TW201343194A - Heparan sulfate production promoter - Google Patents

Abstract

Provided is a heparan sulfate production promoter comprising a mixture of an extract of the Madonna lily (Lilium Candidum) and glucosamine.

Description

Heparin sulfate production promoter

The present invention provides an acesulfate heparin production promoter comprising a mixture of Lilium Candidum extract and glucosamine.

Heparin sulfate is a linear polysaccharide composed of glucosamine, glucuronic acid and iduronic acid, and is often covalently bonded to proteins in the cell surface or extracellular matrix of animals. The form of the formed acesulfate heparin proteoglycan is present. Heptasulfate heparin proteoglycans bind to a variety of physiologically active proteins, such as cell proliferating factors, chemokines, cytokines, morphological factors, etc., and exert cell differentiation, cell proliferation regulation, and cell morphology in vivo. Various functions such as hair growth, cell follow-up, mutual recognition, protection of cell infiltration, and promotion of wound healing.

In particular, heparin sulfate proteoglycan is known to have heparin-binding growth factor (bFGF (basic fibroblast growth factor), HGF (hepatocyte growth factor), VEGF (vascular endothelium). Growth factors, vascular endothelial growth factor, HB-EGF (heparin-binding epidermal growth factor like growth factor), etc.). A basement membrane proteoglycan, which is one type of acesulfate heparin proteoglycan, is also present on the epidermal basement membrane present at the boundary between the epidermis and the dermis, in the skin, by binding heparin-binding growth factor to the epidermal basement membrane. And control the movement of growth factors between the epidermis and the dermis. Moreover, the basement membrane proteoglycan present on the epidermal basement membrane also controls the growth factor binding to the basement membrane to the epidermal basal cells. It is used to clarify that it is necessary for the good proliferation and differentiation of the epidermis. As a result, it is clear that the production of heparin sulfate is also related to the prevention or inhibition of wrinkles (Experimental Dermatology Vol. 20, 10, Oct. 2011, pp. 810-814: Non-Patent Document 1 and WO 2009/123215: Patent Document 1) .

Further, in the heparin sulfate decomposition model, the permeability of VEGF is increased to induce angiogenesis (WO2011/040496: Patent Document 2). If the dermal changes caused by ultraviolet-induced dermal neovascularization are considered to be important for the formation of large wrinkles (Japanese Patent Special Table 2004-526758: Patent Document 3), the decomposition of acesulfate heparin is also closely related to the formation of large wrinkles. . Therefore, it is known that the promotion of the production of heparin sulfate is also effective for the prevention of large wrinkles caused by long-term sun exposure or the like.

It is also clear that the senile pigmented plaque tissue is decomposed in the basement membrane compared to the skin of the exposed portion (Patent Document 2). The publication of Patent Document 2 discloses that the control of vascular endothelial growth factor-A (VEGF-A) expressed in the epidermis is deactivated by the decomposition of acesulfate heparin, thereby changing the blood vessels or lymphatic vessels of the dermis. Inflammation is produced, thereby activating melanocytes to promote the storage of melanin in melanosomes. Further, in the same publication, it is described that melanosomes are promoted from melanocytes in epidermal cells due to failure of control of fibroblast growth factor-7 (FGF-7) expressed in the dermis. That is, it is considered that the decomposition of acetaminophen heparin synergistically accumulates melanosomes in keratinocytes by activation of melanocytes due to inflammation and promotion of melanosomes due to failure of FGF-7 control. Therefore, the production of heparin sulfate is also related to the whitening effect for preventing and inhibiting pigmentation such as chloasma, dark spots, and freckles.

Heparin sulfate is modified by sulfation or epimerization to form various structures. The physiological activity of the acesulfate heparin proteoglycan is the chain length of the oligosaccharide, and the sugar chain length, the sugar chain sequence, and the specific sulfate group are important for cell function, and are displayed according to the nature of the cell. A pattern of specificity that interacts with specific proteins. Therefore, acetaminophen sulfate is responsible for the performance of cell characteristics. Further, it also shows sulfur Acetyl heparin is responsible for the cellular receptors of various viruses, including respiratory syncytial virus.

