JP7598591B2 - HIF-inhibiting composition - Google Patents

Description

The present invention relates to a composition for suppressing HIF. The present invention also relates to a composition for protecting the retina, an anti-aging composition for the eye, a composition for suppressing VEGF in the eye, a composition for suppressing angiogenesis in the eye, a composition for maintaining homeostasis of vascular tissue in the eye, and a composition for reducing the risk of, preventing, or treating posterior ocular diseases.

HIF (Hypoxia Inducible Factor) is a transcription factor that is induced when cells are deficient in oxygen. It has a variety of physiological activities, including hypoxia adaptation response, maintenance of stem cells, and homeostasis such as controlling inflammation. However, overexpression of HIF is thought to be closely related to the onset and progression of cancer.

For this reason, for example, as disclosed in Non-Patent Document 1, HIF is being studied as a target for cancer treatment. On the other hand, as disclosed in Patent Document 1 and Non-Patent Document 2, suppressing HIF is expected to have therapeutic, preventive, and risk-reducing effects not only on cancer, but also on ophthalmological diseases and ocular abnormalities.

International Publication No. 2018/088566

Onnis B, et al. “Development of HIF-1 inhibitors for cancer therapy”J Cell Mol Med. 2009 Kurihara T, et al. “Hypoxia-induced metabolic stress in retinal pigment epithelial cells is sufficient to induce photoreceptor “degeneration” eLife 2016 Wang X, et al. “Mitochondrial ferritin effects mitochondria by stabilizing HIF-1arufwa in retinal pigment epithelium: implications The pathophysiology of age-related macular degeneration” Neurobiol Aging. 2016

However, as disclosed in Non-Patent Document 1, many of the compositions for suppressing HIF are anticancer drugs, which are highly cytotoxic and difficult to apply directly to, for example, eye diseases. Therefore, there is a demand for novel compositions for suppressing HIF that use ingredients that are highly safe and can be applied not only in the field of pharmaceuticals but also in the field of functional foods and supplements.

Therefore, the present invention aims to provide a composition that suppresses HIF, thereby effectively improving associated diseases and ocular abnormalities.

In order to solve the above problems, the inventors conducted intensive research and discovered that ingredients that are safely used in the food and pharmaceutical fields have been recognized to have HIF inhibitory activity, leading to the completion of the present invention.

That is, the present invention provides the compositions described below.
[1] A composition for suppressing HIF, comprising at least one selected from the group consisting of hydroxycitric acid and its salts, Garcinia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine or its derivatives and their salts, niacin, inositol, phytic acid, pantothenic acid, biotin, and folic acid.
[2] A retinal protective composition containing at least one selected from the group consisting of hydroxycitric acid and its salts, Garcinia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine or its derivatives and their salts, niacin, inositol, phytic acid, pantothenic acid, biotin, and folic acid.
[3] An anti-aging composition for the eye, comprising at least one selected from the group consisting of hydroxycitric acid and its salts, Garcinia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine or its derivatives and their salts, niacin, inositol, phytic acid, pantothenic acid, biotin, and folic acid.
[4] A composition for suppressing VEGF in the eye, comprising at least one member selected from the group consisting of hydroxycitric acid and its salts, Garcinia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine or its derivatives and their salts, niacin, inositol, phytic acid, pantothenic acid, biotin, and folic acid.
[5] A composition for inhibiting ocular angiogenesis, comprising at least one selected from the group consisting of hydroxycitric acid and its salts, Garcinia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine or its derivatives and their salts, niacin, inositol, phytic acid, pantothenic acid, biotin, and folic acid.
[6] A composition for maintaining homeostasis of ocular vascular tissue, comprising at least one selected from the group consisting of hydroxycitric acid and its salts, Garcinia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine or its derivatives and their salts, niacin, inositol, phytic acid, pantothenic acid, biotin, and folic acid.
[7] A composition for reducing the risk of, preventing, or treating a posterior eye disease, comprising at least one selected from the group consisting of hydroxycitric acid and its salts, Garcinia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine or its derivatives and their salts, niacin, inositol, phytic acid, pantothenic acid, biotin, and folic acid.
[8] The composition according to [7], wherein the posterior ocular disease is at least one selected from the group consisting of age-related macular degeneration, diabetic retinopathy, diabetic macular edema, retinitis pigmentosa, proliferative vitreous retinopathy, retinal artery occlusion, retinal vein occlusion, uveitis, Leber's disease, retinopathy of prematurity, retinal detachment, retinal pigment epithelial detachment, central serous chorioretinopathy, central exudative chorioretinopathy, polypoidal choroidal vasculopathy, multifocal choroiditis, neovascular maculopathy, retinal aneurysm, retinal angiomatous proliferation, optic neuropathy caused by these diseases, optic neuropathy caused by glaucoma, and ischemic optic neuropathy.

According to the present invention, it is possible to inhibit HIF using ingredients that are safely used in the food and pharmaceutical fields, and effectively improve ocular abnormalities and diseases caused by high levels of HIF.

1(A) to (D) are graphs showing the results of ERG (rod system function) measurement in Test Example 3, in which stimulation (10 cd s/m ² ) was applied under dark adaptation. 2(A) and (B) are graphs showing the results of ERG (cone system function) measurement under light-adapted stimulation (10 cd/m ² ) in Test Example 3. 3(A) and (B) are graphs showing the results of ERG (rod system function) measurement in Test Example 3, in which stimulation (10 cd s/m ² ) was applied under dark adaptation. FIG. 4 is a graph showing the results of ERG (cone system function) measurement under light-adapted stimulation (10 cd/m ² ) in Test Example 3. 5(A) is a graph showing the results of choroidal neovascularization (CNV) induction after laser irradiation on the 42nd day (6 weeks) after the start of administration in Test Example 4. FIG. 5(B) is a graph showing the results of choroidal neovascularization (CNV) induction after laser irradiation on the 56th day (8 weeks) after the start of administration in Test Example 4. FIG. 6 is a graph showing the results of choroidal neovascularization (CNV) induction after laser irradiation in Test Example 5. FIG. 7 is a graph showing the results of choroidal neovascularization (CNV) induction after laser irradiation in Test Example 6. FIG. 8 is a graph showing the results of choroidal neovascularization (CNV) induction after laser irradiation in Test Example 7. FIG. 9 is a graph showing the results of choroidal neovascularization (CNV) induction after laser irradiation in Test Example 8. FIG. 10 is a graph showing the results of confirming the expression of downstream genes of HIF in Test Example 9. FIG. 11 is a graph showing the results of confirming the expression of downstream genes of HIF in Test Example 9.

[HIF suppression composition]
In one embodiment, the composition for suppressing HIF of the present invention comprises a component selected from the group consisting of hydroxycitric acid and its salts, Garcinia cambogia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, and oleanolic acid and its salts. It contains at least one selected from the following.

In this specification, the plant extract may be a crude extract extracted from the whole plant or a necessary part of the plant (flower, inflorescence, flower bud, flower spike, leaf, branch, branch leaf, rhizome, root bark, root, bark, fruit, pericarp, legume, seed, etc.), or it may be a product obtained by further purification or concentration, or may be a product obtained by synthesis, or a commercially available product may be used. There are no particular limitations on the method for obtaining the plant extract, and conventional extraction methods, purification methods, concentration methods, synthesis methods, drying and powdering methods, etc. may be used.

(Hydroxycitric acid and its salts)
Hydroxycitric acid (HCA; 1,2-dihydroxypropane-1,2,3-tricarboxylic acid) is a natural substance that is naturally contained in the pericarp of plants of the Garcinia genus, such as Garcinia cambogia. Its calcium salt (calcium hydroxycitrate) is known to inhibit fatty acid biosynthesis. Garcinia cambogia is a plant of the Hypericaceae family native to Southeast Asia that has long been used as a spice and preservative, and is now well known as a diet ingredient due to its ability to inhibit fat synthesis. Various products containing HCA, including pharmaceuticals that claim to have anti-obesity effects, have been developed.

The salt of hydroxycitric acid is not particularly limited as long as it is medicamentarily, pharmacologically (pharmaceutical) or physiologically acceptable, but examples include alkali metal (particularly sodium) salts and alkaline earth metal (particularly calcium or magnesium) salts. Calcium salts are preferred as salts of hydroxycitric acid.

These hydroxycitric acids and/or salts thereof may be used alone or in any combination of two or more. Food materials containing hydroxycitric acid and/or salts thereof may also be used. Such materials include, but are not limited to, Garcinia cambogia and its extracts.

(Garcinia and its extract)
Garcinia and its extract refer to the fruit or peel of Garcinia campogia, an evergreen tree of the Hypericaceae family, its dried matter, its fine powder, and its extract.

There is no limitation on the method of extraction of Garcinia extract, so long as the effect of the present invention is achieved. As an example, Garcinia extract refers to an extract obtained by immersing Garcinia or its crushed product in water and/or an organic solvent and filtering the residue, or the extract from which the solvent has been removed, or a fine powder of these, or the extract or the product from which the solvent has been removed is dissolved, dispersed, or diluted with an appropriate solvent, and it is also possible to use commercially available products. As an example of another extraction method, Garcinia extract can also be extracted after processing such as steaming the fruit or peel of Garcinia cambogia.

In the present specification, when using a plant extract (also referred to as a plant extract), more specifically, as the extraction solvent, water (including hot water), alcohols such as methanol, ethanol, isopropanol, ethylene glycol, 1,3-butylene glycol, glycerin, etc., esters such as ethyl acetate, ketones such as acetone and methyl ethyl ketone, nitriles such as acetonitrile, ethers such as diethyl ether and tetrahydrofuran, saturated hydrocarbons such as pentane, hexane, cyclopentane, cyclohexane, aromatic hydrocarbons such as toluene, halogenated hydrocarbons such as dichloromethane and chloroform, and other organic solvents such as dimethylformamide and dimethyl sulfoxide (which may all contain water), etc., may be used appropriately, and may be any mixture of one or two of them. Of these solvents, water, ethanol, 1,3-butylene glycol, or a mixture of these is preferred. The extracts described in this specification can be obtained from various raw material companies, and are usually sold in a form including excipients, but are not limited thereto. Hereinafter, the amount of extract refers to the amount including the dry solid content and excipients.

The extraction solvent for Garcinia extract is not limited as long as it achieves the effects of the present invention, but is preferably water, ethanol, or aqueous ethanol, and more preferably aqueous ethanol.

Examples of food materials containing hydroxycitric acid or commercially available Garcinia extract products include, but are not limited to, Super Citrimax (Ryusendo Co., Ltd.), Garcinia Cambogia Extract Powder (Vidya Japan Co., Ltd.), Garcinia Cambogia (BioActives Japan Co., Ltd.), Garcitrin (Sabinsa Japan Corporation), and Garcinia Extract Powder (Koshin Bussan Co., Ltd.).

(Lactoferrin)
Lactoferrin is a glycoprotein with a molecular weight of about 80,000 daltons, which can reversibly bind two molecules of iron per molecule, and is present in mammalian milk.

Examples of lactoferrin used in the present invention include lactoferrin obtained by separating and purifying the raw material from mammalian (e.g., human, cow, goat, sheep, horse, etc.) milk (colostrum, transitional milk, normal milk, terminal milk, etc.) or processed products of such milk (skim milk, whey, etc.) by a standard method (e.g., ion exchange chromatography, etc.) (isolated lactoferrin); apolactoferrin obtained by removing iron from the isolated lactoferrin with hydrochloric acid, citric acid, etc.; and various lactoferrins produced by genetic manipulation in microorganisms, animal cells, transgenic animals, etc. Any one of these lactoferrins may be used, or a mixture of two or more may be used. Commercially available lactoferrin may also be used.

