JP2012062258A - Neovascularization inhibitor and eye disease preventing-treating agent using the same - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a neovascularization inhibitor which uses a new ingredient as an active ingredient.SOLUTION: The characteristics of this invention are as follows: a neovascularization inhibitor (1) using the extract of black rice as an active ingredient; a neovascularization inhibitor (2) using cyanidin and/or a cyanidin glycoside as an active ingredient; a neovascularization inhibitor (3) using peonidin and/or a peonidin glycoside as an active ingredient; a retinal degenerative disease preventing agent (4) using the agent mentioned in any one paragraph from the inhibitor (1) to (3) as an active ingredient; the retinal degenerative disease preventing agent (5) mentioned in the retinal degenerative disease preventing agent (4) in which the retinal degenerative disease is retinitis pigmentosa, cone dystrophy, diabetic retinopathy, age-related macular degeneration, age-related maculopathy, macula edema, retinal detachment, cancer-related retinopathy, retinal vein occlusion or detachment of retinal pigment epithelium; and a cancer preventing-treating agent (6) using the agent mentioned in any one paragraph from the inhibitor (1) to (3) as an active ingredient.

Description

The present invention relates to an angiogenesis inhibitor and an ophthalmic disease preventive / therapeutic agent using the same. The present invention is widely used for foods, pharmaceuticals, cosmetics and the like.

Angiogenesis consists of four processes: vascular basement membrane destruction and thawing, endothelial cell migration, endothelial cell proliferation, endothelial cell differentiation and lumen formation. Here, vascular endothelial growth factor (VEGF) is considered to be involved in the processes after endothelial cell migration. Angiogenesis in cellular tissues plays an important role in development, wound healing, inflammation and the like, and is also known to be related to pathological conditions such as diabetic retinopathy and tumor formation. A typical disease with pathological angiogenesis is solid cancer. In order for a cancer tissue to grow beyond a diameter of 1 to 2 mm in a solid cancer, it is necessary that tumor blood vessels for supplying oxygen and nutrients extend from the existing blood vessels to reach the cancer tissue (J. Folkman et al. al., J. Natl. Cancer Inst., 82: 4 (1990)), and it is known that the proliferation of cancer tissue is explosively accelerated when the blood vessel reaches the cancer tissue. Furthermore, it is known that cancer cell metastasis takes place through this blood vessel.

Vascular endothelial growth factor (VEGF) is thought to play a central role in the neovascularization of this tumor, and therefore, it is expected that a substance that suppresses the expression of VEGF can suppress the growth and metastasis of cancer. Has been. On the other hand, diabetic retinopathy, an eye disease, is recognized as a complication in nearly half of diabetic patients. In diabetic retinopathy, the formation of microcapillaries is promoted due to lack of oxygen, and it is thought that this eventually ruptures and bleeds to form scar tissue and eventually leads to retinal detachment. Therefore, it is expected that the seriousness of retinopathy can be suppressed by suppressing angiogenesis. Miller et al. Reported that VEGF is closely related to the development of proliferative retinopathy based on the results of experiments using monkeys (Miller et al .: Am. J. Pathol. 145: 574 ( 1994)). Therefore, it is considered that a substance that suppresses the expression of VEGF is useful for the prevention or treatment of diabetic retinopathy.

Other diseases such as rheumatoid arthritis, psoriasis, hemangioma, scleroderma, atherosclerosis, and neovascular glaucoma are accompanied by pathological neovascularization, which is one of the main symptoms (J. Folkman et al., N. Engle. J. Med., 320: 1211 (1989)). That is, substances that inhibit angiogenesis (angiogenesis inhibitors), particularly vascular endothelial growth factor (VEGF) expression inhibitors, can be used for the treatment of various diseases in which angiogenesis is involved as described above. Conceivable. From such a background, as angiogenesis inhibitors, for example, compounds described in JP-A-10-259176, JP-A-10-81631 and JP-A-2000-26393 have been proposed.

  Under such a background, the black rice extract, cyanidin and peonidin, and glycosides thereof found an action to suppress angiogenesis, and the present invention was completed. That is, an object of the present invention is to provide an angiogenesis inhibitor containing a novel component as an active ingredient.

The features of the present invention for solving the above-described problems are as follows.
1. An angiogenesis inhibitor comprising black rice extract as an active ingredient.
2. An angiogenesis inhibitor comprising cyanidin and / or cyanidin glycoside as an active ingredient.
3. An angiogenesis inhibitor comprising peonidin and / or peonidin glycoside as an active ingredient.
4). Above 1. Thru 3. A preventive agent for retinal degenerative diseases comprising the agent according to any one of the above as an active ingredient.
5. The retinal degenerative disease is retinitis pigmentosa, cone dystrophy, diabetic retinopathy, age-related macular degeneration, age-related macular degeneration, macular edema, retinal detachment, cancer-related retinopathy, retinal vein occlusion or retinal pigment epithelial detachment 4. above. The preventive agent for retinal degenerative diseases according to 1.
6). Above 1. Thru 3. A cancer preventive / therapeutic agent comprising the agent according to any one of the above as an active ingredient.

1 is a graph showing the effect of black rice extract (PRE) on VEGF-induced human umbilical vein endothelial cell (HUVEC) tube formation. 2 is a graph showing the effect of peonidin on VEGF-induced HUVEC lumen formation. It is a graph which shows the effect | action with respect to VEGF-induced HUVEC proliferation of black rice extract (PRE), cyanidin, and peonidin. 1 is a graph showing the effects of black rice extract (PRE), cyanidin and peonidin on VEGF-induced proliferation of human retinal capillary endothelial cells (HRMEC). It is a graph which shows the effect | action with respect to VEGF-induced HRMEC migration of black rice extract (PRE), cyanidin, and peonidin.

