JP6650954B2 - Skin cosmetics, hair cosmetics and foods and drinks - Google Patents

Description

  The present invention relates to an antiobesity agent, a cyclic AMP phosphodiesterase activity inhibitor, a dipeptidyl peptidase IV activity inhibitor, an antiaging agent, a hair restorer, an antiandrogen, a whitening agent, an anti-inflammatory agent, a skin cosmetic, and a hair cosmetic. , And food and drink.

  In recent years, body fat has increased due to lifestyle habits such as satiety and lack of exercise, and obesity has increased. Such an increase in obesity is observed not only in humans but also in pets and livestock. Obesity causes lifestyle-related diseases such as hyperlipidemia and arteriosclerosis, and is not only a cosmetic problem but also a major health problem.

  Here, cyclic AMP (cAMP) is known to be involved in lipolysis in a living body. cAMP activates a lipase present in a living body, and the activated lipase degrades fat into fatty acids and glycerol. However, activation of cAMP phosphodiesterase induces cAMP degradation and inhibits lipase activation. Therefore, it is considered that by inhibiting the activity of cAMP phosphodiesterase, the amount of cAMP in cells increases, and the degradation of fat can be promoted.

  In addition, platelet aggregation that causes an inflammatory response is related to the concentration of cyclic AMP (cAMP) in platelets. It is known that platelets tend to aggregate when cAMP is degraded by cAMP phosphodiesterase and cAMP concentration is reduced. Have been. Therefore, it is considered that platelet aggregation can be prevented by suppressing the action of cAMP phosphodiesterase to prevent a decrease in cAMP concentration, thereby preventing, treating or improving allergic diseases and inflammatory diseases. Tubeimoside I (see Patent Literature 1) and the like are known as having cAMP phosphodiesterase activity inhibitory activity.

  Obesity causes abnormalities in lipid metabolism and causes lifestyle-related diseases such as dyslipidemia as described above. Here, dyslipidemia is a disease in which one or more components among serum lipids, that is, cholesterol, triglyceride, phospholipid, free fatty acid, and the like are abnormally increased, and various disorders are caused. Cholesterol is a component of animal biological membranes and a starting material such as steroid hormones, and is a very important substance for animals. However, if blood cholesterol is excessive, cholesterol and the like accumulate inside blood vessels, causing arteriosclerosis, and may lead to fatal diseases such as ischemic heart disease and cerebral infarction. On the other hand, free fatty acids are released into the blood from fat cells and used as energy sources in the body, but the surplus is taken up by the liver and re-synthesized into neutral fats. If the free fatty acids in the blood are excessive, the neutral fats will be chronically excessive and the abnormal lipid condition will continue, increasing the risk of arteriosclerosis. In addition, when the blood free fatty acid is excessive, insulin resistance may increase and the pathology of diabetes may progress.

  Therefore, if blood lipids that are too high due to obesity (eg, blood cholesterol, blood free fatty acids, etc.) can be reduced, the symptoms of arteriosclerosis and diabetes that progress with obesity can be prevented, alleviated or ameliorated. It is thought that it can be done. As a component having a blood cholesterol lowering action and a blood free fatty acid lowering action, a roasted yellow leaf extract (Patent Document 2) is known.

  Dipeptidyl peptidase IV (hereinafter, also referred to as “DPPIV”) is one of serine proteases and has an enzymatic activity of recognizing the second proline or alanine from the N-terminal and cleaving the C-terminal side. DPPIV is expressed on epithelial and endothelial cells of tissues such as kidney, liver, intestinal tract, and placenta, and on the cell surface of T cells, and is thought to be involved in various physiological phenomena via its enzymatic activity and the like.

  DPPIV substrates include a hormone called incretin. Incretin is a general term for hormones that are secreted from the intestine by stimulation of nutrients and promote insulin secretion from pancreatic β cells in a blood sugar-dependent manner, and GLP-1 and GIP are known. These incretins not only promote blood glucose-dependent insulin secretion, but also suppress glucagon secretion from pancreatic α-cells, lower blood pressure, suppress gastric emptying, and suppress feeding by acting on the hypothalamus. It has an effect (see Non-Patent Document 1). However, since incretin is degraded by DPPIV, it is known that, for example, the half-life of GLP-1 in vivo is about 1.5 minutes. Therefore, if the enzymatic activity of DPPIV can be inhibited, the half-life of incretin in vivo can be prolonged, whereby the type 2 diabetes, obesity, hypertension, insulin resistance, It is expected to be useful in the treatment of such as.

  DPPIV is the same as CD26, which is one of the T cell activation markers, and is known to regulate its activity using many immunoregulatory peptides as substrates. Therefore, it is considered that by controlling the activity of DPPIV, it is possible to control immune reactions such as autoimmune diseases such as rheumatoid arthritis and rejection by transplantation. Furthermore, DPPIV is known to be involved in metabolism of several neuropeptides and growth hormone; invasion, metastasis, angiogenesis and the like in cancer; and infection of HIV to lymphocytes. Therefore, by inhibiting the activity of DPPIV, neuropathy such as pain, neurodegenerative disease and neuropsychiatric disease (eg, sciatica, Alzheimer's disease, depression, etc.); growth hormone deficiency and growth hormone are used for treatment Diseases; cancer (eg, T-cell lymphoma, acute lymphoblastic leukemia, thyroid cancer, basal cell carcinoma, breast cancer, etc.); and diseases such as HIV infection (AIDS) could be treated.

  The basement membrane exists at the boundary between the epidermis and the dermis that constitute the skin. The basement membrane not only anchors the epidermis and the dermis but also plays an important role in maintaining skin functions (see Non-Patent Document 2). The main skeleton of the basement membrane has a network structure composed of type IV collagen. Various glycoproteins containing laminin 5 as a main component are present at the boundary between the basement membrane and the epidermis and connect the basement membrane and the epidermis. Such laminin 5 is produced from epidermal keratinocytes present in the epidermis. Is done. In the young skin, the interaction of the epidermis and dermis maintains homeostasis by the action of the basement membrane, thereby securing moisture retention, flexibility, elasticity, etc., and the skin is visibly taut and shiny in appearance. Is maintained.

  However, under the influence of certain external factors such as irradiation of ultraviolet rays, remarkable drying of air, and excessive skin washing, and as aging progresses, laminin 5, which is a main component of the basement membrane, is decomposed and deteriorated. And the basement membrane structure is destroyed (see Non-Patent Document 3). As a result, the skin loses its moisturizing function and elasticity, and the keratin starts to peel off abnormally, so that the skin loses its stiffness and luster, and exhibits aging symptoms such as roughness and wrinkles. As described above, changes associated with skin aging, ie, wrinkles, dullness, disappearance of texture, decrease in elasticity, etc., involve a decrease in basement membrane components and structural changes in basement membranes, and the production of laminin 5 Is considered to be able to prevent or ameliorate the aging symptoms of the skin.

  Laminin is composed of various combinations of α chain, β chain and γ chain, and 15 types (laminin 1 to laminin 15) are known at present. Of these, laminin 5 (α3β3γ2) is present in large amounts in the basement membrane of epithelial tissues such as skin, digestive organs, kidneys, and lungs. Genetic disorders (lethal congenital epidermolysis bullosa, Herlitz junctional epidermolysis bullosa) caused by congenital abnormalities of the genes encoding the laminin 5 chains may exhibit fatal symptoms of epidermis detachment of the whole body. Are known. It is known that laminin 5 adheres cells more strongly (has a higher cell adhesion activity) and strongly promotes cell motility (has a higher cell motility activity) than other extracellular matrix molecules.

  As described above, since laminin 5 has high cell motility activity, it is known that it promotes cell migration in damaged skin and promotes wound healing (see Patent Document 3). That is, promoting the production of laminin 5 is important for promoting the healing of skin damage in which the structure of the basement membrane is destroyed.

  In recent years, involvement of matrix metalloproteases (MMPs; Matrix metalloproteinases) has been pointed out as a factor inducing changes associated with skin aging. Among these MMPs, matrix metalloprotease-2 (MMP-2), an enzyme belonging to the group of gelatinases, is known as an enzyme that degrades type IV collagen and laminin 5, which are main components of the basement membrane. It is clear that the expression and activity of MMP-2 greatly increases upon irradiation with ultraviolet light, causes a decrease in basement membrane components due to ultraviolet light, causes a structural change in the basement membrane, and is a major factor such as formation of wrinkles and sagging in the skin. (See Non-Patent Document 4). MMP-2 also degrades type IV collagen and the like constituting the basement membrane present under vascular endothelial cells, and the degraded vascular endothelial cells migrate to the interstitium, proliferate in the interstitium, It creates cavities and builds new blood vessels. It is known that the newly formed blood vessels reach the tumor cells, supply nutrients and oxygen to the tumor cells, and the tumor grows (see Non-Patent Document 5).

  Therefore, by inhibiting the activity of MMP-2, it is possible to suppress a decrease in basement membrane components and a change in the structure of the basement membrane, thereby improving skin functions, suppress angiogenesis, and suppress tumor cell proliferation. It is thought that it can be done. As an agent having an MMP-2 inhibitory action, for example, an extract from black bean adzuki bean (see Patent Document 4) and the like are known.

  On the other hand, among MMPs, matrix metalloprotease-1 (MMP-1), which is an enzyme belonging to the collagenase group, is known as an enzyme that degrades collagen, which is a main component of the above-described extracellular matrix of the dermis. The expression of MMP-1 is greatly increased by irradiation with ultraviolet rays, and is considered to be a cause of a decrease and denaturation of collagen, and a major factor such as formation of skin wrinkles and a decrease in elasticity. When the activity of MMP-1 is enhanced, the extracellular matrix is destroyed. It is known that destruction of the extracellular matrix is associated with various diseases such as cancer invasion / metastasis, rheumatoid arthritis, osteoarthritis of the knee, periodontal disease, and age-related macular degeneration.

  Therefore, it is considered that by inhibiting the activity of MMP-1, it is possible to prevent, treat or ameliorate aging symptoms of the skin and to treat / prevent diseases related to the destruction of extracellular matrix. . As a substance having an MMP-1 activity inhibitory action, for example, corosolic acid (see Patent Document 5) is known.

  The epidermis and dermis of the skin are composed of epidermal cells, fibroblasts, and extracellular matrices such as collagen, elastin, and hyaluronic acid which are outside these cells and support the skin structure. Fibroblasts are actively proliferating in young skin, and the interaction of skin tissues such as fibroblasts and collagen maintains homeostasis, ensuring water retention, flexibility, elasticity, etc. It is also maintained in a fresh state with tension and luster.

  However, under the influence of certain external factors such as irradiation of ultraviolet rays, remarkable drying of air, and excessive skin washing, and as aging progresses, elastin and hyaluronic acid, which are the main components of the extracellular matrix, are reduced. It reduces production and causes degradation and deterioration. As a result, the moisturizing function and elasticity of the skin are reduced, and abnormal exfoliation of the keratin occurs, so that the skin loses its tension and luster, and exhibits aging symptoms such as rough skin and wrinkles. As described above, changes accompanying the aging of the skin, that is, wrinkles, dullness, disappearance of texture, decrease in elasticity, and the like involve reduction and denaturation of matrix components such as elastin and hyaluronic acid. Therefore, promoting the production of elastin or hyaluronic acid is important in preventing, treating or improving skin aging.

  Here, among these extracellular matrix components, hyaluronic acid is a kind of mucopolysaccharide and has a function of retaining cells by being filled in intercellular spaces, and further retains water in intercellular spaces. It has many functions, such as imparting lubricity and flexibility to tissues, and resistance to external forces such as mechanical obstacles. It is thought that if the production of hyaluronic acid can be promoted, skin aging symptoms such as rough skin, wrinkles, dullness, disappearance of texture, decreased elasticity and reduced moisturizing function can be prevented, treated or improved.

  Hyaluronic acid is present in cartilage, synovial fluid, umbilical cord, vitreous humor, and other connective tissues in addition to skin tissue. Of these, hyaluronic acid contained in synovial fluid covers the surface of articular cartilage, and contributes to the smooth operation of joints due to the lubrication function, hyaluronic acid coating / protection function of hyaluronic acid, and the like. On the other hand, in arthritis such as rheumatoid arthritis, it is known that the concentration of hyaluronic acid in synovial fluid is reduced. Therefore, it is thought that by promoting the production of hyaluronic acid, arthritis such as rheumatoid arthritis, osteoarthritis, suppurative arthritis, gouty arthritis, traumatic arthritis, and osteoarthritis can be prevented or treated. Further, it is known that granulation (tissue) is formed during the healing process of a wound or a burn, and hyaluronic acid is significantly increased in the granulation. Therefore, it is considered that by promoting the production of hyaluronic acid, the healing of a wound or a burn can be promoted. As an extract having a hyaluronic acid production promoting effect, an extract from Kusunohagashiwa (see Patent Document 6) and the like are known.

  On the other hand, among the above-mentioned extracellular matrix components, elastin is a fiber that gives elasticity to skin tissue. It is thought that if elastin production can be promoted, wrinkles and sagging are less likely to occur, and skin aging symptoms such as loss of tension and decreased elasticity can be prevented, treated or ameliorated.

  Elastin is widely expressed in tissues requiring elasticity in the body, such as lungs and blood vessels, in addition to skin tissues. It is known that when normal elastin decreases in these tissues with aging, elasticity in the lungs and blood vessels decreases, causing pulmonary diseases such as emphysema and hypertension and vascular diseases such as aneurysms. ing. Therefore, if the production of elastin can be promoted, it is unlikely that the elasticity in the lungs and blood vessels decreases, and lung diseases such as emphysema and high blood pressure, and vascular diseases such as aneurysms can be prevented or treated. . An extract from a plant belonging to the genus Hippophae, for example, is known as having an elastin production promoting action (see Patent Document 7).

