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    • 专利摘要:
    Bifidobacterium lysate for the prevention and/or treatment of skin damage caused by reactive oxygen species generated by visible high energy radiation. The application refers to a lysate of at least one microorganism of the genus Bifidobacterium in effective amount for use in the prevention and/or treatment of skin damage caused by the effect of reactive oxygen species (ROS) generated by the effect of visible high energy radiation, as well as its combinations with other assets and compositions that comprise them for such use. (Machine-translation by Google Translate, not legally binding)
    公开号:ES2725348A1
    申请号:ES201830281
    申请日:2018-03-22
    公开日:2019-09-23
    发明作者:Regueiro Susana Mezquita；Galar Maialen Elizari；Zorzano Eduardo Gonzalez
    申请人:Laboratorios Cinfa SA；
    IPC主号:
    专利说明:

    [0001]
    [0002] Bifidobacterium lysate for the prevention and / or treatment of skin damage caused by reactive oxygen species generated by visible high energy radiation
    [0003]
    [0004] The invention relates to the field of cosmetics and / or dermatological products, more particularly those intended for the protection of the skin against solar radiation and the repair of its harmful effects. In particular, the invention relates to the use of a biotechnologically derived asset for the prevention and treatment of the skin against cellular damage caused by radiation, in particular high energy visible radiation, as well as cosmetic compositions comprising it.
    [0005]
    [0006] STATE OF THE TECHNIQUE
    [0007]
    [0008] It is well known that solar radiation damages human skin, for example causing burns and premature aging of the skin (i.e. photo aging).
    [0009]
    [0010] Sunscreens are the main protectors against solar radiation. They can reflect, absorb or disperse sunlight, protecting the skin from the harmful effects of the sun. In general, an effective sunscreen should avoid burns but also minimize damage to the skin due to the accumulation of radiation received. There are mainly three types of filters used in the protection of solar radiation: physical, chemical and biological filters. The physical filters are inorganic or inert powders of small particles around 180-190 nm in diameter, composed of TiO2, ZnO, ferrous oxide, MgO, mica or talc. They act by reflecting solar radiation regardless of their wavelength. Chemical filters are synthetic chemicals that act by absorbing solar radiation. They function as chromophores that absorb the energy carried by an incident photon, then return to their initial state and release excess energy such as heat (insignificant), fluorescent radiation or chemical transformation into a potentially reactive isomer photoproduct. Biological filters are substances with antioxidant activity that, when applied topically, reduce oxidative stress induced by UV radiation. Improve the protection of the protectors Traditional sunscreens protecting from cellular damage that could be the cause of photoaging and skin cancer. They are vitamins (A, C or E), flavonoids (chelator of Fe) and certain trace elements (enzyme activity).
    [0011]
    [0012] Classic sunscreens protect against ultraviolet (UV) radiation, in particular, UVB and UVA. UV radiation (200-400 nm) accounts for approximately 6% of the solar radiation spectrum and causes the destruction of collagen fibers, as well as the abnormal production of elastic fibers. It also promotes alterations in dermal microvascularization. Depending on the intensity of the ray, the skin can suffer degeneration in its structure and abnormalities in its normal pigmentation process.
    [0013]
    [0014] UVB light causes DNA damage but also inhibits DNA repair. As a consequence of DNA damage, regulation of IL-10 occurs, which in turn leads to inhibition of IL-12 formation. IL-12 is one of the main actors involved in the orchestration of immune responses. Maintaining the balance IL10 / IL12 is a prerequisite to counteract photoaging and chronic radiation damage. In particular, intensive exposure of the skin to UV radiation leads to modifications of DNA bases, in particular, to a dimerization of thymidine by cycloaddition between two adjacent thymidine elements. This change in the structure of the nucleic acid can lead to cell death or inheritable cell damage. The natural mechanism of cell DNA repair, which is based on an enzymatically controlled reaction sequence to cut thymidine dimers out of the DNA chain and replace them with thymidine monomers, has only a limited capacity and can be overcome by the harmful effect of radiation.
    [0015]
    [0016] European patent application EP43128 describes the use of a lysate of microorganisms such as a lysate of Bifidobacterium longum, to promote DNA repair in damaged skin cells by exposure to ultraviolet radiation. It was shown that it inhibits the release of immunosuppressive IL-10, which simultaneously prevents the inhibition of IL-12, and allows to increase the DNA repair capacity of skin cells by the body itself.
    [0017] However, the damage suffered by the skin is not limited to the damage caused in the DNA. The radiation is harmful to the skin by other additional mechanisms, among which the oxidative stress caused by the generation of reactive oxygen species stands out.
    [0018]
    [0019] Approximately 6% of the solar spectrum is UV radiation (200-400 nm), 52% is visible light (400 - 760 nm) and the remaining 42% is infrared radiation (760 nm - 10 nm).
    [0020]
    [0021] It has been described that near infrared radiation, or infrared A (IRA) has the ability to penetrate deep layers of the skin. While the infrared radiation of longer wavelengths (IRB and IRC) does not penetrate deeply into the skin, more than 65% of the shortest wavelength (ARI) reaches the dermis, directly attacking the cellular mitochondria responsible for cell energy supply, so that it produces an increase in free radicals, especially reactive oxygen species (ROS). When free radicals accumulate in the cell, as a consequence of the IRA-induced mitochondrial oxidative stress response, among other things there is an increase in the expression of matrix metalloproteinases (MMP) and a reduced synthesis of collagen and elastin .
    [0022]
    [0023] Until recently it was thought that visible light did not cause damage to skin cells. However, it has been shown that blue light or high energy visible light (HEV), which refers to wavelengths between 390 and 500 nm, can penetrate deep layers of skin tissues. This deep penetration produces a direct attack on cell mitochondria, generating ROS, which produce harmful biological effects on skin cells. Some studies have shown that visible light can induce cell dysfunction and death both in vitro and in vivo, by accumulation of reactive oxygen species, such as the hydroxyl radical, superoxide anion and oxygen singlet, damage that occurs when light Blue excites cell photosensitizers. Thus, although the mechanisms are not fully understood, it seems that visible radiation is also involved in the photoaging of the skin, as well as in processes of hyperpigmentation and degradation of the extracellular matrix. In addition, the incidence of this type of radiation It is due not only to sunlight, but also emitted from artificial light sources, such as blue LED devices.
    [0024]
    [0025] Therefore, in addition to sunscreens to block the effects of UV radiation or molecules that help repair damaged DNA due to UV radiation, new strategies are needed to block the damage caused by other harmful radiation from the solar spectrum or from from artificial light sources to which humans and animals are exposed.
    [0026]
    [0027] Some publications describe the prevention of damage caused by ARI through the use of antioxidant combinations (EP2233127) or through the use of a lysate of Bifidobacterium longum (ES2629910). However, no effective products have been found to prevent skin photo-aging induced by visible high energy light.
    [0028]
    [0029] Thus, there is a need to develop effective strategies to prevent photoaging of the skin more completely, in particular, to prevent or delay, skin signs of photoaging directly induced by visible high energy light. In this same area, and beyond the preventive aspect, it is very desirable to develop strategies to repair the damage caused to the skin by visible high-energy radiation.
