FR3134010A1 - Cosmetic, nutraceutical and/or dermatological use of a strain of Lactobacillus crispatus and/or of a composition comprising it - Google Patents

Abstract

The present invention relates to the cosmetic, nutraceutical use of a strain of Lactobacillus crispatus, in particular that registered under the number CNCM I-5579 and/or of a composition comprising it, to reduce the pigmentation of the skin and/or mucous membranes and /or prevent its increase, in particular to preserve and/or improve the complexion of the skin and/or mucous membranes, preferably to preserve and/or improve the radiance and/or luminosity and/or homogeneity of the skin's complexion and/or mucous membranes, and/or to prevent the appearance and/or reduce the presence of pigment spots on the skin and/or mucous membranes and/or detoxify the skin and/or mucous membranes. It also relates to the strain of Lactobacillus crispatus, or a dermatological composition comprising it, for its use, advantageously topically, for the treatment and/or prevention of pathologies of the skin and/or mucous membranes associated with increased pigmentation. of the skin and/or mucous membranes, in particular to depigment and/or prevent hyperpigmentation of pathological skin and/or pathological mucous membranes.

Description

Cosmetic, nutraceutical and/or dermatological use of a strain of Lactobacillus crispatus and/or of a composition comprising it

The present invention relates to the cosmetic, nutraceutical and/or pharmaceutical, in particular dermatological, use of a strain of Lactobacillus crispatus and/or of a composition comprising it to reduce pigmentation of the skin and/or mucous membranes and/or prevent increased pigmentation of the skin and/or mucous membranes.

The structure and appearance of the skin are modified with age and/or under the influence of extrinsic and/or intrinsic factors. These changes are numerous and include changes in the pigmentation of the skin and/or mucous membranes. The skin becomes dull, less luminous, less radiant, its radiance diminishes and the complexion is less homogeneous with the appearance of pigment spots.

With chronological aging or accelerated aging, particularly under the effect of radiation, typically ultraviolet or pollution, the appearance of these modifications such as pigment spots is accelerated. They are linked to the presence of melanin and lipofuscin, the excess and abnormal distribution of which in the skin causes the appearance of so-called senile or pigmentary brown spots, also called “aged spots”. These can appear as a marker of accelerated melanogenesis and are often considered unsightly. They make the complexion uneven and dull.

Melanogenesis and oxidative stress are the subject of numerous studies and innovations in the field of cosmetics for the development and improvement of cosmetic active ingredients aimed at reducing and/or preventing the increase in skin pigmentation. healthy and/or healthy mucous membranes.

There is a need in the field of cosmetics and dermatology to develop alternative natural active ingredients capable of reducing the pigmentation of the skin and/or mucous membranes and/or preventing the increase in pigmentation of the skin. skin and/or mucous membranes, in particular to preserve and/or improve the complexion of the skin and/or mucous membranes, in particular its radiance and/or its luminosity and/or its homogeneity, particularly within a population more prone to pigment spots, particularly phototype IV to VI skin as defined by the Fitzpatrick classification method.

There is a continuing need to identify agents with one or more of these properties, in particular with proven effectiveness and safety.

Particularly surprisingly, the Applicant has discovered that Lactobacillus crispatus, a beneficial commensal bacteria of the skin and/or mucous membranes, has the property of reducing the pigmentation of the skin and/or mucous membranes and/or preventing its increase, in particular to preserve and/or improve the complexion of the skin and/or mucous membranes, preferably to preserve and/or improve the radiance and/or luminosity and/or homogeneity of the complexion of the skin and/or mucous membranes, and /or to prevent the appearance and/or reduce the presence of pigment spots on the skin and/or mucous membranes.

The Applicant has also unexpectedly discovered that this bacteria also has the property of detoxifying the skin and/or mucous membranes.

The Applicant's invention is all the more interesting since the bacteria Lactobacillus crispatus , and in particular that deposited under the number CNCM I-5579, has the advantage of being naturally present on young skin, unlike bacteria used until now in cosmetic and/or dermatological treatments.

This invention is even more interesting as the Applicant has demonstrated that with age and/or under the influence of intrinsic and/or extrinsic factors, the bacteria Lactobacillus crispatus is less present and less abundant on the skin. Thus, the object of the present invention has the advantage of rebalancing and/or supplementing the microbiota of the skin and/or mucous membranes, in particular with a bacteria of the skin flora which diminishes and becomes rarer over time and/or exposure to intrinsic and/or extrinsic factors.

The object of the present invention is therefore particularly suitable for the treatment of so-called mature skin, in particular people over 50 years old.

The strain according to the invention also has the advantage of being an easily formulated ingredient, and can be easily manufactured on an industrial scale. It is an active ingredient topically acceptable for the skin and mucous membranes, which does not present any risk of allergy.

Applications have already described the use of Lactobacillus crispatus in cosmetics. Application WO9822082 thus discloses the use of sphingomyelinase which can be extracted from lactic acid bacteria to be applied topically in order to increase the levels of ceramides in the skin and mucous membranes, for the prevention or therapeutic treatment of several pathologies.

Application WO2009031099 discloses the use of a probiotic to prevent the appearance and/or to treat the manifestation of sensations of discomfort and/or skin signs associated with a surface skin treatment or an invasive treatment, for aesthetic purposes .

Application WO2019111189 describes the use comprising a probiotic in particular Lactobacillus paracasei for the prevention of inflammatory damage induced by UV radiation.

However, none of these applications discloses or suggests the cosmetic, nutraceutical and/or pharmaceutical, in particular dermatological, use of a strain of Lactobacillus crispatus and/or of a composition comprising it to reduce skin pigmentation and/or mucous membranes, and/or to prevent its increase, in particular to preserve and/or improve the complexion of the skin and/or mucous membranes, preferably to preserve and/or improve radiance and/or luminosity and/or homogeneity of the complexion of the skin and/or mucous membranes, and/or to prevent the appearance and/or reduce the presence of pigment spots on the skin and/or mucous membranes.

Furthermore, none of these applications discloses or suggests the use of a strain of Lactobacillus crispatus and/or a composition comprising it to detoxify the skin and/or mucous membranes.

The first subject of the present invention is the cosmetic and/or nutraceutical use of a strain of Lactobacillus crispatus , preferably the strain of Lactobacillus crispatus deposited under the number CNCM I-5579, to reduce the pigmentation of the skin and/or mucous membranes. and/or to prevent its increase, in particular to preserve and/or improve the complexion of healthy skin and/or healthy mucous membranes, preferably to preserve and/or improve radiance and/or luminosity and/or homogeneity of the complexion of healthy skin and/or healthy mucous membranes, and/or to prevent the appearance and/or reduce the presence of pigment spots on healthy skin and/or healthy mucous membranes, preferably brown spots and/or spots of old age, and/or detoxify healthy skin and/or healthy mucous membranes, in particular phototype IV to VI skin as defined by the Fitzpatrick classification method.

According to a particular embodiment of this use, the strain according to the invention reduces and/or inhibits melanogenesis, in particular by acting on the reduction and/or inhibition of melanin, preferably on the reduction and/or inhibition the synthesis and/or quantity of melanin, and/or by reducing and/or inhibiting the activity of tyrosinase, and/or by detoxifying healthy skin and/or healthy mucous membranes.

According to one aspect of the invention, the strain according to the invention can be used to preserve and/or improve the complexion of healthy skin and/or healthy mucous membranes. The strain according to the invention can also be used to preserve and/or improve the radiance and/or luminosity and/or homogeneity of the complexion, healthy skin and/or healthy mucous membranes. The strain according to the invention can also be used to prevent the appearance and/or reduce the presence of pigment spots on healthy skin and/or healthy mucous membranes, preferably brown spots and/or age spots. The strain according to the invention can also be used to detoxify healthy skin and/or healthy mucous membranes, in particular by an antioxidant action and/or on lipid peroxidation and/or by an increase and/or stimulation of glutathione synthesis. induced by oxidative stress.

It also relates to a cosmetic treatment process characterized in that it comprises the topical application to at least one area of healthy skin and/or healthy mucous membrane of a strain of Lactobacillus crispatus according to the invention, preferably the strain of Lactobacillus crispatus CNCM I-5579, or a cosmetic composition comprising it, to reduce the pigmentation of the skin and/or mucous membranes and/or to prevent its increase, in particular to preserve and/or improve the complexion of healthy skin and/or healthy mucous membranes, preferably to preserve and/or improve the radiance and/or luminosity and/or homogeneity of the complexion of healthy skin and/or healthy mucous membranes, and/or to prevent the the appearance and/or reduce the presence of pigment spots on healthy skin and/or healthy mucous membranes, and/or detoxify the skin and/or mucous membranes.

The invention also relates to a strain of Lactobacillus crispatus according to the invention, preferably the strain of Lactobacillus crispatus CNCM I-5579, or to a pharmaceutical composition, in particular dermatological composition comprising it, for the prevention and/or treatment of pathologies of the skin and/or mucous membranes associated with an increase in pigmentation of the skin and/or mucous membranes, in particular to depigment and/or prevent hyperpigmentation of pathological skin and/or pathological mucous membranes, to reduce pigmentation of the skin and/or mucous membranes and/or to prevent its increase, in particular to preserve and/or improve the complexion of the pathological skin and/or the pathological mucous membranes, preferably to preserve and/or improve the radiance and/or the brightness and/or homogeneity of the complexion of the pathological skin and/or pathological mucous membranes, and/or to prevent the appearance and/or reduce the presence of pigment spots of the pathological skin and/or pathological mucous membranes and/or to detoxify pathological skin and/or pathological mucous membranes.

For the purposes of the present invention, the term “cosmetic use” means a non-pharmaceutical use, that is to say which is not intended for therapeutic use and is applied to a so-called healthy part of the body, in particular on an area of so-called healthy skin and/or mucous membranes.

For the purposes of the present invention, “nutraceutical use” means a use intended for non-pharmaceutical oral use, that is to say which does not require therapeutic treatment.

“Skin” means any area of skin on any part of the body and/or face, including the scalp.

