RU2799324C2 - Olespedea captic extract for hair care applications - Google Patents

Abstract

FIELD: hair care.

SUBSTANCE: use of an extract of Lespedeza capitata obtained after extraction with a water-alcohol mixture or alkyl esters, for the treatment or prevention of alopecia, seborrheic dermatitis of the scalp, erythema of the scalp, seborrhea of the scalp, and combinations thereof are disclosed. Also the use of an extract of Lespedeza capitata and a dermatological composition for the treatment or prevention of androgenetic alopecia, reactive alopecia, postmenopausal alopecia and aerated alopecia, the use of a cosmetic composition for hair and/or scalp care, the use of a dermatological composition for the treatment or prevention of alopecia, seborrheic dermatitis of the scalp skin, scalp erythema, scalp seborrhea, and combinations thereof are disclosed.

EFFECT: group of inventions provides treatment and prevention of alopecia and/or seborrhea of the scalp.

11 cl, 5 tbl, 12 ex

Description

Technical field

The present invention relates to an extract of Lespedeza capitata Michx. for use in cosmetics and/or dermatology in the field of hair care; more specifically, in the treatment and/or prevention of alopecia and/or seborrhea of the scalp.

Prior Art

Lespedeza capitata Michx. is a herbaceous perennial plant of the legume family, native to North America. Lespedeza capitata Michx. cultivated in Europe and especially in France.

Lespedeza capitata Michx. - This is a felt plant, the erect stems of which can reach a height of 1.5 meters. Its leaves are alternate, compound and usually trifoliate; the dimensions of the leaf blades are approximately 4.5×1.8 cm; petiole is short (2-5 mm). The leaves and stems are covered with adpressed hairs that give the plant a silver sheen. Flowers are uriculate, densely clustered in axillary inflorescences. The petals are white, except for the sail, which is pink to purple. The calyx is stable and turns brown over time. The fruits are unexpanded and bear a single seed. Lespedeza capitata Michx. has a deep root system with a main root that can enter the soil to a depth of 2.5 meters.

Lespedeza capitata Michx. is part of the traditional medicine of many Native Americans. Omaha and Ponca used the stems as moxa to treat rheumatism or neuralgia. The Comanche used the leaves as an infusion. The Meskwaki used the root as an antidote for poisoning (USDA-NRCS. Plant Guide Dec 05, 2000). Currently Lespedeza capitata Michx. grown mainly for use in food supplements, pharmaceuticals, homeopathic and veterinary preparations.

Aerial parts of Lespedeza capitata Michx. have been studied mainly in relation to their properties in the urinary area (in particular, in relation to their diuretic and antiazotemic action). Several clinical studies have been documented. Most of them were held in the 60-80s, mainly in France. Clinical studies are particularly concerned with the intravenous administration of Lespedeza capitata Michx extracts. patients with kidney disease. Cholesterol-lowering action of Lespedeza capitata Michx tincture. when administered orally was reported in a clinical study conducted in 39 subjects with stable hypercholesterolemia (Yarnell and Abascal, Alternative and Complementary Therapies. 13(1): 18-23, 2007).

Aerial parts of Lespedeza capitata Michx. are still used for their therapeutic properties. They are sold in the form of powders or extracts for the preparation of tinctures, as dietary supplements, homeopathic or pharmaceutical preparations (azotemia; urinary sphere) or veterinary preparations (urinary sphere).

One recent publication shows that an ethanolic extract of Lespedeza capitata Michx. induces in vitro proliferation of fibroblasts and keratinocytes, inhibition of collagenase and stimulation of collagen synthesis by fibroblasts, and has a lipolytic effect (Pastorino et al. 2017. Industrial Crops and Products. 96: 158-164). Patent application CN 104606122 also claims that Lespedeza capitata, when mixed with other plants, is able to burn pregnancy-related abdominal fat.

The essence of the invention

Unexpectedly and with surprise, Applicants have shown that the extract of Lespedeza capitata Michx. has pharmacological activity of interest in the field of treatment and/or prevention of diseases of the skin and hair, especially for the control of hair loss and for the treatment of seborrhea of the scalp and related diseases.

Indeed, the inventors have shown that the extract of Lespedeza capitata Michx. inhibits the synthesis and release of Dickkopf-1 (DKK1) protein by dermal papilla cells. Thus Lespedeza capitata Michx. It will help delay and prevent hair loss and prolong the life cycle of the hair.

In addition, the inventors have shown that the extract of Lespedeza capitata Michx. inhibits the activity of 5α-reductase in the cells of the dermal papillae. Therefore, the extract of Lespedeza capitata Michx. will be useful in the prevention of hair loss, as well as seborrhea of the scalp and related diseases of the skin and hair.

This is a new application of Lespedeza capitata Michx. in the field of hair care is the subject of the present invention and allows for the use of various products based on Lespedeza capitata Michx. to improve the aesthetic appearance of the hair by caring for it, as well as to prevent and / or treat hair loss and / or seborrhea of the scalp and all associated skin and hair diseases.

Therefore, the invention also relates to an extract of Lespedeza capitata for its use in the treatment and/or prevention of at least one disease of the skin and hair.

Advantageously, said skin and hair disease is selected from alopecia, scalp seborrheic dermatitis, scalp erythema, scalp seborrhea, and combinations thereof.

Oily hair can also be a disease of the skin and hair. Detailed description of the invention

Hair care not only for cosmetic purposes, but also to prevent hair loss and restore hair is always of interest to researchers.

The hair follicle is a mini-organ located in the skin up to the subcutaneous tissue, the main function of which is to produce the hair shaft. The hair follicle is a dynamic structure that produces hair. Hair does not grow continuously, but in accordance with the hair growth cycle, which includes three phases:

- Growth phase (anagen): cells of the dermal papilla (fibroblasts) send a signal to the bulb stem cells, which allows them to proliferate. These cells transform and enclose the dermal papilla to form the sulfur matrix of the hair. They divide and differentiate into keratinocytes, the cells responsible for hair structure. In order for the hair to be properly structured, keratinocytes need sulfur-containing proteins, vitamin B6, and various minerals such as zinc and magnesium. The duration of this phase determines the length of the hair and depends on the proliferation and differentiation of matrix cells at the base of the follicle.

