JP2008207137A - Method for producing capsule of mixed fine particle consisting of particle having different mean particle diameter - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a method for synthesizing nanoparticles having different diameters all at once while improving the production efficiency, which is superior to the conventional method in which nanoparticles having different diameters are synthesized separately and purified in low production efficiency and at a high cost. <P>SOLUTION: A polymerization reaction of a cyanoacrylate monomer is advanced in the coexistence of two or more different stabilizers to synthesize capsules of mixed fine particles having different mean particle diameters all at once. <P>COPYRIGHT: (C)2008,JPO&INPIT

Description

The present invention relates to a method for simultaneously synthesizing a mixed fine particle capsule, that is, a mixture of fine particle capsules composed of fine particle groups having different average particle sizes. Specifically, in the polymerization reaction of a cyanoacrylic monomer, the cyanoacrylic monomer is polymerized in an aqueous solvent in the presence of two or more different stabilizers selected from polysaccharides and surfactants. Fine particle capsules, for example, a method of simultaneously synthesizing mixed nanoparticles (a mixture of two or more types of particles having different particle diameters) containing particles of several tens of nanometers and particles of several hundreds of nanometers simultaneously in one reaction . Furthermore, the present invention relates to the use of the mixed fine particle capsule prepared by the synthesis method of the present invention as a medicine.

In recent years, research on new drugs has been remarkable, and more and more effective drugs have been developed one after another, and the therapeutic results have been steadily increasing. On the other hand, the incidence of side effects of drugs has increased, especially in the treatment of intractable diseases such as life-style related diseases that are difficult to treat with cancer or drastic treatment, and the public's strict attention is paid to the effect of drug use and cost effectiveness. It has come to be. In order to answer these problems, the development of new drug types (new drugs) using fine particle technology such as liposomes, nano micelles, and nanospheres, and drug delivery systems (DDS) for the purpose of sustained drug release and topical administration. ) Research has begun.

Fine particles such as liposomes, nanomicelles, and nanospheres are excellent in binding (for example, adsorption, absorption, uptake) to natural or synthetic substances such as drugs, pharmaceuticals, diagnostic agents, antisense oligonucleotides, proteins, and plasmids, and humans. Or it is clear in the prior art that it is suitable for transporting such substances to target organs such as the brain, liver and kidney in the body of an animal. In particular, in Japan, development of anticancer agents using a cationic liposome system is progressing.

On the other hand, when looking at the disease state from the patient's clinic, as is evident in cancer, the whole diseased organ / organ is not bad, but there is almost no actual condition that some of the organs are dysfunctional and cause disease. In other words, I came to think that the disease is caused by the defect of some organelles of the organ cells.

In gene therapy, which is attracting attention as a new treatment method in the 21st century, it is essential to introduce a target gene or drug into a target cell or organ in a pinpoint manner. To date, methods have been developed that use liposomes, nanomicelles, and other viral vectors as drug and gene carriers, but it is difficult to efficiently target specific cells and organs, and there is no cell / tissue specificity. In addition, there are serious problems such as fear of serious side effects when used in humans.

Therefore, the ultimate treatment method including intractable diseases should aim at cell therapy, and the ultimate drug therapy is the development of drugs with high affinity for each organelle of cells. That is, the development of new DDS for organelles. In the current development of new drugs, most new drug candidates have disappeared due to side effects resulting from pharmacological action in normal organs and tissues in clinical trials. The development of selective cell therapy drug forms not only restores many new drug candidates that are a lot of intelligence and economic products, but also improves the main components that make up the current 25,000 types of drugs into new drug forms. Can greatly improve the current state of medicine.

Research on application of nanotechnology to molecular control and structure construction has just begun in Japan several years ago, but global development competition is tremendous as a national measure of each country. However, research and development of cell therapy drugs with different sizes and physicochemical properties according to the mechanism of action of the drugs have not been conducted yet, and application to cell therapy has not been realized.

The raw material of the nanostructure is divided into a liposome system and an acrylic system. Since the liposome system is a lipid, it is easy to be incorporated into the intraretinal system, and Japan has taken the leadership as a technology for introducing genes into cells. A nanoparticle (nanocapsule), which is one of the nanostructures, is one of composite fine particles and is composed of a plurality of materials. “Gas, liquid, solid” as a constituent component can be combined with substances in various states, and various functions can be imparted to the composite fine particles depending on what is combined.

A nanocapsule generally has a size in the range of several tens of nanometers to several hundreds of nanometers (1 nm = 1,000,000 / mm). The shape and structure of the nanocapsule are determined by the physicochemical properties of the substance used, the preparation method, and the like, and what shape and structure is good depends on the application. A nanocapsule is composed of a container and an internal space. The portion of the container is referred to as a “capsule wall” or “shell”, and a material that can be placed in the internal space is referred to as a “core material” or “core”. Multiple nuclei (cores) can exist depending on the preparation method and application purpose. The function of the composite fine particles and capsules is to protect and isolate the core material for a long time, take out the core material instantaneously when necessary, take out the core material over a long time (sustained release), remove the gas and liquid By encapsulating, it can be apparently treated as a solid.