For the purposes of the present invention, the term "phototype IV to VI skin" means particular skin types classified according to the Fitzpatrick scale which makes it possible to classify individuals according to the reaction of their skin and hair during exposure to UV rays, and/or classified as such by a dermatologist according to the tone of the skin, which depends in particular on the level of activity of the melanocytes, the creation of these cells and their groupings. Preferably, these dark skin types of phototype IV to VI come from characteristic ethnic origins, respectively from India, Asia and/or Africa. Especially,

- Phototype IV (Mediterranean type) corresponds to light matte skin and brown or black hair and eyes. Sunburns are rare, the skin tans well, a dark brown;

- Phototype V (Latino type) corresponds to light matte skin, black hair and eyes. Sunburns are rare, the skin tans well, a dark brown;

- Phototype VI (African type) corresponds to very dark skin, black hair and eyes. Sunburns are very rare.

According to the invention, the term "mucosa(s)" means the ocular mucosa, the nasal mucosa, the ear mucosa, the vaginal mucosa, the urogenital mucosa and/or the oral mucosa, in particular buccal, labial and/or gingival, preferably, the ocular and/or oral mucosa, and even more preferably, the labial and/or ocular mucosa. Advantageously, it does not include the vaginal mucosa.

For the purposes of the present invention, the term "healthy skin" or "healthy mucosa" means an area of skin or mucosa to which the strain according to the invention is applied and called "non-pathological" by a dermatologist, i.e. -meaning no infection, scar, disease, inflammation or skin condition such as candidiasis, impetigo, psoriasis, eczema, acne or dermatitis, or sores or injuries.

For the purposes of the present invention, the term “ Lactobacillus crispatus strain” means the bacterium Lactobacillus crispatus in whole form or in partial form, in particular in the form of lysate, in the form of a fraction or in the form of metabolites. Advantageously, the Lactobacillus crispatus strain is not genetically modified.

The strain of Lactobacillus crispatus according to the invention may be derived from any known species of Lactobacillus crispatus . Preferably, the strain of Lactobacillus crispatus is the species deposited according to the Budapest Treaty at the Institut Pasteur (28 rue du Docteur Roux, F-75024 Paris cedex 15) on 09/09/2020 under the designation CNCM I-5579 .

The advantage of this particular strain of Lactobacillus crispatus is that it is the first species of Lactobacillus crispatus found naturally on the skin.

For the purposes of the present invention, the term "reduce the pigmentation of the skin and/or mucous membranes and/or prevent its increase" means depigmenting, whitening, lightening, preventing the increase, maintaining and/or preserving and/or improving the complexion and/or complexion of the skin and/or mucous membranes, and preferably reduce and/or prevent the increase in melanogenesis, and still preferentially reduce and/or totally or partially inhibit gene and/or protein expression and /or tyrosinase activity and/or reduce and/or inhibit the synthesis and/or quantity of melanin.

The process of skin pigmentation is also called “melanogenesis” and includes numerous stages ranging from the synthesis of melanin in melanocytes to its transport into the keratinocytes of the epidermis via melanosomes. Several proteins are involved in this process. Tyrosinase is the enzyme which intervenes at key stages of melanin synthesis; it cooperates with other proteins to transform melanin into eumelanin and phaeomelanin, the melanins responsible for the pigmentation process.

The term “reduce and/or inhibit melanogenesis” means to reduce and/or totally or partially inhibit gene and/or protein expression and/or tyrosinase activity and/or reduce and/or totally or partially inhibit the synthesis. and/or the amount of melanin.

According to the invention, the term “reduce and/or totally or partially inhibit the activity of tyrosinase” is understood to cause a reduction of at least 25%, preferably of at least 50%, more preferably of at least 75% of the activity of tyrosinase in the presence of the strain of Lactobacillus crispatus according to the invention, compared to the level of activity measured in the absence of the extract.

Preferably, the measurement of tyrosinase activity is carried out on the tyrosinase extracted from so-called normal human melanocytes, still preferentially by measuring the production of melanin by photometrically measuring the absorbance at 540 nm from the tyrosinase extracted from melanocytes so-called normal humans, more preferably in the presence of the strain of Lactobacillus crispatus prepared according to Example 1.e, advantageously under the conditions described in Example 3.

According to the invention, the term "reduce and/or totally or partially inhibit the synthesis and/or the quantity of melanin" means a reduction in melanin of at least 10%, preferably at least 20%, more preferably by at least 30% of the synthesis and/or the quantity of melanin in the presence of the strain of Lactobacillus crispatus according to the invention compared to the synthesis and/or the quantity of melanin detected in the absence of the strain. In a preferred embodiment of the invention, this involves a reduction in the quantity of melanin in the presence of the strain of Lactobacillus crispatus according to the invention prepared according to Example 1.a or according to Example 1 .e. Still preferably, the quantity of melanin is measured by optical density at 475 nm from B16 melanocytes cultured in vitro in a culture medium comprising fetal calf serum, advantageously under the conditions described in Example 2. In another mode preferentially, the quantity of melanin is measured by optical density at 475 nm from a coculture of so-called normal human melanocytes and keratinocytes, cultured in vitro in a culture medium comprising growth factors, advantageously under the conditions described in Example 4.

For the purposes of the present invention, the term "preserve and/or improve the complexion" means maintaining and/or preventing the reduction and/or increasing the radiance and/or luminosity and/or radiance and/or shine of the complexion. and/or the homogeneity of the complexion and/or diminish and/or reduce the dull and/or earthy complexion of the skin and/or mucous membranes.

The measurement of the radiance of the complexion, the luminosity and the homogeneity of the complexion of the skin and/or the mucous membranes can be studied using measurement techniques known to those skilled in the art, in particular can for example be carried out in vivo , by chromametry or by image analysis. This last in vivo measurement method consists of taking high-resolution photographs in a crossed polarized configuration of the faces of volunteers taken at 45° before and after application of the product tested. On the basis of these digital photographs, an image analysis makes it possible to extract and quantify specific parameters (for example: L*, a*, b*, C, h°) related to color, brightness and the homogeneity of the skin.

For the purposes of the present invention, the term “preserve and/or improve the radiance of the complexion” means maintaining and/or preventing the reduction and/or increasing luminosity and/or radiance and/or shine and/or clarity. , and/or diminish and/or reduce the dull and/or earthy complexion of the skin and/or mucous membranes.

For the purposes of the present invention, the term "preserve and/or improve the homogeneity of the complexion" means maintaining and/or preventing the increase and/or reducing the quantity and/or extent of imperfections and/or irregularities of the complexion. complexion color in particular chosen from redness, pigment spots, blackheads, freckles and/or visible pores.

Measuring the evenness of skin tone and the amount of melanin in the skin can be measured by Mexametrie. This in vivo measurement method consists of using the absorption/reflection principle to deduce a melanin index which reflects the quantity of melanin present in the skin.

Advantageously, the measurement of the improvement in complexion is carried out on a population of women having pigment spots on the face, preferably by application of a cream containing the strain of Lactobacillus crispatus prepared according to example 1.e, in the conditions described in Example 7.

The term "maintain and/or prevent the increase and/or reduce imperfections and/or irregularities in the color of the complexion" means to prevent the increase and/or decrease the quantity of melanin in the skin, by at least 2%, advantageously at least 3% in the presence of the strain according to the invention during a 28-day application to the skin relative to the quantity of melanin detected in the absence of the strain and/or at least 2%, advantageously at least 5% in the presence of the strain according to the invention during a 56-day application to the skin compared to the quantity of melanin detected in the absence of the strain.

The term “preventing the appearance and/or reducing the presence of pigment spots” means maintaining and/or preventing the increase and/or reducing the pigmentation of localized pigment spots, in particular the number and/or intensity of the pigment spots. typically on areas of the face, décolleté, neck, back, shoulders and/or hands, particularly spots on the tops of the hands.

Pigment spots include brown and/or dark and/or white spots and/or “aged spots” when they are located on a part of the body exposed to UV radiation and/or linked to chronological or UV-induced aging, for example during exposure to the sun. Pigment spots also include freckles and/or moles and/or post-inflammatory spots and/or which appear in response to attacks, and/or of hormonal origin, in particular in the context of a chloasma or melasma, otherwise called a mask of pregnancy, and/or of medicinal origin, and/or to prevent and/or combat unsightly pigmentary manifestations of the skin and/or mucous membranes accompanying a non-cutaneous pathology.

The Applicant has also discovered that the strain of Lactobacillus crispatus , in particular that deposited under the designation CNCM I-5579, can be used to detoxify the skin and/or mucous membranes, in particular healthy skin and/or mucous membranes.

By “detoxifying the skin and/or mucous membranes” is meant the prevention and/or total and/or partial inhibition of oxidative stress, in particular by the prevention and/or total and/or partial inhibition of the formation of free radicals, and/or lipid peroxidation and/or by the increase and/or stimulation of the synthesis of glutathione which has antioxidant and detoxifying properties.

Detoxifying helps protect the skin against attacks, both external and internal, which cause oxidative stress likely to alter its proper functioning and appearance, in particular a means capable of stimulating the natural protection and repair systems of skin cells. and limit the induction of cellular damage.

“Oxidative stress” means the formation of excessive reactive species, including reactive oxygen species and reactive nitrogen species, which form when the skin is subjected to oxidative stress caused by various factors. such as solar ultraviolet, infrared and visible light, environmental pollution including ozone and particulate matter, and psychological stress.

In addition to impacting the effectiveness of the skin's detoxification systems, oxidative stress can also exacerbate pigmentation, in particular by inducing uneven skin tone or even a pigmentary disorder.

In a preferred embodiment of the invention, this involves reducing and/or inhibiting the formation of DPPH° (2,2-diphenyl-1-picrylhydrazyl) free radicals by at least 10%, preferably by at least 10%. less 30%, in the presence of the Lactobacillus crispatus strain according to the invention compared to the formation of DPPH° free radicals detected in the absence of the strain, and/or reduce and/or inhibit the release of induced intracellular free radicals by oxidative stress, preferably by UVA, of at least 20%, preferably of at least 50%, in the presence of the strain of Lactobacillus crispatus according to the invention in relation to the release of intracellular free radicals detected in absence of the strain.

In a preferred embodiment of the invention, this involves reducing and/or inhibiting the spontaneous formation of DPPH° free radicals, in the presence of the strain of Lactobacillus crispatus according to the invention prepared according to example 1.a and/or 1.e.

Still preferably, the formation of DPPH° free radicals is measured in tubo by optical density at 530 nm, under the conditions described in Example 5.a.

In a preferred embodiment of the invention, this involves reducing and/or inhibiting the release of intracellular free radicals induced by oxidative stress, preferably by UVA, in the presence of the strain of Lactobacillus crispatus according to the invention prepared according to example 1.a.

