KR20160122051A - Soluble microneedle for delivery of peptides regulating neurotransmitters to neuronal cell - Google Patents

Abstract

The present invention relates to a microneedle containing peptide capable of regulating release of neurotransmitter. The present invention further relates to a system for injecting peptide into skin, ensuring increased skin wrinkle amelioration effects by applying the microneedle to skin. According to the present invention, the microneedle shows excellent effectiveness in wrinkle amelioration.

Description

{Soluble microneedle for delivery of peptides to neurotransmitters to neuronal cells}

The present invention relates to a soluble micro needle, a method for administering a skin to an active ingredient using the same, and a method for producing the soluble micro needle.

Various skin delivery drugs have been developed for the improvement of skin diseases and skin moisturizing effects, and studies using peptides have been actively conducted among them.

Delivery of the drug through the skin has been used in a variety of fields and forms due to its ease of use. Drugs that pass through the skin are mainly intended to be delivered to the systemic circulation through the skin. In addition, medicines such as an atopy treatment agent, a whitening agent, or a cosmetic agent for improving wrinkles may be used for the purpose of delivering the drug to the skin's own organs . Despite this convenience and functionality, there are many difficulties in delivering the drug through the skin in the structure of the skin, and it is not easy to develop a drug that passes through the skin. The stratum corneum of the skin is composed of a brick structure composed of keratin-rich keratinocytes and a lipid-filled mortar such as ceramide, fatty acid, or wax. Structure. This structure acts as a barrier and has very low permeability to materials. Only low molecular weight components of 500 Da or less can be delivered into the skin by the diffusion method, and only substances having superior lipid affinity can pass through the skin.

On the other hand, the release of neurotransmitters is caused by the formation of a passage between the two boundaries after fusion of the synaptic vesicle and the presynaptic membrane located at the nerve endings containing the neurotransmitter. SNARE (N-ethylmaleimide-sensitive fusion protein) attachment protein receptor (SNARE) protein, a three-protein complex consisting of the vaprotein VAMP (synaptobrevin) and the plasma membrane-binding proteins Syntaxin 1a and SNAP-25, And synaptic membrane fusion. Here, the neurotransmitter release pathway is opened by membrane fusion between the synaptic vesicle and the synaptic membrane. The synaptic 1a protein attached to the target membrane and the t-SNARE and vesicle (complex of two SNAP-25 proteins) vesicle) of the v-SNARE. During the membrane fusion, rearrangement of the lipid bilayer occurs. Since the biomembranes push strongly against each other, these membranes are supposed to overcome the repulsion between the membranes by giving them strong force from the outside without being spontaneously fused. It is known that SNARE protein provides this force. Thus, the formation of the SNARE complex is a key phenomenon of exocytosis involving the release of neurotransmitters.

In recent years, there has been development of a peptide having a similar role to Botox, which reduces muscle contraction without toxic side effects in a non-invasive manner. These peptides temporarily block the secretion of neurotransmitters such as acetylcholine, which is responsible for the contraction of muscle cells, and play a role in relieving wrinkles.

However, in spite of the development of a peptide having a role similar to that of Botox, there is a problem in that the effect is insufficient when applied to skin as a formulation such as cream, and the original efficacy can not be exhibited.

Therefore, a problem to be solved by the present invention is to provide a method of controlling the release of neurotransmitters to exert the efficacy of a peptide having a wrinkle-reducing effect. That is, it is intended to provide a method for improving the wrinkle reducing effect of skin by applying a micro needle including a peptide that inhibits receptor-ligand binding of a neurotransmitter to the skin.

The present invention relates to a skin administration system capable of regulating the release of a neurotransmitter and delivering a substance with skin wrinkle reducing effect into the skin and a method of manufacturing such a system, which system can be used to control the release of neurotransmitters into the skin ≪ / RTI > peptide.

In order to solve the above problems, the present invention provides a micro needle comprising a peptide. Preferably, the present invention provides a microneedle containing a peptide and having a wrinkle-improving effect. The material forming the micro needle can be dissolved or biodegraded in the skin, and the micro needle is dissolved or collapsed when the skin is applied, thereby releasing the peptide, and the peptide can be stably delivered to the skin.