From the viewpoint of application to pharmaceuticals, its sustained release is considered to be extremely effective. In other words, the pharmacologically active ingredient can be controlled and delivered to the site of action at the optimal rate and dosage for treatment, and the distribution of the active ingredient in the body can be adjusted, thereby improving the therapeutic effect. It will be possible to do this.

The method for producing nanocapsules is basically the same as that for microcapsules. There are a chemical method, a physicochemical method, and a mechanical method depending on how the capsule wall is made as a method for making such a microcapsule. Among these, the chemical method will be described below.

The chemical method is a method in which a capsule wall is formed by using a chemical reaction to form a nanocapsule. This method includes an interfacial polymerization method and an in-situ polymerization method. The interfacial polymerization method is
In this method, nanocapsules are produced by a reaction such as A (reactant) + B (reactant) → C (solid reactive organism = capsule wall). First, the reactant A is dissolved in an aqueous solution in which the core substance is dissolved or dispersed, and when this aqueous solution is poured into the oil phase and vigorously stirred, the aqueous solution is dispersed into the oil phase as small droplets. To do. Such a dispersion system is called a W / O dispersion system, that is, W represents a water phase in the form of water droplets, and O represents an oil phase. When the reactant B that is soluble in the oil phase is injected into such a dispersion, the reaction with the reactant A dissolved in the water droplet occurs at the interface between the water droplet and the oil phase, and the reactant C is generated so as to surround the water droplet. Therefore, encapsulation is performed. In order to complete the encapsulation of the core material by such a mechanism, the reaction must proceed very fast, so that nylon is produced from hexamethylenediamine (reactant A) and sebacoyl dichloride (reactant B). Such a polycondensation reaction is often used. In this way, nanocapsules are produced by a polymerization reaction that occurs at the interface between a liquid (aqueous phase) and a liquid (oil phase), which is called an interfacial polymerization method. Even if the order of the water phase and the oil phase is changed, nanocapsules are produced in the same manner. In this case, the core substance is oil-soluble or dispersed in the oil phase.

The in-situ polymerization method differs from the interfacial polymerization method in that only one type of reactant called a monomer is dissolved in the core material solution, and the monomers are polymerized to form a polymer capsule wall. .
In particular, anionic polymerization is carried out by using a monomer dissolved in a core substance solution at a low pH and using hydroxyl ions as a catalyst. A large number of these monomers combine to form a polymer with a large molecular weight, which becomes a capsule wall and synthesizes nanocapsules. For example, a method of synthesizing a nano-sized polymer from an acrylic monomer using a polymerization stabilizer (such as dextran or polysorbate) is currently known (see Non-Patent Document 1). However, with this method, fine particles cannot be synthesized separately for each 100 nm size, and the degree of polymerization of the polymerization stabilizer dextran and the toxicity of emulsifiers such as polysorbate are major problems for the development of nanocapsule formulations. Yes.

  For the production of nanocapsules as pharmaceuticals and the like, chemical methods are mainly used among the above-mentioned methods, and many reports have been made especially on formulation technology using polycyanoacrylate. Cyanoacrylate is known as an instantaneous adhesive and has been put into practical use as a surgical adhesive or hemostatic agent in the medical field.

For example, JP-T-11-503148 reports the controlled release of biodegradable nanoparticles containing insulin as follows. (See Patent Document 1)

Controlled release drug formulations include nanoparticles formed from biodegradable polycyanoacrylate polymers that have captured insulin, wherein the insulin is complexed to an alkyl polycyanoacrylate. These particles can release bioactive insulin in vivo at a slower rate than nanoparticles from which insulin is free. The formulation may include a mixture of nanoparticles free of insulin and nanoparticles complexed with insulin to obtain the desired release profile. The nanoparticles have a preferred insulin loading of 15-25% w / v and a preferred size of 100-400 nm. Administration may be either oral or parenteral and for oral administration an enteric coating may be applied to target release to the small intestine. Insulin complexation is achieved by polymerizing the cyanoacrylate monomer in the presence of insulin at a low pH, preferably pH = 2.

In the production of the nanoparticles, various dextrans are used as stabilizers, and surfactants such as sorbate are used. Dextran is a high-molecular-weight (100,000 or more) viscous polysaccharide obtained by allowing lactic acid bacteria to act on sucrose. However, the particle size of the obtained nanocapsule is about 200 to 350 nm when dextran is used, and about 10 to 100 nm (peak is about 80 nm) when sorbate (for example, Tween 20 (registered trademark)) is used. It was extremely limited, and the degree of freedom in particle size design was low. Further, there has been no literature that suggests the use of monosaccharide compounds and disaccharide compounds as stabilizers or initiators for polymerization.

There is a fixed size for the functional expression of each organ, cell, and organelle that make up the body. For the transmission of information materials, the size of information materials (including allosteric) and electrical strength in the membrane Depending on the physicochemical properties, the membrane is selectively passed through the membrane by the passive transport system, the active transport system, or the membrane dynamic transport system to express its function.
However, in the above conventional method, it is impossible to perform selective cell therapy because capsules having different physicochemical properties such as size and electric strength at the nanoscale cannot be produced as desired. It was. In addition, it has been impossible to selectively transfer antigens to immune-presenting cells such as dendritic cells, and to develop vaccine methods for various viral diseases such as AIDS, avian influenza and koi herpes.