Hereinafter, the present invention will be described in detail.
The present invention is characterized by using black rice extract as an active ingredient.
Black rice is a kind of ancient rice native to Hanzhong Province, Shaanxi Province, China, and is also called purple black rice or purple rice. The surface of the brown rice is black and contains purple black pigment (anthocyanic pigment) in the pericarp, seed coat and germ. After 5 minutes (a little polished rice), the rice turns purple, and when all the rice cake is removed, it becomes a color close to white rice. There are two types of rice grains: japonica (short) and indica (long), and many of the latter are rice cakes. According to Kampo medicine, black rice has the effects of nourishing and enhancing energy, making the gastrointestines strong, supplementing and activating blood, strengthening lung function, restoring vision, and blackening hair It is considered. Black rice pigment has been reported to have an antioxidant effect that protects blood vessels and prevents arteriosclerosis. Of the black rice pigments, anthocyanidins are preferred. This is because by containing this black rice-derived anthocyanidin, it has a further excellent anti-angiogenic action.
As black rice varieties, Shikokuen, Kurodaen, Konan Kurome: Konan Kurome, Kuromasaki, Unnan Kuromome, Kohoku Murasaki Examples include black rice.

  In the present invention, the raw material for obtaining the pigment extract is not limited to black rice varieties, and only one of these varieties (including those listed above) may be used, or two or more. May be used in combination. The shape of the rice grain may be either Japonica (short grain) or Indica (long grain), or a mixture of these. As for the type of rice, either sticky rice or sticky rice may be used, or a mixture thereof may be used.

  Moreover, when extracting a pigment | dye component from black rice, it is desirable to use defatted rice bran as a raw material. This is because the active ingredient is efficiently extracted from this defatted product by removing the oil in the rice bran.

  Preferred degreasing solvents include N-hexane, acetone and the like. In particular, when N-hexane is used as a degreasing solvent, the extracted oil can be used as edible oil, and the pigment extract can be easily used as a food material.

As an extraction solvent for the pigment component, polar solvents such as water, methanol, ethanol, isopropyl alcohol, 1,3-butylene glycol, ethylene glycol, propylene glycol, glycerin, and ethyl acetate can be used. Two or more of these solvents may be mixed.
Desirably, when water or ethanol is used as the extraction solvent, the active ingredient is efficiently extracted. In particular, hydrous ethanol is an extraction solvent that is less likely to reduce the activity of the active ingredient during extraction and is preferable from the standpoint of safety in the use of the extract in food. The kind of water for extraction is not particularly limited, and tap water, distilled water, mineral water, alkaline ionized water, deep water and the like can be used.

As the extraction temperature, for example, when water-containing ethanol is used, the extraction temperature is 20 to 80 ° C., preferably about 40 to 50 ° C. This is because if the extraction temperature is too low, it becomes difficult to extract the active ingredient, and if the extraction temperature is too high, the activity of the active ingredient tends to decrease.

  The water-containing ethanol as the extraction solvent has an ethanol concentration of 40 to 90% (wt / wt), desirably an ethanol concentration of 60 to 80% (wt / wt). The reason why the ethanol concentration is 40% (wt / wt) or more is that if the ethanol content is too small, the extraction amount of the active ingredient tends to be insufficient. The reason why the ethanol concentration is 90% (wt / wt) or less is that if the ethanol concentration is too high, the oil in the black rice bran is easily dissolved in the water-containing ethanol. In addition, in order to improve the content rate of the active ingredient, the water-containing ethanol extraction may be repeatedly performed while changing the ethanol concentration stepwise.

  As a method for extracting the pigment extract, any method such as continuous extraction, immersion extraction, countercurrent extraction, and supercritical extraction can be adopted, and any apparatus can be used at room temperature or under reflux heating.

  Specifically, the raw material (defatted black rice bran) is put into a treatment tank filled with the extraction solvent, and the active ingredient is eluted while stirring. For example, when water-containing ethanol is used as the extraction solvent, the extraction solvent is used in an amount of about 5 to 100 times (weight ratio) of the extraction raw material, and extraction is performed for about 30 minutes to 2 hours. The active ingredient is eluted in the solvent, and then filtered to remove the extraction residue to obtain an extract.

Thereafter, the extract is subjected to treatments such as dilution, concentration, drying, purification and the like according to a conventional method to obtain an angiogenesis inhibitor according to the present invention.
As a method for purifying the pigment component, the pigment component is adsorbed through a synthetic adsorption resin, a gel filtration resin or the like through an extract, and this is eluted with methanol, ethanol or the like, and concentrated under reduced pressure.

In addition, the present invention is characterized in that cyanidin and / or a glycoside thereof (hereinafter, these are simply referred to as “cyanidine etc.”) as an active ingredient.
Cyanidin is a compound represented by the following chemical formula (1).

As for cyanidin, a commercially available product can be used. In other words, those manufactured by Extrasynthese can be used.
The glycoside of cyanidin is a compound in which a sugar is bonded to the compound of the above chemical formula (1).
Although the cyanidin glycoside used in the present invention is not particularly limited, for example, cyanidin-3-glucoside, cyanidin-3-rutinoid, cyanidin-3-sambubioside, cyanidin-3-sambubioside-5-glucoside or cyanidin-3,5- And diglucoside. In addition, these may use only 1 type and may use 2 or more types together.

In addition, the present invention is characterized in that peonidin and / or peonidin glycoside (hereinafter, these are collectively referred to as “peonidin etc.”) as an active ingredient. Peonidine is a compound represented by the following chemical formula (2).

Although the method of obtaining peonidin is not specifically limited, A commercially available thing can be used. Specifically, those manufactured by Extrasynthese can be used.
The glycoside of peonidin is obtained by binding a saccharide to the compound of the above chemical formula (2).
The peonidin glycoside used in the present invention is not particularly limited. For example, peonidin-3-glucoside, peonidin-3-rutinoid, peonidin-3-sambubioside, peonidin-3-sambubioside-5-glucoside peonidin-3,5- And diglucoside. In addition, these may use only 1 type and may use 2 or more types together.

The angiogenesis inhibitor of the present invention can be used as a material for various foods and drinks. Examples of food and drink include confectionery (gum, candy, caramel, chocolate, cookies, snacks, jelly, gummi, tablet confectionery, etc.), noodles (soba, udon, ramen, etc.), and dairy products (milk, ice cream, yogurt). ), Seasonings (miso, soy sauce, etc.), soups, beverages (juice, coffee, tea, tea, carbonated drinks, sports drinks, etc.), health foods (tablets, capsules, etc.), nutrition Supplementary foods (such as energy drinks) are listed. The angiogenesis inhibitor of the present invention may be appropriately blended with these foods and drinks.

