JP2013194007A - Anti-oxidative stress agent and application of the same - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a novel anti-oxidative stress agent that can be substituted for a conventionally known anti-oxidative stress agent, and an application of the novel agent.SOLUTION: An anti-oxidative stress agent includes a compound 1 and/or a compound 2 described in the specification as an active ingredient. The agent can be incorporated into a drug composition, a skin care composition (cosmetic or the like), and food and drinks.

Description

  The present invention relates to an antioxidant stress agent and use thereof.

  In a living body that obtains energy using oxygen, reactive oxygen species (ROS) are always produced. ROS is highly reactive and modifies functional molecules such as proteins, lipids, DNA, etc. to cause cell dysfunction. Although the cells have a defense mechanism against ROS, oxidative stress occurs when excessive ROS accumulates, and it is thought that the onset and symptoms of diseases associated with oxidative stress such as neurodegenerative diseases and ischemic cerebrovascular disorders are developed. Yes.

  NF-E2 related factor 2 (Nrf2), a transcription factor that plays an important role in xenobiotic metabolism and oxidative stress response, is an antioxidant response element that exists upstream of genes such as phase II enzymes and antioxidant proteins. By binding to (antioxidant response element; ARE), the expression of these genes is regulated.

  Nrf2 binds to cysteine-rich protein Keap1 and localizes in the cytoplasm in the absence of stimulation. However, when a cysteine residue present in Keap1 is oxidatively modified by an electrophilic substance or ROS, a conformational change of Keap1 occurs, and Nrf2 dissociates from Keap1 and moves into the nucleus. Nrf2 transferred into the nucleus forms a heterodimer with a small Maf group transcription factor and binds to ARE, thereby expressing antioxidant proteins such as HO-1 (heme oxygenase 1), glutathione-S-transferase, NQO1 and the like. To enhance.

  Thus, since activation of Nrf2 enhances the expression of a group of antioxidant proteins, the Nrf2-ARE pathway is thought to play a central role in the biological defense mechanism against oxidative stress. Apomorphine (Apo), a dopamine receptor agonist, has a neuronal protective effect against oxidative stress, but it has been revealed that the Nrf2-ARE pathway is involved in this protective action of Apo (non-patented) Reference 1).

  On the other hand, Non-Patent Document 2 reports that 9-oxo-10E, 12Z-octadecadienoic acid and 13-oxo-9Z, 11E-octadecadienoic acid, which are linoleic acid derivatives, activate the Nrf2-ARE pathway. It has been suggested that these linoleic acid derivatives may reduce tissue damage caused by oxidative stress through induction of expression of phase 2 enzymes of drug metabolism and antioxidant proteins. However, it is not known that other linoleic acid derivatives can activate the Nrf2-ARE pathway and that this reduces oxidative stress.

Hirokazu Hara, YAKUGAKU ZASSHI 127 (8) 1199-1205 (2007) Wang et al., Prostaglandins Leukot Essent Fatty Acids. Author Manuscript; available in PMC 2010 July 1.

  An object of this invention is to provide the novel antioxidant stress agent which can substitute for a conventionally well-known antioxidant stress agent, and its use.

  In view of the current state of the prior art as described above, the present inventors have intensively studied. In the process, surprisingly, a linoleic acid derivative different from that disclosed in Non-Patent Document 2 activates the Nrf2-ARE pathway, and thereby the expression of antioxidant proteins such as HO-1. It was found that an antioxidative stress action can be shown through the enhancement of. And based on this knowledge, it came to complete this invention.

  That is, according to one aspect of the present invention, the following compound 1 and / or compound 2:

As an active ingredient, an antioxidant stress agent is provided. In addition, it is preferable that the action expression of the said antioxidant stress agent is an expression enhancement effect of antioxidant protein.

  According to another aspect of the present invention, the pharmaceutical composition contains the above antioxidant stress agent and a pharmaceutically acceptable carrier, and is used for the prevention and / or treatment of a disease or condition associated with oxidative stress. Things are also provided.

  Furthermore, according to still another aspect of the present invention, there is provided a skin external preparation composition containing the above antioxidant stress agent and a pharmaceutically acceptable carrier. Cosmetics comprising the skin external preparation composition are also provided.

  Moreover, according to the further another form of this invention, the food / beverage products containing the said antioxidant stress agent are also provided.

  Furthermore, according to still another aspect of the present invention, there is provided a food or drink containing an effective amount of the above-mentioned antioxidant stress agent, comprising neurodegenerative disease, ischemic cerebrovascular disorder, ischemic heart disease, inflammatory bowel disease, and Food / beverage products having a function of preventing and / or improving eye diseases and having the function indication are also provided. The food or drink is preferably a health food, a functional food, a food for specified health use, a dietary supplement, a food with a disease risk reduction label, or a food for a sick person.