Preferably again, the release of intracellular free radicals induced by oxidative stress, preferentially by UVA, is measured by fluorescence emitted by the formation of DCF (2',7-dichlorofluorescein) from the DCFH-DA probe (2', 7'-dichlorodihydrofluorescein diacetate) at wavelengths of 485 nm excitation and 538 nm emission from normal human fibroblasts, under the conditions described in Example 5.b.

In another embodiment, it involves “preventing and/or inhibiting the peroxidation of membrane lipids”, that is to say preventing and/or totally or partially inhibiting the oxidation of lipids induced by a oxidative stress, preferably by UVA of at least 10%, preferably at least 30% in the presence of the strain of Lactobacillus crispatus according to the invention compared to the lipid peroxidation detected in the absence of the strain.

In a preferred embodiment of the invention, this involves preventing and/or inhibiting the peroxidation of membrane lipids induced by oxidative stress, preferably by UVA, in the presence of the strain of Lactobacillus crispatus according to the invention prepared according to example 1.a.

Preferably again, the peroxidation of membrane lipids induced by oxidative stress, preferentially by UVA, is measured by fluorescence of the formation of MalonDiAldehde (MDA) at wavelengths of 532 nm excitation and 560 nm emission from of normal human fibroblasts, under the conditions described in Example 5.c.

According to another embodiment, it involves maintaining and/or stimulating the synthesis of glutathione, that is to say preventing the decrease and/or increasing the expression and/or quantity of glutathione. in the presence of oxidative stress, preferably induced by UVA, of at least 20%, preferably at least 50%, in the presence of the strain of Lactobacillus crispatus according to the invention in relation to the synthesis of glutathione detected in the absence of the strain.

In a preferred embodiment of the invention, it involves maintaining and/or stimulating the synthesis of glutathione induced by oxidative stress, preferably by UVA, in the presence of the strain of Lactobacillus crispatus according to the invention prepared according to example 1.a.

Still preferably, the synthesis of glutathione is measured in vitro by fluorescence at wavelengths of 355 nm excitation and 430 nm emission from normal human fibroblasts, under the conditions described in Example 6.

According to the present invention, the strain of Lactobacillus crispatus can be used in whole form, in particular viable and/or inactivated, in particular dead, and/or in the form of lysate, and/or in the form of one or more of its fractions and /or in the form of one or more of its metabolites, preferably its secretome.

For each of its forms, the strain according to the invention can be isolated or associated with its culture medium.

Preferably, the strain according to the invention is used associated with its culture medium, that is to say contained in its culture medium.

For the purposes of the present invention, the term “culture medium” means a support containing the nutrient elements allowing the cultivation and/or multiplication of the strain of Lactobacillus crispatus according to the invention. Preferably, it is an MRS medium (Man, Rogosa and Sharp agar).

For the purposes of the present invention, “isolated” means not mixed with one or more compounds likely to be associated with it in its culture medium.

For the purposes of the present invention, the term “whole form” means its native form, a form in which the bacterial envelope is intact, as opposed to a lysed form. When used in whole form, the Lactobacillus crispatus strain may be viable and/or inactivated and/or dead.

For the purposes of the present invention, the term “viable” means a strain of Lactobacillus crispatus according to the present invention capable of forming colonies in culture.

The production of the strain of Lactobacillus crispatus in whole viable form can be carried out according to any method conventionally known to those skilled in the art. Advantageously, it will be carried out according to the protocol as described for example in example 1.a.

For the purposes of the invention, the term “inactivated” means a strain of Lactobacillus crispatus according to the present invention which is no longer capable, temporarily or permanently, of forming colonies in culture.

The inactivation of the Lactobacillus crispatus strain can be carried out using any method conventionally known to those skilled in the art. It can in particular be inactivated by irradiation, heat treatment or, under certain conditions, by high pressure treatment or by extrusion advantageously, it will be carried out by heat treatment, again advantageously according to the protocol as described for example in example 1.b.

Thermal inactivation can be carried out by incubating the Lactobacillus crispatus strain for a given period of time, advantageously from approximately 10 s to 90 min, preferably from approximately 15 minutes to one hour, and at a temperature advantageously of approximately 60 °C to 150°C. Preferably, it will be incubated for around 30 minutes at around 80°C.

For the purposes of the invention, “dead” means a strain of Lactobacillus crispatus according to the present invention which is no longer capable, definitively, of forming colonies in culture.

For the purposes of the invention, the term "lysate" means a material obtained following the destruction or dissolution of biological cells by a phenomenon called cell lysis thus causing the release of intracellular biological constituents naturally contained in the cells of the considered microorganism and fragments of cell membrane components. The lysate used is therefore formed of all the intracellular biological constituents and the constituents of the cell walls and membranes of the Lactobacillus crispatus strain according to the invention.

The lysate can be obtained by any method conventionally known to those skilled in the art. It may in particular be obtained by an osmotic shock, a thermal shock, by ultrasound, by enzymatic lysis, by tyndallization, or even under mechanical stress such as centrifugation, or by increasing the pressure or combinations of these different technologies.

Preferably, the lysate will be obtained by a combination of mechanical action and increased pressure. Still preferably, the lysate will be obtained according to the protocol as described for example in example 1.c.

In a preferred embodiment of the invention, the lysate will be associated with the culture medium of the bacteria.

For the purposes of the present invention, the term "fractions" means a fragment of the strain of Lactobacillus crispatus according to the present invention having an effectiveness in reducing the pigmentation of the skin and/or mucous membranes and/or preventing its increase, in particular to preserve and/or improve the complexion of the skin and/or mucous membranes, preferably to preserve and/or improve the radiance and/or luminosity and/or homogeneity of the complexion of the skin and/or mucous membranes, and/or to prevent the appearance and/or reduce the presence of pigment spots on the skin and/or mucous membranes and/or detoxify the skin and/or mucous membranes. This fraction corresponds to a non-total part of the intracellular biological constituents and the constituents of the cell walls and membranes obtained by lysis of the Lactobacillus crispatus strain according to the invention.

According to a particular embodiment of the invention, the fractions can be the protoplasm of the bacteria. According to a particular embodiment of the invention, the fraction corresponds to all of the intracellular biological constituents, but does not contain constituents of the cell walls and membranes of the Lactobacillus crispatus strain.

In a preferred embodiment of the invention, the fractions will be associated with the culture medium of the bacteria.

For the purposes of the present invention, the term “metabolites” means one or more organic and/or inorganic molecules resulting from the metabolism of the strain of Lactobacillus crispatus according to the present invention and also having an effectiveness in reducing the pigmentation of the skin and /or mucous membranes and/or prevent its increase, in particular to preserve and/or improve the complexion of the skin and/or mucous membranes, preferably to preserve and/or improve radiance and/or luminosity and/or homogeneity of the complexion of the skin and/or mucous membranes, and/or to prevent the appearance and/or reduce the presence of pigment spots on the skin and/or mucous membranes and/or detoxify the skin and/or mucous membranes. Preferably, it is the secretome of the Lactobacillus crispatus strain according to the invention.

In a particular embodiment of the invention, the metabolites will be associated with the culture medium of the bacteria.

For the purposes of the present invention, the term "secretome" means all of the secreted organic and/or inorganic molecules secreted by the strain of Lactobacillus crispatus according to the present invention having an effectiveness in reducing the pigmentation of the skin and/or or mucous membranes and/or to prevent its increase, in particular to preserve and/or improve the complexion of the skin and/or mucous membranes, preferably to preserve and/or improve radiance and/or luminosity and/or homogeneity of the complexion of the skin and/or mucous membranes, and/or to prevent the appearance and/or reduce the presence of pigment spots on the skin and/or mucous membranes and/or detoxify the skin and/or mucous membranes.

The secretome can be obtained by any method conventionally known to those skilled in the art. Advantageously, it will be obtained according to the protocol as described for example in example 1.e.

In a preferred embodiment of the invention, the secretome will be associated with the culture medium of the bacteria.

According to a preferred embodiment of the invention, the strain of Lactobacillus crispatus according to the invention will be used in whole viable form and/or in the form of one or more of its metabolites, preferably its secretome.

According to an advantageous embodiment, the Lactobacillus crispatus strain according to the invention does not contain sphingomyelinase.

According to an advantageous embodiment of the invention, the strain of Lactobacillus crispatus according to the invention is applied topically to healthy skin and/or healthy mucous membranes. Preferably, the skin and/or mucous membranes are phototype IV to VI. Phototypes IV to VI are as defined by the Fitzpatrick classification method.

For the purposes of the present invention, the term "topical route" means the direct local application and/or vaporization of the strain of Lactobacillus crispatus according to the invention or of the composition comprising it according to the invention on the surface of the affected area of skin and/or mucous membrane.

According to the invention, the strain of Lactobacillus crispatus according to the invention can be used alone, in the form of active ingredient and/or in a cosmetic and/or nutraceutical, and/or pharmaceutical composition, in particular dermatological, preferably intended for topical and/or oral application, more preferably for topical application.

When it is used in whole live form according to the invention, and when it is used alone, in the form of active ingredient, the strain according to the invention can be in dry form, that is to say in powder, advantageously in maltodextrin, preferably in a strain content of 10% to 80% (w/w) by weight of the strain, preferably from 30% to 70% (w/w), still advantageously from 40 to 60% (w/w) by weight of the strain relative to the total weight of powder.

Alternatively, the whole living form according to the invention can be used dispersed and/or diluted in a solvent. Preferably, this solvent contains less than 20% (v/v) by volume of water, still preferably less than 5% (v/v) by volume of water, still preferably the solvent does not contain water.

According to an alternative mode, this solvent contains less than 20% (w/w) by weight of water, still preferably less than 5% (w/w) by weight of water, still preferably the solvent does not contain water . Very preferably, this solvent is a cosmetically acceptable oil.

When it is used in whole inactivated form, in particular dead, and/or in the form of lysate, and/or in the form of one or more of its fractions and/or in the form of one or more of its metabolites, in particular of its secretome, and that it is used alone in the form of active ingredient, the strain according to the invention can be in dry form, that is to say in powder form, advantageously in maltodextrin, preferably in a strain content of 1% to 80% (w/w) by weight of the strain, preferably from 3% to 50% (w/w) by weight of the strain, very preferably around 5% or 20% ( p/w) by weight of the strain relative to the total weight of powder, still preferably around 10% (w/w), still preferably 10% (w/w) by weight of the strain relative to the total weight of powder.