It is known in WO 2009/123215 (Patent Document 1) that the lily extract has an acetoinase inhibitory effect. Further, a cosmetic material is known from Japanese Patent Laid-Open Publication No. 2011-225564 (Patent Document 4), which is obtained from a flower bud belonging to the lilium family of Liliaceae, which is obtained for wrinkles, sagging, chloasma, freckles. The skin aging phenomenon is excellent in preventing and improving the skin, keeping the skin in a delicate and healthy state, or improving the skin, and the skin is less irritating, and the safety of the organism is excellent, and the skin is whitening and whitening. An extract having an inhibitory action of elastase activity, an antioxidant action, a inhibition of protein saccharification, and an inhibitory action of tyrosinase activity is used as an active ingredient. Japanese Patent Laid-Open No. 2012-41284 (Patent Document 5) is effective as an extract of Liliaceae (Cardiocrinum) which is excellent in anti-aging effect, whitening action, anti-oxidation action, body-effect, and anti-inflammatory action. Formulated to provide anti-aging agents, whitening agents, antioxidants, body lotions and anti-inflammatory agents. However, it is not known that combination with glucosamine hydrochloride promotes the production of heparin sulfate.

For the glucosamine hydrochloride, a skin external preparation containing a highly glycated end product production inhibitor and a whitening agent (glucosamine is described as one of the components) is known from JP-A-2003-261432 (Patent Document 6). A skin external preparation having a markedly improved skin whitening effect, which is one or more selected from the group consisting of plants and algae extracts, is known in the Japanese Patent Publication No. 2002-128647 (Patent Document 7). And the lavender extract, the extract of the above plant and algae is one or more selected from the group consisting of glucosamine and a derivative thereof, and has melanin production inhibitory action. However, it is not known that it promotes the production of heparin sulfate or inhibits heparinase.

As described above, acesulfate heparin has various physiological effects on human and animal constituents, but the agent for promoting its production is not well known.

[Previous Technical Literature] [Patent Literature]

[Patent Document 1] WO2009/123215

[Patent Document 2] WO2011/040496

[Patent Document 3] Japanese Patent Special Table 2000-503660

[Patent Document 4] Japanese Patent Laid-Open No. 2011-225564

[Patent Document 5] Japanese Patent Special Opening 2012-41284

[Patent Document 6] Japanese Patent Laid-Open No. 2003-261432

[Patent Document 7] Japanese Patent Laid-Open No. 2002-128647

[Non-patent literature]

[Non-Patent Document 1] Experimental Dermatology Vol. 20, 10, Oct. 2011, PP. 810 - 814

An object of the present invention is to provide a novel heparin sulfate production promoter.

As a result of intensive studies, the present inventors have found that a mixture of Lilium Candidum extract and glucosamine exhibits an effect of promoting the production of heparin sulfate.

Accordingly, the present application provides the following invention.

(1) A heparin sulfate production promoter comprising a mixture of Lilium Candidum extract and glucosamine.

(2) The acesulfate heparin production promoter according to (1), wherein the glucosamine is glucosamine hydrochloride.

(3) A whitening agent containing the acesulfate heparin production promoter such as (1) or (2) as a whitening component.

(4) A preventive or inhibitor of wrinkle formation comprising an acesulfate heparin production promoter such as (1) or (2) as an anti-aging component.

(5) A heparin sulfate production promoter according to (1) or (2), which is a whitening agent.

(6) A heparin sulfate production promoter according to (1) or (2), which is a prophylactic or inhibitor of wrinkle formation.

(7) Use of a mixture of Lilium Candidum extract and glucosamine for producing an ethidium sulfate heparin production promoter.

(8) The use of (7), wherein the glucosamine is glucosamine hydrochloride.