  ADVANTAGE OF THE INVENTION According to this invention, the novel antioxidant stress agent which can substitute for a conventionally well-known antioxidant stress agent, and its use are provided.

In an Example, it is the chromatogram obtained when the compound was isolated by HPLC conditions 1. In an Example, it is the chromatogram obtained when the compound was isolated by HPLC conditions 2. In an Example, it is a graph which shows the result of having evaluated the ARE enhancer activity of the compound isolated from the edible part of artichoke by the reporter assay. In an Example, it is an electrophoresis photograph which shows the result evaluated by the western blotting about the effect with respect to the expression of the HO-1 protein of the compound isolated from the edible part of artichoke.

  Hereinafter, specific embodiments for carrying out the present invention will be described in detail, but the technical scope of the present invention is not limited to the following specific embodiments.

  One aspect of the present invention is the following compound 1 and / or compound 2:

Is an anti-oxidative stress agent.

  About the compound 1 and the compound 2 which are the active ingredients of the antioxidant stress agent which concerns on this form, when a commercial item exists, what acquired the said commercial item may be used, and what was synthesize | combined itself is used. Also good. Moreover, you may use what was extracted and refine | purified from the natural product depending on the case. As described in Examples described later, the present inventors extracted the above-mentioned Compound 1 and Compound 2 from the edible portion of Cynara scolymus, a perennial plant belonging to the genus Chrysanthemum. Therefore, it is also possible to obtain the above compound 1 or compound 2 by using a method described in Examples described later, or a method obtained by partially modifying the method.

  The compound represented by the above-mentioned chemical formula 1 may be the compound itself, and includes pharmaceutically acceptable salts, prodrugs and solvates of the compound as long as it is applicable. For example, a salt can also be formed between a cation and a negatively charged group on the compound represented by Chemical Formula 1. Suitable cations include sodium cations, potassium ions, magnesium ions, calcium ions, and ammonium cations such as tetramethylammonium ions. The compounds also include those salts containing quaternary nitrogen atoms. Examples of prodrugs include esters and other pharmaceutically acceptable derivatives, which can provide active compounds upon administration to a subject. Solvate means a complex formed between an active compound and a pharmaceutically acceptable solvent. Pharmaceutically acceptable solvents include water, ethanol, isopropanol, ethyl acetate, acetic acid and ethanolamine.

  As described above, according to one aspect of the present invention, an antioxidative stress agent is provided. “Oxidative stress” is preferable for a living body that is caused by the balance between an oxidative reaction and an antioxidative reaction and leaning toward the former. There is no state. When ROS or the like that cannot be processed by the in-vivo antioxidant system is generated, lipids, proteins, enzymes, and genetic DNA that carry the structure and functions of the living body are oxidized and damaged, The structure and function of the are disturbed. As a result, various diseases are induced, aging is promoted, and cancer and lifestyle-related diseases are easily affected.

  The “antioxidant stress agent” is an agent that repairs the state in which the balance between the oxidation reaction and the antioxidant reaction in the living body is lost and restores the structure and function of the living body. Antioxidant stress agents having such functions can be used for diseases or conditions associated with oxidative stress (eg, neurodegenerative diseases, ischemic cerebrovascular disorders, ischemic heart diseases, inflammatory bowel diseases, ocular diseases and / or rheumatoid arthritis). Etc.) can be used to prevent and / or treat. Here, regarding the diseases or pathologies associated with oxidative stress exemplified above, examples of neurodegenerative diseases include Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, polyglutamine disease, and prion disease. Examples of ischemic cerebrovascular disorders include cerebral infarction and transient cerebral ischemic attack. Furthermore, examples of the ischemic heart disease include angina pectoris and myocardial infarction. Examples of inflammatory bowel disease include ulcerative colitis and Crohn's disease. Examples of eye diseases include retinopathy, cataract, glaucoma, age-related macular degeneration, and the like.

  The antioxidant stress agent by one form of this invention contains the compound 1 and / or compound 2 which were mentioned above as an active ingredient. Of course, both of these may be included as active ingredients. Here, “containing as an active ingredient” means that the antioxidant stress agent according to the present invention is a sufficient amount (that is, an effective amount) of Compound 1 and / or Compound 2 to exert a desired antioxidant stress effect. Is contained.