These foods and drinks can be blended with various ingredients depending on the type, for example, glucose, fructose, sucrose, maltose, sorbitol, stevioside, corn syrup, lactose, citric acid, tartaric acid, malic acid, succinic acid. Acid, lactic acid, L-ascorbic acid, dl-α-tocopherol, sodium erythorbate, glycerin, propylene glycol, glycerin fatty acid ester, polyglycerin fatty acid ester, sucrose fatty acid ester, sorbitan fatty acid ester, propylene glycol fatty acid ester, gum arabic, Food materials such as carrageenan, casein, gelatin, pectin, agar, vitamin Bs, nicotinic acid amide, calcium pantothenate, amino acids, calcium salts, pigments, fragrances and preservatives can be used.

As a specific production method, black rice defatted rice bran pigment extract is spray-dried or freeze-dried together with powdered cellulose, and this is made into powders, granules, tablets, or solutions to easily produce foods and drinks (instant foods, etc.) It can be included. Moreover, the said pigment | dye extract is melt | dissolved, for example in fats and oils, ethanol, glycerin, or these mixtures, it can be made into a liquid state, and it can add to a drink or it can add to a solid food. If necessary, it can be mixed with a binder such as gum arabic or dextrin to form a powder or granules and added to a beverage or a solid food.

When the angiogenesis inhibitor of the present invention is applied to foods and drinks, since the main purpose is disease prevention and health maintenance, the content of active ingredients in the foods and drinks is 1 to 20 wt% or less in total. Preferably there is.

The angiogenesis inhibitor of the present invention may be used as a raw material for drugs (including drugs and quasi drugs). It can be produced by appropriately blending the pigment extract of the present invention with a raw material for a pharmaceutical preparation. Examples of the pharmaceutical raw material that can be blended in the pigment extract of the present invention include excipients (glucose, lactose, sucrose, sodium chloride, starch, calcium carbonate, kaolin, crystalline cellulose, cocoa butter, hydrogenated vegetable oil, kaolin, talc, etc. ), Binder (distilled water, physiological saline, ethanol water, simple syrup, glucose solution, starch solution, gelatin solution, carboxymethylcellulose, potassium phosphate, polyvinylpyrrolidone, etc.), disintegrant (sodium alginate, agar, sodium bicarbonate) , Calcium carbonate, sodium lauryl sulfate, stearic acid monoglyceride, starch, lactose, gum arabic powder, gelatin, ethanol, etc.), disintegration inhibitors (sucrose, stearin, cocoa butter, hydrogenated oil, etc.), absorption enhancers (quaternary) Ammonium base, sodium lauryl sulfate, etc.), Chakuzai (glycerin, starch, lactose, kaolin, bentonite, silicic acid, etc.), lubricants (purified talc, stearates, polyethylene glycol, and the like) and the like.

The method of administering an angiogenesis inhibitor according to the present invention can be generally administered orally in the form of tablets, pills, soft / hard capsules, fine granules, powders, granules, liquids, etc. It may be administered parenterally. When administered as a parenteral agent, it can be administered as a solution or with the addition of a dispersing agent, suspension, stabilizer, etc., by local tissue administration, intradermal, subcutaneous, intramuscular and intravenous injection, etc. it can. Moreover, it is good also as forms, such as a suppository.

The dosage may vary depending on the administration method, medical condition, age of the patient, etc., but for adults, it can be generally administered as 0.5 to 5000 mg as an active ingredient per day, and usually about 0.5 to 3000 mg for children.
The mixing ratio of the angiogenesis inhibitor can be appropriately changed depending on the dosage form, but is usually about 0.3 to 15.0 wt% when administered orally or by mucosal absorption, and when administered parenterally. About 0.01 to 10 wt% is preferable. In addition, since the dose varies depending on various conditions, a dose smaller than the above dose may be sufficient, or it may be necessary to administer beyond the range.

Examples of cosmetics that can contain the angiogenesis inhibitor of the present invention include emulsions, soaps, facial cleansers, bathing agents, creams, emulsions, lotions, eau de cologne, shaving creams, shaving lotions, cosmetic oils, Sunscreen / sunscreen lotion, funny powder, foundation, perfume, pack, nail cream, enamel, enamel remover, eyebrow, blusher, eye cream, eye shadow, mascara, eyeliner, lipstick, lip balm, shampoo, rinse, hair dye , Dispersions, cleaning materials, and the like.
Examples of the form of a pharmaceutical or quasi-drug that can be blended with the pigment extract of the present invention include ointments, creams, and liquids for external use.

