OA21068A - Moringa Peregrina seed cake extract, its production process and its use in cosmetic or nutricosmetic compositions. - Google Patents

Abstract

The invention relates to an extract of Moringa peregrina seed cake comprising mainly a peptide hydrolyzate and a process for obtaining the extract. It also relates to cosmetic or nutricosmetic compositions comprising said extract and the use of said compositions to improve the appearance of the skin, mucous membranes or appendages, prevent and/or combat dryness of the skin and mucous membranes, prevent and /or to fight against the skin signs of aging and/or photo-aging, promote skin whitening and promote thinning.

Description

Description

Title of the invention: Moringa peregrina seed cake extract, its process for obtaining it and its use in cosmetic or nutricosmetic compositions

Technical area

[1] The invention relates to the field of cosmetics and nutricosmetics and more particularly that of active ingredients used in the formulation of compositions for skin care. The invention relates to an extract of Moringa peregrina seed cake comprising mainly a peptide hydrolyzate. The invention also relates to the process for obtaining the particular extract from Moringa peregrina seed cake, the cosmetic or nutricosmetic compositions comprising such extracts, and finally the cosmetic or nutricosmetic use of such compositions for skin care. , scalp and integuments.

Technology background

[2] The Moringaceae constitute a mono-generic family (a single genus, Moringa Adans), element of the Saharo-Sindian flora, made up of between twelve and fourteen species according to the authors, distributed from eastern Africa to Asia . The genus is classically divided into 3 sections which are, however, not confirmed as monophyletic by phylogenetic analyses. The latter rather highlighted clades focused on certain morphological characters: pachycaules (“bottle trees”); tuberous trees and neither pachycaules nor tuberous trees (in English “slender trees”). The species Moringa peregrina (Forssk.) Fiori belongs to the third group. The few genetic studies on the genus or family confirm the reality of the species compared to other species in the genus, particularly with regard to the Indian Moringa, Moringa oleifera Lam. (see in particular the articles: Οί-SON, Μ. E. 2002, Combining Data from DNA Sequences and Morphology for a Phylogeny of Moringaceae (Brassicales), Systematic Botany 27(1): 5573; Hassanein, A. M. A. and AL-SOQEE, A. A., 2018, Morphological and genetic diversity of Moringa oleifera and Moringa peregrina genotypes, Horticulture, Environment and Biotechnology 59(2): 251-261). A recent article on Moringa peregrina sampled from different localities in Saudi Arabia concluded, using ITS markers, that the species is genetically stable (Alaklabi, A., 2015, Genetic diversity of Moringa peregrina species in Saudi Arabia with ITS sequences, Saudi Journal of Biological Sciences 22: 186190) with, however, a high level of intra-population genetic variation.

[3] The species Moringa peregrina is found in rocky environments of Yemen, Oman, and Saudi Arabia, in East Africa, Sudan, Ethiopia, Eritrea, Somalia and Djibouti. Its presence in Iran seems limited to the south-eastern provinces but requires confirmation (PROTA14 = Munyanziza E. and Yongabi K. A., Vegetable oils/Oléagineux, Moringa peregrina (Forssk.) Fiori, http://database.prota.org/protahtml /moringa peregrina_fr.htm, accessed 10/23/2019). In the Levant and Egypt, the species today only represents rare, scattered, relict stations (with the exception of a few altitudinal populations), mainly in the areas of the Sudanian area. Moringa peregrina is also considered rare and endangered today in Sudan and Yemen. Moringa peregrina occupies the driest and most inhospitable habitats compared to the other species of its clade. It is apparently more drought tolerant than Moringa oleifera which is planted commercially on a large scale in tropical and subtropical areas. Recent work has shown that the size and size of seeds has a favorable impact on germination time and the rate and speed of growth of young individuals (Gomaa N. H. and Pico F. X., 2011, Seed germination, seedling traits, and seed bank of the tree Moringa peregrina (Moringaceae) in a hyper-arid environment, American Journal of Botany 98(6); 1024-1030), indicating an adjustment in resource allocation on seed quality rather than number, which allows Moringa peregrina to reproduce effectively in extreme abiotic (hyperarid) environments. Moringa peregrina seeds have a thicker central mesotesta, in terms of cell layer, than those of Moringa oleifera.

[4] There are some historical records which tend to indicate that Moringa peregrina oil was actively traded in the early days of Islam in the AlUla region (Naseef, AAS, 1995, AI- VlS, A study of Cultural and Social Heritage). The oil produced from Moringa peregrina is today mainly intended for self-consumption or local markets. In Saudi Arabia, the leaves were traditionally used as a decoction for internal use to treat diabetes, colon diseases, eye diseases and anemia (Abdel-Kader, MS, Hazazi A. Μ. A., Elmakki OA and Alqasoumi S . L, 2018, Asurvey on the traditional plants used in Al Kobah village, Saudi Pharmaceutical Journal 26(6): 817-821) and as a diuretic, rubefacient and astringent (Aqeel AAM, Tariq M., Mossa JS, Al-YahYA MA ET Al-Said MS, 1984, “Plants used in Arabia Folk medicine”, Report submitted to Saudi Arabian National Genre for Science and Technology, Rtyadh, Saudi Arabia). In Oman, the oil, extracted by women at the end of summer, is used against migraines, fever, burns, lacerations and fractures, constipation and stomach pain, against muscle pain and against hair dryness (Ghazanfar SA, 1994, Handbook of Arabian Medicinal Plants, ^1st ed., CRC Press, Boca Raton, Ann Harbor, su; Ghazanfar SA, 1998, Plants of Economic Importance, cap. 15, in Ghazanfar, SA and Fisher, M. (ed.) Vegetation of the Arabian Peninsula. Geobotany 25, pp. 241-264, Kluwer Academie Publishers, table 11.1, p. 247 and 11.7 p. 251). It was also used in perfume compositions (Ghazanfar SA, 1998, p. 259) and in Oman and Yemen as a facial lotion (Ghazanfar SA and Rechinger B., 1996, Two multi-purpose seed oils form Oman. Plants for Food and Medicine . Paper presented at the joint meeting of the Society for Economic Botany and International Society for Ethnopharmacology, 1 -7 July 1996, London).

[5] Extracts from the seed of Moringa oleifera are known in the cosmetic field. For example, we know from document FR296879 an extract of whole seeds (with integuments) of Moringa oleifera containing per 100 g of dry extract from 5 to 50% oil (including triglycerides, fatty acids and polar lipids) and from 0.01 to 5 % of polyphenols, and its use in cosmetic compositions to fight against skin aging. In this document, an extraction with a medium polar solvent is carried out on the whole seed of Moringa oleifera.

[6] We also know from document FR2776519 that protein extracts of Moringa oleifera seeds, known for their clarifying effects on turbid waters, have a softening, physiological conditioning, hydrating, restructuring, restorative and a softening effect on the skin and mucous membranes. anti-pollution effect. In this document the active ingredients are proteins with molecular weights between 6500 and 8800 Da which are obtained by aqueous extraction on Moringa oleifera cake with 4N sodium hydroxide for one hour at pH 7.5.

[7] We also know from document FR2825267 that extracts of Moringa seeds, whole or crushed, shelled or not shelled, or from which a ground material or delipidated or non-delipidated protein extracts have been produced, have benefits in the fields of deodorization and removal of bad odors, cleanliness, intimate hygiene, oral hygiene and dental care, as well as softening, moisturizing, soothing and anti-fatigue cosmetic properties on the skin, In this document, whole protein extracts are shown to exhibit increased activity when the seed is not deoiled.

[8] Document FR3076460 concerns the use of a protein extract of non-germinated and de-oiled seed of Moringa oleifera for the treatment of sensitive, sensitized, reactive, fragile and/or weakened skin and/or mucous membranes and/or in the treatment and/or prevention of erythema, particularly diaper rash in infants. In this document, the extraction process makes it possible to obtain a major fraction of proteins whose molecular weights are approximately 8,800 Da.

[9] Finally, we know from document KR20130088224 the use of an extract of whole germinated seed of Moringa oleifera in cosmetics, in particular obtained by extraction using a supercritical fluid. This process makes it possible to isolate non-polar carotenoids and amino acids, which are described as active for whitening cosmetic use.

[10] All the aforementioned documents concern the use of the species Moringa oleifera; none describes the use in the cosmetic field of extracts from the Moringa peregrina species.

[11] More specifically for the species Moringa peregrina, it is known that certain phenolic and flavonoid compounds obtained from leaves or the whole seed of Moringa peregrina exhibit antioxidant activity (Al-Dabbas M., 2017, Antioxidant activity of different extracts from the aerial part of Moringa peregrina (Forssk.) Fiori, from Jordan, Pakistan Journal of Pharmaceutical Sciences, 30(6): 2151-2157). These compounds are extracted with solvents such as methanol, ethyl acetate or hexane from the leaves or whole seeds. It appears that it is the leaves which contain the most active compounds. We also know the document by Abu Tarboush el al. XP055753092, 2005 which describes the extraction of a hydrolyzate from shelled Moringa peregrina seeds produced by enzymatic hydrolysis for a period of 10 hours. The aim of this extraction is to obtain a product with a high oil and water absorption capacity. Finally, the document by Nashef et al., ΧΡ0010830,1977, generally describes the consequences of alkaline hydrolysis mainly on pure proteins which can lead to modifications in the physical and chemical properties of proteins as well as their nutritional value. The hydrolysis conditions used with 0.1 M sodium hydroxide are at a temperature of 50°C for a period of 24 hours. The paper claims kidney damage in rats fed alkali-treated proteins under these conditions.

[12] Thus, depending on the species used in the Moringa genus, it is observed that depending on the part of the plants (leaf, seed), the part of the seeds (whole or not, shelled or not) and the extraction process used implemented, in particular the choice of solvent, the extracted molecules turn out to be different. However, the composition of the extract determines the biological activity and consequently the cosmetic effectiveness.

[13] Considering the above, a problem that the invention proposes to solve is to develop new products based on an extract of the species Moringa peregrina of the Moringa genus and the Moringaceae family which can be used in cosmetics and easy to implement.

[14] As a result, the Applicant has demonstrated a new extract obtained from the seed cake of the species Moringa peregrina, which improves the appearance of the skin, mucous membranes and integuments and in particular to prevent and/or or fight against dryness of the skin and mucous membranes, prevent and/or fight against the skin signs of aging and/or photo-aging, promote whitening of the skin and prevent age spots, as well as to promote the slimming and preventing or relieving redness of the skin. The extract according to the invention mainly comprises a peptide hydrolyzate from Moringa peregrina seed cake. The extract obtained specifically from the cake of unshelled seeds of Moringa peregrina is new in two ways in the field of cosmetics compared to the extracts of the prior art, on the one hand by the specific species of origin used and on the other hand by its particular molecular profile.

[15] By intergovernmental agreement of April 10, 2018 between the government of the French Republic and the Kingdom of Saudi Arabia, the plaintiff, Agence Française Pour Le Développement d'AlUla (AFALULA) as well as the Royal Commission for AIUIA (RCU) have for a common project in particular to develop responsible agriculture and the local economy, in particular through the local production of natural products derived from indigenous plants and to protect biodiversity and the rights of the AlUla region of the Kingdom of Saudi Arabia. The Kingdom of Saudi Arabia has been a member of the Nagoya Protocol since October 8, 2020. At the time of writing this patent, the implementing regulations under which the Nagoya Protocol will be incorporated into relevant aspects of local law are in force. exam course. Therefore, at this stage, the Kingdom of Saudi Arabia has no specific requirements with regard to this patent application and the Nagoya Protocol. Thus, on the day the patent application is filed, there is no requirement for a certificate of conformity regarding access to genetic resources.

Summary

[16] The first subject of the invention is an extract of Moringa peregrina seed cake comprising mainly a peptide hydrolyzate which comprises amino acid derivatives, amino acids, peptides and glycopeptides whose molecular weight is between 100 Da and 6,000 Da, preferably between 1,500 Da and 5,000 Da, more preferably between 3,000 and 4,500 Da, and in that it is obtained from unshelled and mature seeds of the fruit of Moringa peregrina by chemical proteolysis at pH greater than 13 for a period of approximately 2 hours at a temperature between 16 and 25°C.

[17] Thanks to its characteristic, the extract according to the invention is unique in the Moringa genus and the Moringaceae family. It will be demonstrated that the extract of the species Moringa peregrina presents a particular peptide hydrolyzate different from those known from other species of the genus, in particular from the species Moringa oleifera, which the applicant was able to demonstrate.

