CN101816632B - Composite functional liposome and preparation method and application thereof - Google Patents

Abstract

The invention discloses a composite functional liposome and a preparation method and application thereof. The composite functional liposome comprises the following components: granulesten, cholesterin, vitamin C and salicylic acid, wherein the mass ratio of the four components is 5-7:1.3-1.5:0.8-1.5:1. The method for preparing the composite functional liposome comprises the following steps of: dissolving the granulesten, the cholesterin and the salicylic acid in an organic solvent to obtain a mixture serving as oil phase; emulsifying the mixture and vitamin C-containing buffer solution which serves as water phase to form water-in-oil microemulsion; emulsifying the water-in-oil microemulsion and buffer solution to finally form water/oil/water multiple emulsion, wherein the emulsification process is completed by ultrasonication; and removing the organic solvent through decompression and distillation to obtain the composite functional liposome. The composite functional liposome has stable property, contains fat-soluble medicaments and water-soluble medicaments, exerts the effects of various medicaments to compound the functions of the liposome, is applied to cosmetics, improves the stability of the medicaments, and reduces the production cost.

Description

A kind of composite functional liposome and its preparation method and application

technical field

The invention relates to a preparation method of liposome, in particular to a composite functional liposome and its preparation method and application.

Background technique

Liposome was discovered and named by Dr. Bangham in 1965. Its structure dimension is a closed vesicle with a bimolecular membrane as a sandwich wall. Various lipids and lipid mixtures can be used to prepare liposomes, with phospholipids being the most commonly used. The main components of phospholipids are phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, phosphatidylglycerol, and phosphatidic acid. Phospholipids are the main components of cell membranes, so liposomes are similar in structure to human cells and have a high affinity for human cells. Phospholipids are amphiphilic molecules whose structure contains a hydrophilic group (a phosphate group and a nitrogenous base) and a hydrophobic group (two longer hydrocarbon chains). Under certain concentration conditions, the polar groups face the water phase on both sides, while the hydrocarbon chains of the non-polar groups face each other and associate into a stable bilayer structure. Utilizing the structural characteristics of liposomes, the lipophilic components can be loaded in the middle of the bilayer molecules, and the hydrophilic components can be sealed and embedded in the center of the sphere. Another type of substance that constitutes lipids is cholesterol, which is more hydrophobic than hydrophilic, embedded in the middle of the bilayer formed by phospholipids, and mainly plays a role in changing the properties of the pure phospholipid layer. It acts like a "buffer" It plays the role of regulating the "fluidity" of the membrane structure.

Liposomes are widely used in the fields of medicine, health food, cosmetics and genetic engineering. One of the important applications is as a drug carrier. The drug is encapsulated in the water phase and membrane phase of the liposome, and the targeting effect of the liposome is controlled so that it is enriched in the lesion to release the drug, thereby reducing the dose of the drug required and greatly avoiding the impact of the drug on the human body. Damage to normal parts. In recent years, the development of sterically stable liposomes has greatly improved the stability of liposomes in vivo, making liposomes as drug carriers moving towards a practical stage in the treatment of cancer and other diseases. With the development of cosmetics towards functionalization, specialization and serialization, bioengineering preparations such as hyaluronic acid, liposomes, ceramides and epidermal growth factors are widely used in skin care products. Liposomal cosmetics from "Capture". After the stability of the liposome is improved, it is used as a delivery system for cosmetic active components to increase the effect, storage and protection components of cosmetics; it can control the release (sustained release) of lipid components during use.

Vitamin C, also called ascorbic acid, is a water-soluble vitamin with good anti-oxidation function. It can be used to promote the synthesis of collagen and mucopolysaccharides, reduce free radical damage to the skin, delay aging, and at the same time inhibit the abnormal pigmentation of the skin. Deposition and tyrosinase activity can reduce the formation of melanin, so it is widely used in whitening and anti-oxidation beauty and skin care cosmetics. But vitamin C is easy to oxidize and deteriorate. If it is directly added to the cream base by ordinary methods, it will lose its effect in three months. Moreover, vitamin C is water-soluble and difficult to penetrate into the stratum corneum of the skin, so it is difficult to fully exert its effect. If it is encapsulated by liposome, it can increase the stability of vitamin C and make it have a longer effect.

