JP2020023465A - Elastic fiber formation promoter and elastic fiber-associated factor production promoter - Google Patents

Abstract

To provide a preparation that promotes the formation of an elastic fiber.SOLUTION: The present invention provides an elastic fiber formation promoter and an elastic fiber-associated factor production promoter that contain at least one selected from the group consisting of cyanocobalamine, and aspartic acid and a salt thereof.SELECTED DRAWING: None

Description

  The present invention relates to an elastic fiber formation promoter and an elastic fiber-related factor production promoter.

  It is said that the conjunctiva sags with age, and almost all people in their 60s and over develop conjunctival relaxation. When the conjunctiva relaxes, the flow path of the tears and the disturbed distribution of the tears occur, causing indefinite complaints such as sore eyes, dry eyes (dry eyes), discomfort, and foreign bodies. In recent years, there is a concern that the age of onset of conjunctival relaxation may be reduced due to the spread of wearing of contact lenses.

  On the other hand, elastic fibers (elastin fibers) are fibers formed by the interaction of elastic fiber-related factors produced by fibroblasts, but in those with conjunctival relaxation, the elastic fibers of the conjunctiva are ruptured and aggregated. In addition to this, it is known that even in healthy adults, the elastic fibers decrease in the age of 50 or older (for example, Non-Patent Document 1).

CORNEA, 23 (3), p294-298, 2004

  When the symptoms of conjunctival flaccidity are strong, it is the reality that the symptoms are alleviated by a surgical technique of removing extra conjunctiva. Under these circumstances, if a preparation that promotes the formation of elastic fibers of the conjunctiva could be provided, it would be easier to improve the conjunctival relaxation without depending on the surgical technique, and to reduce various symptoms caused by conjunctival relaxation. It is expected that it can be relaxed and look beautiful. An object of the present invention is to provide a preparation capable of promoting the formation of elastic fibers.

  The present inventors have conducted intensive studies to solve the above problems, and as a result, have found that cyanocobalamin and aspartic acid or a salt thereof increase gene expression of an elastic fiber-related factor and promote formation of elastic fibers. The present invention is based on this finding, and provides the following inventions.

[1]
An elastic fiber formation promoter comprising at least one selected from the group consisting of cyanocobalamin, and aspartic acid and salts thereof.
[2]
The elastic fiber formation promoter according to [1], further comprising citric acid or a salt thereof.
[3]
The elastic fiber formation promoter according to [1] or [2], which is used for promoting formation of elastic fibers of the conjunctiva.
[4]
An elastic fiber-related factor production promoter comprising at least one selected from the group consisting of cyanocobalamin and aspartic acid and salts thereof.
[5]
The elastic fiber-related factor production promoter according to [4], further comprising citric acid or a salt thereof.
[6]
The elastic fiber-related factor production promoter according to [4] or [5], which is used for promoting production of an elastic fiber-related factor of the conjunctiva.
[7]
The elastic fiber-related factor production promoter according to any one of [4] to [6], wherein the elastic fiber-related factor is Fibulin-5.
[8]
An ophthalmic composition comprising cyanocobalamin, aspartic acid or a salt thereof, citric acid or a salt thereof, and a thickener.
[9]
The ophthalmic composition according to [8], wherein the thickener is at least one selected from the group consisting of hydroxypropylmethylcellulose, hyaluronic acid and salts thereof.
[10]
An ophthalmic composition for improving pupil luminosity, comprising at least one selected from the group consisting of cyanocobalamin, and aspartic acid and salts thereof.

  According to the present invention, it is possible to provide a preparation that promotes formation of elastic fibers and a preparation that promotes production of elastic fiber-related factors.

1 is a graph showing the expression level of Fibulin-5 gene when treated with a medium containing each component in Test Example 1. 10 is a photograph of a localization of an elastic fiber-related factor (Fibulin-5) taken by a fluorescence microscope when treated with a medium containing each component in Test Example 2. 4 is a photograph taken by a fluorescence microscope of the formation of elastic fibers (elastin fibers) when treated with a medium containing each component in Test Example 2.

  Hereinafter, embodiments for carrying out the present invention will be described in detail. However, the present invention is not limited to the following embodiments.

  In this specification, the unit “%” of content means “w / v%” and has the same meaning as “g / 100 mL” unless otherwise specified.

[1. Elastic fiber formation promoter and elastic fiber-related factor production promoter)
The elastic fiber formation promoting agent and the elastic fiber-related factor production promoting agent (also simply referred to as “the agent according to the present embodiment”) according to the present embodiment are cyanocobalamin (also simply referred to as “(A) component”), And at least one member selected from the group consisting of aspartic acid and a salt thereof (also simply referred to as “component (B)”).

  As used herein, the term “elastic fiber” refers to microfibrils (Fiburillin 1, Fiburillin 2), tropoelastin, Fibulin family molecules (for example, Fibulin-5), LTBP family molecules (for example, produced from fibroblasts). LTBP-4), a fiber having elasticity (characteristics of returning to its original state even when stretched) formed by interaction of elastic fiber-related factors such as lysyl oxidase and the like, and has the same meaning as elastin fiber.

  As used herein, "elastic fiber-related factor" means a molecule involved in the formation of elastic fibers. The elastic fiber-related factor production promoter according to this embodiment has an action of promoting the production of the above-mentioned elastic fiber-related factor, and preferably has an action of promoting the production of Fibulin-5. Note that Fibulin-5 is an elastic fiber-related factor also referred to as “FBLN5” or “DANCE”.

  The elastic fiber formation promoting agent according to the present embodiment can be used to promote the formation of elastic fibers in any tissue in a living body, but is preferably used to promote the formation of elastic fibers in the conjunctiva. . Further, the elastic fiber-related factor production promoter according to the present embodiment can be used to promote the production of elastic fiber-related factors in any tissue in a living body. Used to promote production.

Cyanocobalamin is a known compound which is a kind of vitamin B _12.

  The content of the component (A) in the agent according to the present embodiment is not particularly limited, and is appropriately set according to the type and content of other compounding components, the form of preparation, and the like. As the content of the component (A), the total content of the component (A) is, for example, from 0.0002 to the total amount of the agent according to the present embodiment, from the viewpoint of more remarkably exerting the effects of the present invention. It is preferably 2 w / v%, more preferably 0.002 to 0.2 w / v%, even more preferably 0.003 to 0.02 w / v%.

  Aspartic acid is a compound known as an acidic amino acid also called 2-aminobutanedioic acid. Aspartic acid may be any of L-form, D-form and DL-form, but is preferably L-form.

  The salt of aspartic acid is not particularly limited as long as it is pharmaceutically, pharmacologically (pharmaceutically) or physiologically acceptable. Specifically, salts with inorganic bases (eg, ammonium salts; salts with metals such as alkali metals (sodium, potassium, etc.), alkaline earth metals (calcium, magnesium, etc.), aluminum, etc.) and organic bases Salts (eg, salts with organic amines such as methylamine, triethylamine, triethanolamine, morpholine, piperazine, pyrrolidine, tripyridine, picoline, etc.) and the like. Aspartic acid and salts thereof are preferably salts of aspartic acid with an inorganic base, more preferably alkali metal salts and alkaline earth metal salts of aspartic acid, and even more preferably potassium aspartate and magnesium potassium aspartate. Aspartic acid and its salts may be used alone or in combination of two or more.

