WO2022071624A1 - Régulation de la résistance mécanique d'un hydrogel d'acide hyaluronique supramoléculaire auto-assemblé - Google Patents

Régulation de la résistance mécanique d'un hydrogel d'acide hyaluronique supramoléculaire auto-assemblé Download PDF

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WO2022071624A1
WO2022071624A1 PCT/KR2020/016283 KR2020016283W WO2022071624A1 WO 2022071624 A1 WO2022071624 A1 WO 2022071624A1 KR 2020016283 W KR2020016283 W KR 2020016283W WO 2022071624 A1 WO2022071624 A1 WO 2022071624A1
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hyaluronic acid
cyclodextrin
hydrogel
derivative
assembled
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Korean (ko)
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정다함
김문구
김환희
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주식회사 화이바이오메드
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    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08B—POLYSACCHARIDES; DERIVATIVES THEREOF
    • C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
    • C08B37/006—Heteroglycans, i.e. polysaccharides having more than one sugar residue in the main chain in either alternating or less regular sequence; Gellans; Succinoglycans; Arabinogalactans; Tragacanth or gum tragacanth or traganth from Astragalus; Gum Karaya from Sterculia urens; Gum Ghatti from Anogeissus latifolia; Derivatives thereof
    • C08B37/0063—Glycosaminoglycans or mucopolysaccharides, e.g. keratan sulfate; Derivatives thereof, e.g. fucoidan
    • C08B37/0072—Hyaluronic acid, i.e. HA or hyaluronan; Derivatives thereof, e.g. crosslinked hyaluronic acid (hylan) or hyaluronates
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08B—POLYSACCHARIDES; DERIVATIVES THEREOF
    • C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
    • C08B37/0006—Homoglycans, i.e. polysaccharides having a main chain consisting of one single sugar, e.g. colominic acid
    • C08B37/0009—Homoglycans, i.e. polysaccharides having a main chain consisting of one single sugar, e.g. colominic acid alpha-D-Glucans, e.g. polydextrose, alternan, glycogen; (alpha-1,4)(alpha-1,6)-D-Glucans; (alpha-1,3)(alpha-1,4)-D-Glucans, e.g. isolichenan or nigeran; (alpha-1,4)-D-Glucans; (alpha-1,3)-D-Glucans, e.g. pseudonigeran; Derivatives thereof
    • C08B37/0012—Cyclodextrin [CD], e.g. cycle with 6 units (alpha), with 7 units (beta) and with 8 units (gamma), large-ring cyclodextrin or cycloamylose with 9 units or more; Derivatives thereof
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08B—POLYSACCHARIDES; DERIVATIVES THEREOF
    • C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
    • C08B37/0006—Homoglycans, i.e. polysaccharides having a main chain consisting of one single sugar, e.g. colominic acid
    • C08B37/0024—Homoglycans, i.e. polysaccharides having a main chain consisting of one single sugar, e.g. colominic acid beta-D-Glucans; (beta-1,3)-D-Glucans, e.g. paramylon, coriolan, sclerotan, pachyman, callose, scleroglucan, schizophyllan, laminaran, lentinan or curdlan; (beta-1,6)-D-Glucans, e.g. pustulan; (beta-1,4)-D-Glucans; (beta-1,3)(beta-1,4)-D-Glucans, e.g. lichenan; Derivatives thereof
    • C08B37/0027—2-Acetamido-2-deoxy-beta-glucans; Derivatives thereof
    • C08B37/003—Chitin, i.e. 2-acetamido-2-deoxy-(beta-1,4)-D-glucan or N-acetyl-beta-1,4-D-glucosamine; Chitosan, i.e. deacetylated product of chitin or (beta-1,4)-D-glucosamine; Derivatives thereof
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
    • C08J3/00—Processes of treating or compounding macromolecular substances
    • C08J3/02—Making solutions, dispersions, lattices or gels by other methods than by solution, emulsion or suspension polymerisation techniques
    • C08J3/03—Making solutions, dispersions, lattices or gels by other methods than by solution, emulsion or suspension polymerisation techniques in aqueous media
    • C08J3/075—Macromolecular gels

Definitions

  • the present invention relates to the control of the mechanical strength of a supramolecular self-assembled hyaluronic acid hydrogel.
  • a hydrogel is composed of a hydrophilic polymer having a three-dimensional structure, and has a high moisture content without being soluble in water to provide an environment similar to living tissue.
  • the polymer constituting the hydrogel may be composed of a synthetic polymer or a bio-derived polymer, and the bio-derived polymer includes gelatin, collagen, agarose, alginic acid, chitosan, starch including amylose, fullulan, dextran, hyaluronic acid, etc. can be used These bio-derived polymers can constitute a three-dimensional structure of the hydrogel.
  • the polymers such as alginic acid, chitosan, fullulan, dextran, and hyaluronic acid lack self-assembly properties, and a metal crosslinking agent or chemical crosslinking agent is required to form the three-dimensional structure of the hydrogel.
