CN110251421B

CN110251421B - Cyclodextrin inclusion compound as active agent containing protamine and its prepn and application

Info

Publication number: CN110251421B
Application number: CN201910671805.1A
Authority: CN
Inventors: 冯春波; 乔小玲
Original assignee: Shanghai Jahwa United Co Ltd
Current assignee: Shanghai Jahwa United Co Ltd
Priority date: 2019-07-24
Filing date: 2019-07-24
Publication date: 2023-04-14
Anticipated expiration: 2039-07-24
Also published as: CN110251421A

Abstract

The invention discloses an active agent-cyclodextrin inclusion compound, which comprises the following components: an active agent; a cyclodextrin, the molar ratio of the active agent to the cyclodextrin being 1:5 to 1:1; protamine, the weight ratio of protamine to active agent being 2:1 to 1:2. The invention also relates to the preparation of the cyclodextrin inclusion compound which is an active agent containing protamine and the application thereof.

Description

Cyclodextrin inclusion compound as active agent containing protamine and its prepn and application

Technical Field

The invention belongs to the technical field of cosmetics, and particularly relates to a protamine-containing active agent-cyclodextrin inclusion compound, and preparation and application thereof.

Background

Many active agents are chemically unstable and often degrade on storage, or are easily decomposed or isomerized by light. These active agents have poor water solubility (e.g., resveratrol has a solubility of 0.03g/L in water) and low bioavailability, resulting in a major limitation in their use.

Resveratrol (res for short), also called as stilbestrol, is a stilbene compound, is an antitoxin produced by plants under fungal infection, ultraviolet irradiation or pathological conditions, is also used as an antioxidant and an antimutagen, is widely present in plants such as giant knotweed rhizome, peanut, grape, mulberry, sweetberry, chinaroot greenbrier and the like, and is at least found in 72 plants of 31 genera of 21 families at present.

The proven biochemical and pharmacological properties of resveratrol have the effects of resisting atherosclerosis, coronary heart disease, ischemic heart disease and hyperlipidemia, and also have the functions of resisting oxidation and removing free radicals. Research in 1997 of the university of illinois medical school of chicago in the united states shows that resveratrol can effectively inhibit nuclear transcription factor of kappa B (NF-kappa B) protein in tumor cells, so that cancer cells die, malignant tumor diffusion is prevented, and cancer prevention or cancer cell attack is achieved through different biochemical pathways and molecular regulation. Therefore, resveratrol is known as a green anticancer active agent after paclitaxel, and in recent years, resveratrol is widely researched and applied to the industries of medicine and health, health care, beauty treatment and the like. In China, resveratrol is also listed in the catalog of Chinese names of International cosmetic raw material standards (2010 edition) issued by the State food and drug administration.

Resveratrol is an active substance with strong anti-aging effect, is listed as one of 100 most effective anti-aging substances in' anti-aging san Ding (dictionary of san Ding Sheng), also has the effect of ultraviolet radiation prevention, and has very wide application prospect in the field of cosmetics based on the effect.

Resveratrol generally has two configurations (the structure is shown as formula 1), namely cis-Resveratrol (cis-Resveratrol) with the maximum absorption wavelength of 286nm, trans-Resveratrol (trans-Resveratrol) with the maximum absorption wavelength of 306nm, and trans-isomer with relatively strong physiological activity, so that research on Resveratrol mainly takes trans-structure as a main part.

However, resveratrol has poor stability, and decomposition or conversion of resveratrol is caused by temperature, ultraviolet irradiation, continuous heating, acid-base conditions, oxidizing agents, free radicals and the like. Trans-resveratrol has strong physiological activity, but has poor solubility, is only dissolved in organic solvents such as ether, chloroform, methanol, ethanol, acetone, ethyl acetate and the like, is difficult to dissolve in water, has weak capability of penetrating through stratum corneum, and has poor thermal stability and light stability, so that the trans-resveratrol has low effective concentration for really playing a role and low bioavailability.

The literature, namely research on the thermal stability and the photoisomerization reaction of trans-resveratrol in trans-and cis-resveratrol photostability research, shows that the concentration of resveratrol at high temperature is greatly changed, and the effective content loss is high; under the action of ultraviolet light, trans-resveratrol can quickly generate isomerization reaction to generate cis-resveratrol, two benzene rings of a cis-structure cannot be coplanar due to space obstruction, and the conjugation degree is not as good as that of the trans-structure, so that the cis-resveratrol can be in an advantageous conformation when photochemical equilibrium is achieved. In addition to cis-trans isomerization, trans-resveratrol itself also undergoes side reactions, such as removal of hydrogen from phenolic hydroxyl groups, addition of intermediate double bonds to stilbene backbone, etc. Therefore, under the action of ultraviolet light, although the concentration of cis-resveratrol gradually increases in a period of time, the total concentration of the two resveratrol is still reduced because side reactions consume the resveratrol at the same time.

Protamine is an alkaline protein and is obtained from eggs of salmon (Oncorhynchus tschawytscha), trout (Squaliobarbus curriculus), herring (cleupa pallasi), etc. Protamine was discovered in 1870 and studied actively as an antimicrobial agent in 1940-1960. Protamine is now finding more and more widespread use in the food industry. Compared with a chemically synthesized preservative, the protamine has the advantages of high safety, good corrosion resistance, high thermal stability and the like, and has high trophism and functionality, and can reduce blood pressure, assist respiration, promote digestion, inhibit tumor, resist thrombus, strengthen liver function, inhibit blood coagulation and the like. Protamine has also found widespread use in the medical field, either to delay or prevent insulin release, or as a heparin antidote, etc.

Protamine, a cationic active peptide of basic amino acids, mainly consists of arginine and lysine, and the amino acids contain guanidyl (PKa = 12.48), so that protamine carries a large number of positive charges and can interact with phospholipids carrying negative charges on cell membranes or muramic acid carrying negative charges on cell walls, thereby improving the permeability of cell membranes or cell walls and having membrane translocation and membrane transport functions.

The present invention innovatively combines protamine with an active agent-cyclodextrin inclusion compound (e.g., resveratrol as an active agent), and when applied to the cosmetic field, it has been unexpectedly found that the present invention can promote the uptake of the active agent (e.g., resveratrol) into cells, and improve the bioavailability of the active agent (e.g., resveratrol).

Disclosure of Invention

In one aspect, the invention provides an active agent-cyclodextrin inclusion complex comprising:

an active agent;

a cyclodextrin, the molar ratio of the active agent to the cyclodextrin being 1:5 to 1:1;

protamine, the weight ratio of protamine to the active agent is 2:1 to 1:2.

In a preferred embodiment, the cyclodextrin in the active agent-cyclodextrin inclusion complex of the present invention is selected from the group consisting of: hydroxypropyl-beta-cyclodextrin, 2-O-sulfobutyl-beta-cyclodextrin.

In a preferred embodiment, protamine in the active agent-cyclodextrin inclusion complex of the invention is selected from the group consisting of: salmon protamine, trout protamine, herring protamine.

In a preferred embodiment, the active agent-cyclodextrin inclusion compound of the invention has a molar ratio of active agent to cyclodextrin of 1.

In another aspect, the invention relates to the use of protamine in an active agent-cyclodextrin inclusion compound. In a preferred embodiment, the molar ratio of active agent to cyclodextrin is 1:5 to 1:1. In a more preferred embodiment, the weight ratio of protamine to the active agent is 2:1 to 1:2.

In another aspect, the invention also relates to a method for preparing an active agent-cyclodextrin inclusion complex, the method comprising the steps of:

1a) Dissolving an active agent in an organic solvent to form a solution of the active agent;

1b) Dissolving cyclodextrin in an aqueous solvent to form a solution of cyclodextrin;

1c) Adding the solution of the active agent prepared in the step 1 a) into the solution of the cyclodextrin prepared in the step 1 b) to obtain a mixed solution;

1d) Dissolving protamine in an aqueous solvent to prepare a solution of protamine;

1e) Adding the protamine solution prepared in step 1 d) into the mixed solution of step 1 c);

1f) Freeze drying or spray drying to obtain cyclodextrin clathrate of protamine-containing active agent.

2a) Dissolving an active agent in an organic solvent to form a solution of the active agent;

2b) Dissolving cyclodextrin in an aqueous solvent to form a solution of cyclodextrin;

2c) Adding the solution of the active agent prepared in the step 2 a) into the solution of the cyclodextrin prepared in the step 2 b) to obtain a mixed solution;

2d) Freeze drying or spray drying to obtain active agent-cyclodextrin clathrate;

2e) Mixing protamine with the active agent-cyclodextrin inclusion compound prepared in step 2 d) to obtain an active agent-cyclodextrin inclusion compound containing protamine.

