JP5534654B2 - Anti-inflammatory agent - Google Patents

Description

  The present invention uses an antioxidant, an anti-inflammatory agent, an anti-aging agent, and a hair restorer containing a plant extract, and at least one of the above-mentioned antioxidant, anti-inflammatory agent, anti-aging agent, and hair restorer The present invention relates to an external preparation for skin and food and drink.

In recent years, active oxygen has attracted attention as a factor that oxidizes biological components, and its adverse effect on living organisms has become a problem. Reactive oxygen is generated in the process of energy metabolism in living cells, and superoxide [that is, superoxide anion (· O ₂ ⁻ ) generated by one-electron reduction of oxygen molecules, hydrogen peroxide (H ₂ O ₂ ), Singlet oxygen ( ¹ O ₂ ), hydroxy radical (.OH)], and the like. Such active oxygen is essential for the phagocytic sterilization mechanism and plays an important role in removing viruses and cancer cells.
However, excessive generation of the active oxygen attacks in vivo molecules constituting membranes and tissues in the living body and induces various diseases. Superoxide, which is produced in vivo and is a starting material for other active oxygen, is usually eliminated sequentially by the catalytic action of superoxide dismutase (SOD) contained in cells. However, when the production of superoxide is excessive or when the action of SOD is reduced, the superoxide is insufficiently erased and the superoxide concentration becomes high. This is one of the causes of tissue damage such as rheumatoid arthritis, Behcet's disease, myocardial infarction, stroke, cataract, spots, freckles, wrinkles, diabetes, arteriosclerosis, shoulder stiffness, coldness, skin aging, etc. It is considered.
Among these, since the skin is directly affected by environmental factors such as ultraviolet rays, superoxide is likely to be generated. Therefore, when the superoxide concentration is increased, biological tissues such as collagen are decomposed, denatured, or crosslinked. Or oxidize fats and oils to produce lipid peroxides that damage cells, forming skin wrinkles, causing skin aging, inflammation, and skin pigmentation (See Non-Patent Document 1).
Therefore, attempts have been made to obtain active oxygen scavenging substances, radical scavenging substances, hydrogen peroxide scavenging substances, etc. from natural products advantageous in terms of safety, and extracts of Brassica Brassica plants (see Patent Document 1), Efficacy has been confirmed for extracts of plants belonging to the genus Ryukyubekei (see Patent Document 2), extracts of scallops (see Patent Document 3), extracts of sulfur (see Patent Document 4), and the like.

  In addition, there are various causes and onset mechanisms of inflammatory diseases such as contact dermatitis (rash), psoriasis, pemphigus vulgaris, and various other skin diseases with rough skin. Causes include, for example, nitric oxide (NO) production, tumor necrosis factor (TNF-α) production, hyaluronidase activity, hexosaminidase (histamine) release, platelet aggregation, cyclooxygenase-2 (COX-2) activity Etc. are known.

Nitric oxide (NO) is a nitrogen oxide that causes air pollution, acid rain, and the like. In recent years, nitric oxide (NO) is a physiologically active substance that exhibits various functions in vivo, such as vascular endothelium-derived relaxing factor (EDRF), neurotransmitters, microorganisms in biological defense, and tumor cell injury factors. It has been reported. As a physiologically active substance, it is known that nitric oxide produced from macrophages protects against bacterial and viral infections. However, when a large amount of nitric oxide produced from the macrophages is biosynthesized, it is not non-toxic to the living body and causes destruction of the self-tissue, which may cause pathological conditions such as worsening inflammation, rheumatism and diabetes. . Also, large amounts of biosynthesized nitric oxide can cause vascular smooth muscle relaxation and excessive permeability, and can cause endotoxin shock due to a significant decrease in blood pressure.
In such inflammatory diseases, it is important to suppress excessive production of nitric oxide (NO). Examples of herbal medicines having an inhibitory effect on the production of nitric oxide include, for example, rosemary extract, carnosol, carnosic acid, coffee bean extract, primrose extract, auren extract, buckwheat extract, licorice extract, Inu rose extract, Senkyu extract, Tonin extract, Peonies extract, Yakuinin extract, Red grape extract (refer to Patent Document 5), Tang Dynasty, Cod root bark, Japanese continuation, Car fore, Toko Grass roots, half-branches, spikelets, flower buds (see Non-Patent Document 2), and the like have been reported.

  The TNF-α has been found as a factor that necroses a tumor, but recently it is considered to be a cytokine that plays a mediator role not only for tumors but also for regulating the function of normal cells. TNF-α plays an important role in the process from the onset to the end of inflammation, but its continuous and excessive production causes damage to tissues including the skin, and causes systemic fever and cachexia. And cause worsening of inflammation. As such inflammation, for example, chronic inflammatory diseases such as rheumatoid arthritis and osteoarthritis are representative. Therefore, it is important to suppress excessive production of TNF-α in pathological inflammation. Examples of such TNF-α production inhibitors include perilla extract (see Non-patent Document 3), Amaryllidaceae alkaloids ricolin and licorididinol (see Non-Patent Document 4), and the like.

Hyaluronate that maintains affinity for body tissues is decomposed by ultraviolet rays, enzymes, and the like in a water-containing system, and the water retention effect decreases with a decrease in molecular weight. In addition, hyaluronic acid exists as an intercellular tissue and is also involved in vascular permeability. Furthermore, hyaluronidase is considered to be involved in degranulation from mast cells by activation in mast cells. Therefore, by inhibiting the activity of hyaluronidase, which is a hydrolase of hyaluronic acid, it is possible to stabilize hyaluronic acid, prevent the release of various chemical mediators from mast cells, and expect anti-inflammation.
Examples of herbal medicines having an inhibitory action on hyaluronidase activity include, for example, an extract of a genus Osbeckia plant (see Patent Document 6), a Fuji tea extract (see Patent Document 7), a rosemary, a thyme extract, and a Melissa extract ( (See Patent Document 8).

  The histamine release is a phenomenon in which histamine in mast cells is released to the outside of the cells, and the released histamine causes an inflammatory reaction. For this reason, attempts have been made to prevent or treat allergic diseases and inflammatory diseases with substances that inhibit or suppress histamine release. As such histamine release inhibitors, for example, tranilast, sodium cromoglycate, baicalin, baicalein, promethazine hydrochloride and the like have been used. However, all of these substances have side effects and have problems in terms of safety.

Platelet aggregation leads to activation of phospholipase A _{2 in} the arachidonic acid cascade, whereby leukotriene B, prostaglandin E _{2 and the} like are released and become a inflammatory substance. For this reason, attempts have been made to prevent or treat allergic diseases and inflammatory diseases with substances that inhibit or suppress platelet aggregation. Examples of extracts of herbal medicines having an inhibitory action on platelet aggregation include, for example, an extract of a plant belonging to the genus Canalium (see Patent Document 9), an extract of Kangsanfu (see Patent Document 10), and a Fuji tea extract (Patent Document). 11)) and the like have been reported.

Inflammation is a complex reaction with symptoms such as redness, edema, fever, pain, and dysfunction. Microscopically, it consists of common reactions such as blood vessel reaction that causes plasma leakage, leukocyte infiltration, tissue destruction by inflammatory cells, and also causes systemic reactions involving the central nervous system such as fever and hyperalgesia There is. Prostaglandins play an important role in such individual reactions of inflammation, and it has become clear that cyclooxygenase-2, an inducible cyclooxygenase, is mainly involved in the production of prostaglandins during this inflammation. Yes.
For this reason, many cyclooxygenase activity inhibitors represented by aspirin are used for the purpose of preventing and preventing inflammatory reactions (see Non-Patent Document 5). As plant-derived cyclooxygenase activity inhibitors, α-mangostin and γ-mangostin in mangosteen peel extract are known (see Patent Document 12). Further, as a compound having an inhibitory action on cyclooxygenase-2 activity, 2-phenyl-1,2-benzisoselenazol-3 (2H) -one, a salt thereof, or a hydrate thereof is known (patent) Reference 13).

The epidermis and dermis of the skin are composed of epidermal cells, dermal fibroblasts, and extracellular matrices such as collagen and elastin that are outside these cells and support the skin structure. In young skin, the interaction between these skin tissues is kept constant, so that moisture retention, flexibility, elasticity, etc. are secured, and the skin is maintained in a fresh state with its tension and gloss. .
However, when there is an influence of external factors such as irradiation of ultraviolet rays (UV-A, UV-B), significant drying of air, excessive skin washing, contact with hydrogen peroxide, or aging progresses, collagen And the production of extracellular matrix such as elastin is reduced, and the elasticity is reduced by crosslinking. As a result, the skin's moisturizing function and elasticity are lowered, and the horny layer begins to exfoliate abnormally, so that the skin loses its tension and gloss, and exhibits aging symptoms such as roughness and wrinkles.
It is known that the changes associated with aging of the skin, that is, wrinkles, dullness, disappearance of texture, decrease in elasticity, etc., are associated with the decrease or degeneration of dermal matrix components such as collagen and elastin. Yes.

  Elastase is a hydrolase of elastin present in the dermis of the skin, but elastase may be overexpressed by UV exposure or aging, and if elastin is denatured or destroyed by elastase, the elasticity of the skin decreases. It is considered.

In recent years, a factor that induces reduction or denaturation of dermal matrix components includes a group of proteolytic enzymes called matrix metalloproteases (hereinafter sometimes referred to as “MMPs”).
The MMPs are classified into (1) collagenase group (MMP-1, MMP-8 and MMP-13), (2) gelatinase group (MMP-2 and MMP-9), (1) due to differences in primary structure and substrate specificity. 3) Stromelysin group (MMP-3 and MMP-10), (4) Membrane-bound matrix metalloprotease group (MMP-14, MMP-15, MMP-16, and MMP-17), (5) Others ( MMP-7, MMP-11, and MMP-12) (see Patent Document 14).
Among the MMPs, MMP-1 and MMP-14 are known as enzymes that degrade type I collagen, type II collagen, and type III collagen, which are main components of the dermal matrix of the skin. In addition, the expression is greatly increased by the irradiation of ultraviolet rays, which contributes to decrease or denaturation of collagen by ultraviolet rays, and is considered to be a major factor such as the formation of wrinkles on the skin.

  The stratum corneum is formed from keratinocytes formed by terminal differentiation of epidermal keratinocytes and intercellular lipids that fill the space between cells. Intercellular lipids mainly composed of ceramide have a stratum corneum barrier function by forming a lamellar structure. On the other hand, keratinocytes are composed mainly of keratin fibers and covered with a hydrophobic and strong cell membrane-like structure called a cornified envelope (keratin thickened membrane, hereinafter referred to as “CE”) that is a membrane lining protein. . CE is formed by cross-linking and insolubilizing a plurality of CE precursor proteins such as involucrin and loricrin produced in the cells according to the differentiation of epidermal keratinocytes, and this CE is a barrier function of the skin. Intimately involved. Furthermore, ceramide and the like are covalently bonded to a part of them, and the base of the lamellar structure of intercellular lipids is supplied by taking a hydrophobic structure, thereby forming the basis of the stratum corneum barrier function and the skin water retention function .

  However, when the turnover speed becomes abnormal due to the effects of aging, drying, ultraviolet rays (UV-A, UV-B), etc., so-called keratinization, in which the lamellar structure is disturbed or CE is incompletely formed. Is induced, and the structure of corneocytes and intercellular lipids is abnormal, and the water retention function and barrier function of the stratum corneum are reduced. This is thought to lead to skin aging symptoms such as rough skin and dry skin. In patients with psoriasis and atopic dermatitis, immature CE is frequently observed in the skin area where the barrier function is reduced, and the correct formation of CE is very important for the barrier function of the skin. (See Non-Patent Document 6).