    [0030]
    [0031] EXPLANATION OF THE INVENTION
    [0032]
    [0033] The inventors have found that, surprisingly, a lysate of microorganisms of the genus Bifidobacterium, and in particular Bifidobacterium longum, protects against damage caused to the skin by exposure to high energy visible light. This effect is due to the fact that the lysate is able to significantly reduce the generation of ROS due to the effect of visible high energy light. This translates into efficient protection of the visible signs of damage caused to skin cells when applied before exposure to this radiation, that is, it prevents against photoaging directly caused by visible high-energy light.
    [0034] Unexpectedly it has also been found that the lysate of Bifidobacterium longum tested not only protects against visible high-energy radiation, but also exerts a reparative effect on the damage caused by exposure to it. This effect is due to the fact that the lysate reduces the amount of reactive oxygen species that have been generated by the exposure of the skin to the radiation.
    [0035]
    [0036] Therefore, in a first aspect, the invention provides a lysate of at least one microorganism of the genus Bifidobacterium, particularly Bifidobacterium longum, in an amount effective for use in the prevention and / or treatment of skin damage caused by the effect of reactive oxygen species generated by the effect of visible high energy radiation. More particularly, the damage in the skin cells is photoaging induced or produced by high energy visible radiation.
    [0037]
    [0038] This aspect can be reformulated as the use of a lysate of at least one microorganism of the genus Bifidobacterium, in particular of Bifidobacterium longum, in an effective amount for the preparation of a medicament for the prevention and / or treatment of skin damage caused by the effect. of reactive oxygen species generated by the effect of visible high energy radiation. The invention also relates to a method for the prevention and / or treatment of skin damage caused by the effect of reactive oxygen species generated by the effect of visible high energy radiation on a subject in need that comprises administering an effective amount. of a lysate of at least one microorganism of the genus Bifidobacterium, in particular of Bifidobacterium longum. In particular, the damage to the skin cells is caused by prolonged exposure to visible high energy radiation.
    [0039]
    [0040] The specific lysate of Bifidobacterium longum referred to in the present invention is sold under the name Repair Complex CLR by the company K. Richter GmbH. Complex repair refers to a lysate registered under the INCI name: Lysate Bifidat ferment under the name EINECS: Bifidobacterium longum, under the numbers EINECS: 306-168-4 and under the CAS number: 96507-89-0. It is an extract of Bifidobacteria, which contains products of metabolites, cytoplasm fractions, cell wall constituents, as well as polysaccharide complexes. For Sale preserved in paraben (Repair Complex) or paraben free (Repair Complex PF, RCPF).
    [0041]
    [0042] For the purposes of the invention, a "lysate" conventionally represents a material obtained after the destruction or dissolution of biological cells through a phenomenon known as cell lysis, thereby resulting in the release of the intracellular biological constituents contained in natural form in the cells of the microorganism under consideration.
    [0043]
    [0044] For the purposes of the present invention, the term "lysate" is used without preference to indicate the complete lysate obtained through the lysis of the microorganism under consideration or only a fraction thereof. Thus, the lysate is formed wholly or partially from intracellular biological constituents and from the constituents of cell walls and membranes. This cell lysis can be achieved by various techniques, such as an osmotic shock, a thermal shock, by ultrasound application, or alternatively with mechanical centrifugal stress. More particularly, this cell lysate can be obtained in accordance with the technology described in US4464362, and EP43128. The microorganism of the species of Bifidobacterium of the type considered can be grown anaerobically in a suitable culture medium, for example in accordance with the conditions described in US4464362, and EP43128. When the stationary phase is reached, the culture medium can be inactivated by pasteurization, for example, at a temperature of 60 to 65 ° C for 30 min. The microorganisms are then collected by a conventional separation technique, for example membrane filtration, centrifugation and resuspension in a buffer solution or in sterile physiological NaCl solution. The lysate can be obtained by physical means, for example, ultrasonic disintegration or by French press, in order to release its cytoplasmic fractions, cell wall fragments and the products resulting from metabolism. Enzymes, for example, lysozyme, can also be used for cell membrane breakdown. Next, all components in their natural distribution are stabilized in a weakly acidic aqueous solution. In this way, a lysate is generally obtained having a concentration of the order of 0.1 to 50%, in particular 1 to 20 %, and in particular about 5%, by weight of active substances with respect to their total weight.
    [0045]
    [0046] The lysate can be used in various ways, in the form of a solution or in powder form, and could be lyophilized.
    [0047]
    [0048] The microorganism that belongs to the genus Bifidobacterium is selected from among the species: Bifidobacterium longum, Bifidobacterium bifidum, Bifidobacterium breve , Bifidobacterium animalis, Bifidobacterium lactis, Bifidobacterium infantis, Bifidobacterium adolescentis, Bifidobacterium pseudocatemium, and mixtures. The preferred species is Bifidobacterium longum, which turns out to be particularly suitable for the purposes of the invention.
    [0049]
    [0050] The product marketed under the name Repair Complex CLR® is included in the context of the disclosure. According to the information of the supplier, Repair Complex CLR PF is obtained by fermentation of the species Bifidobacterium longum. After growth is complete, the bacteria disintegrate by ultrasound thereby releasing the cytoplasmic fractions and cell wall constituents. After cell disintegration, no particular fraction is isolated, thereby ensuring the presence of all constituents in their natural distribution in the Repair Complex CLR PF product. For the purposes of the present invention both affordable products can be used, the one marketed in CLR form preserved in parabens or the one marketed free of parabens.
    [0051]
    [0052] According to the invention, the lysate of Bifidobacterium , in particular of Bifidobacterium longum, is for use in the prevention and / or treatment of skin damage caused by the effect of reactive oxygen species generated by the effect of visible high energy radiation. More particularly, the damage in the skin cells is photoaging.
    [0053]
    [0054] For the purposes of the present disclosure, the term "photoaging" refers to the effects of exposure of UV ultraviolet light on the skin and / or exposure of IR light on the skin associated with the formation of thick wrinkles, irregular pigmentation of the skin, loss of skin elasticity, impaired skin barrier functions, or a combination thereof. Thus, cutaneous signs of photoaging include changes in pigmentation (mottled pigmentation), yellowing, deep wrinkles, dryness, telangiectasia, premalignant lesions, laxity, atrophy, leather appearance, elastosis (a coarse, yellow, stoned effect of the skin), or actinic purpura (easy bruising related to fragility of the vascular wall in the dermis).
    [0055]
    [0056] For the purposes of the present disclosure, the term "effective amount" means an amount that is sufficient to obtain the expected effect.
    [0057]
    [0058] A particular embodiment of the lysate is for the prevention of skin damage caused by the effect of reactive oxygen species generated by the effect of visible high energy radiation and said prevention comprises reducing the generation of ROS as a result of exposure to the radiation. For the purposes of this disclosure, the term "prevent" means reducing the risk of manifestation of a phenomenon. To obtain this preventive effect, the lysate is applied, preferably topically on the skin, before exposing the skin to visible high energy radiation. Thus, a particular embodiment refers to the lysate of the invention for the prevention of skin damage caused by the effect of reactive oxygen species generated by the effect of visible high energy radiation, in particular by reducing the generation of ROS, where prevention comprises the topical administration of an effective amount of the lysate before exposure to radiation.