In a preferred embodiment, the freeze-drying is arranged to: the sublimation drying is set to be-80 to-40 ℃, the vacuum degree is 1 to 30Pa, the time is 3 to 21 hours, the desorption drying is set to be-10 to +20 ℃, the vacuum degree is 1 to 30Pa, the time is 1 to 10 hours, and the water content is controlled to be 0 to 6 percent.

In a preferred embodiment, the spray drying is arranged to: the air inlet temperature is 40-80 ℃, the atomization pressure is 0.2-0.8 MPa, the feeding speed is 100-2000 ml/h, and the water content is controlled between 0-8%.

In a preferred embodiment, the organic solvent used in the preparation of the active agent-cyclodextrin inclusion compound of the present invention is ethanol.

In yet another aspect, the invention also relates to the use of the active agent-cyclodextrin inclusion compound in cosmetics.

Brief description of the drawings

Fig. 1A-1B show the uptake of resveratrol-conjugated protamine composition of the invention (example 19) in fibroblasts in comparison to the uptake of a common resveratrol composition (example 1) (n = 3).

Fig. 2A-2B show the uptake of a resveratrol-conjugated protamine composition of the invention (example 19) versus a normal resveratrol composition (example 1) in langerhans cells (n = 3).

Fig. 3A-3B show the uptake in melanocytes of resveratrol-conjugated protamine composition of the invention (example 19) compared to the normal resveratrol composition (example 1) (n = 3).

Detailed Description

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, the preferred methods and materials are described herein. For the purposes of the present invention, the following terms are defined below.

The term "about" as used herein refers to an amount, level, value, dimension, size, or amount that differs by up to 30%, 20%, or 10% as compared to the amount, level, value, dimension, size, or amount of a reference. As used herein, unless otherwise indicated, the percentages are by weight.

Throughout the specification and claims, unless the context requires otherwise, the word "comprise", and variations such as "comprises" and "comprising", will be understood to imply the inclusion of a stated integer or step or group of integers or steps but not the exclusion of any other integer or step or group of integers or steps.

The present invention is based on the following unexpected findings: when protamine is used in combination with the active agent-cyclodextrin inclusion compound, a stable active agent-cyclodextrin inclusion compound can be prepared. Moreover, the active agent-cyclodextrin inclusion compound containing protamine can significantly improve the uptake of the active agent, thereby improving the bioavailability of the active agent.

The inventors of the present application have also found that the use of a cyclodextrin inclusion compound containing protamine can achieve a stabilizing effect, promote the uptake of an active agent, and improve the bioavailability of an active agent that is chemically unstable and is easily decomposed by light.

Thus, the inclusion complex of the present invention is particularly useful for chemically unstable active agents, for active agents that are susceptible to degradation during storage, or for active agents that are susceptible to decomposition or isomerization by light. In some embodiments, the active agents employed in the clathrates of the invention are poorly water soluble (e.g., resveratrol has a solubility in water of 0.03 g/L), or have low bioavailability.

Active agent

In some embodiments of the invention, the active agent employed is resveratrol.

Resveratrol (3,4 ',5-trihydroxystilbene (3,4', 5-trihydroxystilbene), also known as resveratrol), is a natural plant extract, is present in 72 plants of at least 21 families and 31 genera, such as red grape, peanut, mulberry, giant knotweed, asteraceae plant (knotted), raspberry (raspberry), blueberry, and is a polyphenol compound. In recent years, more and more scientific research has shown the multifunctional benefits of resveratrol. Resveratrol is found to be an extremely potent antioxidant, regulator of genetic expression via signaling, inhibitor of inflammatory mediators, and anti-aging substance, and is capable of reducing melanin synthesis. Resveratrol can therefore be used in cosmetic compositions as a unique active ingredient in personal care products.

Although resveratrol has the above-mentioned biological activity and excellent skin whitening effect, it is difficult to formulate it into a cosmetic composition. The compound is insoluble in water (the solubility of the compound in water is about 0.03g/Kg (namely 0.003 percent) at 25 ℃), is easy to react with other chemical substances, is unstable to light, and can only be stabilized for several days even under the condition of keeping away from light in a high-purity resveratrol ethanol solution in a Microherb stability test. Therefore, when it is used in cosmetics, it is difficult to stably maintain it in the shelf life.

In addition, resveratrol is also unstable in cosmetic compositions in the form of O/W or W/O compositions, it causes phase separation in the composition and changes the apparent color from white to yellowish brown. Thus, it has not been possible to make resveratrol be present in cosmetics at high levels so far. Furthermore, resveratrol has a tendency to precipitate (crystallize) in aqueous cosmetic compositions.

The present invention is based on the following unexpected findings: the application of the protamine can improve the ingestion of the resveratrol by skin cells and the bioavailability, and experiments in fibroblasts, langerhans cells and melanocytes show that the protamine composition has higher absorption intensity compared with the common composition, and the data have statistical significance.

In certain embodiments of the invention, the weight ratio of resveratrol to protamine is 2:1 to 1:2. In a preferred embodiment, the weight ratio of resveratrol to protamine is 2:1.

In certain embodiments of the invention, oxidized resveratrol, which is the active agent, is employed.

In certain embodiments of the invention, the weight ratio of oxyresveratrol to protamine is 2:1 to 1:2. In a preferred embodiment, the weight ratio of oxyresveratrol to protamine is 2:1.

In certain embodiments of the invention, pterostilbene is employed as the active agent.

Pterostilbene belongs to antifungal active ingredients in a dragon's blood product, and has various biological activities of oxidation resistance, tumor resistance, blood fat reduction, bacteriostasis and the like.

In certain embodiments of the invention, the weight ratio of pterostilbene to protamine is 2:1 to 1:2. In a preferred embodiment, the weight ratio of pterostilbene to protamine is 2:1.

Cyclodextrin

There are many ways to improve the dissolution of the active agent, thereby achieving a rapid dissolution of the active agent and possibly making the active agent more readily absorbed and available after administration by the oral route. These methods include: adding cosolvent, surfactant, cyclodextrin, and salt to obtain amorphous active agent, and making into cocrystal and nanoparticle.

Many methods and processes suffer from one or other of these problems. The research shows that: inclusion by cyclodextrin is a fairly effective option. Based on the structure of the active agent (e.g., resveratrol), the active agent can form an inclusion complex with the cyclodextrin. Literature reports indicate that inclusion complex formation can be achieved by a variety of methods, including solution saturation methods, spray drying methods, freeze drying methods, and the like.

Cyclodextrins are non-reducing cyclic glucose oligosaccharides produced from starch. There are three common cyclodextrins with 6, 7, or 8 glucose units (alpha-, beta-, and gamma-cyclodextrins, respectively) connected by alpha-1,4 glycosidic linkages. Cyclodextrins act as molecular reservoirs by trapping guest molecules (guest molecules) in their internal cavities, thereby forming inclusion complexes. Alpha-cyclodextrin has a smaller cavity and beta-and gamma-cyclodextrin have a larger cavity.

Cyclodextrins suitable for use in the present invention include alpha-, beta-and gamma-cyclodextrins, but beta-and gamma-cyclodextrins are preferred due to the larger internal cavity. Cyclodextrins have been chemically modified, particularly β -cyclodextrins, to improve the solubility of the parent cyclodextrin. Hydroxyethyl β -cyclodextrin, hydroxypropyl β -cyclodextrin (e.g., 2-hydroxypropyl- β -cyclodextrin), methylated β -cyclodextrin, glucosyl β -cyclodextrin, and sulfobutylether β -cyclodextrin are examples of cyclodextrins that have been chemically modified to improve their solubility. In some aspects of the invention, a beta-cyclodextrin will be used that includes a chemically modified beta-cyclodextrin. In some aspects, the chemically modified β -cyclodextrin will be hydroxypropyl β -cyclodextrin or sulfobutylether β -cyclodextrin. In some aspects, the cyclodextrin will be a gamma cyclodextrin, including a chemically modified gamma cyclodextrin. In some aspects, the cyclodextrin is a hydroxypropyl cyclodextrin (e.g., HP4.3- β -cyclodextrin, HP5.5- β -cyclodextrin, HP7.6- β -cyclodextrin, and HP4.5- γ -cyclodextrin). In other aspects, the cyclodextrin will be a sulfobutylether beta-cyclodextrin (e.g., SBE 6.6-beta-cyclodextrin, SBE 6.7-beta-cyclodextrin, SBE 6.8-beta-cyclodextrin, SBE 4.1-beta-cyclodextrin, and SBE 4.6et3.5-beta-cyclodextrin). In other aspects, the cyclodextrin will be a sulfobutylether gamma-cyclodextrin (e.g., SBE 4.3-gamma-cyclodextrin, SBE 4.6-gamma-cyclodextrin, SBE 5.2-gamma-cyclodextrin, and SBE5.6et 6.3-gamma-cyclodextrin). As used herein, a "chemically modified beta cyclodextrin" is a beta cyclodextrin that has been chemically modified to have at least improved solubility compared to its parent cyclodextrin (i.e., unmodified cyclodextrin).

