WO2015181746A1 - Sel de sodium d'heparine sous forme de gel pour administration dermique, et procede de preparation associe - Google Patents

Sel de sodium d'heparine sous forme de gel pour administration dermique, et procede de preparation associe Download PDF

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WO2015181746A1
WO2015181746A1 PCT/IB2015/053974 IB2015053974W WO2015181746A1 WO 2015181746 A1 WO2015181746 A1 WO 2015181746A1 IB 2015053974 W IB2015053974 W IB 2015053974W WO 2015181746 A1 WO2015181746 A1 WO 2015181746A1
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sodium salt
heparin
heparin sodium
salt form
form according
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English (en)
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Krzysztof BILMIN
Paweł GRIEB
Piotr SZOPIŃSKI
Marek LAGNER
Magdalena PRZYBYŁO
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Lipolek Sp. Z O.O.
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Publication of WO2015181746A1 publication Critical patent/WO2015181746A1/fr

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/0012—Galenical forms characterised by the site of application
    • A61K9/0014—Skin, i.e. galenical aspects of topical compositions
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
    • A61K31/715—Polysaccharides, i.e. having more than five saccharide radicals attached to each other by glycosidic linkages; Derivatives thereof, e.g. ethers, esters
    • A61K31/726—Glycosaminoglycans, i.e. mucopolysaccharides
    • A61K31/727—Heparin; Heparan
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/43—Enzymes; Proenzymes; Derivatives thereof
    • A61K38/46—Hydrolases (3)
    • A61K38/48—Hydrolases (3) acting on peptide bonds (3.4)
    • A61K38/4873—Cysteine endopeptidases (3.4.22), e.g. stem bromelain, papain, ficin, cathepsin H
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/24—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite containing atoms other than carbon, hydrogen, oxygen, halogen, nitrogen or sulfur, e.g. cyclomethicone or phospholipids
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/30—Macromolecular organic or inorganic compounds, e.g. inorganic polyphosphates
    • A61K47/36—Polysaccharides; Derivatives thereof, e.g. gums, starch, alginate, dextrin, hyaluronic acid, chitosan, inulin, agar or pectin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K8/00—Cosmetics or similar toiletry preparations
    • A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
    • A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
    • A61K8/64—Proteins; Peptides; Derivatives or degradation products thereof
    • A61K8/66—Enzymes
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K8/00—Cosmetics or similar toiletry preparations
    • A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
    • A61K8/72—Cosmetics or similar toiletry preparations characterised by the composition containing organic macromolecular compounds
    • A61K8/73—Polysaccharides
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/06—Ointments; Bases therefor; Other semi-solid forms, e.g. creams, sticks, gels
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/10—Dispersions; Emulsions
    • A61K9/107—Emulsions ; Emulsion preconcentrates; Micelles
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/10—Dispersions; Emulsions
    • A61K9/127—Liposomes
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P7/00—Drugs for disorders of the blood or the extracellular fluid
    • A61P7/02—Antithrombotic agents; Anticoagulants; Platelet aggregation inhibitors
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q19/00—Preparations for care of the skin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2800/00—Properties of cosmetic compositions or active ingredients thereof or formulation aids used therein and process related aspects
    • A61K2800/40—Chemical, physico-chemical or functional or structural properties of particular ingredients
    • A61K2800/41—Particular ingredients further characterized by their size
    • A61K2800/413—Nanosized, i.e. having sizes below 100 nm

Definitions

  • the invention relates to a gel form of a heparin sodium salt intended for dermal use as a medicinal product or cosmetic, and a method for its preparation.
  • Heparin is a highly water-soluble linear carbohydrate polymer of natural origin, characterized by a high content of sulfonic groups and a high negative charge in aqueous solution. This substance has been widely used in medicine, especially as an active ingredient of drugs and parapharmaceuticals with systemic or local effect, whose task is primarily to prevent blood clotting and the formation of blood clots in blood vessels.