  • a metal crosslinking agent or chemical crosslinking agent is required to form the three-dimensional structure of the hydrogel.
  • the crosslinking agent can react with drugs, enzymes, cells, etc., it is difficult to stably support drugs, enzymes, cells, etc. in the hydrogel.
  • gelatin, agarose, etc. are dissolved in water at a high temperature, and hydrophobic residues in the polymer are physically entangled at a low temperature to form a supramolecular self-assembling hydrogel.
  • a supramolecular self-assembled hydrogel according to such a temperature since a high temperature is required to initially dissolve the polymer, there is a disadvantage in that it is difficult to support drugs, enzymes, cells, etc. having low stability to heat.
  • the formation of the host-guest trapping complex means that the hydrophobic guest compound is trapped in the hydrophobic cavity in the hydrophilic host compound to form the complex through a non-covalent bond.
  • a representative bio-derived host compound includes cyclodextrin, a cyclic oligosaccharide. Cyclodextrins can be divided into alpha (6)- and beta (7)-gamma (8)- according to the number of glucose components. In the case of beta-cyclodextrin, it is known that it is possible to form a strong host-guest supramolecular trapping complex with adamantane.
  • hyaluronic acid-cyclodextrin hyaluronic acid-cyclodextrin
  • adamantane hyaluronic acid-cyclodextrin
  • beta-cyclodextrin beta-cyclodextrin to the glycoskeletal of hyaluronic acid having excellent biocompatibility.
  • a supramolecular self-assembled hydrogel was developed using a substituted (HA-Ad).
  • the hyaluronic acid derivative substituted with cyclodextrin and adamantane is based on the formation of a non-covalent host-guest trapping complex, so it is suitable for loading drugs, enzymes, cells, etc., and can be injected in vivo.
  • the supramolecular self-assembled hyaluronic acid hydrogel is due to the ability to form a complex of cyclodextrin and adamantane, it is possible to control the mechanical strength of the hydrogel by controlling the complex forming ability. Therefore, in the present invention, by controlling the type of cyclodextrin substituted for the hyaluronic acid backbone and the length of the linker between hyaluronic acid and cyclodextrin, the degree of freedom of the cyclodextrin is changed by controlling the formation of adamantane and trapping complexes to increase mechanical strength. It is possible to raise
  • An object of the present invention is to provide a supramolecular self-assembled hyaluronic acid hydrogel prepared by supramolecular self-assembly of cyclodextrin and adamantane.
  • Another object of the present invention is to provide a method for controlling the mechanical strength of a supramolecular self-assembled hyaluronic acid hydrogel.
  • the present invention relates to a hyaluronic acid-cyclodextrin derivative; and hyaluronic acid - prepared from adamantane derivatives,
  • the hyaluronic acid-cyclodextrin derivative is a supramolecular self-assembled hyaluronic acid with controlled mechanical strength in which the carboxyl group of D-glucuronic acid of hyaluronic acid and the hydroxyl group at the 6th carbon position of cyclodextrin are bonded via a linker.
  • a gel is provided.
  • the present invention provides a method for preparing the above-described supramolecular self-assembled hyaluronic acid hydrogel comprising mixing a hyaluronic acid-cyclodextrin derivative and a hyaluronic acid-adamantane derivative.
  • the present invention provides a supramolecular self-assembled hyaluronic acid hydrogel prepared by supramolecular self-assembly of cyclodextrin and adamantane.
  • the hydrogel can control the mechanical strength according to the type of hyaluronic acid-cyclodextrin derivative, so it is possible to provide the optimum mechanical strength required for the application of the supramolecular self-assembled hyaluronic acid hydrogel. In addition, it is possible to control the ability to form a trapping complex.
  • FIG. 1 shows a synthesis diagram of a hyaluronic acid-cyclodextrin derivative.
  • 2 is an NMR spectrum of a hyaluronic acid-cyclodextrin derivative
  • (a) is a 3' derivative having a cyclodextrin substitution degree of 17%
  • (b) is a 3' derivative having a cyclodextrin substitution degree of 49%
  • (c) is The NMR spectrum of the 4' derivative having a cyclodextrin substitution degree of 28% is shown.
  • FIG. 3 shows a synthesis diagram of hyaluronic acid-adamantane derivatives.
  • FIG. 5 shows a comparative graph of Tan ⁇ (loss factor) of the supramolecular self-assembled hyaluronic acid hydrogel.
  • FIG. 6 shows a film formation result and a skin application example.
  • the present invention relates to a hyaluronic acid-cyclodextrin derivative; And it relates to a hydrogel prepared from a hyaluronic acid-adamantane derivative.
  • hyaluronic acid has biocompatibility and biodegradability properties as well as transdermal delivery properties, so it can be safely applied to the human body and can be applied to transdermal drug delivery systems of various protein drugs and chemical drugs, including antigenic proteins.
  • HA hyaluronic acid
  • general formula 1 refers to a polymer having repeating units of N-acetyl-D-glucosamine and D-glucuronic acid. and is used in the sense of including all salts or derivatives of hyaluronic acid.
