LOTUS EXTRACT AND COSMETIC COMPOSITION

Abstract

The present invention relates to an aqueous extract of sacred lotus flowers, of the genus Nelumbo nucifera, characterized in that it is obtained by subcritical water extraction comprising the steps of: a. Recovery of the lotus flowers; b. Extraction with subcritical water, in particular in a lotus flower / water ratio of 1 / 20, preferably a ratio of 1 / 10, at a temperature ranging from 90°C to 110°C, preferably 100°C to 120°C, preferentially 100°C, and at a pressure ranging from 40 to 60 bars, preferably 50 bars; c. Addition of glycerin and a preservative system; and d. Filtration and obtaining the final extract.

Description

Title of the invention: LOTUS EXTRACT AND COSMETIC COMPOSITION FIELD OF THE INVENTION

[0001] The present invention relates to the cosmetic field and in particular to an extract of Sacred Lotus flowers, or Nelumbo nucifera, in particular an extract of Sacred Lotus flowers obtained by subcritical water extraction. The present invention also relates to compositions comprising such an extract as well as the uses of such an extract for promoting and / or improving the skin barrier function, promoting and / or improving the hydration of the epidermis, preventing and / or reducing skin aging and the formation of wrinkles and / or fine lines, promoting and / or improving epidermal renewal. STATE OF THE ART

[0002] The skin is constantly subjected to stress via factors of exogenous origin (e.g. UV radiation, temperature variations, atmospheric pollution, cigarette smoke, etc.) or endogenous origin (e.g. hormones, etc.) likely to induce signs of skin aging, which can be called extrinsic aging and intrinsic (or physiological) aging respectively. The barrier function of the skin represents the first line of defense between the body and external aggressors. It is mainly provided by the epidermis and in particular allows the maintenance of skin hydration, for example by reducing transepidermal water loss, and the protection of the skin from external aggressions.

[0003] The different functions of the skin associated with its role as a barrier are as follows:

[0004] - Physical barrier: it provides the skin with mechanical resistance, control of insensible water loss and the potential entry of external aggressors;

[0005] - Hydration: the skin's hydrolipidic film limits water loss from the epidermis and maintains skin hydration;

[0006] - Antioxidant: the skin is protected from oxidative stress thanks to a defense system which protects cells by destroying excess free radicals.

[0007] Over time, the skin can become diminished or weakened by exogenous and / or endogenous factors that can be a source of stress for the skin. These factors can lead to the reduction or weakening of the skin's barrier function. This weakening can result in the appearance of wrinkles, sagging of the skin, dehydration of the skin which can in turn cause the skin to lose its suppleness and elasticity. With age, the epidermis, which is thinner and more fragile, is also more permeable to these factors which generate stress in its cells and unbalance its natural state of homeostasis.

[0008] We are therefore looking for new anti-aging ingredients capable of strengthening the skin's barrier function and improving its hydration. We already know the use of plant-based cosmetic active ingredients to strengthen the skin's barrier function, or to limit water loss to counter dehydration. However, these plant-based cosmetic active ingredients, such as the active ingredients contained in plants, aromatic herbs and spices, are generally present at very low concentrations in plants and require a step to extract them from their plant matrix. We are therefore looking for new processes for obtaining plant-based cosmetic active ingredients and new plant extracts, in particular processes that are simpler to implement, more effective or that allow the plant-based cosmetic active ingredients to be more concentrated in the plant extract and therefore more effective when used in a cosmetic composition.

[0009] Unexpectedly, the inventors have developed a new active extract of sacred Lotus flowers capable of inducing in vitro the expression of different target genes involved in particular in the barrier function, differentiation, adhesion, cohesion and hydration of the skin.

[0010] Furthermore, the inventors have demonstrated that such an extract is capable of protecting the progenitor cells of the skin, responsible for cell proliferation and renewal of epidermal cells.

[0011] Thus, such an extract of sacred Lotus flowers has the particularity of promoting and / or improving the skin barrier function, skin hydration, skin cell renewal, preventing and / or reducing skin aging and the formation of wrinkles and / or fine lines. Statement of the invention

[0012] In the context of the present invention, the inventors have developed a new extract of sacred Lotus flowers, or Nelumbo nucifera, obtained by subcritical water extraction. This extract contributes in particular to the significant and effective improvement of barrier / differentiation, adhesion / cohesion, hydration functions as well as epidermal renewal.

[0013] The present invention therefore relates to a new aqueous extract of sacred Lotus flowers, of the genus Nelumbo nucifera, characterized in that it is obtained by an extraction with subcritical water comprising the steps of: a. Recovery of lotus flowers; b. Extraction with subcritical water, in particular in a lotus flower / water ratio of 1 / 20, preferably a ratio of 1 / 10, at a temperature ranging from 90°C to 110°C, preferably 100°C to 120°C, preferably 100°C, and at a pressure ranging from 40 to 60 bars, preferably 50 bars; c. Addition of glycerin and a preservative system; and d. Filtration and obtaining the final extract.

[0014] The present invention also relates to a composition, in particular a cosmetic composition, comprising in a physiologically acceptable medium an aqueous extract of sacred Lotus flowers according to the invention.

[0015] The invention further relates to a non-therapeutic cosmetic process for caring for and / or making up the skin and / or lips, comprising the application to said skin and / or said lips, in particular the skin of the face and / or neck, of at least one cosmetic composition according to the invention. The cosmetic process according to the invention makes it possible in particular to promote and / or improve the skin barrier function, to promote and / or improve the hydration of the epidermis, to prevent and / or reduce skin aging and the formation of wrinkles and / or fine lines, to promote and / or improve epidermal renewal.

[0016] The present invention finally relates to the non-therapeutic cosmetic use of the aqueous extract of sacred Lotus flowers according to the invention, as an active ingredient for promoting and / or improving the skin barrier function, promoting and / or improving the hydration of the epidermis, preventing and / or reducing skin aging and the formation of wrinkles and / or fine lines, promoting and / or improving epidermal renewal. DETAILED DESCRIPTION OF THE INVENTION Definitions

[0017] By “keratin material” is meant the skin and / or its appendages, and the lips, in particular the skin of the face and / or neck and / or body and the lips.

[0018] By "sacred lotus extract" or "lotus extract" is meant an extract of a plant of the genus Nelumbo, preferably an extract of Nelumbo nucifera.