- Regression phase (catagen): the matrix dies and, as a result, the dermal papilla is no longer in contact with this matrix. There is no more exchange between cells. The follicle and dermal papilla move upward towards the epidermis.

- Resting phase (telogen): dermal papillae and bulb cells are intact and inactive. Hair fall out. In order for new hair to grow, the cycle must be restarted.

Hair is constantly renewing and of the 100,000-150,000 hairs that make up the hair, most are in the growth phase. Normal physiological hair loss is 60 to 100 hairs per day for healthy hair. Beyond this, the loss is considered pathological, either accidental or permanent.

The term alopecia means partial or total hair loss. Many factors can be involved in alopecia, such as, for example, genetic factors, age, sex, illness, stress, hormonal problems, drug side effects, and trauma. There are several forms of alopecia:

- Hereditary androgenetic alopecia is the most common. Early hair loss occurs in people with a genetic predisposition, especially men. It is manifested by a decrease in hair volume or even baldness and affects 50% of men over 50;

- Postmenopausal alopecia is the most common cause of hair loss in women. In women, hair loss is more diffuse and widespread than in men. Diffuse alopecia in women is a condition that often begins around menopause and affects approximately 40% of older women over 70 years of age. The term diffuse shows that, unlike men, baldness spreads to all the hair on the head evenly;

- Acute or reactive alopecia can be associated with chemotherapy, stress, childbirth, severe nutritional deficiencies, iron deficiency and hormonal disorders. This is a simultaneous and diffuse loss of a significant amount of hair;

- Scarring alopecia can be caused by skin problems (tumor, burn, alopecia areata), acute radiation, lupus erythematosus or parasites (dermatophytosis, lichen);

- Alopecia areata has an autoimmune origin and is characterized by lesions in the form of spots of different sizes in one or more places;

- Congenital alopecia is rare and is the absence of hair roots or anomalies (mutations) of the hair.

Alopecia is mainly associated with a violation of hair renewal, which leads first to an increase in the frequency of the cycle at the cost of hair quality, and then their quantity. The most common occurrence is shortening of the growth cycle (anagen phase) due to arrest of cell proliferation. The consequence of this is premature induction of the catagen phase and more hair follicles in the telogen phase and therefore more hair loss. Therefore, to combat alopecia, it is necessary to restart the hair cycle, for example by activating the anagen phase.

It is now known that the mechanisms responsible for hereditary androgenetic alopecia (also called seborrheic alopecia) include, among others, hormonal components with overexpression of androgen receptors (testosterone and DHT receptors) and with more intense activity of the 5-alpha reductase enzyme. This hormonal dysregulation leads to excessive production of dihydrotestosterone (DHT), the active metabolite of testosterone. In the dermal papilla, this metabolite will stimulate the production of hair cycle inhibitors, resulting in a shortening of the anagen phase, causing the hair to enter the telogen phase too quickly and not giving the hair follicle enough time to produce quality keratin and, necessarily after several cycles, depleting the hair follicle's capacity. to produce a hair shaft.

This alopecia caused by an excess of androgens also affects women during menopause (postmenopausal alopecia) or after androgen treatment. It starts at the temples and crown. This hair loss is more diffuse and widespread than in men. Hair loss evenly spreads to the entire scalp.

Therefore, the 5-alpha reductase inhibitor can treat and/or prevent hair loss in men and/or women.

Seborrhea is an excessive production of sebum by the sebaceous glands. Overall, humans have 2,000,000 sebaceous glands connected to 6,000,000 hairs. The distribution of the sebaceous glands is uneven. The density of sebaceous glands reaches from 300 to 900 sebaceous glands/cm ² on the face and scalp, and this density is about 100 sebaceous glands/cm ² in the upper chest and back. Androgens influence the activity of the sebaceous glands. Androgens are active only under the influence of the enzyme 5-alpha reductase, which ensures the metabolism of androgens in the sebaceous glands, which causes the production of sebum. Overactivation of the 5-alpha reductase enzyme causes seborrhea.

The place of manifestation of seborrhea is the medial region of the face (forehead, nose, chin), where the sebaceous glands are the most numerous and largest. Seborrhea also appears on the scalp, where it predominates in the frontal and frontotemporal regions, as well as in the upper part of the skull.

Scalp seborrhea causes aesthetic and dermatological conditions such as seborrheic dermatitis and scalp erythema.

Seborrheic dermatitis of the scalp (SD) is characterized by diffuse pink inflammatory patches covered with oily, white, hard scales adhering to the epidermis, sometimes purulent and sometimes intensely itchy.

Seborrhea is often associated with androgenetic alopecia.

Thus, an active inhibitor of the enzyme 5-alpha-reductase will reduce sebum secretion, treat seborrhea and solve dermatological and/or aesthetic problems associated with seborrhea.

The hair growth cycle is a highly regulated process during which different parts of the hair follicle (dermal papilla, stem cells found in the bulb, keratinocytes of the hair follicle matrix) come into close interaction through various molecular exchanges. Among the signals involved, the Wnt/β-catenin signal is known to promote hair growth (Leiros et al., Br. J. Dermatol. 166(5)-1035-1042, 2012).

Androgenetic alopecia, which is the most common form of hair loss, is clearly an androgenic disorder. Androgens dysregulate factors secreted by the dermal papilla, resulting in a failure of stem cell differentiation by inhibiting the canonical Wnt signaling pathway.

