Preparation method and application of composition for repairing sensitive skin

Abstract

The invention belongs to the technical field of skin care products, and particularly relates to a preparation method and application of a composition for repairing sensitive skin. The sensitive skin repairing composition is prepared by adopting a staged fermentation preparation method taking the rhizoma bletillae root extract as a carbon source, sequentially performing low-temperature fermentation of bifidobacterium and high-temperature fermentation of thermophilic thermophilic bacteria, and compounding with auxiliary materials, so that the defects of poor stability, low permeability and strong irritation of bifidobacterium fermentation liquor are overcome. Stepped degradation of macromolecular polysaccharide is achieved through double fermentation, and the permeability and stability of the product are improved; high-temperature fermentation is utilized to inactivate infectious microbes and consume residual organic acid, so that the irritation is reduced. The composition is used for skin care products such as water aqua, essence milk and the like, has excellent stability, is mild and non-irritant, has long-acting moisturizing and antioxidant properties, and provides an effective scheme for sensitive skin repair.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of skin care products, and particularly relates to a preparation method of a composition for repairing sensitive skin and application in the preparation of a skin care product for repairing sensitive skin. BACKGROUND

[0002] Sensitive skin, simply referred to as "sensitive skin", is a state of high skin intolerance, which is manifested as subjective symptoms such as burning, stinging, itching and tightness, and objective signs such as erythema, desquamation and capillary dilation when the skin is stimulated by physical, chemical or mental factors. The root cause lies in the damage to the skin barrier function, which leads to the decline of the skin's defense ability to external stimuli, and the nerve immune system inside the skin is in a state of irritability. With the acceleration of modern life pace, the intensification of environmental pollution and the popularity of improper skin care habits, the proportion of sensitive skin population is increasing year by year worldwide. Therefore, the development of safe and efficient sensitive skin repair products has become an important demand in the skin care market.

[0003] At present, there are various types of products on the market for repairing sensitive skin, and their mechanisms of action mainly focus on the following aspects: (1) supplementing intercellular lipids to repair the physical barrier: such as adding ceramide, cholesterol, free fatty acids and other ingredients. (2) increasing skin moisture and reducing trans-epidermal water loss (TEWL): such as using hyaluronic acid, glycerol, butylene glycol and other moisturizing agents. (3) soothing and anti-inflammatory to inhibit inflammatory response: such as using bisabolol, potassium glycyrrhizinate and various plant extracts (such as spilanthes extract, centella asiatica extract, etc.). Commercially known products such as Venuona sensitive skin care cream use plant extracts such as spilanthes and prunus mume oil to soothe and repair the skin.

[0004] However, the existing technology for repairing sensitive skin still has the following technical problems: (1) single function: many products only focus on moisturizing or single anti-inflammatory, and cannot solve the complex problem of coexistence of skin barrier damage, microecological imbalance and chronic inflammation from the root. The formation of sensitive skin is multifactorial, and single-pathway repair has limited effect and is prone to recurrence. (2) It is difficult to balance immediate relief and long-term repair: some products can quickly relieve redness, itching and other symptoms, but they lack the ability to deeply repair and rebuild the skin barrier; while some products focus on long-term repair, but their soothing effect in acute sensitive period is not satisfactory. (3) Lack of ingredient synergy: simple ingredient stacking may not produce a 1+1>2 effect, and even may increase the burden on the skin or cause new irritation. For example, some products containing fermented ingredients may not be suitable for severe sensitive skin. (4) Insufficient defense against environmental damage: the damaged sensitive skin barrier is more vulnerable to environmental hazards such as ultraviolet and infrared rays, and most repair products on the market fail to provide effective protection against these environmental pressures, thereby hindering the repair process.

[0005] Schizochytrium ferment has been proven to have significant application value and advantage in sensitive skin care field due to its comprehensive effects in repairing skin barrier, anti-inflammatory soothing, promoting DNA repair, etc. However, as a complex biological source ingredient, its inherent allergic risk and individual intolerance are its unavoidable shortcomings in sensitive skin application. For example, poor stability, easy to be inactivated by temperature fluctuation and extreme pH value, leading to efficacy attenuation within product shelf life; trace organic acids remaining in fermentation process, which may cause adverse reactions such as skin itching, redness, stinging, swelling, and even acne (papules, pustules), etc.; and strong water solubility of the ingredient, short residence time on the skin surface, difficult to effectively penetrate the stratum corneum, low penetration rate, thereby affecting the full play of repair effect. Therefore, it is urgent to develop a Schizochytrium ferment composition suitable for sensitive skin repair, which is mild in formula, high in performance, and excellent in skin feel. SUMMARY

[0006] The purpose of the present application is to overcome the defects of poor stability, low stratum corneum penetration rate and adverse reactions caused by residual organic acids in fermentation of Schizochytrium ferment in sensitive skin repair in the prior art, and to provide a preparation method and application of a composition for sensitive skin repair.

