KR102532697B1 - A method of preparing polymeric micro particles, polymeric micro particles, medical composition, cosmetic composition, medical articles and cosmetic articles using the same - Google Patents

Abstract

The present invention relates to a method for producing polymeric microparticles capable of realizing high crosslinking efficiency and production yield, as well as excellent mechanical strength and stability, polymeric microparticles, and a medical composition, cosmetic composition, medical product, and cosmetic product including the same.

Description

Method for producing polymeric microparticles, polymeric microparticles, medical composition containing the same, cosmetic composition, medical supplies and cosmetic supplies THE SAME}

The present invention relates to a method for producing polymeric microparticles capable of realizing high crosslinking efficiency and production yield, as well as excellent mechanical strength and stability, polymeric microparticles, and a medical composition, cosmetic composition, medical product, and cosmetic product including the same.

As the field of biopharmaceuticals and regenerative medicine expands, there is an increasing demand for cell mass culture technology capable of efficiently producing cells, tissues, microorganisms, and the like.

Adherent cells are cultured using microcarriers in a 3D bioreactor. Cells, culture medium, and microcarriers are placed in a bioreactor, and the culture medium is agitated to contact the cells and microcarriers, thereby attaching the cells to the surface of the microcarriers and culturing them. Since the microcarrier used at this time provides a high surface area ratio (surface area/volume) on which cells can attach and proliferate, it is suitable for mass culture of cells. However, when the adherent cells are expanded and cultured using microcarriers, a process of recovering the cells through a cell detachment process is necessarily accompanied after the culture is terminated. The cell detachment process induces cell detachment by using a proteolytic enzyme or by changing the temperature. When such a detachment process is added, the manufacturing cost increases, resulting in poor economic feasibility and cell damage. .

In order to solve this problem, the development of new materials or processes is steadily progressing. In particular, in the case of cell therapy products in which cells are injected in vivo, efforts are being made to omit the separation and purification process by securing the biocompatibility of microcarriers for culturing cells. there is. In this case, particles having strength capable of withstanding the stress applied by the fluid surrounding the carrier after the culture process and injecting into the body are required.

In addition, in the case of transdermal drug delivery technology in which a microcarrier containing a drug or physiologically active substance is loaded on a microneedle and delivered, a polymer suitable for application to the living body must be used for the microcarrier, and the stratum corneum of the skin It must have sufficient strength so that the deformation of the particles does not occur in the process of passing through the layer). The microcarrier stably penetrating the transdermally can deliver the loaded drug locally or systemic and act on the necessary lesion.

Hyaluronic acid, which is mainly used as a biocompatible material, is a biopolymer material composed of N-acetyl-D-glucosamine and D-glucuronic acid, and the repeating units are linearly connected. It is abundant in the vitreous humor of the eyeball, synovial fluid of the joint, and chicken comb. Hyaluronic acid is commonly used as a bioinjectable material due to its excellent biocompatibility and viscoelasticity, but its use is limited by itself because it is easily decomposed in vivo or under conditions such as acids and alkalis. In addition, when applied to microcarriers, there was a problem in that cell adhesion was remarkably lowered due to surface negative charge in the biological pH range.

On the other hand, gelatin is a polymer obtained by hydrolyzing collagen, which is a biological connective tissue, and is widely used as a scaffold for cell culture. Cells can be collected or cultured using the cell adhesion performance of gelatin, but its strength is weak and it has phase transition characteristics according to temperature, so there are studies to improve physical properties by introducing functional groups using chemical methods.

Accordingly, there is a need to develop microcarriers or polymeric microparticles that are biocompatible and have excellent physical properties such as physical strength and stability against heat and enzymes.

An object of the present invention is to provide a method for preparing polymeric microparticles capable of realizing excellent mechanical strength and stability as well as high crosslinking efficiency and production yield.

In addition, the present invention is to provide polymeric microparticles prepared by the above production method.

In addition, the present invention is to provide a medical composition containing the polymeric microparticles.

In addition, the present invention is to provide a cosmetic composition containing the polymeric microparticles.

In addition, the present invention is to provide a medical product comprising the above medical composition.

In addition, the present invention is to provide a cosmetic product comprising the above cosmetic composition.

In the present specification, forming crosslinked particles by performing a primary crosslinking reaction of a mixture including a biocompatible polymer in an emulsion state and a first crosslinking agent; and extracting the cross-linked particles and performing a secondary cross-linking reaction in an organic solvent containing a second cross-linking agent.

In the present specification, polymeric microparticles including a polymer matrix in which a biocompatible polymer is crosslinked with a crosslinking agent are provided.

In the present specification, a medical composition comprising the polymeric microparticles and a pharmaceutically effective substance contained in the polymeric microparticles is also provided.

In the present specification, a cosmetic composition comprising the polymeric microparticles and a cosmetically effective substance contained in the polymeric microparticles is also provided.

In the present specification, a medical product containing the above medical composition is also provided.

In the present specification, a cosmetic product containing the above cosmetic composition is also provided.

Hereinafter, a method for manufacturing polymeric microparticles, polymeric microparticles, and medical compositions, cosmetic compositions, medical products, and cosmetic products including the polymeric microparticles according to specific embodiments of the present invention will be described in more detail.

Unless explicitly stated herein, terminology is used only to refer to specific embodiments and is not intended to limit the present invention.

As used herein, the singular forms also include the plural forms unless the phrases clearly dictate the contrary.

As used herein, the meaning of 'comprising' specifies a particular property, domain, integer, step, operation, element, and/or component, and other particular property, domain, integer, step, operation, element, component, and/or group. does not exclude the presence or addition of

In this specification, terms including ordinal numbers such as 'first' and 'second' are used for the purpose of distinguishing one component from another component, and are not limited by the ordinal number. For example, within the scope of the present invention, a first element may also be termed a second element, and similarly, a second element may be termed a first element.

