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US20220175638A1 - Cell activator - Google Patents

Cell activator Download PDF

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Publication number
US20220175638A1
US20220175638A1 US17/601,341 US202017601341A US2022175638A1 US 20220175638 A1 US20220175638 A1 US 20220175638A1 US 202017601341 A US202017601341 A US 202017601341A US 2022175638 A1 US2022175638 A1 US 2022175638A1
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US
United States
Prior art keywords
skin
vitamin
light
ultraviolet light
phycoerythrin
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Pending
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US17/601,341
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English (en)
Inventor
Kazuyuki Miyazawa
Renaud GILLET
Bianca MCCARTHY
Tetsuya Kanemaru
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Shiseido Co Ltd
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Shiseido Co Ltd
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Assigned to SHISEIDO COMPANY, LTD. reassignment SHISEIDO COMPANY, LTD. ASSIGNMENT OF ASSIGNORS INTEREST (SEE DOCUMENT FOR DETAILS). Assignors: GILLET, Renaud, KANEMARU, TETSUYA, MCCARTHY, Bianca, MIYAZAWA, KAZUYUKI
Publication of US20220175638A1 publication Critical patent/US20220175638A1/en
Pending legal-status Critical Current

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Definitions

  • the present invention relates to a cell activator comprising a wavelength conversion substance, to a composition and product comprising the cell activator, and to a method for activating skin using the same.
  • the harm to skin caused by ultraviolet light includes adverse effects such as skin cancer, photoaging, skin spots, wrinkles and inflammation, which are also undesirable from the viewpoint of health and beauty. While ultraviolet light is also utilized for sterilizing purposes, when considered in balance with the harm caused by ultraviolet light there is a greater need to defend against rather than to actively utilize ultraviolet light.
  • UV light Many measures are therefore being taken to protect the skin from ultraviolet light. Such measures include the use of sunscreens, the implementation of indoor spaces for avoidance of sunlight, and the use of head coverings and clothing treated to block UV rays and films designed to block UV rays.
  • the present inventors have conducted active research with the aim of allowing ultraviolet light to be effectively utilized on skin.
  • a cell activator has been devised that converts ultraviolet light wavelengths and activates skin cells.
  • a cell activator comprising a wavelength conversion substance as an active ingredient
  • the wavelength conversion substance converts the wavelength of ultraviolet light contained in incident light to emit emission light having a wavelength longer than the wavelength of the ultraviolet light.
  • the wavelength conversion substance comprises one or more phycobiliproteins selected from among allophycocyanin, C-phycocyanin, R-phycocyanin, phycoerythrocyanin, B-phycoerythrin, b-phycoerythrin, C-phycoerythrin and R-phycoerythrin; one or more inorganic phosphors selected from among zinc oxide phosphors, magnesium titanate phosphors and calcium phosphate phosphors; one or more components selected from among vitamin A, ⁇ -carotene, vitamin K, vitamin B1, vitamin B2, vitamin B6, vitamin B12, folic acid, niacin, lycopene, gardenia, safflower, turmeric, cochineal, perilla, red cabbage, flavonoids, carotenoids, quinoids, porphyrins, anthocyanins, polyphenols, salicylic acid and capsicum extract; and/or
  • the wavelength conversion substance comprises one or more phycobiliproteins selected from among allophycocyanin, C-phycocyanin, R-phycocyanin, phycoerythrocyanin, B-phycoerythrin, b-phycoerythrin, C-phycoerythrin and R-phycoerythrin; one or more inorganic phosphors selected from among zinc oxide phosphors, magnesium titanate phosphors and calcium phosphate phosphors; and/or one or more B vitamins selected from among vitamin B1, vitamin B2, vitamin B6 and vitamin B12.
  • composition comprising the cell activator according to any one of (1) to (5).
  • composition according to (6) wherein the composition is a skin external composition for activating skin by exposing skin to light containing ultraviolet light.
  • a cosmetic method for activating skin of a subject comprising:
  • composition according to (6) or (7) to skin of a subject
  • composition-applied skin with light containing ultraviolet light.