Salicylic acid is a commonly used cosmetic additive, which has the functions of exfoliating, shrinking pores, removing blackheads, and reducing fine lines and wrinkles. The principle of action is that salicylic acid can dissolve the constituent substances between the cuticles and cause the cuticles to fall off, so it can remove the accumulated thick cuticles and promote metabolism. However, the aqueous solution of salicylic acid is acidic and irritating to a certain extent, and is generally dissolved in ethanol first and then added to cosmetics, and a certain amount of ethanol must be added, so it will cause discomfort or even allergies to sensitive skin.

Contents of the invention

The object of the present invention is to provide a composite functional liposome according to the problems of existing cosmetic additives such as easy deterioration or irritation, which can embed a variety of cosmetic additives, delay the oxidative deterioration of drugs, and utilize the liposome Affinity and promote skin penetration to increase the effective absorption of drugs and reduce the irritation of drugs.

Another object of the present invention is to provide a method for preparing the above-mentioned composite functional liposome.

Another object of the present invention is to provide the application of the composite functional liposome.

The above-mentioned purpose of the present invention is achieved through the following technical solutions:

A composite functional liposome, comprising the following components: soybean lecithin, cholesterol, vitamin C and salicylic acid, the mass ratio of the four is 5~7:1.3~1.5:0.8~1.5:1, preferably 9:2 : 1.8: 1.6.

The preparation method of the complex functional liposome of the present invention comprises the following steps: using soybean lecithin, cholesterol, salicylic acid and organic solvent as the oil phase, and a buffer solution containing vitamin C as the water phase, forming a water-in-oil liposome through the first emulsification The emulsion is then emulsified with a buffer solution for the second time to finally form a double emulsion of water/oil/water. The emulsification process is completed by ultrasonic crushing, and then the organic solvent is removed by distillation under reduced pressure to obtain a composite functional liposome.

The present invention is mainly aimed at the application in cosmetics, so among the medicines embedded in liposomes, salicylic acid is selected as the fat-soluble medicine, and vitamin C is used as the water-soluble medicine. However, when the fat-soluble drug and the water-soluble drug are other drugs, the preparation method of the present invention can still be used.

In the above preparation method, the mass ratio of soybean lecithin, cholesterol and salicylic acid is preferably 4.6-8:1-1.7:1, most preferably 5:1.3:1. The concentration of soybean lecithin is one of the important influencing factors in the preparation, and it has a great influence on the time of the second emulsification. Experiments have proved that when other conditions remain unchanged, soybean phospholipids have a wider dosage range, and the higher the concentration of phospholipids, the more difficult it is to form stable liposomes, and the longer the second emulsification time; and the concentration is too small and difficult to achieve Higher embedding rate, because the fat-soluble drug component is loaded in the middle of the bilayer molecule, which has a certain limit on the concentration of phospholipids. When the encapsulation efficiency of fat-soluble drug is too low, the result will be that the drug will separate out in the water phase and aggregate with liposomes to form stratification. Experimental data show that when salicylic acid is used as fat-soluble drug, the mass ratio of soybean lecithin to fat-soluble drug should not be less than 4.6:1.

As a preferred solution, the organic solvent in the preparation method of the present invention is diethyl ether, and the amount added is based on completely dissolving the raw materials.

The liposomes prepared by the double emulsion method can contain more water-soluble drugs, so the concentration of water-soluble drugs has little influence on liposome stability, but the vitamin C aqueous solution is not conducive to the formation of liposomes because of too high acidity, so When preparing the inner water phase, that is, the vitamin C aqueous solution, attention should be paid to adjusting its pH value. In the above preparation method, vitamin C is dissolved in Na ₂ HPO ₄ -K ₂ HPO ₄ buffer solution, adjusted to pH 4~5 with a small amount of Na ₂ HPO ₄ aqueous solution, and prepared to a concentration of 0.04 g/mL for use. Experiments have shown that when the pH of the internal aqueous phase is less than 4, the liposomes formed are easy to separate.

