Non-induced dedifferentiated Lavandula angustifolia plant cells, their extracts and cosmetic uses thereof

Abstract

The present invention relates to non-induced dedifferentiated plant cells of a plant of the species Lavandula angustifolia, or an extract thereof, and also to a cosmetic composition comprising same, its use, and a cosmetic treatment method comprising applying said composition to the skin to improve and / or strengthen the barrier function and also to improve skin moisturization.

Description

[Technical field]

[0001] The present invention relates to the field of skin care.

[0002] The present invention relates to non-induced dedifferentiated plant cells of a plant of the species Lavandula angustifolia, to extracts thereof, and to cosmetic compositions containing same.

[0003] The present invention also relates to a cosmetic method for treating the skin, intended to improve the barrier function, comprising at least one step consisting of applying to the skin at least one composition as defined above.

[0004] In particular, the composition of the present invention is intended to improve and / or strengthen the barrier function of the skin.Furthermore, the present invention finds application in moisturizing the skin, improving skin softness, and improving and / or reducing the microrelief of the skin.The present invention also finds application in treating dry skin. [Background technology]

[0005] Human skin is composed of two compartments: a deeper compartment, the dermis, and a superficial compartment, the epidermis.

[0006] The dermis serves as the solid support for the epidermis. It is also the component that delivers nutrients. The dermis is composed primarily of fibroblasts and extracellular matrix, itself composed primarily of collagen, elastin and a substance known as ground substance, which are synthesized by fibroblasts. Leukocytes, mast cells and tissue macrophages are also found therein. The dermis also contains blood vessels and nerve fibers.

[0007] The epidermis is in contact with the external environment.

[0008] The natural human epidermis is composed mainly of three types of cells: keratinocytes, which make up the majority, melanocytes and Langerhans cells.

[0009] The cells that make up the epidermis are separated by intercellular lipid regions.

[0010] Each of these cell types contributes, by its unique function, to the essential role that skin plays in the body. In particular, keratinocytes undergo a continuous and directional maturation process, leading to the formation of keratinocytes from keratinocytes present in the basal layer of the epidermis. Keratinocytes are fully keratinized dead cells composed of keratinocytes at the terminal stage of their differentiation.

[0011] During differentiation, the phospholipids, whose role consists in generating the fluid structure of the cell membranes of the viable layers of the epidermis, are gradually replaced by a mixture mainly composed of fatty acids, cholesterol and sphingolipids (ceramides). These lipids, organized as specific lamellar liquid crystalline phases, form the intracellular cement of the stratum corneum and are essential for the exchange of water and the barrier function of the epidermis. Thus, the lipid lamellar structure of the lipid regions of the epidermis and keratinocytes participates in the barrier function of the epidermis.

[0012] The skin therefore constitutes a barrier against external aggressions, in particular chemical, mechanical or infectious aggressions, and in this regard, a certain number of defensive reactions take place on it against environmental factors (climate, UV rays, tobacco, etc.) and / or xenobiotic agents (e.g. microorganisms).

[0013] This property, known as the barrier function, is carried out primarily by the outermost layer of the epidermis, the keratinized layer known as the stratum corneum.

[0014] It is clear that the quality and balance of the skin and mucosal barriers depends on complex endogenous biological mechanisms involving numerous growth factors, adhesion molecules, hormones and enzymes of lipid metabolism.

[0015] Thus, disorders of the skin barrier can occur in the presence of external aggressors such as irritants (detergents, acids, bases, oxidants, reducing agents, concentrated solvents, gases or toxic fumes), mechanical stresses (surface friction, impact, abrasion, tearing, dust or particle radiation, shaving or depilation), thermal or climatic imbalances (cold, dryness, UV radiation), xenobiotics (undesirable microorganisms, allergens) or internal aggressors such as psychological stress.

[0016] The following people may be particularly affected by such a disturbance of the barrier function due to external aggressions: - people with "fragile" or "delicate" and sensitive skin that becomes rapidly imbalanced during large amplitude fluctuations in temperature or relative humidity (e.g. baby skin); - humans with "embrittled" skin, including in particular: - in people with a reduced protective hydrolipid membrane composed of sweat, sebum and natural moisturising factors, as is the case in people over 60 years of age, and especially in very elderly people (at least 75 years of age); - Humans with altered composition of hydrolipid membranes; - People who have a lowered reactivity threshold due to neurological hyperactivity; therefore, these skin types often manifest these sensations and clinical signs much more rapidly than other skin types: these are people with sensitive skin.

[0017] Also included may be humans with "attacked" skin, for example shaved skin.

[0018] Impairments in the skin barrier function may be reflected in particular by impaired moisturization, loss of skin elasticity, impaired complexion radiance and a rough appearance of the skin or impaired microrelief thereof.

[0019] It is then appropriate to seek to increase epidermal differentiation in order to strengthen the barrier function of the skin.

[0020] It is therefore desirable to improve and / or strengthen the skin barrier function, in particular in order to: - Overcoming the problem of moisturizing the skin, especially the mucous membranes, and especially treating dry skin; - improving skin elasticity, - maintaining and / or improving the radiance of the complexion; - Preventing and / or treating skin roughness or skin microrelief disorders.

[0021] To counteract the imbalance in barrier function, particular attention should be paid to active agents of natural origin, in particular dedifferentiated plant cells and also to extracts thereof.

[0022] Dedifferentiated plant cells arose from the work of Haberland in 1902. For the last 40 years, plant cell cultures have been used for the production of interesting metabolic products or for the propagation of exactly the same plants (somatic embryogenesis). This plant biotechnology is based on the concept of cytotopotency: "any plant cell is capable of differentiating and regenerating another individual identical to the one from which it is derived". Dedifferentiated plant cells are plant cells derived from an organ (leaf, stem, rhizome, petal, etc.) that are placed in culture and lose their organ specificity, especially their leaf, stem, rhizome or petal specificity, and are potentially capable of generating a whole plant again.

[0023] Undifferentiated plant cells are the equivalent of true plant stem cells, which are derived from meristematic plant cells and have no organ-specific biological past.

[0024] For example, US Pat. No. 5,399,633 and US Pat. No. 5,499,643 describe anti-aging or antioxidant compositions containing undifferentiated plant cells derived from the cambium of Panax ginseng or from plants of the genus Taxus.

[0025] It is a known practice from the prior art to use dedifferentiated plant cells in cosmetic products, mainly in the form of extracts, due to the properties recognized for said plant cells.

[0026] WO 02 / 06336 describes inter alia a cosmetic composition comprising ground material of induced dedifferentiated plant cells from Lavandula angustifolia and Vitex negundo having antioxidant properties. [Prior art documents] [Patent documents]

[0027] [Patent Document 1] International Publication No. 2009 / 151302 Brochure [Patent Document 2] KR2009-0118877 specification [Patent Document 3] European Patent No. 1485064 Summary of the Invention [Problem to be solved by the invention]

[0028] Therefore, there is a need to identify new technological solutions for improving and / or enhancing the barrier function of the skin.

[0029] In particular, there is a need to propose new active agents for improving and / or strengthening the skin's protection against external aggressions, for improving skin moisturization, skin softness and complexion radiance, and / or for reducing the roughness or microrelief of the skin. [Means for solving the problem]

[0030] The present invention aims, inter alia, to meet these needs.

[0031] In particular, the inventors have demonstrated that non-induced dedifferentiated plant cells of a plant of the species Lavandula angustifolia, or an extract thereof, are capable of improving and / or enhancing the barrier function of the skin.

[0032] In particular, the lack of induction of dedifferentiated plant cells of Lavandula angustifolia makes it possible to obtain an increase in the expression of barrier function and moisturizing markers, such as those responsible for the construction of the cornified layer (SPRR1A, CNFN); or those responsible for the synthesis of glycosaminoglycans GAGs (HAS3) and epidermal regeneration (HBEGF), in terms of keratinocyte differentiation (AQP3) (shown in Comparative Example 3b below).

