WO2022260366A1 - Prussian blue/polyvinylpyrrolidone nanoparticle composite and use thereof - Google Patents

Prussian blue/polyvinylpyrrolidone nanoparticle composite and use thereof Download PDF

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WO2022260366A1
WO2022260366A1 PCT/KR2022/007931 KR2022007931W WO2022260366A1 WO 2022260366 A1 WO2022260366 A1 WO 2022260366A1 KR 2022007931 W KR2022007931 W KR 2022007931W WO 2022260366 A1 WO2022260366 A1 WO 2022260366A1
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prussian blue
polyvinylpyrrolidone
disease
nps
pvp
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French (fr)
Korean (ko)
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최원일
오혜련
이진실
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한국세라믹기술원
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    • 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
    • A61K47/50—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 the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/69—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 the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit
    • A61K47/6921—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 the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a particulate, a powder, an adsorbate, a bead or a sphere
    • A61K47/6927—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 the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a particulate, a powder, an adsorbate, a bead or a sphere the form being a solid microparticle having no hollow or gas-filled cores
    • A61K47/6929—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 the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a particulate, a powder, an adsorbate, a bead or a sphere the form being a solid microparticle having no hollow or gas-filled cores the form being a nanoparticle, e.g. an immuno-nanoparticle
    • A61K47/6931—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 the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a particulate, a powder, an adsorbate, a bead or a sphere the form being a solid microparticle having no hollow or gas-filled cores the form being a nanoparticle, e.g. an immuno-nanoparticle the material constituting the nanoparticle being a polymer
    • A61K47/6933—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 the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a particulate, a powder, an adsorbate, a bead or a sphere the form being a solid microparticle having no hollow or gas-filled cores the form being a nanoparticle, e.g. an immuno-nanoparticle the material constituting the nanoparticle being a polymer the polymer being obtained by reactions only involving carbon to carbon, e.g. poly(meth)acrylate, polystyrene, polyvinylpyrrolidone or polyvinylalcohol
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    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
    • A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61F—FILTERS IMPLANTABLE INTO BLOOD VESSELS; PROSTHESES; DEVICES PROVIDING PATENCY TO, OR PREVENTING COLLAPSING OF, TUBULAR STRUCTURES OF THE BODY, e.g. STENTS; ORTHOPAEDIC, NURSING OR CONTRACEPTIVE DEVICES; FOMENTATION; TREATMENT OR PROTECTION OF EYES OR EARS; BANDAGES, DRESSINGS OR ABSORBENT PADS; FIRST-AID KITS
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    • A61K47/69—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 the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit
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    • 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
    • A61K8/40—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing nitrogen
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    • A61K8/72—Cosmetics or similar toiletry preparations characterised by the composition containing organic macromolecular compounds
    • A61K8/81—Cosmetics or similar toiletry preparations characterised by the composition containing organic macromolecular compounds obtained by reactions involving only carbon-to-carbon unsaturated bonds
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    • A61K8/81—Cosmetics or similar toiletry preparations characterised by the composition containing organic macromolecular compounds obtained by reactions involving only carbon-to-carbon unsaturated bonds
    • A61K8/817—Compositions of homopolymers or copolymers of compounds having one or more unsaturated aliphatic radicals, each having only one carbon-to-carbon double bond, and at least one being terminated by a single or double bond to nitrogen or by a heterocyclic ring containing nitrogen; Compositions or derivatives of such polymers, e.g. vinylimidazol, vinylcaprolactame, allylamines (Polyquaternium 6)
    • A61K8/8176—Homopolymers of N-vinyl-pyrrolidones. Compositions of derivatives of such polymers
    • A—HUMAN NECESSITIES
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    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L15/00—Chemical aspects of, or use of materials for, bandages, dressings or absorbent pads
    • A61L15/16—Bandages, dressings or absorbent pads for physiological fluids such as urine or blood, e.g. sanitary towels, tampons
    • A61L15/42—Use of materials characterised by their function or physical properties
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    • A61P17/02—Drugs for dermatological disorders for treating wounds, ulcers, burns, scars, keloids, or the like
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    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
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    • 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
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    • A61L2400/00—Materials characterised by their function or physical properties
    • A61L2400/12—Nanosized materials, e.g. nanofibres, nanoparticles, nanowires, nanotubes; Nanostructured surfaces

Definitions

  • the present invention relates to a Prussian blue/polyvinylpyrrolidone nanoparticle composite and its use for scavenging active oxygen.
  • Prussian Blue is an iron ferrocyanide (Fe 4 [Fe(CN) 6 ] 3 ), which enables charge/electron transfer between high-spin iron (Fe 3+ ) and low-spin iron (Fe 2+ ) ions. It has special physicochemical properties. Prussian Blue's Fe 2+ -CN-Fe 3+ coupling has been used as an important inorganic pigment because it absorbs light in the near-infrared wavelength range and emits a unique blue color. In addition, Prussian Blue can selectively adsorb radioactive cesium, so Radiogardase as a drug for removing radioactive isotopes has received drug approval from the US Food and Drug Administration (FDA). Furthermore, it is proposed as a treatment material for various diseases by using the electrochemical redox of Prussian blue.
  • FDA US Food and Drug Administration
  • Prussian blue can be oxidized and reduced to Prussian white or Prussian green, which is similar to the mechanism of action of antioxidant enzymes (peroxidase (POD), catalase (CAT), superoxide dismutase (SOD), etc.) Possibility has been raised that it will be effective in treating diseases.
  • POD peroxidase
  • CAT catalase
  • SOD superoxide dismutase
  • Prussian blue has excellent biocompatibility and reactive oxygen scavenging ability, it is difficult to use it due to its low in vivo stability. Therefore, various polymer-based Prussian blue composites have been developed to improve the stability of Prussian blue. For example, several polymers such as polyvinylpyrrolidone (PVP), polyvinyl alcohol (PVA), polyallylamine hydrochloride (PAH), poly(diallyldimethyldiammonium chloride) (PDDA), and polystyrenesulfonic acid (PSS) are prepared.
  • PVP polyvinylpyrrolidone
  • PVA polyvinyl alcohol
  • PAH polyallylamine hydrochloride
  • PDDA poly(diallyldimethyldiammonium chloride)
  • PSS polystyrenesulfonic acid
  • the amide functional group in polyvinylpyrrolidone is a bonding site of Prussian Blue and has a great effect on the crystallization of Prussian Blue.
  • PVP polyvinylpyrrolidone
  • the present inventors prepared a Prussian blue/polyvinylpyrrolidone nanoparticle (PB/PVP NP) composite to improve the stability and physicochemical properties of Prussian blue nanoparticles, and its stability, physical
  • PB/PVP NP Prussian blue/polyvinylpyrrolidone nanoparticle
  • the stability of Prussian Blue was further improved after coating with low molecular weight PVP.
  • the present invention was completed by confirming that one PB/PVP NP is the most stable, and has the best antioxidant power, active oxygen removal ability, anti-inflammatory effect and wound healing ability.
  • FIG. 1 A schematic diagram of a method for preparing a Prussian blue/polyvinylpyrrolidone nanoparticle composite according to the present invention is shown in FIG. 1 .
  • the Prussian blue/polyvinylpyrrolidone nanoparticle (PB/PVP NP) complex of the present invention is prepared in polyvinylpyrrolidone (PVP) micelles. It is a spherical structure in which Russian blue nanoparticles (PB NP) are evenly bound. At this time, the amide functional group in polyvinylpyrrolidone acts as a binding site for Prussian Blue.
  • the Prussian blue/polyvinylpyrrolidone nanoparticle (PB/PVP NP) composite of the present invention is composed of Prussian blue nanoparticle (PB NP) as polyvinylpyrrolidone It may be a spherical structure coated with (PVP).
  • An object of the present invention is to provide an antioxidant comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
  • Another object of the present invention is to provide an anti-inflammatory agent comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
  • Another object of the present invention is to provide a pharmaceutical composition comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
  • Another object of the present invention is to provide a quasi-drug composition or medical device comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
  • Another object of the present invention is to provide a cosmetic composition comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
  • Another object of the present invention is to provide a health functional food composition comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
  • the present invention provides an antioxidant comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
  • the molecular weight of the polyvinylpyrrolidone nanoparticles may be 1 to 50 kDa.
  • the molecular weight of the polyvinylpyrrolidone nanoparticles may be 10 kDa.
  • the size of the Prussian blue/polyvinylpyrrolidone nanoparticle complex may be 10 to 300 nm, preferably 20 to 200 nm.
  • the present invention provides an anti-inflammatory agent comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
  • the molecular weight of the polyvinylpyrrolidone nanoparticles may be 1 to 50 kDa.
  • the molecular weight of the polyvinylpyrrolidone nanoparticles may be 10 kDa.
  • the size of the Prussian blue/polyvinylpyrrolidone nanoparticle complex may be 10 to 300 nm, preferably 20 to 200 nm.
  • the present invention provides a pharmaceutical composition for preventing or treating diseases caused by excessive production of active oxygen, comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
  • the molecular weight of the polyvinylpyrrolidone nanoparticles may be 1 to 50 kDa.
  • the molecular weight of the polyvinylpyrrolidone nanoparticles may be 10 kDa.
  • the size of the Prussian blue/polyvinylpyrrolidone nanoparticle complex may be 10 to 300 nm, preferably 20 to 200 nm.
  • diseases caused by excessive production of reactive oxygen species are stroke, Parkinson's disease, Alzheimer's disease, aging, heart disease, ischemia, arteriosclerosis, skin disease, inflammation, rheumatism, autoimmune disease, It is characterized by asthma, hyperlipidemia, liver disease, diabetes, cancer, chronic ulcer, burn or wound.
  • the pharmaceutical composition according to the present invention may be administered to a patient by a general oral or parenteral administration method, and may be in any solid or liquid form.
  • the pharmaceutical composition according to the present invention may further include one or more pharmaceutically acceptable liquid or solid carriers.
  • it may be prepared using diluents or excipients such as commonly used fillers, extenders, binders, wetting agents, disintegrants, and surfactants.
  • the solid formulations may be tablets, pills, powders, granules, capsules, pellets, fine granules or powders, and these solid formulations may be prepared by adding carriers, excipients and/or diluents to the complex.
  • Examples of the carrier, excipient and/or diluent include lactose, sucrose, dextrose, mannitol, malitol, sorbitol, xylitol, erythritol (erithritol), starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, polyvinyl pyrrolidone (polyvinyl pyrrolidone), magnesium stearate and mineral oil.
  • lactose sucrose, dextrose, mannitol, malitol, sorbitol, xylitol, erythritol (erithritol), starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, polyvinyl pyrrolidone (polyvinyl pyrrolidone), magnesium stearate and mineral oil.
  • the liquid formulation may be a solution, suspension, or emulsion, and may include various excipients such as wetting agents, sweeteners, aromatics, and preservatives in addition to water and liquid paraffin, which are commonly used simple diluents.
  • Aqueous suspensions suitable for oral use may be prepared by dispersing the finely divided active ingredient in a viscous material such as natural or synthetic gums, resins, methyl cellulose, sodium carboxymethyl cellulose and other known suspending agents.
  • parenteral administration include injections, drops, infusions, ointments, sprays, suspensions, emulsions, suppositories, and the like.
  • Formulations for parenteral administration include sterilized aqueous solutions, non-aqueous solvents, suspensions, emulsions, freeze-dried formulations, and suppositories.
  • Propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate may be used as non-aqueous solvents and suspending agents.
  • As a base material for suppositories witepsol, macrogol, cacao butter, laurin paper, glycerogelatin, and the like may be used.
  • the present invention provides a quasi-drug composition for preventing or treating diseases caused by excessive production of reactive oxygen species, including a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
  • the molecular weight of the polyvinylpyrrolidone nanoparticles may be 1 to 50 kDa.
  • the molecular weight of the polyvinylpyrrolidone nanoparticles may be 10 kDa.
  • the size of the Prussian blue/polyvinylpyrrolidone nanoparticle complex may be 1 to 300 nm, preferably 20 to 200 nm.
