KR100716802B1 - Preparation of ph-responsive polymer-liposome complexes and the composition of skin external application containing the same - Google Patents

Abstract

The present invention relates to the preparation of a polymer-liposomal nanocomposite using an acidity sensitive polymer and an external composition for skin containing the same, and specifically, using an acidity sensitive polymer composed of methacrylic acid and n-alkyl methacrylate, lipids and cholesterol. As a manufactured polymer-liposomal nanocomposite or nanoemulsion, it has a sensitive sensitivity to changes in acidity, which effectively releases physiologically active ingredients incorporated into the inner phase in vivo, thereby enhancing the effect as a transporter and preparing based on conventional lipids. The present invention relates to a polymer-lipid-based drug carrier and an external dermal composition designed to be more stably present in an aqueous solution and various salt-containing formulations and skin external preparation compositions and cosmetic formulations.

Acidity Sensitive Polymers, Polymer-Liposome Nanocomposites, Liposomes, Nanoemulsions, Formulation Stability

Description

Preparation of polymer-liposome nanocomposites containing acidity sensitive polymers and composition for external preparation containing skin {{{preparation of pH-responsive polymer-liposome complexes and the composition of skin external application containing the same}}

1: Schematic diagram of polymer-liposomal nanocomposite

2 is a graph showing the change in calories with temperature in the aqueous solution of the polymer-liposomal nanocomposite according to the present invention

3: A graph showing a change in calories with temperature when a conventional lipid-cholesterol-based liposome dispersion is mixed according to the composition of a nanoemulsion formulation

4: Graph showing the change in calories with temperature when the polymer-liposomal nanocomposite dispersion according to the present invention is mixed according to the composition of the nanoemulsion formulation

Figure 5: Photograph showing the structural change according to the pH of the solution in the aqueous solution of the conventional lipid-cholesterol-based liposomes and polymer-liposomal nanocomposites according to the present invention

The present invention relates to the preparation of a polymer-liposomal nanocomposite using an acidity sensitive polymer and an external composition for skin containing the same, and specifically, using an acidity sensitive polymer composed of methacrylic acid and n-alkyl methacrylate, lipids and cholesterol. As a manufactured polymer-liposomal nanocomposite or nanoemulsion, it has a sensitive sensitivity to changes in acidity, which effectively releases physiologically active ingredients incorporated into the inner phase in vivo, thereby enhancing the effect as a transporter and preparing based on conventional lipids. The present invention relates to a polymer-lipid-based drug carrier and an external dermal composition designed to be more stably present in an aqueous solution and various salt-containing formulations and skin external preparation compositions and cosmetic formulations.

Recently, as the usefulness of the drug delivery system using liposomes has emerged, researches for producing more effective lipid-based nanocarriers by mixing various lipid components have been attracting attention. In particular, in order to ensure that the physiologically active ingredient incorporated into the liposome inner phase can be delivered to a living cell with high efficiency to maximize the efficacy of the active ingredient, studies are being actively conducted to prepare various components of the liposome. For example, to prepare liposomes containing lipids into which biocompatible polymers have been introduced to increase the in vivo circulation time to enhance the bioavailability of active ingredients, or to produce peptides or drugs of large molecular weight such as proteins or genes to target cells. Liposomes containing lipids into which cell recognition or cell adhesion molecules are introduced for delivery, or liposomes designed to release the bioactive components embedded in liposomes into cells by degrading the stability of liposomes by the specific environment of intracellular organelles. Research is ongoing.

In particular, various systems have been attempted to release physiologically active components embedded in the internal organs in response to changes in the acidity environment of intracellular organelles. For example, dioleolylphosphatidylethanolamine (hereinafter referred to as “DOPE”) vesicles stabilized by the introduction of a pH-sensitive co-surfactant has been extensively studied (Chu). et al., J. Liposome Res., 1994, 4, 361), the actual delivery of vesicles to the cytoplasm of various cells.

The research on the polymer-liposomal nanocomposite, in which the polymer surrounds the liposome, has been studied for the purpose of varying the function of the liposome. In other words, the acid-sensitive polymer was used to change the structure of liposomes to control the amount of drug contained in liposomes according to the environment of the solution. Earlier studies were conducted by Tirrell and co-workers on liposome complexes of poly (ethyl acrylic acid) and phosphatidylcholine. Some changing systems have been developed and proposed (Tirrell et al ., Macromolecules, 1984, 17, 1692). In recent research, a polymer containing a small amount (less than 10 mol%) of long alkyl chains was synthesized by using a polymer having a flexible chain structure such as polyoxyethylene as a main component of liposomes to maintain a stable structure. It is designed to incorporate aliphatic chains into liposomes when dispersed in an aqueous phase, such as liposomes. When the polymer containing acrylic acid is the main component of the system, a double membrane structure of liposomes is partially opened according to the acidity of the solution by adding a monomer having another flexible chain structure (Francis et al., Biomacromolecules) . 2001, 2, 741). However, when the liposomes were prepared using the above polymer, the particle size was observed to increase with time, and after 3 months (pH7), the size was reported to increase by about 30% or more.

To date, the literature reports that polymers containing acrylic acid alone and partially containing aliphatic chains cannot stabilize liposomes due to the high rigidity of acrylic acid in aqueous solution (Hwang et al, Langmuir, 2001, 17, 7713). Other papers report that stability of liposomes using polymers containing monomers containing acrylic acid and aliphatic chains is poor (Vial et al., Langmuir, 2005, 21, 853).

In fact, in order to maximize efficacy by incorporating agonistic components into cells, they maintain a stable structure over a long period of time in a living environment or material storage condition, and then reach a pH of 4.5 to 5.5, which is the pH of ribosomes in cells. The drug is not released slowly, but rather the structure is completely disintegrated and it is advantageous to release a large amount of drug in the cell.

