WO2021070778A1 - Nanodisque - Google Patents

Nanodisque Download PDF

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WO2021070778A1
WO2021070778A1 PCT/JP2020/037729 JP2020037729W WO2021070778A1 WO 2021070778 A1 WO2021070778 A1 WO 2021070778A1 JP 2020037729 W JP2020037729 W JP 2020037729W WO 2021070778 A1 WO2021070778 A1 WO 2021070778A1
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nanodisc
lysolecithin
mass
lecithin
less
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PCT/JP2020/037729
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English (en)
Japanese (ja)
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恵広 柳澤
忠夫 辻
井村　知弘
敏彰 平
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株式会社カネカ
国立研究開発法人産業技術総合研究所
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Publication of WO2021070778A1 publication Critical patent/WO2021070778A1/fr

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    • 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
    • A61K31/12—Ketones
    • A61K31/122—Ketones having the oxygen directly attached to a ring, e.g. quinones, vitamin K1, anthralin
    • 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
    • A61K47/42—Proteins; Polypeptides; Degradation products thereof; Derivatives thereof, e.g. albumin, gelatin or zein
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K8/00—Cosmetics or similar toiletry preparations
    • A61K8/02—Cosmetics or similar toiletry preparations characterised by special physical form
    • A61K8/04—Dispersions; Emulsions
    • 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
    • A61K8/55—Phosphorus compounds
    • 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
    • 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

Definitions

  • the present invention relates to nanodiscs which can be easily produced, which can disperse hydrophobic compounds and the like well in water and have excellent transparency and stability, and cosmetics containing the nanodiscs. ..
  • compositions such as cosmetics and pharmaceutical compositions are hydrophobic and difficult to disperse in aqueous solvents.
  • Organic solvents may be used to disperse hydrophobic compounds, but even ethanol and isopropanol may be reduced or avoided in personal care and pharmaceutical compositions applied to the human body. Tend to do. Therefore, especially in personal care compositions, surfactants are often used to disperse hydrophobic compounds in aqueous solvents.
  • the size of the liposome is relatively large, and the dispersion liquid of the liposome is usually cloudy. Therefore, in recent years, very fine particles called nanodisks having a lipid bilayer structure similar to that of liposomes have been developed.
  • the nanodisc has a structure in which the hydrophobic side of the disc composed of a lipid bilayer is coated with an amphipathic membrane scaffold protein (Non-Patent Documents 1, Patent Documents 4 and 5), and is a layer of nanodiscs.
  • the thickness is several nm and the diameter is less than 10 nm, which is very fine.
  • Patent Document 6 also discloses a composition containing a lipid and a surfactant, which is a polymer aggregate having a diameter of less than 100 nm. Therefore, the dispersion liquid of nanodisc is very transparent.
  • lipid bilayer membrane of nanodiscs is similar to the lipid bilayer membrane of cells, it is considered that the membrane proteins contained in nanodiscs retain the structure and activity of the cell membranes as they are. It is expected that research on membrane proteins will progress dramatically.
  • the present inventors have also developed a nanodisk characterized by containing a lipid bilayer and lipopeptide biosurfactant (Patent Document 7).
  • nanodiscs containing a lipid bilayer have been developed, and it is considered that hydrophobic compounds and the like can be satisfactorily dispersed in an aqueous solvent by using nanodiscs.
  • nanodiscs are expected to be applied to cosmetics having excellent transparency, the transparency may decrease over time and precipitation may occur in some cases. Therefore, the present invention provides nanodiscs which can be easily produced, can disperse hydrophobic compounds and the like well in water, and have excellent transparency and stability, and cosmetics containing the nanodiscs. The purpose is.
  • the present inventors have conducted intensive studies to solve the above problems. As a result, they have found that by using lipopeptide biosurfactant as a surfactant and using a specific ratio of lecithin and lysolecithin as lipids, nanodiscs having excellent transparency and stability can be easily produced, and the present invention has been completed. did. Hereinafter, the present invention will be shown.
  • [1] Contains lipid bilayer and lipopeptide biosurfactant
  • the lipid bilayer contains lecithin and lysolecithin, A nanodisk characterized in that the ratio of the lysolecithin to the total of the lecithin and the lysolecithin is 1% by mass or more and 20% by mass or less.
  • the nanodisc according to the above [1] or [2] which has a thickness of 2 nm or more and 10 nm or less.
  • the lipopeptide biosurfactant is a surfactin represented by the following formula (I) or a salt thereof.