The hair cycle inhibitor Dickkopf protein 1 (DKK1) is known to block canonical activation of the β-catenin mediated Wnt pathway and promote catagen phase progression (Kwack et al., J. Invest. Dermatol 132 (6): 1554- 60, 2012). DKK1 is also described as being regulated by androgens (Kwack et al., J. Invest. Dermatol. 128 (2): 262-9, 2008).

Taken together, these results indicate that DKK1 plays an important role in androgen-induced hair growth suppression and early induction of the catagen phase in androgenetic alopecia (Inui and Itami, J. Dermatol. Sci. 61 (1): 1-6, 2011) .

Extract according to the invention

In the present description, the plant Lespedeza capitata Michx. for brevity, we can designate Lespedeza capitata.

In the context of the present invention, a Lespedeza capitata extract is obtained from one or more parts of the Lespedeza capitata plant, selected from roots and aerial parts such as stems, branches, leaves, fruits, seeds and/or flowers. Predominantly, these are above-ground parts selected from leaves, stems or flowers, either alone or in a mixture.

In a specific embodiment of the invention, the extract is obtained from the leaves and/or stems and/or flowers of Lespedeza capitata, especially when dried.

In a specific embodiment of the invention, the extract is obtained from a cell culture of Lespedeza capitata.

According to a specific embodiment, the extract is a water-alcohol extract, in particular a water-ethanol extract. Preferably, this will be an extract obtained as a result of water-alcohol extraction, in particular water-ethanol.

According to a preferred embodiment, the extract according to the invention can be obtained by the method according to the invention, described below.

The Lespedeza capitata plant, or part thereof, may be fresh or dried, whole, cut or ground, and may then be subjected to an extraction step.

The method for obtaining an extract according to the invention will include the step of extracting the whole Lespedeza capitata plant or part thereof with a hydrophilic or non-polar solvent, preferably a hydrophilic or moderately polar aqueous solvent.

"Moderately polar solvent" means in the context of the present invention a solvent selected from the group consisting of C1-C5 alcohols, glycols (propylene glycol, butylene glycol, pentylene glycol), glycerol, acetone, alkyl esters (such as, in particular, ethyl acetate and isopropyl acetate) , mixtures of water-miscible solvents such as a water-alcohol mixture or an acetone/water mixture, for example, or even hydrotropic solvents. The group of alternative hydrotropic solvents are water-soluble amphiphilic molecules which, at sufficient concentration, can extract moderately polar compounds such as those described in the characterization of the extract.

In the context of the present invention, "alkyl esters" means a compound R ₁ -COO-R ₂ where R ₁ and R ₂ are the same or different (C1-C6) alkyl groups. The ester may in particular be an acetate, i.e. compound CH ₃ COO-R ₂ .

In the context of the present invention, the group "(C1-C6)alkyl" means a straight or branched hydrocarbon chain containing from 1 to 6 carbon atoms, preferably from 1 to 4 carbon atoms. Examples include methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, pentyl or hexyl groups.

In the context of the present invention "non-polar solvent" means a solvent selected from heptane, hexane, limonene, halogenated hydrocarbons (chloroform, dichloromethane), supercritical CO ₂ and supercritical CO ₂ +ethanol.

It is understood that the choice of a non-polar extraction solvent, such as defined above, is not preferred.

"Hydrophilic solvent" means a solvent selected from the group consisting of water, subcritical water, water-miscible alcohols (eg ethanol), C3-C5 glycols, glycerol, acetone and mixtures thereof.

"Aqueous solvent" means a solvent selected from the group consisting of water alone and water in combination with subcritical water, water-miscible alcohols (eg ethanol), C3-C5 glycols, glycerol, acetone and mixtures thereof.

In the context of the present invention, "dry extract" means an extract without an extraction solvent or carrier solvent, or containing only a small trace of the solvent. Thus, such a dry extract contains only material obtained from Lespedeza capitata. It may also contain minor traces of an extraction solvent.

In a preferred embodiment of the invention, the extraction solvent may be selected from ethyl acetate, isopropyl acetate, C1-C5 alcohol, C3-C5 glycol, glycerol or water, or mixtures thereof. Preferably it will be water or a water/alcohol mixture. Preferably it will be an ethanol/water mixture.

Even more preferably, this ethanol/water mixture will have an ethanol/water ratio of 9:1 to 7:3 (v/v).

Even more preferably, the moderately polar solvent is a 9:1 (v/v) ethanol/water mixture.

According to another particular embodiment of the invention, the extraction is carried out under stirring or statically, at reflux, at ambient temperature or at a temperature between ambient and reflux. This can be facilitated by ultrasound, microwaves or extrusion at a plant weight/solvent volume ratio of 1/3 to 1/30 for 1 minute to 48 hours. Extraction can be repeated 2-3 times.

According to another specific embodiment of the invention, the solid phase is then separated by centrifugation or filtration to isolate a clear liquid phase free of particles. Filtration can be carried out through a paper filter or a filter plate with a cutoff of 5 to 20 µm, in particular 10 to 20 µm. Suitable filters or filter plates can be selected from the Pall® K Series Depth Filtration Product Line. These plates consist of a balanced blend of cellulose fibres, diatomaceous earth and perlite to create a well-defined matrix. Therefore, for example, you can choose Pall® series K inserts from K300 to K900. The liquid phase representing the extract may be concentrated to a greater or lesser extent until a dry extract is obtained.

In another embodiment of the invention, the carrier may be added at the stage of concentration to obtain an extract containing from 1 to 75% dry extract. The carrier may be maltodextrin, lactose, silicon dioxide, glycerol, glycol, or any other cosmetically acceptable carrier that solubilizes the extract, preferably of biological origin, such as, for example, bio-based glycols (1,2-pentanediol; 1,3-butanediol; 1,3-propanediol, etc.), as well as hydrotropes, such as, for example, alkyl glycosides (Sepiclear, Apyclean, ARCHS4, etc.).

According to a specific embodiment of the invention, the extract can be decolorized, for example with activated charcoal, to remove all or part of the chlorophyll.