[0019] In the remainder of the description, reference will be made indifferently to “lotus”, “sacred lotus” or “Nelumbo nucifera”.

[0020] By "skin and / or lips" according to the invention, we mean in particular healthy skin and / or lips, that is to say not presenting any troubles or disorders which would be a pathological condition ('unhealthy' subjects, suffering from a pathology). Plant material

[0021] The present application relates to an extract of Sacred Lotus.

[0022] The Sacred Lotus, also called Oriental Lotus or Nelumbo nucifera, is a perennial aquatic plant native to temperate and tropical Asia. It is now found in many Asian countries. The extract according to the invention is preferably Obtained from the flowers of Nelumbo nucifera. Sacred Lotus flowers are distinctive for being large (up to 30 cm in diameter) and solitary. They have numerous petals, around twenty, which are pink to white in color.

[0023] Sacred Lotus flowers can be harvested at two different flowering stages: the flower in bud or the slightly open flower. The flowers in bud are reserved for religious rites and the slightly open flowers are reserved for the cosmetic industry. The flowers can be harvested slightly open and then left to dry in the open air. The petals, sepals and stamens will then be left to dry for different lengths of time so that the water contained in each element is eliminated. Sacred Lotus Extract

[0024] The present invention relates to a novel aqueous extract of sacred Lotus flowers.

[0025] Sacred Lotus extracts are commercially available. These include the Glycolysat® BGV extract of sacred Lotus UP flowers marketed by Solabia.

[0026] The new extract according to the invention is distinguished from commercial extracts by its superior effects on the improvement of the skin barrier function, the hydration of the epidermis, epidermal renewal and the reduction of skin aging, as well as by the reduction of its toxicity for the skin (due in particular to the presence of a smaller number of alkaloids).

[0027] This new extract is obtained from a sample of the Nelumbo nucifera plant, and preferably from the flowers of Nelumbo nucifera.

[0028] This extract can be chosen from the different parts of the plant or their mixtures, in particular a flower extract (petals), a sepal extract, a stamen extract or their mixture.

[0029] Advantageously, it will be an extract of the whole flowers, including the petals, the sepals and the stamen.

[0030] The lotus sample may be a sample of fresh, frozen, freeze-dried lotus, or any mixture thereof. Frozen or freeze-dried samples are advantageously obtained from fresh lotus samples. This makes it possible in particular to preserve the various active ingredients and properties of fresh lotus. The lotus sample is therefore preferably a sample of fresh, freeze-dried lotus, or a mixture of fresh and freeze-dried lotus, more preferably a sample of freeze-dried lotus.

[0031] The sacred Lotus flower extract thus obtained is characterized by its INCI name: nelumbo nucifera flower extract.

[0032] The present aqueous extract of sacred Lotus flowers is obtained by subcritical water extraction.

[0033] The term "aqueous extract of sacred lotus flowers" or "aqueous extract of lotus flowers" means an extract of sacred lotus flowers according to the invention obtained by extraction with subcritical water.

[0034] Subcritical water is a green and innovative extraction technology that allows better valorization of plant raw materials and to obtain original phytochemical profiles in comparison with conventional techniques (maceration, decoction, digestion, etc.). This technique makes it possible in particular to avoid the use of polar and semi-polar solvents of petrochemical origin. This extraction method uses the properties of water, which, in its subcritical state, sees its solvent properties modified and can solubilize weakly polar compounds. In particular, this technique makes it possible to extract molecules of medium polarities, to shorten the extraction time and to increase performance at the qualitative and quantitative level.

[0035] The aqueous extract of sacred Lotus flowers according to the invention can thus be obtained by subcritical water extraction comprising the steps of:

[0036] a. Recovery of lotus flowers;

[0037] b. Extraction with subcritical water;

[0038] c. Addition of glycerin and a preservative system; and

[0039] d. Filtration and obtaining the final extract.

[0040] According to a particular mode, the extraction with subcritical water is carried out: - in a lotus flower / water ratio of 1 / 20, preferably a ratio of 1 / 10; - at a temperature ranging from 90°C to 130°C, preferably 100°C to 120°C, preferably 100°C; and - at a pressure ranging from 40 to 60 bars, preferably 50 bars.

[0041] Optionally, the subcritical water extraction process further comprises a step of static maceration of the lotus flowers in water for 1 to 30 min, preferably 5 to 20 min.

[0042] According to a particular embodiment, the aqueous extract of sacred Lotus flowers according to the invention can be obtained by subcritical water extraction comprising the steps of:

[0043] a. 1 Recovery of lotus flowers;

[0044] a.2 Optionally, static maceration of the lotus flowers in water;

[0045] b. Extraction with subcritical water;

[0046] c. Addition of glycerin and a preservative system; and

[0047] d. Filtration and obtaining the final extract.

[0048] Optionally, the subcritical water extraction process further comprises an evaporation step to concentrate the extract.

[0049] According to another particular embodiment, the aqueous extract of sacred Lotus flowers according to the invention can be obtained by an extraction with subcritical water comprising the steps of:

[0050] a. Recovery of lotus flowers;

[0051] bl Extraction with subcritical water;

[0052] b.2 Optionally, evaporation of the extract obtained;

[0053] c. Addition of glycerin and a preservative system; and

[0054] d. Filtration and obtaining the final extract.

[0055] Advantageously, the preservative system is composed of potassium sorbate and sodium benzoate.

[0056] According to one embodiment, the aqueous extract of sacred Lotus flowers according to the invention can be obtained by subcritical water extraction comprising the steps of:

[0057] a. 1 Recovery of lotus flowers;

[0058] a.2 Optionally, static maceration of the lotus flowers in water;

[0059] bl Subcritical water extraction;

[0060] b.2 Optionally, evaporation of the extract obtained;

[0061] c. Addition of glycerin and a preservative system; and

[0062] d. Filtration and obtaining the final extract.

[0063] According to a particular embodiment, the aqueous extract of sacred Lotus flowers according to the invention is obtained by subcritical water extraction comprising the steps of:

[0064] a. Recovery of lotus flowers;

[0065] b. Extraction with subcritical water, in particular in a lotus flower / water ratio of 1 / 20, preferably a ratio of 1 / 10, at a temperature ranging from 90°C to 110°C, preferably 100°C to 120°C, preferably 100°C, and at a pressure ranging from 40 to 60 bars, preferably 50 bars;

[0066] c. Addition of glycerin and a preservative system; and

[0067] d. Filtration and obtaining the final extract.