[0033] To the best of the inventor's knowledge, there are no documents that refer to the production of non-induced de-differentiated cell lines of plants of the species Lavandula angustifolia, or extracts thereof, having the specific cosmetic properties described herein.

[0034] According to one of its first aspects, the present invention relates to non-induced dedifferentiated plant cells of a plant of the species Lavandula angustifolia, or an extract thereof.

[0035] A second subject of the invention relates to a cosmetic composition comprising, in a physiologically acceptable medium, said cells and / or extracts according to the invention.

[0036] The present invention also relates to the cosmetic use of non-induced dedifferentiated plant cells of a plant of the species Lavandula angustifolia or an extract thereof for improving and / or enhancing the barrier function of the skin.

[0037] According to another aspect, the present invention also relates to the cosmetic use of non-induced dedifferentiated plant cells of a plant of the species Lavandula angustifolia, or an extract thereof, for improving and / or strengthening the protection of the skin against external aggressions.

[0038] The present invention further relates to the cosmetic use of non-induced dedifferentiated plant cells of a plant of the species Lavandula angustifolia, or an extract thereof, for improving skin moisturization, for preventing and / or treating roughness or microrelief, and / or for improving the radiance of the complexion, and / or for improving skin suppleness.

[0039] In addition, the present invention relates to the cosmetic use of non-induced dedifferentiated plant cells of a plant of the species Lavandula angustifolia, or an extract thereof, for preventing and / or treating the cosmetic signs of dry skin.

[0040] Another subject of the invention is a cosmetic skin treatment method comprising the application to the skin of a composition according to the invention to improve and / or strengthen the skin barrier function of the skin.

[0041] The present invention also relates to a cosmetic skin treatment method comprising the application to the skin of a composition according to the invention to improve and / or strengthen the protection of the skin against external aggressions.

[0042] The present invention also relates to a cosmetic skin treatment method comprising the application to the skin of a composition according to the invention to improve skin moisturization, to prevent and / or treat roughness or microrelief and / or to improve the radiance of the complexion and / or to improve the softness of the skin.

[0043] The present invention also relates to a cosmetic method for treating dry skin comprising application to dry skin of a composition according to the invention to treat the cosmetic signs of dry skin.

[0044] The process according to the invention is particularly intended for humans with dry skin, regardless of the age or type of skin of the person, or the cause of the dryness.

[0045] According to another embodiment, the composition may be intended to improve and / or strengthen the barrier function of skin chosen from weak, embrittled, challenged and / or sensitive skin.

[0046] In the context of the present invention, the composition may also be used for application to healthy skin that is or will be subjected to the external aggressions described above. In other specific cases, the composition of the present invention may be applied to the skin when clinical signs of skin barrier deficiency are present.

[0047] definition The term "cosmetic composition" means a composition that comprises a physiologically acceptable medium, ie a medium that is compatible with the skin.

[0048] The term "skin" refers to all skin of the body, preferably the skin of the face, the skin around the collar, neck, arms and forearms or more preferably the skin of the face, especially the skin of the forehead, nose, cheeks, chin and areas around the eyes.

[0049] The term "uninduced cells" refers to cells that have not undergone induction.

[0050] The term "induction" as used herein refers to the triggering by an exogenous elicitor of an under-expressed metabolic pathway in another organism or cell, or the awakening in another organism or cell of a silent metabolic pathway.

[0051] The term "elicitor" as used herein means a molecule or organism capable of inducing an under-expressed metabolic pathway in another organism or awakening a silent metabolic pathway in another organism. Many elicitors are known to those skilled in the art, including biotic elicitors such as jasmonates and their derivatives, as well as abiotic elicitors such as temperature, pH, UV, gases such as CO2, or osmotic shock.

[0052] The term "process that does not include an induction step" refers to a process in which de-differentiated cells are not placed in contact with an elicitor as defined above.

[0053] The term "placed in contact" as used herein means incubation of the dedifferentiated plant cells and the inducer in the same culture medium.

[0054] The term "cosmetic signs of dry skin" refers to the sensation of tightness and / or tension in the skin, the scaly appearance of the skin, and / or the appearance of a rough feeling of the skin.

[0055] Uninduced dedifferentiated Lavandula angustifolia plant cells and their extracts For the purposes of the present invention, the term "dedifferentiated plant cell" refers to any cell type derived from a plant organ of the species Lavandula angustifolia and obtained under specific in vitro culture conditions, which no longer exhibits any specialized characteristics and is capable of any differentiation according to its genome under the influence of induction and is capable of generating alone the whole plant of the plant from which it originated. Such a cell is capable of survival alone and has no dependencies on other cells.

[0056] De-differentiated plant cells are distinct from the undifferentiated plant cells naturally present in the plant.

[0057] For the purposes of the present invention, the term "dedifferentiated plant cells" refers to seeds obtained by in vitro culture derived from organs of plants of the species Lavandula angustifolia, which are capable of differentiating under the influence of induction into any cell type (totipotency) or into several cell types (pluripotency) during specific embryogenesis or division and / or of obtaining novel characteristics of specialized cells.

[0058] Under normal conditions, plant cells express about 20% of their genome, and the remaining 80% is expressed only in response to certain environmental conditions. In vitro culture of these cells under specific culture conditions allows the cells to be "reprogrammed" and thus to utilize parts of this genome that are not expressed in the whole plant. Some compounds that are difficult to obtain by extraction from plants become more available in cell culture.

[0059] Thus, advantageously, the dedifferentiated plant cells of the present invention allow access to novel compounds not present in whole plants or allow significantly increased expression of molecules that are known, but rare in whole plants.

[0060] The present invention also relates to non-induced dedifferentiated cells of a plant of the species Lavandula angustifolia, obtainable by the process detailed herein, contained in the examples.

[0061] The inventors have shown that the process according to the invention makes it possible to obtain a dedifferentiated cell line of a plant of the species Lavandula angustifolia and to culture the cells of this line in the form of dedifferentiated cells for a very long period of time without any detectable alteration of their morphology and without any detectable modification of their properties, in particular with regard to the barrier function and moisturization.

[0062] The uninduced dedifferentiated plant cells of the invention may be obtained from any part of a plant of the species Lavandula angustifolia, or from cells of said plant.

[0063] The term "plant part" means either one or more whole organs of a plant, such as leaves, axes, flowers, petals, sepals, seeds or rhizomes, or one or more fragments of said plant organs, either in vivo cultured or in the wild. Thus, one or more leaves, or one or more leaf fragments of the plant Lavandula angustifolia, may be used to produce the non-induced dedifferentiated plant cells of the invention.

[0064] The term "in vivo culture" refers to any of the conventional types of culture, ie, in soil, outdoors or in a greenhouse, or outside the soil.

[0065] The term "in vitro culture" refers to all techniques known to those skilled in the art for artificially obtaining plants or plant parts in a reproducible manner.

[0066] Preferentially, according to the invention, plants obtained from in vivo culture, and even more preferentially parts of plants obtained from in vivo culture are used.

[0067] Preferably, the uninduced dedifferentiated plant cells of the invention are obtained from at least one leaf or leaf portion of a plant of the species Lavandula angustifolia.

[0068] More preferably, the non-induced dedifferentiated plant cells of the invention are obtained from a plant of the white cultivar of Lavandula angustifolia, which cultivar is specifically registered with the French Pharmacopea.

[0069] The white variety of Lavandula angustifolia plant may be selected from Lavandula angustifolia "Hidcote White", Lavandula angustifolia "Silbermoewe", Lavandula angustifolia "Alba", Lavandula angustifolia "Arctic Snow" (sold by the supplier le jardin du pic vert), or white Lavandula angustifolia cultivated in Drome, France.

[0070] In a highly preferred embodiment, a white cultivar of Lavandula angustifolia is obtained from Drome (France).

[0071] Quite preferably, the non-induced dedifferentiated plant cells according to the invention are obtained using leaves of plants of the white cultivar of Lavandula angustifolia as starting product.

[0072] Advantageously, a culture medium suitable for obtaining the non-induced dedifferentiated plant cells of the invention comprises hormones naturally occurring in plants, and said culture medium does not comprise an elicitor.