Alternatively, the whole inactivated forms, in particular dead, and/or the lysates, and/or the fractions, and/or the metabolites, in particular the secretome according to the invention, can be used soluble and/or diluted and or dispersed in a solvent , in particular polar, such as water, an alcohol, a polyol, a glycol such as pentylene glycol and/or hexylene glycol and/or caprylyl glycol and/or butylene glycol, or one of their mixtures, preferably a hydroglycolic or hydroalcoholic mixture, more preferably comprising a glycol chosen from hexylene glycol, caprylyl glycol and their mixtures.

In another embodiment, the strain according to the invention can be incorporated into a cosmetic and/or nutraceutical composition comprising at least one cosmetically acceptable and/or nutraceutically acceptable excipient. Preferably, it is incorporated into a cosmetic composition comprising at least one cosmetically acceptable excipient.

For the purposes of the present invention, the term “cosmetically acceptable” excipient means a topically acceptable compound and/or solvent, that is to say one which does not induce an allergic response upon contact with the skin, including the scalp. human, and/or mucous membranes, non-toxic, not unstable, chemically.

In a preferred embodiment of the invention, the strain according to the invention is present in the cosmetic composition, at a concentration of 1x10 ^-4 % to 10% (w/w) by weight, preferably between 1x10 ^-4 % and 5% (w/w) by weight, more advantageously between 1x10 ^-3 % and 0.5% (w/w) by weight, relative to the total weight of the composition, said composition further comprising at least one cosmetically acceptable excipient .

Particularly advantageously, when the strain according to the invention is used in the form of one or more of its metabolites, in particular its secretome, it is present in the cosmetic composition, at a concentration of 0.05 to 0.5%. (w/w) by weight, more preferably at a concentration of approximately 0.1% (w/w) by weight, relative to the total weight of the composition, said composition further comprising at least one cosmetically acceptable excipient.

Particularly advantageously, when the strain according to the invention is used in whole viable form, it is present in the cosmetic composition, at a concentration of 0.01 to 0.1% (w/w) by weight, more preferably at a concentration of approximately 0.025% (w/w) by weight, relative to the total weight of the composition, said composition further comprising at least one cosmetically acceptable excipient.

The cosmetic composition of the invention can be chosen from a suspension or a solution, aqueous or oily, an aqueous cream or gel or an oily gel, in particular a shower gel, a shampoo; a milk ; an emulsion, a microemulsion or a nanoemulsion, in particular oil-in-water or water-in-oil or multiple or siliconized; a mask; a serum; lotion; liquid soap; a dermatological bar; an ointment ; a balm; a butter; a foam; a patch; an anhydrous product, preferably liquid, pasty or solid, for example in the form of makeup powders, rods or sticks, in particular in the form of lipstick.

It can also be a makeup product or a makeup remover product.

Advantageously, the strain of Lactobacillus crispatus or the composition of the invention is intended to be applied to healthy skin and/or healthy mucous membranes of all or part of the body and/or the face and/or the scalp, preferably the legs, thighs, arms, stomach, neckline, neck, armpits, lips, preferably all or part of the face, and preferably the cheeks, forehead, chin, lips, eye contour .

Advantageously, the strain and/or the composition comprising it is intended to be applied to an area of healthy skin presenting a dull complexion and/or an area of non-homogeneous healthy skin and/or an area of healthy skin with the appearance pigment spots.

Preferably, the strain of Lactobacillus crispatus according to the invention is particularly suitable for the formulation of so-called neutral and gentle composition for respecting the sebaceous gland, in particular the skin including the scalp and/or mucous membranes.

Alternatively, the Lactobacillus crispatus strain according to the invention can be presented in all the galenic forms conventionally used for oral application, in particular in the form of nutraceutical active ingredient and/or nutraceutical composition.

The compositions according to the invention may contain any suitable solvent and/or any suitable vehicle and/or any suitable excipient, optionally in combination with other compounds of interest.

Therefore, for these compositions, the excipient contains for example at least one compound chosen from the group comprising preservatives, emollients, emulsifiers, surfactants, moisturizers, thickeners, conditioners, mattifying agents, stabilizers , antioxidants, texturing agents, shine agents, film-forming agents, solubilizers, pigments, dyes, perfumes and sun filters. These excipients are preferably chosen from the group consisting of amino acids and their derivatives, polyglycerols, esters, polymers and cellulose derivatives, lanolin derivatives, phospholipids, lactoferrins, lactoperoxidases, stabilizers based on sucrose, vitamins E and its derivatives, natural and synthetic waxes, vegetable oils, triglycerides, unsaponifiables, phytosterols, plant esters, silicones and its derivatives, protein hydrolysates, Jojoba oil and its derivatives derivatives, lipo/water-soluble esters, betaines, aminoxides, plant extracts, sucrose esters, titanium dioxides, glycines, and parabens, and more preferably from the group consisting of butylene glycol, steareth-2, steareth-21, glycol-15 stearyl ether, cetearyl alcohol, phenoxyethanol, methylparaben, ethylparaben, propylparaben, butylparaben, butylene glycol, natural tocopherols, glycerin, dihydroxycetyl sodium phosphate, isopropyl hydroxycetyl ether, glycol stearate, triisononanoin, octyl cocoate, polyacrylamide, isoparaffin, laureth-7, a carbomer, propylene glycol, glycerol, bisabolol, a dimethicone, sodium hydroxide, PEG 30-dipolyhydroxysterate, capric/caprylic triglycerides, cetearyl octanoate, dibutyl adipate, grapeseed oil, jojoba oil, magnesium sulfate, EDTA, cyclomethicone, xanthan gum, citric acid, sodium lauryl sulfate, mineral waxes and oils, isostearyl isostearate, propylene glycol dipelargonate, propylene glycol isostearate, PEG 8, beeswax , hydrogenated palm heart oil glycerides, hydrogenated palm oil glycerides, lanolin oil, sesame oil, cetyl lactate, lanolin alcohol, castor oil, carbon dioxide titanium, lactose, sucrose, low density polyethylene, isotonic saline solution, and mixtures thereof.

Many cosmetically active ingredients are known to those skilled in the art to improve the health and/or physical appearance of the skin (including the scalp) and/or mucous membranes. Those skilled in the art know how to formulate cosmetic, nutraceutical or dermatological compositions to obtain the best effects. On the other hand, these compounds can have a synergistic effect when combined with each other. These combinations are also covered by the present invention. The CTFA Cosmetic Ingredient Handbook, Second Edition (1992) describes various cosmetic and pharmaceutical ingredients commonly used in the cosmetic and pharmaceutical industry, which are particularly suitable for topical use. Examples of these classes of ingredients include, but are not limited to, the following compounds: abrasives, absorbents, compounds for aesthetic purposes such as perfumes, pigments, dyes, essential oils, astringents, etc. (for example: clove oil, menthol, camphor, eucalyptus oil, eugenol, menthyl lactate, witch hazel distillate), anti-acne agents, anti-flocculating agents, anti-foaming agents, antimicrobial agents (for example: iodopropyl butylcarbamate), antioxidants, binders, biological additives, buffering agents, bulking agents, chelating agents, additives, biocidal agents, denaturants, thickeners, and vitamins, and derivatives or equivalents thereof, film-forming materials, polymers, opacifying agents, pH adjusters, reducing agents, depigmenting or lightening agents (for example: hydroquinone, kojic acid, ascorbic acid, magnesium ascorbyl phosphate, ascorbyl glucosamine), conditioning agents (for example: humectants).

The cosmetic composition may also comprise other cosmetic and/or nutraceutical agents having the same properties and inducing a synergistic effect or not with the strain of Lactobacillus crispatus according to the invention, or cosmetic agents with complementary effects.

These may for example be depigmenting and/or lightening ingredients and/or intended to reduce pigment spots on the skin, such as niacinamide or vitamin B3, arbutin, azelaic acid, ascorbic acid or its derivatives, a combination of Saxifraga sarmentosa, Psidium guajava and Carica papaya plant extracts, marketed by BASF Beauty Care Solutions France under the name Dermawhite™ WF, a combination of sulphites and extracts of Camellia sinensis, Scutellaria baicalensis, Cucumis sativus , Pyrus malus, marketed under the name Phytolight ^TM BG, a standardized extract of Lansium domesticum leaves marketed under the name DN-Aura ^TM , a combination of a pea extract and sucrose dilaurate marketed under the name Actiwhite™, derivatives of 4-hydroxyphenoxy acetic acid, in particular 2-(4-hydroxyphenoxy)-propionic acid marketed under the name Radianskin™ by BASF Beauty Care Solutions France, or an extract of purified polysaccharides from Cassia seeds angustifolia sold under the name Hyalurosmooth ^TM , an extract of stabilized proteases isolated from Carica papaya latex sold under the name X-Pressin ^TM , a cationic liposome encapsulating ferulic acid sold under the name Cytovector ferulic ^TM , or even a complex of glycolic acid and L-arginine marketed under the name AH-care ^TM by BASF Beauty Care Solutions France.

Among other active ingredients which may be combined with the strain according to the invention, it may be a plant extract having similar properties of preventing the appearance of pigmentation and/or increasing the radiance of the complexion, in particular an extract of the mushroom Inonotus obliquus marketed under the name Inolixir ^TM by the applicant, an extract of Argania spinosa oil marketed under the name Arganyl™ by the applicant, an extract of Moringa oleifera seeds marketed under the name of Purisoft™ by the applicant, a combination of an extract of Salvia miltiorrhiza and niacinamide marketed under the name of CollRepair™ by the applicant, an extract of Achillea millefolium marketed under the name of Neurobiox™ by the applicant, an extract of Cassia alata leaves sold under the name DN-Age™ and/or an extract of lychee sold under the name Litchiderm™ as anti-oxidant active ingredients, a chicory extract sold under the name Lox-Age™, a yeast extract marketed under the name Vitacell™, an extract of Polygonum bistorta marketed under the name Perlaura™, a galangal extract marketed under the name Hyalufix™, a corn extract marketed under the name of Deliner™ or an extract of Voandzeia subterranea marketed under the name of Epigenist™ by the applicant or even active ingredients promoting skin firmness such as a synthetic tetrapeptide marketed under the name of Dermican™, an extract of Hibiscus abelmoschus marketed under the name Linefactor™, a purified extract of pea marketed under the name Proteasyl™, an extract of Manilkara multinervis marketed under the name Elestan™ by the applicant. An extract of the Origanum majorana plant sold under the name Dermagenist™ and/or an extract of Khaya senegalensis sold under the name Collalift®18 and/or an extract of Eperua falcata sold under the name Eperuline™ or even an extract comprising phytosterols, in particular β-sitosterol and/or campesterol and/or brassicasterol coming from rapeseed oil and marketed by the applicant under the name Phytosoothe™, may also be added to the cosmetic composition.