[18] The second object of the invention is a process for obtaining a meal extract from the seeds of Moringa peregrina according to the invention, characterized in that it comprises the following steps according to which:

a) we collect the unshelled seeds harvested when ripe from the fruit of Moringa peregrina which we dry to obtain an internal humidity level of less than 8%,

b) the dried seeds are pressed so as to separate the oil from the rest of the seed so as to obtain the cake comprising less than 6% by weight of residual oil, c) the cake obtained in step b) is crushed. , d) the ground cake obtained in step c) is dispersed in the aqueous phase),

e) chemical proteolysis is carried out on the aqueous dispersion obtained in step d) for a period of approximately 2 hours, at a pH greater than 13 and at a temperature between 16 and 25°C,

f) the proteolysis is neutralized to stabilize the peptide hydrolyzate obtained, g) the peptide hydrolyzate is recovered by solid/liquid separation, h) the peptide hydrolyzate is purified by ultrafiltration, then optionally, i) the lyophilization is carried out. peptide hydrolyzate obtained in step h).

[19] The third object of the invention is a cosmetic or nutricosmetic composition comprising, as active agent, an effective quantity of an extract of Moringa peregrina seed cake according to the invention and a physiologically acceptable excipient.

[20] Finally, the fourth object of the invention is the cosmetic or nutricosmetic use of a composition according to the invention, to improve the appearance of the skin, mucous membranes or appendages, prevent and/or combat dryness of the skin. the skin and mucous membranes, prevent and/or combat the skin signs of aging and/or photoaging, promote whitening of the skin and promote thinning.

Brief description of the figures

[21] The invention will be better understood, and other aims, details, characteristics and advantages thereof will appear more clearly during the following description of several particular embodiments of the invention, given solely by way of illustration. and non-limiting, with reference to the appended drawings.

[22] [fig. 1 ] represents the comparative results of the moisturizing effect of different products on the skin surface.

Description of embodiments

[23] In this description, unless otherwise specified, it is understood that, when an interval is given, it includes the upper and lower limits of said interval.

[24] In the present invention, the following abbreviations mean:

- MTT: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (the MTT test is a rapid method for counting living cells)

- SDS: Sodium Dodecyl Sulfate

- PBS: Phosphate Buffer Saline

- ELISA: Enzyme-Linked Immunosorbent Assay

- PCR: Polymerase Chain Reaction

- ANOVA: Analysis Of Variance

- MSH: Melanocyte Stimulating Hormone

[25] In the present invention, we mean:

- “mainly a peptide hydrolyzate” a quantity greater than 10%, preferably greater than 20%, even more preferably greater than 30% and which can reach approximately 40% (Mass/Mass) of dry matter in peptides, oligopeptides, glycopeptides and amino acids or their volatile nitrilated derivatives, preferably still 50% of the weight of the dry matter.

- “effective quantity” the necessary quantity of active molecules to obtain the desired result, namely, to improve the appearance of the skin.

- “proteolysis” the segmentation of proteins into peptides, oligopeptides and its basic fragments (amino acids) via chemical or enzymatic hydrolysis.

- “topical application”, the act of applying or spreading the active principle according to the invention, or a composition containing it, to the surface of the skin, a mucous membrane or the integuments.

- “physiologically acceptable” which is suitable for topical use, in contact with human skin, or for use by other routes of administration, for example the oral route or injection into the skin, without risk of toxicity, incompatibility, instability, allergic response.

- “cake”, the deoiled part of the seed after pressing. This is the solid residue from the extraction of oil from the seeds. It is a co-product of crushing, the oil manufacturing process. It generally represents 50 to 75% of the mass of the seeds.

- “unshelled seeds”, the fact that the shell (pericarp) and the integument of the harvested seeds are maintained around the almonds.

- “when the fruit is ripe”, the fact that the fruit is ripe, preferably when the pod is at the beginning of dehiscence and it takes on a dark beige to brown color and when a 180° twist is applied to the lower quarter of the fruit. the pod causes the valves to begin to open.

- “approximately” a margin more or less of 10% to 20% in relation to the information given.

- “pool of active molecules”, also “active principle”, the peptide hydrolyzate extracted according to the process of the invention from Moringa peregrina seed cake. This hydrolyzate is responsible for the biological activities described in the present invention.

- “active agent” a sufficient quantity of an extract according to the invention to obtain the biological activities described. Depending on whether the extract is liquid or dried, concentrated or not, the quantities of the active agent can vary in proportions from 0.002 to 40% by weight relative to the total weight of the composition.

- “signs of skin aging”, all changes in the external appearance of the skin and appendages due to aging such as, for example, wrinkles and fine lines, withered skin, flabby skin, thinning skin, lack of elasticity and/or tone of the skin, dull and lackluster skin or pigmentation spots on the skin, discoloration of the hair or stains on the nails, but also any internal change in the skin that does not appear systematically by a modified external appearance such as, for example, any internal degradation of the skin following exposure to ultraviolet (UV) radiation.

[26] The invention primarily relates to an extract of Moringa peregrina seed cake comprising mainly a peptide hydrolyzate which comprises amino acid derivatives, amino acids, peptides and glyco peptides whose molecular weight is between 100 Da and 6,000 Da, preferably between 1,500 Da and 5,000 Da, more preferably between 3,000 and 4,500 Da, and in that it is obtained from unshelled and mature seeds of the fruit of Moringa peregrina by chemical proteolysis at pH greater than 13 for a period of approximately 2 hours at a temperature between 16 and 25°C.

[27] The peptide hydralyzate, comprising peptides, oligopeptides, glycopeptides and amino acids, such as in the profile defined below, has never been demonstrated in an extract of the seeds of the genus Moringa peregrina. The species Moringa peregrina grows in very arid climates. Thus, its ability to adapt to drought and to reproduce in extremophilic conditions allowed it to acquire particular unique characteristics, which the Applicant was able to identify by implementing a specific extraction process on Moringa peregrina seed cake.

[28] In the context of the present invention, the plant part chosen is Moringa peregrina seed cake. It is known that the seeds of Moringa peregrina are used for the extraction of their oil, useful for self-consumption or in various traditional medicinal treatments. The cake obtained after the seed has been de-oiled is a waste currently used for animal feed in particular.

[29] According to a preferred embodiment, the extract according to the invention is obtained from the cake of unshelled seeds of Moringa peregrina.

[30] According to one embodiment, the peptide hydrolyzate of the extract according to the invention comprises amino acid derivatives, amino acids, peptides and glycopeptides whose molecular weight is between 100 Da and 6000 Da, and more particularly between 1,500 Da and 5,000 Da, and even more particularly between 3,000 Da and 4,500 Da.

[31] According to another embodiment, the extract also comprises between 0.3% and 3% of volatile compounds, including 50% of these compounds, or between 0.15% and 1.5% of the extract according to the invention, consists of light nitrilated compounds, mainly isobutyronitrile and methyl-butanenitrie; of which 5 to 10% of these compounds, or between 0.015 and 0.3% of the extract according to the invention, consists of isothiocyanate derivatives, mainly isopropyl isothiocyanate and isobutyl isothiocyanate; of which 1 to 5% of these compounds, or between 0.003 and 0.15% of the extract according to the invention, consists of essential oil, mainly Eucalyptol, Menthol and Benzaldehyde.

[32] The second object of the invention is a process for an extract of Moringa peregrina seed cake according to the invention comprising the following steps according to which:

a) we collect the unshelled seeds harvested when ripe from the fruit of Moringa peregrina which we dry to obtain an internal humidity level of less than 8%,

b) the dried seeds are pressed so as to separate the oil from the rest of the seed so as to obtain the cake comprising less than 6% by weight of residual oil,

c) we grind the cake obtained in step b), ίο

d) the ground cake obtained in step c is dispersed in the aqueous phase),

e) chemical proteolysis is carried out on the aqueous dispersion obtained in step d) for a period of approximately 2 hours, at a pH greater than 13 and at a temperature between 16 and 25°C,

f) the proteolysis is neutralized to stabilize the peptide hydrolyzate obtained, g) the peptide hydrolyzate is recovered by solid/liquid separation, h) the peptide hydrolyzate is purified by ultrafiltration, then optionally, i) the lyophilization is carried out. peptide hydrolyzate obtained in step h).

[33] We collect the unshelled seeds, that is to say the shell (pericarp) of which we keep, when the fruit is ripe and preferably when the pod is at the beginning of dehiscence.

[34] The seeds are dried to obtain an internal humidity level of less than 8% and preferably around 6%; drying will preferably be done on a ventilated shelf sheltered from the sun's rays, preferably under a shade in the open air.

[35] The dried seeds are then crushed immediately by cold pressing, which allows the oil to be mechanically separated from the rest of the compressed seed, namely the oilcake.

[36] The cake is then crushed mechanically with all types of mechanical crushers such as hammer, flail, knife, crushing/shredding, ball or ball or pestle systems, but also with any type of cryomill.

[37] The dispersion in the aqueous phase according to step d) and the proteolysis according to step e) are advantageously always carried out with stirring, thus allowing dispersion and homogenization of the solid in the liquid, thus improving the exchange surface. overall and consequently proteolysis.

[38] A liquid peptide hydrolyzate is obtained with a density greater than 1 and preferably around 1.1, comprising a dry matter content of between 10 and 15%, preferably around 12.5% comprising between 1 and 6% of nitrogen compounds, in particular volatile nitrile derivatives in a proportion of 0.5 to 1.5%, preferably around 0.8% and 20 mg/liter of polyphenols (0.0002%).

[39] According to a preferred embodiment of the process according to the invention, the solid/liquid separation of step g) is carried out by different processes such as centrifugation, spinning, filtration.

[40] According to another preferred embodiment of the process according to the invention, the ultrafiltration in step h) is carried out with a cut-off threshold of between 100 Da and

000 Da and more particularly between 1,500 Da and 5,000 Da, and even more particularly between 3,000 Da and 4,500 Da.

[41] In one embodiment according to the invention, the process for obtaining a seed cake extract is obtained from the unshelled seed cake by S proteolysis, with stirring, in a proportion of at most 25 % by weight of solid material relative to the total weight used in the aqueous solvent, at a temperature between 16 and 25°C for a period of approximately 2 hours.

[42] In one embodiment of the process for obtaining the liquid Moringa peregrina extract obtained, the peptide hydrolyzate is purified by distillation, microfiltration, ultrafiltration or nanofiltration to concentrate the extract into compounds of interest relative to the volatile nitrite derivatives also extracted. These purification steps make it possible to concentrate the pool of compounds of interest at the expense of other extracted compounds as mentioned.

[43] In another embodiment of the extraction process according to the invention, the liquid extract obtained is dried so as to obtain a dry extract of the seed cake.

Moringa peregrina containing a quantity greater than 10%, preferably greater than 20%, even preferably greater than 30% and which can reach up to approximately 40% (Mass/Mass) of dry matter in peptides, oligopeptides, glycopeptides and amino acids or their volatile nitrilated derivatives, preferably another 50% of the weight of the dry matter.

[44] According to one embodiment of the invention, the liquid Moringa peregrina seed cake extract obtained is preferentially dried for example by atomization, freeze-drying or zeodration so as to obtain a solid Moringa peregrina seed extract, the water being evaporated. Drying can be carried out in the presence of a drying support additive such as maltodextrin, cyclodextrin or inulin, or in the presence of an inorganic support such as phyllosilicate, magnesium silicate or carbonate and its salts.

[45] The invention also relates to the extract of Moringa peregrina seed cake which can be obtained by the extraction process according to the invention.

[46] The third object of the invention is a cosmetic or nutricosmetic composition comprising, as active agent, an effective quantity of an extract of Moringa peregrina seed cake according to the invention, and a physiologically acceptable excipient.

[47] The composition according to the invention can be formulated in the form of different preparations suitable for topical administration or oral administration.

According to a first variant, the different preparations are suitable for topical administration and include creams, oil-in-water and water-in-oil emulsions, milks, ointments, lotions, oils, balms, aqueous or hydro solutions. -alcoholic or glycolic, serums, powders, patches, sprays or any other product for external application such as, but not limited to, medical devices or cosmetic-textile products.

According to a second variant, the different preparations are suitable for oral administration; the plant extract comprising the peptide hydrolyzate which can be used either in a food composition or in a food supplement. The food supplement may be in the form of capsules or soft gelatin or vegetable capsules in the context of the present invention. Said food supplement can then contain from 0.01 to 100% by weight of the plant extract.

[48] In the context of food use, for nutritional or cosmetic purposes (cosmetofood or nutricosmetics), the composition will advantageously be formulated in the form of a preparation suitable for oral administration. It may not include an excipient and may consist entirely of the plant extract comprising the peptide hydrolyzate in dried form.

[49] According to a preferred embodiment, the compositions according to the invention are intended more particularly for topical administration. These compositions must therefore contain a cosmetically acceptable medium, that is to say compatible with the skin and appendages, and cover all cosmetic forms. These compositions may in particular be in the form of creams, oil-in-water emulsions, or water-in-oil emulsions or multiple emulsions, serums, solutions, suspensions, gels, milks, lotions, sticks or even powders, and suitable for application on the skin. skin, lips and/or your appendages. These compositions include the excipients necessary for their formulation, such as solvents, emollients, thickeners, diluents, surfactants, antioxidants, bioactive agents, dyes, preservatives, perfumes. They can be used as a care product and/or as a skin makeup product.

