WO2021015233A1 - エラスターゼ阻害作用を有するローヤルゼリー - Google Patents
エラスターゼ阻害作用を有するローヤルゼリー Download PDFInfo
- Publication number
- WO2021015233A1 WO2021015233A1 PCT/JP2020/028419 JP2020028419W WO2021015233A1 WO 2021015233 A1 WO2021015233 A1 WO 2021015233A1 JP 2020028419 W JP2020028419 W JP 2020028419W WO 2021015233 A1 WO2021015233 A1 WO 2021015233A1
- Authority
- WO
- WIPO (PCT)
- Prior art keywords
- royal jelly
- elastase
- bacillus subtilis
- derived
- proteolytic enzyme
- Prior art date
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- 229960002675 xylitol Drugs 0.000 description 1
- 235000010447 xylitol Nutrition 0.000 description 1
- HEBKCHPVOIAQTA-SCDXWVJYSA-N xylitol Chemical compound OC[C@H](O)[C@@H](O)[C@H](O)CO HEBKCHPVOIAQTA-SCDXWVJYSA-N 0.000 description 1
- 239000011701 zinc Substances 0.000 description 1
- 229910052725 zinc Inorganic materials 0.000 description 1
- UHVMMEOXYDMDKI-JKYCWFKZSA-L zinc;1-(5-cyanopyridin-2-yl)-3-[(1s,2s)-2-(6-fluoro-2-hydroxy-3-propanoylphenyl)cyclopropyl]urea;diacetate Chemical compound [Zn+2].CC([O-])=O.CC([O-])=O.CCC(=O)C1=CC=C(F)C([C@H]2[C@H](C2)NC(=O)NC=2N=CC(=CC=2)C#N)=C1O UHVMMEOXYDMDKI-JKYCWFKZSA-L 0.000 description 1
Classifications
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES, NOT OTHERWISE PROVIDED FOR; PREPARATION OR TREATMENT THEREOF
- A23L21/00—Marmalades, jams, jellies or the like; Products from apiculture; Preparation or treatment thereof
- A23L21/20—Products from apiculture, e.g. royal jelly or pollen; Substitutes therefor
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES, NOT OTHERWISE PROVIDED FOR; PREPARATION OR TREATMENT THEREOF
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES, NOT OTHERWISE PROVIDED FOR; PREPARATION OR TREATMENT THEREOF
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
- A23L33/17—Amino acids, peptides or proteins
- A23L33/18—Peptides; Protein hydrolysates
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/99—Enzyme inactivation by chemical treatment
Definitions
- the present invention relates to a royal jelly food composition containing royal jelly having an elastase inhibitory effect.
- Royal jelly is a milky white paste-like substance secreted by the pharyngeal glands of young worker bees, and its components are rich in amino acids including essential amino acids and are composed of high-quality proteins.
- vitamins include vitamin B1, vitamin B2, vitamin B6, niacin, pantothenic acid, which is effective in promoting growth and preventing aging, vitamin A, vitamin C, and vitamin E.
- minerals include potassium, magnesium, calcium, copper, iron and phosphorus.
- carbohydrates include glucose and fructose.
- royal jelly contains various nutritional components, and is known to have many effects such as antibacterial action, immunopotentiating action, anti-inflammatory action, antiaging action, prevention / therapeutic action of menopausal disorders, and anticancer action. ing.
- Elastin is distributed in the dermis of the skin in the living body. Elastin plays a role in giving elasticity to tissues, and in the skin, it plays a major role in maintaining elasticity or firmness. It is known that the amount of elastin gradually decreases due to aging, ultraviolet rays, active oxygen, stress, and the like. In the skin, elastin depletion has been shown to gradually reduce the elasticity of the skin, causing wrinkles or sagging, which is a major contributor to aging.
- fibroblast-derived elastase is known as an elastase involved in the decomposition of elastin present in the dermis matrix of the skin, and elastic fibers involved in maintaining the elasticity of the skin are used. It is known to break down the constituent elastin. Therefore, it is expected that a high anti-wrinkle effect can be obtained by inhibiting its activity.
