WO2022089559A1 - 修复肌肤屏障的油脂组合物及其应用 - Google Patents
修复肌肤屏障的油脂组合物及其应用 Download PDFInfo
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- 239000000516 sunscreening agent Substances 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 229920001059 synthetic polymer Polymers 0.000 description 1
- 239000000454 talc Substances 0.000 description 1
- 229910052623 talc Inorganic materials 0.000 description 1
- 238000010998 test method Methods 0.000 description 1
- 239000012049 topical pharmaceutical composition Substances 0.000 description 1
- 230000024883 vasodilation Effects 0.000 description 1
- 231100000747 viability assay Toxicity 0.000 description 1
- 238000003026 viability measurement method Methods 0.000 description 1
- 239000004034 viscosity adjusting agent Substances 0.000 description 1
- 239000011782 vitamin Substances 0.000 description 1
- 235000013343 vitamin Nutrition 0.000 description 1
- 229940088594 vitamin Drugs 0.000 description 1
- 229930003231 vitamin Natural products 0.000 description 1
- 239000011787 zinc oxide Substances 0.000 description 1
- XOOUIPVCVHRTMJ-UHFFFAOYSA-L zinc stearate Chemical compound [Zn+2].CCCCCCCCCCCCCCCCCC([O-])=O.CCCCCCCCCCCCCCCCCC([O-])=O XOOUIPVCVHRTMJ-UHFFFAOYSA-L 0.000 description 1
- 229910001928 zirconium oxide Inorganic materials 0.000 description 1
- 239000002478 γ-tocopherol Substances 0.000 description 1
- QUEDXNHFTDJVIY-DQCZWYHMSA-N γ-tocopherol Chemical compound OC1=C(C)C(C)=C2O[C@@](CCC[C@H](C)CCC[C@H](C)CCCC(C)C)(C)CCC2=C1 QUEDXNHFTDJVIY-DQCZWYHMSA-N 0.000 description 1
Images
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K8/00—Cosmetics or similar toiletry preparations
- A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
- A61K8/92—Oils, fats or waxes; Derivatives thereof, e.g. hydrogenation products thereof
- A61K8/922—Oils, fats or waxes; Derivatives thereof, e.g. hydrogenation products thereof of vegetable origin
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K8/00—Cosmetics or similar toiletry preparations
- A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
- A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
- A61K8/33—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing oxygen
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K8/00—Cosmetics or similar toiletry preparations
- A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
- A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
- A61K8/33—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing oxygen
- A61K8/37—Esters of carboxylic acids
- A61K8/375—Esters of carboxylic acids the alcohol moiety containing more than one hydroxy group
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q19/00—Preparations for care of the skin
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q19/00—Preparations for care of the skin
- A61Q19/005—Preparations for sensitive skin
Definitions
- the invention belongs to the field of cosmetics, and in particular relates to an oil composition capable of restoring skin barrier function and its preparation and application.
- the present invention discovers for the first time an oil composition that can effectively repair damaged skin barrier, and the composition can solve the problem of poor water retention capacity of damaged skin epidermis.
- the present invention provides an oil composition for repairing skin barrier function, the composition comprising, based on the weight of the composition:
- the vegetable oil is white chia seed oil
- Synthetic oils and fats are selected from: dioctyl carbonate, caprylic/capric triglycerides or their combinations.
- the weight ratio of vegetable oil to synthetic oil in the composition of the present invention is 1:3 to 1:1.
- the content of vegetable oil in the composition of the present invention is 20-60% by weight.
- the content of synthetic oil in the composition of the present invention is 40-80% by weight.
- the present invention provides an external preparation for skin comprising the oil and fat composition according to claim 1 or 2.
- the external preparation for skin of the present invention is selected from the following forms: essence, cream, lotion, eye cream, essential oil and massage oil.
- the present invention provides the application of the oil composition in repairing skin barrier.
- the present invention provides the application of the oil composition in the preparation of a skin external preparation with a skin barrier function.
- the repairing of the skin barrier is achieved by increasing cell viability, increasing loridin LOR and/or decreasing inflammatory factors.
- the inflammatory factors are IL-1 ⁇ and PGE2.
- Figure 2 shows the tissue viability test results.
- Figure 3 shows the test results of the inflammatory factor IL-1 ⁇ .
- Figure 4 shows the test results of the inflammatory factor PGE2.
- Figure 5 shows histological (H&E) staining pictures.
- Figure 6 shows pictures of loricrin LOR immunofluorescence detection.
- Figure 7 shows the test results of loricrin LOR.
- the oil composition with a specific composition can lock the loss of moisture in the skin epidermis through the oil film formed by the oil itself, thereby helping to maintain the moisture of the skin epidermis.
- the present invention screened out the oil composition with good antioxidant ability and excellent skin feel, and evaluated the ability of the composition in terms of skin damage repairing effect through the "SLS-EpiKutis" skin barrier damage model.
- the present invention provides for the first time an oil composition with anti-allergic, anti-inflammatory, and skin barrier functions, which uses stable vegetable oil white chia seed oil and some synthetic oil (for example, dioctyl carbonate and / or the combination of caprylic acid/capric triglyceride) to achieve anti-allergic, anti-inflammatory and skin barrier functions.
- the oil and fat composition of the present invention can also be applied to skin external preparations such as essence, cream, lotion, eye cream, essence oil, massage oil, etc., to help restore skin barrier function.
- the present invention Compared with the prior art, the present invention has the following beneficial effects: the present invention combines vegetable oil (such as white chia seed oil) and synthetic oil (such as dioctyl carbonate and/or caprylic/capric triglyceride) , to achieve the practical application of modern skin care research.
- vegetable oil such as white chia seed oil
- synthetic oil such as dioctyl carbonate and/or caprylic/capric triglyceride
- the composition has the effect of promoting skin barrier repair, and the effect is better than the single use of white chia seed oil, dioctyl carbonate or caprylic/capric triglyceride.
- SLS-EpiKutis skin barrier damage model
- the present invention aims to disclose a composition that can help reduce skin inflammation and irritation, prevent bacterial invasion, protect skin structural integrity and repair damaged skin barrier, so that it can be applied to modern topical formulations.
- the vegetable oil and fat used in the oil and fat composition of the present invention is white chia seed oil.
- White chia seed oil is a vegetable oil containing more than 98% long-chain fatty acids with antioxidant effects, and has a unique molecular structure that has not been found in other natural vegetable oils.
- the function of white chi flower seed oil is moisturizing first, because it can form a water-locking film on the surface of the skin, locking in moisture and resisting dryness. Second, this unique ingredient makes White Pond Flower Seed Oil a very good antioxidant, while also stretching the skin texture, deeply repairing skin damage and enhancing the overall contour of the skin.
- Chihua oil has 35% more long-chain fatty acids than rapeseed oil and 25% more than cabbage oil. However, its methylene diene (linoleic acid) content is very low, and linoleic acid is the main factor that causes the oxidation of industrial products and affects their stability. Therefore, Chihua seed oil has strong antioxidant activity.
- the oil and fat composition of the present invention comprises 20-60% by weight of Flos japonica seed oil by weight. In a preferred embodiment, the oil and fat composition of the present invention contains 25-60% by weight of Flos japonica seed oil. In a preferred embodiment, the oil and fat composition of the present invention contains 25 to 50% by weight of Flos japonica seed oil.
- Synthetic fats and oils are also contained in the oil-fat composition of this invention.
- the synthetic oils and fats employed include dioctyl carbonate and caprylic/capric triglycerides.
- Dioctyl carbonate is a synthetic oil. It is mainly used as an emollient in cosmetics and skin care products. It has a risk factor of 1 and high safety. Dioctyl carbonate is colorless and highly soluble, used as emollient and dispersant. The ingredient is easily emulsified, which promotes easy application, creates a protective film on the skin, and is smooth and non-greasy with a silky feel. Commonly used in moisturizers/creams, serums, eye creams, cleansers and other products.
