JP2018145281A - Block copolymer, and polymer micelle, and cosmetic composition, and percutaneous absorption preparation - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a block copolymer that makes it possible to obtain a polymer micelle that resists breaking or does not break even when receiving electric stimulus and prevents occurrence of initial burst.SOLUTION: The present invention provides a block copolymer represented by a formula (I) (A is an alkoxy group; Lis a linking group; X-R is a specific group).SELECTED DRAWING: None

Description

  The present invention relates to a block copolymer, a polymer micelle, a cosmetic composition, and a preparation for percutaneous absorption.

  With the development of nanotechnology, transdermal drug delivery using nanoparticles as a carrier has attracted attention. However, the nanoparticles themselves are very difficult to penetrate into the skin through the stratum corneum, and mainly stay on the stratum corneum surface or accumulate in appendages such as hair follicles. Therefore, in order to enable intradermal delivery using nanoparticles, it is necessary to break through the stratum corneum barrier by a physical enhancement method.

In the cosmetic field, it is legally allowed to penetrate the active ingredient into the stratum corneum. At present, many low molecular weight drugs penetrate into the dermis, and there is another problem that high molecular weight drugs do not penetrate into the stratum corneum.
For example, a transdermal cosmetic composition and a transdermal absorbent carrier using a polyethylene glycol polymer have been proposed (see, for example, Patent Documents 1 and 2).
Patent Document 1 discloses a transdermal cosmetic composition comprising polyethylene glycol and a polyamine copolymer polymer micelle containing an active ingredient.
Patent Document 2 discloses a transdermal absorbent using liposome as a carrier.

  An object of this invention is to provide the block copolymer from which the polymer micelle which does not decay | disintegrate easily by electrical stimulation or does not collapse and does not produce an initial burst is obtained.

  The block copolymer of the present invention as a means for solving the problems is represented by the following general formula (I).

[General Formula (I)]
However, in the said general formula (I), A represents an unsubstituted or substituted C1-C12 alkoxy group, and the substituent of the alkoxy group in the case of being substituted is a formyl group and R < ₁ > R < ₂ >. CH— group (wherein R ₁ and R ₂ are each independently an alkoxy group having 1 to 4 carbon atoms, or R ₁ and R ₂ together form —OCH ₂ CH ₂ O—, —O ( CH _{2) 3} O-, or represents any of _{-O (CH 2) 4 O-)} .
L ₁ is a single bond, — (CH ₂ ) _c S—, —CO (CH ₂ ) _c S—, — (CH ₂ ) _c NH—, — (CH ₂ ) _c CO— (where c is 1 to And a linking group selected from —CO—, —COO—, and —CONH—.
R represents one selected from a single bond, — (CH ₂ ) _p —, and — (CH ₂ ) _p CH— (p represents an integer of 1 to 10).
X is methylimino (—CH ₂ NH—), methyliminomethyl (—CH ₂ NHCH ₂ —), methyloxy (—CH ₂ O—), methyloxymethyl (—CH ₂ OCH ₂ —), methyl ester (— CH ₂ OCO-), methyl ester methyl _{(-CH 2} OCOCH 2 -), and represents any of substituents represented by the following structural formulas.
m is 20 to 10,000.
n is 3 to 1,000.

  According to the present invention, it is an object to provide a block copolymer from which polymer micelles that are not easily disintegrated by electrical stimulation or that do not disintegrate and that do not cause an initial burst are obtained.

1 is a ¹ H NMR spectrum diagram of the PEG-b-PCMS block copolymer obtained in Example 1. FIG. 2 is a ¹ H NMR spectrum diagram of PEG-b-PTMSPMA AIBN1 obtained in Example 2. FIG. 3 is a ¹ H NMR spectrum diagram of PEG-b-PTMSPMA AIBN10 obtained in Example 3. FIG. 4 is a ¹ H NMR spectrum diagram of PEG-b-PTMSPMA AIBN100 obtained in Example 4. FIG. FIG. 5 is a TEM image (40,000 times) of the PEG-b-PCMS block copolymer obtained in the example. 6A is a TEM image (10,000 times) after dispersion of a collodion film of the polymer micelle obtained in Example 4. FIG. FIG. 6B is a TEM image (50,000 times) after the collodion film dispersion of the polymer micelle obtained in Example 4. FIG. 6C is a TEM image (50,000 times) after dispersion of the collodion film of the polymer micelle obtained in Example 4. FIG. 7 is a photograph showing the evaluation results of the first disintegration test in the examples. FIG. 8 is a photograph showing the evaluation results of the second disintegration test of Examples 1 and 5. FIG. 9 is a graph showing the results of sustained release tests in Examples. FIG. 10 is a graph showing the results of a drug release test in Examples.

(Block copolymer)
The block copolymer of the present invention is a block copolymer represented by the following general formula (I), and includes a poly (ethylene glycol) chain segment (A) and a silanol group linking segment (B). Is an AB type diblock copolymer, but may be an ABA type triblock copolymer or an ABBA type tetrablock copolymer.