[0073] According to a preferred embodiment, the non-induced dedifferentiated cells of a plant of the species Lavandula angustifolia are cultured by the following steps: i. Providing one or more plant parts of the species Lavandula angustifolia, in particular one or more whole leaves or one or more leaf fragments; ii. cultivating the plant part provided in step i. in a culture medium containing at least one plant hormone to generate dedifferentiated cells; and iii. harvesting the dedifferentiated cells obtained at the end of step ii.; iv. Optionally, a process comprising extracting the de-differentiated cells collected in step iii., The dedifferentiated cells are obtained by a process that does not include a step of inducing the dedifferentiated cells.

[0074] Preferentially, in step ii., plant parts, in particular whole leaves or leaf fragments, are cultivated in anexic form (free from any biological contaminants).

[0075] In step ii. of the process, the plant parts are cultured in a suitable culture medium containing at least one plant hormone, also known as a phytohormone, in certain embodiments, the culture medium contains multiple plant hormones, for example two or three plant hormones.

[0076] The phytohormones contained in the culture medium may be selected from auxins, cytokinins, gibberellins and mixtures thereof.

[0077] Auxins suitable for use in the present invention may be selected from IAA (indole-3-acetic acid), IBA (indolebutyric acid), phenylacetic acid and NAA (naphthaleneacetic acid) and mixtures thereof. Preferentially, 2,4-D (2,4-dichlorophenoxyacetic acid) is excluded from the auxins suitable for use in the present invention, since it is an artificial compound.

[0078] Most particularly suitable cytokinins for use in the present invention may be selected from kinetin (N-(furan-2-ylmethyl)-7H-purin-6-amine), zeatin (2-methyl-4-(7H-purin-6-ylamino)but-2-en-1-ol), and benzyladenine (N-benzyl-7H-purin-6-amine) and mixtures thereof.

[0079] The gibberellin may be selected from gibberellins A3, A1, A12 and mixtures thereof.

[0080] In step ii. of the process, the plant hormones or phytohormones are preferentially selected from indole-3-acetic acid, indolebutyric acid, phenylacetic acid, naphthaleneacetic acid, kinetin, zeatin, benzyladenine, gibberellic acid and gibberellins A1, A3 and GA3.

[0081] In a preferred embodiment, the plant or phytohormones are preferentially selected from naphthaleneacetic acid and kinetin.

[0082] Preferably, the culture medium in step ii. is an aqueous medium.

[0083] For the purposes of the present invention, the term "aqueous medium" means a medium comprising water and, optionally, an additional aqueous solvent that is particularly compatible with the cultivation of plant cells.

[0084] According to certain embodiments, the culture medium of step ii. comprises: - at least one plant hormone, such as 1-naphthaleneacetic acid, kinetin and mixtures thereof; and - optionally at least one salt in hydrated form selected from NH4NO3; KNO3; CaCl2, for example CaCl2·2H2O; MgSO4; KH2PO4; MnSO4, for example MnSO4·4H2O; ZnSO4, for example ZnSO4·7H2O; KI; Na2MoO4, for example Na2MoO4·2H2O; CuSO4, for example CuSO4·5H2O; Na2EDTA, for example Na2EDTA·2H2O; FeSO4, for example FeSO4·7H2O; and mixtures thereof; and at least one carbon source, preferably chosen from mono-, oligo- or polysaccharides and mixtures thereof; in particular chosen from glucose, fructose, sucrose and mixtures thereof, preferably sucrose; and optionally at least one compound selected from myo-inositol, nicotinic acid, pyridoxine HCl, thiamine HCl, and mixtures thereof; and optionally polyvinylpyrrolidone The aqueous medium comprises:

[0085] Advantageously, the amount of carbon source present in the culture medium is between 5 and 40 g / l, preferably between 10 and 30 g / l, and better still, the amount of carbon source present in the culture medium is 20 g / l.

[0086] According to a preferred embodiment, the culture medium is an aqueous medium containing at least NH4NO3; KNO3; CaCl2·2H2O; MgSO4; KH2PO4; MnSO4·4H2O; ZnSO4·7H2O; KI; Na2MoO4·2H2O; CuSO4·5H2O; Na2EDTA·2H2O; FeSO4·7H2O; myo-inositol; nicotinic acid; pyridoxine HCl; thiamine HCl; naphthalene acetic acid; kinetin; sucrose and optionally polyvinylpyrrolidone.

[0087] Advantageously, the culture medium contains the following components: 1200-2000 mg / l NH4NO3; 1500-2100 mg / l KNO3; 300-500 mg / l CaCl2·2H2O; 150-200 mg / l MgSO4; 153-187 mg / l KH2PO4; 10-30 mg / l MnSO4·4H2O; 5-10 mg / l ZnSO4·7H2O; 0.001-0.91 mg / l KI; 0.001-0.30 mg / l Na2MoO4·2H2O; 0.01-0.05 mg / l CuSO4· 5H2O; 10.5-50 mg / L Na2EDTA·2H2O; 10-30 mg / L FeSO4·7H2O; 70-150 mg / L myo-inositol; 0.3-0.6 mg / L nicotinic acid; 0.4-0.6 mg / L pyridoxine; 0.08-0.15 mg / L thiamine; 0.001-11 mg / L naphthalene acetic acid; 0.001-1 mg / L kinetin; 10-30 g / L sucrose; and water, optionally, 0.1-0.5 g / L polyvinylpyrrolidone.

[0088] The concentrations of the various components contained in the culture medium are given as mass concentrations.

[0089] The cultivation in step ii. is advantageously carried out at a temperature range of 20 to 30°C, preferably 24 to 28°C, and even better still 27°C.

[0090] The culturing of step ii. is advantageously carried out in a culture containing a partial pressure of O2 of about 8% to 80%, for example a partial pressure of O2 of 30%.

[0091] The process of step ii. may be carried out by batch, fed-batch (semi-continuous) or continuous fermentation techniques, preferably batch fermentation techniques.

[0092] After cultivation in a suitable medium, in step iii. the non-induced dedifferentiated plant cells of the present invention can be harvested, for example by filtration, and lyophilized or can be subjected to an extraction process.

[0093] According to a particular embodiment, the culturing step ii. is carried out for a period of between 6 and 14 days, and preferentially between 6 and 10 days.

[0094] The present invention also relates to a cell line isolated by the applicant and deposited according to the Budapest Treaty on 28 February 2019 under the reference DSM 33100 with the Deutsche Sammlung von Mikroorganismen und Zellkulturen (DSMZ) [German Collection of Microorganisms and Cell Cultures].

[0095] According to the invention, fresh or freeze-dried dedifferentiated plant cells obtained in step iii, or an extract thereof recovered at the end of step iv., formulated in a composition that stabilises them, may be used.

[0096] According to a particularly preferred embodiment of the present invention, extracts of non-induced dedifferentiated plant cells may be used.The present invention essentially relates to active extracts of non-induced dedifferentiated plant cells with respect to obtaining effects on improving and / or strengthening barrier function and / or improving moisturization.The activity of the extracts of the present invention may be evaluated by various experimental protocols, in particular those detailed in the examples presented below.

[0097] In step iv., any extraction method known to a person skilled in the art may be used to prepare an extract of non-induced dedifferentiated plant cells according to the invention. The process according to the invention may also comprise a step of adding a water-miscible organic solvent between step iii. and the extraction step iv. Step iv. may comprise a step of adding a water-miscible organic solvent.

[0098] Extracts suitable for use in the present invention may include aqueous extracts, organic extracts, or extracts obtained by mixing water with at least one organic extracting solvent that is miscible with water in all proportions, such as aqueous-alcoholic extracts, optionally in the form of a dry extract obtained, in particular, by evaporation, freeze-drying or atomization.