- Caprylic/capric triglycerides are clear to pale yellow, highly refreshing, odorless oils derived from palm or coconut oil. It is widely used in food, medicine, cosmetics and other industries. It can be used as a base material for moisturizing factors, a stabilizer for cosmetics, an antifreeze agent, and a homogenizer.
- the oil and fat compositions of the present invention comprise 40-80% by weight synthetic oil and fat by weight. In a preferred embodiment, the oil and fat composition of the present invention comprises 40-75% by weight of synthetic oil and fat. In a preferred embodiment, the oil and fat composition of the present invention comprises 50-75% by weight of synthetic oil and fat.
- the weight ratio of vegetable oil to synthetic oil in the oil and fat composition of the present invention is 1:3-1:1. In some embodiments of the present invention, the weight ratio of vegetable oil to synthetic oil in the oil and fat composition of the present invention is 1:2-1:1.
- the present invention also provides an external preparation for skin, the external preparation for skin comprising an oil composition having the function of restoring skin barrier.
- the external preparation for skin is a general concept of all ingredients generally used on the outside of the skin, and may be, for example, a cosmetic composition or a pharmaceutical composition.
- the cosmetic composition can be basic cosmetic, facial cosmetic, body cosmetic, hair care cosmetic, etc.
- the dosage form thereof is not particularly limited, and can be reasonably selected according to different purposes.
- the oil and fat composition of the present invention can be added to cosmetics such as essences, creams, lotions, eye creams, essential oils, and massage oils as cosmetic additives.
- the cosmetic composition also contains different medium or base excipients that are permitted in cosmetic aspects according to different dosage forms and purposes.
- Cosmetic, dermatologically or pharmaceutically acceptable excipients that can be used in the skin external preparation composition of the present invention are water phase, oil phase, gel, wax-in-water emulsion, oil-in-water emulsion, or water-in-oil in the form of an emulsion.
- the aqueous phase is a mixture of one or more water-soluble or dispersible components, which may be liquid, semi-solid or solid at room temperature (25°C).
- Excipients include or may be in the form of suspensions, dispersions or solutions in aqueous or hydro-alcoholic vehicles, which may contain thickening or gelling agents. Those skilled in the art can select the appropriate product form and the components contained therein based on the knowledge possessed by those skilled in the art.
- the composition may include an aqueous phase, which may contain water or a mixture of water and at least one hydrophilic organic solvent, such as an alcohol, especially containing 2-5 carbon atoms linear or branched lower monohydric alcohols such as ethanol or propanol; polyols such as propylene glycol, sorbitol, glycerol, panthenol or polyethylene glycol and mixtures thereof.
- an aqueous phase which may contain water or a mixture of water and at least one hydrophilic organic solvent, such as an alcohol, especially containing 2-5 carbon atoms linear or branched lower monohydric alcohols such as ethanol or propanol; polyols such as propylene glycol, sorbitol, glycerol, panthenol or polyethylene glycol and mixtures thereof.
- the composition may also optionally contain a surfactant.
- the composition may also comprise film-forming polymers such as polyurethane, polyacrylic homo- or copolymers, polyesters, hydrocarbon-based resins and/or silicone resins.
- film-forming polymers such as polyurethane, polyacrylic homo- or copolymers, polyesters, hydrocarbon-based resins and/or silicone resins.
- the polymer can be dissolved or dispersed in a cosmetically acceptable vehicle and optionally combined with a plasticizer.
- composition of the present invention may further contain any components commonly used in the cosmetic field. These components include preservatives, aqueous phase thickeners (extract biopolymers, synthetic polymers) and fatty phase thickeners, fragrances, hydrophilic and lipophilic actives, and mixtures thereof.
- compositions of the present invention may also contain additional particulate phases, which may be pigments and/or pearlescent agents and/or fillers used in cosmetic compositions.
- Pigments may be present in the composition, suitable inorganic pigments include titanium oxide, zirconium oxide and cerium oxide and zinc oxide, iron oxide and iron blue; suitable organic pigments include barium, strontium, calcium and aluminum lakes and carbon black.
- Pearlescent agents may be present in the composition, suitable pearlescent agents include mica coated with titanium oxide, iron oxide or natural pigments.
- Fillers may be present in the composition, suitable fillers include talc, silicon dioxide, zinc stearate, mica, kaolin, nylon powder, polyethylene powder, Teflon, starch, boron nitride, copolymers Microspheres, such as silicone resin microbeads.
- compositions of the present invention may be formulated into any suitable product form.
- product forms include, but are not limited to, aerosol sprays, creams, lotions, solids, liquids, dispersions, foams, gels, lotions, mousses, ointments, powders, patches, pomades, solutions , hand pump sprays, sticks, masks and wipes.
- the compositions of the present invention can be conveniently used in the preparation or topical application of products as cosmetic, dermatological or pharmaceutical products by various methods well known in the art.
- the skin external preparation composition of the present invention may include one or more of the following ingredients: antiallergic agent, anti-inflammatory agent, moisturizing agent, antimicrobial agent, antioxidant, chelating agent, colorant depigmenting agent, emollient, emulsifier Agents, exfoliating agents, film formers, fragrances, insect repellants, lubricants, pharmaceutical actives, moisturizing agents, lightfasteners, preservatives, skin care agents, skin penetration enhancers, sunscreens, stabilizers, surfactants agents, thickeners, viscosity modifiers, vitamins, or any combination thereof.
- the oil composition of the present invention has multiple functions such as anti-allergy, anti-inflammatory and skin barrier repair.
- skin barrier damage and repair model experiment Through the skin barrier damage and repair model experiment and the "SLS EpiKutis" skin barrier damage model experiment, the process of human use is simulated, and the composition is uniformly coated on the surface of the "SLS EpiKutis" skin damage model by means of surface administration. Tissue vitality, histomorphological inflammatory factors and barrier-related proteins were used to evaluate the skin damage repair efficacy of the composition.
- test methods without specific conditions are usually in accordance with conventional conditions, or in accordance with the conditions suggested by the manufacturer. All percentages and parts are by weight unless otherwise indicated.
- the white chia seed oils 1-3 used in this application were all purchased from Elementis Specialties.
- the purpose of this example is to screen stable vegetable oils and fats.
- oxidative stability measuring instrument of the OSI type equipment of Omnion Company in the United States, connect the diaphragm air pump, adjust the flow rate to 10L/h accurately, and then turn off the air pump.
- a thyristor contact thermometer or electronic controller to adjust the temperature of the heating block to 110°C. During the test, the temperature should always be kept within the range of ⁇ 0.1°C of the set value.
- Add 50ml of distilled water to the measuring cell with a pipette use a calibrated potentiometer to detect the electrode and adjust the signal to stay on the zero axis of the recording paper. Aspirate and accurately weigh the sample with a pipette.
- sample 1 caprylic/capric triglyceride and sample 2 dioctyl carbonate are synthetic oils with stable structures.
- the oxidation induction time at 110 °C is very long and the structure is very stable.
- White chia seed oil 3 is a vegetable oil, which itself contains unsaturated double bonds, which is relatively easy to be oxidized.
- white chia seed oil 1 FANCOR MEADOWFOAM SEED OIL
- white chia seed oil 2 Limnanthes alba
- white chia seed oil 3 CROPURE MEADOWFOAM.
- the trace components in vegetable oils and the degree of unsaturation of fatty acids have a great influence on the oxidative stability. ⁇ -Tocopherol and phytosterols, etc.).