  • diseases caused by excessive production of active oxygen include stroke, Parkinson's disease, Alzheimer's disease, aging, heart disease, ischemia, arteriosclerosis, skin disease, inflammation, arthritis, rheumatism, and autoimmune diseases. , asthma, hyperlipidemia, liver disease, diabetes, cancer, chronic ulcers, burns or wounds.
  • the quasi-drug composition is characterized in that it is a disinfectant cleanser, shower foam, gargreen, wet tissue, detergent soap, hand wash, humidifier filler, mask, ointment or filter filler.
  • the present invention provides a cosmetic composition for preventing or improving symptoms or diseases caused by active oxygen, comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
  • the symptom or disease caused by the active oxygen is characterized in that skin aging, wrinkle formation, skin pigmentation, atopy, acne, psoriasis or eczema.
  • the molecular weight of the polyvinylpyrrolidone nanoparticles may be 1 to 50 kDa.
  • the molecular weight of the polyvinylpyrrolidone nanoparticles may be 10 kDa.
  • the size of the Prussian blue/polyvinylpyrrolidone nanoparticle complex may be 10 to 300 nm, preferably 20 to 200 nm.
  • the cosmetic may be prepared in the form of an ampoule, cream, lotion, lotion, essence or pack.
  • a carrier used in normal formulation such as dextrin and cyclodextrin, and other arbitrary auxiliary agents may be added.
  • the present invention provides a health functional food composition comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
  • the molecular weight of the polyvinylpyrrolidone nanoparticles may be 1 to 50 kDa.
  • the molecular weight of the polyvinylpyrrolidone nanoparticles may be 10 kDa.
  • the size of the Prussian blue/polyvinylpyrrolidone nanoparticle complex may be 10 to 300 nm, preferably 20 to 200 nm.
  • the present invention provides a medical device containing a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
  • the medical device may be a wound covering material.
  • the Prussian blue/polyvinylpyrrolidone nanoparticle complex of the present invention exhibits excellent tissue regeneration ability even when used at a low concentration.
  • the molecular weight of the polyvinylpyrrolidone nanoparticles may be 1 to 50 kDa.
  • the molecular weight of the polyvinylpyrrolidone nanoparticles may be 10 kDa.
  • the size of the Prussian blue/polyvinylpyrrolidone nanoparticle complex may be 10 to 300 nm, preferably 20 to 200 nm. .
  • the Prussian Blue/Polyvinylpyrrolidone nanoparticle complex according to the present invention is a substance with excellent biocompatibility and stable antioxidant effect, and has excellent active oxygen scavenging ability for inflammatory liver disease, neurodegenerative disease, cancer, diabetes, degenerative arthritis, It can improve and treat active oxygen-derived diseases such as rheumatoid arthritis, autoimmune diseases, and asthma, and has excellent anti-inflammatory effects and can exhibit inflammation relief and improvement effects by controlling oxidative stress that is overexpressed in inflammation. It is expected to be used as a platform that can be widely used in anti-aging cosmetics, atopy-improving cosmetics, health functional foods, pharmaceuticals, and medical devices (including wound dressings) due to its excellent regenerative ability.
  • FIG. 1 is a schematic view of a method for preparing Prussian blue/polyvinylpyrrolidone nanoparticles (PB/PVP NP) by molecular weight and antioxidant efficacy according to the present invention.
  • PB/PVP NP Prussian blue/polyvinylpyrrolidone nanoparticles
  • PB NPs Prussian blue nanoparticles
  • PB/PVP NPs Prussian blue nanoparticles
  • PB NPs Prussian blue nanoparticles
  • PB/PVP NPs Prussian blue nanoparticles
  • PB NPs Prussian blue nanoparticles
  • PB NPs Prussian blue nanoparticles
  • PB/PVP NPs Prussian blue nanoparticles
  • Figure 6 shows the UV-Vis spectrum of Prussian blue nanoparticles (PB NPs) and PB / PVP NPs by molecular weight
  • PB NPs Prussian blue nanoparticles
  • PB/PVP10k NPs PB/PVP360k NPs.
  • PB NPs Prussian blue nanoparticles
  • PB/PVP NPs Prussian blue/polyvinylpyrrolidone nanoparticles
  • PB NPs Prussian blue nanoparticles
  • PB/PVP NPs Prussian blue/polyvinylpyrrolidone nanoparticles
  • FIG. 10 shows changes in dispersion values of Prussian blue nanoparticles (PB NPs) and Prussian blue/polyvinylpyrrolidone nanoparticles (PB/PVP NPs) in tertiary distilled water.
  • PB NPs Prussian blue nanoparticles
  • PB/PVP NPs Prussian blue/polyvinylpyrrolidone nanoparticles
  • PB NPs Prussian blue nanoparticles
  • PB/PVP NPs Prussian blue/polyvinylpyrrolidone nanoparticles
  • Figure 16 compares the results of cell wound healing over time between the control group and the PB/PVP10k NPs of the present invention.
  • prevention refers to any action that suppresses or delays the onset of atopic disease by administration of the pharmaceutical composition according to the present invention.
  • treat means temporarily or permanently alleviating symptoms, removing the cause of symptoms, or preventing or delaying the onset of symptoms of the disease or condition. do.
  • the term “improvement” refers to any action that reduces a parameter associated with an abnormal state, eg, the severity of a symptom.
  • the term "pharmaceutically acceptable” means that the benefit/risk ratio is reasonable without excessive toxicity, irritation, allergic reaction or other problems or complications, so that it is suitable for use in contact with the tissue of a subject (eg, human) and Means within the scope of sound medical judgment
  • Polyvinylpyrrolidone (PVP) of 4 different molecular weights (10, 40, 360, 1300 kDa; 75 mg) was dissolved in 3 ml of tertiary distilled water to prepare a polymer solution, followed by potassium ferricyanide (5 mM). ) was added and reacted while stirring at 400 rpm for 30 minutes at room temperature, then increasing the stirring speed to 530 rpm and slowly adding 1 ml of iron chloride tetrahydrate (5mM) dropwise to the reaction solution while reacting for 1 hour to react for Prussian blue/polyvinyl Pyrrolidone nanoparticles (PB/PVP NP) were prepared.
  • PVP Prussian blue/polyvinyl Pyrrolidone nanoparticles
  • the prepared PB/PVP NPs are named PB/PVP10k NPs, PB/PVP40k NPs, PB/PVP360k NPs, and PB/PVP1300k NPs according to the molecular weight of PVP.
  • the PB/PVP10k NPs refer to Prussian blue/polyvinylpyrrolidone nanoparticles coated with 10 kDa PVP.
  • Prussian blue nanoparticles were prepared in the same manner as in Example 1, except that only 3 ml of tertiary distilled water was used instead of the polymer solution.
  • the prepared PB NPs and PB/PVP NPs were purified using an Amicon Ultra-15 filter, freeze-dried for 3 days, and then their properties were evaluated as follows.
  • the size, dispersion and surface charge of the prepared PB NPs and PB/PVP NPs were analyzed using Zetasizer (ELSZ-2000, Otsuka) equipment.
  • both the PB NPs and the PB/PVP NPs exhibited the unique blue color of Prussian Blue.
  • the size of the PB NPs was about 130 nm.
  • the size of the 10 kDa PVP-coated PB/PVP NPs was about 80 nm, and the size of the 360 kDa and 1300 kDa PVP-coated PB/PVP NPs was about 135 nm, similar to the PB NPs.
  • the dispersion value (PDI) increased after PVP coating, but all were uniform at 0.3 or less.
  • the surface charge of the PB NP was about -40 mV, and the surface charge of the PB/PVP NP showed a surface charge close to neutral as the molecular weight of the polymer increased.
  • the absorbance of PB/PVP NPs in the NIR wavelength range was analyzed using UV-Vis spectroscopy.
  • PB NPs and PB/PVP NPs were observed using a transmission electron microscope (TEM).
  • PB NPs, low molecular weight PB/PVP10k NPs, and high molecular weight PB/PVP360k NPs all had spherical structures.
  • metal nanoparticles were agglomerated and observed as a lump.
  • PB/PVP NPs were prepared by evenly combining PB NPs into PVP micelles. Similar to the analysis through Zetasizer, it can be seen that the low molecular weight PB/PVP NPs are smaller than the high molecular weight PB/PVP NPs.
  • PB NPs and PB/PVP NPs were dispersed in tertiary distilled water, and then stored in an incubator at 37 ° C and 100 rpm for 0 and 1 week. Changes in the properties, size, dispersion and surface charge of nanoparticles Analyzed and the results are shown in Figures 8 to 11.
  • the size of PB/PVP10k NPs did not change even after 1 week, but the sizes of PB NPs and other PB/PVP NPs were reduced compared to before storage in the incubator. This is due to the dissolution of the unstable Prussian Blue. Therefore, the size change of the high molecular weight PB/PVP NPs, which had a large color change due to the dissolution of Prussian blue, was noticeably large.
  • the active oxygen scavenging ability of the prepared PB/PVP NPs was evaluated.
  • 0.1mM EDTA, 0.1mM FeCl 3 , 1mM H 2 O 2 , and 3.75mM 2-deoxy-D-ribose were prepared in tertiary distilled water. 100 ⁇ l of this was sequentially put into a 15 ml tube, and then 1 ml of each of PB NP and PB/PVP NP (0.1 mg/ml) was added. At this time, tertiary distilled water was added instead of the H 2 O 2 and nanoparticle solution to the negative control, and tertiary distilled water was added instead of the nanoparticles to the positive control.
  • the cytotoxicity of the prepared PB/PVP NPs was evaluated.
  • fibroblasts were cultured in DMEM supplemented with 10% FBS and 1% AA.
  • NIH 3T3 (1X10 4 cells/well) was dispensed into a 96-well plate and cultured in an incubator at 37°C and 5% CO 2 environment for 12 hours.
  • PB/PVP10k NPs were treated at concentrations of 0, 0.01, 0.1, 0.5, 1.0, and 5.0 mg/ml and stored in an incubator for 24 hours. Thereafter, the CCK-8 solution was treated and reacted for one hour in an incubator, and the absorbance of the supernatant was measured at 450 nm, and the results are shown in FIG. 13 .
  • the intracellular active oxygen radical scavenging ability of the prepared PB/PVP NPs was evaluated.
  • NIH 3T3 was cultured in DMEM supplemented with 10% FBS. After dispensing NIH 3T3 (1X10 4 cells/well) into 96 well-plates, they were cultured in an incubator for 12 hours. Thereafter, 100 ⁇ l of PB/PVP10k NPs (0, 10, 100 pg/ml) and H 2 O 2 (25 ⁇ M) at various concentrations were treated and reacted in an incubator for 4 hours. At this time, only the cell culture medium was treated instead of the nanoparticles and H 2 O 2 in the control group. After 4 hours, the treated sample was suctioned and the remaining sample was washed once more with PBS.
  • H2DCFDA (10 ⁇ M) was treated and reacted in an incubator for 90 minutes. Then, by measuring the fluorescence (ex / 480nm, em / 535nm) of the DCF generated by reacting with the active oxygen radical to evaluate the active oxygen radical removal ability of the nanoparticles, the results are shown in FIG. 14 .
  • the intracellular anti-inflammatory effect of the prepared PB/PVP NP was evaluated.
  • Raw 264.7 cells were cultured in DMEM supplemented with 10% FBS.
  • Raw 264.7 (1X10 4 cells/well) was dispensed into 96 well-plates and cultured in an incubator for 12 hours. Thereafter, the cells were treated with 100 ⁇ l of PB/PVP10k NPs (0, 10, 100 ⁇ g/ml) and LPS (100 ng/ml) at various concentrations and allowed to react in an incubator for 24 hours. At this time, only the cell culture medium was treated instead of nanoparticles and LPS in the control group. After 24 hours, only the supernatant was transferred to another 96 well-plate and dispensed by 100 ⁇ l.
  • NIH 3T3 was cultured in DMEM supplemented with 10% FBS and 1% AA.
  • NIH 3T3 (1X10 5 cells/well) was dispensed into 24 well-plates and cultured in an incubator for 12 hours.
  • a scratch wound was prepared using a sterile p1000 pipet tip.
