FR2829930A1 - Cosmetic composition comprising essential oils rich in carvacrol, eugenol and thymol for use in protecting the skin against oxidative stress and the effects of free radicals - Google Patents

Abstract

A cosmetic composition comprising a mixture of essential oils rich in carvacrol, eugenol and thymol. The composition contains 9-15 wt.% and preferably 10-12 wt.% of carvacrol, 3-10 wt.% and preferably 5-9 wt.% of thymol and 10-18 wt.% and preferably 12-15 wt.% of eugenol. This content is given by the inclusion of 10-30 and preferably 20 wt.% each of oregano and clove essential oils and 5-25 wt.% and preferably 15 wt.% or thyme essential oil. It additionally contains 15-35 wt.% of camphorated rosemary essential oil, 10-20 wt.% of marjoram essential oil and 0.5-3 wt.% of Curcuma zedoaria (Roscoe) essential oil. An Independent claim is included for a cosmetic vehicle containing 0.2-0.5 wt.% of the claimed composition.

Description

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TOPICAL COSMETIC COMPOSITION BASED ON ESSENTIAL OILS RICH IN CARVACROL, EUGENOL AND THYMOL
The invention relates to a new topical cosmetic composition. More specifically, the subject of the invention is a composition which is characterized in that it comprises a mixture of essential oils rich in carvacrol, eugenol and thymol.

 The Applicant has indeed found that the combination of these three molecules has interesting properties, including anti-oxidants, antiradicalaires, thus making a product able to protect the cutaneous cell and reduce UV-induced cytotoxicity Thanks to its action protective association with the deleterious effects of oxidative stress, such an association thus finds a certain number of topical cosmetic applications, in particular in the field of skin care (protection against exogenous aging, vis-à-vis the environmental stress, anti-aging treatment, treatment of mature skin), sun protection and hair protection.

 According to a first characteristic of the invention, the composition contains: between 9 and 15%, advantageously between 10 and 12% by weight of carvacrol, between 3 and 10%, advantageously between 5 and 9% by weight of thymol, between 10 and 18%, advantageously between 12 and 15% by weight of eugenol.

 Carvacrol, eugenol and thymol are well-known molecules in the phenol family, usually extracted from the relevant aromatic plants. In some cases they can be obtained by synthesis, but the duplicates of synthetic molecules, even if they are known as "identical nature", do not have the same properties.

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Drome), l'huile essentielle de clou de girofle, en particulier Eugenia caryophyllus (spreng) et l'huile essentielle de thym, en particulier Thymus vulgaris (L) thymoliferum. Among the essential oils containing carvacrol, eugenol and thymol, the Applicant has respectively selected the essential oil of oregano, in particular green oregano (Origanum heracleoticum (L) carvacroliferum, thus green

Drome), clove essential oil, especially Eugenia caryophyllus (spreng) and thyme essential oil, especially Thymus vulgaris (L) thymoliferum.

 Each of these essential oils is obtained by conventional methods known to those skilled in the art. Thus, the essential oil of green oregano is obtained by distillation of flowers and leaves; the essential oil of clove is obtained by distillation of the flower buds and the thyme oil is obtained by distillation of the flowering tops.

 In practice, the composition of the invention contains between 10 and 30%, advantageously 20% by weight respectively of essential oil of green oregano and clove, and between 5 and 25%, advantageously 15% by weight of essential oil of thyme.

 To reinforce the anti-radical activity but also to improve the micro-circulation of the blood, the oxygenation of the skin tissue and its detoxification, the regeneration of the epidermis and to provide a decongestant effect, the composition of the invention also contains camphor rosemary extract, including Rosmarinus officinalis (L) camphoriferum. In practice, this rosemary extract is in the form of an essential oil obtained by the distillation of flower heads with water vapor. The monoterpenone molecules, especially camphor, but also verbenone and carvone present an activity and a regenerating power and healing mucosal and skin tissue.

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 In an advantageous embodiment, the composition contains between 15 and 35% by weight of a rosemary essential oil camphor, in practice between 26 and 30%.

 To further improve the protective and detoxifying properties, the composition further contains a marjoram-shell extract (Origanum majorant), in the form of an essential oil in the composition, in a proportion of 10 to 20% by weight, advantageously 15 to 20% by weight. % in weight. The essential oil is obtained by distillation of the flowering plant with water vapor. The richness of this essential oil in monoterpene molecules (myrcenes, a and terpinene y) and monoterpenols (terpinen-1-ol-4, trans-thujanol-4) is remarkable. The latter come to combine their lymphotonic and decongesting activities.