[0099] Preferably, the extract of non-induced dedifferentiated cells of a plant of the species Lavandula angustifolia is selected from: - an aqueous extract of the intracellular media, an aqueous-alcoholic extract of the intracellular media, or an organic extract of the intracellular media, said extracts optionally being in the form of a dry extract; or - an aqueous extract of the insoluble components of the cells, an aqueous-alcoholic extract of the insoluble components of the cells or an organic extract of the insoluble components of the cells (said extract is optionally in the form of a dry extract; said insoluble components of the cells are selected from insoluble intracellular components, pectocellulose walls, cell membranes and mixtures thereof; preferably pectocellulose walls and / or cell membranes).

[0100] Even more preferably, the extract of non-induced dedifferentiated cells of a plant of the species Lavandula angustifolia is selected from: - an aqueous extract of the intracellular media, an aqueous-alcoholic extract of the intracellular media, or an organic extract of the intracellular media, said extracts optionally being in the form of a dry extract; or - aqueous, aqueous-alcoholic or organic extracts of pectocellulose walls and / or cell membranes obtained by enzymatic double digestion, preferably obtained after a first step of enzymatic digestion using one or more carbohydrases followed by a second step of enzymatic digestion using one or more proteases.

[0101] The term "aqueous extract" refers to an extract obtained by means of an aqueous extraction solvent.

[0102] The term "aqueous extraction solvent" means water or a solvent that consists of water.

[0103] The term "mixture of water and at least one water-miscible organic solvent" refers to water / organic solvent mixtures in all proportions.

[0104] The term "aqueous-alcoholic extract" refers to an extract obtained by a mixture of water and ethanol in all proportions.

[0105] The term "organic extract" refers to an extract obtained by means of an organic extraction solvent.

[0106] Among the water-miscible organic extraction solvents, there may be mentioned ethanol, isopropanol, propylene glycol, 1,3-propanediol and mixtures thereof.

[0107] The term "dry extract" refers to an extract containing less than 5% by weight of solvent, preferably less than 3% by weight of solvent, better still less than 1% by weight of solvent, and in a particular embodiment, 0% of solvent. The solvent may be water, an organic solvent or a mixture thereof. Dry extracts suitable for use in the present invention may be obtained, for example, by freeze-drying, atomization or evaporation.

[0108] In a first embodiment, an extraction method suitable for obtaining an extract according to the invention may comprise: i. a first step of splitting non-induced dedifferentiated plant cells in an extraction solvent, said extraction solvent being selected from aqueous and organic extraction solvents or a mixture of water and at least one water-miscible organic solvent; said first step of splitting is carried out, for example, by using a high-pressure homogenizer, in particular at room temperature, ii. A second step of removing components in the cell suspension, preferably by centrifugation followed by a filtration step, so as to recover the concentrated extraction solvent obtained from the first step.

[0109] This extraction method notably leads to the extraction of the intracellular medium of non-induced dedifferentiated plant cells of plants of the species Lavandula angustifolia.

[0110] The term "components of the suspension of non-induced dedifferentiated plant cells of a plant of the species Lavandula angustifolia" means components that are insoluble in the extraction solvent of step i. at a temperature of 25° C. These components may in particular be insoluble intracellular components, pectocellulose walls, cell membranes and mixtures thereof.

[0111] The term "concentrated extractant" refers to an extractant containing intracellular components that are soluble at a temperature of 25°C of uninduced dedifferentiated plant cells of a plant of the species Lavandula angustifolia.

[0112] Step i. of splitting the non-induced dedifferentiated cells can be carried out by any technique known to the person skilled in the art, for example by using ultrasound, or by increasing the temperature to produce a heat-induced split, or by using mechanical constraints on the cells, such as shear, by using ultrasound, or by applying high pressure. Preferably, the splitting of the cells leading to the extract of the invention is carried out by applying high pressure, preferably a pressure between 500 bar and 2000 bar, better still between 1000 bar and 2000 bar, in particular using a high pressure homogenizer.

[0113] In step i., when the extraction solvent is a mixture of water and at least one water-miscible organic extraction solvent, said organic extraction solvent can be ethanol or 1,3-propanediol. Preferably, the weight ratio of [water / organic extraction solvent] is between 1 / 2 and 1 / 1. In addition, the weight ratio of [cells / aqueous+organic extraction solvent] is between 1 / 2 and 0.9 / 1. In addition, a step of drying the extract can be carried out at the end of step ii. by concentrating to dryness, in particular using a rotary evaporator, optionally followed by resuspension in an aqueous solvent and / or optionally followed by a freeze-drying step.

[0114] In step i., when the extraction solvent is an organic extraction solvent, said organic extraction solvent may be ethanol or 1,3-propanediol. In addition, the [cell / organic extraction solvent] mass ratio is 1 / 2 to 0.9 / 1. In addition, a step of drying the extract may be carried out at the end of step ii. by concentrating to dryness, in particular using a rotary evaporator, followed by optional resuspension in an aqueous solvent, followed optionally by a freeze-drying step.

[0115] Step ii. of removing the components in suspension of said cells in suspension may be carried out by any technique known to the person skilled in the art, preferably by centrifugation at 6000×G to 12000×G, better still 8000×G to 10000×G, followed by a step of filtration of the supernatant.

[0116] Centrifugation may be carried out for 20 to 40 minutes.

[0117] Centrifugation may be carried out at a temperature of 4°C.

[0118] Filtration may be carried out by any filtration method known to the person skilled in the art, preferably using a cellulose filter, in particular a 0.1 μm to 1 μm filter, for example a 0.7 μm, in particular a 0.7 μm Whatman filter.

[0119] The extraction method may also include an optional step of sterilizing the concentrated extraction solvent resulting from the second step, for example by autoclaving at 115°C to 130°C, in particular 121°C.

[0120] In a first variant, irrespective of the extraction solvent used, the non-induced dedifferentiated plant cells of a plant of the species Lavandula angustifolia used in step i. are fresh cells, i.e. cells that have not undergone a drying step prior to step i., in particular by evaporation, freeze-drying or atomization.

[0121] In a second variant, irrespective of the extraction solvent used, the non-induced dedifferentiated plant cells of a plant of the species Lavandula angustifolia used in step i. are cells which have undergone a drying step prior to step i., in particular by evaporation, freeze-drying or atomization.

[0122] In a second embodiment, another extraction method suitable for obtaining an extract according to the invention may comprise, instead of the second step ii. above, a second step of removing the concentrated extraction solvent obtained from the first step, so as to recover the components in the cell suspension, for example by centrifugation under the conditions described above.

[0123] Following this second step, the following steps may be performed: i. optionally adjusting the pH; ii. carrying out a first enzymatic digestion of components in said suspension of cells; iii. optionally adjusting the pH; iv. performing a second enzymatic digestion of components in said cell suspension, said second enzymatic digestion being performed with one or more enzymes other than those used for said first enzymatic digestion; v. Inactivating enzyme digestion; vi. optionally adjusting the pH; vii. Optionally, clarifying by centrifugation; viii. Optionally, freeze-drying or atomizing the supernatant obtained from step vii.

[0124] In a preferred embodiment, the components in the cell suspension are subjected to a centrifugation step prior to the double digestion step, and the double digestion step is carried out on the pellet obtained from the centrifugation.

[0125] A possible pH adjustment prior to the step of enzymatic double digestion is optional. If it is carried out, the pH is modified by adding an aqueous solution of an organic or inorganic acid or base. The purpose of this adjustment is to adjust the pH of the pellet, if necessary, to a value that corresponds to the optimal working pH of the enzymes used in the digestion step.

[0126] Preferably, a pH adjustment is carried out, preferably by adding a solution of an organic or inorganic acid to a pH of 3 to 6, more preferentially 3.5 to 4.5, in particular 4. Preferably, the pH is adjusted by a citrate / phosphate buffer at a concentration of 10 to 100 mM, for example 50 mM, or any other composition capable of buffering to the preferred pH of the carbohydrase, from 3 to 6, preferentially 4. The ratio of the components in the cell / buffer solution suspension is between 2 / 100 and 30 / 100, preferably 10 / 100.

