CN116602959B - Composition liposome containing tea active ingredient and having anti-inflammatory and analgesic effects and application thereof - Google Patents
Composition liposome containing tea active ingredient and having anti-inflammatory and analgesic effects and application thereof Download PDFInfo
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- CN116602959B CN116602959B CN202310592122.3A CN202310592122A CN116602959B CN 116602959 B CN116602959 B CN 116602959B CN 202310592122 A CN202310592122 A CN 202310592122A CN 116602959 B CN116602959 B CN 116602959B
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- tea
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Classifications
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/33—Heterocyclic compounds
- A61K31/335—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin
- A61K31/35—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having six-membered rings with one oxygen as the only ring hetero atom
- A61K31/352—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having six-membered rings with one oxygen as the only ring hetero atom condensed with carbocyclic rings, e.g. methantheline
- A61K31/353—3,4-Dihydrobenzopyrans, e.g. chroman, catechin
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/01—Hydrocarbons
- A61K31/015—Hydrocarbons carbocyclic
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/33—Heterocyclic compounds
- A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
- A61K31/40—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with one nitrogen as the only ring hetero atom, e.g. sulpiride, succinimide, tolmetin, buflomedil
- A61K31/409—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with one nitrogen as the only ring hetero atom, e.g. sulpiride, succinimide, tolmetin, buflomedil having four such rings, e.g. porphine derivatives, bilirubin, biliverdine
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/185—Magnoliopsida (dicotyledons)
- A61K36/82—Theaceae (Tea family), e.g. camellia
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
- A61K47/24—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite containing atoms other than carbon, hydrogen, oxygen, halogen, nitrogen or sulfur, e.g. cyclomethicone or phospholipids
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
- A61K47/28—Steroids, e.g. cholesterol, bile acids or glycyrrhetinic acid
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/10—Dispersions; Emulsions
- A61K9/127—Liposomes
- A61K9/1277—Processes for preparing; Proliposomes
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
Landscapes
- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Medicinal Chemistry (AREA)
- Pharmacology & Pharmacy (AREA)
- Animal Behavior & Ethology (AREA)
- General Health & Medical Sciences (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Epidemiology (AREA)
- Natural Medicines & Medicinal Plants (AREA)
- Engineering & Computer Science (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Oil, Petroleum & Natural Gas (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Gastroenterology & Hepatology (AREA)
- Molecular Biology (AREA)
- Pain & Pain Management (AREA)
- Rheumatology (AREA)
- Biophysics (AREA)
- Organic Chemistry (AREA)
- Dispersion Chemistry (AREA)
- Alternative & Traditional Medicine (AREA)
- Biotechnology (AREA)
- Botany (AREA)
- Medical Informatics (AREA)
- Microbiology (AREA)
- Mycology (AREA)
- Medicinal Preparation (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
The invention discloses a composition liposome containing tea active ingredients and having anti-inflammatory and analgesic effects and application thereof. The invention provides a composition containing tea active ingredients with anti-inflammatory and analgesic effects, which mainly comprises epigallocatechin gallate, carotene and chlorophyll, and has excellent synergistic effect, good anti-inflammatory effect and analgesic effect, obvious reduction of inflammatory reaction and small irritation to human skin; the liposome prepared from the active ingredients greatly improves the stability of the active ingredients in aqueous solution, improves the bioavailability, and can exert the efficacy of the composition containing the active ingredients of tea to a greater extent. Compared with the traditional method of using single component of green tea or using crude extract of green tea, the composition containing active ingredients of the tea provided by the invention has better anti-inflammatory and analgesic effects and higher bioavailability, and provides a research thought and basis for preparing anti-inflammatory and analgesic medicines of green tea.
Description
Technical Field
The invention belongs to the technical field of biological medicines, and particularly relates to a composition liposome containing tea active ingredients and having anti-inflammatory and analgesic effects and application thereof.