[0068] According to another particular embodiment, the aqueous extract of sacred Lotus flowers according to the invention is obtained by an extraction with subcritical water comprising the steps of:

[0069] a. 1 Recovery of lotus flowers;

[0070] a.2 Optionally, static maceration of the lotus flowers in water for 1 to 30 min, preferably 5 to 20 min;

[0071] bl Extraction with subcritical water, in particular in a lotus flower / water ratio of 1 / 20, preferably a ratio of 1 / 10, at a temperature ranging from 90°C to 110°C, of preferably 100°C to 120°C, preferably 100°C, and at pressure ranging from 40 to 60 bars, preferably 50 bars;

[0072] b.2 Optionally, evaporation of the extract obtained;

[0073] c. Addition of glycerin and a preservative system; and

[0074] d. Filtration and obtaining the final extract.

[0075] Furthermore, according to a particular embodiment, the aqueous extract of sacred Lotus flowers according to the invention is obtained by subcritical water extraction comprising the steps of:

[0076] a. Recovery of lotus flowers;

[0077] b. Extraction with subcritical water, in particular in a lotus flower / water ratio of 1 / 10, at a temperature of 100°C, and at a pressure of 50 bars;

[0078] c. Addition of glycerin and a preservative system; and

[0079] d. Filtration and obtaining the final extract.

[0080] According to yet another particular embodiment, the aqueous extract of sacred Lotus flowers according to the invention is obtained by subcritical water extraction comprising the steps of:

[0081] al Recovery of lotus flowers;

[0082] a.2 Optionally, static maceration of the lotus flowers in water from 5 to 20 min ;

[0083] bl Extraction with subcritical water, in particular in a lotus flower / water ratio of 1 / 10, at a temperature of 100°C, and at a pressure of 50 bars;

[0084] b.2 Optionally, evaporation of the extract obtained;

[0085] c. Addition of glycerin and a preservative system; and

[0086] d. Filtration and obtaining the final extract.

[0087] In one embodiment, the aqueous sacred lotus flower extract of the invention comprises glycerin, water, a sacred lotus flower extract, and a preservative system comprising sodium benzoate and potassium sorbate.

[0088] In another embodiment, the aqueous extract of sacred lotus flowers according to the invention comprises glycerin in a content ranging in particular from 40 to 60% by weight of the aqueous extract, water in a content ranging in particular from 30 to 40% by weight of the aqueous extract, a crude extract of sacred lotus flowers in a content of crude extract ranging in particular from 10 to 25% by weight of the aqueous extract, and a preservative system comprising sodium benzoate and potassium sorbate in a content ranging in particular from 0.2 to 1% by weight of the aqueous extract.

[0089] By "crude extract" or "crude extract of lotus flowers" within the meaning of the invention, is meant an extract comprising the dry matter originating from the extraction of lotus flowers according to the invention and the constitutional water of the plant.

[0090] According to one embodiment, the crude lotus flower extract comprises 0.92% of dry lotus flower extract in crude lotus flower extract.

[0091] According to yet another embodiment, the aqueous extract of sacred lotus flowers according to the invention comprises glycerin in a content of 50% by weight of the aqueous extract, water in a content of 33.5% by weight of the aqueous extract, a crude extract of sacred lotus flowers in a crude extract content of 16% by weight of the aqueous extract, and a preservative system comprising sodium benzoate and potassium sorbate in a content of 0.5% by weight of the aqueous extract.

[0092] The INCI name of this aqueous extract of sacred Lotus flowers thus obtained is glycerin (and) potassium sorbate (and) sodium benzoate (and) water (and) nelumbo nucifera flower extract.

[0093] According to a particular embodiment, the aqueous extract of sacred Lotus flowers according to the invention further comprises at least one compound chosen from the group comprising minerals, vitamins, amino acids, polyphenols, organic acids, monosaccharide sugars, and mixtures thereof.

[0094] Preferably, the aqueous extract of sacred Lotus flowers according to the invention comprises at least one compound chosen from the group comprising minerals, vitamins, polyphenols, and mixtures thereof.

[0095] Advantageously, the minerals of the aqueous extract of sacred Lotus flowers according to the invention are selected in particular from Phosphorus (P), Zinc (Zn), Sodium (Na), Potassium (K), or a mixture thereof; the vitamins are selected in particular from Folate or vitamin B9, Niacin or vitamin B3, calcium pantothenate or vitamin B5, Pyrodoxine or vitamin B6, or a mixture thereof; the amino acid may be asparagine; the alkaloids are selected in particular from Nuciferine, methylclaurine, or a mixture thereof; the polyphenols are selected in particular from Astragalin, Kaempferol, or a mixture thereof; the organic acid may be malic acid; and the monosaccharide sugars are selected in particular from Fructose, Glucose, or a mixture thereof.

[0096] According to a particular and preferred embodiment, the aqueous extract of sacred Lotus flowers according to the invention is characterized in that it further comprises:

[0097] - from 1300 mg / kg and 2300 mg / kg of minerals, in particular from 1411 mg / kg to 2117 mg / kg of minerals, in particular selected from Phosphorus (P), Zinc (Zn), Sodium (Na), Potassium (K), or a mixture thereof, relative to the total weight of the extract;

[0098] - at least 2 mg / kg of vitamins, in particular at least 2.9 mg / kg of vitamins, in particular selected from Folate or vitamin B9, Niacin or vitamin B3, calcium pantothenate or vitamin B5, Pyrodoxine or vitamin B6, or a mixture thereof, relative to the total weight of the extract; and

[0099] - at least 3.5 mg / kg, in particular at least 4.2 mg / kg by weight of polyphenols, in particular selected from Astragalin, Kaempferol, or a mixture of these, relative to the total weight of the extract.

[0100] In particular, the aqueous extract of sacred Lotus flowers according to the invention is characterized in that it qualitatively comprises fewer (potentially toxic) alkaloids than other commercial extracts of lotus flowers.

[0101] Indeed, the Applicant observed by liquid chromatography coupled with mass spectrometry that the aqueous extract of sacred Lotus flowers according to the invention, of the genus Nelumbo nucifera, comprises less than 15 alkaloids.

