FR2991878A1 - Drink, useful for depigmenting skin of human, comprises extract of vegetables including Physcomitrella patens, maize and Oryza sativa - Google Patents

Abstract

Depigmenting drink comprises at least one vegetable extract including Physcomitrella patens, maize and Oryza sativa. ACTIVITY : Dermatological. MECHANISM OF ACTION : None given.

Description

The present invention relates to a depigmenting beverage for depigmenting the skin, especially human skin, comprising a miRNA or a plant extract. The pigmentation of the skin is an aesthetic problem, especially for women, but it is becoming more and more sensitive in a not insignificant fraction of men. Non-uniform pigmentation may occur, for example, in the form of age spots or pigment irregularities.

Three pigments are responsible for the skin color, by their surface distribution: carotene, hemoglobin and melanin. Among these three pigments, melanin is the most important pigment of the skin: its level and its exact composition (eumelanin or pheomelanin) in the skin determines its color. Melanin is produced from tyrosine with the help of various enzymes involved in melanogenesis, including tyrosinase.

In order to standardize the color of the skin, it is known to apply on the skin depigmenting cosmetic compositions, such as whitening creams. These creams include depigmenting actives.

Examples of such actives are hydroquinone, kojic acid, or azelaic acid.

However, there is still a need for new products to depigment the skin that are more effective and easier to administer.

In addition, in view of the ever-growing quest for non-toxic, non-immunogenic, biologically degradable products and the increasingly burdensome regulatory burdens on chemicals from the chemical industry, it is desirable that these products be of natural origin. It has already been shown that certain human miRNAs can decrease the production of melanin by the enzymes involved in melanogenesis (FR 1161950).

In addition, Zhang et al. demonstrated that expression of certain genes in mice could be regulated by plant miRNAs (Zhang et al., Exogenous plant MIR168a specifically targets mammalian LDLRAP1: evidence of cross-kingdom regulation by microRNA Cell Res 2012 Jan; 22 (1 ): 107-26). However, the Applicant has demonstrated that the level of melanin in human melanocytes was reduced when the cells were treated with plant-produced human miRNA mimetics and that oral ingestion of specific plant miRNAs alters the level of melanin in the cells. human melanocytes in the skin. Therefore, the subject of the present invention is a depigmenting beverage, in particular human skin, comprising at least one of the plant miRNAs chosen from: ppt-miR1039-5p, zma-miR528a *, osa-miR4128e-5p and hsamir330.

By "microRNAs (miRNAs)" is meant non-coding RNAs of about 21 nucleotides in length. They have been shown to play a crucial role in the post-transcriptional regulation of gene expression, for example by affecting the rate of translation or destabilizing the target mRNA (Friedman RC et al., Most mammalian mRNAs are conserved targets of microRNAs, Genome Res 2009, 19: 92-105).

The miRNAs, currently known, are listed in the base www.mirbase.org. Ppt-miR1039-5p is a miRNA found in the whole plant extract, including rhizoids, leaves, stem, and meristems, of Physcomitrella patens, a common moss species in the northern hemisphere. The miRNA ppt-miR1039-5p, has accession number MIMAT0005148 in mirbase and for sequence 5'-UCU UCG UCU UCU UCU CUG UG-3 '(SEQ ID NO: 1). Zma-miR528a * is a miRNA found in the whole plant extract, including the root, leaves, stem, meristems, grains, flowers and fruits of Maize, widely known as maize. The miRNA zma-miR528a * has accession number MIMAT0015362 in mirbase and for sequence 5'-CCU GUG CCU GCC UUC UCU UU-3 '(SEQ ID NO: 2). MiRNA osa-miR1428e-5p is a miRNA found in the whole plant extract, including the root, leaves, stem, meristems, seeds, flowers and fruits of Oryza sativa, a species of rice originally growing in Asia. The miRNA osa-miR4128e-5p has accession number MIMAT0007784 in mirbase and for sequence 5'-AAU UCA CAG GCC UCU UCU UGU G-3 '(SEQ ID NO: 3).