- White chia seed oil has a unique structure and is different from other vegetable oils, containing more than 95% of triglycerides formed by C20-22 fatty acids.
- caprylic/capric acid triglyceride Weigh 23 parts by mass of caprylic/capric acid triglyceride and place it in a beaker, then weigh 50 parts by mass of dioctyl carbonate and 27 parts by mass of Flos japonica seed oil 1 in a beaker, and mix at 200 to 300 rpm. Mix well.
- caprylic/capric acid triglyceride Weigh 30 parts by mass of caprylic/capric acid triglyceride and place it in a beaker, then weigh 35 parts by mass of dioctyl carbonate and 35 parts by mass of Flos japonica seed oil 1 in a beaker, and mix at 200 to 300 rpm. Mix well.
- the skin barrier is disrupted by a stick-tear physical method.
- the main metric for evaluation is the TEWL value.
- the failure criterion for each location was that the TEWL value after avulsion reached 2.5 times the baseline value.
- Barrier repair rate (TEWL immediately after avulsion-TEWL at different time points)/(TEWL immediately after avulsion-basal value TEWL)*100%
- test articles included 30 grams each of Sample 1, Sample 2, Sample 12, and Example 1. The results are shown in Table 2.
- Example 1 has the largest change in the skin barrier repair rate after 72 hours of use, which is better than Sample 1, Sample 2, and Sample 12.
- test items include sample 1, sample 2, sample 12 and 1 gram each of Examples 1-6
- Moisturizing degree After the test subjects used it, they rated the samples, 1 point is not very moisturizing, 9 points are very moisturizing;
- Residual feeling After the subject uses it, the sample is scored, 1 is a lot of residue, 9 is no residue;
- Table 12 Data for calculating the average value of each indicator
- Example 1, Example 3, Example 4, Example 5, Example 6 and Sample 2 are higher than the average value, and Sample 2 has the highest freshness score.
- Example 1, Example 5, and Sample 1 are higher than the average, among which Example 1 and Sample 1 have the highest moisturizing degree score.
- From the score of absorbency, Example 1, Example 3, and Example 5 are higher than the average, and Example 1 has the highest absorbance score.
- Judging from the greasy feeling score, Example 1, Example 3, Example 4, Example 5, and Sample 2 were higher than the average, and Example 1 and Example 3 had the highest greasy feeling score.
- From the score of residual feeling Example 1, Example 3, Example 5, Example 6, Sample 1, and Sample 2 are higher than the average value, and Example 6 has the highest residual feeling score.
- Example 1, Example 2, Example 3, and Sample 2 are higher than the average, and Example 1 has the highest preference score.
- Example 1 is higher than the average in each index, and the overall preference is the highest. It is indicated that Example 1 is the most popular in terms of skin feel experience of consumers.
- SLS-EpiKutis skin barrier damage model The "SLS-EpiKutis skin barrier damage model" was constructed. This model simulates the process of human use, and the sample is evenly coated on the surface of the "SLS-EpiKutis” skin damage model by surface drug delivery. Factors and barrier-related proteins to evaluate the skin damage repair efficacy of the samples of the examples.
- the blank control group that is, the BC group, did not use any substances to stimulate the 3D skin and did not apply any drugs on the surface of the 3D skin.
- the negative control group ie the NC group, used 0.2% SLS to stimulate the 3D skin without applying any drug on the surface of the 3D skin.
- the positive control group namely the PC group, was smeared with 50 ⁇ M of WY 14643 on the surface of the 3D skin for 24 hours after stimulating the 3D skin with 0.2% SLS.
- WY-14643 i.e. pirinic acid
- WY 14643 is a potent peroxisome proliferator and PPAR ⁇ activator.
- WY 14643 (10 ⁇ M) acts on aortic smooth muscle cells, almost completely inhibits IL-1-induced IL-6 and prostaglandin production and cyclooxygenase-2 expression by inhibiting NF- ⁇ B signaling pathway.
- the test group namely the grouping of Example 1, is to apply 2-3 drops of the sample of Example 1 on the surface of the 3D skin for 24 hours after stimulating the 3D skin with 0.2% SLS.
- Detection index LOR, namely loricrin (Loricrin).
- LOR is the product of terminal differentiation of keratinocytes, expressed in the stratum corneum and granular layer, and is usually rich in amino acid residues such as Gly, Ser, Cys, etc.
- LOR is the main component of epidermal keratinocyte encapsulation, accounting for 70-85% of the total protein in the stratum corneum. %, under the action of TGM enzymes, LOR cross-links with itself, or cross-links with SPRRs, which are key reinforcement proteins in the composition of the skin barrier. Decreased LOR content leads to a weakened skin barrier function.
- Detection indicators inflammatory factors IL-1 ⁇ and PGE2.
- IL-1 ⁇ namely Interleukin 1 ⁇ (Interleukin 1 ⁇ )
- Interleukin 1 ⁇ Interleukin 1 ⁇
- IL-1 ⁇ is an important pro-inflammatory cytokine in the process of skin inflammatory response, and normally exists in the cytoplasm of keratinocytes.
- the cell membrane damage caused by external stimuli promotes the release of IL-1 ⁇ into the extracellular space, further induces the expression of inflammatory factors such as IL-6 and IL-8, and triggers an inflammatory cascade.
- PGE2 namely prostaglandin E2 (Prostaglandin E2)
- Prostaglandin E2 Prostaglandin E2
- PLA2 Phospholipase A2, phosphatidic acid A2
- COX- Cyclooxygenase 2, cyclooxygenase
- Duplicate hole 1, duplicate hole 2, and duplicate hole 3, namely R1, R2, R3, are set up to ensure the accuracy of the experimental data.
- MTT incubation After the model is washed, prepare a 24-well plate according to the number of models, and mark them accordingly. Add 0.3 mL of MTT working solution (1 mg/mL) to each well. Place the 24-well plate with the model in an incubator (37 ⁇ 1°C, 5 ⁇ 1% CO2, 95% RH) and incubate for 3h ⁇ 5min.
- Isopropyl alcohol extraction After the MTT incubation, the outer surface of the model was washed with PBS, dried with a sterile cotton swab, transferred to a new 24-well plate, and marked. Add 2 mL of isopropanol to the inner surface of the model, seal the 24-well plate with parafilm, and shake on a plate shaker for 2 h.
- Reading Pierce the model with a 200 ⁇ L pipette tip to allow the isopropanol extract to flow out of the model into a 24-well plate. Discard the pierced model, and mix the isopropanol extract in each well by pipetting 3 times. After mixing, draw 2 parts of 200 ⁇ L isopropanol extract from each well, add them to the corresponding wells of the 96-well plate, and mark them. Isopropanol was used as the zero hole. The absorbance (OD) value was read by a microplate reader (BioTek, Epoch) at a wavelength of 570 nm.
- Tissue viability (%) (OD of administration well-OD of zero-adjusted well)/(OD of control well-OD of zero-adjusted well)
- the model for tissue morphology detection was cut along the edge of the model, placed in 4% paraformaldehyde for 24 hours, embedded in paraffin and sliced, baked, dewaxed to water, stained, and dried at 37°C for half. After 1 hour, the slides were sealed with neutral gum, and after drying, pictures were collected under a 40X microscope under an upright microscope (Olympus, DP26).
- the model culture supernatant before washing was collected and stored at -80°C.
- the test was carried out according to the instructions of the ELISA test kit.
- the kits include IL-1 ⁇ ELISA detection kit (Abeam, ab100560) and PGE2 ELISA detection kit (Abeam, ab133021).