According to a specific embodiment of the invention, the extract of Lespedeza capitata according to the invention is characterized by a content of all polyphenols from 2 to 40%, in particular from 5 to 35%, more specifically from 10 to 30%, even more specifically about 25% (% by weight in relation to to the mass of dry extract), expressed in grams of epicatechin per 100 grams of dry extract.

In the present invention, "about" means that the value in question may be 10% lower or higher, especially 5%, in particular 1%, than the indicated value.

Composition according to the invention

Another aspect of the invention relates to a cosmetic, dermatological or skin and hair composition containing at least one extract of Lespedeza capitata according to the invention with at least one cosmetically or dermatologically acceptable excipient, intended for hair and/or scalp care.

In the present invention, "cosmetically or dermatologically acceptable" means something useful in the preparation of a cosmetic, dermatological, or skin and hair formulation that is generally safe, non-toxic, and not biologically or otherwise undesirable, and that is acceptable for cosmetic, dermatological, or purposes, or for use on the skin and hair, in particular by topical application to the hair and/or scalp.

"Topical application" means application to the skin, especially the scalp, mucous membranes and/or external integuments, especially the hair and scalp.

"Covering" means head and body hair, eyebrows, eyelashes and/or nails, preferably head hair.

In a specific embodiment of the invention, the composition of the present invention is characterized in that it is in a form suitable for topical application to the scalp and/or hair.

In a specific embodiment of the invention, the compositions according to the invention are intended for topical application, especially by applying the composition to the scalp and/or hair and integuments.

Thus, the composition according to the invention can be presented in commonly known topical forms, i.e. as lotions, balms, shampoos, mousses, gels, dispersions, emulsions, sprays, serums, masks or creams, with excipients which provide penetration for the purpose of improving the properties and availability of active ingredients.

Advantageously, the compositions according to the invention may be presented in well-known forms for topical application to the hair and scalp, for example, as a shampoo, conditioner, hair cream, hair lotion, mask or leave-in spray. Therefore, they are cosmetic, dermatological or skin and hair compositions.

Thus, a distinction is made between products that require rinsing and products that do not require rinsing.

In a specific embodiment of the invention, the composition according to the invention is characterized in that it is present in a form suitable for oral administration. In fact, it has been shown that Lespedeza capitata can inhibit the converting enzyme of the renin-angiotensin system even when administered orally (Yarnell, 2012, IJKD, vol 6, 407-418). Therefore, this extract is not completely metabolized in the gastrointestinal tract.

According to another embodiment of the invention, the compositions according to the invention may also be present in conventional oral administration forms, ie in particular tablets, capsules, powders, granules and oral solutions or suspensions. When the solid composition is in the form of tablets, the main active ingredient is mixed with a pharmaceutical carrier such as gelatin, starch, lactose, magnesium stearate, talc, gum arabic or the like. Tablets can be coated with sucrose or other suitable materials, or even processed so that they have prolonged or delayed activity and so that they continuously release a given amount of the active ingredient.

A capsule preparation is obtained by mixing the active ingredient with a diluent and placing the resulting mixture into hard or soft capsules.

In addition to the extract according to the invention, these compositions contain a physiologically acceptable medium, usually water or a solvent based on, for example, alcohols, ethers or glycols. They may also contain surfactants, complexing agents, preservatives, stabilizers, emulsifiers, thickeners, gelling agents, humectants, emollients, trace elements, essential oils, fragrances, colorants, humectants or thermal waters, etc.

In one embodiment, cosmetic or dermatological compositions, in particular compositions for skin and hair, according to the present invention will contain from 0.0001% to 2 wt%, preferably from 0.001 to 1 wt%, preferably from 0.005% to 0.8 wt% , more preferably from 0.01% to 0.5% by weight of the Lespedeza capitata extract by weight of the dry extract relative to the total weight of the composition.

The Lespedeza capitata extracts used in the compositions according to the invention advantageously have a dry extract content of 1% to 10%, preferably 1% to 5% by weight, based on the total weight of the extract.

In a particular embodiment, the Lespedeza capitata extract is the only active agent used in the hair and/or scalp composition. Active agent means a therapeutic or prophylactic agent for diseases of the skin and hair covered by the invention.

The extract according to the invention or the cosmetic or dermatological composition according to the invention can also be used in combination with an alopecia treatment such as finasteride or minoxidil and/or in combination with substances useful for good hair structure, such as, for example, sulfur-containing proteins, vitamin B6 and various minerals such as zinc and/or magnesium.

The extract or composition of the invention may also be combined with at least one antibacterial agent, antifungal agent, anti-inflammatory agent, and mixtures thereof.

The extract according to the invention or the cosmetic or dermatological composition according to the invention can be used by an individual who has had a micrograft.

Preferably, the composition according to the invention has a light texture, also providing optimal penetration without making the hair or scalp greasy. From the very first application, the hair will regain strength and vitality.

Therefore, the invention also relates to the dermatological composition according to the invention for its use in the treatment and/or prevention of at least one disease of the skin and hair.

The composition according to the invention for its use in the treatment and/or prevention of at least one skin and hair disease is characterized in that at least one skin and hair disease is selected from alopecia, seborrheic dermatitis of the scalp, erythema of the scalp, seborrhea of the scalp and their combinations.

The composition according to the invention for the treatment and/or prevention of at least one disease of the skin and hair is characterized in that the alopecia is selected from the group consisting of androgenetic alopecia, reactive alopecia, postmenopausal alopecia and aerated alopecia.

Another aspect of the present invention relates to a dermatological composition containing the extract of Lespedeza capitata according to the invention, for its use in the prevention and/or treatment of alopecia.

In a particular embodiment, the alopecia is selected from the group consisting of androgenetic alopecia, postmenopausal alopecia, reactive alopecia, and aerated alopecia.