 In an advantageous embodiment, the composition additionally contains a zedary extract (Curcuma zedoaria, Roscoe) in the form of an essential oil obtained by distillation of the rhizomes. In practice, the zedary extract enters the composition in a proportion of 0.5 to 3% by weight, of the order of 2% by weight of the composition.

 The composition of the invention can be incorporated into any type of cosmetically acceptable vehicle: cream, milk, gel, serum, microemulsion ... at a rate of 0.2 to 0.5% by weight.

 As already stated, the composition as described above has a number of properties.

 In a particular embodiment, by virtue of its antioxidant, antiradical, anti-inflammatory, stimulating effect of cutaneous microcirculation, the composition will allow the skin to protect itself against any environmental stress inducing the generation of free radicals, especially the

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 UV radiation and the action of many environmental chemical pollutants.

By its scavenger action, the composition protects the skin from the deleterious effects caused by oxidative stress (release of reactive oxygen species, mediators of inflammation and mediators of apoptosis, alteration of biological molecules, DNA, proteins, lipids , modification of the gene expression causing accelerated aging, see phenomena of carcino genesis).

 The invention and the advantages thereof will become more apparent from the following exemplary embodiments.

EXAMPLE 1
Manufacture of the composition of the invention

<Tb>
<tb> Oil <SEP> Essential <SEP>% <SEP> in <SEP> weight
<tb> Oregano <SEP> green <SEP> 20
<tb> Clove <SEP> nail <SEP> 20
<tb> Rosemary <SEP> Camphor <SEP> 29
<tb> Thyme <SEP> thymol <SEP> 15
<tb> Marjoram <SEP> to <SEP> shell <SEP> 15
<tb> Zedoaire <SEP> 1
<Tb>
Procedure: - Weight mixing, filtration and introduction of each ingredient in the order of the formulation.

- Agitation.

- Homogenization.

- Filtration.

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EXAMPLE 2
In Vitro Evaluation of the Antioxidant Potential of the Composition of Example 1 by the DPPH Test a) Principle
The DPPH '(1,1-diphenyl-2-picrylhydrazyl) radical is a stable free radical which strongly absorbs at 517 nm. It is not very specific and can be used to evaluate, in vitro, the antioxidant potential of various chemical structures.

ladite composition. Le taux de réduction du radical DPPH est évalué par le changement d'absorbance d'une solution de DPPH'à 517 nm après 3 heures d'incubation. The antioxidant potential of the composition of the invention was appreciated by measuring the rate of reduction of the DPPH radical in the presence and in the absence of

said composition. The rate of reduction of the DPPH radical is evaluated by the change in absorbance of a solution of DPPH at 517 nm after 3 hours of incubation.

Α-Tocopherol (vitamin E) is chosen as the reference antioxidant. b) Equipment
The optical densities are measured on a device of the Shimadzu spectrophotometer type (UV-2401-PC). c) Results
Each test is done in triplicate.

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The results of the averages of these tests are shown in the table below.

<Tb>
<Tb>

Power
<tb> OD <SEP> to <SEP> 517 <SEP> nm <SEP> Antioxidant <SEP> (%)
<tb> Witness <SEP> 1. <SEP> 24 <SEP> 0
<tb> a-tocopherol <SEP> 50 <SEP> j. <SEP> LM <SEP> (21. <SEP> 5 <SEP> g / ml) <SEP> 0.16 <SEP> 87
<tb>&alpha; -tocopherol <SEP> 10 <SEP> M <SEP> (4.3 <SEG> g / ml) <SEP> 0.92 <SEP> 26
<tb> Example <SEP> 1 <SEP> to <SEP> 10 <SEP> mg / ml <SEP> 0. <SEP> 04 <SEP> 97
<tb> Example <SEP> 1 <SEP> to <SEP> 5 <SEP> mg / ml <SEP> 0.06 <SEP> 96
<tb> Example <SEP> 1 <SEP> to <SEP> 1 <SEP> mg / ml <SEP> 0. <SEP> 08 <SEP> 93
<tb> Example <SEP> 1 <SEP> to <SEP> 500 <SEP><SEP> 0. <SEP> 10 <SEP> 92
<tb> Example <SEP> 1 <SEP> to <SEP> 100 <SEP> g / ml <SEP> 0.21 <SEP> 83
<tb> Example <SEP> 1 <SEP> to <SEP> 50 <SEP> p. <SEP> g / ml) <SEP> 0.35 <SEP> 72
<tb> Example <SEP> 1 <SEP> to <SEP> 25 <SEP><SEP> 0. <SEP> 60 <SEP> 51
<tb> Example <SEP> 1 <SEP> to <SEP> 10 <SEP> jg / ml09424
<tb> Example <SEP> 1 <SEP> to <SEP> 5 <SEP> g / ml <SEP> 1.06 <SEP> 13
<tb> Example <SEP> 1 <SEP> to <SEP> 1 <SEP> g / ml <SEP> 1. <SEP> 19 <SEP> 3
<Tb>