  The content of the component (B) in the agent according to the present embodiment is not particularly limited, and is appropriately set in accordance with the type and content of the other compounding components, the formulation, and the like. As the content of the component (B), the total content of the component (B) is, for example, from 0.01 to more based on the total amount of the agent according to the present embodiment, from the viewpoint of more remarkably exerting the effects of the present invention. It is preferably 20 w / v%, more preferably 0.01 to 2.0 w / v%, even more preferably 0.2 to 2.0 w / v%.

  In the agent according to the present embodiment, the content ratio of the component (B) to the component (A) is not particularly limited, and may be appropriately determined according to the type of the component (B), the type and content of the other compounding components, the dosage form, and the like. Is set. The content ratio of the component (B) to the component (A) is, for example, based on 1 part by mass of the total content of the component (A) contained in the agent according to the present embodiment from the viewpoint of further enhancing the effects of the present invention. The total content of the component (B) is preferably from 0.005 to 100,000 parts by mass, more preferably from 0.05 to 1,000 parts by mass, and still more preferably from 10 to 670 parts by mass. .

  The agent according to the present embodiment may contain only one of the component (A) or the component (B), or may contain both the component (A) and the component (B). It is preferable to contain both component (B) and component (B).

  The agent according to the present embodiment may further contain at least one selected from the group consisting of citric acid and salts thereof (also simply referred to as “component (C)”). Thereby, the effect of the present invention is more remarkably exhibited.

  The salt of citric acid is not particularly limited as long as it is pharmaceutically, pharmacologically (pharmaceutically) or physiologically acceptable. Specifically, salts with inorganic bases (for example, ammonium salts; salts with metals such as alkali metals (sodium, potassium, etc.), alkaline earth metals (calcium, magnesium, etc.), aluminum, etc.) and organic bases Salts (eg, salts with organic amines such as methylamine, triethylamine, triethanolamine, morpholine, piperazine, pyrrolidine, tripyridine, picoline, etc.) and the like. As citric acid or a salt thereof, citric acid and a salt of citric acid with an inorganic base are preferred, citric acid and an alkali metal salt of citric acid are more preferred, and citric acid and sodium citrate are still more preferred. Citric acid and its salts may be hydrates or anhydrides. Citric acid and its salts may be used alone or in combination of two or more.

  The content of the component (C) in the agent according to the present embodiment is not particularly limited, and is appropriately set according to the type and content of other compounding components, the form of preparation, and the like. As the content of the component (C), from the viewpoint of more remarkably exhibiting the effect of the present invention, for example, the total content of the component (C) is 0.0002 to 0.002 to the total amount of the agent according to the present embodiment. It is preferably 5.0 w / v%, more preferably 0.001 to 5.0 w / v%, even more preferably 0.005 to 4.0 w / v%.

  In the agent according to the present embodiment, the content ratio of the component (C) to the component (A) is not particularly limited, and may be appropriately determined according to the type of the component (C), the type and content of the other compounding components, the dosage form, and the like. Is set. The content ratio of the component (C) to the component (A) is, for example, with respect to 1 part by mass of the total content of the component (A) contained in the agent according to the present embodiment, from the viewpoint of further enhancing the effects of the present invention. The total content of the component (C) is preferably 0.0001 to 25,000 parts by mass, more preferably 0.005 to 2500 parts by mass, and 0.25 to 1340 parts by mass. More preferred.

  In the agent according to the present embodiment, the content ratio of the component (C) to the component (B) is not particularly limited, and the types of the components (B) and (C), the types and contents of other components, and the formulation form It is set as appropriate according to the above. The content ratio of the component (C) to the component (B) is, for example, based on 1 part by mass of the total content of the component (B) contained in the agent according to the present embodiment from the viewpoint of further enhancing the effect of the present invention. The total content of the component (C) is preferably 0.00001 to 5000 parts by mass, more preferably 0.0005 to 500 parts by mass, and 0.0025 to 20 parts by mass. More preferred.

  The agent according to the present embodiment may contain only the above-mentioned component (A) and / or component (B), may further contain component (C) as needed, and may contain pharmaceutically as needed. , Pharmacologically (pharmaceutically) or physiologically acceptable additives.

  The administration route of the agent according to the present embodiment is not particularly limited. For example, enteral administration, intravenous administration, transarterial administration, intramuscular administration, subcutaneous administration, intradermal administration, intraperitoneal administration, topical administration to the eye (for example, eye drops) Administration, intravitreal administration, subconjunctival administration, etc.), parenteral administration such as transdermal administration, or oral administration. Among these, parenteral administration is preferred, topical ocular administration and transdermal administration are more preferred, and topical ocular administration is still more preferred, from the viewpoint that the effects of the present invention can be more reliably exerted.

  The agent according to the present embodiment can adopt a formulation suitable for an administration route. Pharmaceutical forms suitable for oral administration include, for example, tablets, capsules, granules, powders, and the like. Pharmaceutical forms suitable for parenteral administration include, for example, ophthalmic compositions (eye drops, eyewash, contact Lens compositions), skin external preparations (solutions, liniments, lotions, creams, ointments, aerosols), injections and the like. These dosage forms can be prepared using ordinary techniques commonly used in the art. Among these, from the viewpoint that the effects of the present invention can be more reliably exerted, an ophthalmic composition and a skin external preparation are preferred, and an ophthalmic composition is more preferred.

  Examples of additives include carriers, chelating agents, bases, pH adjusters, surfactants, fresheners, thickeners, buffers, stabilizers, preservatives, isotonic agents, sugar alcohols, Examples include oils, solvents, dispersants, emulsifiers, excipients, binders, disintegrants, lubricants, humectants, colorants, and fragrances. These additives can be appropriately selected according to the administration route, the formulation form, and the like.