S.J.Douglas, L.Illum, and S.S.Davis; Particle Size and Size Distribution of Poly (butyl2-cyanoacrylate) Nanoparticles, II. Influence of Stabilizers, J. Colloid Inter Science, 103, 154-163, 1985 Japanese National Patent Publication No. 11-503148

The development of a new tide using new particle technology such as liposomes, nanomicelles, and nanospheres (new drugs) and research on drug delivery systems (DDS) to target organs have been conducted. In order to produce a mixed preparation composed of a group of nanoparticles of different sizes, it was necessary to synthesize and mix each particle separately. However, uniform mixing is a very difficult operation, so that production efficiency is poor and high cost is required.
On the other hand, from the viewpoint of bioavailability, a preparation that lasts for a long period of time is desired, and even a fragrance or the like that has a rapid effect in the initial stage is desired. It is rare. For that purpose, a mixed preparation having a broader particle size distribution or different particle sizes is advantageous. Under such a background, it has been strongly desired to develop a synthesis method that can simultaneously synthesize nanoparticles having different sizes and improve production efficiency.

The present inventors have conducted extensive research to solve the above-mentioned problems, and as a result of examining the polymerization reaction of the cyanoacrylate monomer in the synthesis of the nanocapsule, it has been found that an initiator is required for the polymerization reaction. It was. It was also found that conventional stabilizers also serve as initiators in nucleophilic substitution reactions. Thus, it has been found that if different stabilizers are used simultaneously, different polymerization reactions proceed independently during the reaction, and as a result, polymers of different sizes can be obtained simultaneously. In the past, when two or more different stabilizers were used, the particle size of the obtained nanocapsules was thought to be the average value of the capsule particle size obtained using both alone, but surprisingly Unexpectedly, the present invention was completed by finding that a mixture of capsules with different particle sizes was obtained.

Specifically, the present invention is as follows.
1. Different average particle size sizes containing at least one active active ingredient, characterized in that a cyanoacryl monomer is polymerized in an aqueous solvent in the presence of two or more different stabilizers selected from polysaccharides and surfactants A method for producing a mixed fine particle capsule comprising the following particle groups.
2. Stabilizers are alkylaryl poly (oxyethylene) sulfate alkali salts, dextran, cyclodextrins, poly (oxyethylene), poly (oxypropylene) -poly (oxyethylene) -block polymers, ethoxylated fatty alcohols (sets) Macrogol), ethoxylated fatty acids, alkylphenol poly (oxyethylene), copolymers of alkylphenol poly (oxyethylene) and aldehyde, partial fatty acid esters of sorbitan, partial fatty acid esters of poly (oxyethylene) sorbitan, poly (oxyethylene) Fatty acid esters of poly (oxyethylene), fatty alcohol ethers of poly (oxyethylene), fatty esters of sucrose or tuna gol glycerol ester, polyvinyl alcohol, poly (oxyethyl) Emissions) - hydroxy fatty acid ester, a manufacturing method of mixing particulate capsule according to claim 1 which is 2 or more stabilizers selected from macrogol, and partial fatty acid esters of polyhydric alcohols.
3. 3. The method for producing a mixed fine particle capsule according to 2 above, wherein the stabilizer is at least two kinds of stabilizers selected from dextran, β-cyclodextrin and polysorbate.
4. The mixed fine particle capsule according to 3 above, wherein the combination of two or more kinds of stabilizers is dextran and β-cyclodextrin, dextran and polysorbate, β-cyclodextrin and polysorbate, or dextran, β-cyclodextrin and polysorbate. Production method.
5. 5. The cyanoacrylate monomer according to any one of 1 to 4 above, wherein the cyanoacrylate monomer is n-butyl cyanoacrylate monomer, isobutyl cyanoacrylate monomer, isohexyl cyanoacrylate monomer, dextran having different degrees of polymerization, or a mixture thereof. Manufacturing method of mixed fine particle capsule.
6). 1 to 5 above, wherein the active active ingredient is an active active ingredient for a pharmaceutical composition, cosmetic composition, deodorant / fragrance composition, coating composition, adhesive composition, friction reducing composition or food composition The manufacturing method of the mixed fine particle capsule of any one of Claims 1.
7). Active active ingredient is anticancer agent, antisense preparation, antiviral preparation, antibiotic preparation, protein preparation, polypeptide preparation, polynucleotide preparation, antisense nucleotide preparation, vaccine preparation, immunomodulation preparation, steroid preparation, analgesic preparation, anti 7. The method for producing a mixed fine particle capsule according to 6 above, which is an active ingredient for a pharmaceutical composition selected from a morphine preparation, an antifungal preparation and an anthelmintic preparation.
8). 8. A mixed fine particle capsule preparation produced by the method according to any one of 1 to 7 above.

As is apparent from the test results below, by using the method of the present invention, it became possible to simultaneously synthesize mixed fine particle capsules of different sizes, succeeding in increasing production efficiency and reducing production cost. .
In addition, the local area requiring treatment is not necessarily composed of a single tissue or cell group, but is often composed of a plurality of tissues or cell groups. For the transmission of information substances, depending on the physicochemical characteristics such as the size (including allosteric) of the information substance and electrical strength in the membrane, it can be selectively used depending on the passive transport system, active transport system or membrane dynamic transport system. It passes through the membrane and expresses its function. If mixed fine particle capsules of the same constituent and different sizes can be used as pharmaceuticals, it seems that the therapeutic effect can be further increased. Further, from the viewpoint of the release rate of the active active ingredient, the mixed fine particle capsules of different sizes have different release rates, so that they are excellent in sustainability and sustained release.