  The aforementioned extracellular matrix components such as collagen are produced by fibroblasts. In young skin, fibroblasts are actively proliferating, and the interaction of skin tissues such as fibroblasts and collagen maintains homeostasis, ensuring moisture retention, flexibility, elasticity, and the like. It is maintained in a fresh state with tension and gloss. However, under the influence of certain external factors such as ultraviolet rays, remarkable drying of air, excessive skin washing, etc., or as aging progresses, fibroblast proliferation slows down and the skin's moisturizing function and elasticity are reduced. descend. Then, the skin loses its tone and luster, and exhibits aging symptoms such as roughness and wrinkles. Therefore, promoting the proliferation of fibroblasts is considered to be very important in preventing, treating or improving skin aging.

  In addition, fibroblasts play an important role in the healing process of wounds such as trauma and burns, and are also important in the healing process of skin diseases (eg, skin ulcers such as pressure sores, burn ulcers, and diabetic ulcers). is there. Therefore, it is considered that wounds and skin diseases can be treated by promoting the proliferation of fibroblasts. Furthermore, in the field of regenerative medicine, as a method of treating a patient suffering from a wound (severe burn, etc.) that is difficult to heal, a method of culturing and growing skin cells from a patient's own skin fragment and transplanting it into the patient However, the use of a cell growth promoter is expected to shorten the culture period of skin cells. Conventionally, an extract from Kusunohagashiwa (see Patent Document 6 described above) and the like having a fibroblast proliferation promoting action are known.

  The epidermis acts to relieve external stimuli and control the loss of internal components such as water, and has a four-layer structure starting from the lowermost layer, the basal layer, and continuing to the spinous layer, granular layer, and stratum corneum. It is composed of Most cells present in each layer are keratinocytes differentiated from the basal layer. The keratinocytes divided and proliferated in the basal layer, differentiated while passing through the spinous layer and granular layer, became keratinocytes, and constituted the stratum corneum composed of keratin protein fibers with strong cross-linking, Eventually, it will fall off the stratum corneum as dirt.

  The stratum corneum is present in the outermost shell of the skin and acts as a physical barrier to external stimuli. In order for the skin to have this barrier function, the cycle (keratinization) from the production of keratinocytes in the basal layer to the removal of keratinized cells as dirt is usually repeated in a four-week cycle to metabolize the epidermis. . However, the metabolic function of the stratum corneum also deteriorates with aging, and skin troubles such as pricking, dullness, pigmentation, and rough skin occur. Therefore, it is considered that by promoting the proliferation of keratinocytes and restoring the metabolic function of the skin, it is possible to improve skin aging such as fine lines, dullness, and pigmentation. Conventionally, as a substance having an epidermal keratinocyte proliferation promoting action, a shellfish mother extract (see Patent Document 8) and the like are known.

  In skin cells, aquaporins known as water channels are known to be expressed on cell membranes and play a role in taking in low molecular substances such as intercellular water into cells. In humans, the existence of 13 types of aquaporins (AQP0 to AQP12) is known. AQP3 is mainly present in epidermal cells, and is considered to have a role of taking in low-molecular compounds such as glycerol and urea involved in water retention in addition to water.

  However, AQP3 decreases with aging, and it has been suggested that this is one of the causes of a decrease in the water retention function. Therefore, by promoting the expression of AQP3, the water retention ability and barrier function due to aging can be improved. Can be controlled (see Non-Patent Document 6). As a substance having an AQP3 expression promoting action, for example, an extract from the leaf of a star fruit (see Patent Document 9) is known.

  Carbohydrates are very important as energy sources in living organisms including humans. However, on the other hand, saccharides are known to cause glycation reactions with proteins (glycation). The first step of the saccharification reaction is a non-enzymatic reaction between the carbonyl group of a carbohydrate and an amino group of a protein or the like, and finally a final saccharified product (hereinafter referred to as “AGEs”) from a Schiff base via an Amadori compound. Is a series of reactions that form Since a protein is non-enzymatically modified with a sugar by a saccharification reaction, denaturation of the protein or cross-linking between the proteins occurs, thereby reducing the function of the protein.

  The saccharification reaction causes direct damage by modifying or changing the structure of extracellular matrix constituent proteins such as collagen, and has the effect of causing a cellular response by being recognized by a receptor that uses the glycated protein as a ligand. Bring. In particular, the effects of the saccharification reaction are serious for diabetic patients having a high glucose concentration in the blood. It is known that glycation of proteins contributes to diabetic complications such as diabetic neuropathy, diabetic retinopathy, and diabetic nephropathy. In addition, it is known that a saccharification reaction in a blood vessel wall leads to progression of arteriosclerosis due to damage of endothelial cells and accumulation of denatured proteins. In addition, extracellular matrix components such as collagen account for more than half of the dry weight of tissues such as bone and skin. Therefore, for example, when collagen becomes glycated and abnormally cross-linked, osteoporosis and osteoarthritis occur in bone and cartilage tissue, and elasticity in skin and dullness due to yellowing and the like occur. . Further, abnormally cross-linked collagen and the like are hardly susceptible to degradation by collagenase or the like, so that the expression of collagenase or the like is induced and a problem such that normal collagen is degraded occurs.

  Therefore, if the glycation reaction can be suppressed in some way, for example, the formation of AGEs can be suppressed, it is useful for the prevention or treatment of the above-mentioned diseases, such as diabetic complications, arteriosclerosis, osteoporosis, and osteoarthritis. Expected to be. Furthermore, it is expected to be effective in preventing or improving skin elasticity and dullness.

  Many steroid hormones are secreted from producing organs and bind to receptors to express their effects.However, in the case of male hormones collectively called androgens, for example, testosterone enters the cells of target organs and becomes testosterone. After being reduced to 5α-dihydrotestosterone (5α-DHT) by 5α-reductase, it binds to a receptor and exerts an action as an androgen.

  Androgens are important hormones that, when acted excessively, can cause a variety of favorable effects, including androgenetic alopecia, hirsutism, seborrhea, acne (such as acne), benign prostatic hyperplasia, prostate tumors, and precocious boyhood. Induce no symptoms. Therefore, conventionally, a method of suppressing the action of excess androgen in order to improve these various symptoms, specifically, by inhibiting the action of testosterone 5α-reductase, which reduces testosterone to active 5α-DHT. In addition, a method for suppressing the generation of active 5α-DHT is known. So far, an extract from Toshiso (see Patent Document 10) and the like have been known as having testosterone 5α-reductase inhibitory activity.

  Hair grows and falls repeatedly according to a periodic hair cycle (hair cycle) consisting of a anagen, a catagen and a telogen. In this hair cycle, the stage in which a new hair follicle is formed from the telogen to the anagen is considered to be the most important for hair growth, and this stage is responsible for the proliferation and differentiation of hair follicle epithelial cells. It is thought that the dermal papilla cells play an important role. Hair papilla cells are cells located inside the root of the hair follicle wrapped in the basement membrane, inside the hair follicle epithelial cells composed of outer root sheath cells and matrix cells near the hair root. It plays an important role in the growth and differentiation of hair follicle epithelial cells and in the formation of hair, such as by acting on epithelial cells to promote their proliferation (see Non-Patent Document 7).

  Thus, hair papilla cells play an important role in the proliferation and differentiation of hair follicle epithelial cells and in the formation of hair, and it is possible to prevent or improve alopecia by promoting the proliferation of hair papilla cells. It is considered possible. Up to now, as a substance having a hair papilla cell proliferation promoting action, for example, a wild time extract (see Patent Document 11) and the like are known.

  In skin, melanin also serves to protect the body from ultraviolet radiation, but overproduction and uneven accumulation can cause skin darkening and spots. Generally, melanin is converted from tyrosine to dopa and dopa to dopaquinone by the action of an enzyme tyrosinase biosynthesized in pigment cells, and then formed via an intermediate such as 5,6-dihydroxyindophenol. ing. Therefore, in order to prevent, treat or improve skin darkness (pigmentation skin pigmentation), spots, freckles, etc., it is conceivable to suppress the production of melanin.

  Conventionally, for the prevention, treatment or improvement of skin pigmentation, spots, freckles, etc., a treatment using a whitening agent containing a chemically synthesized product such as hydroquinone as an active ingredient has been performed. However, chemically synthesized products such as hydroquinone may cause side effects such as skin irritation and allergies. Therefore, development of a whitening agent containing a highly safe natural raw material as an active ingredient is desired. As an agent having a melanin production inhibitory action, for example, an extract from a plant of the genus South urea (Saussurea) (see Patent Document 12) ) Etc. are known.

  Causes and onset of inflammatory diseases such as contact dermatitis (rash), psoriasis, pemphigus vulgaris, atopic dermatitis, various other skin inflammatory diseases accompanied by rough skin, rheumatoid arthritis, osteoarthritis, asthma, etc. The mechanisms are diverse. It is known that the cause is mainly due to an increase in the activity of hyaluronidase, the release of histamine, the increase in the activity of cyclooxygenase-2 (COX-2), and the like.

  Hyaluronidase is a hyaluronic acid hydrolase. Hyaluronate, which has an affinity for body tissues, is decomposed by ultraviolet rays, oxygen, and the like in a water-containing system, and the water retention effect decreases as the molecular weight decreases. In addition, hyaluronic acid exists as an intercellular tissue in a living body and is involved in vascular permeability. Furthermore, hyaluronidase, which is present in mast cells, is released by degranulation caused by its activation and acts as an inflammatory chemical mediator. Therefore, by inhibiting the activity of hyaluronidase, enhancement of moisturizing and prevention or reduction of inflammation are expected. As a substance having a hyaluronidase activity inhibitory action, for example, an extract from a plant of the genus Osbeckia (see Patent Document 13) is known.

  Histamine release is a phenomenon in which histamine in mast cells is released outside the cells, and the released histamine causes an inflammatory response. Therefore, attempts have been made to prevent or treat allergic diseases and inflammatory diseases with substances that inhibit or suppress histamine release. However, it is difficult to directly evaluate the release of histamine, and the release of histamine can be evaluated using the release of hexosaminidase, which has been confirmed to be released simultaneously with the release of histamine, as an index. Therefore, by suppressing the release of hexosaminidase, the release of histamine can be suppressed at the same time, which is considered to be effective in preventing, treating or ameliorating inflammatory diseases and the like.

  In addition, histamine mediates communication between cells as a local mediator, enhances gastric acid secretion in the digestive tract, functions as a neurotransmitter in the central nervous system, and contributes to maintaining awake state. Have been. Here, excessive histamine release causes ulcers due to excessive gastric acid in the digestive tract and contributes to sleep disorders in the central nervous system. As described above, by suppressing the release of hexosaminidase, the release of histamine can be suppressed at the same time, so that it is considered that this can prevent, treat, or ameliorate gastric ulcers caused by hyperacidity, sleep disorders, and the like. . As a substance having a hexosaminidase release inhibitory action, for example, an extract from wisteria tea (see Patent Document 14) is known.

  Inflammation is a complex reaction that exhibits symptoms such as redness, edema, fever, pain, dysfunction and the like. Microscopically, inflammation consists of common reactions such as vascular reactions that cause plasma leakage, infiltration of leukocytes, tissue destruction by inflammatory cells, and involvement of the central nervous system such as fever and hyperalgesia. Reactions can also be triggered. Prostaglandins play an important role in the individual reactions of such inflammation, and it is clear that the production of prostaglandins during this inflammation mainly involves cyclooxygenase-2, an inducible cyclooxygenase. It has become. For this reason, many cyclooxygenase inhibitors represented by aspirin have been used for the purpose of preventing and preventing inflammatory reactions (see Non-Patent Document 8).

  As for dihydroferulic acid, it is known that when certain lactic acid bacteria are cultured in a medium containing ferulic acid or ethyl ferulate, dihydroferulic acid is detected (Non-Patent Document 9, Patent Document 15). reference).

JP 2006-056855 A JP-A-7-274832 JP 2006-063033 A JP 2007-217352 A JP 2006-265232 A JP-A-2003-146837 JP 2005-022993 A JP 2006-056854 A JP 2009-191039 A JP 2010-184915 A JP 2006-219407 A JP-A-2002-20112 JP-A-2003-055242 JP-A-2003-012532 JP 2014-003929 A

"Japanese Pharmacological Magazine", 2005, Vol.125, p.379-384 J. Cell. Biol., 1992, Vol.199, p.695-703. J. Invest. Dermatol., 1979, Vol. 73, p. 59-66. Nature, 1996, Vol.379, p.335-339 "Angiogenesis and matrix metalloprotease", The 120th Annual Meeting of the Japan Medical Association Symposium, Fundamentals and clinical aspects of angiogenesis, December 13, 2001, p.43-49 "Fragrance Journal", 2006, Vol.34, p.19-23 Trends Genet., 1992, Vol.8, p.55-61 “Atlas of Pharmacology”, translated by Takehiko Fukuhara, Bunkodo, 1995, p.184 J. Sci. Food Agric., 2012, Vol.92, pp.2291-2296

  The present invention provides an anti-obesity effect, a cyclic AMP phosphodiesterase activity inhibitory effect, a dipeptidyl peptidase IV activity inhibitory effect, an anti-aging effect, a hair-growth effect, an anti-androgenic effect, a whitening effect, or an anti-inflammation among compounds derived from natural products. An anti-obesity agent, a cyclic AMP phosphodiesterase activity inhibitor, a dipeptidyl peptidase IV activity inhibitor, an anti-aging agent, a hair restorer, an anti-androgen, a whitening agent, and an anti-obesity agent containing the active ingredient as an active ingredient. An object of the present invention is to provide an inflammatory agent, a skin cosmetic, a hair cosmetic, and a food or beverage containing the compound.