  • porcine heparin API abbreviation of Active Pharmaceutical Ingredient
  • Porcine heparin is a polydisperse polymer, whose chains have a molecular weight within the range of 3-30 kD, and the average molecular weight of 15.5-17.0 kD (Sommers CD., Ye H., Kolinski .E., Nasr M., Buhse L.F., Al-Hakim A., Keire D.A., Characterization of currently marketed heparin products: analysis of molecular weight and heparinase-l digest patterns, Anal. Bioanal. Chem. 2011;401:2445-2454).
  • API heparin is most commonly found in the form of a sodium salt.
  • pharmacological activity is expressed in international units (IU), wherein the activity of porcine heparin API can not be less than 180 lU/mg.
  • Medical applications have also found low molecular weight heparins having an average molecular weight of ⁇ 8.0 kD, obtained by a partial depolymerization of the heparin carried out chemically, physically or enzymatically.
  • An example of a low molecular weight heparin is enoxaparin.
  • heparin and fractionated heparins intended for intravenous or subcutaneous injection are found in the trade in the form of drugs available on prescription, because their anticoagulant effect is strong and can sometimes be accompanied by serious and dangerous side effects, such as autosomal bleeding.
  • side effects such as autosomal bleeding.
  • heparin formulations intended for dermal administration. They are in the form of creams, ointments or sprays and are available without prescription as parapharmaceuticals intended for self-treatment, used e.g. in the case of varicose veins of the lower limbs and the connected risk of superficial venous thrombosis, as well as swellings, pain and feeling of heaviness in the legs, or as dermocosmetics (medical cosmetics) which improve the skin condition.
  • dermocosmetics medical cosmetics
  • heparin preparations for dermal administration in the prophylaxis of superficial venous thrombosis is significantly lower than the efficacy of intravenous or subcutaneous injections of heparin or low molecular weight heparins (Vecchio C, Frisinghelli A., Topically applied heparins for the treatment of vascular disorders: a comprehensive review, Clin. Drug Investig. 2008/28 603-614).
  • An exception is the preparation in the form of an aerosol containing a liposomal form of heparin, whose composition is consistent with patent no. EP0704206 issued to the German company Mika-Pharma.
  • Low molecular weight heparins due to the lower molecular weight of their carbohydrate chains, penetrate the skin better than unfractionated heparin (Betz G., Nowbakht P., Imboden ., Imanidis G., Heparin penetration into and permeation through human skin from aqueous and liposomal formulations in vitro, Int. J. Pharm. 2001; 228:147-159.).
  • low molecular weight heparins are significantly more expensive than unfractionated heparin, which limits their practical applications in formulations for injections.
  • permeability enhancers are often added. Usually these are detergents, amphiphiles, terpenes or alcohols, which are used to destabilize the lipid portion of the horny part of the skin (Kanikkannan, N., Kandimalla, K., Lamba, S. S., and Singh, M. (2000) "Structure- activity relationship of chemical penetration enhancers in transdermal drug delivery” Curr Med Chem 7, 593-608). The use of such substances is often accompanied by skin irritation or the drying of the skin surface. When ethanol is used the patient will experience unpleasant cooling of the skin caused by the evaporation of alcohol.
  • bromelain an enzyme derived from pineapple, is used for cleaning up wounds caused by skin burns and for removal of scabs (Rosenberg L, Krieger Y, Silberstein E, Arnon O, Sinelnikov IA, Bogdanov-Berezovsky A, Singer AJ. Selectivity of a bromelain based enzymatic debridement agent: a porcine study. Burns 2012; 38:1035-1040).
  • Papain is used dermally to improve the condition of the skin by removing or the transient destabilization of a protein intracellular matrix, which allows for an overall improvement of skin condition and removal of some of its heterogeneities or scars (S.J. Baik, B.Y. Kong, E.J. Kim, Cosmetic composition for exfoliating skin keratin, US Patent Application 2010/0254969 Al; Manosroi, A., Chankhampan, C, Manosroi, W., Manosroi, J. Transdermal absorption enhancement of papain loaded in elastic niosomes incorporated in gel for scar treatment, Eur J Pharm Sci 2013; 48: 474-483).