In the present specification, a (co)polymer refers to a polymer or a copolymer, and the polymer refers to a homopolymer composed of a single repeating unit, and the copolymer refers to a multipolymer containing two or more types of repeating units.

In this specification, the molecular weight of the polymer used follows the manufacturer's measurement method, and the method does not deviate from the conventional GPC measurement method. For example, in the present specification, the weight average molecular weight may mean a molecular weight measured by light scattering methods or a viscosity method.

Since the present invention can have various changes and various forms, specific embodiments will be exemplified and described in detail below. However, this is not intended to limit the present invention to a particular disclosed form, and it should be understood to include all modifications, equivalents, and substitutes included in the spirit and technical scope.

In the present specification, microparticles mean that the cross section of the particle is circular or elliptical, and the ratio of the short axis/long axis of the particle (sphericity ratio) is in the range of 0.7 to 1.0. The lengths of the short axis and the major axis of the particle can be derived by taking an optical picture of the particle and calculating an average value of 30 to 100 arbitrary particles in the optical picture.

In the present specification, the particle diameter (Dn) means the particle diameter at the n volume% point of the cumulative distribution of the number of particles according to the particle diameter. That is, D50 is the particle diameter at the 50% point of the cumulative distribution of the number of particles when the particle diameters are accumulated in ascending order, D90 is the particle diameter at the 90% point of the cumulative distribution of the number of particles according to the particle diameter, and D10 is the particle diameter according to the particle diameter It is the particle diameter at the 10% point of the particle number cumulative distribution.

The Dn can be measured using a laser diffraction method. Specifically, after dispersing the powder to be measured in a dispersion medium, it is introduced into a commercially available laser diffraction particle size measuring device (Horiba LA-960) to measure the difference in diffraction pattern according to the particle size when the particles pass through the laser beam to determine the particle size distribution. yield D10, D50, and D90 can be measured by calculating the particle diameter at the point where it becomes 10%, 50%, and 90% of the particle number cumulative distribution according to the particle diameter in the measuring device. More specifically, in the present specification, the particle diameter may mean D50.

In the present specification, an emulsion means a mixed phase in which at least one of oil phase or aqueous phase immiscible liquids is dispersed in another liquid (dispersion medium) in a fine particle state (dispersion medium). Emulsions can be generally divided into macroemulsions, microemulsions, and nanoemulsions according to the particle size of the dispersed phase.

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

One. Manufacturing method of polymeric microparticles

According to one embodiment of the invention, forming cross-linked particles by performing a primary cross-linking reaction of a mixture including a biocompatible polymer in an emulsion state and a first cross-linking agent; and extracting the cross-linked particles and performing a secondary cross-linking reaction in an organic solvent containing a second cross-linking agent.

The emulsion may mean a water-in-oil emulsion, an oil-in-water emulsion, an oil-in-oil emulsion, or multiple emulsions. Specifically, the emulsion of one embodiment may mean a water-in-oil type emulsion.

Conventional polymer microparticles undergo a crosslinking reaction by directly adding the particles to an organic solvent, so the shape of the particles is greatly influenced by the concentration of the polymer and the content of the crosslinking agent. In particular, when the concentration of the polymer is low or the content of the crosslinking agent is large When the particle size is small, the shape of the particles cannot be maintained, so that it is difficult to form microparticles.

Thus, the present inventors maximized the crosslinking density with the same amount of crosslinking agent as the crosslinking reaction proceeded through the second step, as in the manufacturing method of the polymeric microparticles of the embodiment, and the mechanical strength and stability of the microparticles were remarkably improved. was confirmed through experiments and the invention was completed.

In particular, as the secondary crosslinking reaction proceeds in the solid phase among the secondary crosslinking reactions, it was also confirmed through experiments that spherical particles can be implemented even with a small amount of the crosslinking agent and the production yield is improved.

Specifically, the biocompatible polymer refers to a polymer that can be directly injected into the human body for effective drug delivery of drugs applied to the human body. Specifically, the biocompatible polymer is hyaluronic acid (HA), carboxymethyl cellulose (CMC), alginic acid, pectin, carrageenan, chondroitin (sulfate), dextran (sulfate), chitosan, polylysine, collagen, gelatin, carboxymethyl chitin, fibrin, agarose, pullulan, polylactide, polyglycolide (PGA), polylactide-glycolide copolymer (PLGA), poly polyanhydride, polyorthoester, polyetherester, polycaprolactone, polyesteramide, poly(butyric acid), poly(valeric acid), Polyurethane, polyacrylate, ethylene-vinyl acetate polymer, acrylic substituted cellulose acetate, non-degradable polyurethane, polystyrene, polyvinyl chloride, polyvinyl fluoride, poly(vinyl imidazole), chlorosulphonate polyolefins ), polyethylene oxide, polyvinylpyrrolidone (PVP), polyethylene glycol (PEG), polymethacrylate, hydroxypropylmethylcellulose (HPMC), ethylcellulose (EC), hydroxypropylcellulose (HPC), cyclodextrin , poly(N-isopropylamide) (PNIPAam), poloxamer, and polyacrylic acid copolymers, and at least one polymer selected from the group consisting of derivatives thereof.

More specifically, the biocompatible polymer may be hyaluronic acid (HA), gelatin or a mixture of hyaluronic acid and gelatin.

In the present specification, hyaluronic acid may mean including both hyaluronic acid itself and hyaluronic acid salts. Accordingly, the hyaluronic acid aqueous solution may be a concept including all of a hyaluronic acid aqueous solution, a hyaluronic acid salt aqueous solution, and a mixed aqueous solution of hyaluronic acid and a hyaluronic acid salt. The hyaluronic acid salt may be an inorganic salt such as sodium hyaluronate, potassium hyaluronate, calcium hyaluronate, magnesium hyaluronate, zinc hyaluronate, or cobalt hyaluronate, an organic salt such as tetrabutylammonium hyaluronate, or a mixture thereof.