    [0059]
    [0060] The preventive effect of the lysate according to the invention is shown in examples 1 and 2 (Figures 1-4), where it is shown that supplementation with Repair Complex in human fibroblasts and keratinocytes for 24 hours before subjecting the cells to visible high radiation Energy significantly reduced the generation of ROS in these skin cells compared to the untreated control.
    [0061]
    [0062] In another embodiment, the lysate is for the treatment of skin damage induced or caused by visible high energy radiation. For the purposes of the present disclosure the term "treat" means to compensate for a physiological dysfunction and more generally reducing or even eliminating an undesirable disorder, the manifestation of which is especially a consequence of this dysfunction. In this case, the treatment comprises reducing the amount of ROS generated by the visible high energy radiation. In other embodiments of the invention the lysate is, in addition to for use in the treatment of skin damage induced or caused by high energy visible radiation, also for use in the treatment of skin damage induced or produced by radiation. Infrared A, in particular the treatment also includes reducing the amount of ROS generated by Infrared A radiation.
    [0063]
    [0064] To obtain this treatment or repair effect, the lysate is preferably applied topically after exposure to the radiation, that is, when the ROS have already been generated by the effect of radiation. Therefore, in one embodiment, the treatment comprises the topical administration of an effective amount of the lysate after radiation exposure.
    [0065]
    [0066] This treatment effect (or repair effect) is evidenced in Example 3, Figures 5-7. These examples show experimental data obtained from human skin explants, in which the application of the 1% Repair Complex after the explants had been irradiated for 50 min resulted in a significant reduction in ROS levels compared to explants. untreated irradiated. A substantial improvement in the metabolic capacity of skin cells and a reduction in cell cytotoxicity were also observed in treated explants.
    [0067]
    [0068] In general, the invention preferably relates to skin damage induced or caused by radiation comprising photoaging. In other words, the invention preferably relates to photo-aging induced by ROSs generated by exposure to high energy visible radiation. In particular embodiments, skin damage induced or caused by radiation also includes photo aging caused by Infrared radiation A.
    [0069]
    [0070] The lysate of Bifidobacterium, in particular of Bifidobacterium longum, can be formulate in a composition according to the present invention in an amount of 0.1-10% by weight based on the total weight of the composition. Preferably, in an amount of 0.5-5% by weight based on the total weight of the composition. More preferably, in an amount of 0.5-4% by weight based on the total weight of the composition. Even more preferably, in an amount of 0.5-3% by weight based on the total weight of the composition. Even more preferably, in an amount of 0.5-1% by weight based on the total weight of the composition.
    [0071]
    [0072] All percentages mentioned herein are percentages by weight (weight / weight) unless otherwise indicated.
    [0073]
    [0074] The lysate for use according to the invention can be combined with physical, chemical, organomineral, and / or biological filters in order to provide a broad-spectrum photoprotective effect that prevents damage from visible high-energy radiation, allowing exposure to sunlight or devices that emit this type of radiation with less risk. Alternatively, the lysate can be combined with other agents that help repair radiation damage or alleviate its symptoms, such as moisturizing, soothing, anti-radical, antioxidant agents. Sometimes it may be convenient to combine the lysate with a mixture of filters, repair agents and soothing agents.
    [0075]
    [0076] A second aspect of the invention provides a lysate of at least one microorganism of the genus Bifidobacterium, in particular of Bifidobacterium longum, in an amount effective for the prevention and / or treatment of skin damage caused by the effect of reactive oxygen species generated by effect of visible high energy radiation, in combination with at least one filter against solar radiation selected from the group consisting of physical, chemical, organomineral, biological filters and combinations thereof.
    [0077]
    [0078] This aspect can be reformulated as the use of a lysate of at least one microorganism of the genus Bifidobacterium, in particular of Bifidobacterium longum, in an effective amount for the preparation of a medicament for the prevention and / or treatment of skin damage caused by the effect of reactive species of oxygen generated by the effect of visible high energy radiation, in combination with at least one filter against solar radiation selected from the group consisting of physical, chemical, organomineral, biological filters and combinations thereof. The invention also describes a method for the prevention and / or treatment of skin damage caused by the effect of reactive oxygen species generated by the effect of visible high energy radiation on a subject in need that comprises administering an effective amount of a lysate of at least one microorganism of the genus Bifidobacterium, in particular of Bifidobacterium longum, in combination with at least one solar radiation filter selected from the group consisting of physical, chemical, organomineral, biological filters and combinations thereof.
    [0079]
    [0080] By applying these combinations, complete protection against the cumulative damaging effects of sun exposure is achieved, delaying the skin aging process caused by visible high energy light. The combinations also more effectively counteract the damage caused by this type of radiation, improving the repair effect. All this imparts to these combinations a high practical value from the cosmetic point of view.
    [0081]
    [0082] Therefore, the invention also provides in a third aspect a composition, preferably a topical cosmetic composition, comprising an effective amount of a lysate of at least one microorganism of the genus Bifidobacterium, preferably Bifidobacterium longum, and at least one filter against solar radiation selected from the group consisting of physical, chemical, organomineral, biological filters and combinations thereof, together with cosmetically acceptable excipients or vehicles suitable for topical application on a person's skin, for use as defined in the first aspect of the invention.
    [0083]
    [0084] This aspect can also be expressed as a composition, preferably a topical cosmetic composition, as defined above, for the preparation of a medicament for the use defined according to the first aspect of the invention. The description also provides a method for the prevention and / or treatment of skin damage caused by the effect of reactive oxygen species generated by the effect of visible high energy radiation on a subject who needs it, which comprises administering an effective amount of the composition, preferably topical cosmetic composition, as defined above.
    [0085]
    [0086] The presence in the composition of the invention of these filters against solar radiation is particularly suitable when the composition is for use in the prevention of skin damage caused by the effect of reactive oxygen species generated by the effect of visible high radiation energy and said prevention include reducing the generation of ROS as a result of exposure to high energy visible radiation, UV radiation and infrared radiation A. In this way a broad spectrum photoprotector is achieved. Preferably the skin damages referred to are those related to photoaging.
    [0087]
    [0088] The amounts of the lysate in the compositions or combinations of the invention are those described above.
    [0089]
    [0090] In general, the filter is preferably selected from the group consisting of physical, chemical, organomineral, biological, and combinations thereof, for example in an amount of 0.1-50% by weight of the total composition, in particularly in an amount of 5-50%, or 10-50%, or 15-50%, or 10-35%, or 15-35%, or 29-35% by weight of the total composition, preferably in an amount of 29-50% by weight of the total composition, together with cosmetically acceptable excipients or vehicles suitable for topical application to a person's skin.
    [0091]
    [0092] Organic filters, for example, can be selected from those approved by the Council of the European Communities (revised text of the consolidated version of European Directive 76/768 / EEC, Annex VII pages 121-126, published on 04-24-2008 ). Inorganic filters can be selected from a group that includes: metal oxides such as pigments, nanopigments, treated and untreated, such as titanium dioxide (amorphous or crystalline), zinc oxide, iron, zinc, zirconium or cerium.
    [0093] If present, the amount of organic and inorganic filters can vary from about 0.1% to 50%. Generally, the organic and inorganic filters in the cosmetic composition are in an amount of 5-50% by weight of the total composition, in particular in an amount of 10-50%, or 15-50%, or 29- 50%, or 10-35%, or 15-35%, by weight of the total composition, preferably 29-35% by weight of the total composition.