The present invention produces mixed fine particle capsules composed of particles having different average particle sizes by simultaneously using a plurality of different stabilizers when synthesizing a polymer having a size by polymerization reaction of cyanoacrylate monomers. Is.
More specifically, as shown in the following examples, at the time of the polymerization reaction of the cyanoacrylate monomer, by coexisting at least two or more kinds of stabilizers selected from dextran, β-cyclodextrin, sorbate, etc., at least simultaneously. A mixed fine particle capsule composed of a group of particles having two different average particle sizes is manufactured. Further, a low molecular sugar compound may coexist.

Hereinafter, the present invention will be described in detail by explaining terms used in the present invention.
In the present invention, the “cyanoacrylate monomer” means (2-cyanoacrylic acid 2-12C (straight or branched) alkyl). Preferred cyanoacrylate monomers are 4-6C alkyl monomers cyanoacrylate, such as n-butyl cyanoacrylate, isobutyl cyanoacrylate, isohexyl cyanoacrylate, or mixtures thereof.
The polymerization method of the cyanoacrylate monomer for preparing the nanocapsules may be either an interfacial polymerization method or an anionic polymerization method. The interfacial polymerization method is a method in which a monomer and a drug are dissolved in an organic phase, the organic phase is added to an aqueous phase, and a polymerization reaction is performed at an organic-water interface. Nanocapsules are produced in this way. In general, nanoparticles loaded with prior art peptides are prepared by this method.
Anionic polymerization occurs in a low pH aqueous medium and is catalyzed by hydroxyl ions. In this method, it is said that the drug is taken into the nanoparticles, adsorbed onto the nanoparticles, or a combination of both depending on the time for which the drug is added to the polymerization medium.

In the present invention, the “low molecular weight saccharide compound” means a monosaccharide compound, a disaccharide compound or a trisaccharide compound, and preferably a monosaccharide compound or a disaccharide compound.

In the present invention, the “monosaccharide compound” is not particularly limited, but erythrose, threose, ribose, lyxose, xylose, arabinose, allose, talose, gulose, glucose, altrose, mannose, galactose, idose, fructose , Dihydroxyacetone, erythrulose, xylulose, ribulose, psicose, fructose, sorbose, tagatose and the like. Of these, glucose, mannose, ribose and fructose are preferred.

In the present invention, the “disaccharide compound” is not particularly limited, but refers to maltose, trehalose, lactose, sucrose, cellobiose, nigerose, soforose and the like. Of these, maltose, trehalose, lactose and sucrose are preferable.

The “aqueous solvent” is specifically water, preferably distilled water, weak acid salt, physiological saline or hydrous alcohol.

The “mixed fine particle capsule” means a mixture of fine particle capsules composed of fine particle groups having different average particle sizes. In addition, “fine particle capsule” herein means a capsule having a particle size of about 50 to 5000 nm, preferably a capsule having a particle size of 50 to 800 nm.
“Nanoparticles” and “nanocapsules” are synonymous. The capsule means a sheath-like container shape, but is not particularly limited to this and may be a simple nanoparticle.
The amount of the active ingredient conjugated to the biodegradable nanocapsule is preferably 5 to 30% w / v, more preferably 15 to 25% w / v based on the monomer content.

The particle diameter of mixed fine particle capsules obtained by polymerizing cyanoacrylate monomers generally has a certain spread. The “average particle size” in this specification means a particle size that exhibits a peak in these distributions. Therefore, it does not necessarily indicate a statistical average value, but in most cases, it matches the statistical average value.
“Mixed fine particle capsules composed of particles having different average particle sizes” means mixed fine particle capsules composed of two or more particle groups having different average particle sizes, and different particle sizes including at least one kind of nanoparticles. It means a capsule mixture composed of a group, and preferably all of the mixed capsules are nanocapsules.

“Active active ingredient” means an active ingredient (active ingredient) for a pharmaceutical composition, cosmetic composition, deodorant / fragrance composition, coating composition, adhesive composition, friction reducing composition or food composition. Although not particularly limited, preferably, anticancer agent, antisense agent, antiviral agent, antibiotic agent, protein, polypeptide, polynucleotide, antisense nucleotide, vaccine substance, immunomodulator, A steroid, analgesic, antimorphine, antifungal or anthelmintic.
The “pharmaceutical composition” in the “active active ingredient for the pharmaceutical composition” includes anticancer agents, antisense preparations, antiviral preparations, antibiotic preparations, protein preparations, polypeptide preparations, polynucleotide preparations, antisense nucleotide preparations. , Vaccine preparations, immunomodulating preparations, steroid preparations, analgesic preparations, antimorphine preparations, antifungal preparations and anthelmintic preparations.
The form of the “active active ingredient” is not particularly limited and can be various forms such as low molecular weight compounds, proteins, polypeptides, polynucleotides, antisense nucleotides, etc., but from the viewpoint of DDS or vaccine. Preferred are polypeptides, polynucleotides and antisense nucleotides. These active ingredients may be conjugated in the nanocapsule or may be attached and held on the surface of the nanocapsule.