  In order to solve the above problems, an antiobesity agent, a cyclic AMP phosphodiesterase activity inhibitor, a dipeptidyl peptidase IV activity inhibitor, an antiaging agent, a hair restorer, an antiandrogen, a whitening agent, and an antiinflammatory agent of the present invention are provided. Is characterized by using dihydroferulic acid as an active ingredient. Further, the skin cosmetic, the hair cosmetic, and the food and drink of the present invention are characterized by containing dihydroferulic acid.

  According to the present invention, by using dihydroferulic acid as an active ingredient, an anti-obesity agent, a cyclic AMP phosphodiesterase activity inhibitor, a dipeptidyl peptidase IV activity inhibitor, an anti-aging agent, a hair restorer, Androgenic, whitening, and anti-inflammatory agents can be provided. Furthermore, by blending dihydroferulic acid, skin cosmetics, hair cosmetics, and foods and drinks excellent in the above effects can be provided.

Hereinafter, embodiments of the present invention will be described.
The anti-obesity agent, cyclic AMP phosphodiesterase activity inhibitor, dipeptidyl peptidase IV activity inhibitor, anti-aging agent, hair restorer, anti-androgen, whitening agent, and anti-inflammatory agent of the present embodiment are effective for dihydroferulic acid Component. In addition, the skin cosmetic, the hair cosmetic, and the food and drink of the present embodiment contain dihydroferulic acid.

  Dihydroferulic acid is a cinnamic acid derivative having a chemical structure represented by the following formula.

  Dihydroferulic acid is obtained, for example, by fermenting ferulic acid derivatives such as ferulic acid or ethyl ferulate, or a composition containing these (eg, crushed plant extracts or extracts) with a microorganism having phenolic acid reductase. After the conversion of ferulic acid to dihydroferulic acid, the fermented product obtained can be extracted, isolated and purified to produce the fermented product. Examples of the composition containing ferulic acid include crushed products and extracts of plants such as coffee, wheat, corn, tomato, mate, mugwort, and burdock. Furthermore, since ferulic acid is a constituent of lignin in woody plants, lignin or a composition containing the same may be used as the composition containing ferulic acid. On the other hand, examples of microorganisms having a phenolic acid reductase include, for example, Lactobacillus plantarum, Lactobacillus fermentum, Lactobacillus gasseri, Lactobacillus johnsonii, Lactobacillus crispatus, Lactobacillus acidophilus, Lactobacillus amylovorus, Lactobacillus amylovorus, Lactobacillus delcileck, Lactobacillus delcileck, Lactobacillus delcileck, Lactobacillus delbrueckii, Lactobacillus delbrueckii. Lactic acid bacteria.

  The method for extracting, isolating and purifying dihydroferulic acid from the above plant or fermented product is not particularly limited, and can be performed according to a conventional method. For example, in the extraction treatment, the above plant or fermented product as an extraction raw material may be dried and then pulverized as it is or using a crusher, and then subjected to extraction with an extraction solvent. Drying may be performed in the sun or using a commonly used dryer. Alternatively, the material may be subjected to a pretreatment such as degreasing with a nonpolar solvent such as hexane, and then used as an extraction material. By performing a pretreatment such as degreasing, an extraction treatment with a polar solvent can be efficiently performed.

  As the extraction solvent, it is preferable to use a polar solvent, for example, water, a hydrophilic organic solvent, and the like. These may be used alone or in combination of two or more, and used at room temperature or a temperature lower than the boiling point of the solvent. Is preferred.

  The water that can be used as the extraction solvent includes pure water, tap water, well water, mineral spring water, mineral water, hot spring water, spring water, fresh water, and the like, as well as those subjected to various treatments. The treatment applied to water includes, for example, purification, heating, sterilization, filtration, ion exchange, osmotic pressure adjustment, buffering, and the like. Therefore, water that can be used as the extraction solvent in the present embodiment includes purified water, hot water, ion-exchanged water, physiological saline, phosphate buffer, phosphate buffered saline, and the like.

  Examples of the hydrophilic organic solvent that can be used as the extraction solvent include lower aliphatic alcohols having 1 to 5 carbon atoms such as methanol, ethanol, propyl alcohol and isopropyl alcohol; lower aliphatic ketones such as acetone and methyl ethyl ketone; 1,3-butylene Examples thereof include polyhydric alcohols having 2 to 5 carbon atoms such as glycol, propylene glycol, and glycerin.

  When a mixture of two or more polar solvents is used as the extraction solvent, the mixing ratio can be adjusted appropriately. For example, when a mixture of water and a lower aliphatic alcohol is used as an extraction solvent, the mixing ratio of water and the lower aliphatic alcohol is preferably 9: 1 to 1: 9 (volume ratio), More preferably, the ratio is 7: 3 to 2: 8 (volume ratio). When a mixture of water and lower aliphatic ketone is used, the mixing ratio of water and lower aliphatic ketone is preferably 9: 1 to 2: 8 (volume ratio), and water and lower aliphatic ketone are preferably used. When a mixed solution with a polyhydric alcohol is used, the mixing ratio between water and the polyhydric alcohol is preferably 5: 5 to 1: 9 (volume ratio).

  The extraction treatment is not particularly limited as long as the soluble component contained in the extraction raw material can be eluted into the extraction solvent, and can be performed according to a conventional method. For example, the extraction material is immersed in 5 to 15 times (mass ratio) extraction solvent of the extraction material, and the soluble component is extracted at room temperature or under reflux, and then filtered to remove the extraction residue. A liquid can be obtained. When the solvent is distilled off from the obtained extract, a paste-like concentrate is obtained, and when this concentrate is further dried, a dry product is obtained.

  The method for isolating and purifying dihydroferulic acid from the extract obtained as described above, a concentrate of the extract or a dried product of the extract is not particularly limited, and may be performed by a conventional method. Can be. For example, the extract is dissolved in a developing solvent, and subjected to column chromatography using a porous substance such as silica gel or alumina, a porous resin such as styrene-divinylbenzene copolymer or polymethacrylate to obtain dihydroferulic acid. And the like. In this case, the developing solvent may be appropriately selected depending on the stationary phase to be used. For example, when the extract is separated by normal phase chromatography using silica gel as the stationary phase, chloroform: methanol = 95: 5 and the like. Further, the fraction containing dihydroferulic acid obtained by column chromatography can be subjected to optional methods such as reverse phase silica gel chromatography using ODS, recrystallization, liquid-liquid countercurrent extraction, and column chromatography using an ion exchange resin. The organic compound may be purified by using an organic compound purifying means.

[Anti-obesity agent, cyclic AMP phosphodiesterase activity inhibitor, dipeptidyl peptidase IV activity inhibitor, anti-aging agent, hair restorer, anti-androgen, whitening agent, anti-inflammatory agent]
The dihydroferulic acid obtained as described above has excellent anti-obesity action, cyclic AMP (cAMP) phosphodiesterase activity inhibitory action, dipeptidyl peptidase IV (DPPIV) activity inhibitory action, anti-aging action, hair growth action, anti-androgen Anti-obesity agent, cAMP phosphodiesterase activity inhibitor, DPPIV activity inhibitor, anti-aging agent, hair restorer, anti-androgen, whitening agent, and anti-inflammatory agent Can be used as an active ingredient. The anti-obesity agent, cAMP phosphodiesterase activity inhibitor, DPPIV activity inhibitor, anti-aging agent, hair restorer, anti-androgen, whitening agent, and anti-inflammatory agent of the present embodiment include pharmaceuticals, quasi-drugs, cosmetics, and the like. Can be used for a wide range of applications.

  As an active ingredient of the antiobesity agent, cAMP phosphodiesterase activity inhibitor, DPPIV activity inhibitor, antiaging agent, hair restorer, antiandrogen, whitening agent, and antiinflammatory agent according to the present embodiment, A composition containing dihydroferulic acid may be used instead of ferulic acid. Here, the “composition containing dihydroferulic acid” in the present embodiment includes an extract obtained using a plant containing dihydroferulic acid as an extraction raw material, a fermented product containing dihydroferulic acid, and a fermented product containing dihydroferulic acid. The extract obtained as an extraction raw material is included. Further, the “extract” includes an extract obtained by the extraction treatment, a diluent or a concentrate of the extract, or a dried product obtained by drying the extract.

  Contains dihydroferulic acid as an active ingredient of the antiobesity agent, cAMP phosphodiesterase activity inhibitor, DPPIV activity inhibitor, antiaging agent, hair restorer, antiandrogen, whitening agent, and antiinflammatory agent according to the present embodiment. When a composition is used, it is preferable to use a purified one in which the purity of dihydroferulic acid is increased. Anti-obesity agent, cAMP phosphodiesterase activity inhibitor, DPPIV activity inhibitor, anti-aging agent, hair restorer, anti-androgen, which are more excellent in action and effect by using a substance having a higher purity of dihydroferulic acid as an active ingredient Agents, whitening agents, and anti-inflammatory agents can be obtained.

  The anti-obesity effect of dihydroferulic acid is, for example, one or more selected from the group consisting of cAMP phosphodiesterase activity inhibitory effect, blood cholesterol lowering effect, blood free fatty acid lowering effect, and DPPIV activity inhibitory effect. It is exerted based on the action. However, the anti-obesity effect of dihydroferulic acid is not limited to the anti-obesity effect exerted on the basis of the above effect.

  In addition, dihydroferulic acid is a blood lipid-reducing agent, a blood cholesterol-reducing agent, or a blood free fatty acid-reducing agent, utilizing its blood lipid lowering action (particularly, blood cholesterol lowering action and blood free fatty acid lowering action). You may use as an active ingredient of an agent.

  The anti-aging effect of dihydroferulic acid is, for example, a laminin 5 production promoting effect, a matrix metalloproteinase-2 (MMP-2) activity inhibiting effect, a hyaluronic acid production promoting effect, an epidermal keratinocyte proliferation promoting effect, and an elastin production promoting effect. 1 selected from the group consisting of: a matrix metalloprotease-1 (MMP-1) activity inhibitory action, a fibroblast proliferation promoting action, an aquaporin 3 (AQP3) mRNA expression promoting action, and an advanced glycation product (AGEs) formation inhibitory action. It is exerted on the basis of the action of two or more species. However, the anti-aging effect of dihydroferulic acid is not limited to the anti-aging effect exerted based on the above-mentioned effect.

  In addition, dihydroferulic acid has a laminin 5 production promoting action, an MMP-2 activity inhibiting action, a hyaluronic acid production promoting action, an epidermal keratinocyte proliferation promoting action, an elastin production promoting action, an MMP-1 activity inhibiting action, and a fibroblast. Laminin 5 production promoter, MMP-2 activity inhibitor, hyaluronic acid production promoter, epidermal keratinocyte proliferation promoter, elastin production promotion, respectively, utilizing the growth promoting action, AQP3 mRNA expression promoting action, or AGEs formation inhibiting action It may be used as an active ingredient of an agent, an MMP-1 activity inhibitor, a fibroblast proliferation promoter, an AQP3 mRNA expression promoter, or an AGEs formation inhibitor.

  The hair growth effect of dihydroferulic acid is exerted, for example, based on the testosterone 5α-reductase activity inhibitory activity and / or the hair papilla cell proliferation promoting activity. However, the hair-growth action of dihydroferulic acid is not limited to the hair-growth action exerted on the basis of the above-mentioned action. Also, dihydroferulic acid may be used as an active ingredient of a testosterone 5α-reductase activity inhibitor or a hair papilla cell growth promoter, respectively, by utilizing its testosterone 5α-reductase activity inhibitory action or hair papilla cell growth promoting action. Good.

  The antiandrogenic action of dihydroferulic acid is exerted, for example, based on the testosterone 5α-reductase activity inhibitory action. However, the antiandrogenic action of dihydroferulic acid is not limited to the antiandrogenic action exerted based on the above action.

  The whitening effect of dihydroferulic acid is exerted based on, for example, a melanin production inhibitory effect. However, the whitening effect of dihydroferulic acid is not limited to the whitening effect exerted based on the above-mentioned effect. Further, dihydroferulic acid may be used as an active ingredient of a melanin production inhibitor by utilizing its melanin production inhibitory action.

  The anti-inflammatory action of dihydroferulic acid is, for example, one or more selected from the group consisting of a hyaluronidase activity inhibitory action, a hexosaminidase release inhibitory action, and a cyclooxygenase-2 (COX-2) activity inhibitory action. It is exerted based on the action of However, the anti-inflammatory action of dihydroferulic acid is not limited to the anti-inflammatory action exerted based on the above action. In addition, dihydroferulic acid is a hyaluronidase activity inhibitor, a hexosaminidase release inhibitor, or a COX-2 activity inhibitory effect, and is used as a hyaluronidase activity inhibitor, a hexosaminidase release inhibitor, or a COX- (2) It may be used as an active ingredient of an activity inhibitor.

  The antiobesity agent, cAMP phosphodiesterase activity inhibitor, DPPIV activity inhibitor, antiaging agent, hair restorer, antiandrogen, whitening agent, or antiinflammatory agent of the present embodiment contains dihydroferulic acid or dihydroferulic acid The composition may be composed of the composition alone, or may be a composition prepared from dihydroferulic acid or a composition containing dihydroferulic acid.

  The anti-obesity agent, cAMP phosphodiesterase activity inhibitor, DPPIV activity inhibitor, anti-aging agent, hair restorer, anti-androgen, whitening agent, and anti-inflammatory agent of the present embodiment are pharmaceutically acceptable such as dextrin, cyclodextrin and the like. It can be formulated into any dosage form such as powder, granule, tablet, liquid, etc. by using a carrier and other auxiliaries which can be used in accordance with a conventional method. At this time, as an auxiliary agent, for example, an excipient, a binder, a disintegrant, a lubricant, a stabilizer, a flavor / odorant, and the like can be used. Anti-obesity agents, cAMP phosphodiesterase activity inhibitors, DPPIV activity inhibitors, anti-aging agents, hair growth agents, anti-androgens, whitening agents, and anti-inflammatory agents can be used in other compositions (eg, skin cosmetics, hair cosmetics) Etc.), and can be used as ointments, liquids for external use, patches and the like.