In one embodiment of the invention, the molecular weight of hyaluronic acid is not particularly limited, but is preferably 500 g/mol or more and 5,000,000 g/mol or less in order to realize various physical properties and biocompatibility.

In the present specification, gelatin may refer to a protein obtained by treating animal-derived collagen with acid or alkali and subsequently extracting it.

In one embodiment of the invention, the molecular weight of gelatin is not particularly limited, but is preferably 100,000 g/mol or more and 5,000,000 g/mol or less in order to realize various physical properties and biocompatibility.

In one embodiment of the present invention, examples of the first crosslinking agent are not greatly limited. Specifically, the first crosslinking agent is butanediol diglycidyl ether (1,4-butandiol diglycidyl ether: BDDE), ethylene glycol diglycidyl ether (EGDGE), hexanediol diglycidyl ether (1 ,6-hexanediol diglycidyl ether), propylene glycol diglycidyl ether, polypropylene glycol diglycidyl ether, polytetramethylene glycol diglycidyl ether ether), neopentyl glycol diglycidyl ether, polyglycerol polyglycidyl ether, diglycerol polyglycidyl ether, glycerol polyglycidyl ether (glycerol polyglycidyl ether), tri-methylpropane polyglycidyl ether, bis-epoxypropoxyethylene (1,2-(bis(2,3-epoxypropoxy)ethylene), pentaerythritol polyglycy It may include one selected from the group consisting of pentaerythritol polyglycidyl ether, sorbitol polyglycidyl ether, divinylsulfone, and glutaraldehyde. The first cross-linking agent may be butanediol diglycidyl ether (1,4-butandiol diglycidyl ether: BDDE).

In one embodiment of the invention, the step of forming crosslinked particles by primary crosslinking a mixture including the biocompatible polymer in an emulsion state and the first crosslinking agent is added to an aqueous solution in which the biocompatible polymer is dissolved in a hydrophobic solvent to form an emulsion forming; and adding a first cross-linking agent to the mixed solution containing the emulsion.

That is, the primary crosslinking reaction may be a liquid phase reaction that proceeds by adding a first crosslinking agent to the emulsion containing the biocompatible polymer.

In addition, in the step of forming crosslinked particles by performing a primary crosslinking reaction of a mixture including the biocompatible polymer in the emulsion state and the first crosslinking agent, the first crosslinking agent is 30 parts by weight or more 300 parts by weight based on 100 parts by weight of the biocompatible polymer parts by weight or less, 40 parts by weight or more and 250 parts by weight or less, or 50 parts by weight or more and 200 parts by weight or less. As described above, the method for producing polymeric microparticles according to an embodiment of the present invention proceeds with the secondary crosslinking reaction after the primary crosslinking reaction, and 30 parts by weight or more and 300 parts by weight or less with respect to 100 parts by weight of the biocompatible polymer. Even when the first crosslinking agent is added, polymeric microparticles having sufficient mechanical strength and sphericity can be prepared.

When the content of the first crosslinking agent is added at less than 30 parts by weight based on 100 parts by weight of the biocompatible polymer, it is difficult to form particles due to the low degree of crosslinking, and when added in excess of 300 parts by weight, an excessive amount of unreacted crosslinking agent is not purified and crosslinked Technical problems remaining in the particles may arise.

Meanwhile, according to one embodiment of the present invention, in the step of extracting the crosslinked particles and subjecting the secondary crosslinking reaction to the organic solvent containing the second crosslinking agent, the crosslinking particles and the second crosslinking agent may undergo a crosslinking reaction in a solid state. .

That is, according to the method for preparing the polymeric microparticles of the present invention, a secondary crosslinking reaction proceeding in a solid state in an alkaline organic solvent may be included. In addition, the secondary crosslinking reaction proceeding in the solid phase may mean a secondary crosslinking reaction proceeding in a more aggregated state by dehydrating the swollen polymer particles. As in one embodiment of the present invention, in the case of including a secondary crosslinking reaction that proceeds in the solid phase in an organic solvent, that is, a secondary crosslinking reaction that proceeds in a more aggregated state by dehydrating the swollen polymer particles, otherwise Excellent crosslinking efficiency is realized compared to the case, and thus microparticles having high strength can be obtained.

In one embodiment of the invention, the recovery of the cross-linked particles may be performed by using a mesh sieve having an opening size of 30 μm or more. After recovering the crosslinked particles formed through the primary crosslinking reaction, the secondary crosslinking reaction may proceed in a more aggregated solid phase by adding a second crosslinking agent after redispersing in an organic solvent.

In one embodiment of the present invention, examples of the second crosslinking agent are not greatly limited. Specifically, the second crosslinking agent is butanediol diglycidyl ether (1,4-butandiol diglycidyl ether: BDDE), ethylene glycol diglycidyl ether (EGDGE), hexanediol diglycidyl ether (1 ,6-hexanediol diglycidyl ether), propylene glycol diglycidyl ether, polypropylene glycol diglycidyl ether, polytetramethylene glycol diglycidyl ether ether), neopentyl glycol diglycidyl ether, polyglycerol polyglycidyl ether, diglycerol polyglycidyl ether, glycerol polyglycidyl ether (glycerol polyglycidyl ether), tri-methylpropane polyglycidyl ether, bis-epoxypropoxyethylene (1,2-(bis(2,3-epoxypropoxy)ethylene), pentaerythritol polyglycy It may include one selected from the group consisting of pentaerythritol polyglycidyl ether, sorbitol polyglycidyl ether, divinylsulfone, and glutaraldehyde. The second cross-linking agent may be butanediol diglycidyl ether (1,4-butandiol diglycidyl ether: BDDE).