  Moreover, the hair repeats growth and loss according to a periodic hair cycle (hair cycle) consisting of a growth period, a regression period, and a rest period. Of these hair cycles, the stage at which new hair follicles are formed from the resting stage to the growing stage is considered to be the most important for hair growth. In this stage, the proliferation and differentiation of hair follicle epithelial cells are considered. It is believed that the dermal papilla cells play an important role. The dermal papilla cell is a cell located inside the hair follicle epithelial cell composed of outer root sheath cells and matrix cells in the vicinity of the hair root, and located in the root portion of the hair root wrapped in the basement membrane. It plays an important role in the growth and differentiation of hair follicle epithelial cells and the formation of hair, such as by acting on epithelial cells to promote their proliferation (see Non-Patent Document 7). The dermal papilla cell plays the most important role in the growth and differentiation of hair follicle epithelial cells and the formation of hair. The cultured dermal papilla cell is contacted with a target substance, and the presence or absence of the proliferation activity of the cell. A method for testing the hair-growth effect of the target substance by specifying strength is proposed (see Patent Document 15). As herbal medicines having such a dermal papilla cell growth promoting action, for example, an oyster extract, an auren extract, an anemone extract, etc. are known (see Patent Document 16 and Patent Document 17).

  In addition, many steroid hormones are expressed in the molecular form secreted from the production organ and bind to the receptor to express its action. In the case of the male hormones collectively called androgens, for example, testosterone enters the cells of the target organ. It is reduced to 5α-dihydrotestosterone (5α-DHT) by testosterone 5α-reductase, and then binds to the receptor and develops an action as an androgen.

The androgen is an important hormone, but if it acts excessively, it can be affected by various factors such as androgenetic alopecia, hirsutism, seborrhea, acne (such as acne), benign prostatic hyperplasia, prostate tumor, premature male sexuality, etc. Induces undesirable symptoms. Therefore, conventionally, in order to improve these various symptoms, a method of suppressing the action of excess androgen, specifically, by inhibiting the action of testosterone 5α-reductase that reduces testosterone to active 5α-DHT, There have been proposed a method for suppressing the generation of active 5α-DHT and a method for preventing the expression of androgen activity by inhibiting the binding of 5α-DHT generated from testosterone to the receptor.
As a plant extract having an action of inhibiting the binding between 5α-DHT and its receptor, for example, an extract of at least one of majito and cuta is known (see Patent Document 18).

  However, to date, it is a natural product that is easy to obtain, inexpensive, and highly safe, and does not adversely affect the quality of the additive in terms of taste, smell, feeling of use, etc. Antioxidants, anti-inflammatory agents, anti-aging agents, and hair restorers that can be widely used in external preparations and foods and drinks have not yet been provided, and the immediate provision thereof is strongly demanded.

JP 2003-81848 A JP 2005-29483 A JP 2006-321730 A JP 2007-8902 A JP 2002-87975 A Japanese Patent Laid-Open No. 2003-55242 JP 2003-12532 A JP-A-8-333267 JP 2002-53478 A JP 2002-53477 A JP 2001-97873 A JP 2002-47180 A JP 2000-16935 A JP 2000-344672 A JP-A-10-229978 JP-A-9-208431 Japanese Patent Laid-Open No. 11-12134 JP 2002-241297 A “Fragrance Journal” Special Issue No. 14, p156, 1995 “Wakan Journal of Pharmaceutical Sciences”, Vol. 15, p. 302-303, issued in 1998 “Inflammation”, 1993, Vol. 4, p.337-340 “Pharmaceutical Journal” 2001, Vol. 2, p.167-171 Pharmacology Atlas (P184), translated by Takehiko Fukuhara, Bunkodo Experimental Dermatology 12: 591-601 (2003) “Trends Genet”, 1992, Vol. 8, pp. 56-61

  An object of the present invention is to solve the conventional problems and achieve the following objects. That is, the present invention has an excellent antioxidant action and a highly safe antioxidant, an excellent anti-inflammatory action, a highly safe anti-inflammatory agent, and an excellent anti-aging action. And a highly safe anti-aging agent, a hair restoring agent having an excellent hair restoring action and a high safety, and at least one of the above antioxidant, anti-inflammatory agent, anti-aging agent, and hair restoring agent It aims at providing the skin external preparation and food / beverage products using this.

The present inventors have conducted extensive studies to solve the above problems, Mirintonia-Horutenshisu (Millingtonia hortensis), Ria macro filler (Leea macrophylla), Kodariokarikusu-Motoriusu (Codariocalyx motorius), Martini à An'nua ( Martynia annua), Pittosuporumu-Napaurense (Pittosporum napaulense), Hesuperetusa-Kurenurata (Hesperethusa crenulata), Puterosuperumumu-Dibe Cie Folie Umm (Pterospermumu diversifolium), Gurewia-Eriokarupa (Grewia eriocarpa), Tsunberugia-Raurifu Each plant extract rear (Thunbergia laurifolia), and Puremuna integrase Li cordifolia (Premna integrifolia) is excellent antioxidant, anti-inflammatory, anti-aging effects, and found to have a hair-growing action, completed the present invention It came to do.

The present invention is based on the above findings of the present inventors, and means for solving the above problems are as follows. That is,
<1> Mirintonia & Horutenshisu extract (Millingtonia hortensis), extract of Ria macro filler (Leea macrophylla), extract of Kodariokarikusu-Motoriusu (Codariocalyx motorius), extract of Martini A-An'nua (Martynia annua), Pittosuporumu · Napaurense extract (Pittosporum napaulense), extract of Hesuperetusa-Kurenurata (Hesperethusa crenulata), extract of Puterosuperumumu-Dibe Resid Folie um (Pterospermumu diversifolium), extract of Gurewia-Eriokarupa (Grewia eriocarpa), Tsunberugia - Extract Rauriforia (Thunbergia laurifolia), and an antioxidant, characterized in that it contains at least one member selected from extracts of Puremuna integrase Li cordifolia (Premna integrifolia).
<2> The antioxidant according to <1>, which has at least one of a superoxide dismutase (SOD) -like action, a membrane lipid peroxidation inhibiting action by singlet oxygen, a hydrogen peroxide scavenging action, and a radical scavenging action It is.
<3> Mirintonia & Horutenshisu extract (Millingtonia hortensis), extract of Ria macro filler (Leea macrophylla), extract of Kodariokarikusu-Motoriusu (Codariocalyx motorius), extract of Martini A-An'nua (Martynia annua), Pittosuporumu · Napaurense extract (Pittosporum napaulense), extract of Hesuperetusa-Kurenurata (Hesperethusa crenulata), extract of Puterosuperumumu-Dibe Resid Folie um (Pterospermumu diversifolium), extract of Gurewia-Eriokarupa (Grewia eriocarpa), Tsunberugia - Extract Rauriforia (Thunbergia laurifolia), and an anti-inflammatory agent characterized in that it contains at least one member selected from extracts of Puremuna integrase Li cordifolia (Premna integrifolia).
<4> Nitric oxide (NO) production inhibitory action, tumor necrosis factor (TNF-α) production inhibitory action, hyaluronidase activity inhibitory action, hexosaminidase release inhibitory action, platelet aggregation inhibitory action, and cyclooxygenase-2 (COX- 2) The anti-inflammatory agent according to <3>, which has at least one of activity inhibitory actions.
Extract of <5> Mirintonia-Horutenshisu (Millingtonia hortensis), extract of Ria macro Philadelphia (Leea macrophylla), an extract of Kodariokarikusu-Motoriusu (Codariocalyx motorius), extract of Martigny a-An'nua (Martynia annua), Pittosuporumu · Napaurense extract (Pittosporum napaulense), extract of Hesuperetusa-Kurenurata (Hesperethusa crenulata), extract of Puterosuperumumu-Dibe Resid Folie um (Pterospermumu diversifolium), extract of Gurewia-Eriokarupa (Grewia eriocarpa), Tsunberugia - Extract Rauriforia (Thunbergia laurifolia), and an anti-aging agent characterized by containing at least one selected from extracts of Puremuna integrase Li cordifolia (Premna integrifolia).
<6> Elastase activity inhibitory action, MMP-1 activity inhibitory action, skin fibroblast proliferation promoting action, epidermal keratinocyte proliferation promoting action, UV-B damage recovery action, and hydrogen peroxide damage inhibiting action The anti-aging agent according to <5> having any of the above.
Extract of <7> Mirintonia-Horutenshisu (Millingtonia hortensis), extract of Ria macro Philadelphia (Leea macrophylla), an extract of Kodariokarikusu-Motoriusu (Codariocalyx motorius), extract of Martigny a-An'nua (Martynia annua), Pittosuporumu · Napaurense extract (Pittosporum napaulense), extract of Hesuperetusa-Kurenurata (Hesperethusa crenulata), extract of Puterosuperumumu-Dibe Resid Folie um (Pterospermumu diversifolium), extract of Gurewia-Eriokarupa (Grewia eriocarpa), Tsunberugia - Extract Rauriforia (Thunbergia laurifolia), and a hair-growing agent characterized by containing at least one selected from Puremuna integrase extract Li Folia (Premna integrifolia).
<8> The hair restorer according to <7>, which has at least one of an androgen receptor antagonistic action and a hair papillary cell proliferation promoting action.
<9> The antioxidant according to any one of <1> to <2>, the anti-inflammatory agent according to any one of <3> to <4>, and any one of <5> to <6> A skin external preparation characterized by containing at least one of the anti-aging agent described in 1 above and the hair restorer described in any one of <7> to <8> above.
<10> The antioxidant according to any one of <1> to <2>, the anti-inflammatory agent according to any one of <3> to <4>, and any one of <5> to <6> A food or drink comprising the anti-aging agent according to item 1 and at least one of the hair-restoring agent according to any one of <7> to <8>.

  According to the present invention, it is possible to solve the conventional problems, achieve the above-mentioned object, have an excellent antioxidant action, and have a high safety, an excellent anti-inflammatory action, and Anti-inflammatory agent with high safety, anti-aging agent having excellent anti-aging action and high safety, hair-restoring agent having excellent hair-growth action and high safety, and the antioxidant Further, it is possible to provide an external preparation for skin and a food or drink using at least one of an anti-inflammatory agent, an anti-aging agent, and a hair restorer.

(Antioxidants, anti-inflammatory agents, anti-aging agents, and hair restorers)
Antioxidants of the present invention, anti-inflammatory agents, anti-aging agents, and hair growth agents, extracts of Mirintonia-Horutenshisu (Millingtonia hortensis), extract of Ria macro filler (Leea macrophylla), Kodariokarikusu-Motoriusu (Codariocalyx motorius) extract, extract of Martini a-An'nua (Martynia annua), extract of Pittosuporumu-Napaurense (Pittosporum napaulense), extract of Hesuperetusa-Kurenurata (Hesperethusa crenulata), extracted Puterosuperumumu-Dibe Cie Folie um of (Pterospermumu diversifolium) thing, Gurewia-Eriokarupa (Grewia extract eriocarpa), extract of Tsunberugia-Rauriforia (Thunbergia laurifolia), and Puremuna integrase Li cordifolia (and also contains at least one member selected from extracts of Premna integrifolia), other if necessary It contains.