  When the formulation of the agent according to the present embodiment is an ophthalmic composition, for example, a carrier (eg, an aqueous solvent such as water or hydrous ethanol), a chelating agent (eg, ethylenediaminediacetic acid (EDDA), ethylenediaminetriacetic acid, ethylenediamine) Tetraacetic acid (EDTA), N- (2-hydroxyethyl) ethylenediaminetriacetic acid (HEDTA), diethylenetriaminepentaacetic acid (DTPA), etc., bases (eg, octyldodecanol, titanium oxide, potassium bromide, plastibase, etc.), pH adjusters (eg, hydrochloric acid, acetic acid, sodium hydroxide, potassium hydroxide, calcium hydroxide, magnesium hydroxide, triethanolamine, diisopropanolamine, etc.), surfactants (eg, tyloxapol, polyoxyethylene sorbitan fatty acid ester) , Stearic acid Nonionic surfactants such as reoxyl, polyoxyethylene castor oil, polyoxyethylene hardened castor oil, and poloxamers; polyoxyethylene alkyl ether phosphate, polyoxyethylene alkyl ether sulfate, alkylbenzene sulfonate, alkyl sulfate , Anionic surfactants such as N-acyltaurin salts; zwitterionic surfactants such as lauryl dimethylaminoacetate betaine), fresheners (for example, menthol, menthol, camphor, borneol, geraniol, cineol, citronellol, carboxyl) , Anethole, eugenol, limonene, linalool, linalyl acetate, thymol, cymene, terpineol, pinene, camphene, isoborneol, fenchen, nerol, myrcene, myrcenol, linalool acetate, Banjurol, eucalyptus oil, bergamot oil, peppermint oil, cool mint oil, spearmint oil, peppermint oil, fennel oil, cauliflower oil, rose oil, camphor oil, etc.), thickeners (eg, methylcellulose, ethylcellulose, hydroxypropylmethylcellulose) Cellulose polymer compounds such as sodium, hydroxyethylcellulose, sodium carboxymethylcellulose; polyvinyl polymer compounds such as polyvinylpyrrolidone and polyvinyl alcohol; carboxyvinyl polymer; guar gum; hydroxypropyl guar gum; gum arabic; gum karaya; xanthan gum; Salts (sodium salts, etc.); heparin-like substances, heparin, heparin sulfate, heparan sulfate, heparinoids, hyaluronic acid and salts thereof (sodium salts, etc.) ), Mucopolysaccharides of chondroitin sulfate and its salts (such as sodium salts); starch; chitin and its derivatives; chitosan and its derivatives; carrageenan; monosaccharides such as glucose; and the like; buffers (for example, borate buffer, phosphorus Acid buffers, carbonate buffers, acetate buffers, lactate buffers, succinate buffers, Tris buffers, AMPD buffers, etc.), stabilizers (eg, sodium formaldehyde sulfoxylate (Rongalite), sodium bisulfite, Sodium pyrosulfite, aluminum monostearate, glyceryl monostearate, cyclodextrin, monoethanolamine, dibutylhydroxytoluene, etc., preservatives (eg, quaternary ammonium salts of alkyl polyaminoethyl glycines such as benzalkonium chloride, Benzethonium chloride) Chlorhexidine gluconate, polydronium chloride, zinc chloride, sodium benzoate, ethanol, chlorobutanol, sorbic acid, potassium sorbate, sodium dehydroacetate, methyl paraoxybenzoate, ethyl paraoxybenzoate, propyl paraoxybenzoate, propyl paraoxybenzoate, butyl paraoxybenzoate, Oxyquinoline sulfate, phenethyl alcohol, benzyl alcohol, biguanide compounds (specifically, polyhexanide hydrochloride (polyhexamethylene biguanide), alexidine, etc.), Gloquil (trade name, manufactured by Rhodia Co., Ltd.), and the like, tonicity agent (for example, sulfurous acid) Sodium hydrogen sulfite, sodium chloride, potassium chloride, calcium chloride, sodium chloride, magnesium chloride, potassium acetate, sodium acetate, sodium hydrogen carbonate, sodium carbonate, thio Sodium sulfate, magnesium sulfate, glycerin, propylene glycol, etc.), sugar alcohols (eg, xylitol, sorbitol, mannitol, etc.), oils (eg, sesame oil, castor oil, soybean oil, vegetable oils such as olive oil, animal oils such as squalane, Liquid paraffin, mineral oil such as petrolatum, etc.) can be used as an additive.

  The agent according to the present embodiment may be prepared according to a method generally used in the field of pharmaceuticals. For example, component (A), component (B), and if necessary, component (C) and other components Can be prepared by adding and mixing so that the desired content is obtained.

  The dosage and administration of the agent according to the present embodiment is not particularly limited as long as it has an effect and a dosage and dosage with a small side effect.For example, when the agent according to the present embodiment is instilled as an ophthalmic composition, and For adults (15 years and older) and children aged 7 years and older, apply 1 to 2 drops at a time, 4 times a day, use 1 to 2 drops, 1 to 3 drops, or 2 to 3 drops at a time. A method of instilling 5 to 6 times a day can be exemplified.

[2. Ophthalmic composition)
The ophthalmic composition according to the present embodiment is at least one selected from the group consisting of cyanocobalamin (also simply referred to as “component (A)”), aspartic acid and a salt thereof (also simply referred to as “component (B)”). At least one selected from the group consisting of citric acid and salts thereof (also simply referred to as “component (C)”), and a thickener (also simply referred to as “component (D)”). It contains.

Cyanocobalamin is a known compound which is a kind of vitamin B _12.

  The content of the component (A) in the ophthalmic composition according to the present embodiment is not particularly limited, and is appropriately set according to the type and content of other compounding components, the use of the ophthalmic composition, the formulation form, and the like. As the content of the component (A), for example, the total content of the component (A) is 0.0002 to 2 w / v based on the total amount of the ophthalmic composition from the viewpoint of more remarkably exerting the effects of the present invention. %, More preferably 0.002 to 0.2 w / v%, even more preferably 0.003 to 0.02 w / v%.

  Aspartic acid is a compound known as an acidic amino acid also called 2-aminobutanedioic acid. Aspartic acid may be any of L-form, D-form and DL-form, but is preferably L-form.

  The salt of aspartic acid is not particularly limited as long as it is pharmaceutically, pharmacologically (pharmaceutically) or physiologically acceptable. Specifically, salts with inorganic bases (for example, ammonium salts; salts with metals such as alkali metals (sodium, potassium, etc.), alkaline earth metals (calcium, magnesium, etc.), aluminum, etc.) and organic bases Salts (eg, salts with organic amines such as methylamine, triethylamine, triethanolamine, morpholine, piperazine, pyrrolidine, tripyridine, picoline, etc.) and the like. Aspartic acid and a salt thereof are preferably a salt of an aspartic acid with an inorganic base, more preferably an alkali metal salt and an alkaline earth metal salt of aspartic acid, and further preferably potassium aspartate and magnesium potassium potassium aspartate. Aspartic acid and salts thereof may be used alone or in a combination of two or more.

  The content of the component (B) in the ophthalmic composition according to the present embodiment is not particularly limited, and is appropriately set according to the type and content of other compounding components, the use of the ophthalmic composition, the formulation form, and the like. As the content of the component (B), the total content of the component (B) is, for example, 0.01 to 20 w / v based on the total amount of the ophthalmic composition from the viewpoint of more remarkably exerting the effects of the present invention. %, More preferably 0.01 to 2.0 w / v%, even more preferably 0.2 to 2.0 w / v%.

  In the ophthalmic composition according to the present embodiment, the content ratio of the component (B) to the component (A) is not particularly limited, and the type of the component (B), the type and content of other components, and the use of the ophthalmic composition It is appropriately set according to the formulation and the like. As the content ratio of the component (B) to the component (A), for example, from the viewpoint of further enhancing the effect of the present invention, for example, the total content of the component (A) contained in the ophthalmic composition according to the present embodiment is 1 part by mass. On the other hand, the total content of the component (B) is preferably 0.005 to 100000 parts by mass, more preferably 0.05 to 1000 parts by mass, and more preferably 10 to 670 parts by mass. More preferred.

