TWI727206B - Uses of pyrenaria buisanensis extract - Google Patents

Abstract

Uses ofPyrenaria buisanensis extract are provided, comprising the use ofPyrenaria buisanensis extract in moisturizing skin, whitening skin, tightening skin, reducing skin fine lines, anti-skin aging, alleviating dry skin, promoting formation of hyaluronic acid in the skin, assisting in maintaining health of skin, or manufacturing a pharmaceutical composition. The pharmaceutical composition is useful in anti-photodamage, repairing skin tissues, preventing skin diseases, and/or treating skin diseases, or in at least one of preventing cardiovascular diseases, treating cardiovascular diseases, preventing diabetes, treating diabetes, preventing neurodegenerative diseases, and treating neurodegenerative diseases.

Description

Application of Wuwei Mountain Wupi Tea Extract

The present invention relates to the application of Wuwei Mountain Wupi tea ( Pyrenaria buisanensis ) extract, including the use of Wuwei Mountain Wupi tea extract for moisturizing, whitening, firming skin, reducing skin fine lines, anti-aging skin, improving dry skin, and promoting The formation of hyaluronic acid in the skin and/or help maintain skin health; and the use of Wuwei Mountain Wupi tea extract to prevent light damage, repair skin tissues, prevent skin diseases, treat skin diseases, prevent cardiovascular diseases, and treat cardiovascular diseases , Prevention of diabetes, treatment of diabetes, prevention of neurodegenerative diseases, and/or treatment of neurodegenerative diseases.

The skin is the body's first line of defense against external harmful factors, and has important functions such as water retention, warmth and feeling. Ageing, as well as ultraviolet light, radiation, environmental pollution and other factors, may reduce the water content of the skin, cause the loss of collagen and elastin, and cause wrinkles, skin sagging, dullness and roughness, and even cause aging. Damage to DNA in skin cells, accelerate skin cell death, or cause skin lesions and induce skin diseases.

As modern people pay more and more attention to skin whitening, moisturizing and anti-aging skin, the beauty products on the market are becoming more and more diversified, such as oral collagen, applying hyaluronic acid to the skin, and botulinum toxin. In the aforementioned methods, the molecular structure of components such as collagen and hyaluronic acid is too large to be effectively absorbed by the body when administered orally or smeared, so the actual supplementary effect is limited; although the application of Botox can reduce skin fineness Wrinkles, but the effect is short-lived, must be administered regularly, and the cost is high. It may also cause side effects such as swelling or blood stasis at the injection site, drooping eyelids or eyebrows, headache, allergies, asymmetric left and right faces, or unnatural expressions.

Studies have known that the expression enhancement system of genes such as SOD2 , CAT , MSH2 , Tgm1 , Keratin14 , FLG , GBA and HAS3 can help improve cell antioxidant capacity, repair damaged DNA and protein in cells, maintain cell structure, and improve transparency The amount of acid synthesis and/or increase the water content of cells. The inventor of this case conducted research using natural materials in nature and found that Wuwei Mountain Wupi tea extract can improve the expression of SOD2 gene, CAT gene, MSH2 gene, Tgm1 gene, Keratin14 gene, FLG gene, GBA gene, and HAS3 gene in cells. It can also increase skin's antioxidant capacity, reduce skin melanin content, reduce skin sagging, and increase skin moisture content. Therefore, it can be used for moisturizing, whitening, firming skin, reducing skin fine lines, anti-aging skin, improving dry skin, and promoting skin Hyaluronic acid forms, helps maintain skin health, resists light damage, and/or repairs skin tissue, and can be used to prevent, treat or regulate diseases or physiological functions related to the aforementioned genes.

One purpose of the present invention is to provide a use of Wuwei Mountain Wupi tea extract for at least one of the following: moisturizing, whitening, skin tightening, reducing skin fine lines, anti-aging skin, improving dry skin, promoting skin transparency The formation of acid and helps maintain healthy skin. Preferably, the extract is provided by extracting Wuweishan Wupi tea with a polar solvent, and the polar solvent is water, C1-C4 alcohols, or a combination of the foregoing. Preferably, the extract is the leaf extract of Wuwei Mountain Wupi Tea. Preferably, the extract is used by smearing or oral administration.

Another object of the present invention is to provide a use of the Wuwei Mountain Wupi tea extract in the preparation of a pharmaceutical composition for at least one of the following: anti-photodamage, repairing skin tissue, and preventing Skin diseases, and treatment of skin diseases. Preferably, the skin disease is a disease related to dry skin. Preferably, the pharmaceutical composition is in a dosage form selected from the following group: percutaneous administration, oral administration, and subcutaneous injection.

Another object of the present invention is to provide a use of the Wuwei Mountain Wupi tea extract in the preparation of a pharmaceutical composition for at least one of the following: prevention of cardiovascular disease, treatment of cardiovascular disease , Prevention of diabetes, treatment of diabetes, prevention of neurodegenerative diseases, and treatment of neurodegenerative diseases. Preferably, the cardiovascular disease is ischemic stroke, and the neurodegenerative disease is Alzheimer's disease. Preferably, the pharmaceutical composition is in a dosage form selected from the following group: percutaneous administration, oral administration, and subcutaneous injection.

Another object of the present invention is to provide a use of the above Wuwei Mountain Wupi tea extract in the preparation of a pharmaceutical composition for improving the performance of at least one of the following genes: SOD2 gene, CAT gene , MSH2 gene, Tgm1 gene, Keratin14 gene, FLG gene, GBA gene, and HAS3 gene. Preferably, the pharmaceutical composition is in a dosage form selected from the following group: percutaneous administration, oral administration, and subcutaneous injection.

Another object of the present invention is to provide a combination for moisturizing, whitening, firming skin, reducing skin fine lines, anti-aging skin, improving dry skin, promoting the formation of skin hyaluronic acid, and/or helping to maintain skin health It contains an effective amount of the above Wuwei Mountain Wupi tea extract. Preferably, the composition is a skin care product composition or a food composition, and is used in the form of smearing or oral administration.