“Viral diseases” means diseases caused by various viruses such as AIDS, avian influenza and koi herpes.

The “stabilizer” means a substance having an action of stabilizing the nanocapsule, and a stabilizer made of a polysaccharide such as dextran or cyclodextrin and a surfactant such as polysorbate can be used. Specifically, alkylaryl poly (oxyethylene) sulfate alkali salts, dextran, cyclic dextrins such as β-cyclodextrin, poly (oxyethylene), poly (oxypropylene) -poly (oxyethylene) -block polymer, ethoxyl Fatty alcohol (cetomacrogol), ethoxylated fatty acid, alkylphenol poly (oxyethylene), copolymer of alkylphenol poly (oxyethylene) and aldehyde, partial fatty acid ester of sorbitan, partial fatty acid of poly (oxyethylene) sorbitan Esters, poly (oxyethylene) fatty acid esters, poly (oxyethylene) fatty alcohol ethers, sucrose fatty esters or tuna globrel glycerol esters, polyvinyl alcohol Call, poly (oxyethylene) - hydroxy fatty acid esters and macrogol or partial fatty acid esters of polyhydric alcohols. Preferred are polysaccharides and surfactants, particularly preferred as polysaccharides are dextran or β-cyclodextrin, and preferred surfactants as stabilizers are polysorbates and the like, particularly preferably dextran, Tween. (Registered trademark).
As “two or more different stabilizers selected from polysaccharides and surfactants”, preferred combinations include dextran and β-cyclodextrin, dextran and polysorbate, dextran having different degrees of polymerization, β-cyclodextrin and polysorbate, or Dextran, β-cyclodextrin and polysorbate. Particularly preferred is a combination of dextran, polysorbate and dextran having different degrees of polymerization.

“Dextran” is a polysaccharide produced by a kind of lactic acid bacteria using sucrose as a raw material, and is a substance characterized by containing glucose as the only component and containing many α-1,6 glycosidic bonds. Industrially useful dextran is produced by partially hydrolyzing a polymer dextran produced by one kind of lactic acid bacteria, followed by a fractionation / purification process. By adjusting the decomposition and purification process, dextran having different molecular weights is synthesized depending on the application. In the present invention, “dextran” is not particularly limited as long as it is a substance having the above-mentioned characteristics, but dextran 10 (MW10000), dextran 20 (MW20000), dextran 40 (MW40000), dextran 70 (MW70000) Dextran 150 (MW 150,000) is preferred.

The amount of the stabilizer used cannot be generally described because the physical properties differ depending on the stabilizer, but in the case of the stabilizer used, particularly a surfactant, it is preferably within the critical micelle concentration range. Specifically, about 0.05 to 5 (w / w)%, more preferably about 0.1 to 2.5 (w / w)%, most preferably about 1.0% with respect to the whole. About 1.5%.

The nanocapsule particle diameter of the present invention is in the range of 10 to 1000 nm, preferably 50 to 900 nm, particularly preferably 100 to 400 nm, and can be applied for any purpose of sustained release, target therapy, and barrier passage. Moreover, since the particle diameter can be made as small as about 10 nm, it can be used for transdermal absorption and transmucosal, and can also be used as an injection or oral preparation.

For example, for convenient and effective oral administration, a pharmaceutically effective amount of the nanocapsule of the present invention is tableted with one or more excipients, encapsulated in a gel capsule or the like, and water is added. When added, it can be blended with a component that gives a foaming solution and suspended in a solution or the like. For parenteral administration, the nanocapsules can be suspended in a saline solution or the like.

Oral and parenteral formulations can preferably be designed as formulations once to three times a day.

The polymerization method of polycyanoacrylate for preparing the nanocapsules may be either an interfacial polymerization method or an anionic polymerization method. The interfacial polymerization method is a method in which a monomer and a drug are dissolved in an organic phase, the organic phase is added to an aqueous phase, and a polymerization reaction is performed at an organic-water interface. Nanocapsules are produced in this way. In general, nanoparticles loaded with prior art peptides are prepared by this method.
Anionic polymerization occurs in a low pH aqueous medium and is catalyzed by hydroxyl ions. In this method, depending on the time the drug is added to the polymerization medium, the drug is incorporated into the nanoparticles, adsorbed onto the nanoparticles, or a combination of both.

The nanocapsules as described above are prepared by (a) mixing and dissolving an appropriate amount of at least one stabilizer in an acidic aqueous solution, and (b) mixing and dissolving an appropriate amount of a low-molecular-weight saccharide compound; At least one cyanoacrylate monomer may be gradually added to polymerize this monomer. Anionic polymerization of cyanoacrylate is initiated by OH- in aqueous solution. Further, the polymerization rate is affected by the pH of the solution, and becomes relatively fast when the pH is 4 or less because the OH-concentration becomes high, but becomes relatively slow when the pH is 2 or less because the OH-concentration becomes low. The polymerization is carried out at a temperature of -10 ° C to 60 ° C, preferably 0 ° C to 50 ° C, and particularly preferably 10 ° C to 35 ° C.