  • n may be an integer of 25 to 8,000 or 25 to 1,000.
  • HA-TBA tetrabutylammonium salt of hyaluronic acid
  • the 'hyaluronic acid derivative' is an amine group, an aldehyde group, a vinyl group, a thiol group, an allyloxy group, N-succinimidyl-3-(2-pyri) based on the basic structure of hyaluronic acid of Formula 1 above.
  • hyaluronic acid in which functional groups such as dildichio) propionate (N-Succinimidyl-3-(2-pyridyldithio)propionate, SPDP) and N-hydroxysuccinimide (NHS) are introduced refers to
  • hyaluronic acid derivative HA-diaminobutane, HA-hexamethylenediamine, HA-aldehyde, HA-adipic acid dihydrazide (HA) -Adipic Acid Dihydrazide, HA-ADH), HA-2-Aminoethyl methacrylate hydrochloride, HA-Spermine, HA-Spermidine (HA- spermidine), HA-SPDP, or HA-NHS may be used.
  • hyaluronic acid is present in most animals and can be safely applied to the human body as a linear polysaccharide polymer without biodegradability, biocompatibility, and immune response. Because hyaluronic acid performs several different roles in the body depending on its molecular weight, it can be used for many purposes.
  • composition of the hyaluronic acid, salt of hyaluronic acid, or derivative of hyaluronic acid used in the present invention is not limited, but preferably has a molecular weight of 10,000 to 3,000,000 Daltons (Da).
  • hydrogel means a gel using water as a dispersion medium.
  • the hydrogel may be formed when the hydrosol loses fluidity due to cooling or a hydrophilic polymer having a three-dimensional network structure and a microcrystalline structure expands by containing water.
  • Hydrogels of electrolyte polymers exhibit high water absorption and are practically used in various fields as water absorbent polymers. In some hydrogels, the expansion ratio is discontinuously changed due to a phase transition due to temperature, pH, etc.
  • “supramolecule” refers to a molecular complex (or trapping complex) formed by gathering molecules or ions through non-covalent bonds such as hydrogen bonding, electrostatic interaction, or van der Waals attraction. Since representative non-covalent bonds forming the structure of supramolecules are very weak compared to covalent bonds, the structure of supramolecular materials can easily change depending on the surrounding environment, and by using these characteristics, the shape of the material can be arbitrarily controlled. A representative principle of forming a supramolecular structure is self-assembly. Self-assembly refers to a phenomenon in which molecules are assembled through spontaneous interactions.
  • "supramolecular self-assembled hyaluronic acid hydrogel” is a hydrogel produced by the preparation method according to the present invention, and is prepared from hyaluronic acid-cyclodextrin derivatives and hyaluronic acid-adamantane derivatives, cyclodextrin and adamantine. It refers to a hydrogel produced by the supramolecular reaction of thein.
  • the hydrogel according to the present invention may be prepared from a hyaluronic acid-cyclodextrin derivative and a hyaluronic acid-adamantane derivative.
  • the hyaluronic acid-cyclodextrin derivative refers to a derivative in which hyaluronic acid and cyclodextrin are combined.
  • the carboxyl group of D-glucuronic acid of the hyaluronic acid and the hydroxy group at the 6th carbon position of cyclodextrin may form a bond via a linker, specifically, may form a bond through an amide bond.
  • the linker may have a structure of -NH-(CH 2 )m-NH-, and the hyaluronic acid-cyclodextrin derivative may be a compound represented by Formula 1 below.
  • n may be an integer of 25 to 8,000 or 25 to 1,000.
  • m may be an integer of 2 to 8 or an integer of 4 to 6. In the range of m, excellent mechanical properties can be imparted, and in particular, high mechanical strength can be imparted even with a low degree of substitution of cyclodextrin.
  • the degree of substitution of cyclodextrin in the hyaluronic acid-cyclodextrin derivative may be 10 to 50%, 10 to 40%, or 25 to 35%. In general, if the degree of substitution of the cyclodextrin is low, the mechanical properties of the prepared supramolecular hydrogel are poor. In addition, when the degree of substitution is high, the internal hydrophobicity of the cyclodextrin increases, lowering the solubility of the polymer in water, and inhibiting the hydrogen bonding and anionic properties that the carboxy group of hyaluronic acid can have, so that the hyaluronic acid skeleton itself has Structural characteristics may change.
  • the hyaluronic acid-cyclodextrin derivative according to the present invention has an advantage of high solubility in an aqueous solvent while having a low degree of substitution. In this case, the degree of substitution can be calculated as follows.
  • Degree of substitution (molar amount of hyaluronic acid repeating unit to which cyclodextrin is bound) * (molar amount of hyaluronic acid repeating unit to which cyclodextrin is bound and not bound) * 100
  • the hyaluronic acid-adamantane (CD-Ad) derivative refers to a derivative in which hyaluronic acid and adamantane are combined.
  • the hydroxy group of hyaluronic acid and the carboxyl group of adamantane may form a bond through an ester bond or an amide bond.