  • X indicates an amino acid residue selected from leucine, isoleucine and valine; R 1 represents a C 9-18 alkyl group]
  • a cosmetic comprising the nanodisc according to any one of the above [1] to [8].
  • membrane scaffold protein has been used for manufacturing nanodiscs, but it took a lot of time and effort to design and manufacture membrane scaffold protein.
  • the lipopeptide biosurfactant which is the surfactant used in the present invention, is produced by a microorganism and can be produced relatively easily.
  • the nanodisc according to the present invention can be manufactured by a very simple method as compared with the conventional nanodisk which could not be manufactured without going through a complicated process. Further, by using the nanodisc according to the present invention, it is possible to satisfactorily disperse the hydrophobic compound in an aqueous solvent to obtain a visually transparent composition, and the transdermal properties of the hydrophobic compound are remarkably improved. There is a possibility of doing so.
  • the nanodisk of the present invention which has a lipid bilayer structure similar to that of cells, may significantly improve the transdermal properties of hydrophilic compounds existing in the vicinity thereof.
  • the nanodisc of the present invention has excellent stability and can maintain the transparency of the preparation for a long period of time. Therefore, the present invention is industrially excellent as a technique that contributes to the development of personal care compositions and pharmaceutical compositions having excellent properties.
  • FIG. 1 is an external photograph of the lipid preparation according to the present invention after a stability test.
  • FIG. 2 is an external photograph of a lipid preparation outside the scope of the present invention after a stability test.
  • the nanodisc according to the present invention contains a lipid bilayer and lipopeptide biosurfactant.
  • a general nanodisc is stabilized by a structure in which the hydrophobic part of the amphipathic membrane scaffold protein is bound to the side hydrophobic part of the lipid bilayer to cover it, and the hydrophilic part faces outward.
  • the nanodisc according to the present invention is stabilized by a structure in which the hydrophobic portion of the lipopeptide biosurfactant is bound to the side hydrophobic portion of the lipid bilayer to cover it and the hydrophilic portion faces outward. Conceivable.
  • a cyclic peptide is pierced into a lipid bilayer as an ion channel, or a lipid having a bulky hydrophobic portion and a surface active compound having a bulky hydrophilic portion form a bilayer. Therefore, even in the nanodisc according to the present invention, it is possible that some lipopeptide biohydrophobics form a bilayer together with lipids.
  • the size of the nanodisk according to the present invention may be 2 nm or more and 10 nm or less, and the diameter may be 5 nm or more and 40 nm or less.
  • the thickness is preferably 3 nm or more, more preferably 4 nm or more, preferably 8 nm or less, and more preferably 6 nm or less.
  • the diameter is preferably 6 nm or more, more preferably 7 nm or more, preferably 20 nm or less, and more preferably 15 nm or less.
  • the phospholipid constituting the lipid bilayer is a compound having a hydrophilic phosphate base and a hydrophobic tail, and in the present invention, lecithin and lysolecithin are used in combination.
  • Lecithin is a general term for lipid products containing phospholipids, and is produced by separating and drying components that precipitate by adding water to raw materials containing phospholipids, such as soybean oil and egg yolk, or by extracting with ethanol or the like.
  • Lecithin made from soybeans is called soybean lecithin
  • lecithin made from egg yolk is called egg yolk lecithin.
  • the components constituting lecithin are not particularly limited, and examples thereof include glycerophospholipids, sphingolipids, glycolipids, synthetic lipids, and sterols.
  • Glycerophospholipids are double-stranded phospholipids having a complex structure of two fatty acids, glycerin, phosphate and choline, such as dipalmitoylphosphatidylcholine (DOPC), dipalmitoylphosphatidylcholine (DLPC), and dipalmitoylphosphatidylcholine (DMPC). ), Dipalmitoylphosphatidylcholine (DPPC), distearoylphosphatidylcholine (DSPC), 1-palmitoyl-2-oleoylphosphatidylcholine (POPC), 1-stearoyl-2-myristoylphosphatidylcholine, dilinoleylphosphatidylcholine and the like.
  • DOPC dipalmitoylphosphatidylcholine
  • DLPC dipalmitoylphosphatidylcholine
  • DMPC dipalmitoylphosphatidylcholine
  • DPPC dipalmito
  • glycerophospholipid examples include a glycerophospholipid represented by the following formula (II). [In the formula, R 2 and R 3 independently represent a C 10-24 alkyl group or a C 10-24 alkenyl group]
  • examples of the C 10-24 alkyl group include n-decyl, 8-methylnonade, n-undecyl, 9-methyldecyl, n-dodecyl, 10-methylundecyl, n-tridecyl, 11-.