The antioxidant has an antioxidant action based on at least one of a superoxide dismutase (SOD) -like action, a membrane lipid peroxidation inhibiting action by singlet oxygen, a hydrogen peroxide scavenging action, and a radical scavenging action. is there.
The anti-inflammatory agent includes nitric oxide (NO) production inhibitory action, tumor necrosis factor (TNF-α) production inhibitory action, hyaluronidase activity inhibitory action, hexosaminidase release inhibitory action, platelet aggregation inhibitory action, and cyclooxygenase-2 (COX-2) It has an anti-inflammatory action based on at least one of the activity inhibitory actions.
The anti-aging agent inhibits elastase activity, inhibits MMP-1 activity, promotes dermal fibroblast proliferation, promotes epidermal keratinocyte proliferation, recovers from UV-B damage, and suppresses damage to hydrogen peroxide. It has an anti-aging action based on at least one of the actions.
The hair-restoring agent has a hair-restoring action based on at least one of an androgen receptor antagonistic action and a hair papillary cell proliferation promoting action.
The Mirintonia-Horutenshisu (Millingtonia hortensis), Ria macro filler (Leea macrophylla), Kodariokarikusu-Motoriusu (Codariocalyx motorius), Martini A-An'nua (Martynia annua), Pittosuporumu-Napaurense (Pittosporum napaulense), Hesuperetusa-Kurenurata (Hesperethusa crenulata) , Puterosuperumumu-Dibe Cie Folie Umm (Pterospermumu diversifolium), Gurewia-Eriokarupa (Grewia eriocarpa), Tsunberugia-Rauriforia (Thunbergia laurifolia), and (In this specification, collectively referred is to a "plant") Lemna integrase Li cordifolia (Premna integrifolia) each extract contains, antioxidant, anti-inflammatory, anti-aging effects, and hair restoring action The details of the substances that exhibit the above are unclear, but each plant extract has such an excellent action and is useful as an antioxidant, anti-inflammatory agent, anti-aging agent, and hair restorer. This is a new finding by the present inventors that has not been known at all.

The Millingtonia Hortensis ( Millingtonia hortensis ) is a plant of the genus Cranaceae, also known as a sister tree or cork nousen, distributed in Thailand, China, India, Sri Lanka, etc., and easily available from these regions It is.
The site of Miltonia Hortensis used as an extraction raw material is not particularly limited and may be appropriately selected depending on the intended purpose. For example, flowers, persimmons, fruits, pericarps, seeds, seed coats, stems, leaves, branches , Branches, leaves, stems, bark, roots, rhizomes, root barks, and mixtures thereof. Among these, bark and roots are preferable.
It said Ria macro Philadelphia (Leea macrophylla) is a plant of pisonia umbellifera family, Southeast Asia - Africa, China, are distributed in India, are readily available from these areas.
There is no particular limitation on the part of the above-mentioned Leia macrophylla used as an extraction raw material, and it can be appropriately selected according to the purpose. Examples thereof include branches, branches and leaves, trunks, bark, roots, rhizomes, root barks, and mixtures thereof. Among these, leaves are preferable.
Said Kodariokarikusu-Motoriusu (Codariocalyx motorius) is a leguminous plant, also known as Maihagi (Maihagi), are distributed in southern China from Southeast Asia, it is readily available from these areas.
The part of the Kodario calix motrius used as an extraction raw material is not particularly limited and may be appropriately selected depending on the intended purpose. For example, flowers, persimmons, fruits, pericarps, seeds, seed coats, stems, leaves, branches , Branches, leaves, stems, bark, roots, rhizomes, root barks, and mixtures thereof. Among these, leaves and stems are preferable.
The Martini A-An'nua (Martynia annua) is a Matinia family of plants, which is also known as Matinia, are distributed from Central America to the Caribbean coastal region, it is readily available from these areas.
The part of the Martinia Annua used as an extraction raw material is not particularly limited and may be appropriately selected depending on the intended purpose.For example, flowers, persimmons, fruits, pericarps, seeds, seed coats, stems, leaves, Examples thereof include branches, branches and leaves, trunks, bark, roots, rhizomes, root barks, and mixtures thereof. Among these, leaves, stems and roots are preferable.
The Pittosporum napaurense is a plant belonging to the family Toberaceae and is distributed from the Asia-Pacific region such as South and Southeast Asia to Africa, and is easily available from these regions.
The part of the pitsporum napaurense used as an extraction raw material is not particularly limited and may be appropriately selected depending on the intended purpose. For example, flowers, persimmons, fruits, pericarps, seeds, seed coats, stems, leaves, branches , Branches, leaves, stems, bark, roots, rhizomes, root barks, mixtures thereof, and the like. Among these, bark is preferable.
The Hesperetusa crenurata is a citrus family plant also known as citrus, distributed in northern India, Myanmar, southern China and Thailand, and readily available from these regions.
The site of the Hesperetusa krenurata used as an extraction raw material is not particularly limited and may be appropriately selected depending on the intended purpose. For example, flowers, persimmons, fruits, pericarps, seeds, seed coats, stems, leaves, branches , Branches, leaves, stems, bark, roots, rhizomes, root barks, and mixtures thereof. Among these, stems and leaves are preferable.
The Pterosperumum diversifolium is a plant belonging to the family Aogiriaceae and is distributed in Southeast Asia and is easily available from these regions.
The part of Pterosperumum diversifolium used as an extraction raw material is not particularly limited and can be appropriately selected according to the purpose. For example, flowers, persimmons, fruits, pericarps, seeds, seed coats, stems, Examples thereof include leaves, branches, branches and leaves, trunks, bark, roots, rhizomes, rhizomes, and mixtures thereof. Among these, leaves and stems are preferable.
It said Gurewia-Eriokarupa (Grewia eriocarpa) is a linden family of plants, which is also known as the large leaf Tosakana tree, Taiwan, China, Southeast Asia, are distributed in India, are readily available from these areas.
The part of the Grevia eriocarpa used as an extraction raw material is not particularly limited and may be appropriately selected depending on the intended purpose. For example, flowers, persimmons, fruits, pericarps, seeds, seed coats, stems, leaves, branches , Branches, leaves, stems, bark, roots, rhizomes, root barks, and mixtures thereof. Among these, leaves and bark are preferable.
The Thunbergia laurifolia ( Thumbergia laurifolia ) is a plant belonging to the family Vulgare , also known as Laurel Kazura, and is native to the Malay Peninsula from Myanmar and is readily available from these regions.
The portion of the Tumbergia laurifolia used as an extraction raw material is not particularly limited and may be appropriately selected depending on the intended purpose.For example, flowers, persimmons, fruits, pericarps, seeds, seed coats, stems, leaves, Examples thereof include branches, branches and leaves, trunks, bark, roots, rhizomes, root barks, and mixtures thereof. Of these, bark is preferable.
The Premna integrofolia ( Premna integrifolia ) is a plant belonging to the family Oleaceae, also known as the Taiwan whale, which is distributed in southern China, Southeast Asia and India, and is easily available from these regions.
The premuna integrofolia used as an extraction raw material is not particularly limited and may be appropriately selected depending on the intended purpose. For example, flowers, persimmons, fruits, pericarps, seeds, seed coats, stems, leaves , Branches, leaves and leaves, trunks, bark, roots, rhizomes, root barks, mixtures thereof, and the like. Among these, bark is preferable.

  Each plant as the extraction raw material can be subjected to solvent extraction, for example, after being dried, in the state as it is or after being pulverized using a crusher or the like. Among them, the extraction raw material is preferably dried and pulverized immediately after collection. The drying may be performed, for example, in the sun or using a commonly used dryer. In addition, you may use each said plant as an extraction raw material, after giving pretreatments, such as a degreasing | defatting, with nonpolar solvents, such as hexane and benzene. By performing pretreatment such as degreasing, the extraction treatment of each plant with a polar solvent can be performed efficiently.

  Each plant extract can be easily obtained by utilizing a method generally used for plant extraction. Moreover, you may use a commercial item as said plant extract. In addition, the extract of each plant includes any of the extract of each plant, a diluted or concentrated solution of the extract, a dried product of the extract, or a crudely purified product or a purified product thereof. It is.

  There is no restriction | limiting in particular as a solvent used for the said extraction, According to the objective, it can select suitably, For example, water, a hydrophilic organic solvent, or these mixed solvents are room temperature or the temperature below the boiling point of a solvent. It is preferable to use it. Ingredients exhibiting an antioxidant action, an anti-inflammatory action, an anti-aging action, and a hair growth action contained in each plant can be easily extracted by an extraction treatment using a polar solvent as an extraction solvent.

  The water that can be used as the extraction solvent is not particularly limited and can be appropriately selected depending on the purpose. For example, pure water, tap water, well water, mineral water, mineral water, hot spring water, spring water, fresh water, etc. In addition, those subjected to various treatments are included. Examples of the treatment applied to water include purification, heating, sterilization, filtration, ion exchange, adjustment of osmotic pressure, buffering, and the like. Therefore, the water that can be used as the extraction solvent includes purified water, hot water, ion exchange water, physiological saline, phosphate buffer, phosphate buffered saline, and the like.

  The hydrophilic organic solvent that can be used as the extraction solvent is not particularly limited and may be appropriately selected depending on the intended purpose. Examples thereof include lower ones having 1 to 5 carbon atoms such as methanol, ethanol, propyl alcohol, and isopropyl alcohol. Alcohols: lower aliphatic ketones such as acetone and methyl ethyl ketone; polyhydric alcohols having 2 to 5 carbon atoms such as 1,3-butylene glycol, propylene glycol, glycerin, etc., and a mixed solvent of the hydrophilic organic solvent and water Etc. can also be used. In addition, when using the mixed solvent of the said water and the said hydrophilic organic solvent, 1-90 mass parts with respect to 10 mass parts of water in the case of a lower alcohol, and 10 mass parts of water in the case of a lower aliphatic ketone. It is preferable to use what mixed 1-40 mass parts with respect to this. Moreover, in the case of polyhydric alcohol, it is preferable to use what mixed 1-90 mass parts with respect to 10 mass parts of water.

When extracting an extract having an antioxidant effect, an anti-inflammatory effect, an anti-aging effect, and a hair-growth effect from each plant that is an extraction raw material, it is not necessary to adopt a special extraction method at room temperature or under reflux heating. It can be extracted using any extraction device.
Specifically, each of the extraction raw materials is put into a treatment tank filled with an extraction solvent, and the mixture is allowed to stand for 30 minutes to 4 hours with occasional stirring as necessary to elute soluble components, followed by filtration. Then, the solid matter is removed, the extraction solvent is distilled off from the obtained extract, and the extract can be obtained by drying. The amount of the extraction solvent is usually 5 to 15 times (mass ratio) of the extraction raw material. The extraction conditions are usually about 1 to 4 hours at 50 to 95 ° C. when water is used as the extraction solvent. Moreover, when using the mixed solvent of water and ethanol as an extraction solvent, it is about 30 minutes-about 4 hours at 40-80 degreeC normally. The extract obtained by extraction with a solvent is used as an active ingredient of the antioxidant, anti-inflammatory agent, anti-aging agent, and hair restorer of the present invention as long as the extraction solvent is highly safe. be able to.

  The extract of each plant obtained by extraction is diluted, concentrated and dried according to a conventional method in order to obtain a diluted or concentrated solution of the extract, a dried product of the extract, or a crude purified product or purified product thereof. Further, treatment such as purification may be performed. The obtained extract of each plant can be used as an active ingredient of an antioxidant, an anti-inflammatory agent, an anti-aging agent, and a hair restorer as it is, but the concentrated solution or a dried product thereof can be used. Is easy to use. In obtaining the dried extract, a conventional method can be used, and carriers such as dextrin and cyclodextrin may be added to improve hygroscopicity. In addition, each plant as an extraction raw material may have a specific odor and taste. Therefore, the extract of each plant is decolorized and deodorized within a range not causing a decrease in physiological activity. However, since it is not used in large quantities, for example, when it is added to skin cosmetics, there is no practical problem even if it is not purified. Specifically, purification can be performed by activated carbon treatment, adsorption resin treatment, ion exchange resin treatment, or the like.