(9) The use of (7) or (8), wherein the acesulfate heparin production promoter is a whitening agent.

(10) The use of (7) or (8), wherein the heparin sulfate production promoter is a prophylactic or inhibitor of wrinkle formation.

(11) Use of a mixture of Lilium Candidum extract and glucosamine for promoting the production of heparin sulfate.

(12) The use of (11), wherein the glucosamine is glucosamine hydrochloride.

(13) The use of (11) or (12), wherein the acesulfate heparin production promoting system is used for whitening.

(14) The use of (11) or (12), wherein the acesulfate heparin production promoting line is used for prevention or inhibition of wrinkle formation.

The acesulfate heparin production promoter of the present invention can efficiently promote the production of acesulfate heparin, and thus can be suitably used for the following purposes: for various states or symptoms associated with decomposition, reduction, and deficiency of acesulfate heparin Uses for improvement, treatment, prevention, and normalization, such as the use of chloasma, freckles, dark spots, etc. as a whitening agent; wrinkle improvement and the like for anti-aging purposes; for promoting wound healing, in addition For example, it can be suitably used for normalization of abnormalities in cell differentiation, regulation of cell proliferation, morphogenesis of cells, cell adhesion, mutual recognition, and cell infiltration.

Figure 1 shows the effect of each agent having an effect on the production of acesulfate heparin.

The present invention provides an acesulfate heparin production promoter comprising a mixture of Lilium Candidum extract and glucosamine hydrochloride. The lily of the genus is a perennial herb native to the Asian family of Liliaceae. It is naturally grown and cultivated, and its breeding system divides the bulb. It is said that the extract of the lily of the lily is used for the formulation of arthritis or rheumatism, and it is also preferable as a cosmetic effect for preventing wrinkles.

The above-mentioned extract can be obtained by a usual method. For example, a part or all of the plant which is the origin of the plant can be obtained by immersing or heating under reflux at room temperature or heating, followed by filtration and concentration. In the extraction, any of the leaves, flowers, seeds, roots, stems, buds, branches, bark, etc. of the plant may be used, but it is preferably an extract from the bulb. The extraction site can also be dried prior to solvent extraction. The extraction solvent can be used arbitrarily as long as it is a solvent used for usual extraction. For example, an organic solvent such as methanol, ethanol, propylene glycol, 1,3-butylene glycol, or an alcohol such as glycerin, or an alcohol can be used alone. , chloroform, dichloroethane, carbon tetrachloride, acetone, ethyl acetate, hexane, etc.; or an aqueous solvent such as water, physiological saline, phosphate buffer, boric acid buffer, or the like, or a combination of the above solvents. It is preferred to use one or two or more selected from the group consisting of water, methanol, ethanol, and 1,3-butylene glycol as a solvent. It is especially preferred to use a mixture of water, ethanol and 1,3-butanediol as a solvent.

The extract obtained by the extraction with the above solvent may be used as it is, or the extract obtained by extracting the extract obtained by the above solvent by lyophilization or the like may be used, and if necessary, the following may be used. : an impurity is removed by an adsorption method, for example, an ion exchange resin; or it is adsorbed on a column of a porous polymer (for example, Amberlite XAD-2), and then dissolved in a desired solvent and concentrated.

Glucosamine is a chemical name of "2-amino-2-deoxy-D-glucose" which is a kind of "amino sugar" which is known to have a structure in which a hydroxyl group at the 2-position of glucose is substituted with an amine group, and is a carapace. a constituent of chitin contained in a genus or insect, and exists as a carapace Chitosan of N-acetylated sulphate, but in mammalian organisms, glucosamine is also present in the form of N-acetylated glucosamine (N-acetylglucosamine) in various glucosamine aggregates including hyaluronic acid. Glycosaminoglycan or glycoprotein, glycolipid. Further, there are also those derived from plants, such as those derived from the presence of Aspergillus Niger to ferment corn. Glucosamine is basically "glucosamine hydrochloride" obtained by decomposing chitin by hydrochloric acid, and "glucosamine sulfate" is also used in a large amount. In the present invention, glucosamine hydrochloride is preferred.