That is, in one embodiment of the invention, the first cross-linking agent and the second cross-linking agent are each independently butanediol diglycidyl ether (1,4-butandiol diglycidyl ether: BDDE), ethylene glycol diglycidyl ether (ethylene glycol diglycidyl ether: EGDGE), 1,6-hexanediol diglycidyl ether, propylene glycol diglycidyl ether, polypropylene glycol diglycidyl ether , polytetramethylene glycol diglycidyl ether, neopentyl glycol diglycidyl ether, polyglycerol polyglycidyl ether, diglycerol polyglycy Diglycerol polyglycidyl ether, glycerol polyglycidyl ether, tri-methylpropane polyglycidyl ether, bisepoxypropoxyethylene (1,2-(bis(2, 3-epoxypropoxy)ethylene), pentaerythritol polyglycidyl ether, sorbitol polyglycidyl ether, divinylsulfone and glutaraldehyde It may include one selected from, and preferably, both the first and second crosslinking agents may be butanediol diglycidyl ether (1,4-butandiol diglycidyl ether: BDDE).

In addition, the second crosslinking agent may be included in an amount of 30 parts by weight or more and 300 parts by weight or less, 40 parts by weight or more and 250 parts by weight or less, or 50 parts by weight or more and 200 parts by weight or less, based on 100 parts by weight of the biocompatible polymer. In the method for producing polymeric microparticles according to an embodiment of the present invention, as the secondary crosslinking reaction proceeds after the primary crosslinking reaction as described above, 30 parts by weight or more and 300 parts by weight or less based on 100 parts by weight of the biocompatible polymer. It is possible to prepare polymeric microparticles realizing sufficient mechanical strength and sphericity by adding a second crosslinking agent.

When the second crosslinking agent is added in an amount of less than 30 parts by weight based on 100 parts by weight of the biocompatible polymer, it is difficult to form particles due to a low crosslinking degree. There may be technical problems remaining in

On the other hand, in the step of extracting the crosslinked particles and carrying out the secondary crosslinking reaction in an organic solvent containing a second crosslinking agent, the organic solvent is ethanol, N,N-dimethylformamide, N,N-dimethylacetamide, N-methyl- 2-pyrrolidone, N-methylcaprolactam, 2-pyrrolidone, N-ethylpyrrolidone, N-vinylpyrrolidone, dimethylsulfoxide, tetramethylurea, pyridine, dimethylsulfone, hexamethylsulfoxide, Gamma-butyrolactone, 3-methoxy-N,N-dimethylpropanamide, 3-ethoxy-N,N-dimethylpropanamide, 3-butoxy-N,N-dimethylpropanamide, 1,3-dimethyl -Imidazolidinone, ethyl amyl ketone, methyl nonyl ketone, methyl ethyl ketone, methyl isoamyl ketone, methyl isopropyl ketone, cyclohexanone, ethylene carbonate, propylene carbonate, diglyme, 4-hydroxy-4-methyl -2-pentanone, ethylene glycol monomethyl ether, ethylene glycol monomethyl ether acetate, ethylene glycol monoethyl ether, ethylene glycol monoethyl ether acetate, ethylene glycol monopropyl ether, ethylene glycol monopropyl ether acetate, ethylene glycol monoisopropyl It may be one selected from the group consisting of ether, ethylene glycol monoisopropyl ether acetate, ethylene glycol monobutyl ether, and ethylene glycol monobutyl ether acetate.

In addition, in the step of extracting the crosslinked particles and subjecting the secondary crosslinking reaction to the organic solvent containing the second crosslinking agent, the organic solvent containing the second crosslinking agent may be an alkaline mixed solvent. That is, the secondary crosslinking reaction of the present invention can proceed on an organic solvent in which an aqueous alkali solution is mixed with ethanol, which is an organic solvent. Examples of the alkaline solution are not particularly limited, but may be, for example, aqueous sodium hydroxide solution.

As the organic solvent including the second crosslinking agent is an alkaline mixed solvent, it is possible to increase the efficiency of the crosslinking reaction by creating an environment favorable to the nucleophilic substitution reaction ( _SN reaction).

2. Polymeric microparticles

According to one embodiment of the present invention, polymeric microparticles including a polymer matrix in which a biocompatible polymer is crosslinked with a crosslinking agent may be provided.

The present inventors conducted research on polymeric microparticles, and, as described above, after the first crosslinking reaction in the swollen emulsion, the second crosslinking reaction proceeding in the dehydrated and agglomerated solid phase resulted in a crosslinking density with the same amount of crosslinking agent. At the same time as maximizing, it was confirmed through experiments that the mechanical strength and stability of the microparticles were remarkably improved, and the invention was completed.

In particular, as the secondary crosslinking reaction proceeds in a more aggregated solid phase among the secondary crosslinking reactions, it is possible to realize spherical particles even with a small amount of crosslinking agent, and it is also confirmed through experiments that the production yield is improved. has been completed.

Specifically, in the polymer microparticles of the embodiment, the polymer matrix includes a first cross-linked region in which a biocompatible polymer is cross-linked with a first cross-linking agent; and a second cross-linking region in which the biocompatible polymer is cross-linked with a second cross-linking agent.

The first cross-linking region refers to a cross-linking region formed by a primary cross-linking reaction between an emulsion containing a biocompatible polymer and a first cross-linking agent, and the second cross-linking region refers to a cross-linking region formed by a first cross-linking agent and a first cross-linking agent in an emulsion containing a biocompatible polymer. It may refer to a crosslinking region formed by the secondary crosslinking reaction with the second crosslinking agent and a crosslinking region formed through an additional secondary crosslinking reaction between the first crosslinking region and the second crosslinking agent.

Meanwhile, the polymeric microparticles may have an average diameter in distilled water of 1 μm or more and 650 μm or less, 100 μm or more and 600 μm or less, or 200 μm or more and 550 μm or less, or 300 μm or more and 500 μm or less. When the diameter satisfies the aforementioned range, cell adhesion and culture performance are excellent.