[0102] Thus, according to a particular embodiment, the aqueous extract of Lotus flowers according to the invention is characterized in that it comprises less than 15 different alkaloids, in particular less than 14 different alkaloids, and preferably less than 13 different alkaloids.

[0103] The aqueous phase of the composition according to the invention comprises water and optionally a water-soluble solvent.

[0104] According to the invention, the term 'water-soluble solvent' means a compound that is liquid at room temperature and miscible with water (miscibility in water greater than 50% by weight at 25°C and atmospheric pressure). In particular, mention may be made of:

[0105] - lower C1-C5 monoalcohols such as ethanol, isopropanol and their mixtures;

[0106] - C2-C8 glycols such as ethylene glycol, propylene glycol, 1,3-butylene glycol, dipropylene glycol, and mixtures thereof;

[0107] - C2-C32 polyols such as polyglycerols, polyethylene glycols, and their mixtures,

[0108] and mixtures thereof.

[0109] It may also comprise hydrophilic gelling agents, antioxidants, preservatives and mixtures thereof.

[0110] The cosmetic composition according to the invention may further comprise a fatty (solid fatty substances) or oily phase.

[0111] The term "oily phase" means an oil or a mixture of oils which are miscible with each other, or not. For the purposes of the invention, the term "oil" means a fatty substance, insoluble in water, liquid at 25°C and atmospheric pressure. These oils may be volatile or non-volatile, vegetable, mineral or synthetic.

[0112] An oily phase according to the invention may comprise natural, hydrocarbon, silicone oils, and mixtures thereof.

[0113] The composition of the invention may also comprise any additive usually used in cosmetics such as antioxidants, perfumes, active agents cosmetics, such as emollient agents, moisturizing agents, vitamins, anti-aging agents, lifting agents, tightening agents, plumping agents, lightening agents, fillers, pigments, pearlescent agents and mixtures thereof. Composition - galenic

[0114] The invention also relates to a composition comprising, in a physiologically acceptable medium, an aqueous extract of sacred Lotus flowers according to the invention.

[0115] By "physiologically acceptable" is meant according to the invention that the composition is compatible with human keratin materials, and that it has a pleasant appearance (touch, color and / or odor) without generating discomfort when applied to said keratin materials.

[0116] The composition of the invention is preferably a cosmetic composition. Thus, the cosmetic composition according to the invention comprises, in a physiologically acceptable medium, an aqueous extract of sacred Lotus flowers according to the invention.

[0117] The aqueous extract of sacred lotus flowers according to the invention is present in the cosmetic composition of the invention in an amount effective for obtaining the desired effect.

[0118] In particular, the aqueous extract of sacred lotus flowers is present in a content ranging from 0.01% to 25%, preferably from 0.3% to 20%, preferably from 0.5% to 10%, more preferably from 1% to 5%, more preferably from 2% to 4%, more preferably 3%, by weight relative to the weight of the composition.

[0119] The cosmetic composition of the invention may be presented in any galenic form suitable for topical application to keratin materials and in particular the skin, for example a serum, an oil-in-water (O / W) emulsion, a water-in-oil (W / O) emulsion, a multiple emulsion, a microemulsion, a lotion, a suspension, an aqueous gel, a cream, a mousse, a fluid, etc.

[0120] According to a preferred embodiment, the composition according to the invention is characterized in that it is an emulsion, a serum, an oil, a cream.

[0121] According to a preferred embodiment, the composition according to the invention is characterized in that it is an oil-in-water emulsion, a water-in-oil emulsion, a lotion, an aqueous gel, a serum, a cream, a mousse.

[0122] When the compositions are emulsions, they may comprise the emulsifiers and co-emulsifiers generally used in the cosmetic field.

[0123] When the compositions are gels, they may advantageously comprise hydrophilic gelling agents.

[0124] The composition according to the invention may be a care and / or makeup composition for keratin materials (skin, lips, etc.), in particular for the skin of the face and / or neck.

[0125] The composition is preferably intended to be applied to the skin of the face and / or neck and is presented for example in the form of a lotion, a mousse, a serum, a fluid or a cream for the skin of the face and / or neck.

[0126] According to a preferred embodiment, the cosmetic composition according to the invention is characterized in that it is a care and / or makeup composition for the skin and / or lips, in particular a skin care composition, in particular a serum or a cream for the skin of the face and / or neck. Cosmetic process

[0127] The present invention also relates to a cosmetic process for caring for and / or making up keratin materials, in particular the skin and / or the lips, comprising the application, to said keratin materials, of at least one cosmetic composition according to the invention.

[0128] According to one embodiment, the non-therapeutic cosmetic process for caring for and / or making up the skin and / or lips comprises the application to said skin and / or said lips, in particular the skin of the face and / or neck, of the composition according to the invention.

[0129] Advantageously, the cosmetic process is characterized in that the composition is applied to healthy skin and / or lips in need thereof.

[0130] Preferably, the cosmetic process is characterized in that the composition improves and / or promotes the barrier function, the hydration of the skin, epidermal renewal, prevents and / or reduces skin aging and the formation of wrinkles and / or fine lines.

[0131] The composition can be applied to the body, face and / or neck. According to a particular embodiment, the composition is applied to the skin of the face (including the area around the eyes and lips) and / or the neck.

[0132] The cosmetic process according to the invention is a non-therapeutic process. The composition is therefore applied to healthy skin and / or lips, i.e. skin not presenting any disorders or problems which would be considered a pathological condition. Cosmetic use

[0133] The invention further relates to the non-therapeutic cosmetic use of an aqueous extract of sacred Lotus flowers according to the invention, as an active ingredient for promoting and / or improving the skin barrier function, promoting and / or improving the hydration of the epidermis, preventing and / or reducing skin aging and the formation of wrinkles and / or fine lines, promoting and / or improving epidermal renewal.

[0134] According to a particular embodiment, the effective amount of at least one aqueous extract of sacred lotus according to the invention stimulates the expression of genes involved in the barrier / differentiation, adhesion / cohesion, and / or hydration functions of skin cells.

[0135] In particular, the aqueous extract of sacred lotus according to the invention induces the expression of genes coding for the following proteins: Desmoglein-3, Aquaporin-3, Transglutaminase, Involucrin.