  • adamantane may be used to include all salts or derivatives of adamantane.
  • Adamantaneacetic acid Ada-acetic acid
  • (1-adamantyl)phenol in which a carboxyl group is substituted with adamantane may be used.
  • the hyaluronic acid-adamantane derivative may be represented by the following formula (2).
  • n may be an integer of 25 to 8,000 or 25 to 1,000.
  • the degree of substitution of adamantane in the hyaluronic acid-adamantane derivative may be 10 to 50% or 10 to 35%. In the above range, it is possible to prepare a hydrogel having excellent mechanical strength.
  • the hydrogel is the above-mentioned hyaluronic acid-cyclodextrin derivative; and hyaluronic acid-adamantane derivatives.
  • the hydrogel may be prepared by supramolecular reaction of hyaluronic acid-cyclodextrin derivatives of cyclodextrin and hyaluronic acid-adamantane derivatives with adamantane.
  • a supramolecular sieve that is, a hydrogel
  • cyclodextrins are cyclic oligosaccharides with hydrophobic cavities formed through ⁇ -1,4 bonds of 6-8 glucose molecules. dextrin, and eight ⁇ -cyclodextrins.
  • the molecular weight of the cyclodextrin, the size of the hydrophobic cavity, solubility, etc. may vary depending on the number of glucose molecules forming the cyclodextrin.
  • the hydroxyl groups bound to C2 and C3 are spread outward, and the hydroxyl groups bound to C6 are also spread in the opposite direction, so the outside of the ring is hydrophilic as a whole.
  • the inner cavity is hydrophobic. Therefore, the hydrophilic outer shell of the entire structure allows it to dissolve well in a polar solvent such as water, while forming hydrophobic pores with the opposite nature of the outer structure inside the structure. This enables complex formation through host-guest interaction, which is the greatest property of cyclodextrins.
  • the guest material forms a complex by structural fitting while entering the pores of the cyclodextrin having a certain size.
  • the pore height is the same, but the diameter and volume are different.
  • adamantane is used as the guest material.
  • the adamantane has a structure in which four cyclohexane rings are condensed in a basket shape, is highly symmetrical, and is a stable compound, and may form a bond through host-guest interaction with cyclodextrin.
  • a hydrogel can be prepared through host-guest interaction of cyclodextrin of hyaluronic acid-cyclodextrin derivative and adamantane of hyaluronic acid-adamantane derivative.
  • the content ratio of the hyaluronic acid-cyclodextrin derivative and the hyaluronic acid-adamantane derivative may be 1:0.1 to 1:10, 1:0.5 to 1:2, or 1:1.
  • the supramolecular self-assembled hyaluronic acid hydrogel may be a buffer solution based on an aqueous solution.
  • the content of the hydrogel in the buffer may be 1 to 50% by weight, 5 to 20% by weight, or 5 to 15% by weight.
  • the hydrogel of the present invention may contain useful substances.
  • the type of the useful substance is not particularly limited, and for example, it may be one or more selected from the group consisting of drugs, useful substances of functional cosmetics, fluorescent substances, radioactive isotopes, target-directing substances, imaging substances, and culture solutions necessary for cell culture. there is.
  • a hydrogel containing a useful substance may function as a drug delivery system for delivery of the useful substance.
  • the drug is a substance capable of inhibiting, suppressing, alleviating, alleviating, delaying, preventing or treating diseases or symptoms in animals including humans, for example, paclitaxel, doxorubicin, docetaxel, 5- Fluoreuracil (5-fluoreuracil), oxaliplatin (oxaliplatin), cisplatin (cisplatin), carboplatin (carboplatin), berberine (berberine), epirubicin (epirubicin), doxycycline (doxycycline), gemcitabine (gemcitabine), rapamycin, tamoxifen, herceptin, avastin, tysabri, erbitux, campath, zevalin, humira , mylotarg, Xolair, bexxar, raptiva, remicade, siRNA, aptamer, interferon, insulin, reopro ( reopro), rituxan, zenapax,
  • One selected from the group consisting of a wrinkle functional ingredient, a whitening functional ingredient, a UV protection functional ingredient, a natural product-derived ingredient, a skin microbiome, a functional strain, and a fermented product of the skin microbiome and functional strain, as a useful material of the functional cosmetic can be used.
  • adhesives, thickeners, pH adjusters, flavoring agents, wetting agents and preservatives that are included in cosmetic formulations may be additionally used.
  • the fluorescent material may be a fluorescent material generally used in the art to which the present invention belongs, and examples thereof include fluorescein, rhodamine, dansyl, Cy and anthracene. can be used
  • the radioactive isotope may be 3 H, 14 C, 22 Na, 35 S, 33 P, 32 P and 125 I.
  • the target-directing substance refers to any substance capable of selectively recognizing, binding, or delivering a specific target substance, for example, RGD (arginine-leucine-aspartic acid), TAT (threonine-alanine-threonine) and MVm ( Peptides such as methionine-valine-Dmethionine), peptides that recognize specific cells, antigens, and antibodies.