In the following examples, n-butyl cyanoacrylate was Loctite (Ireland). Scanning electron microscopy (SEM) (Leica Cambridge)
(Cambridge) S360). The kinetic characteristics of nanocapsule particles were measured by Brownian diffusion, zeta potential, and molecular weight by the He-Ne laser scattering method. The changes in nanoparticle characteristics in each aqueous solution were observed using colloidal characteristics as an index.
Hereinafter, the present invention will be described in more detail with reference to examples, but it goes without saying that the present invention is not limited only to the embodiments described in the examples.

Reference Example 1 Production of Nanoparticles Using Dextran (Stabilizer) (1) A 1N hydrochloric acid solution (1 ml) was diluted with 100 ml of distilled water to obtain a 0.01N hydrochloric acid aqueous solution at pH 2.21.
(2) 500 mg of dextran having a different degree of polymerization was dissolved in 50 ml of 0.01N hydrochloric acid aqueous solution having pH 2.21 to obtain a 1% dextran 0.01N hydrochloric acid aqueous solution. As dextran, (Dex10 (molecular weight; 10000) 1%, Dex70 (molecular weight; 70000) 1%, Dex150 (molecular weight; 150,000) 1%) were used.
(3) While stirring, n-butyl-2-cyanoacrylate (Histacryl blue (1 tube: 0.5 ml)) was added to the above 1% dextran 0.01N aqueous hydrochloric acid solution at a final concentration of 0.5. It was dripped so that it might become%.
(4) This was stirred at room temperature for 2 hours for polymerization.
(5) A 0.1N aqueous sodium hydroxide solution was mixed for neutralization and further stirred for 30 minutes.
(6) This was washed 4 times with distilled water using a centriprep. (7) The particle size of the obtained nanoparticles was measured by He + Ne laser scattered light analysis.
In addition, the average particle diameter of the (n-butylcyanoacrylate) nanoparticles according to the degree of dextran polymerization was about 150 to 220 nm as shown in the table below.

Reference Example 2: Test using Tween (surfactant) (1) 1 ml of 1N hydrochloric acid solution was diluted with 100 ml of distilled water to obtain a hydrochloric acid aqueous solution of 0.01 N and pH 2.21.
(2) 0.25 ml of Tween-20 was added to 49.25 ml of 0.01N hydrochloric acid aqueous solution with pH 2.21.
(3) While stirring at 600 rpm, cyanoacrylate (Histacryl blue 1 tube: 0.5 ml) was added to the Tween-20 hydrochloric acid aqueous solution so that the final concentration was 0.5%.
(4) An appropriate amount of 0.1N sodium hydroxide aqueous solution was added for neutralization.
(5) This was further polymerized by stirring at room temperature for 2 hours.
(6) This was washed 4 times with distilled water using a centriprep.
(7) The particle size of the obtained nanoparticles was measured by He + Ne laser scattered light analysis.
The result is shown in FIG. Although it performed 3 times with A, B, and C, the peak of the particle diameter was 47.6 nm in all.

Reference Example 3 Test using β-cyclodextrin (1) 1 ml of 1N hydrochloric acid solution was diluted with 100 ml of distilled water to obtain a hydrochloric acid aqueous solution of 0.01 N and pH 2.21.
(2) 1 g of β-cyclodextrin was added and dissolved in 50 ml of 0.01N hydrochloric acid aqueous solution having pH 2.21.
(3) While stirring at 600 rpm, cyanoacrylate (Histacryl blue 1 tube: 0.5 ml) was added to the β-cyclodextrin hydrochloric acid aqueous solution so that the final concentration was 0.5%.
(4) Furthermore, this was stirred at room temperature for 2 hours to polymerize.
(5) The solution was neutralized by adding an appropriate amount of 0.1N sodium hydroxide aqueous solution.
(6) This was washed 4 times with distilled water using a centriprep.
(7) The particle size of the obtained nanoparticles was measured by He + Ne laser scattered light analysis.
The result is shown in FIG. The particle size peaks were 405.3 nm (8.78%%), 1577 nm (86.7%), and 5370 nm (4.5%), and three types of mixed fine particle formulations having different particle diameters were obtained. However, the majority (86.7%) was 1577 nm.

Example 1: Production of mixed fine particle capsules having different particle diameters by the combined use of a stabilizer and a surfactant Manufacture of mixed fine particle capsules by combined use of dextran and Tween
To 50 ml of 0.001N hydrochloric acid aqueous solution, 500 mg of dextran 10 (molecular weight 10,000) was added and dissolved to obtain a dextran solution. To this dextran solution, 500 μl of Tween-20 was added and dissolved, and further 2500 mg of glucose was added and dissolved. Under stirring, n-butyl cyanoacrylate (nBCA)
0.5 ml was added and polymerized by stirring for 3 hours. The reaction solution was neutralized by adding 0.1 N sodium hydroxide solution dropwise, then stirred for 30 minutes, and this was washed four times with distilled water using a centriprep. Finally, the particle diameter of the obtained mixed fine particle capsule was measured by He + Ne laser scattered light analysis.
The result is shown in FIG. The particle size peaks were 27.1 nm (58.9%), 218 nm (37.5%), and 4570 nm (3.5%), and nanoparticles with different particle sizes were obtained.
A similar test was performed using dextran 70 (molecular weight 70,000) instead of dextran 10 (molecular weight 10,000). The result is shown in FIG. The peak of the particle diameter was 34.7 nm (75.9%), 455 nm (23.2%), 4130 nm (0.9%), and nanoparticles having different particle diameters were obtained.