  Therefore, Compound 1 and / or Compound 2 may be used as an antioxidant stress agent as they are, but as long as they contain an active ingredient in such an effective amount and do not impair the antioxidant stress effect, The oxidative stress agent may constitute a composition together with a pharmaceutically acceptable carrier according to a desired product form, other additives, and the like. Further, the antioxidant stress agent according to the present invention is suitable for parenteral administration, oral administration or external administration by mixing with additives such as excipients, in addition to pharmaceutical compositions such as pharmaceuticals and quasi drugs, It can be used in the form of food, cosmetics, etc. For example, according to one preferred embodiment of the present invention, there is provided an external skin composition comprising an antioxidant stress agent comprising the above-mentioned compound 1 and / or compound 2 as active ingredients and a pharmaceutically acceptable carrier. Provided. When the skin external preparation composition is a pharmaceutical product, it can be used for the prevention and / or treatment of a disease or condition associated with oxidative stress in skin tissue. Moreover, when the said skin external preparation composition is cosmetics, it can be used for prevention and / or improvement of the disease or pathological condition which the oxidative stress in a skin tissue is related.

  When the antioxidant stress agent of the present invention is used as a pharmaceutical, the dosage varies greatly depending on the age, symptoms, etc. of the patient, but generally, in the case of oral administration, it is in the range of 1 to 2000 mg / day as a dry weight. It is desirable. When mix | blending with a foodstuff, it is preferable to set it as the density | concentration range of 0.01-50 mass% in view of the effect, the scent at the time of addition, and the point of color tone.

  For use in medicine, a therapeutically effective amount of the above-described compound (Compound 1 and / or Compound 2) is formulated with one or more pharmaceutically acceptable carriers (additives) and / or diluents. As described in detail below, the pharmaceutical composition according to the present invention can be specifically formulated for administration in solid or liquid form. Examples of oral administration include liquid medicine (aqueous solution or non-aqueous solution or suspension), tablet, bolus, powder medicine, granule, paste for application to the tongue. For parenteral administration, for example, as a sterile solution or suspension, for example, a formulation for subcutaneous, intramuscular or intravenous injection, or a dosage form for topical use or administration into the vagina or rectum And can be formulated. In particular, when it is used as an external preparation for skin, its form is not particularly limited. Examples of pharmaceuticals include forms such as sprays, ointments, pastes, creams, lotions, gels, solutions, and patches. For cosmetics, lotion, cosmetic emulsion, cosmetic cream, cosmetic gel, cosmetic liquid, pack, foundation, lipstick, lip balm, lip gloss, facial cleanser, body soap, hand cream, shampoo, rinse Examples of the form of skin care products such as hair styling products or makeup products are illustrated. Of these, lotions, cosmetic gels, cosmetic emulsions, cosmetic creams, and cosmetics are preferred, and lotions, cosmetic emulsions, and cosmetic creams are more preferred because they can be applied to a wide range of sites.

  “Therapeutically effective amount” means the amount of an agent or composition effective to produce any desired therapeutic effect at a reasonable benefit / risk ratio applicable to any medical treatment. For example, the dosage of the antioxidant stress agent according to the present invention varies depending on the target disease, administration subject, administration route, etc., but when the antioxidant stress agent of the present invention is administered orally for the purpose of treating Alzheimer's disease, Since it can easily vary depending on conditions such as the weight of the subject person, it can be appropriately selected by those skilled in the art.

  “Pharmaceutically acceptable” means that within the scope of good medical judgment, commensurate with a reasonable benefit / risk ratio, and without problems and complications such as excessive toxicity, irritation, and allergic reactions, in humans and animals. Used to refer to compounds, materials, compositions, and / or dosage forms suitable for use in contact with tissue.

  “Pharmaceutically acceptable carrier” refers to a liquid or solid filler involved in carrying or transporting the antioxidant stress agent of the present invention from one organ or part of the body to another organ or part of the body. Means a pharmaceutically acceptable material, composition or excipient, such as a diluent, excipient, solvent or encapsulating material. Each carrier must be “acceptable” in the sense of being compatible with the other ingredients of the dosage form and not injurious to the patient. Some of the materials that can function as pharmaceutically acceptable carriers are exemplified below: sugars such as lactose, glucose and sucrose; starches such as corn starch and potato starch; such as sodium carboxymethylcellulose, ethylcellulose and cellulose acetate. Cellulose and its derivatives; powder tragacanth; malt; gelatin; talc; excipients such as cocoa butter and suppository wax; Glycols such as glycol; polyols such as glycerin, sorbitol, mannitol and polyethylene glycol; esters such as ethyl oleate and ethyl laurate; agar; Buffering agents such as magnesium and aluminum hydroxide; including and compatible substances other non-toxic to be used in the drug formulation; alginic acid; pyrogen-free water; isotonic saline; Ringer's solution; ethyl alcohol; phosphate buffer solution. In some embodiments, the drug formulation is non-pyrogenic. That is, those that do not increase the patient's body temperature are preferred.