  • a cosmetic method for activating skin of a subject comprising:
  • the present invention can exhibit a desirable effect on skin by effectively making use of ultraviolet light to activate skin cells. Because it is conventionally undesirable for skin to be exposed to ultraviolet light, it has been common technical knowledge in the field to apply measures to maximally avoid exposing skin to ultraviolet light. It was highly surprising to find, then, that the present invention provides a desirable effect on skin by utilization of ultraviolet light.
  • the invention provides novel uses of the aforementioned compounds that have conventionally been used primarily as dyes, pigments, ultraviolet scattering agents, ultraviolet absorbers, nutrients and antioxidants. The invention also helps to improve quality of life by providing a more positive feeling for persons who have attempted to avoid ultraviolet light as much as possible for beauty or health reasons when outdoors.
  • FIG. 1 is a schematic diagram illustrating Experiments 1 and 2.
  • FIG. 2 shows cellular activity during irradiation of UV using different wavelength conversion substances in Experiment 1.
  • the ordinate represents relative fluorescence intensity (au).
  • FIG. 3 shows cellular activity during irradiation of strong UV using C-phycocyanin at different concentrations in Experiment 2, in terms of relative fluorescence intensity (au).
  • FIG. 4 is a schematic diagram illustrating Experiment 3.
  • FIG. 5 shows cellular activity during irradiation of UV using C-phycocyanin in cells with previously lowered cellular activity in Experiment 3, in terms of relative fluorescence intensity (au) (P test).
  • FIG. 6 shows cellular activity during irradiation of UV using a zinc oxide phosphor in cells with previously lowered cellular activity in Experiment 4, in terms of relative fluorescence intensity (au) (P test).
  • the cell activator of the invention comprises a wavelength conversion substance as an active ingredient.
  • a wavelength conversion substance is a substance that converts the wavelength of ultraviolet light contained in incident light and emits emission light having a wavelength longer than the wavelength of the ultraviolet light.
  • the ultraviolet light may include UVA, UVB and UVC. According to one embodiment, the ultraviolet light is light with a peak wavelength of 200 nm to 400 nm.
  • the ultraviolet light may also be included in incident light such as sunlight, for example. Alternatively, the incident light may be ultraviolet light, and artificially generated ultraviolet light may be used.
  • the emission light emitted by the wavelength conversion substance has a longer wavelength than ultraviolet light, with a peak wavelength of preferably 500 nm to 700 nm.
  • the emission light may have one or more peaks at 510 nm, 520 nm, 530 nm, 540 nm, 550 nm, 560 nm, 570 nm, 580 nm, 590 nm, 600 nm, 610 nm, 620 nm, 630 nm, 640 nm, 650 nm, 660 nm, 670 nm, 680 nm, 690 nm or 700 nm, or in any range within these values, though without being restrictive, or it may be red light, orange light, green light or blue light.
  • the wavelength conversion substance has its main wavelength at 500 nm to 700 nm, for light emitted upon excitation with excitation light of 200 nm to 400 nm.
  • wavelength conversion substances include the following components: phycobiliproteins such as allophycocyanin, C-phycocyanin, R-phycocyanin, phycoerythrocyanin, B-phycoerythrin, b-phycoerythrin, C-phycoerythrin and R-phycoerythrin; natural or synthetic components such as vitamin A, n-carotene, vitamin K, vitamin B1, vitamin B2, vitamin B6, vitamin B12, folic acid, niacin, lycopene, gardenia, safflower, turmeric, cochineal, perilla, red cabbage, flavonoids, carotenoids, quinoids, porphyrins, anthocyanins, polyphenols, salicylic acid and capsicum extract; dyes such as Red No.