In addition, the polymeric microparticles may have a sphericity of 0.9 or more and 1.0 or less, 0.95 or more and 1.0 or less, 0.96 or more and 0.99 or less, or 0.97 or more and 0.99 or less.

The sphericity may be obtained by taking an optical photograph of the polymeric microparticles and calculating an average value of 30 to 100 arbitrary particles in the optical photograph.

In addition, the polymeric microparticles may have a swelling degree of 55 or less, 0.1 or more and 55 or less, 0.1 or more and 50 or less, 0.2 or more and 50 or less, 0.5 or more and 30 or less, or 0.8 or more and 18 or less according to Equation 1 below.

[Equation 1]

Degree of swelling = {(average diameter in distilled water) ³ - (average diameter after drying) ³ } / (average diameter after drying) ³ .

When the degree of swelling of the polymeric microparticles exceeds 55, technical problems such as low crosslinking density and inferior mechanical strength may cause stability.

In addition, the polymeric microparticles have an average compressive strength of 0.1 mN or more, 0.1 mN or more and 100 mN or less, 0.11 mN or more and 100 mN or less, 1 mN or more and 100 mN or less, 1 mN when deformed to a level of 50% of the particle average diameter. It may be 90 mN or less, or 1 mN or more and 60 mN or less.

The average compressive strength may be a value obtained by dividing the compressive force when n polymer microparticles are deformed to 50% of their average diameter by n.

For example, in the present specification, the average compressive strength may be a value obtained by dividing the compressive force by 30 when the 30 polymer microparticles are deformed to 50% of the average diameter.

When the average compressive strength of the polymeric microparticles is less than 0.1 mN, mechanical strength of the polymeric microparticles is inferior, resulting in a significant decrease in cell culturing capacity.

3. Medical composition

According to another embodiment of the present invention, a medical composition comprising the polymeric microparticles of another embodiment and a pharmaceutically effective substance contained in the polymeric microparticles may be provided. Information regarding the polymeric microparticles may include all of the above-described information in the other embodiments.

The pharmaceutically effective substance may exist in a state contained in the polymeric microparticles.

Examples of the pharmaceutically effective substance are not particularly limited, and depending on the application purpose of the polymeric microparticles of the embodiment, an active substance suitable for the purpose may be applied without limitation. That is, specific examples of the pharmaceutically effective substance are not limited, and include ampetaminol, arecolin, atropine, bupranolol, buprenorphine, and capsaicin. ), carisoprodol, chlorpromazine, ciclopirox olamine, cocaine, desipramine, dyclonine, epinephrine , ethosuximide, floxetine, hydromorphine, imipramine, lidocaine, methamphetamine, melproic acid, methylphenidate ( methylpenidate, morphine, oxybutynin, nadolol, nicotine, nitroglycerin, pindolol, prilocaine, procaine ), propanolol, rivastigmine, scopolamine, selegiline, tulobuterol, valproic acid, donepezil, etc. and drugs selected from the group consisting of EPO (Erythropoietin), human growth hormone (hGH), Exenatide, GLP-1 (Glucagon-like peptide-1), insulin, CSF (Granulocyte colony-stimulating factor), There are peptide or protein drugs selected from the group consisting of estrogen, progesterone parathyroid hormone (PTH), etc., and all pharmacological substances whose pharmacological effects have been proven can be applied without limitation.

The addition amount of the pharmaceutically effective substance is also not significantly limited, and the content can be used without limitation depending on the application purpose and subject. For example, the active material may be included in an amount of 0.0001 parts by weight or more and 1000 parts by weight or less based on 100 parts by weight of the polymeric microparticles, and may be included in a small amount or excessive amount relative to the polymeric microparticles.

4. Cosmetic composition

According to another embodiment of the present invention, a cosmetic composition comprising the polymeric microparticles of another embodiment and a cosmetically effective substance contained in the polymeric microparticles may be provided. Information regarding the polymeric microparticles may include all of the above-described information in the other embodiments.

The cosmetically effective substance may exist in a state contained in the polymeric microparticles.

Examples of the effective cosmetic substance are not particularly limited, and depending on the application purpose of the polymeric microparticles of the embodiment, an effective substance suitable for the corresponding purpose may be applied without limitation. That is, specific examples of the cosmetically effective substance are not limited, and there are natural extracts, proteins, vitamins, enzymes, antioxidants, etc., and all substances with proven cosmetic effects can be applied without limitation.

The addition amount of the cosmetically effective substance is also not significantly limited, and the content may be used without limitation depending on the application purpose and subject. For example, the cosmetically effective substance may be included in an amount of 0.0001 parts by weight or more and 1000 parts by weight or less based on 100 parts by weight of the polymeric microparticles, in a small amount or excessive amount relative to the polymeric microparticles.

5. Medical Supplies

According to another embodiment of the present invention, a medical product comprising the medical composition of the other embodiment may be provided. Information about the medical composition may include all of the information described above in the other embodiment.

Although the examples of the above medical supplies are not greatly limited, they are suitable for cases where the properties of the present invention are to be inserted into the body or to be maintained for a long period of time. , wound healing agents, and the like.

6. Beauty Supplies

According to another embodiment of the present invention, the present invention may provide a cosmetic product comprising the cosmetic composition of the other embodiment. Information about the cosmetic composition may include all of the information described above in the other embodiment.

Examples of the cosmetic products are not greatly limited, but in order to implement the characteristics of the present invention, for example, cosmetic creams, lotions, hair gels, packs, and the like.

Although the structure of the cosmetic pack is not greatly limited, for example, it may include a support and a cosmetically effective substance delivery layer formed on the support and including the polymeric microparticles of the other embodiments. Examples of the support include woven fabric, nonwoven fabric, silicone, polyethylene terephthalate, polyethylene, polypropylene, polyurethane, metal mesh, and polyester.