Therefore, the polymer-liposomal nanocomposite has a drug release function due to structural collapse, which is the same function as the acid-sensitive liposome based on lipid-cholesterol, in order to improve the delivery efficiency of the active ingredient and cosmetic formulation. It can be said that it is very important to have a long-term structural stability of the liposome nanocomposite as well as having a system having superior stability than the conventional liposomes.

In general, liposomes based on lipid-cholesterol are also known to be unable to secure long-term stability in the water phase, and are particularly vulnerable to various salts used in the biological environment. In addition, in order for the above compositions to be used in cosmetics and external skin preparations, the stability in the formulation should be guaranteed, but there is a problem in that the structure is easily lost due to various surfactants in the formulation. Therefore, there is an urgent need to develop a new liposome system that can maximize the efficacy of the active ingredient in the body and at the same time stably exist in various biological environments and cosmetic and dermatological formulations.

Therefore, the present inventors have conducted research for the purpose of developing a system that can stably exist in various in vivo environments and cosmetic and external skin preparations while also having a function to improve the delivery and release effect of the existing liposomes. In case of preparing polymer-liposomal nanocomposites using acid-sensitive polymers with specific composition and composition, the drug in the body maintains the same structure of liposomes based on existing lipids and shows sensitivity to specific changes in acidity environment. Not only can increase the delivery efficiency, but also enhance the skin absorption of bioactive substances, maintain the stability of organs in the water phase and stably exist for various salts present in the water phase, as well as in the formulation of cosmetics and external skin preparations It has been found to be stable and the present invention has been completed. It is meaningful that the present invention meets the requirement of maintaining the stability of particles under any storage conditions before use, while inducing the increase of the efficiency of the active ingredient due to rapid structural collapse at a certain acidity which has not been solved by the prior art. In addition, the present invention has proved to have much improved stability for various salts and cosmetic formulations than liposome systems based on lipid-cholesterol, which are widely known.

The present invention relates to an acidity sensitive polymer composed of two monomers of methacrylic acid and n-alkyl methacrylate, and a polymer-liposome nanocomposite composed of lipids and cholesterol, and a polymer thereof. It is an object to provide a manufacturing method.

Another object of the present invention has a particle size of 50 ~ 300nm according to the composition of the polymer and the concentration of the polymer and lipid used, the shape of the particles to maintain the same form as the lipid-based liposomes do not contain the polymer, the hydrophobicity of the polymer In part, the present invention provides a polymer-liposomal nanocomposite having a structure associated with a lipid or a lipid / cholesterol-based bilayer.

It is another object of the present invention to move to a target organ in a state in which the active ingredient is stably infiltrated into the internal organ, and in response to specific changes in the acidic environment of the target organ, disrupting the lipid membrane and releasing a physiologically active ingredient to enhance the effect of the carrier. The present invention provides a polymer-liposomal nanocomposite containing an acidic sensitive polymer that not only provides the maximum efficacy of the active ingredient but also enhances skin absorption of a bioactive substance.

Still another object of the present invention is to bind together with a lipid or lipid / cholesterol bilayer and to bind the bilayer firmly, and to protect the outer wall to destabilize various liposome structures present in the water phase, such as salts or surfactants. The present invention provides a polymer-liposomal nanocomposite containing a polymer that plays a role of stably maintaining the structure of the liposomes from the cosmetics, and thus serves to stably maintain the structure of the cosmetic and skin external preparations.

The present invention relates to a polymer-liposomal nanocomposite using an acidity sensitive polymer and a method for preparing the polymer-liposome nanocomposite, in addition to a lipid or lipid-cholesterol component, methacrylic acid of formula 1 and n-alkyl methacrylate An acid-sensitive polymer composed of two monomers of n-alkyl methacrylate is included, and the inner phase contains a bioactive component and is dispersed in a mixed solvent of water and an organic solvent that can be mixed with water.

[Formula 1]

Hereinafter, the present invention will be described in more detail.

The polymer used in the polymer-liposomal nanocomposite of the present invention is a methacrylic acid monomer introduced to impart acidity sensitivity and an n-alkyl methacrylate monomer introduced to associate with a lipid bilayer. It is composed of acid-sensitive polymers.

Therefore, the polymer-liposomal nanocomposite containing the polymer exhibits sensitivity to acidity, thereby lowering structural stability in the region of acidity below 5.0, and disintegrating at lower acidity, thereby releasing physiologically active ingredients incorporated into the inner phase. As a result, the polymer-liposomal nanocomposite can be applied to the external preparation for skin, which is excellent in the skin absorbency when delivering the bioactive material using the polymer-liposomal nanocomposite as well as the internal wound according to the change in acidity when penetrated into the skin. Since it is possible to effectively release the physiologically active ingredient contained in the cell, it is possible to prepare a skin external preparation that can maximize the efficacy of the physiologically active ingredient.

In general, polymers synthesized with methacrylic acid are known to cause structural changes according to pH change in aqueous solution. The pH sensitivity range and degree are determined by the number of alkyl chains and the molecular weight of the polymer and the molecular weight of the polymer. It is known that it depends on the polydispersity index, which is an index representing a distribution.

The n-alkyl methacrylate has a range of alkyl chains that can be associated according to the type of lipid to which it is associated. Generally, n-alkyl methacrylate has a lipid or lipid-cholesterol bilayer when the length of n> 7, i.e., octyl or more. Association is possible, preferably n = 11-21, and depending on the purpose, a monomer having a cholesterol component may be introduced instead of the n-alkyl chain without using separate cholesterol in the lipid bilayer.

The polymer composed of the methacrylic acid monomer and the n-alkyl methacrylate monomer is prepared by polymerization by a general free radical thermal initiation method, and may also use anionic or cationic polymerization to control molecular weight distribution. . The mixing ratio of methacrylic acid (or acrylic acid, x of formula 1) and n-alkyl methacrylate (or cholesterol substituted methacrylate, y of formula 1) used in the polymer polymerization is 90: 10 to 50: 50, Preferably 85: 15-60: 40 is suitable. If the molar ratio occupied by n-alkyl methacrylate in the nanocomposite composed of methacrylic acid and n-alkyl methacrylate is less than 10%, the polymer may not only partially exist in the water phase but also serve as a surfactant. There is a possibility that, if it exceeds 50%, the hydrophobic portion of the polymer is too high, and as a result, there is a possibility that it acts as an unstable factor of the structure of the liposome when preparing the polymer-liposomal nanocomposite.