- Reagents 4% paraformaldehyde (BioSharp), xylene (Sinopharm), absolute ethanol (Sinopharm), sodium citrate (50X) (Xi'an Hut Biotechnology), Anti-LOR antibody (Abcam, ab198994), Anti-FLG Antibody (Abcam, ab218397), ABC-Peroxidase Kits (VECTASTAIN), Anti-Mouse-488 (goat anti-mouse) (Abcam), Anti-Rabbit-488 (goat anti-rabbit) (Abcam), anti-quencher (Biyuntian) ), Hochest33342 (Biyuntian).
- Fluorescence microscope Inverted microscope (Olympus, CKX41).
- Dewaxing of baked slices The paraffin slices were placed in a baking machine at 70°C, and baked for 4 hours.
- Dewaxing and hydration Immerse the sections in xylene for 10 min, replace the xylene and then soak for 10 min, soak in absolute ethanol for 5 min, soak in 95% ethanol for 5 min, and soak in 75% ethanol for 5 min. Washed with PBS buffer 3 times, 5 min each time.
- Antigen retrieval Put the paraffin sections into 0.01M sodium citrate antigen retrieval solution, use high pressure to repair, and take out the sections after cooling. Washed with PBS buffer solution 3 times, 5 min/time.
- Block peroxidase add 1 drop of 3% H2O2 to each section and incubate for 30min at room temperature to block the activity of endogenous peroxidase. Washed with PBS buffer solution 3 times, 5 min/time.
- Serum blocking dropwise add serum homologous to the secondary antibody for blocking at 37°C for 60 minutes without washing.
- Primary antibody incubation Add primary antibody working solution dropwise and incubate at 4°C overnight. Washed with PBS buffer 3 times, 5 min/time.
- Secondary antibody incubation Add the secondary antibody working solution dropwise and incubate at room temperature for 1 h. Washed with PBS buffer 3 times, 5 min/time.
- washing and photographing washing with PBS buffer 3 times, 5 min/time.
- the PBS solution was wiped off with absorbent paper, and a drop of anti-fade agent was added to mount the slide. Fluorescence microscopy pictures (20X and 40X) were taken within 24h.
- the model was stained with H&E, and the results are shown in Figure 5.
- the pictures are R1, R2, R3 from left to right.
- the stratum corneum of the NC group model was loose and thickened, the living cell layer was damaged, and the cell characteristics of the granular layer, spinous layer and basal layer of the living cell layer were not obvious.
- the damage of the viable cell layer in the PC group WY14643
- the loose and thickening of the stratum corneum was improved to a certain extent.
- the phenomenon of loose and thickening of the stratum corneum in Example 1 was significantly improved, and the phenomenon that the three-layer characteristic structure of the living cell layer was not obvious was significantly improved.
- Table 16 Summary table of LOR mean optical density (IOD/area) values:
- the LOR test pictures are also shown in Figure 6.
- the pictures are R1, R2, R3 from left to right.
- the green fluorescence is the positive site of LOR protein expression, and the blue fluorescence is the location of the nucleus.
- the more green fluorescence in the picture, the more LOR protein, and the increase in LOR content indicates that it can help repair the skin barrier.
- the expression of LOR protein in the NC group was significantly decreased (P ⁇ 0.01); compared with the NC group, the LOR protein expression in the PC(WY14643) group was significantly increased (P ⁇ 0.01); In comparison, the expression of LOR protein in Example 1 was extremely significant, which was increased by 29% (P ⁇ 0.01).