The plant extracts obtained as described above have superoxide dismutase (SOD) -like action, membrane lipid peroxidation inhibition action by singlet oxygen, hydrogen peroxide scavenging action, radical scavenging action, and one Nitric oxide (NO) production inhibitory action, tumor necrosis factor (TNF-α) production inhibitory action, hyaluronidase activity inhibitory action, hexosaminidase release inhibitory action, platelet aggregation inhibitory action, and cyclooxygenase-2 (COX-2) activity inhibitory action And elastase activity inhibitory action, MMP-1 activity inhibitory action, skin fibroblast proliferation promoting action, epidermal keratinocyte proliferation promoting action, UV-B damage recovery action, and hydrogen peroxide damage inhibiting action, And at least one of androgen receptor antagonistic action and hair papillary cell proliferation promoting action And, based on these effects, the antioxidant of the present invention, anti-inflammatory agents, anti-aging agents, and those suitably usable as an active ingredient of hair tonics.
In addition, the extract of each plant is based on each of the above-mentioned actions, a superoxide dismutase (SOD) -like agent, a membrane lipid peroxidation inhibitor by singlet oxygen, a hydrogen peroxide scavenger, and a radical scavenger, Furthermore, nitric oxide (NO) production inhibitor, tumor necrosis factor (TNF-α) production inhibitor, hyaluronidase activity inhibitor, hexosaminidase release inhibitor, platelet aggregation inhibitor, and cyclooxygenase-2 (COX-2) ) Activity inhibitor, and elastase activity inhibitor, MMP-1 activity inhibitor, dermal fibroblast proliferation promoter, epidermal keratinocyte proliferation promoter, UV-B damage recovery agent, and hydrogen peroxide They can also be suitably used as damage inhibitors, androgen receptor antagonists, and hair papilla cell proliferation promoters.

The antioxidant action of the antioxidant of the present invention is based on at least one of superoxide dismutase (SOD) -like action, membrane lipid peroxidation inhibiting action by singlet oxygen, hydrogen peroxide scavenging action, and radical scavenging action. Demonstrated. Among these, it is preferable that the antioxidant has at least a radical scavenging action.
The anti-inflammatory action in the anti-inflammatory agent of the present invention includes a nitric oxide (NO) production inhibitory action, a tumor necrosis factor (TNF-α) production inhibitory action, a hyaluronidase activity inhibitory action, a hexosaminidase release inhibitory action, and a platelet aggregation inhibitory action. This is exhibited based on at least one of the action and the cyclooxygenase-2 (COX-2) activity inhibition action. Among these, the anti-inflammatory agent preferably has at least a nitric oxide (NO) production inhibitory action and a hexosaminidase release inhibitory action.
The anti-aging action of the anti-aging agent of the present invention includes elastase activity inhibitory action, MMP-1 activity inhibitory action, skin fibroblast proliferation promoting action, epidermal keratinocyte proliferation promoting action, recovery action from UV-B damage, and It is demonstrated based on at least one of the damage suppression effect with respect to hydrogen peroxide. Among these, it is preferable that the anti-aging agent has at least an MMP-1 activity inhibitory action.
The hair-restoring action of the hair-restoring agent of the present invention is exhibited based on at least one of an androgen receptor antagonistic action and a hair papillary cell proliferation promoting action. Among these, it is preferable that the hair restorer has at least a dermal papilla cell proliferation promoting action.

The content of the extract of each plant in the antioxidant, anti-inflammatory agent, anti-aging agent, hair restorer is not particularly limited and can be appropriately selected depending on the purpose. The agent, anti-inflammatory agent, anti-aging agent, and hair restorer may be the extracts of the plants.
Moreover, in the said antioxidant, anti-inflammatory agent, anti-aging agent, and hair restorer, the said plant extract may contain only 1 type, or 2 or more types may be contained. Good. The content ratio of each of the antioxidants, anti-inflammatory agents, anti-aging agents, and hair-restoring agents in the case where two or more plant extracts are contained is not particularly limited, and is appropriately selected according to the purpose. be able to.

Moreover, as other components other than the extracts of the respective plants, which can be contained in the antioxidant, anti-inflammatory agent, anti-aging agent, and hair restorer, as long as the effects of the present invention are not impaired, There is no restriction | limiting in particular, According to the objective, it can select suitably, For example, the physiological saline solution etc. for diluting the extract of each said plant to a desired density | concentration etc. are mentioned. Moreover, there is no restriction | limiting in particular also in content of the said other component in the said antioxidant, anti-inflammatory agent, anti-aging agent, and hair restorer, According to the objective, it can select suitably.
Further, the antioxidant, anti-inflammatory agent, anti-aging agent, and hair-restoring agent can be made into any dosage form such as powder, granule, tablet, etc. by formulating as necessary.

  The antioxidant, anti-inflammatory agent, anti-aging agent, and hair restorer of the present invention have excellent antioxidant action, anti-inflammatory action, anti-aging action, and hair growth action, and are excellent in safety. It is suitable for use in the external preparation for skin of the present invention and the food and drink of the present invention.

(External skin preparation)
The skin external preparation of the present invention contains at least one of the above-described antioxidant, anti-inflammatory agent, anti-aging agent, and hair restorer of the present invention, and further contains other components as necessary. Become.
Here, the external preparation for skin means various drugs applied to the skin, and the classification is not particularly limited, and includes, for example, a wide range of skin cosmetics, quasi drugs, pharmaceuticals and the like. It is a waste.
The external preparation for skin may be one obtained by blending the extract of each plant with any external preparation for skin so as not to interfere with the activity thereof, or the external preparation for skin mainly comprising the extract of each plant. An agent may be used. Moreover, the said skin external preparation may be the extract of each said plant itself.

  The external preparation for skin is not particularly limited and may be appropriately selected depending on the intended purpose.For example, an ointment, cream, milky lotion, serum, lotion, foundation, pack, jelly, lip balm, lipstick, bath preparation, Soap, body shampoo, astringent, hair tonic, hair lotion, hair cream, hair liquid, shampoo, pomade, rinse and the like.

  The other components are not particularly limited and may be appropriately selected depending on the purpose. Components usually used in the manufacture of a skin external preparation, such as astringents, bactericides, antibacterial agents, whitening agents, ultraviolet absorption Agent, moisturizer, cell activator, anti-aging agent, anti-inflammatory / anti-allergic agent, antioxidant / active oxygen remover, fats and oils, waxes, hydrocarbons, fatty acids, alcohols, esters, surfactants, Examples include fragrances.

  The content of the antioxidant, anti-inflammatory agent, anti-aging agent, and hair restorer in the external preparation for skin is not particularly limited and can be appropriately selected according to the type of external preparation for skin. For example, the amount of the extract of each plant is preferably 0.0001 to 10% by mass, and more preferably 0.001 to 5% by mass.

(Food)
The food / beverage product of the present invention contains at least one of the above-described antioxidant, anti-inflammatory agent, anti-aging agent, and hair restorer of the present invention, and further contains other components as necessary. .
Here, the food and drink are those which are less likely to harm human health and are taken by oral or gastrointestinal administration in normal social life. It is not limited to the category of goods, etc., and means, for example, a wide range of foods that are taken orally, such as general foods, health foods, health functional foods, quasi drugs, and pharmaceuticals.
The food and drink may be a mixture of any of the above plant extracts in any food or drink so as not to hinder the activity thereof, or a dietary supplement mainly composed of the above plant extracts. There may be. Moreover, the said food / beverage products may be the extract of each said plant itself.

  There is no restriction | limiting in particular as said food-drinks, According to the objective, it can select suitably, For example, drinks, such as a soft drink, a carbonated drink, a nutritive drink, a fruit drink, a lactic acid drink; Ice cream, ice sherbet, shaved ice, etc. Noodles such as buckwheat noodles, udon, harusame, gyoza skin, sweet husk, Chinese noodles, instant noodles; rice cakes, candy, gum, chocolate, tablet confectionery, snack confectionery, biscuits, jelly, jam, cream, baked confectionery, Sweets such as bread; crab, salmon, clams, tuna, sardines, shrimp, skipjack, mackerel, whale, oyster, saury, squid, red sea bream, scallops, abalone, sea urchin, crabs, tocobushi, etc .; kamaboko, ham, sausage Processed fishery and livestock products such as dairy products such as processed milk and fermented milk; salad oil, tempura oil, margarine, mayonnaise, and short nibs Fats and processed oils such as whipped cream and dressings; seasonings such as sauces and sauces; curry, stew, oyakodon, rice cakes, miso cooked rice, Chinese rice cakes, and rice cakes, tempura, eel rice, hayashi rice, oden, marvodorf, Retort pouch foods such as beef bowl, meat sauce, egg soup, omelet rice, dumplings, shumai, hamburger, meatballs; various forms of health foods and nutritional supplements; pharmaceuticals such as tablets, capsules, drinks, troches, etc. Examples include foreign products.

There is no restriction | limiting in particular as said other component, According to the objective, it can select suitably, For example, the auxiliary | assistant raw material or additive etc. which are normally used in manufacturing food-drinks are mentioned.
The auxiliary raw material or additive is not particularly limited and may be appropriately selected depending on the intended purpose. For example, glucose, fructose, sucrose, maltose, sorbitol, stevioside, rubusoside, corn syrup, lactose, citric acid , Tartaric acid, malic acid, succinic acid, lactic acid, L-ascorbic acid, dl-α-tocopherol, sodium erythorbate, glycerin, propylene glycol, glycerin fatty acid ester, polyglycerin fatty acid ester, sucrose fatty acid ester, sorbitan fatty acid ester, Arabia Examples include gum, carrageenan, casein, gelatin, pectin, agar, vitamin Bs, nicotinic acid amide, calcium pantothenate, amino acids, calcium salts, pigments, fragrances, preservatives and the like.

  The content of the antioxidant, anti-inflammatory agent, anti-aging agent, and hair restorer in the food and drink varies depending on the type of the food and drink to be treated, and cannot be generally defined, for example, In addition, when it is intended to be blended in any food or drink within a range that does not impair the original taste of the food or drink, the amount of each plant extract that is an active ingredient is 0.001% by mass to 50% by mass. Is preferable, and 0.01 to 20% by mass is more preferable. In addition, for example, when the purpose is to produce a nutritional supplement food or drink in the form of granules, tablets, capsules, etc., each of which contains an extract of each plant as a main component, the extract of each plant that is an active ingredient As a quantity, 0.01 mass%-100 mass% are preferable, and 5 mass%-100 mass% are more preferable.

(effect)
Antioxidants, anti-inflammatory agents, anti-aging agents, hair-restoring agents, skin external preparations and foods and beverages of the present invention can be used on a daily basis, and are extracts of the respective plants that are active ingredients. By this function, the antioxidant action, anti-inflammatory action, anti-aging action, and hair growth action can be exhibited extremely effectively.

  In addition, although the antioxidant, anti-inflammatory agent, anti-aging agent, hair restorer, skin external preparation, and food and drink of the present invention are suitably applied to humans, their effects are achieved. As long as it is possible, it is also possible to apply to animals other than humans (for example, mice, rats, hamsters, dogs, cats, cows, pigs, monkeys, etc.). In addition, the antioxidant, anti-inflammatory agent, anti-aging agent, hair restorer, and external preparation for skin and food and drink of the present invention are extracts of each plant derived from nature as active ingredients, and are safe. It is also advantageous in terms of excellent properties.