[0007] The present application relates to a preparation method of a composition for sensitive skin repair, comprising the following steps: S1, stirring and mixing of bletilla striata root extract, nitrogen source and inorganic salt to obtain fermentation medium; S2, first step fermentation: sterilizing the fermentation medium, adjusting the pH to 6.5-7.0, inoculating 3-10% volume of bifidobacterium strain for fermentation to obtain primary fermentation liquor of Schizochytrium ferment; S3, second step fermentation: heating the primary fermentation liquor to 60-80℃, adjusting the pH to 7.0-7.5, inoculating 0.5-2% volume of thermophilic thermus strain for fermentation to obtain secondary fermentation liquor of Schizochytrium ferment; S4, post-treatment: high-pressure homogenization of the secondary fermentation liquor at 80-100 MPa, filtration and sterilization, collection of the filtrate to obtain final fermentation liquor of Schizochytrium ferment; S5, stirring and mixing of the final fermentation liquor of Schizochytrium ferment, 1,2-pentanediol, caprylhydroxamic acid, 1,3-propanediol, penetration enhancer and surfactant to obtain the composition for sensitive skin repair.

[0008] Preferably, the preparation of bletilla striata root extract in step S1 comprises the following steps: taking dried bletilla striata root, crushing and passing through a 30-80 mesh sieve, adding 10-20 times mass of deionized water, refluxing extraction at 80-90℃ for 2-5h, and concentrating the filtrate to a solid content of 5-15wt% after filtration to obtain the bletilla striata root extract.

[0009] Preferably, the culture medium contains 97-99.4wt% of the bletilla striata root extract, 0.5-2wt% of the nitrogen source, and 0.1-1wt% of the inorganic salt.

[0010] Preferably, the nitrogen source of the step S1 includes at least one of soybean peptone and tryptone; and the inorganic salt includes at least one of potassium dihydrogen phosphate, magnesium sulfate, and zinc sulfate.

[0011] Preferably, the solid content of the bletilla striata root extract of the step S1 is 5-10wt%.

[0012] Preferably, the fermentation temperature of the step S2 is 30-40℃, the fermentation time is 24-72h, the concentration of the bifidobacterium is 1×10 8 ~1×10 10 CFU / mL.

[0013] Preferably, the fermentation temperature of the step S3 is 50-60℃, the fermentation time is 24-48h, and the concentration of the thermophilic thermus species is 1×10 7 ~1×10 8 CFU / mL.

[0014] Preferably, the filtering is performed by using a polypropylene microfiltration membrane with a pore size of 1.5μm-3.5μm.

[0015] Preferably, the sterilization of the step S4 is low-temperature pasteurization, the sterilization temperature is 80-100℃, and the sterilization time is 30-60min.

[0016] Preferably, the sensitive skin repair composition of the step S5 includes 80-100 parts of the final fermentation broth, 3-10 parts of 1,2-pentanediol, 0.01-0.2 parts of octanoyl hydroxamic acid, 3-7 parts of 1,3-propanediol, 0-10 parts of the penetration enhancer, and 1-5 parts of the surfactant.

[0017] Preferably, the penetration enhancer is at least one of squalane and ceramide, and preferably the penetration enhancer is a mixture of squalane and ceramide in a mass ratio of 1-5:1.

[0018] Preferably, according to some embodiments of the present application, the surfactant is at least one of PEG-40 hydrogenated castor oil, Tween-20, Tween-60, and PEG-100 stearate.

[0019] The present application also provides a composition for sensitive skin repair, which is prepared according to the above method of the present application.

[0020] Preferably, the present application also provides the use of the composition for sensitive skin repair in the preparation of skin care products.

[0021] Preferably, the present application also provides a skin care product containing the composition for sensitive skin repair as described above.

[0022] Preferably, according to some embodiments of the present application, the skin care product further comprises an adjuvant acceptable in skin care products.

[0023] Preferably, according to some embodiments of the present application, the adjuvant comprises at least one of a chelating agent, a humectant, a thickening agent, an emulsifier, a fragrance modulator.