Food materials containing lactoferrin may also be used. Examples of food materials containing lactoferrin include, but are not limited to, commercially available products containing lactoferrin, such as Lactoferrin (Rivason Co., Ltd.), Aura Barrier LF+LPO (Morinaga Milk Industry Co., Ltd.), IDP (Kanematsu Chemical Co., Ltd.), and Bioferrin (Glanbia Japan Co., Ltd.).

(Carrot and its extract)
Carrots and their extracts refer to carrots, finely powdered carrots (carrot powder), and their extracts.

Specifically, ginseng powder is "ginseng" powder as defined in the Japanese Pharmacopoeia, and contains 0.10% or more of ginsenoside Rg1 and 0.20% or more of ginsenoside Rb1 based on the calculated dried matter of the herb. Ginseng powder that meets the standards set out in the Japanese Pharmacopoeia is commercially available and can be used.

The carrot extract is not limited to a specific extraction method as long as it provides the effects of the present invention. As an example, the carrot extract refers to an extract obtained by immersing and extracting carrots or crushed carrots in water and/or an organic solvent and filtering the residue, or the extract from which the solvent has been removed, or a fine powder of these, or the extract or the solvent-removed product dissolved, dispersed, or diluted with an appropriate solvent, and commercially available products can also be used. As another example of an extraction method, the carrot extract can be extracted after processing the carrots, such as by steaming them.

The extraction solvent for carrot extract is not limited as long as it achieves the effects of the present invention, but is preferably water, ethanol, or aqueous ethanol, and more preferably aqueous ethanol.

In addition, the origin of the ginseng and its extract is not limited as long as it produces the effects of the present invention, but it is preferable that it is Panax ginseng (Korean ginseng), American ginseng (Western ginseng), Siberian ginseng (Eleuthero), Sanshichi ginseng (Tanshichi ginseng, Denshichi ginseng), or a combination of these, and more preferably Panax ginseng (Korean ginseng).

Commercially available products of ginseng and its extracts include, but are not limited to, the following. Examples of Panax ginseng include Black Ginseng Extract Powder (Nihon Yakuhin Co., Ltd.), Korean Ginseng Extract Powder (Frontier Foods Co., Ltd.), Korean Ginseng Extract (Sankt Co., Ltd.), Ginseng Extract Powder S (Matsuura Yakugyo Co., Ltd.), and Ginseng Cansou Extract 10% (Indena Japan Co., Ltd.). Examples of American ginseng include Western Ginseng Extract Powder (FAP Japan Co., Ltd.), Western Ginseng Extract Powder (Matsuura Yakugyo Co., Ltd.), Wisconsin Ginseng (Kokan Pharmaceutical Research Institute Co., Ltd.), and American Ginseng Extract (Tama Biochemical Co., Ltd.). Examples of Siberian ginseng include Eleuthero Extract (Matsuura Yakugyo Co., Ltd.), Siberian Ginseng Extract Powder (Koshin Bussan Co., Ltd.), Eleuthero Extract Powder (FAP Japan Co., Ltd.), Eleuthero Extract Powder (Kokan Pharmaceutical Research Institute Co., Ltd.), and Siberian Ginseng Dried Extract (Ask Pharmaceutical Co., Ltd.). Examples of Sanchi ginseng (sanchi ginseng, danchi ginseng) include danchi ginseng extract powder (Nakahara Co., Ltd.), danchi ginseng extract powder (Matsuura Pharmaceutical Co., Ltd.), danchi ginseng (Adaptogen Pharmaceutical Co., Ltd.), etc., but commercially available products of ginseng and its extracts are not limited to these.

(Pyridoxine and its salts)
Pyridoxine is a compound also known as 5-hydroxy-6-methylpyridine-3,4-dimethanol. Pyridoxine and/or its salts are known compounds as vitamin _B6 , a water-soluble vitamin, and may be synthesized by known methods or may be commercially available.

Salts of pyridoxine are not particularly limited as long as they are medicamentally, pharmacologically (pharmaceutical), or physiologically acceptable, but specific examples include organic acid salts [e.g., monocarboxylates (acetate, trifluoroacetate, butyrate, palmitate, stearate, etc.), polycarboxylates (fumarate, maleate, succinate, malonate, etc.), oxycarboxylates (lactate, tartrate, citrate, etc.), organic sulfonates (methanesulfonate, toluenesulfonate, tosylate, etc.)], inorganic acid salts (e.g., hydrochloride, sulfate, nitrate, hydrobromide, phosphate, etc.), etc.

These pyridoxines and/or salts thereof may be used alone or in any combination of two or more. From the viewpoint of improving the foreign body sensation in the eye, the (A) component is preferably an inorganic acid salt of pyridoxine, more preferably pyridoxine hydrochloride or phosphate, and even more preferably pyridoxine hydrochloride.

Food materials containing pyridoxine and/or a salt thereof may also be used. Examples of food materials containing pyridoxine and/or a salt thereof or commercially available products thereof include, for example, red pepper, rice bran, garlic, basil, parsley, wheat germ, etc., and examples of commercially available products thereof include, but are not limited to, vitamin _B6- containing yeast (Ceti Co., Ltd.), vitamin _B6 (DSM Nutrition Japan, Inc.), Capsicum oleoresin (Vidya Japan Co., Ltd.), selected dried defatted rice bran (Tsukuno Foods Co., Ltd.), Oryza Dorim (Oryza Oil & Fat Chemical Co., Ltd.), and garlic extract (BioActives Japan Co., Ltd.).

Among food materials containing pyridoxine and/or a salt thereof, rice bran is preferred from the viewpoint of achieving a significant effect of the present invention, and defatted rice bran (also called defatted rice bran extract) is more preferred.

Defatted rice bran is obtained by defatting rice bran obtained by polishing brown rice. It can also be used to make extracts with various extraction solvents. In the present invention, rice bran obtained from the seeds of rice (Oryza sativa) belonging to the genus Oryza in the family Poaceae is used as the raw material, and rice bran derived from any variety of rice, such as japonica or indica, can be used.

(Oleanolic acid and its salts)
Oleanolic acid is a type of β-amyrin triterpene, and is known to exist as a free form or saponin in olive leaves, Swertia japonica, Clove, Grape, Beet, Persimmon, Plum, Loquat, Sea Buckthorn, Argan, and Sugar Beet. Oleanolic acid can be extracted and purified from the above-mentioned raw materials by known methods, or can be obtained by chemical synthesis or as a commercially available product.

Food materials containing oleanolic acid may also be used. Examples of food materials containing oleanolic acid or commercially available products thereof include, but are not limited to, Oleanolic-55 (Tama Biochemical Co., Ltd.), Oleavita (Nutrition Act Co., Ltd.), Nippum Pamis Extract GR, Nippum Pamis Extract OA (Nippon Flour Mills Co., Ltd.), and Palo Asul (IHM Co., Ltd.).

There are no particular limitations on the oleanolic acid, so long as it is medicamentarily, pharmacologically (pharmaceutical) or physiologically acceptable, but examples include alkali metal salts such as sodium salt and potassium salt, alkaline earth metal salts such as calcium salt, and organic acid salts including basic amino acid salts such as arginine salt.

(Thiamin or its derivatives and their salts)
Thiamine is a compound known as vitamin _B1 , a water-soluble vitamin, and can be synthesized by known methods or obtained commercially.

There are no particular limitations on the salts of thiamine so long as they are medicamentally, pharmacologically (pharmaceutical), or physiologically acceptable, but specific examples include organic acid salts [e.g., monocarboxylates (acetate, trifluoroacetate, butyrate, palmitate, stearate, etc.), polycarboxylates (fumarate, maleate, succinate, malonate, etc.), oxycarboxylates (lactate, tartrate, citrate, etc.), organic sulfonates (methanesulfonate, toluenesulfonate, tosylate, etc.)], inorganic acid salts (e.g., hydrochloride, sulfate, nitrate, hydrobromide, phosphate, etc.), etc.

These thiamine or its derivatives and their salts may be used alone or in any combination of two or more. Examples of thiamine or its derivatives and their salts include dibenzoylthiamine, dibenzoylthiamine hydrochloride, thiamine cetyl sulfate, thiamine thiocyanate, thiamine naphthalene-1,5-disulfonate, thiamine lauryl sulfate, and bisbentiamine. Among these, inorganic acid salts of thiamine are preferred, with thiamine hydrochloride or nitrate being more preferred, and nitrate being even more preferred.

Food materials containing thiamine or its derivatives and their salts may also be used. Examples of food materials containing thiamine or its derivatives and their salts or commercially available products thereof include, but are not limited to, thiamine nitrate (Nakahara Co., Ltd.), thiamine hydrochloride (Sankt Co., Ltd.), vitamin B1-containing yeast (Seti Co., Ltd.), vitamin B1 (DSM Co., Ltd.), selected dried defatted rice bran (Tsukuno Foods Co., Ltd.), Oryza Dorim (Oryza Oil & Fat Chemical Co., Ltd.), germ powder (Nippon Flour Mills Co., Ltd.), and sesame (BioActives Japan Co., Ltd.).

(Niacin)
Niacin is a general term for nicotinic acid (vitamin B3, niacin) and its amide compounds (nicotinamide, nicotinamide, niacinamide), and is a known compound as a water-soluble vitamin. It can be synthesized by known methods or obtained as a commercially available product.

Niacin may be used alone or in any combination of two or more. From the viewpoint of improving the sensation of foreign body in the eye, niacinamide is preferred as component (A).

Food materials containing niacin may also be used. Examples of food materials containing niacin or commercially available products thereof include, but are not limited to, rice bran, niacinamide-containing yeast (Seti Co., Ltd.), nicotinic acid (DSM Co., Ltd.), selected dried defatted rice bran (Tsukuno Foods Co., Ltd.), and Oryza Dorim (Oryza Oil & Fat Chemical Co., Ltd.).

(Inositol)
Inositol is a known compound that is a type of cyclitol and has a structure (1,2,3,4,5,6-cyclohexanehexaol) in which one hydrogen atom on each carbon of cyclohexane is replaced with a hydroxyl group. It can be synthesized by known methods or obtained commercially.

In addition, food materials containing inositol may be used. Examples of food materials containing inositol or commercially available products thereof include, but are not limited to, rice bran, inositol (Sankt Co., Ltd.), inositol (Tsukuno Foods Industry Co., Ltd.), selected dried defatted rice bran (Tsukuno Foods Industry Co., Ltd.), Oryza Dorim (Oryza Oil & Fat Chemical Co., Ltd.), and Kittle Syrup (Pures Japan Co., Ltd.).

(Phytic acid and its salts)
Phytic acid is a biological substance that is a hexaphosphate ester of myo-inositol, also known as myo-inositol-1,2,3,4,5,6-hexaphosphate (myo-inositol-1,2,3,4,5,6-hexaphosphate, hexakisphosphate, or hexakis(dihydrogenphosphate)). It can be synthesized by known methods or obtained commercially. Salts of phytic acid include calcium and magnesium salts of phytic acid (phytin), which are prepared from plant seeds and grains and are obtained as white powders that are almost insoluble in water.

Food materials containing phytic acid may also be used. Examples of food materials containing phytic acid or commercially available products thereof include, but are not limited to, rice bran, phytic acid (IP6) (Tsukuno Foods Industry Co., Ltd.), phytic acid FC (FC Chemical Co., Ltd.), phytic acid (Fuso Chemical Co., Ltd.), selected dried defatted rice bran (Tsukuno Foods Industry Co., Ltd.), and Oryza Dorim (Oryza Oil & Fat Chemical Co., Ltd.).

(Pantothenic acid and/or its salts)
Pantothenic acid, D(+)-N-(2,4-dihydroxy-3,3-dimethylbutyryl)-β-alanine, is a known compound that is a water-soluble vitamin belonging to the vitamin B group, and can be synthesized by known methods or obtained commercially.