When liposomes are prepared by the double emulsion method, the aqueous solution added to the oil phase is dispersed into small water droplets under the action of ultrasound. Phospholipids and cholesterol are adsorbed on the surface of water droplets, with the hydrophilic groups facing inward and the hydrophobic groups facing outward to form a monomolecular film, thus forming water-in-oil (W/O) colostrum. After the buffered aqueous solution is added to the emulsion, the organic solvent is volatilized under ultrasound, and the excess phospholipids and cholesterol in the oil phase quickly form a monomolecular film at the oil-water interface with the buffer solution, and the small water droplets in the oil phase pass through the single molecule of the oil-water interface. The membrane is surrounded by it, the hydrophobic groups associate face-to-face, and the hydrophilic groups face the water phase on both sides, forming a bilayer liposome structure in the water phase.

The first emulsification is easy to achieve, the ultrasonic time is generally 1~2min, and the water bath temperature is 25~35℃. The key to emulsification is the ratio of water to oil. The water phase should not exceed half of the oil phase, usually one-third or one-fourth of the oil phase. Because diethyl ether is used as the organic solvent, the boiling point is only 34.5°C, and the ultrasonic wave can volatilize diethyl ether. When the water phase is too much, it is easy to reach the phase inversion point. When more than 2 times the drug-free water phase is added, the phase inversion may occur, that is, the formation of O/ W emulsion instead of double emulsion. The colostrum formed by the first emulsification is a uniform turbid liquid, which does not coagulate and separate after being placed for 2 minutes.

The second emulsification depends on the ultrasonic power, time and concentration of soybean lecithin. When emulsifying, the colostrum is added to the water phase containing more than 2 times the volume of the drug, and the Na ₂ HPO ₄ -K ₂ HPO ₄ buffer solution with pH=7.4 is used in the experiment. The colostrum floats on the upper layer with the oil phase, and the oil phase is reduced by ultrasonic stirring, emulsification appears in the phase interface, and the emulsion is formed and diffused into the water phase. The time required is 5-10 minutes. Ultrasound can make the ether volatilize quickly, and the colostrum particles move violently, and can pass through the monomolecular membrane in a short time to form a single-chamber liposome with a smaller particle size. If the ultrasonic power is too small, the time for the colostrum particles to pass through the monomolecular membrane will increase, and it is easy to form heterogeneous multi-lamellar liposomes with larger particle size and shorter layering time, so the second emulsification power should be kept Above 100W. The oil phase disappears and there are no fat blocks, forming a uniform and slightly transparent emulsion to end the ultrasonication, avoiding liposome rupture caused by prolonged ultrasonication, and reducing the embedding rate.

During the preparation, vacuum distillation was used to remove the remaining ether, the vacuum degree was 0.08-0.09Mpa, the distillation time was 5-15min, and the water bath temperature was 30-35°C. If it is adjusted to a higher vacuum degree, heating is not required, which is more conducive to protecting liposomes and drugs from oxidation.

The composite functional liposome of the present invention can contain fat-soluble drugs and water-soluble drugs at the same time, and when used as a drug carrier, it can exert the effects of various drugs and reduce the irritation of drugs.

Compared with the prior art, the present invention has the following beneficial effects:

(1) With reference to the literature records of the preparation of existing liposomes, in the past, liposomes mostly added a small amount of antioxidants for embedding a drug, and the realized function was relatively single; the composite functional liposomes of the present invention can fill this deficiency , which can realize the inclusion of drugs with greatly different properties in the same liposome, and save the various processing steps required when adding cosmetics and drugs due to the different properties of the drugs on the basis of the original excellent properties of the liposomes;

(2) The compound functional liposome of the present invention is stable in property, has good encapsulation efficiency to the medicine of various different properties, has improved the stability of medicine, has reduced production cost, can be widely used in medicine, health food, cosmetics and genetic engineering;

(3) The composite functional liposome of the present invention embeds vitamin C and salicylic acid in the liposome simultaneously, delays the oxidative deterioration process of the medicine, and utilizes the cell affinity of the liposome and promotes skin permeability, It increases the effective absorption of drugs, and when used in the preparation of cosmetics, it can achieve various functions such as anti-oxidation and whitening.