[0024] Preferably, according to some embodiments of the present application, the adjuvant comprises at least one of a chelating agent 0.05-0.5%, a humectant 1-10%, a thickening agent 0.5-2%, an emulsifier 1-5%, a fragrance modulator 0.1-0.5% by weight percentage.

[0025] Preferably, the chelating agent is selected from at least one of disodium ethylenediaminetetraacetate, trisodium ethylenediaminetetraacetate, tetrasodium ethylenediaminetetraacetate, trisodium ethylenediamine disuccinate, sodium citrate.

[0026] Preferably, the humectant comprises at least one of glycerin, sodium hyaluronate, nicotinamide, butylene glycol, sorbitol.

[0027] Preferably, the thickening agent is at least one of sodium polyacrylate, carboxymethyl cellulose, hydroxyethyl cellulose, and xanthan gum.

[0028] Preferably, the emulsifier is at least one of cocamidopropyl betaine, polysorbate-20, polysorbate-60, polyglyceryl-10 laurate, potassium lauryl phosphate.

[0029] Preferably, the fragrance modulator is selected from one or a combination of menthol, rose flower essential oil, jasmine flower essential oil, carnation flower essential oil, gardenia flower essential oil, violet flower essential oil.

[0030] Preferably, according to some embodiments of the present application, the composition for sensitive skin repair is added to the skin care product in an amount of 0.01-2wt%.

[0031] Preferably, according to some embodiments of the present application, the skin care product is one of an aqueous agent, an essence, an essence cream, a mask liquid, and a cream.

[0032] The core component of the Bletilla striata root extract is Bletilla striata polysaccharide. The present application uses the Bletilla striata root extract as a unique carbon source to perform low-temperature fermentation in the first stage by using Bifidobacterium; then, the temperature is raised and Thermus thermophilus is introduced to perform high-temperature fermentation in the second stage. Compared with the traditional Bifidobacterium fermentation product lysate (Saccharomyces boulardii fermentation lysate), the present application solves the key problems of the Saccharomyces boulardii fermentation lysate, such as poor stability, low permeability and strong irritation: (1) the double fermentation gradually decomposes the large molecular polysaccharide in the Bletilla striata root extract into small molecular active substances which are more easily permeable to the skin; (2) the high-temperature fermentation stage of the Thermus thermophilus not only inactivates the Bifidobacterium and harmful microorganisms in the first stage and terminates the continuous generation of organic acids, but also utilizes the Thermus thermophilus to metabolize part of the residual lactic acid, thereby reducing the irritation risk; (3) the Thermus thermophilus introduced in the second stage can produce heat-stable superoxide dismutase SOD, which can effectively scavenge free radicals and resist the damage to the skin caused by ultraviolet rays and environmental stress, thereby endowing the sensitive skin repair composition with stronger antioxidant and repair functions.

[0033] Further, for sensitive skin, the content of ceramides in the skin is often significantly lower than that of healthy skin, leading to barrier dysfunction, and the damaged barrier can be repaired by supplementing ceramides, thereby providing a stable penetration environment for the Saccharomyces boulardii product. The added squalane penetrant is a lipophilic component, which is similar in structure to the lipids (such as cholesterol and fatty acids) in the stratum corneum, can quickly integrate into the lipid bilayer of the stratum corneum, reduce the arrangement density of the lipid molecules, and reduce the physical resistance when the Saccharomyces boulardii fermentation product filtrate (containing water-soluble and fat-soluble active molecules) penetrates. The present application uses squalane and ceramides in a mass ratio of 1-5:1, which synergistically promotes the penetration efficiency of various small molecular active ingredients in the Saccharomyces boulardii product along the concentration gradient to the deep layer of the skin, thereby providing an important guarantee for repairing sensitive skin.

[0034] Further, the added octanoyl hydroxamic acid not only buffers the pH of the system, chelates harmful metal ions to inhibit chemical degradation, but also has antibacterial effect, and together with 1,2-pentanediol, it forms a broad-spectrum microbial defense network to maintain the structural integrity and activity of the sensitive skin repair composition in complex environments (such as temperature and pH fluctuations), thereby delaying the inactivation and improving the stability of the skin care product.

