JP2020002027A - Composition for promoting hair growth - Google Patents

Abstract

To provide a composition for promoting hair growth.SOLUTION: A composition for promoting hair growth contains at least one selected from the group consisting of a compound represented by the following formula (I), a physiologically acceptable salt thereof and a physiologically acceptable hydrate thereof. In formula (I), R, R, Rand Rindependently represent hydrogen, a hydroxy group or -OCH, R, Rand Rindependently represent, hydrogen, -CHor -CHor -OCH).SELECTED DRAWING: Figure 5

Description

  The present invention relates to compositions for promoting hair growth.

  Hair extends from hair follicles present in the skin. The hair follicle has a complex structure, and at the bottom, cells called hair papilla cells, in which mesenchymal-derived fibroblasts are differentiated, are present. On the outside of the hair papilla cells, there are hair cells that have differentiated keratinocytes surrounding them. As the differentiation of keratin protein progresses, the synthesis of keratin protein progresses and the hair grows.

  Hair grows at a slower rate of elongation with age and may not grow. In addition, hair may not be able to grow due to factors such as stress and hormonal imbalance. The size of the hair growth market is increasing year by year, and various hair growth methods are being developed.

  Papaya (scientific name: Carica Papaa Linn) is widely used in various fields as a compounding ingredient in foods and pharmaceuticals because it contains not only edible tropical fruits but also a protein hydrolase known as papain. It has been. It has also been used topically on the skin as an external preparation for skin, such as cosmetics, burn treatment and chronic skin ulcers.

  It is suggested that the health functionality may be based on the ROS scavenging activity based on the antioxidant ability of the product, especially the hydroxyl radical scavenging activity (Non-Patent Document 1), and the effect on the skin may be based on the singlet oxygen scavenging activity. .

  Fermented papaya fruit products are known to be effective against various diseases due to the above antioxidant action. For example, it is suggested that it may be effective against diseases such as epilepsy, amnesia, Alzheimer's disease, and depression, may enhance immunity of skin and colon mucosa, and may suppress allergic inflammation.

In addition, the effectiveness against the skin aging due to the effects of ultraviolet rays, such as skin spots and wrinkles, is not limited to the hydroxyl radical scavenging effect of fermented papaya fruit products, but is especially improved by the ability to scavenge singlet oxygen. It is suggested that At that time, by eliminating active oxygen generated in the skin, the production of lipid peroxide is suppressed, and the moisturizing function of the stratum corneum at the top layer of the skin is maintained, so atopic dermatitis, dry skin, It is suggested that prevention and / or improvement can also be achieved with accompanying itching.
As an extension of these effects, cosmetic effects such as skin whitening and moist skin are also expected.

  The use of papaya fermentation products in foods (Patent Literatures 1 to 4), detergents and cleaning fields (Patent Literatures 5 to 8), soaps (Patent Literatures 9 and 10), and cosmetic agents for external use (Patent Literature 11) is considered. Has been put into practical use.

Neuroscience143 2006 P63-72

JP 2001-120224 A

JP 2003-24009 A

JP 2004-267026 A

JP 2006-75086 A

JP-A-7-82597

JP 2001-152188 A

JP 2001-181687 A

JP 2001-247896 A

JP-A-53-54207

JP-A-2004-123585

JP 2007-302664 A

  However, it was unknown which compound in the fermented papaya had antioxidant activity.

  Therefore, the present inventors have found that the compounds found in the process of isolating and purifying the compounds contained in the papaya fermented product have not only an antioxidant ability but also an unexpected effect of promoting hair growth. The present inventors have further studied and found that isopropyl 2,5-dihydroxybenzoate has an effect of promoting hair growth, and thus completed the present invention.

According to the present invention,
The following formula (I)
(In the formula (I),
R ₁ , R ₂ , R ₃ and R ₄ are each independently hydrogen, a hydroxyl group or -OCH ₃ ;
R _5, R ₆ and R ₇ are each independently hydrogen, -CH ₃ or -C ₂ H ₅ or -OCH ₃₎
A composition for promoting hair growth, comprising at least one selected from the group consisting of a compound represented by the formula, a physiologically acceptable salt thereof and a physiologically acceptable hydrate thereof: Is provided. By using such a compound, hair growth can be promoted.

According to the present invention,
A fermented papaya is provided for promoting hair growth. By using such a fermented product, hair growth can be promoted. According to the present invention,
There is provided a composition comprising the papaya fermentation. The use of such a composition can promote hair growth.