[50] The composition according to the invention may in particular consist of a composition for hair care, and in particular a shampoo, a conditioner, a treatment lotion, a styling cream or gel, a restructuring lotion for the hair, a mask , etc. The cosmetic composition according to the invention can be used in particular in treatments involving an application which is followed or not by rinsing, or even in the form of shampoo. The composition according to the invention can advantageously be used in anti-dandruff treatments. It can also be in the form of a dye or mascara to be applied with a brush or comb, particularly on the eyelashes, eyebrows or hair.

[51] The compositions according to the invention further comprise any additive commonly used in the envisaged field of application as well as the adjuvants necessary for their formulation, such as solvents, thickeners, diluents, anti-oxidants, dyes, sun filters, self-tanning agents, pigments, fillers, preservatives, perfumes, odor absorbers, cosmetic or pharmaceutical active ingredients, essential oils, vitamins, essential fatty acids, surfactants, film-forming polymers, etc.

[52] The INCI Dictionary & Handbook (“International Nomenclature of Cosmetic Ingredients” ( ^13th ed. 2010) published by The Personal Care Products Council Inc., Washington, DC) describes a wide variety, without limitation, of cosmetic ingredients and pharmaceuticals commonly used in the skin care industry, which are suitable for use as additional ingredients in the compositions according to the present invention.

[53] In all cases, those skilled in the art will ensure that these adjuvants as well as their proportions are chosen in such a way as not to harm the desired advantageous properties of the composition according to the invention.

[54] According to an advantageous embodiment of the invention, the quantity or content of plant extract in the cosmetic composition according to the invention is from 0.002 to 20% by weight, and in particular from 0.01 to 10% by weight relative to to the total weight of the composition.

[55] According to another advantageous embodiment of the invention, the quantity or content of plant extract in the nutricosmetic composition according to the invention is from 0.01 to 100% by weight relative to the total weight of the composition. More preferably, the quantity of plant extract is 0.02 to 40% by weight, and in particular 0.2 to 20% by weight relative to the total weight of the composition.

[56] The fourth object of the invention is the cosmetic or nutricosmetic use of a composition according to the invention to improve the appearance of the skin, mucous membranes or appendages, prevent and/or combat dryness of the skin and mucous membranes, prevent and/or combat the skin signs of aging and/or photoaging, promote whitening of the skin and promote thinning.

[57] Although the invention has been described in connection with several particular embodiments, it is obvious that it is in no way limited there and that it includes all the technical equivalents of the means described as well as their combinations if these fall within the scope of the Invention.

[58 1 The use of the verb “include”, “understand” or “include” and its conjugated forms does not exclude the presence of other elements or other steps than those set out in a claim.

Examples

[59] Example 1: Preparation of a plant extract according to the invention from Moringa peregrina cake

[60] Seeds of Moringa peregrina (Forssk.) Fîori obtained when the fruit was ripe were dried to obtain an internal humidity level of less than 8% and preferably around 6%, then pressed with a mechanical screw press. , so as to separate the oil from the rest of the seed to obtain on the one hand virgin peregrina oil (INC name! “Moringa peregrina seed oil”) and on the other hand a cake. The cake is then isolated in the form of sausages pre-cut into pieces of 1 to 2 cm on which the extraction is carried out. The composition of raw materials used in the process is given below.

[61] [Table 1]

Materials Supplier % Actual weighed quantity Peregrina cake *pre-cut into pieces of 1 to 2 cm 8.4% 33.6 Network water 80.68% 322.7 Sodium hydroxide PROCH1MIA 3.36% 13.5 Citric acid monohydrate F6000 ADETIS 7.14% 28.7 Sodium benzoate ADETÎS 0.42% 1.7 __Total 100 400.2

[62] Protocol:

[63] a) A concentrated solution of 1 molar of a strong alkaline agent such as sodium hydroxide, potassium hydroxide or calcium hydroxide is prepared, or of an aqueous mixture concentrating 1 molar of strong alkaline agents, but preferably sodium hydroxide; this strong alkaline solution has a pH greater than 13,

[64] b) We weigh in a ratio [1:10] (mass/mass) peregrina cake precut into pieces of approximately 1cm in alkaline solution,

[65] c) we grind the cake obtained in step b),

[66] d) The ground cake obtained in step c) is dispersed in the aqueous phase.

[67] e) chemical proteolysis is carried out on the aqueous dispersion obtained in step d) for a period of approximately 2 hours at a temperature of 20°C,

[68] f) proteolysis is neutralized to stabilize the peptide hydrolyzate obtained with a weak acid so as to buffer the solution in a pH range 4.5-5.5,

[69] g) the peptide hydrolyzate is recovered by solid/liquid separation,

[70] h) the peptide hydrolyzate is purified by ultrafiltration.

[71] A translucent yellow filtrate containing approximately 12.48% dry matter is obtained. The liquid extract obtained is subsequently named “peregrina peptide hydrolyzate according to the invention” or “peregrina peptide hydrolyzate”. This liquid extract is subsequently used for effectiveness tests.

[72] This peregrina peptide hydrolyzate according to the invention has a density greater than 1 and preferably around 1.1, comprising a dry matter content of between 10 and 15%, preferably around 12.5% comprising between 1 and 6 % of nitrogen compounds, mainly peptides and glycopeptides and also volatile nitrile derivatives in a proportion of 0.5 to 1.5%, preferably around 0.8% and mg/liter of polyphenols (0.0002%). The composition of the volatile compounds of the peregrina hydrolyzate according to the invention is found below.

[73] [Table 2]

Tr CAS No. Compounds % DM 4.42. 64-17-5 Ethanol 0.809 5.58 67-64-1 Acetone 0.444 6.10 75-15-0 Carbon disulfide 0.327 7.83 78-93-3 2-Butanone 0.165 8.57 141-78-6 Ethyl acetate 0.047 8.99 78-82-0 Isobutyronitrile 20,720 10.39 78-82-0 Isobutyronitrile 0.127 12.06 547-63-7 Methyl isobutyrate 0.201 14.10 18936-17-9 2-Methyl l-b utanen itri le 2.194 14.56 625-28-5 3-Methyl-butanenitriie 27,495 18.77 66-25-1 Hexanal 0.186 21.09 2253-73-8 Isopropyl isothiocyanate 4,590 23.65 628-73-9 Hexanenitrile 0.198 24.43 11 CM 3-0 2-Heptanone 0.093 27.18 4426-79-3 2-isoth rocyan ato-Butane 1,058 27.59 80-56-8 Alpha-Pinene 0.048 28.45 591-82-2 Isobutyl isothiocyanate 2,299 28.99 100-52-7 Benzaldehyde 1,753 29.77 108-95-2 Phenol 0.413

30.80 13475-82-6 2,2,4,6,6-pentamethyl-Heptane θ-318 32.80 99-87-6 Para-Cymene 0.232 33.10 138-86-3 Limonene 0.133 __ 33.34 470-82-6 Eucalyptol 0.918 36.45 1195-32-0 Para-Cymenene 0.035__ 36.88 124-19-6 Nonanal 0.080 __ 40.26 65-85-0 Benzoic acid 19,150 41.07 1490-04-6 Menthol 0.918 56.29 96-76-4 2,4-Di-telt-butylphenol 0.086 __________________________Total 85.035

The peregrina peptide hydrolyzate contains isopropyl isothiocyanate and isobutyl isothiocyanate at relatively high levels, confirming previous publications on the species (Kjaer et al. 1979, Isothiocyanates in Myrosinase-treated seed extracts of Moringa peregrina, Phytochemistry, 18, pp. 1485-1487; Afsharypuor et al., 2010, Volatile Constituents of the Seed Kemel and Leaf of Moringa peregrina (Forssk.) Fiori, Agricolt. Cultivated in Chabahar (Iran), Iranian Journal of Pharmaceutical Sciences 6(2): 141-144; Dehshahri S. et al., 2012, Determination of volatile glucosinate degradation products in seed coat, stem and in vitro cultures of Moringa peregrina (Forssk.) Fiori, ScienceOpen, Research in Pharmaceutical Sciences 7 (1): 51-56). Isothiocyanates are compounds produced by various plants belonging to the order Brassicales, notably in the families Brassicaceae, Capparaceae, Caricaceae and Moringaceae as a defense system against attack by pathogens. In the genus Moringa, they have been identified notably in M. oleifera and M. stenopetala (Baker f.) Cufold. (Abd Rani N.Z., Kharaina, H. and Kumolosasi, E., 2018, Moringa genus; A review of Phytochemistry and Pharmacology, Frontiers in Pharmacology, vol. 9, art. 108). Isothiocyanates come from the hydrolysis of glucosinolates by the enzyme myrosinase when plant tissues are damaged. Isothiocyanates have been reported to have various biological effects, such as antifungal activity (Troncoso-Rojas, R. and Tiznado-Hernandez, Μ. E., 2007, Natural compounds to control fungal diseases in fruits & vegetables, in Troncoso -Rojas, R,, Tiznado-Hernandez, Μ. E., Gonzalez-Leon, A. (ed) Recent advances in alternative postharvest technologies to control fungal diseases in fruits & vegetables. Editorial. Transworld Research Network, Kerala, India, pp 127-156; Troncoso-Rojas, R. et al., 2005, Analysis of the isothiocyanates present in cabbage leaves extract and their potential application to control Aiternaria rot in bell peppers, Food Research International 38:701708), anti-microbial effects, anti-cancer and anti-inflammatory (Park, EJ. et al., 2011, Inhibition of lipopolysaccharide induced cyclooxygenase-2 expression and inducible nitric oxide synthase by 4-[(2 '-O-acetyl-a-L-rhamnosyloxy)benzyl]isothiocyanatefrom M. oleifera, Nutrition and Cancer 63(6): 971-982; Rajan, T. S. et al., 2016, Anticancer activity of glucomoringin isothiocyanate in human malignant astrocytoma cells, Fitoterapa 110:17; Radia E. P. et al., 2012, Antimicrobial isothiocyanates from the seeds of Moringa oleifera Lam., Zeitschrift für Naturforschung C, 67, 557-564; WATERMAN, C, et al., 2014, Stable, water extractable isothiocyanates from Moringa oleifera leaves attenuate inflammation in vitro. Phytochemistry 103: 114-122). Furfural is present at a very standard concentration, found in this type of seeds and in many dried fruits. Carbon disulfide, isobutyronitrile, methyl isobutyrate, methyl butanenitrile and hexanenitrile are volatile compounds derived from amino acids. These are markers of protein degradation. These degradation compounds represent approximately 50% of the volatile compounds in the hydrolyzed peregrina cake extract. This result indicates that the extract consists mainly of proteins or peptides. Only very small traces of fats or free sugars were detected in the hydrolyzed extract of Moringa peregrina, and essential oils, these representing 4% of the volatile compounds with the presence of Menthol at approximately 1%. , the presence of Eucalyptol at 1% and the presence of Benzaldhéyde at approximately 2%. In the dry matter, the citrate buffer (which is an saccharide compound) represents approximately 50% of the remainder and includes the glycosylated (saccharide) compounds resulting from the degradation by proteolysis of proteins, oligopeptides and amino acids. Finally, it should be noted the presence of approximately 20 mg/liter of polyphenols (0.0002%) from peregrina seeds. Benzoic acid should not be considered in the characterization because it is a stabilizing agent added to the extract. This liquid filtrate composes the extract according to the invention and is used pure in the tests which follow.

[74] The dry matter of the extract described above is obtained by a gravimetric method based on the mass before and after evaporation present in the liquid extract. This dry mass is mainly composed of proteins and glycoproteins, whose molecular weight is between 100 and 6000 Da, partially degraded and water-soluble which are called oligopeptides or glyco-oligopeptides or peptide hydrolyzate according to the invention, resulting from the hydrolysis of Moringa peregrina cake. The dry extract also includes a sodium citrate buffer which stabilizes these oligopeptides and glycooligopeptides in a static hydrolyzed state. Finally, this extract is microbiologically stabilized with a water-soluble preservative such as sodium benzoate.

[75] Example 2: Effect of the peregrina peptide hydrolyzate according to the invention as an anti-oxidant

[76] The objective of this study is to evaluate the modulation of antioxidant activity by the peptide hydrolyzate peregrina in an acellular in vitro colorimetric model using the DPPH radical (2,2-diphenyl-1-picrylhydrazyl ) as well as the reference antioxidant, ascorbic acid. The method used is called inhibition. It is based on the degradation of the oxidizing radical DPPH, purple in color absorbing at 540 nm, by a reference antioxidant, ascorbic acid. This reaction serves as a positive control and leads to the formation of the compound DPPH which is colorless or even light yellow. The peregrina peptide hydrolyzate according to the invention and the reference product “Ascorbic Acid” are placed in contact with the DPPH solution for 30 min at 40°C. The antioxidant activity is then evaluated by measuring absorbance at 540 nm. The modulation of this activity is expressed as a percentage of stimulation of the antioxidant activity by the active ingredient tested, with the maximum antioxidant activity obtained in the presence of ascorbic acid (T ⁺ ) as a reference.