  • PB/PVP10k NPs (0, 0.01, 0.1, 0.5, 1mg/ml) are dispersed in the culture medium without FBS, respectively, treated with cells to create a starvation environment, and then placed in an incubator for a set period of time. After culturing and observing the degree of cell migration through a microscope, the results are shown in FIG. 16.

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Abstract

The present invention relates to a Prussian blue/polyvinylpyrrolidone nanoparticle composite for removal of reactive oxygen species and use thereof. The Prussian blue/polyvinylpyrrolidone nanoparticle composite according to the present invention is a substance having excellent biocompatibility and stable antioxidative efficacy and can be effectively used in the development of an antioxidant, a pharmaceutical composition, a quasi-drug composition, a cosmetic composition, or a food composition due to superior reactive oxygen species scavenging capacity.

Description

프러시안 블루/폴리비닐피롤리돈 나노입자 복합체 및 이의 용도Prussian blue/polyvinylpyrrolidone nanoparticle composite and uses thereof
본 발명은 활성산소 제거를 위한 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체 및 이의 용도에 관한 것이다.The present invention relates to a Prussian blue/polyvinylpyrrolidone nanoparticle composite and its use for scavenging active oxygen.
프러시안 블루는 철분 페로시안 화합물(ferric ferrocyanide, Fe4[Fe(CN)6]3)로 고스핀 철 (Fe3+) 이온과 저스핀 철 (Fe2+) 이온 간 전하/전자 이동이 가능해 특별한 물리화학적 특성을 갖는다. 프러시안 블루의 Fe2+-CN-Fe3+ 커플링은 근적외선 파장대의 빛을 흡수해 특유의 파란빛을 띄어 중요한 무기안료로 사용되어 왔다. 또한 프러시안 블루는 방사성 세슘과 선택적으로 흡착할 수 있어 방사능 동위원소 제거용 의약으로서 라디오가르다제(Radiogardase)가 미국 식품의약국 (FDA)로부터 의약품 승인을 받았다. 더 나아가 프러시안 블루의 전기화학적 산화 환원을 이용해 다양한 질병의 치료 소재로 제시되고 있다.Prussian Blue is an iron ferrocyanide (Fe 4 [Fe(CN) 6 ] 3 ), which enables charge/electron transfer between high-spin iron (Fe 3+ ) and low-spin iron (Fe 2+ ) ions. It has special physicochemical properties. Prussian Blue's Fe 2+ -CN-Fe 3+ coupling has been used as an important inorganic pigment because it absorbs light in the near-infrared wavelength range and emits a unique blue color. In addition, Prussian Blue can selectively adsorb radioactive cesium, so Radiogardase as a drug for removing radioactive isotopes has received drug approval from the US Food and Drug Administration (FDA). Furthermore, it is proposed as a treatment material for various diseases by using the electrochemical redox of Prussian blue.
인간의 약 80%의 질병이 활성산소로부터 유래되는데 최근 프러시안 블루의 전기화학적 산화 환원이 활성산소 제거에 효과적이라는 연구 결과가 있었다. 즉, 프러시안 블루는 프러시안 화이트나 프러시안 그린으로 산화 및 환원할 수 있는데 이는 항산화 효소(peroxidase(POD), catalase(CAT), superoxide dismutase(SOD) 등)의 작용기작과 유사해 활성산소 유래 질병 치료에 효과적일 것이라는 가능성이 제기됐다.About 80% of human diseases are caused by active oxygen, and a recent study showed that the electrochemical oxidation-reduction of Prussian blue is effective in removing active oxygen. In other words, Prussian blue can be oxidized and reduced to Prussian white or Prussian green, which is similar to the mechanism of action of antioxidant enzymes (peroxidase (POD), catalase (CAT), superoxide dismutase (SOD), etc.) Possibility has been raised that it will be effective in treating diseases.
그러나 프러시안 블루는 생체적합성 및 활성산소제거능이 뛰어남에도 불구하고 생체 내 안정성이 낮아 그 활용에 어려움이 있다. 따라서 프러시안 블루의 안정성을 향상시키기 위해 다양한 고분자 기반 프러시안 블루 복합체가 개발되었다. 예를 들어, 폴리비닐피롤리돈(PVP), 폴리비닐알코올(PVA), 폴리알릴아민 수화염화물(PAH), poly(diallyldimethyldiammonium chloride)(PDDA), 폴리스티렌설폰산(PSS)와 같은 여러 고분자를 프러시안 블루의 템플레이트(template)로 사용하는 프러시안 블루 복합체가 개발되었다. 이 중 생체적합성이 뛰어난 폴리비닐피롤리돈을 템플레이트로 사용하는 프러시안 블루 복합체의 경우 폴리비닐피롤리돈 내의 아마이드 관능기가 프러시안 블루의 결합부위로 프러시안 블루의 결정화에 많은 영향을 미친다. 이와 관련하여 프러시안 블루 대비 폴리비닐피롤리돈(PVP)의 양을 조절하여 복합체의 크기를 조절한 연구 결과가 있다(H. Ming, N. L. K. Torad, Y. Chiang, K. C. W. Wu, and Y. Yamauchi, "Size- and shape-controlled synthesis of Prussian blue nanoparticles by a polyvinylpyrrolidone-assisted crystallization process," CrystEngComm, vol. 14, pp. 3387-3396, 2012.)However, although Prussian blue has excellent biocompatibility and reactive oxygen scavenging ability, it is difficult to use it due to its low in vivo stability. Therefore, various polymer-based Prussian blue composites have been developed to improve the stability of Prussian blue. For example, several polymers such as polyvinylpyrrolidone (PVP), polyvinyl alcohol (PVA), polyallylamine hydrochloride (PAH), poly(diallyldimethyldiammonium chloride) (PDDA), and polystyrenesulfonic acid (PSS) are prepared. A Prussian Blue complex was developed to use as a template for Russian Blue. Among them, in the case of the Prussian Blue complex using polyvinylpyrrolidone having excellent biocompatibility as a template, the amide functional group in polyvinylpyrrolidone is a bonding site of Prussian Blue and has a great effect on the crystallization of Prussian Blue. In this regard, there are studies in which the size of the composite was adjusted by adjusting the amount of polyvinylpyrrolidone (PVP) compared to Prussian Blue (H. Ming, N. L. K. Torad, Y. Chiang, K. C. W. Wu, and Y. Yamauchi, "Size- and shape-controlled synthesis of Prussian blue nanoparticles by a polyvinylpyrrolidone-assisted crystallization process," CrystEngComm, vol. 14, pp. 3387-3396, 2012.)
이러한 고분자 템플레이트를 이용한 프러시안 블루의 안정성 향상 및 물리화학적 특성 조절은 복합체의 효능에도 영향이 있어 최적화가 필요하다. The stability improvement and physicochemical property control of Prussian Blue using such a polymer template also affect the efficacy of the composite, so optimization is required.
[선행기술][Prior art]
H. Ming, N. L. K. Torad, Y. Chiang, K. C. W. Wu, and Y. Yamauchi, "Size- and shape-controlled synthesis of Prussian blue nanoparticles by a polyvinylpyrrolidone-assisted crystallization process," CrystEngComm, vol. 14, pp. 3387-3396, 2012.H. Ming, N. L. K. Torad, Y. Chiang, K. C. W. Wu, and Y. Yamauchi, "Size- and shape-controlled synthesis of Prussian blue nanoparticles by a polyvinylpyrrolidone-assisted crystallization process," CrystEngComm, vol. 14, p. 3387-3396, 2012.
[관련된 국가정부과제][Related national government tasks]
과제고유번호 : 2021023867Assignment identification number: 2021023867
과제번호 : 2021M3E5E7023867Assignment number: 2021M3E5E7023867
부처명 : 과학기술정보통신부Department Name: Ministry of Science and ICT
연구관리전문기관 : 한국연구재단Research management institution: National Research Foundation of Korea
연구사업명 : 바이오·의료기술개발사업Research Project Name: Bio/Medical Technology Development Project
연구과제명 : 주입형 프러시안블루 복합체 기반 퇴행성관절염 치료제 개발Title of research project: Development of treatment for degenerative arthritis based on injectable Prussian Blue complex
주관기관 : 한국세라믹기술원Organized by : Korea Institute of Ceramic Technology
연구기간 : 2021.04.01 ~ 2023.12.31Research period: 2021.04.01 ~ 2023.12.31
기여율(%) : 100%Contribution rate (%): 100%
이에 본 발명자들은 프러시안 블루 나노 입자들의 안정성 및 물리화학적 특성을 향상시키기 위해 프러시안 블루/폴리비닐피롤리돈 나노입자(Prussian blue/polyvinylpyrrolidone nanoparticle, PB/PVP NP) 복합체를 제조하고 이의 안정성, 물리화학적 특성, 활성산소제거능, 세포독성, 활성산소라디칼 제거능, 항염증 효과, in vitro 상처 치유 효과를 시험한 결과, 저분자량 PVP 코팅 후 프러시안 블루의 안정성이 더 향상되었고 그 중 10kDa의 PVP를 코팅한 PB/PVP NP가 가장 안정하고, 항산화력, 활성산소제거 능력, 항염증 효과 및 상처 치유 능력이 가장 우수함을 확인함으로써 본 발명을 완성하였다. Accordingly, the present inventors prepared a Prussian blue/polyvinylpyrrolidone nanoparticle (PB/PVP NP) composite to improve the stability and physicochemical properties of Prussian blue nanoparticles, and its stability, physical As a result of testing chemical properties, active oxygen scavenging ability, cytotoxicity, active oxygen radical scavenging ability, anti-inflammatory effect, and in vitro wound healing effect, the stability of Prussian Blue was further improved after coating with low molecular weight PVP. The present invention was completed by confirming that one PB/PVP NP is the most stable, and has the best antioxidant power, active oxygen removal ability, anti-inflammatory effect and wound healing ability.
본 발명에 따른 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체의 제조 방법에 대한 모식도를 도 1에 나타내었다.A schematic diagram of a method for preparing a Prussian blue/polyvinylpyrrolidone nanoparticle composite according to the present invention is shown in FIG. 1 .
도 1 및 도 7을 참조하면 본 발명의 프러시안 블루/폴리비닐피롤리돈 나노입자(Prussian blue/polyvinylpyrrolidone nanoparticle, PB/PVP NP) 복합체는 폴리비닐피롤리돈(PVP) 마이셀(micelle) 안에 프러시안 블루 나노입자(Prussian blue nanoparticle, PB NP)가 고르게 결합된 구형 구조체이다. 이때 폴리비닐피롤리돈 내의 아마이드 관능기는 프러시안 블루의 결합부위로 작용한다. 결과적으로, 본 발명의 프러시안 블루/폴리비닐피롤리돈 나노입자(Prussian blue/polyvinylpyrrolidone nanoparticle, PB/PVP NP) 복합체는 프러시안 블루 나노입자(Prussian blue nanoparticle, PB NP)를 폴리비닐피롤리돈(PVP)로 코팅한 구형 구조체일 수 있다.1 and 7, the Prussian blue/polyvinylpyrrolidone nanoparticle (PB/PVP NP) complex of the present invention is prepared in polyvinylpyrrolidone (PVP) micelles. It is a spherical structure in which Russian blue nanoparticles (PB NP) are evenly bound. At this time, the amide functional group in polyvinylpyrrolidone acts as a binding site for Prussian Blue. As a result, the Prussian blue/polyvinylpyrrolidone nanoparticle (PB/PVP NP) composite of the present invention is composed of Prussian blue nanoparticle (PB NP) as polyvinylpyrrolidone It may be a spherical structure coated with (PVP).