    [0094]
    [0095] In particular, filters suitable for use in the present invention are UVA or UVB filters.
    [0096]
    [0097] The term "UVA protector" means a chemical compound that blocks UV radiation at the wavelength of approximately 320,440 nm. Some examples of UV sunscreens are 2-hydroxy-4-methoxybenzophenone (oxybenzone, INCI name: benzophenone-3); 3,3 '- (1,4-phenylenedimethylene) bis [7,7-dimethyl-2-oxo-bicyclo- (2,2,1) hept-1-yl methanesulfonic acid] or its cosmetically acceptable salts (INCI name: acid terephthalidendial camphorsulfonic acid); 1- (4-tert-Butyl-phenyl) -3- (4-methoxyphenyl) propan-1,3-dione (INCI name: butyl methoxy dibenzoyl methane); phenol, 2- (2H-benzotriazol-2-yl) -4-methyl-6- (2methyl-3- (1,3,3,3-tetramethyl-1- (trimethylsilyl) oxy) -disiloxani) propyl) (name INCI: drometrizol trisiloxane); 2-hydroxy4-methoxybenzophenone-5-sulfonic acid or its sodium salt (INCI name: benzophenone-4 or sulisobenzone); 2,2’-methylene-bis-6- (2H-benzotriazol-2-yl) -4- (tetramethyl-butyl) -1,1,3,3-phenol (INCI name: methylene-bis-benzotriazolyl tetramethylbutylphenol); monosodium salt of 2-2’-bis- (1,4-phenylene) 1 H -benzoimidazol-4,6-disulfonic acid (INCI name: phenyl dibenzoimidazole tetrasulfonate disodium); (1,3,5) -triazin-2,4-bis {[4- (2-ethyl-hexyloxy) -2-hydroxy] -phenyl) -6- (4-methoxyphenyl) (INCI name: bisethylhexyloxyphenyl methoxyphenyl triazine) ; or 2 - (- 4- (diethylamino) -2-hydroxybenzoyl) -benzoic acid hexyl ester (INCI name: hexyl diethylamino hydroxybenzoyl benzoate). If present, the UVA protector may vary from about 0.01% to 20%, preferably, from 1 to 10%, more preferably, from 2 to 5%, by weight of the total composition.
    [0098]
    [0099] As UVA filters, butyl methoxy dibenzoyl methane, methylene-bisbenzotriazolyl tetramethylbutylphenol, bis-ethylhexyloxyphenyl methoxy phenyl triazine, or combinations thereof can be used. Generally, the amount of butyl methoxy dibenzoyl Methane is between 1-5% by weight of the total composition. In addition, generally, the amounts of methylene-bis-benzotriazolyl tetramethylbutylphenol and bis-ethylhexyloxyphenyl methoxy phenyl triazine are between 1-10% by weight of the total composition.
    [0100]
    [0101] The term "UVB protector" means a chemical compound that blocks UV radiation at the wavelength of approximately 290 to 320 nm. There is a variety of UVB chemical sunscreens and it can be used for the purposes of the present invention. Some examples of UVB sunscreens are several esters of alpha-cyano-beta, beta-diphenyl acrylic acid such as 2-ethylhexyl 2-cyano-3,3-diphenylacrylate (INCI name: octocrylene). Other suitable UVB sunscreens are benzylidene camphor derivatives such as 3-4'-methylbenzylidene-d-1-camphor (INCI name: 4-methylbenzylidene camphor) or 3-benzylidenecamphor; cinnamate derivatives such as 2-ethylhexyl 4-methoxycinnamate (INCI name: ethylhexyl methoxycinnamate); Isopentyl 4-methoxycinnamate (INCI name: isoamyl p-methoxycinnamate); benzophenone derivatives such as oxybenzone or sulisobenzone, or sodium sulisobenzone; salicylate derivatives such as 2-ethyl salicylate (INCI name: ethylhexyl salicylate), 3,3,5-trimethylcyclohexyl salicylate (INCI name; homosalate); various aminobenzoic acid derivatives such as p-aminobenzoic acid (INCI name: PABA), ethyl 2-hexyl 4-dimethyl amino benzoate (INCI name: ethylhexyl dimethyl PABA), 4,4 - ((6 ( ((1,1-dimethylethyl) amino) carbonyl) phenyl) amino) 1,3,5-triazin-2,4-di) diimino) bis-, bis (2-ethylhexyl) (INCI name: diethylhexyl butamide triazone); drometrizol trisiloxane; bis-ethylhexyloxyphenol methoxy phenyl triazine; or 2,4,6-trianilino-p- (carbo-2’ethylhexyl-1’oxy) -1,3,5-triazine (INCI name: ethylhexyl triazone). If present, the UVB protector may vary from about 0.01% to 20%, preferably, from 1 to 10%, more preferably, from 2 to 5%, by weight of the total composition.
    [0102]
    [0103] Preferred UVB protectors are: octocrylene, ethylhexyl methoxycinnamate, diethylhexyl butamido triazone or combinations thereof. Generally, the content of octocrylene is between 1 and 10% by weight of the total composition, preferably 6-10% by weight of the total composition. Generally, the amount of ethylhexyl methoxycinnamate is between 5-10% by weight of the total composition. Generally, the amount of diethylhexyl butamide triazone is between 1-10% by weight of the total composition.
    [0104]
    [0105] Preferred protectors according to the invention are diethylhexyl butamido triazone, octocrylene, ethylhexyl methoxycinnamate, butyl methoxy dibenzoylmethane, diethylamino hydroxybenzoyl benzoate, or ethylhexyl salicylate.
    [0106]
    [0107] The compositions used for the purposes of the invention can be formulated to have certain sun protection factor (SPF) values ranging from about 1-100, in particular 2-80, or 2-45, or 5-30, or 10-70, or 15-70, or 15 35. The SPF indicates the increase in time that the skin can be exposed to the sun without suffering adverse effects: redness, erythema and burns. The calculation of SPF values is well known in the art.
    [0108]
    [0109] If present, a preferred physical protector according to the invention is micronized titanium dioxide or nano size and / or micronized zinc oxide or nano size.
    [0110]
    [0111] If present, preferred organomineral filters include methylene-bisbenzotriazolyl tetramethylbutylphenol (tinosorb® m) or bis-ethylhexyloxyphenol methoxyphenyl triazine (tinosorb® s).
    [0112]
    [0113] If present, the preferred biological protectors are substances with antioxidant activity such as vitamins (A, C or E), flavonoids (chelators of Fe) and other trace elements (enzymatic activity). The most preferred biological filters are vitamins (A, C, or E).
    [0114]
    [0115] In a particular embodiment, the compositions used for the purposes of the invention comprise the lysate of Bifidobacterium longum in combination with at least one chemical filter and at least one organomineral filter. In another particular embodiment, said composition further comprises at least one physical filter. Preferably, the chemical and organomineral filters are selected from those mentioned above.
    [0116] In another particular embodiment, the compositions used for the purposes of the invention comprise the lysate of Bifidobacterium longum in combination with at least one chemical filter and at least one physical filter. Preferably, the chemical and physical filters are selected from those mentioned above.