In addition to the pigment extract according to the present invention, the external preparation for skin of the above form includes ingredients, oils, and higher alcohols that are blended in external preparations for skin such as cosmetics and quasi-drugs as long as the angiogenesis inhibitory effect is not impaired. Fatty acids, ultraviolet absorbers, powders, pigments, surfactants, polyhydric alcohols / sugars, polymers, physiologically active ingredients, solvents, antioxidants, fragrances, preservatives, and the like can be blended.
Examples are listed below, but the present invention is not limited to these examples.
(1) Examples of oil [ester-based oil phase components]: glyceryl tri-2-ethylhexanoate, cetyl 2-ethylhexanoate, isopropyl myristate, butyl myristate, isopropyl palmitate, ethyl stearate, octyl palmitate, Isocetyl isostearate, butyl stearate, butyl myristate, ethyl linoleate, isopropyl linoleate, ethyl oleate, isocetyl myristate, isostearyl myristate, isostearyl palmitate, octyldodecyl myristate, isocetyl isostearate, diethyl sebacate , Diisopropyl adipate, isoaralkyl neopentanoate, glyceryl tri (capryl / caprate), trimethylolpropane tri-2-ethylhexanoate, trimethylotriisostearate Propane, tetra-2-ethylhexanoate pentaerythritol, cetyl caprylate, decyl laurate, hexyl laurate, decyl myristate, myristyl myristate, cetyl myristate, stearyl stearate, decyl oleate, cetyl ricinoleate, isolaurate Stearyl, isotridecyl myristate, isocetyl myristate, isostearyl myristate, isocetyl palmitate, isostearyl palmitate, octyl stearate, isocetyl stearate, isodecyl oleate, octyldodecyl linoleate, octyldodecyl isoleate, isopropyl isostearate Cetostearyl 2-ethylhexanoate, stearyl 2-ethylhexanoate, hexyl isostearate, ethylene glycol dioctanoate Cole, ethylene glycol dioleate, propylene glycol dicaprate, propylene glycol di (capryl / capric acid), propylene glycol dicaprylate, neopentyl glycol dicaprate, neopentyl glycol dioctanoate, glyceryl tricaprylate, glyceryl triundecylate, triisopalmitine Glyceryl acid, glyceryl triisostearate, octyldodecyl neopentanoate, isostearyl octanoate, octyl isononanoate, hexyl decyl neodecanoate, octyldodecyl neodecanoate, isocetyl isostearate, isostearyl isostearate, octyldecyl isostearate, polyglycerin oleic acid Ester, polyglycerol isostearate, dipropyl carbonate Dialkyl carbonate (C12-18), triisocetyl citrate, triisoaralkyl citrate, triisooctyl citrate, lauryl lactate, myristyl lactate, cetyl lactate, octyl decyl lactate, triethyl citrate, acetyl triethyl citrate, acetyl tributyl citrate , Trioctyl citrate, diisostearyl malate, 2-ethylhexyl hydroxystearate, di-2-ethylhexyl succinate, diisobutyl adipate, diisopropyl sebacate, dioctyl sebacate, cholesteryl stearate, cholesteryl isostearate, cholesteryl hydroxystearate, Cholesteryl oleate, dihydrocholesteryl oleate, phytosteryl isostearate, phytosteryl oleate, 12-stearoylhydroxystearin Isocetyl, 12-stearoyl-hydroxystearic acid stearyl 12-stearoyl-hydroxystearic acid isostearate, and the like.
[Hydrocarbon-based oil phase component]: Squalane, liquid paraffin, α-olefin oligomer, isoparaffin, ceresin, paraffin, liquid isoparaffin, polybutene, microcrystalline wax, petrolatum and the like.
[Animal and vegetable oil and its hardened oil, and naturally derived wax]: beef tallow, hardened beef tallow, pork tallow, hardened tallow, horse oil, hardened horse oil, mink oil, orange luffy oil, fish oil, hardened fish oil, egg yolk oil, etc. Animal oil and its hardened oil, avocado oil, almond oil, olive oil, cacao butter, apricot oil, kukui nut oil, sesame oil, wheat germ oil, rice germ oil, rice bran oil, safflower oil, shea butter, soybean oil, evening primrose oil, Perilla oil, tea seed oil, camellia oil, corn oil, rapeseed oil, hardened rapeseed oil, palm kernel oil, hardened palm kernel oil, palm oil, hardened palm oil, peanut oil, hardened peanut oil, castor oil, hardened castor oil, Vegetable oil such as sunflower oil, grape seed oil, jojoba oil, hardened jojoba oil, macadamia nut oil, medhome oil, cottonseed oil, hardened cottonseed oil, palm oil, hardened palm oil and the hardened oil thereof, beeswax High acid value beeswax, lanolin, reduced lanolin, hydrogenated lanolin, liquid lanolin, carnauba wax and montan wax.
[Silicone-based oil phase component]: dimethylpolysiloxane, methylphenylpolysiloxane, methylcyclopolysiloxane, octamethylpolysiloxane, decamethylpolysiloxane, dodecamethylcyclosiloxane, methylhydrogenpolysiloxane, polyether-modified organopolysiloxane , Dimethylsiloxane / methylcetyloxysiloxane copolymer, dimethylsiloxane / methylstearoxysiloxane copolymer, alkyl-modified organopolysiloxane, terminal-modified organopolysiloxane, amino-modified silicone oil, amino-modified organopolysiloxane, dimethiconol, silicone gel , Acrylic silicone, trimethylsiloxysilicic acid, silicone RTV rubber and the like.
[Fluorine oil phase component]: Perfluoropolyether, fluorine-modified organopolysiloxane, fluorinated pitch, fluorocarbon, fluoroalcohol, fluoroalkyl / polyoxyalkylene co-modified organopolysiloxane, and the like.
(2) Examples of higher alcohols include lauryl alcohol, myristyl alcohol, cetyl alcohol, stearyl alcohol, isostearyl alcohol, oleyl alcohol, behenyl alcohol, 2-ethylhexanol, hexadecyl alcohol, octyldodecanol and the like.
(3) Examples of fatty acids Caprylic acid, capric acid, undecylenic acid, lauric acid, myristic acid, palmitic acid, palmitoleic acid, stearic acid, isostearic acid, oleic acid, linoleic acid, linolenic acid, arachidic acid, arachidonic acid, behenic acid , Erucic acid, 2-ethylhexanoic acid and the like.
(4) Examples of UV absorbers Paraaminobenzoic acid, amyl paraaminobenzoate, ethyldihydroxypropyl paraaminobenzoate, glyceryl paraaminobenzoate, ethyl paraaminobenzoate, octyl paraaminobenzoate, octyldimethyl paraaminobenzoate, ethylene glycol salicylate, salicylic acid Octyl, triethanolamine salicylate, phenyl salicylate, butylphenyl salicylate, benzyl salicylate, homomenthyl salicylate, benzyl cinnamate, octyl paramethoxycinnamate, 2-ethylhexyl paramethoxycinnamate, mono-2-diparamethoxycinnamate Glyceryl ethylhexanoate, isopropyl paramethoxycinnamate, diethanolamine salt of paramethoxyhydrocinnamic acid, diisopropyl diisopropylcinnamic acid Stealth mixture, urocanic acid, ethyl urocanate, hydroxymethoxybenzophenone, hydroxymethoxybenzophenone sulfonic acid and its salts, dihydroxymethoxybenzophenone, dihydroxymethoxybenzophenone sodium disulfonate, dihydroxybenzophenone, dihydroxydimethoxybenzophenone, hydroxyoctoxybenzophenone, tetrahydroxybenzophenone, Butylmethoxydibenzoylmethane, 2,4,6-trianilino-p- (carbo-2-ethylhexyl-1-oxy) -1,3,5-triazine, 2- (2-hydroxy-5-methylphenyl) benzotriazole , Methyl-O-aminobenzoate, 2-ethylhexyl-2-cyano-3,3-diphenyl acrylate, phenylbenzimidazole sulfate, 3- (4-methylbenzylidene) camphor, Isopropyl dibenzoylmethane, 4- (3,4-dimethoxyphenyl methylene) 2-ethylhexyl-2,5-dioxo-1-imidazolidin propionate,, and polymer derivatives and silane derivatives thereof.