  • phycobiliproteins such as allophycocyanin, C-phycocyanin, R-phycocyanin, phycoerythrocyanin, B-phycoerythrin, b
  • cerium and phosphorus and/or magnesium, and red phosphors comprising compounds obtained by europium activation of mixed crystals consisting of the alkaline earth metal sulfides described in Japanese Patent No. 6361416 combined with gallium compounds, the zinc oxide phosphors mentioned in International Patent Publication No. 2018/004006, the zinc oxide phosphors mentioned in Japanese Unexamined Patent Publication No. 2018-131422, and the inorganic phosphors mentioned in Japanese Unexamined Patent Publication HEI No. 5-117127.
  • the inorganic phosphor is one or more phosphors selected from among phosphors obtained by doping zinc oxides represented by ZnO: Zn, Zn 1+z , ZnO 1 ⁇ x with the sulfur-containing compounds mentioned in International Patent Publication No. 2018/004006, including sulfides and/or sulfates such as zinc sulfide or zinc sulfate, magnesium titanate phosphors obtained by doping magnesium titanates such as MgTiO 3 or Mg 2 TiO 4 with manganese, and calcium phosphate phosphors obtained by doping calcium phosphates such as Ca(H 2 PO 4 ) 2 , CaHPO 4 or Ca 3 (PO 4 ) 2 with cerium.
  • phosphors obtained by doping zinc oxides represented by ZnO: Zn, Zn 1+z , ZnO 1 ⁇ x with the sulfur-containing compounds mentioned in International Patent Publication No. 2018/004006, including sulfides and/or sulfates such as zinc
  • the wavelength conversion substance may be obtained by extraction from a natural source such as an animal, plant or algae, or it may be obtained by an artificial method such as chemical synthesis.
  • phycobiliproteins can be prepared by extraction from algae, including blue-green algae such as Spirulina ( Spirulina platensis ) or red algae such as porphyridiophylla ( Porphyridium purpureum ), by the method described in Japanese Patent No. 4048420, Japanese Patent No. 4677250 or Japanese Patent No. 3303942, for example.
  • Zinc oxide phosphors can be produced by the method described in International Patent Publication No. 2018/004006, Japanese Unexamined Patent Publication No. 2018-131422 or Japanese Unexamined Patent Publication HEI No.
  • Magnesium titanate phosphors can be produced by the method described in Japanese Unexamined Patent Publication No. 2017-88719.
  • Calcium phosphate phosphors can be produced by the method described in International Patent Publication No. 2018/117117.
  • these wavelength conversion substances may be composed of, or may include, the components mentioned above, and they may be single components alone or combinations of more than one of the components.
  • the aforementioned phycobiliproteins or inorganic material phosphors may be mixed with other wavelength conversion substances such as B vitamins (vitamin B1, vitamin B2, vitamin B6 or vitamin B12) to exhibit synergistic effects.
  • B vitamins vitamin B1, vitamin B2, vitamin B6 or vitamin B12
  • the wavelength conversion substance content in the cell activator, composition or product of the invention is not particularly restricted so long as the wavelength conversion effect of the invention is not impaired, and it may be appropriately determined for the type of wavelength conversion substance and the purpose of use of the cell activator or composition. It may be any content in the range of 0.01 to 99.99 wt % or 0.1% to 999 wt %, for example.
  • the cell activation is not restricted but may be accelerated metabolism or turnover of cells of an animal such as a human, such as skin fibroblasts and/or keratinocytes, or improvement of their function, or accelerated proliferation, inhibited oxidation, improved resistance against fatigue or external stimulation, suppressed reduction in function or activity, or cellular repair.
  • Activation of skin cells is expected to provide an effect of preventing or ameliorating wrinkles, blemishes, skin aging and photoaging.
  • Measurement of the cell activation effect may be carried out by measuring cell viability or proliferation using Alamar Blue, as in the Examples, or other methods such as dye assays, mitochondrial membrane potential-dependent dye assay, elastase cleavage dye assay, ATP and ADE assay, glycolysis flux and oxygen consumption assay.
  • any form of administration may be used for the cell activator and composition of the invention, but an external preparation for skin will often be preferred, such as a drug, quasi drug or cosmetic, for activating skin by exposing the skin to light containing ultraviolet light.