They may also be anti-aging active ingredients and/or tightening agents for a synergistic effect with the strain of Lactobacillus crispatus . Advantageously, these are active ingredients increasing the gene and/or protein expression of collagen or active ingredients preventing the degradation of collagen such as retinol, vitamin C, an extract of Davilla rugosa marketed under the name Collguard ^TM by BASF Beauty Care Solutions, a Hibiscus abelmoschus seed extract marketed under the name Linefactor ^TM , a soy protein hydrolyzate extract marketed under the name Phytokine ^TM , a peptide marketed under the name Dermican ^TM by the applicant or the products marketed under the names Matrixyl ^TM , Matrixyl 3000 ^TM and Regestril ^TM by the company Sederma or Neuroguard by the company Codif, or even Eternaline ^TM by the company Silab.

The tensing agents which can be used in the invention can be chosen from synthetic polymers, such as polyurethane latexes or acrylic latexes, polymers of natural origin, in particular polyholosides in the form of starch or in the form of carrageenans, alginates, agars, gellans, cellulose polymers and pectins; plant proteins and protein hydrolysates; mixed silicates; wax microparticles; colloidal particles of inorganic filler chosen for example from silica, silica-alumina composites; as well as their mixtures.

Advantageously, the Lactobacillus crispatus strain according to the invention may be combined with a cosmetic agent chosen from benzoic acid and/or its salts, levulinic acid and/or its salts such as sodium levulinate, acid sorbic and/or its salts, anisic acid and/or its salts, and their mixture.

Finally, it may be cosmetic ingredients such as for example antimicrobial agents, anti-radical agents, soothing, calming or relaxing agents, agents acting on microcirculation to improve the radiance of the complexion, in particular of the face, healing agents or slimming agents. Advantageously, the cosmetic and/or dermatological composition of the present invention also contains one or more tensing agents and/or one or more antimicrobial agents and/or one or more anti-radical agents and/or one or more soothing agents and/or one or more agents. slimming agents and/or one or more agents active on the microcirculation.

Among the antimicrobial agents associated with the Lactobacillus crispatus strain in non-viable form in a preferred embodiment of the present invention, mention may be made of 2,4,4'-trichloro-2'-hydroxy diphenyl ether (or triclosan), 3 ,4,4'-trichlorobanilide, phenoxyethanol, phenoxypropanol, phenoxyisopropanol, hexamidine isethionate, metronidazole and its salts, miconazole and its salts, itraconazole, terconazole, econazole, ketoconazole, saperconazole , fluconazole, clotrimazole, butoconazole, oxiconazole, sulfaconazole, sulconazole, terbinafine, undecylenic acid and its salts, benzoyl peroxide, 3-hydroxy benzoic acid, 4-hydroxy acid benzoic acid, phytic acid, N-acetyl-L-cysteine acid, lipoic acid, azelaic acid and its salts, arachidonic acid, resorcinol, octoxyglycerin, octanoylglycine, caprylyl glycol , 10-hydroxy-2-decanoic acid, farnesol, phytosphingosines and their mixtures.

Anti-free radical agents can be vitamin C and its derivatives including ascorbyl glucoside, phenols and polyphenols, in particular tannins, ellagic acid and tannic acid; epigallocatechin and natural extracts containing it, in particular green tea extracts; anthocyanins; phenolic acids, stilbenes; active scavengers of mono- or polycyclic aromatic compounds, tannins such as ellagic acid and indole derivatives and/or active scavengers of heavy metals such as EDTA, anti-radical active ingredients such as vitamin E and its derivatives such as tocopheryl acetate; bioflavonoids; coenzyme Q10 or ubiquinone.

As soothing agents included in the composition of the invention, pentacyclic triterpenes, ursolic acid and its salts, oleanolic acid and its salts, betulinic acid and its salts, salts of salicylic acid can be used. and in particular zinc salicylate, bisabolol, Allantoin, unsaturated omega 3 oils, cortisone, hydrocortisone, indomethacin and beta methasone, anti-inflammatory active ingredients, and in particular those described in the application FR2847267, in particular the root extract of Pueraria lobata marketed under the name Inhipase™ by the applicant, the extracts of Theobroma cacao.

The active ingredients acting on microcirculation, vasoprotectors or vasodilators, can be chosen from flavonoids, ruscogenins, nicotinates and essential oils.

The slimming active ingredients can be chosen in particular from lipoprotein lipase inhibitor agents such as those described in patent US2003086949 (Coletica) and in particular an extract of Peruvian liana ( Uncaria tomentosa ); draining active ingredients, notably hesperitin laurate (Flavagrum™), or quercitin caprylate (Flavenger™); agents inhibiting the enzyme phosphodiestarase, activating agents of adenylate cyclase, cAMP and/or active ingredients capable of trapping spermine and/or spermidine. We can cite an extract of Coleus Forskohlii root, an extract of Cecropia obtusa , Uva lactuca , caffeine, forskolin, theophylline, theobromine and/or their derivatives, a hydrolyzed kappa carrageenan product called Slimexcess™ marketed by the applicant and/or their mixtures.

The present invention also relates to a cosmetic treatment process comprising the topical application to at least one area of healthy skin and/or healthy mucous membrane of a strain of Lactobacillus crispatus according to the invention, or of a composition cosmetic comprising it, to reduce the pigmentation of the skin and/or mucous membranes and/or prevent its increase, in particular to preserve and/or improve the complexion of healthy skin and/or healthy mucous membranes, preferably to preserve and/or improve radiance and/or luminosity and/or homogeneity of the complexion of healthy skin and/or healthy mucous membranes, and/or to prevent the appearance and/or reduce the presence of pigment spots on healthy skin and/or or healthy mucous membranes and/or detoxify the skin and/or mucous membranes.

Advantageously, the Lactobacillus crispatus strain according to the invention, preferably in the form of a cosmetic composition according to the invention, is used in regular topical application and preferably at least once a day, advantageously twice a day, for at least 10 days, preferably for 20 days, and even more preferably for at least 28 days.

Advantageously, the topical application of a strain of Lactobacillus crispatus according to the invention or of a cosmetic composition comprising it, is carried out on healthy skin and/or healthy mucous membranes of all or part of the body and/or or the face and/or scalp, preferably the legs, thighs, arms, stomach, neckline, neck, armpits, lips, even preferably all or part of the face, and preferably the cheeks, forehead , chin, lips, eye contour.

In an advantageous embodiment of the cosmetic process according to the invention, the cosmetic composition comprises the strain of Lactobacillus crispatus according to the invention at a concentration of between 1x10 ^-4 % and 10% (w/w) by weight, preferably between 1x10 ^-4 % and 5% (w/w) by weight, still advantageously between 1x10 ^-3 % and 0.5% (w/w) by weight, relative to the total weight of the composition, said composition further comprising an excipient cosmetically acceptable.

The cosmetic treatment process according to the invention aims to reduce the pigmentation of the skin and/or mucous membranes and/or to prevent its increase, in particular to preserve and/or improve the complexion of healthy skin and/or healthy mucous membranes, preferably to preserve and/or improve the radiance and/or luminosity and/or homogeneity of the complexion of healthy skin and/or healthy mucous membranes, and/or to prevent the appearance and/or reduce the presence of pigment spots on healthy skin and/or healthy mucous membranes, advantageously includes the following steps:

- The identification on the individual of an area of healthy skin and/or healthy mucous membrane, for which it is desired to reduce the pigmentation of the skin and/or mucous membranes and/or prevent its increase, in particular to preserve and/or improve the complexion of healthy skin and/or healthy mucous membranes, preferably to preserve and/or improve the radiance and/or luminosity and/or homogeneity of the complexion of healthy skin and/or healthy mucous membranes, and/ or to prevent the appearance and/or reduce the presence of pigment spots on healthy skin and/or healthy mucous membranes, and

- Topical application to this area of healthy skin and/or healthy mucous membrane, of a cosmetic composition comprising the strain of Lactobacillus crispatus according to the invention, in an effective quantity to reduce the pigmentation of the skin and/or mucous membranes and/or prevent its increase, in particular to preserve and/or improve the complexion of healthy skin and/or healthy mucous membranes, preferably to preserve and/or improve the radiance and/or luminosity and/or homogeneity of the complexion healthy skin and/or healthy mucous membranes, and/or to prevent the appearance and/or reduce the presence of pigment spots in healthy skin and/or healthy mucous membranes, namely in a content of Lactobacillus crispatus strain included advantageously between 1x10 ^-4 % and 10% (w/w) by weight, preferably between 1x10 ^-4 % and 5% (w/w) by weight, still advantageously between 1x10 ^-3 % and 0.5% (w/w ) by weight, relative to the total weight of the composition.

For the purposes of the present invention, the term "topically and/or orally acceptable" means an ingredient suitable for application respectively topically and/or orally, non-toxic, non-irritating to the skin including the scalp and does not does not induce an allergic response, and which is not chemically unstable.

The subject of the invention is also the strain of Lactobacillus crispatus according to the invention, alone or in a pharmaceutical composition, preferably dermatological, comprising it, for its use, advantageously topically for the treatment and/or prevention of skin pathologies. and/or mucous membranes involving hyperpigmentation and/or an increase in pigmentation, in particular a reduction in the radiance and/or luminosity and/or homogeneity of the complexion of the pathological skin and/or the pathological mucous membranes, and/ or an increase in the appearance of pigment spots in pathological skin and/or pathological mucous membranes, and/or an increase in the activity of melanogenesis, in particular through an increase in melanin and/or activity tyrosinase, and/or an increase in oxidative stress of pathological skin and/or pathological mucous membranes.

In particular, the subject of the invention is the strain of Lactobacillus crispatus according to the invention or a pharmaceutical composition, preferably dermatological, comprising it, for its use, advantageously topically, for the treatment and/or prevention of skin pathologies. and/or mucous membranes associated with an increase in pigmentation of the skin and/or mucous membranes, in particular to depigment and/or prevent hyperpigmentation of pathological skin and/or pathological mucous membranes, and/or for the treatment and/or mucous membranes. or the prevention of skin and/or mucous membrane pathologies associated with oxidative stress.