In the above-mentioned Patent Document 1 (Patent No. 5183378), the block copolymer of the present invention is sufficient for the uniform distribution of the active ingredient, the stability of the polymer micelle in water or in an organic solvent, and the sustained release property. It is based on the knowledge that it is not.
In the above-mentioned Patent Document 2 (Patent No. 6026339), the block copolymer of the present invention is not sufficient in terms of uniform distribution, stability and sustained release of the active ingredient since it penetrates from the pores. This is based on the knowledge that when an external stimulus is applied, a significant defect occurs.

[General Formula (I)]

In the general formula (I), A represents an unsubstituted or substituted alkoxy group having 1 to 12 carbon atoms, and when substituted, the substituent of the alkoxy group is a formyl group, and R ₁ R ₂ CH— Groups (provided that R ₁ and R ₂ are each independently an alkoxy group having 1 to 4 carbon atoms, or R ₁ and R ₂ together form —OCH ₂ CH ₂ O—, —O (CH ₂ ) ₃ O- or -O (CH ₂ ) ₄ O-).
The alkyl moiety in the alkoxy group having 1 to 12 carbon atoms of A is preferably linear or branched alkyl, such as methyl, ethyl, propyl, iso-propyl, butyl, iso-butyl, sec-butyl, Examples include tert-butyl, pentyl, hexyl, heptyl, dodecanyl and the like.
A represents an alkoxy group substituted with the formula: R ₁ R ₂ CH—, and R ₁ and R ₂ are independently alkoxy having 1 to 4 carbon atoms or R ₁ and R ₂ are combined together. _{-OCH 2 CH 2 O -, -} O (CH 2) 3 O- or -O _{when (CH 2)} represents a 4 O-is, such substituents formyl group readily cleaved under acidic conditions Therefore, proteins (for example, antibodies and other ligands) can be easily covalently bonded via their amino groups. If present, such substituents are preferably present at the unbound end of the alkoxy group.

In the general formula (I), L ₁ is a single bond, — (CH ₂ ) _c S—, —CO (CH ₂ ) _c S—, — (CH ₂ ) _c NH—, — (CH ₂ ) _c CO -Represents a linking group selected from-(where c is an integer of 1 to 5), -CO-, -COO-, and -CONH-. c is preferably 1 or 2.

In the general formula (I), R is any one selected from a single bond, — (CH ₂ ) _p —, and — (CH ₂ ) _p CH— (p represents an integer of 1 to 10). Represents. p is preferably 1 or 2.

In the general formula (I), X represents methylimino (—CH ₂ NH—), methyliminomethyl (—CH ₂ NHCH ₂ —), methyloxy (—CH ₂ O—), methyloxymethyl (—CH ₂ OCH). _2- ), methyl ester (—CH ₂ OCO—), methyl ester methyl (—CH ₂ OCOCH ₂ —), and a substituent represented by the following structural formula.

In the said general formula (I), m is 20-10,000, and 20-500 are preferable.
In the said general formula (I), n is 3-1,000, and 3-50 are preferable.

In the general formula (I), L ₁ is —CH ₂ S—, X is a substituent represented by the following structural formula, and R is —CH ₂ CH—, which is not easily disintegrated by electrical stimulation. Or a polymer micelle that does not collapse and does not generate an initial burst is preferable.

<Method for producing block copolymer>
The block copolymer represented by the general formula (I) can be synthesized, for example, as follows.
First, a heterobifunctional polyethylene glycol (PEG-SH) having a methoxy group at the α-terminal and a thiol group at the ω-terminal and toluene are added to the reaction vessel.
Next, azobisisobutyronitrile and chloromethylstyrene are added to the reaction vessel, heated to 60 ° C., and stirred for 24 hours. When the resulting reaction mixture is poured into hexane, a white precipitate is formed. In order to remove the polychloromethylstyrene homopolymer, the obtained precipitate was washed three times with diethyl ether, which is a good solvent for the polychloroethylstyrene homopolymer, and lyophilized with benzene. Polyethylene glycol-b-polychloromethylstyrene (PEG-b-PCMS) having a methoxy group at the α-terminal is synthesized.
Next, the obtained PEG-b-PCMS is put in a reaction vessel, 3-aminopropyltrimethoxysilane is dissolved in dimethyl sulfoxide (DMSO), added to the reaction vessel, and stirred at room temperature for 2 hours. After completion of the reaction, the reaction mixture is poured into 2-propanol cooled to −15 ° C., then re-precipitated three times with 2-propanol, and benzene lyophilized to obtain PEG-b-PTMSPAMS. PEG-Block- (Trimethoxysilyl) propylamine-polymethylstyrene can be synthesized.

The number average molecular weight (Mn) of the block copolymer represented by the general formula (I) is preferably 5,000 or more and 1,000,000 or less. The weight average molecular weight (Mw) of the block copolymer is preferably 5,000 or more and 1,000,000 or less. The Mw / Mn of the block copolymer is preferably 1.0 or more and 2.0 or less.
The number average molecular weight (Mn), the weight average molecular weight (Mw), and Mw / Mn of the block copolymer can be measured by, for example, size exclusion chromatography (SEC).

