CN116211909B - Chinese mugwort polyphenol extract with lipid-lowering effect and preparation method and application thereof - Google Patents
Chinese mugwort polyphenol extract with lipid-lowering effect and preparation method and application thereof Download PDFInfo
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- CN116211909B CN116211909B CN202310291691.4A CN202310291691A CN116211909B CN 116211909 B CN116211909 B CN 116211909B CN 202310291691 A CN202310291691 A CN 202310291691A CN 116211909 B CN116211909 B CN 116211909B
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- Prior art keywords
- mugwort
- lipid
- lowering effect
- polyphenol extract
- polyphenol
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Classifications
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- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
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- A61K36/185—Magnoliopsida (dicotyledons)
- A61K36/28—Asteraceae or Compositae (Aster or Sunflower family), e.g. chamomile, feverfew, yarrow or echinacea
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
- A23L33/105—Plant extracts, their artificial duplicates or their derivatives
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- A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
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- A61K47/26—Carbohydrates, e.g. sugar alcohols, amino sugars, nucleic acids, mono-, di- or oligo-saccharides; Derivatives thereof, e.g. polysorbates, sorbitan fatty acid esters or glycyrrhizin
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- A—HUMAN NECESSITIES
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- A61K2236/30—Extraction of the material
- A61K2236/33—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones
- A61K2236/333—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones using mixed solvents, e.g. 70% EtOH
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- A—HUMAN NECESSITIES
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- A—HUMAN NECESSITIES
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- Proteomics, Peptides & Aminoacids (AREA)
- Bioinformatics & Cheminformatics (AREA)
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Abstract
The invention discloses a Chinese mugwort polyphenol extract with lipid-lowering effect, and a preparation method and application thereof. The preparation method comprises the following steps: ultrasonic-assisted extraction is carried out on mugwort powder by taking eutectic solvent as extraction solvent, and then, the mugwort polyphenol extract with lipid-lowering effect is obtained by filtering, concentrating and drying; wherein the eutectic solvent is choline chloride-sorbitol. Compared with a chemical synthesis method, the method has the advantages of good biodegradability, low cost, high safety and the like, and provides a basis for large-scale production and high-value utilization of the mugwort; the invention takes the Chinese mugwort Ai Aishe as a development object, extracts the polyphenol substances therein, proves that the Chinese mugwort has the lipid-lowering effect, expands the application of the Chinese mugwort Ai Aishe, improves the economic benefit of the Chinese mugwort Ai Aishe, and has important significance for the intensive processing of the Chinese mugwort and the application in the health field.
Description
Technical Field
The invention relates to the field of foods, medicines or health care products, in particular to a Chinese mugwort polyphenol extract with lipid-lowering effect, and a preparation method and application thereof.
Background
With the improvement of living standard, people have more and more intake of high-sugar and high-fat food, which causes metabolic disorder of human body, and thus causes increased risks of obesity, diabetes, cardiovascular diseases and the like. According to the report of nutrition and chronic disease status of Chinese residents (2020), the overweight rate and the obesity rate of the Chinese residents are respectively 34.3 percent and 16.4 percent by 2020, and the overall overweight rate of adult residents is more than 50 percent. Overweight and obesity can cause a series of chronic diseases and even death, and obesity can induce diseases such as hypertension, hyperlipidemia, atherosclerosis and the like, causing serious disease burden and economic burden.