According to the present invention,
The following formula (I)
(In the formula (I),
R ₁ , R ₂ , R ₃ and R ₄ are each independently hydrogen, a hydroxyl group or -OCH ₃ ;
R _5, R ₆ and R ₇ are each independently hydrogen, -CH ₃ or -C ₂ H ₅ or -OCH ₃₎
A composition for antioxidation comprising at least one selected from the group consisting of a compound represented by formula (I), a physiologically acceptable salt thereof and a physiologically acceptable hydrate thereof is provided. By using such a composition, oxidation can be prevented.

FIG. 1 shows the HMBC correlation of compound PP-6. FIG. 2 shows a linear regression equation (Y = aX + b) with the average value of Trolox netAUC on the X-axis and the Trolox concentration (μg) on the Y-axis. FIG. 3 shows the results of the antioxidant activity (ORAC) of PP-6 (mean ± SD, n = 4). FIG. 4 shows a schematic of the human hair follicle dermal papilla cell (HFDPC) assay. FIG. 5 shows the cell viability (mean ± SD, n = 3) of HFDPC cells after addition of PP-6.

  Hereinafter, embodiments of the present invention will be described. The following embodiments are exemplifications, and the scope of the present invention is not limited to those shown in the following embodiments. It should be noted that, for the same contents, the description is omitted to avoid repetition.

Definitions For convenience, certain terms used in this application are collected here. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Unless otherwise indicated by context, the singular forms “a”, “an”, and “the” include plural references.

  The term "patient" refers to an animal, including a human, capable of promoting hair growth or treating, treating, ameliorating, or preventing a disease associated with hair growth with a composition of the present invention. The terms "subject" or "patient" are intended to refer to both male and female gender, unless the gender is specified. Thus, the term "subject" or "patient" includes any mammal that can benefit from the compositions of the present invention. Examples of "subjects" or "patients" include, but are not limited to, humans, rats, mice, guinea pigs, monkeys, pigs, goats, cows, horses, dogs, cats, birds and chickens. In an exemplary embodiment, the "subject" or "patient" is a human.

  The numerical ranges and parameters shown in the invention are approximations, but the numerical values given in specific examples are as accurate as possible. However, any numerical value inherently contains certain errors necessarily resulting from the standard deviation found in each test measurement. Also, as used herein, the term "about" generally means within 10%, 5%, 1% or 0.5% of a given value or range. Alternatively, the term "about" means within an acceptable standard error, as considered by one skilled in the art.

DETAILED DESCRIPTION OF EMBODIMENTS Composition for Promoting Hair Growth In this embodiment,
The following formula (I)
(In the formula (I),
R ₁ , R ₂ , R ₃ and R ₄ are each independently hydrogen, a hydroxyl group or -OCH ₃ ;
R _5, R ₆ and R ₇ are each independently hydrogen, -CH ₃ or -C ₂ H ₅ or -OCH ₃₎
A composition for promoting hair growth, comprising at least one selected from the group consisting of a compound represented by the formula, a physiologically acceptable salt thereof and a physiologically acceptable hydrate thereof: Is provided.

Also,
The compound represented by the above formula (I) has the following formula (II)
(In the formula (II),
R ₁ , R ₂ , R ₃ and R ₄ are each independently hydrogen, a hydroxyl group or -OCH ₃ )
The compound represented by

Also,
Compounds represented by the above formula (II) are represented by the following formulas (III) to (V)
And at least one compound selected from the group of

Also,
The compound represented by the above formula (IV) has the following formula (VI)
(Isopropyl 2,5-dihydroxybenzoate).

  The composition may be a pharmaceutical, cosmetic or food composition. When the composition is used as a pharmaceutical or cosmetic composition, it may be applied to areas of the skin in which hair growth is desired. When applied to the skin, the composition may be applied directly to the skin, or the composition may be applied directly to the skin via an appropriate substrate impregnated with the composition (eg, gauze or gel). May be. When the composition is applied to the skin, the composition may be in the form of a lotion (liquid), mousse, gel, jelly, emulsion, suspension, cream, ointment, sheet, aerosol. Or it may be in the form of a spray. When the present composition is used as a pharmaceutical composition, the present composition may be administered subcutaneously in an area where hair growth is to be promoted.