  • the present invention is a gel containing heparin with an admixture of proteolytic enzymes, intended for use as a medicinal product or cosmetic, which is characterized by the durability required for such preparations, and at the same time increases the efficiency of heparin passing through the skin.
  • Heparin gel may contain excipients in the form of polymers (e.g. hydrogels) or lipid structures, whose function is to slow down the evaporation of water and to increase the efficiency of the active substance (heparin) passing through the skin.
  • a suspension of neutral lipid aggregates in water has a tendency to aggregation and possible fusion, which significantly alters its properties and thus affects the release profiles of the active substance.
  • To stabilize the suspension of lipid aggregates two solutions are used; placing the electrostatic charge on the aggregates surface and the use of polymers, which sterically stabilize the lipid aggregates suspension. These two modifications alone or in combination prevent an aggregation and possible fusion of liposomes ( Allen, T. M. and E. H.
  • the addition to the formulation of the hydrogel preferably 0.5%, further stabilizes the aggregates by their spatial separation (Mufamadi, M. S., V. Pillay, et al.: "A review on composite liposomal technologies for specialized drug delivery.” J. Drug Delivery, 2011).
  • the most effective liposome preparations containing heparin (e.g. Lipohep) on the market do not meet the criterion of physico-chemical stability, which significantly lowers their efficacy.
  • the penetration yield of active substances through the skin depends on two factors; the state of the stratum corneum and the concentration of active substance in the aqueous solution on the skin surface.
  • the use of heparin alone results in a lack of its penetration into the dermis because it is not capable of penetrating the stratum corneum barrier.
  • heparin alone results in a lack of its penetration into the dermis because it is not capable of penetrating the stratum corneum barrier.
  • water evaporation it precipitates quickly on the skin surface, which results in a low functional stability of the preparation on the skin surface. For this reason preparations of this type have minimal efficacy.
  • Another approach is a combination of heparin with a hydrophobic base, which limits the precipitation of the heparin on the skin surface, however, in this case, a low contact angle causes the preparation's contact surface with the skin to be limited, which reduces or even prevents the transport of heparin.
  • the combination of heparin with a hydrogel slows down the evaporation of water and ensures good contact with the skin, however, both in this and previous solutions the stratum corneum barrier stays intact. This solution is also characterized by low efficacy resulting from a small stream of heparin penetrating the dermis.
  • heparin beneficially influences the permeability of the stratum corneum, however, rapid evaporation limits the amount of heparin penetrating the dermis (Cevc, G., G. Blume, et al.: "The skin: A pathway for systemic treatment with patches and lipid-based agent carriers.” Advanced Drug Delivery Reviews, 1996, 18(3): 349-378).
  • the available liposomal preparation with the trade name LipoHep contains considerable amounts of ethanol (4-6%) in its composition, which may result in skin irritation.
  • the solution used in the present invention is characterized by the fact that the heparin may be adsorbed on the surface of stable liposomes or contained in a hydrogel matrix, which makes the process of water evaporation slow down considerably (Fig. 7). Additionally, excipients, such as lipids in the form of liposomes, a hydrogel, or both together, in each case together with a proteolytic enzyme act synergistically on the stratum corneum temporarily increasing its permeability for hydrophilic substances including heparin. This fact is illustrated in Fig. 9 where the penetration of hydrophilic substances through the skin is illustrated by the example of carboxy-fluorescein.
  • OECD-# 4208 Liposomes modify the lipid fraction of the skin while the proteolytic enzymes destabilize the protein fraction of the epidermis. All this causes the penetration of heparin into the dermis to increase.
  • Fig. 8 and Fig 10 Preparation of the invention (Example 4) provides a much higher stream of heparin through the skin, measured on an in vitro model.
  • Fig. 8 and Fig. 10 show cumulative amounts of heparin in the trans range measured on the Franz model for the preparation of the invention, and the most effective so far on the market, Lioton (OECD- #428).