Currently, the main prevention and treatment modes for obesity include surgery, medicines, sports, diet improvement and the like. Research shows that a series of weight-reducing medicines in the current market, including phentermine, fenframine and the like, are forbidden due to the fact that the weight-reducing medicines can cause side effects of cardiovascular diseases, hyperthyroidism and the like, while lipid-reducing medicines approved for eating, such as orlistat, luo Kaxi lin and the like, can cause adverse reactions, such as insomnia, constipation, dizziness and the like, so that the weight-reducing and lipid-reducing medicines have good effects, but the side effects caused by the weight-reducing and lipid-reducing medicines cannot be ignored, and the long-term taking safety is low. Thus, lipid-lowering active ingredients derived from natural plants are attracting attention. Compared with the side effect brought by the traditional medicine, the natural plant active ingredient not only has good lipid-lowering effect, but also has higher safety and other health benefits. Therefore, the development of natural plant-derived lipid-lowering products is one of the effective measures for lowering lipid. Natural active ingredients that have been shown to have lipid lowering effects include polyphenols, flavones, terpenes, alkaloids and the like. Wherein the polyphenol compounds are widely reported functional components with various activities, and have remarkable effects on lipid lowering, antioxidation, anti-inflammatory and other aspects.
The mugwort is an economic crop planted in Qichun county of Hubei province in China, is a national geographic marking product in China, is used as a traditional medicine and food homologous plant in China, and has long medicinal and edible histories. At present, the research on the mugwort is mainly focused on the volatile oil of the mugwort, which is one of the main curative effect components of the mugwort for Chinese medicine moxibustion, and the report on the main physiological activity of the mugwort is mainly focused on the aspects of antioxidation, anti-inflammation, bacteriostasis, immunity enhancement and the like, and the activity of the mugwort on lipid reduction is not reported.
Disclosure of Invention
The invention aims to overcome the technical defects, and provides a mugwort polyphenol extract with lipid-lowering effect, a preparation method and application thereof, and the technical problem of insufficient activity of mugwort in lipid-lowering in the prior art is solved.
In a first aspect, the invention provides a preparation method of a Chinese mugwort polyphenol extract with lipid-lowering effect, which comprises the following steps: ultrasonic-assisted extraction is carried out on mugwort powder by taking eutectic solvent as extraction solvent, and then, the mugwort polyphenol extract with lipid-lowering effect is obtained by filtering, concentrating and drying; wherein the eutectic solvent is choline chloride-sorbitol.
In a second aspect, the invention provides a mugwort polyphenol extract with lipid-lowering effect, which is obtained by the preparation method of the mugwort polyphenol extract with lipid-lowering effect provided by the first aspect.
In a third aspect, the invention provides application of the mugwort polyphenol extract with lipid-lowering effect in the fields of foods, medicines or health care products.
Compared with the prior art, the invention has the beneficial effects that:
The invention adopts ultrasonic wave auxiliary eutectic solvent method to extract polyphenol substances in mugwort leaf, the used eutectic solvent is choline chloride-sorbitol mixture, compared with chemical synthesis method, the method has the advantages of good biodegradability, low cost, high safety and the like, and provides basis for mugwort mass production and high-value utilization; the invention takes the Chinese mugwort Ai Aishe as a development object, extracts the polyphenol substances therein, proves that the Chinese mugwort has the lipid-lowering effect, expands the application of the Chinese mugwort Ai Aishe, improves the economic benefit of the Chinese mugwort Ai Aishe, and has important significance for the intensive processing of the Chinese mugwort and the application in the health field.
Drawings
FIG. 1 is a photograph of C.elegans oil red O-stain from a blank control group, a high fat control group, and a group treated with an extract of Agents.
Detailed Description
The present invention will be described in further detail with reference to the drawings and examples, in order to make the objects, technical solutions and advantages of the present invention more apparent. It should be understood that the specific embodiments described herein are for purposes of illustration only and are not intended to limit the scope of the invention.
In a first aspect, the invention provides a preparation method of a Chinese mugwort polyphenol extract with lipid-lowering effect, which comprises the following steps: ultrasonic-assisted extraction is carried out on mugwort powder by taking eutectic solvent as extraction solvent, and then, the mugwort polyphenol extract with lipid-lowering effect is obtained by filtering, concentrating and drying; wherein the eutectic solvent is choline chloride-sorbitol.