Another object of the present invention is to provide a pharmaceutical composition for resisting light damage, repairing skin tissue, preventing skin diseases, and/or treating skin diseases, which contains an effective amount of the above Wuwei mountain ebony tea extract . Preferably, the skin disease is a disease related to dry skin. Preferably, the pharmaceutical composition is in a dosage form selected from the following group: percutaneous administration, oral administration, and subcutaneous injection.

Another object of the present invention is to provide a pharmaceutical composition for preventing cardiovascular diseases, treating cardiovascular diseases, preventing diabetes, treating diabetes, preventing neurodegenerative diseases, and/or treating neurodegenerative diseases, which contains An effective amount of the above Wuwei Mountain Wupi tea extract. Preferably, the cardiovascular disease is ischemic stroke, and the neurodegenerative disease is Alzheimer's disease. Preferably, the pharmaceutical composition is in a dosage form selected from the following group: percutaneous administration, oral administration, and subcutaneous injection.

Another object of the present invention is to provide a pharmaceutical composition for enhancing the expression of SOD2 gene, CAT gene, MSH2 gene, Tgm1 gene, Keratin14 gene, FLG gene, GBA gene, and/or HAS3 gene, which contains an effective The amount of the above Wuwei Mountain Wupi Tea Extract. Preferably, the pharmaceutical composition is in a dosage form selected from the following group: percutaneous administration, oral administration, and subcutaneous injection.

Another object of the present invention is to provide a method for moisturizing, whitening, firming the skin, reducing fine lines, anti-aging skin, improving dry skin, promoting the formation of skin hyaluronic acid, and/or helping to maintain skin health, which It comprises administering an effective amount of the above-mentioned Wuweishan ebony tea extract to an individual in need, wherein the Wuweishan ebony tea extract can be administered to the above-mentioned skin care product composition or food composition in the form of Individuals in need.

Another object of the present invention is to provide a method for resisting light damage, repairing skin tissues, preventing skin diseases, treating skin diseases, preventing cardiovascular diseases, treating cardiovascular diseases, preventing diabetes, treating diabetes, preventing neurodegenerative diseases, and/ Or a method for treating neurodegenerative diseases, which comprises administering to an individual in need an effective amount of the above Wuwei Mountain ebony tea extract, wherein the Wuwei Mountain ebony tea extract can be in the form of the above-mentioned pharmaceutical composition Give to the needy individual. For example, the method can be used to prevent dry skin related diseases, treat dry skin related diseases, prevent ischemic stroke, treat ischemic stroke, prevent Alzheimer's disease, and/or treat Alzheimer's disease.

Another object of the present invention is to provide a method for enhancing the performance of SOD2 gene, CAT gene, MSH2 gene, Tgm1 gene, Keratin14 gene, FLG gene, GBA gene, and/or HAS3 gene. The individual in need is administered an effective amount of the above Wuwei mountain ebony tea extract, wherein the Wuwei mountain ebony tea extract can be administered to the individual in need in the form of the above-mentioned pharmaceutical composition.

The detailed technical content and some specific implementation aspects of the present invention will be described in the following content, so that those skilled in the art to which the present invention belongs can understand the features of the present invention.

The following will describe some specific implementation aspects of the present invention; however, without departing from the spirit of the present invention, the present invention can still be practiced in many different forms, and the protection scope of the present invention should not be construed as limited to those stated in the specification. . In addition, unless otherwise specified in the text, the "a", "the" and similar terms used in this specification (especially in the scope of the patent application described later) should be understood to include the singular and plural forms; the so-called "treatment" does not Should be interpreted as treating an individual until complete recovery, but should include maintaining an individual’s disease progression or symptoms to a substantially static level, increasing the recovery rate of an individual, improving the severity of a specific condition, or improving a patient The so-called "prevention" refers to inhibiting or preventing the onset of a specific disease, or maintaining the good health of a sensitive individual or establishing the individual's tolerance to the disease; the so-called "regulation" refers to positive regulation (including induction, Stimulates, enhances) or negatively regulates (including inhibition and weakening) to make the individual move toward the normal state of the physiological function; the so-called "individual" refers to a human or non-human mammal.

Studies have shown that SOD2 (Superoxide Dismutase Gene) and CAT (Hydrogen Peroxide Decomposition Enzyme Gene) performance increase system can help enhance the antioxidant capacity of cells. In addition, it is known that low or missing S OD2 and/or CAT gene expression is related to the occurrence of skin aging, cardiovascular disease, diabetes, Alzheimer’s disease and other diseases. For example, see: Anti-oxidant gene expression imbalance, aging and Down syndrome. Life Sciences . 76: 1407-1426 (2005), Catalase Deficiency and Type 2 Diabetes. Diabetes Care. 31(12):e93 (2008), SOD2 deficiency promotes aging phenotypes in mouse skin. AGING. 4(2) : 116-118 (2012), SOD2 in Mitochondrial Dysfunction and Neurodegeneration. Free Radic Biol Med . 62:4-12 (2013), and Sunlight damage to cellular DNA: Focus on oxidatively generated lesions. Free Radical Biology and Medicine . 107: 110-124 (2017), the full text of these documents is incorporated here for reference. Therefore, if the expression of S OD2 and/or CAT gene can be effectively improved, it can achieve anti-aging of skin (such as anti-skin photoaging), prevent cardiovascular disease, treat cardiovascular disease, prevent diabetes, treat diabetes, and prevent Azhai Alzheimer’s disease, and/or the effect of treating Alzheimer’s disease.