There are no particular limitations on the salts of pantothenic acid, so long as they are medicamentally, pharmacologically (pharmaceutical), or physiologically acceptable, but specific examples include calcium salts, sodium salts, ammonium salts, etc.

These pantothenic acids and/or salts thereof may be used alone or in any combination of two or more. As component (A), calcium salts of pantothenic acid are preferred.

Food materials containing pantothenic acid and/or a salt thereof may also be used. Examples of food materials containing pantothenic acid and/or a salt thereof or commercially available products thereof include, but are not limited to, pantothenic acid (DSM Co., Ltd.), pantothenic acid-containing yeast (Cetie Co., Ltd.), selected dried defatted rice bran (Tsukuno Foods Co., Ltd.), Oryza Dorim (Oryza Oil & Fat Chemical Co., Ltd.), and Capsicum oleoresin (Vidya Japan Co., Ltd.).

(Biotin)
It is a water-soluble vitamin classified as a B vitamin, and can be synthesized by known methods or obtained commercially.

Food materials containing biotin may also be used. Examples of food materials containing phytic acid or commercially available products thereof include, but are not limited to, rice bran, biotin-containing yeast (Cetie Co., Ltd.), selected dried defatted rice bran (Tsukuno Foods Co., Ltd.), and Oryza Dorim (Oryza Oil & Fat Chemical Co., Ltd.).

(Folic Acid)
Folic acid is a water-soluble vitamin classified as a B vitamin, which has a structure in which pteridine is bound to para-aminobenzoic acid and one or more glutamic acids, and is a physiologically active substance classified as a water-soluble vitamin. It can be synthesized by a known method or is available commercially.

Food materials containing folic acid may also be used. Examples of food materials containing folic acid or commercially available products thereof include, but are not limited to, rice bran, folic acid-containing yeast (Cetie Co., Ltd.), and folic acid (DSM Co., Ltd.).

The total content of at least one selected from the group consisting of hydroxycitric acid and its salts, Garcinia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine and its salts, niacin, inositol, phytic acid and its salts, pantothenic acid and its salts, biotin, folic acid, and rice bran is appropriately set depending on the type and content of other blended ingredients and the formulation form of the composition, etc., and is preferably 0.1 to 95% by mass, more preferably 0.3 to 80% by mass, even more preferably 0.5 to 75% by mass, and particularly preferably 1 to 70% by mass, based on the total amount of the composition.

When hydroxycitric acid and its salts are contained, the content of hydroxycitric acid and its salts alone is not particularly limited, but is, for example, preferably 0.1 to 95% by mass, more preferably 1 to 80% by mass, even more preferably 10 to 75% by mass, and particularly preferably 20 to 70% by mass, relative to the total amount of the composition.

When Garcinia and its extract are contained, the content of Garcinia and its extract alone is not particularly limited, but is, for example, preferably 0.1 to 95% by mass, more preferably 1 to 80% by mass, even more preferably 10 to 75% by mass, and particularly preferably 20 to 70% by mass, relative to the total amount of the composition.

When carrot and its extract are contained, the content of carrot and its extract alone is not particularly limited, but is, for example, preferably 0.1 to 95% by mass, more preferably 1 to 80% by mass, even more preferably 10 to 75% by mass, and particularly preferably 20 to 70% by mass, relative to the total amount of the composition.

When pyridoxine and its salts are contained, the content of pyridoxine and its salts alone is not particularly limited, but is, for example, preferably 0.1 to 95% by mass, more preferably 0.3 to 70% by mass, even more preferably 0.5 to 50% by mass, and particularly preferably 1 to 30% by mass, relative to the total amount of the composition.

When oleanolic acid and its salts are contained, the content of oleanolic acid and its salts alone is not particularly limited, but is, for example, preferably 0.1 to 95% by mass, more preferably 1 to 80% by mass, even more preferably 10 to 75% by mass, and particularly preferably 20 to 70% by mass, relative to the total amount of the composition.

When lactoferrin is contained, the content of lactoferrin alone is not particularly limited, but is, for example, preferably 0.1 to 95% by mass, more preferably 1 to 80% by mass, even more preferably 10 to 75% by mass, and particularly preferably 20 to 70% by mass, relative to the total amount of the composition.

When thiamine is contained, the content of thiamine alone is not particularly limited, but is, for example, preferably 0.001 to 95% by mass, more preferably 0.01 to 50% by mass, and even more preferably 0.1 to 10% by mass, relative to the total amount of the composition.

When niacin is contained, the content of niacin alone is not particularly limited, but is, for example, preferably 0.01 to 95% by mass, more preferably 0.1 to 80% by mass, and even more preferably 1 to 75% by mass, relative to the total amount of the composition.

When inositol is contained, the content of inositol alone is not particularly limited, but is, for example, preferably 1 to 95% by mass, more preferably 5 to 80% by mass, and even more preferably 10 to 75% by mass, relative to the total amount of the composition.

When phytic acid is contained, the content of phytic acid alone is not particularly limited, but is, for example, preferably 1 to 95% by mass, more preferably 10 to 80% by mass, and even more preferably 20 to 75% by mass, relative to the total amount of the composition.

When pantothenic acid is contained, the content of pantothenic acid alone is not particularly limited, but is, for example, preferably 0.001 to 95% by mass, more preferably 0.01 to 50% by mass, and even more preferably 0.1 to 10% by mass, relative to the total amount of the composition.

When biotin is contained, the content of biotin alone is not particularly limited, but is, for example, preferably 0.00005 to 50% by mass, more preferably 0.0001 to 10% by mass, and even more preferably 0.001 to 1% by mass, relative to the total amount of the composition.

When folic acid is contained, the content of folic acid alone is not particularly limited, but is, for example, preferably 0.00005 to 50% by mass, more preferably 0.0001 to 10% by mass, even more preferably 0.001 to 1% by mass, and particularly preferably 20 to 70% by mass, relative to the total amount of the composition.

[Application]
The HIF-suppressing composition of the present invention is also suitable for use in improving conditions, symptoms, and diseases associated with the inhibition of HIF. As mentioned above, Non-Patent Document 2 and the like show the relationship between HIF and ocular abnormalities. Furthermore, Non-Patent Document 3 and the like show that abnormally high levels of HIF are observed in retinal pigment epithelial cells (RPE cells) with aging. Therefore, inhibiting HIF in retinal cells is also effective for anti-aging of the eye.

It is also known that VEGF expression is regulated by HIF-1α and HIF-2α, which are types of HIF. Therefore, inhibiting HIF in retinal cells is effective in suppressing VEGF in the eye and suppressing angiogenesis induced by VEGF. In this way, inhibiting HIF in retinal cells suppresses angiogenesis, and is therefore also effective in maintaining the homeostasis of vascular tissue in the eye.

In addition, it is known that inhibiting HIF has a therapeutic effect on retinal ganglion cells and photoreceptor cell disorders in the posterior segment of the eye, and a protective effect on retinal ganglion cells and photoreceptor cells. Therefore, the above-mentioned components are effective against posterior segment diseases and abnormalities, including glaucoma and age-related macular degeneration.

In this specification, the posterior segment of the eye refers to the area inside the back surface of the crystalline lens of the eye, including the retina, macula, sclera, choroid, vitreous body, and optic nerve. Of the areas of the posterior segment of the eye, it is preferable to target the retina in order to achieve a significant effect of the present invention.

In this specification, a posterior segment abnormality refers to a state in which some abnormality or finding occurs in at least one selected from the group consisting of the retina, macula, sclera, choroid, vitreous body, and optic nerve, although it does not correspond to a disease in the posterior segment. Abnormalities and (clinical) findings in the posterior segment can be found, for example, through a health check, a medical examination by a doctor, a clinical test, etc. Examples of examinations of the posterior segment include fundus examination, intraocular pressure examination, fundus three-dimensional image analysis (OCT) examination, visual field examination, refraction examination, slit lamp microscopy, etc. Among these examinations, for example, in fundus examination, abnormal findings in the posterior segment are evaluated from the viewpoint of hypertensive changes and arteriosclerotic changes using "Scheie's classification". In addition, in addition to the "Scheie classification", fundus examination may show findings such as enlarged optic nidus, leopard-striated fundus, hard exudates, drusen, suspected glaucoma, hemorrhage, suspected hemorrhage, retinal choroidal atrophy, suspected cataract, crossing phenomenon, exudates, retinal degeneration, meandering, suspected macular degeneration, conus, optic disc findings, etc. The present invention may also be used for reducing the risk, preventing, or treating posterior segment abnormalities.

In this specification, prevention of posterior ocular abnormalities means preventing or delaying the onset of posterior ocular abnormalities, or reducing the risk of onset of posterior ocular abnormalities.

As used herein, improvement of posterior ocular abnormalities refers to alleviation or improvement of a condition in which there is an abnormality in the posterior ocular segment, prevention or delay of the worsening of the abnormal condition, or prevention, delay or reversal of the progression of a condition in which there is an abnormality in the posterior ocular segment.

The posterior ocular disease targeted by the composition for reducing, preventing or treating the risk of posterior ocular diseases of the present invention includes, but is not limited to, at least one selected from the group consisting of age-related macular degeneration, diabetic retinopathy, diabetic macular edema, retinitis pigmentosa, proliferative vitreous retinopathy, retinal artery occlusion, retinal vein occlusion, uveitis, Leber's disease, retinopathy of prematurity, retinal detachment, retinal pigment epithelial detachment, central serous chorioretinopathy, central exudative chorioretinopathy, polypoidal choroidal vasculopathy, multifocal choroiditis, neovascular maculopathy, retinal aneurysm, retinal angiomatous proliferation, optic nerve disorders caused by these diseases, optic nerve disorders caused by glaucoma and ischemic optic nerve disorders. Among these posterior ocular diseases, it is preferable to target age-related macular degeneration and/or retinitis pigmentosa from the viewpoint of significantly exerting the effects of the present invention.

As used herein, prevention of posterior ocular disease means preventing or delaying the onset of a posterior ocular disease or symptom, or reducing the risk of onset of a posterior ocular disease or symptom.

As used herein, treatment of a posterior ocular disease refers to alleviating or improving a posterior ocular disease or symptom, preventing or delaying the worsening of a disease or symptom, or preventing, delaying, or reversing the progression of a posterior ocular disease or symptom.

Improvement of posterior segment abnormalities or prevention of posterior segment diseases includes, but is not limited to, reducing the risk of developing at least one selected from the group consisting of age-related macular degeneration, diabetic retinopathy, diabetic macular edema, retinitis pigmentosa, proliferative vitreous retinopathy, retinal artery occlusion, retinal vein occlusion, uveitis, Leber's disease, retinopathy of prematurity, retinal detachment, retinal pigment epithelium detachment, central serous chorioretinopathy, central exudative chorioretinopathy, polypoidal choroidal vasculopathy, multifocal choroiditis, neovascular maculopathy, retinal aneurysm, retinal angiomatous proliferation, optic neuropathy caused by these diseases, optic neuropathy caused by glaucoma, and ischemic optic neuropathy.

The composition of the present invention can be added to or mixed with food, beverages, feed, or pet food. Alternatively, it can be used as a beverage or food as it is. Alternatively, it can be added to or blended with foods and beverages that are labeled as having functional properties such as retinal protection, anti-aging of the eye, VEGF inhibition, angiogenesis inhibition, maintenance of vascular tissue homeostasis, and risk reduction or prevention of posterior ocular diseases, i.e., health foods, functional foods, foods for the sick, and foods for specified health uses.

Health foods, functional foods, foods for the sick, and foods for specified health uses can be used in various formulation forms, such as solid preparations (tablets, granules, fine granules, powders, capsules, chewable tablets, etc.), liquid preparations (syrups, suspensions), and liquid diets. Foods in formulation form can be produced in the same manner as known pharmaceutical preparations, and can be produced by mixing the active ingredient with a carrier acceptable for use as a food, such as a suitable excipient, and then using conventional means.