In order to solve the above problems, the present invention provides a microneedle containing a substance having a wrinkle-reducing effect to regulate the release of neurotransmitter, more preferably a peptide which inhibits receptor-ligand binding of a neurotransmitter / RTI > The material forming the micro needle is dissolved in the skin to dissolve or collapse the micro needle at the time of application of the skin to the micro needle, thereby controlling the discharge of the neurotransmitter so that the material having the wrinkle reducing effect can be stably delivered to the skin.

The present inventors have studied various administration systems and it has not been easy to envisage any system capable of enhancing the effect of a substance having a wrinkle-reducing effect, as mentioned above, by controlling neurotransmitter release. That is, peptides and derivatives thereof capable of controlling the release of acetylcholine and the like, which are neurotransmitters, may add an alkyl chain or the like to increase the lipophilicity. In this case, the problem that the molecular weight is increased and the skin permeability is lowered Occurs. That is, instead of increasing the lipid affinity, the molecular weight is increased, so that it is difficult to pass through the stratum corneum to achieve the original purpose.

It has not been easy in the industry to envision a peptide delivery system that can enhance the lipid affinity of low-lipid-affinity peptides and allow the skin to permeate stably even when the molecular weight increases. The inventors of the present invention have found that it is possible to control the release of neurotransmitters in the soluble micro-needle after various efforts, and to prevent wrinkles by effectively suppressing the transmission of neurotransmitters by containing a substance having a wrinkle- Respectively.

The material constituting the micro needle according to the present invention (the biocompatible material forming the structure of the micro needle) may be made of a material dissolvable in the skin, and a water-soluble polymer, a saccharide and the like may be used. The water-soluble polymer may be selected from natural, semi-synthetic and synthetic polymers. Natural polymers include gelatin, pectin, dextran, hyaluronic acid or its salts, collagen, agar, Arabic Gum, , Sorghum such as Xanthan Gum, Acacia Gum, Karaya Gum, Traganth Gum and Guar Gum, carrageenic acid, alginic acid, alginate Sodium alginate), and the like can be used.

Examples of the semi-synthetic polymer include methyl cellulose, ethyl cellulose, hydroxy ethyl ethyl cellulose, sodium carboxy methyl cellulose, soluble starch, Pullulan, Dextrine, Carboxy methyl starch, Dialdehyde starch and the like can be used. Examples of the synthetic polymer include polyvinyl alcohol, polyvinyl pyrrolidone, polyvinyl methacrylate, polyacrylic acid and salt thereof, polyethylene oxide oxide, polypropylene oxide, polyethylene oxide and polypropylene oxide copolymer, carboxy-containing acrylic resin, carboxy-containing polyester resin, polyamide (polyamide) ), Water-soluble polyurethane, maltodextrin, and polydextrose, or a water-soluble synthetic polymer prepared by using a radical polymerizable monomer can be selected. The polymer synthesized by the radical polymerizable monomer may be a homopolymer of an ionic monomer and a nonionic monomer, and in some cases, a copolymer thereof may be used.

The sugar may be xylose, sucrose, maltose, lactose, trehalose or a mixture thereof.

Also, in addition to the water-soluble polymer, a material forming the structure of the micro needle can be a poly (lactide), a poly (glycolide), a poly (lactide-co-glycolide), a polyanhydride, biodegradable polymers such as polyorthoesters, polyetherester, polycaprolactone, polyesteramide, poly (butyric acid), poly (valeric acid), polyurethane or copolymers thereof ; And polyolefins such as polyacrylates, ethylene-vinyl acetate polymers, acrylic substituted cellulose acetates, non-degradable polyurethanes, polystyrenes, polyvinyl chlorides, polyvinyl fluorides, poly (vinylimidazoles), chlorosulphonate polyolefins, Polyethylene oxide or copolymers thereof may be used singly or in combination, but the present invention is not limited thereto.

Considering the skin penetration strength of the micro needles and the dissolution rate in the skin in general, it is preferable to use an oligosaccharide such as oligo-hyaluronic acid, sodium carboxymethyl cellulose (Na-CMC), and saccharide ) (More preferably Trehalose) is more preferable, and it is further preferable that the mixture is mixed up to glycerin to be described later. Preferably, the microneedle according to the present invention may further include a plasticizer, a surfactant, a preservative, an anti-inflammatory agent, etc. in addition to the above-mentioned components for forming the micro needle.

Examples of the plasticizer include polyols such as ethylene glycol, propylene glycol, dipropylene glycole, butylene glycol, and glycerine, Or may be used in combination.