The molecular weight of a polymer composed of a methacrylic acid monomer and an n-alkyl methacrylate monomer affects the particle size of the polymer-liposomal nanocomposite, the structural stability of the polymer-liposomal nanocomposite, and in particular, the long-term structural stability. The molecular weight that can be used in the composite is in the range of 5,000 to 200,000 based on the number average molecular weight, preferably 10,000 to 100,000.

In addition, as the lipid component used to prepare the polymer-liposomal nanocomposites of the present invention, phospholipids or nitrogenous lipids having fatty acid chains having 12 to 24 carbon atoms may be used. It is generally more preferable to use phospholipids, for example, egg yolk lecithin (phosphatidylcholine), soybean lecithin, lysolecithin, sphingomyelin, phosphatidine acid, phosphatidylserine, phosphatidylglycerol, phosphatidyl inositol Natural phospholipids selected from phosphatidylethanolamine, diphosphatidylglycerol, cardiolipin, plasmogen; Hydrogenated products of natural phospholipids; Dicetylophosphate, distearoylphosphatidylcholine, dioleoylphosphatidylethanolamine, dipalmitoylphosphatidylcholine, dipalmitoylphosphatidylethanolamine, dipalmitoylphosphatidylserine, eleostaroylphosphatidylcholine, eleostaloylolamine, Synthetic lipids of osteoaroylphosphatidylserine; Derivatives of synthetic lipids; And one or two or more mixtures selected from fatty acid mixtures obtained by hydrolysis of synthetic lipids.

When two kinds of phospholipids are used in combination, for example, a combination of phosphatidylcholine: phosphatidylethanolamine, phosphatidylcholine: phosphatidylglycerol, phosphatidyl choline: phosphatidyl inositol, phosphatidylcholine: phosphatidine acid or phosphatidylcholine: diole oil phosphatidylethanolamine, etc. Can be used as Although the mixing ratio varies depending on the component composition to be mixed, the mixing ratio of the maximum component to the minimum component is preferably 1: 5 or less. For example, in the case of phosphatidylcholine: dioleoylphosphatidylethanolamine, 1: 1, 2: 1, 3: 1, 4: 1, 5: 1, 1 in the molar ratio of 5: 1 to 1: 5, respectively. It may be mixed in various ways, such as: 5, 1: 4, 1: 3, or 1: 2. In addition, when a mixture of three phospholipids is used, for example, phosphatidylcholine: dioleoylphosphatidyl ethanolamine: phosphatidylserine in a molar ratio range of 1: 5, respectively, 1: 1: 1, 2: 1: 1, It can be mixed and used in various mixing ratios such as 3: 1: 2, 3: 2: 1, 3: 2: 2, 4: 1: 1, or 4: 2: 1.

The cholesterol-liposome nanocomposites according to the present invention may be further mixed with cholesterol as needed, and the addition of cholesterol may interfere with the self-association of the lipid-polymer bilayer but improve the curvature of the liposomes to help form a stable spherical liposome. Will be given.

The physiologically active ingredient incorporated into the polymer-liposomal nanocomposite inner phase of the present invention is mainly water-soluble, and known as a whitening substance, N-butyldeoxynojirimycin, N-butyldeoxynojirimycin, 1-dioxynojirimycin, and castanospermine ( castanospermin, actinomycetes culture extract (SCE), calcium pentatheine sulfonate, arbutin, ascorbic acid, ethyl ascorbyl ether, ascorbyl phosphate sodium One or more whitening products selected from vitamin C derivatives such as ascorbyl phosphate, a-ketoglutaric acid and epigallocatechin gallate (EGCG), known as anti-wrinkle substances. Antioxidants and anti-wrinkle materials can be used. In addition, the amount of physiologically active ingredient incorporated into the polymer-liposomal nanocomposite phase of the present invention may be used in an amount of 0.1 to 10% based on the total weight of the polymer-liposomal nanocomposite solution including water, and preferably 1 to 5%.

The ratio of the poly (methacrylic acid-co-n-alkyl methacrylate) copolymer, lipids and cholesterol, which is an acidity sensitive polymer used to maintain the acidity sensitivity and structural stability of the polymer-liposomal nanocomposite according to the present invention, is poly ( 1-50% by weight of poly (methacrylic acid-co-n-alkyl methacrylate) copolymer, preferably based on the total weight of the methacrylic acid-co-n-alkyl methacrylate) copolymer, lipids and cholesterol 5-30% by weight, 50-99% by weight lipids, preferably 50-90% by weight, and 0-30% by weight cholesterol, preferably 10-25% by weight.

In general, liposomes are prepared using ultrasonic waves or using a high pressure emulsifier, and the polymer-liposomal nanocomposite according to the present invention uses a high pressure emulsifier.

a) heating and dissolving the poly (methacrylic acid-co-n-alkyl methacrylate) copolymer and the lipid of Formula 1 in an organic solvent that can be mixed with water to 50-70 ° C .;

b) 50 to 70 of the mixed solution of a) Mixing with warmed water to < RTI ID = 0.0 > C < / RTI >

c) nano-particles of the mixed solution of b) with a high pressure emulsifier.

[Formula 1]

Cholesterol may be further mixed in step a), and the organic solvent that can be mixed with water used is not limited, but is preferably selected from ethanol, methanol, isopropyl alcohol, acetone and tetrahydrofuran. In addition, the water heated in step b) may further contain a water-soluble active ingredient. Step C) of the nanoparticles is a high pressure emulsifier, and the pressure can be adjusted to 100 to 1000 bar according to the desired particle size and dispersion degree, preferably 500 to 1000 bar.

The method may further include removing the residual organic solvent using a rotary evaporator after step c) according to the organic solvent used.