For example, tablets can be prepared by mixing a powdered active ingredient with a pharma- ceutically acceptable carrier component (such as an excipient) and compressing the mixture, and confectionery tablets such as candy can be prepared by pouring the mixture into a mold. Tablets may be sugar-coated to form sugar-coated tablets. Furthermore, tablets may be single-layer tablets or layered tablets such as double-layer tablets.

Granules and other powdered granules may be prepared by various granulation methods (extrusion granulation, milling granulation, dry compaction granulation, fluidized bed granulation, rolling granulation, high speed stirring granulation, etc.), and tablets can be prepared by an appropriate combination of the above granulation methods and tableting methods (wet tableting, direct tableting), etc.

Capsules can be prepared by filling a capsule (soft or hard capsule) with a powdered agent (powder, granules, etc.) in a conventional manner.

The liquid preparation can be prepared by dissolving or dispersing each component in an aqueous medium (purified water, purified water containing ethanol, etc.) which is a carrier component, filtering or sterilizing the mixture as necessary, filling the mixture into a specified container, and sterilizing the mixture. The preferred dosage form of the solid preparation of the present invention is a capsule or tablet, and a soft capsule (soft capsule) is more preferred.

Soft capsules have a smooth surface and are easy to swallow, making them popular with users. Common methods for manufacturing soft capsules include the flat plate method, the rotary method, and the seamless method.

In the rotary method (punching method) manufacturing, a sheet-like capsule shell sandwiches the flowing filling contents and forms a capsule shape along the holes of a rotating cylindrical die. On the other hand, in the seamless method (dropping method) manufacturing, the capsule shell composition and contents are simultaneously ejected from multiple concentric nozzles to form a seamless capsule shape.

The base material for the soft capsule shell is not particularly limited, but may be starch, pullulan, cellulose, polyvinyl alcohol, gelatin, succinated gelatin, etc., with starch, gelatin, and succinated gelatin being preferred, and gelatin and succinated gelatin being more preferred. These may be used alone or in combination of two or more.

The compositions of the present invention can also be manufactured into liquid beverages such as soups, juices, fruit juice drinks, milk, dairy drinks, whey drinks, lactic acid bacteria drinks, tea drinks, alcoholic beverages, coffee drinks, carbonated drinks, soft drinks, water drinks, cocoa drinks, jelly drinks, sports drinks, and diet drinks; semi-solid foods such as puddings and yogurt; noodles, confectioneries, spreads, etc.

When the composition of the present invention is prepared as a food composition, various food additives may be blended. Examples of food additives include antioxidants, colorants, flavorings, seasonings, sweeteners, acidulants, pH adjusters, quality stabilizers, preservatives, etc.

When the composition of the present invention is prepared as a pharmaceutical composition, it is prepared as a formulation containing at least one active ingredient selected from the group consisting of hydroxycitric acid and its salts, Garcinia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, and oleanolic acid and its salts, and preferably a pharmaceutical acceptable carrier. A pharmaceutical acceptable carrier generally refers to an inactive, non-toxic, solid or liquid bulking agent, diluent, or encapsulating material that does not react with the active ingredient, such as water, ethanol, polyols, appropriate mixtures thereof, and solvents or dispersion media such as vegetable oils.

The pharmaceutical composition is administered orally or parenterally, for example, into the oral cavity, into the digestive tract, or into the nasal cavity. Oral preparations include solid preparations (tablets, granules, fine granules, powders, capsules, chewable tablets, etc.) and liquid preparations (syrups, suspensions, inhalants, etc.). Parenteral preparations include eye drops, drops, nasal drops, and injections, etc.

The pharmaceutical composition may further contain additives commonly used in the pharmaceutical field. Such additives include, for example, excipients, binders, disintegrants, lubricants, antioxidants, colorants, flavoring agents, etc., and can be used as needed. The composition may also be coated with a known retardant or the like to achieve sustained release so as to act for a long time. The pharmaceutical composition may further contain other additives or drugs, such as antacids and gastric mucosa protectors, as needed.

The pharmaceutical composition can be applied in the form of an oral composition, an oral composition, etc. The pharmaceutical composition can be used therapeutically or non-therapeutically.

The daily oral intake or dosage of hydroxycitric acid and/or its salts for an adult can be appropriately determined depending on the individual's condition, body weight, sex, age, activity of the material, intake or administration route, intake or administration schedule, formulation form, or other factors. The daily oral intake or dosage of hydroxycitric acid and/or its salts for an adult is, for example, preferably 0.01 mg/kg body weight/day or more, more preferably 0.1 mg/kg body weight/day or more, even more preferably 0.5 mg/kg body weight/day or more, particularly preferably 1 mg/kg body weight/day or more, and most preferably 10 mg/kg body weight/day or more. The daily oral intake or dosage of hydroxycitric acid and/or its salts for an adult is, for example, preferably 400 mg/kg body weight/day or less, more preferably 300 mg/kg body weight/day or less, even more preferably 100 mg/kg body weight/day or less, particularly preferably 50 mg/kg body weight/day or less, and most preferably 30 mg/kg body weight/day or less. The oral intake or dosage of hydroxycitric acid and/or its salt per day for an adult is, for example, preferably 0.01 to 400 mg/kg body weight/day, more preferably 0.1 to 300 mg/kg body weight/day, even more preferably 0.5 to 100 mg/kg body weight/day, particularly preferably 1 to 50 mg/kg body weight/day, and most preferably 10 to 30 mg/kg body weight/day. The content of hydroxycitric acid and/or its salt can be the amount that results in the above intake or dosage. The oral intake or dosage per day for an adult may be divided into 1 to 6 capsules, 1 to 4 capsules, 1 to 3 capsules, or 1 to 2 capsules depending on the dosage form, for example, in the case of capsules.

The daily oral intake or administration amount of Garcinia and/or its extract for an adult can be appropriately determined depending on the individual's condition, body weight, sex, age, activity of the material, intake or administration route, intake or administration schedule, formulation form, or other factors. The daily oral intake or administration amount of Garcinia and/or its extract for an adult is, for example, preferably 0.02 mg/kg body weight/day or more, more preferably 0.2 mg/kg body weight/day or more, even more preferably 1.0 mg/kg body weight/day or more, particularly preferably 2 mg/kg body weight/day or more, and most preferably 20 mg/kg body weight/day or more. The daily oral intake or administration amount of Garcinia and/or its extract for an adult is, for example, preferably 800 mg/kg body weight/day or less, more preferably 600 mg/kg body weight/day or less, even more preferably 200 mg/kg body weight/day or less, particularly preferably 100 mg/kg body weight/day or less, and most preferably 60 mg/kg body weight/day or less. The daily oral intake or administration amount of Garcinia cambogia and/or its extract for an adult is, for example, preferably 0.02 to 800 mg/kg body weight/day, more preferably 0.2 to 600 mg/kg body weight/day, even more preferably 1.0 to 200 mg/kg body weight/day, particularly preferably 2 to 100 mg/kg body weight/day, and most preferably 20 to 60 mg/kg body weight/day. The content of Garcinia cambogia and/or its extract can be the amount that results in the above intake or administration amount. The daily oral intake or administration amount for an adult may be divided according to the dosage form, for example, in the case of capsules, into 1 to 6 capsules, 1 to 4 capsules, 1 to 3 capsules, or 1 to 2 capsules.

The daily oral intake or dosage of lactoferrin for adults can be determined appropriately depending on the individual's condition, body weight, sex, age, activity of the material, intake or administration route, intake or administration schedule, formulation form, or other factors. The daily oral intake or dosage of lactoferrin for adults is, for example, preferably 0.01 mg/kg body weight/day or more, more preferably 0.1 mg/kg body weight/day or more, even more preferably 0.5 mg/kg body weight/day or more, particularly preferably 1 mg/kg body weight/day or more, and most preferably 2 mg/kg body weight/day or more. The daily oral intake or dosage of lactoferrin for adults is, for example, preferably 200 mg/kg body weight/day or less, more preferably 150 mg/kg body weight/day or less, even more preferably 100 mg/kg body weight/day or less, particularly preferably 50 mg/kg body weight/day or less, and most preferably 20 mg/kg body weight/day or less. The oral intake or dosage of lactoferrin for an adult per day is, for example, preferably 0.01 to 200 mg/kg body weight/day, more preferably 0.1 to 150 mg/kg body weight/day, even more preferably 0.5 to 100 mg/kg body weight/day, particularly preferably 1 to 50 mg/kg body weight/day, and most preferably 2 to 20 mg/kg body weight/day. The lactoferrin content can be an amount that results in the above intake or dosage. The oral intake or dosage for an adult per day may be divided into 1 to 6 capsules, 1 to 4 capsules, 1 to 3 capsules, or 1 to 2 capsules depending on the dosage form, for example, in the case of capsules.

The daily oral intake or administration amount of carrot and/or its extract for an adult can be appropriately determined depending on the individual's condition, body weight, sex, age, activity of the material, intake or administration route, intake or administration schedule, formulation form, or other factors. The daily oral intake or administration amount of carrot and/or its extract for an adult is, for example, preferably 0.02 mg/kg body weight/day or more, more preferably 0.2 mg/kg body weight/day or more, even more preferably 1.0 mg/kg body weight/day or more, particularly preferably 2 mg/kg body weight/day or more, and most preferably 20 mg/kg body weight/day or more. The daily oral intake or administration amount of carrot and/or its extract for an adult is, for example, preferably 800 mg/kg body weight/day or less, more preferably 600 mg/kg body weight/day or less, even more preferably 200 mg/kg body weight/day or less, particularly preferably 100 mg/kg body weight/day or less, and most preferably 60 mg/kg body weight/day or less. The daily oral intake or administration amount of ginseng and/or its extract for an adult is, for example, preferably 0.02 to 800 mg/kg body weight/day, more preferably 0.2 to 600 mg/kg body weight/day, even more preferably 1.0 to 200 mg/kg body weight/day, particularly preferably 2 to 100 mg/kg body weight/day, and most preferably 20 to 60 mg/kg body weight/day. The content of ginseng and/or its extract can be an amount that results in the above intake or administration amount. The daily oral intake or administration amount for an adult may be divided according to the dosage form, for example, in the case of capsules, into 1 to 6 capsules, 1 to 4 capsules, 1 to 3 capsules, or 1 to 2 capsules.

The daily oral intake or dosage of pyridoxine and/or its salts for an adult can be appropriately determined depending on the individual's condition, body weight, sex, age, activity of the material, intake or administration route, intake or administration schedule, formulation form, or other factors. The daily oral intake or dosage of pyridoxine and/or its salts for an adult is, for example, preferably 0.0001 mg/kg body weight/day or more, more preferably 0.001 mg/kg body weight/day or more, even more preferably 0.005 mg/kg body weight/day or more, particularly preferably 0.01 mg/kg body weight/day or more, and most preferably 0.02 mg/kg body weight/day or more. The daily oral intake or dosage of pyridoxine and/or its salts for an adult is, for example, preferably 50 mg/kg body weight/day or less, more preferably 10 mg/kg body weight/day or less, even more preferably 5 mg/kg body weight/day or less, particularly preferably 1 mg/kg body weight/day or less, and most preferably 0.5 mg/kg body weight/day or less. The oral intake or dosage of pyridoxine and/or its salt per day for an adult is, for example, preferably 0.0001 to 50 mg/kg body weight/day, more preferably 0.001 to 10 mg/kg body weight/day, even more preferably 0.005 to 5 mg/kg body weight/day, particularly preferably 0.01 to 1 mg/kg body weight/day, and most preferably 0.02 to 0.5 mg/kg body weight/day. The content of pyridoxine and/or its salt can be the amount that results in the above intake or dosage. The oral intake or dosage per day for an adult may be divided into 1 to 6 capsules, 1 to 4 capsules, 1 to 3 capsules, or 1 to 2 capsules depending on the dosage form, for example, in the case of capsules.