The invention utilizes ultrasonic wave auxiliary eutectic solvent method to separate polyphenol-rich extract from mugwort leaf, uses choline chloride-sorbitol eutectic solvent as extraction solvent, and obtains mugwort polyphenol extract through ultrasonic extraction, concentration and drying. Compared with chemical synthesis method, the method has the advantages of good biodegradability, low cost, high safety, and the like, and provides a basis for large-scale production and high-value utilization of mugwort.
In this embodiment, the mugwort powder is obtained by drying and grinding mugwort Ai Aishe into powder.
In this embodiment, choline chloride-sorbitol is prepared by mixing choline chloride and sorbitol in a molar ratio of 1:1 to 1:3, mixing and stirring until the mixture is colorless and transparent.
Preferably, the molar ratio of choline chloride to sorbitol is 1:2.
Preferably, the temperature of mixing and stirring is 70-90 ℃, preferably 80 ℃; the mixing and stirring time is 5 to 7 hours, preferably 5 hours.
In this embodiment, the feed liquid ratio of mugwort powder to eutectic solvent is 1: 20-1: 50g/mL, preferably 1:30g/mL.
In the embodiment, in the ultrasonic-assisted extraction process, the extraction temperature is 40-55 ℃, preferably 45 ℃; the ultrasonic time is 2-4 hours, preferably 3 hours; the ultrasonic power is 300-450W, preferably 350W.
In this embodiment, the drying method is freeze drying.
In a second aspect, the invention provides a mugwort polyphenol extract with lipid-lowering effect, which is obtained by the preparation method of the mugwort polyphenol extract with lipid-lowering effect provided by the first aspect.
The main components of the mugwort polyphenol extract provided by the invention are chlorogenic acid, apigenin, luteolin, quercetin, kaempferol, brown cyanidin, eupatorium adenophorum and caffeic acid, and the mugwort polyphenol extract has good lipid-lowering effect. The extract is derived from natural plants, and has the advantages of safety, no toxicity, less side effects, etc.
In a third aspect, the invention provides application of the mugwort polyphenol extract with lipid-lowering effect in the fields of foods, medicines or health care products.
The mugwort polyphenol extract disclosed by the invention can obviously reduce the fat content in a high-fat caenorhabditis elegans model body, has a good lipid-lowering effect, and has potential application in the aspects of preparing foods, medicines and health-care products with lipid-lowering effects.
In the present embodiment, the dosage forms of the food, the medicine, the health care product include pills, powders, capsules, tablets, syrups, soft capsules, granules and the like, and the present invention is not limited thereto.
Example 1: preparation of mugwort polyphenol extract
Step 1: treatment of mugwort raw materials: drying fresh folium Artemisiae Argyi in oven at 60deg.C for 24 hr, pulverizing, and sieving with 60 mesh sieve to obtain folium Artemisiae Argyi powder.
Step 2: preparation of the eutectic solvent: choline chloride and sorbitol are in a molar ratio of 1:1, and stirring at 80deg.C for 5 hr to colorless and transparent to obtain extractive solvent.
Step 3: extracting: 500g of mugwort powder is weighed, and the feed liquid ratio is 1: adding 20g/mL of the extraction solvent in the step 2, extracting at 40 ℃ and ultrasonic power of 300W for 2 hours, and combining and concentrating the obtained extracting solution.
Step 4: and (3) drying: and (3) placing the concentrated extract in the step (3) into a freezing plate, pre-freezing for 12 hours at the temperature of minus 20 ℃, and drying in a freeze dryer to obtain the Chinese mugwort polyphenol freeze-dried powder.
Example 2: preparation of mugwort polyphenol extract
Step 1: treatment of mugwort raw materials: drying fresh folium Artemisiae Argyi in oven at 60deg.C for 24 hr, pulverizing, and sieving with 60 mesh sieve to obtain folium Artemisiae Argyi powder.
Step 2: preparation of the eutectic solvent: choline chloride and sorbitol are in a molar ratio of 1:2, mixing, stirring at 80deg.C for 5 hr to colorless and transparent to obtain extractive solvent.
