medical care value.
At present, the extraction of folium artemisiae argyi volatile oil is reported more, and the extraction is mainly carried out by organic solvent extraction, ultrasonic-assisted extraction, enzyme-assisted extraction and the like. Chinese patent CN111454767A discloses an extraction method of folium artemisiae argyi essential oil, wherein an enzyme is adopted to assist in extraction to obtain folium artemisiae argyi volatile oil crude product, and folium artemisiae argyi is not comprehensively utilized in the whole process, so that the waste of total flavonoids is caused. Chinese patent CN201410445559.5 discloses an extraction method and application of folium artemisiae argyi volatile oil, wherein the folium artemisiae argyi volatile oil is extracted by a reduced pressure distillation method assisted by a biological enzymolysis technology, and the volatile oil is suitable for preparing microcapsule products. Chinese patent CN111358821A discloses a method for performing optimized enzymolysis by a response surface method to assist ultrasonic extraction of folium artemisiae argyi total flavonoids, drying and crushing folium artemisiae argyi, adding enzyme into folium artemisiae argyi powder for enzymolysis, obtaining filter residue and filtrate by using the optimized enzymolysis process conditions of the response surface method, adding ethanol into the filter residue for ultrasonic extraction to obtain ethanol extract, combining the ethanol extract and the enzymolysis filtrate, concentrating by a rotary evaporator, and drying in a vacuum drying oven to obtain the folium artemisiae argyi total flavonoids.
Firstly, extracting folium artemisiae argyi volatile oil by steam distillation through water-proof distillation, and then extracting folium artemisiae argyi total flavonoids from folium artemisiae argyi residues; the antioxidant application of the folium artemisiae argyi volatile oil is considered, and no report or patent for simultaneously extracting the folium artemisiae argyi volatile oil and the folium artemisiae argyi total flavonoids exists at present. The method synchronously extracts the volatile oil and the total flavonoids from the folium artemisiae argyi, is easy to operate, mild in extraction conditions, convenient in post-treatment, low in cost, suitable for expanded production, fully utilizes folium artemisiae argyi resources, and has important significance for application and development of the folium artemisiae argyi in the fields of medicines, health care products, cosmetics and the like.
Disclosure of Invention
The invention aims to provide a method for comprehensively extracting folium artemisiae argyi volatile oil and total flavonoids and an oxidation resistance evaluation of the folium artemisiae argyi volatile oil. The preparation method is simple, the process is green and environment-friendly, large-scale production can be realized, the post-treatment is convenient, and the cost is low; the yield of the volatile oil and the total flavonoids of the folium artemisiae argyi is high, the purity of a crude extract is high, and the comprehensive development and utilization of folium artemisiae argyi resources are improved; the extracted volatile oil of folium Artemisiae Argyi has good oxidation resistance, and can be applied in the fields of medicine, health product and cosmetic.
The invention comprises the following steps:
1. extraction of volatile argyi leaf oil
Cutting folium artemisiae argyi into small sections, crushing the small sections by using a crusher, sieving the small sections by using a sieve of 100 meshes, weighing 10-100 g of folium artemisiae argyi powder, adding the folium artemisiae argyi powder into a water-proof distillation device, adding 2L of distilled water, heating and distilling the folium artemisiae argyi powder in a water-proof way, extracting the folium artemisiae argyi volatile oil for 1-3 hours when the first drop of liquid flows down, performing extraction separation to obtain folium artemisiae argyi volatile oil, and performing GC-MS (;
2. evaluation of antioxidant Properties of volatile oil of Artemisia princeps Pampanini
(1) DPPH (1, 1-diphenyl picryl phenylhydrazine) free radical scavenging capacity experimental method
Preparing folium artemisiae argyi volatile oil solutions with certain concentrations by using absolute ethyl alcohol respectively as sample solutions; respectively taking 2.0 mL of sample liquid and 2.0 mL of DPPH free radical solution with a certain concentration, uniformly mixing, standing in the dark for 0.5-1 h, taking absolute ethyl alcohol as a reference, and determining the light absorption value A; 2.0 mL of sample solutionMixing with 2.0 mL of absolute ethyl alcohol, and measuring the light absorption value of the mixture to be A0(ii) a The light absorption value measured by 2.0 mL of DPPH solution and 2.0 mL of absolute ethyl alcohol mixed solution is A1. By VC (ascorbic acid) as a positive control.
(2) Total reducing power determination experimental method
Respectively preparing folium Artemisiae Argyi volatile oil solution and V with anhydrous ethanolCThe solution was used as a sample solution. Taking 3-5 mL of two sample solutions, adding 3-5 mL of PBS (0.1 mol. L)-1Phosphate buffer solution) buffer and 1% K3Fe(CN)6 And (3) preserving the heat of the solution for 20-30 min in a water bath at 50-70 ℃, and quickly taking out and cooling after heat preservation is finished. Adding 2-4 mL of 10% trichloroacetic acid solution, and centrifuging at 2000-4000 r/min for 10-15 min. Taking 2-4 mL of supernatant, and adding 2-4 mL of distilled water and 0.1% FeCl respectively3The solution (0.5 mL) was allowed to stand for 10 min, and the absorbance at 700 nm was measured.