[0127] Alternatively, the adjustment is carried out by adding a solution of an organic or inorganic base, preferably to a pH of 6.5 to 8.5, more preferentially 7.5 to 8.5, in particular a pH of 8. Preferably, the pH is adjusted by a potassium hydroxide or sodium hydroxide solution at a concentration of 0.1 M to 3 M, preferably 1 M, or any other composition capable of buffering to the preferential pH of the protease, 6.5 to 8.5, preferentially 8. The ratio of the components in the cell / buffer solution suspension is 2 / 100 to 30 / 100, preferably 10 / 100.

[0128] In a particular embodiment, the enzymatic double digestion of the components of the cell suspension is carried out using a carbohydrase and a protease enzyme.

[0129] According to a first variant, the digestion is carried out successively, after optional adjustment of the pH to a given value, by the action of a carbohydrase enzyme and then by the action of a protease enzyme.

[0130] According to a second variant, digestion is carried out successively, after optional adjustment of the pH to a given value, by the action of a protease enzyme and then by the action of a carbohydrase enzyme.

[0131] According to one embodiment, an inactivation step, in particular a heat inactivation, occurs between the two enzymatic digestion steps. According to a first variant of this embodiment, the optionally centrifuged components in the suspension of cells are optionally adjusted to a given pH, then first treated with a protease enzyme, then subjected to an inactivation step and treated with a carbohydrase enzyme. According to a second variant of this embodiment, the optionally centrifuged components in the suspension of cells are optionally adjusted to a given pH, then first treated with a carbohydrase enzyme, then subjected to an inactivation step and treated with a protease enzyme.

[0132] According to a preferred embodiment of the invention, the term "carbohydrase enzymes" refers to enzymes such as cellulases (endoglucanases, cellobiohydrolases, β-glucosidases), hemicellulases, xylanases, pectinases and mixtures thereof, such as the enzyme Viscozyme L® sold by the company Novozyme. Viscozyme L is a mixture of carbohydrases isolated from the genus Aspergillus. The amount of carbohydrase used is between 0.01% and 5% by weight, more preferentially between 2% and 3% by weight, for example 2.5% by weight. The working temperature is between 40° C. and 60° C., preferentially 50° C.; the working pH is between 3 and 6 (limits included), preferably 4, and the treatment time is between 30 minutes and 24 hours, preferably 90 minutes, in particular with stirring at 50 rpm and 250 rpm, for example 150 rpm. The ratio of the enzyme solution / buffer suspension of the cellular components ranges from 0.5 / 100 to 20 / 100, preferably 2.5 / 100.

[0133] The term "protease enzyme" refers to enzymes of the classification EC 3.4, such as, for example, exoproteases, endoproteases and mixtures thereof.

[0134] According to a preferred embodiment of the invention, the term "protease enzyme" refers to the enzyme Alcalase 2.4L sold by the company Novozyme. Alcalase 2.4L is a broad spectrum endoprotease, purified and isolated from Bacillus licheniformis. The amount of protease used is between 0.01% and 5% by weight, more preferentially between 2% and 3% by weight, for example 2.5% by weight. The working temperature is between 40°C and 60°C, preferentially 50°C; the working pH is between 6.5 and 8.5 (limits included), preferentially between 7.5 and 8.5, for example 8, and the treatment time is between 30 minutes and 24 hours, preferably 90 minutes, in particular with stirring at 50 rpm and 250 rpm, preferably 150 rpm. The enzyme solution / buffer suspension ratio of the components of the cells ranges from 0.5 / 100 to 20 / 100, preferably 2.5 / 100.

[0135] According to a preferred embodiment, digestion of the components in the cell suspension is carried out sequentially by the action of Viscozyme L® sold by the company Novozyme and then by the action of Alcalase 2.4L sold by the company Novozyme.

[0136] According to a particular embodiment, the enzymatic digestive inactivation is a heat inactivation which may be carried out for 10 to 30 minutes, preferably 15 minutes.

[0137] In a particular embodiment, the inactivation of the enzymatic digestion is carried out at a constant temperature between 80°C and 90°C, preferably at 90°C.

[0138] The term "about 15 minutes" refers to a period of 15 minutes ± 5 minutes.

[0139] In a particular embodiment, after the inactivation step is completed, the pH of the reaction medium is adjusted to 7 with an organic or inorganic acid or an organic or inorganic base, preferably an organic or inorganic acid.

[0140] The hydrolysis product resulting from the enzymatic double digestion may be subjected to a centrifugation step, in particular at 10000×G for 30 minutes at 4° C. The supernatant obtained after this centrifugation step is dried by any drying technique known to the person skilled in the art, preferably by freeze-drying or spraying.

[0141] The extract so obtained may be called a hydrolysate of the insoluble components of said cells, in particular a hydrolysate of the pectocellulose walls and / or of the cell membranes.

[0142] Another extract suitable for use in the present invention may be a dry extract of non-induced dedifferentiated plant cells of a plant of the species Lavandula angustifolia, in particular obtained from an extract such as those described in the first and second embodiments above, by any conventional drying method such as evaporation, freeze-drying or atomization. A powder is thus obtained which may be used directly or mixed with a suitable solvent before use.

[0143] The non-induced dedifferentiated plant cells of the invention, or extracts thereof, may also be used in the form of a lyophilizate. Such a lyophilizate may be obtained by any lyophilization method known to those skilled in the art.

[0144] In principle, freeze-drying consists in removing water from a liquid, pasty or solid product by the combined action of cold and vacuum. When water in the solid state is heated at very low pressure, it sublimes, i.e. it goes directly from the solid state to the gaseous state. The water vapor (or the vapor of any other solvent) leaves the product, which is captured by refrigeration, using a cooler or a trap. With this technique it is possible to preserve the volume and the appearance of the processed product. This technique can be carried out using a freeze dryer.

[0145] Freeze drying involves at least two steps: freezing, sublimation, and optionally secondary drying.

[0146] Freezing consists of bringing a substance to a temperature of -20°C to -80°C very rapidly, so as to seal in the water in the form of ice where it was in a liquid state.

[0147] Sublimation consists of removing the "free" water. Heat is supplied to the product and the ice undergoes sublimation under a vacuum that is in the range of 100 μbar to 1000 μbar but can vary greatly from one product to another. Depending on the product and the needs of the production, the temperature can be changed during the cycle. The water vapor is captured in a "trap" or "cooler" and the dehydration of the product proceeds continuously. If the majority of the water has undergone sublimation, the product loses about 80% to 90% of its water.

[0148] The secondary drying consists in removing the trapped water from the product. In this step the vacuum is high, between 5 μbar and 100 μbar. After this step the product is dried, in particular between 90% and 99%, for example 95%.

[0149] After recovering the cells from the culture medium, for example by filtration through gauze of controlled porosity (approximately 50 μm), the cells are frozen at low temperatures, preferably between −20° C. and −80° C. The frozen cells are then subjected to a step of ice sublimation under a vacuum ranging from 100 to 1000 μbar, followed by a step of secondary drying under a vacuum ranging from 5 μbar to 100 μbar.

[0150] The lyophilized uninduced dedifferentiated plant cells may be supplemented with water or a water containing mixture prior to use.

[0151] Cosmetic Composition Advantageously, the non-induced dedifferentiated plant cells and / or extracts thereof are used in an amount representing a solids content of between 0.01% and 40% by weight relative to the total weight of the composition containing them, preferentially in an amount representing a solids content of between 0.01% and 20% by weight relative to the total weight of the composition and preferably in an amount representing a solids content of between 0.01% and 10% by weight relative to the total weight of the composition.

[0152] The composition according to the invention contains a physiologically acceptable medium.

[0153] Such a physiologically acceptable medium may in particular consist of water and, optionally, a physiologically acceptable organic solvent selected from: lower alcohols, for example containing from 1 to 8 carbon atoms, in particular from 1 to 6 carbon atoms, such as ethanol, isopropanol, propanol or butanol; polyethylene glycols containing from 6 to 80 ethylene oxide units; polyols, such as propylene glycol, isoprene glycol, butylene glycol, glycerol, sorbitol or 1,3-propanediol.

[0154] It may also be an anhydrous medium, especially an oily medium containing oils and / or fatty substances other than oils.