  • the dosage form, coating method and number of doses may be determined as desired. For example, it may be applied onto skin in the form of cosmetic water or a spray, oil, cream, latex, gel, sunscreen or suntan lotion either periodically or irregularly, once or several times per day at morning, noon or evening, or before going out or engaging in outdoor activities, marine sports or skiing, for example, when exposure to sunlight is expected.
  • the cell activator and composition of the invention may also be used in combination with an additive such as an excipient, preservative, thickener, binder, disintegrator, dispersing agent, stabilizer, gelling agent, antioxidant, surfactant, preservative, oil, powder, water, alcohol, thickener, chelating agent, silicone, antioxidant, humectant, aromatic, drug component, antiseptic agent, pH adjustor or neutralizer, selected as necessary or desired. It may also be used in combination with other cell activators to increase the effect of the invention.
  • an additive such as an excipient, preservative, thickener, binder, disintegrator, dispersing agent, stabilizer, gelling agent, antioxidant, surfactant, preservative, oil, powder, water, alcohol, thickener, chelating agent, silicone, antioxidant, humectant, aromatic, drug component, antiseptic agent, pH adjustor or neutralizer, selected as necessary or desired. It may also be used in combination with other cell activators to increase the effect of the invention.
  • the present invention further provides products such as sun visors, caps, clothing, gloves, screen films, window sprays or creams, window materials or wall materials, for example, that comprise a cell activator of the invention and are intended to activate skin by exposing the skin to light containing ultraviolet light.
  • products such as sun visors, caps, clothing, gloves, screen films, window sprays or creams, window materials or wall materials, for example, that comprise a cell activator of the invention and are intended to activate skin by exposing the skin to light containing ultraviolet light.
  • the usage of additives in the products of the invention and the forms of the products may also be as desired.
  • the present invention further provides a method for producing the cell activator, composition or product of the invention.
  • a method for activating skin of a subject is also provided, the method comprising application of a cell activator or composition of the invention onto the skin of a subject and exposing the skin to light containing ultraviolet light after application of the cell activator or composition; or passing light containing ultraviolet light through the product of the invention, and exposing the skin to the transmitted light; wherein the cell activator, composition or product converts the wavelength of ultraviolet light in the incident light and emits emission light with a longer wavelength than the wavelength of the ultraviolet light, transmitting the ultraviolet light with a peak wavelength of preferably 200 nm to 400 nm as light with a peak wavelength of 500 nm to 700 nm.
  • the method for activating skin will often be for the purpose of beautifying, instead of treatment by a doctor or health care professional.
  • the invention further provides a cosmetic counseling method for supporting cosmetology, which includes providing a cosmetic method, cell activator, composition or product of the invention to a subject.
  • Wavelength conversion substances were prepared in the following manner.
  • B-phycoerythrin is obtained from porphyridiophylla ( Porphiridium cruentum ) extract, the absorption spectrum having a peak wavelength at 305 nm and the emission spectrum having peak wavelengths at 570 nm and 610 nm.
  • C-phycocyanin is obtained from Spirulina ( Spirulina platensis ) extract, the absorption spectrum having a peak wavelength at 350 nm and the emission spectrum having peak wavelengths at 640 nm and 700 nm.
  • Lumate R by Sakai Chemical Industry Co., Ltd. was used.
  • Lumate R is a magnesium titanate phosphor obtained by doping MgTiO 3 with manganese, the absorption spectrum having a peak wavelength at 365 nm and the emission spectrum having peak wavelengths in the range of 660 to 680 nm.
  • the wavelength conversion substances of (1) and (2) were dissolved in water to prepare solutions at concentrations of 1% and 5%.
  • the wavelength conversion substances of (3) and (4) were dispersed in alcohol to prepare 5% and 10% dispersions.
  • Human skin fibroblasts purchased from Kurabo
  • human skin keratinocytes purchased from Kurabo or PromoCell
  • a cell suspension (1 mL) stored with liquid nitrogen was placed in a hot water bath (37° C.) and thawed until small ice pellets remained, and then diluted with 9 mL of warm medium. 2.