  • the essence of the solution according to the invention is the form of the heparin sodium salt for dermal applications, having a viscosity of less than 7000 cP, characterized by the fact that it consists of: heparin, or a pharmaceutically acceptable salt thereof; a hydrogel or liposomes, or both together; a proteolytic enzyme of plant origin or its stabilized derivative; and also chitosan, positive polyions, organic solvents, buffer and optionally: preservatives, antioxidants and gelling agents.
  • the lipid nanostructural form of heparin sodium salt is characterized by the fact that the overall concentration of the positive polyions is between 0 and 10 % by weight.
  • the lipid nanostructural form of heparin sodium salt is characterized by the fact that the overall concentration of the chitosan is between 0 and 10 % by weight.
  • the lipid nanostructural form of heparin sodium salt is characterized by the fact that the overall concentration of heparin in the formulation, ensuring the activity of a substance in the bloodstream as a result of dermal administration is between 0.1 and 10 % by weight, preferably 2.4 % by weight.
  • the lipid nanostructural form of heparin sodium salt is characterized by the fact that the building block of the liposomes is phosphatidylcholine, preferably 1.2-palmitoyl-sn-glycero-3- phosphatidylcholine.
  • the lipid nanostructural form of heparin sodium salt is characterized by the fact that the overall concentration of 1.2-palmitoyl-sn-glycero-3-phosphatidylcholine is between 0 and 30 % by weight, preferably 10 % by weight.
  • the lipid nanostructural form of heparin sodium salt is characterized by the fact that the building block of liposomes is phosphatidylcholine mixed with the quaternary ammonium compound from the group of tensides, preferably with benzalkonium chloride.
  • the lipid nanostructural form of heparin sodium salt is characterized by the fact that the overall concentration of cationic substance is between 0 - 80 mol% relative to phosphatidylcholine, preferably 40 %.
  • the lipid nanostructural form of heparin sodium salt is characterized by the fact that it contains single-layer liposomes.
  • the lipid nanostructural form of heparin sodium salt is characterized by the fact that it contains multi-layer liposomes.
  • the lipid nanostructural form of heparin sodium salt is characterized by the fact that it contains a mixture of single-layer and multi-layer liposomes.
  • the lipid nanostructural form of heparin sodium salt is characterized by the fact that the heparin is in an unbound form with liposomes.
  • the lipid nanostructural form of heparin sodium salt is characterized by the fact that the heparin is in a bound form with liposomes.
  • the lipid nanostructural form of heparin sodium salt is characterized by the fact that the heparin is in a bound and an unbound form with liposomes.
  • the lipid nanostructural form of heparin sodium salt is characterized by the fact that the factor supporting penetration through the skin is a cysteine-rich proteolytic enzyme of plant origin, papain or bromelain, or a stabilized derivative of this enzyme.
  • the lipid nanostructural form of heparin sodium salt is characterized by the fact that the proteolytic enzyme or stabilized derivative thereof is in an aqueous phase outside the liposomes.
  • the lipid nanostructural form of heparin sodium salt is characterized by the fact that the proteolytic enzyme is located both inside and outside the liposomes.
  • the lipid nanostructural form of heparin sodium salt is characterized by the fact that the concentration of the proteolytic enzyme is in the range of 0.01 - 10 % by weight.
  • the lipid nanostructural form of heparin sodium salt is characterized by the fact that the organic solvents are compounds from the group of dihydroxyl alcohols.
  • the lipid nanostructural form of heparin sodium salt is characterized by the fact that the organic solvents are compounds from the group of short chain alcohols, preferably with a chain length of c2-c4.
  • the lipid nanostructural form of heparin sodium salt is characterized by the fact that the overall concentration of the dihydroxyl alcohol is between 0 and 30 % by weight, preferably 10 %.
  • the lipid nanostructural form of heparin sodium salt is characterized by the fact that the organic solvents are compounds from the group of alkyl halides.
  • the lipid nanostructural form of heparin sodium salt is characterized by the fact that the overall concentration of organic solvent is not more than 0.5 % by weight of the formulation.