  When the antiobesity agent, cAMP phosphodiesterase activity inhibitor, DPPIV activity inhibitor, antiaging agent, hair restorer, antiandrogen, whitening agent, or antiinflammatory agent of the present embodiment is formulated, dihydroferulic acid or dihydroferula The content of the composition containing an acid is not particularly limited, and can be appropriately set according to the purpose.

  The anti-obesity agent, cAMP phosphodiesterase activity inhibitor, DPPIV activity inhibitor, anti-aging agent, hair restorer, anti-androgen, whitening agent, or anti-inflammatory agent of the present embodiment may have an anti-obesity effect, if necessary. And other natural extracts having cAMP phosphodiesterase activity inhibitory action, DPPIV activity inhibitory action, anti-aging action, hair growth action, anti-androgen action, whitening action, or anti-inflammatory action, containing dihydroferulic acid or dihydroferulic acid And used as an active ingredient.

  The method of administering the anti-obesity agent, cAMP phosphodiesterase activity inhibitor, DPPIV activity inhibitor, anti-aging agent, hair restorer, anti-androgen, whitening agent, or anti-inflammatory agent of the present embodiment to a patient includes transdermal administration, Oral administration and the like can be mentioned, but a method suitable for prevention or treatment thereof may be appropriately selected depending on the type of the disease. In addition, the dosage of the anti-obesity agent, cAMP phosphodiesterase activity inhibitor, DPPIV activity inhibitor, anti-aging agent, hair restorer, anti-androgen, whitening agent, or anti-inflammatory agent of the present embodiment depends on the type of the disease, The dose may be appropriately increased or decreased depending on the degree, individual difference between patients, administration method, administration period, and the like.

  The antiobesity agent of the present embodiment is selected from the group consisting of cAMP phosphodiesterase activity inhibitory action, blood cholesterol lowering action, blood free fatty acid reducing action, and DPPIV activity inhibitory action of dihydroferulic acid as an active ingredient. Through the action of one or more species, it can promote cAMP production, promote fat breakdown in adipocytes, reduce blood lipid levels, and prolong the half-life of incretin in vivo can do. As a result, the anti-obesity agent of the present embodiment can prevent or ameliorate various diseases such as obesity, associated arteriosclerosis, diabetes, and metabolic syndrome. However, the anti-obesity agent of the present embodiment has a blood cholesterol-reducing action or a blood free fatty acid-reducing action in addition to the above-mentioned use as a cAMP phosphodiesterase activity inhibitor or a DPPIV activity inhibitor. It can be used for any purpose that is meaningful.

  For example, the anti-obesity agent of the present embodiment or the blood lipid-lowering agent, blood cholesterol-lowering agent, or blood free fatty acid-lowering agent described above is used for a lipid disorder not caused by obesity (for example, blood cholesterol or blood cholesterol). It is possible to prevent or ameliorate various diseases such as arteriosclerosis without obesity, diabetes, and metabolic syndrome without obesity.

  The cAMP phosphodiesterase activity inhibitor of the present embodiment promotes cAMP production through the inhibitory activity of cAMP phosphodiesterase activity of dihydroferulic acid as an active ingredient, and thus can suppress aggregation of platelets. Various inflammatory diseases and the like can be prevented, treated, or improved. In addition, the cAMP phosphodiesterase activity inhibitor of the present embodiment can promote the degradation of fat in adipocytes through the cAMP phosphodiesterase activity inhibitory action of dihydroferulic acid, resulting in obesity and arteriosclerosis associated therewith. Can prevent or improve various lifestyle-related diseases such as diabetes, metabolic syndrome and the like. However, the cAMP phosphodiesterase activity inhibitor of the present embodiment can be used for all other uses that are significant in exhibiting a cAMP phosphodiesterase activity inhibitory action in addition to these uses.

  The DPPIV activity inhibitor of the present embodiment can prevent or treat type 2 diabetes, obesity, hypertension, insulin resistance, etc. through the DPPIV activity inhibitory action of dihydroferulic acid, and autoimmune such as rheumatoid arthritis. Disease or transplant rejection can be prevented or treated. However, the DPPIV activity inhibitor of the present embodiment can be used for all purposes other than those uses that are significant in exhibiting a DPPIV activity inhibitory action. For example, the DPPIV activity inhibitor of the present embodiment uses the DPPIV activity inhibitory action of dihydroferulic acid to exert a neuropathy such as pain, neurodegenerative disease, and neuropsychiatric disease (eg, sciatica, Alzheimer's disease, depression, etc.); Growth hormone deficiency and diseases where growth hormone is used for treatment; cancer (eg, T-cell lymphoma, acute lymphoblastic leukemia, thyroid cancer, basal cell carcinoma, breast cancer, etc.); diseases such as HIV infection (AIDS) Can be prevented or treated.

  The anti-aging agent of this embodiment has a laminin 5 production promoting action, an MMP-2 activity inhibiting action, a hyaluronic acid production promoting action, an epidermal keratinocyte proliferation promoting action, an elastin production promoting action of dihydroferulic acid as an active ingredient, Through one or more actions selected from the group consisting of MMP-1 activity inhibitory action, fibroblast proliferation promoting action, AQP3 mRNA expression promoting action, and AGEs formation inhibitory action, formation of skin wrinkles, elasticity It can prevent, treat or ameliorate skin aging symptoms such as a decrease and a decrease in moisturizing function. However, in addition to these uses, the anti-aging agent of the present embodiment has a laminin 5 production promoting action, an MMP-2 activity inhibiting action, a hyaluronic acid production promoting action, an epidermal keratinocyte proliferation promoting action, an elastin production promoting action, an MMP -1 activity inhibitory action, fibroblast proliferation promoting action, AQP3 mRNA expression promoting action or AGEs formation inhibitory action.

  For example, the anti-aging agent of the present embodiment or the above-mentioned laminin 5 production promoter can promote laminin 5 production by the action of dihydroferulic acid. Thereby, reconstruction of the basement membrane structure can be induced, and wounds on the skin can be treated and improved. In addition, the anti-aging agent of the present embodiment or the above-described laminin 5 production promoter can be used as a preventive or therapeutic agent for a disease (eg, epidermolysis bullosa) caused by laminin 5 deficiency (deficiency).

  In addition to the above-mentioned uses, the anti-aging agent of the present embodiment or the above-mentioned MMP-2 activity inhibitor, or the above-mentioned MMP-1 activity inhibitor has an MMP-2 activity inhibitory action or an MMP-1 activity inhibitory action of dihydroferulic acid. And other diseases whose destruction of the extracellular matrix is known to be associated with the pathology (eg, cancer invasion / metastasis, rheumatoid arthritis, knee osteoarthritis, periodontal disease, age-related macular degeneration, etc.) ) Can be used for treatment and prevention. Further, the anti-aging agent or the MMP-2 activity inhibitor of the present embodiment can suppress angiogenesis and suppress the growth of tumor cells by its MMP-2 activity inhibitory action.

  In addition to the above-mentioned uses, the anti-aging agent of the present embodiment or the above-mentioned hyaluronic acid production promoting agent, due to its hyaluronic acid production promoting action, has rheumatoid arthritis, osteoarthritis, purulent arthritis, gouty arthritis, traumatic arthritis. For preventing, treating or ameliorating arthritis such as osteoarthritis; promoting the healing of wounds or burns; Further, the anti-aging agent of the present embodiment or the above-mentioned epidermal keratinocyte proliferation promoter restores the metabolism of the skin through the epidermal keratinocyte proliferation promoting action of dihydroferulic acid, and produces fine lines, dullness, pigmentation, etc. For prevention, treatment or improvement of reproductive medicine; regenerative medicine; and the like.

  In addition to the above-mentioned uses, the anti-aging agent of the present embodiment or the above-mentioned elastin production promoting agent prevents and treats pulmonary diseases such as emphysema; vascular diseases such as hypertension and aneurysm due to its elastin production promoting action. Or it can be used for improvement applications. In addition, the anti-aging agent of the present embodiment or the above-mentioned fibroblast proliferation promoter promotes healing of wounds such as trauma and burns through the fibroblast proliferation promotion action of dihydroferulic acid; skin diseases (eg, pressure ulcer, It can be used for the treatment or improvement of burn ulcers, diabetic ulcers and other skin ulcers), regenerative medicine, and the like.

  In addition to the above-mentioned uses, the anti-aging agent of the present embodiment or the above-mentioned AQP3 mRNA expression promoter can improve aging-related water retention ability and barrier function through the AQP3 mRNA expression promoting action of dihydroferulic acid.

  In addition to the above-mentioned uses, the anti-aging agent of the present embodiment or the above-mentioned AGEs formation inhibitor can be used to reduce the AGEs formation inhibitory effect of dihydroferulic acid, such as diabetic neuropathy, diabetic retinopathy, and diabetic nephropathy. Complications; arteriosclerosis caused by protein saccharification; osteoporosis and osteoarthritis caused by protein saccharification can be prevented or treated. In addition, the anti-aging agent of the present embodiment or the above-mentioned AGEs formation inhibitor can prevent or suppress hair damage due to a saccharification reaction of a protein through its AGEs formation inhibitory action, whereby the hair becomes rough. Can be prevented or suppressed, the elasticity or suppleness of the hair can be restored, and firmness and firmness can be imparted to the hair.

  The hair restorer of the present embodiment is capable of reducing the activity of dihydroferulic acid, which is an active ingredient, to inhibit the activity of testosterone 5α-reductase and / or promote the growth of hair dermal papilla cells. It can prevent, treat or ameliorate the symptoms and the like, and is particularly suitable for preventing, treating or ameliorating male pattern baldness. However, the hair restorer of the present invention can be used for all other uses that are significant in exhibiting a testosterone 5α-reductase activity inhibitory action or a hair papilla cell growth promoting action.

  For example, the hair restorer of the present embodiment or the above-described hair papilla cell growth promoter activates hair papilla cells through the hair papilla cell growth promotion action of dihydroferulic acid, and proliferates and differentiates hair follicle epithelial cells and hair. Can be promoted, and the transition from the anagen to the catagen and the telogen in the hair cycle can be prevented, and the anagen can be prolonged. Thereby, alopecia can be prevented or ameliorated. Further, the hair restorer of the present embodiment or the above-mentioned hair papilla cell growth promoter is used for application in the field of regenerative medicine, such as hair regeneration using hair papilla cells, through the hair papilla cell growth promotion action of dihydroferulic acid. You can also.

  The anti-androgen agent of the present embodiment or the testosterone 5α-reductase activity inhibitor described above is a disease in which androgen is involved, such as male pattern hair loss, through the testosterone 5α-reductase activity inhibitory activity of dihydroferulic acid as an active ingredient. It can prevent, treat, or ameliorate sickness, hirsutism, seborrhea, acne (such as acne), benign prostatic hyperplasia, prostate tumor, precocious boyhood, and the like. However, the antiandrogens of the present invention can be used for all other uses that are significant in exhibiting an inhibitory effect on testosterone 5α-reductase activity in addition to these uses.

  The whitening agent of the present embodiment can prevent or improve skin darkening, pigmentation such as spots and freckles through the melanin production inhibitory action of dihydroferulic acid as an active ingredient. However, the whitening agent of the present embodiment can be used for any purpose other than these uses that is significant in exhibiting a melanin production inhibitory action.

  The anti-inflammatory agent of the present embodiment is one or two selected from the group consisting of a hyaluronidase activity inhibitory action, a hexosaminidase release inhibitory action, and a COX-2 activity inhibitory action of dihydroferulic acid as an active ingredient. Through the above actions, contact dermatitis (rash), psoriasis, pemphigus vulgaris, atopic dermatitis, and various other skin inflammatory diseases associated with rough skin can be prevented, treated, or ameliorated. However, the anti-inflammatory agent of the present embodiment is used for all purposes that are significant in exhibiting a hyaluronidase activity inhibitory action, a hexosaminidase release inhibitory action, or a COX-2 activity inhibitory action in addition to these uses. be able to.

  For example, the anti-inflammatory agent of the present embodiment or the above-mentioned hyaluronidase activity inhibitor, hexosaminidase release inhibitor, or COX-2 activity inhibitor is a dihydroferulic acid-containing hyaluronidase activity inhibitory action, hexosaminidase release inhibition It can prevent, treat or ameliorate rheumatoid arthritis, osteoarthritis, asthma, etc. through its action or COX-2 activity inhibitory action. In addition, the anti-inflammatory agent of the present embodiment or the above-mentioned hexosaminidase release inhibitor prevents and treats gastric ulcers, sleep disorders, etc. caused by excessive gastric acid through the hexosaminidase release inhibitory action of dihydroferulic acid. Or can be improved.

  Further, the anti-obesity agent, cAMP phosphodiesterase activity inhibitor, DPPIV activity inhibitor, anti-aging agent, hair restorer, anti-androgen, whitening agent, or anti-inflammatory agent of the present embodiment has excellent anti-obesity effect, cAMP phosphodiesterase Since it has an activity inhibitory action, a DPPIV activity inhibitory action, an anti-aging action, a hair-growth action, an anti-androgen action, a whitening action, or an anti-inflammatory action, it is suitable to be incorporated into, for example, a skin external preparation. In this case, dihydroferulic acid or a composition containing dihydroferulic acid may be blended as it is, or an anti-obesity agent formulated from a composition containing dihydroferulic acid or dihydroferulic acid, a cAMP phosphodiesterase activity inhibitor , A DPPIV activity inhibitor, an anti-aging agent, a hair restorer, an anti-androgen, a whitening agent, or an anti-inflammatory agent.