In one embodiment of the present invention, the length of the microneedles is not limited to a specific size. However, since the peptide of the present invention which forms the SNARE complex through the horny layer is a muscle and a nerve cell under the subcutaneous fat located deep from the surface of the skin, the length of the micro needle is the most protruding of the micro needle, It is preferable that the length penetrating the skin is 250 占 퐉 to 900 占 퐉, more preferably 300 占 퐉 to 800 占 퐉, still more preferably 350 占 퐉 to 700 占 퐉.

The micro needle in the above range can easily reach the target site through the stratum corneum of the skin. Also, when the micro-needle of the present invention is within the above-mentioned range, it is preferable in terms of the structural stability of the peptide, the delivery efficiency to the target cell, and the stability against degradation by bio enzyme have. In one aspect, when the length of the micro needle is shorter than the length range, it may be difficult to reach the target site. If the length of the micro needle is longer than the range, the probability of being absorbed into the body is higher.

Preferably, the microneedles of the present invention contain 0.0001 to 20% by weight, preferably 0.001 to 10% by weight, of the peptides having the effect of controlling neurotransmitter release and having a wrinkle-reducing effect, based on the total weight of the micro needle. Even more preferably 0.1 to 5% by weight based on the total weight of the micro needle.

The micro-needle having excellent skin wrinkle reducing effect can be produced within the above content range, and the stability of the peptide contained in the micro needle can be excellent in the above content range, which is advantageous for attaining the object of the present invention.

The amount of the peptide of the present invention contained per micro needle of the present invention may be 0.0001 to 20 mg, preferably 0.001 to 10 mg. Even more preferably from 0.1 to 5 mg, based on the total weight of the micro needle.

The microneedles having the above content ranges can be administered once every two days to reduce the skin wrinkles of adult individuals (based on 50 kg).

Use once every 2 days rather than daily use maximizes the wrinkle-reducing effect while minimizing skin side effects. The above 2-day interval means administration on day 1, day 2 is rest, and administration on day 3.

The microneedles may contain a high content of peptide components that regulate neurotransmitter release, and may provide a polyphenol skin delivery system in which the amount delivered to the skin is much greater than that of a typical cosmetic composition.

The microneedles of the present invention may comprise up to 1% by weight of water relative to the total weight of the microneedles. When a peptide is included in a cosmetic composition, the peptide may be in contact with moisture to cause denaturation of the peptide. However, the micro-needle of the present invention has a very low moisture content and can be delivered into the skin without denaturation of the peptide component. In addition, since the amount of water contained in the micro needle is small, the possibility of contamination by microorganisms can be reduced.

A substance having a wrinkle-reducing effect which can control the neurotransmitter discharge which can be used in the present invention can prevent the neurotransmitter from being delivered to the receptor of the neurotransmitter to suppress the muscle contraction effect, It is a substance that can be. More specifically, in order to move the muscles, neurotransmitters must be transferred from neurons to muscle cells through the synapse. In order to secrete the neurotransmitter acetylcholine in the synapse, a SNARE complex is formed at the end of the neuron and released to the synapse. Botox, commonly known, breaks down the SNAP-forming component (SNAP-25), preventing acetylcholine from being released into the synapse. Botox-like peptides, on the other hand, have a structure similar to that of SNAP-25 (SNAP-25). SNAP-25 is involved in the formation of complexes and prevents the release of acetylcholine.

The peptides having the wrinkle-improving effect of the present invention can be widely known in the art and are not particularly limited to the types of peptides. Natural peptides as well as chemically synthesized peptides may be included, and preparations containing peptides as well as pure peptides may be used as the peptides contained in the micro needle of the present invention. Peptide derivatives of peptides known to have wrinkle-reducing effects can also be included within the scope of the present invention.

For example, the peptide derivatives may be selected from the group consisting of X50 Myocept ™ from Infinitec, (Pal) -Tyr-Pro-Trp-The-Gln-Arg ([Pal] DdPQRR), Palmitoyl-hexapeptide-52 -Phe ([Pal] YPWTQRF), Argireline (TM), Acetyl-EEMQRR), GABA (gamma-amino butyric acid), botulinum toxin, ≪ / RTI >

Also included are peptides having the formula 1, isomers thereof, racemic compounds, their cosmetic or pharmaceutically acceptable salts, and the like.