(5) Examples of powders and pigments Red 104, Red 201, Yellow 4, Blue 1, Black 401 and other dyes, Yellow 4 AL lake, Yellow 203 BA lake and other lake dyes, nylon powder , Silk powder, urethane powder, Teflon (registered trademark) powder, silicone powder, polymethyl methacrylate powder, cellulose powder, starch, silicone elastomer spherical powder, polyethylene powder, yellow iron oxide, red iron oxide, black Colored pigments such as iron oxide, chromium oxide, carbon black, ultramarine and bitumen, white pigments such as zinc oxide, titanium oxide and cerium oxide, extender pigments such as talc, mica, sericite, kaolin and barium sulfate plate, titanium mica Pearl pigments such as barium sulfate, calcium carbonate, magnesium carbonate, aluminum silicate, metal salts such as magnesium silicate, Inorganic powders such as Rica and alumina, metal soaps such as aluminum stearate, magnesium stearate, zinc palmitate, zinc myristate, magnesium myristate, zinc laurate, zinc undecylenate, bentonite, smectite, boron nitride, etc. It is done. There are no particular restrictions on the shape of these powders (spherical, rod-like, needle-like, plate-like, irregular shape, flake-like, spindle-like, etc.) and particle diameter. These powders are conventionally known surface treatments such as fluorine compound treatment, silicone treatment, silicone resin treatment, pendant treatment, silane coupling agent treatment, titanium coupling agent treatment, oil agent treatment, N-acylated lysine treatment, The surface treatment may or may not be performed in advance by polyacrylic acid treatment, metal soap treatment, amino acid treatment, lecithin treatment, inorganic compound treatment, plasma treatment, mechanochemical treatment or the like.
(6) Examples of surfactant [anionic surfactant]: fatty acid soap, α-acyl sulfonate, alkyl sulfonate, alkyl allyl sulfonate, alkyl naphthalene sulfonate, alkyl sulfate, POE alkyl ether Sulfates, alkylamide sulfates, alkyl phosphates, POE alkyl phosphates, alkylamide phosphates, alkyloyl alkyl taurine salts, N-acyl amino acid salts, POE alkyl ether carboxylates, alkyl sulfosuccinates, alkyls Examples include sodium sulfoacetate, acylated hydrolyzed collagen peptide salt, and perfluoroalkyl phosphate.
[Cationic surfactant]: alkyltrimethylammonium chloride, stearyltrimethylammonium chloride, stearyltrimethylammonium bromide, cetostearyltrimethylammonium chloride, distearyldimethylammonium chloride, stearyldimethylbenzylammonium chloride, behenyltrimethylammonium bromide, benzaza chloride Examples thereof include luconium, amidopropyldimethylhydroxypropylammonium chloride behenate, diethylaminoethylamide stearate, dimethylaminopropylamide stearate, and lanolin derivative quaternary ammonium salts.
[Amphoteric surfactant]: Carboxybetaine type, amide betaine type, sulfobetaine type, hydroxysulfobetaine type, amide sulfobetaine type, phosphobetaine type, aminocarboxylate type, imidazoline derivative type, amidoamine type and the like.
[Nonionic surfactant]: Propylene glycol fatty acid ester, glycerin fatty acid ester, polyglycerin fatty acid ester, sorbitan fatty acid ester, POE sorbitan fatty acid ester, POE sorbitol fatty acid ester, POE glycerin fatty acid ester, POE alkyl ether, POE fatty acid ester, POE Examples thereof include hydrogenated castor oil, POE castor oil, POE / POP copolymer, POE / POP alkyl ether, polyether-modified silicone lauric acid alkanolamide, alkylamine oxide, and hydrogenated soybean phospholipid.
[Natural surfactant]: Examples include lecithin, saponin, and sugar surfactant.
(7) Examples of polyhydric alcohols and sugars Ethylene glycol, diethylene glycol, polyethylene glycol, propylene glycol, dipropylene glycol, polypropylene glycol, glycerin, diglycerin, polyglycerin, 3-methyl-1,3-butanediol, 1,3 -Butylene glycol, sorbitol, mannitol, raffinose, erythritol, glucose, sucrose, fructose, xylitol, lactose, maltose, maltitol, trehalose, alkylated trehalose, mixed isomerized sugar, sulfated trehalose, pullulan and the like. These chemically modified compounds can also be used.
(8) Examples of polymers Acrylic ester / methacrylic acid ester copolymer (plus size, manufactured by Kyoyo Chemical), vinyl acetate / crotonic acid copolymer (resin 28-1310, manufactured by NSC), vinyl acetate / croton Acid / vinyl neodecanate copolymer (28-2930, manufactured by NSC), methyl vinyl ether maleic acid half ester (Gantrez ES, manufactured by ISP), T-butyl acrylate / ethyl acrylate / methacrylic acid copolymer (rubymer) , BASF), vinyl pyrrolidone / vinyl acetate / vinyl propionate copolymer (Rubicol VAP, BASF), vinyl acetate / crotonic acid copolymer (Rubiset CA, BASF), vinyl acetate / croton Acid / vinyl pyrrolidone copolymer (Rubiset CAP, manufactured by BASF), vinyl pyrrolidone / acrylate copolymer (Rubiflex, BAS) F), acrylate / acrylamide copolymer (Ultrahold, BASF), vinyl acetate / butyl maleate / isobornyl acrylate copolymer (Advantage, ISP), carboxyvinyl polymer (Carbopol, BF Goodrich), anionic polymer compounds such as acrylic acid / alkyl methacrylate copolymer (Pemulene, BF Goodrich), and acetic acid amphoteric products of dialkylaminoethyl methacrylate polymers (Yukaformer, Mitsubishi Chemical) Amphoteric polymer compounds such as octyl acrylate acrylate / hydroxypropyl acrylate / butylaminoethyl methacrylate copolymer (AMPHOMER, NSC), quaternized vinylpyrrolidone / dimethylaminoethyl methacrylate (GAFQUAT, ISP) , Methyl vinyl imidazolium chloride / Cationic polymer compounds such as vinylpyrrolidone copolymer (rubicoat, manufactured by BASF), polyvinylpyrrolidone (rubiscol K, manufactured by BASF), vinylpyrrolidone / vinyl acetate copolymer (rubiscol VA, manufactured by BASF) Nonionic polymer compounds such as vinylpyrrolidone / dimethylaminoethyl methacrylate copolymer (copolymer 937, manufactured by ISP), vinylcaprolactam / vinylpyrrolidone / dimethylaminoethyl methacrylate copolymer (copolymer VC713, manufactured by ISP), etc. is there. Cellulose or derivatives thereof, keratin and collagen or derivatives thereof, calcium alginate, pullulan, agar, gelatin, tamarind seed polysaccharide, xanthan gum, carrageenan, high methoxyl pectin, low methoxyl pectin, guar gum, gum arabic, crystalline cellulose, arabino Naturally derived polymer compounds such as galactan, gum karaya, gum tragacanth, alginic acid, albumin, casein, curdlan, gellan gum and dextran can also be suitably used.