(Polymer micelle)
The polymer micelle of the present invention contains the block copolymer of the present invention, and further contains other components as necessary.
The polymer micelle of the present invention comprising the block copolymer represented by the general formula (I) is excellent in “crosslinking of the core portion of the micelle” and can stably maintain the micelle structure in water or in an organic solvent. In addition, inhibition of disintegration by external stimulation and long-term sustained release of the active ingredient can be enhanced. Furthermore, since it can penetrate into the stratum corneum and be stored, it can be uniformly distributed.

The block copolymer represented by the general formula (I) has a core-shell structure, and the shell portion is A- (CH ₂ CH ₂ O) _m- (where A and m are the general formula (I)). And the core part is preferably a silanol part.
The average particle diameter of the polymer micelle is preferably 1 nm to 1 μm, and more preferably 10 nm to 500 nm.
The zeta potential of the polymer micelle is preferably −30 mV to 50 mV, and more preferably −10 mV to 40 mV.
The average particle diameter and zeta potential can be measured using, for example, Malvern Zeta Sizer ZSP manufactured by Spectrosys.

An active ingredient can be included in the polymer micelle.
Examples of the method for encapsulating the active ingredient include stirring the polymer micelle and the active ingredient and removing the unincorporated active ingredient by gel filtration, or encapsulating in the dialysis membrane during dialysis when preparing the polymer micelle. Examples thereof include a method of introducing an active ingredient.

There is no restriction | limiting in particular as said active ingredient, According to the objective, it can select suitably, For example, ascorbic acid, vitamin C ethyl, vitamin C glycoside, ascorbyl palmitate, kojic acid, lucinol, tranexamic acid, oil property Whitening ingredients such as licorice extract, vitamin A derivative, placenta extract; anti-wrinkle ingredients such as retinol, retinoic acid, retinol acetate, retinol palmitate, EGF, cell culture extract, acetylglucosamine; tocopherol acetate, capsine, vanillyl amide, etc. Blood circulation promoting ingredients; diet ingredients such as raspberry ketone, evening primrose extract and seaweed extract; antibacterial ingredients such as isopropylmethylphenol, photosensitizer and zinc oxide; vitamins such as vitamin D2, vitamin D3 and vitamin K; glucose and trehalose Such as maltose, and the like.
Examples of the high molecular medicinal component include bioactive peptides or derivatives thereof, nucleic acids, oligonucleotides, various antigen proteins, bacteria, virus fragments, and the like.

  Further, for the purpose of promoting percutaneous absorption, nonionic surfactants such as glyceryl monostearate and sucrose fatty acid ester; water-soluble polymer compounds such as carboxylic acid; water-soluble chelating agents such as EDTA, salicylic acid or Aromatic carboxylic acid compounds such as derivatives; Aliphatic carboxylic acid compounds such as capric acid and oleic acid; Bile salts, propylene glycol, hydrogenated lanolin, isopropyl myristate, diethyl sebacate, urea, lactic acid, aison, etc. added can do.

Further, an ultraviolet absorber or an ultraviolet scattering agent may be included.
Examples of the ultraviolet absorber include octyl cinnamate, ethyl-4-isopropyl cinnamate, methyl-2,5-diisopropyl cinnamate, ethyl-2,4-diisopropyl cinnamate, methyl-2,4-diisopropyl cinnamate. Propyl-p-methoxycinnamate, isopropyl-p-methoxycinnamate, isoamyl-p-methoxycinnamate, octyl-p-methoxycinnamate, 2-ethoxyethyl-p-methoxycinnamate, cyclohexyl-p-methoxycinnamate Cinnamate UV absorbers such as mate, ethyl-α-cyano-β-phenylcinnamate, 2-ethylhexyl-α-cyano-β-phenylcinnamate, glyceryl mono-2-ethylhexanoyl-diparamethoxycinnamate 2,4- Hydroxybenzophenone, 2,2'-dihydroxy-methoxybenzophenone, 2,2'-dihydroxy-4,4'-dimethoxybenzophenone, 2,2 ', 4,4'-tetrahydroxybenzophenone, 2-hydroxy-4-methoxybenzophenone 2-hydroxy-4-methoxy-4′-methylbenzophenone, 2-hydroxy-4-methoxybenzophenone-5-sulfonate, 4-phenylbenzophenone, 2-ethylhexyl-4′-phenyl-benzophenone-2-carboxylate Benzophenone ultraviolet absorbers such as 2-hydroxy-4-n-octoxybenzophenone and 4-hydroxy-3-carboxybenzophenone; PABA monoglycerin ester, N, N-dipropoxy PABA ethyl ester, N, N-diethoxy Paraaminobenzoic acid UV absorbers such as ABA ethyl ester, N, N-dimethyl PABA ethyl ester, N, N-dimethyl PABA butyl ester, N, N-dimethyl PABA methyl ester; amyl salicylate, menthyl salicylate, homomenthyl salicylate, Salicylic acid UV absorbers such as octyl salicylate, phenyl salicylate, benzyl salicylate, p-isopropanol phenyl salicylate; 3- (4′-methylbenzylidene) -d-camphor, 3-benzylidene-d, 1-camphor, urocanic acid, urocanin Examples include acid ethyl ester, octyl triazone, 4-methoxy-4′-t-butyldibenzoylmethane and the like.
Examples of the ultraviolet scattering agent include fine particle titanium oxide, zinc oxide, cerium oxide and the like.