    [0117]
    [0118] In a preferred embodiment, the composition comprises the lysate of Bifidobacterium longum in combination with ethylhexyl methoxycinnamate, octocrylene, butyl methoxydibenzoylmethane, diethylhexyl butamido triazone, and bisethylhexyloxyphenyl methoxyphenyl triazine.
    [0119]
    [0120] The compositions used for the purposes of the invention further comprise excipients or vehicles suitable for topical application to a person's skin. Among these excipients or vehicles, the following are preferred: moisturizing agents such as lychee extract; emollient agents; antioxidant agents such as argan extract, vitamin E acetate, or green tea oil; revitalizing agents such as rice protein, SPF elevators such as wheat protein derivatives or amino acids with skin compatibility; water resistant polymers; preservatives; emulsifiers; volatile silicones; gelling agents such as xanthan gum or sclerotium gum; perfumes; or dyes.
    [0121]
    [0122] The invention also contemplates that the compositions of the invention comprise, in addition or alternatively to the sunscreen, an additional active compound, for example, a repairing, moisturizing, antiradical, antioxidant, calming, anti-inflammatory, antibiotic or combinations thereof. The presence of these additional agents in the composition may be beneficial in any case, but it is particularly suitable when the composition is for use in the treatment of skin damage caused by the effect of reactive oxygen species generated by the effect of visible radiation. of high energy and such prevention comprises reducing the amount of ROS generated as a result of exposure to high energy visible radiation, and also of UV radiation and infrared radiation A.
    [0123] Therefore, the invention also contemplates a composition, preferably a topical cosmetic composition, comprising an effective amount of a lysate of at least one microorganism of the genus Bifidobacterium, preferably Bifidobacterium longum, and at least one additional active compound, selected from a repairing agent , moisturizing, antiradical, antioxidant, calming, anti-inflammatory, and combinations thereof, together with cosmetically acceptable excipients or vehicles suitable for topical application on a person's skin, for use as defined in the first aspect of the invention, where the additional compound is in an amount of 0.1-50% by weight with respect to the weight of the total composition. Preferably these compositions are for use in the treatment of skin damage caused by the effect of reactive oxygen species generated by the effect of visible high energy radiation and / or Infrared A radiation.
    [0124]
    [0125] In this sense, non-limiting examples of the additional active compounds are: Allantoin, Bisabolol, panthenol, vitamins (pe niacinamide, vitamin C and its derivatives), plant extracts (marigold, Licorice, Mimosa, green tea, Centella Asiatica, Rosehip, algae and microalgae extracts), enzymes, peptides (hexapeptide-3, Tripeptide-38, hexapeptide-9), hyaluronic acid, ectoin, urea, cholesterol, hydrolyzed milk proteins, soybeans, wheat, silk, oligosaccharides such as beta glucans and alpha glucans, vegetable oils (eg Borage, Chamomile, Jojoba), essential oils (eg chamomile), polyunsaturated fatty acids (eg omega 3, omega 6, omega 9), silanoles (derived from organic silicon and derivatives), ceramides.
    [0126]
    [0127] The appropriate amounts of lysate in the composition, as well as the excipients or vehicles suitable for topical application are as described in previous embodiments.
    [0128]
    [0129] The cosmetic compositions used for the purposes of the invention may be in the form of an emulsion, cream, milk, lotion, ointment, solid bar, foam, spray, oil, ointment and fluid, among others. They can be in anhydrous form, in an aqueous solution, in the form of a suspension, or in the form of a water-in-oil or oil-in-water emulsion or water in silicone or water in silicone oils or water emulsion. in powdery form. In general, any composition used for the purposes of the invention can be applied to the skin, in any part of the skin, in any part of the body.
    [0130]
    [0131] Finally, the present application also describes a lysate of at least one microorganism of the genus Bifidobacterium, in particular of Bifidobacterium longum, in an amount effective to inhibit the production of MMP-1 induced by visible high energy radiation. This can also be expressed as the use of a lysate of at least one microorganism of the genus Bifidobacterium, in particular of Bifidobacterium longum, for the preparation of a medicament to inhibit the production of MMP-1 induced by high energy visible radiation. It can also be expressed as a method to inhibit the production of MMP-1 induced by high energy visible radiation, which comprises administering an effective amount of a lysate of at least one microorganism of the genus Bifidobacterium, in particular of Bifidobacterium longum, together with vehicles or cosmetically acceptable excipients, to a subject in need.
    [0132]
    [0133] Throughout the description and the claims the word "comprises" and its variants are not intended to exclude other technical characteristics, additives, components or steps. For those skilled in the art, other objects, advantages and features of the invention will be derived partly from the description and partly from the practice of the invention. The following examples and drawings are provided by way of illustration, and are not intended to be limiting of the present invention. In addition, the present invention covers all possible combinations of particular and preferred embodiments indicated herein.
    [0134]
    [0135] BRIEF DESCRIPTION OF THE DRAWINGS
    [0136]
    [0137] FIG. 1 Generation of ROS in human keratinocytes (HaCaT) for control without irradiation (C) and samples irradiated 60 minutes with HEV (C + HEV), in relation to control without irradiation. **** Represents the statistical significance with the value of p <0.0001.
    [0138]
    [0139] FIG. 2 ROS generation in human keratinocytes (HaCaT) for control without irradiation (C), samples irradiated 60 minutes with HEV (C + HEV) and samples irradiated 60 minutes with HEV after being supplemented with Repair Complex at
    [0140] 2.5% (A) and 5% (B). Standardized samples with respect to the HEV Control group. ** Represents statistical significance with p-value <0.01. *** Represents the statistical significance with the value of p <0.001. **** Represents the statistical significance with p value <0.0001.
    [0141]
    [0142] FIG. 3 ROS generation in human fibroblasts (NHDF) for control without irradiation (C) and samples irradiated 60 minutes with HEV (C + HEV), in relation to control without irradiation. **** Represents the statistical significance with the value of p <0.0001.
    [0143]
    [0144] FIG. 4 ROS generation in human fibroblasts (NHDF) for control without irradiation (C), samples irradiated 60 minutes with HEV (C + HEV) and samples irradiated 60 minutes with HEV after being supplemented with Repair Complex at
    [0145] 1.5% (D) and 2.5% (F). Standardized samples with respect to the HEV Control group. ** Represents statistical significance with p-value <0.01. *** Represents the statistical significance with the value of p <0.001. **** Represents the statistical significance with p value <0.0001.
    [0146]
    [0147] FIG. 5 Percentage of LDH enzyme compared to the Healthy Control group (C, considered 100%). A, skin explants irradiated for 24 h. B, explants irradiated for 24 hours and treated with 1% Repair Complex.
    [0148]
    [0149] FIG. 6 Percentage of resazurin with respect to the Healthy Control group (C, considered as 100%). A, skin explants irradiated for 24 h. B, explants irradiated for 24 hours and treated with 1% Repair Complex.
    [0150]
    [0151] FIG. 7 Percentage of ROS positive cells. C, sound control. A, skin explants irradiated for 24 h. B, explants irradiated for 24 hours and treated with 1% Repair Complex.
    [0152] EXAMPLES
    [0153]
    [0154] In the examples the product Repair Complex CLR PF (paraben free) was used, although it is noted that the presence or absence of parabens does not affect the results in question. The examples will refer to this product simply as Repair complex.