[77] Protocol: A solution of DPPH is incubated for 30 minutes at 40°C, in the absence (control), in the presence of the peptide hydrolyzate of peregrina according to the invention (T*) and at decreasing concentrations of sample to be tested. At the end of the incubation period, the antioxidant activity in the presence of the reference product and in the presence or absence of the peregrina peptide hydrolyzate was revealed by coloring after 30 minutes at 40°C. It was thus evaluated by measuring the absorbance of the reaction medium at 540 nm. For each concentration tested, the modulation of the antioxidant activity by the product tested is calculated according to the following formula.

[78] [Math. 1] Percentage of modulation of antioxidant activity = 100 x [(ODsm Control - DÛ540 Product to be tested) / DOmc Reference product].

[79] If the result is negative, the product to be tested will be considered as oxidizing; if the result is positive, the percentage will be expressed as stimulation of anti-radical activity. The results obtained are given below for the inhibition of DPPH by the peregrina peptide hydrolyzate according to the invention.

[80] [Table 3]

DPPH inhibition peregrina peptide hydrolyzate according to the invention 2% 12 1.0% 13 0.1% 2

[81] Conclusion: The peregrina peptide hydrolyzate according to the invention is capable of protecting free radicals, it presents significant anti-oxidant properties from a concentration of 1%.

[82] Example 3: Effect of the peregrina extract according to the invention as an inhibitor of metalloproteinases

[83] The objective of this study is to evaluate the modulation of the anti-Metalloprotease activity by the peregrina peptide hydrolyzate according to the invention in an acellular in vitro model using a type I collagenase and a hyaluronidase, a substrate complex and a chromophore, Ninhydrin. A buffered solution of Collagenase type I and Hyaluronidase reacts with a specific substrate complex and transforms it to form a compound capable of activating a chromophore by incubation at 80°C for 15 min. The activity of Collagenase and Hyaluronidase can thus be evaluated by measuring the absorbance at 565 nm. The sample is placed in contact with the Collagenase and Hyaluronidase solution at the same time as the enzyme substrate complex at 37°C for 5 minutes. The substrate transformed by the enzymes is capable of activating the chromophore by incubation at 80 ^° C for 15 minutes. The activity of Collagenase and Hyaluronidase in the presence/absence of the sample is then evaluated by measuring the absorbance at 565 nm. The modulation of this activity is expressed as a percentage of inhibition or activation of the activity of Collagenase and Hyaluronidase in the absence of active ingredient, that is to say only in the presence of the enzyme substrate.

[84] Protocol: A solution of Collagenase type I and Hyaluronidase enzymes is incubated in its substrate, for 5 minutes, in the absence or presence of the peregrina extract according to the invention tested. The solutions are then brought into contact with the chromogen Ninhydrin, before incubation for 15 minutes at 80°C. At the end of the incubation period, the activity of the Collagenase and Hyaluronidase enzymes with and without test or reference product was evaluated by measuring the absorbance of the reaction media at 565 nm. For each concentration tested, the modulation of the activity of the Collagenase and Hyaluronidase enzymes by the product under test is calculated according to the following formula.

[85] [Math. 2] Percentage of modulation of the activity of the Collagenase/Hyaluronidase enzymes = 100 x [(OD test or reference product - OD Collagenase/Hyaluronidase alone)/D0 Collagenase/Hyaluronidase alone].

[86] If the result is negative, the percentage is expressed as enzyme inhibition; if the result is positive, the percentage is expressed in enzyme activation. The results of the inhibition of Metalloproteases are given below.

[87] [Table 4]

Inhibition versus Control (%) 1% 100

Peregrina peptide hydrolyzate according to the invention 0.5% 100 0.1% 92 0.01% 50

[88] Conclusion: The peregrina peptide hydrolyzate according to the invention generates a strong inhibition of Metailoproteases (Collagenase! Hyaluronidase). It is capable of completely inhibiting these Metalloproteases from a concentration of 0.5% and has a good potential to protect with great effectiveness the extra-cellular matrix of the skin and, through this inhibition, it shows an anti-aging effect.

[89] Comparative tests are reported below with the extract obtained according to patent FR2946879 of Pierre Fabre, of which here are the results, according to the same test simply involving collagenase:

[90] [Table 5]

Percentage inhibition versus Control {%) Moringa oleîfera extract according to Pierre Fabre patent 1% Concentration not compatible with testing system 0.5% 4 0.1% 24 0.01% 42

[91] Conclusion; the extract according to the Pierre Fabre patent shows a slight inhibitory action on collagenase activity in inverse dose dependence with an inhibition peak of 42% for all concentrations combined compared to an inhibition peak of 100% for the peptide hydrolyzate seion the invention.

[92] The anti-aging activity on this parameter appears different and new compared to the effects observed with the extract according to the Pierre Fabre patent.

[93] Example 4: Effect of the peregrina peptide hydrolyzate according to the invention to inhibit the enzymes Histone Desacetylases (HDACs) and Sirtuin I

[94] The objective of this study is to demonstrate the inhibitory activity of the peregrina peptide hydrolyzate according to the invention on the HDACs and Sirtuin I enzymes. A buffered solution of HDACs & Sirtuin I reacts with a substrate for 20 minutes at 37°C and transforms it to form a compound which colors in the presence of a developer after incubation at 37°C for 10 minutes. The maximum deacetylation activity of Sirtuins can thus be evaluated by measuring the absorbance at 405 nm. The peregrina extract according to the invention or the reference product “Trichostatin A (STA) inhibitor 1μΜ” are brought into contact with the solution. of Sirtuins at the same time as the enzyme substrate for 20 minutes at 37°C, the substrate transformed by the enzyme is colored by the addition of a developer. The deacetylating activity of HDACs and Sirtuine i in the presence of the active ingredient is then evaluated by measuring the absorbance at 405 nm. The modulation of this activity is expressed as a percentage of inhibition or activation of the maximum activity of HDACs & Sirtuin! in the absence of active ingredients, that is to say only in the presence of the substrate of the HDACs & Sirtuin I enzymes.

[95] Protocol: A solution of Sirtuin enzymes is incubated in its substrate for 20 min in the absence (control) or presence of the reference product, or increasing concentrations of the products under test. The peregrina peptide hydrolyzate according to the invention is tested at the following concentrations: 2%; 1%; 0.1% (V/V). At the end of the incubation period, the activity of the Sirtuin enzymes with and without test or reference product was revealed by staining a developer solution (10 min at 37°C) and evaluated. by measuring the absorbance of the reaction media at 405 nm. For each concentration tested, the modulation of the deacetylating activity of the enzymes Histone Deacetysases and Sirtuins I by the product under test is calculated according to the following formula.

[96] [Math. 3] Percentage of modulation of the activity of Sirtuin enzymes = 100 x [(OD405 test or reference product) - (DO405 HDACs & Sirtuin I alone)]/D04os Sirtuins alone.

[97] If the result is negative, the percentage is expressed as inhibition of the enzymatic reaction; if the result is positive, the percentage is expressed as activation of the enzymatic reaction. The results of inhibition of Histone Desacetylase (HDACs) enzymes are given below.

[98] [Table 6]

Peregrina peptide hydrolyzate according to the invention percentage Inhibition versus Control (%) 2% 15 1% ns 0.10% -18

[99] Conclusion: At 2% The peregrina peptide hydrolyzate according to the invention shows significant inhibition of HDACs; this inhibition means the ability to promote the self-protection of skin cells against genetic drift, particularly linked to the aging process. Thus the extract appears to be useful against one of the most common genetic deviations on the surface of the skin, namely fibrosis which manifests itself by the appearance of flesh “pimples” (fibrotic protuberance). The extract can advantageously interfere with fibrosis phenomena on the surface of the skin and thus prevent aging of the skin.

[100] Example 5: Tensor effect of the peregrina peptide hydrolyzate according to the invention

[101] The objective of this study is to evaluate the tensor effect of the active peptide hydrolyzate peregrina in an in vitro acecular model of lyophilized collagen discs. Lyophilized collagen disks of 8 mm diameter were used. The contraction of these discs in response to the different treatments was measured by image analysis. The more the surface area of the collagen discs decreases, the greater the tightening effect of the active ingredients. The reference product used in this study is bovine serum albumin at 100 mg/mL.

[102] Protocol: Collagen disks 8 mm in diameter were soaked with 40 μΙ of ultra-pure water (control), of the reference product, or of increasing concentrations of the peregrina peptide hydrolyzate according to the invention: 0.05 ; 0.1 and 0.5% (v/v). The peregrina peptide hydrolyzate was solubilized directly in ultrapure water. Digital images of each of the discs were taken using a scanner, before and 30 minutes after imbibition. The surface area of the collagen discs was measured on the images using image analysis software. The results of the tensor effect are given below.

[103] [Table?]

Reduction Versus Control (%) Peregrina peptide hydrolyzate according to the invention 0.5% 51.1 0.1% 38.4 0.05% ns

[104] Conclusion: The peregrina peptide hydrolyzate according to the invention shows a very significant tightening effect at low concentration (less than 1%). These results demonstrate a “lifting” effect and therefore a short-term anti-aging effect.

[105] Examples 2 to 5 demonstrate that the peregrina peptide hydrolyzate according to the invention has the profile of a good anti-aging active ingredient, in the short or long term.

[106] Example 6: Effect of the peregrina peptide hydrolyzate according to the invention to inhibit the action of endothelin 1

[107] It has been reported in recent years that endothelin promotes an increase in intracellular calcium concentration in epidermal melanocytes (melanin cells) to facilitate cell growth through the signal transduction system! intracellular. This action improves the activity of tyrosinase which is a rate-determining enzyme in the synthesis of melanin (Kadono, S. et al., 2001, The Role of the Epidermal Endothelin Cascade in the Hyperpigmentation Mechanism of Lentigo Senilis, Journal of Investigative Dermatology , 116 (4): 571 -577). It has also been reported (Kawahara N. et al., 2006, About the Component of Snow Tea, Collection of Pharmaceutical, Society of Japan annual convention Subject matter, abstract 30-0823, p. 159) that endothelin is a factor of melanocyte activation produced by epidermal keratinocytes, as well as an important factor in ultraviolet light-induced pigmentation or senile dark spot (Pang, Y., Geng, 1, Qin, Y. et al. Endothelin-l increases melanin synthesis in an established sheep skin melanocyte culture. In Vitro Cellular Æ Developmenlal Biology - Animal 52, 749-756 (2016) or HACHIYA, AKIRA etal. Biochemical Characterization of Endothelin-converting Enzyme-1a in Cultured Skinderived Cells and Its Postulated Role in the Stimulation of Melanogenesis in Human Epidermis. Journal of Biological Chemistry, Volume 277, Issue 7,5395-540). These biological actions of endothelin suggest that active ingredients capable of inhibiting the action of endothelin may be useful in reducing or preventing melanin production or pigmentation.

[108] Endothelin is the most potent vasoconstrictor known in the human body. On the other hand, the decrease in endothelin is also known to create a vasodilator effect (Hirata, Y. et al, 1988, Cellular mechanism of action by a novel vasoconstrictor endothelin in cultured rat vascular smooth muscle cells, Biochemical and Biophysical Research Communications, 154(3): 868-875; Shalinkumar P. et al., 2008, H2S Mediates the Vasodilator Effect of Endothelin-1 in the Cerebral Circulation. American Journal ofPhysiology. Heart Circulatory Physiology, 315, pp. 1759-1764 ).

[109] The objective is to determine type 1 endothelin in human microvascular endothelial cells after exposure for 24 hours to the peregrina peptide hydrolyzate according to the invention.

[110] Protocol: Human microvascular endothelial cells were supplied by the company PELOBiotech and cultured in 96-well plates according to the supplier's production procedures. This involves letting the extracts act at different concentrations on endothelial cells at 80% confluence for 24 hours, then quantifying endothelin 1 using the PicoKine ELISA kit (EDN1) in the cell supernatants. A viability test is carried out beforehand to define the non-toxic doses to be used when measuring endothelin 1. The negative control is carried out using the cells in culture medium without treatment. The positive control in the viability test is S DS at 0.5%.

All conditions are prepared in culture media, and the cells are then incubated at 36.5°C 15% CO2 for 24 hours.

a) Application of the solutions to be tested on endothelial cells:

The products to be tested are placed in contact with the sub-confluent endothelial cells in 96-well plates. For each concentration the test is carried out on 3 wells. The plates are incubated for 24 hours ± 1 hour at 36.5°C / 5% CO2.

b) Viability test:

Cell viability is evaluated by the MTT method on the cells after incubation with the products. After 24 hours of incubation, the supernatants are collected and stored at -20°C for assays. The wells are then rinsed once with 200 μL of PBS. In each well, 50 μL of MTT solution at 0.5 mg/ml are added: incubation for 3 hours at 36.5°C i 5% CO2. In each well 100 μL of isopropanol are added. After homogenization, an absorbance reading is taken at 550 nm. For each condition, the ratio of the average optical densities of the cells to the average optical densities of the negative controls will determine the viability rate.

c) Dosage of endothelin 1:

The assay is carried out using the ELISA kit. The results of inhibition of endothelin action are given below.