[0035] The advantages or beneficial effects of the composition for repairing sensitive skin of the present application at least include: The present application uses Bletilla striata root extract as a unique carbon source, uses Bifidobacterium for the first stage of low-temperature fermentation and Thermus thermophilus for the second stage of high-temperature fermentation, and the double fermentation process not only degrades macromolecular polysaccharides in Bletilla striata root extract into small molecule active substances in a step-by-step manner, but also inactivates miscellaneous bacteria and metabolizes residual organic acids, reduces LPS-induced pro-inflammatory factor mRNA expression to resist inflammation and soothe, promotes HaCaT cell migration to repair skin barrier, and has excellent mildness, penetration and stability. In skin care products, only 0.01-2wt% of the sensitive skin repair composition can achieve long-term moisturizing with 4-hour moisturizing rate ≥61.8% and DPPH free radical scavenging rate of up to 79.3%, effectively overcoming the defects of traditional Schizosaccharomyces fermentate such as poor stability, low penetration rate and strong irritation. BRIEF DESCRIPTION OF DRAWINGS

[0036] Figure 1 HaCaT cell migration ability of the sensitive skin repair composition of Example 1.

[0037] Figure 2 Microscopic view of the HaCaT cell migration ability of the sensitive skin repair composition of Example 1.

[0038] Figure 3 Effect of the sensitive skin repair composition of Example 1 on RAW264.7 cell activity.

[0039] Figure 4 Effect of the sensitive skin repair composition of Example 1 on LPS-induced RAW264.7 cell inflammatory factor mRNA expression. DETAILED DESCRIPTION

[0040] In order to more clearly illustrate the purpose, technical scheme and advantages of the present application, the following will be a detailed description of the technical scheme of the present application through specific examples. It should be pointed out that these examples are only used to illustrate the present application, and do not limit the scope of protection, the actual protection scope of the present application should be limited by the claims.

[0041] The materials, reagents and the like used in the following examples and comparative examples are commercially available reagents and materials unless otherwise specified. The amount of components in the following examples is 1g per weight part or per part unless otherwise specified.

[0042] 1. Experimental materials: Bifidobacterium, accession number: CGMCC 1.5014, purchased from China General Microbiological Culture Collection Center.

[0043] Thermus thermophilus, accession number: CGMCC 1.15099, purchased from China General Microbiological Culture Collection Center.

[0044] PEG-40 hydrogenated castor oil: CO40, CAS No. 61788-85-0, purchased from Prinova.

[0045] Squalane: Baisheme Biotechnology (Zhejiang) Co., Ltd.

[0046] Ceramide: Shenzhen Dickman Biotechnology Co., Ltd.

[0047] 1,2-pentanediol; purchased from Deqing Fragrance & Flavor (Nantong) Co., Ltd.

[0048] Suberohydroxamic acid: purchased from Guangdong Hainuokey Biological Medicine Co., Ltd.

[0049] 1,3-propanediol: purchased from Shandong Xinheng Chemical Co., Ltd.

[0050] Preparation of Bletilla striata root extract: Take dried Bletilla striata root, crush it through a 40-mesh sieve, weigh 100 g, add 10 times the mass of deionized water, reflux extract at 85°C for 3 h, extract 3 times, filter and concentrate to a solid content of 8wt%, to obtain Bletilla striata root extract, ready for use.

[0051] 2. Examples and comparative examples Example 1 S1, preparation of fermentation medium: mix 98wt% Bletilla striata root extract, 1.2wt% soybean peptone, and 0.8wt% potassium dihydrogen phosphate by weight percentage, and stir uniformly.

[0052] S2, first step fermentation: sterilize the medium at 121°C for 20 min, adjust the pH to 6.8 after cooling, inoculate 5% by volume of Bifidobacterium strain (concentration 5×10 9 CFU / mL), ferment at 37°C for 48 h to obtain primary fermentation broth.

[0053] S3, second step fermentation: warm the primary fermentation broth to 65°C, adjust the pH to 7.2, inoculate 1% by volume of Thermus thermophilus strain (concentration 5×10 7 CFU / mL), ferment at 50°C for 36 h to obtain secondary fermentation broth.

[0054] S4, post-treatment: break the secondary fermentation broth by high-pressure homogenization at 90 MPa, filter with a 2.0 μm polypropylene microfiltration membrane, and pasteurize at 90°C for 40 min, collect the filtrate to obtain the final fermentation broth.

[0055] S5, composition compounding: take 90 parts of final fermentation broth, 5 parts of 1,2-pentanediol, 0.1 part of suberohydroxamic acid, 5 parts of 1,3-propanediol, and 3 parts of Tween-60 by weight, stir and mix uniformly to obtain the sensitive skin repair composition.

[0056] Example 2 S1, fermentation medium preparation: mix 98.5wt% Bletilla striata root extract, 1wt% tryptone, 0.5wt% magnesium sulfate by weight percentage, stir uniformly.