  The salt of citric acid is not particularly limited as long as it is pharmaceutically, pharmacologically (pharmaceutically) or physiologically acceptable. Specifically, salts with inorganic bases (for example, ammonium salts; salts with metals such as alkali metals (sodium, potassium, etc.), alkaline earth metals (calcium, magnesium, etc.), aluminum, etc.) and organic bases Salts (eg, salts with organic amines such as methylamine, triethylamine, triethanolamine, morpholine, piperazine, pyrrolidine, tripyridine, picoline, etc.) and the like. As citric acid or a salt thereof, citric acid and a salt of citric acid with an inorganic base are preferred, citric acid and an alkali metal salt of citric acid are more preferred, and citric acid and sodium citrate are still more preferred. Citric acid and its salts may be hydrates or anhydrides. Citric acid and its salts may be used alone or in combination of two or more.

  The content of the component (C) in the ophthalmic composition according to the present embodiment is not particularly limited, and is appropriately set according to the type and content of the other compounding components, the use of the ophthalmic composition, the formulation form, and the like. As the content of the component (C), the total content of the component (C) is, for example, 0.0002 to 5.0 w, based on the total amount of the ophthalmic composition, from the viewpoint of more remarkably exerting the effects of the present invention. / W%, more preferably 0.001 to 5.0 w / v%, even more preferably 0.005 to 4.0 w / v%.

  In the ophthalmic composition according to the present embodiment, the content ratio of the component (C) to the component (A) is not particularly limited, and the type of the component (C), the type and content of other compounding components, and the use of the ophthalmic composition It is appropriately set according to the formulation and the like. As the content ratio of the component (C) to the component (A), for example, from the viewpoint of further enhancing the effect of the present invention, for example, the total content of the component (A) contained in the ophthalmic composition according to the present embodiment is 1 part by mass. The total content of the component (C) is preferably 0.0001 to 25000 parts by mass, more preferably 0.005 to 2500 parts by mass, and 0.25 to 1340 parts by mass. Is more preferred.

  In the ophthalmic composition according to the present embodiment, the content ratio of the component (C) to the component (B) is not particularly limited, and the types of the components (B) and (C), the types and the contents of other components, It is set appropriately according to the use of the ophthalmic composition, the formulation form, and the like. As the content ratio of the component (C) to the component (B), for example, from the viewpoint of further enhancing the effects of the present invention, for example, the total content of the component (B) contained in the ophthalmic composition according to the present embodiment is 1 part by mass. The total content of the component (C) is preferably 0.00001 to 5000 parts by mass, more preferably 0.0005 to 500 parts by mass, and 0.0025 to 20 parts by mass. Is more preferred.

  Examples of the thickener include cellulose polymer compounds (eg, methyl cellulose, ethyl cellulose, hydroxypropyl methyl cellulose, hydroxyethyl cellulose, sodium carboxymethyl cellulose, etc.), polyvinyl polymer compounds (polyvinyl pyrrolidone, polyvinyl alcohol, etc.), carboxyvinyl polymer Guar gum, hydroxypropyl guar gum, gum arabic, gum karaya, xanthan gum, agar, alginic acid and its salts (such as sodium salts), mucopolysaccharides (for example, heparin analogs, heparin, heparin sulfate, heparan sulfate, heparinoid, hyaluronic acid and its Salts (such as sodium salts), chondroitin sulfate and its salts (such as sodium salts)), starch, chitin and its derivatives, chitosan and its derivatives Carrageenan, monosaccharides (glucose, etc.) and the like.

  As the thickener, cellulose-based polymer compounds, polyvinyl-based polymer compounds, mucopolysaccharides are preferred, cellulose-based polymer compounds, mucopolysaccharides are more preferred, hydroxypropylmethylcellulose, hydroxyethylcellulose, chondroitin sulfate and salts thereof, Hyaluronic acid and its salts and glucose are more preferred, and hydroxypropyl methylcellulose and sodium hyaluronate are particularly preferred.

  As the thickener, those commercially available may be used. As the thickener, one type may be used alone, or two or more types may be used in combination.

  The content of the component (D) in the ophthalmic composition according to the present embodiment is not particularly limited, and is appropriately set according to the type and content of the other compounding components, the use of the ophthalmic composition, the formulation form, and the like. As the content of the component (D), the total content of the component (D) is, for example, 0.0001 to 5 w / v based on the total amount of the ophthalmic composition from the viewpoint of more remarkably exerting the effects of the present invention. %, More preferably 0.0005 to 3 w / v%, even more preferably 0.001 to 1.2 w / v%.

  In the ophthalmic composition according to the present embodiment, the content ratio of the component (D) to the component (A) is not particularly limited, and the type of the component (D), the type and content of other components, and the use of the ophthalmic composition It is appropriately set according to the formulation and the like. As the content ratio of the component (D) to the component (A), for example, from the viewpoint of further enhancing the effect of the present invention, for example, the total content of the component (A) contained in the ophthalmic composition according to the present embodiment is 1 part by mass. On the other hand, the total content of the component (D) is preferably 0.00005 to 25,000 parts by mass, more preferably 0.0025 to 1500 parts by mass, and 0.05 to 400 parts by mass. Is more preferred.

  In the ophthalmic composition according to the present embodiment, the content ratio of the component (D) to the component (B) is not particularly limited, and the types of the components (B) and (D), the types and the contents of other components, It is set appropriately according to the use of the ophthalmic composition, the formulation form, and the like. As the content ratio of the component (D) to the component (B), for example, from the viewpoint of further enhancing the effect of the present invention, for example, the total content of the component (B) contained in the ophthalmic composition according to the present embodiment is 1 part by mass. On the other hand, the total content of the component (D) is preferably 0.000005 to 5000 parts by mass, more preferably 0.00025 to 300 parts by mass, and 0.0005 to 6 parts by mass. Is more preferred.

(Buffer)
It is preferable that the ophthalmic composition according to the present embodiment further contains a buffering agent other than the component (C). When the ophthalmic composition further contains a buffer, the effect of the present invention is more remarkably exhibited. The buffer is not particularly limited as long as it is pharmaceutically, pharmacologically (pharmaceutically) or physiologically acceptable.

  Examples of the buffer include an inorganic buffer which is a buffer derived from an inorganic acid, and an organic buffer which is a buffer derived from an organic acid or an organic base.

  Examples of the inorganic buffer include a borate buffer, a phosphate buffer, and a carbonate buffer. Examples of the borate buffer include boric acid or a salt thereof (such as an alkali metal borate and an alkaline earth metal borate). Examples of the phosphate buffer include phosphoric acid and salts thereof (eg, alkali metal phosphate and alkaline earth metal phosphate). Examples of the carbonate buffer include carbonic acid and salts thereof (eg, alkali metal carbonate and alkaline earth metal salt). Further, a borate, a phosphate or a carbonate hydrate may be used as the borate buffer, the phosphate buffer or the carbonate buffer. As a more specific example, boric acid or a salt thereof (sodium borate, potassium tetraborate, potassium metaborate, ammonium borate, borax, etc.) as a borate buffer; phosphoric acid or a salt thereof as a phosphate buffer Salts (disodium hydrogen phosphate, sodium dihydrogen phosphate, potassium dihydrogen phosphate, trisodium phosphate, tripotassium phosphate, calcium monohydrogen phosphate, calcium dihydrogen phosphate, etc.); Or salts thereof (sodium hydrogen carbonate, sodium carbonate, ammonium carbonate, potassium carbonate, calcium carbonate, potassium hydrogen carbonate, magnesium carbonate, etc.).