[111] [Table 8]

Extract concentration Cell growth versus control (%) Endothelin 1 versus control (%) Endothelin 1 versus control (pg/ml) Peregrina extract according to the invention 2% +8.46 -34.90 -47.09 1% 1.54 -13.03 -17.58 0.10% -1.15 -12.44 -16.79

[112j Conclusion: The viability test carried out at the end of treatment did not show any toxic effect for the concentrations tested.

Endothelin 1 is assayed in cell supernatants at non-toxic concentrations. The quantity of endothelin 1 for each condition is measured using the ELISA kit.

For the negative control cells, the values are around 134.94 pg/ml. For cells treated with different concentrations of extracts, the values are from 87.85 pg/ml (with 2% of the extract according to the invention) to 118.15 pg/ml (with 0.1% of the extract according to the invention) , which shows very significant inhibitions from 0.1% of the extract according to the invention with approximately 12.44% inhibition of type 1 endothelin production and up to 34.90% inhibition with 2% of the extract according to the invention. 'invention.

The results on endothelin and its specific dose-dependent inhibition demonstrate that the peptide hydrolyzate according to the invention is a powerful skin lightener.

[113] Example 7: Effect of the peregrina peptide hydrolyzate according to the invention to release free fatty acids.

[114] Lipolysis in adipocytes, or the hydrolysis of triacylglycerol (TAG) to release fatty acids and glycerol for use by other organs, is a unique function of white adipose tissue. Lipolysis in adipocytes occurs on the surface of cytosolic lipid droplets, which have recently attracted much attention as dynamic organelles, an integral part of lipid metabolism. The recent identification of AdPLA as the primary adipose phospholipase A(2) led to the discovery of dominant autocrine/paracrine regulation of lipolysis by PGE (2). The above mechanisms are key players in lipolysis and its regulation. Recent discoveries link lipolysis to genetic alterations or mutations and predict the activation of lipolysis in adipocytes as a potential therapeutic target (Ahmadjan M. et al., 2010, Lipolysis in Adipocytes, The International Journal of Biochemistry and CellBiology, 42( 5); 555-559).

[115] The objective of the study is to evaluate the capacity of the peptide hydrolyzate according to the invention to increase the release of glycerol in the culture of human cells (adipocytes). Glycerol content correlates with lipolysis. The increase in glycerol content implies a lipolysis effect. The cosmetic slimming effect is associated with the lipolysis effect.

[116] Protocol: Human adipocytes are cultured in 24- and 12-well plates according to internal procedures. This involves letting the samples act at defined concentrations on confluent adipocytes for 24 hours. A preliminary viability test with MTT after 24 hours makes it possible to assess cytotoxicity and choose the concentrations for the “lipolytic effect” test. This “lipolytic effect” is evaluated by measuring the glycerol in the supernatants after 24 hours of exposure to the samples.

[117] The negative control is carried out using the cells in culture medium without treatment.

The positive control for the viability test is 0.5% SDS. The positive control for the “lipolytic effect” test is caffeine at 0.05%. All conditions are prepared in culture media, and the cells are then incubated at 36.5°C l 5% CO2 for 24 ± hours.

• Application of solutions to be tested on adipocytes:

- The products to be tested are placed in contact with the confluent adipocytes in the 24-well (cytotoxicity test) and 12-well plates (“lipolytic effect” test).

- For each concentration, the test is carried out on 3 wells.

- The plates are incubated for 24 ± 1 hours at 36.5°C 15% CO2.

• Viability test:

Cell viability is evaluated by the MTT method on the cells after 24 hours of incubation with the products.

- After 24 hours of incubation, the planned wells are rinsed once with 200pL of PBS.

In each well, 300 μl of MTT solution at 0.5 mg/ml are added: incubation for 3 hours at 36.5°C/5% CO2.

- In each well 800 μΙ of isopropanol are added.

- After homogenization, an absorbance reading is taken at 550 nm.

- For each condition, the ratio of the average optical densities of the cells to the average optical densities of the negative controls will determine the viability rate.

- A viability threshold value of 70% compared to the negative control value is used to classify the substances tested as cytotoxic or non-cytotoxic.

• “Lipolytic effect” test:

- The lipolytic effect is evaluated by the dosage of glycerol in the culture media after 24+1 hours of incubation with the products.

- The culture media are mixed with the “free glycerol reagent” according to the supplier’s protocol.

The glycerol solution is used to make the standard range at concentrations from 0 to 6250 μΜ.

The OD reading is carried out at 550 nm. The results of the release of free fatty acids are given below.

[118] [Table 9]

Concentration Cell growth versus control (Caffeine, %) I Increase in glycerol production versus control (%) Glycerol Production Production (pM) peregrina peptide hydrolyzate 2% -6.16 +40.00 8.32 0.5% -7 +3.00 0.63 0.10% +9.03 NS 0

[119] Conclusion: The peptide hydrolyzate according to the invention showed that it promoted the release of glycerol from human adipocytes and therefore induced a slimming effect.

[120} Example 8: Effect of the peregrina extract according to the invention to stimulate the ZAG protein

[121] Zinc alpha-2-glycoprotein (ZAG) is a plasma glycoprotein that takes its name from its electrophoretic mobility and its ability to be precipitated by Zn salts. ZAG is a member of the immunoglobulin gene superfamily and has a three-dimensional structure highly homologous to MHC class I and II molecules. ZAG was detected immunohistochemically in normal secretory epithelial cells of the breast, prostate and liver, in salivary, bronchial, gastrointestinal and sweat glands and in normal stratified epithelia, including the epidermis. ZAG mRNA remains uniformly distributed in different cell types [Za7]. Due to its production by the secretory epithelium, ZAG is present in most body secretions and constitutes 2.5% of saliva proteins and 30% respectively. % of seminal fluids.

[122] The standard functions of ZAG are unclear. However, ZAG has been isolated from the urine of human cancer patients with cachexia and may function as a lipid mobilization factor. Purified ZAG extracted from human or mouse serum, or extracted from human urine in cancer patients, induces lipolysis, resulting in the release of glycerol. The latter also increases the use of lipids in human and mouse adipocytes (Hirai K. et al., 1998, Biological Evaluation of a LipidMobilizing Factor Isolated from the Urine of Cancer Patients, Cancer Research, 58(11): 2359- 2365). ZAG activates guanosine triphosphate (GTP)-dependent adenylate cyclase activity on adipocyte membranes, increasing cellular cyclic adenosine monophosphate (cAMP) levels. This can potentially lead to the activation of multiple cellular pathways.

[123] To further investigate the biological properties of ZAG, stable transfers of recombinant human ZAG (rh) were created in the murine melanoma cell line B16F10. The effect of ZAG transfection on melanin production was determined in vitro and in vivo. Finally, the effect of ZAG on tyrosinase expression and activity was determined. Overall, these studies show that ZAG inhibits melanin production in normal and malignant melanocytes. Mechanisms include post-transcriptional effects on protein tyrosinase, with the potential for additional indirect effects. These studies made it possible to identify a previously unknown biological function of ZAG and made possible a method of modulating melanin production, thus preventing and/or reducing pigmentation of the skin and hair due to the increase in melanin production (Ηλε L, Method of Modulating Melanin Production, United States Patent No. US7.803.750 B2, 2010) and (Ghada F. M. et al., 2015, Highlights in Pathogenesis of Vitiligo, World Journal ofClinical Cases 3(3) ; 221-230).

[124] The objective of this study is to evaluate the capacity of the peregrina peptide hydrolyzate according to the invention to increase the release of ZAG in its culture of human cells (keratinocytes). The ZAG content is correlated with a whitening effect, a slimming effect, an anti-fibrotic effect and a soothing effect.

[125] Protocol: Normal human keratinocytes are isolated from foreskins and cultured in 24- and 96-well plates according to internal procedures.

[126] This involves letting the samples act at defined concentrations on the keratinocytes at 80% confluence for 48 hours, then quantifying the ZAG using the ELISA kit in the cell supernatants.

[127] A preliminary viability test is carried out to define the non-toxic doses to be used when measuring ZAG.

[128] The negative control is carried out using the cells in culture medium without treatment.

[129] The positive control for the viability test is 0.5% SDS.

[130] All conditions are prepared in culture media, and the cells are then incubated at 36.5°C / 5% CO2 for 24 hours for the viability test and 48 hours for the ZAG assay.

• Application of solutions to be tested on keratinocytes;

The products to be tested are placed in contact with the subconfluent keratinocytes in the 24 and 96 well plates.

For each concentration, the test is carried out on 3 wells.

The plates are incubated for 24 hours and 48 hours at 36.5°C / 5% CO2.

• Viability test:

Cell viability is evaluated by the MTT method on the cells after incubation with the products.

- After 24 and 48 hours of incubation, the planned wells are rinsed once with 200 μL of PBS.

In each well, 50 μl of MTT solution at 0.5 mg/ml are added: incubation for 3 hours at 36.5°C/5% CO2.

In each well 100 μ) of isopropanol are added.

After homogenization, an absorbance reading is taken at 550 nm.

- For each condition, the ratio of the average optical densities of the cells to the average optical densities of the negative controls will determine the viability rate.

• ZAG protein dosage:

- After 48 hours of incubation, all supernatants are collected and stored at -20°C for assays.

The assay is carried out using an ELISA kit. The results of the ZAG assay are given below.

[131] [Table 10]

Concentration Cell growth versus control (%) ZAG versus control (%) ZAG versus control (ng/ml) peregrina peptide hydrolyzate 2% -17.88 +337.80 0.390__ 1% -12.54 +195.73 0.157 0.1 -3.88 +151.83 0.085

[132] Conclusion: Peregrina peptide hydrolyzate can significantly increase ZAG production, with good dose dependence and low toxicity to human cells. It therefore has a whitening effect, a slimming effect, an antifibrotic effect and a soothing effect based on its ability to significantly increase ZAG.

[133] Example 9: Effect of the peregrina peptide hydrolyzate according to the invention to inhibit the production of melanin

[134] Skin pigmentation is a complex process that, in both the epidermis and hair follicles, begins with the synthesis of melanin in melanosomes within melanocytes, followed by the transfer of melanosomes to surrounding keratinocytes, which in turn transport the pigment and eventually degrade it. In humans, the entire melanocyte population is located in the hair follicles and in the basal layer of the epidermis. Regardless of their location in the skin, melanocytes have a common embryological origin, the neural crest from which they derive in the form of melanoblasts (non-pigmented cells). There are two types of melanin in epidermal cells, eumelanin, a brown-black pigment, and pheomelanin, a yellow-red pigment. In melanocytes, eumelanosomes and pheomelanosomes coexist. Tyrosinase is the key enzyme that regulates the first steps in the synthesis of pheomelanin and eumelanin: the conversion of Ltyrosine to L-3,4-dihydroxyphenylalanine (L-DOPA) and the oxidation of this compound to dopaquinone. From dopaquinone, the synthesis pathways differ for eumelanin and pheomelanin. The main role of melanin is to protect the skin from the harmful effects of UV rays, thus preventing the development of skin cancer (Brenner M. et al., 2008,The Protective Role of Melanin Against UVdamage in human skin, Photochemistry and Photobiology 84(3): 539-549).

[135] The objective is to bring together all the data used, as well as the results obtained, for carrying out the melanin modulation test on human melanocytes after exposure for 5 days to the peregrina extract according to the invention .

Protocol: Human melanocytes are cultured in 96- and 24-well plates.

This involves letting the peregrina extract according to the invention act at concentrations of 5%; 2%; 1% and 0.1% on confluent melanocytes for 5 days. An MTT viability pre-test after 24 hours allows cytotoxicity to be assessed and concentrations to be chosen for the melanin modulation test. This modulation is evaluated by measuring melanin in cell lysates after 5 days of exposure to the extracts. The negative control is carried out using the cells in culture medium without treatment. The positive control for the viability test is 0.5% SDS. For the test of melanin modulation media with and without a -MSH are used as negative controls.

All conditions are prepared in culture media, and the cells are then incubated at 36.5°C / 5% CO2 for 24 hours for the cytotoxicity test and 5 days for the melanin assay.

a) Application of the solutions to be tested on the melanocytes: The concentrations to be tested are brought into contact with the confluent melanocytes in the 96 (cytotoxicity test) and 24 (melanin dosage) well plates. For each concentration the test is carried out on 3 wells. The plates are incubated for 24 ± 1 hours and 5 days at 36.5C/5% CO2.

b) Viability test: Cell viability is evaluated by the MTT method on the cells after 24 hours of incubation with the products. After 24 hours of incubation, the planned wells are rinsed once with 200 gL of PBS. In each well, 50 μΙ of MTT solution at 0.5 mg/mi are added and the incubation is carried out for 3 hours at 36.5°C / 5% CO2. In each well 150 μΙ of isopropanol are added. After homogenization, an absorbance reading is taken at 550 nm. For each condition, the ratio of the average optical densities of the cells to the average optical densities of the negative controls will determine the viability rate.