- an elastase inhibitor for example, an elastase inhibitor characterized by containing an extract of a flower of pomegranate (Punica granatum) (Patent Document 1) or an extract of Saxifraga stolonifera Meerburg (Patent Document 2) as an active ingredient.
- the agent is disclosed.
- An object of the present invention is to provide a new technique capable of obtaining royal jelly having an elastase inhibitory action.
- the present invention It comprises a food composition for elastase inhibition containing enzyme-treated royal jelly obtained by a step of reacting a proteolytic enzyme derived from Bacillus subtilis as an active ingredient.
- a royal jelly having an elastase inhibitory action can be produced by a safer and simpler method, and a royal jelly composition having this elastase inhibitory action can be provided.
- the royal jelly having an elastase inhibitory action is produced by allowing a proteolytic enzyme derived from Bacillus subtilis to act on the royal jelly.
- the country of origin of royal jelly can be, for example, Japan, China, Taiwan, Thailand, Brazil, European countries, Oceania countries, the United States, etc., and royal jelly from any country of origin may be used.
- royal jelly from a plurality of countries of origin may be appropriately mixed and used.
- the royal jelly is preferably a liquid, and when the royal jelly in a freeze-dried state is used, it can be dissolved in purified water, tap water, an appropriate buffer solution, or the like before use. In addition, frozen royal jelly can be thawed and used.
- the royal jelly may be processed by heating, centrifugation, alcohol extraction, filtration, freeze-drying or hot-air drying, and added with various nutrients.
- the proteolytic enzyme derived from Bacillus subtilis that acts on royal jelly is obtained by culturing Bacillus subtilis, and may be a culture solution or a refined product. Further, the proteolytic enzyme derived from Bacillus subtilis may be obtained by gene recombination, or may be modified with, for example, sugar or polyethylene glycol.
- the solid content concentration of royal jelly after being mixed with a proteolytic enzyme derived from Bacillus subtilis can be 0.1 to 30 W / W%, preferably 3 to 10 W / W%.
- 10 W / W% means that when 100 g of royal jelly is dried by freeze-drying or the like, 10 g of a dried product can be obtained.
- the conditions for treating royal jelly with a Bacillus subtilis-derived proteolytic enzyme are not particularly limited as long as the desired elastase inhibitor is produced, and the stability of royal jelly and It can be appropriately set in consideration of the stability and reactivity of the proteolytic enzyme derived from Bacillus subtilis.
- the time for allowing the proteolytic enzyme derived from Bacillus subtilis to act on the royal jelly is not particularly limited as long as the target elastase inhibitor is produced, but it is preferably several hours to one day, preferably 3 hours to 10 hours. Prolonged reactions had little effect on the production of elastase inhibitors.
- the pH at which the proteolytic enzyme derived from Bacillus subtilis is allowed to act on royal jelly is not particularly limited as long as it produces the target elastase inhibitor, but pH 3 to 11, preferably pH 5 to 11, is appropriate.
- the temperature at which the proteolytic enzyme derived from Bacillus subtilis acts on the royal jelly is not particularly limited as long as the target elastase inhibitor is produced, but is 30 ° C to 80 ° C, preferably 40 ° C to 60 ° C. Appropriate.
- Bacillus subtilis-derived proteolytic enzyme While the Bacillus subtilis-derived proteolytic enzyme is reacted with royal jelly, it may be allowed to stand, and it may be shaken or stirred.
- a proteolytic enzyme may act, and those derived from microorganisms, animals, or plants are exemplified.
- Microbial-derived proteolytic enzymes are obtained by microbial culture, and plant- or animal-derived proteolytic enzymes are obtained by homogenizing plant or animal organs.
- These proteolytic enzymes may be those that have been further purified, or those that have been modified with polyethylene glycol or the like. Further, two or more kinds of proteolytic enzymes may be used in combination.