As other components, an additive component included in the polymer micelle can be included, and the type and amount thereof can be appropriately selected within a range that does not impair the desired effects of stability and active ingredients.
Examples of the additive component include nonionic polymers such as guar gum and tamarind gum; cationic polymers such as cationized cellulose and diallyldimethylammonium chloride polymer; anionic polymers such as xanthan gum and sodium alginate; other natural water-soluble compounds or Derivatives thereof: surfactants, oils, colorants, preservatives, chelating agents, antioxidants, humectants, lower alcohols, polyhydric alcohols, fragrances, refreshing agents, pH adjusting agents, and the like.

  The surfactant is added for the purpose of emulsification, solubilization, and dispersion. For example, nonionic surfactants such as POE fatty acid ester, polyglycerin fatty acid ester, POE higher alcohol ether, POE.POP block polymer; Anionic surfactants such as potassium, fatty acid sodium, higher alkyl sulfates, alkyl ether sulfates, acyl sarcosinates, sulfosuccinates; alkyltrimethylammonium salts, dialkyldimethylammonium salts, alkylpyridinium salts, benzalco chloride Cationic surfactants such as nium; imidazoline-based and betaine-based amphoteric surfactants.

  Examples of the oil include vegetable oils such as olive oil, jojoba oil, castor oil, rice bran oil, and palm oil; animal oils such as squalane, beef tallow, and lanolin; synthetic oils such as silicone oil, polyisobutene, fatty acid ester, and fatty acid glycerin; Waxes such as mole, candelilla wax and carnauba wax; hydrocarbons such as liquid paraffin, ceresin, microcrystalline wax and petrolatum; higher alcohols such as cetanol, stearyl alcohol and octyldodecanol; stearic acid, lauric acid, myristic acid and oleic acid Higher fatty acids such as silicone resin, silicone rubber, polyether-modified silicone, perfluoroether, and the like.

  Examples of the colorant include organic dyes such as Blue No. 1, Green No. 3, Red No. 202, Red No. 227, Yellow No. 4, chlorophyll, β-carotene, and natural dyes.

  Examples of the humectant include vitamins A, B, C, E or derivatives thereof, various amino acids, sodium hyaluronate, trimethylglycine, and the like.

  The polymer micelle is preferably obtained by condensation in an acidic or basic solution at the time of forming the polymer micelle from the viewpoint that the internal cross-linking is improved by the progress of condensation of the silanol portion which is the core portion of the polymer micelle.

<Method for producing polymer micelle>
The polymer micelle is prepared by, for example, stirring and suspending a PEG block copolymer in 200 mL of ultrapure water, and then putting the suspension in an ultrahigh pressure homogenizer (Microfluidizer, Microfluidics Corporation, Massachusetts, USA) at a pressure of 1,000 bar. , And can be produced under conditions of a flow rate of 150 mL / min. Alternatively, it can be produced by dialysis.

The polymer micelle of the present invention is excellent in dispersibility in water and organic solvents and can be stored in the stratum corneum with respect to the skin and the like, and is excellent in sustained release of the active ingredient and stability of the polymer micelle. In addition, by utilizing the crosslinking of the silanol moiety, stable long-term sustained release can be obtained by imparting or encapsulating functionality to the particles themselves by crosslinking the drug with the polymer.
The polymer micelle is a biocompatible polymer micelle useful in the transdermal field such as pharmaceuticals, quasi-drugs, and cosmetics. The pharmaceutical composition of the present invention described below, the formulation for transdermal absorption of the present invention, etc. Is preferably used.

(Cosmetics composition)
The cosmetic composition of the present invention contains the polymer micelle of the present invention, and further contains other components as necessary.
There is no restriction | limiting in particular as said other component, According to the objective, it can select suitably.
The dosage form of the cosmetic composition is not particularly limited as long as the polymer micelle does not disintegrate, and examples thereof include an ointment, a plaster, a coating agent, a spray, a lotion, a cream, an inhalant, and a nasal drop. It is done.

(Percutaneous absorption preparation)
The preparation for transdermal absorption of the present invention contains the polymer micelle of the present invention, and further contains other components as necessary.
There is no restriction | limiting in particular as said other component, According to the objective, it can select suitably.
The dosage form of the preparation for transdermal absorption is not particularly limited as long as the polymer micelle does not disintegrate, and examples thereof include an ointment, a plaster, a coating agent, a spray, a lotion, a cream, an inhalant, and a nasal drop. Can be mentioned.

  Examples of the present invention will be described below, but the present invention is not limited to these examples.