  Here, the external preparation for skin is not limited in its category, and includes a wide range of quasi-drugs, pharmaceuticals, and the like, which are used transdermally, in addition to the skin cosmetics described below.

  Further, the anti-obesity agent, cAMP phosphodiesterase activity inhibitor, DPPIV activity inhibitor, anti-aging agent, hair restorer, anti-androgen, whitening agent, or anti-inflammatory agent of the present embodiment has excellent anti-obesity effect, cAMP phosphodiesterase It has an activity inhibitory action, DPPIV activity inhibitory action, anti-aging action, hair-growth action, anti-androgen action, whitening action, or anti-inflammatory action, so that it can be suitably used as a reagent for studies on these action mechanisms. it can.

[Skin cosmetics, hair cosmetics]
Dihydroferulic acid has excellent anti-aging, hair-growth, anti-androgen, whitening, anti-inflammatory, cAMP phosphodiesterase activity, DPPIV activity, and anti-obesity effects. It is suitable for blending into cosmetics or hair cosmetics. In this case, dihydroferulic acid or a composition containing dihydroferulic acid may be blended as it is, or an anti-aging agent, a hair restorer, an anti-androgen formulated from a composition containing dihydroferulic acid or dihydroferulic acid An agent, a whitening agent, an anti-inflammatory agent, a cAMP phosphodiesterase activity inhibitor, a DPPIV activity inhibitor, or an anti-obesity agent may be added.

  Skin cosmetics or hair by blending dihydroferulic acid or the above anti-aging agent, hair restorer, anti-androgen, whitening agent, anti-inflammatory agent, cAMP phosphodiesterase activity inhibitor, DPPIV activity inhibitor, or anti-obesity agent Cosmetics have anti-aging, laminin 5 production promoting, MMP-2 activity inhibiting, hyaluronic acid production promoting, epidermal keratinocyte proliferation promoting, elastin production promotion, MMP-1 activity inhibiting, fibroblast proliferation Promoting action, AQP3 mRNA expression promoting action, AGEs formation suppressing action, hair growth action, testosterone 5α-reductase activity inhibitory action, hair papilla cell growth promoting action, anti-androgen action, whitening action, melanin production inhibitory action, anti-inflammatory action, hyaluronidase activity Inhibitory action, hexosaminidase release inhibitory action, COX-2 activity Adverse effects, cAMP phosphodiesterase inhibitory activity, DPPIV inhibitory activity, anti-obesity effect can be imparted to blood cholesterol lowering action, or blood free fatty acid lowering action.

  All of these effects exert favorable effects when applied to skin cosmetics or hair cosmetics. Among them, anti-aging effect, laminin 5 production promoting effect, MMP-2 activity inhibitory effect, hyaluronic acid Acid production promoting action, epidermal keratinocyte proliferation promoting action, elastin production promoting action, MMP-1 activity inhibiting action, fibroblast proliferation promoting action, AQP3 mRNA expression promoting action, AGEs formation suppressing action, whitening action, melanin production suppressing action, An anti-inflammatory action, a hyaluronidase activity inhibitory action, a hexosaminidase release inhibitory action, a COX-2 activity inhibitory action, a cAMP phosphodiesterase activity inhibitory action, a testosterone 5α-reductase activity inhibitory action, or an antiandrogenic action is imparted to skin cosmetics. Is particularly preferable because their effects are easily exhibited.

  In addition, among the above-mentioned actions, hair growth action, testosterone 5α-reductase activity inhibitory action, hair papilla cell proliferation promoting action, antiandrogen action, anti-inflammatory action, hyaluronidase activity inhibitory action, hexosaminidase release inhibitory action, COX- 2 activity inhibitory action, cAMP phosphodiesterase activity inhibitory action, laminin 5 production promotion action, hyaluronic acid production promotion action, elastin production promotion action, fibroblast cell growth promotion action, or AGEs formation suppression action is imparted to the hair cosmetic, It is particularly preferable because those functions are easily exerted.

  Skin cosmetics or hair makeup which can contain dihydroferulic acid, or the above-mentioned anti-aging agent, hair restorer, anti-androgen, whitening agent, anti-inflammatory agent, cAMP phosphodiesterase activity inhibitor, DPPIV activity inhibitor, or anti-obesity agent The type of ingredients is not particularly limited, and examples of skin cosmetics include ointments, creams, emulsions, lotions, packs, foundations, and the like. Examples of hair cosmetics include hair tonics and hair Cream, hair liquid, shampoo, pomade, rinse and the like.

  Dihydroferulic acid, or the above anti-aging agent, hair restorer, anti-androgen, whitening agent, anti-inflammatory agent, cAMP phosphodiesterase activity inhibitor, DPPIV activity inhibitor, or anti-obesity agent mixed in skin cosmetics or hair cosmetics In this case, the compounding amount can be appropriately adjusted according to the type of the skin cosmetic or the hair cosmetic, but the preferable compounding ratio is about 0.0001 to 10% by mass, and the particularly preferable compounding ratio is Is about 0.001 to 1% by mass in terms of a standard extract.

  The skin cosmetic or the hair cosmetic of the present embodiment has an anti-aging effect, a laminin 5 production promoting effect, an MMP-2 activity inhibiting effect, a hyaluronic acid production promoting effect, an epidermal keratinocyte proliferation promoting effect, and elastin possessed by dihydroferulic acid. Production promoting action, MMP-1 activity inhibiting action, fibroblast proliferation promoting action, AQP3 mRNA expression promoting action, AGEs formation inhibiting action, hair growth action, testosterone 5α-reductase activity inhibitory action, hair papilla cell growth promoting action, antiandrogen action Whitening action, melanin production inhibitory action, anti-inflammatory action, hyaluronidase activity inhibitory action, hexosaminidase release inhibitory action, COX-2 activity inhibitory action, cAMP phosphodiesterase activity inhibitory action, DPPIV activity inhibitory action, anti-obesity action, blood Cholesterol lowering action or blood free fatty acid reduction As long as it does not hinder, the main ingredients, auxiliaries or other ingredients used in the manufacture of normal skin cosmetics or hair cosmetics, such as astringents, bactericidal / antibacterial agents, whitening agents, ultraviolet absorbers, humectants, cell activation Agents, anti-inflammatory / antiallergic agents, antioxidant / active oxygen removers, oils and fats, waxes, hydrocarbons, fatty acids, alcohols, esters, surfactants, fragrances and the like can be used in combination. The combined use in this way results in a more general product, and a synergistic effect with the other active ingredient used in combination may bring about an excellent effect more than expected.

  The skin cosmetic of this embodiment has an anti-aging effect, a laminin 5 production promoting effect, an MMP-2 activity inhibiting effect, a hyaluronic acid production promoting effect, an epidermal keratinocyte proliferation promoting effect, an elastin production promoting effect of dihydroferulic acid, MMP-1 activity inhibitory action, fibroblast proliferation promotion action, AQP3 mRNA expression promotion action, AGEs formation suppression action, whitening action, melanin production suppression action, anti-inflammatory action, hyaluronidase activity inhibition action, hexosaminidase release inhibition action, COX -2 activity inhibitory action, cAMP phosphodiesterase activity inhibitory action, hair growth action, testosterone 5α-reductase activity inhibitory action, hair papilla cell growth promoting action, anti-androgen action, DPPIV activity inhibitory action, anti-obesity action, blood cholesterol lowering action, And blood free fatty acid reduction Prevention, treatment or amelioration of skin aging symptoms, such as wrinkling of skin, reduced elasticity, reduced moisturizing function, through one or more selected actions; promotion of wound or burn healing; , Dry skin, etc., prevention, treatment or improvement of dry skin diseases (eg, atopic dermatitis, psoriasis, ichthyosis etc.); prevention, treatment or improvement of skin darkening, pigmentation such as spots, freckles, etc. Prevention, treatment or amelioration of contact dermatitis (rash), psoriasis, pemphigus vulgaris and other various skin inflammatory diseases associated with rough skin; prevention, treatment or amelioration of seborrhea, acne (such as acne) and the like; For preventing, treating or ameliorating obesity and associated lifestyle-related diseases such as arteriosclerosis, diabetes, and metabolic syndrome.

  In addition, the hair cosmetic composition of the present embodiment is characterized in that the hair growth action, testosterone 5α-reductase activity inhibitory action, hair papilla cell growth promotion action, antiandrogenic action, anti-inflammatory action, hyaluronidase activity inhibitory action, and hexosa which dihydroferulic acid has Minidase release inhibitory action, COX-2 activity inhibitory action, cAMP phosphodiesterase activity inhibitory action, anti-aging action, laminin 5 production promotion action, MMP-2 activity inhibition action, hyaluronic acid production promotion action, epidermal keratinocyte proliferation promotion action, Elastin production promoting action, MMP-1 activity inhibiting action, fibroblast proliferation promoting action, AQP3 mRNA expression promoting action, AGEs formation suppressing action, whitening action, melanin production suppressing action, DPPIV activity inhibitory action, anti-obesity action, blood cholesterol reduction Action and blood free fatty acid reducing action Prevention, treatment or amelioration of alopecia such as androgenetic alopecia, alopecia areata, and trichothyromania through one or more actions selected from the group consisting of: rough skin, dry skin, etc., and dry skin diseases ( For example, treatment of atopic dermatitis, psoriasis, ichthyosis, etc.); prevention, treatment or improvement of contact dermatitis (rash), psoriasis, pemphigus vulgaris and other various skin inflammatory diseases associated with rough skin; It can prevent or suppress sticking, restore the elasticity and suppleness of hair, and give the hair firmness and firmness.

(Food and drink)
Dihydroferulic acid has excellent anti-obesity action, cAMP phosphodiesterase activity inhibitory action, DPPIV activity inhibitory action, anti-aging action, hair growth action, anti-androgen action, whitening action, and anti-inflammatory action. It is suitable for blending in. In this case, dihydroferulic acid may be directly added, or an antiobesity agent formulated from dihydroferulic acid, a cAMP phosphodiesterase activity inhibitor, a DPPIV activity inhibitor, an antiaging agent, a hair restorer, an antiandrogen, a whitening agent Agents and anti-inflammatory agents.

  Here, foods and drinks are foods, drugs, and quasi-drugs under the administrative divisions that are not likely to cause harm to human health and are ingested in normal social life by oral or gastrointestinal administration. It is not limited to such categories. Therefore, “foods and drinks” in the present embodiment include general foods, health foods (functional foods and drinks), health foods (foods for specified health use, nutritional foods), quasi-drugs, and pharmaceuticals Etc. are widely included.

  By adding dihydroferulic acid or the above-mentioned anti-obesity agent, cAMP phosphodiesterase activity inhibitor, DPPIV activity inhibitor, anti-aging agent, hair restorer, anti-androgen agent, anti-inflammatory agent, or whitening agent to food and drink, Action, blood cholesterol lowering action, blood free fatty acid lowering action, cAMP phosphodiesterase activity inhibitory action, DPPIV activity inhibitory action, anti-aging action, laminin 5 production promoting action, MMP-2 activity inhibiting action, hyaluronic acid production promoting action, epidermis Keratinocyte proliferation promoting action, elastin production promoting action, MMP-1 activity inhibiting action, fibroblast proliferation promoting action, AQP3 mRNA expression promoting action, AGEs formation inhibitory action, hair growth action, testosterone 5α-reductase activity inhibitory action, hair papilla cells Growth promotion, anti-androgen, anti-inflammatory Hyaluronidase inhibitory effect, hexosaminidase release inhibitory effect, COX-2 inhibitory activity, can impart whitening effect, or the melanin production inhibitory effect on food products. These effects are preferable because the effects are easily exerted by being imparted to foods and drinks.

  Dihydroferulic acid or an anti-obesity agent formulated from dihydroferulic acid, a cAMP phosphodiesterase activity inhibitor, a DPPIV activity inhibitor, an anti-aging agent, a hair restorer, an anti-androgen, a whitening agent, or an anti-inflammatory agent in food and drink When blended, the amount of the active ingredient in them can be appropriately changed in consideration of the purpose of use, symptoms, gender, etc., but in consideration of the general intake of food or drink to be added, Preferably, the daily extract intake is about 1 to 1000 mg. When the food or drink to be added is granular, tablet or capsule food or drink, dihydroferulic acid or an antiobesity agent formulated from dihydroferulic acid, cAMP phosphodiesterase activity inhibitor, DPPIV activity inhibitor, anti-aging The amount of the agent, hair restorer, antiandrogen, whitening agent or anti-inflammatory agent is usually 0.1 to 100% by mass, preferably 5 to 100% by mass, based on the food or drink to be added.

  The food or drink of the present embodiment may be one in which dihydroferulic acid is blended with any food or drink that does not hinder its activity, or may be a dietary supplement containing dihydroferulic acid as a main component. .

  When producing the food or drink of the present embodiment, for example, saccharides such as dextrin and starch; proteins such as gelatin, soy protein, and corn protein; amino acids such as alanine, glutamine, and isoleucine; Sugars: Any auxiliaries such as oils and fats such as soybean oil and medium-chain fatty acid triglyceride can be added to make foods and drinks of any shape.

  Foods and beverages to which dihydroferulic acid can be added are not particularly limited. Specific examples thereof include soft drinks, carbonated drinks, nutritional drinks, fruit drinks, lactic acid drinks and the like (concentrated undiluted solution and powder for adjustment of these drinks). Ice cream, ice sorbet, shaved ice, etc .; noodles such as buckwheat, udon, harusame, gyoza skin, sweet potato skin, Chinese noodles, instant noodles; candy, chewing gum, candy, gum, chocolate, tablet confectionery, Snacks, biscuits, jellies, jams, creams, baked goods and other confectionery products; fish and livestock processed foods such as kamaboko, ham and sausage; dairy products such as processed milk and fermented milk; salad oil, tempura oil, margarine, mayonnaise, shortening Fats and oils and fats and processed foods such as whipped cream and dressing; seasonings such as sauces and sauces; Soups, stews, salads, side dishes, pickles; other various forms of health and nutritional supplements; tablets, capsules, drinks, etc., which are usually used when dihydroferulic acid is added to these foods and drinks. Auxiliary materials and additives can be used in combination.