  • RGD arginine-leucine-aspartic acid
  • TAT threonine-alanine-threonine
  • MVm Peptides such as methionine-valine-Dmethionine
  • peptides that recognize specific cells, antigens, and antibodies.
  • Folic acid, nucleic acids, aptamers, or carbohydrates eg, glucose, fructose, mannose, galactose, ribose, etc.
  • the imaging material is a spectroscopy such as NMR (Nuclear Resonance Spectrometer), MRI (Magnetic Resonance Image), PET (Positron Emission Tomography), CT (Nuclear Resonance Spectrometer), fluorescence microscope, confocal laser scanning microscope, etc.
  • the Ga-complex includes Ga-DTPA, Ga-DTPA-BMA, Ga-DOT and Ga-cyclam
  • the nanoparticles include gold, silver, manganese, cadmium, selenium, tellurium and zinc
  • the nanoparticles are 1-200 nm in size, and single-walled nanotubes, multi-walled nanotubes, fullerene, graphene, and the like may be used as the carbon nanomaterial.
  • components used in the art can be used without limitation as a culture medium required for cell culture.
  • the present invention relates to a method for preparing the above-described supramolecular self-assembled hyaluronic acid hydrogel.
  • the supramolecular self-assembled hyaluronic acid hydrogel according to the present invention may include mixing a hyaluronic acid-cyclodextrin derivative and a hyaluronic acid-adamantane derivative.
  • the hyaluronic acid-cyclodextrin derivative can be prepared by reacting hyaluronic acid with cyclodextrin.
  • the hyaluronic acid-cyclodextrin derivative is prepared by (a) reacting a hydroxy group at the 6th carbon position of the cyclodextrin with a diamine compound to prepare a cyclodextrin having an amine group;
  • a cyclodextrin having an amine group may be prepared by forming a reactive group in the hydroxyl group at the 6th carbon position of cyclodextrin and then reacting it with a diamine compound.
  • diamine compound from the group consisting of 1,2-ethylenediamine, 1,3-propanediamine, 1,4-butanediamine, 1,5-pentanediamine, 1,6-hexanediamine, 1,7-heptanediamine and 1,8-octanediamine One or more selected may be used.
  • the cyclodextrin having an amine group may be used in 0.5 to 1.5 equivalents, 0.5 to 1.0 equivalents, or 0.6 to 0.8 equivalents relative to 1 equivalent of hyaluronic acid.
  • the content of the cyclodextrin used affects the degree of substitution, and the degree of substitution can be adjusted to 10 to 50% in the content range.
  • Step (b) may be performed in the presence of a solvent and a coupling reagent.
  • Water, DMSO (dimethyl sulfoxide) or PBS (Phosphate-buffered saline) may be used as the solvent, and DMTMM (4-(4,6-Dimethoxy-1,3,5-triazin-2-yl) as a coupling reagent) -4-methylmorpholinium chloride), EDC(N-(3-Dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride, NHS(N-Hydroxysuccinimide), Pyridine, HBTU(N′-Tetramethyl-O-(1H-benzotriazol-1-yl) uronium hexafluorophosphate) or BOP ((Benzotriazol-1-yloxy)tris(dimethylamino)phosphonium hexafluorophosphate) may be used.
  • PBS or MES (2-(N-morpholino)ethanesulfonic acid) may be used as a buffer.
  • the step (b) may be carried out at 25 to 50 °C, room temperature for 20 to 48 hours or 20 to 30 hours.
  • the degree of substitution of the hyaluronic acid-cyclodextrin derivative prepared by adjusting the equivalent weight and reaction time of the cyclodextrin having an amine group can be adjusted.
  • the degree of substitution of the hyaluronic acid-cyclodextrin derivative may be 10 to 50%, 10 to 40%, or 25 to 35%.
  • the hyaluronic acid-adamantane derivative can be prepared by reacting hyaluronic acid with adamantane.
  • it can be prepared by dissolving hyaluronic acid and adamantane in a solvent and then reacting in the presence of a reaction reagent.
  • the reaction reagent is a reagent that causes an esterification reaction, such as 4-DMAP (4-Dimethylaminopyridine), DVA (Divinyl acetate), N,N'-dicyclohexylcarbodiimide (N,N'-dicyclohexylcarbodiimide; DCC), Adamantane anhydride or Di-tert-butyl You can use dicarbonate.
  • PBS or MES (2-(N-morpholino)ethanesulfonic acid) may be used as a buffer. The reaction may be carried out in a vacuum state.
  • the degree of substitution of adamantane in the hyaluronic acid-adamantane derivative prepared above may be 10 to 50%.
  • a hydrogel can be prepared through a supramolecular reaction of hyaluronic acid-cyclodextrin derivative cyclodextrin and hyaluronic acid-adamantane derivative adamantane, that is, host-guest interaction.
  • the content ratio of the hyaluronic acid-cyclodextrin derivative and the hyaluronic acid-adamantane derivative may be 1:0.1 to 1:10, 1:0.5 to 1:2, or 1:1.