(9) Examples of physiologically active ingredients Examples of physiologically active ingredients include substances that impart some physiological activity to the skin when applied to the skin. For example, whitening ingredients, anti-inflammatory agents, anti-aging agents, UV protection agents, slimming agents, tanning agents, antioxidants, hair growth agents, hair restorers, moisturizers, blood circulation promoters, antibacterial agents, bactericides, desiccants, A cooling sensation agent, a warming sensation agent, vitamins, an amino acid, a wound healing promoter, an irritation relaxation agent, an analgesic agent, a cell activator, an enzyme component, etc. are mentioned. Examples of these suitable ingredients include, for example, ashitaba extract, avocado extract, amateur extract, altea extract, arnica extract, aloe extract, apricot extract, apricot kernel extract, ginkgo biloba extract, fennel extract, turmeric extract, oolong tea extract, ages Extract, Echinacea leaf extract, Ogon extract, Oat extract, Oen extract, Barley extract, Hypericum extract, Oyster extract, Dutch mustard extract, Orange extract, Seawater dried product, Seaweed extract, Hydrolyzed elastin, Hydrolyzed wheat powder, Hydrolyzed silk , Chamomile extract, carrot extract, cormorant extract, licorice extract, calcade extract, oyster extract, kiwi extract, kina extract, cucumber extract, guanosine, gardenia extract, kumazasae Kiss, Clara extract, Walnut extract, Grapefruit extract, Clematis extract, Chlorella extract, Mulberry extract, Gentian extract, Black tea extract, Yeast extract, Burdock extract, Rice bran ferment extract, Rice germ oil, Comfrey extract, Collagen, Cowberry extract, Saisin extract , Psycho extract, Saitai extract, Salvia extract, Soybean extract, Sasa extract, Hawthorn extract, Salamander extract, Shiitake extract, Giant extract, Shikon extract, Perilla extract, Linden extract, Shimotake extract, Japanese peony extract, Ginger root extract, Birch extract, Japanese cedar Extract, Kizuta extract, Hawthorn extract, Elderberry extract, Achillea millefolium extract, Atlantic mint extract, Sage extract, Zeni mallow Extract, Senkyu Extract, Assembly Extract, Soybean Extract, Taiso Extract, Thyme Extract, Tea Extract, Fresh Coffee Bean Extract, Clove Extract, Chigaya Extract, Chimpi Extract, Toki Extract, Tokinsenka Extract, Tonin Extract, Spruce Extract, Dokudami Extract, Tomato Extract, Natto Extract, carrot extract, garlic extract, wild rose extract, hibiscus extract, buckwheat extract, parsley extract, honey, hamamelis extract, parietaria extract, toad extract, bisabolol, loquat extract, dandelion extract, burdock extract, butterfly extract, butcher bloom extract, Grape extract, propolis, loofah extract, safflower extract, peppermint extract, bodaiju extract, button extract, hop extract, matsue , Maronnier extract, Citrus extract, Mukuroji extract, Melissa extract, Peach extract, Cornflower extract, Eucalyptus extract, Yukinoshita extract, Yuzu extract, Yakuinin extract, Artemisia extract, Lavender extract, Apple extract, Lettuce extract, Lemon extract, Forsythia extract, Rose extract , Rosemary extract, roman chamomile extract, royal jelly extract and the like.
Biopolymers such as deoxyribonucleic acid, mucopolysaccharide, sodium hyaluronate, sodium chondroitin sulfate, collagen, elastin, chitin, chitosan, hydrolyzed eggshell membranes, amino acids, hydrolyzed peptides, sodium lactate, urea, sodium pyrrolidonecarboxylate , Moisturizing ingredients such as betaine, whey, trimethylglycine, oily ingredients such as sphingolipid, ceramide, phytosphingosine, cholesterol, cholesterol derivatives, phospholipids, ε-aminocaproic acid, glycyrrhizic acid, β-glycyrrhetinic acid, lysozyme chloride, guaiazulene, Anti-inflammatory agents such as hydrocortisone, vitamin A, vitamin B2, vitamin B6, vitamin C, vitamin D, vitamin E, calcium pantothenate, biotin, nicotinamide, vitamin C ester, etc. Active ingredients such as vitamins, allantoin, diisopropylamine dichloroacetate, 4-aminomethylcyclohexanecarboxylic acid, antioxidants such as tocopherol, carotenoid, flavonoid, tannin, lignan, saponin, α-hydroxy acid, β-hydroxy acid, etc. Cell activators, blood circulation promoters such as γ-oryzanol and vitamin E derivatives, wound healing agents such as retinol and retinol derivatives, whitening such as arbutin, kojic acid, placenta extract, sulfur, ellagic acid, linoleic acid, tranexamic acid, glutathione Agent, cephalanthin, licorice extract, red pepper tincture, hinokitiol, garlic iodide extract, pyridoxine hydrochloride, DL-α-tocopherol, DL-α-tocopherol acetate, nicotinic acid, nicotinic acid derivative, calcium pantothenate, D-pan Tenenyl alcohol, acetyl pantothenyl ethyl ether, biotin, allantoin, isopropylmethylphenol, estradiol, ethinyl estradiol, capronium chloride, benzalkonium chloride, diphenhydramine hydrochloride, takanal, camphor, salicylic acid, nonylic acid vanillylamide, nonanoic acid vanillylamide, piroctone Mines, glyceryl pentadecanoate, L-menthol, mononitroguaiacol, resorcin, γ-aminobutyric acid, benzethonium chloride, mexiletine hydrochloride, auxin, female hormone, cantalis tincture, cyclosporine, zinc pyrithione, hydrocortisone, minoxidil, polymonostearate Examples include hair restorers such as oxyethylene sorbitan, mint oil, and Sasanishiki extract.
(10) Examples of antioxidants: sodium bisulfite, sodium sulfite, erythorbic acid, sodium erythorbate, dilauryl thiodipropionate, tocopherol, tolylbiguanide, nordihydroguaiaretic acid, parahydroxyanisole, butylhydroxyanisole, dibutylhydroxy Examples include plant extracts having an antioxidant effect such as toluene, ascorbyl stearate, ascorbyl palmitate, octyl gallate, propyl gallate, carotenoid, flavonoid, tannin, lignan, saponin, apple extract and clove extract.
(11) Examples of solvents Examples include purified water, ethanol, lower alcohol, ethers, LPG, fluorocarbon, N-methylpyrrolidone, fluoroalcohol, volatile linear silicone, and next-generation fluorocarbon.