Once the active agent (e.g., resveratrol) is fully included with the β -cyclodextrin, its dissolution is significantly improved and bioavailability is improved compared to the active agent itself.

In a preferred embodiment of the invention, the cyclodextrin employed is selected from: hydroxypropyl-beta-cyclodextrin, 2-O-sulfobutyl-beta-cyclodextrin, or a combination thereof.

In certain embodiments of the invention, the molar ratio of active agent to cyclodextrin is 1:5 to 1:1. In a preferred embodiment, the molar ratio of active agent to cyclodextrin is 1.3 to 1.5.

Absorption enhancer

The invention adopts protamine to improve the absorption of the active agent-cyclodextrin inclusion compound. Surprisingly, it was found that protamine can significantly increase the uptake of active agents in the active agent-cyclodextrin inclusion compound, which is statistically significant. The application of the protamine can improve the uptake of active agents (such as resveratrol) by skin cells and improve the bioavailability, and experiments in fibroblasts, langerhans cells and melanocytes show that the protamine composition has higher absorption intensity compared with the common composition, and the data have statistical significance.

In certain embodiments of the invention, protamine is selected from the group consisting of: salmon protamine, trout protamine, herring protamine, or combinations thereof.

Preparation method

The active agent-cyclodextrin inclusion compound of the present invention can be prepared by the following procedure. For example, 1 a) dissolving an active agent in an organic solvent to form a solution of the active agent; 1b) Dissolving cyclodextrin in an aqueous solvent to form a solution of cyclodextrin; 1c) Adding the solution of the active agent prepared in the step 1 a) into the solution of the cyclodextrin prepared in the step 1 b) to obtain a mixed solution; 1d) Dissolving protamine in an aqueous solvent to prepare a solution of protamine; 1e) Adding the protamine solution prepared in step 1 d) into the mixed solution of step 1 c); 1f) Freeze drying or spray drying to obtain active agent-cyclodextrin clathrate.

Alternatively, the active agent-cyclodextrin inclusion compound of the present invention can be prepared by the following procedure. For example, 2 a) dissolving an active agent in an organic solvent to form a solution of the active agent; 2b) Dissolving cyclodextrin in an aqueous solvent to form a solution of cyclodextrin; 2c) Adding the solution of the active agent prepared in the step 2 a) into the solution of the cyclodextrin prepared in the step 2 b) to obtain a mixed solution; 2d) Freeze drying or spray drying to obtain active agent-cyclodextrin clathrate; 2e) Mixing protamine with the active agent-cyclodextrin inclusion compound prepared in step 2 d) to obtain the desired active agent-cyclodextrin inclusion compound.

In certain embodiments of the invention, the active agent is resveratrol. For example, resveratrol can be dissolved in an organic solvent such as methanol, ethanol, or a polyhydric alcohol. In certain embodiments of the invention, the organic solvent may be ethanol. For example, the organic solvent-soluble active agent is added in an amount of 10 to 20 times the weight of resveratrol. In certain preferred embodiments, ultrasonic solubilization may be employed.

In certain embodiments of the present invention, the aqueous solvent is a cosmetically acceptable aqueous solvent. For example, the aqueous solvent may be purified water or deionized water.

In certain embodiments of the present invention, the mixture obtained in step 1 c) is filtered and the filtrate is collected. In certain embodiments of the invention, the collected filtrate is distilled under reduced pressure to remove the organic solvent. For example, in a preferred embodiment, the organic solvent is distilled off at 45 ℃ under reduced pressure.

In certain embodiments of the invention, the resulting active agent-cyclodextrin inclusion compound is frozen in a freezer at-80 ℃. Then, freeze-drying was performed with the following settings: setting sublimation drying at-80-40 deg.c and vacuum degree of 1-30 Pa for 3-21 hr, setting desorption drying at-10-20 deg.c and vacuum degree of 1-30 Pa for 1-10 hr, and controlling water content in 0-6%.

In certain embodiments of the invention, the resulting active agent-cyclodextrin inclusion compound is frozen in a freezer at-80 ℃. Then, spray drying was performed with the following settings: the air inlet temperature is 40-80 ℃, the atomization pressure is 0.2-0.8 MPa, the feeding speed is 100-2000 ml/h, and the water content is controlled between 0-8%.

Application method

The active agent-cyclodextrin inclusion compounds of the present invention can be topically applied to mammalian skin. In one embodiment, the skin is subject to acne problems. In another embodiment, the skin requires an exfoliating treatment. The active agent-cyclodextrin inclusion complex can be applied to skin in need of treatment according to a suitable treatment regimen, e.g., from up to 2 times per day to as little as 1 time per week (e.g., once per day, once every two days, once per week), etc.

In certain embodiments, the active agent-cyclodextrin inclusion compounds of the present invention may be used to treat other desirable conditions associated with the skin. For example, the active agent-cyclodextrin inclusion compounds of the present invention are useful for treating post-inflammatory hyperpigmentation, for reducing pore size, for reducing sebum production, and for reducing scarring.

The technical aspects of the present invention will be described in detail below with reference to preferred embodiments, but the scope of the present invention is not limited to these embodiments, and the technical aspects of the present invention are intended to be described and not limited. Test methods in which specific conditions are not specified in the following examples are generally carried out under conventional conditions or under conditions recommended by the manufacturer. All percentages and parts are by weight unless otherwise indicated.

The experimental materials used in the examples of the present invention are as follows:

resveratrol: supplied by Dezhi (Shanghai) Co., ltd;

oxidizing resveratrol: purchased from sabinda corporation, usa;

pterostilbene: purchased from sabinda corporation, usa;

hydroxypropyl- β -cyclodextrin: (CAS No 128446-35-5) available from Zhiyuan Biotech, inc., shandong Binshony;

salmon protamine, trout protamine, herring protamine, and coumarin are all purchased from SIGMA-ALORICH company.

Flow cytometer, model FACSCANTO 10C, BD Biosciences

Vacuum freeze dryer model FD-2, beijing Bo Yi kang laboratory instruments Ltd

Spray dryer, model QFN-9000Y, shanghai Qiao Fengshi industries Ltd

In vitro fibroblast, melanocyte, and Langerhans cell culture method adopts enzyme digestion method, i.e., using MEM complete culture solution, at 37 deg.C and 5% CO ₂ Culturing in the incubator.

Example 1

1g of resveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a resveratrol solution. 9g of hydroxypropyl-beta-cyclodextrin is taken and dissolved in 20ml of purified water. Slowly dripping the resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1-30 Pa for 3-21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1-30 Pa for 1-10 h, and controlling water content between 0 and 6 percent to obtain white loose porous spongy amorphous powder. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 2

1g of resveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a resveratrol solution. Taking 11g of hydroxypropyl-beta-cyclodextrin, and dissolving in 20ml of purified water. Slowly dripping the resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1-30 Pa for 3-21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1-30 Pa for 1-10 h, and controlling water content between 0 and 6 percent to obtain white loose porous spongy amorphous powder. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 3

1g of resveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a resveratrol solution. 6.5g of hydroxypropyl-beta-cyclodextrin is taken and dissolved in 20ml of purified water. Slowly dripping the resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1-30 Pa for 3-21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1-30 Pa for 1-10 h, and controlling water content between 0 and 6 percent to obtain white loose porous spongy amorphous powder. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 4

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 5.0g of beta-cyclodextrin is taken and dissolved in 15ml of purified water. Slowly dripping the resveratrol solution into beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1-30 Pa for 3-21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1-30 Pa for 1-10 h, and controlling water content between 0 and 6 percent to obtain white loose porous spongy amorphous powder. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 5

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 6.5g of beta-cyclodextrin is taken and dissolved in 15ml of purified water. Slowly dripping the resveratrol solution into beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1-30 Pa for 3-21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1-30 Pa for 1-10 h, and controlling water content between 0 and 6 percent to obtain white loose porous spongy amorphous powder. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 6

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 8.0g of beta-cyclodextrin is taken and dissolved in 15ml of purified water. Slowly dripping the resveratrol solution into beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1-30 Pa for 3-21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1-30 Pa for 1-10 h, and controlling water content between 0 and 6 percent to obtain white loose porous spongy amorphous powder. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 7

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 5.0g of 2-O-sulfobutyl-beta-cyclodextrin is dissolved in 30ml of purified water. Slowly dripping the resveratrol solution into the 2-O-sulfobutyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain a clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1-30 Pa for 3-21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1-30 Pa for 1-10 h, and controlling water content between 0 and 6 percent to obtain white loose porous spongy amorphous powder. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 8

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 6.5g of 2-O-sulfobutyl-beta-cyclodextrin is dissolved in 30ml of purified water. Slowly dripping the resveratrol solution into the 2-O-sulfobutyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain a clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1-30 Pa for 3-21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1-30 Pa for 1-10 h, and controlling water content between 0 and 6 percent to obtain white loose porous spongy amorphous powder. Grinding, and sieving with a 300-mesh sieve to obtain a white composition for later use.