Background
Inflammation is a defensive response of human tissue with vascular system to injury factors, mainly manifested by local tissue degeneration (denaturation, necrosis), exudation (vascular response, fluid and cell exudation) and proliferation changes, and simultaneously, the inflammation tissue can cause swelling congestion and increased exudates of local tissue, press nerve end, and cause pain. It was found that the anti-inflammatory effect of the natural product is mainly expressed as that inflammatory swelling or granuloma caused by chemical and physical stimulus can be significantly suppressed, and that the general inducibility of blood capillaries of the ventral brain and hand of the mouse after inflammation can be reduced daily, and that the analgesic effect of suppressing the expression of the inflammatory factor and its gene can be generally expressed as that of prolongation of pain threshold of various pain-causing factors such as thermal pain, tenderness and chemical stimulus. However, the existing clinical anti-inflammatory and analgesic drugs have certain toxic and side effects or adverse reactions, such as coagulation disorders, cardiovascular and cerebrovascular diseases, nephrotoxicity, liver injury, granulocyte deficiency, gastrointestinal injury and the like, besides exerting anti-inflammatory and analgesic effects.
Tea leaves are the main source of beneficial plant ingredients in the human diet and are processed from camellia japonica. Tea originates in China and is one of the most widely consumed nonalcoholic beverages at present. Green tea, one of the main teas in china, accounts for 20% of the global tea consumption. The green tea is prepared from new leaves or buds of tea tree without fermentation, and through the processes of deactivation of enzymes, shaping, drying and the like, and retains natural substances of the fresh leaves, and contains nutrient components such as tea polyphenol, catechin, chlorophyll, caffeine, amino acids, vitamins and the like. The green color and tea soup are more preserved with the green pattern of fresh tea, thus obtaining the name. Green tea contains many bioactive substances, and key bioactive components include catechin, theanine, caffeine, etc., and catechin substances have been widely studied for their health protection effects against oxidation, inflammation, cancer, obesity, diabetes, etc.
The tea extract is widely applied to the fields of food, health care, medicine, daily chemical industry and the like, but most of active ingredients in the tea are volatile and unstable in property and are easily influenced by factors such as temperature, pH, metal ions and the like to undergo degradation, oxidization, isomerization and other reactions, so that the original physiological activity of the tea is changed, and the effect is very limited. Therefore, the use of green tea is still lacking in medicine, and the green tea crude extract is usually used in the use process, so that the use efficiency and pertinence are not high, and the effect is still to be improved.
Disclosure of Invention
The invention aims to solve the problems in the prior art, and provides a composition liposome containing tea active ingredients and an application thereof, wherein the composition liposome containing the tea active ingredients can be prepared to effectively improve the stability of the composition in aqueous solution, so that the bioavailability of the composition is improved, and the composition has a strong anti-inflammatory and analgesic effect.
The first object of the invention is to provide a composition liposome containing tea active ingredients with anti-inflammatory and analgesic effects.
The second object of the invention is to provide a preparation method of a composition liposome containing tea active ingredients and having anti-inflammatory and analgesic effects.
The third object of the invention is to provide the application of the liposome of the composition containing the tea active ingredient and having the anti-inflammatory and analgesic effects.
A fourth object of the present invention is to provide a medicament with anti-inflammatory and/or analgesic properties
The invention provides a composition liposome containing tea active ingredients with anti-inflammatory and analgesic effects, which consists of liposome and a composition containing tea active ingredients encapsulated in the liposome, wherein the composition containing tea active ingredients comprises the following components in parts by weight: 5-7 parts of epigallocatechin gallate, 1-4 parts of carotene and 1-4 parts of chlorophyll.
Epigallocatechin gallate (Epigallocatechin gallate, EGCG) is polyphenols in tea, and has anticancer effect. The catechin monomer is separated from tea leaves, has very strong antioxidant activity, can protect cells and DNA from damage, and also has the functions of resisting tumor, deodorizing, preventing caries, resisting ultraviolet radiation and inhibiting bacteria.
Carotene (Carotene) is a main vitamin A source substance, is an organic compound with large molecular weight, can be dissolved in most organic reagents, is insoluble in water, has extremely poor stability in water, and is easy to oxidize and change color. The carotene enters the body and can be converted into vitamin A, so that the functions of improving eyesight, protecting and nourishing the liver can be achieved, the metabolism of cells in the body can be promoted, the ultraviolet-resistant function is achieved, sunburn can be prevented in summer, and the effects of resisting oxidization and delaying aging can be achieved.