[0136] According to a particular embodiment, the effective quantity of at least one sacred lotus extract according to the invention makes it possible to protect the progenitor cells of the skin, responsible for cell proliferation and renewal of the epidermis. DESCRIPTION OF FIGURES

[0137] [Fig.l]: Protein expression of desmoglein-3, aquaporin-3, transglutaminase and involucrin in keratinocytes treated or not with the aqueous extract of lotus flowers according to the invention, in total intensity per cell.

[0138] [Fig.2]: Protein expression of desmoglein-3, aquaporin-3, transglutaminase and involucrin in keratinocytes treated or not with BGV glycolysate of commercial sacred lotus flowers (comparative), in total intensity per cell.

[0139] [Fig.3]: Quantification of the number of nuclei evaluated 3 days and 6 days after differentiation of keratinocytes treated or not with the aqueous extract of lotus flowers according to the invention, in number of cells per field. EXAMPLES

[0140] EXAMPLE 1: Preparation of the aqueous extract of Lotus

[0141] The aqueous extract of sacred Lotus flowers according to the invention is obtained by subcritical water extraction comprising the steps of:

[0142] a. Recovery of lotus flowers;

[0143] b. Extraction with subcritical water, in particular in a lotus flower / water ratio of 1 / 10, at a temperature of 100°C, and at a pressure of 50 bars;

[0144] c. Addition of glycerin and a preservative system; and

[0145] d. Filtration and obtaining the final extract.

[0146] The aqueous extract of lotus flowers thus obtained comprises less than 15 alkaloids and further comprises: - 1411 mg / kg of minerals among Phosphorus (P), Zinc (Zn), Sodium (Na), Potassium (K); - 2.91 mg / kg of vitamins including Folate or vitamin B9, Niacin or vitamin B3, calcium pantothenate or vitamin B5, Pyrodoxine or vitamin B6; and - 4.2 mg / kg of polyphenols among Astragalin and Kaempferol.

[0147] EXAMPLE 2: Effect of the aqueous extract of Lotus flowers according to the invention on the barrier function of the skin 2.1 - Objectives of this study

[0148] The present study consisted of investigating the properties of the aqueous extract of lotus flowers according to the invention on the formation of the physical barrier and the maintenance of skin hydration. The impact of the aqueous extract of lotus flowers according to the invention, as well as of a commercial extract Glycolysat® BGV of sacred Lotus UP flowers (hereinafter called glycolysate) marketed by Solabia (comparative extract), was evaluated on the renewal of skin keratinocytes and on the expression of differentiation markers linked to the barrier function.

[0149] The markers evaluated in this study are: - Desmoglein-3: cell adhesion protein involved in cell-cell function; - Aquaporin-3: protein involved in the exchange of water molecules between cells and their environment in the deep layers of the epidermis; - Transglutaminase: protein involved in the differentiation of epidermal cells; - Involucrin: protein involved in the formation of the cell envelope that protects corneocytes, surface cells of the skin.

[0150] The aqueous extract of lotus flowers according to the invention comprises glycerin (50% by weight of the aqueous extract), potassium sorbate (0.2% by weight of the aqueous extract), sodium benzoate (0.3% by weight of the aqueous extract), water (33.5% by weight of the aqueous extract) and the crude extract of lotus flowers according to the invention (16% by weight of the aqueous extract). The content of dry extract of lotus flowers in the crude extract of lotus flowers represents 0.92% by weight of the crude extract of lotus flower.

[0151] Sacred Lotus Flower Glycolysat® BGV comprises butylene glycol (68% by weight of glycolysate) and crude lotus flower extract (32% by weight of glycolysate). The content of dry lotus flower extract in the crude lotus flower extract represents 2% by weight of crude lotus flower extract glycolysate. 2.2. Materials and methods

[0152] 2.2.1. Cell culture

[0153] Passage 5 normal human keratinocytes (NHK) from a 47-year-old female donor are cultured in 96-well plates on complete Epilife medium. After 24 hours of culture, the cells are treated according to two conditions: - with 500 pM of CaCl2, in the presence or absence of aqueous extract of Lotus flowers according to the invention at 0.3% (corresponding to 0.00044% of dry extract of lotus flowers) in the culture medium; - with 500 pM of CaCl2, in the presence or absence of Glycolysat® BGV of commercial UP sacred Lotus flowers (Solabia) at 0.1% (corresponding to 0.00064% of dry extract of lotus flowers) in the culture medium.

[0154] 2.2.2. Cell fixation and immunofluorescence labeling

[0155] After 72 hours of treatment, each well is rinsed three times with 200 pL of PBS (phosphate buffered saline). The cells are then fixed with 100 pL of a commercial solution of 10% Formalin (Formalin solution, neutral buffered, Sigma, HT5014) per well for 10 min. Each well is then rinsed with 200 pL of PBS, three times.

[0156] Once fixed, the cells are permeabilized with a 0.1% PBS and triton solution, adding 100 μL of the PBS / triton solution per well for 10 min at room temperature. Each well is then rinsed with 200 μL of PBS, three times.

[0157] The permeabilized cells are then saturated with 100 μL of a 1% PBS / BSA solution per well for at least 30 min at room temperature.

[0158] The cells are then labeled with one of four primary antibodies: rabbit antibody against transglutaminase I (Thermo - Invitrogen Life Technologies, 1 / 500 dilution), rabbit antibody against aquaporin-3 (Abcam, 1 / 200 dilution), mouse antibody against desmoglein-3 (Thermo - Invitrogen Life Technologies, 1 / 200 dilution), and mouse antibody against involucrin (Thermo - Invitrogen Life Technologies, 1 / 200 dilution). 70 pL of primary antibody is placed in each well and left to incubate overnight at 4°C. Each well is then rinsed with 200 pL of PBS, three times. Then a DAPI DNA marker (4',6-diamidino-2-phenylindole, Thermo - Invitrogen Life Technologies, dilution 1 / 500) and one of the two secondary antibodies are added: either Anti-Mouse AF568 (Thermo - Invitrogen Life Technologies, dilution 1 / 500) or Anti-Rabbit AF568 (Thermo - Invitrogen Life Technologies, dilution 1 / 500).70 pL of secondary antibody solution and DAPI marker are placed in each well and left to incubate for 45 min in the dark at room temperature. Each well is then rinsed with 200 pL of PBS, three times.

[0159] Each well is then filled with 200 pL of PBS and the cells are stored at 4°C.