The oral intake or dosage of oleanolic acid and/or its salts per day for an adult can be appropriately determined depending on the individual's condition, body weight, sex, age, activity of the material, intake or administration route, intake or administration schedule, formulation form, or other factors. The oral intake or dosage of oleanolic acid and/or its salts per day for an adult is, for example, preferably 0.01 mg/kg body weight/day or more, more preferably 0.1 mg/kg body weight/day or more, even more preferably 0.5 mg/kg body weight/day or more, particularly preferably 1 mg/kg body weight/day or more, and most preferably 10 mg/kg body weight/day or more. The oral intake or dosage of oleanolic acid and/or its salts per day for an adult is, for example, preferably 2000 mg/kg body weight/day or less, more preferably 1500 mg/kg body weight/day or less, even more preferably 1000 mg/kg body weight/day or less, particularly preferably 800 mg/kg body weight/day or less, and most preferably 500 mg/kg body weight/day or less. The oral intake or dosage of oleanolic acid and/or its salt per day for an adult is, for example, preferably 0.01 to 2000 mg/kg body weight/day, more preferably 0.1 to 1500 mg/kg body weight/day, even more preferably 0.5 to 1000 mg/kg body weight/day, particularly preferably 1 to 800 mg/kg body weight/day, and most preferably 10 to 500 mg/kg body weight/day. The content of oleanolic acid and/or its salt can be the amount that results in the above intake or dosage. The oral intake or dosage per day for an adult may be divided into 1 to 6 capsules, 1 to 4 capsules, 1 to 3 capsules, or 1 to 2 capsules depending on the dosage form, for example, in the case of a capsule.

The daily oral intake or dosage of thiamine for an adult can be determined appropriately depending on the individual's condition, body weight, sex, age, activity of the material, intake or administration route, intake or administration schedule, formulation form, or other factors. The daily oral intake or dosage of thiamine for an adult is, for example, preferably 0.0001 mg/kg body weight/day or more, more preferably 0.001 mg/kg body weight/day or more, and even more preferably 0.005 mg/kg body weight/day or more. The daily oral intake or dosage of thiamine for an adult is, for example, preferably 10 mg/kg body weight/day or less, more preferably 1 mg/kg body weight/day or less, and even more preferably 0.1 mg/kg body weight/day or less. The daily oral intake or dosage of thiamine for an adult is, for example, preferably 0.0001 to 10 mg/kg body weight/day, more preferably 0.001 to 1 mg/kg body weight/day, and even more preferably 0.005 to 0.1 mg/kg body weight/day. The amount of thiamine contained may be the amount that results in the above-mentioned intake or dosage. The oral intake or dosage for an adult per day may be divided into 1 to 6 capsules, 1 to 4 capsules, 1 to 3 capsules, or 1 to 2 capsules depending on the dosage form.

The daily oral intake or dosage of niacin for an adult can be determined appropriately depending on the individual's condition, body weight, sex, age, activity of the material, intake or administration route, intake or administration schedule, formulation form, or other factors. The daily oral intake or dosage of niacin for an adult is, for example, preferably 0.01 mg/kg body weight/day or more, more preferably 0.1 mg/kg body weight/day or more, and even more preferably 0.5 mg/kg body weight/day or more, as nicotinic acid. The daily oral intake or dosage of niacin for an adult is, for example, preferably 100 mg/kg body weight/day or less, more preferably 50 mg/kg body weight/day or less, and even more preferably 10 mg/kg body weight/day or less, as nicotinic acid. The oral intake or dosage of niacin per day for an adult is, for example, preferably 0.01 to 100 mg/kg body weight/day, more preferably 0.1 to 50 mg/kg body weight/day, and even more preferably 0.5 to 10 mg/kg body weight/day. The content of niacin can be an amount that results in the above intake or dosage. The oral intake or dosage per day for an adult may be divided into 1 to 6 capsules, 1 to 4 capsules, 1 to 3 capsules, or 1 to 2 capsules depending on the dosage form, for example, in the case of capsules.

The daily oral intake or dosage of inositol for an adult can be determined appropriately depending on the individual's condition, body weight, sex, age, activity of the material, intake or administration route, intake or administration schedule, formulation form, or other factors. The daily oral intake or dosage of inositol for an adult is, for example, preferably 0.01 mg/kg body weight/day or more, more preferably 0.1 mg/kg body weight/day or more, and even more preferably 0.5 mg/kg body weight/day or more. The daily oral intake or dosage of inositol for an adult is, for example, preferably 200 mg/kg body weight/day or less, more preferably 150 mg/kg body weight/day or less, and even more preferably 100 mg/kg body weight/day or less. The daily oral intake or dosage of inositol for an adult is, for example, preferably 0.01 to 200 mg/kg body weight/day, more preferably 0.1 to 150 mg/kg body weight/day, even more preferably 0.5 to 100 mg/kg body weight/day, particularly preferably 1 to 50 mg/kg body weight/day, and most preferably 2 to 20 mg/kg body weight/day. The inositol content can be an amount that results in the above intake or dosage. The daily oral intake or dosage for an adult may be divided into 1 to 6 capsules, 1 to 4 capsules, 1 to 3 capsules, or 1 to 2 capsules depending on the dosage form, for example, in the case of a capsule.

The daily oral intake or dosage of phytic acid for an adult can be determined appropriately depending on the individual's condition, body weight, sex, age, activity of the material, intake or administration route, intake or administration schedule, formulation form, or other factors. The daily oral intake or dosage of phytic acid for an adult is, for example, preferably 0.01 mg/kg body weight/day or more, more preferably 0.1 mg/kg body weight/day or more, and even more preferably 1 mg/kg body weight/day or more. The daily oral intake or dosage of phytic acid for an adult is, for example, preferably 200 mg/kg body weight/day or less, more preferably 150 mg/kg body weight/day or less, and even more preferably 100 mg/kg body weight/day or less. The oral intake or dosage of phytic acid per day for an adult is, for example, preferably 0.01 to 200 mg/kg body weight/day, more preferably 0.1 to 150 mg/kg body weight/day, even more preferably 0.5 to 100 mg/kg body weight/day, particularly preferably 1 to 50 mg/kg body weight/day, and most preferably 2 to 20 mg/kg body weight/day. The content of phytic acid can be an amount that results in the above intake or dosage. The oral intake or dosage per day for an adult may be divided into 1 to 6 capsules, 1 to 4 capsules, 1 to 3 capsules, or 1 to 2 capsules depending on the dosage form, for example, in the case of a capsule.

The daily oral intake or dosage of pantothenic acid for an adult can be appropriately determined depending on the individual's condition, body weight, sex, age, activity of the material, intake or administration route, intake or administration schedule, formulation form, or other factors. The daily oral intake or dosage of pantothenic acid for an adult is, for example, preferably 0.001 mg/kg body weight/day or more, more preferably 0.005 mg/kg body weight/day or more, and even more preferably 0.01 mg/kg body weight/day or more. The daily oral intake or dosage of pantothenic acid for an adult is, for example, preferably 100 mg/kg body weight/day or less, more preferably 10 mg/kg body weight/day or less, and even more preferably 1 mg/kg body weight/day or less. The daily oral intake or dosage of pantothenic acid for an adult is, for example, preferably 0.001 to 100 mg/kg body weight/day, more preferably 0.005 to 10 mg/kg body weight/day, and even more preferably 0.01 to 1 mg/kg body weight/day. The content of pantothenic acid can be an amount that results in the above-mentioned intake or dosage. The daily oral intake or dosage for an adult may be divided into 1 to 6 capsules, 1 to 4 capsules, 1 to 3 capsules, or 1 to 2 capsules depending on the dosage form, for example, in the case of a capsule.

The oral intake or dosage of biotin per day for an adult can be appropriately determined depending on the individual's condition, body weight, sex, age, activity of the material, intake or administration route, intake or administration schedule, formulation form, or other factors. The oral intake or dosage of biotin per day for an adult is, for example, preferably 0.01 mg/kg body weight/day or more, more preferably 0.1 mg/kg body weight/day or more, and even more preferably 0.5 mg/kg body weight/day or more. The oral intake or dosage of biotin per day for an adult is, for example, preferably 100 mg/kg body weight/day or less, more preferably 50 mg/kg body weight/day or less, and even more preferably 10 mg/kg body weight/day or less. The oral intake or dosage of biotin per day for an adult is, for example, preferably 0.01 to 100 mg/kg body weight/day, more preferably 0.1 to 50 mg/kg body weight/day, and even more preferably 0.5 to 10 mg/kg body weight/day. The amount of biotin contained can be the amount that results in the above-mentioned intake or administration amount. Note that the oral intake or administration amount per day for an adult may be divided into 1 to 6 capsules, 1 to 4 capsules, 1 to 3 capsules, or 1 to 2 capsules depending on the dosage form, for example, in the case of capsules.

The daily oral intake or dosage of folic acid for adults can be determined appropriately depending on the individual's condition, body weight, sex, age, activity of the material, intake or administration route, intake or administration schedule, formulation form, or other factors. The daily oral intake or dosage of folic acid for adults is, for example, preferably 0.01 μg/kg body weight/day or more, more preferably 0.1 μg/kg body weight/day or more, and even more preferably 0.5 μg/kg body weight/day or more. The daily oral intake or dosage of folic acid for adults is, for example, preferably 200 μg/kg body weight/day or less, more preferably 150 μg/kg body weight/day or less, and even more preferably 100 μg/kg body weight/day or less. The oral intake or dosage of folic acid per day for an adult is, for example, preferably 0.01 to 200 mg/kg body weight/day, more preferably 0.1 to 150 μg/kg body weight/day, and even more preferably 0.5 to 100 μg/kg body weight/day. The content of folic acid can be an amount that results in the above intake or dosage. The oral intake or dosage per day for an adult may be divided into 1 to 6 capsules, 1 to 4 capsules, 1 to 3 capsules, or 1 to 2 capsules depending on the dosage form, for example, in the case of a capsule.

The oral intake or administration amount of at least one selected from the group consisting of hydroxycitric acid and its salts, Garcinia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine, niacin, inositol, phytic acid, pantothenic acid, biotin, and folic acid can be appropriately determined depending on the individual's condition, body weight, sex, age, activity of the material, intake or administration route, intake or administration schedule, formulation form, or other factors. The oral intake or administration amount of the above-mentioned components per time is, for example, preferably 0.01 mg or more, more preferably 0.1 mg or more, even more preferably 0.5 mg or more, particularly preferably 1 mg or more, and most preferably 10 mg or more. The oral intake or administration amount of the above-mentioned components per time is, for example, preferably 500 mg or less, more preferably 400 mg or less, even more preferably 300 mg or less, particularly preferably 200 mg or less, and most preferably 100 mg or less. The oral intake or administration amount of the above-mentioned component per time is, for example, preferably 0.01 to 500 mg, more preferably 0.1 to 400 mg, even more preferably 0.5 to 300 mg, particularly preferably 1 to 200 mg, and most preferably 10 to 100 mg.

The food composition for preventing and/or improving posterior ocular abnormalities, or the agent for preventing and/or treating posterior ocular diseases of the present invention can be ingested or administered once or several times a day, typically 1 to 6 times a day, 1 to 3 times a day, 1 to 2 times a day, or at any period and interval.