B. Properties of Nanoparticles The mixed fine particle capsules produced in the above test were analyzed in more detail. As a result, there were three particle size peaks, and nanoparticle mixtures having three different particle sizes were obtained.
As is apparent from FIGS. 3 and 4, nanoparticles having different particle sizes could be produced simultaneously by using dextran and Tween in combination.
In the case of dextran 10 + Tween-20 + monosaccharide (glucose), three different types of nanoparticles of 27.1 nm (58.9%), 218 nm (37.5%) and 4570 nm (3.5%) were obtained. SEM images of these particles are shown in FIG.
In the case of dextran 70 + Tween-20 + monosaccharide (glucose), three different types of nanoparticles of 34.7 nm (75.9%), 455 nm (23.2%) and 4130 nm (0.9%) were obtained. SEM images of these particles are shown in FIG.

C. Discussion As is apparent from the test results of A and B above, by using dextran and a surfactant (for example, Tween) as stabilizers, mixed microparticle capsules having different particle diameters can be simultaneously produced. However, in any case using dextran 10 and dextran 70, the third peak is negligible, so it can be said that it is a mixed particle having two different particle sizes.

A mixed fine particle capsule in which ampicillin and DNA were conjugated as active active ingredients was manufactured, and the particle size and distribution showed the same results as described above. The same was true in the following tests.

Example 2 Production of mixed fine particle capsules having different particle diameters by the combined use of a stabilizer and β-cyclodextrin Production of mixed fine particle capsules by combined use of dextran and β-cyclodextrin
Add 50 mg of dextran 10 (molecular weight 10000) and 50 mg of β-cyclodextrin to 50 ml of 0.001N hydrochloric acid solution, dissolve, add 0.5 ml of n-butylcyanoacrylate (nBCA) with stirring, and stir for 3 hours. Polymerized. The reaction solution was neutralized by adding 0.1 N sodium hydroxide solution dropwise, then stirred for 30 minutes, and this was washed four times with distilled water using a centriprep. Finally, the particle diameter of the obtained mixed fine particle capsule was measured by He + Ne laser scattered light analysis. Moreover, the same test was conducted by changing the blending amount of dextran 10 (molecular weight 10,000) to 500 mg.
The results are shown in the table below.

Example 3 Production of mixed fine particle capsules having different particle diameters by combined use of surfactant and β-cyclodextrin Production of mixed fine particle capsules by combined use of β-cyclodextrin and Tween
To 50 ml of 0.001N hydrochloric acid aqueous solution, 0.25 ml of Tween-20 and 50 mg of β-cyclodextrin were added and dissolved. While stirring this solution, 0.5 ml of n-butyl cyanoacrylate (nBCA) was added, and polymerization was carried out by stirring for 3 hours. The reaction solution was neutralized by adding 0.1 N sodium hydroxide solution dropwise, then stirred for 30 minutes, and this was washed four times with distilled water using a centriprep. Finally, the particle diameter of the obtained mixed fine particle capsule was measured by He + Ne laser scattered light analysis. Further, the same test was conducted by changing the amount of β-cyclodextrin to 500 mg.
The results are shown in the table below.

Example 4: Production of mixed fine particle capsules having different particle diameters by the combined use of a stabilizer and a surfactant (effect of blending amount)
A. Manufacture of mixed fine particle capsules by combined use of dextran and Tween
To 50 ml of 0.001 normal hydrochloric acid aqueous solution, 0.25 ml of Tween-20 and 50 mg of dextran 10 (molecular weight 10,000) were added and dissolved to obtain a dextran solution. While stirring this dextran solution, 0.5 ml of n-butylcyanoacrylate (nBCA) was added, and the mixture was stirred for 3 hours for polymerization. The reaction solution was neutralized by adding 0.1 N sodium hydroxide solution dropwise, then stirred for 30 minutes, and this was washed four times with distilled water using a centriprep. Finally, the particle diameter of the obtained mixed fine particle capsule was measured by He + Ne laser scattered light analysis.
Moreover, the same test was conducted by changing the blending amount of dextran 10 (molecular weight 10,000) to 500 mg.
The results are shown in the table below.

Example 5: Production of mixed fine particle capsules having different particle diameters by the combined use of a stabilizer, a surfactant and β-cyclodextrin Manufacture of mixed fine particle capsules using dextran, Tween and β-cyclodextrin in combination
To 50 ml of 0.001 normal hydrochloric acid aqueous solution, 0.25 ml of Tween-20, 500 mg of dextran 10 (molecular weight 10,000) and 50 mg of β-cyclodextrin were added and dissolved. While stirring this solution, 0.5 ml of n-butylcyanoacrylate (nBCA) was added, and the mixture was stirred for 3 hours for polymerization. The reaction solution was neutralized by adding 0.1 N sodium hydroxide solution dropwise, then stirred for 30 minutes, and this was washed four times with distilled water using a centriprep. Finally, the particle diameter of the obtained mixed fine particle capsule was measured by He + Ne laser scattered light analysis. The results are shown in FIG. The particle size peaks were 45.3 nm (54.5%), 130 nm (15.8%), and 4430 nm (29.7%), and nanoparticles with different particle sizes were obtained.
Further, the same test was conducted by changing the amount of β-cyclodextrin to 500 mg.
The results are shown in the table below.