The polymer-liposomal nanocomposite of the present invention obtained by the above-described manufacturing method has a particle size of 50-300 nm according to the concentration of the polymer and the lipid, and maintains the same structure and form as the lipid-based liposome without the polymer. As shown in FIG. 1, the hydrophobic portion of the polymer not only has a structure associated with lipids or lipid / cholesterol-based bilayers, but also has a sensitive sensitivity to changes in acidity, thereby effectively releasing a biologically active component incorporated into the inner stomach. This can enhance the effect as a carrier.

The external preparation composition for skin containing the polymer-liposomal nanocomposite of the present invention is not particularly limited in its formulation, and it is softening longevity, astringent longevity, nourishing longevity, nourishing cream, massage cream, eye cream, eye essence, essence, Cleansing cream, cleansing lotion, cleansing foam, cleansing water, pack, powder, makeup base, foundation, body lotion, body cream, body oil, body essence, body cleanser, hair dye, shampoo, rinse, toothpaste, mouthwash, hair conditioner , Wool, lotions, ointments, gels, creams, patches and sprays and the like.

Hereinafter, the present invention will be described in more detail with reference to Examples and Experimental Examples, but the present invention is not limited only to these examples.

Example 1-17 Preparation of Polymer-Liposome Nanocomposite Using Acid Sensitive Polymer

Poly (methacrylic acid-co-n-alkyl methacrylate) copolymer, 100% hydrogenated oleyl-palmitoyl / oleylyl-stearyl phosphatidylcholine mixture (Lipoid S100-3) and cholesterol are shown in Table 1 below. The mixture was prepared by heating and dissolving it in ethanol at 60 ° C. using the weight composition ratio, and added to 60 ° C. water and stirred for 5 minutes at 5000 rpm using an homogenizer to prepare the first dispersed polymer-liposome complex. After the preparation was nanoparticles using a high pressure homogenizer (1000bar, 3cycle). The residual ethanol solution was removed using a rotary evaporator to prepare a polymer-liposomal nanocomposite using an acid sensitive polymer containing the active ingredient.

TABLE 1

Example 18-23 Preparation of Polymer-Liposome Nanocomposites Containing Active Ingredients

Poly (methacrylic acid-co-n-alkyl methacrylate) copolymer (mol composition 70:30, number average molecular weight 26,000), 100% hydrogenated oleyl-palmitoyl / oleylyl-stearyl phosphatidylcholine mixture (Lipoid S100-3), cholesterol and water-soluble active ingredients were dissolved in ethanol using the weight composition ratio shown in Table 2 below, and then the mixed solution was heated to 60 ℃ aqueous solution of the active ingredient dissolved in 300g of water. The mixture was added to an active ingredient solution heated to 60 ° C., mixed and stirred for 5 minutes at 5000 rpm using an emulsifier to prepare a first dispersed polymer-liposomal complex, followed by a high pressure homogenizer (1000 bar, 3 cycle) to nanoparticles. The residual ethanol solution was removed using a rotary evaporator to prepare a polymer-liposomal nanocomposite containing the active ingredient.

TABLE 2

Comparative Example 1-4 Preparation of Liposomes Comprised of Lipid-Cholesterol

100% hydrogenated oleorel-palmitoyl / oleoreyl-stearyl phosphatidyl choline mixture (Lipoid S100-3), cholesterol and water-soluble active ingredient using a weight ratio of the composition shown in Table 3 after warming and dissolving in ethanol After the mixture was prepared, the mixture was added to water at 60 ° C., mixed and stirred for 5 minutes at a speed of 5000 rpm with an homogenizer, and then nanoparticles were formed using a high pressure homogenizer (1000 bar, 3 cycle). . Liposome consisting of lipid-cholesterol was prepared by removing residual ethanol solution using a rotary evaporator.

TABLE 3

Experimental Example 1 Structural Analysis of Polymer-Liposome Nanocomposites and Liposomes

Of the polymer-liposomal nanocomposite using the acidity-sensitive polymer of Example 1-17, the polymer-liposomal nanocomposite containing the water-soluble active ingredient of Example 18-23 and the lipid and lipid-cholesterol-based liposomes of Comparative Example 1-4 The nanoemulsion particle size was measured using a dynamic light scattering apparatus (Zetasizer 3000HSa Malvern Instruments). The scattering angle was fixed at 90 ° and the temperature was maintained at 25 ° C., and the average particle diameter was calculated from the Stokes-Einstein equation, and the results are shown in Table 4.

TABLE 4

As shown in Table 4, it can be seen that the particle size of the polymer-liposomal complex including the acidity sensitive polymer of Example 1-23 is larger than that of the liposome of Comparative Example 1-4, which does not contain the acidity sensitive polymer. The particle size of the polymer-liposomal composite of Example 1-23 was increased as the polymer content was increased.The particle size of the polymer-liposomal nanocomposites of Examples 6 and 7 was 150 nm and 80 nm, respectively. It can be seen that the size depends on the concentration of lipids, cholesterol and polymers contained in ethanol.

In addition, although a large change in particle size was not observed with increasing molecular weight of the poly (methacrylic acid-co-n-alkyl methacrylate) copolymer, which is an acidic polymer, the polymer-liposome nanocomposite of Example 9 having a number average molecular weight of 68,000 In the case of, the particle size was observed to increase slightly to 300 nm compared to the case of Examples 4 and 8 using molecular weights of 17,000 and 26,000.

Particularly, in Comparative Example 1, in which a liposome is prepared using only a lipid single component, precipitation does not occur, whereas liposomes are not prepared. On the other hand, when the polymer is properly mixed with a single lipid component, the particles are successfully formed. This could be observed because the polymer plays a stable role in the formation of lipid bilayers.