Step 3: extracting: 500g of mugwort powder is weighed, and the feed liquid ratio is 1:30g/mL of the extraction solvent in the step 2 is added, the extraction is carried out for 3 hours at 45 ℃ under the ultrasonic power of 350W, and the obtained extracting solutions are combined and concentrated.
Step 4: and (3) drying: and (3) placing the concentrated extract in the step (3) into a freezing plate, pre-freezing for 12 hours at the temperature of minus 20 ℃, and drying in a freeze dryer to obtain the Chinese mugwort polyphenol freeze-dried powder.
Example 3: preparation of mugwort polyphenol extract
Step 1: treatment of mugwort raw materials: drying fresh folium Artemisiae Argyi in oven at 60deg.C for 24 hr, pulverizing, and sieving with 60 mesh sieve to obtain folium Artemisiae Argyi powder.
Step 2: preparation of the eutectic solvent: choline chloride and sorbitol are in a molar ratio of 1:2, mixing, stirring at 80deg.C for 7 hr to colorless and transparent to obtain extractive solvent.
Step 3: extracting: 500g of mugwort powder is weighed, and the feed liquid ratio is 1: adding 40g/mL of the extraction solvent in the step 2, extracting for 4 hours at 50 ℃ under the ultrasonic power of 400W, and combining and concentrating the obtained extracting solution.
Step 4: and (3) drying: and (3) placing the concentrated extract in the step (3) into a freezing plate, pre-freezing for 12 hours at the temperature of minus 20 ℃, and drying in a freeze dryer to obtain the Chinese mugwort polyphenol freeze-dried powder.
Example 4: preparation of mugwort polyphenol extract
Step 1: treatment of mugwort raw materials: drying fresh folium Artemisiae Argyi in oven at 60deg.C for 24 hr, pulverizing, and sieving with 60 mesh sieve to obtain folium Artemisiae Argyi powder.
Step 2: preparation of the eutectic solvent: choline chloride and sorbitol are in a molar ratio of 1:3 mixing, stirring at 80deg.C for 5 hr to colorless and transparent to obtain extractive solvent.
Step 3: extracting: 500g of mugwort powder is weighed, and the feed liquid ratio is 1: adding 50g/mL of the extraction solvent in the step 2, extracting for 4 hours at 55 ℃ under the ultrasonic power of 450W, and combining and concentrating the obtained extracting solution.
Step 4: and (3) drying: and (3) placing the concentrated extract in the step (3) into a freezing plate, pre-freezing for 12 hours at the temperature of minus 20 ℃, and drying in a freeze dryer to obtain the Chinese mugwort polyphenol freeze-dried powder.
Test group 1: determination of polyphenol content in mugwort polyphenol extract
Step 1: drawing a standard curve: gallic acid (GALLIC ACID) is used as a standard substance, methanol solution with the volume fraction of 50% is used for preparing standard substances with different concentrations, 500 mu L of standard substance solution is mixed with 250 mu L of 50% Fu Lin Fen reagent and then reacted for 5min, 500 mu L of 5% sodium carbonate solution is added for light-proof reaction for 1h, absorbance is measured at 765nm, the concentration of gallic acid is taken as an abscissa, and absorbance is taken as an ordinate, so that a standard curve is drawn.
Step 2: determination of polyphenol content in mugwort polyphenol extract: after mixing 500. Mu.L of the sample with 250. Mu.L of 50% Fu Lin Fen reagent, reacting for 5min, adding 500. Mu.L of 5% sodium carbonate solution, reacting for 1h in the dark, measuring absorbance at 765nm, and calculating total phenol content in the sample according to a standard curve, wherein the total phenol content is expressed as MG GALLIC ACID/g sample. The results are shown in Table 1:
TABLE 1 total phenol content in extracts of different examples
As can be seen from Table 1, the choline chloride-sorbitol molar ratio, the extraction temperature, the feed liquid ratio and the ultrasonic power in the examples have a great influence on the total phenol content in the prepared mugwort polyphenol extract, wherein the total phenol content in the mugwort extract prepared in the example 2 is the highest.