본 발명은 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체를 유효 성분으로 포함하는 항산화제를 제공하는 것을 목적으로 한다.An object of the present invention is to provide an antioxidant comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
본 발명은 또한 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체를 유효 성분으로 포함하는 항염증제를 제공하는 것을 목적으로 한다.Another object of the present invention is to provide an anti-inflammatory agent comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
본 발명은 또한 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체를 유효 성분으로 포함하는 약제학적 조성물을 제공하는 것을 목적으로 한다.Another object of the present invention is to provide a pharmaceutical composition comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
본 발명은 또한 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체를 유효 성분으로 포함하는 의약외품 조성물 또는 의료기기를 제공하는 것을 목적으로 한다.Another object of the present invention is to provide a quasi-drug composition or medical device comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
본 발명은 또한 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체를 유효 성분으로 포함하는 화장료 조성물을 제공하는 것을 목적으로 한다.Another object of the present invention is to provide a cosmetic composition comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
본 발명은 또한 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체를 유효 성분으로 포함하는 건강기능식품 조성물을 제공하는 것을 목적으로 한다.Another object of the present invention is to provide a health functional food composition comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
제1 구현예에 따르면, According to the first embodiment,
본 발명은 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체를 유효 성분으로 포함하는 항산화제를 제공한다.The present invention provides an antioxidant comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
본 발명의 일 실시예에 따르면, 상기 폴리비닐피롤리돈 나노입자의 분자량은 1 내지 50kDa일 수 있다. 바람직하게는, 상기 폴리비닐피롤리돈 나노입자의 분자량은 10kDa일 수 있다. According to one embodiment of the present invention, the molecular weight of the polyvinylpyrrolidone nanoparticles may be 1 to 50 kDa. Preferably, the molecular weight of the polyvinylpyrrolidone nanoparticles may be 10 kDa.
본 발명의 일 실시예에 따르면, 상기 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체의 크기는 10 내지 300 nm, 바람직하게는 20 내지 200 nm일 수 있다.According to one embodiment of the present invention, the size of the Prussian blue/polyvinylpyrrolidone nanoparticle complex may be 10 to 300 nm, preferably 20 to 200 nm.
제2 구현예에 따르면, According to the second embodiment,
본 발명은 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체를 유효 성분으로 포함하는 항염증제를 제공한다.The present invention provides an anti-inflammatory agent comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
본 발명의 일 실시예에 따르면, 상기 폴리비닐피롤리돈 나노입자의 분자량은 1 내지 50kDa일 수 있다. 바람직하게는, 상기 폴리비닐피롤리돈 나노입자의 분자량은 10kDa일 수 있다. According to one embodiment of the present invention, the molecular weight of the polyvinylpyrrolidone nanoparticles may be 1 to 50 kDa. Preferably, the molecular weight of the polyvinylpyrrolidone nanoparticles may be 10 kDa.
본 발명의 일 실시예에 따르면, 상기 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체의 크기는 10 내지 300 nm, 바람직하게는 20 내지 200 nm일 수 있다.According to one embodiment of the present invention, the size of the Prussian blue/polyvinylpyrrolidone nanoparticle complex may be 10 to 300 nm, preferably 20 to 200 nm.
제3구현예에 따르면,According to the third embodiment,
본 발명은 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체를 유효 성분으로 포함하는 활성산소 과잉생성으로 인해 유발되는 질환의 예방 또는 치료용 약제학적 조성물을 제공한다.The present invention provides a pharmaceutical composition for preventing or treating diseases caused by excessive production of active oxygen, comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
본 발명의 일 실시예에 따르면, 상기 폴리비닐피롤리돈 나노입자의 분자량은 1 내지 50kDa일 수 있다. 바람직하게는, 상기 폴리비닐피롤리돈 나노입자의 분자량은 10kDa일 수 있다.According to one embodiment of the present invention, the molecular weight of the polyvinylpyrrolidone nanoparticles may be 1 to 50 kDa. Preferably, the molecular weight of the polyvinylpyrrolidone nanoparticles may be 10 kDa.
본 발명의 일 실시예에 따르면, 상기 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체의 크기는 10 내지 300 nm, 바람직하게는 20 내지 200 nm일 수 있다.According to one embodiment of the present invention, the size of the Prussian blue/polyvinylpyrrolidone nanoparticle complex may be 10 to 300 nm, preferably 20 to 200 nm.
본 발명에 따른 약제학적 조성물에 있어서, 상기 활성산소 과잉생성으로 인해 유발되는 질환은 뇌졸중, 파킨슨병, 알츠하이머병, 노화, 심장질환, 허혈, 동맥경화, 피부질환, 염증, 류마티스, 자가면역질환, 천식, 고지혈증, 간질환, 당뇨병, 암, 만성궤양, 화상 또는 창상인 것을 특징으로 한다.In the pharmaceutical composition according to the present invention, diseases caused by excessive production of reactive oxygen species are stroke, Parkinson's disease, Alzheimer's disease, aging, heart disease, ischemia, arteriosclerosis, skin disease, inflammation, rheumatism, autoimmune disease, It is characterized by asthma, hyperlipidemia, liver disease, diabetes, cancer, chronic ulcer, burn or wound.
본 발명에 따른 약제학적 조성물은 일반적인 경구 또는 비경구 투여 방법으로 환자에게 투여될 수 있으며, 고체 또는 액체 형태 어떠한 형태로도 가능하다. 또한, 본 발명에 따른 약제학적 조성물은 약제학적으로 허용 가능한 1종 이상의 액체 또는 고체 담체를 더 포함할 수 있다. 또한 보통 사용하는 충진제, 증량제, 결합제, 습윤제, 붕해제, 계면활성제 등의 희석제 또는 부형제를 사용하여 조제될 수 있다. 상기 고체 형태의 제제에는 정제, 환제, 산제, 과립제, 캡슐제, 펠렛제, 세립제 또는 분제일 수 있으며, 이러한 고형제제는 상기 복합체에 담체, 부형제 및/또는 희석제를 첨가하여 조제할 수 있다. 상기 담체, 부형제 및/또는 희석제로는 락토오스(lactose), 수크로오스(sucrose), 덱스트로오스(dextrose), 만니톨(mannitol), 말리톨(malitol), 소르비톨(sorbitol), 자일리톨(xylitol), 에리트리톨(erithritol), 전분(starch), 아카시아고무(acacia rubber), 알지네이트(alginate), 젤라틴(gelatin), 칼슘 포스페이트(calcium phosphate), 칼슘 실리케이트(calcium silicate), 셀룰로오스(cellulose), 폴리비닐 피롤리돈(polyvinyl pyrrolidone), 마그네슘 스테아레이트(magnesium stearate) 및 광물유 등이 있다. 상기 액체 형태의 제제는 용액, 현탁액 또는 유탁액일 수 있으며, 흔히 사용되는 단순 희석제인 물, 리퀴드 파라핀 이외에 여러 가지 부형제, 예를 들면 습윤제, 감미제, 방향제, 보존제 등이 포함될 수 있다. 경구용으로 적당한 수성 현탁제로는 미분된 활성 성분을 천연 또는 합성검, 수지, 메틸셀룰로오스(methyl cellulose), 소디움카르복시메틸셀룰로오스(Sodium carboxymethyl cellulose) 및 공지의 현탁제와 같은 점성 물질에 분산시켜 제조될 수 있다. 비경구 투여제로는 주사제, 점적제, 수액, 연고, 스프레이제, 현탁제, 유제, 좌제 등을 들 수 있다. 비경구 투여를 위한 제제에는 멸균된 수용액, 비수성용제, 현탁제, 유제, 동결건조 제제 및 좌제가 포함된다. 비수성용제, 현탁제로는 프로필렌글리콜(propylene glycol), 폴리에틸렌글리콜(polyethylene glycol), 올리브 오일과 같은 식물성 오일, 에틸올레이트(ethyl oleate)와 같은 주사 가능한 에스테르 등이 사용될 수 있다. 좌제의 기제로는 위텝솔(witepsol), 마크로골(macrogol), 카카오지, 라우린지, 글리세로젤라틴(glycerogelatin) 등이 사용될 수 있다.The pharmaceutical composition according to the present invention may be administered to a patient by a general oral or parenteral administration method, and may be in any solid or liquid form. In addition, the pharmaceutical composition according to the present invention may further include one or more pharmaceutically acceptable liquid or solid carriers. In addition, it may be prepared using diluents or excipients such as commonly used fillers, extenders, binders, wetting agents, disintegrants, and surfactants. The solid formulations may be tablets, pills, powders, granules, capsules, pellets, fine granules or powders, and these solid formulations may be prepared by adding carriers, excipients and/or diluents to the complex. Examples of the carrier, excipient and/or diluent include lactose, sucrose, dextrose, mannitol, malitol, sorbitol, xylitol, erythritol (erithritol), starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, polyvinyl pyrrolidone (polyvinyl pyrrolidone), magnesium stearate and mineral oil. The liquid formulation may be a solution, suspension, or emulsion, and may include various excipients such as wetting agents, sweeteners, aromatics, and preservatives in addition to water and liquid paraffin, which are commonly used simple diluents. Aqueous suspensions suitable for oral use may be prepared by dispersing the finely divided active ingredient in a viscous material such as natural or synthetic gums, resins, methyl cellulose, sodium carboxymethyl cellulose and other known suspending agents. can Examples of parenteral administration include injections, drops, infusions, ointments, sprays, suspensions, emulsions, suppositories, and the like. Formulations for parenteral administration include sterilized aqueous solutions, non-aqueous solvents, suspensions, emulsions, freeze-dried formulations, and suppositories. Propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate may be used as non-aqueous solvents and suspending agents. As a base material for suppositories, witepsol, macrogol, cacao butter, laurin paper, glycerogelatin, and the like may be used.
제4구현예에 따르면, 본 발명은 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체를 유효 성분으로 포함하는 활성산소 과잉생성으로 인해 유발되는 질환의 예방 또는 치료용 의약외품 조성물을 제공한다.According to a fourth embodiment, the present invention provides a quasi-drug composition for preventing or treating diseases caused by excessive production of reactive oxygen species, including a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
본 발명의 일 실시예에 따르면, 상기 폴리비닐피롤리돈 나노입자의 분자량은 1 내지 50kDa일 수 있다. 바람직하게는, 상기 폴리비닐피롤리돈 나노입자의 분자량은 10kDa일 수 있다.According to one embodiment of the present invention, the molecular weight of the polyvinylpyrrolidone nanoparticles may be 1 to 50 kDa. Preferably, the molecular weight of the polyvinylpyrrolidone nanoparticles may be 10 kDa.
본 발명의 일 실시예에 따르면, 상기 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체의 크기는 1 내지 300 nm, 바람직하게는 20 내지 200 nm일 수 있다.According to one embodiment of the present invention, the size of the Prussian blue/polyvinylpyrrolidone nanoparticle complex may be 1 to 300 nm, preferably 20 to 200 nm.
본 발명에 따른 의약외품 조성물에 있어서, 상기 활성산소 과잉생성으로 인해 유발되는 질환은 뇌졸중, 파킨슨병, 알츠하이머병, 노화, 심장질환, 허혈, 동맥경화, 피부질환, 염증, 관절염, 류마티스, 자가면역질환, 천식, 고지혈증, 간질환, 당뇨병, 암, 만성궤양, 화상 또는 창상인 것을 특징으로 한다.In the quasi-drug composition according to the present invention, diseases caused by excessive production of active oxygen include stroke, Parkinson's disease, Alzheimer's disease, aging, heart disease, ischemia, arteriosclerosis, skin disease, inflammation, arthritis, rheumatism, and autoimmune diseases. , asthma, hyperlipidemia, liver disease, diabetes, cancer, chronic ulcers, burns or wounds.
본 발명에 따른 의약외품 조성물에 있어서, 상기 의약외품 조성물은 소독 청결제, 샤워폼, 가그린, 물티슈, 세제 비누, 핸드 워시, 가습기 충진제, 마스크, 연고제 또는 필터 충진제인 것을 특징으로 한다.In the quasi-drug composition according to the present invention, the quasi-drug composition is characterized in that it is a disinfectant cleanser, shower foam, gargreen, wet tissue, detergent soap, hand wash, humidifier filler, mask, ointment or filter filler.
제5구현예에 따르면,According to the fifth embodiment,
본 발명은 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체를 유효 성분으로 포함하는 활성산소로 유발되는 증상 또는 질환의 예방 또는 개선용 화장료 조성물을 제공한다.The present invention provides a cosmetic composition for preventing or improving symptoms or diseases caused by active oxygen, comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
본 발명에 따른 화장료 조성물에 있어서, 상기 활성산소로 유발되는 증상 또는 질환은 피부노화, 주름생성, 피부색소침착, 아토피, 여드름, 건선 또는 습진인 것을 특징으로 한다.In the cosmetic composition according to the present invention, the symptom or disease caused by the active oxygen is characterized in that skin aging, wrinkle formation, skin pigmentation, atopy, acne, psoriasis or eczema.