  • the lipid nanostructural form of heparin sodium salt is characterized by the fact that the buffer substance is 4-2-hydroxyethyl-l-piperazineethanesulfonic acid.
  • the lipid nanostructural form of heparin sodium salt form is characterized by the fact that the pH of the HEPES buffer (4-2-hydroxyethyl-l-piperazineethanesulfonic acid) is in the range of 6.0 to 8.0.
  • the lipid nanostructural form of heparin sodium salt is characterized by the fact that the antioxidant is a disodium edetate.
  • the lipid nanostructural form of heparin sodium salt form is characterized by the fact that the preservative is a methyl parahydroxybenzoate.
  • the lipid nanostructural form of heparin sodium salt form is characterized by the fact that the gelling agent is a copolymer of ammonium acrylamidomethylpropanosulfate and vinylpyrrolidone.
  • the lipid nanostructural form of heparin sodium salt is characterized by the fact that the overall concentration of the copolymer of ammonium acrylamidomethylpropanosulfate and vinylpyrrolidone is between 0 and 5 % by weight.
  • the method for the preparation of the lipid nanostructural form of heparin sodium salt for dermal applications is characterized by the fact that the heparin, or its pharmaceutically acceptable salts, is partially complexed in a two-phase system with an organic solvent in the presence of cationic polyions in a system with/without chitosan, and is stabilized with a lipid system in an organic solvent and with the elimination of the solvent there occurs a transfer of a stabilized aggregate to an aqueous phase.
  • the water portion which is of low viscosity, by means of high pressure processes, different from the process of homogenization, turns to a high viscosity phase of not less than 7000 cPs.
  • the proteolytic enzyme, an antioxidant, a preservative and a surplus portion of heparin are added, giving a product with a viscosity of not less than 7000 cPs, suitable for dermal administration in the form of a gel.
  • the mixture of thus prepared liposomes is mixed with a gelling agent.
  • the method for the preparation of the lipid nanostructural form of heparin sodium salt is characterized by the fact that the acquisition of its final, stable form does not require the use of gelling agents, and is provided by a suitable qualitative, quantitative composition and the manufacturing process.
  • the method for the preparation of lipid nanostructural form of heparin sodium salt is characterized by the fact that the complexation of heparin with lipid aggregate is carried out using positive polyions.
  • the method for the preparation of lipid nanostructural form of heparin sodium salt is characterized by the fact that the complexation of heparin with the lipid aggregate is carried out using chitosan.
  • the method for the preparation of lipid nanostructural form of heparin sodium salt is characterized by the fact that the complexation of heparin with the lipid aggregate is carried out using both chitosan ions and positive polyions.
  • the method for the preparation of lipid nanostructural form of heparin sodium salt is characterized by the fact that the process of complexation of heparin with the lipid aggregate occurs as a result of the thermodynamic phase separation process.
  • lipid nanostructures in dermal gel composition increases the bioavailability of heparin, and also the nanostructures used form a thin lipid film, modifying the lipid portion of the epidermis and protecting the ointment from drying too quickly, extending at the same time the absorption time of heparin and increasing its efficiency.
  • a proteolytic enzyme destabilizes the protein components of the epidermis aiding the absorption process of heparin.
  • Nanostructures produced according to the following examples were subjected to standard size measurement using a dynamic light scattering method showing their homogeneity, and the sign and value of the surface charge is determined via the measurement of the zeta-potential.
  • a nanostructure was obtained, the construction of which is schematically shown in Fig. 6.
  • Negatively charged heparin is condensed on the surface of the chitosan. Due to the anisotropic nature of charge distribution on the polymer (heparin) surface after binding to chitosan, the aggregate still shows a negative charge (Fig. 4).
  • a lipid from an organic solvent with a positive net charge is added, stabilizing the nanoaggregate. Transferring the aggregate to the aqueous phase takes place in the vicinity of positively charged polyions, stabilizing the nanoaggregate in the presence of neutral amphiphilic particles, ensuring a uniform distribution of the complex.