MiRNA hsa-mir330 is a miRNA found in human RNA. The miRNA hsa-mir330 has accession number MI0000803 in mirbase and for sequence 5'- CUU UGG CGA UCA CUG CCU CUC UGG GCC UGU GUC UUA GGC UCU GCA AGA UCA ACC GAG CAA AGC ACA CGG CCU GCA GAG AGG CAG CGC UCU GCC C-3 '(SEQ ID NO: 4). The beverage according to the invention may advantageously comprise said miRNA in a proportion ranging from 0.0001% to 10% and preferably from 0.001% to 1% by weight, relative to the total weight of the beverage. The present invention also relates to a depigmenting beverage comprising at least one of the plant extracts selected from: Physcomitrella patens, Maize and Oryza sativa. The drink according to the invention may advantageously comprise the three plant extracts: Physcomitrella patens, Maize and Oryza sativa. The beverage according to the invention may advantageously comprise said plant extract in a proportion ranging from 0.0001% to 10% and preferably from 0.001% to 1% by weight, relative to the total weight of the beverage. The plant extract that can be used in the present invention can be derived from whole plants, in particular the root, the leaves, the stem, the meristems, the seeds, the flowers and the fruits. This extract can be obtained using any solvent and preferably using an aqueous solvent, alcoholic or oily. By "aqueous solvent" is meant a solvent containing predominantly water. "Alcoholic solvent" means a solvent selected from monoalcohols such as methanol, ethanol or isopropanol, and glycols such as propylene glycol or dipropylene glycol, optionally in the presence of water "Oily solvent" means a solvent chosen from edible oils.

In addition to the miRNA previously described, the beverage according to the present invention may also comprise at least one additive that is customary in the agri-food field. In particular, according to a preferred embodiment, the beverage according to the invention may comprise, in a nonlimiting manner, one or more of the following additives: one or more sweeteners such as glucose, and / or alternative sweeteners (in English, "alternative sweeteners ") such as aspartame, sucralose, sorbitol, other polyols, or acesulfame potassium. Said alternative sweeteners and / or sweeteners may be present in the beverage at a content of the order of 0.001 to 30% by weight, relative to the total weight of the beverage; one or more texturizers and / or gelling agents such as xanthan gum, carrageenan, pectin, or cellulosic compounds. Said texturizers and / or gelling agents may be present in the beverage at a content of the order of 0.001 to 30% by weight, relative to the total weight of the beverage; one or more nutraceuticals such as minerals (Ca2 +, Mg2 +), vitamins (A, B, C, D, K), probiotics, fruit juices and nectars, coconut water, or antioxidants such as pomegranate extract.

Said nutraceuticals can be present in the beverage in a content of the order of 0.001 to 30% by weight, relative to the total weight of the beverage; one or more emulsifiers. Said emulsifiers may be present in the beverage at a content of the order of 0.001 to 30% by weight, relative to the total weight of the beverage; one or more preservatives such as branched phenols or biocides (sorbates, propionates, or benzoates). Said preservatives may be present in the beverage at a content of the order of 0.001 to 30% by weight, relative to the total weight of the beverage; one or more acids such as citric acid. Said acids may be present in the beverage in a content of the order of 0.001 to 30% by weight, relative to the total weight of the beverage; one or more dyes and / or food flavorings. Said dyes and / or food flavors may be present in the beverage at a content of the order of 0.001 to 30% by weight, relative to the total weight of the beverage; - one or more perfumes and / or flavor enhancers. Said perfumes and / or flavor enhancers may be present in the beverage at a content of the order of 0.001 to 30% by weight, relative to the total weight of the beverage; one or more amino acids and / or proteins such as collagen or elastin. Said amino acids and / or proteins may be present in the beverage at a content of the order of 0.001 to 30% by weight, relative to the total weight of the beverage; caffeine, which may be present in the beverage in a content of the order of 0.001 to 30% by weight, relative to the total weight of the beverage; - and their mixtures, without this list being exhaustive. Examples of such additives are cited in particular in Food and Beverage Additives - US Industry Study with Forecasts for 2014 & 2019 (Freedonia group) which describes a wide variety of food additives commonly used in the food industry, which are suitable for used as additional ingredients in the depigmenting beverage according to the present invention. The present invention also relates to the use of the depigmenting beverage according to the invention, for depigmenting the skin, in particular human skin.

The beverage of the present invention may for example be taken once or twice daily, before and / or after the meal.

This drink can be provided to consumers in various forms such as can, bottle or cardboard brick. EXAMPLES Example 1: Treatment with ppt-miR1039-5p Materials and Methods Cells Human MNT-1 melanoma pigmented cells (Reish et al., 1995) were cultured in RPMI1640 medium (# 21875, Gibco), with 10% fetal calf serum (# 10270, Gibco), 1% penicillin-streptomycin (# 15140, Gibco) and 1% L-glutamine (# 25030, Gibco), at 37 ° C and 5% CO2 in a humid atmosphere. 4x105 miRNA mimetics of MNT-1 cells were inoculated per well in a 6-well plate (# 353046, Falcon) at day -1. At day 0, the cells were transfected with 50 and 100 nM miRNA mimetic (ppt-miR1039-5p, Qiagen MSY0005148), using 5uL of Lipofectamine 2000 (# 1668-019, Invitrogen).