  When the present composition is used as a cosmetic composition, specific embodiments of the cosmetic composition include basic cosmetics such as lotions, emulsions, skin creams, lotions, oils, and packs; face wash, cleansing, body soap, and the like. Skin cleansing agents; hair cosmetics such as shampoos, rinses, conditioners, hair styling agents, and hair restorers; bath salts such as bath salts, bath tablets, and bath liquids; massage agents;

  When the present composition is used as a food composition, the food composition is obtained by processing a food that inherently contains the above-described compound, and is obtained by processing the above-mentioned compound by biological action (eg, fermentation). It may be a food composition or a food composition to which the above compound or a food containing the compound is added. Examples of the food composition include a papaya fermented product. The form of the food composition may be in its original form (e.g., liquid, sol, gel, powder and granules), and may be in the form of capsules, tablets, granules using appropriate carriers and auxiliaries. For example, it may be in a form easy to take.

  The present composition may contain a substance having an effect of promoting hair growth, which is different from the compound of the present embodiment. As such a substance, a conventionally known substance can be used, and a single substance or a combination of plural substances may be used. The content of these is not limited, but may be any range as long as the effect of promoting hair growth can be exhibited.

  Hair can also be read as body hair and includes, for example, head hair. Hair growth means growth through the activation of cells involved in hair, and may be hair growth, hair growth or both. Hair growth refers to the growth of currently existing hair, and may be hair elongation, increased hair diameter, or both. Hair growth means growing new hair (including regrowth), and growing one or more hairs from one pore and newly growing from one pore where one or more hairs have already grown. Includes growing one or more hairs. Cells involved in hair may be mesenchymal-derived fibroblasts, hair papilla cells, keratinocytes, and hair mother cells. Cell activation includes cell differentiation and proliferation (eg, proliferation of dermal papilla cells).

  The compositions can also treat, treat, ameliorate or prevent diseases related to hair growth (eg, alopecia). Specific examples include androgenetic alopecia, female alopecia, alopecia areata, seborrheic alopecia, pityriasis alopecia, age-related alopecia and the like.

In this embodiment,
Fermented papaya, to promote hair growth,
Contains at least one selected from the group consisting of compounds represented by any of the above formulas (I) to (VI), physiologically acceptable salts thereof and physiologically acceptable hydrates thereof And a composition comprising the papaya fermentation product or the papaya fermentation product. The papaya fermented product according to the present embodiment is a step of crushing papaya, a step of adding carbohydrate to the papaya, a step of maturing or fermenting the papaya to which the carbohydrate has been added, and a maturing or fermentation. A step of obtaining an undiluted solution from the papaya, a step of adjusting the concentration of the saccharide in the undiluted solution, a step of allowing the undiluted solution in which the concentration of the saccharide is adjusted to be aged to obtain an intermediate raw material, And a fermentation step.

  Papaya refers to `` papaya (scientific name: Carica papaya Linn) '' which is a fruit of the genus papaya of the papaya family, not only the yellow mature papaya that is lining up in the store as a fruit, but also the green immature used for cooking as a vegetable Also includes papaya. Papaya is widely cultivated from the tropics to the subtropics, but the origin is not particularly limited, and any of them can be used. In addition, any papaya, whether mature or immature, can be used as a fermentation target, but it is preferable to use wild immature papaya.

  The papaya is preferably crushed to facilitate fermentation. As the papaya to be crushed, it is preferable to use one excluding the scab part and the bottom of the fruit. This is because the bottom portion and the bottom portion are hard and may remain as foreign matter after fermentation. The crushing method is not particularly limited as long as the papaya after crushing can be mashed or pasted, and examples thereof include crushing, grinding, and a combination thereof.

  The amount of carbohydrate added to papaya may be from 10% to 50% by weight of the weight of papaya used. The papaya may be crushed before or after the sugar is added. In addition, sugar may be added during papaya crushing.

  The papaya crushed and added with carbohydrate is subjected to fermentation or standing aging. The fermentation or standing aging may be between two temperatures selected from any of 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 ° C. Such temperature control may be performed using a constant temperature incubator, a constant temperature chamber, or the like, and may be room temperature if the above temperature conditions are satisfied. In addition, the period of fermentation or standing aging may be a period between two numbers selected from any of 4, 5, 6, 7, 8, 9, and 10 days.

  The method for obtaining a stock solution from fermented or aged papaya is not particularly limited as long as it can separate a solid and a liquid, and examples thereof include a method by filtration. In the case of filtration, it is also possible to perform pressure filtration or centrifugal filtration.

  Adjust the carbohydrate concentration of the stock solution. Preferably, the concentration of the carbohydrate is between 40% and 60%. The carbohydrate concentration may be adjusted based on the results obtained by measuring the carbohydrate concentration during production, or may be adjusted based on past measurement data without measuring the carbohydrate concentration.