- the production of royal jelly having an elastase inhibitory effect generally includes a step of inactivating the enzyme after treatment of the royal jelly with a proteolytic enzyme derived from Bacillus subtilis.
- a proteolytic enzyme derived from Bacillus subtilis may be inactivated to the extent that there is no problem as a food.
- the method of inactivation include a method of inactivating by heating, a method of inactivating with a drug, and a method of removing the proteolytic enzyme derived from Bacillus subtilis by filtration, and these methods may be used alone or. Two or more types may be combined.
- heating inactivates the proteolytic enzyme derived from Bacillus subtilis.
- a heating temperature of 60 ° C. or higher, preferably 80 ° C. or higher.
- a food sterilization step it may also serve as a step of inactivating a proteolytic enzyme derived from Bacillus subtilis.
- Bacillus subtilis-derived proteolytic enzyme when allowed to act on the royal jelly, a stabilizer or reaction accelerator of the Bacillus subtilis-derived proteolytic enzyme may be added.
- the royal jelly produced by the action of a proteolytic enzyme derived from Bacillus subtilis is a royal jelly having an elastase inhibitory effect.
- the elastase inhibitor may be purified by treating the royal jelly having the elastase inhibitory action according to the present invention using various chromatographies.
- the purification method include gel filtration chromatography, ion exchange chromatography, affinity chromatography, hydrophobic chromatography, reverse phase chromatography, normal phase chromatography, ultrafiltration, and electrophoresis. These can be used for purification alone or in combination.
- a carrier for gel filtration chromatography capable of separating proteins of various molecular weights
- a carrier for gel filtration chromatography capable of separating proteins having a molecular weight of about 10,000 or less is preferable.
- the ion exchange group used in ion exchange chromatography include an anion exchanger and a cation exchanger.
- the anion exchanger include a diethylaminoethyl group (DEAE group) and a quaternary aminoethyl group (QAE group).
- a cation exchanger a carboxymethyl group (CM group) and a sulfopropyl group (SP group) can be exemplified.
- Examples of the carrier used for hydrophobic chromatography include a carrier to which a butyl group (Butyl group), an ethyl group (Ethyl group), and a phenyl group (Phenyl group) are bonded.
- Examples of the carrier used for the reverse phase chromatography include an octadecyl group (C18) and a carrier to which C30, C8, C4 and the like having an alkyl group length different from that of the octadecyl group are bonded.
- Examples of the carrier used for normal phase chromatography include silica gel, a carrier to which a cyanopropyl group, a functional group having a diol structure, an aminopropyl group, a polyamine, and the like are bonded.
- various components include foods, sugars, lipids, emulsifiers, thickeners, seasonings, flavors, acidity regulators, preservatives, fruit juices, flavors, various nutritional components, etc., as long as the effects of the present invention are not impaired. Can be used in. Further, various components may be used alone or in combination of two or more.
- examples of sugar include sucrose, high fructose corn syrup, glucose, fructose, palatinose, trehalose, lactose, and xylose.
- Examples of the emulsifier include sucrose fatty acid ester, glycerin fatty acid ester, and lecithin.
- Examples of the thickener include carrageenan, gum arabic, xanthan gum, guar gum, pectin, locust bean gum thickener starch, and gellan gum.
- Examples of the acidity regulator include citric acid, lactic acid, malic acid, fumaric acid, gluconic acid, tartaric acid and the like.
- Examples of the preservative include benzoic acid and its salt, sorbic acid and its salt, paraben, sodium sulfite, pectin decomposition product, glycine and the like.
- fruit juice examples include tomato juice, plum juice, apple juice, lemon juice, orange juice, and berry juice.
- fragrance examples include spices such as herbs and spices, fruit-based fragrances, and fragrances such as vanilla.
- other preferable nutritional components include vitamins such as vitamin D and minerals such as calcium, magnesium, iron, manganese, and zinc.
- the royal jelly composition according to the present invention can also be provided as food.