[Chemical Formula 1]

R ₁ -AA-R ₂

Wherein AA may be an amino acid sequence comprising 3-40 amino acids,

R ₁ can be any one selected from H or C ₃ to C ₂₄ alkyl, aryl, or acyl groups;

R ₁ of the peptide of formula 1 may be an acyl group having 3 to 24 carbon atoms and the acyl group may be saturated or unsaturated and may be linear, branched or cyclic ).

More preferably, R ₁ is an acyl group of the formula CH ₃ - (CH ₂ ) m -CO-, and m may be any one selected from 1 to 22.

More preferably, R < ₁ > may be a polyethylene glycol polymer having a molecular weight of 200 to 35,000 Da.

Preferably the AA is MAEDADMRNELEEMQRRADQL, ADESLESTRRMLQLVEESKDAGI, ELEEMQRRADQLA, ELEEMQRRADQL, ELEEMQRRADQ, ELEEMQRRAD, ELEEMQRRA, ELEEMQRR, LEEMQRRADQL, LEEMQRRADQ, LEEMQRRAD, LEEMQRRA, LEEMQRR, EEMQRRADQL, EEMQRRADQ, EEMQRRAD, EEMQRRA, EEMQRR, LESTRRMLQLVEE, NKDMKEAEKNLT, KNLTDL, IMEKADSNKTRIDEANQRATKMLGSG, SNKTRIDEANQRATKMLGSG , TRIDEANQRATKMLGSG, DEANQRATKMLGSG, NQRATKMLGSG and QRATKMLGSG.

Wherein R ₂ of the peptide of Formula 1 is a C ₁ to C ₂₄ aliphatic or cyclic group and the C ₁ to C ₂₄ aliphatic or cyclic group is unsubstituted or substituted with one or more substituents selected from the group consisting of An amino group, a hydroxyl group or a thiol group.

The AA may also include an amino acid sequence derived from the amino and carboxy domain of SNAP-25 protein.

For example, the substituents of R 1 and R 2 in the above formula (1) may be those known from US 2010-0021510 A1, and the entire contents of US 2010-0021510 A1 are incorporated herein. The compounds of the above formula (1) can be used in the examples disclosed in US 2010-0021510 A1.

Preferably, the peptide contained in the micro needle according to an embodiment of the present invention is selected from the group consisting of Palmitoyl-hexapeptide-52 ([Pal] -Asp-Asp-Met-Gln-Arg- -18 ([Pal] -Tyr-Pro-Trp-The-Gln-Arg-Phe ([Pal] YPWTQRF).

When the above-mentioned Palmitoyl-DDMQRR and Palmitoyl-YPWTQRF are used together, they can be more preferably used in terms of compatibility with the micro needle. In addition, the microneedles containing Palmitoyl-DDMQRR and Palmitoyl-YPWTQRF directly inhibit the formation of SNARE and block the inflow of Ca ^{2 +} ions into the nerve cells, thereby indirectly inhibiting the formation of SNARE. Excellent skin wrinkle reduction effect can be obtained.

Examples of the neurotransmitter include dopamine, serotonin, histamine, acetylcholine, adrenaline, noradrenaline, gamma amino butyric acid (GABA), L-glutamic acid, glycine and the like.

The present invention also provides a microneedle patch system for administration (delivery) of a neurotransmitter release modulating substance to which the above-described microneedles are attached.

The microneedles of the present invention can be produced by a method for producing conventional soluble micro-needles in the art, and the manufacturing method is not particularly limited.

The present invention also provides a skin administration method of a neurotransmitter release controlling substance with improved skin permeation efficiency of a peptide using a micro needle according to the present invention.

The present invention also provides a wrinkle-improving use of a micro needle which contains a substance having a wrinkle-reducing effect to control the release of the neurotransmitter.

In accordance with one embodiment of the present invention, the present invention provides a microneedle containing a substance that inhibits SNARE complex formation.

According to an embodiment of the present invention, there is provided a skin administration method of a neurotransmitter release controlling substance having a skin wrinkle reducing effect by inhibiting SNARE complex formation using the micro needle.

The micro needle according to one embodiment of the present invention may coat a part of the neurotransmitter emission controlling substance having a skin wrinkle reducing effect on the surface of the microparticles by inhibiting formation of the SNARE complex or may be included in microparticles. In other words, some peptides can be coated on the microparticle surface or transferred into the microparticles in order to effectively transfer the desired amount to the target site while stabilizing the structure of the peptides included in the micro needle. In one embodiment of the invention, the peptide coated microparticles can be dissolved in the skin and comprise at least one polymeric material or a polymer with amphipathic properties, wherein the polymer is not limited to hydroxyethylcellulose (HEC), hydroxypropylmethylcellulose (HPMC), hydroxypropylcellulose (HPC), methylcellulose (MC), hydroxyethylmethylcellulose (HEMC), or ethylhydroxy-ethylcellulose (EHEC) Derivatives as well as pluronic.