  An intermediate material is obtained by allowing the undiluted solution in which the carbohydrate concentration is adjusted to stand and ripen. The temperature of the standing aging at this time may be between two temperatures selected from any of 15, 16, 17, 18, 19, and 20 ° C. Such temperature control may be performed using a constant temperature incubator, a constant temperature chamber, or the like, and may be room temperature if the above temperature conditions are satisfied. Further, the period of the stationary ripening may be a period between two numbers selected from any of 7, 8, 9, 10, 11, 12, 13, and 14 days.

  The intermediate material is preferably fermented by adding one or more fermented bodies selected from the group consisting of enzymes for miso, lactic acid bacteria and amylase.

  In the present embodiment, yeast can be used for fermentation of papaya. The yeast to be used is not particularly limited as long as it can ferment papaya.Examples include Candida spp. (Candida) and Pichia spp. ) And the like, and preferably yeast of the genus Saccharomyces. Moreover, the natural yeast adhering or existing on papaya may be used for fermentation as it is. As long as the papaya can be fermented, it may be prepared by freeze-drying by a usual method or may be stored refrigerated. Optimum conditions for yeast fermentation depend on each yeast, and the method of use and conditions can be appropriately selected according to the conditions of the production process and the state of the target product. Furthermore, it is also possible to use one or more yeasts simultaneously or sequentially.

  In this embodiment, lactic acid bacteria can be used for fermentation of papaya. The lactic acid bacterium to be used is not particularly limited as long as it can ferment papaya, and examples thereof include Streptococcus, Lactobacillus, Lactococcus, Bifidobacterium, and Leuco bacterium. The genus Nostock (Leuconostoc), the genus Pediococcus (Pediococcus) and the like can be mentioned. Preferably, it is a lactic acid bacterium of the genus Streptococcus. Natural lactic acid bacteria attached or present on papaya may be used for fermentation as it is. Any lactic acid bacterium may be prepared by freeze-drying by a conventional method or stored refrigerated as long as papaya can be fermented. The optimum conditions for lactic acid bacteria fermentation are dependent on each lactic acid bacterium, and the method of use and conditions can be appropriately selected according to the conditions of the production process and the state of the target product. Furthermore, it is also possible to use one or more lactic acid bacteria simultaneously or sequentially.

  In the present embodiment, amylase can be used for fermentation of papaya. Amylase is a digestive enzyme that can hydrolyze the glycosidic bonds of amylose and amylopectin contained in starch.If it has sufficient activity in the production process of the target product, it does not have any particular restrictions. Absent. The optimal conditions for amylase depend on the enzyme used, and the enzyme to be used can be appropriately selected according to the conditions of the production process and the state of the target product. Furthermore, one or more amylase can be used simultaneously or sequentially.

  In this embodiment, an enzyme for miso can be used for fermentation of papaya. The enzyme for miso includes secretions of Aspergillus and the like used in the production of fermented foods such as miso and soy sauce. Examples of microorganisms contained in the genus Aspergillus include Aspergillus oryzae, Aspergillus niger, Aspergillus soae, and the like. Preferably, it is Aspergillus oryzae. The enzyme for miso does not necessarily need to be a purified enzyme as long as it meets the required quality of the target product, and may be a crudely purified enzyme, and it is also possible to use the cells themselves It is. The optimum conditions for the enzyme for miso are enzyme-dependent, and the method of use and conditions can be appropriately selected according to the conditions of the production process and the state of the target product. Furthermore, it is also possible to use one or more kinds of miso enzymes simultaneously or sequentially.

  Prior to fermentation, papaya is pasteurized (steam or hot water, etc.) or high temperature sterilized (heated steam or hot water, etc.), fermented, microwave, high frequency, infrared, far infrared, ultraviolet, γ-ray, X-ray, Sterilization treatment such as electron beam and ozone sterilization can also be performed. In the case of fermentation using mature papaya, it is preferable to perform a sterilization treatment because it is easy to cause spoilage by microorganisms and the like. On the other hand, when a wild immature papaya is used, it is preferable not to perform a sterilization treatment when microorganisms (such as yeast and lactic acid bacteria) attached to or present on the immature papaya are useful for fermentation.

  Carbohydrates added for the purpose of promoting fermentation can be starch, malt, dextran, sucrose (sucrose), maltose (maltose), glucose (glucose), etc., but inactivate papaya enzymes and prevent decay In view of the above, it is preferable to use a low-molecular-weight saccharide capable of increasing intracellular osmotic pressure and causing plasma membrane separation even with a saccharide having the same mass, and in particular, using glucose which is a monosaccharide. Is preferred.