  Examples of the organic buffer include an acetate buffer, a lactate buffer, a succinate buffer, a Tris buffer, an AMPD buffer and the like. Examples of the acetic acid buffer include acetic acid or a salt thereof (eg, an alkali metal acetate, an alkaline earth metal acetate, and the like). Examples of the lactic acid buffer include lactic acid or a salt thereof (eg, an alkali metal lactate and an alkaline earth metal lactate). Examples of the succinic acid buffer include succinic acid or a salt thereof (such as an alkali metal succinate). In addition, acetate, lactate, or succinate hydrate may be used as the acetate buffer, the lactate buffer, or the succinate buffer. As more specific examples, acetic acid or a salt thereof (ammonium acetate, sodium acetate, potassium acetate, calcium acetate, etc.) as an acetic acid buffer; lactic acid or a salt thereof (sodium lactate, potassium lactate, calcium lactate, etc.) as a lactic acid buffer ); Examples of the succinic acid buffer include succinic acid and salts thereof (monosodium succinate, disodium succinate, etc.). Examples of the Tris buffer include tromethamol or a salt thereof (tromethamol hydrochloride or the like). Examples of the AMPD buffer include 2-amino-2-methyl-1,3-propanediol or a salt thereof.

  Examples of the buffer include a borate buffer (for example, a combination of boric acid and borax), a phosphate buffer (for example, a combination of disodium hydrogen phosphate and sodium dihydrogen phosphate), and a tris buffer (for example, , Tromethamol) is preferred, a borate buffer is more preferred, boric acid and its salts are more preferred, and a combination of boric acid and borax is even more preferred.

  A commercially available buffer may be used. One type of buffer may be used alone, or two or more types may be used in combination.

  The content of the buffer other than the component (C) in the ophthalmic composition according to the present embodiment is not particularly limited, and the type of the buffer, the type and content of the other components, the use and the form of the ophthalmic composition, and the form of the ophthalmic composition Is appropriately set in accordance with. The content of the buffer other than the component (C) is, for example, based on the total amount of the ophthalmic composition, from the viewpoint of more remarkably exerting the effects of the present invention. , 0.001 to 20 w / v%, more preferably 0.01 to 4.0 w / v%, even more preferably 0.05 to 3.0 w / v%.

  The pH of the ophthalmic composition according to the present embodiment is not particularly limited as long as it is within a pharmaceutically, pharmacologically (pharmaceutically) or physiologically acceptable range. The pH of the ophthalmic composition according to the present embodiment may be, for example, 4.0 to 9.5, preferably 4.0 to 9.0, and more preferably 4.5 to 9.0. Is more preferably 4.5 to 8.5, still more preferably 5.0 to 8.5, still more preferably 5.0 to 8.0.

  The ophthalmic composition according to the present embodiment can be adjusted to an osmotic pressure ratio within a range acceptable for a living body, if necessary. The appropriate osmotic pressure ratio can be appropriately set according to the use, formulation form, method of use, etc. of the ophthalmic composition, and can be, for example, 0.4 to 5.0, and 0.6 to 3.0. It is preferably 0.8 to 2.2, more preferably 0.8 to 2.0. The osmotic pressure ratio is the ratio of the osmotic pressure of the sample to 286 mOsm (the osmotic pressure of a 0.9 w / v% aqueous sodium chloride solution) based on the 17th Revised Japanese Pharmacopoeia. (Freezing point descent method). The osmotic pressure ratio measurement standard solution (0.9 w / v% sodium chloride aqueous solution) is obtained by drying sodium chloride (Japanese Pharmacopoeia standard reagent) at 500 to 650 ° C for 40 to 50 minutes and then in a desiccator (silica gel). After allowing to cool, 0.900 g thereof is accurately measured and dissolved in purified water to prepare exactly 100 mL, or a commercially available standard solution for measuring osmotic pressure ratio (0.9 w / v% aqueous sodium chloride solution) can be used.

  The viscosity of the ophthalmic composition according to the present embodiment is not particularly limited as long as it is within a pharmaceutically, pharmacologically (pharmaceutically) or physiologically acceptable range. As the viscosity of the ophthalmic composition according to the present embodiment, for example, the viscosity at 20 ° C. measured with a rotational viscometer (RE550 type viscometer, manufactured by Toki Sangyo Co., Ltd., rotor: 1 ° 34 ′ × R24) is 0.5. The pressure is preferably 10 to 10 mPa · s, more preferably 1 to 5 mPa · s, and still more preferably 1 to 3 mPa · s.

The ophthalmic composition according to the present embodiment contains an appropriate amount of a combination of components selected from various pharmacologically active components and physiologically active components in addition to the above components as long as the effects of the present invention are not impaired. Is also good. The component is not particularly limited, and examples thereof include an active component in an ophthalmic drug described in the OTC Pharmaceuticals Manufacturing and Marketing Approval Standards 2012 edition (supervised by the Society of Regulatory Science). Specific examples of the components used in ophthalmic drugs include the following components.
Antiallergic agents: for example, sodium cromoglycate, tranilast, potassium pemirolast and the like.
Antihistamines: for example, diphenhydramine hydrochloride, iproheptin, chlorpheniramine maleate, levocabastine hydrochloride, ketotifen fumarate, olopatadine hydrochloride and the like.
(A) amino acids other than the component (B): for example, monoaminomonocarboxylic acids such as glycine, alanine, aminobutyric acid, aminovaleric acid and aminocaproic acid; monoaminodicarboxylic acids such as glutamic acid and salts thereof; diaminomono acids such as arginine and lysine Carboxylic acids or salts thereof; aminoethylsulfonic acid (taurine) or salts thereof, and the like.
Anti-inflammatory agents: For example, methyl salicylate, glycol salicylate, allantoin, dipotassium glycyrrhizinate, tranexamic acid, lysozyme, lysozyme chloride, indomethacin, planoprofen, ibuprofen, ibuprofen piconol, ketoprofen, felbinac, bendazac, piroxicam, bufexamac, flufenam Butyl acid, epsilon-aminocaproic acid, berberine chloride, berberine sulfate, sodium azulene sulfonate and the like.
Steroids: For example, fluticasone propionate, fluticasone furoate, mometasone furoate, beclomethasone propionate, flunisolide and the like.
Decongestant: For example, tetrahydrozoline hydrochloride, tetrahydrozoline nitrate, naphazoline hydrochloride, naphazoline nitrate, epinephrine, epinephrine hydrochloride, ephedrine hydrochloride, phenylephrine hydrochloride, dl-methylephedrine hydrochloride and the like.
Ocular muscle modulators: for example, cholinesterase inhibitors having an active center similar to acetylcholine, such as neostigmine methyl sulfate, tropicamide, helenien, atropine sulfate and the like.
Vitamins other than the component (A): retinol acetate, retinol palmitate, tocopherol acetate, sodium flavin adenine dinucleotide, pyridoxine hydrochloride, panthenol, calcium pantothenate, ascorbic acid, sodium ascorbate and the like.
Inorganic salts: chlorides of metals such as calcium chloride, magnesium chloride, sodium chloride and potassium chloride; ammonium chloride; sulfates of metals such as calcium sulfate, magnesium sulfate, sodium sulfate, potassium sulfate and ammonium sulfate.
Astringents: for example, zinc white, zinc lactate, zinc sulfate and the like.
Others: For example, sulfamethoxazole, sulfisoxazole, sulfisomidine, and salts thereof.