Studies have also pointed out that the MSH2 gene expression enhancement system in cells can help repair damaged DNA and proteins in cells. This can be seen for example: DNA repair mechanisms in dividing and non-dividing cells. DNA repair (Amst). 12( 8): 620-636 (2013), and Repairing Double-Strand DNA Breaks. Nature Education . 3(9):26 (2010). The full texts of these documents are incorporated here for reference. In addition, it is known that the low or missing MSH2 gene expression is related to skin aging. For example, see: Rejuvenation of Gene Expression Pattern of Aged Human Skin by Broadband Light Treatment: A Pilot Study. Journal of Investigative Dermatology . 133: 394-402 (2013 ), the full text of the document is incorporated here for reference. Therefore, if the expression of MSH2 gene can be effectively improved, the effect of anti-aging skin can be achieved.

Tgm1 and Keratin14 gene expression enhancement lines help maintain cell structure, see for example: A keratin scaffold regulates epidermal barrier formation, mitochondrial lipid composition, and activity. J. Cell Biol . 211(5):1057-1075 (2015), and Transglutaminase Function in Epidermis. J Invest Dermatol . 124(3):481-492 (2005), the full texts of these documents are incorporated here for reference. In addition, it is known that the low or missing expression of Tgm1 and/or Keratin14 genes is related to the occurrence of skin aging and dry skin-related diseases. For example, see: Mutations in the gene for transglutaminase 1 in autosomal recessive lamellar ichthyosis. Nat Genet . 9( 3):279-283 (1995), Transglutaminase Function in Epidermis. J Invest Dermatol . 124(3):481-492 (2005), Novel Mutations of the Transglutaminase 1 Gene in Lamellar Ichthyosis. J Invest Dermatol . 117(8) :214-218 (2001), and Disorders of keratinisation: from rare to common genetic diseases of skin and other epithelial tissues. Ulster Med J. 76 (2):72-82 (2007), the full texts of these documents are incorporated here Place for reference. Therefore, if the Tgm1 and/or Keratin14 gene expression can be effectively improved, it can help maintain skin health, moisturize, tighten the skin, reduce fine lines, improve dry skin, resist skin aging, prevent dry skin related diseases, and/ Or the effect of treating dry skin related diseases.

The increase in gene expression of GBA, FLG and HAS3 helps to form the skin barrier, increase the synthesis of hyaluronic acid, and increase the water content of cells. For example, see: Hyaluronan Synthase 3 Regulates Hyaluronan Synthesis in Cultured Human Keratinocytes. The Journal of Investigative Dermatology. 118: 43-48 (2002), Toll-like receptor 3 activation is required for normal skin barrier repair following UV damage. J Invest Dermatol . 135(2):569-578 (2015), Expression of differential genes involved in the maintenance of water balance in human skin by Piptadenia colubrina extract. J Cosmet Dermatol . 9(1):35-43 (2010)、New concept of the pathogenesis of atopic dermatitis: Interplay among the barrier, allergy, and pruritus as a trinity. J Dermatol Sci . 70(1): 3-11 (2013), and Filagrin in the frontline: role in skin barrier function and disease. Journal of Cell Science . 122(9): 1285-1294 (2009) , The full texts of these documents are incorporated here for reference. In addition, it is known that underexpression or deletion of GBA , FLG and/or HAS3 genes is related to skin aging and dry skin. For example, see: HAS3 underexpression as an indicator of poor prognosis in patients with urothelial carcinoma of the upper urinary tract and urinary bladder. Tumor Biol . 36(7):5441-5450 (2015)、Specifically Neuropathic Gaucher's Mutations Accelerate Cognitive Decline in Parkinson's. Ann Neurol . 80(5):674-685 (2016)、Filaggrin in the frontline: role in skin barrier function and disease. Journal of Cell Science . 122(9):1285-1294 (2009), and The filaggrin story: novel insights into skin-barrier function and disease. Trends Mol Med . 14(1):20-27 ( 2008), the full texts of these documents are incorporated here for reference. Therefore, if the expression of GBA , FLG and/or HAS3 genes can be effectively improved, it can help maintain skin health, moisturize, tighten skin, reduce fine lines, resist skin aging, and/or improve dry skin.

Wuwei Mountain Wupi Tea ( Pyrenaria buisanensis ) is a plant endemic to Taiwan and belongs to the Camellia family and the genus Wupi Tea in taxonomy. The inventor of this case discovered that Wuwei Mountain Wupi tea extract can effectively improve the performance of genes such as SOD2 , CAT , MSH2 , Tgm1 , Keratin14 , FLG , GBA , HAS3 and so on. Therefore, the present invention relates to the application of Wuwei mountain ebony tea extract, including the use of Wuwei mountain ebony tea extract for moisturizing, whitening, firming skin, reducing fine lines, anti-aging skin, improving dry skin, and promoting skin transparency Formation of plasmic acid and/or help maintain skin health, use Wuwei Mountain Wupi tea extract to prepare a pharmaceutical composition, provide a skin care composition containing an effective amount of Wuwei Mountain Wupi tea extract, food composition or A pharmaceutical composition and a method for administering an effective amount of the aforementioned skin care composition, food composition or pharmaceutical composition to an individual in need. The skin care product composition provided according to the present invention can be used for moisturizing, whitening, skin tightening, reducing skin fine lines, anti-aging skin, and/or improving skin dryness; the food composition provided according to the present invention can be used To promote the formation of skin hyaluronic acid and/or help maintain skin health; the pharmaceutical composition provided according to the present invention can be used to resist light damage, repair skin tissue, prevent skin diseases, and/or treat skin diseases, and It can be used for at least one of the following: prevention of cardiovascular disease, treatment of cardiovascular disease, prevention of diabetes, treatment of diabetes, prevention of neurodegenerative diseases, and treatment of neurodegenerative diseases. For example, the pharmaceutical composition can be used to prevent dry skin related diseases (including ichthyosis), treat dry skin related diseases, prevent ischemic stroke, treat ischemic stroke, prevent Alzheimer's disease, and/or treat Alzheimer's disease. In addition, the pharmaceutical composition and method provided according to the present invention can also be used to enhance the performance of SOD2 gene, CAT gene, MSH2 gene, Tgm1 gene, Keratin14 gene, FLG gene, GBA gene, and/or HAS3 gene.