In addition, the particle size change according to the length (n) of the alkyl chain of the n-alkyl methacrylate of the acidity sensitive polymer used in Examples 1-17 was carried out in the case of lauryl (n = 11) of Example 10. The stearyl (n = 17) of Examples 1-9 and 14-17 showed similar particle size, whereas the octyl of Example 11 (n = 7) greatly increased the particle size. In the case of butyl (n = 3), particles did not form and precipitated. Also similar to the lauryl (n = 11) of Example 10 and stearyl (n = 17) of Examples 1-9 and 14-17 for Example 13 using cholesterol instead of n-alkyl methacrylate The particle size is shown.

In addition, the precipitation occurs in Examples 15-17, so that the appropriate methacrylic acid and stearyl methacrylate to form particles from the polymer-liposomal nanocomposites containing different molar ratios of methacrylic acid and stearyl methacrylate. The molar ratio can confirm that the molar ratio occupied by stearyl methacrylate in the nanocomposite composed of methacrylic acid and stearyl methacrylate is 10% or more and 50% or less, the poly (methacrylic acid-co-n- Alkyl Methacrylate) The molar ratio of methacrylic acid and stearyl methacrylate constituting the copolymer and the length (n) of the alkyl chain of n-alkyl methacrylate have a great influence on the particle formation of the polymer-liposomal nanocomposite. I could see that.

In the case of the polymer-liposomal nanocomposite of Example 18-23 containing a water-soluble active ingredient, the change in particle size by the kind of active ingredient was hardly observed within the same composition, and Example 1- containing no active ingredient It showed almost the same particle size as the polymer-liposomal nanocomposite of 17.

The thermal properties of the polymer-liposomal nanocomposite of Example 5 and the liposome of Comparative Example 4 were measured by a microcalorimeter, and the results are shown in FIG. 2. The liposome of Comparative Example 4 showed a gel-liquid crystal transition point at around 50 ° C., whereas the polymer-liposomal complex of Example 5 had a gel-liquid crystal transition point at around 43 ° C. The gel-liquid crystal transition point in the polymer-liposomal nanocomposite is that it maintains the self-association structure, which is a unique property of liposomes, and therefore, the components of the acid-sensitive polymer of the polymer-liposomal nanocomposite prepared according to the present invention. Since the aliphatic chain of instearyl methacrylate is self-associating like a lipid, as shown in FIG. 1, an acid sensitive polymer used in a polymer-liposomal nanocomposite is associated with a lipid-cholesterol to induce liposome intrinsic. It was found that it surrounds the outer wall of liposomes while maintaining its properties.

Experimental Example 2 Evaluation of Stability of Polymer-Liposome Nanocomposites

The solution prepared for observing the long-term stability of lipid-cholesterol-based liposomes of 2 and 3 compared to the polymer-liposomal nanocomposites of Examples 1, 4, 10, 11 and 19 was a low temperature (2 ° C.) and room temperature (25 The change in particle size with time was stored at each ℃ and measured using a dynamic light scattering device (Dynamic light scattering, Zetasizer 3000HSa Malvern Instruments) is shown in Table 5 below.

TABLE 5

As shown in Table 5, in the case of the polymer-liposomal nanocomposite using the poly (methacrylic acid-co-n-alkyl methacrylate) copolymer, the length of the alkyl chain used in the n-alkyl methacrylate was prepared. It was found that the particle stability of the polymer-liposomal nanocomposite varies. In the case of octyl (n = 7) of Example 11, the particle size was 420 nm to form large particles, but precipitates formed after 3 weeks of storage at low temperature and room temperature. It was very bad.

In addition, in the case of the polymer liposome nanocomposites of Examples 1, 4, and 10 using stearyl (n = 17) and lauryl (n = 11) methacrylate as monomers, as shown in Table 5, not only 8 weeks but also 12 months It was found that the particle size did not increase significantly even when stored at low temperature and room temperature. This can be said that when the alkyl chain is short in length, as described in the prior art, the addition of the polymer acts as an element that destabilizes the structure of the liposome, and the polymer used to prepare a highly stable polymer-liposomal nanocomposite It has been found that they have a specific composition and specific solution properties of the polymer in aqueous solution.

In comparison of the stability of the lipid-cholesterol-based liposome of Comparative Example 3 containing the actinomycete extract and the polymer-liposomal nanocomposite of Example 19, in Comparative Example 3, the particle size gradually increased at low temperature and room temperature. After 8 weeks, some precipitates occurred, but the particle size of the Example 19 containing the same amount of actinomycetes extract almost did not change during 8 weeks. Therefore, it was observed that the polymer-liposomal nanocomposite according to the present invention maintains a longer-term stable structure than the liposomes based on lipid-cholesterol when the active ingredient is included.

Salts of the polymer-liposomal nanocomposites of Example 4 and the lipid-cholesterol-based liposomes of Comparative Example 2 in sodium phosphate buffer and HEPES (N-2-hydroxyethyl-piperazine-N'-2-ethan sulfonate) buffer The results of observing the long-term stability according to the addition of (salt) is shown in Table 6 below.

TABLE 6

As shown in Table 6, in the case of Example 4, the polymer-liposomal nanocomposite according to the present invention has a long-term difference in particle size and polydispersity regardless of the case in which the distilled water and the salt are stored in the buffer solution. I didn't see it. In contrast, Comparative Example 2, which is a lipid-cholesterol-based liposome, has an effect on particle size and polydispersity when stored in a buffer solution containing salt, compared to the case of storage in distilled water, and thus sodium phosphate and sodium chloride (NaCl). In the sodium phosphate buffer solution with), the polydispersity increased with increasing the particle size and the structure became unstable.In addition, the structure collapsed after 2 weeks when stored in the HEPES buffer solution. A precipitate could be observed.

Therefore, it can be seen from Table 5 and Table 6 that the polymer-liposomal nanocomposites according to the present invention maintain a much more stable structure in the presence of water-soluble active ingredients and other salts than liposomes based on lipid-cholesterol.