- Specific forms of such foods include, for example, beverages, confectionery, candy, gum, bread, livestock meat products, dairy products, retort foods, instant foods, frozen foods, jelly-like foods, bee products, pickles, seasonings. And so on.
- These foods are also useful as so-called health foods, functional foods, foods for specified health uses, nutritionally functional foods, dietary supplements, supplements and the like.
- examples of these food shapes include granules, powders, tablets, capsules, chewables, drinks, jellies, pastes, and grains.
- the royal jelly having an elastase inhibitory effect according to the present invention can be produced as a pharmaceutical composition containing a pharmaceutically acceptable carrier (medicinal carrier), if necessary.
- a pharmaceutical composition can be used as a hypoglycemic agent.
- the pharmaceutical carrier include fillers, bulking agents, binders, wetting agents, disintegrants, lubricants, diluents and excipients, which are usually used depending on the usage pattern of the preparation. These are appropriately selected and used according to the administration form of the obtained preparation.
- water pharmaceutically acceptable organic solvents, collagen, polyvinyl alcohol, polyvinylpyrrolidone, carboxyvinyl polymers, sodium alginate, water-soluble dextran, sodium carboxymethyl starch, pectin, xanthan gum, gum arabic, casein,
- examples thereof include gelatin, agar, glycerin, propylene glycol, polyethylene glycol, vaseline, paraffin, stearyl alcohol, stearic acid, human serum albumin, mannitol, sorbitol, and lactose. These may be used alone or in combination of two or more as appropriate depending on the dosage form of the target drug.
- stabilizers examples include human serum albumin, ordinary L-amino acids, sugars, and cellulose derivatives.
- L-amino acid is not particularly limited, and may be, for example, glycine, cysteine, glutamic acid, or the like.
- the saccharides are also not particularly limited, for example, monosaccharides such as glucose, mannose, galactose and fructose, sugar alcohols such as mannitol, inositol and xylitol, disaccharides such as sucrose, maltose and lactose, dextran, hydroxypropyl starch, chondroitin sulfate, etc. It may be any of polysaccharides such as hyaluronic acid and derivatives thereof.
- the cellulose derivative is not particularly limited, and may be any of methyl cellulose, ethyl cellulose, hydroxyethyl cellulose, hydroxypropyl cellulose, hydroxypropyl methyl cellulose, sodium carboxymethyl cellulose and the like.
- the surfactant is not particularly limited, and either an ionic surfactant or a nonionic surfactant can be used.
- Surfactants include, for example, polyoxyethylene glycol sorbitan alkyl ester type, polyoxyethylene alkyl ether type, sorbitan monoacyl ester type, fatty acid glyceride type and the like.
- Buffers include boric acid, phosphoric acid, acetic acid, citric acid, ⁇ -aminocaproic acid, glutamate and / or their corresponding salts (eg, alkali metal salts such as their sodium, potassium, calcium, magnesium salts and the like. Alkaline earth metal salt) can be exemplified.
- Examples of the tonicity agent include sodium chloride, potassium chloride, saccharides, and glycerin.
- the chelating agent include sodium edetate and citric acid.
- the royal jelly having an elastase-inhibiting effect according to the present invention is expected to obtain a high anti-wrinkle effect by inhibiting the activity of decomposing elastin constituting elastic fibers involved in maintaining the elasticity of the skin.
- the enzyme-treated royal jelly solution and the raw royal jelly solution obtained in Example 1 were examined for elastase inhibitory activity according to the method described below.
- (1) Acquisition of Elastase Crude Enzyme Solution The inhibition rate of elastase activity derived from human fibroblasts was tested as follows. As cells, normal human adult-derived skin fibroblasts (NHDF; Kurabo Industries Ltd.) were used. The growth medium was prepared using Kurabo Industries' FibroLife S2 Comp kit according to the instruction manual. The cells were seeded in T-75 flasks (TC-treated by Eppendorf Co., Ltd.) at a density of 2,500 cells / cm 2 , and cultured in a 5% CO 2 incubator at 37 ° C. for 24 hours, and the growth medium was changed.