For the purposes of the present invention, from a dermatological point of view, “treatment and/or prevention of pathologies of the skin and/or mucous membranes to depigment and/or prevent hyperpigmentation” means a pigmentary disturbance under the effect pathologies of the skin and/or mucous membranes, such as for example Addison's disease, liver failure, purpura, melanoma, chloasma, dermatosis papulosa nigra, which is called pathological hyperpigmentation, or well again, acne, eczema, atopic dermatitis, rosacea, rosacea, redness.

For the purposes of the present invention, from a dermatological point of view, “treatment and/or prevention of pathologies of the skin and/or mucous membranes associated with oxidative stress” means the treatment and/or prevention of numerous skin diseases. linked to excessive oxidative stress, including psoriasis, certain eczemas, acne, vitiligo, and even certain skin cancers.

The Lactobacillus crispatus strain according to the invention can be found in the form of a pharmaceutical composition, preferably dermatological, comprising at least one pharmaceutically and/or dermatologically acceptable excipient. In one embodiment of the invention, said composition is applied topically and/or orally, preferably topically.

Advantageously, it is present in the pharmaceutical composition, preferably dermatological, at a concentration of 1x10 ^-4 % to 10% (w/w) by weight, preferably between 1x10 ^-4 % and 5% (w/w) by weight, again advantageously between 1x10 ^-3 % and 0.5% (w/w) by weight relative to the total weight of the composition, said composition further comprising at least one pharmaceutically, preferably dermatologically acceptable excipient.

The invention will be better understood on reading the description of the figures and examples which follow.

Examples referring to the description of the invention are presented below. These examples are given for illustration purposes and in no way limit the scope of the invention. Each of the examples has a general scope.

The examples form an integral part of the present invention and any characteristic appearing new compared to any state of the prior art from the description taken as a whole, including the examples, forms an integral part of the invention.

On the other hand, in the examples, all percentages are given by weight unless otherwise indicated, the temperature is expressed in degrees Celsius unless otherwise indicated, and the pressure is atmospheric pressure unless otherwise indicated.

Examples

Example 1. Preparation of different forms of Lactobacillus crispatus

Example 1.a: production of whole viable form of Lactobacillus crispatus .

The isolated Lactobacillus crispatus (CNCM I-5579) bacteria were seeded in an MRS type culture medium (Man's Agar, Rogosa, Sharpe), incubated at 37°C anaerobically. The pH was maintained at a value of pH = 6, and the medium was incubated for 24 h. At the end of incubation, the medium was centrifuged, the cell concentrate was lyophilized in the presence of maltodextrin, for a final quantity of maltodextrin of 50% (w/w) by weight relative to the total weight of the ingredient.

Example 1.b: production of the entire inactivated form of Lactobacillus crispatus

The isolated Lactobacillus crispatus (CNCM I-5579) bacteria were seeded in an MRS type culture medium, incubated at 37°C anaerobically. The pH was maintained at a value of pH = 6, and the medium was incubated for 24 h. At the end of incubation the medium was centrifuged, the cell concentrate was inactivated by heating it at 80°C for 30 minutes. The cell concentrate was then resuspended in glycerin.

Example 1.c: production of lysate of the bacteria Lactobacillus crispatus

The isolated Lactobacillus crispatus (CNCM I-5579) bacteria were seeded in an MRS type culture medium, incubated at 37°C anaerobically. The pH was maintained at a value of pH = 6, and the medium was incubated for 24 h. At the end of incubation the whole was passed through the high pressure homogenizer. The lysate was then resuspended in glycerin.

Example 1.d: production of the secretome of the bacterium Lactobacillus crispatus

The isolated Lactobacillus crispatus (CNCM I-5579) bacteria were seeded in an MRS type culture medium, incubated at 37°C anaerobically. The pH was maintained at a value of pH = 6, and the medium was incubated for 24 h. At the end of incubation the medium was filtered to recover the secretome of the bacteria. The secretome was then diluted in glycerin.

Example 1.e: production of the secretome of the bacteria Lactobacillus crispatus in powder form

The isolated Lactobacillus crispatus (CNCM I-5579) bacteria were seeded in an MRS type culture medium, incubated at 37°C anaerobically. The pH was maintained at a value of pH = 6, and the medium was incubated for 24 h. At the end of incubation the medium was filtered to recover the secretome of the bacteria. The secretome was then atomized with maltodextrin, for a final amount of maltodextrin of 90% (w/w) by weight relative to the total weight of the ingredient.

Example 2: Inhibition of melanin synthesis by melanocytes in the presence of the Lactobacillus crispatus strain according to the invention.

Melanocytes from a B-16 murine line were seeded in a 96-well plate at a rate of 8000 cells/cm ² , then cultured in MEM medium (Minimum Essential Medium) supplemented with 10% FBS (Fetal Bovine Serum) at 37°C, 5% CO ₂ and 95% Relative Humidity for 3 days.

The medium was then removed and replaced with 200 μL of the medium previously described with the addition of 1 μM of NDP-α-MSH (analog of melanotropic hormone) used as a stimulator of melanin synthesis and in the presence of the Lactobacillus crispatus strain according to Example 1.a at a concentration of 0.1% (w/w), for 5 days.

The same culture medium without addition of the strain according to the invention but with the addition of 1 μM of NDP-α-MSH or with the addition of kojic acid at 0.01% (w/w) were used as manipulation controls, respectively as melanin stimulatory control and melanin inhibitory control.

The same culture medium without addition of the strain according to the invention but with ethanol was used as a control (Control).

After 5 days of incubation, the cells were lysed with a solution of sodium hydroxide (NaOH) or potassium hydroxide (KOH). The absorbance was measured at 475nm.

The quantity of melanin was deduced from a standard range curve made up of different concentrations of commercial synthetic melanin.

The absorbance results were normalized in relation to the absorbance obtained with the same cellular medium in the absence of the Lactobacillus crispatus strain according to the invention (Control). The results presented correspond to the average of 6 replicates (n=6).

Results

Control (ethanol) NDP-α-MSH 1µM
(Melanin simulation control) Kojic acid 0.01% (w/w) (Melanin inhibition control) Product of the invention Lactobacillus crispatus (Example 1.a at 0.1% w/w) Average melanin (%) 24 100 18 48 Standard deviation 4 8 4 5 Statistics / NDP-α-MSH (*)
P<0.001 (Student T test) N / A (*)
P<0.001 (Student T test) (*) p<0.05 (One way ANOVA)

The strain of Lactobacillus crispatus according to Example 1.a product of the invention showed a significant inhibition of 52% of melanin synthesis by melanocytes of the B-16 murine line cultured in vitro . It can therefore be used to reduce pigmentation of the skin and/or mucous membranes and/or prevent its increase.

Example 3: Inhibition of tyrosinase activity in the presence of the Lactobacillus crispatus strain according to the invention.

Tyrosinase is a key enzyme in the melanin synthesis process responsible for skin tone. The tyrosinase used is extracted from normal human melanocytes.

The strain of Lactobacillus crispatus according to Example 1.e at a concentration of 0.05% (w/w) or a reference compound (kojic acid) at a concentration ranging from 0.25 mM to 8 mM, were prepared -incubated with the enzyme for 10 minutes on ice. Then, the tyrosinase substrate, L-Tyrosine at a final concentration of 1 mM was added to the medium and the plates were incubated at 37°C for 24 hours.

The same enzymatic extract without addition of the strain according to the invention but with ethanol was used as a control (Control).

Enzyme activity was assessed by measuring melanin production by optical density (OD) at 540 nm using a microplate reader.

The optical density measurements were also carried out without addition of substrate (n=1) in order to detect possible interference from the strain of Lactobacillus crispatus according to example 1.e or from the reference compound for each concentration tested.

The percentage inhibition of tyrosinase activity was calculated according to the following formula:

% inhibition: 100-[OD value/Average OD of control x 100]

All experimental conditions were carried out in n=3. Intergroup comparisons were performed by an unpaired Student's t test.

Unprocessed control
(ethanol) Product of the invention Lactobacillus
crispatus (Example 1.e at 0.05% (w/w)) Mean human tyrosinase inhibition (%) 0 99 Deviation from the average 3 0 Statistics/control N / A *** (p<0.001)

The Lactobacillus crispatus strain according to Example 1.e of the invention showed a significant inhibition of the activity of tyrosinase extracted from human melanocytes by 99%. By this action of inhibiting the activity of tyrosinase, the Lactobacillus crispatus strain can therefore be used to reduce the pigmentation of the skin and/or mucous membranes and/or prevent its increase.

Example 4: Inhibition of melanin synthesis in a co-culture of melanocytes and keratinocytes in the presence of the Lactobacillus crispatus strain according to the invention.

Normal human keratinocytes and human melanocytes from newborns were seeded at 20,000 cells/cm ² and 10,000 cells/cm ² respectively, then cultured for 7 days at 37°C under 5% CO ₂ and at 95% humidity. relative in a mixture of 50%/50% (v/v) of culture medium for melanocytes and culture medium for keratinocytes, supplemented with growth factors.

After one week of culture, the culture medium was replaced with a mixture of DMEM culture medium (Dulbecco's modified Eagle's minimum essential medium) without antibiotics and melanocyte medium without growth factors, and supplemented with 40µM oleic acid. .

This culture medium without addition of the strain according to the invention was used as an untreated control (Control).

To this same medium, the secretome of the bacteria Lactobacillus crispatus obtained according to Example 1.e was added at a concentration of 0.1% (w/w) relative to the total weight of the culture medium and the secretome.

The same culture medium without the strain, but in the presence of Rucinol at 0.002% (w/w) and was used as a positive control.

The cultures were continued for 72 hours at 37°C under 5% CO ₂ and at 95% relative humidity, then the culture medium was renewed. After 48 hours of additional culture, the medium was then eliminated, then the cells rinsed with PBS (phosphate buffer). Then, a 1N sodium hydroxide solution containing 10% Dimethylsulfoxide (DMSO) was added to each well of the plate. 200µL of each well were transferred to a transparent plate. The optical density reflecting the quantity of melanin was read at 475nm.