On day 1, the medium was changed. At day 3, the cells were trypsinized and seeded at 4x105 cells per well.

At day 4, cells were re-transfected with 50 nM miRNA mimetic (see day 0). On day 5, the medium was changed. At day 7, the cells were trypsinized and seeded at 4x105 cells per well. At day 8, a six-well plate was used for RNA lysis and the other for protein lysis.

Western blot The proteins were extracted from a six-well plate for 30 minutes at 4 ° C using 75 μL of radioimmunoprecipitation buffer (RIPA) (1% NP40, 0.5% sodium deoxycholate, 0.1% SDS in PBS) with protease inhibitors (Roche). Protein lysates were centrifuged for 30 min at 14,000 rpm at 4 ° C. The supernatant was transferred to a 1.5 ml Eppendorf tube pre-cooled and placed on ice. Protein concentration was determined with BCA Protein Assay (ThermoFisher). Proteins (50 μg) were separated on a 10% SDS-polyacrylamide gel and transferred to ProTron nitrocellulose membranes (Whatman) at 100V for 1h at room temperature. The membranes were saturated with skim milk powder in buffered saline Tris (SCT) solution. The membranes were probed with the diluted primary antibody (see below) in 0.5% (v / v) Tween / SCT (TTBS) overnight, and then subjected to three washes of five minutes each in TTBS. Secondary antibodies were diluted in 1: 10,000 TTBS and applied for 1h at room temperature. The membranes are then washed three times in TTBS for 10 min each and developed with SuperSignal WestPico Chemiluminescent Solution (Thermoscientific) using Amersham Hyperfilm (GE Healthcare). The primary antibodies used are the following: - Anti-Tyr goat polyclonal (C-19), SC-7833 (Santa Cruz), diluted to 1 / 1,000 - Anti-13-actin mouse monoclonal (CA-15), A5441 (Sigma), 1 / 7,500 The secondary antibodies used are: - HRP anti-mouse IgG, W402B (Promega), 1 / 10,000 - Goat Anti-IgG HRP, 705-035-147 (Jackson ImmunoResearch), 1 / 10,000.

Results It should be noted that the addition of 50nM of ppt-miR1039-5p reduced the amount of tyrosinase compared to the control test, and that a greater decrease in the amount of tyrosinase is observed in case of addition. 100nM of ppt-miR1039-5p (Figure 1). These results show that pptmiR1039-5p decreases the amount of tyrosinase in MNT1 cells.

Example 2: A depigmenting beverage composition A depigmenting beverage with the following composition a) or b) was prepared. miRNA (ppt-miR1039-5p) Composition a) Coconut water 25 Reconstituted whey 25% Pomegranate and red fruit juice 5% Stabilizer: pectin 0.1% Acidifier: citric acid 0.1% Water QSP 100m1 Composition b) Caffeine 80 mg Taurine 1000 mg D-glucuronalactone 600 mg Vitamin B2 1.5 mg Vitamin B3 20.5 mg Vitamin B5 5 mg Vitamin B6 5 mg Vitamin B12 5 pg Water QSP 100m1

Claims (8)

REVENDICATIONS1. A depigmenting beverage comprising at least one of the plant extracts selected from: Physcomitrella patens, Maize and Oryza sativa. 2. Beverage according to claim 1, characterized in that it comprises the three plant extracts: Physcomitrella patens, Maize and Oryza sativa. 3. Drink according to claim 1 or 2, characterized in that it comprises said plant extract in a proportion ranging from 0.0001% to 10% by weight, relative to the total weight of the beverage. 4. Beverage according to any one of the preceding claims, characterized in that said plant extract is an aqueous extract, alcoholic or oily. A depigmenting beverage comprising at least one of the miRNAs selected from: ppt-miR1039-5p, zma-miR528a *, osa-miR4128e-5p, and hsa-mir330. 25 6. Drink according to claim 5, characterized in that it comprises said miRNA in a proportion ranging from 0.0001% to 10% by weight, relative to the total weight of the beverage. 30 7. Beverage according to any one of the preceding claims, characterized in that it further comprises at least one additive chosen from: several sweeteners, alternative sweeteners, texturizers, gelling agents, nutraceuticals, emulsifiers, preservatives, acids, colorants, food flavorings, fragrances, flavor enhancers, amino acids, proteins, caffeine and their mixtures. 8. Use of a beverage according to any one of the preceding claims for depigmenting the skin.