In another embodiment, the peptide-coated microparticles are selected from the group consisting of poly (vinyl alcohol), poly (ethylene oxide), poly (2-hydroxyethyl methacrylate), poly (n-vinyl pyrrolidone), polyethylene glycol Its mixtures and similar polymers. Stabilizers necessary for stabilization of other coated peptides may include stabilizers commonly used in the art, and may include, for example, polysaccharides, non-reducing sugars, and the like.

In the present invention, the substance that forms microparticles together with the peptides should be a substance capable of stably trapping the peptide without causing structural modification of the peptide during the production of the micro needle. Particularly, the material forming the microparticles can form a hydrophobic core, so that it can provide stability without structural modification of the peptide. Particularly, in consideration of the amino acid structure of Pal-52 and Pal-18 used as the preferred examples of the present invention, it is preferable to use a substance forming a hydrophobic core.

As the material for forming the microparticles, a polymer capable of forming a hydrophobic core may be used, and a polymer such as a poly (lactide), a poly (glycolide), a poly (lactide-co-glycolide), a polyanhydride, (Polyacrylic acid), poly (vinylidene chloride), poly (vinylidene chloride), poly (vinylidene fluoride), poly (vinylidene fluoride) A biodegradable polymer such as a copolymer thereof; And polyolefins such as polyacrylates, ethylene-vinyl acetate polymers, acrylic substituted cellulose acetates, non-degradable polyurethanes, polystyrenes, polyvinyl chlorides, polyvinyl fluorides, poly (vinylimidazoles), chlorosulphonate polyolefins, Polyethylene oxide or copolymers thereof may be used singly or in combination, but the present invention is not limited thereto.

Considering the ability to inhibit peptides, particularly SNARE complex formation and release in the skin, the hydrophobic core-forming polymers can be poly (lactide), poly (glycolide), and poly (lactide-co-glycolide) ) And mono-methoxypolyethylene glycol-polycaprolactone (MPEG-PCL).

Such a microparticle may be of a matrix type or a reservoir type so long as the object of the present invention can be achieved.

Microparticles that can be used in the present invention can be prepared by a variety of methods well known in the art. For example, microparticles usable in the present invention can be prepared by using a solvent exchange method, a solvent evaporation method, a membrane permeation method, a spray drying method, Can be manufactured. For example, the methods described in the Journal of Controlled Release 70 (2001) 1-20 and International Journal of PharmTech Research, 3 (2011) 1242-1254 may be used. Preferably by the general emulsification and solvent evaporation method.

Preferably, the diameter of the microparticles according to the present invention is 0.01 to 10 mu m. If the particle size is more than 10 μm, the strength of the needle becomes weaker when it is impregnated into the micro needle, which makes skin penetration difficult. The diameter of the microparticles according to the present invention is measured by a laser light scattering (LLS) method, for example, using a Zetasizer 2000 ^™ from Malvern.

Preferably, the microparticles included in the micro needle of the present invention may be 30 to 60% by weight based on the total weight of the peptide contained in the micro needle. When the content is in the above range, the effect of improving the wrinkles of the skin appears quickly, but the duration may be prolonged.

The micro-particles can be coated on the surface of the microparticles, or the micro-needle containing the peptide in the micro-particles can be prepared and then dried on the micro-needle by various means. Various temperature and humidity levels may be used to dry the microneedles according to one embodiment of the present invention. In addition, the micro needle can remove water by various means.

The present invention provides a micro needle having an effect of improving skin wrinkles.

The present invention provides a skin micro-needle which can improve the skin permeation efficiency of peptides having a wrinkle-improving effect.

The present invention provides a method for improving skin wrinkles by administering a peptide, which uses such a micro needle.