Examples of the present invention will be described below. In addition, the Example shown below is demonstrated for confirmation of the various effect | actions, effects, etc. of the black rice extract obtained by this invention, and the scope of the present invention is not limited to these products and production methods. Absent.

Example 1
The varieties “Shikokuen” were used as the black rice used as a preparation raw material for the angiogenesis inhibitor (black rice extract).
Black rice bran (400 g) was degreased with 5 times the weight ratio of n-hexane, and then the rice bran was air-dried.
Next, 50% ethanol was used for stirring extraction at room temperature. The extract was concentrated under reduced pressure at 40 ° C. or less to obtain 60 g of an angiogenesis inhibitor (black rice extract). As a result of analyzing the black rice extract of Example 1 by HPLC, it was confirmed that it contained 2.9% cyanidin-3-O-glucoside and 0.2% peonidin-3-O-glucoside.

Example 2 and Example 3
Example 2 Cyanidine and Peonidine Here, cyanidin (manufactured by Extrasynthese) was used as Example 2, and peonidin (manufactured by Extrasynthese) was used as Example 3.

Test Example 1: Measurement of evaluation of VEGF-induced human umbilical vein endothelial cell (HUVEC) on luminal formation An angiogenesis kit (Kurabo) was used for evaluation of luminal formation. This kit consists of co-culture of HUVEC and fibroblasts, and the addition of VEGF promotes HUVEC tube formation. The black rice extract (PRE) of Example 1 and the peonidin of Example 3 were dissolved in a VEGF-containing medium, and the medium was replaced on the first, fourth, seventh and ninth days of culture. On day 11, the cells were fixed with 70% ethanol and reacted with the primary antibody and the secondary antibody. Thereafter, the lumen of HUVEC was stained, 50 mm ² per well was photographed with a CCD camera (Keyence) (FIG. 1A), and analyzed with angiogenesis quantification software Ver. 2 (Kurabo). Area (luminal area, FIG. 1B), length (luminal length, FIG. 1C), joints (number of branch points, FIG. 1D) and paths (number of branches forming the lumen network, FIG. 1E) calculated by the quantitative software ) Was evaluated.

Results and effects of examples
Addition of VEGF promoted HUVEC lumen formation. Black rice extract (PRE) is effective for VEGF-induced HUVEC lumen formation in areas (lumen area), length (lumen length), joints (number of branch points) and paths (number of branches forming the lumen network). In both cases, a concentration-dependent significant inhibitory effect was observed (Fig. 1). In addition, the addition of peonidin has a significant inhibitory effect on area formation (luminal area), length (luminal length), joints (number of branch points) and paths (number of branches forming the luminal network). (Figure 2).

Test Example 2: Evaluation method for VEGF-induced HUVEC proliferation of black rice extract (PRE), cyanidin and peonidin: HUVEC was seeded at a density of 2000 cells / well in a 96-well plate, cultured for 24 hours, and then growth factors were removed. The medium was replaced and cultured for 24 hours. Each sample was dissolved in VEGF-containing medium. This medium was added to HUVEC and cultured for 72 hours. Thereafter, CCK-8 was added to each well, incubated for 3 hours at 37 ° C., 5% CO ₂ , and then directly measured for absorbance at 492 nm (reference wavelength: 660 nm) to count viable cells. The result is shown in FIG.

Results: With the addition of VEGF, HUVEC increased cells compared to the control group (V in FIG. 3). Addition of black rice extract (PRE) showed a significant concentration-dependent inhibitory effect on the growth of VEGF-induced HUVEC (FIG. 3A). Furthermore, a significant inhibitory action was similarly observed for cyanidin (FIG. 3B) and peonidin (FIG. 3C).

Test Example 3: VEGF-induced human retinal capillary endothelial cells (HRMEC: Human Retinal Microvascular Endothelial
Cell) Measurement of evaluation for growth promotion
HRMEC was seeded in a 96-well plate at a density of 2000 cells / well, cultured for 24 hours, replaced with a medium excluding growth factors, and cultured for 24 hours. Each sample was dissolved in VEGF-containing medium. This medium was added to HRMEC and cultured for 24 hours. Thereafter, CCK-8 was added to each well, incubated for 3 hours at 37 ° C., 5% CO ₂ , and then directly measured for absorbance at 492 nm (reference wavelength: 660 nm) to count viable cells. The result is shown in FIG. 4A. Further, the same test was performed for cyanidin (FIG. 4B) and peonidin (FIG. 4C).

Results: With the addition of VEGF, HRMEC increased cells compared to the control group. Addition of black rice extract (PRE) showed a significant concentration-dependent inhibitory effect on the growth of VEGF-induced HRMEC (FIG. 4A). Furthermore, significant inhibitory action was confirmed in a concentration-dependent manner for cyanidin (FIG. 4B) and peonidin (FIG. 4C).
Based on the above results, black rice extract is followed by HUVEC, followed by human retinal capillary endothelial cells.
(HRMEC) also showed a significant anti-angiogenic effect. From this, the black rice extract was confirmed to be effective against eye diseases that lead to blindness such as diabetic retinopathy and age-related macular degeneration associated with angiogenesis.