  • examples thereof include methyldodecyl, n-tetradecyl, n-pentadecyl, n-hexadecyl, n-heptadecyl, n-octadecyl, n-nonadecylic, n-icosyl, n-docosyl, n-tetracosyl and the like.
  • Examples of the C 10-24 alkenyl group include decenyl, dodecenyl, tetradecenyl, hexadecenyl, octadecenyl, icosenyl, docosenyl, tetracosenyl and the like.
  • Sphingolipid has a structure in which phosphoric acid and a base are bound to ceramide in which a fatty acid is amide-bonded to sphingoid. That is, the sphingolipid is a double-chain phospholipid having a long-chain hydrocarbon group derived from sphingosine and a long-chain hydrocarbon group derived from a long-chain fatty acid. Examples of sphingolipids include sphingomyelin.
  • glycolipids examples include mannosyl erythritol lipid, sophorolipid, sulfoxyribosyl glyceride, diglycosyl diglyceride, digalactosyl diglyceride, galactosyl diglyceride, glycosyl diglyceride, galactosyl cerebroside, lactosyl cerebroside, and ganglioside.
  • sterols examples include animal-derived sterols such as cholesterol succinic acid, lanosterol, dihydrolanosterol, desmosterol, and dihydrocholesterol; plant-derived sterols such as stigmasterol, cytosterol, campesterol, and brushcasterol (phytosterols); thymosterol. , Sterols derived from microorganisms such as ergosterol and the like.
  • Lysolecithin is a single-stranded phospholipid that has only one hydrophobic tail, whereas major lecithin such as glycerophospholipids and sphingolipids are double-stranded phospholipids that have two hydrophobic tails. is there. Lysolecithin is produced, for example, by treating glycerophospholipids and sphingolipids with phospholipase A2 to remove one of the hydrophobic tails.
  • lysolecithin examples include lysophosphatidylcholine having a structure in which the fatty acid at the 2-position of phosphatidylcholine is removed by phospholipase A2, lysophosphatidic acid having a structure in which choline is removed from lysophosphatidylcholine, sphingosine monophosphate, sphingosine phosphorylcholine and the like. Can be mentioned.
  • Lecithin and lysolecithin generally contain a carbon-carbon double bond in the long-chain fatty acid moiety, but lecithin and lysolecithin used in the present invention may be saturated hydrogenated additives to which hydrogen is added or unsaturated. A mixture of mold and saturated mold may be used.
  • the lipid bilayer of the nanodisc according to the present invention is mainly composed of lecithin and lysolecithin.
  • a general lipid bilayer is mainly composed of double-stranded phospholipids, which are planar because they have a well-balanced hydrophilic and hydrophobic parts and occupy an elongated rectangular parallelepiped space. It has the property of easily forming an infinite aggregate.
  • a membrane scaffold protein which is a polymer, forms a disc state by surrounding the lipid bilayer.
  • the shape is maintained by the molecular aggregate instead of the polymer, stabilization is necessary for practical use, and it was considered that the addition of the third component is effective for that purpose.
  • lysolecithin is a single chain, the hydrophobic part is small with respect to the hydrophilic part, the shape is different from lecithin, and it fills the intermolecular gap that causes the instability of nanodiscs. It was thought that nanodiscs could be stabilized by using an appropriate amount of lysolecithin together. Therefore, in the present invention, the ratio of lecithin to lysolecithin is adjusted to improve the stability of nanodiscs.
  • the ratio of lysolecithin to the total of lecithin and lysolecithin is 1% by mass or more and 20% by mass or less.
  • the ratio within this range, as shown in the results of the examples described later, the nanodisc stabilizing effect can be obtained by using lysolecithin in combination with lecithin.
  • 1.5% by mass or more is preferable, 2% by mass or more is more preferable, 3% by mass or more is further preferable, 15% by mass or less is preferable, and 10% by mass or less is more preferable.
  • the total amount of lecithin and lysolecithin can be appropriately adjusted within the range in which nanodiscs can be satisfactorily produced.
  • the total amount of lecithin and lysolecithin with respect to the solvent containing the nanodisk of the present invention can be 0.01% by mass or more and 10% by mass or less.
  • Lipopeptide biosurfactant has a hydrophobic group and a peptide containing a hydrophilic moiety, exhibits a surface-active effect, and is produced by a microorganism.
  • examples of the lipopeptide biosurfactant include surfactin, arslovactin, itulin, fendisin, serawettin, leikecin, and viscocin.
  • surfactin represented by the above formula (I) or a salt thereof (hereinafter referred to as “surfactin (I)" is preferable.