Example 1
Synthesis of polyethylene glycol-b-polychloromethylstyrene (PEG-b-PCMS) having a methoxy group at the α-terminal Heterobifunctional polyethylene glycol having a methoxy group at the α-terminal and a thiol group at the ω-terminal (PEG-SH) (Mn: 5000; 0.2 mmol, 1 g) and 10 mL of toluene were added to the reaction vessel.
Next, azobisisobutyronitrile (2 mmol, 164.2 mg) and chloromethylstyrene (10 mmol, 1.38 mL) were added to the reaction vessel, heated to 60 ° C., and stirred for 24 hours. The resulting reaction mixture was poured into hexane and a white precipitate formed. In order to remove the polychloromethylstyrene homopolymer, the obtained precipitate was washed three times with diethyl ether, which is a good solvent for the polychloroethylstyrene homopolymer, and then freeze-dried with benzene. A white powder (PEG-b-PCMS) was obtained.
According to size exclusion chromatography (SEC) measurement of the obtained PEG-b-PCMS, the number average molecular weight (Mn) = 9,504, the weight average molecular weight (Mw) = 1,173, and Mw / Mn = 1.386. It was.
^The result of measuring the ¹ H NMR spectrum is shown in FIG.
The size exclusion chromatography (SEC) measurement was performed using HPLC NANO SPACE manufactured by Shiseido Co., Ltd.
¹ H NMR spectra were performed using a Bruker AVANCE III 500 MHz N0MR spectrometer.

Synthesis of PEG-b-PTMSPAMS PEG-Block- (Trimethoxysilyl) propylamine-polymethylstyrene PEG-b-PCMS (Mn: 9504; 1 g) was added to the reaction vessel.
Next, 3-aminopropyltrimethoxysilane (2.5 g, 10 mmol) was dissolved in 10 mL of dimethyl sulfoxide (DMSO), added to the reaction vessel, and stirred at room temperature for 2 hours. After completion of the reaction, the reaction mixture was poured into 2-propanol cooled to −15 ° C., then re-precipitated three times with 2-propanol, and then lyophilized with benzene.

-Preparation of polymer micelles-
Put 2 mL of N, N-dimethylformamide (DMF) that passed through a 0.2 μm syringe filter into the screw and 10 mg of amphiphilic block polymer: PEG-b-PTMSPAMS, and apply heat using a dryer to completely remove the polymer. Dissolved in. The polymer DMF solution was transferred with a Pasteur pipette to a dialysis membrane with a molecular weight cut off of 3,500, which had been immersed in water in advance, and dialyzed against 2 L of distilled water. Distilled water was exchanged every few hours and dialyzed for 24 hours. Distilled water was added to the collected solution to make a total volume of 6.5 mL.
About the obtained polymer micelle, the average particle diameter and the zeta potential were measured using Malvern Zeta Sizer ZSP manufactured by Spectrosys. The results are shown in Table 3.

(Example 2) AIBN1
Synthesis of PEG-b-PTMSPMA PEG-Block- (3- (trimethylsilyl) propylmethacrylate) PEG-SH (Mn: 4,600; 0.2 mmol, 1 g) and 10 ml of toluene were added to the reaction vessel. Next, azobisisobutyronitrile (0.2 mmol, 16.42 mg) was added to the reaction vessel.
Next, 3- (trimethoxysilyl) propyl methacrylate (2.5 g, 10 mmol) was dissolved in 10 mL of toluene, added to the reaction vessel, and stirred at room temperature for 2 hours.
^The result of measuring the ¹ H NMR spectrum is shown in FIG.
A TEM image (40,000 times) of the obtained PEG-b-PCMS block copolymer is shown in FIG.

-Preparation of polymer micelles-
Put 2 mL of N, N-dimethylformamide (DMF) through a 0.2 μm syringe filter and 10 mg of amphiphilic block polymer polymer: PEG-b-PTMSPAMS through a screw, apply heat using a dryer, and completely remove the polymer. Dissolved.
The polymer DMF solution was transferred with a Pasteur pipette to a dialysis membrane with a molecular weight cut off of 3,500, which had been immersed in water in advance, and dialyzed against 2 L of distilled water. Distilled water was exchanged every few hours and dialyzed for 24 hours.
About the obtained polymer micelle, the average particle diameter and the zeta potential were measured in the same manner as in Example 1. The results are shown in Table 3.

(Example 3) AIBN10
A polymer was synthesized and polymer micelles were prepared in the same manner as in Example 2, except that the azobisisobutyronitrile (2 mmol, 164.2 mg) was used.
^The result of measuring the ¹ H NMR spectrum is shown in FIG.
About the obtained polymer micelle, the average particle diameter and the zeta potential were measured in the same manner as in Example 1. The results are shown in Table 3.

(Example 4) AIBN100
A polymer was synthesized and polymer micelles were prepared in the same manner as in Example 2 except that the azobisisobutyronitrile (20 mmol, 1,642 mg) was used.
^The result of measuring the ¹ H NMR spectrum is shown in FIG.
About the obtained polymer micelle, the average particle diameter and the zeta potential were measured in the same manner as in Example 1. The results are shown in Table 3.
TEM images of the obtained polymer micelles after dispersion of the collodion film are shown in FIG. 6A (10,000 times), FIG. 6B (50,000 times), and FIG. 6C (50,000 times).