Example 9

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 7.5g of 2-O-sulfobutyl-beta-cyclodextrin is dissolved in 30ml of purified water. Slowly dripping the resveratrol solution into the 2-O-sulfobutyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain a clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1-30 Pa for 3-21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1-30 Pa for 1-10 h, and controlling water content between 0 and 6 percent to obtain white loose porous spongy amorphous powder. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 10

1g of resveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a resveratrol solution. 9g of hydroxypropyl-beta-cyclodextrin is taken and dissolved in 20ml of purified water. Slowly dripping the resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, and then spray-drying at the air inlet temperature of 40-80 ℃, the atomization pressure of 0.2-0.8 MPa, the feeding speed of 100-2000 ml/h and the water content of 0-8 percent to obtain the microsphere composition. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 11

1g of resveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a resveratrol solution. Taking 11g of hydroxypropyl-beta-cyclodextrin, and dissolving in 20ml of purified water. Slowly dripping the resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, and then spray-drying at the air inlet temperature of 40-80 ℃, the atomization pressure of 0.2-0.8 MPa, the feeding speed of 100-2000 ml/h and the water content of 0-8 percent to obtain the microsphere composition. Grinding, and sieving with a 300-mesh sieve to obtain a white composition for later use.

Example 12

1g of resveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a resveratrol solution. 6.5g of hydroxypropyl-beta-cyclodextrin is taken and dissolved in 20ml of purified water. Slowly dripping the resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove the ethanol, continuously stirring for 0.5h, and then carrying out spray drying at the air inlet temperature of 40-80 ℃, the atomization pressure of 0.2-0.8 MPa, the feeding speed of 100-2000 ml/h and the water content of 0-8% to obtain the microspherical composition. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 13

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 6.5g of beta-cyclodextrin is taken and dissolved in 15ml of purified water. Slowly dripping the resveratrol solution into beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, and then spray-drying at the air inlet temperature of 40-80 ℃, the atomization pressure of 0.2-0.8 MPa, the feeding speed of 100-2000 ml/h and the water content of 0-8 percent to obtain the microsphere composition. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 14

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 5.0g of beta-cyclodextrin is taken and dissolved in 15ml of purified water. Slowly dripping the resveratrol solution into beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, and then spray-drying at the air inlet temperature of 40-80 ℃, the atomization pressure of 0.2-0.8 MPa, the feeding speed of 100-2000 ml/h and the water content of 0-8 percent to obtain the microsphere composition. Grinding, and sieving with a 300-mesh sieve to obtain a white composition for later use.

Example 15

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 8.0g of beta-cyclodextrin is taken and dissolved in 15ml of purified water. Slowly dripping the resveratrol solution into beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove the ethanol, continuously stirring for 0.5h, and then carrying out spray drying at the air inlet temperature of 40-80 ℃, the atomization pressure of 0.2-0.8 MPa, the feeding speed of 100-2000 ml/h and the water content of 0-8% to obtain the microspherical composition. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 16

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 5.0g of 2-O-sulfobutyl-beta-cyclodextrin is dissolved in 30ml of purified water. Slowly dripping the resveratrol solution into the 2-O-sulfobutyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain a clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, and then spray-drying at the air inlet temperature of 40-80 ℃, the atomization pressure of 0.2-0.8 MPa, the feeding speed of 100-2000 ml/h and the water content of 0-8 percent to obtain the microsphere composition. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 17

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 6.5g of 2-O-sulfobutyl-beta-cyclodextrin is dissolved in 30ml of purified water. Slowly dripping the resveratrol solution into the 2-O-sulfobutyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain a clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, and then spray-drying at the air inlet temperature of 40-80 ℃, the atomization pressure of 0.2-0.8 MPa, the feeding speed of 100-2000 ml/h and the water content of 0-8 percent to obtain the microsphere composition. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 18

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 7.5g of 2-O-sulfobutyl-beta-cyclodextrin is taken and dissolved in 30ml of purified water. Slowly dripping the resveratrol solution into the 2-O-sulfobutyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain a clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, and then spray-drying at the air inlet temperature of 40-80 ℃, the atomization pressure of 0.2-0.8 MPa, the feeding speed of 100-2000 ml/h and the water content of 0-8 percent to obtain the microsphere composition. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 19

1g of resveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a resveratrol solution. 9g of hydroxypropyl-beta-cyclodextrin is taken and dissolved in 20ml of purified water. Slowly dripping the resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Ethanol was removed by distillation under reduced pressure at 45 ℃. 0.5g of salmon protamine was weighed and dissolved in 5ml of purified water. Slowly dripping salmon protamine into cyclodextrin under continuous stirring, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1 to 30Pa for 3 to 21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1 to 30Pa for 1 to 10 h, and controlling water content to be 0 to 6 percent to obtain white loose porous spongy amorphous powder. Grinding, and sieving with a 300-mesh sieve to obtain a white composition for later use.

Example 20

1g of resveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a resveratrol solution. Taking 11g of hydroxypropyl-beta-cyclodextrin, and dissolving in 20ml of purified water. Slowly dripping the resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. The ethanol was removed by distillation under reduced pressure at 45 ℃. 0.5g of salmon protamine was weighed and dissolved in 5ml of purified water. Slowly dripping salmon protamine into cyclodextrin under continuous stirring, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1 to 30Pa, and time of 3 to 21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1 to 30Pa, and time of 1 to 10 h, and controlling water content between 0 and 6 percent to obtain white loose porous spongy amorphous powder. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 21

1g of resveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a resveratrol solution. Taking 6.5g of hydroxypropyl-beta-cyclodextrin, and dissolving in 20ml of purified water. Slowly dripping the resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. The ethanol was removed by distillation under reduced pressure at 45 ℃. 0.5g of salmon protamine was weighed and dissolved in 5ml of purified water. Slowly dripping salmon protamine into cyclodextrin under continuous stirring, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1 to 30Pa, and time of 3 to 21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1 to 30Pa, and time of 1 to 10 h, and controlling water content between 0 and 6 percent to obtain white loose porous spongy amorphous powder. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 22

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 5.0g of beta-cyclodextrin is taken and dissolved in 15ml of purified water. Slowly dripping the resveratrol solution into beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Ethanol was removed by distillation under reduced pressure at 45 ℃. 0.5g of salmon protamine was weighed and dissolved in 5ml of purified water. Slowly dripping salmon protamine into cyclodextrin under continuous stirring, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1 to 30Pa for 3 to 21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1 to 30Pa for 1 to 10 h, and controlling water content to be 0 to 6 percent to obtain white loose porous spongy amorphous powder. Grinding, and sieving with a 300-mesh sieve to obtain a white composition for later use.

Example 23

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 6.5g of beta-cyclodextrin is taken and dissolved in 15ml of purified water. Slowly dripping the resveratrol solution into the beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. The ethanol was removed by distillation under reduced pressure at 45 ℃. 0.5g of salmon protamine was weighed and dissolved in 5ml of purified water. Slowly dripping salmon protamine into cyclodextrin under continuous stirring, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1 to 30Pa for 3 to 21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1 to 30Pa for 1 to 10 h, and controlling water content to be 0 to 6 percent to obtain white loose porous spongy amorphous powder. Grinding, and sieving with a 300-mesh sieve to obtain a white composition for later use.