  • the hydrogel can be prepared by physical mixing of the hyaluronic acid-cyclodextrin derivative and the hyaluronic acid-adamantane derivative.
  • the present invention provides the aforementioned hyaluronic acid-cyclodextrin derivative; And it relates to a drug delivery system comprising a hyaluronic acid-adamantane derivative.
  • hydrogel according to the present invention is prepared through self-assembly of the above-mentioned derivatives, useful substances, specifically drugs, can be loaded inside the hydrogel.
  • useful substances specifically drugs
  • the above-mentioned types may be used.
  • the useful material is supported in the hydrogel by mixing the useful material with the hyaluronic acid-cyclodextrin derivative and the hyaluronic acid-adamantane derivative in the production process of the hydrogel, so that the useful material can be easily supported in the hydrogel. .
  • the useful substance in the preparation of the drug delivery system, that is, when the useful substance is supported in the hydrogel, is used in the form of an HA-useful substance conjugate in which a useful substance is bound to hyaluronic acid, or hyaluronic acid-a It can be used in the form of an HA-Ad-useful substance conjugate in which a useful substance is bound to a damantane derivative. Alternatively, it may be used as a useful substance itself.
  • the HA-useful substance conjugate is prepared by introducing an aldehyde group to hyaluronic acid, and binding a useful substance (the useful substance may include an amine group or modified with an amine group) to hyaluronic acid through an amine-aldehyde reaction.
  • a useful substance the useful substance may include an amine group or modified with an amine group
  • the HA-Ad-useful substance conjugate introduces an aldehyde group to the hyaluronic acid-adamantane (HA-Ad) derivative, and a useful substance to the HA-Ad derivative through an amine-aldehyde reaction (the useful substance contains an amine group or , may be modified with an amine group).
  • the drug delivery system may be used to deliver useful substances in vivo or in vitro to animals including humans.
  • the present invention provides a pharmaceutical composition comprising the drug delivery system according to the present invention.
  • the pharmaceutical composition may further include a pharmaceutically acceptable carrier or diluent and the like, and the pharmaceutical composition may be administered by any method known to those skilled in the art, for example, oral or parenteral administration, such as injection, infusion, implantation.
  • parenteral routes include intravascular, intratumoral, peripheral cancer, transmucosal, transdermal, intramuscular, intranasal, intravenous, intradermal, subcutaneous, intraperitoneal, intraventricularly, intracranially, intravaginal, inhalation. , work, etc.
  • the pharmaceutical composition may be used as it is, or formulated into a form suitable for the route of administration, that is, a solid formulation, a liquid formulation, or a hydrogel.
  • the present invention relates to a composition for transdermal delivery comprising the above-described supramolecular self-assembled hyaluronic acid hydrogel.
  • hyaluronic acid-cyclodextrin derivatives according to the present invention.
  • hyaluronic acid - When the supramolecular self-assembled hyaluronic acid hydrogel prepared from the adamantane derivative is applied to the skin, a film is formed, which enables maintenance of the active substance in the formulation and effective transdermal delivery.
  • the content of the hyaluronic acid-cyclodextrin derivative and the hyaluronic acid-adamantane derivative included in the composition for transdermal delivery may be 0.5 to 10 wt%, 1 to 5 wt%, or 1 to 2 wt%, respectively .
  • a film is formed when applied to the skin, etc., and the maintenance and transdermal delivery of useful substances can be effectively achieved. If the content of the derivative is too small, a problem occurs in the storage of useful substances, and the film is not formed and exists only as a fluid gel, so it is better to adjust the content range.
  • the composition for transdermal delivery may include the above-mentioned useful substances, specifically, wrinkle functional ingredient, whitening functional ingredient, UV blocking functional ingredient, natural product-derived ingredient, skin microbiome, functional strain, and skin It may include one or more selected from the group consisting of fermentation products of microbiome and functional strains.
  • the present invention provides a hyaluronic acid-cyclodextrin derivative; And it relates to a cosmetic composition comprising a hyaluronic acid-adamantane derivative.
  • the cosmetic composition according to the present invention may have an effect of improving wrinkles and may be used as a cosmetic composition for improving wrinkles.
  • the synthetic process of the hyaluronic acid-cyclodextyrin derivative is shown as a schematic diagram in FIG. 1 .
  • hyaluronic acid 100 mg was dissolved by stirring in 20 ml of 0.1M MES buffer (pH 5.5). After that, 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)4-methoxymorpholinium chloride (DMTMM) corresponding to 4 times the number of moles of hyaluronic acid was added to the hyaluronic acid solution. and stirred for 1 to 2 hours.
  • DTMM 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)4-methoxymorpholinium chloride
  • a hyaluronic acid-cyclodextrin derivative was synthesized by adding 0.75 times equivalent of the cyclodextrin prepared in 1 to the hyaluronic acid reaction solution and stirring at room temperature for 24 hours.
  • the prepared derivative (4') was purified through dialysis and obtained through freeze-drying.