According to another aspect, the invention relates to the cosmetic use of an extract according to the invention in accordance with the method described above, or a cosmetic or dermatological composition according to the invention in accordance with the method described above, for hair and/or scalp care to limit hair loss, and/ or to enhance hair growth and/or to increase the density of hair follicles and/or to obtain more hair coverage and/or to enhance follicle regeneration and/or to reduce seborrhea of the scalp and/or to reduce oiliness of the hair.

In another aspect, the invention relates to a method for cosmetically treating the hair and/or scalp to limit hair loss and/or to enhance hair growth and/or to increase the density of hair follicles and/or to obtain more hair coverage and/or to to enhance follicle regeneration, and/or to reduce scalp seborrhea, and/or to reduce oily hair, comprising administering an extract according to the invention to a person in need thereof. This administration may be oral or topical.

The invention thus relates to the cosmetic use of the Lespedeza capitata extract according to the invention to limit hair loss and/or to enhance hair growth and/or to increase the density of hair follicles and/or to obtain more hair coverage and/or to enhance regeneration follicles, and / or to reduce seborrhea of the scalp, and / or to reduce oily hair.

Alternatively, the invention relates to the cosmetic use of the cosmetic composition according to the invention for hair and/or scalp care to limit hair loss and/or to enhance hair growth and/or to increase the density of hair follicles and/or to obtain more hair coverage, and/or to enhance the regeneration of follicles, and/or to reduce seborrhea of the scalp, and/or to reduce oily hair.

The invention also relates to a cosmetic method for hair and/or scalp care to limit hair loss and/or to enhance hair growth and/or to increase the density of hair follicles and/or to obtain more hair coverage and/or to enhance regeneration follicles, and/or to reduce seborrhea of the scalp, comprising administering to an individual in need thereof an effective amount of an extract according to the invention in accordance with the method described above, or a cosmetic or dermatological composition according to the invention in accordance with the method described above.

In another aspect, the invention relates to a cosmetic treatment of the hair and/or scalp to limit hair loss and/or to enhance hair growth and/or to increase the density of hair follicles and/or to obtain more hair coverage and/or to enhance regeneration follicles, and/or to reduce seborrhea of the scalp, and/or to reduce oily hair, including the introduction of a cosmetic composition according to the invention to a person in need of it. This administration is preferably carried out as a topical application to the hair and/or scalp.

The extract or cosmetic or dermatological composition according to the invention will preferably be applied topically and/or administered orally.

The composition according to the invention, such as described here, is characterized in that it is present in a form suitable for topical application to the scalp and/or hair.

The composition according to the invention, such as described here, is characterized in that it is present in a form suitable for oral administration.

The extract according to the invention or the cosmetic or dermatological composition according to the invention also helps to stop hair loss by lengthening its cycle, so that the hairline is preserved quantitatively and qualitatively.

In accordance with another aspect, the present invention also relates to a cosmetic hair care method for improving hair aesthetics by enhancing hair growth and/or obtaining more hair coverage and/or limiting hair loss, characterized in that it comprises applying to the hair and/or scalp with an effective amount of the Lespedeza extract according to the invention or the composition according to the invention, leaving the composition in contact with the hair and/or scalp, and optionally rinsing the hair and scalp.

In accordance with another aspect, the present invention also relates to a cosmetic hair care method for improving the aesthetics of hair by combating hair oiliness and/or restoring shine to dull hair, characterized in that it comprises applying to the hair and/or scalp an effective amount of the Lespedeza extract according to the invention or the composition according to the invention, leaving the composition in contact with the hair and/or scalp, and optionally rinsing the hair and scalp.

The following examples are intended to illustrate the invention without limiting its scope. They are given as a non-limiting example.

Examples

Preparation of the extract used in the invention.

Example 1 Extraction with ethanol 90 by reflux

2.5 kg of dried and ground aerial parts of Lespedeza capitata are extracted at reflux with stirring with 25 liters of ethanol/water 90:10 (v/v) for 1 hour in a reactor. The extract is then filtered through K900 and the solvent is evaporated to give 250 grams of green powder in 10% mass yield. The resulting dry extract contains 26% polyphenols expressed as epicatechin.

Example 2: Extraction with ethyl acetate by boiling under reflux

100 grams of dried and ground aerial parts of Lespedeza capitata are extracted at reflux with stirring with 1 liter of ethyl acetate for 1 hour in a reactor. The extract is then filtered through K900 and the solvent is evaporated to give 2 grams of a green paste in a 2% mass yield. The resulting dry extract contains 3% polyphenols expressed as epicatechin.

Example 3: Extraction with ethanol 30 at ambient temperature

20 kg of dried and ground aerial parts of Lespedeza capitata are extracted at ambient temperature with stirring with 350 liters of ethanol/water 30:70 (v/v) for 12 hours in a reactor. The extract is then filtered through K900 and the solvent is evaporated to give 3.4 grams of a brown powder in a 17% mass yield. The resulting dry extract contains 36% polyphenols expressed as epicatechin.

Example 4 Extraction with ethanol 90 using ultrasound followed by decolorization with activated charcoal.

26 grams of dried and ground aerial parts of Lespedeza capitata are brought into contact with 260 milliliters of a mixture of ethanol/water 90:10 (v/v), then extracted under the action of ultrasound (20 kHz) 3 times for 1 min with an amplitude of 100%. After filtration through K900, the extract is concentrated and decolorized with activated charcoal (0.2% w/v). After filtration, the solvent is evaporated to give 1.5 grams of a brown powder in a 6% mass yield. The resulting dry extract contains 25% polyphenols expressed as epicatechin.

Example 5 Extraction with Ethanol 90 by Reflux and Decolorization with Activated Charcoal

280 g of dried and crushed aerial parts of Lespedeza capitata are extracted by boiling under reflux with stirring with 2800 ml of a mixture of ethanol/water 90:10 (v/v) for 1 hour in the reactor. After filtration through K900, the extract is concentrated and decolorized with activated charcoal (0.2% w/v). After filtration, the solvent is evaporated to give 25 grams of extract as a brown powder in a mass yield of 9%. The resulting extract contains 25% polyphenols expressed as epicatechin.