According to the method of the present invention, mixed fine particle capsules of different sizes can be simultaneously synthesized, and production efficiency can be increased and production cost can be reduced. In addition, the local area requiring treatment is not necessarily composed of a single tissue or cell group, but is often composed of a plurality of tissues or cell groups. For the transmission of information substances, depending on the physicochemical characteristics such as the size (including allosteric) of the information substance and electrical strength in the membrane, it can be selectively used depending on the passive transport system, active transport system or membrane dynamic transport system. It passes through the membrane and expresses its function. By using the method of the present invention, it becomes possible to efficiently synthesize nanocapsules having the same constituents and different sizes, and if these can be used as pharmaceuticals, the therapeutic effect can be further increased. . This is also preferable from the viewpoint of time-lag release of active ingredients.
In other words, the development of new drugs using nanotechnology is expected, and the development of new diagnostic techniques and new vaccine techniques is also possible. Furthermore, it is expected to be applied to cosmetics, deodorants and fragrances, friction reducing compositions, new coating techniques, and the like.

It is a figure which shows the particle size distribution of the nanocapsule at the time of using Tween20 as a stabilizer (reference example 2). It is a figure which shows the particle size distribution of the capsule particle | grains which used (beta) -cyclodextrin as a stabilizer (reference example 3). (Example 1) which is a figure which shows the particle diameter distribution of the nanocapsule at the time of using Tween20 and dextran 10 as a stabilizer. (Example 1) which is a figure which shows the particle diameter distribution of the nanocapsule at the time of using Tween20 and dextran 70 as a stabilizer. It is a SEM image of a nanocapsule at the time of using Tween20 and dextran 10 as a stabilizer (Example 1). 5A is 25KV × 20,000, and the scale is 1 μm. . 5B is 25KV × 35,000, and the scale is 0.5 μm. It is a SEM image of a nanocapsule at the time of using Tween20 and dextran 70 as a stabilizer (Example 1). 6A is 25KV × 10,000, and the scale is 1 μm. . 6B is 25KV × 35,000, and the scale is 0.5 μm. It is a SEM image of the nanocapsule at the time of using dextran, Tween, and (beta) -cyclodextrin as a stabilizer (Example 5).

Claims (8)

Different average particle size sizes containing at least one active active ingredient, characterized in that a cyanoacryl monomer is polymerized in an aqueous solvent in the presence of two or more different stabilizers selected from polysaccharides and surfactants A method for producing a mixed fine particle capsule comprising the following particle groups. Stabilizers are alkylaryl poly (oxyethylene) sulfate alkali salts, dextran, cyclodextrins, poly (oxyethylene), poly (oxypropylene) -poly (oxyethylene) -block polymers, ethoxylated fatty alcohols (sets) Macrogol), ethoxylated fatty acids, alkylphenol poly (oxyethylene), copolymers of alkylphenol poly (oxyethylene) and aldehyde, partial fatty acid esters of sorbitan, partial fatty acid esters of poly (oxyethylene) sorbitan, poly (oxyethylene) Fatty acid esters of poly (oxyethylene), fatty alcohol ethers of poly (oxyethylene), fatty esters of sucrose or tuna gol glycerol ester, polyvinyl alcohol, poly (oxyethyl) Emissions) - hydroxy fatty acid esters, polyhydric manufacturing method of mixing particulate capsule according to claim 1 alcohol is 2 or more stabilizers selected from macrogol and partial fatty acid esters of.   The method for producing a mixed fine particle capsule according to claim 2, wherein the stabilizer is at least two kinds of stabilizers selected from dextran, β-cyclodextrin and polysorbate.   The combination of two or more kinds of stabilizers is dextran and β-cyclodextrin, dextran and polysorbate, β-cyclodextrin and polysorbate, dextran having a different degree of polymerization, or dextran, β-cyclodextrin and polysorbate. A method for producing mixed fine particle capsules. The method for producing a mixed fine particle capsule according to any one of claims 1 to 4, wherein the cyanoacrylate monomer is n-butyl cyanoacrylate monomer, isobutyl cyanoacrylate monomer, isohexyl cyanoacrylate monomer, or a mixture thereof. . The active active ingredient is an active active ingredient for a pharmaceutical composition, cosmetic composition, deodorant / fragrance composition, paint composition, adhesive composition, friction reducing composition or food composition. The method for producing a mixed fine particle capsule according to any one of the above. Active active ingredient is anticancer agent, antisense preparation, antiviral preparation, antibiotic preparation, protein preparation, polypeptide preparation, polynucleotide preparation, antisense nucleotide preparation, vaccine preparation, immunomodulation preparation, steroid preparation, analgesic preparation, anti The method for producing a mixed microcapsule according to claim 6, which is an active ingredient for a pharmaceutical composition selected from a morphine preparation, an antifungal preparation and an anthelmintic preparation. A mixed fine particle capsule preparation produced by the method according to any one of claims 1 to 7.

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