The Wuwei Mountain Wupi tea extract used in the present invention can be provided by extracting Wuwei Mountain Wupi tea raw materials with a polar solvent, wherein the polar solvent can be water, C1-C4 alcohols, or a combination thereof. Among them, there are no special restrictions on the use parts of Wuwei Mountain Wupi tea raw materials. It can be the whole plant of Wuwei Mountain Wupi Tea or different parts of Wuwei Mountain Wupi Tea, such as the roots, stems, leaves of Wuwei Mountain Wupi Tea. And/or flower department. In some embodiments of the present invention, the leaves of Wuwei Mountain Wupi Tea are used to prepare the Wuwei Mountain Wupi Tea Extract. In addition, the amount of extraction solvent is not particularly limited, and the amount that can disperse the material is usually used. For example, a polar solvent can be used in the extraction step: Wuwei Mountain Wupi Tea Leaf = 1-20:1 by weight. In a specific embodiment of the present invention, the extraction solvent: Wuwei Mountain Wupi tea leaves = 10:1 by weight ratio is used for extraction.

In the extraction step, a suitable extraction time can also be selected depending on the polar solvent used. Taking water as the polar solvent and the weight ratio of water: Wuwei Mountain Wupi tea leaves of about 10-20:1 as an example, the extraction usually takes 0.5 to 2 hours. In addition, if necessary, before or during the extraction step, one or more operations such as heating, cooling, stirring, ultrasonic wave, etc. may be supplemented, or the raw materials of Wuwei Mountain Wupi tea may be finely crushed before the extraction step to improve the extraction. effect. For example, extraction can be performed at 75 to 95°C. In a specific embodiment of the present invention, extraction is performed at 85°C for 0.5 hours.

The Wuwei Mountain Wupi tea extract used according to the present invention may be the raw extract obtained by extraction, or may be provided by performing one or more steps such as filtering, sterilizing, concentrating, and diluting the raw extract as needed, In order to enhance the convenience of using the extract. In order to achieve the maximum extraction efficiency as much as possible, if necessary, the Wuwei Mountain Wupi tea raw materials can be repeatedly extracted with the same or different polar solvents, and the extracts obtained from the multiple extractions can be combined. For example, the obtained extract can be concentrated under reduced pressure to maintain the stability of the active ingredients in the Wuwei Mountain Wupi tea extract during the storage period. If necessary, the temperature during concentration under reduced pressure can be adjusted. For example, it can be concentrated under reduced pressure at 45 to 70°C. In a specific embodiment of the present invention, concentration under reduced pressure is performed at 55 to 65°C.

The skin care composition provided according to the present invention can be used for whole body or topical use, and can be in any suitable form without any special restrictions. Depending on the intended use, it can be in a corresponding suitable dosage form. For example, it can be In the form of lotion, cream, gel (such as hydrogel), or solution (such as essence, lotion) for direct external use, but not limited to this.

The food composition provided according to the present invention can be in any suitable form without any special restrictions. For example, the food composition can be prepared into a form that can be swallowed or drunk, such as health food, beauty drink, etc., but not limited to this.

The pharmaceutical composition provided according to the present invention can be administered systemically or locally, and can be delivered through various drug delivery systems (DDS), including oral drug delivery systems (oral drug delivery systems) and transdermal drug delivery systems (Transdermal drug delivery system), injection delivery system (injection delivery system), etc. For example, but not limited to this, the pharmaceutical composition provided according to the present invention can be made of liposomes, microcapsules, nanoparticles, microneedles, etc. The system is delivered to achieve the effects of improving the bioavailability, controlling the drug release rate, accurately administering drugs for the lesion, and reducing drug side effects.

The pharmaceutical composition provided according to the present invention can be in any suitable form without any special restrictions. It is in a corresponding suitable dosage form depending on the intended use; for example, but not limited to this, the pharmaceutical composition The drug can be administered orally or parenterally (for example: transdermal administration, subcutaneous injection) to individuals in need. Depending on the use form and purpose, a suitable carrier can be selected to provide the pharmaceutical composition, wherein the carrier includes excipients, diluents, adjuvants, stabilizers, absorption delay agents, disintegrating agents, solubilizers, Emulsifiers, antioxidants, binders, binders, tackifiers, dispersants, suspending agents, lubricants, hygroscopic agents, etc.

Taking a dosage form suitable for oral administration as an example, the pharmaceutical composition provided according to the present invention may contain any pharmaceutically acceptable substance that does not adversely affect the desired benefits of the active ingredient (ie, Wuwei Mountain Wupi tea extract) Carriers, such as: water, saline, dextrose, glycerin, ethanol or the like, oils (such as olive oil, castor oil, cottonseed oil, peanut oil, corn oil, and germ oil), polyethylene glycol, Starch, kaolinite, bentonite, sodium citrate, gelatin, agar, carboxymethyl cellulose, acacia, alginic acid and its salts, glyceryl monostearate, calcium stearate (Calcium stearate), and a combination of the foregoing. Any convenient method can be used to provide the pharmaceutical composition in a dosage form suitable for oral administration, such as: lozenges (such as dragees), pills, capsules, granules, powders, liquid extracts, solutions, syrups, Suspensions, tinctures, etc.

Taking a dosage form suitable for transdermal administration as an example, the pharmaceutical composition provided according to the present invention may also contain any medicine that does not adversely affect the desired benefits of the active ingredient (ie, Wuwei Mountain Wupi tea extract) Acceptable carriers include water, mineral oil, propylene glycol, polyethylene oxide, liquid paraffin, sorbitan monostearate, and polysorbate 60. Any suitable method can be used to provide the pharmaceutical composition in a dosage form suitable for transdermal administration, such as patches, lotions, creams, gels (such as hydrogels), and pastes (such as dispersions) for direct external use. Paste, ointment), spray, or solution (such as suspension), but not limited to this.