The present invention may also have the following aspects.
A composition for suppressing HIF, comprising at least one member selected from the group consisting of hydroxycitric acid and its salts, Garcinia cambogia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine and its salts, niacin, inositol, phytic acid and its salts, pantothenic acid and its salts, biotin, folic acid, and rice bran;
A composition for retinal protection containing at least one selected from the group consisting of hydroxycitric acid and its salts, Garcinia cambogia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine and its salts, niacin, inositol, phytic acid and its salts, pantothenic acid and its salts, biotin, folic acid, and rice bran;
an anti-aging composition for the eye, comprising at least one selected from the group consisting of hydroxycitric acid and its salts, Garcinia cambogia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine and its salts, niacin, inositol, phytic acid and its salts, pantothenic acid and its salts, biotin, folic acid, and rice bran;
A composition for suppressing ocular VEGF, comprising at least one member selected from the group consisting of hydroxycitric acid and its salts, Garcinia cambogia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine and its salts, niacin, inositol, phytic acid and its salts, pantothenic acid and its salts, biotin, folic acid, and rice bran;
A composition for inhibiting ocular angiogenesis, comprising at least one member selected from the group consisting of hydroxycitric acid and its salts, Garcinia cambogia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine and its salts, niacin, inositol, phytic acid and its salts, pantothenic acid and its salts, biotin, folic acid, and rice bran;
A composition for maintaining homeostasis of ocular vascular tissue, comprising at least one member selected from the group consisting of hydroxycitric acid and its salts, Garcinia cambogia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine and its salts, niacin, inositol, phytic acid and its salts, pantothenic acid and its salts, biotin, folic acid, and rice bran;
A composition for reducing the risk of, preventing or treating a posterior eye disease, comprising at least one selected from the group consisting of hydroxycitric acid and its salts, Garcinia cambogia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine and its salts, niacin, inositol, phytic acid and its salts, pantothenic acid and its salts, biotin, folic acid and rice bran;
Use of a composition containing at least one selected from the group consisting of hydroxycitric acid and its salts, Garcinia cambogia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine and its salts, niacin, inositol, phytic acid and its salts, pantothenic acid and its salts, biotin, folic acid, and rice bran, for the manufacture of a HIF suppressant;
Use of a composition containing at least one selected from the group consisting of hydroxycitric acid and its salts, Garcinia cambogia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine and its salts, niacin, inositol, phytic acid and its salts, pantothenic acid and its salts, biotin, folic acid, and rice bran, for the manufacture of a retinal protective agent;
Use of a composition containing at least one selected from the group consisting of hydroxycitric acid and its salts, Garcinia cambogia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine and its salts, niacin, inositol, phytic acid and its salts, pantothenic acid and its salts, biotin, folic acid, and rice bran, for the manufacture of an anti-aging agent for the eye;
Use of a composition containing at least one selected from the group consisting of hydroxycitric acid and its salts, Garcinia cambogia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine and its salts, niacin, inositol, phytic acid and its salts, pantothenic acid and its salts, biotin, folic acid, and rice bran, for the manufacture of an ocular VEGF inhibitor;
Use of a composition containing at least one selected from the group consisting of hydroxycitric acid and its salts, Garcinia cambogia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine and its salts, niacin, inositol, phytic acid and its salts, pantothenic acid and its salts, biotin, folic acid, and rice bran, for the manufacture of an ocular angiogenesis inhibitor;
Use of a composition containing at least one selected from the group consisting of hydroxycitric acid and its salts, Garcinia cambogia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine and its salts, niacin, inositol, phytic acid and its salts, pantothenic acid and its salts, biotin, folic acid, and rice bran, for the manufacture of an agent for maintaining homeostasis of ocular vascular tissue;
Use of a composition containing at least one selected from the group consisting of hydroxycitric acid and its salts, Garcinia cambogia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine and its salts, niacin, inositol, phytic acid and its salts, pantothenic acid and its salts, biotin, folic acid, and rice bran, for the manufacture of an agent for reducing the risk of, preventing, or treating a posterior ocular disease;
A method for suppressing HIF, comprising having a human ingest a composition containing at least one member selected from the group consisting of hydroxycitric acid and its salts, Garcinia cambogia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine and its salts, niacin, inositol, phytic acid and its salts, pantothenic acid and its salts, biotin, folic acid, and rice bran;
A method for protecting the retina, comprising having a human ingest a composition containing at least one selected from the group consisting of hydroxycitric acid and its salts, Garcinia cambogia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine and its salts, niacin, inositol, phytic acid and its salts, pantothenic acid and its salts, biotin, folic acid, and rice bran;
an anti-aging method for the eye, comprising having a person ingest a composition containing at least one selected from the group consisting of hydroxycitric acid and its salts, Garcinia cambogia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine and its salts, niacin, inositol, phytic acid and its salts, pantothenic acid and its salts, biotin, folic acid, and rice bran;
A method for suppressing ocular VEGF, comprising having a human ingest a composition containing at least one member selected from the group consisting of hydroxycitric acid and its salts, Garcinia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine and its salts, niacin, inositol, phytic acid and its salts, pantothenic acid and its salts, biotin, folic acid, and rice bran;
A method for inhibiting ocular neovascularization, comprising having a human ingest a composition containing at least one member selected from the group consisting of hydroxycitric acid and its salts, Garcinia cambogia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine and its salts, niacin, inositol, phytic acid and its salts, pantothenic acid and its salts, biotin, folic acid, and rice bran;
A method for maintaining homeostasis of ocular vascular tissue, comprising having a human ingest a composition containing at least one selected from the group consisting of hydroxycitric acid and its salts, Garcinia cambogia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine and its salts, niacin, inositol, phytic acid and its salts, pantothenic acid and its salts, biotin, folic acid, and rice bran;
A method for reducing the risk of, preventing or treating a posterior ocular disease, comprising having a person ingest a composition containing at least one selected from the group consisting of hydroxycitric acid and its salts, Garcinia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine and its salts, niacin, inositol, phytic acid and its salts, pantothenic acid and its salts, biotin, folic acid, and rice bran;
the composition, use, or method, wherein the posterior ocular disease is at least one selected from the group consisting of age-related macular degeneration, diabetic retinopathy, diabetic macular edema, retinitis pigmentosa, proliferative vitreous retinopathy, retinal artery occlusion, retinal vein occlusion, uveitis, Leber's disease, retinopathy of prematurity, retinal detachment, retinal pigment epithelial detachment, central serous chorioretinopathy, central exudative chorioretinopathy, polypoidal choroidal vasculopathy, multifocal choroiditis, neovascular maculopathy, retinal aneurysm, retinal angiomatous proliferation, optic neuropathy caused by these diseases, optic neuropathy caused by glaucoma, and ischemic optic neuropathy;
the composition, use or method, wherein the salt of hydroxycitric acid is a calcium salt;
the composition, use or method, wherein the salt of pyridoxine is a hydrochloride or a phosphate;
the composition, use, or method, wherein the rice bran is defatted rice bran;
the composition, use, or method, wherein said niacin is niacinamide;
the composition, use, or method, wherein the salt of pantothenic acid is a calcium salt;
the composition, use, or method, wherein the total content of at least one selected from the group consisting of hydroxycitric acid and its salts, Garcinia and its extract, lactoferrin, carrot and its extract, pyridoxine and its salts, oleanolic acid and its salts, thiamine and its salts, niacin, inositol, phytic acid and its salts, pantothenic acid and its salts, biotin, folic acid, and rice bran is 0.1 to 95% by mass;
the composition, use, or method, wherein the daily oral intake of the hydroxycitric acid and its salts for an adult is 0.01 to 400 mg/kg body weight/day;
the composition, use or method, wherein the daily oral intake of the Garcinia cambogia and/or extract thereof per adult is 0.02 to 800 mg/kg body weight/day;
the composition, use or method, wherein the daily oral intake of the ginseng and/or its extract for an adult is 0.02 to 800 mg/kg body weight/day;
the composition, use, or method, wherein the daily oral intake of the pyridoxine and/or a salt thereof per adult is 0.0001 to 50 mg/kg body weight/day;
the composition, use, or method, wherein the daily oral intake of the oleanolic acid and/or a salt thereof for an adult is 0.01 to 2000 mg/kg body weight/day;
the composition, use, or method, wherein the daily oral intake of lactoferrin for an adult is 0.01 to 200 mg/kg body weight/day;
the composition, use, or method, wherein the daily oral intake of thiamine per adult is 0.0001 to 10 mg/kg body weight/day;
the composition, use, or method, wherein the daily oral intake of niacin for an adult is 0.01 to 100 mg/kg body weight/day;
the composition, use, or method, wherein the daily oral intake of the inositol per adult is 0.01 to 200 mg/kg body weight/day;
the composition, use, or method, wherein the daily oral intake of phytic acid and/or a salt thereof per adult is 0.01 to 200 mg/kg body weight/day;
the composition, use, or method, wherein the daily oral intake of the pantothenic acid and/or a salt thereof per adult is 0.001 to 100 mg/kg body weight/day;
the composition, use, or method, wherein the daily oral intake of biotin per adult is 0.01 to 100 mg/kg body weight/day;
the composition, use, or method, wherein the daily oral intake of folic acid for an adult is 0.01 to 200 mg/kg body weight/day;
the composition, use, or method, wherein the amount of the ingredient taken orally per dose is 0.01 to 500 mg;
The composition, use or method as described above, wherein the ingredient is taken 1 to 6 times a day;

Next, the present invention will be specifically explained using examples, but the present invention is not limited to the following examples. Furthermore, the amount of extract shown in the examples is the amount calculated as a dry solid content and including excipients, unless otherwise specified.

[Test Example 1: Screening of HIF-inhibiting substances 1]
A mouse retinal cone cell line (661W) was transfected with both HIF activity-dependent-Firefly-Luciferase and endogenous control CMV-Renilla-Luciferase using lentivirus to create a stable expression line RH661W. HIF was stabilized by adding cobalt chloride (CoCl ₂ ), a prolyl hydroxylase (PHD) inhibitor, to this RH661W, and a luminescence signal correlated with HIF activity was obtained by further adding luciferin. Various samples (candidate substances) were added to this RH661W, and the HIF inhibitory effect of the candidate substances on cobalt chloride-induced HIF activity was confirmed.

Therefore, RH661W was seeded at 0.8 x 10 ⁴ cells/well in a 96-well plate. After 24 hours, the cells were attached to the bottom of the plate, and then 200 μM CoCl ₂ was added to induce HIF activity. At the same time, candidate substances were added at 1 mg/ml. The candidate substances used were (-)-hydroxycitric acid calcium salt, Garcinia cambogia extract (Bioactives Japan Co., Ltd.), carrot extract (Matsuura Pharmaceutical Co., Ltd.), pyridoxine hydrochloride, 20-23% vitamin B ₆ , defatted rice bran (Oryza Oil & Fat Chemical Co., Ltd.), oleanolic acid, and lactoferrin.
After 24 hours, the luminescence intensity was measured using the Dual-Glo Luciferase assay system manufactured by Promega, and the HIF suppression effect was verified.

The activity of each was calculated as follows. The number of photons emitted by the cells was measured using a plate reader (BioTek, SYNERGYHTX), the ratio was calculated, and the average value of each candidate substance was calculated. The ratio of the addition of the candidate substance was divided by the ratio when only cobalt chloride was added to calculate the cell + _CoCl2 ratio. The results are shown in Table 1.

[Test Example 2: Screening of HIF-inhibiting substances 2]
A human retinal pigment epithelial cell line (ARPE) was transfected with both HIF activity-dependent-Firefly-Luciferase and endogenous control CMV-Renilla-Luciferase using lentivirus to create a stable expression line RHARPE. HIF was stabilized by adding cobalt chloride (CoCl ₂ ), a prolyl hydroxylase (PHD) inhibitor, to this RHARPE, and a luminescence signal correlated with HIF activity was obtained by further adding luciferin. Various samples (candidate substances) were added to this RHARPE, and the HIF inhibitory effect of the candidate substances on cobalt chloride-induced HIF activity was confirmed.