  Examples of the present invention will be described below, but the present invention is not limited to these examples.

(Production Example 1)
-Production of water extract of each plant-
To the pulverized product of each plant shown in Table 1 below, 10 times the amount of water was added by mass ratio, heated at 80 ° C. for 2 hours, and filtered. The same amount of water was added to the residue, heated at 80 ° C. for 2 hours, and filtered. The filtrate was concentrated and lyophilized to obtain an aqueous extract (lyophilized product) of each plant.

(Production Example 2)
-Production of 50 mass% ethanol extract of each plant-
To the pulverized product of each plant shown in Table 1 below, 10 times by mass 50% by mass ethanol was added, heated at 80 ° C. for 2 hours, and filtered. The same amount of 50% by mass ethanol was added to the residue, heated at 80 ° C. for 2 hours, and filtered. The filtrate was concentrated and freeze-dried to obtain a 50% by mass ethanol extract (lyophilized product) of each plant.
In addition, in each Example mentioned later, it tested using the 50 mass% ethanol extract of each plant obtained by this manufacture example 2 as a test sample. Table 1 below shows the “extraction rate” of the 50% by mass ethanol extract of each plant obtained and the “extract No.” of the 50% by mass ethanol extract of each plant obtained.

(Production Example 3)
-Production of 80 mass% ethanol extract of each plant-
To the pulverized product of each plant shown in Table 1 below, 10 times by mass (mass ratio) of 80 mass% ethanol was added, heated at 80 ° C. for 2 hours, and filtered. The same amount of 80% by mass ethanol was added to the residue, heated at 80 ° C. for 2 hours, and filtered. The filtrate was concentrated and freeze-dried to obtain an 80% by mass ethanol extract (lyophilized product) of each plant.

(Example 1: Superoxide dismutase (SOD) -like action test)
Each plant extract was used as a test sample, and the superoxide dismutase (SOD) -like action was tested by the following test method.

In a test tube, 0.05 mL / L sodium bicarbonate buffer (pH 10.2) 2.4 mL, 2 mmol / L xanthine 0.1 mL, 3 mmol / L EDTA 0.1 mL, 50 μg / mL bovine serum albumin ( BSA) 0.1 mL and 0.75 mmol / L nitro blue tetrazolium 0.1 mL were added. To this, 0.1 mL of each test sample solution was added and left at 25 ° C. for 10 minutes.
Next, 0.1 mL of xanthine oxidase solution was added, stirred rapidly, and reacted at 25 ° C. for 20 minutes. Thereafter, 0.1 mL of 6 mmol / L copper chloride was added to stop the reaction, and the absorbance at a wavelength of 560 nm was measured. A blank test was performed and corrected in the same manner.
The calculation method of the SOD like action (superoxide elimination rate (%)) is as follows.
Superoxide erasure rate (%) = {1− (St−Sb) / (Ct−Cb)} × 100
St: Absorbance at a wavelength of 560 nm of the test sample solution Sb: Absorbance at a wavelength of 560 nm of the test sample solution blank Ct: Absorbance at a wavelength of 560 nm of the control solution Cb: Absorbance at a wavelength of 560 nm of the control solution blank

  The results are shown in Table 2. In addition, Extract No. 1-2, 4-1, 4-2, 5, 6-1, 6-2, and 7-2 were used at a sample concentration of 100 μg / mL. Extract No. 1-1, 2, 7-1, 8-1, 8-2, 9, and 10 were used at a sample concentration of 25 μg / mL.

(Example 2: Peroxidation inhibition test of membrane lipid by singlet oxygen)
Each plant extract was used as a test sample, and the membrane lipid peroxidation inhibitory action of singlet oxygen was tested by the following test method.

10 mL of a clear sample bottle (made of glass), 5 mL of a 2% by weight erythrocyte suspension, 5 mL of PBS (−), or 5 mL of PBS (−) containing each test sample solution, and 10 mmol / L hematoporphyrin—20 mmol / L 0.01 mL of sodium hydroxide solution was added and mixed well. This reaction solution was irradiated uniformly on a merry-go-round with a 15 W halogen lamp for 35 minutes to generate ¹ O ₂ and hemolyze erythrocytes.
After completion of the reaction, 1 mL of the reaction solution was collected, and 2 mL of PBS (−) was added and mixed well. After centrifugation (1660 × g, 5 minutes, 4 ° C.), the absorbance of the supernatant at a wavelength of 540 nm was measured. As a control, 2 mL of purified water was added to 1 mL of the reaction solution to prepare a mixed solution in which red blood cells were completely hemolyzed. The absorbance of the mixture at a wavelength of 540 nm was measured. A blank test was performed and corrected in the same manner.
The calculation method of the peroxidation inhibitory action of membrane lipid by singlet oxygen is as follows.
Permeation inhibition rate of membrane lipid (%) = (1−B / A) × 100
A: Absorbance at a wavelength of 540 nm of completely dissolved blood B: Absorbance at a wavelength of 540 nm of the supernatant of the reaction solution

  The results are shown in Table 3. In addition, Extract No. 1-1, 1-2, 2, 4-2, 6-1, 7-1, 9, and 10 were used at a sample concentration of 200 μg / mL. Extract No. 4-1 was used at a sample concentration of 100 μg / mL. Extract No. 7-2 and 8-2 were used at a sample concentration of 50 μg / mL. Extract No. 8-1 was used at a sample concentration of 25 μg / mL.

(Example 3: Hydrogen peroxide scavenging action test)
Using each plant extract as a test sample, the hydrogen peroxide scavenging action was tested by the following test method.

After adding 10 μL of each test sample solution to 10 μL of a 1.5 mmol / L hydrogen peroxide solution and reacting at 37 ° C. for 20 minutes, a coloring solution [100 μmol / L DA-64, 0.5 mass% Triton X- 100 mL of 100 mols / L PIPES buffer (pH 7.0) (100 mL / mL peroxidase 1 mL was added) 2.98 mL was added and reacted at 37 ° C. for 5 minutes. After completion of the reaction, absorbance at a wavelength of 727 nm was measured. A blank test was performed and corrected in the same manner.
The calculation method of hydrogen peroxide scavenging action is as follows.
Hydrogen peroxide elimination rate (%) = {1− (St−Sb) / (Ct−Cb)} × 100
St: Absorbance at a wavelength of 727 nm of the test sample solution Sb: Absorbance at a wavelength of 727 nm of the test sample solution blank Ct: Absorbance at a wavelength of 727 nm of the control solution Cb: Absorbance at a wavelength of 727 nm of the control solution blank

  The results are shown in Table 4. In addition, Extract No. 1-1, 1-2, 2, 3, 4-1, were used at a sample concentration of 100 μg / mL.

(Example 4: Radical scavenging action test)
Each plant extract was used as a test sample, and the radical scavenging action was tested using 1,1-diphenyl-2-picrylhydradyl radical (DPPH), which is a very stable radical, by the following test method.

3 mL of each test sample solution was added to 3 mL of 150 μmol / L DPPH ethanol solution, the container was immediately sealed and shaken and allowed to stand for 30 minutes, and then the absorbance at a wavelength of 520 nm was measured. A blank test was performed and corrected in the same manner.
The calculation method of the DPPH radical scavenging action is as follows.
DPPH radical scavenging rate (%) = {C− (St−Sb)} / C × 100
St: Absorbance at a wavelength of 520 nm of the test sample solution Sb: Absorbance at a wavelength of 520 nm of the test sample solution blank C: Absorbance at a wavelength of 520 nm of the control solution

  The results are shown in Table 5. In addition, Extract No. 1-1, 1-2, 2, 3, 4-1, 4-2, 5, 6-1, and 6-2 were used at a sample concentration of 200 μg / mL. Extract No. 7-1, 7-2, 8-1, 8-2, 9, 10 were used at a sample concentration of 50 μg / mL.

From the results of Examples 1-4, extract of Mirintonia-Horutenshisu (Millingtonia hortensis), extract of Ria macro filler (Leea macrophylla), extract of Kodariokarikusu-Motoriusu (Codariocalyx motorius), Martini A-An'nua (Martynia annua extract), extract of Pittosuporumu-Napaurense (Pittosporum napaulense), extract of Hesuperetusa-Kurenurata (Hesperethusa crenulata), extract of Puterosuperumumu-Dibe Resid Folie um (Pterospermumu diversifolium), extracted Gurewia-Eriokarupa of (Grewia eriocarpa) , Extract of Tsunberugia-Rauriforia (Thunbergia laurifolia), and Puremuna integrase Li cordifolia extract (Premna integrifolia) is superoxide dismutase (SOD) mimetic actions peroxidation inhibitory effect of membrane lipids by singlet oxygen, peroxide It was confirmed to have at least one of hydrogen oxide scavenging action and radical scavenging action, and it was suggested that each of these plant extracts can be suitably used as an active ingredient of an antioxidant.

(Example 5: Nitric oxide (NO) production inhibitory effect test)
Each plant extract was used as a test sample, and the nitric oxide (NO) production inhibitory action was tested by the following test method.

Mouse macrophage cells (RAW264.7) were cultured using Dulbecco's MEM containing 10% fetal bovine serum (FBS), and then the cells were collected with a cell scraper. The collected cells are diluted with 10% FBS-containing phenol red-free Dulbecco MEM to a concentration of 3.0 × 10 ⁶ cells / mL, and then seeded at 100 μL per well in a 96-well microplate for 4 hours. Cultured. After completion of the culture, the medium was removed, 100 μL of a test sample solution dissolved in 10% by mass of FBS-containing phenol red-free Dulbecco MEM containing DMSO having a final concentration of 2% by mass was added to each well, and the final concentration was 1 μg / mL. 100 μL of lipopolysaccharide (LPS, E. coli 0111; B4, manufactured by DIFCO) dissolved in Dulbecco's MEM containing no phenol red containing FBS at a concentration of% FBS was added and cultured for 48 hours. The amount of NO production was measured using the amount of nitrite ion (NO ₂ ⁻ ) as an index. After completion of the culture, the same amount of grease reagent (1% by mass of sulfanilamide, 0.1% by mass of N-1-naphthyl ethylenedihydride in 5% by mass of phosphoric acid solution) was added to the culture solution in each well, Reacted for 10 minutes at room temperature. After the reaction, absorbance at a wavelength of 540 nm was measured.
The calculation method of NO production inhibitory action is as follows.
NO production inhibition rate (%) = {1− (A−B) / (C−D)} × 100
A: Absorbance at a wavelength of 540 nm when a test sample is added and LPS is stimulated B: Absorbance at a wavelength of 540 nm when a test sample is added and LPS is not stimulated C: Absorbance at a wavelength of 540 nm when LPS is stimulated by a control D: When LPS is not stimulated by a control Absorbance at a wavelength of 540 nm

  The results are shown in Table 6. In addition, Extract No. 1-1, 1-2, 2, 3, 4-1, 4-2, 5, 7-1, 8-1, 9, and 10 were used at a sample concentration of 200 μg / mL. Extract No. 6-1, 7-2, and 8-2 were used at a sample concentration of 50 μg / mL.

(Example 6: Tumor necrosis factor (TNF-α) production inhibitory effect test)
Using each plant extract as a test sample, the tumor necrosis factor (TNF-α) production inhibitory action was tested by the following test method.