  In another embodiment, the method further comprises a step of adding carbohydrate to the intermediate raw material after fermentation and then drying. Thereby, a dried papaya fermented product can be obtained.

  The amount of saccharide added to the intermediate material after fermentation is preferably 1 to 5 times the weight of the intermediate material before fermentation. The drying method is not particularly limited, but air drying which can suppress denaturation by heat and the like is preferable. The temperature at the time of air drying is preferably in the range of 15 to 20 ° C.

  In another embodiment, the step of fermenting the intermediate raw material comprises dividing the intermediate raw material into three parts, a first intermediate raw material, a second intermediate raw material, and a third intermediate raw material. 0 to 5.0% by weight of the enzyme for miso is added to the first intermediate material, mixed and fermented, and 3.0 to 30.0% by weight of lactic acid bacteria and 0.2 to 1% by weight of the second intermediate material. The enzyme for miso was added to the second intermediate material at a concentration of 0.0% by weight, mixed and fermented, and 0.5 to 4.0% by weight of an amylase of the third intermediate material was added to the third intermediate material. This is the step of fermenting after mixing. Thereby, a plurality of fermented products having different fermented bodies can be obtained from one intermediate raw material.

  Such fermentation is preferably carried out between 35 and 45 ° C. Such temperature control may be performed using a constant temperature incubator, a constant temperature chamber, or the like. Further, the fermentation time is preferably 20 to 72 hours, more preferably 40 to 50 hours.

  In another embodiment, the method further comprises the step of aging the dried, fermented intermediate material for at least six months.

  Such aging is preferably performed by enclosing in an aluminum packaging material having a high gas barrier property.

  The form of the papaya fermented product obtained by the above production method is a milky white powder, but it can be used as it is. As other forms, the composition can be prepared in any form such as powders, tablets, capsules, soft capsules, liquids, syrups, and pills, but is not particularly limited thereto.

According to the present embodiment,
Contains at least one selected from the group consisting of compounds represented by any of the above formulas (I) to (VI), physiologically acceptable salts thereof and physiologically acceptable hydrates thereof , An antioxidant composition is provided.

Functional food or health food According to this embodiment,
Contains at least one selected from the group consisting of compounds represented by any of the above formulas (I) to (VI), physiologically acceptable salts thereof and physiologically acceptable hydrates thereof And a functional food or health food for promoting hair growth.
A functional food or health food having an effect of promoting hair growth can be provided by appropriately adding the above compound as needed.

  As used herein, the term "functional food" refers to a food that contains components that can provide more benefits to those who have consumed the food due to nutrients originally contained in the food itself.

  In this specification, the term "healthy food" is a general term for foods that are generally sold and used as foods that contribute to maintaining and promoting health, and also include supplements that assist in ingesting nutrients that are often in short supply.

Method for treating, treating, ameliorating or preventing a disease related to hair growth According to the present invention,
A method for treating, treating, ameliorating or preventing a disease relating to hair growth, the method comprising:
To a patient suffering from the above-mentioned disease relating to hair growth, a compound represented by any of the above formulas (I) to (VI), a physiologically acceptable salt thereof and a physiologically acceptable hydrate thereof Administering a composition comprising at least one or more selected from the group consisting of:
A method is provided.

  When the above composition is administered to a patient, the dosage varies depending on various conditions such as the severity of the patient's symptoms, age, weight, and physical condition. In general, it is sufficient to administer once or more times a day with the above-mentioned dose and usage, and the number and amount of administrations may be appropriately increased or decreased according to the above-mentioned various conditions.

  The daily dose, administration period and number of administrations of the composition may be the same as those of the therapeutic agent described above. The administration of the composition may be terminated at the discretion of the physician, or may be terminated at the patient's own discretion.

Method for promoting hair growth According to this embodiment,
A method for promoting hair growth, the method comprising:
A group consisting of a compound represented by any of the above formulas (I) to (VI), a physiologically acceptable salt thereof and a physiologically acceptable hydrate thereof, for a subject who desires to promote hair growth. Applying a composition containing at least one or more selected from those to the skin area in need of promoting hair growth,
A method is provided.

  Although the embodiments of the present invention have been described above, these are merely examples of the present invention, and various configurations other than the above can be adopted.