본 발명의 일 실시예에 따르면, 상기 폴리비닐피롤리돈 나노입자의 분자량은 1 내지 50kDa일 수 있다. 바람직하게는, 상기 폴리비닐피롤리돈 나노입자의 분자량은 10kDa일 수 있다.According to one embodiment of the present invention, the molecular weight of the polyvinylpyrrolidone nanoparticles may be 1 to 50 kDa. Preferably, the molecular weight of the polyvinylpyrrolidone nanoparticles may be 10 kDa.
본 발명의 일 실시예에 따르면, 상기 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체의 크기는 10 내지 300 nm, 바람직하게는 20 내지 200 nm일 수 있다.According to one embodiment of the present invention, the size of the Prussian blue/polyvinylpyrrolidone nanoparticle complex may be 10 to 300 nm, preferably 20 to 200 nm.
본 발명에 따른 화장료 조성물에 있어서, 상기 화장료는 앰플, 크림, 로션, 화장수, 에센스 또는 팩의 형태로 제조될 수 있다. 또한, 보존이나 취급을 용이하게 하기 위하여 덱스트린(dextrin), 사이클로덱스트린(cyclodextrin) 등의 통상 제제화에 사용되는 캐리어, 그 밖의 임의의 조제를 부가하여도 좋다.In the cosmetic composition according to the present invention, the cosmetic may be prepared in the form of an ampoule, cream, lotion, lotion, essence or pack. In addition, in order to facilitate preservation and handling, a carrier used in normal formulation, such as dextrin and cyclodextrin, and other arbitrary auxiliary agents may be added.
제6구현예에 따르면,According to the sixth embodiment,
본 발명은 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체를 유효 성분으로 포함하는 건강기능식품 조성물을 제공한다.The present invention provides a health functional food composition comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
본 발명의 일 실시예에 따르면, 상기 폴리비닐피롤리돈 나노입자의 분자량은 1 내지 50kDa일 수 있다. 바람직하게는, 상기 폴리비닐피롤리돈 나노입자의 분자량은 10kDa일 수 있다.According to one embodiment of the present invention, the molecular weight of the polyvinylpyrrolidone nanoparticles may be 1 to 50 kDa. Preferably, the molecular weight of the polyvinylpyrrolidone nanoparticles may be 10 kDa.
본 발명의 일 실시예에 따르면, 상기 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체의 크기는 10 내지 300 nm, 바람직하게는 20 내지 200 nm일 수 있다.According to one embodiment of the present invention, the size of the Prussian blue/polyvinylpyrrolidone nanoparticle complex may be 10 to 300 nm, preferably 20 to 200 nm.
제7구현예에 따르면,According to the seventh embodiment,
본 발명은 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체를 유효 성분으로 포함하는 의료기기를 제공한다.The present invention provides a medical device containing a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
본 발명의 일 실시예에 따르면, 상기 의료기기는 창상 피복재일 수 있다. According to one embodiment of the present invention, the medical device may be a wound covering material.
도 16 및 도 17을 참조하면, 본 발명의 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체는 저농도로 사용할 경우에도 뛰어난 조직 재생 능력을 나타냄을 확인하였다.Referring to FIGS. 16 and 17 , it was confirmed that the Prussian blue/polyvinylpyrrolidone nanoparticle complex of the present invention exhibits excellent tissue regeneration ability even when used at a low concentration.
본 발명의 일 실시예에 따르면, 상기 폴리비닐피롤리돈 나노입자의 분자량은 1 내지 50kDa일 수 있다. 바람직하게는, 상기 폴리비닐피롤리돈 나노입자의 분자량은 10kDa일 수 있다.According to one embodiment of the present invention, the molecular weight of the polyvinylpyrrolidone nanoparticles may be 1 to 50 kDa. Preferably, the molecular weight of the polyvinylpyrrolidone nanoparticles may be 10 kDa.
본 발명의 일 실시예에 따르면, 상기 본 발명의 일 실시예에 따르면, 상기 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체의 크기는 10 내지 300 nm, 바람직하게는 20 내지 200 nm일 수 있다.According to one embodiment of the present invention, the size of the Prussian blue/polyvinylpyrrolidone nanoparticle complex may be 10 to 300 nm, preferably 20 to 200 nm. .
본 발명에 따른 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체는 생체적합성이 뛰어나고 안정적인 항산화 효능을 지닌 물질로서, 활성산소제거 능력이 뛰어나 염증성 간 질환, 신경 퇴행성 질환, 암, 당뇨병, 퇴행성관절염, 류마티스관절염, 자가면역질환, 천식 등의 활성산소 유래 질병의 개선 및 치료가 가능할 수 있고, 항염증 효과가 뛰어나 염증에서 과다 발현되는 산화적 스트레스를 조절함으로써 염증 완화 및 개선 효과를 나타낼 수 있고, 조직재생 능력이 우수하여 항노화 화장품, 아토피 개선 화장품, 건강기능식품, 의약품, 의료기기(창상피복재 포함)에 널리 사용될 수 있는 플랫폼으로 사용될 수 있을 것으로 기대된다.The Prussian Blue/Polyvinylpyrrolidone nanoparticle complex according to the present invention is a substance with excellent biocompatibility and stable antioxidant effect, and has excellent active oxygen scavenging ability for inflammatory liver disease, neurodegenerative disease, cancer, diabetes, degenerative arthritis, It can improve and treat active oxygen-derived diseases such as rheumatoid arthritis, autoimmune diseases, and asthma, and has excellent anti-inflammatory effects and can exhibit inflammation relief and improvement effects by controlling oxidative stress that is overexpressed in inflammation. It is expected to be used as a platform that can be widely used in anti-aging cosmetics, atopy-improving cosmetics, health functional foods, pharmaceuticals, and medical devices (including wound dressings) due to its excellent regenerative ability.
도 1은 본 발명에 따른 분자량별 프러시안 블루/폴리비닐피롤리돈 나노입자(PB/PVP NP)의 제조방법 및 항산화 효능에 대한 모식도이다.1 is a schematic view of a method for preparing Prussian blue/polyvinylpyrrolidone nanoparticles (PB/PVP NP) by molecular weight and antioxidant efficacy according to the present invention.
도 2는 프러시안 블루 나노입자(PB NP) 및 분자량별 PB/PVP NP의 사진이다.2 is a photograph of Prussian blue nanoparticles (PB NPs) and PB/PVP NPs by molecular weight.
도 3은 프러시안 블루 나노입자(PB NP) 및 분자량별 PB/PVP NP의 크기를 나타낸 것이다.3 shows the size of Prussian blue nanoparticles (PB NPs) and PB/PVP NPs by molecular weight.
도 4는 프러시안 블루 나노입자(PB NP) 및 분자량별 PB/PVP NP의 분산값을 나타낸 것이다.4 shows Prussian blue nanoparticles (PB NPs) and dispersion values of PB/PVP NPs by molecular weight.
도 5는 프러시안 블루 나노입자(PB NP) 및 분자량별 PB/PVP NP의 표면전하를 나타낸 것이다.5 shows the surface charge of Prussian blue nanoparticles (PB NPs) and PB/PVP NPs for each molecular weight.
도 6는 프러시안 블루 나노입자(PB NP) 및 분자량별 PB/PVP NP의 UV-Vis 스펙트럼을 나타낸 것이다Figure 6 shows the UV-Vis spectrum of Prussian blue nanoparticles (PB NPs) and PB / PVP NPs by molecular weight
도 7은 (a) 프러시안 블루 나노입자(PB NP), (b) PB/PVP10k NP 및 (c) PB/PVP360k NP의 TEM 사진이다.7 is a TEM image of (a) Prussian blue nanoparticles (PB NPs), (b) PB/PVP10k NPs, and (c) PB/PVP360k NPs.
도 8은 프러시안 블루 나노입자(PB NP) 및 프러시안 블루/폴리비닐피롤리돈 나노입자(PB/PVP NP)의 3차 증류수 내 색상 변화를 나타낸 것이다.8 shows the color change of Prussian blue nanoparticles (PB NPs) and Prussian blue/polyvinylpyrrolidone nanoparticles (PB/PVP NPs) in tertiary distilled water.
도 9는 프러시안 블루 나노입자(PB NP) 및 프러시안 블루/폴리비닐피롤리돈 나노입자(PB/PVP NP)의 3차 증류수 내 크기 변화를 나타낸 것이다.9 shows the size change of Prussian blue nanoparticles (PB NPs) and Prussian blue/polyvinylpyrrolidone nanoparticles (PB/PVP NPs) in tertiary distilled water.
도 10은 프러시안 블루 나노입자(PB NP) 및 프러시안 블루/폴리비닐피롤리돈 나노입자(PB/PVP NP)의 3차 증류수 내 분산값 변화를 나타낸 것이다.FIG. 10 shows changes in dispersion values of Prussian blue nanoparticles (PB NPs) and Prussian blue/polyvinylpyrrolidone nanoparticles (PB/PVP NPs) in tertiary distilled water.
도 11은 프러시안 블루 나노입자(PB NP) 및 프러시안 블루/폴리비닐피롤리돈 나노입자(PB/PVP NP)의 3차 증류수 내 표면전하 변화를 나타낸 것이다.11 shows surface charge changes of Prussian blue nanoparticles (PB NPs) and Prussian blue/polyvinylpyrrolidone nanoparticles (PB/PVP NPs) in tertiary distilled water.
도 12는 프러시안 블루 나노입자(PB NP) 및 프러시안 블루/폴리비닐피롤리돈 나노입자(PB/PVP NP)의 OH 활성산소제거 능력을 나타낸 것이다.12 shows the OH active oxygen scavenging ability of Prussian blue nanoparticles (PB NPs) and Prussian blue/polyvinylpyrrolidone nanoparticles (PB/PVP NPs).
도 13은 PB/PVP10k NP의 농도별 세포독성 결과를 나타낸 것이다.13 shows the results of cytotoxicity of PB/PVP10k NPs by concentration.
도 14는 PB/PVP10k NP의 농도별 세포 내 활성산소라디칼 제거 능력을 나타낸 것이다.14 shows the intracellular active oxygen radical removal ability according to the concentration of PB/PVP10k NP.
도 15는 PB/PVP10k NP의 농도별 세포 내 항염증 효과 결과를 나타낸 것이다.15 shows the intracellular anti-inflammatory effect results according to the concentration of PB/PVP10k NP.
도 16은 대조군과 본 발명의 PB/PVP10k NP의 시간별 세포 상처 치유 결과를 비교한 것이다.Figure 16 compares the results of cell wound healing over time between the control group and the PB/PVP10k NPs of the present invention.
도 17는 PB/PVP10k NP의 24시간 후 농도별 세포 상처 치유 결과를 나타낸 것이다.17 shows the results of cell wound healing by concentration after 24 hours of PB/PVP10k NPs.
이하, 본 발명의 구현예를 상세히 설명하기로 한다. 다만, 이는 예시로서 제시되는 것으로, 이에 의해 본 발명이 제한되지는 않으며 본 발명은 후술할 청구항의 범주에 의해 정의될 뿐이다.Hereinafter, embodiments of the present invention will be described in detail. However, this is presented as an example, and the present invention is not limited thereby, and the present invention is only defined by the scope of the claims to be described later.
달리 정의되지 않는 한, 본원에서 사용되는 모든 기술적 및 과학적 용어는 본 발명이 속하는 당업자에 의해 통상적으로 이해되는 것과 동일한 의미를 갖는다.Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.
본 명세서에서 사용된 용어, "예방"은 본 발명에 따른 약학적 조성물의 투여에 의해 아토피성 질환을 억제시키거나 발병을 지연시키는 모든 행위를 의미한다.As used herein, the term "prevention" refers to any action that suppresses or delays the onset of atopic disease by administration of the pharmaceutical composition according to the present invention.
본 명세서에서 사용된 용어, "치료하다", "치료" 등의 용어들은 일시적 또는 영구적으로 증상을 완화하거나, 증상의 원인을 제거하거나, 또는 상기 질병이나 병태의 증상의 발현을 방지하거나 늦추는 것을 의미한다.As used herein, the terms "treat", "treatment" and the like mean temporarily or permanently alleviating symptoms, removing the cause of symptoms, or preventing or delaying the onset of symptoms of the disease or condition. do.