[0160] 2.2.3 Quantification of the number of nuclei

[0161] The acquisitions are made with the plate scanner (ArrayScan XTI, Thermo Cellomics) with the xl0 objective (25 images per well). The images are then analyzed using the Spot detector image analysis software, a bio-application developed for the arrayscan system. This program (pipeline) allows the detection of the red labeling of the targeted protein (transglutaminase I, aquaporin-3, desmoglein-3 or involucrin) corresponding to its expression. The surface of the measurement area is delimited as the entire surface of the image. The number of cells is determined by counting the nuclei by detecting the blue labeling (DAPI marker). The results are transferred to a Microsoft Excel file.

[0162] Statistical analysis was performed using the Student test. Results are considered significant when the p-value is less than 0.05: p<0.05(*), p<0.01(**), p<0.001 (***).

[0163] The results are expressed as intensity of the detected marking (Thresholding) / number of cells. 2.3. Results

[0164] 2.3.1 Effect of the aqueous extract of lotus flowers according to the invention on the function barrier in differentiated KHNs

[0165] The expression of specific protein markers of the barrier function was evaluated in keratinocytes treated or not with the aqueous extract of lotus flowers according to the invention. These results are represented in Table 1 and in [Fig.l].

[0166] The expression of specific protein markers of barrier function was also evaluated in keratinocytes treated or not with Glycolysat® BGV of commercial UP sacred lotus flowers (Solabia) (comparative extract). These results are shown in Table 2 and in [Fig.2].

[0167] Table 1: results of the expression of protein markers of the barrier function in keratinocytes treated with the aqueous extract of lotus flowers according to the invention

[0168] [Tables 1] Untreated Control (NT) Aqueous extract of Lotus flowers according to the invention % vs Control p-value Desmoglein-3 333271 464727 +39% 0.002 Aquaporin-3 173383 849908 +390% 0.000 Transglutaminase 811614 1294162 +59% 0.000 Involucrin 320320 396580 +24% 0.048

[0169] Table 2: Results of the expression of protein markers of barrier function in keratinocytes treated with Glycolysat® BGV of sacred lotus flowers UP

[0170] [Tables2] Untreated Control (NT) Glycolysate (comparative) % vs Control p-value Aquaporin-3 1313723 1990989 +52% 0.000 Transglutaminase 440474 409541 -7% 0.014 Involucrine 58179 85379 +47% 0.000

[0171] Note that for BGV Glycolysate, the desmoglein-3 marker was not evaluated.

[0172] The results in Table 1 and [Fig.l] show an increase in protein markers associated with barrier function (Desmoglein-3, Aquaporin-3, Transglutaminase, Involucrin), in keratinocytes treated with the aqueous extract of lotus flowers, compared to untreated keratinocytes. The aqueous lotus extract according to the invention shows a positive effect on the expression of early differentiation markers, indicating an improvement in keratinocyte differentiation necessary for an effective barrier function.

[0173] The results in Table 2 and [Fig.2] show a more moderate increase in the Aquaporin-3 marker, and even a decrease in the expression of the Transglutaminase marker in keratinocytes treated with Glycolysat® BGV of sacred lotus flowers UP (comparative extract), compared to untreated keratinocytes. The aqueous lotus extract according to the invention therefore shows a reinforced positive effect on the expression of early differentiation markers, compared to the glycolysate (Glycolysat® BGV of sacred lotus flowers UP).

[0174] 2.3. 2. Effect of the aqueous extract of lotus flowers according to the invention on the proliferation of normal human keratinocytes

[0175] The quantification of the number of nuclei was evaluated 3 days and 6 days after differentiation for keratinocytes treated or not with the aqueous extract of lotus flowers according to the invention. These results are represented in Table 3 and in [Fig.3].

[0176] Table 3: results of the quantification of the number of nuclei of keratinocytes treated with the aqueous extract of lotus flowers according to the invention

[0177] [Tables 3] Untreated Control (NT) Aqueous extract of Lotus flowers according to the invention % vs Control p-value Number of kernels (J3) 1883 1930 +3% 0.028 Number of kernels (J6) 1564 1817 +16% 0.000

[0178] The results in Table 3 and [Fig. 3] show an increase in the number new nuclei in keratinocytes treated with the aqueous extract of lotus flowers according to the invention, compared to untreated keratinocytes, by 3% after the 3rd day of differentiation and by 16% after the 6th day of differentiation. The new Aqueous extract of lotus flowers therefore shows a positive effect on the number of nuclei observed, indicating an increase in the proliferation of keratinocytes.

[0179] EXAMPLE 3: Protective effect of aqueous extract of lotus flowers on skin progenitor cells 3.1 - Objectives of this study

[0180] This technique, developed by Y. Barrandon and H. Green (1987), makes it possible to characterize keratinocytes in relation to their capacity to form clones (CFE = Clony Forming Efficiency) during particular culture conditions.

[0181] The ability of keratinocytes to form clones depends on their proliferation potential as well as their state of differentiation and age (Lecardonnel, 2013). Three types of clones can be analyzed: Holoclones, large clones with regular edges; Meroclones, medium clones with irregular edges; and Paraclones, small heterogeneous clones (Barrandon and H. Green, 1987).

[0182] Stem keratinocytes are located at the level of the basal membrane of the epidermis and are therefore preferentially available when isolated from a surgical residue (Germain, 1993). These progenitor cells have a higher growth capacity than transient keratinocytes and therefore form more Holoclones (Webb, 2004).

[0183] The aim of this study is to evaluate the impact of the active ingredients (aqueous extract of lotus flowers according to the invention and Glycolysat® BGV of sacred lotus flowers UP used as a comparative extract) on the protection of progenitor cells and not their impact on increasing the capacity of progenitor cells to proliferate. 3.2. Materials and methods

[0184] 3.2.1. Cell culture and active ingredients

[0185] Human Fibroblasts (HF) from a surgical foreskin residue of a 1-year-old male patient are irradiated using a gamma ionizing ray source (Fhiy) to cause cell division arrest. These Fhiy are seeded and cultured in 6-well plates at a density of 5 xlO3 cells / cm in FAD culture medium (DMEM 1 g / L glucose with Glutamax-I supplemented with ham F12 with Glutamax-I, fetal calf serum (FCS), hydrocortisone, adenine, penicillin-streptomycin, amphoterecin B, isoproterenol, insulin and triiodo-L-thyronine).