Chlorophyll (Chlorophyl) is a green pigment that higher plants and all other organisms capable of photosynthesis contain. Free chlorophyll is very unstable and can be decomposed by light, acid, alkali, oxygen, oxidant, etc., and is soluble in solvents such as ethanol, diethyl ether and acetone, and insoluble in water. Chlorophyll has anticancer, antibacterial, and antioxidant effects, and can be used in medical, food, and cosmetic industries.
The composition containing the tea active ingredients adopts epigallocatechin gallate, carotene and chlorophyll, has poor stability of three substances, is extremely easy to oxidize or degrade, is in aqueous solution and unstable, and can greatly improve the stability of the three substances in aqueous solution and remarkably improve the treatment effect by coating the three main active ingredients in liposome. The components in the liposome of the composition containing the tea active ingredients have good synergistic effect, and provide a basis for preparing anti-inflammatory and analgesic medicines containing the tea active ingredients.
Preferably, the mass fraction ratio of the liposome to the composition containing the tea active ingredient is (85-110): (7-15).
Preferably, the composition containing the tea active ingredients comprises the following components in parts by mass: 5-6 parts of epigallocatechin gallate, 1-3 parts of carotene and 2-4 parts of chlorophyll.
Preferably, the liposome adopts the following mass ratio (4-8): 1 and cholesterol, and is prepared by a film dispersion method.
Preferably, the liposome is prepared from the following raw materials in parts by weight: 70-80 parts of lecithin, 10-20 parts of cholesterol and 5-10 parts of tween 80.
In particular, a thin film dispersion method is adopted to prepare liposome, and the ester-alcohol ratio (the mass ratio of phospholipid to cholesterol) of the liposome is controlled to be 4-8:1, the particle size of the prepared liposome is 0.13-0.2 mu m, the encapsulation rate is as high as 87%, the prepared green tea liposome has better anti-inflammatory and analgesic effects, the stability of the components can be enhanced through the encapsulation of the liposome, the problem of unstable water solubility of the green tea composition is solved, the bioavailability is improved, and the efficacy of the green tea composition can be exerted to a greater extent.
More preferably, the mass ratio of the phospholipid to the cholesterol is 4:1.
Preferably, the liposome comprises the following raw materials in parts by mass: 80 parts of lecithin, 20 parts of cholesterol and 10 parts of tween 80.
The invention provides a preparation method of a composition liposome containing tea active ingredients, which comprises the steps of mixing the composition containing the tea active ingredients with liposome raw materials, adding an organic solvent, oscillating and dissolving, removing the organic solvent, adding a water-soluble buffer solution after the lipid forms a film, oscillating and shaking uniformly, carrying out ultrasonic treatment, filtering by a membrane, centrifuging, and obtaining the green tea composition liposome, and storing in a dark place.
Preferably, chloroform is used as the organic solvent; the water-soluble buffer solution was a PBS buffer solution having a pH of 7.4.
Preferably, the organic solvent is removed by rotary evaporation at a temperature of 40-55deg.C and a rotation speed of 45-70rpm/min.
More preferably, the condition of the rotary evaporation is that the temperature is 45 ℃ and the rotating speed is 70rpm/min.
Preferably, the condition of the ultrasonic wave is that the ultrasonic power is 250-300w and the time is 3-5min.
More preferably, the condition of the ultrasonic treatment is ultrasonic power 300w for 3min.
Preferably, the filter membrane is a filter membrane of 0.45 μm and 0.22 μm in this order.
Preferably, the centrifugation conditions are 7000-10000rpm/min and the centrifugation time is 20-40min.
More preferably, the centrifugation conditions are 8000rpm/min and the centrifugation time is 30min.
As a most preferred embodiment, the present invention provides a method for preparing a liposome of a composition containing tea active ingredients having anti-inflammatory and analgesic effects, comprising the steps of:
epigallocatechin gallate (5-7 parts), chlorophyll (1-4 parts), carotene (1-4 parts), lecithin (80 parts), cholesterol (20 parts) and tween 80 (10 parts) are uniformly mixed, added into chloroform solution (20 parts) under the condition of room temperature, oscillated and dissolved, and subjected to negative pressure evaporation by a rotary evaporator under a certain environment, organic solvent is removed, after lipid forms a film on the wall, PBS buffer solution with PH=7.4 is added, oscillation and shaking are carried out, then the obtained liposome solution is subjected to ultrasonic treatment, and then the prepared liposome solution is sequentially subjected to a filter membrane with 0.45 mu m and a filter membrane with 0.22 mu m, and finally the liposome composition containing tea active ingredients is obtained through centrifugation. Specifically, the rotary evaporation temperature is set to 45 ℃, the rotary speed is 70rpm/min, the ultrasonic power is 300w, the time is 3min, the centrifugal condition is 8000rpm/min, the centrifugal time is 30min, and the materials are stored at 4 ℃ and protected from light. After the preparation, the prepared liposome composition containing the tea active ingredient is stored in a glass bottle and stored in a refrigerator.