Example 6 Extraction with ethanol 90 by reflux followed by decolorization with activated charcoal and placement in a carrier

2.5 kg of dried and ground aerial parts of Lespedeza capitata are extracted by refluxing with stirring with 25 liters of ethanol/water 90:10 (v/v) for 1 hour in a reactor. After filtration through K900, the extract is concentrated and decolorized with activated charcoal (0.2% w/v). After filtration, the extract was dried in 1,2-pentanediol to give 46 grams of extract as a dark viscous liquid. The resulting extract contains 50% 1,2-pentanediol and 12% polyphenols expressed as epicatechin.

Example 7 Extraction with ethanol 90 by reflux

100 g of dried and crushed leaves of Lespedeza capitata are extracted by boiling under reflux with stirring with 1 liter of a mixture of ethanol/water 90:10 (v/v) for 1 hour in a reactor. The extract is then filtered through K900, the solvent is evaporated to give 17 grams of a green powder in a 17% mass yield. The resulting dry extract contains 28% polyphenols expressed as epicatechin.

Pharmacological study

Example 8 Inhibition of DKK1 Synthesis and Release by Dermal Papilla Cells by Extract of Aerial Parts of Lespedeza capitata.

This study aims to evaluate the effect of the extract of the present invention on the production and secretion of DKK1 by human hair follicle dermal papilla cells and to evaluate the dermatological and cosmetic value of this extract as a hair loss prevention agent. DKK1 has been shown to be a key factor in androgenetic alopecia (see Kwack et al, VMV Reports, Volume 43, Issue 10, 2010, pp. 688-692).

Experiments were performed on dermal papilla cells of human hair follicles obtained from three different donors.

Cells are seeded on 96-well plates and cultured for 24 hours in standard culture medium. The medium is then replaced with a test medium with or without the test compound (control). Lespedeza capitata is an extract obtained in accordance with example 5 of the present invention, and this extract is tested at two concentrations, 3 μg/ml and 10 μg/ml; it is a dry extract diluted with DMSO. Twenty-four hours later, culture supernatants were collected and stored at -80°C.

DKK1 protein was quantified using Luminex (kit reference: HBNMAG-51K, Millipore) according to the manufacturer's instructions.

The raw data is analyzed using Microsoft Excel and Prism software. Statistical analysis is performed by comparison between groups using ANOVA followed by Dunnett's test.

results

Table 1 below shows the expression of DKK1 in the presence or absence of Lespedeza capitata and its percentage of inhibition.

The dermal papilla cells of the human hair follicle naturally secrete DKK1 at a level of approximately 150 pg/mL. When these cells are treated with Lespedeza capitata extract, the expression of this protein is reduced in a concentration dependent manner and significantly at concentrations as low as 3 µg/mL. In the presence of Lespedeza capitata extract at a concentration of 10 μg/ml, the decrease in DKK1 reaches 72%.

Thus, the inventors have demonstrated that the Lespedeza extract has inhibitory activity against the DKK1 target.

Therefore, this extract has an interesting activity for use as an agent for the prevention and/or treatment of hair loss.

Example 9: Inhibition of 5α-Reductase Synthesis by Dermal Papilla Cells with Extract of Aerial Parts of Lespedeza capitata.

This study aims to evaluate the effect of the extract of the present invention on 5α-reductase activity in dermal papilla cells in order to evaluate the value of this extract in preventing hair loss and/or scalp seborrhea.

Experiments were performed on dermal papilla cells of human hair follicles from two different donors. Cells are seeded (150,000 cells/well) in 24-well plates and cultured for 24 hours in standard culture medium. The medium is then replaced with test medium containing or without test compound (control) or reference product (finasteride at a concentration of 10 μM) for 24 hours. The extract of Lespedeza capitata is an extract prepared in accordance with example 5 of the present invention, and this extract is tested at two concentrations: 30 μg/ml and 100 μg/ml; it is a dry extract diluted with DMSO. The cells are then treated for 24 hours with media containing testosterone C ¹⁴ and test compound or not. The culture supernatants are then harvested for analysis of testosterone metabolism.

Steroid molecules of the supernatant are extracted with a mixture of chloroform/methanol. The organic phase is collected and the DHT is separated by thin layer chromatography using a solvent system containing dichloromethane, ethyl acetate and methanol. The chromatogram is then autoradiographed and the transformed testosterone is evaluated by densitometric analysis. The raw data is analyzed using Microsoft Excel software. Statistical analysis is performed by intergroup comparison with repeated measures ANOVA followed by Dunnett's test.

In this study, the DHT/testosterone ratio is chosen to represent 5α-reductase enzyme activity. If this ratio decreases when a compound is added, that compound is considered a 5α-reductase inhibitor. To be sure that this compound is indeed specific for this enzyme, it is important to check for a decrease in the 5α-androstane-3α,17β-diol/testosterone ratio and androstenedione/testosterone ratio, as well as an increase in the 4-androsten-3,17-dione/testosterone ratio.

results

Table 2 below shows the 5α-reductase activity (DHT/testosterone ratio) after 24 plus 24 hours incubation with the reference compound (finasteride) and Lespedeza capitata extract.

Finasteride treatment of human hair follicle dermal papilla cells shows a very strong reduction in the conversion of testosterone to DHT (decrease in the DHT/testosterone ratio). These results were expected and allow the test to be validated.

In the presence of Lespedeza capitata extract, the DHT/testosterone ratio decreases in a concentration dependent manner; this decrease (which reaches 60%) is statistically significant at 100 µg/ml Lespedeza capitata. Although the activity at 30 μg/ml was not qualified as statistically significant, it is still indicative.

Thus, the inventors have clearly demonstrated that the Lespedeza extract has 5α-reductase inhibitory activity.