A viability threshold value of 70% relative to the negative control value is used to classify the substances tested as cytotoxic or non-cytotoxic. A “non-cytotoxic” classification is given on the in vitro results for viability >

70% and a “cytotoxic” classification for viability <70%.

The 5% concentration of the extract according to the invention was found to be cytotoxic at 5% under the test conditions. The concentrations 2%, 1% and 0.1% are therefore used for the melanin modulation test. The quantity of melanin present in the cells is measured after cell lysis. The results of inhibition of melanin production are given below.

[136] [Table 11]

Extract concentration Cell growth versus control (%) Melanin inhibition versus control (%) Melanin Content ____(l&ml)____ Peptide hydrolyzate according to the invention 2% -24.48 +16.50 152 1.0% -17.15 +55.00 82 0.10% -18.06 +71.40 52

[137] Conclusion: The peregrina peptide hydrolyzate according to the invention inhibits the production of melanin in cells, which gives it a depigmenting effect and a skin whitening property.

[138] Example 10: Effect of the peregrina peptide hydrolyzate according to the invention on the modulation of the dosage of DKK1 and DKK3

[139] The involvement of interactions between melanocytes and fibroblasts in the regulation of melanogenesis is well known and has been studied intensively. Although these interactions are not yet fully understood, they are the cause of the whiteness of the palmar-plantar areas and are now used in cosmetics for the development of depigmenting products Yamaguchi and colleagues (Yamaguchi Y. et al., 2004, Mesenchymal-Epithelial Interactions in the Skin: Increased Expression of Dickkopf by Palmoplantar Fibroblasts Inhibits Melanocyte Growth and Differentiation, Journal of Cell Biology 165(2): 275-285) demonstrated that a soluble messenger produced by fibroblasts of the palmoplantar regions , was able to modify the differentiation program of melanocytes in these regions, leading to a decrease in melanin production. This messenger was identified by the team as a protein called Dikkopf1 (DKK-1).

[140] The signaling pathways used by DKK-1 to produce these results are well identified today. Thanks to its antagonistic action on the Wnt receptor, DKK-1 is in fact capable of shunting the intracellular signaling pathways activated by βcatenin, generally responsible for the regulation of genes involved in melanogenesis. Yamaguchi and colleagues also demonstrated that DKK-3, a molecule close to DKK-1, but without effect on the Wnt receptor, could play a regulatory role on the effect of DKK-1. In fact, the greater the quantity of DKK-3 in the vicinity of this Wnt receptor, the weaker the interactions between DKK-1 and this receptor. Increased DKK3 reduces the inhibitory effects of DKK-1 on melanogenesis. The work of Yamaguchi and colleagues (see above) suggests that the identification of agents having an influence on the DKK1/DKK3 ratio in cultures of normal human dermal fibroblasts of non-palmo-plantar origin would make it possible to control melanin production. from normal human non-palmoplantar melanocytes

[141] The objective of this study is to evaluate the effect of the compound called “Peregrina peptide hydrolyzate” on the synthesis and release of DKK-1 in a model composed of normal human fibroblasts in monolayer culture.

[142] Protocol: Human fibroblast cells are obtained from a 68-year-old donor. To carry out the experiments, the fibroblasts are cultured in monolayer until they reach confluence. Dexamethasone at 100 nm was used as a reference inducer of DKK-1 synthesis and release.

The skin discs were incubated for 48 hours in the absence (control) or presence of reference product or product to be tested: “Peregrina peptide hydrolyzate”: 0.01; 0.1 and 0.5% (v/v).

[143] At the end of the incubation, the incubation media were removed to perform the DKK-1 release measurement.

The test compound “peregrina peptide hydrolyzate” was diluted directly in the incubation medium to achieve the different concentrations described above.

At the end of the 48-hour incubation period, DKK-1 released into the incubation media was quantified using a sensitive and specific ELISA kit.

At the end of the incubation period, the proteins contained in the cell lysates were quantified using a spectro-colorimetric method (Bradford method),

Results are expressed as ng of DKK-1 per mg of protein (mean ± S.D.).

The level of significance between the “control” and the “reference product” was evaluated using a Student t test (*: p <0.05).

The level of significance between the “control” and the “test product” was assessed by a one-way analysis of variance (one-way ANOVA) followed by a Holm-Sidak test (*: p < 0.05).

[144] Under our experimental conditions, the reference product named “Dexamethasone”, tested at 100 nm, significantly increased the DKK-1 released by 181.8% (p <0.01) compared to “Control”. The results on modulation of DKK1 dosage are given below.

[145] [Table 12]

Concentration Cell growth versus control (%) DKK1 versus control (%) peregrina peptide hydrolyzate 0.5% +1.9 +131.50 0.1% That. 0 +32.30 0.05% -0.6 +26.10

Study on DKK3

The aim of this study is to evaluate the effect of compounds called “Peregrina Peptide Hydrolyzate” on the synthesis and release of DKK-3 in a model composed of normal human fibroblasts in monolayer culture.

Human fibroblast cells were obtained from a 68-year-old donor.

To perform the experiments, fibroblasts were cultured in monolayer until reaching confluence.

Human fibroblast cells were obtained from a 68-year-old donor.

To perform the experiments, fibroblasts were cultured in monolayer until reaching confluence.

At the end of the 48-hour incubation period, DKK-3 released into the incubation media was quantified using a sensitive and specific ELISA kit.

At the end of the incubation period, the proteins contained in the cell lysates were quantified using a spectro-colorimetric method (Bradford method).

Results are expressed as ng of DKK-3 per mg of protein (mean ± S.D.).

The level of significance between the “control” and the “reference product” was evaluated using a Student t test (*: p <0.05).

The level of significance between the “control” and the “test product” was assessed by a one-way analysis of variance (one-way ANOVA) followed by a Holm-Sidak test (*: p < 0.05). The results on modulation of DKK3 dosage are given below.

[146] [Table 13]

Concentration Cell growth versus control (%) DKK3 versus control (%) peregrina peptide hydrolyzate 0.5% 101.9 -20.7 0.1% approx. 100 -8 0.05% 99.4 -3.9

[147] Conclusion: The peregrina peptide hydrolyzate according to the invention can considerably increase the production of DKK1, and tends to reduce the production of DKK3 in its human cells. It has a strong capacity to reduce skin pigmentation thanks to the principle of palmar-plantar inhibition, through its ability to considerably increase DKK1 and significantly decrease DKK3, which increases the DKK1/DKK3 ratio. The increase in this ratio leads to limiting the pre-melanocyte/melanocyte differentiation pathway; thus the number of operational melanocytes decreases over time, reducing the skin's ability to tint.

[148] Example 11: Effect of the peregrina peptide hydrolyzate according to the invention to preserve DNA

[149] Telomere length dynamics are very important for the regulation of replicative lifespan in cells, particularly in long-lived species. Telomere shortening and telomerase activity are important factors in aging and tumorigenesis (Shay, J. W., 2005, Senescence an Immortalization: Role of Telomeres and Telomerase, Carcinogenesis 26(5): 867-874). Telomeres are complex nucleotide sequences that cover the ends of chromosomes from degradation, unwanted fusion-recombination, inappropriate activation of the DNA damage response. They also play an essential role in cell division and chromosome stability. There is growing evidence that telomere stability may be affected by occupational and environmental exposure, as some of these factors have been associated with increased inflammation, oxidative stress, DNA damage , chromosomal aberration and epigenetic alterations. Extremely short and long telomeres have been associated with neurodegenerative diseases, cardiovascular diseases (CVD), and cancer risk.

[150] Telomerase is a ribonucioprotein that catalyzes the addition of telomeric repeats to the ends of telomeres. Telomeres are long stretches of repeated sequences that cap the ends of chromosomes and are thought to stabilize the chromosome. In humans, telomeres are generally 7 to 10 kb long and include several repeats of the -TTAGGG- sequence.

[151] Telomerase is not expressed in most adult cells and telomere length decreases with successive replication cycles. After a certain number of replication cycles, progressive shortening of telomeres causes cells to enter a phase of telomeric crisis, which in turn leads to cellular senescence. Certain diseases are associated with rapid telomeric loss, leading to premature cellular senescence. It has been demonstrated that the expression of the gene encoding the human telomerase protein in human cells (Blasco M., 2007, Telomere Length, Stem Cells and Aging, Nature Chemical Biology 3, p. 640-649) confers a phenotype immortal, probably by bypassing the cells' natural senescence pathway. Additionally, telomerase gene expression in aging cells with short telomeres has been shown to produce an increase in telomere length and restore a phenotype typically associated with younger cells,

[152] The objective of this study is to evaluate the effect of the compound called “Peregrina peptide hydrolyzate” on telomere shortening in a model composed of normal human fibroblasts in monolayer culture. It is well known that the telomere corresponds to a biological clock. Telomere length gradually decreases with cell divisions, ultimately leading to the cell becoming unable to replicate. Measurement of telomere length was performed using quantitative PCR and in comparison with telomere length between cells at passages 2 and 5.

[153] Protocol: Human fibroblast cells were obtained from a 44-year-old donor. To perform the experiments, cells were used at passage 2 and 5. Fibroblasts were cultured for 3 consecutive passages in the absence (control) or presence of an increasing concentration to be tested of peregrina peptide hydrolyzate: 0.01; 0.1 and 0.5% (v/v).

[154] Preparation of test compound; The test compound “Peregrina Peptide Hydrolyzate” was diluted directly in the incubation medium in order to reach the different concentrations described above,

[155] At the end of the incubation, the cells were trypsinized. DNA was extracted from the cells using a dedicated DNA extraction kit. DNA was quantified by nanodrop,

[156] Telomere length was measured by quantitative PCR (q-PCR). For each sample, variation in telomere length was measured by relative quantification using the SCR (single copy reference) gene as the gene. reference.

For each sample, a q-PCR is performed using a telomere primer set that recognizes and amplifies telomere sequences and a second q-PCR is performed using the SCR primer set that recognizes and amplifies a telomere region. 100 bp on human chromosome 17 and serves as a reference for data normalization.

[157] Results are expressed in relative units corresponding to the length of telomeres relative to cells at passage 2 (mean ± S.D.). The level of significance between “control” at passages 2 and 5 was assessed using a Student t test (*p < 0.05). The level of significance between “control” and “test compound” was assessed independently for each product by a one-way analysis of variance (one-way ANOVA) followed by a Holm-Sîdak test (*: p < 0.05).

[158] Results: Under our experimental conditions, the peregrina peptide hydrolyzate tested at 0.05%, 0.1% and 0.5% (v! v), significantly reduced the shortening of telomeres in normal human fibroblasts.

[159] Telomere shortening: inhibition (compared to control) at 0.05% (v/v) + 8.9% (p <0.05) and at 0.1% (v/v) + 15.1% (p <0.01) and at 0.5% (v/v) + 16.6% (p <0.01).

[160] [Table 14]

Concentration Cell growth versus control (%) Telomere length versus control (%) Hydrosol 0.5% +1.9 +16.60 peptide 0.1% That. 0 +15.10 peregrina 0.05% -0.6 +8.90

Conclusion: In a context of normal multiplication or division of human cells, the peregrina peptide hydrolyzate according to the invention demonstrates an ability to considerably increase the length of telomeres. Telomeres are caps involved in the protection of DNA material; increasing telomere size preserves the integrity of the DNA material over time. Increased telomere length correlates with the ability to preserve DNA material. The peregrina peptide hydrolyzate, which can increase this length, is therefore involved in the preservation of human genetic material (DNA).

[161] Example 12: Comparative tests of different products on hydration on the surface of the skin

[162] The corneometry study allows the measurement of the moisturizing effect of a product on the surface of the skin. The device used is a corneometer equipped with a probe measuring the humidity of the skin by impedancemetry. A panel made up of 6 people (2 men and 4 women) tested under similar hydrometry and temperature conditions on the internal surfaces of their forearms. The inner side of the left forearm carries zones 1 and 2, the inner side of the right forearm carries zones 3 and 4.

[163] Zone 1 corresponds to the application zone of the reference moisturizing gel; zone 2 corresponds to the application zone of the reference gel + 2% of the peptide hydrolyzate according to the invention; zone 3 corresponds to the application zone of the reference gel + 2% of the peptide hydrolyzate with Moringa oleifera; zone 4 corresponds to the application zone of the reference gel + 2% of Purisoft® according to patent FR3076460 of BASF Beauty Care Solutions.

[164] The quantity of product applied to each zone is the same, and the measurement times are the same for each zone. An initial measurement is made before applying the product to each area so as to know the zero state of humidity of the skin. Then, for each area and each product, a first measurement is carried out 5 minutes after application of the product to the area, always by the same operator. Finally, a final measurement is carried out 30 minutes after the second measurement above, still by the same operator.