BRIEF DESCRIPTION OF THE DRAWINGS The accompanying drawings, which are incorporated in and constitute a part of the specification, illustrate exemplary embodiments of the invention and, together with the description of the invention, It should not be construed as limited.
FIG. 1 is a view showing an example of various methods for manufacturing a micro needle according to the present invention.
2 shows a Franz diffusion cell for evaluating drug release behavior of a micro needle according to the present invention.
FIG. 3 is a graph showing the skin permeation amount of a peptide contained in a cream and a peptide impregnated in a micro needle.
4 shows the effect of the micro needles of the present invention, Botox-like cosmetics, and Botox-like peptides in improving skin wrinkles.
Fig. 5 shows the effect of inhibiting muscle contraction of substances having SNARE Complex formation inhibitory effect.
Fig. 6 shows the results of confirming the wrinkle-improving effect of the peptide-containing cream and the peptide-containing micro needle.
7 is a graph showing a wrinkle reducing effect according to the length of the micro needle according to the present invention.

Hereinafter, embodiments of the present invention will be described in detail to facilitate understanding of the present invention. However, the embodiments according to the present invention can be modified into various other forms, and the scope of the present invention should not be construed as being limited to the following embodiments. Embodiments of the invention are provided to more fully describe the present invention to those skilled in the art.

Production Example 1. Peptide-coated and impregnated microparticle preparation

The peptides can be coated on the surface of microparticles using methoxy poly (ethylene glycol) -b-poly (caprolactone), mPEG-PCL) ≪ / RTI > First, a solution obtained by dissolving 10 g of mPEG-PCL (5 k-10 k mixture) in 40 mL of ethanol was mixed with 20 g of dipropylene dissolved in 4 g of peptide (2 g of palmitoyl hexapeptide-52 and 2 g of palmitoyl heptapeptide-18). The solution was slowly added to 100 mL of a 0.5% aqueous solution of polyvinyl alcohol while stirring. The ethanol solution was left to stir for a certain period of time to evaporate the ethanol solvent, and the remaining ethanol was removed by using a rotary evaporator to prepare a solution having a total peptide content of 4%.

Analysis by liquid chromatography revealed that palmitoyl hexapeptide-52 and palmitoyl heptapeptide-18 contents were 1.9%, respectively. As a result of particle size analyzer (Malvern Zetasizer 2000) ^TM , the average microparticle size was 120 nm .

Production Example 2. Production of Soluble Micro Needle

Soluble Micro Needle is manufactured by solution casting method. It is manufactured by casting solution into mold, filling solution into micro mold with vacuum or centrifuge, and then drying. In this case, common synthetic and natural water-soluble polymer is used for forming micro needle structure .

Manufacturing example  3. Peptides  And Peptides Micro-particle  Contained Micro needle  Produce

Microneedles containing peptides (Palmitoyl Hexapeptide-52 and Palmitoyl Heptapeptide-18) (added as a solution form) or peptides (Palmitoyl Hexapeptide-52 and Palmitoyl Heptapeptide-18) microparticles were prepared as shown in Table 1 below.

P-S MN P-MP MN oligo-HA 6.0 6 Na-CMC 6.0 6 Trehalose 10.0 10 Glycerin 5.0 5 HCO-40 0.2 0.2 Peptide-DPG solution (10%)
(Palmitoyl Hexapeptide-52 / Palmitoyl Heptapeptide-18: 1/1) 1.0 - Peptide microparticle (4%)
(Palmitoyl Hexapeptide-52 / Palmitoyl Heptapeptide-18: 1/1) - 2.5 water To 100 To 100

(1) soluble micro needles impregnated with peptides (P-S MN)

Specifically, soluble micro-needles impregnated with a peptide were prepared as follows. Hyaluronic acid (Hyaluronic acid), Na-CMC (sodium carboxymethyl cellulose) and Trehalose were dissolved in purified water, and then Glycerin, HCO-40 and peptide-DPG solution were added to prepare solutions. The prepared peptide solution was cast in a silicon microneedle mold, and the solution was filled in a fine mold by centrifugation at 3000 rpm for 10 minutes. After the solution was filled, it was dried in a drying oven (70 ° C) for 3 hours, and the micro needle was separated from the silicone mold using an adhesive film.

(2) soluble micro-needle (P-MP MN) impregnated with peptide microparticles

Specifically, soluble micro-needles impregnated with peptide microparticles were prepared as follows. Hyaluronic acid (Hyaluronic acid), Na-CMC (sodium carboxymethyl cellulose) and Trehalose were dissolved in purified water, and then Glycerin, HCO-40 and peptide microparticles (peptide 4%) were added to prepare solutions. The prepared solution was cast into a silicone microneedle mold, and the solution was filled in a fine mold by centrifugation at 3000 rpm for 10 minutes. After the solution was filled, it was dried in a drying oven (70 ° C) for 3 hours, and the micro needle was separated from the silicone mold using an adhesive film. The manufacturing process is shown in Fig.