Therefore, RHARPE was seeded at 0.8 x 10 ⁴ cells/well in a 96-well plate. After 24 hours, the cells were attached to the bottom of the plate, and then 200 μM CoCl ₂ was added to induce HIF activity. At the same time, candidate substances were added at 1 mg/ml. The candidate substances used were carrot extract (Matsuura Pharmaceutical Co., Ltd.), Garcinia cambogia extract (Bioactives Japan Co., Ltd.), (-)-hydroxycitric acid calcium salt, lactoferrin, vitamin B ₁ , vitamin B ₆ , niacinamide (nicotinamide), inositol, phytic acid, D-calcium pantothenate, d-biotin, folic acid, defatted rice bran (Oryza Oil & Fat Chemical Co., Ltd.), and oleanolic acid.
After 24 hours, the luminescence intensity was measured using the Dual-Glo Luciferase assay system manufactured by Promega, and the HIF suppression effect was verified.

The activity of each was calculated as follows. The number of photons emitted by the cells was measured using a plate reader (BioTek, SYNERGYHTX), the ratio was calculated, and the average value of each candidate substance was calculated. The ratio of the addition of the candidate substance was divided by the ratio when only cobalt chloride was added to calculate the cell + _CoCl2 ratio. The results are shown in Table 2.

As shown in Tables 1 and 2, HIF inhibitory activity was observed for hydroxycitric acid and its salts, Garcinia and its extract, carrot and its extract, pyridoxine and its salts, oleanolic acid, lactoferrin, thiamine, niacin, inositol, phytic acid, pantothenic acid, biotin, defatted rice bran, and folic acid.

It is known that abnormally high levels of HIF are observed in retinal pigment epithelial cells (RPE cells) with age. In Test Example 1, retinal cone cells, which also play an important role as photoreceptor cells in the retina, were used to screen for HIF inhibitors, and in Test Example 2, retinal pigment epithelial cells were used to screen for HIF inhibitors. The above ingredients that were confirmed to suppress HIF are capable of inhibiting HIF in retinal cells, suggesting that they are effective for anti-aging of the eye.

It is also known that VEGF expression is regulated by HIF-1α and HIF-2α, which are types of HIF. Therefore, inhibiting HIF in retinal cells is effective in suppressing VEGF in the eye and suppressing angiogenesis induced by VEGF. In this way, it has been suggested that inhibiting HIF in retinal cells suppresses angiogenesis and is therefore also effective in maintaining the homeostasis of vascular tissue in the eye.

In addition, it is known that inhibiting HIF has a therapeutic effect on retinal ganglion cells and photoreceptor cell disorders in the posterior segment of the eye, and a protective effect on retinal ganglion cells and photoreceptor cells. Therefore, it has been suggested that the above-mentioned components are effective against posterior segment diseases and abnormalities, including glaucoma and age-related macular degeneration.

[Test Example 3: Electroretinogram (ERG) Test (Protection of Retinal Photoreceptor Cells by Inhibiting the Decrease in Retinal Action Potential)]
Six-week-old male BALB/cA mice were fed a control diet (MF, Oriental Yeast Co., Ltd.) for the control group, and a diet containing 3 g/kg/day of Garcinia cambogia extract (Bioactives Japan Co., Ltd.) mixed with the control diet for a total of 10 weeks for the Garcinia group. After dark adaptation from the previous day, the mice were exposed to 3000 Lux light for 1 hour on the 8th week of administration. ERG was measured 1 and 2 weeks after the exposure.
Eight-week-old BALB/cA mice (male) were intraperitoneally administered corn oil (SIGMA-ALDRICH corn oil 2303205, the same below) for the control group, and (-)-hydroxycitric acid suspended in corn oil at 300 mg/kg/day/1000 μL for the hydroxycitric acid group, 5 days/week for a total of two weeks. One week after administration, the mice were dark adapted from the previous day, and then irradiated with 3000 Lux light for one hour. One week after irradiation, ERG was measured.
For ERG, a Ganzfeld dome, measuring device, and LED stimulator (PuREC, MAYO Corporation, Japan) were used to record full-field flash ERG responses. After overnight dark adaptation, mice were anesthetized with MMB. The relevant electrode was recorded with a contact lens electrode, and the indifferent electrode was placed subcutaneously between the eyes. A clipping electrode to the tail was used as ground.
ERG responses were obtained from both eyes of each animal.
First, scotopic responses to stimulation (10 cd s/m ² ) under dark adaptation were recorded.
Next, the photopic response was recorded. The photopic response was recorded by stimulating the stimulus (20 cd/m ² ) under a white background (30 cd/m ² ).
All mice were kept warm during the procedure using a heat pad.
The a-wave of the ERG was fitted to the following equation:
Based on the Lamb and Pugh model, responses (R) were calculated as a function of time (T) and flash intensity (I). Rmax is the maximum amplitude, S is the sensitivity, and Td is the delay before the onset of the a-wave, respectively. S and Rmax data were transformed to logarithmic values for data analysis.

[Formula 1]

R (I, T) = {1-exp [-I・S・(T−Td)2]・Rmax

After 30 Hz low-pass filtering, ERG b-waves were fitted to the Naka-Rushton equation: where R is the amplitude, Rmax is the maximum amplitude, I is the flash intensity, K is the light intensity at half-maximum amplitude, and n is a dimensionless compression constant.

[Formula 2]

R = In / In + Kn・Rmax

Figures 1 (A) to (D) show the results of ERG (rod system function) measurement by stimulation (10 cd s/m ² ) under dark-adapted conditions in the Garcinia administration group and the control administration group. Figures 2 (A) to (B) show the results of ERG (cone system function) measurement by stimulation (10 cd s/m ² ) under light-adapted conditions in the Garcinia administration group and the control administration group. In this specification, the number n in the figures of the electroretinogram (ERG) test results indicates the number of mice evaluated.

As shown in Figures 1 and 2, the decrease in ERG amplitude was suppressed in the Garcinia administration group compared to the control administration group at both the 1st and 2nd week of irradiation, and the suppressive effect of Garcinia administration on retinal light damage was confirmed. Therefore, the retinal protective effect of Garcinia in the retinal light damage model was confirmed.

Figures 3(A) and (B) show the results of ERG (rod system function) measurement by stimulation (10 cd s/ ^m2 ) under dark-adapted conditions in the hydroxycitric acid administration group and the corn oil (control) administration group. Figure 4 shows the results of ERG (cone system function) measurement by stimulation (10 cd s/ ^m2 ) under light-adapted conditions in the hydroxycitric acid administration group and the corn oil (control) administration group.

As shown in Figures 3 and 4, the decrease in ERG amplitude was suppressed in the hydroxycitric acid group compared to the corn oil group at 1 week after irradiation, and the suppressive effect of hydroxycitric acid on retinal light damage was confirmed. Therefore, the retinal protective effect of hydroxycitric acid in the retinal light damage model was confirmed.
Hydroxycitric acid is an active ingredient contained in large amounts in Garcinia, so it was found that the results of Garcinia administration in Figures 1 and 2 are consistent with the results of hydroxycitric acid administration in Figures 3 and 4.

[Test Example 4: Laser CNV test method (inhibition of neovascularization) 1]
The diet was orally administered to 4-week-old C57BL6/J mice (male) for a total of 49 and 63 days, respectively, and on the 42nd day after the start of administration, a choroidal neovascularization (CNV) model was created by laser irradiation at 200 mW, 0.1 sec, and 75 μm, and the CNV volume was quantified 7 days after irradiation. The control group was administered a control diet (MF, Oriental Yeast Co., Ltd.), and the Garcinia administration group was administered a diet containing Garcinia cambogia extract (Bioactives Japan Co., Ltd.) mixed with the control diet at 300 mg/kg/day.

Specifically, C57BL/6 mice were anesthetized with MMB, and 5 sites per eye were irradiated near the optic disc with an argon laser (spot size 75 μm, duration 0.1 sec, intensity 200 mW). Rupture of Bruch's membrane was confirmed by the generation of air bubbles during laser application. In this specification, the number n in the figures showing the results of the laser CNV test indicates the number of sites evaluated for laser irradiation.
Laser irradiation was performed on the 42nd and 56th days after the start of feeding, respectively. On the 7th day after laser irradiation, the mice were anesthetized with Somnopentyl (Kyoritsu Seiyaku Co., Ltd., Tokyo, Japan), and the eyeballs were enucleated. Only the retinal pigment epithelium was flat-mounted and fixed for 1 hour with 4% PFA (prepared by mixing 4% paraformaldehyde, 16% PFA (ELECTRON MICROSCOPY SCIENCE), 20x PBS, and MQ in a ratio of 5:1:14), washed three times with PBS, and stained with isolectin (Isolectin GS-IB ₄ From Griffonia simplicifolia, Alexa Fluor ^TM 594 Conjugate: diluted with PBS = 1:200) at 4°C for 2 nights. Then, the sections were washed three times with PBS and mounted with Fluoromount-G (SouthernBiotech). The sections were observed under a Zweiss LSM 710 confocal microscope, and the volume of CNV was quantified using Imaris (Bitplane).

5A is a graph showing the results of choroidal neovascularization (CNV) induction after laser irradiation on the 42nd day (6 weeks) after the start of administration. FIG. 5B is a graph showing the results of choroidal neovascularization (CNV) induction after laser irradiation on the 56th day (8 weeks) after the start of administration.
As shown in Figures 5 (A) and (B), even when choroidal neovascularization (CNV) was induced by laser irradiation, the Garcinia administration group showed an inhibitory effect on the induction of CNV. As predicted in Test Examples 1 and 2, it was suggested that if a component that has been confirmed to have HIF inhibitory activity in retinal cells is used, it would have an inhibitory effect on the induction of CNV when choroidal neovascularization (CNV) is induced by laser irradiation.

[Test Example 5: Laser CNV test method (inhibition of neovascularization) 2]
Six-week-old C57BL6/J mice (male) were intraperitoneally administered corn oil (control group) and (-)-hydroxycitric acid suspended in corn oil (30 mg/kg/day/1000 μl) (-)-hydroxycitric acid (hydroxycitric acid) (hydroxycitric acid group) for 2 weeks, 5 days/week. Choroidal neovascularization (CNV) models were created by irradiating the mice with a laser at 200 mW, 0.1 sec, and 75 μm one week after administration, and the CNV volume was quantified 7 days after irradiation.

Specifically, C57BL/6 mice were anesthetized with MMB, and 5 sites per eye were irradiated near the optic disc with an argon laser (spot size 75 μm, duration 0.1 sec, intensity 200 mW). Rupture of Bruch's membrane was confirmed by the generation of air bubbles during laser irradiation.
Laser irradiation was performed one week after the start of administration, and on the seventh day after laser irradiation, the mice were anesthetized with Somnopentyl (Kyoritsu Seiyaku Co., Ltd., Tokyo, Japan), and the eyeballs were enucleated. Only the retinal pigment epithelium was flat-mounted and fixed for 1 hour with 4% PFA (prepared by mixing 4% paraformaldehyde, 16% PFA (ELECTRON MICROSCOPY SCIENCE), 20x PBS, and MQ in a ratio of 5:1:14), washed three times with PBS, and stained with isolectin (Isolectin GS-IB4 from Griffonia simplicifolia, Alexa Fluor ^™ 594 Conjugate: diluted with PBS = 1:200) at 4°C for two nights. Then, the sections were washed three times with PBS and mounted with Fluoromount-G (SouthernBiotech). The sections were observed under a Zweiss LSM 710 confocal microscope, and the volume of CNV was quantified using Imaris (Bitplane).

FIG. 6 is a graph showing the results of choroidal neovascularization (CNV) induction after laser irradiation one week after the start of administration.
As shown in Figure 6, even when choroidal neovascularization (CNV) was induced by laser irradiation, the hydroxycitric acid administration group showed an inhibitory effect on CNV induction. Considering the results of Test Examples 1, 2, and 5, it was suggested that a component that has been confirmed to have HIF inhibitory activity in retinal cells would have an inhibitory effect on CNV induction when choroidal neovascularization (CNV) was induced by laser irradiation.