[165] The combination of differences observed before and after application does not follow a normal law studied in ANOVA (analysis of variance). The value obtained by difference of the measurement before and after is therefore the parameter studied. The results obtained constitute a qualitative and not quantitative trend of the values obtained;

[166] [Table 15]

Area Average estimated differences 1 = reference (placebo) 1,286 2 = ref. + peptide hydrolyzate according to invention 2% 1,190 3 = Ref. + peptide hydrolyzate of M. oleifera 2% -0.214 4 = ref. + Purisoft® BASF extract 2% -0.667

467] Conclusion: Zone 2 presents a positive hydration value for the skin between before and after application of the product containing the extract according to the invention, while zones 3 and 4 containing the extracts of Moringa oleifera according to 2 extraction modes, present negative values, that is to say that the skin is less hydrated after than before application of the products. The peptide hydrolyzate according to the invention makes it possible to maintain the benefit of hydration in a topical application product, which distinguishes the peregrina peptide hydrolyzate according to the invention from the other extracts tested. Figure 1 shows the results obtained.

[168] Example 13: Formulation of a makeup product

[169] [Table 16]

Ingredients % Water Q5P Caprylîc / Cap rie triglyceride 19.0000 Gum Acacia Senegal 7.0000 Charcoal 6.0000 Glycerin 5.0000 Propanedioi 5.0000 Bentonite 3.1500 Cetearyl glucoside 3x0000 Cetearyl alcohol 3.0000 Benzyl alcohol 1.0000 Peregrina peptide hydrolyzate according to invention 2.0000 Cellulose gum 0x0000 Xanthan gum 0.1750 Citric acid 0.1750

170] Example 14: Formulation of a cleansing product

[1711 [Table 17]

Ingredients % Water QSP Sodium coco sulfate 5.0000 Sodium cocoyl isethionate 4.0000 Bentonite 3.7800 Caprylic / Capric triglyceride 2.0000 peregrina peptide hydrolyzate according to the invention 1.0000 Gluconolactone 0.7500 Sodium benzoate 0.5450 Scent 0.5000

Xanthan gum 0.2700 Sodium stearcyl glutamate 0.2250 Citric acid 0.2250 Calcium gluconate 0.0050

[172] Example 15: formulation of a care product

[173] [Table 18]

Ingredient % Water Q5P Caprylic / Capric triglyceride 18.0000 Bentonite 4.2000 Cetearyl alcohol 1.5000 Peregrina peptide hydrolyzate according to invention 2.0000 Glu canota clone 0.7500 Sodium benzoate 0.5450 Xanthan gum 0.5000 Scent 0.5000 Sodium stearoyl glutamate 0.2500 Citric acid 0.2500 Calcium gluconate 0.0050

174] Example 16: relaxing or slimming tablet of 1 g: 2% dry extract according to the invention (containing 60% peptide hydrolyzate on an inulin support) + 47% calcium carbonate containing 200 IU of vitamin D + 25% magnesium gluconate + 23% inulin + 3% magnesium stearate).

[175] Example 17: Slimming powder in 750 mg capsule containing 200 mg of caffeine, 200 mg of dry extract according to the invention (containing 60% peptide hydrolyzate on an inulin support), 200 mg of chitosan and 150 mg of calcium carbonate.)

[176] Example 18: Appetite suppressant & slimming powder in 750 mg capsule containing 200 mg of caffeine, 200 mg of dry extract according to the invention (containing 60% peptide hydrolyzate on an inulin support), 200 mg of chitosan and 150 mg of Konjac (glucomannan).

[177] Example 19: Sublingual relaxing spray containing 2% extract according to example 1. 1% mother tincture of Rhodiola, 1% mother tincture of Melissa, 1% mother tincture of Valerian,

1% Hawthorn mother tincture, 2% horsetail extract, 0.15% salycilic acid, 7% sorbitol, and QSP in water).

[178] Example 20: Anti-wrinkle cream containing 2% hydrolyzed extract according to Example 1

[179] [Table 19]

Ingredients % INCI Water 70.95 Aqua Salicylic acid 0.15 Salicylic acid Sodium benzoate 0.5 Sodium benzoate Frametime CXG 3.2 Bentonite & Xanthan gum & Sodium stearoyl glutamate & Citric acid Xanthan gum FF 0.2 Xanthan gum Vegetable glycerin Palmera G995E 3.5 Glycerin Cetostearyl alcohol 50/50 1 Cetearyl alcohol DUB MCT 5545 18 Caprylic / Capric triglyceride Frangipani fragrance DIV/02356 0.5 Scent Hydrolyzed extract Peregrina 2 Moringa peregrina hydrolyzed seed extract

[180] Example 21: Toxicological tests of the hydrolyzate according to the invention

Preparation of the peptide hydrolyzate in accordance with Example 1: Unshelled seeds of Moringa peregrina (Forssk.) Fiori harvested when the fruit was ripe were dried to obtain an internal humidity level of less than 8% and preferably around 6 %, then pressed with a mechanical endless screw press, so as to separate the oil from the rest of the seed to obtain on the one hand the virgin oil and on the other hand a cake. The cake is then isolated in the form of sausages pre-cut into pieces of 1 to 2 cm. By following the protocol described in Example 1, we obtain the liquid extract which is used pure in the tests which follow.

[181 ] 1. Determination of mutagenic activity on the bacterial strain Salmonella typhimurium (TA 100) - Bacterial reverse mutation test

The test took place in 3 main phases:

- A preliminary experiment is carried out in order to evaluate the cytotoxicity of the element to be tested and to select the dose range for subsequent experiments,

- A first genotoxicity experiment (Test 1), with and without metabolic activation, with direct incorporation of the test system and the test (or controls) on minimal agar, over the dose range defined by the preliminary study,

-A second experiment (Test 2), with pre-incubation of the test system and test item (or controls), with and without metabolic activation, with dose levels defined by the study manager after analysis of the results of the first experiment. This second experiment was carried out in order to confirm or complete the results of the first, in particular when equivocal or negative results were obtained.

The dilutions of the extract according to Example 1 were prepared in water to carry out the cytotoxicity test.

[182] The latter was carried out on the Salmonella typhimurium TA100 strain at concentrations of 5000, 1600, 500, 160 and 50 pg/plate, with and without S9-Mix.

The reagents used for the preparation of S9-MÎX were prepared according to the following instructions:

[183] [Table 201

Final concentration MgCI2 (0.4 M) + KCI (1.65 M) 8mM + 33mM Glucose 6 Phosphate (0.2 M) 5mM NADP (0.1M) 4mM Phosphate buffer for S9-Mix (pH 7.4 - 0.2 M) 0.1M S 9 fraction 10% Water Adjust for final concentration

[184] Bacteria were exposed to the test extract with and without a metabolic activation system. The metabolic system used is a cofactor-enhanced post-mitochondrial fraction (S9). This S9 fraction, a microsomal fraction of Sprague Dawley rat liver homogenate treated with an enzyme inducer, is prepared according to Maron, D.M. and Ames, B.N. (1983) and was supplied by MOLTOX TM. It is stored at a temperature below -70°C. The S9 microsomal fraction was used at a concentration of 10% in S9-Mix. The protocol applied was as follows;

- In 3 hemolysis tubes, was introduced:

- dosage without metabolic activation:

• 0.1 ml of the different concentrations of the test elements.

• 0.5 ml of sterile 0.2 M phosphate buffer, pH 7.4, • 2 ml of top agar for S. typhimurium, • 0.1 ml of bacterial inoculum (TA100).

- dosage with metabolic activation:

• 0.1 ml of the different concentrations of the test elements, • 2 ml of top agar for S. typhimurium, • 0.1 ml of bacterial inoculum (TA100), • 0.5 ml of S9-Mix.

- Mix and pour onto the surface of the background agar previously distributed in Petri dishes.

- Incubate at 37°C ± 2°C for 48 to 72 hours.

These assays were carried out for each test: preliminary cytotoxicity test, test 1 and test 2. The untreated control, the negative controls and the positive controls carried out during the pre-incubation method were incubated for 20 to 30 minutes at 37°C±2°C before pouring the top agar.

The protocol applied was as follows:

- In 4 fractions of 2 ml of top agar for S. typhimurium, introduce:

o 0.1 ml of 0.2 M phosphate buffer, pH 7.4, o 0.1 ml of solvent, o 0.1 ml of S9-Mix. o 0.1 ml of the preparation of the element to be tested at the highest concentration,

- A fraction of 2 ml of top agar for S. typhimurium is used to check its sterility.

- Mix and pour onto the surface of the background agar previously distributed in Petri dishes.

- Incubate at 37°C + 2°C for 48 to 72 hours.

- The test is carried out in triplicate.

- No bacterial growth should be observed.

For at least 5 concentrations of the extract to be tested, a test without metabolic activation and a test with metabolic activation were carried out.

[185] Expression of results and interpretation

Many criteria make it possible to determine whether a result is positive, in particular an increase in the number of revertants correlated with the dose of item to be tested, or a reproducible increase in the number of revertants at one or more concentrations, with or without metabolic activation.

The element to be tested is considered mutagenic if, at the end of the verification steps, a dose-effect relationship has been obtained, in a reproducible manner, on one or some of the 5 strains with and/or without metabolic activation. Mutagenicity is only taken into account for a given concentration when the number of revertants is at least equal to twice the spontaneous reversion rate for strains TA98, TA100 and TA102 (R > 2) and three times the spontaneous reversion rate for strains TA1535 and TA1537 (R > 3).

- The test item is considered non-mutagenic if, after test 1 and test 2, the revertant rate has always remained less than twice the spontaneous reversion rate for all concentrations of the test item tested, for strains TA98, TA100 and TA102 (R <2) and less than three times the spontaneous reversion rate for strains TA1535 and TA1537 (R <3), with and without metabolic activation, and provided that it has been verified that the absence of the mutagenic effect is not linked to the toxicity of the concentrations tested.

The preliminary study showed no cytotoxicity of the test element; therefore, this concentration range was used for genotoxicity test 1.

[186] Depending on the result obtained for test 1, it was decided to use the same dilution range for test 2. The analysis of the revertants shows that:

- No cytotoxic effect was observed,

- No concentration of the extract to be tested showed an R ratio greater than or equal to at least double the spontaneous reversion rate for TA98, TA100 and TA102 and triple the spontaneous reversion rate for TA1535 and TA1537, with and without metabolic activation,

- No dose response was observed, regardless of test system or test conditions.

[187] In view of the results obtained during this study, the peptide hydrolyzate according to Example 1 can be considered as having no mutagenic or promotagenic activity.

[188] 2. In vitro phototoxicity test 3T3 N RU

The principle of the test is based on the comparison of the cytotoxicity of the peptide hydrolyzate according to Example 1 in the presence and absence of a non-cytotoxic dose of UVA, on cells in culture. Cytotoxicity is evaluated by determining cell viability using a vital dye: neutral red, 24 hours after treatment with the reference elements and the M. peregrina extract according to the invention with or without irradiation. at UVA. The cells used are Balb/c 3T3 lineage mouse embryo fibroblasts clone 31 (ATCC - CCL163). The positive control is a solution of chlorpromazine (CAS number: 6909-0). The negative control is a diluent of the tested and reference extract (buffered saline +/-1% solvent). The peptide hydrolyzate was tested at 8 concentrations on at least four culture wells per concentration studied, in the presence or absence of UVA. The fibroblasts were trypsinized and two 96-well culture plates were seeded with 100 μΙ of a cell suspension at 2.10 ⁶ cells/ml (i.e. 2.10 ⁶ cells per well) in complete culture medium.

The seeded plates were incubated in the oven for 24 hours at 37°C, 5% CO ₂ . At the end of the incubation, the semi-confluence of the cell layer was checked. The dilutions were prepared just before their deposition on the cells. The PH of the highest concentration was measured; it is between 6.5 and 7.8. The culture medium was removed, each well was carefully rinsed beforehand with 150 μl of PBS maintained at room temperature before being treated with 100 μl of each dilution of the extract or reference. The culture plates were incubated in the dark for 1 hour ± 5 minutes at 37°C and 5% CO ₂ . Irradiation was carried out using a BIO SUN solar irradiator (Vilber Lourmat RMX3W). The BIO SUN is a system that controls UV irradiation using a programmable microprocessor. The system continuously tracks the UV light emission. Irradiation stops automatically when the energy delivered is equal to the programmed energy. The spectral irradiation of the device under test was measured in the wavelength range 250 to 700 nanometers with a calibrated spectroradiometer.