According to the present invention, a method for producing polymeric microparticles capable of realizing high crosslinking efficiency and production yield, as well as excellent mechanical strength and stability, and polymeric microparticles using the same can be provided.

The invention is explained in more detail in the following examples. However, the following examples are merely illustrative of the present invention, and the contents of the present invention are not limited by the following examples.

Example 1: Preparation of polymeric microparticles

0.15 g of hyaluronate (weight average molecular weight: 500 kDa, manufacturer: SK Bioland) and 1 g of gelatin (gel strength: 300 g Bloom, manufacturer: Sigma, product name: G2500) were mixed in distilled water at 3 wt.% and 20 wt. %, and then the two solutions are mixed in a 1:1 volume ratio. This mixed solution (11.15 g) was mixed with a liquid paraffin solution (40 g) to prepare a mixed solution containing a water-in-oil (W/O) microemulsion. Thereafter, 1.1 g (1 ml) of 1,4-butandiol diglycidyl ether (BDDE) was added as a cross-linking agent to 51.15 g of the mixed solution, followed by a cross-linking reaction at room temperature for 5 days, Crosslinked particles were prepared by washing with acetone, dichloromethane, and distilled water in that order. At this time, the prepared crosslinked particles were recovered using a net having a sieve size of 45 μm.

A solution in which 1.1 g (1 ml) of 1,4-butandiol diglycidyl ether (BDDE) was added to a mixture of 31.56 g of ethanol and 9.88 g of 0.1N NaOH aqueous solution was prepared, and this solution The cross-linked particles were added to the mixture, and a cross-linking reaction was performed at room temperature for 3 days to prepare polymeric microparticles. After washing the prepared particles in the order of ethanol and distilled water, the prepared cross-linked particles were recovered using a sieve having a size of 45 μm. The recovered cross-linked particles were filtered using a 300 μm mesh sieve, and the remaining cross-linked particles were analyzed.

Example 2: Preparation of polymeric microparticles

Polymer microparticles were prepared in the same manner as in Example 1, except that only 1 g of gelatin was added without adding hyaluronate.

Comparative Example 1: Preparation of polymeric microparticles

0.15 g of hyaluronate (weight average molecular weight: 500 kDa, manufacturer: SK Bioland) and 1 g of gelatin (gel strength: 300 g Bloom, manufacturer: Sigma, product name: G2500) were mixed in distilled water at 3 wt.% and 20 wt. %, and then the two solutions are mixed in a 1:1 volume ratio. This mixed solution (11.15 g) was mixed with a liquid paraffin solution (40 g) to prepare a mixed solution containing a water-in-oil (W/O) microemulsion. Thereafter, 1.1 g (1 ml) of 1,4-butanediol diglycidyl ether (BDDE) was added as a cross-linking agent to 51.15 g of the mixed solution, followed by a cross-linking reaction at room temperature for 5 days.

Thereafter, 1.1 g (1 ml) of 1,4-butandiol diglycidyl ether (BDDE) was added to the mixed solution as a cross-linking agent, followed by a cross-linking reaction at room temperature for 3 days to obtain polymeric microparticles. was manufactured. After washing the prepared particles in the order of acetone, dichloromethane, and distilled water, the prepared cross-linked particles were recovered using a sieve having an opening size of 45 μm. The recovered cross-linked particles were filtered using a 300 μm mesh sieve, and the remaining cross-linked particles were analyzed.

Comparative Example 2: Preparation of polymeric microparticles

0.15 g of hyaluronate (weight average molecular weight: 500 kDa, manufacturer: SK Bioland) and 1 g of gelatin (gel strength: 300 g Bloom, manufacturer: Sigma, product name: G2500) were mixed in distilled water at 3 wt.% and 20 wt. %, and then the two solutions are mixed in a 1:1 volume ratio. This mixed solution (11.15 g) was mixed with a liquid paraffin solution (40 g) to prepare a mixed solution containing a water-in-oil (W/O) microemulsion. Thereafter, 1.1 g (1 ml) of 1,4-butanediol diglycidyl ether (BDDE) was added as a cross-linking agent to 51.15 g of the mixed solution, followed by a cross-linking reaction at room temperature for 5 days. After washing the prepared particles in the order of acetone, dichloromethane, and distilled water, the prepared cross-linked particles were recovered using a sieve having an opening size of 45 μm.

The crosslinked particles were added to a mixture of 31.56 g of ethanol and 9.88 g of 0.1N NaOH aqueous solution, and crosslinked at room temperature for 3 days to prepare polymeric microparticles. After washing the prepared particles in the order of ethanol and distilled water, the prepared cross-linked particles were recovered using a sieve having a size of 45 μm. The recovered cross-linked particles were filtered using a 300 μm mesh sieve, and the remaining cross-linked particles were analyzed.

Comparative Example 3: Preparation of polymeric microparticles

0.15 g of hyaluronate (weight average molecular weight: 500 kDa, manufacturer: SK Bioland) and 1 g of gelatin (gel strength: 300 g Bloom, manufacturer: Sigma, product name: G2500) were mixed in distilled water at 3 wt.% and 20 wt. %, and then the two solutions are mixed in a 1:1 volume ratio. This mixed solution (11.15 g) was mixed with a liquid paraffin solution (40 g) to prepare a mixed solution containing a water-in-oil (W/O) microemulsion. Thereafter, 1.1 g (1 ml) of 1,4-butanediol diglycidyl ether (BDDE) was added as a cross-linking agent to 51.15 g of the mixed solution, followed by a cross-linking reaction at room temperature for 5 days. After washing the prepared particles in the order of acetone, dichloromethane, and distilled water, the prepared cross-linked particles were recovered using a sieve having an opening size of 45 μm. The recovered cross-linked particles were filtered using a 300 μm mesh sieve, and the remaining cross-linked particles were analyzed.

Experimental Example: Measurement of physical properties of polymeric microparticles

With respect to the polymeric microparticles prepared in Examples and Comparative Examples, swelling, sphericity, strength, cell culture suitability, and stability of the polymeric microparticles were evaluated in the following manner.