Liposomes are an artificial membrane. The hydrophilic head of phospholipid molecule is inserted into water, the hydrophobic tail of liposome is extended to air, and after stirring, spherical liposome of double-layer lipid molecule is formed, and its diameter is 25-1000 nm. The liposome can be used for transgenosis or preparing medicines, and the characteristics that the liposome can be fused with cell membranes are utilized to send the medicines into cells. The composition containing the tea active ingredients is prepared into the liposome, so that the liposome has excellent anti-inflammatory and analgesic effects, has less stimulation to human bodies, greatly improves the stability of three substance components in aqueous solution, simultaneously improves the bioavailability, has better effect, and provides a foundation for preparing the green tea anti-inflammatory and analgesic medicines.
Therefore, the invention also provides application of the composition liposome containing the tea active ingredient in preparing anti-inflammatory and/or analgesic drugs.
The invention also provides an anti-inflammatory and/or analgesic drug comprising the composition liposome containing the tea active ingredient.
The invention has the following beneficial effects:
The invention provides a composition liposome containing tea active ingredients with anti-inflammatory and analgesic effects, which mainly comprises epigallocatechin gallate, carotene and chlorophyll as active ingredients; the three water-insoluble tea extracts are combined, the stability of the three substances in aqueous solution is greatly improved by a liposome wrapping method, the treatment effect is remarkably improved, the liposome prepared after wrapping has excellent anti-inflammatory and analgesic effects, and the wrapped composition reduces the irritation to human bodies. In addition, the composition containing the tea active ingredients has a synergistic enhancement effect, and has better anti-inflammatory and analgesic effects and higher bioavailability than the traditional method using a single green tea ingredient or using a green tea crude extract.
Detailed Description
The invention is further illustrated below in connection with specific examples which are intended to be illustrative of the invention and are not to be construed as limiting the invention, the specific techniques or conditions not identified in the examples being according to the techniques or conditions described in the literature in this field or according to the product specifications.
The purity of the epigallocatechin gallate adopted by the invention is 98%, and CAS number 989-51-5; chlorophyll purity 85%, CAS number 479-61-8; the purity of the carotene is 96%, and CAS number 7235-40-7; lecithin, cholesterol, tween 80 were all purchased from commercial products.
Example 1
A preparation method of a composition liposome containing tea active ingredients with anti-inflammatory and analgesic effects comprises the following steps: the following components in parts by mass: 5 parts of epigallocatechin gallate, 2 parts of chlorophyll, 3 parts of carotene, 80 parts of lecithin, 10 parts of cholesterol and 5 parts of tween 80 are uniformly mixed, added into 20 parts of chloroform solution at room temperature, oscillated and dissolved, and subjected to negative pressure evaporation by a rotary evaporator, wherein the rotary evaporation temperature is 45 ℃, the rotating speed is 70rpm/min, the organic solvent is removed, 50 parts of PBS buffer solution with PH of 7.4 is added after the lipid forms a film on the wall, oscillation and shaking are carried out, then the obtained liposome solution is subjected to ultrasonic treatment, the ultrasonic power is 300w for 3min, the prepared liposome solution is sequentially subjected to a filter membrane with 450 mu m and 200 mu m, and then subjected to centrifugation for 30min at 8000rpm/min, so that the composition liposome containing tea active ingredients is obtained as an experiment group 1.