Test group 2: lipid-lowering activity test of mugwort polyphenol extract
Step 1: accurately weighing appropriate amount of the powder of the mugwort polyphenol extract obtained in the example 2, dissolving with DMSO to prepare 60mg/mL mother liquor, and storing at-20deg.C for use.
Step 2: enlarged culture of caenorhabditis elegans: and (3) coating a proper amount of E.coli OP50 bacterial suspension on the central position of the NGM flat plate, drying by a super clean bench, and placing in a 37 ℃ incubator for inversion culture for 24 hours. After the long bacterial NGM plate is cooled to normal temperature, the nematode needle is used to pick the needed nematode strain, and the nematode strain is inoculated on the NGM plate and cultured at 20 ℃. When a large number of nematodes grow on the NGM plate, the end of the metal spoon is burned by using an alcohol burner flame, the culture medium with the nematodes is cut and connected to a new NGM plate, and the culture is continued at 20 ℃.
Step 3: synchronization of caenorhabditis elegans: the nematodes in spawning period are washed from the NGM plate with M9 buffer solution, put into a 1.5mL centrifuge tube, the supernatant is discarded after standing, and the bacteria adhered to the nematodes are removed by repeatedly washing with M9 three times. After the last time, adding 1mL of nematode lysate into a centrifuge tube, rapidly and severely oscillating for 15min until the nematode body is completely cracked, centrifuging at 10000rpm/min for 1min at normal temperature, discarding the supernatant, washing the obtained precipitate with M9 buffer solution for 4 times, finally dripping M9 buffer solution containing nematode eggs on an NGM plate coated with E.coli OP50, airing in an ultra-clean bench, and culturing in a culture box at 20 ℃ to obtain the nematode offspring at the same period.
Step four: establishment of a high-fat dietary nematode model: culturing in 96-well plates, adding 100 mu LS-complex culture medium into each well, picking up wild nematodes synchronized to the L4 stage into the S-complex culture medium by using a picking needle, and picking up 10-15 nematodes per well, wherein the time for picking up the nematodes is set as day 0. 200 mu L M of buffer solution is added to each of the outermost circles of the 96-well plates to prevent water evaporation, and the plates are sealed and then placed into 20 ℃ for culture. Wherein a blank control group containing only DMSO, a high-fat control group containing only 2% D-glucose, and a mugwort polyphenol-treated group containing 240 μg/mL mugwort polyphenol extract and 2% D-glucose were set. And culturing nematodes until adults are third day, collecting nematodes of different treatment groups, washing with M9 buffer solution for 3 times to remove adhered mixed bacteria, fully grinding on ice, centrifuging at 10000rpm/min for 5min at 4 ℃, measuring triglyceride content in supernatant of nematode homogenate according to Triglyceride (TG) kit specification steps of Nanjing built biosciences, and repeating each experiment three times, wherein the results are shown in Table 2.
As shown in Table 2, compared with the blank control group, the content of TG in the caenorhabditis elegans in the high-fat control group added with 2% of D-glucose in the diet is obviously improved by 2.15 times that of the blank control group, and after 240 mug/mL of the mugwort polyphenol extract is added, the content of TG in the caenorhabditis elegans is obviously reduced by 68.3% of that of the high-fat control group, so that the mugwort polyphenol extract has better lipid-lowering effect and can be applied to product development with lipid-lowering effect.
TABLE 2 Triglycerides in caenorhabditis elegans of different treatment groups
| TG(mmol/g Protein) | |
| Blank control group | 0.98±0.04a |
| High fat control group | 2.15±0.18c |
| Chinese mugwort polyphenol treatment group | 1.47±0.26b |
Test group 3: determination of lipid-lowering effect of mugwort polyphenol extract
Step 1: a proper amount of the mugwort polyphenol extract powder obtained in the example 2 is accurately weighed, dissolved by DMSO to prepare 60mg/mL mother liquor, and stored at-20 ℃ for standby.