본 명세서에서 사용된 용어, "개선"은 비정상적인 상태와 관련된 파라미터, 예를 들면 증상의 정도를 감소시키는 모든 행위를 의미한다.As used herein, the term "improvement" refers to any action that reduces a parameter associated with an abnormal state, eg, the severity of a symptom.
본 명세서에서 사용된 용어, "약제학적으로 허용 가능한"은 과도한 독성, 자극, 알레르기 반응 또는 기타 문제점 또는 합병증 없이 이득/위험 비가 합리적이어서 대상체(예: 인간)의 조직과 접촉하여 사용하기에 적합하며 건전한 의학적 판단의 범주 이내인 것을 의미한다As used herein, the term "pharmaceutically acceptable" means that the benefit/risk ratio is reasonable without excessive toxicity, irritation, allergic reaction or other problems or complications, so that it is suitable for use in contact with the tissue of a subject (eg, human) and Means within the scope of sound medical judgment
실시예 1. 프러시안 블루/폴리비닐피롤리돈 나노 입자(PB/PVP NP) 제조Example 1. Preparation of Prussian Blue/Polyvinylpyrrolidone Nanoparticles (PB/PVP NP)
4종의 상이한 분자량의 폴리비닐피롤리돈(PVP)(10, 40, 360, 1300kDa; 75mg)을 각각 3ml의 3차 증류수에 녹여 고분자 용액을 제조한 후 포타슘 페리시아나이드(potassium ferricyanide)(5mM) 1ml을 첨가하고 상온에서 30분 동안 400rpm으로 교반하며 반응시켜 준 후, 530rpm으로 교반 속도를 높이고 반응용액에 iron chloride tetrahydrate(5mM) 1ml을 천천히 적가하면서 1시간 동안 반응시켜 프러시안 블루/폴리비닐피롤리돈 나노입자(PB/PVP NP)를 제조하였다. 제조된 PB/PVP NP를 PVP의 분자량에 따라 PB/PVP10k NP, PB/PVP40k NP, PB/PVP360k NP, PB/PVP1300k NP로 명명하기로 한다. 상기 PB/PVP10k NP는 10kDa의 PVP로 코팅한 프러시안 블루/폴리비닐피롤리돈 나노 입자를 의미한다.Polyvinylpyrrolidone (PVP) of 4 different molecular weights (10, 40, 360, 1300 kDa; 75 mg) was dissolved in 3 ml of tertiary distilled water to prepare a polymer solution, followed by potassium ferricyanide (5 mM). ) was added and reacted while stirring at 400 rpm for 30 minutes at room temperature, then increasing the stirring speed to 530 rpm and slowly adding 1 ml of iron chloride tetrahydrate (5mM) dropwise to the reaction solution while reacting for 1 hour to react for Prussian blue/polyvinyl Pyrrolidone nanoparticles (PB/PVP NP) were prepared. The prepared PB/PVP NPs are named PB/PVP10k NPs, PB/PVP40k NPs, PB/PVP360k NPs, and PB/PVP1300k NPs according to the molecular weight of PVP. The PB/PVP10k NPs refer to Prussian blue/polyvinylpyrrolidone nanoparticles coated with 10 kDa PVP.
비교예 1. 프러시안 블루 나노 입자(PB NP)의 제조Comparative Example 1. Preparation of Prussian Blue Nanoparticles (PB NP)
고분자 용액 대신 3ml의 3차 증류수만 사용한 것을 제외하고는 실시예 1과 동일한 방법으로 프러시안 블루 나노 입자(PB NP)를 제조하였다.Prussian blue nanoparticles (PB NP) were prepared in the same manner as in Example 1, except that only 3 ml of tertiary distilled water was used instead of the polymer solution.
제조한 PB NP 및 PB/PVP NP를 Amicon Ultra-15 filter를 이용하여 정제하고 3일간 동결 건조한 후 하기와 같이 특성을 평가하였다.The prepared PB NPs and PB/PVP NPs were purified using an Amicon Ultra-15 filter, freeze-dried for 3 days, and then their properties were evaluated as follows.
[특성 평가][Attribute evaluation]
실험예 1. 나노 입자의 크기, 분산도 및 표면전하Experimental Example 1. Size, dispersion and surface charge of nanoparticles
제조한 PB NP 및 PB/PVP NP의 크기, 분산도 및 표면전하를 Zetasizer (ELSZ-2000, Otsuka) 장비를 이용하여 분석하였다. The size, dispersion and surface charge of the prepared PB NPs and PB/PVP NPs were analyzed using Zetasizer (ELSZ-2000, Otsuka) equipment.
도 2에서 볼 수 있듯이 PB NP 및 PB/PVP NP 모두 프러시안 블루 고유의 파란색을 나타냈다.As can be seen in FIG. 2, both the PB NPs and the PB/PVP NPs exhibited the unique blue color of Prussian Blue.
도 3에서 볼 수 있듯이 PB NP의 크기는 약 130nm이었다. 10kDa PVP 코팅 PB/PVP NP의 크기는 약 80nm로 작게 제조되었고 360kDa 및 1300kDa PVP 코팅 PB/PVP NP의 크기는 약 135nm로 PB NP와 유사하였다.As can be seen in Figure 3, the size of the PB NPs was about 130 nm. The size of the 10 kDa PVP-coated PB/PVP NPs was about 80 nm, and the size of the 360 kDa and 1300 kDa PVP-coated PB/PVP NPs was about 135 nm, similar to the PB NPs.
도 4에서 볼 수 있듯이 분산값(PDI)의 경우 PVP 코팅 후 증가했으나 모두 0.3 이하로 균일하였다.As can be seen in FIG. 4, the dispersion value (PDI) increased after PVP coating, but all were uniform at 0.3 or less.
도 5에서 볼 수 있듯이 PB NP의 표면전하는 약 -40mV였고 PB/PVP NP의 표면전하는 고분자의 분자량이 높아질수록 중성에 가까운 표면전하를 보였다.As can be seen in FIG. 5, the surface charge of the PB NP was about -40 mV, and the surface charge of the PB/PVP NP showed a surface charge close to neutral as the molecular weight of the polymer increased.
실험예 2. 흡광도Experimental Example 2. Absorbance
PB/PVP NP의 NIR 파장대에서 흡광도를 UV-Vis spectroscopy를 이용해 분석하였다. The absorbance of PB/PVP NPs in the NIR wavelength range was analyzed using UV-Vis spectroscopy.
도 6에서 볼 수 있듯이 PVP 코팅 후 PB 나노 입자 모두 700nm의 파장대에서 강한 흡광도를 보였다. 700nm의 파장대에서 흡광도는 프러시안 블루를 코팅하는 PVP의 분자량에 따라 달랐다. 즉, 저분자량 PVP(10kDa PVP, 40kDa PVP) 코팅 PB/PVP NP의 흡광도보다 고분자량 PVP(360kDa PVP, 1300kDa PVP) 코팅 PB/PVP NP가 더 낮았다. 이는 고분자량 PVP가 결합 부위(binding site)를 더 많이 포함하기 때문이다.As can be seen in Figure 6, after the PVP coating, all of the PB nanoparticles showed strong absorbance in the wavelength range of 700 nm. The absorbance in the wavelength range of 700 nm was different depending on the molecular weight of PVP coating Prussian Blue. That is, the absorbance of high molecular weight PVP (360 kDa PVP, 1300 kDa PVP) coated PB/PVP NPs was lower than that of low molecular weight PVP (10 kDa PVP, 40 kDa PVP) coated PB/PVP NPs. This is because high molecular weight PVP contains more binding sites.
실험예 3. 나노 입자의 형태Experimental Example 3. Shape of nanoparticles
투과전자현미경(TEM)을 이용하여 PB NP, PB/PVP NP의 형태를 관찰하였다.The morphologies of PB NPs and PB/PVP NPs were observed using a transmission electron microscope (TEM).
도 7에서 볼 수 있듯이 PB NP, 저분자량의 PB/PVP10k NP, 고분자량의 PB/PVP360k NP 모두 구형 구조를 가졌다. PB NP의 경우 금속 나노 입자들이 뭉쳐서 한 덩어리로 관찰되었다. PB/PVP NP는 PVP 마이셀 안에 PB NP가 고르게 결합되어 제조되었다. Zetasizer를 통해 분석했을 때와 유사하게 저분자량 PB/PVP NP가 고분자량 PB/PVP NP보다 작은 것을 확인할 수 있다.As can be seen in FIG. 7, PB NPs, low molecular weight PB/PVP10k NPs, and high molecular weight PB/PVP360k NPs all had spherical structures. In the case of PB NP, metal nanoparticles were agglomerated and observed as a lump. PB/PVP NPs were prepared by evenly combining PB NPs into PVP micelles. Similar to the analysis through Zetasizer, it can be seen that the low molecular weight PB/PVP NPs are smaller than the high molecular weight PB/PVP NPs.
실험예 4. 안정성 평가Experimental Example 4. Stability evaluation
안정성 평가를 위해 동결 건조된 PB NP 및 PB/PVP NP를 3차 증류수에 분산시킨 후 37℃, 100rpm의 인큐베이터에서 0, 1주간 보관 후 나노 입자의 성상, 크기, 분산도 및 표면전하의 변화를 분석하여 그 결과를 도 8 내지 도 11에 나타내었다.For stability evaluation, freeze-dried PB NPs and PB/PVP NPs were dispersed in tertiary distilled water, and then stored in an incubator at 37 ° C and 100 rpm for 0 and 1 week. Changes in the properties, size, dispersion and surface charge of nanoparticles Analyzed and the results are shown in Figures 8 to 11.
도 8에서 볼 수 있듯이 인큐베이터 내 보관 후 PB NP 및 PB/PVP NP의 성상을 관찰했을 때, PB NP의 경우 프러시안 블루 고유의 푸른색을 잃었고, 고분자량의 PVP가 코팅된 PB/PVP NP도 색을 잃었다. 그러나 저분자량의 PVP가 코팅된 PB/PVP NP의 경우 프러시안 블루의 안정성이 향상되어 1주 뒤에도 색을 유지하였다. 특히 PB/PVP10k NP의 경우 눈에 띄는 색 변화가 없었다. As can be seen in FIG. 8, when the properties of PB NPs and PB/PVP NPs were observed after storage in an incubator, PB NPs lost the blue color inherent to Prussian blue, and PB/PVP NPs coated with high molecular weight PVP also lost color However, in the case of PB/PVP NPs coated with low molecular weight PVP, the stability of Prussian blue was improved and the color was maintained even after 1 week. In particular, there was no noticeable color change in the case of PB/PVP10k NP.
도 9에서 볼 수 있듯이 PB/PVP10k NP의 크기는 1주 후에도 변화가 없었으나 PB NP 및 다른 PB/PVP NP의 크기는 인큐베이터 내 보관하기 전보다 줄었다. 이는 불안정한 프러시안 블루가 용해되었기 때문이다. 따라서 프러시안 블루의 용해에 의해 색 변화가 컸던 고분자량 PB/PVP NP의 크기 변화가 눈에 띄게 컸다.As can be seen in FIG. 9, the size of PB/PVP10k NPs did not change even after 1 week, but the sizes of PB NPs and other PB/PVP NPs were reduced compared to before storage in the incubator. This is due to the dissolution of the unstable Prussian Blue. Therefore, the size change of the high molecular weight PB/PVP NPs, which had a large color change due to the dissolution of Prussian blue, was noticeably large.
도 10에서 볼 수 있듯이 PB NP의 분산값이 1주 후 크게 증가하였으나 PB/PVP NP의 분산값은 큰 변화없이 유지되었다. As can be seen in FIG. 10, the variance value of PB NPs increased significantly after 1 week, but the variance values of PB/PVP NPs were maintained without significant change.