Example 2
A preparation method of a composition liposome containing tea active ingredients with anti-inflammatory and analgesic effects comprises the following steps: the following components in parts by mass: mixing epigallocatechin gallate 5 parts, chlorophyll 3 parts, carotene 2 parts, lecithin 80 parts, cholesterol 10 parts and tween 80 5 parts uniformly, adding into chloroform solution 20 parts at room temperature, oscillating for dissolution, performing negative pressure evaporation by using a rotary evaporator, removing organic solvent at a rotary evaporation temperature of 45 ℃ and a rotation speed of 70rpm/min, adding PBS buffer solution with pH of 7.4 into 50 parts after lipid forms a film on the wall, oscillating for shaking uniformly, performing ultrasonic treatment on the obtained liposome solution for 3min at an ultrasonic power of 300w, sequentially passing the prepared liposome solution through a filter membrane with a thickness of 450 mu m and a filter membrane with a thickness of 200 mu m, and centrifuging to obtain a composition liposome containing tea active ingredients as an experimental group 2.
Example 3
A preparation method of a composition liposome containing tea active ingredients with anti-inflammatory and analgesic effects comprises the following steps: the following components in parts by mass: 6 parts of epigallocatechin gallate, 2 parts of chlorophyll, 2 parts of carotene, 80 parts of lecithin, 10 parts of cholesterol and 5 parts of tween 80 are uniformly mixed, 20 parts of chloroform solution is added under the condition of room temperature, vibration dissolution is carried out, a rotary evaporator is used for negative pressure evaporation, the rotary evaporation temperature is 45 ℃, the rotating speed is 70rpm/min, the organic solvent is removed, after the lipid forms a film on the wall, 50 parts of PBS buffer solution with PH of 7.4 is added for shaking, the obtained liposome solution is subjected to ultrasonic treatment for 3min at the ultrasonic power of 300w, the prepared liposome solution is sequentially filtered by a filter membrane with the wavelength of 450 mu m and 200 mu m, and is centrifuged for 30min at 8000rpm/min, so that the composition liposome containing the tea active ingredients is obtained as an experiment group 3.
Example 4
A preparation method of a composition liposome containing tea active ingredients with anti-inflammatory and analgesic effects comprises the following steps: the following components in parts by mass: 6 parts of epigallocatechin gallate, 1 part of chlorophyll, 3 parts of carotene, 80 parts of lecithin, 10 parts of cholesterol and 5 parts of tween 80 are uniformly mixed, 20 parts of chloroform solution is added under the condition of room temperature, vibration dissolution is carried out, a rotary evaporator is used for negative pressure evaporation, the rotary evaporation temperature is 45 ℃, the rotating speed is 70rpm/min, the organic solvent is removed, after the lipid forms a film on the wall, 50 parts of PBS buffer solution with PH of 7.4 is added for shaking, the obtained liposome solution is subjected to ultrasonic treatment for 3min at the ultrasonic power of 300w, the prepared liposome solution is sequentially filtered by a filter membrane with the wavelength of 450 mu m and 200 mu m, and is centrifuged for 30min at 8000rpm/min, so that the composition liposome containing tea active ingredients is obtained as experiment group 4.
Example 5
A preparation method of a composition liposome containing tea active ingredients with anti-inflammatory and analgesic effects comprises the following steps: the following components in parts by mass: 7 parts of epigallocatechin gallate, 2 parts of chlorophyll, 1 part of carotene, 80 parts of lecithin, 20 parts of cholesterol and 10 parts of tween 80 are uniformly mixed, 20 parts of chloroform solution is added at room temperature, vibration dissolution is carried out, a rotary evaporator is used for negative pressure evaporation, the rotary evaporation temperature is 45 ℃, the rotating speed is 70rpm/min, the organic solvent is removed, after the lipid forms a film on the wall, 50 parts of PBS buffer solution with PH of 7.4 is added, vibration shaking is carried out, then the obtained liposome solution is subjected to ultrasonic treatment, the ultrasonic power is 300w for 3min, the prepared liposome solution is sequentially subjected to a filter membrane with 450 mu m and 200 mu m, and then subjected to centrifugation for 30min at 8000rpm/min, so that a composition liposome containing tea active ingredients is obtained as experiment group 5.