- NHDF normal human adult-derived skin fibroblasts
- the growth medium was prepared using Kurabo Industries' FibroLife S2 Comp kit according to the instruction manual.
- the cells were seeded in T-75 flasks (TC-treated
- the substrate was prepared by dissolving the synthetic substrate succinyl-alanyl-alanyl-alanyl-p-nitroanilide (manufactured by Peptide Institute) in 0.1 M Tris-hydrochloric acid buffer (pH 8.0) so as to have a concentration of 1.5 mM.
- the elastase inhibition rate was calculated as follows.
- the absorbance value when 0.1 M Tris-hydrochloric acid buffer (pH 8.0) was added instead of the test substance was CA, and 0.1 M Tris-hydrochloric acid buffer (pH 8.0) was added instead of the test substance to replace the substrate.
- the absorbance when 0.1 M Tris-hydrochloric acid buffer (pH 8.0) was added was defined as CB.
- the absorbance value when the test substance is added is SA
- the absorbance when the test substance is added and 0.1 M Tris-hydrochloric acid buffer (pH 8.0) is added instead of the substrate is SB
- the enzyme-treated royal jelly solution had a higher elastase inhibitory activity than the raw royal jelly solution.
- royal jelly with high elastase inhibitory activity can be produced by allowing a proteolytic enzyme derived from Bacillus subtilis to act on royal jelly.
- the royal jelly composition containing royal jelly having an elastase inhibitory action produced by the present invention can be suitably used for foods and drinks, and can be particularly preferably used for beauty foods and the like.
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- Nutrition Science (AREA)
- Bioinformatics & Cheminformatics (AREA)
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- General Chemical & Material Sciences (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
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| JP2021534068A JPWO2021015233A1 (enrdf_load_stackoverflow) | 2019-07-23 | 2020-07-22 | |
| KR1020217039031A KR20220035328A (ko) | 2019-07-23 | 2020-07-22 | 엘라스타아제 저해작용을 갖는 로열젤리 |
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| JP (1) | JPWO2021015233A1 (enrdf_load_stackoverflow) |
| KR (1) | KR20220035328A (enrdf_load_stackoverflow) |
| WO (1) | WO2021015233A1 (enrdf_load_stackoverflow) |
Citations (8)
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| JPH0150882B2 (enrdf_load_stackoverflow) * | 1984-11-30 | 1989-11-01 | Asahi Optical Co Ltd | |
| JP2006061038A (ja) * | 2004-08-25 | 2006-03-09 | Api Co Ltd | 酵素処理ローヤルゼリー含有ドリンク剤 |
| JP2006273857A (ja) * | 2005-03-07 | 2006-10-12 | Access Business Group Internatl Llc | 皮膚の保湿性、肌理、及び外観を改善するための、植物を主成分とする調合物 |
| JP2007204418A (ja) * | 2006-02-01 | 2007-08-16 | Yamada Bee Farm Corp | 皮膚外用剤 |
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| JPH11246386A (ja) | 1998-03-03 | 1999-09-14 | Shiseido Co Ltd | エラスターゼ阻害剤 |
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| JP6059536B2 (ja) * | 2013-01-08 | 2017-01-11 | 森川健康堂株式会社 | コラゲナーゼ阻害作用を有するローヤルゼリーの製造方法 |
| JP6450882B1 (ja) * | 2018-06-11 | 2019-01-09 | アピ株式会社 | 水分蒸散量抑制用組成物 |
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2020
- 2020-07-22 KR KR1020217039031A patent/KR20220035328A/ko active Pending
- 2020-07-22 WO PCT/JP2020/028419 patent/WO2021015233A1/ja active Application Filing
- 2020-07-22 JP JP2021534068A patent/JPWO2021015233A1/ja active Pending
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| KR20220035328A (ko) | 2022-03-22 |
| JPWO2021015233A1 (enrdf_load_stackoverflow) | 2021-01-28 |
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