One of the 2 plates was irradiated with its lid at room temperature, and the other plate was protected from UV rays and kept at room temperature during the irradiation. After irradiation, the treatment medium was aspirated and the cells were rinsed. Then 100 μl of complete culture medium were added gently and the plates were incubated for 18 to 22 hours at 37°C and 5% CO ₂ . The next day, cell viability (growth, morphology, integrity of the monolayer) was evaluated by observations on a phase contrast microscope. The culture medium was removed, each well was previously rinsed and maintained at room temperature before being treated with 100 μl of the staining solution. The plates were returned to the incubator for 3 h under the same conditions. The dye solution was removed and the cells were rinsed, then the rinsing solution was removed and 150 μl of desorption solution was added to each well. The plates were shaken until the crystals were completely solubilized. Absorbances were measured at 450 nm.

[189] Validation of the test;

The sensitivity of the cells to UVA is monitored approximately every 10 passages, by evaluating their viability after exposure to increasing doses of irradiation. The cells are cultured at the density used in the assay. They are irradiated the following day at doses of 2.5 and 9 J/cm ² and cell viability is determined one day later using the NRU test. The cells meet the quality criteria if their viability after irradiation at 5 J/cm ² UVA is greater than or equal to 80% of the viability of the controls kept in the dark; at the highest dose of 9 J/cm ² UVA, the viability must be at least equal to 50% of that of the controls kept in the dark.

[190] Results;

The negative control has an absorbance greater than or equal to 0.4. Chlorpromazine, a positive control, has an Clso of between 0.1 and 2 pg/ml in the presence of UVA and between 7 and 90 pg/ml in the absence of UVA. These results allow the test to be validated. The concentration of the peptide hydrolyzate of peregrina cake giving 50% cell death in the presence _or absence of UVA cannot be estimated. Mortality never reached 50%. The concentration of the peptide hydrolyzate of peregrina cake giving 50% cell viability in the presence or absence of UVA cannot be estimated. Viability is always above 50%.

Conclusion: Under the experimental conditions retained, the peptide hydrolyzate of peregrina cake can be considered non-phototoxic.

[191] 3. Evaluation of the ocular irritant potential by studying in vitro cytotoxicity using the neutral red detection method on SIRC cell line

This in vitro study is based on the evaluation of the cytotoxicity of the peptide hydrolyzate of peregrina cake by determining the concentration leading to 50% cell mortality (Clso) on a cell monolayer using the release technique. neutral red. The cells used are SIRC rabbit corneal fibroblasts (ATCC - CCL60) free of mycoplasmas.

The peptide hydrolyzate was diluted in 25% and 50% physiological saline. The fibroblasts were trypsinized and two 24-well culture plates were seeded with 1 ml of a cell suspension at 2.10 ⁵ cells/ml in complete culture medium. The seeded plates were incubated in the oven overnight at 37°C and 5% CO ₂ . At the end of the incubation, the confluence of the cell layer was checked. The staining solution was prepared at 0.5 mg/ml in complete culture medium. The culture medium was removed; 1 ml of the coloring solution was placed in each well. The plates were returned to the incubator at 37°C and 5% CO2 for 3 hours +/- 15 minutes. After this contact time, the dye solution was removed and replaced with 1 ml of complete culture medium per well. The plates were kept at room temperature for at least 30 minutes to stabilize the system before contact with the extract or reference. Each well was rinsed with 2 ml of PBS, maintained at room temperature, then 500 μl of each dilution of peptide hydrolyzate or reference were deposited in contact with the cell layer. The contact time was 60 seconds (30 seconds for the positive control). The treatment was carried out well by well with the stopwatch triggered at the time of deposition of the peptide hydrolyzate or the reference. The plate was shaken manually throughout the treatment. After 55 seconds (or 25 seconds for the positive control), the dilution was aspirated. At precisely 60 or 30 seconds, 5 successive rinses were carried out (5 x 2 ml of PBS maintained at room temperature). The supernatant was aspirated after each rinse and after the last rinse the wells remained devoid of medium while awaiting the revelation phase. After complete treatment of the culture plate, 1 ml of the desorption solution was placed in each well. The plate is shaken for approximately 15 minutes until a uniform color is obtained. The solutions obtained for each culture well were taken and distributed into 2 wells of a 96-well plate, i.e. 150 μΙ/well.

[192] Results:

The concentration of the peptide hydrolyzate giving 50% cell death was evaluated at >50%. The percentage of cell death at 50% peptide hydrolyzate was estimated at 17%.

Conclusion: Under the experimental conditions chosen, the cytotoxicity of the peptide hydrolyzate of peregrina cake can be classified as negligible cytotoxicity.

[193] 4. Evaluation of the skin compatibility of a peptide hydrolyzate after single application under an occlusive dressing for 48 hours under dermatological control.

The aim of this study is to evaluate the degree of skin compatibility of the peptide hydrolyzate by patch test, carried out on the antero-external side of the arm for 48 hours; and generally evaluate the capacity of the peptide hydrolyzate to maintain the skin in good condition. 10 healthy volunteers, female or male, aged 18 to 65 years, having neither dry nor sensitive skin and free from any dermatological lesion on the treatment area were to be included in the study. The skin compatibility of the peptide hydrolyzate, prepared in the form of a lotion with 5% of the peregrina extract according to Example 1 and 95% of a Propanediol/Sorbitol mixture, was evaluated 48 hours after the initial application between 30 40 minutes after dressing removal. Skin reactions (erythema and edema) were counted from 0 to 3 according to the following scales:

[194] [Table 21]

Quoting Erythema (E) Edema (Oe) 0 no erythema no edema 0.5 Barely noticeable erythema, very slightly pink coloring in part of the patching area barely noticeable palpable edema 1 Light erythema, pink discoloration over the entire patching surface palpable and visible edema 2 Moderate erythema, clear discoloration over the entire patching surface marked edema with or without papules or vesicles 3 Significant erythema, intense coloring over the entire patching area significant edema spreading outside the patching area with or without papules or vesicles

Any other skin reaction (bubbles, papules, vesicles, dryness, scaling, roughness, soap effect, etc.) was evaluated according to the following scale and reported descriptively:

- 0: no reaction,

- 0.5: very light: light

- 2: moderate

- 3: important.

At the end of the study, an average irritation index (LLM.) was calculated according to the following formula 10:

[195] [Math. 4]

LLM. = Sum of skin reactions (E + Oe + bubbles + papules + vesicles) / Number of volunteers analyzed

The I.LM. obtained made it possible to classify the extract tested according to the scale presented in table 15 below:

LLM. <0.20

Non-irritating

0.20 < I.I.M. <0.50

Slightly irritating

0.50 < I.LM. <2

Moderately irritating < LLM. <3

Very irritating

Results: The Average Irritation Index (LLM.) of the peptide hydrolyzate of peregrina cake is equal to: 0.

Conclusion: The peptide hydrolyzate of peregrina cake can be considered non-irritating after 48 consecutive hours of application in 12 volunteers.

[196] General conclusion of the tests:

The results of the tests carried out above are conclusive and demonstrate for the peptide hydrolyzate according to Example 1:

) Eye and skin irritation tests are negative

2) Phototoxicity tests are negative

3) Mutagenicity tests are negative.

The safety of the peptide hydrolyzate according to the invention is demonstrated and ideal for large-scale topical cosmetic use without restriction on target populations.

[197] Example 22: Skin depigmentation activity of the hydrolyzed oilcake extract

The aim of the test is to evaluate the depigmenting potential and the acceptability of a cosmetic product according to the invention after 28 days of application. The product tested is the anti-wrinkle cream described in Example 20.

Population analyzed:

Number of volunteers included and analyzed: 23

Female gender and average age of 64 years, between 44 and 70 years, with lentigo on the face, neck, décolleté and/or hands.

Instructions for use used for the study: twice a day, for 28 days, apply a small amount of product to clean skin on the face, neck, décolleté and hands then massage until the product is completely absorbed; Avoid contact with eyes ; apply the sunscreen product provided before exposure to the sun.

Evaluation criteria :

- Evaluation of the depigmenting potential: mexametric measurements at three sites (treated pigmented zone, treated non-pigmented zone, untreated non-pigmented control zone), on D1 and D28.

- Reporting of feelings of discomfort by volunteers.

- Photographs for illustrative purposes of a lentigo using the C-Cube videodermoscope (Pixience SAS, Toulouse, France) on D1 and D28 (sent in the form of contact sheets by WeTransfer when the final report is made available).

- Cosmetic acceptability: questionnaire completed by the volunteer on D28.

Study results:

[198] fTable 22]

Melanic index __________(UA)_______________________ Treated pigmented area Non-pigmented area treated Untreated non-pigmented area D1 J28 D1 J28 D1 J28 Average 200.9 183.8 143.5 133. 8 122.9 121.4 Standard deviation 42.8 35.3 27.2 227 33.4 32.7 Median 185.0 180.5 149.5 131. 0 116.0 117.5 Minimum 126.0 116.5 95.5 91.5 79.0 84.5 Maximum 289.5 251.5 197.0 173. 5 228.0 214.5 % change -8.5% -6.8% -1.2% p-value* 0.000 0.001 0.148 Significance** S S NS

Wilcoxon test for paired data;**S: Significant (p < 0.05) NS: No

Significant (p >0.05)

The analysis of the mexametric measurements made it possible to highlight a statistically significant reduction in the melanin index on the 2 treated areas (pigmented zone and non-pigmented zone) on D28 compared to D1. The variations recorded between D28 and D1 in the untreated area were found to be statistically insignificant.

Conclusion: Under the conditions of the study, the product tested showed significant depigmenting effectiveness after 28 days of use.

Claims (9)

[Claim 1] [Moringa peregrina seed cake extract mainly comprising a peptide hydrolyzate which comprises amino acid derivatives, amino acids, peptides and glycopeptides whose molecular weight is between 100 Da and 6,000 Da, preferably between 1,500 Da and 5,000 Da, preferably still between 3,000 and 4,500 Da, characterized in that it is obtained from unshelled and mature seeds of the fruit of Moringa peregrina by chemical proteolysis at pH greater than 13 for a period approximately 2 hours at a temperature between 16 and 25°C. [Claim 2] Extract according to claim 1, characterized in that it also comprises between 0.3% and 3% of volatile compounds, of which 50% of these compounds, i.e. between 0.15% and 1.5% of the extract, consists of nitrilated compounds. light, with mainly isobutyronitrile and methyl-butanenitrile; of which 5 to 10% of these compounds, or between 0.015 and 0.3% of the extract, is made up of isothiocyanate derivatives, mainly isopropyl isothiocyanate and isobutyl isothiocyanate; of which 1 to 5% of these compounds, or between 0.003 and 0.15%, is made up of essential oil, mainly Eucalyptol, Menthol and Benzaldehyde. [Claim 3] Process for obtaining a meal extract from Moringa peregrina seeds according to one of claims 1 or 2, characterized in that it comprises the following steps according to which: a) we collect the unshelled seeds harvested when ripe from the fruit of Moringa peregrina which we dry to obtain an internal humidity level of less than 8%, b) the dried seeds are pressed so as to separate the oil from the rest of the seed so as to obtain the cake comprising less than 6% by weight of residual oil, c) the cake obtained in step b) is crushed. , d) the ground cake obtained in step c is dispersed in the aqueous phase), e) chemical proteolysis is carried out on the aqueous dispersion obtained in step d) for a period of approximately 2 hours, at a pH greater than 13 and at a temperature between 16 and 25°C, f) the proteolysis is neutralized to stabilize the peptide hydrolyzate obtained, g) the peptide hydrolyzate is recovered by solid/liquid separation, h) the peptide hydrolyzate is purified by ultrafiltration, then optionally, i) the lyophilization is carried out. peptide hydrolyzate obtained in step h). [Claim 4] Method according to claim 3, characterized in that the solid/liquid separation of step g) is carried out by different processes such as centrifugation, spinning, filtration. [Claim 5] Method according to one of claims 3 or 4, characterized in that in step h) the ultrafiltration is carried out with a cut-off threshold of between 100 Da and 6000 Da and more particularly between 1500 Da and 5000 Da, and even more particularly between 3000 Da and 4500 Da. [Claim 6] Cosmetic or nutricosmetic composition, characterized in that it comprises, as active agent, an effective quantity of an extract of Moringa peregrina seed cake according to one of claims 1 or 2, and a physiologically acceptable excipient. [Claim 7] Composition according to claim 6, characterized in that it is a cosmetic composition formulated to be applied topically to the skin and in that the extract of Moringa peregrina seed cake is present in the composition at a concentration from 0.002 to 20% by weight relative to the total weight of the composition, preferably from 0.01 to 10%. [Claim 8] Composition according to claim 6, characterized in that it is a nutricosmetic composition formulated to be ingested and in that the extract of Moringa peregrina seed cake is present in the composition at a concentration of 0.01 to 100 % by weight relative to the total weight of the composition. [Claim 9] Cosmetic or nutricosmetic use of a composition according to one of claims 6 to 8, to improve the appearance of the skin, mucous membranes or appendages, prevent and/or combat dryness of the skin and mucous membranes, prevent and/or combat the skin signs of aging and/or photoaging, promote whitening of the skin and promote thinning.)| |