  The anti-obesity agent, cAMP phosphodiesterase activity inhibitor, DPPIV activity inhibitor, anti-aging agent, hair restorer, anti-androgen, whitening agent, anti-inflammatory agent, skin cosmetic, hair cosmetic, and food and beverage of the present embodiment The product is suitably applied to humans, but as long as the respective effects are exhibited, animals other than humans (eg, mice, rats, hamsters, dogs, cats, cows, pigs, monkeys, etc.) ) Can also be applied.

  Hereinafter, the present invention will be described specifically by showing test examples, but the present invention is not limited to the following examples. In this test example, dihydroferulic acid (Sample 1 manufactured by Tokyo Chemical Industry Co., Ltd.) was used as a test sample.

[Test Example 1] Cyclic AMP phosphodiesterase activity inhibitory test Cyclic AMP phosphodiesterase activity inhibitory activity of dihydroferulic acid (sample 1) was tested as follows.

  0.2 mL of 50 mmol / L Tris-HCl buffer (pH 7.5) containing 5 mmol / L magnesium chloride, 0.1 mL of 2.5 mg / mL bovine serum albumin solution, 0.1 mg / mL cyclic AMP phosphodiesterase solution 0.1 mL and 0.05 mL of a test sample solution (Sample 1, see Table 1 below for sample concentration) were added, and the mixture was allowed to stand at 37 ° C. for 5 minutes. Thereafter, 0.05 mL of a 0.5 mg / mL cyclic AMP solution was added and reacted at 37 ° C. for 60 minutes. After completion of the reaction, the reaction was stopped by boiling on a boiling water bath for 3 minutes, and the mixture was centrifuged (2260 × g, 10 minutes, 4 ° C.). Analysis was performed under high performance liquid chromatography condition 3. Further, as a control, the same operation was performed by adding only the solvent to which no sample was added.

<High performance liquid chromatography condition 3>
Product name: Chromatocoder 12 (manufactured by SYSTEM INSTRUMENTS)
Stationary phase: Wakosil C ₁₈ -ODS 5 μm (manufactured by Wako Pure Chemical Industries, Ltd.)
Column length: 250mm
Mobile phase: 1 mmol / L TBAP in 25 mmol / L KH ₂ PO ₄ : CH ₃ CN = 90: 10
Mobile phase flow rate: 1.0 mL / min
Detection: 260 nm

  Next, the peak area of the cyclic AMP standard (A), the peak area of the supernatant of the reaction solution between the cyclic AMP standard and the cyclic AMP phosphodiesterase when no sample was added (B1), and the peak area when the test sample was added. The peak area (B2) of the supernatant of the reaction solution between the click AMP standard and the cyclic AMP phosphodiesterase was determined. From the obtained results, the decomposition rate (C) of the cyclic AMP standard when no sample was added and the decomposition rate (D) of the cyclic AMP standard when the test sample was added were calculated by the following equations.

Degradation rate of standard without sample (C,%) = (1-B1 / A) × 100
Degradation rate (D,%) of standard product upon addition of test sample = (1-B2 / A) × 100

Then, based on each decomposition rate (C, D) calculated by the above formula, the cyclic AMP phosphodiesterase activity inhibition rate (%) was calculated by the following formula.
cAMP phosphodiesterase activity inhibition rate (%) = (1-D / C) × 100
Table 1 shows the results.

  As shown in Table 1, it was confirmed that dihydroferulic acid (sample 1) has an excellent cyclic AMP phosphodiesterase activity inhibitory action.

[Test Example 2] Blood lipid lowering effect test Dihydroferulic acid (sample 1) was tested for blood lipid lowering effect as follows.

  After 4-week-old TSOD mice (purchased from Shimizu Experimental Materials Co., Ltd.) were fully quarantined and acclimated, their general condition was observed and their weight was measured. Animals with good health were selected and used at 5 weeks of age. Animals are housed in plastic breeding cages under the following conditions: temperature: 22 ± 3 ° C., humidity: 55 ± 15%, ventilation: all-fresh, lighting 12 hours / day (6 am to 6 pm). Bred. The feed was freely inoculated with a laboratory animal solid feed laboratory MR Stock (manufactured by Nihon Nosan Kogyo Co., Ltd.), and drinking water was allowed to freely ingest tap water with an automatic water supply device.

Five-week-old mice were divided into a 50 mg / kg / day dihydroferulic acid administration group and a control group (consisting of 7 mice in each group, 14 mice in total), and intragastric administration once a day using a sonde. And bred for 7 weeks. Fasting was performed for 16 hours from the day before the end of the test, and blood was collected under isoflurane anesthesia at the end of the test. Plasma was prepared from the collected blood by centrifugation (3,000 rpm, 10 minutes), and the amounts of total cholesterol and free fatty acids were measured. In addition, DRI-CHEM3030 (manufactured by FUJIFILM Corporation) was used for measurement of total cholesterol (T-CHO), and NEFA-C Test Wako (manufactured by Wako Pure Chemical Industries) was used for measurement of free fatty acid (NEFA). .
Table 2 shows the results.

  As shown in Table 2, it was confirmed that dihydroferulic acid (sample 1) was excellent in blood cholesterol lowering action and blood free fatty acid lowering action.

[Test Example 3] DPPIV activity inhibitory activity test Dihydroferulic acid (sample 1) was tested for dipeptidyl peptidase IV (DPPIV) inhibitory activity as follows.

  In a 96-well plate, 25 μL of a test sample (sample 1, see Table 3 below for final concentration) prepared with 25 mM Tris-HCl buffer (pH 8.0) and 0.4 μg / Then, 25 μL of a solution of mL DPPIV (rhCD26, manufactured by R & D System) was mixed, and preincubation was performed at 37 ° C. for 5 minutes. Thereafter, 50 μL of 0.5 mM Gly-Pro-p-NA • Tos (manufactured by Peptide Research Laboratories) prepared with the above buffer solution was added, and reacted at 37 ° C. for 90 minutes. After the completion of the reaction, the absorbance at a wavelength of 415 nm was measured. From the obtained results, the DPPIV inhibition rate (%) was calculated by the following equation.

DPPIV inhibition rate (%) = {1- (CD) / (AB)} × 100
Each term in the formula represents the following.
A: Absorbance at a wavelength of 415 nm with no sample added / enzyme added B: Absorbance at a wavelength of 415 nm without sample added / no enzyme added C: Absorbance at a wavelength of 415 nm with test sample added / enzyme added D: Test sample added / enzyme The absorbance at a wavelength of 415 nm without addition is shown in Table 3.

  As shown in Table 3, dihydroferulic acid (Sample 1) exhibited an excellent DPPIV inhibitory effect.

[Test Example 4] Laminin 5 production promoting action test Laminin 5 production promoting action was tested on dihydroferulic acid (sample 1) as follows.

Human normal neonatal epidermal keratinocytes (NHEK) were precultured in an 80 cm ² flask using a human normal epidermal keratinocyte medium (KGM) at 37 ° C. and 5% CO ₂ -95% air. The cells were collected by trypsinization. The collected cells were diluted with KGM-BPE-free medium (KGM-BPE) so as to have a cell density of 1.0 × 10 ⁵ cells / mL, and were seeded at 500 μL / well in a 24-well plate. Cultivation was performed for 2 days under the conditions of ° C and 5% CO ₂ -95% air.

After completion of the culture, the medium was removed, and a test sample dissolved in a KGM-BPE medium (Sample 1, see Table 4 below for sample concentration) was added to each well in an amount of 500 μL, and the mixture was added at 37 ° C. and 5% CO ₂ -95%. The cells were cultured under air conditions for 48 hours. In addition, it culture | cultivated similarly using the KGM-BPE culture medium without a sample as a control. After completion of the culture, 100 μL of the supernatant was transferred to an ELISA plate, and adsorbed on the plate at 37 ° C. for 2 hours. The amount of the adsorbed laminin 5 was measured using a monoclonal anti-human laminin 5 antibody (mouse IgG, manufactured by Chemicon). It was measured by the ELISA method. From the obtained measurement results, the laminin 5 production promotion rate (%) was calculated by the following equation.

Laminin 5 production promotion rate (%) = A / B × 100
Each term in the formula represents the following.
A: Absorbance at a wavelength of 405 nm when a test sample was added B: Absorbance at a wavelength of 405 nm when no sample was added The results are shown in Table 4.

  As shown in Table 4, it was confirmed that dihydroferulic acid (sample 1) has an excellent laminin 5 production promoting action.

[Test Example 5] MMP-2 activity inhibitory activity test MMP-2 used was prepared by the method described in JP-A-2007-217352. That is, the MMP-2 protein was expressed in large amounts as a recombinant proMMP protein having six histidine residues at the C-terminus using an Escherichia coli gene expression system, purified and refolded using Ni-NTA resin, and then activated. Moved to the mold. This was used as an enzyme preparation, which was appropriately diluted to prepare an enzyme solution.
Using the obtained enzyme solution, dihydroferulic acid (sample 1) was tested for MMP-2 activity inhibitory activity as follows.

Using a 96-well plate for measuring fluorescence intensity, 20 μL of a test sample (Sample 1, see Table 5 below for sample concentration), 40 μL of enzyme solution, and buffer solution (0.05 mol / L Tris, 150 mmol / L NaCl, 10 mmol / L CaCl) ₂ , 20 μL of 50 μmol / L ZnSO ₄ , 0.02% NaN ₃ , 0.05% Brij 35 (pH 7.5)) were mixed, and allowed to stand at 37 ° C. for 15 minutes. Thereafter, 120 μL of a fluorescent substrate peptide (MOCAc / DNP peptide) adjusted to 4.16 μmol / L was added, and the fluorescence intensity at an excitation wavelength of 340 nm and a fluorescence wavelength of 420 nm was measured immediately, and this was defined as the fluorescence intensity immediately after the addition of the substrate. Immediately after the measurement, the reaction was carried out at 37 ° C. for 120 minutes. During this time, the fluorescence intensity at an excitation wavelength of 340 nm and a fluorescence wavelength of 420 nm was measured every 15 minutes, and these were defined as the fluorescence intensity after the addition of the substrate. In addition, a blank test was performed in the same manner to make corrections. From the obtained results, the MMP-2 activity inhibition rate (%) was calculated by the following equation.

MMP-2 activity inhibition rate (%) = {1- (CD) / (AB)} × 100
Each term in the formula represents the following.
A: Fluorescence intensity at 420 nm 120 minutes after substrate addition without sample addition B: Fluorescence intensity at 420 nm immediately after substrate addition without sample addition C: Fluorescence intensity at 420 nm 120 minutes after substrate addition with test sample addition D : Fluorescence intensity at 420 nm immediately after addition of the substrate in the test sample addition Table 5 shows the results.

  As shown in Table 5, it was confirmed that dihydroferulic acid (sample 1) had an excellent MMP-2 activity inhibitory action.

[Test Example 6] Test for promoting dermal hyaluronic acid production Human normal skin fibroblasts (NB1RGB) were cultured using an α-MEM medium containing 10% FBS, and then the cells were recovered by trypsin treatment. The collected cells were diluted with an α-MEM medium containing 0.5% FBS to a cell density of 1.6 × 10 ⁵ cells / mL, and then seeded in a 96-well plate at 100 μL / well and cultured overnight. .

  After completion of the culture, the medium was removed, and a test sample (sample 1, see Table 6 below for the sample concentration) dissolved in α-MEM medium containing 0.5% FBS was added to each well, and the cells were cultured for 3 days. In addition, it culture | cultivated similarly using the (alpha) -MEM culture medium containing 0.5% FBS which does not add a sample as a control. After the culture, the amount of hyaluronic acid in the medium in each well was measured by a sandwich method using hyaluronic acid binding protein (HABP). From the obtained results, the hyaluronic acid production promotion rate (%) was calculated by the following equation.

Dermal hyaluronic acid production promotion rate (%) = A / B × 100
Each term in the formula represents the following.
A: Amount of hyaluronic acid when test sample was added B: Amount of hyaluronic acid when test sample was not added The results are shown in Table 6.

  As shown in Table 6, it was confirmed that dihydroferulic acid (sample 1) has an excellent dermal hyaluronic acid production promoting action.

Test Example 7 Test for Promoting Epidermal Keratinocyte Proliferation The dihydroferulic acid (Sample 1) was tested for its promotion of epidermal keratinocyte proliferation as follows.

Normal human neonatal epidermal keratinocytes (NHEK) were cultured using a normal human epidermal keratinocyte growth medium (KGM), and the cells were recovered by trypsinization. The collected cells were diluted with a KGM medium so as to have a cell density of 3.0 × 10 ⁴ cells / mL, and then seeded in a collagen-coated 96-well plate at 100 μL / well and cultured overnight. After completion of the culture, 100 μL of a test sample dissolved in the KGM medium (sample 1, see Table 7 below for sample concentration) was added to each well, and the cells were cultured for 3 days. In addition, it culture | cultivated similarly using the KMG culture medium without a sample as a control.

  The epidermal keratinocyte proliferation promoting effect was measured using the MTT assay. That is, after culturing for 3 days, the medium was removed, and 100 μL of MTT dissolved in PBS (−) buffer at a final concentration of 0.4 mg / mL was added to each well. After culturing for 2 hours, intracellular blue formazan was extracted with 100 μL of 2-propanol. After the 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 taken as the amount of blue formazan produced. From the obtained results, the epidermal keratinocyte proliferation promotion rate (%) was calculated by the following equation.