In the ophthalmic composition according to the present embodiment, as long as the effects of the present invention are not impaired, various additives are appropriately selected according to a usual method, depending on the use and the form of the preparation, and one or more kinds are added. May be used together in an appropriate amount. Examples of such additives include various additives described in the Pharmaceutical Excipients Dictionary 2007 (edited by the Japan Pharmaceutical Excipients Association). Representative additives include the following additives.
Carrier: For example, an aqueous solvent such as water and hydrous ethanol.
Chelating agents: for example, ethylenediaminediacetic acid (EDDA), ethylenediaminetriacetic acid, ethylenediaminetetraacetic acid (EDTA), N- (2-hydroxyethyl) ethylenediaminetriacetic acid (HEDTA), diethylenetriaminepentaacetic acid (DTPA) and the like.
Base: for example, octyldodecanol, titanium oxide, potassium bromide, plastibase and the like.
pH adjusters: for example, hydrochloric acid, acetic acid, sodium hydroxide, potassium hydroxide, calcium hydroxide, magnesium hydroxide, triethanolamine, diisopropanolamine and the like.
Nonionic surfactants: for example, polyoxyethylene sorbitan fatty acid esters, polyoxyethylene hydrogenated castor oil, polyoxyethylene castor oil, polyethylene glycol monostearate, poloxamers and the like.
Anionic surfactants: for example, polyoxyethylene alkyl ether phosphate, polyoxyethylene alkyl ether sulfate, alkylbenzene sulfonate, alkyl sulfate, N-acyl taurate and the like.
Amphoteric surfactant: for example, betaine lauryldimethylaminoacetate and the like.
Cooling agents: For example, menthol, menthol, camphor, borneol, geraniol, cineol, citronellol, carvone, anethole, eugenol, limonene, linalool, linalyl acetate, thymol, cymene, terpineol, pinene, camphene, isoborneol, fenchen, nerol , Myrcene, myrsenol, linalool acetate, lavandulol, eucalyptus oil, bergamot oil, peppermint oil, cool mint oil, spearmint oil, peppermint oil, fennel oil, cinnamon oil, rose oil, camphor oil and the like.
Stabilizers: for example, sodium formaldehyde sulfoxylate (Rongalit), sodium bisulfite, sodium pyrosulfite, aluminum monostearate, glyceryl monostearate, cyclodextrin, monoethanolamine, dibutylhydroxytoluene and the like.
Preservatives: For example, quaternary ammonium salts of alkyl polyaminoethyl glycines (eg, benzalkonium chloride, benzethonium chloride, etc.), chlorhexidine gluconate, polydronium chloride, zinc chloride, sodium benzoate, ethanol, chlorobutanol, sorbic acid, Potassium sorbate, sodium dehydroacetate, methyl paraoxybenzoate, ethyl paraoxybenzoate, propyl paraoxybenzoate, butyl paraoxybenzoate, oxyquinoline sulfate, phenethyl alcohol, benzyl alcohol, biguanide compounds (specifically, polyhexanide hydrochloride (poly Hexamethylene biguanide), alexidine, etc.), Gloquil (trade name, manufactured by Rhodia) and the like.
Isotonicity agents: for example, sodium bisulfite, sodium sulfite, potassium chloride, calcium chloride, sodium chloride, magnesium chloride, potassium acetate, sodium acetate, sodium hydrogen carbonate, sodium carbonate, sodium thiosulfate, magnesium sulfate, glycerin, propylene glycol etc.
Sugar alcohols: for example, xylitol, sorbitol, mannitol, glycerin and the like. These may be d-form, l-form or dl-form.
Oils: for example, vegetable oils such as sesame oil, castor oil, soybean oil, olive oil, animal oils such as squalane, and mineral oils such as liquid paraffin and petrolatum.

  When the ophthalmic composition according to the present embodiment contains water, as the content of water, from the viewpoint of more remarkably exerting the effects of the present invention, for example, based on the total amount of the ophthalmic composition, the content of water is , 80 w / v% or more and less than 100 w / v%, more preferably 85 w / v% or more and 99.5 w / v% or less, and 90 w / v% or more and 99.2 w / v% or less. Is more preferred.

  The water used in the ophthalmic composition according to the present embodiment may be pharmaceutically, pharmacologically (pharmaceutically) or physiologically acceptable. Examples of such water include distilled water, ordinary water, purified water, sterilized purified water, water for injection, and distilled water for injection. These definitions are based on the 17th revised Japanese Pharmacopoeia.

  The ophthalmic composition according to the present embodiment is, for example, a component (A), a component (B), a component (C), a component (D), and other components as required so as to have a desired content. It can be prepared by adding and mixing. Specifically, for example, it can be prepared by dissolving or suspending the above components in purified water, adjusting the pH and osmotic pressure to a predetermined value, and sterilizing by filtration sterilization or the like.

  The ophthalmic composition according to the present embodiment can take various dosage forms depending on the purpose, and examples thereof include a liquid preparation, a gel preparation, and a semi-solid preparation (such as an ointment). Among them, liquid preparations are preferable, and aqueous liquid preparations are more preferable.

  The ophthalmic composition according to the present embodiment includes, for example, eye drops (also referred to as eye drops or eye drops. In addition, the eye drops include artificial tears, eye drops that can be instilled while wearing a contact lens), and eye wash. (Also referred to as an eyewash or an eyewash. The eyewash includes an eyewash that can be washed while wearing a contact lens.) A composition for a contact lens [a solution for wearing a contact lens, a composition for contact lens care (a contact lens) Disinfectant, contact lens preservative, contact lens cleaning agent, contact lens cleaning preservative), contact lens mounting ophthalmic solution used for both contact lens mounting solution and eye drops during contact lens mounting] Can be used as Preferred examples of the ophthalmic composition according to the present embodiment include eye drops, eyewashes, and compositions for contact lenses, and more preferred examples are eye drops, eye drops that can be instilled while wearing contact lenses, and eyewashes. And eye drops that can be washed while wearing a contact lens, contact lens mounting solutions, and eye drops that are worn with contact lenses. More preferred examples include eye drops and eye drops that can be dropped while wearing contact lenses. The “contact lens” includes a hard contact lens and a soft contact lens (including both ionic and nonionic, and including both silicone hydrogel contact lenses and non-silicone hydrogel contact lenses).