Example 24

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 8.0g of beta-cyclodextrin is taken and dissolved in 15ml of purified water. Slowly dripping the resveratrol solution into the beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. The ethanol was removed by distillation under reduced pressure at 45 ℃. 0.5g of salmon protamine was weighed and dissolved in 5ml of purified water. Slowly dripping salmon protamine into cyclodextrin under continuous stirring, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1 to 30Pa for 3 to 21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1 to 30Pa for 1 to 10 h, and controlling water content to be 0 to 6 percent to obtain white loose porous spongy amorphous powder. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 25

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 5.0g of 2-O-sulfobutyl-beta-cyclodextrin is dissolved in 30ml of purified water. Slowly dripping the resveratrol solution into the 2-O-sulfobutyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain a clear, colorless and transparent solution. The ethanol was removed by distillation under reduced pressure at 45 ℃. 0.5g of salmon protamine was weighed and dissolved in 5ml of purified water. Slowly dripping salmon protamine into cyclodextrin under continuous stirring, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1 to 30Pa for 3 to 21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1 to 30Pa for 1 to 10 h, and controlling water content to be 0 to 6 percent to obtain white loose porous spongy amorphous powder. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 26

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 6.5g of 2-O-sulfobutyl-beta-cyclodextrin is taken and dissolved in 30ml of purified water. Slowly dripping the resveratrol solution into the 2-O-sulfobutyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain a clear, colorless and transparent solution. The ethanol was removed by distillation under reduced pressure at 45 ℃. 0.5g of salmon protamine was weighed and dissolved in 5ml of purified water. Slowly dripping salmon protamine into cyclodextrin under continuous stirring, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1 to 30Pa, and time of 3 to 21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1 to 30Pa, and time of 1 to 10 h, and controlling water content between 0 and 6 percent to obtain white loose porous spongy amorphous powder. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 27

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 7.5g of 2-O-sulfobutyl-beta-cyclodextrin is dissolved in 30ml of purified water. Slowly dripping the resveratrol solution into the 2-O-sulfobutyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain a clear, colorless and transparent solution. The ethanol was removed by distillation under reduced pressure at 45 ℃. 0.5g of salmon protamine was weighed and dissolved in 5ml of purified water. Slowly dripping salmon protamine into cyclodextrin under continuous stirring, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1 to 30Pa, and time of 3 to 21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1 to 30Pa, and time of 1 to 10 h, and controlling water content between 0 and 6 percent to obtain white loose porous spongy amorphous powder. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 28

1g of resveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a resveratrol solution. 9g of hydroxypropyl-beta-cyclodextrin is taken and dissolved in 20ml of purified water. Slowly dripping the resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Ethanol was removed by distillation under reduced pressure at 45 ℃. 0.5g of salmon protamine was weighed and dissolved in 5ml of purified water. Slowly dripping the salmon protamine into the cyclodextrin under the continuous stirring, continuously stirring for 0.5h, and then carrying out spray drying at the air inlet temperature of 40-80 ℃, the atomization pressure of 0.2-0.8 MPa, the feeding speed of 100-2000 ml/h and the water content of 0-8 percent to obtain the microspherical composition. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 29

1g of resveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a resveratrol solution. Taking 11g of hydroxypropyl-beta-cyclodextrin, and dissolving in 20ml of purified water. Slowly dripping the resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. The ethanol was removed by distillation under reduced pressure at 45 ℃. 0.5g of salmon protamine was weighed and dissolved in 5ml of purified water. Slowly dripping the salmon protamine into the cyclodextrin under the continuous stirring, continuously stirring for 0.5h, then carrying out spray drying, wherein the air inlet temperature is 40-80 ℃, the atomization pressure is 0.2-0.8 MPa, the feeding speed is 100-2000 ml/h, and the water content is controlled between 0-8 percent, thus obtaining the microspherical composition. Grinding, and sieving with a 300-mesh sieve to obtain a white composition for later use.

Example 30

1g of resveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a resveratrol solution. 6.5g of hydroxypropyl-beta-cyclodextrin is taken and dissolved in 20ml of purified water. Slowly dripping the resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Ethanol was removed by distillation under reduced pressure at 45 ℃. 0.5g of salmon protamine was weighed and dissolved in 5ml of purified water. Slowly dripping the salmon protamine into the cyclodextrin under the continuous stirring, continuously stirring for 0.5h, then carrying out spray drying, wherein the air inlet temperature is 40-80 ℃, the atomization pressure is 0.2-0.8 MPa, the feeding speed is 100-2000 ml/h, and the water content is controlled between 0-8 percent, thus obtaining the microspherical composition. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 31

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 6.5g of beta-cyclodextrin is taken and dissolved in 15ml of purified water. Slowly dripping the resveratrol solution into beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Ethanol was removed by distillation under reduced pressure at 45 ℃. 0.5g of salmon protamine was weighed and dissolved in 5ml of purified water. Slowly dripping the salmon protamine into the cyclodextrin under the continuous stirring, continuously stirring for 0.5h, then carrying out spray drying, wherein the air inlet temperature is 40-80 ℃, the atomization pressure is 0.2-0.8 MPa, the feeding speed is 100-2000 ml/h, and the water content is controlled between 0-8 percent, thus obtaining the microspherical composition. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 32

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 5.0g of beta-cyclodextrin is taken and dissolved in 15ml of purified water. Slowly dripping the resveratrol solution into the beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. The ethanol was removed by distillation under reduced pressure at 45 ℃. 0.5g of salmon protamine was weighed and dissolved in 5ml of purified water. Slowly dripping the salmon protamine into the cyclodextrin under the continuous stirring, continuously stirring for 0.5h, then carrying out spray drying, wherein the air inlet temperature is 40-80 ℃, the atomization pressure is 0.2-0.8 MPa, the feeding speed is 100-2000 ml/h, and the water content is controlled between 0-8 percent, thus obtaining the microspherical composition. Grinding, and sieving with a 300-mesh sieve to obtain a white composition for later use.

Example 33

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 8.0g of beta-cyclodextrin is taken and dissolved in 15ml of purified water. Slowly dripping the resveratrol solution into the beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. The ethanol was removed by distillation under reduced pressure at 45 ℃. 0.5g of salmon protamine was weighed and dissolved in 5ml of purified water. Slowly dripping the salmon protamine into the cyclodextrin under the continuous stirring, continuously stirring for 0.5h, then carrying out spray drying, wherein the air inlet temperature is 40-80 ℃, the atomization pressure is 0.2-0.8 MPa, the feeding speed is 100-2000 ml/h, and the water content is controlled between 0-8 percent, thus obtaining the microspherical composition. Grinding, and sieving with a 300-mesh sieve to obtain a white composition for later use.

Example 34

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 5.0g of 2-O-sulfobutyl-beta-cyclodextrin is dissolved in 30ml of purified water. Slowly dripping the resveratrol solution into the 2-O-sulfobutyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain a clear, colorless and transparent solution. Ethanol was removed by distillation under reduced pressure at 45 ℃. 0.5g of salmon protamine was weighed and dissolved in 5ml of purified water. Slowly dripping the salmon protamine into the cyclodextrin under the continuous stirring, continuously stirring for 0.5h, and then carrying out spray drying at the air inlet temperature of 40-80 ℃, the atomization pressure of 0.2-0.8 MPa, the feeding speed of 100-2000 ml/h and the water content of 0-8 percent to obtain the microspherical composition. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 35

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 6.5g of 2-O-sulfobutyl-beta-cyclodextrin is dissolved in 30ml of purified water. Slowly dripping the resveratrol solution into the 2-O-sulfobutyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain a clear, colorless and transparent solution. The ethanol was removed by distillation under reduced pressure at 45 ℃. 0.5g of salmon protamine was weighed and dissolved in 5ml of purified water. Slowly dripping the salmon protamine into the cyclodextrin under the continuous stirring, continuously stirring for 0.5h, and then carrying out spray drying at the air inlet temperature of 40-80 ℃, the atomization pressure of 0.2-0.8 MPa, the feeding speed of 100-2000 ml/h and the water content of 0-8 percent to obtain the microspherical composition. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 36

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 7.5g of 2-O-sulfobutyl-beta-cyclodextrin is dissolved in 30ml of purified water. Slowly dripping the resveratrol solution into the 2-O-sulfobutyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain a clear, colorless and transparent solution. The ethanol was removed by distillation under reduced pressure at 45 ℃. 0.5g of salmon protamine was weighed and dissolved in 5ml of purified water. Slowly dripping the salmon protamine into the cyclodextrin under the continuous stirring, continuously stirring for 0.5h, then carrying out spray drying, wherein the air inlet temperature is 40-80 ℃, the atomization pressure is 0.2-0.8 MPa, the feeding speed is 100-2000 ml/h, and the water content is controlled between 0-8 percent, thus obtaining the microspherical composition. Grinding, and sieving with a 300-mesh sieve to obtain a white composition for later use.

Example 37

1g of resveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a resveratrol solution. 9g of hydroxypropyl-beta-cyclodextrin is taken and dissolved in 20ml of purified water. Slowly dripping the resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1-30 Pa for 3-21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1-30 Pa for 1-10 h, and controlling water content between 0 and 6 percent to obtain white loose porous spongy amorphous powder. 0.5g of salmon protamine is weighed, mixed with the white amorphous powder uniformly, ground together and sieved by a 300-mesh sieve to obtain a white composition for later use.