HRMEC run test
A 12-well plate was coated with collagen, seeded at a density of 4 × 10 ⁴ cells / well, and cultured for 48 hours at 37 ° C. and 5% CO ₂ . Thereafter, the medium was replaced with a medium excluding growth factors and cultured for 24 hours. Thereafter, the cells present on the center line of the well were detached using a 1 mL chip, washed with PBS, and the medium was changed. Immediately after that, I took a picture using a CCD camera (before running). VEGF and black rice extract (PRE) were added to the desired concentration and incubated for 24 hours. Thereafter, each well was photographed in the same manner (after migration, see FIG. 5A), and the number of cells that had moved to the detached location compared with that before migration was counted. The result is shown in FIG. Further, the same test was performed for peonidin and cyanidin. The result is shown in FIG. 5C.

Results: By adding VEGF, HRMEC increased the number of cells (number of migration) compared to the control group. Addition of black rice extract (PRE) showed a significant concentration-dependent inhibitory effect on VEGF-induced HRMEC migration (FIG. 5B). Furthermore, a significant inhibitory action was similarly confirmed for peonidin and cyanidin (FIG. 5C).

The compounding example of the angiogenesis inhibitor by this invention is shown.
Formulation Example 1: Chewing gum
53.0wt% sugar
Gum base 20.0
Glucose 10.0
Minamata 16.0
Fragrance 0.5
Angiogenesis inhibitor 0.5
100.0wt%

Formulation Example 2: Gummy
Reduced water tank 40.0wt%
Granulated sugar 20.0
Buto sugar 20.0
Gelatin 4.7
Water 9.68
Yuzu fruit juice 4.0
Yuzu Flavor 0.6
Dye 0.02
Angiogenesis inhibitor 1.0
100.0wt%

Formulation Example 3: Candy
50.0 wt% sugar
Minamata 33.0
Water 14.4
Organic acid 2.0
Fragrance 0.2
Angiogenesis inhibitor 0.4
100.0wt%

Formulation Example 4: Yogurt (hard / soft)
Milk 41.5wt%
Nonfat dry milk 5.8
Sugar 8.0
Agar 0.15
Gelatin 0.1
Lactic acid bacteria 0.005
Angiogenesis inhibitor 0.4
Perfume
Water residue
100.0wt%

Formulation Example 5: Soft drink
Fructose glucose liquid sugar 30.0wt%
Emulsifier 0.5
Angiogenesis inhibitor 0.3
Perfume
Purified water residue
100.0wt%

Formulation Example 6: Tablets
76.4 wt% sugar
Glucose 19.0
Sucrose fatty acid ester 0.2
Angiogenesis inhibitor 0.5
Purified water 3.9
100.0wt%

Formulation Example 7: Soft capsule
Brown rice germ oil 47.0wt%
Yuzu seed oil 40.0
Emulsifier 12.0
Angiogenesis inhibitor 1.0
100.0wt%

Formulation Example 8: Tablet
Lactose 54.0wt%
Crystalline cellulose 30.0
Starch degradation product 10.0
Glycerin fatty acid ester 5.0
Angiogenesis inhibitor 1.0
100.0wt%

Formulation Example 9: Eye drops Ketotifen fumarate 0.7 wt%
Sodium azulene sulfonate 0.2
Cromoglycate sodium 9.8
L-potassium aspartate 8.5
Allantoin 3.0
Tetrahydrozoline hydrochloride 0.5
Methyl sulfate neostigmine 0.05
Benzalkonium chloride solution 0.1
Glycerin 25.0
Angiogenesis inhibitor 1.0
pH regulator
Purified water balance
100.0wt%

Formulation Example 10: Cosmetic cream
Squalane 20.0wt%
Beeswax 5.0
Refined jojoba oil 5.0
Glycerin 5.0
Glycerol monostearate 2.0
Polyoxyethylene (20) sorbitan
Monostearate 2.0
Angiogenesis inhibitor 2.0
Preservative appropriate amount
Perfume
Purified water residue
100.0wt%

Formulation Example 11: Lotion
Ethanol 5.0wt%
Glycerin 2.0
1,3-butylene glycol 2.0
Polyethylene oleyl ether 0.5
Sodium citrate 0.1
Citric acid 0.1
Angiogenesis inhibitor 0.1
Purified water residue
100.0wt%

Formulation Example 12: Body gel
Macadamia nut oil 2.0wt%
Octyldodecyl myristate 10.0
Methylphenylpolysiloxane 5.0
Behenyl alcohol 3.0
Stearic acid 3.0
Batyl alcohol 1.0
Glyceryl monostearate 1.0
Tetraoleic acid polyoxyethylene sorbit
2.0
Hydrogenated soybean phospholipid 1.0
Ceramide 0.1
Retinol palmitate 0.1
Preservative appropriate amount
Camellia extract 1.0
Angiogenesis inhibitor 1.0
1,3-butylene glycol 5.0
Purified water residue
100.0wt%

Formulation Example 13: Latex
Squalane 4.0wt%
Vaseline 2.5
Cetanol 2.0
Glycerin 2.0
Lipophilic glyceryl monostearate
1.0
Stearic acid 1.0
L-Arginine 1.0
Angiogenesis inhibitor 0.5
Potassium hydroxide 0.1
Fragrance
Purified water residue
100.0wt%

Formulation Example 14: Bath agent (liquid)
Propylene glycol 50.0wt%
Ethanol 20.0
Sodium sulfate 5.0
Angiogenesis inhibitor 0.5
Lanolin 0.5
Avocado oil 0.5
Dye 1.5
Fragrance 22.0
100.0wt%

  As described above, the present invention can provide an angiogenesis inhibitor having a novel component that can inhibit angiogenesis and thereby effectively prevent cancer and eye diseases.

Claims (6)

An angiogenesis inhibitor comprising black rice extract as an active ingredient. An angiogenesis inhibitor comprising cyanidin and / or cyanidin glycoside as an active ingredient. An angiogenesis inhibitor comprising peonidin and / or peonidin glycoside as an active ingredient. A preventive agent for retinal degenerative disease comprising the agent according to any one of claims 1 to 3 as an active ingredient. The retinal degenerative disease is retinitis pigmentosa, cone dystrophy, diabetic retinopathy, age-related macular degeneration, age-related macular degeneration, macular edema, retinal detachment, cancer-related retinopathy, retinal vein occlusion or retinal pigment epithelial detachment The preventive agent for retinal degenerative disease according to claim 4. A cancer preventive / therapeutic agent comprising the agent according to any one of claims 1 to 3 as an active ingredient.