(Example 5)
-Formation of polymer micelles-
In Example 1, when forming a polymer micelle, an appropriate amount of triethylamine was dropped into 2 L of distilled water, dialysis was performed after confirming that it became alkaline with pH test paper, Polymer micelles were formed in the same manner as in Example 1 except that ordinary distilled water was used.
About the obtained polymer micelle, the average particle diameter and the zeta potential were measured in the same manner as in Example 1. The results are shown in Table 3.

(Example 6)
In Example 1, when polymer micelles were formed, the same procedure as in Example 1 was conducted except that 0.4 mL of ethyl tetraethyl orthosilicate (TEOS) was added to the 5 mL dialysis membrane in the dialysis membrane. Thus, polymer micelles were formed.
About the obtained polymer micelle, the average particle diameter and the zeta potential were measured in the same manner as in Example 1. The results are shown in Table 3.

(Example 7)
In Example 1, in the same manner as in Example 6, when forming a polymer micelle, 0.4 mL of ethyl tetraethyl orthosilicate (TEOS) was added to a 5 mL dialysis membrane in the dialysis membrane, In the same manner as in Example 5, polymer micelles were formed in the same manner as in Example 1 except that TEA (triethylamine) was added to the outer layer.
About the obtained polymer micelle, the average particle diameter and the zeta potential were measured in the same manner as in Example 1. The results are shown in Table 3.

(Example 8)
-Method of linking drug to polymer micelle-
In Example 1, polymer micelles were formed in the same manner as in Example 1 except that an appropriate amount of rhodamine B isothiocyanate was added to the dialysis membrane when forming the polymer micelles.
About the obtained polymer micelle, the average particle diameter and the zeta potential were measured in the same manner as in Example 1. The results are shown in Table 3.

Example 9
-Method of encapsulating drug in polymer micelle-
In Example 1, polymer micelles were formed in the same manner as in Example 1 except that an appropriate amount of rhodamine B was added to the dialysis membrane when forming polymer micelles.
About the obtained polymer micelle, the average particle diameter and the zeta potential were measured in the same manner as in Example 1. The results are shown in Table 3.

(Comparative Example 1)
<Liposome production>
(1) The lipid concentration and volume to be prepared were determined, the required amount of the lipid stock solution was calculated, and a vial of an appropriate size was prepared.
(2) The lipid chloroform solution was measured with a pipette or a microsyringe and transferred to a vial.
(3) Chloroform is blown off with a nitrogen gas spraying device. Chloroform evaporated in a few minutes.
(4) In order to completely evaporate residual chloroform, it was left in a vacuum desiccator for 1 hour or more.
(5) Distilled water was put in a vial, rhodamine B was added, and liposomes were prepared by ultrasonic waves.
(6) The vial was held by hand so that the liquid level in the vial was lower than the liquid level of the ultrasonic water tank, and ultrasonic waves were applied for about 30 seconds with the maximum ultrasonic output. Immediately after removal, the mixture was vigorously stirred with a test tube mixer. This operation was repeated 3 to 4 times. As a result, the lipid film peeled off the wall surface, and liposomes were generated.
About the obtained liposome, it carried out similarly to Example 1, and measured the average particle diameter and the zeta potential. The results are shown in Table 4.

(Comparative Example 2)
<Synthesis of mPEG-PLGA>
Micelles were produced using Sigma-aldrich m-PEG-PLGA (5,000 Da-10,000 Da) 76513 (manufactured by Sigma-aldrich).
M-PEG-PLGA was dissolved in dichloromethane so as to be 5% (w / v).
Further, n-butanol was used as a solvent with rhodamine B, and heated in the presence of p-toluenesulfonic acid. Thereafter, a fat-soluble fluorescent dye to which sodium p-toluenesulfonate was added during the extraction of the treatment was synthesized and dissolved in dichloromethane so as to be 2.5% (w / v). It was mixed with 1 mL and 0.1 mL respectively, the scintillation vial was filled with 20 mL of distilled water, and the mixed solution was added dropwise while stirring at a speed of 2,000 rpm using an overhead stirrer, and the mixture was stirred for 1 hour or more. Then, it filtered through the PVDF filter with an average hole diameter of 0.45 micrometer.
About the obtained mPEG-PLGA, it carried out similarly to Example 1, and measured the average particle diameter and the zeta potential. The results are shown in Table 4.

  Next, various properties of the obtained polymer and polymer micelle were evaluated as follows. The results are shown in Tables 3 and 4.