  • the roots of the carboxymethyl group and the carboxyethyl group represent the optically active sites.
  • X represents any one amino acid residue selected from leucine, isoleucine and valine.
  • the amino acid residue as X may be L-form or D-form, but L-form is preferable.
  • R 1 represents a C 9-18 alkyl group.
  • the "C 9-18 alkyl group” refers to a linear or branched monovalent saturated hydrocarbon group having 9 or more and 18 or less carbon atoms.
  • N-tetradecyl n-pentadecylic, n-hexadecyl, n-heptadecyl, n-octadecyl and the like.
  • the above-mentioned surfactin (I) may be used alone or in combination of two or more.
  • the C 9-18 alkyl group of R 1 may contain a plurality of different surfactins (I).
  • Surfactin (I) can be used by culturing a microorganism, for example, a strain belonging to Bacillus subtilis, and separating it from the culture solution according to a known method, and it can be used as a refined product or an unrefined product.
  • the culture solution can be used as it is.
  • those obtained by a chemical synthesis method can also be used in the same manner.
  • a salt of surfactin (I) can also be used.
  • the counter cation constituting the salt is not particularly limited, and examples thereof include alkali metal ions and ammonium ions.
  • the alkali metal ion that can be used for the salt of surfactin (I) is not particularly limited, but represents lithium ion, sodium ion, potassium ion, or the like. Further, the two alkali metal ions may be the same as each other or may be different from each other.
  • Examples of the substituent of the ammonium ion include alkyl groups such as methyl, ethyl, n-propyl, isopropyl, n-butyl and t-butyl; aralkyl groups such as benzyl, methylbenzyl and phenylethyl; phenyl, toluyl, xsilyl and the like. Examples include organic groups such as the aryl group of. Examples of the ammonium ion include tetramethylammonium ion, tetraethylammonium ion, pyridinium ion and the like.
  • the two counter cations may be the same or different from each other. Also, one of the carboxy group -COOH or -COO - shall may be in the state of.
  • the amount of lipopeptide biosurfactant used may be appropriately adjusted within the range in which nanodisks are formed.
  • the amount of lipopeptide biosurfactant with respect to the total of lecithin and lysolecithin can be 0.5 mass times or more and 5 mass times or less. If the amount is 0.5 mass times or more, nanodisks are formed more reliably, and if the amount is 5 mass times or less, the amount of lipopeptide biosurfactant that is not involved in the formation of nanodisks is so excessive. Not economical.
  • the amount is preferably 1 mass times or more, preferably 4 mass times or less, and more preferably 3 mass times or less.
  • the nanodisc according to the present invention may contain components other than lecithin, lysolecithin and lipopeptide biosurfactant.
  • an aqueous solvent is mainly used as a solvent for cosmetics and pharmaceutical compositions, but there is a problem that it is difficult to disperse a hydrophobic compound in the aqueous solvent.
  • a hydrophobic compound can probably be taken in and dispersed in an aqueous solvent, so that a highly transparent composition can be obtained.
  • the "aqueous solvent” refers to water and a mixed solvent of a water-miscible organic solvent and water.
  • the water-miscible organic solvent refers to an organic solvent that can be miscible with water without limitation, and examples thereof include C 1-3 alcohol, preferably ethanol or isopropanol.
  • the proportion of the water-miscible organic solvent in the mixed solvent is preferably 10% by mass or less, more preferably 5% by mass or less, still more preferably 2% by mass or less.
  • the aqueous solvent may be a buffer solution, and its pH is not particularly limited, but is preferably 5.0 or more and 13.0 or less.
  • Hydrophobic compound refers to a compound that has a hydrophobic group, is sparingly soluble in water, and is soluble in oil.
  • poorly soluble means that 100 mL or more of water is required to dissolve 1 g of the hydrophobic compound at 20 ⁇ 5 ° C.
  • the hydrophobic compound used in the present invention is not particularly limited as long as it is an active ingredient to be blended in cosmetics, pharmaceutical compositions, etc., but for example, a whitening agent, an anti-aging agent, an antioxidant, a moisturizer, a hair restorer, and cell activation. Examples include agents, vitamins and amino acids.
  • fat-soluble vitamins such as tocopherol, coenzyme Q10, reduced coenzyme Q10, and derivatives thereof; oils such as squalane; steroidal anti-inflammatory agents such as hydrocortisone, prednisolone, dexamethasone, betamethasone; acetyl.
  • Non-steroidal anti-inflammatory agents such as salicylic acid, ibuprofen, indomethacin, loxoprofen; antibiotics; antifungal agents; preservatives such as methylparaben; fragrances such as menthol; pigments such as alizanin.