Preparation Example 4. Preparation of microneedles of various lengths

To determine the optimal length of the optimal micro needle, 150, 250, 300, 400, 500 ㎛ microneedle molds with a length of 150 ㎛ or more penetrating the skin based on the protruding and pointed end of the micro needle were manufactured and used After the preparation of soluble micro needle, the release behavior and the wrinkle - reducing effect of the drug were compared. The results are shown in Fig. As the length of the microneedles increased up to 400 ㎛, the skin permeability increased and the wrinkle reduction rate increased.

Production Example 5. Manufacturing of an emulsion type cream containing a peptide

In order to compare the skin permeation amount of the peptide impregnated with the micro needle, the peptides were compared in the oil in water cream of the common emulsifier type as a comparative example shown in Table 2 below.

ingredient Content wt (%) C14-22 Alcohol / C12-20 alkyl glucoside
(Mixture C14-22 alcohol: C12-20 alkyl glucoside = 80:20 weight ratio) 1.5 Glyceryl stearate, phage-100 stearate
(Mixture 50:50 weight ratio) 1.2
Glyceryl stearate 0.9 Cetearyl alcohol 1.5 Polyglyceryl-3 methyl glucoside distearate 1.5 Hydrogenated polydecene 4.5 Cyclohexasiloxane 3.5 Carbomer 0.2 Tromethamine 0.2 glycerin 3.0 Dipropylene glycol 5.0 1,2-hexanediol 2.0 Palmitoyl hexapeptide-52 1.0 Palmitoyl heptapeptide-18 1.0 Purified water Remaining amount (To 100)

[Drug Release Behavior]

The release of peptides (palmitoyl hexapeptide-52 and palmitoyl heptapeptide-18) from the micro-needle prepared above was evaluated using a Franz diffusion cell equipped with pig skin (see Figure 2). As an acceptor solution, a PBS solution containing 30% by weight of DPG was used.

In other words, using Franz diffusion cell, peptide content of pig skin tissue and acceptor solution with time was measured by Liquid Chromatography.

The amount of peptide contained in the cream was 0.1 ㎍, which was insignificant, but the peptide impregnated in the micro needle was directly penetrated to the skin by the needles, The skin permeation rate was high.

These results are shown in Fig.

[Wrinkle improvement effect]

Peptide cream was treated daily for 4 weeks for wrinkles, and Microneedle for peptide application was treated twice a week for 4 weeks. Wrinkle improvement was confirmed by silicone replica and wrinkle image analysis method . (N = 20)

It was confirmed that the peptide was more effective than microneedle, which is more effective than peptides, in improving the wrinkle. These results are shown in Fig.

[Inhibition of muscle contraction]

C2C12 cells were cultured in DMEM medium supplemented with 10% fetal calf serum and 1% antibiotics. The neuroblasts were then co-cultured further in the same plate. When C2C12 cells began to contract, the number of contractions of C2C12 cells was measured for 30 seconds. After the medium was completely removed, the cells were washed three times with PBS, and the medium containing no calf serum and 50 ppm of peptide were added and reacted for 2 hours. Thereafter, the number of contractions of C2C12 cells was measured again for 30 seconds, and the degree of muscle contraction inhibition was confirmed. Peptide inhibited the release of neurotransmitters from neurons and decreased the number of contractions of C2C12 cells. Among them, X50 (palmitoyl hexapeptide-52, palmitoyl heptapeptide-18) was relatively higher in inhibiting the release of neurotransmitters than BLP (BoNT-L peptide) or Dex (Dextrane).