<Electrophoretic test in vitro>
As a model dispersion, an aqueous dispersion of liposomes encapsulating rhodamine B was prepared.
As the current generator, a DC voltage / current source was used, and ADC 6242 was used.
It was connected to the lead electrode part produced on the urethane sheet, and an electric current was passed.
The agarose gel used agarose KANTO of Kanto Chemical Co., Inc. Dilution with 1 × TAE buffer produced a gel with a final concentration of 3 mass%.
A silver / silver chloride electrode patch 1 cm long × 1 cm wide was placed on the negative electrode side in a state of being affixed to an agarose gel, and a current of 0.45 mA / cm ² was passed at 20 ° C.
The distance between the plus electrode and the minus electrode was 3 cm apart.
Then, the cross section on the negative electrode side of the agarose gel was cut, and the distance permeated from the negative side to the positive side was observed visually or using a fluorescence microscope, and evaluated according to the following criteria. The moving distance is the moving distance of the aqueous dispersion of liposomes encapsulating rhodamine B, which is a model dispersion.
[Evaluation criteria]
1: The travel distance of Rhodamine B after 120 minutes has passed is 2 cm or more 2: The travel distance of Rhodamine B after 120 minutes has passed is 1.5 cm or more and less than 2 cm 3: Rhodamine B after 120 minutes have passed The moving distance of 1 cm or more and less than 1.5 cm 4: The moving distance of Rhodamine B after 120 minutes has passed is 0 cm or more and less than 1 cm

-Experimental conditions-
・ Agarose gel made with 3% (final concentration) TAE buffer ・ Amount added: 50 μm
・ Current: 2mA × 2hr
・ Zeta potential drug set: minus

* Polystyrene (PS) particles (Fluoresbrite YG Carboxylate Microspheres 0.05 μm, manufactured by Polysciences) were not electrophoresed.

It was found that the polymer micelle of Example 1 of the present invention migrates in a wide range as compared with calcein.
In the case of polystyrene particles, even when electricity is applied, they do not migrate and remain stored, while the polymer micelles of the present invention can behave widely.
In addition, small molecules such as calcein and rhodamine B exhibit a behavior that migrates widely apart from the movement distance.

<First disintegration test>
1.5 mL of each sample was added to a 2 mL tube. Next, the platinum electrode was inserted into the tube using a direct current generator (manufactured by ADC, 6421A), and current was applied for 1 hour. A constant current of 5 mA was passed. The degree of cloudiness was visually observed and evaluated according to the following criteria. The results are shown in Table 1 and FIG. In addition, in FIG. 7, the result of Examples 1-5 was shown as 1-5. Only particle size data that could be measured are shown in Table 1.
[Evaluation criteria]
1: No turbidity 2: Slight turbidity 3: White turbidity 4: Severe turbidity 5: Severe turbidity and agglomeration

* In Table 3, “-” means unmeasured.

* In Table 4, “-” means unmeasured.

<Second disintegration test>
-Disintegration-
1.5 mL of each sample of Comparative Example 2, Example 1, and Example 5 was added to a 2 mL tube. Next, the platinum electrode was inserted into the tube using a direct current generator (manufactured by ADC, 6421A), and current was applied for 1 hour. A constant current of 1 mA was applied. Then, DLS measurement was performed and the average particle diameter was measured. The results are shown in Table 5.
In addition, the result of the 2nd disintegration test of Example 1 and Example 5 was shown in FIG.

<Sustained release test>
At the time of polymer micelle production, 1 mL of ultrapurely diluted rhodamine B (4 μM) was added to the dialysis membrane, and rhodamine B was included during micelle formation. After one day, lyophilization was performed overnight. After drying, it was dissolved ultrapure to 10 mg / mL, and micelles were formed again. The dialysis membrane in which the solution was put into the dialysis membrane was put into 37 ° PBS (pH 7) as an outer layer and stirred. The outer layer was collected every other day, and the amount of fluorescence of rhodamine B leaked into the outer layer was measured.
A solution obtained by diluting Rhodamine B (4 μM) 10 times or 100 times with ultrapure water was evaluated as a comparison. The results are shown in FIG.
From the results of FIG. 9, it was found that the greater the amount of rhodamine B encapsulated, the more sustained release, and the smaller the amount, the less sustained release. It was found that the amount of sustained release can be controlled by the amount of inclusion.

<Drug release test>
3 mL of each sample was added to the dialysis membrane, and dialyzed against 3 L of distilled water. The water in the outer layer was collected every other day and measured for fluorescence. Distilled water was changed daily. The fluorescence intensity was measured using a fluorescence spectrometer (manufactured by HITACHI, Fluorescence Spectrophotometer F-2500).
The aim of TEOS addition (added into the dialysis membrane) is to promote internal crosslinking.
The aim of adding TEA (added to the outer layer liquid) is to promote internal crosslinking.
The results of the drug release test are shown in FIG.
From the results of FIG. 10, the ease of promoting sustained release was RhB>RhB-TEOS-TES>RhB-TEOS>RhB-iso>RhB-iso-TEOS-TES> RhB-iso-TEOS.
Although the effect of addition of TEA was not observed, it was found that the controlled release can be controlled by promoting the crosslinking.