- the measures to repair the barrier damage mainly include: 1) by increasing the content of the key components (brick ash structure) of the barrier composition to achieve the effect of barrier damage repair; To achieve a certain degree of damage repair effect.
- the experiments of the present application comprehensively determine the barrier damage repair efficacy of the compounds to be tested by detecting barrier-related indicators: macroscopic tissue morphology and barrier-related protein (LOR) staining, as well as inflammatory response-related indicators.
- the experimental results show that Example 1 has better skin barrier damage repairing effect.
- compositions of Examples 1-6 can be used for the preparation of external preparations for skin.
- the external preparation for skin is preferably a cosmetic composition, such as essence, cream, lotion, eye cream, essence oil, massage oil and the like.
- the weight percentage of the compositions of Examples 1-6 in the external preparation for skin is 0.0001%-20% (w/w).
- the preferred weight percent is 0.001% to 10% (w/w).
- a more preferred weight percentage is 0.001%-5% (w/w).
- the most preferred weight percent is 0.01%-5% (w/w).
- compositions of application examples 1-7 specific application examples in external preparations for skin, and formulations and preparation methods of these dosage forms.
- "-" means no addition.
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Abstract
本发明提供了一种修复皮肤屏障功能的油脂组合物，以所述组合物的重量计，所述组合物包含：植物油脂，所述植物油脂是白池花籽油；和合成油脂，所述合成油脂选自：碳酸二辛酯、辛酸/癸酸甘油三酯或它们的组合。本发明还涉及包含该油脂组合物的皮肤外用剂及其在修复肌肤屏障方面的应用。
Description
本发明属于化妆品领域，具体涉及能够修复肌肤屏障功能的油脂组合物及其制备与应用。
随着人们生活水平的提升以及生活节奏的不断提速，越来越多的人开始关注自己的肌肤问题，对于护肤的需求越来越大。但是大部分消费者没有一个专业的科学认知，从而形成了许多不良的护肤习惯。例如，文献刘青，皮肤屏障功能修复及相关，皮肤科学通报[J],2017,34(4):432-436指出，存在过度去角质，过度清洁肌肤等。加上恶劣的天气环境、药物使用不当、遗传等因素，导致越来越多的消费者皮肤屏障受损，以至于皮肤水分加速流失，使得皮脂膜变薄，有害及刺激物质更容易进入表皮层，导致过敏、痘痘闭口、皮肤红痒、爆皮等肌肤问题。文献：何黎，皮肤屏障与保湿，实用医院临床杂志[J],2009,6(2):25-27指出，皮肤屏障是由皮脂膜和角质层组成，主要起着保持皮肤水分、防止体内水分蒸发、抵御外界异物侵入体内的作用。过度频繁的用力剥脱、去除，无异于对皮肤的天然屏障进行“强拆”，容易造成各种肌肤问题。由此可见，本领域需要保持皮肤结构完整，修复受损皮肤的方法，例如可以在保湿剂中添加含有抗炎、抗敏的组分，或者调节角质形成细胞的正常代谢。
本发明首次发现了一种能够有效修复受损肌肤屏障的油脂组合物，该组合物能够解决受损肌肤表皮保水能力不佳的问题。
发明内容
一方面，本发明提供了一种修复皮肤屏障功能的油脂组合物，以所述组合物的重量计，所述组合物包含：
植物油脂，所述植物油脂是白池花籽油；和
合成油脂，所述合成油脂选自：碳酸二辛酯、辛酸/癸酸甘油三酯或它们的组合。
在优选的实施方式中，本发明所述组合物中植物油脂与合成油脂的重量比为1:3至1:1。在优选的实施方式中，本发明所述组合物中植物油脂的含量为20-60重量％。在优选的实施方式中，本发明所述组合物中合成油脂的含量为40-80重量％。
另一方面，本发明提供了一种皮肤外用剂，其包含如权利要求1或2所述的油脂组合物。在优选的实施方式中，本发明所述皮肤外用剂选自以下形式：精华液、膏霜、乳液、眼霜、精华油和按摩油。
另一方面，本发明提供了所述油脂组合物在修复肌肤屏障方面的应用。
另一方面，本发明提供了所述油脂组合物在制备具有修复肌肤屏障功能的皮肤外用剂中的应用。在优选的实施方式中，所述修复肌肤屏障通过提高细胞活力、增加兜甲蛋白LOR和/或降低炎症因子实现。在优选的实施方式中，所述炎症因子是IL-1α和PGE2。
图1显示了使用样品1、样品2、样品12和实施例1之后皮肤屏障修复率变化。
图2显示了组织活力测试结果。
图3显示了炎症因子IL-1α的测试结果。
图4显示了炎症因子PGE2的测试结果。
图5显示了组织结构(H&E)染色图片。
图6显示了兜甲蛋白LOR免疫荧光检测图片。
图7显示了兜甲蛋白LOR的测试结果。
发明详述
除非另有定义，本文所用的所有技术和科学术语具有本发明所属领域普通技术人员共同理解的相同含义。虽然与本文所述相似或等同的任何方 法和材料可用于实施或测试本发明，但本文描述的是优选的方法和材料。对于本发明的目的，下面定义了以下术语。
本文所用术语“约”指与参比品的数量、水平、数值、维度、大小或用量相比，差异可高达30％、20％或10％的数量、水平、数值、维度、大小或用量。本文所使用的百分含量，除非另有说明，均以重量计。
在全篇说明书和权利要求书中，除非另有要求，以下词语“包含”和其变体“含有”和“包括”应理解为意指包括所述的整体或步骤，或一组整体或步骤，但不排除任何其它整体或步骤，或其它一组整体或步骤。
为了有效修复受损肌肤，首先需要解决受损肌肤表皮保水能力不佳的困难。本申请的发明人意外地发现，采用特定组成的油脂组合物能够通过油脂本身形成的油膜来锁住肌肤表皮水分的流失，从而帮助保持肌肤表皮的水分。而且，本发明通过筛选出具有良好抗氧化能力及优异肤感的油脂组合物，并通过“SLS-EpiKutis”皮肤屏障损伤模型，评价了该组合物在皮肤损伤修复功效方面的能力。
因此，本发明首次提供了一种具有抗敏、抗炎、修复肌肤屏障功能的油脂组合物，该油脂组合物采用稳定的植物油脂白池花籽油与一些合成油脂(例如，碳酸二辛酯和/或辛酸/癸酸甘油三酯)的组合，实现了抗敏、抗炎、修复肌肤屏障功能。而且，本发明的油脂组合物还可以应用于精华液、膏霜、乳液、眼霜、精华油、按摩油等皮肤外用剂中，帮助修复皮肤屏障功能。