In the acesulfate heparin production promoter of the present invention, the content of the mixture of the genus Lilium sinensis and the glucosamine is not particularly limited as long as it is sufficient to promote the effect of promoting the production of acesulfate heparin, as long as it is based on acesulfate heparin. The use of the accelerator can be appropriately selected. In general, the blending amount of the virgin lily of the entire heparin sulfate production promoter is usually 0.000001% by mass to 1% by mass based on the dry weight (solid content of the solid content), and preferably 0.00001% by mass. %~0.1% by mass, particularly preferably 0.0001 to 0.01, and the amount of glucosamine is usually set to 0.00001% by mass to 0.1% by mass, and more preferably 0.00005% by mass to 0.05% by mass, more preferably set to 0.005% by mass. 0.0001% by mass to 0.01% by mass. The ratio of the virgin lily to the glucosamine in the acesulfate heparin production promoter of the present invention can be set to 10:1 to 1:100, preferably 1:1 to 1: when the amount of the virgin lily is set to dry weight. 10, more preferably 1:2.

The acesulfate heparin production promoter of the present invention can be formulated according to a conventional method, or can be prepared as a component constituting a skin external preparation, but can be appropriately formulated as needed for cosmetics or medicines including quasi-drugs. A component such as a skin external preparation, such as an oil component, a surfactant, a powder, a coloring matter, water, an alcohol, a viscosity increasing agent, a chelating agent, a polyoxin, an antioxidant, an ultraviolet absorber, a moisturizing agent, a fragrance, Various medicinal ingredients, preservatives, pH adjusters, neutralizers, and the like. The other components are not particularly limited, and may be appropriately selected depending on the use, the dosage form, the administration form, and the like of the pharmaceutical composition, and examples thereof include pharmaceutically acceptable carriers and/or auxiliary agents. As an auxiliary, for example, it can be listed Lifting agents, adhesives, disintegrating agents, tackifiers, dispersing agents, resorption enhancers, flavoring agents, buffers, surfactants, dissolution aids, preservatives, emulsifiers, isotonic agents, stabilizers, pH adjuster, etc.

As a specific example, when the acesulfate heparin production promoter of the present invention is used as a skin external preparation, a component generally used for an external preparation such as a whitening agent, a moisturizer, an antioxidant, an oil component, or an ultraviolet absorbing agent may be appropriately formulated as needed. Agents, surfactants, tackifiers, alcohols, powder ingredients, colorants, aqueous ingredients, water, various skin nutrients, and the like. Further, a metal ion blocking agent such as disodium oxalic acid tetraacetate, trisodium oxalate tetraacetate, sodium citrate, sodium polyphosphate, sodium metaphosphate or gluconic acid may be appropriately formulated, and methylparaben, Preservatives such as ethyl p-hydroxybenzoate and butyl p-hydroxybenzoate, heat of caffeine, tannic acid, verapamil, tranexamic acid and its derivatives, licorice extract, licorice, and papaya Water extract, various natural medicines, vitamin E acetate, glycyrrhizic acid and its derivatives or its salts, vitamin C, magnesium ascorbyl phosphate, ascorbyl glucoside, arbutin, citric acid and other whitening agents, glucose, fructose, A saccharide such as mannose, sucrose or trehalose; a vitamin A derivative such as retinoic acid, retinol, retinyl acetate or retinyl palmitate.

The above components are exemplified and are not limited to these. Further, the components may be formulated in combination according to the formulation according to the desired form.