Aspects of the present invention are as follows, for example.
<1> A block copolymer represented by the following general formula (I).
[General Formula (I)]
However, in the said general formula (I), A represents an unsubstituted or substituted C1-C12 alkoxy group, and the substituent of the alkoxy group in the case of being substituted is a formyl group and R < ₁ > R < ₂ >. CH— group (wherein R ₁ and R ₂ are each independently an alkoxy group having 1 to 4 carbon atoms, or R ₁ and R ₂ together form —OCH ₂ CH ₂ O—, —O ( CH _{2) 3} O-, or represents any of _{-O (CH 2) 4 O-)} .
L ₁ is a single bond, — (CH ₂ ) _c S—, —CO (CH ₂ ) _c S—, — (CH ₂ ) _c NH—, — (CH ₂ ) _c CO— (where c is 1 to And a linking group selected from —CO—, —COO—, and —CONH—.
R represents one selected from a single bond, — (CH ₂ ) _p —, and — (CH ₂ ) _p CH— (p represents an integer of 1 to 10).
X is methylimino (—CH ₂ NH—), methyliminomethyl (—CH ₂ NHCH ₂ —), methyloxy (—CH ₂ O—), methyloxymethyl (—CH ₂ OCH ₂ —), methyl ester (— CH ₂ OCO-), methyl ester methyl _{(-CH 2} OCOCH 2 -), and represents any of substituents represented by the following structural formulas.
m is 20 to 10,000.
n is 3 to 1,000.
<2> In the above general formula (I), L ₁ is —CH ₂ S—, X is a substituent represented by the following structural formula, and R is —CH ₂ CH—. It is a block copolymer.
<3> A polymer micelle comprising the block copolymer according to any one of <1> to <2>.
<4> The block copolymer represented by the general formula (I) has a core / shell structure, and the shell portion is A— (CH ₂ CH ₂ O) _m — (where A and m are the above general formulas). (It represents the same meaning as (I).) The polymer micelle according to <3>.
<5> The polymer micelle according to any one of <3> to <4>, wherein the average particle diameter is 1 nm or more and 1 μm.
<6> The polymer micelle according to any one of <3> to <5>, wherein an active ingredient is included in the polymer micelle.
<7> The polymer micelle according to any one of <3> to <6> obtained by condensation in an acidic or basic solution at the time of forming the polymer micelle.
<8> A cosmetic composition comprising the polymer micelle according to any one of <3> to <7>.
<9> A percutaneous absorption preparation comprising the polymer micelle according to any one of <3> to <7>.

  The block copolymer according to any one of <1> to <2>, the polymer micelle according to any one of <3> to <7>, the cosmetic composition according to <8>, and the According to the preparation for transdermal absorption described in <9>, the conventional problems can be solved and the object of the present invention can be achieved.

Japanese Patent No. 5183378 Japanese Patent No. 6026339

Claims (9)

A block copolymer represented by the following general formula (I):
[General Formula (I)]
However, in the said general formula (I), A represents an unsubstituted or substituted C1-C12 alkoxy group, and the substituent of the alkoxy group in the case of being substituted is a formyl group and R < ₁ > R < ₂ >. CH— group (wherein R ₁ and R ₂ are each independently an alkoxy group having 1 to 4 carbon atoms, or R ₁ and R ₂ together form —OCH ₂ CH ₂ O—, —O ( CH _{2) 3} O-, or represents any of _{-O (CH 2) 4 O-)} .
L ₁ is a single bond, — (CH ₂ ) _c S—, —CO (CH ₂ ) _c S—, — (CH ₂ ) _c NH—, — (CH ₂ ) _c CO— (where c is 1 to And a linking group selected from —CO—, —COO—, and —CONH—.
R represents one selected from a single bond, — (CH ₂ ) _p —, and — (CH ₂ ) _p CH— (p represents an integer of 1 to 10).
X is methylimino (—CH ₂ NH—), methyliminomethyl (—CH ₂ NHCH ₂ —), methyloxy (—CH ₂ O—), methyloxymethyl (—CH ₂ OCH ₂ —), methyl ester (— CH ₂ OCO-), methyl ester methyl _{(-CH 2} OCOCH 2 -), and represents any of substituents represented by the following structural formulas.
m is 20 to 10,000.
n is 3 to 1,000.
_2. The block copolymer according to claim 1, wherein, in the general formula (I), L ₁ is —CH ₂ S—, X is a substituent represented by the following structural formula, and R is —CH ₂ CH—. .
  A polymer micelle comprising the block copolymer according to claim 1. The block copolymer represented by the general formula (I) has a core-shell structure, and the shell portion is A- (CH ₂ CH ₂ O) _m- (where A and m are the general formula (I)). The polymer micelle according to claim 3, which represents the same meaning as   The polymer micelle according to any one of claims 3 to 4, having an average particle diameter of 1 nm or more and 1 µm.   The polymer micelle according to any one of claims 3 to 5, wherein an active ingredient is encapsulated in the polymer micelle.   The polymer micelle according to any one of claims 3 to 6, obtained by condensation in an acidic or basic solution at the time of polymer micelle formation.   A cosmetic composition comprising the polymer micelle according to any one of claims 3 to 7. A preparation for transdermal absorption, comprising the polymer micelle according to any one of claims 3 to 7.