[0155] When the physiologically acceptable medium is an aqueous medium, it is compatible with the skin, preferably having a pH in the range of 3-8, and better still in the range of 4-7.

[0156] When the composition comprises an aqueous or aqueous-alcoholic medium, it is possible to add a fatty (or oily) phase to this medium.

[0157] The compositions according to the invention are in particular compositions intended for topical application to the skin.

[0158] Thus, the compositions according to the invention containing dedifferentiated plant cells of a plant of the species Lavandula angustifolia or an extract thereof as defined above may be in any of the conventionally used presentation forms for topical application, in particular in the form of an aqueous, aqueous-alcoholic or oily solution, an oil-in-water (O / W), water-in-oil (W / O) or multi-layer (triple layer: W / O / W or O / W / O) emulsion, in the form of an aqueous or oily gel, in the form of a liquid, pasty or solid anhydrous product, or in the form of a dispersion of a fatty phase in an aqueous phase using small globules that are polymeric nanoparticles such as nanospheres and nanocapsules, or lipid vesicles of ionic and / or non-ionic type. These compositions are prepared according to the usual methods.

[0159] Furthermore, the compositions used according to the invention may have a certain degree of fluidity and may have the appearance of a white or colored cream, a pomade, a milk, a lotion, a serum, a paste or a mousse. They may optionally be applied to the skin in aerosol form. They may also be in solid form, for example in the form of a stick.

[0160] When the composition used according to the invention comprises an oily phase, it preferably contains at least one oil. It may also contain other fatty substances.

[0161] Oils that may be used in the compositions of the present invention may include the following examples: - Hydrocarbon oils of animal origin; Hydrocarbon oils of vegetable origin; in particular synthetic esters and ethers of fatty acids, such as oils of the formulae R1COOR2 and R1OR2, in which R1 represents a fatty acid residue containing from 8 to 29 carbon atoms and R2 represents a branched or unbranched hydrocarbon-based chain containing from 3 to 30 carbon atoms; - Linear or branched hydrocarbons of mineral or synthetic origin; - fatty alcohols containing from 8 to 26 carbon atoms, - Fluoro oils that are partly hydrocarbon-based, and / or silicone oils, such as volatile or non-volatile polymethylsiloxanes (PDMS) with linear or cyclic silicone chains, which can be liquid or pasty at room temperature; - and mixtures thereof.

[0162] In the above list of oils, the term "hydrocarbon-based oil" means any oil that contains mainly carbon and hydrogen atoms and, optionally, ester, ether, fluoro, carboxylic acid and / or alcohol groups.

[0163] Other fatty substances which may be present in the oily phase are, for example, fatty acids containing from 8 to 30 carbon atoms, waxes, silicone resins and silicone elastomers.

[0164] These fatty substances may be selected in various ways by a person skilled in the art in order to prepare a composition having the desired properties, for example in the sense of consistency or texture.

[0165] According to a particular embodiment of the invention, the composition according to the invention is a water-in-oil (W / O) or oil-in-water (O / W) emulsion, in particular an O / W emulsion. The proportion of the oily phase of the emulsion may range from 5% to 80% by weight, preferably from 5% to 50% by weight, relative to the total weight of the composition. The oils, emulsifiers and coemulsifiers used in the composition in the form of an emulsion are chosen from those conventionally used in cosmetics or dermatology. The emulsifiers and coemulsifiers are generally present in the composition in a proportion ranging from 0.3% to 30% by weight, preferably from 0.5% to 20% by weight, relative to the total weight of the composition. The emulsion may contain lipid vesicles.

[0166] The emulsions generally contain at least one emulsifier, used alone or in a mixture, chosen from amphoteric, anionic, cationic or nonionic emulsifiers, which are selected in a suitable manner depending on the emulsion to be obtained (W / O or O / W emulsion).

[0167] The cosmetic composition of the invention may contain auxiliaries that are common in the cosmetic field, such as hydrophilic or lipophilic gelling or thickening agents, such as xanthan gum, hydrophilic or lipophilic active agents, preservatives, antioxidants, solvents, fragrances, fillers, screening agents, odor absorbents, dyes and salts. The amounts of these various auxiliaries are those conventionally used in the field in question, for example between 0.01% and 20% of the total mass of the composition. Depending on their nature, these auxiliaries may be introduced into the fatty phase, the aqueous phase and / or into the lipid globules. [Brief description of the drawings]

[0168] [Figure 1] FIG. 2 HPLC chromatogram of an extract of non-induced de-differentiated plant cells of the species Lavandula angustifolia obtained according to Example 1a (according to the invention) versus an extract of UV-induced de-differentiated plant cells of the species Lavandula angustifolia obtained according to Example 2 (not according to the invention). DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0169] Working Example Example 1 Example 1a - Preparation of an aqueous extract of the intracellular medium of non-induced dedifferentiated cells of Lavandula angustifolia (cultivated in Drome, France) according to the invention Recovery of the airborne fraction in calcium hypochlorite (50 g / l or 40 g / l containing 60% active chlorine) for 30 min and subsequent decontamination. Successive rinsing in three sterile osmosed water baths (5 min / bath).

[0170] Cutting the aerial part to the graft and collecting only the leaves.

[0171] The leaves are cultured in the culture medium shown in Table 1 below on agar in light and darkness.

[0172] [Table 1]

[0173] Following successive subcultures in the presence of a culture medium, dedifferentiated plant cells were obtained that could be cultured in a fermenter. The parameters used to control the cultivation in the bioreactor were: - T°:27℃; - Aeration: pO2 30%, controlled by sterile air intrusion and / or by stirring, while avoiding any shear stress on the cells. - Agitation: 150 rpm

[0174] The batch production lasts for about 10 days. The resulting culture medium is then separated from the dedifferentiated cells by filtration through gauze with a porosity of 50 μm.

[0175] At the end of the filtration step, the cells obtained were milled using a high-pressure homogenizer (2000 bar) in the presence of water at a cell / water mass ratio of 1 / 1.The particles in the suspension were then removed by filtration at 10000×G for 30 minutes at 4° C., followed by filtration of the supernatant using a 0.7 μm Whatman cellulose filter to obtain an aqueous extract of the intracellular media.

[0176] The aqueous extract so obtained is dried by freeze-drying under the following conditions: freezing the sample at -40°C, followed by sublimation by placing under vacuum (<1 mbar) at 20°C, then secondary drying achieved at a low pressure of 100 μbar by excluding air injection into the system; introduction into the dry extract.

[0177] Example 1b - Preparation of an alcoholic extract of the intracellular medium of non-induced dedifferentiated cells of Lavandula angustifolia according to the invention At the end of the filtration step through 50 μm gauze of Example 1, the cells obtained were milled using a high-pressure homogenizer (2000 bar) in the presence of ethanol: [cell / solvent] ratio 1 / 1. The particles in the suspension were then removed by filtration at 10000×G for 30 minutes at 4° C., followed by filtration of the supernatant using a 0.7 μm Whatman cellulose filter to obtain an alcoholic extract of the intracellular media.

[0178] The extract thus obtained was purified in two steps: by concentration on a rotary evaporator at -50°C followed by redissolution in water; then - Drying by freeze-drying under the following conditions: freezing the sample at -40°C, followed by sublimation by placing under vacuum (<1 mbar) at 20°C, then secondary drying achieved at a low pressure of 100 μbar by excluding air injection into the system; introduction into the dry extract.

[0179] Example 1c - Preparation of membranes of pectocellulose wall hydrolysates according to the invention and of non-induced dedifferentiated cells of Lavandula angustifolia At the end of the filtration step through 50 μm gauze of Example 1, the cells obtained are milled using a high-pressure homogenizer (2000 bar) in the presence of water at a [cell / water] mass ratio of 1 / 1. The particles or debris in the suspension are then collected by centrifugation at 10000×G for 30 minutes at 4° C.