1. Average diameter and degree of swelling

The average diameter of the polymeric microparticles of Examples and Comparative Examples was measured, and swelling degree was calculated therefrom.

The degree of swelling of the particles according to the present invention was calculated by Equation 1 below.

[Equation 1]

Degree of swelling = {(average diameter in distilled water) ³ - (average diameter after drying) ³ } / (average diameter after drying) ³ .

At this time, the closer the swelling degree value is to 0, the more swelling does not occur, and the larger the value, the larger the swelling is.

The average diameter in distilled water was measured using a laser particle size analyzer (Horiba, Partica LA-960), and after drying from the diameters of 30 random particles in the SEM (Hitach, S-4800) photograph of the dried microparticles. The average diameter was calculated.

2. Sphericity

Optical (Olympus, BX53) photographs of the polymeric microparticles of Examples and Comparative Examples were taken, and the degree of sphericity was calculated therefrom.

The degree of sphericity according to the present invention was calculated as the average value of the ratio of the longest diameter to the shortest diameter (long axis ratio) of 30 random particles in the optical photograph.

At this time, the closer the sphericity value is to 1, the closer it is to a sphere.

3. robbery

The strength of the polymer microparticles of Examples and Comparative Examples was measured using a texture analyzer. Thirty microparticles swollen with distilled water were placed in a single layer on the area below the flat cylindrical probe of the equipment equipped with a 5 N load cell. The initial trigger force was set to 1 mN, and the particles were compressed at a speed of 1 mm/s. The force when the average particle diameter was deformed to 50% of the original diameter was determined as the compressive force.

The average compressive strength is a value calculated by dividing the compressive force by 30, which is the number of microparticles to be measured.

4. Cell culture compatibility

The cell culture medium was filled in a 6-well plate, and the cells were cultured by plate-rocking method by adding polymeric microparticles and cells. At this time, the temperature of the culture solution was maintained at 37 ℃, and the number of cells cultured in the polymer microparticles was confirmed by culturing for 3 days.

At this time, cell culture suitability was evaluated according to the following criteria.

Appropriate: When the number of cultured cells is more than 150% compared to the number of input cells

Unsuitable: When the number of cultured cells is less than 150% compared to the number of input cells

5. Stability

The stability of polymeric microparticles immersed in phosphate-buffered saline solution during sterilization using a high-temperature and high-pressure sterilizer (autoclave) and particle stability following long-term culture were evaluated according to the following criteria.

Suitable: When the weight reduction rate of the dried polymer microparticles before and after using the autoclave is less than 10%

Unsuitable: When the weight reduction rate of the dried polymer microparticles before and after using the autoclave exceeds 10%

division Example 1 Example 2 Comparative Example 1 Comparative Example 2 Comparative Example 3 Average diameter in distilled water (㎛) 323±24 471±10 482±62 497±12 498±16 Average diameter after drying (㎛) 123±11 386±24 125±25 121±15 123±21 degree of swelling 17.19 0.82 56.51 132.43 127.79 sphericity 0.98±0.06 0.91±0.17 0.99±0.01 0.98±0.09 0.98±0.03 Average compressive strength (mN) 1.97 1.17 0.90 0.30 0.23 cell culture compatibility fitness fitness fitness fitness fitness stability fitness fitness incongruity incongruity incongruity

As shown in Table 1, the polymer microparticles of the Example prepared by the manufacturing method of the present invention, which includes the step of recovering the particles after the first crosslinking and performing the second crosslinking, have 150% or more of the number of cultured cells compared to the number of cells introduced first. As a result, it was confirmed that it was not only suitable for cell culture, but also suitable for sterilization treatment and long-term culture as the weight reduction rate of the dried polymer microparticles before and after autoclave treatment was less than 10%.

In addition, it was confirmed that the polymer microparticles of Example exhibited a high degree of sphericity with a degree of sphericity of 0.9 or more, and at the same time, a degree of swelling of 0.82 or more and 17.19 or less, exhibiting an excellent degree of crosslinking. In addition, it was confirmed that the average compressive strength was 1.17 mN or more, realizing excellent mechanical properties, and that the particles had a high crosslinking density.

That is, the polymer microparticles of Examples prepared by the manufacturing method of the present invention, which includes recovering particles after primary crosslinking and performing secondary crosslinking, are suitable for cell culture, sterilization treatment, and long-term culture, yet have excellent crosslinking density and mechanical properties It was confirmed that

On the other hand, the polymeric microparticles of Comparative Example 1, in which the particles were not recovered after the primary crosslinking and the secondary crosslinking was performed immediately, showed a weight reduction rate of more than 10% of the dried polymeric microparticles before and after autoclave treatment, making them unsuitable for sterilization treatment and long-term culture. In addition, it was confirmed that the swelling degree was 56.51, indicating an inferior degree of crosslinking compared to the present example. In addition, the average compressive strength was 0.9 mN, indicating inferior mechanical properties, indicating that the particles had a low crosslinking density.

In addition, the polymer microparticles of Comparative Example 2, in which the particles were recovered after the primary crosslinking and subjected to only dehydration without adding a secondary crosslinking agent, showed a weight reduction rate of more than 10% of the dried polymeric microparticles before and after autoclave treatment, making it suitable for sterilization and long-term culture. In addition to being unsuitable, it was confirmed that the swelling degree was 132.43, indicating an inferior degree of crosslinking compared to the examples. In addition, the average compressive strength was 0.3 mN, indicating inferior mechanical properties, and it was confirmed that the particles had a low crosslinking density.