At the concentrations tested, the composition of the invention has a dose-dependent reducing power with respect to the DPPH radical. * More precisely, the higher the concentration of the composition of the invention increases and the greater the antioxidant power is important.

 As shown in this table, for a concentration of 100 g / ml of composition, the power scavenger antioxidant is similar to that of vitamin E tested at 50 M (83% against 87%).

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de l'invention vis-à-vis des effets délétères induits par les radiations WB. Aussi, afin d'évaluer l'effet protecteur de cet actif, une approche expérimentale basée sur la mesure de la cytotoxicité et de la péroxydation lipidique induites par les UV B sur des kératinocytes humains cultivés en absence et en présence de l'extrait est réalisée. EXAMPLE 3 1-Demonstration of the Protective Effect of the Composition of the Invention with Respect to the Deleterious Effects of UV Radiation,
The purpose of this study is to evaluate the protective activity of the composition

of the invention with respect to the deleterious effects induced by WB radiation. Also, in order to evaluate the protective effect of this active agent, an experimental approach based on the measurement of UV B-induced cytotoxicity and lipid peroxidation on human keratinocytes cultured in the absence and in the presence of the extract is carried out. .

milieu KGM sérum free à 37 C en atmosphère humide air-C02 (95%-5 %). 2 - Used cells
Normal human keratinocytes isolated from prepuce skin obtained during surgical procedures in young children are cultured in

medium KGM serum free at 37 C in humid atmosphere air-C02 (95% -5%).

3-Evaluation of the Activity of the Composition on UVB Induced Toxicity
3.1. Principle
The keratinocytes are seeded at 20 * 103 cells / well in 96-well microplates. At 80-90% confluency of the cell monolayers, the culture medium is removed and replaced by new medium containing the product under study (Preventive treatment). The cells are placed in an oven at 37 ° C. and incubated for 48 hours in an air-CO2 atmosphere (95% -5%).

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After 48 hours of treatment, the plates are emptied and the cells are exposed to UV B (50 and 150 mJ / cm) through a solution of PBS (200 11) in the absence of the product under study. UVB is generated by Philips TL 20 W / 12 tubes. Light intensities (in mW / cm) are measured and monitored using a UVX (UVX Digital Radiometer, ULTRA-VIOLET PRODUCTS, INC) radiometer.

After irradiation of the cells, the PBS solution is removed and replaced with new culture medium not containing the product under study. The cells are returned to the oven at 37 ° C. and incubated for 24 hours in an airCO (95% -5%) atmosphere. Cell viability is assessed at the end of the test by a Neutral Red test.

A = [ (D. O -D. O n. uv)/ (D. O ,)] X 100 B = [ (D. O -D. O. . uv)/D. O )] x 100

Le pourcentage d'activité protectrice (A. P.) est calculé selon la formule : A. P [ (A-B)/A] xlOO 3. 2. Résultats La composition de l'invention a été testée à trois concentrations retenues après une étude préalable de cytotoxicité : CI = 5 ug/ml C2 = 10 g/ml C3= 25 ug/ml The percentage of cytotoxicity of UV B is calculated, at the level of the control cultures and treated cultures, according to the formulas:

A = [(D.O.D.O.sub.N u.sub.nu) / (D.O.)] X 100 B = [(D.O-OD.sub.v) / D. O)] x 100

The percentage of protective activity (AP) is calculated according to the formula: A. P [(AB) / A] × 100 3. 2. Results The composition of the invention was tested at three concentrations selected after a preliminary cytotoxicity study. : CI = 5 μg / ml C2 = 10 g / ml C3 = 25 μg / ml

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Vitamin E (dl-a-tocopherol) was tested in parallel as a reference antiradical.