  • the outer layer is formed of a neutral lipid with a result that the nanostructure is readily dissolved in water and shows a uniform size distribution (Fig. 3, Fig.5).
  • Fig. 1 contains a left panel showing the size distribution of chitosan in a buffer with pH 6.5 and a right panel showing the fitting of correlation function to the experimental data for a solution of chitosan in a buffer with pH 6.5.
  • Fig. 2 contains a left panel showing the size distribution of heparin condensed on the surface of chitosan in a buffer with pH 6.5 and a right panel showing the fitting of correlation function to the experimental data for a solution of heparin with chitosan in a buffer with pH 6.5.
  • Fig. 3 contains a left panel showing the size distribution of heparin condensed on the surface of chitosan in envelopes from a lipid bilayer in a buffer with pH 6.5 with the addition of calcium ions a nd a right panel showing the fitting of the correlation function to the experimental data for a solution of heparin with chitosan in envelopes from a lipid bilayer in a buffer with the addition of calcium ions with pH 6.5.
  • Fig. 4 contains a left panel showing a zeta-potential distribution of heparin aggregates with chitosan in aqueous solution (the clearly negative (-40Mv) average value of the zeta potential is visible) and a right panel showing a phase diagram corresponding to the distribution from the left panel.
  • Fig. 5 contains a left panel showing a zeta-potential distribution of heparin aggregates with chitosan in aqueous solution (it is visible that the clearly increased value of the zeta-potential has shifted towards positive values (-6.8mV) average value of the zeta-potential) and a right panel showing a phase diagram corresponding to the distribution from the left panel.
  • Fig. 6 shows a block diagram of the lipid nanoaggregates "elastosome” containing condensed heparin on chitosan stabilized with positive polyions and a constant charge on the surface of the lipid.
  • Fig. 7 shows evaporation curves obtained for the heparin formulation of liposomal form, having the composition described in example 1 ( - A -) in comparison to the commercial preparation LipoHep (- ⁇ -
  • Fig. 8 shows cumulative amounts of heparin in a trans range measured on the Franz model for the preparation of the invention (Example 4) and the market equivalent (LipoHep)
  • Fig. 9 shows cumulative amounts of hydrophilic fluorescent marker carboxy-fluorescein in a trans range measured on the Franz model for the preparation of the invention (P2 - Formulation of Example 4) and the market equivalent (Lioton)
  • the obtained result shows increased penetration through a skin model (for the formulation from Example 4; P2) of hydrophilic substances in the presence of a proteolytic enzyme (5 % by weight of papain).
  • Fig. 10 shows cumulative amounts of heparin in a trans range measured on the Franz model for the preparation of the invention (Example 4) and the market equivalent (Lioton)
  • the obtained result shows increased penetration through a skin model by heparin in the presence of a proteolytic enzyme (5 % by weight of papain).
  • the graph also shows that increasing the amount of heparin (P3 - 2400 units of heparin) also increases its amount in the trans range.
  • a cationic lipid (DOTAP) (Avanti Lipids, USA) was dissolved in chloroform and added to the aqueous solution of heparin and chitosan. As a result, two immiscible phases were formed. Methanol was added to the sample so as to form one homogeneous phase.
  • DOTAP cationic lipid
  • the mixture was stirred at a temperature of about 25 °C until a homogeneous solution was obtained.
  • Purified phosphatidylcholine dissolved in chloroform was added to a solution containing heparin, chitosan and a cationic lipid, then all was topped up with a HEPES buffer to the final volume (5 ml). This was all centrifuged in order to accelerate phase separation.
  • the lower phase contained mostly chloroform with an excess of purified phosphatidylcholine and methanol.
  • the upper phase containing nanostructures and the buffer was separated, taking care that it was not contaminated by the lower phase in order to determine the size distribution of nanostructures.
  • the remaining methanol in the buffer phase was filtered using a MilliPore system. Finally, a proteolytic enzyme was added to the aqueous phase.