Preparation of compounds
15 kg of papaya fermented product (SAIDO-PS501, Carica Therapy, Japan) was extracted with methanol at room temperature. The extract was concentrated to dryness under reduced pressure using a rotary evaporator to obtain 450 g of a methanol extract. The total methanol extract was chromatographed on a Diaion HP-20 stationary phase eluting with 100% MeOH from water to give the methanol fraction (7.0 g). The methanol fraction was added to the top of a silica gel column (500 g, 30 × 6 cm) previously packed in n-hexane. Flow n-hexane through the column with increasing ratio of EtOAc (n-hexane: EtOAc = 50: 50 → 0: 100) and then flow through the column with increasing ratio of MeOH (EtOAc: MeOH = 100: 0). → 50: 50) to perform gradient elution. The effluent was fractionated every 250 mL. Each fraction was concentrated at 40 ° C. under reduced pressure and confirmed by thin layer chromatography (TLC). Fractions with the same chromatographic pattern were pooled together and evaporated to dryness. TLC was performed on the pre-coated RP-C18F ₂₅₄ plate using MeOH-H ₂ O (50:50) for the sub-fractions 34-35 eluted with EtOAc-methanol (80:20), and the compound (hereinafter referred to as PP -6) was obtained in an amount of 3.8 mg.

Compound identification
¹ H and ¹³ C-NMR spectra of PP-6 were measured using a Bruker DRX 600 NMR spectrometer (Bruker Daltonics Inc., MA, USA). TMS was used as an internal standard for chemical shifts. Chemical shifts (δ) were expressed in ppm relative to the TMS resonance. The HR-FAB-MS of PP-6 was measured using a JEOL JMS 700 spectrometer (JEOL, Japan). Dimethyl sulfoxide (DMSO) and organic solvent were purchased from Wako Pure Chemical Industries (Osaka, Japan). Diaion HP-20 was purchased from Mitsubishi Chemical Co. (Tokyo, Japan). Silica gel (75-120 mesh) was purchased from Wako Pure Chemical Industries (Osaka, Japan). Thin layer chromatography (TLC) silica gel 60F ₂₅₄ plates were purchased from Merck Co., Darmstadt, Germany. The plate on which the sample was developed was developed in a solvent mixture of different organic solvents. The developed chromatograms were visualized under UV light 254 nm, were visualized spot by spraying vanillin / H ₂ SO ₄ reagent. After visualization, the plates were warmed for 5 minutes in an oven preheated to 110 ° C. Preparative TLC was performed on precoated silica gel 60GF ₂₅₄ (20 × 20 cm × 0.2 mm thickness) or pre-coated RP-C18F ₂₅₄ plate (5 × 7.5 cm × 0.2 mm thickness).

It was analyzed by ¹ H and ¹³ C-NMR spectra as well as HR-FAB-MS spectra of PP-6. As a result, a protonated molecule [M + H] ⁺ peak was shown at m / z 197.0813 corresponding to the molecular formula of C ₁₀ H ₁₂ O ₄ . ^{The 1} H NMR spectrum clearly indicated the presence of two sets of proton signals. The set of proton signals from 6.78 ppm to 7.22 ppm (hereinafter the first set) consists of three aromatic signals, δ _H 6.78 (d, J = 9.0 Hz) and δ _H 6.96 (dd, J = 9.0, 3.0 Hz). And δ _H 7.22 (d, J = 3.0 Hz). This was confirmed by ¹³ C NMR spectral data showed downfield signal caused by benzene ring resonance between [delta] _C 156.2 from [delta] _C 113.8. The doublet observed at δ _H 6.78 (d, J = 9.0 Hz) couples with one of the doublets at δ _H 6.96 (dd, J = 9.0, 3.0 Hz). Attributable to two ortho-coupled protons at H-3 and H-4. Another of the doublets at δ _H 6.96 (dd, J = 90, 3.0 Hz) was meta-coupled with the remaining aromatic protons at δ _H 7.22 (d, J = 3.0 Hz). Therefore, they were located at H-4 and H-6, respectively. This indicated the substitution pattern of the trisubstituted aromatic ring. This was confirmed by assigning the presence of two oxygenated carbons at δ _C 150.7 and 156.2 in the ¹³ C NMR spectrum to C-5 and C-2, respectively. The second set of protons relates to the proton signal of the aliphatic moiety with a doublet at δ _H 1.38 (J = 6.0 Hz) associated with the quartet oxymethine proton at δ _H 5.26 (J = 6.6 Hz). . ^{The 13} C NMR spectrum further showed two characteristic peaks, δ _C 170.9 belonging to the carbonyl ester moiety and δ _C 70.5 belonging to the carbon attached to the acyloxy oxygen. The complete structure of PP-6 was assembled based on the HMBC correlation as shown in Table 1 and FIG. As a result, PP-6 was identified as isopropyl 2,5-dihydroxybenzoate (isopropyl 2,5-dihydroxybenzoate), which is an aliphatic ester of hydroxysalicylic acid.