The percentage of cytotoxicity induced by UV B was calculated in each batch.

<Tb>
<Tb>

50 <SEP> mJ / cm2 <SEP> UVB <SEP> 150 <SEP> mJ / cm2 <SEP> UV-B
<tb> Composition <SEP> of <SEP> the invention <SEP> Vit. <SEP> E <SEP> Composition <SEP> of <SEP> the invention <SEP> Vit. <SEP> E
<tb> Conc. <SEP> 0 <SEP> 5 <SEP> 10 <SEP> 25 <SEP> 215 <SEP> 0 <SEP> 5 <SEP> 10 <SEP> 25 <SEP> 215
<tb> (g / ml)
<tb> Cytotoxicity <SEP> 20.9% <SEP> 8.9% <SEP> 4.0% <SEP> 0.7% <SEP> 0.0% <SEP> 25.0% <SEP> 22.8% <SEP> 13.8% <SEP> 0. <SEP> 0% <SEP> 8.6%
<tb> Activity <SEP> - <SEP> + 57% <SEP> + 81% <SEP> + 97% <SEP> + <SEP> 1 <SEP> 00% <SEP> - <SEP> + 9% <SEP> + 45% <SEP> + <SEP> 100% <SEP> + 66%
<tb> Protective
<Tb>

Exposure of the cells to UV B induces a cytotoxic effect: cell viability is reduced by 21% and 25% respectively when the cells are exposed to 50 and 150 mJ / cm 2.

 Whatever the dose applied, there is a very clear decrease in UV-induced cytotoxicity B in the cultures treated with the composition of the invention. It should be noted that the protective effect is dosedpendeant and that this active is capable, under the experimental conditions selected, of providing complete protection of the cells.

 Under the same experimental conditions, vitamin E (215 μg / ml) is able to completely or partially oppose (66%) UV B-induced cell damage when exposed to 50 and 150mJ / cm2 cells. .

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4 - Evaluation of the activity of the composition on lipid peroxidation induced by UVB
4.1. Principle
Human keratinocytes are inoculated at 500 * 103 cells / well in 6-well plates. At 80-90% confluency of the cell monolayers, the culture medium is removed and replaced by new medium containing the product under study (Preventive treatment). The cells are placed in an oven at 37 ° C. and incubated for 48 hours in an airCO (95% -5%) atmosphere.

du produit à l'étude. Les W B sont générés par des tubes Philips TL 20 W/12. Les intensités lumineuses (en mW/cm) sont mesurées et contrôlées à l'aide d'un radiomètre UVX (UVX Digital Radiometer, ULTRA-VIOLET PRODUCTS, INC). After 48 hours of treatment, the plates are emptied and the cells are exposed to WB (50 mJ / cnr) through a solution of PBS (2 ml) in the absence

of the product under study. WBs are generated by Philips TL 20 W / 12 tubes. Light intensities (in mW / cm) are measured and monitored using a UVX (UVX Digital Radiometer, ULTRA-VIOLET PRODUCTS, INC) radiometer.

 After irradiation of the cells, the PBS solution is removed and replaced with new culture medium not containing the product under study. The cells are placed in an oven at 37 ° C. and incubated for 24 hours in an air-CO (95% -5%) atmosphere. At the end of the test, the cells are detached from their support. After centrifugation, the cell pellets are taken up in distilled water and are lysed by sonication. The various cell extracts are removed and processed for the MDA assay according to the instructions provided by the Calbiochem laboratories (Lipid peroxidation assay kit, LPO test). An aliquot of each extract is taken and processed for the assay of cellular proteins (BioRad Protein assay).

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The protective activity (A. P) is calculated according to the formula: A. P = [(MDA c A. P = [(MDA ,, rol + uv-MDA t ,, uv) / MDA, ,, uv] x 100 4. 2. Results The composition of the invention was tested at three concentrations: C, -5 g / ml ClOug / ml C, = 25 ug / ml Vitamin E was tested in parallel.
The following table groups all the results: MDA level (nmol / mg prot) and protective activity (A. P).