In addition, the polymer microparticles of Comparative Example 3, in which only the primary crosslinking was performed and the secondary crosslinking step was omitted, showed a weight reduction rate of more than 10% of the dried polymeric microparticles before and after autoclave treatment, making them unsuitable for sterilization treatment and long-term culture. It was confirmed that the swelling degree was 127.79, indicating an inferior degree of crosslinking compared to the present example. In addition, the average compressive strength was 0.23 mN, indicating inferior mechanical properties, confirming that the particles had a low crosslinking density.

Claims (18)

Forming crosslinked particles by performing a primary crosslinking reaction of a mixture including a biocompatible polymer in an emulsion state and a first crosslinking agent; and
Extracting the cross-linked particles and subjecting them to a secondary cross-linking reaction in an organic solvent containing a second cross-linking agent;
The step of forming crosslinked particles by performing a primary crosslinking reaction of a mixture including the biocompatible polymer in the emulsion state and the first crosslinking agent,
forming an emulsion by adding an aqueous solution in which the biocompatible polymer is dissolved to a hydrophobic solvent; and
Adding a first crosslinking agent to the mixed solution containing the emulsion; Including,
In the step of extracting the crosslinked particles and performing a secondary crosslinking reaction in an organic solvent containing a second crosslinking agent, the crosslinking particles and the second crosslinking agent perform a crosslinking reaction in a solid state.
delete delete According to claim 1,
In the step of forming crosslinked particles by performing a primary crosslinking reaction of a mixture including the emulsion state biocompatible polymer and a first crosslinking agent,
The first crosslinking agent is included in an amount of 30 parts by weight or more and 300 parts by weight or less based on 100 parts by weight of the biocompatible polymer.
According to claim 1,
In the step of extracting the crosslinked particles and performing a secondary crosslinking reaction in an organic solvent containing a second crosslinking agent,
The second crosslinking agent is included in an amount of 30 parts by weight or more and 300 parts by weight or less based on 100 parts by weight of the biocompatible polymer.
According to claim 1,
In the step of extracting the crosslinked particles and performing a secondary crosslinking reaction in an organic solvent containing a second crosslinking agent,
The organic solvent containing the second crosslinking agent is an alkaline mixed solvent, a method for producing polymeric microparticles.
According to claim 1,
The organic solvent is ethanol, N,N-dimethylformamide, N,N-dimethylacetamide, N-methyl-2-pyrrolidone, N-methylcaprolactam, 2-pyrrolidone, N-ethylpyrrolidone , N-vinylpyrrolidone, dimethylsulfoxide, tetramethylurea, pyridine, dimethylsulfone, hexamethylsulfoxide, gamma-butyrolactone, 3-methoxy-N,N-dimethylpropanamide, 3-ethoxy- N,N-dimethylpropanamide, 3-butoxy-N,N-dimethylpropanamide, 1,3-dimethyl-imidazolidinone, ethyl amyl ketone, methyl nonyl ketone, methyl ethyl ketone, methyl isoamyl ketone, Methylisopropylketone, cyclohexanone, ethylene carbonate, propylene carbonate, diglyme, 4-hydroxy-4-methyl-2-pentanone, ethylene glycol monomethyl ether, ethylene glycol monomethyl ether acetate, ethylene glycol monoethyl ether , consisting of ethylene glycol monoethyl ether acetate, ethylene glycol monopropyl ether, ethylene glycol monopropyl ether acetate, ethylene glycol monoisopropyl ether, ethylene glycol monoisopropyl ether acetate, ethylene glycol monobutyl ether, ethylene glycol monobutyl ether acetate One selected from the group, a method for producing polymeric microparticles.
According to claim 1,
The first crosslinking agent and the second crosslinking agent are each independently butanediol diglycidyl ether, ethylene glycol diglycidyl ether, hexanediol diglycidyl ether, propylene glycol diglycidyl ether, polypropylene glycol diglycidyl Dyl ether, polytetramethylene glycol diglycidyl ether, neopentyl glycol diglycidyl ether, polyglycol polyglycidyl ether, diglycerol polyglycidyl ether, glycerol polyglycidyl ether, trimethylpropane polyglycidyl A method for producing polymeric microparticles comprising one selected from the group consisting of dilether, bisepoxypropoxyethylene, pentaerythritol polyglycidyl ether and sorbitol polyglycidyl ether, divinylsulfone, and glutaraldehyde.
A biocompatible polymer includes a polymer matrix in the form of cross-linking with a cross-linking agent,
The polymer matrix includes a first cross-linked region in which a biocompatible polymer is cross-linked with a first cross-linking agent; and
a second cross-linking region in which the biocompatible polymer is cross-linked with a second cross-linking agent; including,
The first crosslinking region is a crosslinking region formed by a primary crosslinking reaction between the emulsion containing the biocompatible polymer and the first crosslinking agent,
The second crosslinking region is a crosslinking region formed by a secondary crosslinking reaction between an emulsion containing a biocompatible polymer and a second crosslinking agent, and a crosslinking region formed through an additional secondary crosslinking reaction between the first crosslinking region and the second crosslinking agent, polymeric microparticles.
According to claim 9,
The polymeric microparticles have an average diameter of 1 μm or more and 650 μm or less in distilled water.
According to claim 9,
The polymeric microparticles have a swelling degree of 55 or less according to Equation 1 below:
[Equation 1]
Degree of swelling = {(average diameter in distilled water) ³ - (average diameter after drying) ³ }/(average diameter after drying) ³ .
According to claim 9,
The polymeric microparticles
Polymer microparticles having an average compressive strength of 0.1 mN or more when deformed to a level of 50% of the average particle diameter.
delete According to claim 9,
The polymeric microparticles are prepared by the method of claim 1, the polymeric microparticles.
A medical composition comprising the polymeric microparticles of claim 9 and a pharmaceutically effective substance contained in the polymeric microparticles.
A cosmetic composition comprising the polymeric microparticles of claim 9 and a cosmetically effective substance contained in the polymeric microparticles.
A medical article comprising the medical composition of claim 15 .
A cosmetic article comprising the cosmetic composition of claim 16 .