[Test Example 6: Laser CNV test method (inhibition of neovascularization) 3]
The diet was orally administered to 4-week-old C57BL6/J mice (male) for a total of 63 days, and on the 56th day after the start of administration, a choroidal neovascularization (CNV) model was created by irradiating the mice with a laser at 200 mW, 0.1 sec, and 75 μm, and the CNV volume was quantified 7 days after irradiation. The control group was administered a control diet (MF, Oriental Yeast Co., Ltd.), and the carrot extract group was administered a diet containing carrot extract (Matsuura Pharmaceutical Co., Ltd.) mixed with the control diet at 200 mg/kg/day.

Specifically, C57BL/6 mice were anesthetized with MMB, and 5 sites per eye were irradiated near the optic disc with an argon laser (spot size 75 μm, duration 0.1 sec, intensity 200 mW). Rupture of Bruch's membrane was confirmed by the generation of air bubbles during laser application. In this specification, the number n in the figures showing the results of the laser CNV test indicates the number of sites evaluated for laser irradiation.
Laser irradiation was performed on the 56th day after the start of feeding, and on the 7th day after laser irradiation, the mouse was anesthetized with Somnopentyl (Kyoritsu Seiyaku Co., Ltd., Tokyo, Japan), and the eyeball was enucleated. Only the retinal pigment epithelium was flat-mounted and fixed for 1 hour with 4% PFA (prepared by mixing 4% paraformaldehyde, 16% PFA (ELECTRON MICROSCOPY SCIENCE), 20x PBS, and MQ in a ratio of 5:1:14), washed three times with PBS, and stained with isolectin (Isolectin GS-IB4 from Griffonia simplicifolia, Alexa FluorTM 594 Conjugate: diluted with PBS = 1:200) at 4°C for 2 nights. Then, the sections were washed three times with PBS and mounted with Fluoromount-G (SouthernBiotech). The sections were observed under a Zweiss LSM 710 confocal microscope, and the volume of CNV was quantified using Imaris (Bitplane).

FIG. 7 is a graph showing the results of choroidal neovascularization (CNV) induction after laser irradiation 8 weeks after the start of administration.
As shown in Figure 7, even when choroidal neovascularization (CNV) was induced by laser irradiation, the ginseng extract administration group showed an inhibitory effect on CNV induction. Considering the results of Test Examples 1, 2, and 6, it was suggested that a component that has been confirmed to have HIF inhibitory activity in retinal cells would have an inhibitory effect on CNV induction when choroidal neovascularization (CNV) is induced by laser irradiation.

[Test Example 7: Laser CNV test method (inhibition of neovascularization) 4]
Three-week-old C57BL6/J mice (male) were administered PBS (control group) or lactoferrin suspended in PBS (1600 mg/kg/day) (6 days/week for a total of 5 weeks) via gastric tube. Choroidal neovascularization (CNV) models were created by irradiating the mouse with a laser at 200 mW, 0.1 sec, and 75 μm one week after administration, and the CNV volume was quantified one week after irradiation.

Specifically, C57BL/6 mice were anesthetized with MMB, and 5 sites per eye were irradiated near the optic disc with an argon laser (spot size 75 μm, duration 0.1 sec, intensity 200 mW). Rupture of Bruch's membrane was confirmed by the generation of air bubbles during laser irradiation.
Four weeks after the start of administration, the mouse was irradiated with a laser. One week after the laser irradiation, the mouse was anesthetized with Somnopentyl (Kyoritsu Seiyaku Co., Ltd., Tokyo, Japan), and the eyeball was removed. Only the retinal pigment epithelium was flat-mounted and fixed with 4% PFA (prepared by mixing 4% paraformaldehyde, 16% PFA (ELECTRON MICROSCOPY SCIENCE), 20x PBS, and MQ in a ratio of 5:1:14) for one hour. The mouse was then washed three times with PBS and stained with isolectin (Isolectin GS-IB4 from Griffonia simplicifolia, Alexa Fluor ^™ 594 Conjugate: diluted with PBS = 1:200) for two nights at 4°C. Then, the sections were washed three times with PBS and mounted with Fluoromount-G (SouthernBiotech). The sections were observed under a Zweiss LSM 710 confocal microscope, and the volume of CNV was quantified using Imaris (Bitplane).

FIG. 8 is a graph showing the results of choroidal neovascularization (CNV) induction after laser irradiation 4 weeks after the start of administration.
As shown in Figure 8, even when choroidal neovascularization (CNV) was induced by laser irradiation, the lactoferrin administration group showed an inhibitory effect on CNV induction. Considering the results of Test Examples 1, 2 and 7, it was suggested that if a component that has been confirmed to have HIF inhibitory activity in retinal cells has an inhibitory effect on CNV induction when choroidal neovascularization (CNV) is induced by laser irradiation.

[Test Example 8: Laser CNV test method (inhibition of neovascularization) 5]
The food was orally administered to 3-week-old C57BL6/J mice (male) for a total of 63 days. On the 56th day after the start of administration, a choroidal neovascularization (CNV) model was created by laser irradiation at 200 mW, 0.1 sec, and 75 μm, and the CNV volume was quantified 7 days after irradiation. The control group was fed a diet consisting of vitamin _B6- deficient AIN-93G (Oriental Yeast Co., Ltd.) mixed with vitamin _B6 at 0.17 mg/kg/day, the vitamin _B6- administered groups were fed a diet consisting of vitamin _B6 -deficient AIN-93G mixed with vitamin _B6 at 1.53 mg/kg/day or 5.96 mg/kg/day, and the rice bran-administered groups were fed a diet consisting of vitamin _B6 -deficient AIN-93G mixed with vitamin _B6 at 0.17 mg/kg/day and defatted rice bran (Oryza Oil & Fat Chemical Co., Ltd.) at 100 mg/kg/day.

Specifically, C57BL/6 mice were anesthetized with MMB, and 5 sites per eye were irradiated near the optic disc with an argon laser (spot size 75 μm, duration 0.1 sec, intensity 200 mW). Rupture of Bruch's membrane was confirmed by the generation of air bubbles during laser application. In this specification, the number n in the figures showing the results of the laser CNV test indicates the number of sites evaluated for laser irradiation.
The mice were irradiated with a laser on the 56th day after the start of feeding. On the 7th day after laser irradiation, the mice were anesthetized with Somnopentyl (Kyoritsu Seiyaku Co., Ltd., Tokyo, Japan), and the eyeballs were enucleated. Only the retinal pigment epithelium was flat-mounted and fixed with 4% PFA (prepared by mixing 4% paraformaldehyde, 16% PFA (ELECTRON MICROSCOPY SCIENCE), 20x PBS, and MQ at a ratio of 5:1:14) for 1 hour. The mice were then washed three times with PBS and stained with isolectin (Isolectin GS-IB4 from Griffonia simplicifolia, Alexa FluorTM 594 Conjugate: diluted with PBS = 1:200) at 4°C for 2 nights. Then, the sections were washed three times with PBS and mounted with Fluoromount-G (SouthernBiotech). The sections were observed under a Zweiss LSM 710 confocal microscope, and the volume of CNV was quantified using Imaris (Bitplane).

FIG. 9 is a graph showing the results of choroidal neovascularization (CNV) induction after laser irradiation 8 weeks after the start of administration.
As shown in Figure 9, even when choroidal neovascularization (CNV) was induced by laser irradiation, the vitamin _B6 administration group and the rice bran administration group were found to have an inhibitory effect on the induction of CNV. Considering the results of Test Examples 1, 2, and 8, it was suggested that if a component that has been confirmed to have HIF inhibitory activity in retinal cells is used, it will have an inhibitory effect on the induction of CNV when choroidal neovascularization (CNV) is induced by laser irradiation.

[Test Example 9: Test to confirm expression of HIF downstream genes]
Human retinal pigment epithelial cell line (ARPE) was seeded at 1.0 x 10 ⁶ cells/well in a 12-well plate. After 24 hours, the cells were attached to the bottom of the plate, and the medium was replaced and 200 μM CoCl ₂ was added to induce HIF activity. At the same time, Garcinia cambogia extract (Bioactives Japan Co., Ltd.) and (-)-hydroxycitric acid calcium salt were added at 1 mg/ml.

Eight hours after addition, RNA was extracted and gene expression was confirmed by RT-PCR (ReverTra Ace qPCR RT Master Mix (TOYOBO)) and real-time PCR (THUNDERBIRD SYBO qPCR Mix (TOYOBO)). The results for Garcinia cambogia extract (Bioactives Japan Co., Ltd.) and (-)-hydroxycitric acid calcium salt are shown in Figures 10 and 11, respectively.

As shown in Figures 10 and 11, the expression of HIF downstream genes VEGF, GLUT1, PDK1, and BNIP3, which were increased by _CoCl2 , was significantly suppressed by the addition of Garcinia cambogia extract and (-)-hydroxycitric acid calcium salt.

In this specification, the MMB anesthetic was prepared by mixing three drugs: midazolam (4 mg/kg, SANDOZ, Yamagata, Japan), medetomidine (0.75 mg/kg, Fukushima, Japan), and butorphanol tartrate (5 mg/kg, Meiji Seika Pharmaceutical Co., Tokyo, Japan). Each drug was contained in the MMB anesthetic in a ratio of 1:1.07:1.33.

All procedures herein have been approved by the Ethics Committee for Animal Research at Keio University School of Medicine, which adheres to the ARVO Statement for the Use of Animals in Ophthalmic and Vision Research, the Keio University Institutional Guidelines for Animal Experimentation, and the Guidelines for the Use of Animals in Research (ARRIVE). C57BL/6J mice and BALB/cA mice were obtained from CLEA Japan, Inc.

Claims (8)

A composition for suppressing HIF, comprising at least one of hydroxycitric acid and its salts, Garcinia cambogia containing hydroxycitric acid and/or its salts, and/or an extract thereof. A retinal protection composition containing at least one of hydroxycitric acid and its salts, Garcinia cambogia containing hydroxycitric acid and/or its salts, and/or an extract thereof. A composition for suppressing high levels of HIF in retinal pigment epithelial cells, comprising at least one of hydroxycitric acid and its salts, Garcinia cambogia containing hydroxycitric acid and/or its salts, and/or an extract thereof. A composition for suppressing VEGF in the eye, comprising at least one of hydroxycitric acid and its salts, Garcinia cambogia containing hydroxycitric acid and/or its salts, and/or an extract thereof. A composition for inhibiting ocular angiogenesis, comprising at least one of hydroxycitric acid and its salts, Garcinia cambogia containing hydroxycitric acid and/or its salts, and/or an extract thereof. A composition for maintaining homeostasis of vascular tissue by suppressing VEGF in the eye and angiogenesis induced by VEGF, which contains at least one of hydroxycitric acid and its salts, Garcinia cambogia containing hydroxycitric acid and/or its salts, and/or an extract thereof. A composition for reducing the risk of, preventing, or treating a posterior ocular disease, comprising at least one of hydroxycitric acid and its salts, and Garcinia cambogia and/or an extract thereof containing hydroxycitric acid and/or its salts (with the exception of cases where the posterior ocular disease is a diabetic complication) . The composition according to claim 7, wherein the posterior ocular disease is at least one selected from the group consisting of age-related macular degeneration, retinitis pigmentosa, proliferative vitreoretinopathy, retinal artery occlusion, retinal vein occlusion, uveitis, Leber's disease, retinopathy of prematurity, retinal detachment, retinal pigment epithelial detachment, central serous chorioretinopathy, central exudative chorioretinopathy, polypoidal choroidal vasculopathy, multifocal choroiditis, neovascular maculopathy, retinal aneurysm, retinal angiomatous proliferation, optic neuropathy caused by these diseases, optic neuropathy caused by glaucoma, and ischemic optic neuropathy.

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