Detailed ways

The present invention is further explained below in conjunction with the examples, but the examples do not limit the present invention in any form.

Example 1

Dissolve vitamin C in a small amount of Na ₂ HPO ₄ -K ₂ HPO ₄ buffer solution, add Na ₂ HPO ₄ aqueous solution dropwise to adjust the pH to 5, and use the buffer solution to prepare a concentration of 0.04 g/mL. Dissolve 0.15g of soybean lecithin, 0.04g of cholesterol, and 0.03g of salicylic acid in 4mL of ether, add 1mL of the above-mentioned vitamin C aqueous solution, the power is 90W, and the temperature of the water bath is 30°C. for colostrum. Add colostrum into 10mL of Na ₂ HPO ₄ -K ₂ HPO ₄ buffer solution with pH=7.4, use 100W power, and ultrasonically break at 45°C for 6 minutes until the oil layer completely disappears, forming a uniform and slightly transparent emulsion, which is W /O/W double milk. Thereafter, rotary steaming was performed under reduced pressure at 35°C for 8 min until there was no ether smell. The prepared liposome emulsion is packed into a centrifuge tube with a volume of 2mL, centrifuged for 1h, and the rotating speed is 10000rad/min, and the supernatant is taken for ultraviolet absorption measurement, and the encapsulation efficiency of vitamin C is 45%. The encapsulation efficiency of acid was 52%.

Example 2

Dissolve 0.15g of soybean lecithin, 0.04g of cholesterol, and 0.03g of salicylic acid in 4mL of ether, add 1mL of vitamin C aqueous solution with a pH of 3.8 and a concentration of 0.04g/mL, with a power of 90W and a water bath temperature of 25°C for 2min to A homogeneous turbid liquid formed by the disappearance of the layer. Add the prepared colostrum into 10 mL of Na ₂ HPO ₄ -K ₂ HPO ₄ buffer solution with pH=7.4, and ultrasonically break it for 7 minutes at a power of 100 W and a water bath temperature of 40°C until the oil layer disappears to form a uniform slightly transparent double emulsion. Thereafter, rotary steaming was performed under reduced pressure at 30°C for 10 min until there was no ether smell. The prepared liposome emulsion is packed into a centrifuge tube with a volume of 2mL, centrifuged for 1h, and the rotating speed is 10000rad/min, and the supernatant is taken for ultraviolet absorption measurement, and the encapsulation efficiency of vitamin C is 32%. The encapsulation efficiency of acid is 38%.

Example 3

Take 1.5mL vitamin C aqueous solution with a pH of 5 and a concentration of 0.04g/mL, add 4mL of ether solution dissolved with 0.2g soybean lecithin, 0.04g cholesterol, and 0.04g salicylic acid, with a power of 90W and a water bath temperature of 30°C Ultrasound for 2 minutes until the stratification disappears and uniform turbid colostrum is formed. Add colostrum into 10mL of Na ₂ HPO ₄ -K ₂ HPO ₄ buffer solution with pH=7.4, and ultrasonically crush it for 6 minutes at a power of 100W and a water bath temperature of 45°C until the oil layer completely disappears, forming a uniform and slightly transparent double emulsion. Then, it was rotary evaporated under reduced pressure at 35°C for 10 min until there was no ether smell. The prepared liposome emulsion is packed into a centrifuge tube with a volume of 2mL, centrifuged for 1h, and the rotating speed is 10000rad/min, and the supernatant is taken for ultraviolet absorption measurement, and the encapsulation efficiency of vitamin C is 40%. The encapsulation efficiency of acid was 39%.