The results were the average of the tests performed ± the standard deviation, expressed as a percentage, relative to the untreated control normalized to 100%. Statistical analysis of the results was carried out in relation to the untreated control using the One-Way ANOVA test.

Results

Untreated control Rucinol 0.002% (w/w) (Positive control) Product of the invention L actobacillus crispatus at 0.1% (w/w) (Example 1.e) Average amount of melanin (%) 100 72 68 Standard deviation 10 3 7 % inhibition N / A 28 32 Statistics/control N / A P<0.01 P<0.01

The strain of Lactobacillus crispatus according to Example 1.e of the invention showed a significant reduction of 32% in the synthesis of melanin by melanocytes cocultured in vitro with human skin keratinocytes. Thus, the strain of Lactobacillus crispatus , and in particular its secretome can therefore be used to reduce the pigmentation of the skin and/or mucous membranes and/or prevent its increase.

Example 5: Reduction of oxidative stress in the presence of the Lactobacillus crispatus strain according to the invention.

Example 5.a: Reduction in the formation of DPPH° free radicals in tubo in the presence of the Lactobacillus crispatus strain according to the invention.

The assay of the antiradical activity of the Lactobacillus crispatus strain was evaluated by a test using DPPH° (2,2-diphenyl-1-picrylhydrazyl).

The strain of Lactobacillus crispatus according to Example 1.a was mixed in PBS (phosphate buffer saline) at a concentration of 0.05% (w/w), and according to Example 1.e at a concentration of 0.7% (w/w).

A negative control was carried out using the strain dissolution solvent, PBS, and a positive control was carried out with vitamin C at 1 mM.

Each of the solutions containing the invention, the negative control or the positive control was mixed at 50%/50% (v/v) in a DPPH° solution, in wells of 96-well plates.

Each of the solutions containing the invention, the negative control or the positive control was also mixed at 50%/50% (v/v) in an ethanol solution which is the solvent for DPPH°, in order to produce a blank, in this same plate.

The whole was incubated for 30 minutes with stirring. At the end of incubation, the optical density (OD) was measured at a wavelength of 530 nm to estimate the quantity of free radicals formed in each of the conditions tested.

The results of measuring the DPPH° radicals to obtain the anti-radical activity of the invention were the average of 6 tests (n=6) and were expressed as an average percentage +/- a standard deviation. The product of the invention was compared to its negative control (PBS alone in the absence of the Lactobacillus crispatus strain). The positive vitamin C control was compared to its negative control, water. The statistical analysis was carried out by a One Way ANOVA analysis of variance for the invention and by a Student test for vitamin C.

Negative control (PBS) Vitamin C 1mM Product of the invention (example 1e at 0.7% (w/w)) Product of the invention Lactobacillus crispatus (Example 1a at 0.05% (w/w)) Average radicals (%) 100 0 68.81 66.19 Standard deviation 4.33 0 3.44 2.21 Statistics / control N / A P<0.001 (Student t test) Not tested P<0.001 (One way ANOVA)

The strain of Lactobacillus crispatus according to example 1.a of the invention showed a significant in tubo anti-radical activity of 34%. The Lactobacillus crispatus strain according to Example 1.e of the invention showed an in tubo anti-radical activity of 31%. Through this action of reducing the production of free radicals, the Lactobacillus crispatus strain can therefore be used to detoxify the skin and/or mucous membranes, to reduce the pigmentation of the skin and/or mucous membranes and/or prevent its increase.

Example 5.b: Decrease in the release of cellular free radicals induced by UVs within fibroblasts in the presence of the Lactobacillus crispatus strain according to the invention.

So-called “normal” human fibroblasts, that is to say not presenting pathology, from a healthy donor were seeded in a 24-well plate at a rate of 30,000 cells/cm², then cultured in a DMEM medium (medium essential minimum of modified Eagle Dulbecco) / Ham's F-12, supplemented with 10% FCS and 0.5% antibiotic, for 96 hours at 37°C, under 5% CO ₂ and 95% relative humidity.

Then, 0.5 mL of the cell suspension was incubated for 72 hours in the presence of the Lactobacillus crispatus strain according to Example 1.a at a concentration of 0.0125% (w/w), or an antioxidant control (Vitamin E at 0.0003% (v/v)) in EMEM medium (Eagle's minimum essential medium) supplemented with 1% FCS and 0.5% antibiotic.

This culture medium without addition of the strain according to the invention was used as an untreated control (UVA control alone - Oxidative stress control).

A DCFH-DA (2',7'-dichlorodihydrofluorescein diacetate) probe solubilized at 10 μM in PBS (phosphate buffer) was incubated with the cells. After the first hour of incubation at 37°C under 5% CO ₂ and 95% relative humidity, the cells were rinsed with PBS, and irradiated with UVA at 20J/cm² for 3 hours. A non-irradiated control was also produced. After irradiation, the irradiated medium was removed, and the cells were rinsed with PBS.

The reading of fluorescence on the cell layer linked to the formation of DCF (2',7-dichlorofluorescein) from the probe (DCFH-DA) was read at 485 nm excitation and 538 nm emission.

The results were the average of 12 replicates (n=12) and were expressed as Mean +/- one standard deviation. The results were compared against the UVA alone control normalized to 100% and statistically compared using the One Way ANOVA analysis of variance test.

UVA control alone (Oxidative stress control) Vitamin E 0.0003% (v/v) Product of the invention Lactobacillus crispatus (Example 1a at 0.0125% (w/w)) Average radicals measured by DCF fluorescent probe (%) 100 71 44 Standard deviation 8 25 21 Statistics / control N / A P<0.05 (Student's t-test) P<0.05 (student's t test)

The Lactobacillus crispatus strain according to example 1.a of the invention showed a significant reduction at the cellular level of 56% in the release of intracellular free radicals induced by UVA oxidative stress. Through this action of reducing the release of intracellular free radicals induced by UVA, the Lactobacillus crispatus strain can therefore be used to detoxify the skin and/or mucous membranes and/or preserve and/or improve radiance and/or luminosity and/or homogeneity of the skin and/or mucous membranes, preferably reducing the presence and/or preventing the appearance of pigment spots on the skin and/or mucous membranes, particularly when these undergo oxidative stress, preferably by UVA.

Example 5.c: Decrease in the peroxidation of membrane lipids induced by UVs within fibroblasts in the presence of the Lactobacillus crispatus strain according to the invention.

So-called “normal” human fibroblasts, that is to say not presenting pathology, from a healthy donor were seeded in a 24-well plate at a rate of 30,000 cells/cm², then cultured in a DMEM medium. /Ham's F-12, supplemented with 10% FCS and 0.5% antibiotic, for 96 hours at 37°C, under 5% CO ₂ and 95% relative humidity.

Then, 0.5 mL of the cell suspension was incubated for 72 hours in the presence of the Lactobacillus crispatus strain according to Example 1.a at a concentration of 0.00625% (w/w), or an antioxidant control (Vitamin E at 0.0003% (v/v)) in EMEM medium supplemented with 1% FCS and 0.5% antibiotic.

This culture medium without addition of the strain according to the invention was used as an untreated control (after irradiation, constitutes the UVA control alone - Oxidative stress control).

After the first hour of incubation at 37°C under 5% CO ₂ and 95% relative humidity, the cells were rinsed with PBS, and irradiated with UVA at 20J/cm² for 3 hours. A non-irradiated control was also produced. After irradiation, the supernatants were collected and the cells were rinsed with PBS.

A quantity of 0.5 mL of the supernatants was transferred into a Pyrex glass tube to which a solution of 40% trichloroacetic acid and 2% thiobarbituric acid was added. The mixture obtained was heated for 30 minutes at 100°C. Then, the formation of MDA (MalonDiAldehde), which is an oxidation product released following UVA-induced lipid peroxidation, was measured by fluorescence at 532nm excitation and 560nm emission.

The results were the average of the tests carried out and were expressed as Mean +/- one standard deviation. The results were compared against the UVA alone control normalized to 100% and statistically compared using the One Way ANOVA analysis of variance test.

UVA control alone (Oxidative stress control) Vitamin E 0.0003% (v/v) Product of the invention Lactobacillus crispatus (Example 1a at 0.00625% w/w) Average MDA released (%) 100 48 68 Standard deviation 11 10 12 Statistics / control N / A P<0.001 (Student's t-test) P<0.001 (student's t test)

The Lactobacillus crispatus strain according to example 1.a of the invention showed a significant reduction at the cellular level in the formation of MDA induced by UVA of 32%. Through this action of reducing lipid peroxidation in the presence of UV stress, the Lactobacillus crispatus strain can therefore be used to detoxify the skin and/or mucous membranes, particularly when they undergo oxidative stress, particularly stress. by UVA.

Example 6: Stimulation of cellular antioxidant defenses after UV irradiation within fibroblasts in the presence of the Lactobacillus crispatus strain according to the invention.

So-called “normal” human fibroblasts, that is to say not presenting pathology, from a healthy donor were seeded in a 24-well plate at a rate of 30,000 cells/cm², then cultured in a DMEM/Ham's medium. F-12, supplemented with 10% FCS and 0.5% antibiotic, for 96 hours at 37°C, under 5% CO ₂ and 95% relative humidity.

This culture medium without addition of the strain according to the invention was used as an untreated control (after irradiation, constitutes the UVA control alone - Oxidative stress control).

Then, 0.5 mL of the cell suspension was incubated for 72 hours with the Lactobacillus crispatus strain according to Example 1.a at a concentration of 0.0125% (w/w), or an antioxidant control (Vitamin E at 0.0003% (v/v)) in EMEM medium supplemented with 1% FCS and 0.5% antibiotic.

After the first hour of incubation at 37°C under 5% CO ₂ and 95% relative humidity, the cells were rinsed with PBS, and irradiated with UVA at 20J/cm² for 3 hours. A non-irradiated control was also produced. After irradiation, the irradiated medium was removed, and the cells were rinsed with PBS.

1N sodium hydroxide (NaOH) was added to each well, followed by a solution of o-phthaldialdehyde (OPT) diluted ^1/15 in GSH buffer (reduced L-glutathione). The whole was incubated for 15 minutes at room temperature. Then the quantity of glutathione was measured by fluorescence at 355 nm excitation and 430 nm emission.

The results were the average of the tests carried out and were expressed as Mean +/- one standard deviation. The results were compared against the UVA alone control normalized to 100% and statistically compared using the One Way ANOVA analysis of variance test.