与现有技术相比，本发明具有的有益效果如下：本发明通过对植物油脂(例如白池花籽油)和合成油脂(例如碳酸二辛酯和/或辛酸/癸酸甘油三酯)搭配组合，达到现代护肤研究的实际应用。通过皮肤屏障损伤与修复模型实验发现，该组合物有促进皮肤屏障修复的作用，比单独使用白池花籽油、碳酸二辛酯或辛酸/癸酸甘油三酯上，效果更优。另外，通过“SLS-EpiKutis”皮肤屏障损伤模型，发现该组合物具有一定抗敏抗炎效果。本发明旨在公开的组合物能帮助减少皮肤炎症和刺激，防止细菌侵入，保护皮肤结构完整和修复受损皮肤屏障，从而应用到现代外用制剂中。
植物油脂
本发明的油脂组合物中采用的植物油脂是白池花籽油。白池花籽油是一种植物油脂，含有98％以上具有抗氧化作用的长链脂肪酸，具有独特的分子结构，在其他天然植物油中没有发现过这种结构。白池花籽油的功能首先是保湿，因为他可以在肌肤表面形成锁水性薄膜，紧锁水分抵御干燥。其次，这种独特的成分使白池花籽油具有非常好的抗氧化剂，同时也能舒展肌肤纹理，深层修复肌肤损失，提升肌肤整体轮廓。从化学组成上来看，池花油的长链脂肪酸比菜籽油多35％，比海甘蓝油多25％。而它的亚甲基二烯(亚油酸)含量却很低，亚油酸是造成工业产品发生氧化而影响其稳定性的主要因素。故而池花籽油具有极强的抗氧化活性。
在一些实施方式中，以重量计，本发明油脂组合物包含20-60重量％的白池花籽油。在优选的实施方式中，本发明油脂组合物包含25-60重量％的白池花籽油。在优选的实施方式中，本发明油脂组合物包含25-50重量％的白池花籽油。
合成油脂
本发明的油脂组合物中还包含合成油脂。在本发明的一些实施方式中，采用的合成油脂包括碳酸二辛酯和辛酸/癸酸甘油三酯。
碳酸二辛酯是一种合成油脂，在化妆品、护肤品里主要作用是柔润剂，风险系数为1，安全性高，对于孕妇一般没有影响，没有致痘性。碳酸二辛酯无色且溶解性强，作柔润剂和分散剂使用。该成分易乳化，能够促进产品易于涂抹，可以使皮肤形成保护膜，并且光滑不油腻，留有丝滑触感。常用于保湿乳/霜，精华液、眼霜、洁面乳等产品中。
辛酸/癸酸甘油三酯为清晰至淡黄色的高清爽度无味油脂，属棕榈油或椰子油的衍生物。广泛应用于食品、医药、化妆品等行业，可作为保湿因子的基料，化妆品的稳定剂，防冻剂，均质剂。
在一些实施方式中，以重量计，本发明油脂组合物包含40-80重量％的合成油脂。在优选的实施方式中，本发明油脂组合物包含40-75重量％的合成油脂。在优选的实施方式中，本发明油脂组合物包含50-75重量％的合成油脂。
在本发明的一些实施方式中，本发明的油脂组合物中植物油脂与合成油脂的重量比为1:3-1:1。在本发明的一些实施方式中，本发明的油脂组合物中植物油脂与合成油脂的重量比为1:2-1:1。
皮肤外用剂
本发明还提供了一种皮肤外用剂，所述皮肤外用剂包含具有修复肌肤屏障功能的油脂组合物。
所述皮肤外用剂是通常用于皮肤外部的所有成分的统称概念，例如可以是化妆品组合物或药学组合物。所述化妆品组合物中可以是基础化妆品、面部妆容化妆品、身体用化妆品、头发护理用化妆品等，对其剂型无特殊限制，根据不同目的可合理选择。
本发明的油脂组合物可以作为化妆品添加剂添加到精华液、膏霜、乳液、眼霜、精华油、按摩油等化妆品中。
所述化妆品组合物中根据剂型和目的的不同还含有不同的化妆品学层面允许的介质或基质赋形剂。
可以用于本发明皮肤外用剂组合物的化妆品、皮肤病学或药学上可接受的赋形剂为水相、油相、凝胶、水包蜡型乳液、水包油型乳液或油包水型乳液的形式。水相为一种或多种水溶性或分散性组分的混合物，其在室温(25℃)下可以为液体、半固体或固体。赋形剂包括或可以为在水或水-醇赋形剂中的混悬液、分散液或溶液的形式，其可以含有增稠剂或凝胶剂。本领域技术人员可以基于本领域技术人员掌握的知识选择合适的产品形式，其中包含的组分。
所述的组合物可以包括水相，该水相可以含有水或水与至少一种亲水性有机溶剂的混合物，所述的亲水性有机溶剂诸如醇，尤其是含有2-5个碳原子的直链或支链低级一元醇，如乙醇或丙醇；多元醇，如丙二醇、山梨醇、甘油、泛醇或聚乙二醇及其混合物。
当发明的组合物为乳液形式时，该组合物还可以任选包含表面活性剂。
所述的组合物还可以包含成膜聚合物，如聚氨基甲酸酯、聚丙烯酸均聚物或共聚物、聚酯、基于烃的树脂和/或硅氧烷树脂。可以将聚合物溶于 或分散于化妆品可接受的赋形剂中并且任选与增塑剂合并。
本发明的组合物可以进一步包含常用于化妆品领域中的任何组分。这些组分包括防腐剂、水相增稠剂(提取物生物聚合物、合成聚合物)和脂肪相增稠剂、芳香剂、亲水性和亲脂性活性剂及其混合物。
本发明的组合物还可以包含另外的颗粒相，所述的颗粒相可以为化妆品组合物中使用的颜料和/或珠光剂和/或填充剂。
颜料可以存在于组合物中，合适的无机颜料包括氧化钛、氧化锆和氧化铈以及氧化锌、氧化铁和铁蓝；合适的有机颜料包括钡、锶、钙和铝色淀和碳黑。
珠光剂可以存在于组合物中，合适的珠光剂包括涂覆了氧化钛、氧化铁或天然颜料的云母。
填充剂可以存在于组合物中，合适的填充剂包括滑石粉、二氧化硅、硬脂酸锌、云母、高岭土、尼龙粉末、聚乙烯粉末、特氟龙、淀粉、一氮化硼、共聚物微球，例如硅氧烷树脂微珠。
可以将本发明的组合物配制成任何合适的产品形式。这类产品形式包括，但不限于气溶胶型喷雾剂、霜剂、乳液、固体、液体、分散体、泡沫、凝胶、化妆水、摩丝、软膏、粉剂、贴剂、润发油、溶液、手按泵型喷雾剂、棒状物、面膜和湿纸巾。可以将本发明的组合物通过本领域众所周知的各种方法便利地用于制备或作为化妆品、皮肤病学或药物局部施用产品。
本发明的皮肤外用剂组合物可以包括一种或多种下列成分：抗过敏剂、抗炎剂、保湿剂、抗微生物剂、抗氧化剂、螯合剂、着色剂去色素剂、润肤剂、乳化剂、表皮脱落剂、成膜剂、香料、昆虫驱避剂、润滑剂、药物活性剂、增湿剂、耐光剂、防腐剂、护肤剂、皮肤渗透增强剂、防晒剂、稳定剂、表面活性剂、增稠剂、粘度调节剂、维生素或其任意组合。
油脂组合物的功效
本发明的油脂组合物具有抗敏、抗炎、修复肌肤屏障等多重功效。通过皮肤屏障损伤与修复模型实验和“SLS EpiKutis”皮肤屏障损伤模型实验，模拟人体使用过程，采用表面给药的方式，将组合物均匀涂布于“SLS EpiKutis”皮肤损伤模型表面，通过检测模型组织活力、组织形态学炎症因子和屏障相关蛋白，评价组合物的皮肤损伤修复功效。
以下，通过优选的实施方式对本发明的技术方案进行详细说明，但本发明的范围并不局限于这些实施方式，其旨在说明本发明的技术方案而不是限制本发明的范围。下列实施例中未注明具体条件的试验方法，通常按照常规条件，或按照制造厂商所建议的条件。除非另有说明，所有的百分比和份数按重量计。
以下实施例中使用的各种油脂的来源和名称如下：
本申请中使用的白池花籽油1-3均购自海明斯特殊化学公司(Elementis Specialties)。
测试例1：油脂的氧化稳定性测试
本实施例的目的是筛选稳定的植物油脂。
取用以下14种不同的油脂各20g，辛酸/癸酸甘油三酯、碳酸二辛酯、鳄梨油、橄榄油、榛子油、杏仁油、米糠油、红花籽油、初榨橄榄油、小麦胚芽油、澳洲坚果油、白池花籽油1、白池花籽油2、白池花籽油3，根据GB-T21121-2007的方法，在110℃下测试其本身的氧化稳定性，测试样品110℃氧化诱导时间。最后将油脂的氧化稳定性数据进行比较。
使用美国Omnion公司OSI型设备的氧化稳定性的测定仪器，连接好隔膜式气泵，将流量精准调至10L/h，然后再关掉气泵。用可控硅触点式温度计或电子控制器将加热块温度调节至110℃，在测试过程中，温度应一直保持在设定值±0.1℃的范围内。用移液管在测量池内加入50ml蒸馏水，用校准的电位计检测电极并调节信号使其停留在记录纸的零轴线上。用移液管吸取并准确称取试样。打开隔膜式气泵，将流量精准设置为10L/h，用连接软管将通气管的进、出气口分别与泵和测量池相连接。盖好通气管密封塞，并将通气管置于已达到110℃的加热块上相应的孔中，当信号达到记录仪满刻度时结束测量。仪器通过曲线的二阶导数的最大值自动计算出诱导时间。结果如表1所示。
表1：油脂氧化稳定性测试比较
从实验数据上可以看到，样品1辛酸/癸酸甘油三酯和样品2碳酸二辛酯 为合成油脂，其结构稳定，在110℃下的氧化诱导时间很长，结构非常稳定。样品3-14的鳄梨油、橄榄油、榛子油、杏仁油、米糠油、红花籽油、初榨橄榄油、小麦胚芽油、澳洲坚果油、白池花籽油1、白池花籽油2、白池花籽油3为植物油脂，其本身含有不饱和双键，比较容易被氧化。