<Tb>
<Tb>

Witness <SEP> Invention <SEP> Vit. <SEP> E
<tb> without <SEP> + UV <SEP> 5 <SEP> g / ml <SEP> 10 <SEP> g / ml + <SEP> 25 <SEP> g / ml + <SEP> 430 <SEP> g / ml
<tb> UV <SEP> + UV <SEP> UV <SEP> UV <SEP> + UV
<tb> MDA
<tb> 0.382 <SEP> 1.070 <SEP> 1.098 <SEP> 0.856 <SEW> 0. <SEP> 521 <SEP> 0.752
<tb> (nmol / mg <SEP> prot <SEP>)
<tb> Activity
<tb> 0 <SEP>% <SEP> 20% <SEP> 51 <SEP>% <SEP> 30%
<tb> Protective
<Tb>

Exposure of keratinocytes to W B results in a marked increase in the level of cellular MDA. Indeed, the basal rate is multiplied by approximately 3 after irradiation of the cells at a dose of 50 mJ / cm 2.

 The levels of MDA measured at the level of the cultures treated with the composition of the invention at 10 and 25 μg / ml are lower than that recorded in FIG.

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 level of irradiated control cultures. It should be noted that this protective effect against lipid peroxidation is dose-dependent. At a dose of 25 p. g / ml, the composition of the invention is capable of reducing by more than 50% the degree of UVB-induced peroxidation.

vis-à-vis des effets délétères induits par les W B. La viabilité cellulaire (test au Rouge Neutre) a été mesurée après exposition des cellules à deux doses d'UVB (50 mJ/cm2 et 150 mJ/cn2). Le degré de péroxydation lipidique induite par les UV B a été apprécié par la mesure du taux de MDA cellulaire après irradiation UVX (50 mI/cm2) des cellules. Under the same experimental conditions, vitamin E at a much higher concentration (430 μg / ml) reduces the UVX-induced MDA level by only 30%.
5-Conclusion
The antioxidant potential of the composition of the invention was evaluated on human keratinocytes in culture by evaluating its protective effect

The cell viability (Neutral Red test) was measured after exposure of the cells to two doses of UVB (50 mJ / cm 2 and 150 mJ / cm 2). The degree of lipid peroxidation induced by UV B was appreciated by measuring the level of cellular MDA after UVX irradiation (50 mI / cm 2) of the cells.

 Under the experimental conditions selected, this study has shown that the treatment of the cells with the composition of the invention makes it possible to significantly reduce the cytotoxicity induced by WB. This protective effect could be explained, in part, by an anti-inflammatory activity. oxidizing at the cell membranes. In fact, the level of MDA measured after irradiation at the level of the cultures treated with the composition of the invention are clearly lower than that recorded at the level of the irradiated control cultures.

Claims (9)

1 / Topical cosmetic composition characterized in that it comprises a mixture of essential oils rich in carvacrol, eugenol and thymol.  2 / A composition according to claim 1, characterized in that it contains: between 9 and 15%, advantageously between 10 and 12% by weight of carvacrol, - between 3 and 10%, advantageously between 5 and 9% by weight of thymol between 10 and 18%, advantageously between 12 and 15% by weight of eugenol.  3 / Composition according to one of claims 1 to 2, characterized in that it contains a mixture of essential oils of green oregano (Origanum heracleoticum (L) carvacroliferum, done green Drome), clove (Eugenia

 caryophyllus (spreng)) and thyme (Thymus vulgaris (L) thymoloferum).  4 / Composition according to Claim 3, characterized in that it contains between 10 and 30%, advantageously 20% by weight respectively of essential oil of green oregano and clove and between 5 and 25%, advantageously 15% by weight of essential oil of thyme.  5 / composition according to one of claims 1 to 4, characterized in that it further contains a camphor rosemary essential oil (Romarinus officinalis (L) camphoriferum), used at a rate of 15 to 35% by weight.  6 / Composition according to one of claims 1 to 5, characterized in that it further contains a marjoram essential oil shell (Origanum majorana), used in a proportion of 10 to 20% by weight. <Desc / Clms Page number 14>  7 / Composition according to one of claims 1 to 6, characterized in that it further contains a zedoary essential oil (Curcuma zedoaria (Roscoe)), used in a proportion of 0.5 to 3% by weight.  8 / Cosmetic vehicle characterized in that it contains 0.2 to 0.5% by weight of the composition according to one of claims 1 to 7.  9 / Use of the composition object of claims 1 to 7 as an antioxidant, anti-radical, as a protective vis-à-vis oxidative stress. LABORATOIRESANOFLORESA REPRESENTATIVE: Cabinet LAURENT and CHARRAS  DEPOSITORS: GA TTEFOSSE HOLDING