[0186] After 24 hours of culture, normal human keratinocytes (NHK) from an abdominal surgical residue of a 37-year-old female patient are seeded at passage 1 at a density of 5.5 xlO3 cells / cm2 on the Fhiy. The cells are treated when they reach 50 to 60% confluence with the active ingredients alone. (in biological replicate of n = 4) for 24 hours then subjected to oxidative stress. Untreated conditions (NT) are also made, in which the cells are not put in contact with the active ingredients.

[0187] The active ingredients are the aqueous extract of lotus flowers according to the invention and the commercial extract Glycolysat® BGV of sacred lotus flowers UP (hereinafter called glycolysat) marketed by Solabia (comparative extract).

[0188] 3.2.2. Induction of oxidative stress by treatment of cells with H 2O 2

[0189] After the 24 hours of pre-treatment with the active ingredients, each well containing KHN on Fhiy in a 6-well plate is rinsed with PBS before inducing oxidative stress with a 100 pM hydrogen peroxide (H2O2) solution in PBS. Each active ingredient is prepared in the presence of the H2O2 solution (“scavenger” effect) before being deposited or not on the cells (in the NT 'Untreated' condition, the cells are not put in contact with the active ingredients). Oxidative stress is induced for 15 minutes at 37°C.

[0190] The KHN on Fhiy are trypsinized immediately after hydrogen peroxide stress and then seeded in Petri dishes at very low density on a feeder layer (150 KHN / cm2) to allow them to distribute homogeneously on the culture medium, adhere and multiply in the form of a clone from a single cell.

[0191] 3.2.3. Fixation and staining of clones

[0192] Eight days after cell seeding, the cell lawn of the Petri dishes is rinsed with PBS. A solution of rhodamine B (Merck Millipore; Ref. 107599; RC-1425) at a concentration of 10 mg / mL is prepared in 10% formalin containing 4% Formaldehyde (Sigma Aldrich) under a fume hood. This dye is fluorescent at / .cm = 570 nm and binds to the amine groups of proteins.

[0193] This solution is deposited on the cell mat in large excess and then incubated with stirring at room temperature for at least 30 minutes. The excess rhodamine solution is then recovered and the cell mat rinsed under a stream of cold tap water until the wash water becomes clear again. The Petri dishes are left to air dry on absorbent paper for at least 24 hours.

[0194] 3.2.4. Image acquisition with the iBright FL1500

[0195] Each Petri dish is imaged with the iBright FL1500 Imaging System (Invitrogen Thermo Fisher) equipped with a high-sensitivity 9.1 MegaPixel CCD camera and 5 filters to detect different fluorophores (from green: 500 nm to near infrared: 850 nm). A dish is placed in the center of the tray and then illuminated by white LEDs to detect the fluorescence of Rhodamine B (Xem = 570 nm). The camera is programmed according to the following parameters on the “Protein Gels / Fluorescent” acquisition method: Optical zoom: 2x / Focus: 455 / Binning: 1x1 / Acquisition time: 60 ms. The images obtained are in .Tiff and the pixel area is 1244 pm2.

[0196] 3.2.5. Image analysis on Cell Profiler

[0197] Images from each Petri dish are analyzed using the open source image analysis software Cell Profiler version 2.0, which allows quantitative measurements to be programmed automatically. A pipeline program is created to detect and segment clones so that they can be counted and their surface areas calculated.

[0198] For each replica, the surface data is classified to define the 3 types of clones: • Small clones: [100-1000] pixels or 0.124 - 1.244 mm2 • Medium clones: [1000-5000] pixels or 1.244 - 6.22 mm2 • Large clones: [5000-20000] pixels or 6.22 - 24.88 mm2

[0199] 3.2.6. Statistical analyses

[0200] For each of the conditions, carried out in 4 biological replicates, descriptive statistics tests are performed to determine the mean, the standard deviation and the confidence interval (a = 0.05). In order to evaluate statistically significant variations between the control condition and the treated conditions, the Student t test is used. The Fisher F test is first applied by comparing the two data matrices: • When the value is a > 0.05 then the variance for the Student t test is 2 • If the value is a < 0.05 then the variance will be equal to 3.

[0201] The variations retained are those which are significant for the Student t test (* p<0.05; ** p<0.01; *** p<0.001). 3.3. Results

[0202] The protective effect of the aqueous extract of lotus flowers on clones derived from epidermal stem cells was evaluated on KHN on Fhiy, treated or not with the aqueous extract of lotus flowers according to the invention. These results are shown in Table 4.

[0203] The protective effect of the glycolysate on clones derived from epidermal stem cells was evaluated in KHN on Fhiy, treated or not with Glycolysat® BGV from commercial UP sacred lotus flowers (Solabia) (comparative extract). These results are shown in Table 5.

[0204] Table 4: results of the protective effect of the aqueous extract of Lotus flowers according to the invention on clones derived from epidermal stem cells

[0205] [Tables4] Control H2O2 Aqueous extract of Lotus flowers according to the invention % vs Control n p-value Number of clones 26 41 +60% 0.004 Total surface area of ​​clones 217500 365014 +68% 0.021

[0206] Table 5: results of the protective effect of Glycolysat® BGV of sacred Lotus flowers UP on clones derived from epidermal stem cells

[0207] [Tables5] Control H2O2 Glycolysate (comparative) % vs Control n p-value Number of clones 26 38 +49% 0.038 Total clone area 217500 267488 +23% 0.652

[0208] The results in Table 4 show a significant increase in the number of clones when the cells are treated with the aqueous extract of lotus flowers according to the invention, compared to untreated cells. The aqueous extract of lotus according to the invention shows a positive effect on the protection of progenitor cells and on their ability to proliferate.

[0209] The results in Table 5 show an increase in the number of clones when the cells are treated with Glycolysat® BGV of sacred Lotus flowers UP (comparative), compared to untreated keratinocytes. This increase is however less significant than with the aqueous extract of lotus flowers according to the invention.