  • albutin and its derivatives L-ascorbic acid and its derivatives; hydroquinone and its derivatives; glutathione and its derivatives; placenta extract; pantothenic acid and its derivatives; tranexamic acid and its derivatives; Derivatives; Plant extracts such as chamomile extract and citrus extract; Carotenoids such as beta-carotene; Astaxanthin and its derivatives; Flavonoids; Catechin and its derivatives; Vitamin A and its derivatives; ⁇ -lipoic acid and its derivatives; Glycyrrhizic acid And its derivatives; thiotaurine and its derivatives; urea and its derivatives; glycerin derivatives; salicylic acid and its derivatives; nicotinic acid and its derivatives; L-amino acids and its derivatives; adenosine and its derivatives; plant female hormone-like components such as isoflavone; Grabridine and its derivatives; magnolignan and its derivative
  • the nanodisc according to the present invention may contain a membrane protein.
  • Membrane proteins are proteins that are bound to biological membranes, and are mainly biological membranes other than covalent bonds such as hydrophobic interactions and electrostatic interactions with endogenous membrane proteins, of which at least a part of them is present in the biological membrane. It is classified as a superficial membrane protein that is bound to. Conventionally, it has been considered extremely difficult to study the function of a membrane protein because the higher-order structure changes and the original function is lost when a membrane protein existing in a biological membrane such as a cell membrane is isolated.
  • the nanodisc according to the present invention has a lipid bilayer similar to that of a biological membrane, and it is considered that the membrane protein maintains the structure in the biological membrane in the lipid bilayer. Research on functions will progress dramatically, and it is possible that the difficult industrial use of membrane proteins will be possible.
  • the nanodisk according to the present invention may contain a hydrophilic compound.
  • the hydrophilic compound is not particularly limited as long as it is a compound having an affinity for water molecules.
  • Many hydrophilic compounds are water-soluble, and water-soluble hydrophilic compounds can be dissolved in aqueous solvents, but they may be adsorbed on the hydrophilic part of nanodiscs to further stabilize them. is there.
  • an active ingredient such as a metal oxide, which is hydrophilic but insoluble in water, on the hydrophilic portion of the nanodisk, it may be possible to disperse the active ingredient in an aqueous solvent.
  • a plurality of nanodisks are laminated with each other to form a multilayer structure.
  • the hydrophilic compound may be present between the lipid bilayer layers.
  • the hydrophilic compound refers to a compound in which the amount of water for dissolving 1 g at 20 ⁇ 5 ° C. is less than 30 mL.
  • hydrophilic compounds include ultraviolet scattering agents such as titanium oxide particles and zinc oxide particles; inorganic pigments such as red iron oxide, titanium oxide and iron oxide; water-soluble vitamins such as ascorbic acid and its derivatives; locust bean gum and guar gum.
  • ultraviolet scattering agents such as titanium oxide particles and zinc oxide particles
  • inorganic pigments such as red iron oxide, titanium oxide and iron oxide
  • water-soluble vitamins such as ascorbic acid and its derivatives
  • locust bean gum and guar gum examples include thickeners such as derivatives, carrageenan, pectin, xanthan gum, gellan gum, and alginic acid.
  • the nanodisc according to the present invention may contain a surfactant other than lipopeptide biosurfactant.
  • a surfactant is not particularly limited, such as a nonionic surfactant, an anionic surfactant, a cationic surfactant, and an amphoteric surfactant.
  • the nonionic surfactant is not particularly limited, but for example, polyoxyethylene (hereinafter, also referred to as POE) -polyoxypropylene (hereinafter, also referred to as POP) block copolymer; POE-POP block copolymer addition of ethylene diamine such as poloxamine.
  • POE polyoxyethylene
  • POP polyoxypropylene
  • POE sorbitan fatty acid esters such as monolauric acid POE (20) sorbitan, monooleic acid POE (20) sorbitan, POE sorbitan monostearate, POE sorbitan tristearate; POE (5) hardened castor oil, POE (10) ) Hardened castor oil, POE (20) Hardened castor oil, POE (40) Hardened castor oil, POE (50) Hardened castor oil, POE (60) Hardened castor oil, POE (100) Hardened castor oil, etc.
  • the numbers in parentheses indicate the average number of moles of POP or POE added.
  • the anionic surfactant is not particularly limited, and examples thereof include alkylbenzene sulfonate, alkyl sulfate, polyoxyethylene alkyl sulfate, aliphatic ⁇ -sulfomethyl ester, and ⁇ -olefin sulfonic acid.