  In addition, wetting agents such as sodium lauryl sulfate and magnesium stearate, emulsifiers and lubricants, and colorants, release agents, coatings, sweeteners, flavors and fragrances, preservatives and antioxidants are also included in the composition. May be present.

  Examples of pharmaceutically acceptable antioxidants include: water-soluble antioxidants such as ascorbic acid, cysteine hydrochloride, sodium bisulfate, sodium disulfite, sodium sulfite, etc .; ascorbyl palmitate, butylhydroxy Oil-soluble antioxidants such as anisole (BHA), butylhydroxytoluene (BHT), lecithin, propyl gallate, α-tocopherol and the like; and citric acid, ethylenediaminetetraacetic acid (EDTA), sorbitol, tartaric acid, phosphoric acid and the like Such a metal chelating agent can also be contained as needed.

  Suitable dosage forms of the invention for oral administration may be in the form of capsules, cachets, pills, tablets, lozenges (with seasoned active ingredients, usually sucrose and gum arabic or tragacanth), powders, granules, or As a solution or suspension in an aqueous or non-aqueous liquid, or as an oil-in-water or water-in-oil liquid emulsion, or as an elixir or syrup, or pastilles (gelatin and glycerin, or inert groups such as sucrose and gum arabic) And / or a gargle and the like, each containing a predetermined amount of the compound of the present invention as an active ingredient. The agent of the present invention may be administered as a bolus, electuary or paste.

  In solid dosage forms of the invention for oral administration (capsules, tablets, pills, dragees, powders, granules, etc.), the active ingredient is one or more pharmaceuticals such as sodium citrate or dicalcium phosphate. A top acceptable carrier and / or mixed with any of the following: fillers or extenders such as starch, lactose, sucrose, glucose, mannitol, and / or silicic acid; eg carboxymethylcellulose, alginate, gelatin , Polyvinylpyrrolidone, binders such as sucrose and / or gum arabic; humectants such as glycerol; disintegration such as agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate Agents; dissolution retardants such as paraffin; 4 Absorption enhancers such as ammonium compounds; wetting agents such as cetyl alcohol and glycerol monostearate; absorbents such as kaolin and bentonite clay; talc, calcium stearate, magnesium stearate, solid polyethylene glycol, sodium lauryl sulfate, and Lubricants such as mixtures thereof; and colorants. In the case of capsules, tablets and pills, the drug composition may comprise a buffer. Similar types of solid compositions can also be used as fillers in soft and hard-filled gelatin capsules with excipients such as lactose or lactose and high molecular weight polyethylene glycols and the like.

  A tablet may be made by compression or molding, optionally with one or more accessory ingredients. Compressed tablets can be binders (eg, gelatin or hydroxypropylmethylcellulose), lubricants, inert diluents, preservatives, disintegrants (eg, sodium glycolate starch or crosslinked sodium carboxymethylcellulose), surfactants Alternatively, it can be prepared using a dispersant. Molded tablets can be made by molding in a suitable machine a mixture of the powdered compound moistened with an inert liquid diluent.

  Solid dosage forms such as tablets of the pharmaceutical composition according to the invention, such as sugar-coated tablets, capsules, pills and granules, are optionally scored or coated with coatings such as enteric coatings well known in the pharmaceutical dispensing arts and It may be prepared using a shell. They use, for example, hydroxypropyl methylcellulose, other polymer matrices, liposomes and / or microspheres in various ratios to provide the desired release profile, and slow or controlled release of the internal active ingredient. May be formulated to provide. They may be sterilized, for example, by filtration through a bacteria retaining filter, or by incorporating a sterilant in the form of a sterile solid composition that can be dissolved in a sterile injectable medium such as sterile water immediately before use. . These compositions may optionally contain opacifiers, in compositions that release one or more active ingredients only in certain parts of the gastrointestinal tract or preferentially there, optionally in a delayed manner. There may be. Examples of embedding compositions that can be used are polymeric substances and waxes. The active ingredient may be in microencapsulated form, if appropriate, with one or more of the above-described excipients.

  Liquid dosage forms for oral administration of the compounds of the invention include pharmaceutically acceptable emulsions, microemulsions, solutions, suspensions, syrups and elixirs. Liquid dosage forms include, in addition to the active ingredient, inert diluents commonly used in the art, such as water and other solvents, ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzoic acid Like fatty acid esters of benzyl, propylene glycol, 1,3-butadiene glycol, oils (especially cottonseed oil, peanut oil, corn oil, embryo oil, olive oil, castor oil and sesame oil), glycerol, tetrahydrofuryl alcohol, polyethylene glycol and sorbitan Solubilizers and emulsifiers, and mixtures thereof.