白池花籽油1的商品名为FANCOR MEADOWFOAM SEED OIL、白池花籽油2的商品名为Limnanthes alba、白池花籽油3的商品名为CROPURE MEADOWFOAM。植物油脂中的微量成分与脂肪酸不饱和程度对氧化稳定性起到很大的影响，白池花籽油的氧化稳定性主要与其本身含有的微量成分有关(如α-生育酚、β-生育酚、γ-生育酚及植物甾醇等)。白池花籽油结构独特，不同于其它植物油，含95％以上的C20-22脂肪酸形成的甘油三酯。从样品12-14的白池花籽油1、白池花籽油2、白池花籽油3可以看出，白池花籽油在110℃下的氧化诱导时间比其他植物油脂要长，说明其在植物油脂中相对比较稳定。尤其是样品12，其氧化诱导时间接近合成油脂，说明在植物油中，白池花籽油1的氧化稳定性最好。
实施例1：混合油脂组合物的制备
称取23质量份的辛酸/癸酸甘油三酯置于烧杯中，再称取27质量份的碳酸二辛酯和50质量份的白池花籽油1于烧杯中，在200～300rpm下，混合搅拌均匀配用。
实施例2：混合油脂组合物的制备
称取27质量份的辛酸/癸酸甘油三酯置于烧杯中，再称取23质量份的碳酸二辛酯和50质量份的白池花籽油1于烧杯中，在200～300rpm下，混合搅拌均匀配用。
实施例3：混合油脂组合物的制备
称取23质量份的辛酸/癸酸甘油三酯置于烧杯中，再称取50质量份的碳酸二辛酯和27质量份的白池花籽油1于烧杯中，在200～300rpm下，混合搅拌均匀配用。
实施例4：混合油脂制备
称取35质量份的辛酸/癸酸甘油三酯置于烧杯中，再称取35质量份的碳酸二辛酯和30质量份的白池花籽油1于烧杯中，在200～300rpm下，混合搅拌均匀配用。
实施例5：混合油脂组合物的制备
称取30质量份的辛酸/癸酸甘油三酯置于烧杯中，再称取35质量份的碳酸二辛酯和35质量份的白池花籽油1于烧杯中，在200～300rpm下，混合搅拌均匀配用。
实施例6：混合油脂组合物的制备
称取35质量份的辛酸/癸酸甘油三酯置于烧杯中，再称取30质量份的碳酸二辛酯和35质量份的白池花籽油1于烧杯中，在200～300rpm下，混合搅拌均匀配用。
测试例2：皮肤屏障损伤与修复评估
采用Corneofix20胶带，用黏贴-撕脱的物理方式破坏皮肤屏障。评估的主要指标是TEWL值。每个位置的破坏标准为撕脱后的TEWL值达到基础值的2.5倍。
屏障修复率＝(撕脱后即刻的TEWL-不同时间点的TEWL)/(撕脱后即刻的TEWL-基础值TEWL)*100％
选用年龄在25～35岁左右的年轻女性，人数为8人，每天早晚各使用一次，每次测试前用清水轻轻擦洗手前臂内侧，自然晾干。再用胶带黏贴手前臂内侧，使得撕脱后的TEWL值达到基础值的2.5倍后。检测手前臂内侧的即刻、2h、24h、48h、72h、168h的TEWL值。空白对照为用胶带黏贴手前臂内侧，但不使用任何样品。
测试品包括样品1、样品2、样品12和实施例1各30克。结果如表2所示。
表2：使用后皮肤屏障修复率变化
从实验结果来看，样品1在使用72h后，对于皮肤屏障修复变化率达到80.3％，样品2在使用72h后，对于皮肤屏障修复变化率达到79.6％，样品12在使用72h后，对于皮肤屏障修复变化率达到82.4％，实施例1在使用72h后，对于皮肤屏障修复变化率达到83.3％。故样品1、样品2、样品12、实施例1都有一定的促进皮肤屏障修复的作用。其中，实施例1在使用72h后对于皮肤屏障修复率的变化最大，优于样品1、样品2、样品12。
测试例3：消费者测试评估
选用年龄在25～50岁左右的女性，人数为10人，先使用洁面乳清洁两手臂内侧，自然晾干。再分别涂抹样品于手臂内侧，涂抹至吸收后，对使用的样品进行分数评估。
测试品包括样品1、样品2、样品12以及实施例1-6各1克
测试指标
清爽度：受试者使用后，给样品打分，1分为非常不清爽，9分为非常清爽；
滋润度：受试者使用后，给样品打分，1分为非常不滋润，9分为非常滋润；
吸收度：受试者使用后，给样品打分，1分为非常不好吸收，9分为非常好吸收；
油腻感：受试者使用后，给样品打分，1分为非常油腻，9分为非常不油腻；
残留感：受试者使用后，给样品打分，1分为有很多残留，9分为没有残留感；
总体喜好度：受试者使用后，给样品打分，1分为非常不喜欢，9分为非常喜欢。
评估数据
表3：实施例1评估数据
表4：实施例2评估数据
表5：实施例3评估数据
表6：实施例4评估数据
表7：实施例5评估数据
表8：实施例6评估数据
表9：样品1评估数据
表10：样品2评估数据
表11：样品12评估数据
表12：各项指标计算平均值的数据
从测试结果上来看，从清爽度的评分上来看，实施例1、实施例3、实施例4、实施例5、实施例6以及样品2高于平均值，其中样品2清爽度评分最高。从滋润度的评分上来看，实施例1、实施例5、样品1高于平均值，其中实施例1和样品1滋润度评分最高。从吸收度的评分上来看，实施例1、实施例3、实施例5高于平均值，其中实施例1吸收度评分最高。从油腻感的评分上来看，实施例1、实施例3、实施例4、实施例5、样品2高于平均值，其中实施例1和实施例3油腻感评分最高。从残留感的评分上来看，实施例1、实施例3、实施例5、实施例6、样品1、样品2高于平均值，其中实施例6残留感评分最高。从对测试样品整体喜好度上来看，实施例1、实施例2、实施例3、样品2高于平均值，其中实施例1在喜好度评分最高。
综上指标数据，实施例1在各项指标上都高于平均值，且总体喜好度最高。说明实施例1在消费者使用肤感体验上最受欢迎。
测试例4：“SLS-EpiKutis”损伤模型测试评估
采用十二烷基硫酸钠(SLS)刺激3D皮肤模型
构建“SLS-EpiKutis皮肤屏障损伤模型。该模型模拟人体使用过程，采用表面给药的方式，将样品均匀涂布于“SLS-EpiKutis”皮肤损伤模型表面，通过检测模型组织活力、组织形态学炎症因子和屏障相关蛋白，评价实施例样品的皮肤损伤修复功效。
空白对照组，即BC分组，为不采用任何物质刺激3D皮肤并且不涂抹任何药物于3D皮肤表面。
阴性对照组，即NC分组，为采用0.2％SLS刺激3D皮肤后，不涂抹任何药物于3D皮肤表面。
阳性对照组，即PC分组，为采用0.2％SLS刺激3D皮肤后，涂抹50μM的WY 14643药物于3D皮肤表面24小时。
WY-14643，即匹立尼酸，是一种有效的过氧化物酶体增殖子和PPARα激活剂。WY 14643(10μM)作用于主动脉平滑肌细胞，通过抑制NF-κB信号通路，几乎完全抑制IL-1诱导的IL-6和前列腺素的产生及环氧合酶-2的表达。
测试组，即实施例1分组，为采用0.2％SLS刺激3D皮肤后，涂抹2～3滴实施例1样品于3D皮肤表面24小时。
检测指标：LOR，即兜甲蛋白(Loricrin)。
LOR是角质细胞末端分化的产物，表达在角质层和颗粒层，通常富含Gly、Ser、Cys等氨基酸残基，LOR是表皮角质化胞封的主要组件，占角质层总蛋白的70～85％，在TGM酶的作用下，LOR与自身交联，或与SPRRs交联，是皮肤屏障组成的关键加固蛋白。LOR含量的降低会导致皮肤屏障功能减弱。
检测指标：炎症因子IL-1α和PGE2。
IL-1α，即白介素1α(Interleukin 1α)是皮肤炎症反应过程中的重要前炎性细胞因子，正常情况下存在于角质形成细胞的细胞质中。当皮肤屏障受损，外来刺激物造成细胞膜损伤促使IL-1α释放至胞外，进一步诱导IL-6、IL-8等炎性因子的表达，引发炎症级联反应。
PGE2，即前列腺素E2(Prostaglandin E2)是花生四烯酸级联反应中产生的前列腺素类介质，可以诱发血管扩张，导致皮肤炎症的形态学改变。在PLA2(Phospholipase A2，磷脂酸A2)、COX-(Cyclooxygenase 2，环氧合酶)等因子调控下由角质形成细胞生成。
复孔1、复孔2、复孔3，即R1、R2、R3，设置复孔是为了保证实验数据的准确性。
组织活力检测
MTT孵育：模型清洗结束后，根据模型数量准备24孔板，并做相应标记。每孔加入0.3mL的MTT工作液(1mg/mL)。将装有模型的24孔板放置于培养箱(37±1℃、5±1％CO2、95％RH)中，孵育3h±5min。