In order to compare the structural stability in the cosmetic formulation of liposomes based on the polymer-liposomal nanocomposite and lipid-cholesterol according to the present invention, the solutions prepared in Example 4 and Comparative Example 2 were prepared according to the composition and content of Table 7 below. Into the formulated nanoemulsion formulation having an average particle size of 144nm at 10 ℃, 30% and 50% by weight at 60 ℃ stirred for 5 minutes at a speed of 7200rpm with a homogenizer and then cooled and degassed After the process, the formulation containing the vesicles was prepared. The prepared formulation-vesicle mixed solution was observed using a differential scanning calorimetry (Differential scanning calorimetry, DSC Q1000, TA Instruments) to observe the change in calories according to temperature and the results are shown in FIGS. 3 and 4.

TABLE 7

FIG. 3 shows the change of calorie according to temperature when liposomes based on lipid-cholesterol of Comparative Example 2 were contained in various amounts in the nanoemulsion formulation. The liposomes showed a gel-liquid crystal transition point near 51 ° C. In case of nanoemulsion, the transition phenomenon of emulsion droplet was observed around 62 ° C. However, in the case of the liposome-nanoemulsion mixture, it was observed that the transition point to be released from the liposome solution and the nanoemulsion formulation disappeared, and that one transition point occurred, and the transition point was changed as the composition was changed. . If the vesicle and nanoemulsion fish sauce are stably and independently present in the mixed liquor, the transition point to be drawn from the vesicle solution and nanoemulsion formulation will have to come out with only a peak size ratio at each transition point. However, only one transition point from FIG. 3 means that new particles are reconstituted as the liposomes and nanoemulsions are mixed, and when mixed with the nanoemulsions, the structure of the initial lipid-cholesterol-based liposomes is no longer present in the formulation. I could see that it doesn't exist.

Figure 4 shows the change of calorie according to the temperature of the polymer-liposomal nanocomposite of Example 4 polymer-liposomal nanocomposite showed a gel-liquid crystal transition point at 45 ℃, mixed polymer-liposomal nanocomposite and nanoemulsion In the case of the polymer-liposomal nanocomposite, the gel-liquid crystal transition point and the transition point caused by the fish sauce of the nanoemulsion were maintained as it is, indicating two transition points. This is because, unlike the case of liposome-nanoemulsion, when the polymer-liposome nanocomposite and the nanoemulsion emulsion are mixed and stirred, the polymer-liposome nanocomposite and the nanoemulsion are uniformly maintained while maintaining the unique structure of each of the polymer-liposome nanocomposite and the nanoemulsion.

Therefore, the polymer-liposomal nanocomposite according to the present invention has solved the storage stability problem of the conventional polymer-liposomal complex, and compared to the conventional lipid-cholesterol-based liposomes, water-soluble active ingredients, salts and surfactants. It can be seen that it is maintaining a very stable structure in the cosmetic formulation containing.

Experimental Example 3 Observation of Acidity Sensitivity of Polymer-Liposome Nanocomposites

In order to observe the acidity sensitivity of the polymer-liposomal nanocomposites according to the present invention, the liposomes of the polymer-liposomal nanocomposites of Example 1 and 4 and the comparative example 2 were converted into pH 3, 4, 5, 6, and 7.4 with a hydrochloric acid solution. The result of observing the state of the solution by diluting at a concentration of 0.1% by weight in an appropriate sodium phosphate buffer solution is shown in Figure 5, the change in particle size according to the acidity of the solution is shown in Table 8 It was. 5, the liposomes composed of lipid-cholesterol of Comparative Example 2 were well dispersed in the aqueous solution regardless of the acidity of the solution, but the polymer-liposomal nanocomposites of Examples 1 and 4 according to the present invention had a pH of 5 or more. The solution was well dispersed in the aqueous phase, but in aqueous solutions of pH 4 and 3, it was observed that it was no longer stable and collapsed.

In addition, as shown in Table 8, when the polymer-liposomal nanocomposites of Examples 1 and 4 are dispersed in a solution of pH 7.4 ~ 5 when the dispersion in distilled water and the variation in particle size and polydispersity is not large It has a pH sensitivity that the structure is rapidly collapsed at the lower pH, which reflects the property that the acid-sensitive polymer used in the polymer-liposomal nanocomposite rapidly changes its shape in the aqueous phase based on the specific pH. I think.

On the other hand, the liposome of Comparative Example 2 seems to be well dispersed regardless of the acidity of the solution in Figure 5, the particle size and polydispersity as the acidity of the solution is different from that dispersed in distilled water as shown in Table 8 below I could see it getting very large.

Therefore, the polymer-liposomal nanocomposite according to the present invention is stably maintained in the particle form at pH 5 or above, but the pH of the polymer-liposomal nanocomposite is rapidly lowered at a stability of the polymer-liposomal nanocomposite so that the biologically active ingredient contained therein is in the cell. It can be confirmed that it can be effectively released in the interior to enhance the delivery effect as a transporter.

Table 8 Results of Particle Size Measurement According to Acidity of Polymer-Liposome Nanocomposites and Lipid-Cholesterol-Based Liposomes

Experimental Example 4 Observation of Drug Release Behavior According to Acidity of Polymer-Liposome Nanocomposites

In order to enhance the intracellular efficiency of the water-soluble active ingredient, the carrier should be used to increase the rate of incorporation of the active ingredient into the cell, and at the same time, the carrier must exhibit a function of releasing drugs in a specific environment within the cell. The intracellular ribosome is maintained at a pH lower than the pH seen in normal physiological conditions (pH 4.5 ~ 5.5) and is water-soluble only if it has the ability to release the drug in response to such intracellular changes. It can enhance the intracellular delivery efficiency of the active ingredient. Therefore, in order to observe the selective release of the water-soluble drug as the acidity of the aqueous solution containing the polymer-liposomal nanocomposite according to the present invention was tested in the following way.

0.455 g of 8-hydroxypyrene-1,3,6-trisulfonic acid trisodium salt (hereinafter referred to as “HPTS”) and 0.525 g of para -Xylene-bis-pyridium bromide (hereinafter referred to as "DPX") was dissolved in 300 ml of an aqueous solution containing 10 mM HEPES and 11 mM sodium phosphate.