  • the diluted suspension was gently mixed and transferred to a T75 flask, and incubated overnight at 37° C. 3. On the following day, the medium was exchanged with 10 mL of fresh medium. 4.
  • the medium was periodically exchanged (once every 2 days for fibroblasts, or once every 2 to 3 days for keratinocytes), while continuing growth of the cells.
  • the fibroblasts were resuspended in 5 mL of warm FGM (containing 10% serum), sterilized and transferred to a 50 mL Falcon tube. The flask was further rinsed with 5 mL of warm FGM and added to the Falcon tube, to ensure transfer of all of the cells. 9. The cells were centrifuged at 10,000 rpm for 5 minutes (4° C.), and the supernatant was carefully removed while avoiding disturbing the cell pellet. 10.
  • the cells were resuspended in FGM or KGM at a concentration of 2 ⁇ 10 4 cells/well (500 ⁇ L) for fibroblasts or 4 ⁇ 10 4 cells/well (500 ⁇ L) for keratinocytes, and plated in a 24-well plate. 11.
  • the cells were seeded in the 24-well plate, with periodic exchange of the medium (once every 2 days for fibroblasts, or once every 2 to 3 days for keratinocytes), growing the cells until they reached 60 to 70% confluence (differing for the type of experiment). (Note: fibroblasts are expected to reach desired confluence by 24 hours assuming a cell density of 2 ⁇ 10 4 cells/well.
  • the medium was changed to non-supplemented medium (for keratinocytes) or low-concentration serum-containing (0.5% FCS) medium (for fibroblasts).
  • the power source of a solar simulator was activated to warm up the lamp.
  • the solar simulator used was a UG11 filter.
  • a UG11 filter is a filter that allows passage of UVB alone while cutting light of other wavelengths.
  • the UV light passing through the UG11 filter had a peak wavelength of 300 nm to 385 nm.
  • the temperature control plate was turned on and set to 33° C. 3.
  • the cells prepared in Experiment 1-2 were rinsed once with warm PBS. 4.
  • a 0.5 mL portion of warmed Martinez solution (145 mM NaCl, 5.5 mM KCl, 1.2 mM MgCl 2 .6H 2 O, 1.2 mM NaH 2 PO 4 .2H 2 O, 7.5 mM HEPES, 1 mM CaCl 2 and 10 mM D-glucose) was added to each well. 5. As shown in FIG.
  • the cell-containing wells were set on a plate, 0.4 ml of each solution containing the wavelength conversion substances (1) to (4) prepared in Experiment 1-1 was injected into the wells of a 24-well plate, the cell-containing wells were placed over it in a manner covering the wells, and UV light was irradiated into the cell solution through the wavelength conversion substance solution without allowing direct contact between the wavelength conversion substance solution and the cell solution. 6. The irradiation was carried out to a total radiation dose of 100 mJ/cm 2 .
  • the cells kept at 48 hours in an incubator after Experiment 1-3 were used for activity measurement by the following method.
  • UV irradiation lowered the cellular activity compared to the non-irradiated control.
  • the activity of the cells irradiated with UV through the wavelength conversion substances was higher compared to non-irradiation, for all of the wavelength conversion substances.
  • UV irradiation was carried out with a radiation dose of 100 mJ/cm 2 , either with the cell culture covered with a film prepared by dispersing a zinc oxide phosphor as a wavelength conversion substance in an acrylic polymer to 5%, or without using a film.
  • a wavelength conversion substance has an effect of not only inhibiting reduction in cellular activity by UV irradiation, but also of activating cells utilizing UV light. Activation of skin cells is expected to prevent or ameliorate wrinkles, blemishes, skin aging and photoaging.

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US17/601,333 Pending US20220175637A1 (en) 2019-04-05 2020-04-03 Composition cotaining ultraviolet wavelength-converting substance and hydrophobized silica and/or hydrophobized starch
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