As for injection or drip dosage forms suitable for subcutaneous injection, the pharmaceutical composition may contain one or more such as isotonic solutions, salt buffers (such as phosphate buffer or citrate buffer), solubilizers, and emulsifiers. Ingredients, 5% sugar solution, and other carriers. Alternatively, the pharmaceutical composition is prepared as a pre-injection solid, the pre-injection solid is provided in a dosage form that is soluble in other solutions or suspensions, or an emulsifiable dosage form, and the pre-injection solid is provided before being administered to an individual in need The solid is dissolved in other solutions or suspensions or emulsified before injection to provide the desired injection.

Optionally, the skin care composition, food composition, or pharmaceutical composition provided according to the present invention may further contain additives in a suitable amount, for example, to improve the palatability of the food composition or pharmaceutical composition when taken. Flavoring agents, toners, coloring agents, etc. for the sense and visual perception, as well as buffers, preservatives, preservatives, antibacterial agents that can improve the stability and storage of the skin care composition, food composition, or pharmaceutical composition Agents, antifungal agents, etc. In addition, the skin care composition or the pharmaceutical composition may optionally contain one or more other active ingredients to further enhance the efficacy of the skin care composition or the pharmaceutical composition or increase the flexibility and the degree of formulation of the formulation, as long as the Other active ingredients do not have any adverse effects on the benefits of the active ingredients of the present invention (ie, Wuwei Mountain Wupi Tea Extract).

The skin care composition, food composition, or pharmaceutical composition provided according to the application of the present invention can be administered at different frequencies such as once a day, multiple times a day, or once a few days, depending on the needs and age of the individual. , Body weight, health condition, and purpose of administration. In addition, the content of Wuwei Mountain Wupi tea extract in the skin care composition, food composition or pharmaceutical composition can also be adjusted according to actual application requirements.

In accordance with the present invention, the Wuwei Mountain Wupi tea extract is used for moisturizing, whitening, firming the skin, reducing fine lines, anti-aging skin, improving dry skin, promoting the formation of skin hyaluronic acid, and/or helping to maintain skin health In the application, the Wuwei Mountain Wupi tea extract used can be provided in the form of a skin care composition or a food composition. The administration mode, administration route, administration form, application frequency, and related applications of the skin care composition and food composition are also as described above.

As mentioned above, the present invention also provides a method for moisturizing, whitening, firming skin, reducing skin fine lines, anti-aging skin, improving dry skin, promoting the formation of skin hyaluronic acid, and/or helping to maintain skin health, which includes An effective amount of Wuwei Mountain Wupi Tea Extract is administered to an individual in need. The aforementioned system in need refers to individuals who need to improve skin quality and/or skin condition, or prevent skin quality and/or skin condition from getting worse; in particular, the system has thickened skin keratin and skin wrinkles , Skin spots, dull skin, dry skin, sagging skin, and/or skin aging, or an individual who works outdoors for a long time, but not limited to this. In the aforementioned method, the Wuwei Mountain Wupi tea extract used can be administered to the individual in need in the form of the aforementioned skin care product composition or food composition. The administration mode, administration route, administration form, application frequency, and related applications of the skin care composition and food composition are also as described above.

The present invention also provides an anti-light damage, repair of skin tissue, prevention of skin diseases, treatment of skin diseases, prevention of cardiovascular diseases, treatment of cardiovascular diseases, prevention of diabetes, treatment of diabetes, prevention of neurodegenerative diseases, and/or treatment of neurodegenerative diseases The method of disease includes administering an effective amount of Wuweishan Wupi tea extract to an individual in need. The aforementioned needful system refers to people with skin diseases, people with skin diseases, and/or high-risk groups of skin diseases; or, people with cardiovascular disease, people with diabetes, and people with neurodegenerative diseases Persons, high-risk groups for cardiovascular disease, high-risk groups for diabetes, and/or high-risk groups for neurodegenerative diseases. In the aforementioned method, the Wuwei Mountain Wupi tea extract used can be administered to the individual in need in the form of the above-mentioned pharmaceutical composition. The administration mode, administration route, administration form, administration frequency, and related applications of the pharmaceutical composition are also as described above.

The present invention also provides a method for enhancing the performance of SOD2 gene, CAT gene, MSH2 gene, Tgm1 gene, Keratin14 gene, FLG gene, GBA gene, and/or HAS3 gene, which comprises administering to an individual in need An effective amount of Wuwei Mountain Wupi Tea Extract. The aforementioned needful system refers to individuals whose SOD2 gene, CAT gene, MSH2 gene, Tgm1 gene, Keratin14 gene, FLG gene, GBA gene, and/or HAS3 gene have deletions, mutations or low performance. In the aforementioned method, the Wuwei Mountain Wupi tea extract used can be administered to the individual in need in the form of the above-mentioned pharmaceutical composition. The administration mode, administration route, administration form, administration frequency, and related applications of the pharmaceutical composition are all as described above.

The following examples further illustrate the present invention. The embodiments are only provided as illustrations, and are not used to limit the protection scope of the present invention. The scope of protection of the present invention is shown in the attached patent scope.

Example

[ preparation Example ] ：Preparation of Wuwei Mountain Wupi Tea Extract

The Wuwei Mountain Wupi tea provided by Ku Yan Zhuoyun Plant Conservation Center (KBCC) of Taiwan is processed as follows to provide Wuwei Mountain Wupi tea extract: I. Wash the Wuwei Mountain Wupi tea leaves with reverse osmosis (RO) water If necessary, repeat the aforementioned washing operation; II. Mix the cleaned Wuwei Mountain Wupi tea leaves with water (Wuwei Mountain Wupi tea leaves and water weight ratio is 1:10), and at 85° Perform extraction at C for 0.5 hours to provide an extract; III. After the extract is cooled to room temperature, filter with a 400 mesh filter to provide a filtrate; and IV. At 55 to 65°C , The filtrate is concentrated under reduced pressure to provide a concentrated extract (that is, the Wuwei mountain ebony tea extract of the present invention).