0.6 g of poly (methacrylic acid-co-stearyl methacrylate) (mol composition 70:30, number average molecular weight 26,000) and 100% hydrogenated oleyl-palmitoyl / oleyl-stearyl phosphatidyl choline mixture (S100) -3) 1.2 g and 0.15 g of cholesterol were warmed and dissolved in 30 g of ethanol, and then added to a buffer solution containing HPTS-DPX, which was heated to 65 ° C, and mixed at a speed of 5000 rpm using a homogenizer for 5 minutes. . The first dispersed polymer-liposomal complex was pulverized into a nanoparticle by using a high pressure homogenizer (high pressure homogenizer; 1000 bar, 3 cycle) to make a polymer-liposomal nanocomposite containing HPTS-DPX. Particle size is shown.

Meanwhile, the polymer-liposome nano after dissolving 1.8 g of oleorel-palmitoyl / oleoreyl-stearyl phosphatidyl choline mixture (S100-3) and 0.2 g of cholesterol in 30 g of ethanol was compared to the comparative group. Liposomes containing HPTS-DPX were prepared in the same manner as the complex, and showed a particle size of 200 nm.

200 ml of the polymer-liposomal nanocomposite and the lipid-cholesterol-based liposome solution containing HPTS-DPX contained 20 ml of 20 mM HEPES buffer solution (pH 7.1, 144 mM sodium chloride) and 100 mM MES buffer solution (pH 3.45, 144 mM sodium chloride). ) Into 20 ml and dispersed. 2 ml of the dispersion solution was encapsulated in a dialysis tube and then inserted into a 30 ml isotonic solution (pH 7.1, 3.45) having the same pH, and then placed in a constant temperature stirrer to detect the solution according to time, followed by fluorescence spectrometer. (Flourescence spectroscopy, Hitachi 4500) The amount of HPTS-DPX in the detection solution was detected and shown in Table 9 below. On the other hand, the total amount of HPTS-DPX present in the isotonic solution was added to the polymer-liposomal nanocomposite and liposomes in 10% Triton X-100 solution to completely disintegrate the structure and detected by a fluorescence spectrometer (Flourescence spectroscopy, Hitachi F-4500). .

[Table 9] Changes in HPTS-DPX Release Rate with Changes in Acidity of Polymer-Liposome Nanocomposites and Lipid-Cholesterol Liposomes

Table 9 shows that both the polymer-liposomal nanocomposite and the lipid-cholesterol liposome have higher drug release at pH 3.45 than at pH 7.1. As shown in Table 8, the pH of the solution is low. It can be said that this is due to the increase in particle size and structural collapse of the polymer-liposomal nanocomposite and lipid-cholesterol liposome.

In addition, the effect of increasing the drug release as the pH of the solution is lowered from 7.1 to 3.45 from Table 9 was observed that the polymer-liposomal nanocomposite is much larger than the lipid-cholesterol liposome, which is the polymer-liposomes at pH 5 or less While the structure of the nanocomposite is completely collapsed and HPTS-DPX can be released freely, lipid-cholesterol liposomes are due to the difference that only a slight structure is unstable and the complete release of HPTS-DPX is impossible.

Therefore, it can be seen that the polymer-liposomal nanocomposite according to the present invention has the ability to control and release the drug according to the change in acidity when the structure changes rapidly with pH change and contains a water-soluble drug. .

Experimental Example 5 Observation of Skin Transmittance of Polymer-Liposomal Nanocomposites Containing Active Ingredients

In order to evaluate the skin absorption ability of the polymer-liposomal nanocomposite according to the present invention, the skin permeability of the arbutin solution and the arbutin solution containing no carrier was measured.

The polymer-liposome nanocomposite solution containing 2% by weight of arbutin and 2% by weight of arbutin aqueous solution, respectively, were prepared. At the same time, the skin collected from the Guinea pig was mounted on Franz-cell (Hanson Research), a skin permeation testing device, and filled with the polymer-liposomal solution containing arbutin and the aqueous solution of arbutin, respectively. Using a high performance liquid chromatography (Hewlett Packard), the solution underneath the skin was taken every 6 hours to see how the polymer-liposomal solution containing arbutin and the aqueous solution of arbutin penetrated the skin. The concentration of arbutin is quantified and shown in Table 10 below.

[Table 10] Changes in Skin Permeation of Arbutin-Containing Polymer-Liposome Nanocomposites and Arbutin Solution over Time

As time passed from Table 10, the skin permeation rate of the polymer-liposome nanocomposite solution containing arbutin gradually increased compared to the arbutin aqueous solution, and after 18 hours, about 70% more arbutin contained the skin than the arbutin aqueous solution. The thing which permeated was observed. Therefore, it can be seen that the polymer-liposomal nanocomposite containing the active ingredient has an excellent skin permeation enhancement function as compared with the aqueous solution of the active ingredient containing no carrier.

Experimental Example 6 Intracellular Whitening Efficacy of Polymer-Liposomal Nanocomposites Containing Active Ingredients

In order to evaluate whether the polymer-liposomal nanocomposite containing the active ingredient can exert excellent intracellular efficacy compared to the aqueous solution of the active ingredient without the carrier, an actinomycete extract known to possess melanin production inhibitory ability was used. Polymer-liposomal nanocomposite containing the actinomycetes extract of Example 19 and the actinomycetes extract aqueous solution were maintained in the cell culture to maintain the total actinomycetes concentration in the medium in which Human Melanoma HM3KO cells were cultured. Injected. After culturing the culture medium containing the cells for 24 hours (incubation), the cells are removed, the cells are separated by centrifugation, and the remaining filtrate is collected, and the filtrate is UV-vis spectroscopy. The amount of melanin produced was quantified from the size of the absorption peaks generated at 490 nm using the following, and the ratios are shown in Table 11 below.