Step 2: the amplification and synchronization of caenorhabditis elegans was identical to that described in test group 2.
Step 3: staining of the high-fat caenorhabditis elegans oil red O: when the nematodes are cultured until the third day, N2 nematodes of different treatment groups are collected, washed three times with M9 buffer solution, added with 2% paraformaldehyde for fixing by 100 mu L, then the centrifuge tube is put into liquid nitrogen, freeze thawing is repeated three times, after the last thawing, 1500rpm/min at 4 ℃, centrifuging for 1min, washing 2 times with M9 buffer solution, after the supernatant is sucked off, 1mL of 60% isopropanol is added for dewatering for 15min, the supernatant is centrifuged off, 1mL of 60% oil red O staining solution (100 mL of isopropanol is dissolved in 60 ℃ water bath for 0.5g of oil red O powder to prepare stock solution, and then water is added for preparing 60% working solution), shaking is carried out overnight, and light-shielding staining is carried out. After staining, the centrifuge tube was removed, centrifuged at 1500rpm/min at 4℃for 1min, the staining solution was removed, and M9 was added for 2 washes to remove excess flooding, the nematodes were transferred to a slide, and the staining of the nematode droplets was observed under a microscope and photographed. The results are shown in FIG. 1.
As can be seen from figure 1, the mugwort polyphenol extract of the invention has better lipid-lowering effect.
In conclusion, the polyphenol extracted by utilizing the ultrasonic-assisted eutectic solvent method has a good extraction effect, and the extract can obviously reduce the triglyceride content in the in-vivo model biological caenorhabditis elegans, which shows that the extract has the potential of improving lipid metabolism disorder and losing weight. The invention proves that the mugwort polyphenol extract has good lipid-lowering effect in-vivo model organisms for the first time. The invention not only realizes the recycling of the by-products of the Chinese mugwort processing industry, but also has positive regulation effect on lipid metabolism disorder in a organism and improves the health.
The above-described embodiments of the present invention do not limit the scope of the present invention. Any other corresponding changes and modifications made in accordance with the technical idea of the present invention shall be included in the scope of the claims of the present invention.
Claims (6)
1. A preparation method of a Chinese mugwort polyphenol extract with lipid-lowering effect is characterized by comprising the following steps: ultrasonic-assisted extraction is carried out on mugwort powder by taking eutectic solvent as extraction solvent, and then, the mugwort polyphenol extract with lipid-lowering effect is obtained by filtering, concentrating and drying; wherein,
The eutectic solvent is prepared by mixing choline chloride and sorbitol according to a molar ratio of 1:1 to 1:3, mixing and stirring until the mixture is colorless and transparent; the temperature of the mixing and stirring is 70-90 ℃, and the time of the mixing and stirring is 5-7 h; the feed liquid ratio of the mugwort powder to the eutectic solvent is 1: 20-1: 50 g/mL; in the ultrasonic auxiliary extraction process, the extraction temperature is 40-55 ℃, the ultrasonic time is 2-4 h, and the ultrasonic power is 300-450W.
2. The method for preparing the mugwort polyphenol extract with lipid-lowering effect according to claim 1, wherein the molar ratio of choline chloride to sorbitol is 1:2.
3. The method for preparing a mugwort polyphenol extract with lipid-lowering effect according to claim 1, wherein the drying mode is freeze drying.
4. The method for preparing a mugwort polyphenol extract having lipid-lowering effect according to claim 1, wherein the mugwort powder is obtained by drying mugwort leaves and grinding the mugwort leaves into powder.
5. The mugwort polyphenol extract with lipid-lowering effect, which is characterized by being obtained by the preparation method of the mugwort polyphenol extract with lipid-lowering effect according to any one of claims 1 to 4.
6. The use of the extract of argyi polyphenol with lipid-lowering effect as claimed in claim 5 for preparing lipid-lowering medicines.
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