Example 6
A preparation method of a composition liposome containing tea active ingredients with anti-inflammatory and analgesic effects comprises the following steps: the following components in parts by mass: 7 parts of epigallocatechin gallate, 1 part of chlorophyll, 2 parts of carotene, 80 parts of lecithin, 20 parts of cholesterol and 10 parts of tween 80 are uniformly mixed, added into 20 parts of chloroform solution at room temperature, oscillated and dissolved, and subjected to negative pressure evaporation by a rotary evaporator, wherein the rotary evaporation temperature is 45 ℃, the rotating speed is 70rpm/min, the organic solvent is removed, 50 parts of PBS buffer solution with PH of 7.4 is added after the lipid forms a film on the wall, oscillation and shaking are carried out, then the obtained liposome solution is subjected to ultrasonic treatment for 3min at ultrasonic power of 300w, the prepared liposome solution is sequentially subjected to a filter membrane with 450 mu m and 200 mu m, and then subjected to centrifugation for 30min at 8000rpm/min, so that a composition liposome containing tea active ingredients is obtained as experiment group 6.
Comparative example 1
The comparative example was conducted in the same manner as in example 1 except that chlorophyll and carotene were not used, and only 10 parts of epigallocatechin gallate was used, except that the same conditions were used as in control 1.
Comparative example 2
The comparative example was conducted in the same manner as in example 1 except that no epigallocatechin gallate and no carotene were used, and only 10 parts of chlorophyll was used, except that the other conditions were the same, as in control group 2.
Comparative example 3
The comparative example was conducted in the same manner as in example 1 except that no epigallocatechin gallate and chlorophyll were used, and only 10 parts of carotene was used, except that the other conditions were the same, as in control group 3.
Comparative example 4
The comparative example was conducted in the same manner as in example 1 except that 5 parts and 5 parts of chlorophyll were used as the control group 4 without using epigallocatechin gallate.
Comparative example 5
The comparative example was conducted in the same manner as in example 1 except that chlorophyll was not used, and only 5 parts of epigallocatechin gallate and 5 parts of carotene were used, except that the same conditions were used as in control group 5.
Comparative example 6
The comparative example was conducted in the same manner as in example 1 except that no carotenes were used, and only 5 parts of epigallocatechin gallate and 5 parts of chlorophyll were used, except that the same conditions were used as in control 6.
Comparative example 7
The comparative example the method for preparing the liposome composition containing the tea active ingredient was the same as in example 1, except that 80 parts of lecithin, 20 parts of cholesterol, 10 parts of tween 80 were not used for encapsulation, and the other conditions were the same as in control group 7.
Test experiment 1 anti-inflammatory Performance index detection
The anti-inflammatory properties of the liposomes of the compositions containing tea active ingredient prepared in examples 1 to 6 (i.e., experimental groups 1 to 6) and comparative examples 1 to 7 (i.e., control groups 1 to 7) were tested. Healthy Kunming male mice (purchased from Guangzhou carrot biotechnology Co., ltd.) were selected 78 and randomly divided into 13 groups of 6. In the experiment, 0.1 part of dimethylbenzene is smeared on the inner side and the outer side of the right ear canal of each animal in each group until the animal is inflamed, and the left ear is not treated to be used as a blank control group. After 30min of xylene application, 0.1 g/animal sample of each group was uniformly applied to the inner and outer surfaces of the right ear canal of each animal. After only administering the test object for 1h, the mice were killed by dislocation of the cervical vertebrae, the two ear galleries were cut off, the test object sample of the right ear gallery was washed off with physiological saline, and the two round ear pieces were chiseled at the same position with a puncher, and the weight of the right ear piece minus the weight of the left ear piece was the swelling value, and statistical analysis was performed.
The results are shown in table 1, and show that the auricle swelling value of the experimental group 3 is obviously lower than that of other groups, the anti-inflammatory effect of the experimental group is better in each experimental group, and the auricle swelling value of each experimental group is obviously lower than that of the control group, so that the combined and compounded epigallocatechin gallate, chlorophyll and carotene can obviously improve inflammation, and the improvement effect of the experimental group is better than that of the control group 7 which is not subjected to liposome wrapping by adopting a single component or two components.