  • Benzalkonium chloride was dissolved in chloroform and added to the aqueous solution of heparin, resulting in the formation of two immiscible phases. Methanol was added to the sample so as to form one homogeneous phase.
  • the mixture was stirred at a temperature of about 25 °C until a homogeneous solution was obtained.
  • Purified phosphatidylcholine dissolved in the chloroform was added to a solution containing heparin, chitosan, calcium chloride and benzalkonium chloride, then the whole mixture was topped up with purified water to the final volume.
  • the whole mixture was centrifuged in order to accelerate phase separation.
  • the lower phase contained mostly chloroform with an excess of purified phosphatidylcholine.
  • the upper phase containing mostly water was separated, taking care that it was not contaminated by the lower phase in order to determine the size of the nanostructures.
  • the remaining methanol in the buffer phase was filtered using a MilliPore system.
  • the aqueous portion was subjected to the gelation process involving gelling agents.
  • composition of the ready formulation according to example 2 ACTIVE INGREDIENTS:
  • the whole mixture was stirred at a temperature of about 25 °C until a homogeneous suspension was obtained.
  • the resulting suspension was extruded at 60°C under a pressure of 150 psi. As a result of the extrusion a gel with a viscosity of 9000 cPs was obtained.
  • composition of the ready formulation according to example 3 ACTIVE INGREDIENTS:
  • Sample P3 with papain Heparin was added to purified water and dissolved at room temperature. To the prepared solution was added 1 g of ammonium acryloyldimethyltaurate and polyvinylpyrrolidone copolymer and it was stirred at 25°C until complete crosslinking of the gel. To the thus prepared gel was added 3 g of papain and it was stirred until the enzyme dispersed in the gel structure.

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  • Organic Chemistry (AREA)
  • Inorganic Chemistry (AREA)
  • Medicinal Preparation (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
  • Polysaccharides And Polysaccharide Derivatives (AREA)

Abstract

La présente invention concerne la forme nanostructurale lipidique du sel de sodium d'héparine destinée à des applications dermiques, qui présente une viscosité d'au moins 7000 cP et est caractérisée par le fait qu'elle est constituée : d'héparine ou d'un sel pharmaceutiquement acceptable de ce composé, de liposomes et/ou d'un hydrogel, d'enzymes protéolytiques d'origine végétale ou de leur dérivé stabilisé, de préférence le chitosane, de polyions positifs, de solvants organiques, d'agents tampon et gélifiants, et éventuellement de conservateurs et d'antioxydants. L'invention concerne également un procédé de préparation d'une forme nanostructurale lipidique du sel de sodium d'héparine destinée à des applications dermiques.
PCT/IB2015/053974 2014-05-29 2015-05-27 Sel de sodium d'heparine sous forme de gel pour administration dermique, et procede de preparation associe WO2015181746A1 (fr)

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PLPL408371 2014-05-29
PL408371A PL229532B1 (pl) 2014-05-29 2014-05-29 Żelowy preparat soli sodowej heparyny do podawania naskórnego

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CN114452248A (zh) * 2020-11-06 2022-05-10 上海帕尼生物科技有限公司 牛肺肝素钠水凝胶制剂、水凝胶贴片及其应用
CN114557928A (zh) * 2022-03-01 2022-05-31 上海联衡生物科技有限公司 一种美白祛黄护肤品及其制备方法

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Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN110204632A (zh) * 2019-04-04 2019-09-06 姜德亮 一种肠粘膜提取肝素钠粗品盐解工艺
CN114452248A (zh) * 2020-11-06 2022-05-10 上海帕尼生物科技有限公司 牛肺肝素钠水凝胶制剂、水凝胶贴片及其应用
CN114557928A (zh) * 2022-03-01 2022-05-31 上海联衡生物科技有限公司 一种美白祛黄护肤品及其制备方法
CN114557928B (zh) * 2022-03-01 2023-09-29 上海联衡生物科技有限公司 一种美白祛黄护肤品及其制备方法

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