  In addition to inert diluents, oral compositions may include adjuvants such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, coloring, flavoring and preserving agents.

  Suspensions may be suspended in addition to the active compound, for example, ethoxylated isostearyl alcohol, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar and tragacanth, and mixtures thereof. A turbidity agent may be included.

  Dosage forms of the drug composition of the present invention for rectal or vaginal administration may be presented as a suppository. This suppository is a mixture of one or more suitable non-irritating excipients or carriers including, for example, cocoa butter, polyethylene glycol, suppository wax or salicylate, and one or more agents of the present invention. Which is solid at room temperature but liquid at body temperature, it will melt in the rectum or vaginal cavity and release the active compound.

  The dosage forms of the present invention suitable for vaginal administration also include pessaries, tampons, creams, gels, pastes, foams or spray dosage forms containing carriers as known to be suitable in the art.

  Dosage forms for topical or transdermal administration of one or more antioxidant stress agents of the present invention include powders, sprays, ointments, pastes, creams, lotions, gels, solutions, patches and inhalants. The active agent may be mixed under sterile conditions with a pharmaceutically acceptable base material and, if necessary, preservatives, buffers, or propellants.

  Ointments, pastes, creams and gels, in addition to the active compounds according to the invention, contain animal or vegetable fats, oils, waxes, paraffins, starches, tragacanth, cellulose derivatives, polyethylene glycols, silicones, bentonite, silicic acid, talc and oxidation An excipient may be included such as zinc, or a mixture thereof.

  Powders and sprays may contain, in addition to the compounds of this invention, excipients such as lactose, talc, silicic acid, aluminum hydroxide, calcium silicates and polyamide powder, or mixtures of these substances. The spray may further comprise customary high pressure gases such as chlorofluorinated hydrocarbons and volatile unsubstituted hydrocarbons such as butane and propane.

  Transdermal patches have the further advantage of controlled delivery of the antioxidant stress agent of the present invention to the body. Such dosage forms can be made by dissolving or dispersing the antioxidant stress agent of the present invention in a suitable medium. Absorption enhancers can also be used to increase the flux of substances containing the antioxidant stress agent of the present invention across the skin. The speed of such flux can be controlled by either providing a speed control membrane or by dispersing the compound in a polymer matrix or gel.

  A pharmaceutical composition having an antioxidant stress agent of the present invention suitable for parenteral administration comprises one or more pharmaceutically acceptable sterile isotonic or non-aqueous solutions together with one or more active compounds of the present invention, Dispersants, suspensions or emulsions, or sterile powders that can be reconstituted in a sterile injectable solution or dispersion just prior to use, which contain antioxidants, buffers, bacteriostats, preparations for the intended recipient Solutes that are isotonic with blood, or suspensions or concentrates.

  Examples of suitable aqueous and non-aqueous carriers that can be used in the pharmaceutical composition having the antioxidant stress agent of the present invention include water, ethanol, polyol (eg, glycerol, propylene glycol, polyethylene glycol, etc.), and suitable Mixtures, vegetable oils such as olive oil, and injectable organic esters such as ethyl oleate. The inherent fluidity can be maintained, for example, by the use of a coating material such as lecithin, by the maintenance of the required particle size in the case of dispersants and by the use of surfactants.

  These compositions may contain adjuvants such as preservatives, wetting agents, emulsifying agents and dispersing agents. Prevention of the activity of microorganisms can be ensured by the inclusion of various antibacterial and antifungal agents such as parabens, chlorobutanol, phenol sorbate and the like. It is preferable to include isotonic agents such as sugar and sodium chloride in the composition. In addition, prolonged absorption of the injectable drug form can be brought about by the inclusion of agents that delay absorption such as aluminum monostearate and gelatin.

  According to this invention, the food / beverage products containing the antioxidant stress agent mentioned above are also provided. Here, it is preferable that the said food / beverage products contain the said antioxidant stress agent in an appropriate quantity so that the effective component of the antioxidant stress agent may be included. Here, “including an effective amount of an active ingredient” means that the active ingredient is contained in such an amount that an effect as an active ingredient can be exhibited as a result of ingesting an amount of each food or drink normally consumed. You may mix | blend the active ingredient which concerns on this invention with the food / beverage products which concern on this invention as it is or in the form of the antioxidant stress agent as mentioned above. In addition, the food and drink according to the present invention are prepared as food and drink by adding conventional additives such as stabilizers to the active ingredient according to the present invention, various proteins, sugars, fats, trace elements, vitamins, etc. It may be prepared by further blending them, liquid, semi-liquid or solid, paste, or a general food or drink with active ingredients added.