异丙醇浸提：MTT孵育结束后，PBS清洗模型外表面，并用无菌棉签擦干，转移到新的24孔板中，并做好标记。在模型内表面加入2mL异丙醇，用封口膜密封24孔板，在孔板振摇器上震荡2h。
读数：用200μL移液器枪头刺穿模型，使异丙醇浸提液从模型内流出到24孔板中。丢弃被刺穿的模型，吹打每孔内的异丙醇浸提液3次使其充分混匀。混匀后，从每孔中吸取2份200μL异丙醇浸提液，分别加入96孔板的对应孔中，做好标记。采用异丙醇作为调零孔。酶标仪(BioTek，Epoch)570nm波长读取吸光度(OD)值。
计算：按以下公式计算各实验分组的相对组织活力：
组织活力(％)＝(给药孔OD-调零孔OD)/(对照孔OD-调零孔OD)
组织形态检测
将用于组织形态检测的模型沿模型边缘环切取下，放置于4％的多聚甲醛进行固定24h后，石蜡包埋切片，经过烤片，切片脱蜡至水，染色，37℃烘干半小时后，中性树胶封片，干燥后，正置显微镜(Olympus，DP26)下40X镜下，采集图片。
炎症因子检测
收集清洗前模型培养上清液，保存于-80℃。依据ELISA检测试剂盒说明书进行测试。试剂盒包括IL-1αELISA检测试剂盒(Abcam，ab100560)和PGE2ELISA检测试剂盒(Abcam，ab133021)。
免疫荧光检测
试剂：4％多聚甲醛(BioSharp)、二甲苯(国药)、无水乙醇(国药)、柠檬酸钠(50X)(西安赫特生物)、Anti-LOR抗体(Abcam，ab198994)、Anti-FLG抗体(Abcam，ab218397)、ABC-Peroxidase Kits(VECTASTAIN)、 Anti-Mouse-488(山羊抗鼠)(Abcam)、Anti-Rabbit-488(山羊抗兔)(Abcam)、抗淬灭剂(碧云天)、Hochest33342(碧云天)。
荧光显微镜：倒置显微镜(Olympus，CKX41)。
烤片脱蜡：石蜡切片置于70℃烤片机中，烤片4小时。
脱蜡水化：将切片置于二甲苯中浸泡10min，更换二甲苯后再浸泡10min，无水乙醇中浸泡5min，95％乙醇中浸泡5min，75％乙醇中浸泡5min。PBS缓冲液清洗3次，每次5min。
抗原修复：将石蜡切片放入0.01M柠檬酸钠抗原修复溶液，采用高压修复，冷却后取出切片。PBS缓冲溶液清洗3次，5min/次。
阻断过氧化物酶：每张切片加1滴3％H2O2，室温下孵育30min，以阻断内源性过氧化物酶的活性。PBS缓冲溶液清洗3次，5min/次。
血清封闭：滴加与二抗同源的血清37℃封闭60min，无需冲洗。
一抗孵育：滴加一抗工作液，4℃孵育过夜。PBS缓冲液清洗3次，5min/次。
二抗孵育：滴加二抗工作液，室温孵育1h。PBS缓冲液清洗3次，5min/次。
核染：二抗孵育结束后，PBS缓冲液清洗3次，5min/次。甩净玻片上附着的PBS溶液，每张切片滴加100μL的Hochest33342工作液，室温孵育5min。
清洗拍照：PBS缓冲液清洗3次，5min/次。用吸水纸擦去PBS溶液，加一滴抗淬灭剂封片。24h内荧光显微镜拍照(20X和40X)。
结果分析：利用Image-pro Plus(IPP)软件分析目的蛋白荧光强度。
数据统计分析
应用GraphPad Prism Program软件作图，各组间采用T-test统计分析，*p＜0.05表示差异显著，**p＜0.01表示差异极显著。
实验结果：
表13：组织活力MTT数据结果
结合表13的结果可知，与BC组相比，NC组细胞活力极显著下降(P<0.01)；与NC组相比，PC(WY14643)组细胞活力显著提升(P<0.01)；与NC组相比，实施例1细胞活力显著性提升(P<0.05)。
表14：炎症因子IL-1α数据结果
结合表14的结果可知，与BC组相比，NC组IL-1α含量极显著上升(P<0.01)；与NC组相比，PC(WY14643)组IL-1α含量极显著下降(P<0.01)；与NC组相比，实施例1IL-1α含量极显著下降(P<0.01)。
表15：PGE2实验数据结果
结合表15的结果可知，与BC组相比，NC组PGE2含量极显著上升(P<0.01)；与NC组相比，PC(WY14643)组PGE2含量极显著下降(P<0.01)；与NC组相比，实施例1两组PGE2含量极显著下降(P<0.01)。
组织结构(H&E)的图片
培养结束后模型进行H&E染色，结果如图5所示。图片自左向右分别为R1、R2、R3。与BC相比，NC组模型角质层疏松增厚，活细胞层受损，活细胞层颗粒层、棘层、基底层细胞特征不明显。与NC组相比，PC组(WY14643)活细胞层受损情况明显改善，角质层疏松增厚现象有一定改善。与NC组相比，实施例1角质层疏松增厚现象有明显改善，活细胞层三层特征结构不明显现象有明显改善。
表16：LOR平均光密度(IOD/面积)值汇总表：
结合表16的结果可知，与BC组相比，NC组LOR蛋白表达极显著下降(P<0.01)；与NC组相比，PC(WY14643)组LOR蛋白表达极显著提升(P<0.01)；与NC组相比，实施例1的LOR蛋白表达极显著提升(P<0.01)。
此外，LOR试验图片还在图6中显示。图片自左向右分别为R1、R2、R3。绿色荧光为LOR蛋白表达阳性部位，蓝色荧光为细胞核所在部位。图片中绿色荧光越多代表LOR蛋白量越多，LOR含量的提升说明能帮助皮肤屏障的修复。与BC组相比，NC组LOR蛋白表达极显著下降(P<0.01)；与NC组相比，PC(WY14643)组LOR蛋白表达极显著提升(P<0.01)；实施例1与NC组相比，实施例1在LOR蛋白表达极显著，相比提升29％(P<0.01)。
屏障损伤修复的措施主要包括：1)通过提升屏障组成的关键成份(砖灰结构)的含量来达到屏障损伤修复的作用；2)因为屏障损伤过程会伴随着炎症反应的发生，因此抗炎可以达到一定程度的损伤修复功效。
本申请的实验通过检测屏障相关指标：宏观层面的组织形态和屏障相关蛋白(LOR)染色，以及炎症反应相关的指标，综合判定待测化合物的屏障损伤修复功效。实验结果显示，实施例1具有较好的皮肤屏障损伤修复功效。
应用例
取实施例1-6的组合物，可用于皮肤外用剂的制备。
所述皮肤外用剂优选为化妆品组合物，例如精华液、膏霜、乳液、眼霜、精华油、按摩油等。所述实施例1-6的组合物，在皮肤外用剂中的重量百分比为0.0001％-20％(w/w)。优选的重量百分比为0.001％-10％(w/w)。更优选的重量百分比为0.001％-5％(w/w)。最优选的重量百分比为0.01％-5％(w/w)。
以下是应用例1-7的组合物，在皮肤外用剂中的具体应用的实施例，及其这些剂型的配方和制备方法。以下各表中“-”表示无添加。
应用例1：面霜的制备
应用例2：乳液的制备
应用例3：精华液的制备
应用例4：面膜的制备
应用例5：眼霜的制备
应用例6：精华油的制备
应用例7：按摩油的制备
以上所有皮肤外用剂中的具体应用例中，实施例1-6的组合物均可部分或者全部取代配方中的油脂部分直接应用于配方中。
Claims (10)
- 一种修复皮肤屏障功能的油脂组合物，以所述组合物的重量计，所述组合物包含：植物油脂，所述植物油脂是白池花籽油；和合成油脂，所述合成油脂选自：碳酸二辛酯、辛酸/癸酸甘油三酯或它们的组合。
- 如权利要求1所述的油脂组合物，其特征在于，所述组合物中植物油脂与合成油脂的重量比为1:3至1:1。
- 如权利要求1或2所述的油脂组合物，其特征在于，所述组合物中植物油脂的含量为20-60重量％。
- 如权利要求1或2所述的油脂组合物，其特征在于，所述组合物中合成油脂的含量为40-80重量％。
- 一种皮肤外用剂，其包含如权利要求1或2所述的油脂组合物。
- 如权利要求5所述的皮肤外用剂，所述皮肤外用剂选自以下形式：精华液、膏霜、乳液、眼霜、精华油和按摩油。
- 如权利要求1所述的油脂组合物在修复肌肤屏障方面的应用。
- 如权利要求1所述的油脂组合物在制备具有修复肌肤屏障功能的皮肤外用剂中的应用。
- 如权利要求7或8所述的应用，所述修复肌肤屏障通过提高细胞活力、增加兜甲蛋白LOR和/或降低炎症因子实现。
- 如权利要求9所述的应用，所述炎症因子是IL-1α和PGE2。
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