Murine macrophage cells (RAW264.7) were cultured using Dulbecco's MEM containing 10% FBS, and then the cells were collected with a cell scraper. The collected cells were diluted with Dulbecco's MEM containing 10% FBS to a concentration of 1.0 × 10 ⁶ cells / mL, then seeded at 100 μL per well on a 96-well plate, and cultured for 4 hours. After completion of the culture, the medium was removed, 100 μL of a test sample dissolved in 10% FBS-containing Dulbecco MEM containing a final concentration of 2% DMSO was added to each well, and lipolysis dissolved in 10% FBS-containing Dulbecco MEM at a final concentration of 1 μg / mL. 100 μL of polysaccharide (LPS, E. coli 0111; B4, DIFCO) was added and cultured for 24 hours. After completion of the culture, the amount of TNF-α in the culture supernatant of each well was measured using a sandwich ELISA method.
The calculation method of the TNF-α production inhibitory action is as follows.
TNF-α production inhibition rate (%) = {(B−A) / B} × 100
A: TNF-α amount when test sample is added B: TNF-α amount when test sample is not added

  The results are shown in Table 7. In addition, Extract No. 1-1, 2, 4-1, 4-2, 5, 6-1, 6-2, 7-1, 7-2, 8-1, 9, and 10 were used at a sample concentration of 12.5 μg / mL. .

(Example 7: Hyaluronidase activity inhibitory action test)
Each plant extract was used as a test sample, and the hyaluronidase activity inhibitory action was tested by the following test method.

Add 0.1 mL of a hyaluronidase solution (Type IV-S (Type bovine testis; SIGMA 400 NF units / mL)) to 0.2 mL of 0.1 mol / L acetate buffer (pH 3.5) in which the test sample is dissolved. Then, 2.5 mL of 2.5 mmol / L calcium chloride was added as an activator and reacted for 20 minutes at 37 ° C. To this, 0.4 mg / mL potassium hyaluronate solution (from robster comb) was added. 5 mL was added and reacted for 40 minutes at 37 ° C. After that, 0.2 mL of 0.4 mol / L sodium hydroxide was added to stop the reaction and cooled, and then 0.2 mL of boric acid solution was added to each reaction solution for 3 minutes. After cooling with ice, 6 mL of p-DABA reagent was added and reacted at 37 ° C. for 20 minutes. The absorbance was measured at Length 585 nm. Corrected Perform a blank determination in the same manner.
The calculation method of the hyaluronidase activity inhibitory action is as follows.
Hyaluronidase activity inhibition rate (%) = {1− (St−Sb) / (Ct−Cb)} × 100
St: Absorbance at a wavelength of 585 nm of the test sample solution Sb: Absorbance at a wavelength of 585 nm of the test sample solution blank Ct: Absorbance at a wavelength of 585 nm of the control solution Cb: Absorbance at a wavelength of 585 nm of the control solution blank

  The results are shown in Table 8. In addition, Extract No. 5, 7-1, 7-2, 8-1, 8-2, 9, and 10 were used at a sample concentration of 400 μg / mL.

(Example 8: Hexosaminidase release inhibitory action test)
Each plant extract was used as a test sample, and the hexosaminidase release inhibitory action was tested by the following test method.

Rat basophil leukemia cells (RBL-2H3) were cultured using 15% FBS-added S-MEM, and then cells were collected by trypsin treatment. The collected cells are diluted with a medium to a concentration of 4.0 × 10 ⁵ cells / mL, added to a final concentration of 0.5 μg / mL of DNP-specific IgE, and then seeded at 100 μL per well on a 96-well plate, Cultured overnight. After completion of the culture, the medium was removed, and washing was performed twice with 500 μL of Siraganian buffer. Next, 30 μL of the same buffer solution and 10 μL of a test sample prepared with the same buffer solution were added, and the mixture was allowed to stand at 37 ° C. for 10 minutes. Thereafter, 10 μL of a 100 ng / mL DNP-BSA solution was added, and the mixture was allowed to stand at 37 ° C. for 15 minutes to release hexosaminidase. Thereafter, the release was stopped by allowing the 96-well plate to stand on ice. 10 μL of the cell supernatant of each well and 10 μL of 1 mmol / L p-NAG (p-nitrophenyl N-acetyl β-D-glucosamine) solution were added to a new 96-well plate and reacted at 37 ° C. for 1 hour. After completion of the reaction, 250 μL of 0.1 mol / L Na ₂ CO ₃ / NaHCO ₃ was added to each well, and the absorbance at a wavelength of 415 nm was measured. Moreover, as a blank test, the absorbance at a wavelength of 415 nm of a mixed solution of 10 μL of cell supernatant and 250 μL of 0.1 mol / L Na ₂ CO ₃ / NaHCO ₃ was measured and corrected.
The calculation method of the hexosaminidase release inhibitory action is as follows.
Inhibition rate of hexosaminidase release (%) = {1− (BC) / A} × 100
A: Absorbance at a wavelength of 415 nm when no test sample is added B: Absorbance at a wavelength of 415 nm when a test sample is added C: Absorbance at a wavelength of 415 nm when a test sample is added and p-NAG is not added

  The results are shown in Table 9. In addition, Extract No. 1-1, 1-2, 2, 3, 4-1, 4-2, 5, 6-1, 6-2, 7-1, 7-2, 8-1, 8-2, 9, 10 Used at a sample concentration of 400 μg / mL.

(Example 9: Platelet aggregation inhibitory effect test)
Each plant extract was used as a test sample, and the platelet aggregation inhibitory action was tested by the following test method.

First, rabbit blood collected by adding 1/10 volume of heparin sodium injection solution (Japanese Pharmacopoeia) was centrifuged (180 × g, 10 minutes, room temperature), and platelet suspension (Platelet Rich Plasma; PR) P.).
Next, 1 μL of a 200 mmol / L calcium chloride solution was added to 223 μL of platelet suspension (PRP) and reacted at 37 ° C. for 1 minute. 1 μL of the test sample solution was added thereto, reacted for another 2 minutes, and after stirring for 1 minute with a stir bar added, 25 μL of collagen solution was added and the platelet aggregation rate was measured at 37 ° C. for 10 minutes. Separately, the platelet aggregation rate was measured in the same manner as above except that the solvent of the test sample solution was added instead of the test sample solution as a control.
The calculation method of the platelet aggregation inhibitory action is as follows.
Platelet aggregation inhibition rate (%) = [(A−B) / A] × 100
A: Platelet aggregation rate of control B: Platelet aggregation rate upon addition of test sample solution

  The results are shown in Table 10. In addition, Extract No. 1-1, 1-2, 2, 5, 6-1, 7-1, 7-2, 8-1, 8-2, and 9 were used at a sample concentration of 400 μg / mL. Extract No. 3 was used at a sample concentration of 200 μg / mL.

(Example 10: Cyclooxygenase-2 (COX-2) activity inhibition test)
Using each plant extract as a test sample, cyclooxygenase-2 (COX-2) activity inhibitory action was tested by the following test method.

Murine macrophage cells (RAW264.7) were cultured using Dulbecco's MEM containing 10% FBS, and then the cells were collected with a cell scraper. The collected cells were diluted with Dulbecco MEM containing 10% FBS to a concentration of 2.0 × 10 ⁵ cells / mL, then seeded at 100 μL per well in a 96-well plate, and cultured for 18 hours. After culturing, in order to acetylate and inactivate COX-1 already present and COX-2 expressed in a small amount, the medium was replaced with a 500 μmol / L aspirin-containing medium and cultured for 4 hours. The cells were washed three times with PBS (−), 100 μL of a test sample dissolved in Dulbecco's MEM containing 10% FBS containing final concentration 0.5% DMSO was added to each well, and then 10% FBS at a final concentration of 1 μg / mL. 100 μL of lipopolysaccharide (LPS, E. coli 0111; B4, DIFCO) dissolved in containing Dulbecco's MEM was added and cultured for 16 hours. After completion of the culture, the amount of prostaglandin E _{2 in} the culture supernatant of each well was quantified using PGE ₂ EIA Kit (Cayman Chemical).
The calculation method of COX-2 activity inhibitory action is as follows.
COX-2 activity inhibition rate (%) = {1− (A−C) / (B−C)} × 100
A: Prostaglandin _{E 2} amount when the test specimen · LPS stimulated B: amount prostaglandin _{E 2} at the time of a test sample without additives · LPS stimulated C: prostaglandins during a test sample without additives · LPS unstimulated Jin _{E 2} amount

  The results are shown in Table 11. In addition, Extract No. 2 and 4-1 were used at a sample concentration of 200 μg / mL. Extract No. 4-2 was used at a sample concentration of 50 μg / mL.

From the results of Examples 5-10, an extract of Mirintonia-Horutenshisu (Millingtonia hortensis), extract of Ria macro filler (Leea macrophylla), extract of Kodariokarikusu-Motoriusu (Codariocalyx motorius), Martini A-An'nua (Martynia annua extract), extract of Pittosuporumu-Napaurense (Pittosporum napaulense), extract of Hesuperetusa-Kurenurata (Hesperethusa crenulata), extract of Puterosuperumumu-Dibe Resid Folie um (Pterospermumu diversifolium), extracted in Gurewia-Eriokarupa (Grewia eriocarpa) Things, extracts of Tsunberugia-Rauriforia (Thunbergia laurifolia), and Puremuna-extract integrase Li Folia (Premna integrifolia) nitric oxide (NO) production inhibitory activity, tumor necrosis factor (TNF-alpha) production inhibitory activity , Hyaluronidase activity inhibitory action, hexosaminidase release inhibitory action, platelet aggregation inhibitory action, and cyclooxygenase-2 (COX-2) activity inhibitory action, each of these plant extracts, It was suggested that it can be suitably used as an active ingredient of an anti-inflammatory agent.

(Example 11: Elastase activity inhibitory action test)
Each plant extract was used as a test sample, and the elastase activity inhibitory action was tested by the following test method.

In a 96-well plate, 50 μL of a test sample prepared with 0.2 mol / L Tris-HCL buffer (pH 8.0) and 50 μL of a 20 μg / mL elastase type III solution were mixed. Then, 0.4514 mg / mL N-SUCCINYL-ALA-ALA-ALA-p-NITROANILIDE 100 μL prepared in the above buffer solution was added and reacted at 25 ° C. for 15 minutes. After completion of the reaction, absorbance at a wavelength of 415 nm was measured. A blank test was performed and corrected in the same manner.
The calculation method of the elastase activity inhibitory action is as follows.
Elastase activity inhibition rate (%) = {1− (C−D) / (A−B)} × 100
A: Absorbance at a wavelength of 415 nm without addition of a test sample and addition of an enzyme B: Absorbance at a wavelength of 415 nm without addition of a test sample and addition of an enzyme C: Absorbance at a wavelength of 415 nm with addition of a test sample and addition of an enzyme D: Addition of a test sample Absorbance at a wavelength of 415 nm with no enzyme added

  The results are shown in Table 12. In addition, Extract No. 2, 4-2, 7-1, 7-2 and 8-1 were used at a sample concentration of 400 μg / mL.

(Example 12: MMP-1 activity inhibitory action test)
Using each plant extract as a test sample, MMP-1 activity inhibitory action was tested by the following test method.