3. Method for extracting total flavonoids from folium artemisiae argyi residues
(1) Adding folium artemisiae argyi residues and an ethanol solution in a certain ratio into a 250 mL round-bottom flask, heating the mixture in a water bath at the temperature of 60-80 ℃ for 1-3 hours to obtain an folium artemisiae argyi residue extracting solution, wherein the material-liquid ratio is 1: 20-1: 30, and the mass concentration of ethanol is 50% -80%;
(2) filtering folium Artemisiae Argyi residue extractive solution, performing rotary evaporation to obtain concentrated solution, and oven drying the concentrated solution in vacuum drying oven to obtain crude flavone extract.
(3) The content of the crude flavone extract is measured by a rutin standard curve, and the optimal extraction process obtained by an orthogonal test is as follows.
Description of the drawings:
FIG. 1 is a schematic diagram showing the effect of extraction time on the yield of volatile oil of folium Artemisiae Argyi.
FIG. 2 shows a chromatogram obtained by gas chromatography-mass spectrometry of volatile folium Artemisiae Argyi oil.
FIG. 3 is a schematic diagram of DPPH free radical scavenging ability of folium Artemisiae Argyi volatile oil.
FIG. 4 is a schematic diagram of the determination of the total reducing power of the volatile oil of folium Artemisiae Argyi.
The specific implementation mode is as follows:
the technical solutions provided by the present invention are described in detail below with reference to examples, and it should be understood that the following detailed description is only illustrative and not intended to limit the scope of the present invention.
Example 1: extraction of folium artemisiae argyi volatile oil and antioxidant evaluation thereof
(1) Cutting folium Artemisiae Argyi into small segments, pulverizing with a pulverizer, sieving with 100 mesh sieve, weighing 50 g folium Artemisiae Argyi powder, adding into a water-proof distillation device, adding 2L distilled water, heating and distilling over water, timing with the first drop of liquid flow, extracting for 2 h, extracting and separating to obtain folium Artemisiae Argyi volatile oil, and performing GC-MS qualitative and quantitative analysis on the folium Artemisiae Argyi volatile oil;
(2) an experimental method for DPPH free radical scavenging ability comprises preparing folium Artemisiae Argyi volatile oil solution (8 mg. mL) with anhydrous ethanol-1) As a sample liquid; 2.0 mL of the sample solution and 2.0 mL of the DPPH radical solution (1.0X 10)- 4mol·L-1) Mixing, standing in dark for 0.5 hr, and measuring absorbance A with anhydrous ethanol as reference; 2.0 mL of sample liquid is mixed with 2.0 mL of absolute ethyl alcohol, and the light absorption value is measured to be A0(ii) a The light absorption value measured by 2.0 mL of DPPH solution and 2.0 mL of absolute ethyl alcohol mixed solution is A1. By VCUsed as a positive control.
(3) Total reducing power determination experiment method, preparing folium Artemisiae Argyi volatile oil solution (8 mg. mL) with anhydrous ethanol-1) And VCSolution (0.8 mg. mL)-1) As a sample liquid. 4 mL of PBS (0.1 mol. L) was added to 4 mL of the two sample solutions-1) Buffer and 1% K3Fe(CN)6 And (3) preserving the temperature of the solution in a water bath at 60 ℃ for 30 min, and quickly taking out the solution for cooling after the heat preservation is finished. 3 mL of 10% trichloroacetic acid solution was added, followed by centrifugation at 4000 r/min for 15 min. 2 mL of the supernatant was taken, and 2 mL of distilled water and 0.1% FeCl were added to the supernatant3The solution (0.5 mL) was allowed to stand for 10 min, and the absorbance at 700 nm was measured.
Example 2: method for extracting total flavonoids from folium artemisiae argyi residues
(1) After drying the folium artemisiae argyi residues in the embodiment 1, adding 5 g of the folium artemisiae argyi residues into a 250 mL round-bottom flask, heating the mixture in a water bath at 70 ℃ for 2 hours at a material-liquid ratio of 1:30 and an ethanol mass concentration of 60% to obtain an folium artemisiae argyi residue extracting solution;
(2) filtering folium Artemisiae Argyi residue extractive solution, performing rotary evaporation to obtain concentrated solution, and oven drying the concentrated solution in vacuum drying oven to obtain crude flavone extract with purity of 44.1%;
(3) the content of the crude flavone extract is measured by a rutin standard curve, and the optimal extraction process obtained by an orthogonal test is as follows: the mass concentration of ethanol is 60%, the water bath heating time is 2 h, the extraction temperature is 70 ℃, and the feed-liquid ratio is 1:30 g/ml, so that the yield of the folium artemisiae argyi residue total flavonoids is 9.21%.
The effect of extraction time on extraction yield can be seen in figure 1, with an optimal time of 2.5 h.