Thus, this extract of Lespedeza capitata shows interesting activity for its use as an agent for the prevention and/or treatment of skin and hair diseases, in particular hair loss and/or seborrhea of the scalp and related diseases.

Example 10: Inhibition of metabolic activity and/or proliferation of dermal papilla cells by extract of aerial parts of Lespedeza capitata.

The dermal papilla is located at the base of the hair follicle and is deeply anchored in the skin tissue. It has long been known that dermal papilla cells play a key role in hair follicle growth. The highly vascularized dermal papilla delivers nutrients to the hair and provides the molecular information needed for epithelial cell proliferation and differentiation to produce new hair every cycle. Studies have shown that in healthy human hair follicles and in rat models of injury-induced mustache regeneration, there is a correlation between dermal papilla size (as well as dermal papilla cell count and hair shaft size). This correlation holds true in the context of progressive hair loss, in which the size of hair follicles and hair shafts consistently decrease with hair growth cycles. Moreover, the increase in the number of dermal papilla cells in the follicle may occur in part due to the recruitment of new cells to the dermal papilla, but the proliferation of dermal papilla cells may also contribute to this increase.

This study is aimed at evaluating the effect of the extract according to the present invention on the metabolic activity and/or proliferation of dermal papilla cells in order to determine the value of this extract in the field of preventing hair loss.

Experiments were performed on dermal papilla cells of human hair follicles obtained from three different donors.

Cells are seeded on 96-well plates and cultured for 24 hours in standard culture medium. The medium is then replaced with test medium (Follicle Dermal Papilla Cell Basal (HFDPC) Medium, Ref C-26500 from Promocell) with or without test compound (control). The same medium group supplemented with 5% supplemented medium (Follicle Dermal Papilla Cell Growth Medium SupplementPack, Ref C-39620, Promocell) acted as a positive control. Lespedeza capitata extract is an extract prepared in accordance with example 2 of the present invention, and this extract is tested at two concentrations: 3 μg/ml and 10 μg/ml; it is a dry extract diluted with DMSO. Seventy-two hours later, culture supernatants were collected and stored at -80°C.

results

Table 3 shows the effect of Lespedeza extract on metabolic activity and cell proliferation.

Treatment of dermal papilla cells with the supplemented medium causes a significant activation of cell proliferation and/or metabolic activity. This result confirms this model. 3 µg/ml of Lespedeza capitata extract causes a statistically significant increase in cell proliferation and/or metabolic activity. This activation does not reach significance at a concentration of Lespedeza capitata of 10 μg/ml, no doubt because of the great variability in results, but nevertheless it at least indicates an effect of the extract.

Thus, the inventors have clearly demonstrated that the Lespedeza extract has a stimulating activity in relation to cell proliferation and/or metabolic activity. Thus, this Lespedeza capitata extract has an interesting action as it promotes hair growth by strengthening and stimulating the life cycle of the hair, as well as promoting hair regeneration.

Example 11: Study of the influence of aerial parts of Lespedeza capitata on the inhibition of human JAK1, JAK2 and JAK3.

JAK-STAT is a transduction signaling pathway regulating growth, survival, differentiation, and resistance to pathogens. This pathway mediates the action of cytokines, interferons, and growth factors. In mammals, the JAK family consists of four members: JAK1, JAK2, JAK3, and TYK2. In a study done on mouse hair follicles, the JAK-STAT pathway was shown to be dynamically regulated in the hair cycle; the JAK-STAT pathway is upregulated during the catagen and telogen phases and downregulated early in the anagen phase. Moreover, it has been demonstrated in mice that topical treatment during the telogen phase with inhibitors of the JAK-STAT pathway, including tofacitinib (JAK1/3>JAK2>TYK2) and ruxolitinib (JAK1/2>TYK2>JAK3), resulted in a rapid return to the beginning of the phase. anagen (Harel et al., 2015 Sci. Adv.1, e1500973). The same study also showed that JAK-STAT inhibition in human hair follicles increased ex vivo hair growth. These data suggest that the JAK-STAT signaling pathway may be a novel target for hair growth stimulation.

The purpose of this example is to evaluate whether the extract according to the present invention can inhibit the JAK-STAT signaling pathway. This inhibition is assessed on human recombinant JAK1, JAK2 or JAK3 proteins.

Methods

Lespedeza capitata extract is an extract obtained in accordance with example 5 of the present invention. This extract is diluted with DMSO and tested at various concentrations (0.1-1000 μg/ml). Positive controls (tofacitinib and ruxolitinib) are also evaluated in this test.

Test products are incubated with human recombinant JAK1, JAK2 or JAK 3 protein with reaction buffer (Tris/HCl for JAK1, MOPS (3-morpholino-1-propane sulfonic acid) for JAK2 and JAK3), EDTA and certain peptide substrates. The phosphorylation reaction is then initiated by adding a mixture of magnesium acetate and radiolabeled ATP (45 μM for JAK1 and JAK2 and 10 μM for JAK3). After incubation for 40 minutes at ambient temperature, the reaction is stopped by adding phosphoric acid. Four washes with phosphoric acid and one with methanol were performed to elute small molecules, including labeled ATP. Finally, the radioactivity of the particular phosphorylated substrate is measured. Compounds are tested in three separate experiments and duplicates are made for each experiment.

IC ₅₀ values in μg/ml for each recombinant JAK1, JAK2 and JAK3 protein in relation to the tested products are presented below in Table 4:

The results obtained with the reference compounds (ruxolitinib and tofacitinib) are as expected. Indeed, these two compounds strongly inhibit JAK-STAT activity (97% to 100% maximum inhibition at very low IC ₅₀ values, about a nanogram per milliliter). Ruxolitinib exhibits greater affinity for JAK1 or JAK2 than for JAK3. Tofacitinib has a greater affinity for JAK1 or JAK3 than for JAK2. These expected results support this test.