  In the present invention, the “food or drink” is not a drug and is not particularly limited as long as it is in a form that can be taken orally by mammals, and the form is also a liquid (solution, suspension, emulsion). Liquid, etc.), semi-liquid, powder, or solid molded product. Therefore, the food and drink may be in the form of a beverage, for example, or may be in the form of a dietary supplement tablet such as a supplement.

  Specific examples of food and drink include instant noodles, retort foods, canned foods, microwave foods, instant soups and miso soups, freeze-dried foods, etc .; soft drinks, fruit juice drinks, vegetable drinks, soy milk drinks, coffee Beverages such as beverages, tea beverages, powdered beverages, concentrated beverages, nutritional beverages, alcoholic beverages; flour products such as bread, pasta, noodles, cake mixes, fried flour, bread crumbs; rice cakes, caramels, chewing gums, chocolates, cookies, Confectionery such as biscuits, cakes, pies, snacks, crackers, Japanese confectionery, dessert confectionery; sauces, processed tomato seasonings, flavor seasonings, cooking mixes, sauces, dressings, soups, curry and stew Seasonings; processed fats and oils, butter, margarine, mayonnaise, etc .; milk drinks, yogurts, lactic acid bacteria drinks Dairy products such as ice cream and cream; processed fishery products such as fish ham and sausages and fish paste products; processed livestock products such as livestock ham and sausages; canned agricultural products, jams and marmalades, pickles, boiled beans, cereals, etc. Processed agricultural products; frozen foods; nutritional foods.

  The food and drink according to the present invention is suitable for a person suffering from or suspected of suffering from a disease associated with oxidative stress as described above, or a person having a high risk of suffering from the disease. Can be used for Here, as a person having a high risk of suffering from the disease, for example, taking into account various indicators including body composition and eating habits, or from a diagnosis / diagnosis such as a health checkup, the risk is high And those who have been perceived by the person or others as having such a high risk.

  In the present invention, “food and beverage” includes foods classified as health foods, functional foods, foods for specified health use, dietary supplements, foods with a disease risk reduction label, or foods for the sick. Is done. Furthermore, the term “food or drink” can be used to include feed when used for mammals other than humans. The food for specified health here refers to any legal restrictions in each country (for example, Japan) from the viewpoint of health when manufacturing or selling food for the purpose of prevention and / or improvement of hypertension. It means food that may be received. Such a food may be a food that indicates that the food may reduce the risk of disease, that is, a food with a disease risk reduction label. Here, the disease risk reduction label is a label for foods that may reduce the disease risk, and is based on or based on the standard established by the FAO / WHO Joint Food Standards Committee (Codex Committee). The display may be a prescribed display or an approved display with reference to FIG.

  In the food / beverage products of this invention, in addition to the active ingredient mentioned above, you may further add the component which has another function. Further, for example, the active ingredient of the present invention is added to foods, health foods, functional foods and supplements (for example, foods containing one or more vitamins such as minerals such as calcium and magnesium and vitamin K) consumed in daily life. By mix | blending, in addition to the effect by this invention, the food / beverage products which have the function based on another component can be provided.

  According to another aspect of the present invention, there is provided a food or drink containing the above-described antioxidant stress agent, which is a neurodegenerative disease, ischemic cerebrovascular disorder, ischemic heart disease, inflammatory bowel disease, and / or eye disease. There is provided a food or drink having a function of preventing and / or improving odors and having the function indication. Here, the function display attached to the food or drink can be attached to, for example, any of the main body of the product, the container, the packaging, the instruction, the attached document, or the promotional material.

  In the production of foods and drinks according to the present invention, sugars, fragrances, fruit juices, food additives, stabilizers and the like that are used in the usual food and beverage product design can be added as appropriate. Manufacture of food and drink products can be carried out with reference to manufacturing techniques known in the art. The food / beverage products according to the present invention can take various forms, and the food / beverage products according to the present invention may be manufactured according to known pharmaceutical manufacturing techniques. In that case, it can be produced using a carrier or additive as described in the item of the production of the antioxidant stress agent or pharmaceutical composition according to the present invention. Moreover, in a manufacturing stage, it is good also as multifunctional food / beverage products by combining with the other component which exhibits functions other than the function in this invention, or another functional food.