Example 1 ：The effect of Wuwei Mountain Wupi Tea Extract in enhancing the antioxidant capacity of cells

It is known that when the content of Reactive Oxygen Species (ROS) in skin cells is too high, it will cause cell tissue damage and DNA damage, leading to skin aging. To prevent ROS from being toxic to skin cells, organisms secrete enzymes such as superoxide dismutase (SOD) to decompose excessive oxidized substances in the body. The following experiments were performed to confirm the effectiveness of the Wuwei Mountain Wupi tea extract of the present invention in inhibiting the generation of ROS in skin cells and improving the SOD activity of skin cells.

( 1-1 ) ROS Content detection

Human skin fibroblasts (CCD-966SK; purchased from ATCC) were cultured in MEM medium (Minimum essential medium, purchased from Gibco, product number: 61100-061) for 24 hours, after which the cells were divided into four groups, and Carry out the following treatments: (1) Control group: Place the cells in MEM medium and culture for 1 hour. (2) "H ₂ O ₂ "group: The cells were cultured in MEM medium for 1 hour, and then H ₂ O ₂ was added to the medium to make the final concentration in the medium reach 1 millimolar, and continue to culture 1 hour. (3) "H ₂ O ₂ + extract (2)" group: The cells were cultured in MEM medium containing 2 mg per ml of Wuwei Mountain Wupi tea extract provided by [Preparation Example] for 1 hour, and then , Add H ₂ O ₂ to the culture medium to make the final concentration in the culture medium reach 1 millimolar concentration, and continue culturing for 1 hour. (4) "H ₂ O ₂ + extract (1)" group: The cells were cultured in MEM medium containing 1 mg per ml of Wuwei Mountain Wupi tea extract provided by [Preparation Example] for 1 hour, and then , Add H ₂ O ₂ to the culture medium to make the final concentration in the culture medium reach 1 millimolar concentration, and continue culturing for 1 hour.

Take the above groups of cells and treat them with DCFH-DA dye (purchased from Sigma, product number: D6883) for 15 minutes, and then detect the excitation light wavelength of each group at 450-490 nm and emission light with a flow cytometer. Fluorescence intensity of wavelength 510-550nm. Among them, since ROS can convert DCFH-DA (without fluorescence) to DCF (with fluorescence), the measured fluorescence intensity can represent the ROS content in the cell, and the higher the fluorescence intensity, the ROS content in the cell. Higher. Finally, the Student t-test was used for statistical analysis, and the results of the control group were used as a benchmark to calculate the ROS content in the cells of other groups. The results are shown in Figure 1.

It can be seen from Figure 1 that compared with the control group, _{the ROS content of the "H 2} O ₂ "group was significantly increased, which shows that the "H ₂ O ₂ " group can simulate the state of high levels of peroxides in skin cells. However, compared with the "H ₂ O ₂ "group, _{the ROS content of the "H 2} O ₂ + extract (2)" group and the "H ₂ O ₂ + extract (1)" group were significantly reduced. The foregoing results show that Wuwei Mountain Wupi tea extract can effectively reduce the ROS content in skin cells and has an antioxidant effect.

( 1-2 ) SOD Activity detection

Human skin fibroblasts were cultured in MEM medium for 24 hours. After that, the cells were divided into four groups and the following treatments were performed: (1) Control group: cells were cultured in MEM medium for 24 hours. (2) "H ₂ O ₂ "group: The cells were cultured in MEM medium for 24 hours, and then H ₂ O ₂ was added to the medium to make the final concentration in the medium reach 1 millimolar, and continue to culture 6 hours. (3) "H ₂ O ₂ + extract (2)" group: The cells were cultured for 24 hours in MEM medium containing 2 mg per ml of Wuwei Mountain Wupi tea extract provided by [Preparation Example], and then , Add H ₂ O ₂ to the culture medium to make the final concentration in the culture medium reach 1 millimolar concentration, and continue culturing for 6 hours. (4) "H ₂ O ₂ + extract (1)" group: The cells were cultured in a MEM medium containing 1 mg per ml of Wuwei Mountain Wupi tea extract provided by [Preparation Example] for 24 hours, and then , Add H ₂ O ₂ to the culture medium to make the final concentration in the culture medium reach 1 millimolar concentration, and continue culturing for 6 hours.

Then, using the principle of enzyme action and colorimetric determination, the SOD activity test kit (purchased from Cayman, product number: 706002) was used to perform the following treatments on the above groups of cells: (a) Remove the medium and use phosphate buffered saline (PBS) ) After washing the cells, add Trypsin (purchased from Thermo, product number: 15400-054) and react for 3 minutes. After the cells are separated from the culture plate, collect the cells in a centrifuge tube for centrifugation (400 g, 5 minutes), and remove the supernatant After washing with PBS, centrifuge (400 g, 5 minutes), and remove the supernatant; (b) Add 50 microliters of cell extraction solution and centrifuge (12000 g, 30 minutes) at 4°C, Remove the insoluble matter to obtain a cell extract; (c) Combine 10 microliters of the cell extract obtained from step (b) with 200 microliters of tetrazolium salt solution and 20 microliters of xanthine oxidized After the enzyme reaction solution (xanthine oxidase solution) is evenly mixed, react for 30 minutes; (d) Next, measure the product of step (c) with an enzyme immunoassay reader (ELISA reader; purchased from Epoch, product number: 1212171) The absorbance at a wavelength of 450 nm; and (e) Replace the cell extract used in step (c) with the SOD standard, repeat the processing of steps (c) to (d), and record the absorbance (hereinafter referred to as , SOD standard absorbance value).

After that, use the absorbance value measured with the SOD standard in step (e) to calculate the SOD activity of each group of cells according to the formula provided in the instruction manual of the SOD activity test kit (purchased from Cayman, product number: 706002), The results are shown in Figure 2.