Chemical shifts (δ) are expressed in ppm, and coupling constants (J) are expressed in Hz. ¹ H and ¹³ C NMR were measured in heavy methanol (CD ₃ OD) at 600 and 150 MHz, respectively.

Oxygen Radical Absorption Capacity (ORAC) Assay
ORAC of PP-6 was measured by ORAC assay. The ORAC assay is described in Garrett, AR, et al, (Measuring Antioxidant Capacity Using the ORAC and TOSC Assays.In Methods in molecular biology (Clifton, NJ); 2010; Vol. 594, pp. 251-262) and Roy, MK, It was performed based on et al, (ORAC and DPPH assay comparison to assess antioxidant capacity of tea infusions: Relationship between total polyphenol and individual catechin content. Int. J. Food Sci. Nutr. 2010, 61, 109-124). Fluorescein is excited by light at a wavelength of 485 nm and emits light at a wavelength near 520 nm. Light is quenched when fluorescein is oxidized by peroxy radicals generated from AAPH (2,2'-azobis (2-aminopropane) dihydrochloride). The presence of antioxidants in the sample scavenges peroxy radicals and slows quenching of fluorescein. In the ORAC method, the ability to delay the quenching speed is evaluated as the antioxidant ability of a sample.

  PP-6 was dissolved in a 75 mmol / L phosphate buffer (pH 7.4) and diluted to an appropriate concentration (sample solution). Standard curves were prepared using 50, 25, 12.5 and 6.25 μmol / L trolox solutions (Trolox standard sample solutions). The experiments were performed in sets of four identical solutions. A Trolox standard sample solution and a sample solution were added to a 96-well plate, and Blank was filled with 20 μL of a 75 mmol / L phosphate buffer (pH 7.4). 200 μL of a 94.4 nM fluorescein solution was added to each well. The 96-well plate was warmed in a 37 ° C. water bath for 10 minutes. Next, 75 μL of a 31.7 mM AAPH solution heated at 37 ° C. for 10 minutes was added to each well. Using a fluorescence spectrophotometer having an excitation wavelength of 485 nm and an emission wavelength of 515 nm, the fluorescence intensity was measured at 37 ° C. for 30 minutes at intervals of 30 seconds from 30 seconds after the addition of AAPH. The AUC of each sample was calculated from the obtained measurement results of the fluorescence intensity in each well. AUC was determined by the following equation.

  The data of 0 μM trolox was used as a blank, and the increase in AUC (AUCtrolox-AUCBlank) of the data obtained with each trolox standard sample solution (50, 25, 12.5 and 6.25 μmol / L) was defined as netAUC. Table 2 shows the average of the obtained values.

  Table 3 shows the increase in AUC of the data obtained with the sample solution (PP-6) as netAUC.

A linear regression equation (Y = aX + b) was created by taking the average value of Trolox netAUC on the X axis and the Trolox concentration (μg) on the Y axis (FIG. 2). The relative ORAC value was calculated using this regression equation.

Results The results are expressed in mg TE / mg PP-6 as mg Trolox equivalents (TE) of PP-6 (FIG. 3). PP-6 exhibited antioxidant activity of 0.19 mg TE / mg, about one fifth of that of Trolox, a potent antioxidant. All chemicals and reagents used in this assay were of analytical grade and were purchased from Wako Chemical, Osaka, Japan.

Human hair follicle dermal papilla cell (HFDPC) assay
An overview of the HFDPC cell assay is shown in FIG. HFDPC cells were cultured in a 96-well plate under the following conditions.
Medium: Follicular dermal papilla cell (FDPC) medium (PromoCell, Heiderberg) containing 10% FBS (GE Healthcare Life Sciences Hyclone lab, Utah, USA) and 1% streptomycin / penicillin (Fujifilm Wako Pure Chemical Industries, Osaka, Japan) , Germany)
Temperature: 37 ° C
Cell concentration: 2 × 10 ⁴ cells / well Atmosphere: Humidified atmosphere under 5% CO ₂ conditions Culture time: 24 hours

  Next, the above medium was replaced with an FDPC medium containing 0.5% FBS and PP-6 (final concentrations of 11 μg / mL and 110 μg / mL, respectively) and cultured for 3 days. Thereafter, the cells were cultured for 4 hours in an FDPC medium containing 10% FBS and MTT [3- (4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide)] (5 mg / mL), and acid isopropanol was added. Was added to each well to dissolve the purple-blue insoluble formazan precipitate. After keeping in the dark for 4 hours, the absorbance was measured at 570 nm using a plate reader.