The extract of the aerial part of Lespedeza capitata shows strong JAK1 inhibition (100% maximal inhibition) with an IC ₅₀ value of 24.8 µg/mL, strong JAK2 inhibition (100% maximal inhibition) with a slightly higher IC ₅₀ value of 146.5 µg/mL, and also strong inhibition of JAK3 (100% maximal inhibition) with an IC ₅₀ value of 3.9 μg/ml.

This test demonstrates that Lespedeza capitata extract induces inhibition of tyrosine kinase activity (with a high affinity for JAK3), exhibiting pharmacological activity of interest in promoting hair growth.

Example 12: Confirmation of the inhibitory activity of the extract of the aerial part of Lespedeza capitata against the JAK-STAT pathway at the cellular level

The purpose of this example is to confirm the inhibitory activity of Lespedeza capitata extract on the JAK-STAT signaling pathway in a cell model, follicular keratinocytes of the outer epithelial sheath of the hair follicle (outer root sheath (ORS) model). This model uses the interleukin IL-6 to activate the JAK1/2-STAT3 pathway by activating IL-6 and GP130 receptors, both of which are expressed in the epithelial lining of the hair follicle. IL-6 is a cytokine that acts as an inhibitor of the hair growth cycle. Its overexpression in a transgenic mouse model results in delayed hair growth. Moreover, in androgenetic alopecia, IL-6 will be overexpressed in dermal papilla cells under the influence of androgens (Kwack et al., 2012, J Invest Dermatology 132 (1): 43-9). IL-6 has also been reported to delay the growth of hair follicles in humans.

Method

This study is carried out on follicle keratinocytes of the outer epithelial sheath of the hair follicle (outer root sheath (ORS) model). Keratinocytes are cultured in 96-well plates in the appropriate medium (CnT-PR from CellNTech). Twenty-four hours later, the cells are washed with alkaline phosphate buffer (PBS) and the medium is changed to CnT-PR-H (standard maintenance medium for primary human keratinocytes). After 48 hours, cells are treated for 1 hour with test substance (Lespedeza capitata extract at 30 and 100 μg/ml, diluted in DMSO) and reference compounds (ruxolitinib and tofacitinib at 5 μM, both diluted in DMSO). The tested extract of Lespedeza capitata is an extract prepared in accordance with example 5 of the present invention. Then IL-6 stimulation is carried out at a concentration of 100 ng/ml for 15 minutes.

All conditions were tested on cells from η equal to 3 different donors. Lespedeza capitata extracts are tested in n equal to 3 different experiments, control conditions, stimulation and with reference compounds are carried out in n equal to 6 experiments. For each experiment, data is obtained in triplicate.

JAK activity is determined by assessing the degree of phosphorylation of the STAT3 protein. Statistical analysis is performed by a parametric test after checking the normality and equivalence of the variance, otherwise choose a non-parametric test. results

The results are shown below in Table 5.

Treatment of keratinocytes with IL-6 causes a significant increase in the degree of STAT3 phosphorylation. This increase reaches 16% and is statistically significant (p less than 0.05). The reference compounds (ruxolitinib and tofacitinib tested at 5 μM) significantly reduced the extent of STAT3 phosphorylation by 85% and 104%, respectively, p less than 0.05 for ruxolitinib and p less than 0.01 for tofacitinib. These expected results support this test.

Lespedeza capitata extract at both concentrations tested significantly (p less than 0.01 for both concentrations) and very markedly inhibited the extent of STAT3 phosphorylation induced by IL-6 stimulation.

Thus, the inventors demonstrate the value of the extract of the aerial parts of Lespedeza capitata for stimulating hair regrowth.

Claims (11)

1. Use of Lespedeza capitata extract obtained after extraction with a water-alcohol mixture or alkyl esters for the treatment or prevention of alopecia, seborrheic dermatitis of the scalp, erythema of the scalp, seborrhea of the scalp and combinations thereof. 2. Use according to claim 1, wherein the Lespedeza capitata extract is obtained from one or more parts of the Lespedeza capitata plant selected from roots and aerial parts such as stems, branches, leaves, fruits, seeds and/or flowers. 3. Use according to claim 1, wherein the extract of Lespedeza capitata is obtained after extraction with an ethanol/water mixture. 4. Application according to any one of paragraphs. 1-3, where the extract of Lespedeza capitata contains from 2 to 40 wt.% of the total amount of polyphenols in relation to the weight of the dry extract. 5. The use of Lespedeza capitata extract obtained after extraction with a water-alcohol mixture or alkyl esters for the treatment or prevention of androgenetic alopecia, reactive alopecia, postmenopausal alopecia and aerated alopecia. 6. The use of a cosmetic composition containing as an active ingredient an effective amount of an extract of Lespedeza capitata as defined in any one of paragraphs. 1-4 with at least one cosmetically acceptable excipient for hair and/or scalp care. 7. Use according to claim 6, wherein said composition contains from 0.0001 to 2 wt.%, preferably from 0.001 to 1 wt.% of Lespedeza capitata extract, by weight of dry extract relative to the total weight of the composition. 8. Use according to claim 6 or 7, wherein said composition is in a form suitable for topical application to the scalp and/or hair. 9. Application according to any one of paragraphs. 6-8, wherein said composition is in a form suitable for oral administration. 10. The use of a dermatological composition containing as an active ingredient an effective amount of an extract of Lespedeza capitata as defined in any one of paragraphs. 1-4 with at least one dermatologically acceptable excipient for the treatment or prevention of alopecia, scalp seborrheic dermatitis, scalp erythema, scalp seborrhea, and combinations thereof. 11. The use of a dermatological composition containing as an active ingredient an effective amount of an extract of Lespedeza capitata as defined in any one of paragraphs. 1-4, with at least one dermatologically acceptable excipient, for the treatment or prevention of androgenetic alopecia, reactive alopecia, postmenopausal alopecia, and aerated alopecia.