The administration route and dosage form of the acesulfate heparin production promoter of the present invention are not limited, and may be appropriately selected depending on the use thereof. Examples of the administration route include topical administration (external skin use, etc.), oral administration, parenteral administration (intravenous administration, intraperitoneal administration, etc.), and the like, and are used as an anti-aging agent or whitening agent. In the case of the agent, it is preferably used as a skin external preparation. As a dosage form, when it is administered topically (external skin material), a solution system, a dissolution system, an emulsification system, a powder dispersion system, a water-oil two-layer system, and a water-oil-powder three-layer system can be used. Patches, ointments, creams, lotions, lotions, gels, aerosols, etc. In the case of oral administration, examples include: tablets, coated tablets, sugar-coated tablets, granules. A solid preparation such as a preparation, a powder, a capsule (for example, a hard gelatin capsule or a soft gelatin capsule), or a liquid preparation (solution, suspension) such as an internal liquid preparation or a syrup preparation. In the case of non-oral administration, a form such as an injection solution can be mentioned.

In the present specification, "whitening" means inhibiting the blackening of the skin and improving melasma, freckles, dark spots, etc. by promoting the production of acesulfate heparin, which is caused by the following reasons: Decomposition and reduction of acesulfate heparin in the basement membrane, activation of melanocytes, and the accumulation of melanosomes in keratinocytes. In the present invention, the "whitening method" is intended for beauty purposes unless otherwise specified, but depending on the circumstances, there is a case of medical treatment.

In the present specification, the term "anti-aging" means inhibiting the skin change accompanying the heparin sulfate-binding growth factor by promoting the production of heparin sulfate, specifically, inhibiting epidermal differentiation abnormality, dermal angiogenesis, lymphatic Tube expansion, elastin decomposition, thereby preventing and improving wrinkles, sagging, and hardening of the skin, while maintaining a skin condition that is elastic and delicate and healthy, and the skin changes accompanying the heparin sulfate-binding growth factor are caused by the following reasons : Decomposition and reduction of acetyl sulfate heparin of the basement membrane proteoglycan due to aging or photoaging. In the present invention, the "anti-aging method" is intended for beauty purposes unless otherwise specified. However, depending on the circumstances, there is a case of medical treatment.

The acesulfate heparin production promoter of the present invention is not limited to the use as a whitening agent, the use for anti-aging such as wrinkle improvement, and the like for promoting wound healing, such as improvement of chloasma, freckles, and dark spots. It can also be used for the treatment, improvement, prevention, normalization, etc. of various other states or symptoms associated with the decomposition, reduction, and deficiency of acesulfate heparin. Here, as a state or symptom associated with decomposition, reduction, or deficiency of acesulfate heparin, it is considered that there are abnormalities in cell differentiation, regulation of cell proliferation, morphogenesis of cells, cell follow-up, mutual recognition, and cell infiltration, etc. Examples thereof include proliferation, metastasis, and angiogenesis of cancer cells.

The present invention has been described above by way of specific examples, but the above specific examples are intended to be illustrative, and the invention may be practiced without departing from the scope of the invention.

Hereinafter, the present invention will be specifically described by way of examples, but the present invention is not limited to the following examples.

[Examples]

40,000 HaCaT cells (obtained: German Cancer Research Center, Heidelberg, Germany) as human epidermal cells were inoculated into each well of a 24-well plate, and DMEM containing 10% FBS (fetal bovine serum) was used. The medium was cultured for 3 days. The medium of each well was replaced with a sample containing only 0.0005% of the extract of the Madonna lily (acquired: GATTEFOSSE) as an active ingredient, and containing only 0.001% of glucosamine hydrochloride (TSI Health Sciences) as an active ingredient. A sample or a medium containing a mixture of 0.0005% of the extract of the Madonna lily and 0.001% of glucosamine hydrochloride as a sample of the active ingredient was further cultured for 2 days. After washing once with PBS (Phosphate Buffered Saline), 200 μl of the cell disrupted solution of the following composition was added to each well and piped, followed by freezing.