  • the cationic surfactant is not particularly limited, and examples thereof include benzalkonium chloride and benzethonium chloride.
  • amphoteric tenside is not particularly limited, and examples thereof include glycine type such as alkyldiaminoethylglycine, betaine type acetate such as lauryldimethylaminoacetic acid betaine, and imidazoline type amphoteric tenside.
  • glycine type such as alkyldiaminoethylglycine
  • betaine type acetate such as lauryldimethylaminoacetic acid betaine
  • imidazoline type amphoteric tenside imidazoline type amphoteric tenside.
  • nanodiscs are manufactured by a complicated method in which a membrane scaffold protein is added to a phospholipid solution containing sodium cholic acid as a surfactant, and then the surfactant is removed by dialysis or the like.
  • the nanodisc according to the present invention can be produced by an extremely simple method of mixing at least lecithin, lysolecithin and lipopeptide biosurfactant in an aqueous solvent.
  • the total concentration of lecithin and lysolecithin in the reaction solution may be appropriately adjusted, but can be, for example, 0.001% by mass or more and 40% by mass or less, preferably 0.1% by mass or more and 30% by mass or less. ..
  • the concentration of lipopeptide biosurfactant in the reaction solution can also be 0.001 part by mass or more and 50% by mass or less, preferably 0.1% by mass or more and 45% by mass or less.
  • the amounts of the hydrophobic compound and the hydrophilic compound are preferably 0.1% by mass or more and 50% by mass or less, respectively, based on the total amount of lecithin and lysolecithin, respectively. It is more preferably 5.5% by mass or more and 20% by mass or less, and even more preferably 1% by mass or more and 10% by mass or less.
  • the reaction conditions of lecithin, lysolecithin and lipopeptide biosurfactant may be appropriately adjusted within a range in which the formation of nanodisks progresses satisfactorily.
  • the reaction temperature may be room temperature.
  • the temperature can be set to 20 ° C. or higher and 30 ° C. or lower, which is near the general phase transition temperature.
  • the reaction time can be 2 minutes or more and 50 hours or less.
  • nanodiscs may not be formed simply by mixing each component, but even in such a case, the reaction temperature is relatively high, for example, 40 ° C. or higher.
  • Nanodiscs may be formed by lowering the temperature to 90 ° C. or lower.
  • the temperature is preferably 85 ° C. or lower, more preferably 80 ° C. or lower.
  • the nanodisk according to the present invention can also be produced by first preparing liposomes from lecithin and lysolecithin, and then adding lipopeptide biosurfactant. That is, after dissolving lecithin and lysolecithin in a lower alcohol solvent such as methanol, ethanol and isopropanol or an organic solvent such as chloroform, the organic solvent is distilled off to form a lipid film, and an aqueous solvent is added and stirred to form liposomes. ..
  • a lower alcohol solvent such as methanol, ethanol and isopropanol or an organic solvent such as chloroform
  • the concentration of the liposome dispersion liquid at this time may be appropriately adjusted, and for example, it can be 0.01% by mass or more and 40% by mass or less with respect to the organic solvent, and 0.05% by mass or more and 35% by mass or less. The following is preferable.
  • nanodiscs can be obtained.
  • the concentration of the lipopeptide biosurfactant in the reaction solution may be appropriately adjusted, and may be, for example, 0.01% by mass or more and 40% by mass or less, 0.05% by mass or more, based on the amount of the liposome solution. It is preferably 35% by mass or less.
  • ingredients other than lecithin, lysolecithin and lipopeptide biosurfactant may be added in a timely manner.
  • the hydrophilic compound may be added to the aqueous solvent in a timely manner.
  • Hydrophobic compounds, membrane proteins and water-insoluble hydrophilic compounds are preferably allowed to coexist with at least lecithin and lysolecithin in aqueous solvents.
  • the amounts of the hydrophobic compound and the hydrophilic compound are preferably 0.1% by mass or more and 100% by mass or less, more preferably 0.5% by mass or more and 50% by mass or less, based on the total of lecithin and lysolecithin, respectively. More preferably, it is by mass% or more and 20% by mass or less.
  • biomolecules lecithin and lysolecithin exhibit a surface-active action, they assist the dispersion of these compounds in an aqueous solvent. Further, in the case of a membrane protein, it is also possible to bind the membrane protein to the nanodisk by cell-free synthesis of the membrane protein in the presence of the nanodisk according to the present invention.
  • the nanodisks may be freeze-dried and then the freeze-dried nanodisks may be dissolved again in an aqueous solvent.