  • HA-CD hyaluronic acid-cyclodextrin
  • the synthetic process of the hyaluronic acid-adamantane derivative is shown as a schematic diagram in FIG. 3 .
  • a TBA (Tetrabutylammonium) salt of hyaluronic acid was prepared.
  • a hyaluronic acid solution was prepared by dissolving 1 g of hyaluronic acid in 100 ml of deionized tertiary distilled water.
  • the Na (sodium) salt of hyaluronic acid was removed by passing the hyaluronic acid solution through a DOWEX 50Wx 8,400 or cation-substituted resin column.
  • the pH of the hyaluronic acid eluent from which Na was removed was adjusted to 7-8 using 40% TBA-OH (Tetrabutylammonium hodroxide). After that, the mixture was stirred for 1 hour using a magnetic stirrer and then freeze-dried to prepare a TBA salt of hyaluronic acid (6).
  • TBA-OH Tetrabutylammonium hodroxide
  • the TBA salt of hyaluronic acid (6) and adamantane acetic acid (1-adamataneacetic acid, 7) corresponding to 2-3 times the standard equivalent of hyaluronic acid and 4-DMAP (4) corresponding to 0.7 to 1 times the equivalent of hyaluronic acid -dimethylaminopyridine) was dissolved in DMSO (dimethyl sulfoxide). Before and after dissolution, oxygen in the reaction vessel was removed using vacuum and argon gas. After that, di-tert-butyl-dicarbonate (an amount equivalent to ⁇ 0.5 times the hyaluronic acid standard equivalent) was added to the reaction solution, and then sufficiently stirred for 24 hours using a magnetic stirrer.
  • the reaction solution not the sediment recovery method, was recovered through the addition of isopropyl alcohol, put into a dialysis membrane, and dialysis was performed using 20% (v/v) DMSO solution for 24 hours. Thereafter, the obtained product may be obtained by further dialysis through the above method.
  • the substitution degree of the prepared hyaluronic acid-adamantane derivative was 20%.
  • the supramolecular self-assembled hyaluronic acid hydrogel was prepared by mixing the two hyaluronic acid derivatives in a 1:1 ratio.
  • Cyclodextrin (3A-Amino-3A-deoxy-(2AS,3AS)-beta-cyclodextrin, 2) in which the hydroxy group at carbon 3 is substituted with an amine group was prepared by TOKYO CHEMICAL INDUSTRY CO., LTD, (Product No.: A1916). ) was purchased and used.
  • hyaluronic acid 100 mg was dissolved by stirring in 10 ml of 0.1M MES buffer (pH 5.5). After that, 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)4-methoxymorpholinium chloride (DMTMM) corresponding to 4 times the number of moles of hyaluronic acid was added to the hyaluronic acid solution. and stirred for 1 to 2 hours.
  • DTMM 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)4-methoxymorpholinium chloride
  • Example 1 (1) of Example 1, except that 0.7 times equivalent of the cyclodextrin of 1 was added to the hyaluronic acid reaction solution and reacted with stirring at room temperature for 24 hours to synthesize a hyaluronic acid-cyclodextrin derivative
  • a hyaluronic acid-cyclodextrin derivative (2') was prepared in the same manner as described above.
  • the degree of substitution of the prepared hyaluronic acid-adamantane derivative was 50%.
  • a hyaluronic acid-adamantane derivative was prepared in the same manner as in Example 1 (2).
  • a hydrogel was prepared in the same manner as in (3) of Example 1, except that 5 wt% of the hyaluronic acid-cyclodextrin derivative prepared in (1) was dissolved in PBS solution to prepare a hyaluronic acid-cyclodextrin derivative solution. did
  • hyaluronic acid 100 mg was dissolved by stirring in 20 ml of 0.1M MES buffer (pH 5.5). After that, 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)4-methoxymorpholinium chloride (DMTMM) corresponding to twice the number of moles of hyaluronic acid was added to the hyaluronic acid solution. and stirred for 1 to 2 hours.
  • DTMM 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)4-methoxymorpholinium chloride
  • Example 1 A hyaluronic acid-cyclodextrin derivative (3') was prepared in the same manner as in 1).
  • the substitution degree of the prepared hyaluronic acid-adamantane derivative was 17%.
  • a hyaluronic acid-adamantane derivative was prepared in the same manner as in Example 1 (2).
  • a hydrogel was prepared in the same manner as in (3) of Example 1, except that 5 wt% of the hyaluronic acid-cyclodextrin derivative prepared in (1) was dissolved in PBS solution to prepare a hyaluronic acid-cyclodextrin derivative solution. did
  • hyaluronic acid 100 mg was dissolved by stirring in 15 ml of 0.1M MES buffer (pH 5.5). After that, 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)4-methoxymorpholinium chloride (DMTMM) corresponding to twice the number of moles of hyaluronic acid was added to the hyaluronic acid solution. and stirred for 1 to 2 hours.
  • DTMM 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)4-methoxymorpholinium chloride
  • the degree of substitution of the prepared hyaluronic acid-adamantane derivative was 49%.