10 mmol/L Tris-HCl pH 7.4, 150 mmol/L NaCl, 2 mmol/L ethylenediaminetetraacetic acid, 250 lmol/L benzylsulfonium fluoride, 0.1 mmol/L N-B Isobutyleneimine, 0.3% Nonidet P40, 0.05% Triton X-100, 0.3% sodium deoxycholate, 0.1% sodium lauryl sulfate, 0.1% BSA (bovine serum Albumin, bovine serum albumin)

After 24 hours, the thawed sample was supplied to a heparin sulfate ELISA kit (Seikagaku Biobusiness Co., Ltd.) to determine the amount of heparin sulfate. The results are shown in Fig. 1. The amount of heparin sulfate was expressed as 100% when the culture was carried out using only DMEM medium containing 10% FBS without adding a sample. The data was statistically analyzed by t-test.

Table 1 shows the raw data of the results shown in Fig. 1 and the results of the statistical processing. The original data of the control group were 99.0565, 98.5847, 97.1694, and 105.189. The original data of the extract of the lily of the lily was 102.673, 100.157, 118.556, and 120.601. The P value of the student's t-test (both sides) was statistically processed. 0.108734", at a meaningful level of 5% (P = 0.05), 0.108734>0.05, that is, there is no significant difference between the two on the production of heparin sulfate.

Therefore, it is clear that the extract of the Madonna lily itself does not have any effect on the amount of heparin sulfate produced. Further, as shown in Fig. 1 or Table 1, it can be seen that a combination of the extract of the genus Lilium sinensis and the glucosamine which does not have the promoting activity of acesulfate heparin is used, firstly, at a meaningful level of 1% compared with the control group. Or a meaningful difference (P = 0.000004) at 5% to increase the activity of heparin sulfate production. It can be seen that compared with glucosamine hydrochloride alone, the combination of the extract of Madonna lily and glucosamine also has a meaningful difference at a meaningful level of 5% to increase the production of heparin sulfate (P=0.029460), and thus compared with the lily of the lily alone. There was a significant difference at a meaningful level of 1% or 5% to increase the production of heparin sulfate (P = 0.001386). Furthermore, it can be confirmed from the table that between the control group and glucosamine hydrochloride (P=0.000424), between the Madonna lily and glucosamine hydrochloride (P=0.025858), glucosamine hydrochloride and the extract of the Madonna lily + glucosamine ( P=0.029460), and all were P<0.01 or P<0.05, and there was a difference in significance.

As described above, in the case of the extract of the Madonna lily, there was no significant difference compared with the control group, which proved that although the heparinase was known to be inhibited, heparin sulfate was not necessarily produced. As for the mixture of the extract of the Madonna lily and the glucosamine hydrochloride, it is less meaningful than the control group, and it is also less meaningful than the glucosamine hydrochloride alone, and it can also be meaningful compared with the lily of the lily alone. difference. Therefore, it is considered that a combination of Madonna lily and glucosamine hydrochloride can exert a significant effect of promoting the production of acesulfate heparin.

[Industrial availability]

The acesulfate heparin production promoter of the present invention can efficiently promote the production of acesulfate heparin, and thus can be suitably used for the following purposes: for various states or symptoms associated with decomposition, reduction, and deficiency of acesulfate heparin Uses for improvement, treatment, prevention, and normalization, such as the use of chloasma, freckles, dark spots, etc. as a whitening agent; for anti-aging purposes such as wrinkle improvement; for promoting wound healing; For example, it can be suitably used for normalization of abnormalities in cell differentiation, regulation of cell proliferation, morphogenesis of cells, cell adhesion, mutual recognition, and cell infiltration.

Claims (4)

A heparin sulfate production promoter comprising a mixture of Lilium Candidum extract and glucosamine. The acesulfate heparin production promoter according to claim 1, wherein the glucosamine is glucosamine hydrochloride. A whitening agent containing the acesulfate heparin production promoter as claimed in claim 1 or 2 as a whitening ingredient. A prophylactic or inhibitor of wrinkle formation comprising the acesulfate heparin production promoter according to claim 1 or 2 as an anti-aging component.