FIG. 2 shows a chromatogram obtained by gas chromatography-mass spectrometry of volatile folium Artemisiae Argyi oil, wherein the content of caryophyllene is 14.37% at most.
FIG. 3 is a schematic diagram of the DPPH free radical scavenging ability of folium Artemisiae Argyi volatile oil, and the obtained folium Artemisiae Argyi volatile oil has good oxidation resistance and DPPH scavenging IC50It was 5.7 mg/mL.
FIG. 4 is a schematic diagram of the determination of the total reducing power of the volatile oil of folium Artemisiae Argyi, and the determination shows that the volatile oil of folium Artemisiae Argyi has good total reducing power.
In conclusion, according to the invention, the folium artemisiae argyi volatile oil is extracted by water-proof distillation, the folium artemisiae argyi volatile oil is obtained through different extraction time and extraction modes, qualitative and quantitative analysis is carried out on the folium artemisiae argyi volatile oil through GC-MS to obtain the components of the folium artemisiae argyi volatile oil, and then the antioxidant determination is carried out on the folium artemisiae argyi volatile oil to obtain the folium artemisiae argyi volatile oil which has good inoxidizability and is IC for removing50It was 5.7 mg/mL. Because the damage of steam distillation to the folium artemisiae argyi is low, orthogonal optimization ethanol heating reflux extraction conditions are carried out on folium artemisiae argyi residues, folium artemisiae argyi total flavonoids are obtained through rotary evaporation, concentration and drying, and under the optimized extraction conditions, the purity of the crude folium artemisiae argyi total flavonoids reaches 44.1%, and the yield reaches 9.21%. The process has high extraction efficiency, simple process and no toxicity, fully utilizes folium Artemisiae Argyi resources, improves the added value of products, and provides theoretical and experimental basis for comprehensive utilization of folium Artemisiae Argyi.
Claims (3)
1. A comprehensive extraction method of folium artemisiae argyi volatile oil and folium artemisiae argyi total flavonoids and an oxidation resistance evaluation of the folium artemisiae argyi volatile oil are characterized by comprising the following steps:
and (3) extracting folium artemisiae argyi volatile oil, cutting folium artemisiae argyi into small sections, crushing the small sections by using a crusher, sieving the small sections with a 100-mesh sieve, weighing 10-100 g of folium artemisiae argyi powder, adding the folium artemisiae argyi powder into a water-proof distillation device, adding 2L of distilled water, heating and distilling the folium artemisiae argyi powder in a water-proof way, extracting the folium artemisiae argyi volatile oil for 1-3 hours from the first drop of liquid flow, performing extraction separation to obtain the folium.
2. Evaluation of antioxidant Properties of the Artemisia princeps Pampanini volatile oil obtained in claim 1
Evaluation of DPPH radical scavenging ability: respectively preparing folium artemisiae argyi volatile oil solution with a certain concentration by using absolute ethyl alcohol to serve as sample liquid, respectively and uniformly mixing 2.0 mL of sample liquid and 2.0 mL of DPPH free radical solution with a certain concentration, standing in the dark for 0.5-1 h, using the absolute ethyl alcohol as reference, determining the light absorption value to be A, mixing 2.0 mL of sample liquid and 2.0 mL of absolute ethyl alcohol, and determining the light absorption value to be A0And the light absorption value measured by a mixed solution of 2.0 mL of DPPH solution and 2.0 mL of absolute ethyl alcohol is A1By VCAs a positive control, comparing the DPPH free radical scavenging capacity of the folium artemisiae argyi volatile oil and Vc; evaluation of total reducing power: respectively preparing folium Artemisiae Argyi volatile oil solution and V with anhydrous ethanolCTaking 3-5 mL of two sample solutions as sample solutions, adding 3-5 mL of 0.1 mol.L-1PBS buffer and 1% K3Fe(CN)6 The solution is subjected to heat preservation in a water bath at 50-70 ℃ for 20-30 min, and is rapidly taken out and cooled after the heat preservation is finished, 2-4 mL of 10% trichloroacetic acid solution is added, then the solution is centrifuged at 2000-4000 r/min for 10-15 min, 2-4 mL of supernatant is taken, and 2-4 mL of distilled water and 0.1% FeCl are respectively added3Standing for 10 min with 0.5 mL of solution, measuring absorbance at 700 nm, and comparing folium Artemisiae Argyi volatile oil with Vc total reducing ability.
3. The method for extracting total flavonoids from folium Artemisiae Argyi residue after extracting volatile oil from folium Artemisiae Argyi of claim 1
Adding a certain proportion of dried folium artemisiae argyi residues and an ethanol solution into a 250 mL round-bottom flask, heating in a water bath at the temperature of 60-80 ℃ for 1-3 hours to obtain an folium artemisiae argyi residue extracting solution, wherein the material-liquid ratio is 1: 20-1: 30, and the mass concentration of ethanol is 50-80%; filtering folium Artemisiae Argyi residue extractive solution to obtain filtrate, performing rotary evaporation to obtain concentrated solution, and oven drying the concentrated solution in vacuum drying oven to obtain crude flavone extract.
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