Epidermal keratinocyte proliferation promotion rate (%) = St / Ct × 100
Each term in the formula represents the following.
St: the amount of blue formazan produced in cells to which the test sample was added Ct: the amount of blue formazan produced in cells to which no sample was added The results are shown in Table 7.

  As shown in Table 7, it was confirmed that dihydroferulic acid (sample 1) has an excellent epidermal keratinocyte proliferation promoting effect.

[Test Example 8] Elastin production promoting action test Elastin production promoting action was tested on dihydroferulic acid (sample 1) as follows.

After normal human fibroblasts (NB1RGB) were cultured using Dulbecco's MEM medium containing 10% FBS, the cells were recovered by trypsinization. The collected cells were diluted with the above medium to a cell density of 2.2 × 10 ⁵ cells / mL, and then seeded in a 96-well microplate at 100 μL / well and cultured overnight.

  After completion of the culture, the medium was removed, 150 μL of a test sample (sample 1, see Table 8 below for sample concentration) dissolved in Dulbecco's MEM medium containing 0.25% FBS was added to each well, and the cells were cultured for 2 days. As a control, a Dulbecco MEM medium containing 0.25% FBS containing no sample was similarly cultured. After completion of the culture, the supernatant was recovered, and the amount of elastin released into the culture supernatant was measured by ELISA. From the measurement results, the elastin production promotion rate (%) was calculated by the following equation.

Elastin production promotion rate (%) = A / B × 100
Each term in the formula represents the following.
A: The amount of elastin when the test sample was added B: The amount of elastin when the sample was not added The results are shown in Table 8.

  As shown in Table 8, dihydroferulic acid (sample 1) exhibited an excellent elastin production promoting action.

[Test Example 9] MMP-1 activity inhibitory activity test Dihydroferulic acid (sample 1) was tested for MMP-1 activity inhibitory activity as follows.

  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 / L calcium chloride (see Table 9 below for sample 1 and sample concentration) , 50 μL of MMP-1 solution (manufactured by Sigma, COLLAGENASE Type IV from Clostridium histolyticum) and 400 μL of Pz peptide solution (manufactured by BACHEM Feinchemikalien AG, Pz-Pro-Leu-Gly-Pro-D-Arg-OH). After reaction at 37 ° C. for 30 minutes, 1 mL of a 25 mmol / L citric acid solution was added to stop the reaction.

  Thereafter, 5 mL of ethyl acetate was added, and the mixture was vigorously shaken. This was centrifuged (1600 × g, 10 minutes), and the absorbance of the ethyl acetate layer at a wavelength of 320 nm was measured. In addition, a blank test was performed in the same manner to make correction. From the obtained results, the MMP-1 activity inhibition rate (%) was calculated by the following equation.

MMP-1 activity inhibition rate (%) = {1- (CD) / (AB)} × 100
Each term in the formula represents the following.
A: Absorbance at a wavelength of 320 nm with no sample added / enzyme added B: Absorbance at a wavelength of 320 nm without sample added / no enzyme added C: Absorbance at a wavelength of 320 nm with sample added / enzyme added D: Sample added / no enzyme added Table 9 shows the results of the absorbance at a wavelength of 320 nm.

  As shown in Table 9, dihydroferulic acid (Sample 1) was found to have an excellent MMP-1 activity inhibitory action.

[Test Example 10] Test for promoting skin fibroblast proliferation The dihydroferulic acid (Sample 1) was tested for its promotion of skin fibroblast proliferation as follows.

Normal human skin fibroblasts (NB1RGB) were cultured using an α-MEM medium containing 10% FBS, and then the cells were recovered by trypsinization. The collected cells were diluted with an α-MEM medium containing 5% FBS to a cell density of 7.0 × 10 ⁴ cells / mL, and then seeded in a 96-well plate at 100 μL / well and cultured overnight. After completion of the culture, the medium was removed, and a test sample (sample 1, see Table 10 below for the sample concentration) dissolved in an α-MEM medium containing 5% FBS was added to each well in an amount of 100 μL, followed by culturing for 3 days. In addition, it culture | cultivated similarly using the (alpha) -MEM culture medium containing 5% FBS without a sample as a control.

  The fibroblast proliferation promoting effect was measured using the MTT assay. That is, after culturing for 3 days, the medium was removed, and 100 μL of MTT dissolved in PBS (−) buffer at a final concentration of 5 mg / mL was added to each well. After culturing for 2 hours, intracellular blue formazan was extracted with 100 μL of 2-propanol. After the 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 taken as the amount of blue formazan produced. From the obtained results, the fibroblast proliferation promotion rate (%) was calculated by the following equation.

Fibroblast growth promotion rate (%) = St / Ct × 100
Each term in the formula represents the following.
St: the amount of blue formazan produced when the test sample was added Ct: the amount of blue formazan produced without the addition of the sample The results are shown in Table 10.

  As shown in Table 10, it was confirmed that dihydroferulic acid (sample 1) had an excellent fibroblast proliferation promoting action.

[Test Example 11] Aquaporin 3 (AQP3) mRNA expression promoting action test AQP3 mRNA expression promoting action was tested on dihydroferulic acid (Sample 1) as follows.

Human normal neonatal epidermal keratinocytes (NHEK) were pre-cultured in an 80 cm ² flask using a human normal epidermal keratinocyte medium (KGM) at 37 ° C. and 5% CO ₂ -95% air. The cells were collected by trypsinization. After diluting the collected cells with a KGM medium to a cell density of 20 × 10 ⁴ cells / mL, the cells were seeded at a rate of 2 mL each on a 35 mm Petri dish (manufactured by FALCON) (40 × 10 ⁴ cells / Petri dish) at 37 ° C. The cells were cultured for 24 hours under the conditions of 5% CO _{2 and} 95% air.

After the culture, the medium was removed, and 2 mL of a KGM medium of a test sample dissolved in the KGM medium (Sample 1, see Table 11 below for sample concentration) was added to each dish, and the mixture was heated at 37 ° C. and 5% CO ₂ -95% air. For 24 hours. In addition, it culture | cultivated similarly using the KGM culture medium without a sample as a control. After the culture, the medium was removed, total RNA was extracted with ISOGEN (Nippon Gene, Cat. No. 311-02501), the amount of each RNA was measured with a spectrophotometer, and the amount was adjusted to 200 ng / μL. Total RNA was prepared.

  Using this total RNA as a template, the expression level of mRNA was measured for AQP3 and GAPDH as an internal standard. Detection was performed by a real-time 2 Step RT-PCR reaction using a TaKaRa SYBR Prime Script RT-PCR kit (Perfect Real Time) (manufactured by Takara Bio Inc., code No. RR063A) using a real-time PCR device Smart Cycler (manufactured by Cepheid). . The correction value of the expression level of AQP3 mRNA was determined by the value of GAPDH based on the total RNA preparation prepared from the cells cultured in “addition of test sample” and “without addition of sample”. From the obtained values, the AQP3 mRNA expression promotion rate (%) was calculated by the following equation.

AQP3 mRNA expression promotion rate (%) = A / B × 100
Each term in the formula represents the following.
A: Correction value when test sample was added B: Correction value when no sample was added The results are shown in Table 11.

  As shown in Table 11, it was confirmed that dihydroferulic acid (sample 1) has an excellent AQP3 mRNA expression promoting action.

[Test Example 12] AGEs formation inhibitory effect test The inhibitory effect of dihydroferulic acid (Sample 1) on the formation of the final saccharified product (AGEs) was tested as follows.

  In a 96-well type I collagen-coated plate (manufactured by Asahi Glass Co., Ltd.), 0.2M D (-)-ribose prepared in PBS (-) buffer and a test sample (see Table 14 for sample 1 and sample concentration). After the mixture was added to 100 μL, the mixture was allowed to stand at 37 ° C. for 2 weeks to form AGEs. The PBS (-) buffer alone as a negative control and the 0.2M D (-)-ribose solution prepared with the PBS (-) buffer as a positive control were left to stand, respectively. Two weeks later, the amount of AGEs was measured by an ELISA method using an anti-AGEs antibody (manufactured by Transgenic) to evaluate the AGEs formation inhibitory effect. From the obtained results, the AGEs formation inhibition rate (%) was calculated by the following equation.

AGEs formation suppression rate (%) = {(BC) / (BA)} × 100
Each term in the formula represents the following.
A: Absorbance at a wavelength of 405 nm with a negative control B: Absorbance at a wavelength of 405 nm with a positive control C: Absorbance at a wavelength of 405 nm with addition of a test sample The results are shown in Table 12.

  As shown in Table 12, dihydroferulic acid (Sample 1) exhibited an excellent AGEs formation inhibitory action.

[Test Example 13] Testosterone 5α-reductase inhibitory activity test Dihydroferulic acid (Sample 1) was tested for testosterone 5α-reductase inhibitory activity as follows.

  In a V-bottom test tube with a lid, 20 μL of a 4.2 mg / mL testosterone (manufactured by Wako Pure Chemical Industries, Ltd.) solution prepared with propylene glycol and 5 mmol / L Tris-HCl (pH 7.13) containing 1 mg / mL NADPH were used. 825 μL of the buffer solution was mixed.

  Further, 80 μL of a test sample (sample 1, see Table 12 below for sample concentration) solution prepared in 50% ethanol and 75 μL of S-9 (rat liver homogenate, manufactured by Oriental Yeast Co., Ltd.) were added and mixed. Incubated at 37 ° C. for 30 minutes. Thereafter, 1 mL of methylene chloride was added to stop the reaction. This was centrifuged (1600 × g, 10 minutes), the methylene chloride layer was separated, and the separated methylene chloride layer was subjected to gas chromatography analysis under the following conditions to obtain 3α-androstanediol, 5α. -The concentrations of dihydrotestosterone (5α-DHT) and testosterone were quantified. As a control, the same treatment was carried out using the same amount (80 μL) of the sample solvent instead of the test sample solution, and subjected to gas chromatography analysis.

<Gas chromatography conditions>
Equipment used: Shimadzu GC-2010 (manufactured by Shimadzu Corporation)
Column: DB-1701 (inner diameter: 0.53 mm, length: 30 m, film thickness: 1.0 μm, manufactured by J & W Scientific)
Column temperature: 240 ° C
Inlet temperature: 300 ° C
Detector: FID
Sample injection volume: 1 μL
Split ratio: 1: 2
Carrier gas: nitrogen gas Carrier gas flow rate: 3 mL / min

The quantification of the concentrations of 3α-androstanediol, 5α-DHT and testosterone was performed by the following method.
Standards of 3α-androstanediol, 5α-DHT and testosterone were dissolved in methylene chloride, and the solution was subjected to gas chromatography analysis. From the concentration (μg / mL) and peak area of these compounds, the peak area and compound The corresponding relationship with the concentration was determined in advance. Then, the concentrations per peak area of 3α-androstanediol, 5α-DHT, and testosterone after the reaction of testosterone with S-9 were determined using the following relationship (1) using the correspondence relationship determined in advance. Was determined based on

A = B × C / D (1)
Each term in the formula represents the following.
A: Concentration of 3α-androstanediol, 5α-DHT or testosterone B: Peak area of 3α-androstanediol, 5α-DHT or testosterone C: Concentration of standard D: Peak area of standard

  Using the compound concentration calculated based on the formula (1), based on the following formula (2), the conversion rate (the concentration of 3α-androstanediol and 5α-DHT formed by the reduction of testosterone by testosterone 5α-reductase) And the initial concentration of testosterone).

Conversion rate = (E + F) / (E + F + G) (2)
Each term in the formula represents the following.
E: Concentration of 3α-androstanediol (μg / mL)
F: Concentration of 5α-DHT (μg / mL)
G: Concentration of testosterone (μg / mL)

The testosterone 5α-reductase inhibition rate (%) was calculated based on the following equation (3) using the conversion rate calculated based on the equation (2).
Testosterone 5α-reductase inhibition rate (%) = (1-H / I) × 100 (3)
Each term in the formula represents the following.
H: Conversion rate with addition of test sample I: Conversion rate without addition of sample Table 13 shows the results.

  As shown in Table 13, it was confirmed that dihydroferulic acid (sample 1) has an excellent testosterone 5α-reductase inhibitory action.

[Test Example 14] Test for promoting hair papilla cell proliferation The effect of promoting hair papilla cell proliferation was tested for dihydroferulic acid (Sample 1) as follows.

Normal human hair papilla cells (HFDPC, derived from male parietal region) were cultured using a hair papilla cell growth medium (PCGM, manufactured by Toyobo Co., Ltd.) containing 1% FCS and a growth additive, and then treated with trypsin. Was recovered. The collected cells were diluted to a cell density of 1.0 × 10 ⁴ cells / mL using 10% FBS-containing DMEM medium, and then seeded at 200 μL / well in a collagen-coated 96-well plate. Cultured for days. In addition, it culture | cultivated similarly using the serum-free DMEM medium without a sample as a control.

  Thereafter, the medium was removed, and 200 μL of a test sample dissolved in serum-free DMEM medium (Sample 1, see Table 16 for sample concentration) was added to each well, and the cells were further cultured for 4 days. After the completion of the culture, the effect of promoting hair papilla cell proliferation was measured by MTT assay. That is, the medium was removed, 200 μL of 0.4 mg / mL MTT prepared in a serum-free DMEM medium was added, and the cells were further cultured for 2 hours. Then, blue formazan generated in the cells was extracted with 100 μL of 2-propanol. With respect to this extract, the absorbance at 570 nm where the absorption maximum point of blue formazan 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 taken as the amount of blue formazan produced. From the measurement results, the hair papilla cell growth promotion rate (%) was calculated based on the following equation.