Example 38

1g of resveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a resveratrol solution. Taking 11g of hydroxypropyl-beta-cyclodextrin, and dissolving in 20ml of purified water. Slowly dripping the resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1-30 Pa for 3-21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1-30 Pa for 1-10 h, and controlling water content between 0 and 6 percent to obtain white loose porous spongy amorphous powder. 0.5g of salmon protamine is weighed, mixed with white amorphous powder evenly, ground together and sieved by a 300-mesh sieve to obtain a white composition for later use.

Example 39

1g of resveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a resveratrol solution. 6.5g of hydroxypropyl-beta-cyclodextrin is taken and dissolved in 20ml of purified water. Slowly dripping the resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1-30 Pa for 3-21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1-30 Pa for 1-10 h, and controlling water content between 0 and 6 percent to obtain white loose porous spongy amorphous powder. 0.5g of salmon protamine is weighed, mixed with the white amorphous powder uniformly, ground together and sieved by a 300-mesh sieve to obtain a white composition for later use.

Example 40

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 5.0g of beta-cyclodextrin is taken and dissolved in 15ml of purified water. Slowly dripping the resveratrol solution into beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1-30 Pa for 3-21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1-30 Pa for 1-10 h, and controlling water content between 0 and 6 percent to obtain white loose porous spongy amorphous powder. 0.5g of salmon protamine is weighed, mixed with white amorphous powder evenly, ground together and sieved by a 300-mesh sieve to obtain a white composition for later use.

EXAMPLE 41

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 6.5g of beta-cyclodextrin is taken and dissolved in 15ml of purified water. Slowly dripping the resveratrol solution into beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1-30 Pa for 3-21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1-30 Pa for 1-10 h, and controlling water content between 0 and 6 percent to obtain white loose porous spongy amorphous powder. 0.5g of salmon protamine is weighed, mixed with white amorphous powder evenly, ground together and sieved by a 300-mesh sieve to obtain a white composition for later use.

Example 42

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 8.0g of beta-cyclodextrin is taken and dissolved in 15ml of purified water. Slowly dripping the resveratrol solution into beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1-30 Pa for 3-21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1-30 Pa for 1-10 h, and controlling water content between 0 and 6 percent to obtain white loose porous spongy amorphous powder. 0.5g of salmon protamine is weighed, mixed with white amorphous powder evenly, ground together and sieved by a 300-mesh sieve to obtain a white composition for later use.

Example 43

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 5.0g of 2-O-sulfobutyl-beta-cyclodextrin is taken and dissolved in 30ml of purified water. Slowly dripping the resveratrol solution into the 2-O-sulfobutyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain a clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1-30 Pa for 3-21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1-30 Pa for 1-10 h, and controlling water content between 0 and 6 percent to obtain white loose porous spongy amorphous powder. 0.5g of salmon protamine is weighed, mixed with white amorphous powder evenly, ground together and sieved by a 300-mesh sieve to obtain a white composition for later use.

Example 44

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 6.5g of 2-O-sulfobutyl-beta-cyclodextrin is dissolved in 30ml of purified water. Slowly dripping the resveratrol solution into the 2-O-sulfobutyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain a clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1-30 Pa for 3-21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1-30 Pa for 1-10 h, and controlling water content between 0 and 6 percent to obtain white loose porous spongy amorphous powder. 0.5g of salmon protamine is weighed, mixed with white amorphous powder evenly, ground together and sieved by a 300-mesh sieve to obtain a white composition for later use.

Example 45

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 7.5g of 2-O-sulfobutyl-beta-cyclodextrin is taken and dissolved in 30ml of purified water. Slowly dripping the resveratrol solution into the 2-O-sulfobutyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain a clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1-30 Pa for 3-21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1-30 Pa for 1-10 h, and controlling water content between 0 and 6 percent to obtain white loose porous spongy amorphous powder. 0.5g of salmon protamine is weighed, mixed with the white amorphous powder uniformly, ground together and sieved by a 300-mesh sieve to obtain a white composition for later use.

Example 46

1g of resveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a resveratrol solution. 9g of hydroxypropyl-beta-cyclodextrin is taken and dissolved in 20ml of purified water. Slowly dripping the resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove the ethanol, continuously stirring for 0.5h, and then carrying out spray drying at the air inlet temperature of 40-80 ℃, the atomization pressure of 0.2-0.8 MPa, the feeding speed of 100-2000 ml/h and the water content of 0-8% to obtain the microspherical composition. 0.5g of salmon protamine is weighed, mixed with white amorphous powder evenly, ground together and sieved by a 300-mesh sieve to obtain a white composition for later use.

Example 47

1g of resveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a resveratrol solution. Taking 11g of hydroxypropyl-beta-cyclodextrin, and dissolving in 20ml of purified water. Slowly dripping the resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, and then spray-drying at the air inlet temperature of 40-80 ℃, the atomization pressure of 0.2-0.8 MPa, the feeding speed of 100-2000 ml/h and the water content of 0-8 percent to obtain the microsphere composition. 0.5g of salmon protamine is weighed, mixed with white amorphous powder evenly, ground together and sieved by a 300-mesh sieve to obtain a white composition for later use.

Example 48

1g of resveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a resveratrol solution. 6.5g of hydroxypropyl-beta-cyclodextrin is taken and dissolved in 20ml of purified water. Slowly dripping the resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove the ethanol, continuously stirring for 0.5h, and then carrying out spray drying at the air inlet temperature of 40-80 ℃, the atomization pressure of 0.2-0.8 MPa, the feeding speed of 100-2000 ml/h and the water content of 0-8% to obtain the microspherical composition. 0.5g of salmon protamine is weighed, mixed with white amorphous powder evenly, ground together and sieved by a 300-mesh sieve to obtain a white composition for later use.

Example 49

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 6.5g of beta-cyclodextrin is taken and dissolved in 15ml of purified water. Slowly dripping the resveratrol solution into the beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, and then spray-drying at the air inlet temperature of 40-80 ℃, the atomization pressure of 0.2-0.8 MPa, the feeding speed of 100-2000 ml/h and the water content of 0-8 percent to obtain the microsphere composition. 0.5g of salmon protamine is weighed, mixed with white amorphous powder evenly, ground together and sieved by a 300-mesh sieve to obtain a white composition for later use.

Example 50

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 5.0g of beta-cyclodextrin is taken and dissolved in 15ml of purified water. Slowly dripping the resveratrol solution into beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, and then spray-drying at the air inlet temperature of 40-80 ℃, the atomization pressure of 0.2-0.8 MPa, the feeding speed of 100-2000 ml/h and the water content of 0-8 percent to obtain the microsphere composition. 0.5g of salmon protamine is weighed, mixed with white amorphous powder evenly, ground together and sieved by a 300-mesh sieve to obtain a white composition for later use.

Example 51

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 8.0g of beta-cyclodextrin is taken and dissolved in 15ml of purified water. Slowly dripping the resveratrol solution into the beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, and then spray-drying at the air inlet temperature of 40-80 ℃, the atomization pressure of 0.2-0.8 MPa, the feeding speed of 100-2000 ml/h and the water content of 0-8 percent to obtain the microsphere composition. 0.5g of salmon protamine is weighed, mixed with the white amorphous powder uniformly, ground together and sieved by a 300-mesh sieve to obtain a white composition for later use.

Example 52

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 5.0g of 2-O-sulfobutyl-beta-cyclodextrin is taken and dissolved in 30ml of purified water. Slowly dripping the resveratrol solution into the 2-O-sulfobutyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain a clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, and then spray-drying at the air inlet temperature of 40-80 ℃, the atomization pressure of 0.2-0.8 MPa, the feeding speed of 100-2000 ml/h and the water content of 0-8 percent to obtain the microsphere composition. 0.5g of salmon protamine is weighed, mixed with white amorphous powder evenly, ground together and sieved by a 300-mesh sieve to obtain a white composition for later use.

Example 53

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 6.5g of 2-O-sulfobutyl-beta-cyclodextrin is dissolved in 30ml of purified water. Slowly dripping the resveratrol solution into the 2-O-sulfobutyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain a clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove the ethanol, continuously stirring for 0.5h, and then carrying out spray drying at the air inlet temperature of 40-80 ℃, the atomization pressure of 0.2-0.8 MPa, the feeding speed of 100-2000 ml/h and the water content of 0-8% to obtain the microspherical composition. 0.5g of salmon protamine is weighed, mixed with white amorphous powder evenly, ground together and sieved by a 300-mesh sieve to obtain a white composition for later use.