도 11에서 볼 수 있듯이 표면전하(Zeta potential)의 경우 1주 후에도 큰 변화가 관찰되지 않았으나, 고분자량 PVP(360kDa PVP, 1300kDa PVP) 코팅된 PB/PVP NP는 1주 후에는 중성 전하를 가지는 경향을 나타냈다. 따라서 저분자량 PVP 코팅 후 프러시안 블루의 안정성이 더 향상되었고 그 중 PB/PVP10k NP가 가장 안정함을 알 수 있었다.As can be seen in FIG. 11, no significant change was observed in the surface charge (Zeta potential) even after 1 week, but high molecular weight PVP (360 kDa PVP, 1300 kDa PVP) coated PB / PVP NPs tend to have neutral charges after 1 week showed Therefore, the stability of Prussian Blue was further improved after coating with low molecular weight PVP, and among them, PB/PVP10k NP was found to be the most stable.
실험예 5. 활성산소제거능 평가Experimental Example 5. Evaluation of reactive oxygen scavenging ability
제조한 PB/PVP NP의 활성산소제거능을 평가하였다.The active oxygen scavenging ability of the prepared PB/PVP NPs was evaluated.
먼저, 0.1mM EDTA, 0.1mM FeCl3, 1mM H2O2, 3.75mM 2-deoxy-D-ribose을 3차 증류수에 준비하였다. 이를 15ml 튜브에 순서대로 100μl 씩 넣어준 후 PB NP 및 PB/PVP NP (0.1mg/ml)을 각각 1ml 넣어주었다. 이 때 네거티브 대조군(negative control)에는 H2O2와 나노 입자 용액 대신 3차 증류수를, 포지티브 대조군(positive control)에는 나노 입자 대신 3차 증류수를 넣어주었다. 그 후 0.5ml의 phosphate buffer(20Mm)와 0.1ml의 ascorbic acid(0.1mM)를 넣어주었다. 그 후 37℃, 100rpm의 인큐베이터에서 한시간 동안 반응시켰다. 그 후 1ml의 2-thiobarbituric acid(1% w/v)와 1ml의 trichloroacetic acid (2% w/v)를 넣고 85℃의 수조에서 15분간 중탕시켜 반응을 종결시켰다. 분홍색으로 변한 반응용액을 상온에서 충분히 식혀주고 96-well plate에 300μl씩 분주한 후 파장대 535nm에서 흡광도를 측정하였다. 얻은 흡광도 측정값을 하기 수학식 1에 대입해 활성산소제거능을 평가하여 그 결과를 도 12에 나타내었다.First, 0.1mM EDTA, 0.1mM FeCl 3 , 1mM H 2 O 2 , and 3.75mM 2-deoxy-D-ribose were prepared in tertiary distilled water. 100 μl of this was sequentially put into a 15 ml tube, and then 1 ml of each of PB NP and PB/PVP NP (0.1 mg/ml) was added. At this time, tertiary distilled water was added instead of the H 2 O 2 and nanoparticle solution to the negative control, and tertiary distilled water was added instead of the nanoparticles to the positive control. After that, 0.5 ml of phosphate buffer (20 Mm) and 0.1 ml of ascorbic acid (0.1 mM) were added. After that, it was reacted for one hour in an incubator at 37° C. and 100 rpm. Then, 1 ml of 2-thiobarbituric acid (1% w/v) and 1 ml of trichloroacetic acid (2% w/v) were added and the reaction was terminated by heating in a water bath at 85° C. for 15 minutes. The reaction solution that turned pink was sufficiently cooled at room temperature, and 300 μl was dispensed into a 96-well plate, and then absorbance was measured at a wavelength of 535 nm. The obtained absorbance measurement value was substituted into the following Equation 1 to evaluate the active oxygen scavenging ability, and the results are shown in FIG. 12.
[수학식 1][Equation 1]
Figure PCTKR2022007931-appb-img-000001
Figure PCTKR2022007931-appb-img-000001
도 12에서 볼 수 있듯이 PVP 코팅 후 프러시안 블루의 활성산소제거능력이 향상되었다. PB NP의 경우 0.1mg/ml의 농도에서 10%의 항산화력을 보였다. 흥미롭게도 PVP 고분자 자체에서도 활성산소제거능력을 가져 분자량이 낮을수록 높은 항산화력을 보였다. 가장 안정하고 입자의 크기가 작아 표면적이 가장 넓은 PB/PVP10k NP의 항산화력은 36%로 가장 높았다. 이러한 결과로부터 높은 항산화력을 갖는 PVP와 PB NP는 항산화력에 있어서 시너지 효과를 나타냄을 알 수 있다. 특히 PB/PVP10k NP가 항산화력이 가장 우수함을 알 수 있었다.As can be seen in FIG. 12, the active oxygen scavenging ability of Prussian Blue was improved after PVP coating. In the case of PB NP, 10% of antioxidant power was shown at a concentration of 0.1 mg/ml. Interestingly, the PVP polymer itself also has active oxygen scavenging ability, and the lower the molecular weight, the higher the antioxidant power. PB/PVP10k NP, which is the most stable and has the widest surface area due to its small particle size, had the highest antioxidant capacity at 36%. From these results, it can be seen that PVP and PB NPs, which have high antioxidant power, exhibit synergistic effects in antioxidant power. In particular, it was found that PB/PVP10k NP had the best antioxidant activity.
[세포 실험 (In vitro assay)][Cell test ( In vitro assay)]
실험예 6. 세포독성시험(Cell viability)Experimental Example 6. Cell viability
제조한 PB/PVP NP의 세포독성을 평가하였다.The cytotoxicity of the prepared PB/PVP NPs was evaluated.
먼저, 섬유아세포, NIH 3T3를 10% FBS와 1% AA가 첨가된 DMEM에서 배양하였다. NIH 3T3(1Ⅹ104cells/well)를 96-well plate에 분주하고 12시간 동안 37℃, 5% CO2 환경의 인큐베이터에서 배양하였다. 0, 0.01, 0.1, 0.5, 1.0, 5.0mg/ml의 농도로 PB/PVP10k NP을 처리하고 24시간 동안 인큐베이터에 보관하였다. 그 후 CCK-8 용액을 처리하고 인큐베이터에서 한시간 동안 반응시킨 뒤 상층액의 흡광도를 450nm에서 측정하여 그 결과를 도 13에 나타내었다.First, fibroblasts, NIH 3T3, were cultured in DMEM supplemented with 10% FBS and 1% AA. NIH 3T3 (1Ⅹ10 4 cells/well) was dispensed into a 96-well plate and cultured in an incubator at 37°C and 5% CO 2 environment for 12 hours. PB/PVP10k NPs were treated at concentrations of 0, 0.01, 0.1, 0.5, 1.0, and 5.0 mg/ml and stored in an incubator for 24 hours. Thereafter, the CCK-8 solution was treated and reacted for one hour in an incubator, and the absorbance of the supernatant was measured at 450 nm, and the results are shown in FIG. 13 .
도 13에서 볼 수 있듯이 생체적합성이 우수한 소재 기반의 PB/PVP10k NP 처리 후 NIH 3T3 내에서 세포 독성을 일으키지 않았다. 고농도인 5.0mg/ml의 나노 입자를 처리했을 때도 세포 생존율에 의미있는 변화가 일어나지 않았다. 따라서 PB/PVP10k NP는 의약품 및 화장품으로 활용함에 있어 안전성 측면에서 문제가 없음을 알 수 있었다.As shown in FIG. 13, after treatment with PB/PVP10k NP based on a material having excellent biocompatibility, cytotoxicity was not caused in NIH 3T3. Even when the nanoparticles were treated at a high concentration of 5.0 mg/ml, there was no significant change in cell viability. Therefore, it was found that there is no problem in terms of safety in using PB/PVP10k NP as pharmaceuticals and cosmetics.
실험예 7. 세포 내 활성산소라디칼 제거능 평가Experimental Example 7. Intracellular reactive oxygen radical scavenging ability evaluation
제조한 PB/PVP NP의 세포 내 활성산소라디칼 제거능을 평가하였다.The intracellular active oxygen radical scavenging ability of the prepared PB/PVP NPs was evaluated.
먼저, NIH 3T3를 10% FBS가 첨가된 DMEM에서 배양하였다. NIH 3T3(1Ⅹ104 cells/well)를 96 well-plate에 분주 후 12시간 동안 인큐베이터에서 배양하였다. 그 후 다양한 농도의 PB/PVP10k NP(0, 10, 100pg/ml)와 H2O2(25μM)를 100μl 씩 처리하고 4시간동안 인큐베이터 내에서 반응시켰다. 이 때 대조군에는 나노 입자와 H2O2 대신 세포배양액만 처리하였다. 4시간 후 처리한 시료를 석션하고 잔여하는 시료를 한 번 더 PBS로 세척하였다. 그리고 H2DCFDA(10μM)를 처리하고 90분간 인큐베이터에서 반응시켰다. 그 후 활성산소라디칼과 반응해 생성된 DCF의 형광도(ex/480nm, em/535nm)를 측정하여 나노 입자의 활성산소라디칼 제거능을 평가하여 그 결과를 도 14에 나타내었다.First, NIH 3T3 was cultured in DMEM supplemented with 10% FBS. After dispensing NIH 3T3 (1Ⅹ10 4 cells/well) into 96 well-plates, they were cultured in an incubator for 12 hours. Thereafter, 100 μl of PB/PVP10k NPs (0, 10, 100 pg/ml) and H 2 O 2 (25 μM) at various concentrations were treated and reacted in an incubator for 4 hours. At this time, only the cell culture medium was treated instead of the nanoparticles and H 2 O 2 in the control group. After 4 hours, the treated sample was suctioned and the remaining sample was washed once more with PBS. Then, H2DCFDA (10 μM) was treated and reacted in an incubator for 90 minutes. Then, by measuring the fluorescence (ex / 480nm, em / 535nm) of the DCF generated by reacting with the active oxygen radical to evaluate the active oxygen radical removal ability of the nanoparticles, the results are shown in FIG. 14 .
도 14에서 볼 수 있듯이 산화 스트레스 제재인 H2O2을 처리했을 때 생성된 ROS%를 100%라고 봤을 때, 10pg/ml와 100pg/ml의 PB/PVP10k NP를 처리한 세포 내 ROS%는 각각 45%와 15%로 나노 입자의 농도가 증가함에 따라 ROS%이 낮아짐을 알 수 있었다. 100pg/ml의 고농도로 PB/PVP10k NP를 처리한 경우에도 산화 스트레스 제재를 처리하지 않는 대조군 그룹(CTL)보다 낮아 PB/PVP10k NP가 매우 뛰어난 활성산소제거 물질임을 알 수 있었다.As can be seen in FIG. 14, when the ROS% generated when the oxidative stress agent H 2 O 2 was treated was 100%, the ROS% in cells treated with 10 pg/ml and 100 pg/ml PB/PVP10k NPs were respectively It was found that ROS% decreased as the concentration of nanoparticles increased to 45% and 15%. Even when PB/PVP10k NPs were treated at a high concentration of 100 pg/ml, it was lower than that of the control group (CTL) not treated with the oxidative stress agent, indicating that PB/PVP10k NPs were very excellent active oxygen scavengers.
실험예 8. 세포 내 항염증 효과 평가Experimental Example 8. Intracellular anti-inflammatory effect evaluation
제조한 PB/PVP NP의 세포 내 항염증 효과를 평가하였다.The intracellular anti-inflammatory effect of the prepared PB/PVP NP was evaluated.