  • a hyaluronic acid-adamantane derivative was prepared in the same manner as in Example 1 (2).
  • a hydrogel was prepared in the same manner as in (3) of Example 1, except that 5 wt% of the hyaluronic acid-cyclodextrin derivative prepared in (1) was dissolved in PBS solution to prepare a hyaluronic acid-cyclodextrin derivative solution. did
  • hyaluronic acid-cyclodextrin derivatives prepared in Examples and Comparative Examples were analyzed by NMR (DPX300, Bruker, Germany).
  • the storage modulus (G') and loss modulus (G'') of the hydrogel through the MCR 92 rheometer (Anton Paar, USA) were measured at an angular frequency of 0.1 to 100 rad/s and 25 mm in diameter at 25°C. Measured by plate.
  • the measurement results are shown in FIG. 4 .
  • FIG 4 (a) is composed of 5wt% HA-CD (2', substitution degree 50%) and 5wt% HA-Ad (6', substitution degree 20%) prepared in Comparative Example 1
  • (b) is a comparison Consisting of 5wt% HA-CD (3', substitution degree 17%) and 5wt% HA-Ad (6', substitution degree 20%) prepared in Example 2
  • (c) is 5wt% HA prepared in Comparative Example 3 -CD (3', substitution degree 49%) and 5wt% HA-Ad (6', substitution degree 20%)
  • (d) is 4wt% HA-CD (4', substitution degree) prepared in Example 1 28%) and 5wt% HA-Ad (6', degree of substitution 20%) shows the rheological properties of the supramolecular self-assembled hyaluronic acid hydrogel.
  • the hydrogel prepared in Comparative Example 1 had a higher degree of substitution of cyclodextrin compared to Comparative Example 2, and thus the elasticity of the gel was high. It can be seen that Comparative Example 3 having a similar degree of substitution of cyclodextrin has higher elasticity than Comparative Example 1. That is, the hydrogel is bonded to the 6th carbon position of the cyclodextrin, and a high degree of substitution has excellent elasticity.
  • Example 1 has a lower degree of cyclodextrin substitution as compared to Comparative Examples 1 and 3, and has an elasticity of equal to or greater than that of Comparative Example 3 despite having a lower concentration of hyaluronic acid.
  • the measurement results are shown in FIG. 5 .
  • Comparative Example 2 exhibits a higher Tan ⁇ value than other examples in all angular frequency ranges, and a Tan ⁇ value of 1 or less at an angular frequency of 30 rad/s or more.
  • Comparative Example 1 with a high degree of substitution, a lower Tan ⁇ value than that of Comparative Example 2 was shown at the initial angular frequency, and a Tan ⁇ value of 1 or less was shown at an angular frequency of 10 rad/s or more, resulting in superior mechanical strength than Comparative Example 2 confirmed that.
  • a Tan ⁇ value lower than 1 means that the elasticity of the hydrogel is higher than the viscosity, and a lower Tan ⁇ value means higher mechanical strength. Therefore, through this experimental example, it can be confirmed that the hydrogel prepared in Example 1 has excellent mechanical strength while having a low degree of substitution.
  • a cosmetic formulation for transdermal delivery was prepared by dissolving the hyaluronic acid-cyclodextrin derivative and the hyaluronic acid-adamantane derivative prepared in Example 1 at a concentration of 1 wt% or 2 wt%, respectively, in the cosmetic composition as shown in Table 1 below.
  • Cosmetic formulation includes water, thickener, pH adjuster, moisturizer, preservative solvent, etc., whitening functional ingredient, wrinkle improvement functional ingredient, natural product-derived functional ingredient, skin microbiome and functional strain, skin microbiome and functional strain fermentation Water and the like may be further included.
  • the film formation results and skin application examples of the prepared cosmetic formulations are shown in FIG. 6 .
  • the content of the hyaluronic acid-cyclodextrin derivative and the hyaluronic acid-adamantane derivative in the cosmetic formulation may be 1 to 5 wt%, and a film may be formed when applied to the skin together with each cosmetic ingredient.
  • the active ingredient is maintained and efficient transdermal delivery can be achieved.
  • the supramolecular self-assembled hyaluronic acid hydrogel according to the present invention can control the mechanical strength according to the type of hyaluronic acid-cyclodextrin derivative, it is possible to provide the optimum mechanical strength required for the field of application of the hyaluronic acid hydrogel.

Abstract

La présente invention concerne un hydrogel d'acide hyaluronique supramoléculaire auto-assemblé. L'hydrogel d'acide hyaluronique supramoléculaire auto-assemblé, selon la présente invention, permet de réguler la résistance mécanique en fonction du type de dérivé d'acide hyaluronique-cyclodextrine, et par conséquent, peut fournir la résistance mécanique optimale requise dans le domaine de l'application d'hydrogels d'acide hyaluronique.
PCT/KR2020/016283 2020-09-29 2020-11-18 Régulation de la résistance mécanique d'un hydrogel d'acide hyaluronique supramoléculaire auto-assemblé WO2022071624A1 (fr)

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