Example 54

1g of resveratrol and 0.02g of coumarin are weighed out and dissolved in 10ml of ethanol. 7.5g of 2-O-sulfobutyl-beta-cyclodextrin is dissolved in 30ml of purified water. Slowly dripping the resveratrol solution into the 2-O-sulfobutyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain a clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove the ethanol, continuously stirring for 0.5h, and then carrying out spray drying at the air inlet temperature of 40-80 ℃, the atomization pressure of 0.2-0.8 MPa, the feeding speed of 100-2000 ml/h and the water content of 0-8% to obtain the microspherical composition. 0.5g of salmon protamine is weighed, mixed with the white amorphous powder uniformly, ground together and sieved by a 300-mesh sieve to obtain a white composition for later use.

Example 55

1g of resveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a resveratrol solution. 9.0g of hydroxypropyl-beta-cyclodextrin is taken and dissolved in 20ml of purified water. Slowly dripping the resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. The ethanol was removed by distillation under reduced pressure at 45 ℃. 0.5g of trout protamine was weighed and dissolved in 5ml of purified water. Slowly dripping the trout protamine under the continuous stirring of the cyclodextrin, continuously stirring for 0.5h, then carrying out freeze drying, setting the sublimation drying at-80 to-40 ℃, the vacuum degree at 1 to 30Pa and the time at 3 to 21 h, setting the desorption drying at-10 to +20 ℃, the vacuum degree at 1 to 30Pa and the time at 1 to 10 h, and controlling the water content at 0 to 6 percent to obtain white loose porous spongy amorphous powder. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 56

1g of resveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a resveratrol solution. 9.0g of hydroxypropyl-beta-cyclodextrin is taken and dissolved in 20ml of purified water. Slowly dripping the resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1-30 Pa for 3-21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1-30 Pa for 1-10 h, and controlling water content between 0 and 6 percent to obtain white loose porous spongy amorphous powder. Weighing 0.5g of trout protamine, uniformly mixing with the white amorphous powder, grinding together, and sieving by a 300-mesh sieve to obtain a white composition for later use.

Example 57

1g of resveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a resveratrol solution. 9.0g of hydroxypropyl-beta-cyclodextrin is taken and dissolved in 20ml of purified water. Slowly dripping the resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. The ethanol was removed by distillation under reduced pressure at 45 ℃. 0.5g of trout protamine was weighed and dissolved in 5ml of purified water. Slowly dripping trout protamine into the mixture under the continuous stirring of cyclodextrin, continuously stirring for 0.5h, then carrying out spray drying, wherein the air inlet temperature is 40-80 ℃, the atomization pressure is 0.2-0.8 MPa, the feeding speed is 100-2000 ml/h, and the water content is controlled to be 0-8%, thus obtaining the microspherical composition. Grinding, and sieving with a 300-mesh sieve to obtain a white composition for later use.

Example 58

1g of resveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a resveratrol solution. 9.0g of hydroxypropyl-beta-cyclodextrin is taken and dissolved in 20ml of purified water. Slowly dripping the resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, and then spray-drying at the air inlet temperature of 40-80 ℃, the atomization pressure of 0.2-0.8 MPa, the feeding speed of 100-2000 ml/h and the water content of 0-8 percent to obtain the microsphere composition. Weighing 0.5g of trout protamine, uniformly mixing with the microspherical powder, grinding together, and sieving by a 300-mesh sieve to obtain a white composition for later use.

Example 59

1g of resveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a resveratrol solution. 9.0g of hydroxypropyl-beta-cyclodextrin is taken and dissolved in 20ml of purified water. Slowly dripping the resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. The ethanol was removed by distillation under reduced pressure at 45 ℃. 0.5g of herring protamine was weighed and dissolved in 5ml of purified water. Slowly dripping herring protamine into cyclodextrin under continuous stirring, continuously stirring for 0.5h, freeze drying, sublimation drying at-80 to-40 deg.C under vacuum degree of 1-30 Pa for 3-21 h, desorption drying at-10 to +20 deg.C under vacuum degree of 1-30 Pa for 1-10 h, and controlling water content at 0-6% to obtain white loose porous sponge-like amorphous powder. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 60

1g of resveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a resveratrol solution. 9.0g of hydroxypropyl-beta-cyclodextrin is taken and dissolved in 20ml of purified water. Slowly dripping the resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1-30 Pa for 3-21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1-30 Pa for 1-10 h, and controlling water content between 0 and 6 percent to obtain white loose porous spongy amorphous powder. Weighing 0.5g herring protamine, mixing with white amorphous powder, grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 61

1g of resveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a resveratrol solution. 9.0g of hydroxypropyl-beta-cyclodextrin is taken and dissolved in 20ml of purified water. Slowly dripping the resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. The ethanol was removed by distillation under reduced pressure at 45 ℃. 0.5g herring protamine was weighed out and dissolved in 5ml purified water. Slowly dripping herring protamine into cyclodextrin under continuous stirring, continuously stirring for 0.5h, spray drying, wherein the air inlet temperature is 40-80 ℃, the atomization pressure is 0.2-0.8 MPa, the feeding speed is 100-2000 ml/h, and the water content is controlled between 0-8 percent to obtain the microsphere composition. Grinding, and sieving with 300 mesh sieve to obtain white composition.

Example 62

1g of resveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a resveratrol solution. 9.0g of hydroxypropyl-beta-cyclodextrin is taken and dissolved in 20ml of purified water. Slowly dripping the resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. Distilling under reduced pressure at 45 ℃ to remove ethanol, continuously stirring for 0.5h, and then spray-drying at the air inlet temperature of 40-80 ℃, the atomization pressure of 0.2-0.8 MPa, the feeding speed of 100-2000 ml/h and the water content of 0-8 percent to obtain the microsphere composition. Weighing 0.5g herring protamine, mixing with the microspherical powder, grinding together, and sieving with 300 mesh sieve to obtain white composition.

Example 63

1g of pterostilbene and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain a pterostilbene solution. 9g of hydroxypropyl-beta-cyclodextrin is taken and dissolved in 20ml of purified water. Slowly dripping the pterostilbene solution into the hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain clear, colorless and transparent solution. The ethanol was removed by distillation under reduced pressure at 45 ℃. 0.5g of salmon protamine was weighed and dissolved in 5ml of purified water. Slowly dripping salmon protamine into cyclodextrin under continuous stirring, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1 to 30Pa for 3 to 21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1 to 30Pa for 1 to 10 h, and controlling water content to be 0 to 6 percent to obtain white loose porous spongy amorphous powder. Grinding, and sieving with a 300-mesh sieve to obtain a white composition for later use.

Example 64

1g of oxyresveratrol and 0.02g of coumarin are weighed and dissolved in 10ml of ethanol to obtain the oxyresveratrol solution. 9g of hydroxypropyl-beta-cyclodextrin is taken and dissolved in 20ml of purified water. Slowly dripping the oxidized resveratrol solution into hydroxypropyl-beta-cyclodextrin under magnetic stirring (300 r/min), and continuously stirring for 2h to obtain a clear, colorless and transparent solution. The ethanol was removed by distillation under reduced pressure at 45 ℃. 0.5g of salmon protamine was weighed and dissolved in 5ml of purified water. Slowly dripping salmon protamine into cyclodextrin under continuous stirring, continuously stirring for 0.5h, freeze-drying, setting sublimation drying at-80 to-40 ℃, vacuum degree of 1 to 30Pa for 3 to 21 h, setting desorption drying at-10 to +20 ℃, vacuum degree of 1 to 30Pa for 1 to 10 h, and controlling water content to be 0 to 6 percent to obtain white loose porous spongy amorphous powder. Grinding, and sieving with a 300-mesh sieve to obtain a white composition for later use.

Example 65

Fibroblasts, plates (6-well plates) were cultured by conventional methods. Examples 1-18 are compositions without added roe (blank group) and examples 19-62 are compositions with added roe (roe extract group). The blank group was added with a resveratrol clathrate-medium solution (resveratrol concentration 50 ug/ml) with a fluorescent probe (coumarin). Adding resveratrol clathrate-culture medium solution (resveratrol concentration of 50ug/ml and roe extract concentration of 25 ug/ml) with fluorescent probe (coumarin) and roe extract. After 4h incubation of the cells, the cells were treated by the following procedure: after incubation, the medium was removed, 300ul of pancreatin was added to each well, digestion was carried out for 2min, the medium was added to stop digestion, pipetting three times, centrifugation (1200 r/min,3 min), supernatant was discarded, 1ml of PBS was added to the precipitate, centrifugation (1200 r/min,3 min) was carried out, supernatant was discarded, 0.3ml of PBS was added to the precipitate, cell uptake was measured using a cell flow meter, and the results were shown in table 1.