  When the ophthalmic composition according to the present embodiment is an ophthalmic solution, its usage and dosage are not particularly limited as long as it is effective and has few side effects, for example, for adults (15 years old or older) and 7 years old In the case of the above children, a method of instilling 1 to 2 drops at a time 4 times a day, using 1 to 2 drops, 1 to 3 drops, or 2 to 3 drops 5 to 6 times a day The method used can be exemplified.

  The ophthalmic composition according to the present embodiment is provided in an arbitrary container. The container for containing the ophthalmic composition according to the present embodiment is not particularly limited, and may be, for example, made of glass or plastic. It is preferably made of plastic. Examples of the plastic include polyethylene terephthalate, polyarylate, polyethylene naphthalate, polycarbonate, polyethylene, polypropylene, polyimide, a copolymer of the monomers constituting these, and a mixture of two or more of these. Preferred are polypropylene, polyethylene and polyethylene terephthalate, and more preferred is polyethylene terephthalate. In addition, the container that stores the ophthalmic composition according to the present embodiment may be a transparent container in which the inside of the container can be visually recognized, or may be an opaque container in which the inside of the container is difficult to visually recognize. Preferably, it is a transparent container. Here, the “transparent container” includes both a colorless transparent container and a colored transparent container.

  A nozzle that accommodates the ophthalmic composition according to the present embodiment may be equipped with a nozzle. The material of the nozzle is not particularly limited, and may be, for example, glass or plastic. It is preferably made of plastic. Examples of the plastic include polyethylene terephthalate, polybutylene terephthalate, polyethylene, polypropylene, polyethylene naphthalate and a copolymer of the monomers constituting these, and a mixture of two or more of these. As the material of the nozzle, from the viewpoint of further enhancing the effects of the present invention, polypropylene, polyethylene, polyethylene terephthalate, polybutylene terephthalate, and polyethylene naphthalate are preferable, and polyethylene is more preferable.

  The container for storing the ophthalmic composition according to the present embodiment may be a multi-dose type in which a plurality of usage amounts are stored, or may be a unit dose type in which a single usage amount is stored. Since the effects of the present invention can be more remarkably exhibited, a multi-dose type is preferable.

  Since the ophthalmic composition according to the present embodiment contains at least one selected from the group consisting of cyanocobalamin, and aspartic acid and a salt thereof, the ophthalmic composition has an effect of promoting the formation of elastic fibers and a production of an elastic fiber-related factor. It has the effect of doing. Therefore, the ophthalmic composition according to the present embodiment can be used as an elastic fiber formation promoter and an elastic fiber-related factor production promoter.

[3. Ophthalmic composition for improving conjunctival relaxation, ophthalmic composition for improving pupil brightness, ophthalmic composition for anti-aging]
Since the ophthalmic composition according to the present embodiment contains at least one selected from the group consisting of cyanocobalamin, and aspartic acid and salts thereof, formation of elastic fibers of the conjunctiva is promoted, and conjunctival relaxation is improved. Therefore, as one embodiment of the present invention, there is provided an ophthalmic composition for improving conjunctival relaxation, comprising at least one selected from the group consisting of cyanocobalamin, and aspartic acid and salts thereof. In this specification, “conjunctival relaxation” means a state in which the conjunctiva is relaxing. Conjunctival relaxation may be a state in which the conjunctiva is relaxed due to aging (eg, conjunctival relaxation with aging), specifically, conjunctival flaccidity, conjunctival wrinkles (white-eye wrinkles), Includes conjunctival sagging (white eye sagging) and the like.
Also, when the conjunctiva relaxes or the elastic fibers of the conjunctiva decrease, tears cannot stay on the surface of the eye, and the “glow of the pupil” is lost. On the other hand, when the formation of elastic fibers of the conjunctiva is promoted, the relaxation of the conjunctiva is improved, the tears are more likely to stay on the surface of the eyes, and the “brightness of the pupils” is improved. Therefore, as one embodiment of the present invention, there is provided an ophthalmic composition for improving pupil luminosity, comprising at least one selected from the group consisting of cyanocobalamin and aspartic acid and salts thereof.
Further, improving conjunctival relaxation generally leads to anti-aging of the eyes and conjunctiva. Therefore, as one embodiment of the present invention, there is provided an ophthalmic composition for anti-aging comprising cyanocobalamin and at least one selected from the group consisting of aspartic acid and salts thereof. In addition, improving the conjunctival relaxation leads to improvement of indefinite complaints such as dryness of the eyes (dry eyes), eye fatigue, blurred eyes, foreign body sensation, difficulty in focusing, and hyperemia.

  In these embodiments, regarding the type and content of the component (A), the type and content of the component (B), the type and content of other components, the formulation of the ophthalmic composition, and the like, [2. Ophthalmic composition].

  Hereinafter, the present invention will be specifically described based on test examples, but the present invention is not limited thereto.

[Test Example 1: Measurement of Fibulin-5 gene expression in human conjunctival fibroblasts]
After expanding HCConf (human conjunctival fibroblasts, Sciencecell # 6570, Lot 5965) using FM (Sciencecell # 2301), a 12-well plate was prepared so as to have a cell number of 2 × 10 ⁵ cells / well (1 mL). And cultured at 37 ° C. under 5% CO ₂ for 1 day. Thereafter, the medium was changed to DMEM / F12 (Thermo Fisher Scientific 11039021) to which an antibiotic (Thermo Fisher Scientific 15240062) was added, and the cells were further cultured for one day. After the medium in each well was removed by suction, 1 mL of a medium containing the components described in Examples and Comparative Examples shown in Table 1 was added, and the cells were cultured for 48 hours. Here, DMEM / F12 to which an antibiotic was added was used for dissolving the components described in Examples and Comparative Examples shown in Table 1. The numerical values of each component shown in Table 1 are expressed in w / v% based on the total amount of the medium. Thereafter, the cells were collected using QIAS Shredder (QIAGEN 79656), and then the mRNA was collected using RNeasy Mini Kit (250) (QIAGEN 74106). Using the obtained mRNA solution, ReverseTra Ace (registered trademark) (TOYOBO TRT-101), a reverse transcription reaction was carried out with a thermal cycler (Applied Biosystems 2720). Using the obtained cDNA solution, TaqMan (registered trademark) probe (Thermo Fisher Scientific), TaqMan (registered trademark) Fast Advanced Master Mix (ThermoFisherResearchFisher-Third-QuantitativeResearchFisher-Third-Quantitative-Research). The reaction was performed. 18 s was used for the internal standard. All methods of using the kit followed the attached protocol. After the quantitative Real-Time PCR, the CT value of each sample was calculated by an automatic analysis of QuantStudio (registered trademark) Design & Analysis Software. Using the calculated CT values, the average value and SD value of the relative mRNA expression levels of each Example and Comparative Example with respect to the untreated group (Comparative Example 1) were calculated by the following procedures 1) to 3).
1) The ΔCT value of Fibulin-5 in each Example and Comparative Example was calculated from Expression (1).
Equation (1): ΔCT value = (CT value of Fibulin-5 in each example and comparative example) − (CT value of 18s)
2) The average value of the ΔCT values of the non-treated group (Comparative Example 1) was calculated, and the ΔΔCT values of the respective Examples and Comparative Examples were calculated from Expression (2).
Equation (2): ΔΔCT value = (ΔCT value of each example and comparative example) − (average ΔCT value of non-treated group (Comparative example 1))
3) The relative mRNA expression level of each Example and Comparative Example relative to the untreated group (Comparative Example 1) was calculated from Equation (3).
Formula (3): Relative mRNA expression level of each Example and Comparative Example = 2− ^ΔΔCT value The results are shown in Table 1 and FIG.