    [0155]
    [0156] Example 1. Determination of the protective capacity against visible light of high energy of the Repair Complex in human keratinocytes (HaCaT) in vitro
    [0157]
    [0158] OBJECTIVE: Analysis of the effects of the repair complex to counteract oxidative stress induced by visible high energy light (HEV) in human keratinocytes (HaCaT), through the quantification of reactive oxygen species (ROS).
    [0159]
    [0160] REAGENTS: Distilled water (Braun), DMEM (Dulbecco's Modified Eagles Medium), FBS (fetal bovine serum) chelated, penicillin, streptomycin, buffered saline phosphate solution (Sigma), trypan blue solution (Bio-Rad), trypsin (Sigma), ethanol (Sigma Aldrich), ROS detection kit (Sigma Aldrich). The immortalized HaCaT keratinocyte cell line was obtained as usual in the field (Wilson, "Growth and Differentiation of HaCaT Keratinocytes", Methods Mol Biol. 2014; 1195: 33-41).
    [0161]
    [0162] METHODOLOGY: HaCaT keratinocytic cells were cultured in the presence of different concentrations (2.5% and 5%) of Repair Complex for 24 hours. After treatment, the cells were exposed to high energy visible light (HEV) to induce the generation of ROS. The cells were exposed to HEV for 60 minutes (1-2 J / cm) and the effects were evaluated. ROS concentration was measured by spectrofluorimetry. The Repair Complex was used at concentrations of 2.5% and 5%.
    [0163]
    [0164] Live cells were counted in a Bürker chamber under the microscope. HaCaT cells were grown overnight at a density of 10,000 cells / well in a 96-well black plate, in growth medium. 24 hours later, the culture medium was removed and replaced with a new culture medium supplemented with 2.5% and 5% Repair Complex. After 24 hours of incubation period, the culture medium of all wells was replaced by PBS and ROS master mix in all cultured wells, including 2-3 control wells without cells to obtain blank control. The cells were exposed to HEV for 60 minutes (theoretical lamp power = 66.990 mW / m2, total dose received) around 1-2 J / cm2 at a wavelength between 420-500 nm, peak at 420 nm). The non-irradiated controls were incubated at 26 ° C during this time in the dark. One hour after the irradiation period, the ROS levels were measured in all samples. ROS generated and accumulated inside the cells reacted with a fluorogenic sensor located in the cytoplasm, which resulted in an amount of a fluorometric product proportional to the amount of ROS generated in the cell. Fluorescence was measured at Aex = 490 / Aem = 525. Three biological replicates were made with eight technical repeats for each concentration of 2.5% and 5%.
    [0165]
    [0166] The blank control mean was subtracted from all sample data and, after that, all data was normalized to the irradiated control and represented as mean ± SEM. Data were analyzed comparing statistically the control treated against irradiated, and irradiated versus non-irradiated samples. The Student's t-test was applied for the analysis. Statistical significance was established at p <0.05, 95% confidence. Data is represented in two forms of normalization. In the first normalization process, the "untreated control" was used as the reference "control" and these data define the efficacy of the irradiation treatment in the generation of ROS. In the second case, the "irradiated control" was used as a reference control to determine the effectiveness of the Repair Complex to avoid the generation of ROS. The formula used to analyze the raw data and to obtain the data used in the graphs was "X / M", where X is the gross value of the absorbance and M is the average of the corresponding control for each biological replica.
    [0167]
    [0168] RESULTS: The results showed that HEV (blue light) radiation for 60 minutes significantly induced the generation of ROS in HaCaT cells at levels at 69.7 ± 6.9 times higher than the non-irradiated control (Figure 1). When HaCaT cells were treated with Repair Complex for 24 hours at 2.5% and 5%, the Results indicated that ROS levels generated by HEV were lower by 39.7 ± 11.5% and 47.0 ± 11.4%, respectively, compared to HEV Control (Figure 2).
    [0169]
    [0170] CONCLUSION: Supplementation with 2.5% and 5% Repair Complex in human keratinocytes (HaCaT) for 24 hours before subjecting the cells to HEV radiation significantly reduced ROS generation by 39.7 ± 11.5% and 47.0 ± 11.4%, compared With untreated control. This indicates that the Repair Complex exerts a potent protective effect on the damage induced in keratinocytes by the generation of ROS due to the effect of HEV radiation.
    [0171]
    [0172] Example 2. Determination of the protective capacity against visible light of high energy of the Repair Complex in human fibroblasts (NHDF) in vitro
    [0173]
    [0174] OBJECTIVE: Analysis of the effects of the repair complex to counteract oxidative stress induced by visible high energy light (HEV) in human fibroblasts (NHDF), through the quantification of reactive oxygen species (ROS).
    [0175]
    [0176] REAGENTS AND METHODOLOGY: the reagents used and the methodology were as described in example 1, except that fibroblast cells (NHDF primary fibroblast cell line (PromoCell) were used and the Repair Complex supplementation was done in concentrations of 1.5 and 2.5%.
    [0177]
    [0178] RESULTS: The results showed that HEV (blue light) radiation for 60 minutes significantly induced the generation of ROS in NHDF cells at levels 38.9 ± 3.6 times higher than the non-irradiated control (Figure 3). When NHDF cells were treated with Repair Complex for 24 hours at 1.5% and 2.5%, the results indicated that ROS levels generated by HEV were lower by 10.6 ± 3.8% and 14.0 ± 3.7, respectively, compared to HEV Control (Figure 4).
    [0179]
    [0180] CONCLUSION: Supplementation with 1.5% and 2.5% Repair Complex in human fibroblasts (NHDF) for 24 hours before subjecting cells to HEV radiation significantly reduced the generation of ROS by 10.6 ± 3.8% and 14.0 ± 3.7, respectively, compared to the untreated control. This indicates that the Repair Complex exerts a potent protective effect on the damage induced in the fibroblasts by the generation of ROS due to the effect of HEV radiation.
    [0181]
    [0182] Example 3. Repair effect of the Repair Complex on radiation damage in skin explants.
    [0183]
    [0184] OBJECTIVE: To assess the repair capacity of the Repair Complex on the harmful effects on the skin caused by exposure to solar radiation. The parameters analyzed in this study were performed in organotypic cultures of human skin explants.
    [0185]
    [0186] Human skin explants have great advantages over in vitro cell cultures, since the characteristic architecture of ex vivo tissue is maintained for at least 14 days, maintaining its three-dimensional structure and differentiated cell types. They are therefore a good replica of the fabric of origin. The explants do not lose cell heterogeneity, spatial organization, or systemic regulatory components as is the case in cell cultures in vitro.
    [0187]
    [0188] To assess the repair capacity of the Repair Complex for damage caused by the effect of high-energy solar radiation and infrared A, the study was carried out on irradiated human skin explants (Solar Simulator), where species production was induced oxygen reagents (ROS). In this regard, it should be noted that the skin damage caused by ROS generated by the effect of exposure to sunlight is directly related to visible high-energy light and infrared A, but not to UV radiation. This is evidenced in, P. Schroeder et al., "Cellular response to infrared radiation involves retrograde mitochondrial signaling", Free Radie. Biol. Med. 2007, vol. 43, pp. 128-135, where it is explained that the generation of ROS by mitochondria is not affected by UVA or UVB radiation Several studies indicate that the main inducers of ROS generation and its accumulation in cells are visible high energy radiation and the increase in intracellular temperature caused by radiation infrared A.