  When administering or ingesting the pharmaceutical composition and food or drink according to the present invention, the dose or intake of the active ingredient according to the present invention is the recipient, the age and weight of the recipient, symptoms, administration time, dosage form, administration It can be determined depending on the method, the combination of drugs and the like. In the present invention, the dose or intake of the composition or food / drink per day is taken into consideration so that at least the amount of the active ingredient per day necessary for obtaining the antioxidant stress effect can be administered or taken. It is preferable to appropriately set the content in the composition or food or drink.

  Therefore, the pharmaceutical composition or food or drink according to the present invention preferably contains Compound 1 and / or Compound 2 as active ingredients, preferably 30 to 6000 mg per day per adult in terms of the active ingredient, more preferably 50 -2000 mg, more preferably in an amount provided in the range of 100-1000 mg.

  Hereinafter, preferred embodiments of the present invention will be described in more detail by way of examples. However, the technical scope of the present invention should not be construed as being limited to the following examples.

<< Extraction and Purification of Compound 1 and Compound 2 from Artichoke >>
To 600 g of the edible portion of dried artichoke, 2300 mL of ethyl acetate was added and extracted at 80 ° C. for 2 hours. After the extract was filtered, the solvent was removed under reduced pressure to obtain 17.5 g of an artichoke ethyl acetate extract. Similar extraction was performed again to obtain a total of 32.3 g of extract. After dissolving 32.3 g of the obtained extract in 10 mL of ethyl acetate, 30 g of silica gel was added and dried under reduced pressure with an evaporator to prepare a sample layer. 700 g of silica gel was packed in a chromatographic tube, and the previously prepared sample layer was laminated to prepare dry column chromatography. Elution was started with a hexane: ethyl acetate mixed solvent (mixing ratio, 2: 3), and 1400 mL of the eluate was collected per fraction and separated into four fractions. Subsequently, the second fraction was sequentially subjected to dry column chromatography using hexane: ethyl acetate: acetic acid mixed solvent (mixing ratio, 80: 20: 1) and hexane: acetone: methanol (mixing ratio, 40: 8: 1). Fractionation gave 300 mg of crude fraction. Subsequently, about 300 mg of the obtained crude fraction was dissolved in 2.0 mL of dichloromethane, and then 2.0 mL of acetonitrile was added. The solution was filtered through a 0.45 μm membrane filter and subjected to the following separation operation by HPLC.

(HPLC condition 1)
Column: Capcell Pak C ₁₈ column (UG80, 250 × 10 mm id, Shiseido Co., Ltd.)
Solvent: acetonitrile-0.1% formic acid 70:30
Flow rate: 4.0 mL / min
Temperature: 40 ° C
Detection wavelength: 254 nm
Load amount: 0.4 mg
The chromatogram obtained under these conditions is shown in FIG. As shown in FIG. 1, three main peaks were observed in the crude fraction. These are hereinafter referred to as peak 1 (Rt 10.9), peak 2 (Rt 11.8) and peak 3 (Rt 12.5), respectively.

  Subsequently, as a result of the fractionation operation for the three main peaks, each peak was purified (peak 1 45 mg; peak 2 22.9 mg; peak 3 22.1 mg).

  Furthermore, the peak 1 obtained above was purified by subjecting it to the following separation operation by HPLC.

(HPLC condition 2)
Column: Cosmosil cholester (250 × 10 mm id, Nacalai Tesque)
Solvent: methanol-acetonitrile-0.1% formic acid 65:12:23
Flow rate: 3.5 mL / min
Temperature: 40 ° C
Detection wavelength: 254 nm
Load: 1.0 mg
The chromatogram obtained under these conditions is shown in FIG. As is clear from FIG. 2, peak 1 was found to be a mixture, and two main peaks were observed. Hereinafter, these are referred to as peak 1-1 (Rt 19.8) and peak 1-2 (Rt 22.1), respectively.

  Then, as a result of performing fractionation operation about said peak 1-1 and peak 1-2, it came to the refinement | purification of each peak (peak 1-1 19.5 mg; peak 1-2 17.2 mg).

≪ARE enhancer activity of isolated compound (reporter assay) ≫
An ARE reporter gene in which an ARE (antioxidant response element) is placed upstream of a thymidine kinase gene promoter that controls the expression of the firefly luciferase gene, and a control gene in which the expression of the Renilla luciferase gene is controlled by the thymidine kinase gene promoter, Using TransIT-LT1 (manufactured by Mirus Bio), it was introduced into rat pheochromocytoma PC12 cells, which is one type of model cell of neuronal cells.