Example 4

Dissolve 0.18g of soybean lecithin, 0.05g of cholesterol, and 0.03g of salicylic acid in 5mL of ether, add 1mL of vitamin C aqueous solution with a pH of 5 and a concentration of 0.04g/mL, with a power of 100W and a water bath temperature of 25°C for 1.5min to The stratification disappears, and the uniform turbid liquid formed is colostrum. Add colostrum into 15mL Na ₂ HPO ₄ -K ₂ HPO ₄ buffer solution with pH=7.4, use 100W power, 40°C water bath temperature, and ultrasonically crush for 8 minutes until the oil layer completely disappears, forming a uniform and slightly transparent emulsion, which is W /O/W double milk. Thereafter, the mixture was rotary evaporated under reduced pressure at 30°C for 12 minutes until there was no ether smell. The prepared liposome emulsion is packed into a centrifuge tube with a volume of 2mL, centrifuged for 1h, and the rotating speed is 10000rad/min, and the supernatant is taken for ultraviolet absorption measurement, and the encapsulation efficiency of vitamin C is 42%. The encapsulation efficiency of acid is 45%.

Example 5

Weigh 0.16g of soybean lecithin, 0.03g of cholesterol, 0.03g of salicylic acid and dissolve in 4mL of ether, add 1mL of vitamin C aqueous solution with a pH of 5 and a concentration of 0.04g/mL. 1min until the layering disappears, and the uniform turbid liquid formed is colostrum. Add colostrum into 12mL of Na ₂ HPO ₄ -K ₂ HPO ₄ buffer solution with pH=7.4, power at 100WA, water bath temperature at 50°C, and ultrasonically crush for 5 minutes until the oil layer completely disappears and a uniform slightly transparent emulsion is formed, which is W/ O/W complex milk. Thereafter, the solution was rotary evaporated under reduced pressure at 35°C for 12 minutes until there was no ether smell. The prepared liposome emulsion is packed into a centrifuge tube with a volume of 2mL, centrifuged for 1.2h, and the rotating speed is 8000rad/min, and the supernatant is taken for ultraviolet absorption measurement, and the encapsulation efficiency of vitamin C is 40%. The encapsulation efficiency of salicylic acid was 43%.

Claims (3)

1. A composite functional liposome simultaneously carrying vitamin C and salicylic acid, comprising the following components: soybean lecithin, cholesterol, vitamin C and salicylic acid, the mass ratio of the four being 5～7: 1.3～1.5 : 0.8～1.5:1; its preparation method comprises the following steps: take soybean lecithin, cholesterol, salicylic acid and organic solvent as oil phase, the buffer solution containing vitamin C as water phase, form water-in-oil microstructure through first emulsification Emulsion colostrum is emulsified with buffer solution for the second time to finally form water/oil/water double emulsion. The emulsification process is completed by ultrasonic crushing, and then the organic solvent is removed by vacuum distillation to obtain composite functional liposomes; wherein, The mass ratio of the soybean lecithin, cholesterol and salicylic acid is 4.6～8:1～1.7:1; The organic solvent is diethyl ether, and the amount added is subject to completely dissolving the raw materials; the water phase containing vitamin C is vitamin C dissolved in Na ₂ HPO ₄ -K ₂ HPO ₄ buffer solution, the concentration is 0.04g/ml, pH 4-5; the buffer solution is Na ₂ HPO ₄ -K ₂ HPO ₄ buffer solution with pH=7.4; The temperature of the water bath for the first emulsification is 25-35°C, the ultrasonic time is 1-2min, and the power is 90-100W; the temperature of the water bath for the second emulsification is 40-50°C, and the ultrasonic time is 5-8min. The power is 100W; The colostrum is a uniform turbid liquid, which is not coagulated and stratified after being placed for 2 minutes; the double emulsion is a uniform and slightly transparent emulsion, the dispersed phase is the particles of the colostrum, and the continuous phase is the water phase without water-soluble drugs; The vacuum degree of the vacuum distillation is 0.08-0.09Mpa, the temperature of the water bath is 30-35°C, and the distillation time is 5-15min. 2. according to the composite functional liposome of claim 1, which simultaneously entraps vitamin C and salicylic acid, it is characterized in that the mass ratio of soybean lecithin, cholesterol, vitamin C and salicylic acid is 9: 2: 1.8: 1.6. 3. the application of the complex function liposome of claim 1 or 2 that simultaneously entraps vitamin C and salicylic acid in the preparation of a drug carrier that reduces drug irritation.