[0180] The first step of enzymatic hydrolysis of the pellets obtained is carried out using carbohydrases, which allows the hydrolysis of cellulosic compounds: - Suspension of wall debris in a pH 4 buffered solution (50 mM citrate / phosphate buffer) to debris / buffer solution ratio of 10 / 100. - Hydrolysis of the wall debris by adding an enzyme solution of carbohydrase (Viscozyme L from Novozyme) in a ratio of enzyme solution / buffered debris suspension ranging from 2.5 / 100. This mixture is placed at the optimum working temperature of the carbohydrase at 50°C, with stirring at 150 rpm for 90 minutes.

[0181] A second step of protein hydrolysis with a protease is then carried out: - Adjustment of the pH of the reaction mixture to the preferred pH of the protease of 8 by simple addition of a concentrated base solution such as potassium hydroxide. - Hydrolysis of the wall debris by adding an enzyme solution of protease (Alcalase 2.4L from Novozyme) in a ratio of enzyme solution / buffered debris suspension ranging from 2.5 / 100. This mixture is placed at the optimum working temperature of the protease at 50°C, with stirring at 150 rpm for 90 minutes.

[0182] To complete these hydrolysis steps, the hydrolysis product is subjected to enzyme inactivation at 90° C. for 15 min, followed by separation of the particles remaining in suspension from the hydrolysis product obtained from the above two hydrolysis steps by centrifugation (10,000×G, 4° C., 30 min).

[0183] To concentrate and preserve the supernatant obtained from the centrifugation, it is dried by lyophilization under the following conditions: freezing the sample at -40°C, followed by sublimation by placing under vacuum (<1 mbar) at 20°C, then secondary drying achieved at a low pressure of 100 μbar by excluding air injection into the system.

[0184] The resulting product is called a cell wall hydrolysate; it is a compound rich in sugars.

[0185] Example 2 - Preparation of an aqueous extract of UV-induced dedifferentiated cells of Lavandula angustifolia other than that of the present invention During their cultivation in the production medium (see Table 1, culture medium composition), induce the Lavandula angustifolia cell lines 10 days after inoculation by UV light (280-400 nm) using four Philips CLEO performance sunlamps (40-0-14 / 2.6) positioned at a distance of 50 cm under direct illumination on the cells for 15 h.

[0186] This induction procedure does not apparently form any impurities during cell culture. After the end of the culture, i.e. 24 hours after induction, the plant cells are filtered through gauze with a porosity of 50 μm to remove the remaining culture medium. Thus, at the end of the filtration step, a fresh biomass is obtained; said cells obtained are milled in a high-pressure homogenizer at 2000 bar in the presence of water to extract the intracellular medium of the cells. The extract thus obtained is dried by freeze-drying under the following conditions: freezing the sample at -40°C, followed by a sublimation step by placing it under vacuum (<1 mbar) at 20°C, then a secondary drying achieved at a low pressure of 100 μbar by excluding the injection of air into the system.

[0187] Example 3 Example 3a: Analysis of the UPLC chromatographic profiles of the extracts obtained according to Example 1a (according to the invention) and Example 2 (other than according to the invention) A) Materials and Methods To perform a pseudo-quantitative analysis of extracts 1a and 2, they are prepared in identical concentrations and the solutions are doped with a standard absorbance in the UV range: caffeine.

[0188] The first step consists of dissolving the sample in water to obtain a solution at 5 g / L. The solution so obtained is gently heated and sonicated for 30 minutes.

[0189] The second step consists of preparing an aqueous solution of caffeine at a concentration of 0.17 g / L.

[0190] The final step consisted of mixing 1.8 mL of the sample solution (cell extract 1a or 2) with 0.2 mL of the caffeine solution, homogenizing the mixture, then filtering through a 0.45 μm filter and then a 0.2 μm filter.

[0191] Analyses are performed on an Acquity UPLC Additol H-class system (Waters) containing a diode array detector, a corona detector (CAD) and a single quadrupole mass spectrometer.

[0192] Chromatography Systems - Acquity UPLC BEH Shield RP18 Column - Length 50mm - Inner diameter 2.1mm - Column volume 0ml - Particle diameter 1.8μm - Mobile phase: A = water 0.1% HCOOH; B = ACN (acetonitrile) 0.1% HCOOH - Flow rate=0.5mL / min - Injection volume 2μL - Temperature: T column = 30°C; T sample = 20°C

[0193] Slope 1 is shown in Table 2 below.

[0194] [Table 2]

[0195] detection - Diode array detector (DAD): records chromatograms in the range 200-700 nm. - Corona Charged Aerosol Detector (CAD): Set the pressure at P = 35.1 psi, which corresponds to a nitrogen flow rate set at D = 1.21 bar. - Mass Spectrometer (MS): Coupling of HPLC to mass spectrometry is performed by a single quadrupole mass spectrometer equipped with an electrospray ionization (ESI) source; the mass spectrometer operates simultaneously in positive and negative ionization mode over a 100-2000 amu mass range.

[0196] B) Result The two chromatograms of extracts 1a and 2 were overlaid (see FIG. 1).

[0197] Among the 23 target compounds: - One compound is reduced when cells are induced by UV light, - Six compounds were not affected by induction. - 11 compounds were enriched when cells were induced with UV light. - 5 compounds are newly synthesized (not detectable in the non-induced sample at the end of the method).

[0198] The two extracts 1a and 2 therefore differ from each other in their composition.

[0199] Example 3b: Evaluation of the effect on barrier function / moisture retention markers of an extract outside the scope of the invention of UV-induced dedifferentiated cells of Lavandula angustifolia obtained according to Example 1a (according to the invention) versus an extract of non-induced dedifferentiated cells of Lavandula angustifolia obtained according to Example 2 (other than according to the invention). A) Materials and Methods cytotoxicity Healthy human epidermal keratinocytes were seeded in 96-well culture plates and then cultured in culture medium at 37°C and 5% CO2 for 24 hours. The medium was then replaced with culture medium containing or not containing test compound (control) (8 concentrations were tested), and then incubated for 24 hours. All conditions were performed in duplicate. After the end of incubation, cell viability was measured by a standard test that measures mitochondrial activity by Alamar Blue®.

[0200] Analysis of gene expression related to barrier function / moisture retention by RT-qPCT Healthy human epidermal keratinocytes (NHEK) were seeded in 48-well culture plates and then cultured in culture medium at 37°C and 5% CO2 for 3 days, with the culture medium being refreshed after the first 24 hours of culture. After the end of incubation, the culture medium was replaced with test medium containing or not containing the test compound (control) (supplemented with 1.5 mM CaCl2), and the cells were then incubated for 24 hours. All conditions were performed in duplicate.

[0201] After the treatment, the culture medium was removed and the cells were rinsed twice with PBS (without CaCl2, without MgCl2). Total RNA was then isolated using a magnetic bead extraction kit according to the supplier's recommendations (MagMAXTM-96 Total RNA Isolation Kit, Ambion). RNA quantification and its quality control were analyzed by Labchip GX (Perkin Elmer).

[0202] The expression of selected transcripts was analyzed by quantitative PCR in two steps. First, cDNA was reverse transcribed from RNA using the Quantitect® Reverse Transcription Kit (Qiagen) according to the supplier's recommendations. Then, quantitative PCR experiments were carried out using the LightCycler 480 Real-Time PCR System in 384-well plates (Roche) according to SYBR® Green (Roche) integration technology. The primers used are shown in Table 2 below.

[0203] [Table 3]

[0204] B) Result

[0205] [Table 4]

[0206] The extract of non-induced cells obtained according to Example 1a (extract according to the present invention) significantly promoted the expression of transcripts related to stratum corneum construction (SPRR1A, CNFN), keratinocyte differentiation (AQP3) and epidermal regeneration (HBEGF) compared to the extract of UV-induced cells according to Example 2 (extract other than the present invention).

[0207] Thus, the extract 1a of non-induced dedifferentiated cells from Lavandula angustifolia according to the invention proves to be particularly effective for preventing and treating skin dehydration and for improving and strengthening the barrier function.