Table 1 mouse swelling value
Test experiment 2 analgesic effect detection
The liposome compositions containing the tea active ingredients prepared in the examples 1-6 and the comparative examples 1-7 are subjected to a mouse wound torsion experiment, and the specific treatment method is as follows: mice were anesthetized with 5% chloral hydrate, full-cortical wounds with a diameter of 5mm were made on the backs of the mice after anesthesia, drug was administered at the wounds at a concentration of 0.5mg/ml for each group of drug liposome, the dose was 30 μl, after 30min of treatment, the wound sites of the mice were dropped with 10 parts per kg of 1% glacial acetic acid solution for observation of the average value of the torsion response times of the mice within 15min, and the analgesic rate was calculated, with the analgesic rate=1- (drug administration torsion response times/blank torsion response times) ×100%.
The statistical results are shown in table 2, and show that the pain rate of the experimental group 3 is obviously higher than that of the other groups, and the pain rate of each experimental group is obviously higher than that of the control group, so that the experimental group has a synergistic effect after the epigallocatechin gallate, the chlorophyll and the carotene are compounded for use, and the effect is better than that of the control group 7 which is not wrapped in liposome or one or two of the components.
Table 2 statistics of analgesic rate for each treatment group
Further detection shows that the particle size of the liposome prepared in the embodiments 1-6 is 0.13-0.2 mu m, the encapsulation efficiency is as high as 87%, and compared with the comparison group 7 without liposome encapsulation, the liposome composition containing the tea active ingredient provided by the invention has better anti-inflammatory and analgesic effects, and the encapsulation of the liposome can enhance the stability of the components, especially solve the water-solubility problem of carotene and chlorophyll, and improve the stability of epigallocatechin gallate, chlorophyll and carotene in water solution, and the bioavailability, and can play the effect of the green tea composition to a greater extent.
The invention provides a research flow of a preparation method of a composition liposome containing tea active ingredients with anti-inflammatory and analgesic effects, and a plurality of methods and ways for realizing the technical scheme are provided, the above is only a feasible implementation mode of the invention, and it should be pointed out that a plurality of improvements and modifications can be made by one of ordinary skill in the art without departing from the principle of the invention, and the improvements and the modifications are also considered as the protection scope of the invention. The components not explicitly described in this embodiment can be implemented by using the prior art.
Claims (10)
1. The composition liposome containing the tea active ingredient with the anti-inflammatory and analgesic effects is characterized by comprising the liposome and a composition containing the tea active ingredient, wherein the composition containing the tea active ingredient is encapsulated in the liposome and comprises the following components in parts by weight: 5-7 parts of epigallocatechin gallate, 1-4 parts of carotene and 1-4 parts of chlorophyll.
2. The tea active ingredient-containing composition liposome according to claim 1, wherein the tea active ingredient-containing composition comprises the following components in parts by mass: 5-6 parts of epigallocatechin gallate, 1-3 parts of carotene and 2-4 parts of chlorophyll.
3. The tea active ingredient-containing composition liposome according to claim 1, wherein the mass fraction ratio of the liposome to the tea active ingredient-containing composition is (85-110): (7-15).
4. A liposome composition containing a tea active ingredient according to any one of claims 1 to 3, wherein the liposome is prepared by the following steps (4-8): 1 and cholesterol, and is prepared by a film dispersion method.
5. A tea active ingredient-containing composition liposome according to any one of claims 1 to 3, wherein the liposome is prepared from the following raw materials in parts by mass: 70-80 parts of lecithin, 10-20 parts of cholesterol and 5-10 parts of tween 80.
6. A method for preparing a liposome of a composition containing tea active ingredients according to any one of claims 1 to 3, characterized in that the composition containing tea active ingredients and liposome raw materials are mixed, an organic solvent is added, shaking and dissolving are carried out, the organic solvent is removed, after the lipid forms a film, a water-soluble buffer solution is added, shaking and shaking are carried out, ultrasound, a filter membrane and centrifugation are carried out, and the composition liposome containing tea active ingredients is obtained and stored in a dark place.
7. The method of claim 6, wherein the condition of the ultrasound is an ultrasound power of 250-300w for 3-5min.
8. The method according to claim 6, wherein the filter membrane is a filter membrane of 0.45 μm followed by a filter membrane of 0.22 μm.
9. Use of a liposome of a composition containing tea active ingredients as claimed in any one of claims 1 to 3 in the manufacture of an anti-inflammatory and/or analgesic medicament.
10. A medicament with anti-inflammatory and/or analgesic properties, characterized in that it comprises a liposome of a composition containing a tea active ingredient according to any one of claims 1 to 3.
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