Claims (17)

Micro needles containing peptides that regulate the release of neurotransmitters. The method of claim 1, wherein the peptide that modulates the release of the neurotransmitter is selected from the group consisting of botulinum toxin, palmitoyl-DDMQRR, palmitoyl-YPWTQRF, acetyl-EEMQRR, And the micro needle. 2. The method of claim 1, wherein the peptide that modulates the release of the neurotransmitter
Claims 1. A micro needle, characterized in that it is a peptide having the formula (1), an isomer thereof, a racemic compound, or a cosmetic or pharmaceutically acceptable salt thereof.
[Chemical Formula 1]
R ₁ -AA-R ₂
Wherein AA is MAEDADMRNELEEMQRRADQL, ADESLESTRRMLQLVEESKDAGI, ELEEMQRRADQLA, ELEEMQRRADQL, ELEEMQRRADQ, ELEEMQRRAD, ELEEMQRRA, ELEEMQRR, LEEMQRRADQL, LEEMQRRADQ, LEEMQRRAD, LEEMQRRA, LEEMQRR, EEMQRRADQL, EEMQRRADQ, EEMQRRAD, EEMQRRA, EEMQRR, LESTRRMLQLVEE, NKDMKEAEKNLT, KNLTDL, IMEKADSNKTRIDEANQRATKMLGSG, SNKTRIDEANQRATKMLGSG, TRIDEANQRATKMLGSG , DEANQRATKMLGSG, NQRATKMLGSG and QRATKMLGSG, wherein the amino acid sequence is selected from the group consisting of SEQ ID NO:
Wherein R < ₁ > is any one selected from H or an alkyl, aryl, or acyl group having 3 to 24 carbon atoms,
Wherein R ₂ is a C ₁ to C ₂₄ aliphatic or cyclic group and the C ₁ to C ₂₄ aliphatic or cyclic group is unsubstituted or substituted with an amino group, Or a thiol group. The microneedle of claim 1, wherein the peptide that modulates the release of the neurotransmitter is Palmitoyl-DDMQRR, Palmitoyl-YPWTQRF, or a mixture thereof. The neurotransmitter according to claim 1, wherein the neurotransmitter is any one or more selected from the group consisting of dopamine, serotonin, histamine, acetylcholine, adrenaline, noradrenaline, gammaaminobutyric acid, L-glutamic acid, . The microneedle of claim 1, wherein the material forming the microneedle is dissolved in the skin. 7. The method of claim 6, wherein the material forming the micro needle is selected from the group consisting of Hyaluronic acid, sodium carboxymethyl cellulose, sodium carboxymethyl cellulose (Na-CMC), vinyl pyrrolidone-vinyl acetate copolymer, polyvinyl alcohol Polyvinyl alcohol, polyvinyl pyrrolidone, saccharides, or a mixture thereof. 7. The microneedle of claim 6, wherein the material forming the microneedle comprises hyaluronic acid, sodium carboxymethylcellulose and a saccharide. The micro needle according to claim 1, wherein the peptide contained in the micro needle is coated on the micro particle surface or contained in the micro particle. The method of claim 9, wherein the microparticles coated with the peptide on the surface are selected from the group consisting of poly (vinyl alcohol), poly (ethylene oxide), poly (2-hydroxyethyl methacrylate) Polyethylene glycol, and mixtures thereof. ≪ RTI ID = 0.0 > 11. < / RTI > 10. The method of claim 9, wherein the microparticles in which the peptide is incorporated are selected from the group consisting of poly (lactide), poly (glycolide), poly (lactide-co-glycolide), polyanhydride, polyorthoesters, poly (Methacrylic acid), poly (butyric acid), polyurethane, or copolymers thereof, such as, for example, polyesters, ether esters, polycaprolactones, mono-methoxypolyethylene glycol-polycaprolactone At least one biodegradable polymer selected from the group consisting of: And polyolefins such as polyacrylates, ethylene-vinyl acetate polymers, acrylic substituted cellulose acetates, non-degradable polyurethanes, polystyrenes, polyvinyl chlorides, polyvinyl fluorides, poly (vinylimidazoles), chlorosulphonate polyolefins, Biodegradable polymer selected from the group consisting of polyethylene oxide, polyethylene oxide, and copolymers thereof. The microneedle is produced by adding microparticles to the microneedle. The microneedle according to claim 1, wherein the microneedles have a length of 250 탆 to 900 탆 penetrating the skin with respect to the protruding end of the microneedles. 13. A skin patch for improving skin wrinkles comprising the micro-needle according to any one of claims 1 to 12. 14. The skin patch for skin wrinkle improvement according to claim 13, wherein 0.1 to 5 mg of a peptide is administered to adult skin to control the release of neurotransmitter once every two days. A wrinkle-improving micro-needle containing a substance that inhibits SNARE complex formation. A method for reducing skin wrinkles comprising applying to a skin a micro needle comprising a peptide that modulates the release of a neurotransmitter. 17. The method of claim 16, wherein the skin wrinkle reduction method inhibits SNARE complex formation in the skin.

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