[0208] Example 4 - Evaluation of moisturizing ability of isolated stratum corneum by measurement using a corneometer Tests were carried out to evaluate the moisturizing capacity of the extract of the present invention, formulated in a solvent (80% / 20% water / n-propanol) in an amount of 5% by weight based on the total weight of the composition.

[0209] This technique makes it possible to measure the dielectric capacitance of the stratum corneum (SC), which depends on the average dielectric constant of the tissue, which varies greatly with the amount of water contained in the SC.

[0210] Before / during measurements and processing, the SC samples are conditioned at 75% relative humidity and 25° C. Volume measurements are performed using a Corneometer™ (Courage & Khazaka, Germany).

[0211] The test extract, extracts 1a, 1b and 1c according to the invention, or moisturizing active agents such as glycerol were dissolved in a water / n-propanol mixture (80 / 20) and the solution was applied at 10 μl / cm on the SC. 2 followed by air drying for a total duration of 4 hours.

[0212] Measurements are taken at T0 before treatment and at Ttreat(4h) after complete drying of the treatment.

[0213] Each treatment is systematically compared to its control (solvent) and to its TO.

[0214] Measure 4-5 SC samples per treatment using at least two different batches of SCs.

[0215] For each SC sample, the variation of the corneometer signal (HCM) after treatment is first calculated: DHCMi=HCMi(Ttreat)-HCMi(T0). The mean of the DHCMi(vehicle) variation is then calculated for the control samples (treated with solvent). This mean is subtracted from all DHCMi(active agent) and DHCMi(positive control) variations to correct for systematic bias.

[0216] For each sample i, the following is determined: For vehicle (control): DHCMi(veh) = HCMiveh(Ttreat) - HCMiveh(T0) Regarding active agents: DHCMiactive agent = HCMiactive agent (Ttreat) - HCMiactive agent (T0) For the positive control (glycerol): DHCMipositive control = HCMipositive control (Ttreat) - HCMipositive control (T0)

[0217] To correct for the systematic bias associated with the solvent, the correction value DHCMicorr. active agent is considered for the active agent according to the following: DHCMicorr. active agent =DHCMiactive agent-M(veh) where M(veh) corresponds to the average of the DHCMi(veh) variation observed on n solvent control samples. [Formula 1]

number

[0218] To correct for solvent-related systematic bias, the correction value DHCMicorr. positive control is considered for the positive control as follows: DHCMicorr. positive control=DHCMipositive control-M(veh) where M(veh) is as defined above.

[0219] The DHCPCorr. positive control and DHCPCorr. active agent values ​​are then normalized by the following calculation: %Normalized=[(DHCMicorr. positive control) / (M(positive control)-M(veh))]×100 %Normalized=[(DHCMicorr. active agent) / (M(positive control)-M(veh))]×100 where M(veh) is as defined above, M(positive control) corresponds to the mean of the DHC Mi positive control variation observed on n positive control samples. [Formula 2]

number

[0220] For the extract according to the invention tested at 5% compared to glycerol at 5%, the % normalized values ​​obtained are reported in the table below.

[0221] [Table 5]

[0222] This test shows that the dielectric capacitance of the stratum corneum (SC) obtained with extracts 1a or 1b is similar to that obtained with glycerol at the same concentration, and in addition, the dielectric capacitance of the stratum corneum (SC) obtained with extract 1c is much better than that obtained with glycerol at the same concentration.

[0223] Unexpectedly, extracts 1a, 1b and 1c according to the invention allow good moisturization of the stratum corneum and therefore of the skin, the moisturizing effect proving to be similar (extracts 1a and 1b) or even greater (extract 1c) than that of glycerol.

[0224] Example 5 - Cosmetic Composition The following compositions were prepared:

[0225] [Table 6]

[0226] The above compositions were applied to the skin to enhance the barrier function and / or moisturize the skin.

Claims

1. A cosmetic composition for improving and / or enhancing the skin barrier function of the skin, comprising non-induced dedifferentiated plant cells of a plant of the species Lavandula angustifolia, or an extract thereof, in a physiologically acceptable medium, The extract of non-induced dedifferentiated cells of a plant of the species Lavandula angustifolia has the following properties: an aqueous extract of the intracellular media or an ethanolic extract of the intracellular media, said extract optionally in the form of a dry extract; or an aqueous extract of the insoluble components of said cells, said extract optionally in the form of a dry extract; said insoluble components of said cells being selected from insoluble intracellular components, pectocellulose walls, cell membranes and mixtures thereof; preferably pectocellulose walls and / or cell membranes; The composition is characterized in that it is selected from:

2. 2. The composition according to claim 1, characterized in that the non-induced dedifferentiated plant cells of a plant of the species Lavandula angustifolia, or an extract thereof, are obtained from plant material obtained from a part of a plant of the species Lavandula angustifolia, the part of the plant being one or more whole organs of the plant selected from leaves, axes, flowers, petals, sepals, seeds or rhizomes, or one or more fragments of said organs of said plant in vivo cultured or in the wild.

3. 3. The composition according to claim 1 or 2, characterized in that the non-induced dedifferentiated plant cells of a plant of the species Lavandula angustifolia, or an extract thereof, are obtained from a part of the plant chosen from leaves or leaf fragments of the plant Lavandula angustifolia.

4. The non-induced dedifferentiated plant cells of a plant of the species Lavandula angustifolia, or an extract thereof, are subjected to the following steps: i. providing one or more parts of a plant of the species Lavandula angustifolia; ii. Cultivating the plant part provided in step i. in a culture medium containing at least one plant hormone to generate dedifferentiated cells; and iii. Harvesting the de-differentiated cells obtained at the end of step ii.; iv. Optionally, extracting the de-differentiated cells recovered in step iii. The composition according to any one of claims 1 to 3, characterized in that it is obtained by a method comprising the steps of:

5. wherein the culture medium of step ii. comprises: at least one plant hormone, such as 1-naphthaleneacetic acid, kinetin and mixtures thereof; and -NH ４ NO ３ ;KNO ３ ; CaCl ２ , e.g. CaCl ２ ・2H ２ O; MgSO ４ ;KH ２ P.O. ４ MnSO4, for example MnSO4.4H ２ O; ZnSO ４ , e.g. ZnSO ４ ・7H ２ O; K; Na ２ MoO ４ , for example Na ２ MoO ４ ・2H ２ O; CuSO ４ , e.g. CuSO ４ ・5H ２ O;Na ２ EDTA, e.g. Na ２ EDTA 2H ２ O; FeSO ４ , for example FeSO ４ ・7H ２ O; and mixtures thereof, optionally in hydrated form; and at least one carbon source, preferably chosen from mono-, oligo- or polysaccharides and mixtures thereof; in particular chosen from glucose, fructose, sucrose and mixtures thereof, preferably sucrose; and optionally at least one compound selected from myo-inositol, nicotinic acid, pyridoxine HCl, thiamine HCl and mixtures thereof; - and optionally polyvinylpyrrolidone The composition according to claim 4, characterized in that it is an aqueous culture medium comprising:

6. The non-induced dedifferentiated plant cells of a plant of the species Lavandula angustifolia, or an extract thereof, ４ NO ３ ;KNO ３ ; CaCl ２ ・2H ２ O; MgSO ４ ;KH ２ P.O. ４ MnSO4.4H ２ O; ZnSO ４ ・7H ２ O; K; Na ２ MoO ４ ・2H ２ O; CuSO ４ ・5H ２ O;Na ２ EDTA 2H ２ O; FeSO ４ ・7H ２ 6. The composition according to any one of claims 1 to 5, characterized in that it is obtained by culturing parts of plants from the species Lavandula angustifolia in an aqueous medium containing: O; myo-inositol; nicotinic acid; pyridoxine HCl; thiamine HCl; naphthalene acetic acid; kinetin; sucrose and optionally polyvinylpyrrolidone.

7. 7. The composition according to any one of claims 1 to 6, wherein the dedifferentiated plant cells and / or extracts thereof are used in an amount representing between 0.01% and 40% by weight of solids relative to the total weight of the composition containing them, preferentially in an amount representing between 0.01% and 20% by weight of solids relative to the total weight of the composition.

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