In Examples 1-1, 1-2 and 1-3 treated with a medium containing cyanocobalamin, potassium aspartate or potassium magnesium aspartate, the mRNA of Fibulin-5 was lower than that of the untreated comparative example 1-1. The expression level increased. On the other hand, in Comparative Example 1-2 treated with a medium containing anhydrous citric acid, the expression level of Fibulin-5 mRNA was lower than that in Comparative Example 1-1 without treatment.
Further, in Examples 1-4 and 1-5 in which the medium was treated with cyanocobalamin or potassium aspartate mixed with anhydrous citric acid, Examples 1 and 2 in which the medium was treated with cyanocobalamin or potassium aspartate alone. In Example 1-6 in which the expression level of Fibulin-5 mRNA was increased as compared with that in Example 1-6, which was treated with a medium containing cyanocobalamin, magnesium / potassium aspartate, and citric anhydride, the expression of Fibulin-5 mRNA was increased. It was confirmed that the amount was significantly increased.

[Test Example 2: Evaluation of elastic fiber-related factor production and elastic fiber formation]
HConf (human conjunctival fibroblasts, Sciencecell # 6570, Lot 5965) was expanded and cultured using FM (Sciencecell # 2301), and then collagen 1 was coated so as to obtain 2 × 10 ⁴ cells / well (100 μL). The cells were inoculated into a 96-well plate (FALCON 354407) and cultured at 37 ° C. under 5% CO ₂ for 2 days. The medium in each well was removed by suction, and 200 µL of the medium containing the components described in Examples and Comparative Examples shown in Tables 2 and 3 was added to each well, and the cells were cultured at 37 ° C under 5% CO ₂ for 3 weeks. Here, the components described in Examples and Comparative Examples shown in Tables 2 and 3 were dissolved in a fibrogenesis medium (DMEM / F12 (Thermo Fisher Scientific 11039021), FGS (Science cell # 2301), FBS (MP Bio IC2917354F). ) And an antibiotic (Thermo Fisher Scientific 15240062), which was filtered through a 0.2 μm filter.The numerical values of each component shown in Tables 2 and 3 are w / v% based on the total amount of the medium. After 3 weeks, the medium containing the components described in each Example and Comparative Example was removed by decantation, and the plate was washed with 200 µL of D-PBS (-) (Kojin Bio, 16220015). Methanol (WAKO 137-01823) 5 0 μL was slowly added, and the mixture was fixed for 20 minutes at −20 ° C. After the fixation, methanol was removed, and the plate was washed with 200 μL of PBS, 200 μL of 1% BSA (Sigma A3059-100G) -PBS was added, and the reaction was performed at room temperature for 1 hour. Thereafter, 1% BSA-PBS was removed, and the plate was washed with 200 μL of PBS.
Primary antibody (Rabbit anti human FBLN5 antibody (Proteintech 12188-1-AP) or Mouse anti human ELN antimicrobial prepared by adding Rabbit anti human FBLN5 antibody diluted 200-fold with Canget Signal A (TOYOBO NKB-501) And incubated for 2 hours at room temperature. The primary antibody was removed and washed twice with 200 μL of PBS. Then, a secondary antibody (Goat anti-rabbit IgG 2nd antibody Alexa 546 (Thermo Fisher Scientific A11010) or Goat anti-mouse of first-generation anti-aging drug was added to 0.1% BSA-PBS and then diluted with 0.1% BSA-PBS. This was added in an amount of 50 μL, and allowed to react for 1.5 hours in a dark place. After removing the secondary antibody and washing with 200 μL of PBS, the localization of elastic fiber-related factor (Fibulin-5) and the formation of elastic fibers (elastin fibers) were observed with a fluorescence microscope, and photographs were taken (FIG. 2 and FIG. 2). 3). Using ImageJ, the ratio (%) of the elastic fiber-related factor (Fibulin-5) or the elastic fiber (elastin fiber) in the obtained image was calculated.
The results are shown in Tables 2 and 3, and FIGS.

As shown in Table 2 and FIG. 2, in Examples 2-1 to 2-4, which were treated with the medium containing potassium aspartate having different contents, the elastic fiber-related factor (Fibulin-5) increases in a concentration-dependent manner. It was confirmed that. In Example 2-5, which was treated with a medium in which citric acid was added to potassium aspartate, Example 2-3 and Comparative Example 2-2, each of which was treated with a medium containing potassium aspartate or citric anhydride alone. It was confirmed that the elastic fiber-related factor (Fibulin-5) was remarkably increased as compared with.
As shown in Table 3 and FIG. 3, it was confirmed that in Examples 2-3 and 2-4 treated with media containing potassium aspartate having different contents, elastic fibers (elastin fibers) increased in a concentration-dependent manner. Was done. In Example 2-5, which was treated with a medium in which potassium aspartate was mixed with anhydrous citric acid, Example 2-3 and Comparative Example 2-, in which each of the cells was treated with a medium containing potassium aspartate or anhydrous citric acid alone. It was confirmed that elastic fibers (elastin fibers) increased remarkably as compared with 2.

(Formulation example)
Formulation examples are shown in Tables 4 and 5 below. The units of each component in Tables 4 and 5 are w / v% except for those specified in the tables. Formulation Examples 1 to 19 are all eye drops.

Claims (10)

  An elastic fiber formation promoter comprising at least one selected from the group consisting of cyanocobalamin, and aspartic acid and salts thereof.   The elastic fiber formation promoting agent according to claim 1, further comprising at least one selected from the group consisting of citric acid and salts thereof.   The elastic fiber formation promoter according to claim 1 or 2, which is used for promoting formation of elastic fibers of the conjunctiva.   An elastic fiber-related factor production promoter comprising at least one selected from the group consisting of cyanocobalamin and aspartic acid and salts thereof.   The elastic fiber-related factor production promoter according to claim 4, further comprising at least one selected from the group consisting of citric acid and salts thereof.   The agent for promoting production of an elastic fiber-related factor according to claim 4 or 5, which is used for promoting production of an elastic fiber-related factor of the conjunctiva.   The elastic fiber-related factor production promoter according to any one of claims 4 to 6, wherein the elastic fiber-related factor is Fibulin-5.   An ophthalmic composition comprising at least one selected from the group consisting of cyanocobalamin, aspartic acid and salts thereof, at least one selected from the group consisting of citric acid and salts thereof, and a thickener.   The ophthalmic composition according to claim 8, wherein the thickener is at least one selected from the group consisting of hydroxypropylmethylcellulose, and hyaluronic acid and salts thereof. An ophthalmic composition for improving pupil luminosity, comprising at least one selected from the group consisting of cyanocobalamin, and aspartic acid and salts thereof.