  • the nanodisc concentration of the redissolved solution can be adjusted arbitrarily.
  • the nanodisc according to the present invention can take in a hydrophobic compound therein and disperse it well in an aqueous solvent and is extremely fine, the dispersion in the aqueous solvent is visually sufficient. It looks transparent and is considered to have high permeability to skin tissues.
  • a state in which the dispersoid cannot be visually confirmed and the dispersion liquid becomes visually transparent may be referred to as "solubilization". Therefore, it can be said that the nanodisc according to the present invention is particularly effective as a component of a personal care composition such as a cosmetic or a pharmaceutical composition containing a hydrophobic compound as an active ingredient.
  • Example 1 Production of Nanodiscs Containing Coenzyme Q10 According to the composition shown in Table 1, only soy lecithin (“Basis LP-20” manufactured by Nisshin Oillio Group) or soy lecithin and hydrogenated lysolecithin (“SLP-LPC70H”” A solution was prepared by stirring and mixing a combination with Tsuji Oil Co., Ltd.), coenzyme Q10 (manufactured by Kaneka Co., Ltd.) and an appropriate amount of chloroform with a vortex mixer. During the draft, chloroform was distilled off by spraying nitrogen gas on the obtained solution, and the solution was dried.
  • Base LP-20 manufactured by Nisshin Oillio Group
  • SLP-LPC70H soy lecithin and hydrogenated lysolecithin
  • the phosphate buffer solution and paraben as a preservative were added to the obtained dried product, the mixture was heated to 60 to 70 ° C., and the mixture was stirred and mixed with a vortex mixer to obtain a solution.
  • Sodium surfactin and the remaining amount of phosphate buffer were added to the obtained solution, the mixture was heated to 60 to 70 ° C., and stirred and mixed with a vortex mixer to obtain a solution.
  • the hydrogenated lysolecithin product used contains about 70% by mass of lysophosphatidylcholine as lysolecithin.
  • Test Example 1 Measurement of Nanodisc Particle Size According to the composition shown in Table 2, soy lecithin (“Basis LP-20” manufactured by Nisshin Oillio Group) alone, or soy lecithin and hydrogenated chloroform (“SLP-LPC70H” Tsuji) A solution was prepared by stirring and mixing the combination with (manufactured by Oil & Refinery Co., Ltd.) and an appropriate amount of chloroform with a vortex mixer. During the draft, chloroform was distilled off by spraying nitrogen gas on the obtained solution, and the solution was dried.
  • the particle size increases as the proportion of lysolecithin increases, but if the proportion of lysolecithin / (lysolecithin + lecithin) is 25% by mass or less, the appearance of the mixed solution is transparent and the average particle size.
  • the particle size of general liposomes is said to be 0.1 to 0.2 ⁇ m, and the mixed solution was somewhat opaque, so the ratio of lysolecithin / (lysolecithin + lecithin).
  • the main component was considered to be liposomes.
  • Test Example 2 Measurement of particle size of nanodisc According to the composition shown in Table 3, hydrogenated lecithin (“SLP-PC70HS” manufactured by Tsuji Oil Co., Ltd.) only, or hydrogenated lecithin and hydrogenated chloroform (“SLP-LPC70H” Tsuji Oil Co., Ltd.) A solution was prepared by stirring and mixing the combination with (manufactured by the company) and an appropriate amount of chloroform with a vortex mixer. During the draft, chloroform was distilled off by spraying nitrogen gas on the obtained solution, and the solution was dried.
  • SLP-PC70HS hydrogenated lecithin
  • SLP-LPC70H hydrogenated lecithin and hydrogenated chloroform
  • Test Example 3 Method of obtaining nanodiscs directly from powder According to the composition shown in Table 3, hydrogenated lecithin (“SLP-PC70HS” manufactured by Tsuji Oil Co., Ltd.) and hydrogenated lecithin (“SLP-LPC70H” manufactured by Tsuji Oil Co., Ltd.)
  • SLP-PC70HS hydrogenated lecithin
  • SLP-LPC70H hydrogenated lecithin
  • the phosphate buffer solution was directly added to the powder of sodium surfactin, and the mixture was stirred with a vortex for 5 minutes, and then stirred at 180 rpm and 25 ° C. for 20 hours using a shaker (“TAITEC BioShaker BR-23FP”). did.
  • the mixture was stirred and mixed with a vortex mixer for 3 minutes to obtain an opaque liquid.
  • Test Example 4 Formation of Nanodiscs Using Different Hydrogenated Lecithin
  • SLP-PC92H manufactured by Tsuji Oil Co., Ltd.