Hair papilla cell growth promotion rate (%) = A / B × 100
Each term in the formula represents the following.
A: The amount of blue formazan produced when the test sample was added. B: The amount of blue formazan produced without the sample added. The results are shown in Table 14.

  As shown in Table 14, dihydroferulic acid (Sample 1) was found to have an excellent hair papilla cell growth promoting action.

[Test Example 15] Test of melanin production inhibitory action on B16 melanoma cells Melanin production inhibitory action on B16 melanoma cells was tested for dihydroferulic acid (Sample 1) as follows.

After culturing B16 melanoma cells using Dulbecco's MEM medium containing 10% FBS, the cells were recovered by trypsinization. The collected cells were diluted with Dulbecco's MEM medium containing 10% FBS and 1 mmol / L theophylline to a cell density of 24.0 × 10 ⁴ cells / mL, and then seeded in a 48-well plate at 300 μL / well. Cultured for hours.

  After completion of the culture, 300 μL of a test sample (Sample 1, see Table 13 below for the sample concentration) dissolved in Dulbecco's MEM medium containing 10% FBS and 1 mmol / L theophylline was added to each well and cultured for 4 days. As a control, Dulbecco's MEM medium containing 10% FBS and 1 mmol / L theophylline containing no sample was similarly cultured. After completion of the culture, the medium was removed, 200 μL of a 2 mol / L NaOH solution was added, the cells were disrupted by an ultrasonic crusher, and the absorbance at a wavelength of 475 nm was measured. From the measured absorbance value, the amount of melanin was calculated based on a calibration curve created using synthetic melanin (manufactured by SIGMA).

  In order to measure the cell viability, the cells were cultured in the same manner as above, the medium was removed, washed with 400 μL of PBS buffer, and dissolved in Dulbecco MEM containing 10% FBS at a final concentration of 0.05 mg / mL. 200 μL of the neutral red thus added was added to each well, and the cells were cultured for 2.5 hours. After the culture, the neutral red solution was removed, and 200 μL of an ethanol / acetic acid solution (ethanol: acetic acid: water = 50: 1: 49) was added to each well to extract a dye. After the extraction, the absorbance at a wavelength of 540 nm was measured. From the obtained results, the melanin production inhibition rate (%) corrected by the cell viability according to the following formula was calculated.

Melanin production inhibition rate (%) = {1- (B / D) / (A / C)} × 100
Each term in the formula represents the following.
A: Melanin content without sample addition B: Melanin content with test sample addition C: Absorbance at 540 nm without sample addition D: Absorbance at 540 nm with test sample addition The results are shown in Table 15.

  As shown in Table 15, dihydroferulic acid (sample 1) was found to have an excellent melanin production inhibitory action.

[Test Example 16] Hyaluronidase activity inhibitory test A test sample (Sample 1, see Table 14 below for the sample concentration) dissolved in 0.1 mol / L acetate buffer (pH 3.5) was added to a hyaluronidase solution (Type 0.1 mL of IV-S (from bovine testes), 400 NF units / mL, manufactured by SIGMA was added, and the mixture was allowed to stand at 37 ° C. for 20 minutes. Further, 0.2 mL of 2.5 mmol / L calcium chloride was added as an activator, and the mixture was allowed to stand at 37 ° C. for 20 minutes. To this, 0.5 mL of a 0.8 mg / mL sodium hyaluronate solution (from rooster comb) was added and reacted at 37 ° C. for 40 minutes. Thereafter, 0.2 mL of 0.4 mol / L sodium hydroxide was added to stop the reaction, and after cooling, 0.2 mL of a boric acid solution was added to each reaction solution and the mixture was boiled for 3 minutes. After cooling on ice, 6 mL of a p-DABA reagent was added and reacted at 37 ° C. for 20 minutes. Thereafter, the absorbance at a wavelength of 585 nm was measured.

The same operation and measurement of the absorbance were performed for a blank where no enzyme solution was added. Further, as a control, the same measurement was performed when distilled water was added without adding the sample solution. From the obtained results, the hyaluronidase activity inhibition rate (%) was calculated by the following formula.
Hyaluronidase activity inhibition rate (%) = {1- (St-Sb) / (Ct-Cb)} × 100
Each term in the formula represents the following.
St: Absorbance of test sample solution at wavelength 585 nm Sb: Absorbance of test sample solution blank at wavelength 585 nm Ct: Absorbance of control solution at wavelength 585 nm Cb: Absorbance of control solution blank at wavelength 585 nm The results are shown in Table 16.

  As shown in Table 16, it was confirmed that dihydroferulic acid (sample 1) had an excellent hyaluronidase activity inhibitory action.

[Test Example 17] Test for inhibition of hexosaminidase release The inhibitory effect of hexosaminidase release on dihydroferulic acid (Sample 1) was tested as follows.

Rat basophil leukemia cells (RBL-2H3) were cultured using S-MEM medium containing 15% FBS, and then recovered by trypsinization. The collected cells were diluted with S-MEM medium containing 15% FBS to a cell density of 4.0 × 10 ⁵ cells / mL, and DNP-specific IgE was added to a final concentration of 0.5 μL / mL. Thereafter, the cells were seeded in a 96-well plate at 100 μL per well and cultured overnight.

  After the culture, the medium was removed, and washing was performed twice with 100 μL of a Shiraganian buffer. Next, 10 μL of a test sample dissolved in the same buffer (Sample 1, see Table 15 below for sample concentration) and 30 μL of the same buffer were added to each well, and the mixture was allowed to stand at 37 ° C. for 10 minutes. As a control, the same operation was performed using 40 μL of a sample-free Shiraganian buffer solution. Subsequently, 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, release was stopped by placing the 96-well plate on ice. 10 μL of the cell supernatant in each well was collected in a new 96-well plate, and 10 μL of a 1 mmol / L p-NAG (p-nitrophenyl-N-acetyl-β-D-glucosaminide) solution was added to each well, and the mixture was added 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, the absorbance at wavelengths of 415 nm and 650 nm was measured, and the value obtained by subtracting the absorbance at 650 nm from the absorbance at 415 nm was used as a correction value. Further, as a blank, the absorbance at a wavelength of 415 nm and a wavelength of 650 nm of a mixed solution of 10 μL of the cell supernatant and 250 μL of 0.1 mol / L Na ₂ CO ₃ / NaHCO ₃ was measured, and a correction value was calculated. From the obtained measurement results, the hexosaminidase release inhibition rate (%) was calculated by the following equation.

Hexosaminidase release inhibition rate (%) = {1- (BC) / A} × 100
Each term in the formula represents the following.
A: Correction value without addition of sample B: Correction value with addition of test sample C: Correction value without addition of test sample / no addition of p-NAG Table 17 shows the results.

  As shown in Table 17, it was confirmed that dihydroferulic acid (sample 1) had an excellent hexosaminidase release inhibitory action.

[Test Example 18] Test for inhibiting COX-2 activity in mouse macrophages (Test for inhibiting PGE ₂ production)
Mouse macrophage cells (RAW 264.7) were cultured using Dulbecco's MEM medium containing 10% FBS, and then the cells were collected using a cell scraper. The collected cells were diluted with Dulbecco's MEM medium containing 10% FBS to a concentration of 2.0 × 10 ⁵ cells / mL, and then seeded on a 96-well plate at 100 μL / well and cultured for 18 hours.

After completion of the culture, the medium was replaced with a 500 μmol / L aspirin-containing medium and cultivated for 4 hours in order to acetylate and inactivate the existing COX-1 and a small amount of expressed COX-2. Thereafter, the cells were washed three times with a PBS (-) buffer solution and dissolved in Dulbecco's MEM medium containing 10% FBS containing 0.5% DMSO (final concentration, see Table 16 for sample concentrations). Was added to each well, and 100 μL of lipopolysaccharide (LPS, E. coli 0111; B4, manufactured by DIFCO) dissolved in Dulbecco MEM containing 10% FBS at a final concentration of 1 μg / mL was added, followed by culturing for 16 hours. . As a control, Dulbecco's MEM medium containing 10% FBS and containing 0.5% DMSO without addition of a sample was cultured in the same manner. After completion of the culture, the amount of prostaglandin E _{2 in} the culture supernatant of each well was quantified using a PGE ₂ EIA Kit (Cayman Chemical). From the obtained results, the COX-2 activity inhibition rate (%, PGE ₂ production inhibition rate) was calculated by the following equation.

Macrophage COX-2 activity inhibition rate (%) = {1- (AC) / (BC)} × 100
Each term in the formula represents the following.
A: Prostaglandin E ₂ amount when test sample was added and LPS was stimulated B: Prostaglandin E ₂ amount when no sample was added and LPS was stimulated C: Prostaglandin E ₂ amount when no sample was added and LPS was not stimulated Result Is shown in Table 18.

  As shown in Table 18, it was confirmed that dihydroferulic acid (sample 1) had an excellent COX-2 activity inhibitory effect on macrophages.

[Formulation Example 1]
An emulsion having the following composition was produced by a conventional method.
0.01 g of dihydroferulic acid
Jojoba oil 4.00g
1,3-butylene glycol 3.00 g
Arbutin 3.00 g
2.50 g of polyoxyethylene cetyl ether (20E.O.)
Olive oil 2.00g
Squalane 2.00g
Cetanol 2.00g
Glyceryl monostearate 2.00 g
2.00 g of polyoxyethylene sorbitan oleate (20E.O.)
Methyl paraoxybenzoate 0.15g
Stearyl glycyrrhizinate 0.10 g
Huang Gui Extract 0.10g
Dipotassium glycyrrhizinate 0.10 g
Ginkgo biloba leaf extract 0.10g
Conchiolin 0.10g
Oubaku extract 0.10g
Chamomile extract 0.10g
Spice 0.05g
Remaining purified water (total amount is 100 g)

[Formulation Example 2]
A cream having the following composition was produced by a conventional method.
Dihydroferulic acid 0.05g
Kujin extract 0.1g
Ougon extract 0.1g
Liquid paraffin 5.0 g
4.0 g of beeswax
Squalane 10.0g
3.0 g of cetanol
Lanolin 2.0g
1.0 g of stearic acid
1.5 g of polyoxyethylene sorbitan oleate (20E.O.)
Glyceryl monostearate 3.0 g
Oil-soluble licorice extract 0.1 g
1,3-butylene glycol 6.0 g
1.5 g of methyl paraoxybenzoate
Spice 0.1g
Remaining purified water (total amount is 100 g)

[Formulation Example 3]
A serum having the following composition was produced by a conventional method.
0.01 g of dihydroferulic acid
Chamomile extract 0.1g
Carrot extract 0.1g
Xanthan gum 0.3g
0.1 g of hydroxyethyl cellulose
Carboxyvinyl polymer 0.1 g
1,3-butylene glycol 4.0 g
Dipotassium glycyrrhizinate 0.1 g
Glycerin 2.0g
Potassium hydroxide 0.25g
Spice 0.01g
Preservative (methyl paraoxybenzoate) 0.15g
2.0 g of ethanol
Remaining purified water (total amount is 100 g)

[Formulation Example 4]
A hair tonic having the following composition was produced by a conventional method.
0.4 g of dihydroferulic acid
Tocopherol acetate suitable amount Cephalatin 0.002g
0.1 g of isopropylmethylphenol
Sodium hyaluronate 0.15g
Glycerin 15.0g
15.0 g of ethanol
Perfume Appropriate amount Chelating agent (sodium edetate) Appropriate amount Preservative (Hinokitiol) Appropriate amount Solubilizing agent (Polyoxyethylene cetyl ether) Appropriate amount

[Formulation Example 5]
A shampoo having the following composition was produced by a conventional method.
0.5 g of dihydroferulic acid
1.0 g of marjoram extract
Plum fruit extract 0.2g
Coconut oil fatty acid sodium methyltaurine 10.0g
Palm oil fatty acid amidopropyl betaine 10.0 g
Sodium polyoxyethylene alkyl ether sulfate 20.0 g
Palm oil fatty acid diethanolamide 4.0g
Propylene glycol 2.0g
Appropriate amount of fragrance Purified water Remainder (total amount is 100g)

[Formulation Example 6]
A tablet having the following composition was produced by a conventional method.
Dihydroferulic acid 5.0mg
Dolomite (containing 20% calcium and 10% magnesium) 83.4mg
16.7mg casein phosphopeptide
Vitamin C 33.4mg
Maltitol 136.8mg
Collagen 12.7mg
12.0mg sucrose fatty acid ester

[Formulation Example 7]
An oral liquid preparation having the following composition was produced by a conventional method.
<Composition in one ampule (100 mL per bottle)>
0.3% by mass of dihydroferulic acid
Sorbit 12.0% by mass
Sodium benzoate 0.1% by mass
Spice 1.0% by mass
0.5% by mass of calcium sulfate
Purified water balance (100% by mass)

  The anti-obesity agent, cAMP phosphodiesterase activity inhibitor, DPPIV activity inhibitor, anti-aging agent, hair restorer, anti-androgen, whitening agent, and anti-inflammatory agent of the present invention can prevent, treat or ameliorate aging symptoms of the skin; Promotion of wound or burn healing; prevention, treatment or amelioration of dry skin diseases; prevention, treatment or amelioration of alopecia, especially alopecia of androgenetic alopecia; pigmentation of skin darkening, spots, freckles, etc. Prevention or improvement; prevention, treatment or improvement of type 2 diabetes, obesity, hypertension, insulin resistance, etc .; prevention or improvement of various skin inflammatory diseases; prevention, treatment or improvement of obesity;

Claims (3)

A melanin production inhibitor comprising dihydroferulic acid as an active ingredient. Melanin production suppressing skin cosmetic which is characterized in that blended dihydro ferulic acid. Melanin production inhibition food or drink, characterized in that blended with dihydro ferulic acid.