The FL1H median (detection channel corresponding to coumarin) in the flow cytometer data was statistically averaged, and the results showed statistical significance (P < 0.01).

Table 1 shows the composition of each of the compositions prepared in examples 1-62 and the results in the fibroblast experiment.

TABLE 1

Examples 1-18 are mean FL1H median values without added roe extract (common inclusion compound) and examples 19-62 are mean FL1H median values with added roe extract samples (added roe inclusion compound), the values indicating that the fibroblast uptake is significantly higher with added roe inclusion compound than with added roe inclusion compound. The addition of the fish egg extract can promote the resveratrol composition to be absorbed by fibroblasts.

As shown in fig. 1A-1B, the uptake of resveratrol-conjugated protamine composition of the invention (example 19) was compared to the uptake of the common resveratrol composition (example 1) in fibroblasts (n = 3). The results show that the mean value of the median FL1H of example 1 without the addition of roe extract (common inclusion compound) was about 53581, and that the mean value of the median FL1H of example 19 with the addition of roe extract sample (with the addition of roe inclusion compound) was about 96385, which is significantly higher than the former. The addition of the fish egg extract can promote the resveratrol composition to be absorbed by fibroblasts.

Example 66

Langerhans cells, plates (6-well plates) were cultured by conventional methods. Examples 1-18 are compositions without added roe (blank group) and examples 19-62 are compositions with added roe (roe extract group). The blank group is added with resveratrol clathrate-culture medium solution (resveratrol concentration is 50 ug/ml) with fluorescent probe (coumarin), and roe extract group is added with resveratrol clathrate-culture medium solution (resveratrol concentration is 50ug/ml, roe extract concentration is 25 ug/ml) with fluorescent probe (coumarin). After 4h incubation of the cells, the cells were treated by the following procedure: after incubation, removing the culture medium, adding 300ul of pancreatin into each well, digesting for 2min, adding the culture medium to stop digestion, beating for three times, centrifuging (1200 r/min,3 min), discarding the supernatant, taking the precipitate, adding 1ml of PBS to beat, centrifuging (1200 r/min,3 min), discarding the supernatant, adding 0.3ml of PBS to beat, measuring the cell uptake by using a cell flow meter, and detecting on a computer, wherein the results are shown in Table 2.

Table 2 shows the composition of each of the compositions prepared in examples 1-62 and the results in the langerhans cell experiment.

TABLE 2

Examples 1-18 are mean values for the median FL1H without added roe extract (common inclusion compound) and examples 19-62 are mean values for the median FL1H with added roe extract samples (added roe inclusion compound), indicating that the uptake of langerhans cells is significantly higher with added roe inclusion compound than with added roe inclusion compound. The addition of the fish egg extract can promote the absorption of the resveratrol composition by Langerhans cells.

As shown in fig. 2A-2B, the resveratrol conjugated protamine composition of example 19 of the present invention was compared with the uptake of the ordinary resveratrol composition of example 1 in langerhans cells (n = 3). The results showed that the mean value of the median FL1H of example 1 without the addition of roe extract (common inclusion compound) was about 7896, and the mean value of the median FL1H of example 19 with the addition of roe extract (with the addition of roe inclusion compound) was about 15627, which is significantly higher than the former. The addition of the fish egg extract can promote the absorption of the resveratrol composition by Langerhans cells.

Example 67

Melanocytes, plates (6-well plates) were cultured by conventional methods. Examples 1-18 are compositions without added roe (blank group) and examples 19-62 are compositions with added roe (roe extract group). The blank group is added with resveratrol clathrate-culture medium solution (resveratrol concentration is 50 ug/ml) with fluorescent probe (coumarin), and roe extract group is added with resveratrol clathrate-culture medium solution (resveratrol concentration is 50ug/ml, roe extract concentration is 25 ug/ml) with fluorescent probe (coumarin). After 4h incubation of the cells, the cells were treated by the following procedure: after incubation, the medium was removed, 300ul of pancreatin was added to each well, digestion was carried out for 2min, the medium was added to stop digestion, pipetting three times, centrifugation (1200 r/min,3 min), supernatant was discarded, 1ml of PBS was added to the precipitate and pipetting, centrifugation (1200 r/min,3 min) was discarded, 0.3ml of PBS was added and pipetting, cell uptake was measured using a cell flow meter, and the results were shown in table 3.

Table 3 shows the composition of each of the compositions prepared in examples 1-62 and the results in the melanocyte uptake assay.

TABLE 3

Examples 1-18 are mean values for the median FL1H without added roe extract (common inclusion complex) and examples 19-62 are mean values for the median FL1H with added roe extract samples (added roe inclusion complex), indicating that the uptake of melanocytes is significantly higher with added roe inclusion complex than with the former. The addition of the fish egg extract can promote the absorption of the resveratrol composition by melanocytes.

As shown in fig. 3A-3B, the resveratrol conjugated protamine composition of example 19 of the present invention was compared with the conventional resveratrol composition of example 1 in terms of the uptake of melanocytes (n = 3). The results showed that the mean value of the FL1H median of example 1 without the addition of roe extract (common inclusion) was about 6018, and the mean value of the FL1H median of the roe extract-added sample (with the addition of roe inclusion) was about 9917, which is significantly higher than the former. The addition of the fish egg extract can promote the absorption of the resveratrol composition by melanocytes.

Application example

The composition of the present invention can be used as an intermediate material for preparing a skin external preparation, preferably a cosmetic composition, including but not limited to the preparation of products in the form of cream, lotion, jelly, lotion, essence, mask, eye cream, aerosol (cleansing foam), spray, body wash, face wash, etc., and the compositions obtained in examples 1 to 62 have a weight percentage of 0.0001% to 20% (w/w) in the skin external preparation. The preferred weight percentage is 0.001% -10% (w/w). More preferably 0.001-5% (w/w). The most preferred weight percentage is 0.01% -5% (w/w). The following are specific examples of the application of the compositions obtained in examples 1 to 62 to external preparations for skin, and the formulation and preparation methods of these preparations. The specific application examples are as follows:

application example 1: preparation of face cream

Application example 2: preparation of the emulsion

Application example 3: preparation of jelly

Application example 4: preparation of astringent

Application example 5: preparation of essence

Application example 6: preparation of facial mask

/>

Application example 7: preparation of eye cream

Application example 8: preparation of an aerosol (cleaning foam)

Application example 9: preparation of the spray

Application example 10: preparation of shower gel

/>

Application example 11: preparation of facial cleanser

/>

/>

Claims

1. An active agent-cyclodextrin inclusion complex comprising:

an active agent which is resveratrol;

cyclodextrin which is hydroxypropyl-beta-cyclodextrin, the molar ratio of the active agent to the cyclodextrin being 1:5 to 1:1;

protamine, the weight ratio of the protamine to the active agent is 2:1 to 1:2,

the active agent-cyclodextrin inclusion compound is prepared by the following method:

1c) Adding the solution of the active agent prepared in the step 1 a) into the solution of the cyclodextrin prepared in the step 1 b) to obtain a mixed solution, filtering the mixed solution, collecting filtrate, and distilling the collected filtrate under reduced pressure to remove the organic solvent;

1e) Adding the protamine solution prepared in step 1 d) to the mixture of step 1 c);

1f) Freeze drying or spray drying to obtain cyclodextrin clathrate of protamine-containing active agent;

alternatively, the first and second electrodes may be,

2e) Mixing protamine with the active agent-cyclodextrin inclusion compound prepared in step 2 d) to obtain an active agent-cyclodextrin inclusion compound containing protamine,

wherein the protamine is salmon protamine.

2. The active agent-cyclodextrin inclusion complex of claim 1, wherein the molar ratio of the active agent to the cyclodextrin is from 1.3 to 1.5.

3. The active agent-cyclodextrin inclusion complex of claim 1, wherein the freeze-drying is configured to: sublimation drying is carried out at the temperature of-80 to-40 ℃ and the vacuum degree of 1 to 30Pa for 3 to 21 hours, desorption drying is carried out at the temperature of-10 to +20 ℃ and the vacuum degree of 1 to 30Pa for 1 to 10 hours, and the water content is controlled between 0 and 6 percent.

4. The active agent-cyclodextrin inclusion complex of claim 1, wherein the spray drying is configured to: the air inlet temperature is 40-80 ℃, the atomization pressure is 0.2-0.8 MPa, the feeding speed is 100-2000 ml/h, and the water content is controlled between 0-8%.

5. The active agent-cyclodextrin inclusion complex of claim 1, wherein the organic solvent in step 1 a) or step 2 a) is ethanol.

6. Use of the active agent-cyclodextrin inclusion compound according to claim 1 for the preparation of cosmetics.