As a test method, a method in which the Wunsch and Heidrich method was partially modified was used. That is, in a test tube with a lid, 50 μL of a test sample dissolved in 0.1 mol / L Tris-HCl buffer (pH 7.1) containing 20 mmol / mL calcium chloride, MMP-1 (COLLAGENASE Type IV from Clostridium histolyticum (Sigma) )) 50 μL of the solution and 400 μL of Pz-peptide (Pz-Pro-Leu-Gly-Pro-D-Arg-OH (BACHEM Feinchemikaline AG)) solution were mixed and reacted at 37 ° C. for 30 minutes, and then 25 mmol / L 1 mL of citric acid solution was added to stop the reaction. Thereafter, 5 mL of ethyl acetate was added and shaken vigorously. This was centrifuged (1600 × g, 10 minutes), and the absorbance of the ethyl acetate layer at a wavelength of 320 nm was measured. A blank test was performed and corrected in the same manner.
The calculation method of MMP-1 activity inhibitory action is as follows.
MMP-1 activity inhibition rate (%) = {1− (C−D) / (A−B)} × 100
A: Absorbance at a wavelength of 320 nm without addition of a test sample and addition of an enzyme B: Absorbance at a wavelength of 320 nm without addition of a test sample and addition of an enzyme C: Absorbance at a wavelength of 320 nm with addition of a test sample and addition of an enzyme D: Addition of a test sample Absorbance at 320 nm with no enzyme added

  The results are shown in Table 13. In addition, Extract No. 1-1, 1-2, 2, 3, 4-1, 4-2, 5, 6-1, 6-2, 7-1, 7-2, 8-1, 8-2, 9, 10 are Used at a sample concentration of 400 μg / mL.

(Example 13: Skin fibroblast proliferation promoting action test)
Each plant extract was used as a test sample, and the skin fibroblast proliferation promoting action was tested by the following test method.

Human normal skin fibroblasts (NB1RGB) were cultured using α-MEM containing 10% FBS, and then cells were collected by trypsin treatment. The recovered cells were diluted with α-MEM containing 5% FBS to a concentration of 7.0 × 10 ⁴ cells / mL, then seeded at 100 μL per well in a 96-well plate, and cultured overnight. After completion of the culture, 100 μL of a test sample dissolved in α-MEM containing 5% FBS was added to each well and cultured for 3 days. The skin fibroblast proliferation promoting action was measured using the MTT assay. After completion of the culture, 100 μL of the medium was removed from each well, and 20 μL of MTT dissolved in PBS (−) at a final concentration of 5 mg / mL was added to each well. After culturing for 4.5 hours, 100 μL of 0.01 mol / L hydrochloric acid solution in which 10% SDS was dissolved was added to each well and cultured overnight, and then the absorbance at a wavelength of 570 nm was measured. At the same time, the absorbance at a wavelength of 650 nm was measured as turbidity, and the difference between the two was used as the amount of blue formazan produced. In addition, a blank test was performed and corrected in the same manner.
The calculation method of the dermal fibroblast proliferation promoting action is as follows.
Skin fibroblast proliferation promotion rate (%) = (St−Sb) / (Ct−Cb) × 100
St: Absorbance in cells with added test sample Sb: Absorbance in blank test with added test sample Ct: Absorbance in cells without added test sample Cb: Absorbance in blank test without added test sample

  The results are shown in Table 14. In addition, Extract No. 2, 4-1, 4-2, 9, 10 were used at a sample concentration of 400 μg / mL. Extract No. 3 was used at a sample concentration of 200 μg / mL. Extract No. 5 and 7-1 were used at a sample concentration of 100 μg / mL. Extract No. 7-2, 8-1, and 8-2 were used at a sample concentration of 50 μg / mL.

(Example 14: Epidermal keratinocyte proliferation promoting test)
Each plant extract was used as a test sample, and the epidermal keratinocyte proliferation promoting action was tested by the following test method.

Normal human neonatal foreskin keratinocytes (NHEK) were cultured using a normal human epidermal keratinocyte long-term culture growth medium (EpiLife-KG2), and then cells were collected by trypsin treatment. The collected cells were diluted with EpiLife-KG2 to a concentration of 1.5 × 10 ⁴ cells / mL, then seeded at 100 μL per well on a collagen-coated 96-well plate, and cultured overnight. After completion of the culture, 100 μL of a test sample dissolved in EpiLife-KG2 was added to each well and cultured for 3 days. The epidermal keratinocyte proliferation promoting action was measured using the MTT assay. After completion of the culture, the medium was removed, and 100 μL of MTT dissolved in PBS (−) at a final concentration of 0.4 mg / mL was added to each well. After culturing for 2 hours, blue formazan produced in the cells was extracted with 100 μL of 2-propanol. After extraction, the absorbance at a wavelength of 570 nm was measured. At the same time, the absorbance at a wavelength of 650 nm was measured as turbidity, and the difference between the two was used as the amount of blue formazan produced.
The calculation method of the epidermal keratinocyte proliferation promoting action is as follows.
Epidermal keratinocyte proliferation promotion rate (%) = St / Ct × 100
St: Absorbance in cells to which test sample was added Ct: Absorbance in cells to which test sample was not added

  The results are shown in Table 15. In addition, Extract No. 1-1 was used at a sample concentration of 12.5 μg / mL. Extract No. 1-2 was used at a sample concentration of 3.125 μg / mL.

(Example 15: Test for recovery from UV-B damage)
Each plant extract was used as a test sample, and the recovery action from UV-B damage was tested by the following test method.

Human normal skin fibroblasts (NB1RGB) were cultured using α-MEM containing 10% FBS, and then cells were collected by trypsin treatment. The collected cells were diluted to a concentration of 2.0 × 10 ⁵ cells / mL using α-MEM, and then seeded on a 48-well plate at 200 μL per well. After culturing for 24 hours, the medium was replaced with 100 μL of PBS (−) and irradiated with 1.0 J / cm ² of UV-B. Immediately after irradiation, PBS (-) was removed, 400 μL of a test sample dissolved in D-MEM containing 10% FBS was added to each well, and cultured for 24 hours. The recovery effect from UV-B damage was measured using an MTT assay. After completion of the culture, the medium was removed, and 200 μL of MTT dissolved at a final concentration of 0.4 mg / mL was added to each well. After culturing for 2 hours, blue formazan produced in the cells was extracted with 200 μL of 2-propanol. After extraction, the absorbance at a wavelength of 570 nm was measured. At the same time, the absorbance at a wavelength of 650 nm was measured as turbidity, and the difference between the two was used as the amount of blue formazan produced. Similarly, after cell seeding, cells that were not irradiated with UV-B, and cells that were irradiated with UV-B after cell seeding and were not added with the test sample were also measured in the same manner, and were set as a non-irradiated group and an irradiated group, respectively.
The calculation method of the recovery action from UV-B damage is as follows.
UV-B damage recovery rate (%) = {(Nt−C) − (Nt−Sa)} / (Nt−C) × 100
Nt: Absorbance in cells not irradiated with UV-B C: Absorbance in cells irradiated with UV-B and no test sample added Sa: Absorbance in cells irradiated with UV-B and added with test sample

  The results are shown in Table 16. In addition, Extract No. 1-1, 2, 3, 4-1, 4-2, 5, 7-1, 7-2, 8-1, 8-2, 9, 10 were used at a sample concentration of 100 μg / mL. Extract No. 6-2 was used at a sample concentration of 6.25 μg / mL.

(Example 16: Damage suppression test for hydrogen peroxide)
Each plant extract was used as a test sample, and the damage suppressing action against hydrogen peroxide was tested by the following test method.

Human normal newborn dermal fibroblasts (NBIRGB) were cultured using α-MEM medium (GIBCO BLR, pH 7.2), and then cells were collected by trypsin treatment. The collected cells were diluted with α-MEM medium to a concentration of 2.5 × 10 ⁵ cells / mL, seeded in a 48-well plate, seeded at 200 μL per well, and cultured overnight. After completion of the culture, the medium was removed, 200 μL of a test sample dissolved in α-MEM containing 1% FBS was added to each well, and cultured for 24 hours. After completion of the culture, the medium was removed and washed with 400 μL of PBS (−). After washing, 200 μL of hydrogen peroxide (1 mmol / L) dissolved in Hanks buffer or Hanks buffer alone was added to each well and incubated for 2 hours. After incubation, the plate was washed with 400 μL of PBS (−), and 200 μL of neutral red dissolved in α-MEM containing 1% FBS at a final concentration of 0.05 mg / mL was added to each well. After culturing for 2.5 hours, the neutral red solution was discarded, and 200 μL of ethanol / acetic acid solution (ethanol: acetic acid: water = 50: 1: 49) was added to each well to extract the dye. After extraction, the absorbance at a wavelength of 540 nm was measured.
The calculation method of the damage suppression action with respect to hydrogen peroxide is as follows.
Hydrogen peroxide damage suppression rate (%) = {1− (C−A) / (C−B)} × 100
A: Absorbance of hydrogen peroxide treatment / test sample treatment,
B: Absorbance of hydrogen peroxide treatment / no test sample treatment,
C: Absorbance of hydrogen peroxide untreated / test sample untreated

  The results are shown in Table 17. In addition, Extract No. 2, 4-1, 7-1, 7-2, 8-1, 8-2 and 9 were used at a sample concentration of 400 μg / mL. Extract No. 4-2, 5 and 6-2 were used at a sample concentration of 100 μg / mL. Extract No. 6-1 was used at a sample concentration of 25 μg / mL.

From the results of Examples 11 to 16, an extract of Mirintonia-Horutenshisu (Millingtonia hortensis), extract of Ria macro filler (Leea macrophylla), extract of Kodariokarikusu-Motoriusu (Codariocalyx motorius), Martini A-An'nua (Martynia annua extract), extract of Pittosuporumu-Napaurense (Pittosporum napaulense), extract of Hesuperetusa-Kurenurata (Hesperethusa crenulata), extract of Puterosuperumumu-Dibe Resid Folie um (Pterospermumu diversifolium), Gurewia-Eriokarupa of (Grewia eriocarpa) Extract, extract of Tsunberugia-Rauriforia (Thunbergia laurifolia), and Puremuna integrase extract Li Folia (Premna integrifolia) elastase inhibitory activity, MMP-1 inhibitory activity, dermal fibroblast proliferation promoting action, It has been confirmed that it has at least any one of epidermal keratinocyte proliferation promoting action, UV-B damage recovery action, and hydrogen peroxide damage inhibiting action. Each of these plant extracts is an active ingredient of an anti-aging agent. It was suggested that it can be suitably used.

(Example 17: Androgen receptor antagonism test)
Each plant extract was used as a test sample, and androgen receptor antagonism was tested by the following test method.

First, after culturing SC-3 cells cloned from mouse spontaneous breast cancer (Shionogi cancer, SC115) using 2% DCC-FBS and ^10-8 mol / L testosterone-containing MEM (MEM / 2). Cells were collected by trypsinization. The collected cells were diluted with activated carbon-treated FBS-containing MEM (MEM / 2) to a concentration of 1.0 × 10 ⁵ cells / mL, seeded at 100 μL per well in a 96-well microplate, and cultured overnight. After completion of the culture, the medium was removed, and 100 μL of a test sample solution dissolved in 0.5% by mass of BSA-containing Ham F12 + MEM (HMB) containing 10 ⁻⁹ mol / L dihydrotestosterone (DHT) was added and cultured for 48 hours. . Thereafter, 100 μL of MTT dissolved in MEM / 2 containing FBS containing activated carbon at a final concentration of 0.4 g / mL was added to each hole. After culturing for 2 hours, blue formazan produced in the cells was extracted with 200 μL of 2-propanol. After extraction, the absorbance at a wavelength of 570 nm was measured. At the same time, the absorbance at a wavelength of 650 nm was measured as turbidity, and the difference between the two was used as the amount of blue formazan produced. As a blank test, a cell cultured with HMB alone was used as a positive control, and a cell cultured with HMB containing only 10 ⁻⁹ mol / L DHT was used as a positive control.
The calculation method of androgen receptor antagonism is as follows.
Androgen receptor antagonist ratio (%) = [1- (C−D) / (A−B)] × 100
A: Absorbance at 570 nm to 650 nm without addition of DHT and no sample solution B: Absorbance at 570 nm to 650 nm without addition of DHT and no sample solution C: Absorbance at 570 nm to 650 nm with addition of DHT and addition of sample solution D: Absorbance at 570 nm to 650 nm with no DHT added and sample solution added

  The results are shown in Table 18. In addition, Extract No. 1-1, 1-2, 2, 3, 4-1, 4-2, 5, 6-2, 7-1, 8-1, 9, and 10 were used at a sample concentration of 50 μg / mL. Extract No. 6-1 and 7-2 were used at a sample concentration of 12.5 μg / mL. Extract No. 8-2 was used at a sample concentration of 3.13 μg / mL.