Raw 264.7 세포를 10% FBS를 첨가한 DMEM에서 배양하였다. Raw 264.7(1Ⅹ104 cells/well)를 96 well-plate에 분주하고 12시간 동안 인큐베이터에서 배양하였다. 그 후 세포에 다양한 농도의 PB/PVP10k NP(0, 10, 100μg/ml)와 LPS(100ng/ml)를 100μl씩 처리하고 24시간동안 인큐베이터 안에서 반응시켰다. 이 때 대조군에는 나노 입자와 LPS 대신 세포배양액만 처리하였다. 24시간 후 상층액만 다른 96 well-plate에 옮겨 100μl씩 분주하였다. 그리고 Griess reagent 100μl를 처리한 후 상온에서 10분간 반응시켰다. Nitric oxide(NO)와 반응하면서 분홍색으로 변한 용액의 흡광도를 560nm의 파장대에서 plate reader를 이용해 분석해 PB/PVP10k NP의 항염증 효과를 평가하여 그 결과를 도 15에 나타내었다.Raw 264.7 cells were cultured in DMEM supplemented with 10% FBS. Raw 264.7 (1Ⅹ10 4 cells/well) was dispensed into 96 well-plates and cultured in an incubator for 12 hours. Thereafter, the cells were treated with 100 μl of PB/PVP10k NPs (0, 10, 100 μg/ml) and LPS (100 ng/ml) at various concentrations and allowed to react in an incubator for 24 hours. At this time, only the cell culture medium was treated instead of nanoparticles and LPS in the control group. After 24 hours, only the supernatant was transferred to another 96 well-plate and dispensed by 100 μl. And after processing 100 μl of Griess reagent, it was reacted at room temperature for 10 minutes. The absorbance of the solution, which turned pink while reacting with nitric oxide (NO), was analyzed using a plate reader in a wavelength range of 560 nm to evaluate the anti-inflammatory effect of PB/PVP10k NP, and the results are shown in FIG. 15 .
도 15에서 볼 수 있듯이 PB/PVP10k NP를 처리했을 때 lipopolysaccharide(LPS) 자극에 의해 대식세포 내에서 과량으로 생성된 NO가 줄어드는 것을 확인하였다. Raw 264.7세포에 LPS를 처리하였을 때 생성된 NO의 농도를 100%라고 했을 때, 처리한 PB/PVP10k NP의 농도가 10 ㎍/㎖에서 100 ㎍/㎖로 증가함에 따라 NO의 농도가 73%에서 19%로 확연히 감소함을 확인하였다. 또한 LPS를 처리하지 않는 대조군(CTL)의 NO 농도가 6%인 것을 고려했을 때 PB/PVP10k NP가 10 ㎍/㎖의 낮은 농도에서도 뛰어난 NO 생성억제 효과를 가진다는 것을 확인하였다. 산화 스트레스로 과량으로 생성된 NO는 염증반응을 촉진해서 조직 손상을 유발할 수 있다. 따라서 본 발명의 효과적인 NO 생성억제능을 가진 PB/PVP10k NP는 뛰어난 항염증성 물질임을 알 수 있었다.As can be seen in FIG. 15, it was confirmed that NO produced in excess in macrophages by lipopolysaccharide (LPS) stimulation was reduced when PB/PVP10k NPs were treated. When the concentration of NO produced when Raw 264.7 cells were treated with LPS was 100%, as the concentration of PB/PVP10k NP treated increased from 10 μg/ml to 100 μg/ml, the concentration of NO increased from 73% to 100 μg/ml. It was confirmed that it significantly decreased to 19%. In addition, considering that the NO concentration of the control group (CTL) not treated with LPS was 6%, it was confirmed that the PB/PVP10k NP had an excellent NO production inhibitory effect even at a low concentration of 10 μg/ml. Excessive production of NO due to oxidative stress promotes an inflammatory response and can lead to tissue damage. Therefore, it was found that the PB/PVP10k NP having an effective NO production inhibitory activity of the present invention is an excellent anti-inflammatory substance.
실험예 9. In vitro 상처 치유 실험Experimental Example 9. In vitro wound healing experiment
제조한 PB/PVP NP에 대해 in vitro 상처 치유 실험을 수행하였다.In vitro wound healing experiments were performed on the prepared PB/PVP NPs.
NIH 3T3를 10% FBS와 1% AA가 첨가된 DMEM에서 배양하였다. NIH 3T3(1Ⅹ105 cells/well)를 24 well-plate에 분주해 12시간 인큐베이터에서 배양하였다. 세포단층이 형성되면 멸균된 p1000 pipet tip을 이용해 scratch wound를 제조하였다. 떨어진 세포들을 석션으로 제거해준 뒤 FBS를 포함하지 않는 배양액에 PB/PVP10k NP (0, 0.01, 0.1, 0.5, 1mg/ml)를 각각 분산시켜 세포에 처리해 starvation 환경을 만든 후, 정해진 시간동안 인큐베이터에서 배양하며 세포이동 정도를 현미경을 통해 관찰한 후 그 결과를 도 16에 나타내었다.NIH 3T3 was cultured in DMEM supplemented with 10% FBS and 1% AA. NIH 3T3 (1Ⅹ10 5 cells/well) was dispensed into 24 well-plates and cultured in an incubator for 12 hours. When the cell monolayer was formed, a scratch wound was prepared using a sterile p1000 pipet tip. After removing the fallen cells by suction, PB/PVP10k NPs (0, 0.01, 0.1, 0.5, 1mg/ml) are dispersed in the culture medium without FBS, respectively, treated with cells to create a starvation environment, and then placed in an incubator for a set period of time. After culturing and observing the degree of cell migration through a microscope, the results are shown in FIG. 16.
도 16에서 볼 수 있듯이 세포배양액만 처리한 대조군의 scratch wound가 회복되지 않는 조건에서 PB/PVP10k NP를 처리했을 때 세포 간의 거리가 시간 별로 줄어듦을 확인했다. 즉, 나노 입자 처리 후 4시간 후 세포 간의 거리가 127μm에서 95μm로 좁아져 유의미한 세포이동을 보였고, 24시간 후에는 wound gap이 69μm로 PB/PVP10k NP가 뛰어난 조직 재생 능력을 보유하고 있음을 알 수 있었다. As can be seen in FIG. 16, it was confirmed that the distance between cells decreased over time when PB/PVP10k NP was treated under conditions where the scratch wound of the control group treated only with cell culture medium was not recovered. In other words, after 4 hours of nanoparticle treatment, the distance between cells narrowed from 127μm to 95μm, showing significant cell migration, and after 24 hours, the wound gap was 69μm, indicating that PB/PVP10k NPs have excellent tissue regeneration ability. there was.
또한 도 17에서 볼 수 있듯이 처리되는 PB/PVP10k의 농도가 증가할수록 세포 간의 거리가 줄어듦을 확인하였다. 그러나, 낮은 농도인 0.01mg/ml의 PB/PVP10k NP 처리 후에도 wound gap이 109μm로 줄어드는 뛰어난 상처 치유 결과를 보임을 알 수 있다.In addition, as shown in FIG. 17, it was confirmed that the distance between cells decreased as the concentration of PB/PVP10k to be treated increased. However, even after treatment with PB/PVP10k NP at a low concentration of 0.01 mg/ml, it can be seen that excellent wound healing results were shown, with the wound gap reduced to 109 μm.

Claims (16)

  1. 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체를 유효 성분으로 포함하는 항산화제.An antioxidant comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
  2. 제1항에 있어서,According to claim 1,
    상기 폴리비닐피롤리돈 나노입자의 분자량은 1 내지 50kDa인 것인, 항산화제.The polyvinylpyrrolidone nanoparticles have a molecular weight of 1 to 50 kDa, an antioxidant.
  3. 제1항에 있어서,According to claim 1,
    상기 폴리비닐피롤리돈 나노입자의 분자량은 10kDa인 것인, 항산화제.The polyvinylpyrrolidone nanoparticles have a molecular weight of 10 kDa, an antioxidant.
  4. 제1항에 있어서,According to claim 1,
    상기 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체의 크기는 10 내지 300 nm인 것인, 항산화제.The size of the Prussian blue / polyvinylpyrrolidone nanoparticle complex is 10 to 300 nm, the antioxidant.
  5. 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체를 유효 성분으로 포함하는 항염증제.An anti-inflammatory agent comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
  6. 제5항에 있어서,According to claim 5,
    상기 폴리비닐피롤리돈 나노입자의 분자량은 1 내지 50kDa인 것인, 항염증제.The polyvinylpyrrolidone nanoparticles have a molecular weight of 1 to 50 kDa, anti-inflammatory agent.
  7. 제5항에 있어서,According to claim 5,
    상기 폴리비닐피롤리돈 나노입자의 분자량은 10kDa인 것인, 항염증제.The polyvinylpyrrolidone nanoparticles have a molecular weight of 10 kDa, anti-inflammatory agent.
  8. 제5항에 있어서,According to claim 5,
    상기 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체의 크기는 10 내지 300 nm인 것인, 항염증제.The size of the Prussian blue / polyvinylpyrrolidone nanoparticle complex is 10 to 300 nm, the anti-inflammatory agent.
  9. 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체를 유효 성분으로 포함하는 활성산소 과잉생성으로 인해 유발되는 질환의 예방 또는 치료용 약제학적 조성물로서,A pharmaceutical composition for preventing or treating diseases caused by excessive production of active oxygen, comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient,
    상기 활성산소 과잉생성으로 인해 유발되는 질환은 뇌졸중, 파킨슨병, 알츠하이머병, 노화, 심장질환, 허혈, 동맥경화, 피부질환, 염증, 관절염, 류마티스, 자가면역질환, 천식, 고지혈증, 간질환, 당뇨병, 암, 만성궤양, 화상 또는 창상인 것인, 약제학적 조성물.Diseases caused by the excessive production of reactive oxygen species include stroke, Parkinson's disease, Alzheimer's disease, aging, heart disease, ischemia, arteriosclerosis, skin disease, inflammation, arthritis, rheumatism, autoimmune disease, asthma, hyperlipidemia, liver disease, diabetes , Cancer, chronic ulcers, burns or wounds, the pharmaceutical composition.
  10. 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체를 유효 성분으로 포함하는 활성산소 과잉생성으로 인해 유발되는 질환의 예방 또는 치료용 의약외품 조성물로서,A quasi-drug composition for preventing or treating diseases caused by excessive production of active oxygen, comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient,
    상기 활성산소 과잉생성으로 인해 유발되는 질환은 뇌졸중, 파킨슨병, 알츠하이머병, 노화, 심장질환, 허혈, 동맥경화, 피부질환, 염증, 류마티스, 자가면역질환, 천식, 고지혈증, 간질환, 당뇨병, 암, 만성궤양, 화상 또는 창상인 것인, 의약외품 조성물.Diseases caused by the excessive production of reactive oxygen species include stroke, Parkinson's disease, Alzheimer's disease, aging, heart disease, ischemia, arteriosclerosis, skin disease, inflammation, rheumatism, autoimmune disease, asthma, hyperlipidemia, liver disease, diabetes, and cancer , Chronic ulcers, burns or wounds, quasi-drug composition.
  11. 제10항에 있어서,According to claim 10,
    상기 의약외품 조성물은 소독 청결제, 샤워폼, 가그린, 물티슈, 세제 비누, 핸드 워시, 가습기 충진제, 마스크, 연고제 또는 필터 충진제인 것인, 의약외품 조성물.The quasi-drug composition is a disinfectant cleanser, shower foam, gargreen, wet tissue, detergent soap, hand wash, humidifier filler, mask, ointment or filter filler, quasi-drug composition.
  12. 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체를 유효 성분으로 포함하는 활성산소로 유발되는 증상 또는 질환의 예방 또는 개선용 화장료 조성물로서,A cosmetic composition for preventing or improving symptoms or diseases caused by active oxygen containing a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient,
    상기 활성산소로 유발되는 증상 또는 질환은 피부노화, 주름생성, 피부색소침착, 아토피, 여드름, 건선 또는 습진인 것을 특징으로 하는 것인, 화장료 조성물.Symptoms or diseases caused by the active oxygen are skin aging, wrinkle formation, skin pigmentation, atopy, acne, psoriasis or eczema, characterized in that the cosmetic composition.
  13. 제12항에 있어서,According to claim 12,
    상기 화장료는 앰플, 크림, 로션, 화장수, 에센스 또는 팩인 것을 특징으로 하는 것인, 화장료 조성물.The cosmetic composition is characterized in that the cosmetic is an ampoule, cream, lotion, lotion, essence or pack.
  14. 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체를 유효 성분으로 포함하는 건강기능식품 조성물.A health functional food composition comprising a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
  15. 프러시안 블루/폴리비닐피롤리돈 나노입자 복합체를 유효 성분으로 포함하는 의료기기.A medical device containing a Prussian blue/polyvinylpyrrolidone nanoparticle complex as an active ingredient.
  16. 제15항에 있어서, 상기 의료기기는 창상 피복재인 것인 의료기기.The medical device according to claim 15, wherein the medical device is a wound covering material.
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