Pre-culture of cells HFDPC cells were used in this test. 10 μL of a culture solution containing HFDPC cells cultured on a Petri dish was added to a hemocytometer, and the number of cells was measured using a microscope. Thereafter, Dulbecco's Modified Eagle Medium (DMEM) medium was added to the cell culture solution, and the cell concentration was adjusted to 1.0 × 105 cells / mL. This was inoculated at 1 mL per well of a 96-well plate, and cultured statically at 5% CO ₂ and 37 ° C. for 24 hours.

Addition of Sample The medium in the 96-well plate was removed, and 1 mL of DMEM medium was newly added. Thereafter, 2 μL of the previously prepared sample solution was added to the 96-well plate. The 96-well plate was cultured at room temperature for 24 hours at 37 ° C. in 5% CO ₂ . Twenty-four hours later, remove the medium from the 96-well plate again, add 1 mL of DMEM medium to the 96-well plate, add 2 μL of the sample solution, and incubate at 5% CO ₂ at 37 ° C for 48 hours. Cultured.

Measurement of Cell Viability by MTT Method To each well, 50 μL of an MTT staining solution (in 5 mg / mL PBS) was added, and the cells were incubated at 5% CO ₂ at 37 ° C. for 4 hours. The medium was removed, and 1 mL of a hydrochloric acid-isopropanol solution was added to each well. The absorbance at 570 nm was measured with a microplate reader and used as an index of cell viability.

result
To examine the effect of PP-6 on hair growth, the proliferation ability of HFDPC (hair papilla cell model) when PP-6 was added was evaluated by cell viability. FIG. 5 shows the cell viability of HFDPC cells. The addition of PP-6 (11, 110 μg / mL) increased the cell viability of HFDPC (FIG. 5). Therefore, it was revealed that PP-6 has a function of promoting the proliferation of hair papilla cells. From the above results, it was revealed that PP-6 contains a component that contributes to the proliferation of hair papilla cells.

Claims (9)

The following formula (I)
(In the formula (I),
R ₁ , R ₂ , R ₃ and R ₄ are each independently hydrogen, a hydroxyl group or -OCH ₃ ;
R _5, R ₆ and R ₇ are each independently hydrogen, -CH ₃ or -C ₂ H ₅ or -OCH ₃₎
A composition for promoting hair growth, comprising at least one selected from the group consisting of a compound represented by the formula, a physiologically acceptable salt thereof and a physiologically acceptable hydrate thereof: . The compound represented by the formula (I) is represented by the following formula (II)
(In the formula (II),
R ₁ , R ₂ , R ₃ and R ₄ are each independently hydrogen, a hydroxyl group or -OCH ₃ )
2. The composition according to claim 1, which is a compound represented by the formula: The compound represented by the formula (II) is represented by the following formulas (III) to (V)
The composition according to claim 2, which is at least one compound selected from the group consisting of:   4. The composition according to claim 3, wherein the compound represented by the formula (II) is a compound represented by the formula (IV). The compound represented by the formula (IV) is represented by the following formula (VI)
5. The composition according to claim 4, which is a compound represented by the formula:   Fermented papaya to promote hair growth. The following formula (I)
(In the formula (I),
R ₁ , R ₂ , R ₃ and R ₄ are each independently hydrogen, a hydroxyl group or -OCH ₃ ;
R _5, R ₆ and R ₇ are each independently hydrogen, -CH ₃ or -C ₂ H ₅ or -OCH ₃₎
7. The papaya fermented product according to claim 6, comprising at least one selected from the group consisting of a compound represented by the formula, a physiologically acceptable salt thereof, and a physiologically acceptable hydrate thereof.   A composition comprising the papaya fermented product according to claim 6 or 7. The following formula (I)
(In the formula (I),
R ₁ , R ₂ , R ₃ and R ₄ are each independently hydrogen, a hydroxyl group or -OCH ₃ ;
R _5, R ₆ and R ₇ are each independently hydrogen, -CH ₃ or -C ₂ H ₅ or -OCH ₃₎
An antioxidant composition comprising at least one compound selected from the group consisting of a compound represented by the formula, a physiologically acceptable salt thereof, and a physiologically acceptable hydrate thereof.