  • SLP-LPC70H manufactured by Tsuji Oil Co., Ltd.
  • Test Example 5 Formation of Nanodiscs Using Different Soy Lecithin According to the composition shown in Table 5, soy lecithin (“SLP-White” manufactured by Tsuji Oil Co., Ltd.) and hydrogenated lecithin (“SLP-LPC70H” manufactured by Tsuji Oil Co., Ltd.) Nanodiscs were prepared with the combination of. Specifically, soybean lecithin, hydrogenated lysolecithin, and an appropriate amount of chloroform were stirred and mixed with a vortex mixer to prepare a solution. During the draft, chloroform was distilled off by spraying nitrogen gas on the obtained solution, and the solution was dried.
  • Test Example 6 Stability test The mixture (10 mL) prepared in Example 1 was placed in a 50 mL falcon tube and placed in a constant temperature and humidity chamber (manufactured by Yamato) at 50 ° C, 25 ° C or 5 ° C for 30 days (1M). Alternatively, it was stored for 60 days (2M), or it was stored for 30 days or 60 days even under the condition that the cycle of raising and lowering the temperature in the range of ⁇ 5 ° C. to 45 ° C. was repeated 3 times in 2 days. The appearance was observed after 30 days or 60 days, and the stability was evaluated according to the following criteria. The results are shown in Table 6.
  • ⁇ Transparent
  • ⁇ Slightly reduced transparency is observed, but overall transparent
  • ⁇ Clear turbidity and precipitation are observed.
  • the transparency of the nanodisc solution is maintained for 30 days even at a relatively high temperature of 50 ° C., and the ratio is 20% by mass. If it is% or less, the transparency is maintained even after 60 days except when it is held at 50 ° C. or 25 ° C., and if it is 10% by mass or less, the transparency is maintained after 60 days even if it is held at 50 ° C. Therefore, the stability of the nanodisk according to the present invention was demonstrated.

Abstract

L'objectif de la présente invention est de fournir : un nanodisque qui peut être produit facilement et permet de disperser de manière appropriée un composé hydrophobe ou similaire dans de l'eau, et qui a une excellente transparence et stabilité ; et un cosmétique contenant le nanodisque. Selon la présente invention, un nanodisque comprend une bicouche lipidique et un biotensioactif lipopeptidique et est caractérisé en ce que la bicouche lipidique comprend une lécithine et une lysolécithine. Le rapport de la lysolécithine au total de la lécithine et de la lysolécithine est de 1 à 20 % en masse.
PCT/JP2020/037729 2019-10-10 2020-10-05 Nanodisque WO2021070778A1 (fr)

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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2023161179A1 (fr) * 2022-02-24 2023-08-31 Evonik Operations Gmbh Nouvelle composition contenant des liposomes et des biosurfactants

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JP2010511032A (ja) * 2006-11-29 2010-04-08 マルベルン コスメセウチクス リミテッド 脂質及び界面活性剤の高分子集合体を含む組成物
JP2013034829A (ja) * 2011-07-08 2013-02-21 Dunlop Sports Co Ltd ゴルフボール用樹脂組成物およびゴルフボール
WO2017090740A1 (fr) * 2015-11-26 2017-06-01 株式会社コーセー Composition contenant une structure bicellulaire
WO2018169954A1 (fr) * 2017-03-13 2018-09-20 Sdg, Inc. Nanoparticules à base de lipides à stabilité améliorée
WO2018181538A1 (fr) * 2017-03-31 2018-10-04 株式会社カネカ Nanodisque et son procédé de production

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Publication number Priority date Publication date Assignee Title
JP2010511032A (ja) * 2006-11-29 2010-04-08 マルベルン コスメセウチクス リミテッド 脂質及び界面活性剤の高分子集合体を含む組成物
JP2013034829A (ja) * 2011-07-08 2013-02-21 Dunlop Sports Co Ltd ゴルフボール用樹脂組成物およびゴルフボール
WO2017090740A1 (fr) * 2015-11-26 2017-06-01 株式会社コーセー Composition contenant une structure bicellulaire
WO2018169954A1 (fr) * 2017-03-13 2018-09-20 Sdg, Inc. Nanoparticules à base de lipides à stabilité améliorée
WO2018181538A1 (fr) * 2017-03-31 2018-10-04 株式会社カネカ Nanodisque et son procédé de production

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* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2023161179A1 (fr) * 2022-02-24 2023-08-31 Evonik Operations Gmbh Nouvelle composition contenant des liposomes et des biosurfactants

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