It can be seen from Figure 2 that _{the SOD activity of the "H 2} O ₂ "group is extremely low. However, the groups treated with Wuwei Mountain Wupi tea extract (including the "H ₂ O ₂ + extract (2)" group and the "H ₂ O ₂ + extract (1)" group) were induced by _{H 2} O ₂ Under the oxidative pressure, the SOD activity is significantly improved. The foregoing results show that Wuwei Mountain Wupi tea extract can effectively improve the SOD activity of skin cells, help the decomposition of peroxides, and have an antioxidant effect.

Example 2 : Wuwei Mountain Wupi Tea Extract Yu Sheng SOD2 , CAT and MSH2 Gene expression The benefit of

The human skin fibroblasts were cultured in MEM medium for 24 hours. Then, the cells were divided into six groups and the following treatments were performed: (1) Control group: cells were cultured in MEM medium for 6 hours. (2) "UVA" group: The cells were cultured in MEM medium for 6 hours, and then the cells were irradiated with UVA (15 joules/cm²) for 6 hours. (3) "Extract (1)-6hr" group, and "Extract (1)-24hr" group: put the cells in each milliliter containing 1 mg [Preparation Example] Wuwei Mountain Wupi tea extract provided Cultured in MEM medium for 6 hours and 24 hours respectively, and then irradiated the cells with UVA (15 Joules/cm²) for 6 hours. (4) "Extract (2)-6hr" group, and "Extract (2)-24hr" group: put the cells in each milliliter containing 2 mg [Preparation Example] Wuwei Mountain Wupi tea extract provided Cultured in MEM medium for 6 hours and 24 hours respectively, and then irradiated the cells with UVA (15 Joules/cm²) for 6 hours.

After that, the above-mentioned cells were collected separately, RNA extraction kit (RNA Extraction Kit, purchased from Geneaid) was used for RNA extraction, and then reverse transcriptase (SuperScript® III Reverse Transcriptase, purchased from Invitrogrn) was used to reverse the RNA. Transcribe into cDNA. Then, use the ABI Step One Plus instrument and the KAPA SYBR FAST qPCR kit to perform qPCR (quantitative polymerase chain reaction) on the provided cDNA to detect the gene expression levels of SOD2 , CAT and MSH2 in each group of cells. Finally, the Student t-test was used for statistical analysis, and the control group was used as a benchmark (that is, the gene performance of the control group was set to 1 times) to calculate the relative gene expression of the remaining groups. The results are shown in Figures 3 to 5.

As can be seen from Figures 3 to 5, compared with the "UVA" group, the "extract (1)-6hr" group, the "extract (1)-24hr" group, the "extract (2)-6hr" group, and The expression levels of SOD2 and MSH2 genes in the cells of the "extract (2)-24hr" group were significantly improved, and the CAT genes of the "extract (2)-6hr" group and the "extract (2)-24hr" group of cells Performance has also increased significantly. The foregoing results show that Wuwei Mountain Wupi tea extract can effectively enhance the expression of SOD2 , CAT and MSH2 genes, so it can be used to enhance the antioxidant capacity of cells, help repair damaged DNA and proteins in cells, and achieve anti-aging, It has the effect of resisting light damage and repairing skin tissue, and can be used to prevent cardiovascular disease, treat cardiovascular disease, prevent diabetes, treat diabetes, prevent Alzheimer's disease, and/or treat Alzheimer's disease.

Example 3 ：The Wuwei Mountain Wupi Tea Extracts to enhance FLG , HAS3 , GBA , Tgm1 and Keratin14 Gene expression The benefit of

Culture human epidermal primary keratinocytes (HPEK-50; purchased from CELLnTEC, product number: PR3D-HPEK-50) in a serum-free medium for keratinocytes (Keratinocyte-SFM; purchased from Thermo, product number: 17005042) for 24 hours Then, divide the cells into two groups and perform the following treatments: (1) Control group: Place the cells in Keratinocyte-SFM medium for 6 hours. (2) "Extract" group: Cells were placed in a Keratinocyte-SFM medium containing 0.03125 mg/ml [Preparation Example] Wuwei Mountain Wupi tea extract provided for 6 hours.

After that, the above-mentioned cells were collected separately, RNA extraction kit (RNA Extraction Kit, purchased from Geneaid) was used for RNA extraction, and then reverse transcriptase (SuperScript® III Reverse Transcriptase, purchased from Invitrogrn) was used to reverse the RNA. Transcribe into cDNA. Then, use the ABI Step One Plus instrument and the KAPA SYBR FAST qPCR kit to perform qPCR on the aforementioned cDNA to detect the gene expression levels of FLG , HAS3 , GBA , Tgm1 , and Keratin14 in each group of cells. Finally, the SCORE method is used for statistical analysis, and the control group is used as a benchmark (that is, the gene performance of the control group is set to 1 times) to calculate the relative gene expression of the remaining groups. The results are shown in Figures 6 to 8.

It can be seen from Fig. 6 to Fig. 8 that compared with the control group, the expression levels of Tgm1 , Keratin14 , FLG , GBA , and HAS3 genes of the cells in the "extract" group were significantly improved. The foregoing results show that Wuwei Mountain Wupi tea extract can indeed increase the expression of Tgm1 , Keratin14 , FLG , GBA , and HAS3 genes, so it can be used to help maintain cell structure, help form skin barriers, increase hyaluronic acid synthesis, and Increase the moisture content of cells to achieve the effect of moisturizing, firming the skin, reducing fine lines, and/or improving dry skin, and can be used to prevent skin aging, prevent dry skin-related diseases, and/or treat dry skin-related diseases.