UVA control alone (Oxidative stress control) Vitamin E 0.0003% (v/v) Product of the invention Lactobacillus crispatus (Example 1a at 0.0125% (w/w)) Average Glutathione (%) 100 100 154 Standard deviation 11 13 41 Statistics / control N / A NS P<0.05 (student's t test)

The Lactobacillus crispatus strain according to example 1.a of the invention showed a significant increase in cellular antioxidant defenses by an increase in glutathione of 54% after UVA irradiation.

By this action of increasing cellular antioxidant defenses in the presence of oxidative stress by UVA, the strain of Lactobacillus crispatus can therefore be used to detoxify the skin and/or mucous membranes, particularly when they undergo oxidative stress, notably a UVA stress.

Example 7: In vivo measurement of the reduction in the quantity of melanin in the skin in the presence of the Lactobacillus crispatus strain according to the invention.

A population of 30 Chinese women of phototype II to IV, aged 40 to 50 years old, with pigment spots on the face, applied to the half-face a cream in the form of an emulsion comprising a final concentration by weight of the strain Lactobacillus crispatus according to example 1.e of 1% (w/w) relative to the total weight of the cream (Formulation example 8.b), or without said extract replaced by water (placebo), at a rate of 2 applications per day for 2 months.

The assessment of the skin's melanin content was carried out using a device called Mexameter which uses the absorption/reflection principle. From the Mexameter measurements, a melanin index is deduced. A decrease in this index corresponds to a decrease in melanin.

The effectiveness of the composition containing the strain of Lactobacillus crispatus according to Example 1 was compared with that of the so-called placebo emulsion.

The melanin index was measured at the start of the study (D0) after one month of application (D28) and finally after 2 months of application (D56).

The effectiveness of the formulation containing the strain of Lactobacillus crispatus according to example 1.e and of the placebo formula were expressed as an average and were compared with respect to the values at the start of the study (D0) and compared between the formulation containing the Lactobacillus crispatus strain with that containing the placebo, using the Student t test or the Wilcoxon test.

Product of the invention Lactobacillus crispatus (Example 1e at 1% (w/w)) Placebo Melanic index at D28 (in % vs D0) -3.1% (p<0.001 vs D0; p<0.001 vs placebo) -0.1% Melanic index at D56 (in % vs D0) -5.5% (p<0.01 vs D0; p<0.01 vs placebo) +1%

The formulation of the Lactobacillus crispatus strain according to Example 1 significantly reduced the quantity of skin melanin measured via the melanin index using a Mexametre.

These results show the beneficial effect of the Lactobacillus crispatus strain formulated to preserve and/or improve the radiance and/or luminosity and/or homogeneity of healthy skin and/or healthy mucous membranes.

Example 8: Example of cosmetic composition comprising the Lactobacillus crispatus strain according to the invention

Example 8.a: Example of cosmetic composition comprising the Lactobacillus crispatus strain according to Example 1.a

The Lactobacillus crispatus strain obtained according to Example 1.a was added just before application to the skin in a formulation as defined below according to Table 8, at a final concentration of 0.05% (w/w ) relative to the total weight of the formulation.

Denomination Quantity
(% by total weight) Phase A Cetearyl alcohol, lecithin, sodium cetearyl sulfate, olus oil 3.00 Hydrogenated Vegetable Glycerides 2.50 Caprylyl Caprylate/Caprate 6.50 Dicaprylyl carbonate 4.00 Phase B Water 66.35 Butylene Glycol 10.00 Sodium Benzoate 0.25 Phase C Glycerin 5.00 Xanthan gum 1.00 Phase D Glycerin, water, Sodium Levulinate, Sodium Anisate 1.00 Citric acid 0.40

Example 8.b: Example of cosmetic composition comprising the Lactobacillus crispatus strain according to Example 1.e

The proportions are expressed in % and the names in capital letters correspond to the INCI names of the ingredients.

Phase A GLYCERIN 5.00 BUTYLENE GLYCOL 10.00 XANTHAN GUM 2.00 Phase B Water 58.95 Sodium benzoate 0.25 GLYCERIN, WATER, SODIUM LEVULINATE, SODIUM ANISATE 1.00 Phase C POLYGLYCERYL-2 DIPOLYHYDROXYSTEARATE 1.00 DIPROPYLHEPTYL CARBONATE 3.00 DICAPRYLYL CARBONATE 3.00 CAPRYLYL CAPRYLATE/CAPRATE 10.00 LAURETH-7 CITRATE 0.50 Phase D Citric acid 0.30 Phase E Extract of Lactobacillus crispatus obtained according to example 1e 1.00 Water 4.00

We proceed according to methods known to those skilled in the art to mix together the different phases A, B, C at room temperature, by adding B to A while stirring, then C to AB. The pH is adjusted to 4.9 with D. Then phase E is added. Finally, the mixture is homogenized using Ultra Turrax to prepare a composition according to the present invention.

Claims (21)

Cosmetic and/or nutraceutical use of a strain of Lactobacillus crispatus , preferably the strain of Lactobacillus crispatus being the species registered under the designation CNCM I-5579, to reduce pigmentation of the skin and/or mucous membranes and/or prevent its increase. Use according to claim 1, in which the strain reduces and/or inhibits melanogenesis, in particular by acting on the reduction and/or inhibition of melanin, preferably on the reduction and/or inhibition of the synthesis and/or the quantity of melanin, and/or by reducing and/or inhibiting the activity of tyrosinase, and/or by detoxifying healthy skin and/or healthy mucous membranes. Use according to any one of the preceding claims to preserve and/or improve the complexion of healthy skin and/or healthy mucous membranes. Use according to any one of the preceding claims, to preserve and/or improve the radiance and/or luminosity and/or homogeneity of the complexion, healthy skin and/or healthy mucous membranes. Use according to any one of the preceding claims, to prevent the appearance and/or reduce the presence of pigment spots on healthy skin and/or healthy mucous membranes, preferably brown spots and/or age spots. Use according to any one of the preceding claims, for detoxifying healthy skin and/or healthy mucous membranes, in particular by an antioxidant action and/or on lipid peroxidation and/or by an increase and/or stimulation of glutathione synthesis. induced by oxidative stress. Use according to any one of the preceding claims, in which the strain is used in whole form, in particular viable and/or inactivated, in particular dead, and/or in lysate form, and/or in the form of one or more of its fractions, and/or in the form of one or more of its metabolites, preferably its secretome. Use according to any one of the preceding claims, in which the strain of Lactobacillus crispatus is that deposited under the number CNCM I-5579. Use according to one of the preceding claims, in which the Lactobacillus crispatus strain is used in whole viable form and/or in the form of one or more of its metabolites, preferably its secretome. Use according to one of the preceding claims, in which the strain of Lactobacillus crispatus is associated with its culture medium. Use according to one of the preceding claims, in which the strain of Lactobacillus crispatus is applied topically to healthy skin and/or healthy mucous membranes. Use according to claim 11 in which the skin is phototype IV to VI. Use according to any one of the preceding claims, such that the strain of Lactobacillus crispatus is present in a cosmetic composition at a concentration of 1x10 ^-4 % to 10% by weight, preferably between 1x10 ^-4 % and 5% by weight, again advantageously between 1x10 ^-3 % and 0.5% by weight, relative to the total weight of the composition, said composition further comprising at least one cosmetically acceptable excipient. Cosmetic treatment process characterized in that it comprises the topical application to at least one area of healthy skin and/or healthy mucous membrane of a strain of Lactobacillus crispatus as defined according to one of claims 7 to 10 , or a cosmetic composition according to claim 13, to reduce the pigmentation of the skin and/or mucous membranes and/or prevent its increase, in particular to preserve and/or improve the complexion of healthy skin and/or healthy mucous membranes , preferably to preserve and/or improve the radiance and/or luminosity and/or homogeneity of the complexion of healthy skin and/or healthy mucous membranes, and/or to prevent the appearance and/or reduce the presence of pigment spots on healthy skin and/or healthy mucous membranes and/or detoxify the skin and/or mucous membranes. Cosmetic treatment method according to claim 14, in which the topical application of a strain of Lactobacillus crispatus as defined according to one of claims 7 to 10, or of a cosmetic composition according to claim 13, is carried out on healthy skin and/or healthy mucous membranes of all or part of the body and/or face and/or scalp, preferably the legs, thighs, arms, stomach, décolleté, neck, armpits, the lips, again preferably all or part of the face, and preferably the cheeks, the forehead, the chin, the lips, the eye contour. Cosmetic care process according to any one of claims 14 to 15, in which the cosmetic composition comprises the strain of Lactobacillus crispatus as defined according to one of claims 7 to 10 at a concentration of 1x10 ^-4 % to 10% in weight, preferably between 1x10 ^-4 % and 5% by weight, more advantageously between 1x10 ^-3 % and 0.5% by weight, relative to the total weight of the composition, said composition further comprising at least one cosmetically acceptable excipient. Strain of Lactobacillus crispatus , or a pharmaceutical composition, preferably dermatological comprising it, for its use, advantageously topically, for the treatment and/or prevention of pathologies of the skin and/or mucous membranes associated with an increase in pigmentation of the skin and/or mucous membranes, in particular to depigment and/or prevent hyperpigmentation of pathological skin and/or pathological mucous membranes, and/or for the treatment and/or prevention of pathologies of the skin and/or mucous membranes associated with oxidative stress. Strain according to claim 17 in which the strain is used in whole form, in particular viable and/or inactivated, in particular dead, and/or in lysate form, and/or in the form of one or more of its fractions, and/ or in the form of one or more of its metabolites, preferably its secretome, and/or in which the strain is associated with its culture medium. Strain of Lactobacillus crispatus for its use according to any one of claims 17 to 18, characterized in that it is in the form of a pharmaceutical composition, preferably dermatological, comprising at least one dermatologically acceptable excipient. Strain of Lactobacillus crispatus for its use according to any one of claims 17 to 19, characterized in that it is present in the pharmaceutical composition, preferably dermatological, at a concentration of 1x10 ^-4 % to 10% by weight, preferably between 1x10 ^-4 % and 5% by weight, still advantageously between 1x10 ^-3 % and 0.5% by weight, relative to the total weight of the composition, said composition further comprising at least one pharmaceutically, preferably dermatologically, acceptable excipient. Strain of Lactobacillus crispatus for its use according to any one of claims 17 to 20, characterized in that the strain is as defined according to one of claims 7 to 10.