(Example 18: Test of promoting dermal papilla cell proliferation)
Each plant extract was used as a test sample, and the hair papilla cell growth promoting effect was tested by the following test method.

After culturing using a hair papilla cell growth medium (TOYOBO, TPGM-250) containing normal human hair papilla cells (TOYOBO, CA60205), the cells were collected by trypsin treatment. After the collected cells were diluted to a concentration of 2.0 × ¹⁰ 4 cells / mL with 10% fetal bovine serum (FBS) containing Dulbecco's MEM (DMEM), 1 hole per 100μL of a 96-well microplate coated with collagen Seeding and culturing for 3 days. After the culture, the medium was removed, 200 μL of each test sample solution dissolved in serum-free Dulbecco's MEM (DMEM) was added to each well, and further cultured for 4 days. The dermal papilla cell proliferation promoting effect was measured using MTT assay. Specifically, after completion of the culture, the medium was removed, and MTT [3- (4,5-Dimethyl-2-thiazolyl) -2,5-diphenyl- dissolved in serum-free DMEM at a final concentration of 0.4 mg / mL. 2H-tetrazolium Bromide] was added to each well in an amount of 100 μL. After culturing for 2 hours, blue formazan produced in the cells was extracted with 100 μL of 2-propanol. After extraction, the absorbance at a wavelength of 570 nm was measured. At the same time, the absorbance at a wavelength of 650 nm was measured as turbidity, and the difference between the two was used as the amount of blue formazan produced. In addition, a blank test was performed and corrected in the same manner.
The calculation method of the dermal papilla cell proliferation promoting action is as follows.
Growth rate of dermal papilla cells (%) = A / B × 100
A: Absorbance when test sample solution is added B: Absorbance when test sample solution is not added

  The results are shown in Table 19. In addition, Extract No. 1-1, 2-1, 4-1, 4-2, 5, 6-2, 7-1, 7-2, 8-1, 8-2, 9, 10 were used at a sample concentration of 25 μg / mL. Extract No. 3 and 6-1 were used at a sample concentration of 6.25 μg / mL.

From the results of Examples 17-18, extract of Mirintonia-Horutenshisu (Millingtonia hortensis), extract of Ria macro filler (Leea macrophylla), extract of Kodariokarikusu-Motoriusu (Codariocalyx motorius), Martini A-An'nua (Martynia annua extract), extract of Pittosuporumu-Napaurense (Pittosporum napaulense), extract of Hesuperetusa-Kurenurata (Hesperethusa crenulata), extract of Puterosuperumumu-Dibe Resid Folie um (Pterospermumu diversifolium), Gurewia-Eriokarupa of (Grewia eriocarpa) Extract, extract of Tsunberugia-Rauriforia (Thunbergia laurifolia), and extract of Puremuna integrase Li cordifolia (Premna integrifolia) is androgen receptor antagonism, and have at least one of dermal papilla cell growth promoting action It was confirmed and it was suggested that each of these plant extracts can be suitably used as an active ingredient of a hair restorer.

(Formulation example 1)
-Emulsion-
A milky lotion was produced by a conventional method according to the following composition.
· Mirintonia-Horutenshisu (Millingtonia hortensis L.f.) 50 wt% ethanol extract of the bark of (extract No. 1-1) · · · 0.10 g
・ Jojoba oil: 4.00 g
・ 1,3-Butylene glycol ・ ・ ・ 3.00g
・ Polyoxyethylene cetyl ether (20E.O.) ... 2.50 g
・ Olive oil ... 2.00g
・ Squalane ... 2.00g
・ Cetanol ... 2.00g
・ Glyceryl monostearate ... 2.00g
・ Oleic acid polyoxyethylene sorbitan (20E.O.) ... 2.00g
・ Methyl paraoxybenzoate 0.15 g
・ Twilight extract ... 0.10g
・ Dipotassium glycyrrhizinate ... 0.10g
・ Ginkgo biloba extract ... 0.10g
・ Conchiolin ... 0.10g
・ Oat extract ... 0.10g
・ Chicken extract ... 0.10g
・ Fragrance ... 0.05g
・ Purified water: remainder (total 100.00 g)

(Formulation example 2)
-Lotion-
In accordance with the following composition, lotion was produced by a conventional method.
- Ria macro filler (Leea macrophylla Roxb.) 50 wt% ethanol extract of the leaves of (extract No.2) · · · 0.10 g
・ Glycerin ... 3.00g
・ 1,3-Butylene glycol ・ ・ ・ 3.00g
・ Oleic acid polyoxyethylene sorbitan (20E.O.) ... 2.00g
・ Methyl paraoxybenzoate 0.15 g
・ Citric acid ... 0.10g
・ Sodium citrate: 0.10 g
・ Oil-soluble licorice extract ... 0.10g
・ Seaweed extract ... 0.10g
・ Cudin extract ... 0.10g
・ Xylobiose Mixture ... 0.05g
・ Fragrance ... 0.05g
・ Purified water: remainder (total: 100.00 g)

(Formulation example 3)
-Cream-
According to the following composition, the cream was manufactured by a conventional method.
· Kodariokarikusu-Motoriusu (Codariocalyx motorius (Hout.) H.Ohashi ) 50 wt% ethanol extract of leaves and stems of (extract No.3) · · · 0.10 g
・ Squalane ... 10.00g
・ 1,3-butylene glycol ... 6.00g
・ Liquid paraffin ・ ・ ・ 5.00g
・ Salah beeswax 4.00 g
・ Cetanol ... 3.00g
・ Glyceryl monostearate ... 3.00 g
・ Lanoline ... 2.00g
・ Oleic acid polyoxyethylene sorbitan (20E.O.) ... 1.50 g
・ Methyl paraoxybenzoate ... 1.50g
・ Stearic acid: 1.00 g
・ Yeast extract ... 0.10g
・ Perilla extract ... 0.10g
-Linden extract ... 0.10g
・ Jiuyu extract ... 0.10g
・ Fragrance ... 0.10g
・ Purified water: remainder (total: 100.00 g)

(Formulation example 4)
−Pack−
According to the following composition, the pack was produced by a conventional method.
-50% ethanol extract (extract No. 4-1) of leaves and stems of Martynia annua L. 0.20 g
・ Polyvinyl alcohol: 15.00g
・ Ethanol ... 10.00g
・ Propylene glycol: 7.00 g
・ Polyethylene glycol ... 3.00g
・ Sage extract ... 0.10g
・ Toki extract ... 0.10g
・ Carrot extract ... 0.10g
・ Ethyl paraoxybenzoate 0.05g
・ Fragrance ... 0.05g
・ Purified water: remainder (total: 100.00 g)

(Formulation example 5)
-Hair tonic-
A hair tonic was produced by a conventional method according to the following composition.
・ Pyridoxine hydrochloride ... 0.1g
・ Resorcin ・ ・ ・ 0.01g
・ D-pantothenyl alcohol ... 0.1g
・ Dipotassium glycyrrhizinate ... 0.1g
・ L-Menthol 0.05g
・ 1,3-Butylene glycol ・ ・ ・ 4.0g
・ Carrot extract ... 0.5g
・ Ethanol ... 25.0g
-50% by weight ethanol extract (extract No. 7-1) of leaves of Pterospermumu diversifolium Bl.
-Fragrance-appropriate amount-Purified water-the balance (total: 100.00 g)

(Formulation example 6)
-Shampoo-
A shampoo was produced by a conventional method according to the following composition.
・ Polyoxyethylene alkyl ether sodium sulfate 30.0 g
・ Polyoxyethylene alkyl ether ammonium sulfate ... 20.0g
・ Palm oil fatty acid amidopropyl betaine ... 6.0g
・ Palm oil fatty acid modiethanolamide: 4.0 g
・ Ethylene glycol distearate ... 2.0g
・ Preservative (methyl paraoxybenzoate) ... 0.15g
· Gurewia-Eriokarupa (Grewia eriocarpa Juss.) 50 wt% ethanol extract of the leaves of (Extract No. 8-1) · · · 0.2 g
・ Muguroji extract ... 0.2g
・ Cotton extract ... 0.5g
・ Oat extract ... 0.3g
・ Rosemary extract 0.5g
・ Fragrance: 0.01g
・ 1,3-Butylene glycol ・ ・ ・ 3.0g
・ Purified water: remainder (total: 100.00 g)

(Formulation example 7)
-Rinse-
According to the following composition, the rinse was manufactured by the conventional method.
・ Stearyltrimethylammonium chloride 1.5g
・ Polyoxyethylene cetyl ether ... 1.0g
・ Cetyl alcohol: 2.0g
・ Octyldodecanol ・ ・ ・ 1.0g
・ Cationized cellulose: 0.5g
・ Propylene glycol ... 5.0g
· Tsunberugia-Rauriforia 50 wt% ethanol extract of the bark of (Thunbergia laurifolia) (extract No.9) · · · 0.2 g
・ Muguroji extract ... 0.2g
・ Cotton extract ... 0.5g
・ Oat extract ... 0.3g
・ Rosemary extract 0.5g
・ Fragrance ... 3.0g
・ Purified water: remainder (total: 100.00 g)

(Formulation example 8)
-Tablet dietary supplements-
The following mixture was tableted to produce a tablet-shaped dietary supplement.
-50% by weight ethanol extract (extract No. 5) of bark of Pittosporum napaulense (extract No. 5) ... 30 g
・ Powdered sugar (sucrose) 178g
・ Sorbit ・ ・ ・ 10g
・ Glycerin fatty acid ester: 12g

(Formulation example 9)
-Granular dietary supplement-
The following mixture was formed into granules to produce a granular dietary supplement.
-50 mass% ethanol extract (extract No. 6-1 ) of stem of Hesperetusa crenurata (Extract No. 6-1) ... 20 g
・ Beat oligosaccharide ... 1000g
・ Vitamin C ... 167g
・ Stevia extract ... 10g

The external preparation for skin containing at least one of the antioxidant, anti-inflammatory agent, anti-aging agent, and hair restorer of the present invention has at least one of excellent antioxidant effect, anti-inflammatory effect, anti-aging effect, and hair-restoring effect. For example, ointments, creams, milky lotions, serums, lotions, foundations, packs, jellies, lip balms, lipsticks, bath salts, soaps, body shampoos, astringents, hair It can be suitably used for tonic, hair lotion, hair cream, hair liquid, shampoo, pomade, rinse and the like.
In addition, a food or drink containing at least one of the antioxidant, anti-inflammatory agent, anti-aging agent, and hair restorer of the present invention has an excellent antioxidant effect, anti-inflammatory effect, anti-aging effect, and Since it has at least one of hair-growth action and is excellent in safety, it can be suitably used for, for example, health foods, dietary supplements and the like.

Claims (1)

It contains an extract of Millingtonia hortensis and has an inhibitory action on nitric oxide (NO) production, an inhibitory action on tumor necrosis factor (TNF-α) production, an inhibitory action on hexosaminidase release, and an inhibitory action on platelet aggregation. An anti-inflammatory agent comprising at least one of them.