[0210] The aqueous extract of lotus flowers according to the invention therefore protects progenitor cells more effectively, compared to the glycolysate (comparative extract). EXAMPLE 4#: Formulations 4.1 Composition in the form of an emulsion

[0211] Aqueous phase:

[0212] Demineralized water qsp 100.0%

[0213] Glycols 20.0%

[0214] Preservatives 0.6%

[0215] Chelating agent 0.04%

[0216] Sodium Hydroxide 0.15%

[0217] Aqueous extract of sacred lotus flowers, of the genus Nelumbo nucifera 0.2%

[0218] Fatty phase:

[0219] Vegetable oil, esters, silicones 10%

[0220] Antioxidant 0.2%

[0221] Perfume concentrate 0.4%

[0222] Steareth-2 0.5%

[0223] Steareth-21 2%

[0224] The cosmetic composition is prepared according to the following procedure:

[0225] - the emulsifiers (Steareth-2 and Steareth-21) are dispersed in the aqueous phase which is brought to 70°C;

[0226] - the fatty phase (excluding aqueous extract of sacred Lotus flowers, of the genus Nelumbo nucifera, perfume concentrate, antioxidant) is heated to 70°C;

[0227] - the emulsion is produced by introducing the fatty phase into the aqueous phase under strong agitation;

[0228] - the emulsifiers are neutralized by adding sodium hydroxide and the emulsion is cooled under moderate stirring with the introduction of the aqueous extract of sacred Lotus flowers, of the genus Nelumbo nucifera, the perfume concentrate, the antioxidant and at low temperature.

[0229] Applied to the skin, particularly the skin of the face, this composition provides tone, elasticity and firmness, the features are enhanced, the face is reshaped.

[0230] 4.2: Composition in the form of an aqueous gel for the face

[0231] Aqueous extract of sacred lotus flowers, of the genus Nelumbo nucifera 1%

[0232] Preservatives 0.50%

[0233] Sugar 0.20%

[0234] Carbomer 0.70%

[0235] Purified water Qsp 100%

[0236] Tetrasodium EDTA powder 0.20%

[0237] Sodium hydroxide 0.17%

[0238] The cosmetic active ingredients of the composition according to the invention and the preservatives are mixed and homogenized at room temperature with stirring. The sugars are added with stirring, then the carbomer, then the water and the EDTA with stirring. The aqueous gel is then neutralized with the addition of sodium hydroxide with stirring until a homogeneous gel is obtained.

[0239] Applied to facial skin, this aqueous gel corrects skin dryness, increases firmness and helps slow down skin aging. REFERENCES

[0240] Barrandon and Green (1987). Three clonal types of keratinocyte with different capacities for multiplication. National Acad Sciences, 84: 2302-2306.

[0241] Lecardonnel (2013). Aging and colony-forming efficiency of human hair follicle keratinocytes. Wiley Online Library.

[0242] Germain (1993). Improvement of human kératinocyte isolation and culture using thermolysin. Burns 19 (2) : 99-104

[0243] Webb (2004). Location and phenotype of human adult kératinocyte stem cells of the skin. Différentiation; Research In Biological Diversity, 72 (8)

Claims

Claims

1. Aqueous extract of sacred lotus flowers, of the genus Nelumbo nucifera, characterized in that it is obtained by subcritical water extraction comprising the steps of: a. Recovery of the lotus flowers; b. Extraction with subcritical water, in particular in a lotus flower / water ratio of 1 / 20, preferably a ratio of 1 / 10, at a temperature ranging from 90°C to 110°C, preferably 100°C to 120°C, preferentially 100°C, and at a pressure ranging from 40 to 60 bars, preferably 50 bars; c. Addition of glycerin and a preservative system; and d. Filtration and obtaining the final extract.

2. The aqueous extract according to claim 1, characterized in that it comprises glycerin in a content ranging in particular from 40 to 60% by weight of the aqueous extract, water in a content ranging in particular from 30 to 40% by weight of the aqueous extract, a crude extract of sacred lotus flowers in a content of crude extract ranging in particular from 10 to 25% by weight of the aqueous extract, and a preservative system comprising sodium benzoate and potassium sorbate in a content ranging in particular from 0.2 to 1% by weight of the aqueous extract.

3. The aqueous extract of sacred Lotus flowers according to claim 1 or 2, characterized in that it further comprises: - from 1300 mg / kg to 2300 mg / kg of minerals, in particular from 1411 mg / kg to 2117 mg / kg of minerals, in particular selected from Phosphorus (P), Zinc (Zn), Sodium (Na), Potassium (K), or a mixture thereof; - at least 2 mg / kg of vitamins, in particular at least 2.9 mg / kg of vitamins, in particular selected from Folate or vitamin B9, Niacin or vitamin B3, calcium pantothenate or vitamin B5, Pyrodoxine or vitamin B6, or a mixture thereof; and - at least 3.5 mg / kg, in particular at least 4.2 mg / kg of polyphenols, in particular selected from Astragalin, Kaempferol, or a mixture thereof.

4. The aqueous extract of Sacred Lotus flowers according to any one of claims 1 to 3, characterized in that it comprises less than 15 alkaloids.

5. Cosmetic composition comprising, in a physiologically acceptable medium, an aqueous extract of sacred Lotus flowers as defined in any one of claims 1 to 4.

6. Cosmetic composition according to claim 5, characterized in that the aqueous extract of lotus flowers is present in the composition in a content ranging from 0.01% to 25% by weight relative to the weight of the composition.

7. Cosmetic composition according to claim 5 or 6, characterized in that it is in the form of an oil-in-water emulsion, a water-in-oil emulsion, a lotion, an aqueous gel, a serum, a cream, a mousse.

8. Cosmetic composition according to any one of claims 5 to 7, characterized in that it is a care and / or makeup composition for the skin and / or lips, in particular a skin care composition, in particular a serum or cream for the skin of the face and / or neck.

9. Non-therapeutic cosmetic process for the care and / or makeup of healthy skin and / or healthy lips comprising the application to said skin and / or said lips, in particular the skin of the face and / or neck, of a composition as defined in any one of claims 5 to 8.

10. Cosmetic process according to claim 9, characterized in that the composition promotes and / or improves the barrier function and / or hydration of healthy skin, prevents and / or reduces skin aging and the formation of wrinkles and / or fine lines, promotes and / or improves epidermal renewal.

11. Non-therapeutic cosmetic use of the aqueous extract of Sacred Lotus flowers as defined according to any one of claims 1 to 4, as an active ingredient for promoting and / or improving the skin barrier function, promoting and / or improving the hydration of the epidermis, preventing and / or reducing skin aging and the formation of wrinkles and / or fine lines, promoting and / or improving epidermal renewal, healthy skin and / or healthy lips.

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