Example 4 : Human test

This experiment was conducted in a self-control method. At week 0, 10 volunteers used the DermaLab® Combo skin moisture analyzer to measure and record the skin moisture content. In addition, C+K electronic Mexameter® MX18 red pigment measuring probe is used to measure melanin content, and C+K electronic MPA580 skin elasticity tester is used to measure skin laxity and record. Then, each subject applied Wuwei Mountain Wupi Tea Essence (containing 1% of the Wuwei Mountain Wupi Tea Extract provided by [Preparation Example], based on the total weight of the essence) on half of the face in the morning and evening. , And smear the other half of the face with the essence containing only the base solvent (that is, without the Wuwei Mountain ebony tea extract of the present invention, but other ingredients are the same as the Wuwei Mountain ebony tea essence) for 6 weeks; thereafter , And then measure and record the skin melanin content, skin laxity, and skin moisture content of the left and right sides of the face with a multifunctional skin detector. Next, statistical analysis was performed with Student t-test, and the results of week 0 were used as a benchmark (ie, week 0 was set as 100%) to calculate the melanin content, skin laxity, and skin laxity after 6 weeks. Skin moisture content. The results are shown in Figures 9-11.

It can be seen from Figures 9 to 11 that after 6 weeks of using the essence containing Wuwei Mountain Wupi tea extract of the present invention, the subject's skin melanin content and skin laxity were significantly reduced, and the skin moisture content was significantly increased. The foregoing results show that the Weishan ebony tea extract of the present invention does have the effects of moisturizing, whitening, firming the skin, and reducing skin fine lines.

Figure 1 shows the effect of the Wuwei mountain ebony tea extract of the present invention in reducing the ROS content in cells. The cell line of the control group was cultured in a medium without the Wuwei mountain ebony tea extract for 1 hour, "H ₂ O _{The cell lines of the 2} " group were cultured for 1 hour in the medium without Wuwei Mountain Wupi tea extract, and then _{treated with H 2} O ₂ for 1 hour. The "H ₂ O ₂ + extract (1)" group and "H ₂ Cells in the "O ₂ + extract (2)" group were cultured in a medium containing 1 and 2 mg of Wuweishan Wupi tea extract per ml for 1 hour, and then _{treated with H 2} O ₂ for 1 hour;

Figure 2 shows the effect of the Wuwei mountain ebony tea extract of the present invention in enhancing the SOD activity of cells. Among them, the cell line of the control group was cultured in a medium without the Wuwei mountain ebony tea extract for 24 hours, "H ₂ O _{The cell lines of group 2} were cultured for 24 hours in the medium without Wuwei Mountain Wupi tea extract, and then _{treated with H 2} O ₂ for 6 hours. The _{cell lines of group "H 2} O ₂ + extract (1)" and "H ₂ The cells of the "O ₂ + extract (2)" group were cultured in a medium containing 1 and 2 mg of Wuweishan Wupi tea extract per ml for 24 hours, and then _{treated with H 2} O ₂ for 6 hours;

Figure 3, Figure 4 and Figure 5 show the effect of the Wuwei Mountain Wupi tea extract of the present invention in enhancing the expression of SOD2 , CAT and MSH2 genes. Among them, Figure 3 shows the expression of SOD2 genes in each group of cells, and Figure 4 shows The expression level of CAT gene of each group of cells, Figure 5 shows the expression level of MSH2 gene of each group of cells, and among them, the cell line of the control group was cultured in the medium without Wuwei Mountain Wupi tea extract for 6 hours, "UVA Cell lines in the "group" were cultured in a medium without Wuwei Mountain Wupi tea extract for 6 hours, and then irradiated with UVA for 6 hours, the "extract (1)-6hr" group and the "extract (1)-24hr" group The cell lines were cultured for 6 hours and 24 hours in a medium containing 1 mg of Wuwei Mountain Wupi tea extract per milliliter, and then irradiated with UVA for 6 hours. The "extract (2)-6hr" group and the "extract ( 2) Cells in the "-24hr" group were cultured in a medium containing 2 mg of Wuwei Mountain Wupi tea extract per ml for 6 hours and 24 hours, and then irradiated with UVA for 6 hours;

Figure 6, Figure 7, and Figure 8 show the effect of Wuwei Mountain Wupi tea extract of the present invention in enhancing the expression of Tgm1 , Keratin14 , FLG , GBA and HAS3 genes. Among them, Figure 6 shows the Tgm1 gene and Keratin14 of each group of cells Gene expression level, Figure 7 shows the FLG gene expression level of each group of cells, Figure 8 shows the GBA gene and HAS3 gene expression level of each group of cells, and among them, the cell line of the control group does not contain Wuwei Shanwu Cultured in the medium of the extract of skin tea for 6 hours, the cells of the "extract" group were cultured in the culture medium containing the extract of Wuweishan Wupi tea for 6 hours;

Figure 9, Figure 10, and Figure 11 respectively show the effects of the Wuwei Mountain Wupi tea extract of the present invention in reducing skin melanin content, reducing skin laxity, and increasing skin moisture content.

Claims (6)

A use of Wuwei Mountain Wupi tea ( Pyrenaria buisanensis ) extract in preparing a composition, wherein the extract is provided by extracting Wuwei Mountain Wupi tea with a polar solvent, and the composition is used in at least one of the following : Promote the formation of skin hyaluronic acid, moisturize, whiten, tighten skin, reduce skin fine lines, anti-aging skin, and improve dry skin. Such as the use of claim 1, wherein the composition is a food composition or a skin care product composition. Such as the use of claim 1, wherein the polar solvent is selected from the following group: water, C1-C4 alcohols, and a combination of the foregoing. The use according to any one of claims 1 to 3, wherein the composition is applied by smearing or oral administration. A use of Wuwei Mountain Wupi tea extract in the preparation of a pharmaceutical composition, wherein the pharmaceutical composition is used for at least one of the following: anti-photodamage, repairing skin tissue, preventing dry skin diseases, and treating dry skin diseases . A use of Wuwei Mountain Wupi tea extract in the preparation of a medical composition, wherein the medical composition is used to enhance the performance of at least one of the following genes: SOD2 gene, CAT gene, MSH2 gene, Tgm1 gene, Keratin14 gene , FLG gene, GBA gene, and HAS3 gene.

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