    [0189]
    [0190] REAGENTS: Dulbecco's Modified Eagle's Medium (Sigma D-5546), Penicillin / streptomycin (Gibco 15140-122), Hank’s Salts (Sigma H-2387), PBS (10x). (Roche 11 666 789001), 10 mm diameter discs, Resazurin (Sigma R7017), CytoTox 96 (Promega G1780), DFFDA (Invitrogen C-13293), Phospho-Histone H2A.X pSer140 Antibody (3F2) (ThermoFisher MA1- 2022), Goat anti-Mouse IgG (H + L) Secondary Antibody, Alexa Fluor® 488 conjugate (ThermoFisher A-11001), Collagenase Type I (Invitrogen 17018-029) and abdominal human skin implants.
    [0191]
    [0192] The explants were obtained as follows: pieces of skin of the abdomen were obtained with the informed consent of healthy women between 40 and 55 years undergoing plastic surgery. Up to 2 h after surgery, the skin was cut into 0.8 cm 2 pieces and samples were placed with the dermis down and the epidermis up on culture plates containing Dulbecco's modified Eagle's medium (DMEM) with antibiotics (penicillin / streptomycin 1%). Characteristics of skin samples: Female, Caucasian, Abdomen, 40-55 years, Phenotype II or III. Skin samples were kept in culture medium at 37 ° C under 5% CO 2 . The cellular medium was replaced every two business days. Similarly, the resazurin and lactate dehydrogenase assays were performed every two business days to determine the acceptable condition of the skin explants during the study.
    [0193]
    [0194] The product under study, Repair Complex CLR PF, was prepared on the same day of the 1% test in culture medium.
    [0195]
    [0196] METHODOLOGY: Both healthy and irradiated human skin explants were used as an experimental system. Damaged skin is obtained by exposure of healthy skin to solar radiation, UV / vis irradiation (295-780 nm) emitted by the SOL 500 simulator (Dr. Honle). A 10 J radiation was applied to the explants. The intensity used in the study was 3.2-3.6 mW / cm2, exposing the explants to said solar intensity for 50 minutes. The study groups were the following:
    [0197]
    [0198] 1. Healthy Control Group: 4 skin explants without being injured.
    [0199] 2. Irradiated Group: 4 skin explants subjected to solar radiation for 24 hours.
    [0200] 3. Irradiated treated group: 4 skin explants subjected to solar radiation for 24 hours and incubated artificially after irradiation with 1% Repair Complex for 24 h.
    [0201]
    [0202] A single experiment was carried out, where each experimental group presented 4 replications. The Repair Complex under study was tested at a concentration of 1%.
    [0203] The following parameters were evaluated: Cytotoxicity (lactate dehydrogenase, LDH) and Alamar Blue (resazurin) and ROS production.
    [0204]
    [0205] Resazurin Assay
    [0206]
    [0207] Resazurin dye (7-hydroxy-3H-phenoxazin-3-one 10-oxide) is widely used as an indicator of cell viability in proliferation and cytotoxicity assays. The assay is based on the fact that viable and metabolically active cells reduce resazurin to resorufin (fluorescent compound), which is released into the culture medium. This conversion is intracellular, facilitated by mitochondrial, microsomal and cytosolic oxidoreductases.
    [0208]
    [0209] Resazurin is a non-toxic and stable compound in the cellular medium that, when reduced, is transformed into resorufin (fluorescent), a soluble and permeable compound that is released into the culture medium. Therefore, it allows continuous measurements of cell proliferation and metabolism without the loss of the experimental system.
    [0210]
    [0211] The skin explants of the control, irradiated and irradiated treated groups were incubated with 6 pM resazurin for at least 1 hour. Subsequently, 100 pl of supernatant was taken from each sample and transferred to a 96-well plate. The resorufin produced was measured by fluorimetry at an excitation wavelength of 530-560 nm and an emission wavelength of 590 nm.
    [0212]
    [0213] LDH test
    [0214]
    [0215] The LDH cytotixicity test is a colorimetric test where the Lactate dehydrogenase (LDH) enzyme, LDH is a stable cytosolic enzyme that is emitted to the culture medium when the cell membrane is damaged. An increase in cell death or cells with the damaged cytoplasmic membrane during the test would mean an increase in the LDH enzyme in the culture medium.
    [0216]
    [0217] 100 ql of the culture medium of each sample of the groups tested was transferred to a 96-well plate. The presence of LDH in the culture medium was detected by the following enzymatic reaction: LDH oxidizes lactate in pyruvate which reacts with the tetrazolium salt WST-1 to form formazan. The increase in the amount of formazan correlates directly with the number of lysed (damaged) cells. Formazan is a colored compound soluble in water and can be measured using an ELISA plate reader at 490-500 nm.
    [0218]
    [0219] Dermal tissue breakdown
    [0220]
    [0221] Disintegration of human skin explants from the groups tested was carried out in order to obtain a cell suspension of each explant. Briefly, the tissue was digested with 3.5 mg / ml collagenase in DMEM for 4 hours at 37 ° C and subsequently filtered. The cell filtrate was enriched by centrifugation (1700 rpm and 5 minutes) and the resulting pellet was resuspended in DMEM.
    [0222]
    [0223] ROS Determination
    [0224]
    [0225] Intracellular ROS levels were determined through the fluorescent probe, 2 ', 7'-difluorescein diacetate (DFFH-DA). This method is based on intracellular hydrolysis of DFFH-DA by intracellular esterases becoming a non-fluorescent derivative, DFFH. After entering the DFFH-DA compound in the cell, esterases hydrolyze it with the non-fluorescent derivative carboxy-2 ', 7'-dichlorodihydrofluorescein, which is retained inside the cell due to its negative charge. Oxidation of this compound by ROS present in the cell implies a conversion of this compound to carboxy-2 ', 7'-dichlorofluorescein, which is a fluorescent compound.
    [0226] The skin cell suspensions were washed with 300 pl phosphate buffer (PBS), with a minimum quantity of approximately 100,000 cells per sample. Subsequently, the cells were labeled with DFFH-DA at a final concentration of 10 pM and incubated for 30 minutes at 37 ° C and 5% CO2. After incubation, the cells were washed with PBS and analyzed by flow cytometry (Cytomics FC500-MCL).
    [0227]
    [0228] RESULTS:
    [0229]
    [0230] LDH test
    [0231]
    [0232] The results of this trial were calculated considering the Healthy Control group (skin explants that were not irradiated) as 100% LDH. As already indicated above, an increase in the amount of formazan in the culture medium correlates directly with an increase in lysed (damaged) cells.
    [0233]
    [0234] As shown in Figure 5, an increase in the amount of LDH with respect to the control group was observed in the experimental group with irradiation. The samples treated with Repair Complex at 1% after irradiation showed a marked decrease in the percentage of LDH, with these values approaching the healthy control group. These data suggest that the Repair Complex at 1% concentration in human skin explants has a reparative effect on plasma membrane damage (damaged or dead skin cells) generated by irradiation under the conditions of this test.