  For the culture of PC12 cells, DMEM medium supplemented with 5% fetal calf serum and 10% horse serum was used. Cells into which the ARE reporter gene and control gene have been introduced are seeded in a 24-well culture plate and cultured for 24 hours, and then the test sample (the compound of peak 1-2 or the compound of peak 3 isolated above) having a final concentration of 20 μM. Added in concentration. As a control, only the solvent DMSO was added. After 24 hours, ARE enhancer activity was evaluated as a value of firefly luciferase activity / Renilla luciferase activity by measuring the activity of luciferase using Dual Luciferase Assay System (Promega) and a luminometer. The results are shown in FIG.

  From the results shown in FIG. 3, it can be seen that both the compound of peak 1-2 (compound 1) and the compound of peak 3 (compound 2) isolated above show excellent ARE enhancer activity.

≪Identification of compounds by instrumental analysis≫
The compound of peak 1-2 (compound 1) and the compound of peak 3 (compound 2) that showed excellent ARE enhancer activity were identified by instrumental analysis as follows.

<< Effect of isolated compound on expression of HO-1 protein (Western blot) >>
PC12 cells were cultured in DMEM medium supplemented with 5% fetal bovine serum and 10% horse serum, and human normal dermal fibroblasts (HDF) were cultured in DMEM medium supplemented with 10% fetal bovine serum and seeded in 12-well plates. . After 24 hours, test samples (Compound 1 or Compound 2) were treated at a final concentration of 20 μM. As a control, it was treated only with DMSO as a solvent. The cells were added with RIPA buffer (10 mM Tris-HCl, pH 7.5, 1% NonidetP-40, 0.1% Sodium deoxycholate, 0.1% SDS, 150 mM NaCl, 1 mM EDTA) supplemented with protease inhibitor cocktail (Sigma). After lysis and centrifugation at 15,000 × g, 4 ° C. for 15 minutes, the supernatant was used as a cell lysis protein sample. Each 10 mg of this protein sample was developed by 10% SDS polyacrylamide gel electrophoresis according to a conventional method, transferred from the gel to a PVDF membrane using a semi-dry type transfer device, and subjected to Western blotting. As a washing solution, 0.1% Tween 20-containing Tris buffer (TBS-T) is used, as a blocking solution, a 5% skim milk TBS-T solution is used, and as a primary antibody, a rabbit-derived anti-HO-1 polyclonal antibody (manufactured by Stressgen) ) Was used at a 1,000-fold dilution, and a sheep-derived anti-mouse IgG antibody (HRP labeled, manufactured by GE Healthcare) was used as a secondary antibody at a 3000-fold dilution. Detection was performed by a chemiluminescence method using an ECL Plus Western Blotting Detection System (GE Healthcare). The results are shown in FIG. “PC12” shown in FIG. 4 is a result showing the expression of HO-1 protein in PC12 cells, and “HDF” is a result showing the expression of HO-1 protein in human normal dermal fibroblasts (HDF).

  From the results shown in FIG. 4, Compound 1 and Compound 2 showed antioxidant protein activity in a wide range of cells including at least PC12 cells and human normal dermal fibroblasts (HDF) via the ARE enhancer activity shown above. It can be seen that the expression of one type of HO-1 protein is enhanced.

  From the above, it was proved that the antioxidant stress agent according to the present invention exhibits an excellent antioxidant stress action by containing Compound 1 or Compound 2 as an active ingredient. The antioxidant stress agent according to the present invention can prevent and / or prevent neurodegenerative diseases, ischemic cerebrovascular disorders, ischemic heart diseases, inflammatory bowel diseases, eye diseases and / or rheumatoid arthritis related to oxidative stress. Or it can be used for treatment and has a very high advantage.

Claims (8)

Compound 1 and / or Compound 2 below:
Antioxidant stress agent containing as an active ingredient.   The antioxidant stress agent according to claim 1, wherein the action expression of the antioxidant stress agent is an action of enhancing the expression of the antioxidant protein.   A pharmaceutical composition comprising the antioxidant stress agent according to claim 1 or 2 and a pharmaceutically acceptable carrier, which is used for prevention and / or treatment of a disease or condition associated with oxidative stress.   A skin external preparation composition comprising the antioxidant stress agent according to claim 1 or 2 and a pharmaceutically acceptable carrier.   Cosmetics comprising the skin external preparation composition according to claim 4.   Food / beverage products containing the antioxidant stress agent of Claim 1 or 2. A food or drink containing an effective amount of the antioxidant stress agent according to claim 1 or 2,
A food and drink having a function of preventing and / or improving neurodegenerative diseases, ischemic cerebrovascular disorders, ischemic heart diseases, inflammatory bowel diseases, and / or eye diseases and provided with function indications.   The food or drink according to claim 7, which is a health food, a functional food, a food for specified health use, a dietary supplement, a food with a disease risk reduction label, or a food for a sick person.