Table 11 Relative melanin production rate (%) compared to the control (100%) according to the concentration of the polymer-liposomal nanocomposite containing the aqueous actinomycetes extract and actinomycetes extract

As can be seen in Table 11, when the concentration of the actinomycetes extract of 0.5mM and 0.25mM, it was found that the melanin production inhibitory effect is excellent when the polymer-liposomal nanocomposite is collected, the concentration of actinomycetes of 0.5mM Even though it has excellent melanin production inhibitory effect compared to 1mM aqueous solution, the polymer-liposomal nanocomposite of 0.25mM actinomycete extract concentration was also superior to melanin production inhibitory effect compared to 0.5mM aqueous solution. Therefore, it was found that the case of using the polymer-liposomal nanocomposite containing the active ingredient was superior to the effect ingredient transfer ability to the cell than the case of not using the carrier.

As described above, the polymer-liposomal nanocomposite using the acidity-sensitive polymer according to the present invention is designed to enable controlled release of the drug as the acidity of the solution changes by using the characteristic of changing the structure according to the acidity of the aqueous solution. . In addition, the stability of the polymer-liposomal nanocomposite rapidly decreases at a certain acidity, thereby effectively releasing the physiologically active ingredients contained therein into the cell, thereby improving the incorporation rate and efficacy of the active ingredients. Absorbance is shown. In addition, the polymer-liposomal nanocomposite using the acidity sensitive polymer according to the present invention has much improved stability for various salts and cosmetic formulations than the liposome system based on lipid-cholesterol, which is widely known.

Claims (16)

A polymer-liposomal nanocomposite comprising a poly (methacrylic acid-co-n-alkyl methacrylate) copolymer and a lipid of Formula 1 below. [Formula 1]

The method of claim 1, Wherein said poly (methacrylic acid-co-n-alkyl methacrylate) copolymer and lipids are dispersed in a mixed solvent of water and an organic solvent that can be mixed with water. The method of claim 2, Polymer-liposomal nanocomposite, further comprising cholesterol. The method of claim 2, Polymer-liposomal nanocomposite, characterized in that the water-soluble active ingredient is embedded in the polymer nanocomposite. The method of claim 3, wherein Poly (methacrylic acid-co-n-alkyl methacrylate) copolymer, 1-50% by weight poly (methacrylic acid-co-n-alkyl methacrylate) copolymer, lipids based on total weight of lipids and cholesterol A polymer-liposomal nanocomposite comprising 50-99% by weight and 0-30% by weight of cholesterol. The method of claim 3 , A polymer-liposomal nanocomposite, wherein the poly (methacrylic acid-co-n-alkyl methacrylate) copolymer has a number average molecular weight of 5,000 to 200,000. The method of claim 1, The lipids include yolk lecithin (phosphatidylcholine), soybean lecithin, lysolecithin, sphingomyelin, phosphatidine acid, phosphatidylserine, phosphatidylglycerol, phosphatidyl inositol, and phosphatidyl having 12 to 24 fatty acid chains. Natural phospholipids selected from ethanolamine, diphosphatidylglycerol, cardiolipin, plasmogen; Hydrogenated products obtainable from natural phospholipids; Dicetylphosphate, distearoylphosphatidylcholine, dioleoylphosphatidylethanolamine, dipalmitoylphosphatidylcholine, dipalmitoylphosphatidylethanolamine, dipalmitoylphosphatidylserine, eleostaroylphosphatidylcholine, eleostearoylolamine and Synthetic lipids of eleostaroylphosphatidylserine; Derivatives of synthetic lipids; And fatty acid mixtures obtainable by hydrolysis of synthetic lipids. Polymer-liposomal nanocomposites, characterized in that one or two or more mixtures selected from the group consisting of. The method of claim 4, wherein Water-soluble active ingredients include N-butyldeoxynojirimycin, 1-deoxynojirimycin, castanospermin, actinomycetes culture extract (SCE), calcium pentatheine sulfonate , Arbutin, vitamin C (ascorbic acid), vitamin C derivatives of ethyl ascorbyl ether, a-ketoglutaric acid, epigallocatechin gallate , EGCG) polymer-liposome nanocomposite, characterized in that one or two or more kinds of mixed components. The method of claim 8, The polymer-liposomal nanocomposite is a polymer-liposomal nanocomposite, characterized in that to release the water-soluble active ingredient embedded in the inner phase at pH 5 or less. The method of claim 1, Polymer-liposomal nanocomposite, characterized in that the particle size of the polymer-liposomal nanocomposite is 50 ~ 300nm. a) heating and dissolving the poly (methacrylic acid-co-n-alkyl methacrylate) copolymer and the lipid of Formula 1 in an organic solvent that can be mixed with water to 50-70 ° C .; b) mixing the mixture of a) with water warmed to 50 to 70 ° C. and dispersing it first with an emulsifier; And c) nanoparticle-forming the mixture of b) with a high pressure emulsifier; Method for producing a polymer-liposomal nanocomposite using an acidity-sensitive polymer, characterized in that consisting of. [Formula 1]

The method of claim 11, Method for producing a polymer-liposomal nanocomposite using an acidity-sensitive polymer, characterized in that further mixing cholesterol in step a). The method of claim 11, Method for producing a polymer-liposomal nanocomposite using an acidity sensitive polymer, characterized in that the water-soluble active ingredient is contained in the step b). The method of claim 11, The method of preparing a polymer-liposomal nanocomposite using an acidity-sensitive polymer, characterized in that the organic solvent that can be mixed with water in step a) is selected from ethanol, methanol, isopropyl alcohol, acetone, tetrahydrofuran. An external preparation composition for skin, comprising the polymer-liposomal nanocomposite according to any one of claims 1 to 10. The method of claim 15, The composition is softening cream, astringent makeup, nourishing cream, nourishing cream, massage cream, eye cream, eye essence, essence, cleansing cream, cleansing lotion, cleansing foam, cleansing water, pack, powder, makeup base, foundation, body lotion , Body cream, body oil, body essence, body cleanser, hair dye, shampoo, rinse, toothpaste, mouthwash, hair dresser, wool, lotion, ointment, gel, cream, patch and spray The external preparation composition for skin.

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