    [0235]
    [0236] Resazurin Assay
    [0237]
    [0238] To corroborate the results obtained in the LDH test, the resazurin test was carried out. The resazurin marker is widely used as an indicator of cell viability in numerous cytotoxic assays. But it is also an indicator of the metabolic activity of cells since this assay is based on the ability of metabolically active cells to reduce resazurin in resofurin and dihydroresorufin by oxidoreductases enzymes.
    [0239] The results are described in Figure 6. The results of this test were calculated considering the Healthy Control group (skin explants that were not irradiated) as 100% of the fluorescent compound derived from Resazurine. In the experimental group with irradiation a decrease in the amount of markedzinin was observed with respect to the control group. This indicates that irradiation affected the ability of the tissue to reduce resazurin, and this decrease in its reducing capacity is directly related to a decrease in the metabolic activity of the tissue (cells that make up the skin explant), and therefore with appearance of damage to the dermal tissue. The post-treated samples with 1% Repair Complex showed resazurin values very similar to the non-irradiated Control group.
    [0240]
    [0241] These data suggest that the 1% Repair Complex product in human skin explants has a reparative effect on the damage caused by irradiation, maintaining the metabolic activity (mitochondrial reducing capacity).
    [0242]
    [0243] ROS Determination
    [0244]
    [0245] The human skin samples (healthy control of unirradiated skin, irradiated skin for 24 h and irradiated skin and treated with 1% Repair Complex) were disintegrated with collagenase. Subsequently, the cell suspensions generated were incubated with DFFH-DA and the presence of ROS determined by flow cytometry. The results of ROS are described in Figure 7. The figure shows that the percentage of ROS-positive cells in the irradiated skin cell suspensions indicated an increase of this percentage with respect to the Healthy Control group. However, the ROS percentage values in the cell suspensions of the irradiated skin samples and subsequently treated with Repair Complex were very similar to the values observed in the Healthy Control group. This indicates that the compound under study at 1% has a restorative effect on the reactive oxygen species produced in explants of irradiated skin, under the conditions of the study.
    [0246] APPOINTMENT LIST
    [0247]
    [0248] Patent Bibliography
    [0249] EP43128
    [0250] EP2233127
    [0251] ES2629910
    [0252]
    [0253] Non-patent bibliography:
    [0254] P. Schroeder et al., "Cellular response to infrared radiation involves retrograde mitochondrial signaling", Free Radie. Biol. Med. 2007, vol. 43, pp. 128-135.
    [0255] Wilson, "Growth and Differentiation of HaCaT Keratinocytes", Methods Mol Biol.
    [0256] 2014; 1195: 33-41.
    权利要求:
    Claims (23)
    [1]
    1. Lysate of at least one microorganism of the genus Bifidobacterium in effective amount for use in the prevention and / or treatment of skin damage caused by the effect of reactive oxygen species (ROS) generated by the effect of visible high radiation Energy.
    [2]
    2. Listed for use according to claim 1, wherein the use is for prevention and where prevention comprises reducing the generation of ROS.
    [3]
    3. Listed for use according to any of claims 1-2, wherein the prevention comprises the topical administration of an effective amount of the lysate before exposure to radiation.
    [4]
    4. Listed for use according to claim 1, wherein the use is for the treatment of skin damage induced or caused by visible high energy radiation.
    [5]
    5. Listed for use according to claim 4, wherein the treatment comprises reducing the amount of ROS generated by the visible high energy radiation.
    [6]
    6. Listed for use according to any of claims 4-5, wherein the use further comprises the treatment of skin damage induced or caused by Infrared A radiation.
    [7]
    7. Listed for use according to claim 6, wherein the treatment comprises reducing the amount of ROS generated by the Infrared A radiation.
    [8]
    8. Listed for use according to any of claims 4-7, wherein the treatment comprises the topical administration of an effective amount of the lysate after radiation exposure.
    [9]
    9. Listed for use according to any of claims 1-8, wherein the skin damage induced or caused by radiation comprises the photoaging caused by visible high-energy radiation.
    [10]
    10. Listed for use according to claim 9, wherein the skin damage induced or caused by the radiation further comprises photo aging caused by the Infrared radiation.
    [11]
    11. Listed for use according to any of claims 1-10, wherein the lysate is at least one microorganism of the species Bifidobacterium longum.
    [12]
    12. Listed for use according to any of claims 1-11, wherein the effective amount of the lysate is 0.1 to 10% by weight with respect to the total weight of a composition comprising it.
    [13]
    13. Listed for use according to claim 12, wherein the effective amount of the lysate is 0.5 to 5% by weight with respect to the total weight of a composition comprising it.
    [14]
    14. Listed for use according to claim 13, wherein the effective amount of the lysate is 1 to 2.5% by weight with respect to the total weight of a composition comprising it.
    [15]
    15. Listed for use according to any of claims 1-14 which is for topical application.
    [16]
    16. Listed for use according to any one of claims 1-15, in combination with at least one solar radiation filter selected from the group consisting of physical, chemical, organomineral, biological filters and combinations thereof.
    [17]
    17. Listed for use according to any of claims 1-15, in combination with at least one solar radiation filter selected from the group consisting of physical, chemical, organomineral, biological and combinations thereof,
    where the use also includes the prevention and / or treatment of skin damage induced or produced by Infrared A and Ultraviolet radiation.
    [18]
    18. Topical cosmetic composition comprising an effective amount of a lysate of at least one microorganism of the genus Bifidobacterium and an effective amount of at least one solar radiation filter selected from the group consisting of physical, chemical, organomineral, biological filters and combinations thereof, together with cosmetically acceptable excipients or vehicles suitable for topical application on a person's skin, for use according to any of claims 1-10.
    [19]
    19. Topical cosmetic composition for use according to claim 18, wherein the filter is in an amount of 10 to 50% by weight with respect to the weight of the total composition.
    [20]
    20. Topical cosmetic composition for use according to any of claims 18-19, wherein the lysate is at least one microorganism of the species Bifidobacterium longum.
    [21]
    21. Cosmetic composition for use according to any of claims 19 20, wherein the effective amount of the lysate is 0.1 to 10% by weight with respect to the total weight of a cosmetic composition comprising it.
    [22]
    22. Cosmetic composition for use according to claim 21, wherein the effective amount of the lysate is 0.5 to 5% by weight with respect to the total weight of a cosmetic composition comprising it.
    [23]
    23. Cosmetic composition for use according to claim 22, wherein the effective amount of the lysate is 1 to 2.5% by weight with respect to the total weight of a cosmetic composition comprising it.
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    ES2725348B2|2020-04-22|
    引用文献:
    公开号 | 申请日 | 公开日 | 申请人 | 专利标题
    EP2033627A2|2007-09-04|2009-03-11|L'oreal|Use of a Bifidobacterium species lysate for treating sensitive skin|
    WO2009053564A2|2007-09-04|2009-04-30|L'oreal|Cosmetic use of a lysate of bifidobacterium species for reinforcing the skin barrier function|
    ES2586453T3|2008-10-28|2016-10-14|L'oreal|Use of a microorganism lysate for the treatment of oily skin|
    ES2629910T3|2011-02-11|2017-08-16|Laboratorios Cinfa, S.A.|Active ingredient obtained biotechnologically in cosmetic compositions useful to protect the skin from damage induced or caused by infrared radiation|
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