[0057] S2, first step fermentation: sterilize the medium at 120℃ for 30min, adjust the pH to 6.5 after cooling, inoculate 3% volume of Bifidobacterium strain (concentration 1×10 8 CFU / mL), ferment at 35℃ for 72h to obtain primary fermentation broth.

[0058] S3, second step fermentation: warm the primary fermentation broth to 75℃, adjust the pH to 7.0, inoculate 0.5% volume of Thermus thermophilus strain (concentration 1×10 7 CFU / mL), ferment at 60℃ for 48h to obtain secondary fermentation broth.

[0059] S4, post-treatment: break the secondary fermentation broth by high pressure homogenization at 80MPa, filter with 1.5μm polypropylene microfiltration membrane, pasteurize at 85℃ for 60min, collect the filtrate to obtain the final fermentation broth.

[0060] S5, composition compounding: take 85 parts of final fermentation broth, 3 parts of 1,2-pentanediol, 0.05 parts of octanoyl hydroxamic acid, 3 parts of 1,3-propanediol, 2 parts of PEG-40 hydrogenated castor oil, stir and mix uniformly to obtain the sensitive skin repair composition.

[0061] Example 3 S1, fermentation medium preparation: mix 97.5wt% Bletilla striata root extract, 2wt% soybean peptone, 0.5wt% zinc sulfate by weight percentage, stir uniformly.

[0062] S2, first step fermentation: sterilize the medium at 121℃ for 25min, adjust the pH to 7.0 after cooling, inoculate 10% volume of Bifidobacterium strain (concentration 1×10 10 CFU / mL), ferment at 40℃ for 24h to obtain primary fermentation broth.

[0063] S3, second step fermentation: warm the primary fermentation broth to 70℃, adjust the pH to 7.5, inoculate 2% volume of Thermus thermophilus strain (concentration 1×10 8 CFU / mL), ferment at 55℃ for 24h to obtain secondary fermentation broth.

[0064] S4, post-treatment: break the secondary fermentation broth by high pressure homogenization at 100MPa, filter with 3.5μm polypropylene microfiltration membrane, pasteurize at 100℃ for 30min, collect the filtrate to obtain the final fermentation broth.

[0065] S5, composition compounding: taking 100 parts of final fermentation liquor, 10 parts of 1,2-pentanediol, 0.2 parts of octanoyl hydroxamic acid, 7 parts of 1,3-propanediol, 5 parts of PEG-100 stearate by weight, stirring and mixing uniformly to obtain a sensitive skin repair composition.

[0066] Example 4 The difference from Example 1 is that the amount of penetration enhancer (squalane: ceramide = 1:1) is 3 parts, and the remaining steps and parameters are consistent with Example 1.

[0067] Example 5 The difference from Example 1 is that the amount of penetration enhancer (squalane: ceramide = 3:1) is 5 parts, and the remaining steps and parameters are consistent with Example 1.

[0068] Example 6 The difference from Example 5 is that the penetration enhancer is 5 parts of squalane, and the remaining steps and parameters are consistent with Example 1.

[0069] Example 7 The difference from Example 5 is that the penetration enhancer is 5 parts of ceramide, and the remaining steps and parameters are consistent with Example 1.

[0070] Comparative Example 1 Compared with Example 1, Comparative Example 1 omits the second fermentation of Thermus thermophilus, and the remaining conditions are the same as Example 1.

[0071] Comparative Example 2 Compared with Example 1, Comparative Example 2 uses a sucrose solution with a solid content of 8wt% instead of white root extract, and the remaining conditions are the same as Example 1.

[0072] Comparative Example 3 Compared with Example 1, an equal amount of 1,2-pentanediol is used instead of octanoyl hydroxamic acid in Comparative Example 3, and the other conditions are the same as Example 1.

[0073] Comparative Example 4 Compared with Example 1, an equal amount of octanoyl hydroxamic acid is used instead of 1,2-pentanediol in Comparative Example 4, and the other conditions are the same as Example 1.

[0074] 3, Performance characterization of sensitive skin repair composition.

[0075] (1) Migration ability to HaCaT cells (I) Experimental conditions: Cell strain: HaCaT cells.

[0076] Density: 3*10 5 / ml.

[0077] Tested sample: sensitive skin repair raw material of Example 1.

[0078] Positive control: 8% FBS.

[0079] (II) Experimental method: logarithmic growth period HaCaT cells were taken to 12-hole plates, and after 24 h when the cells grew on the bottom, 200 μL of the gun head was used to gently scratch the bottom to simulate human skin damage; the culture medium was aspirated and washed with PBS to remove the cells; then the culture medium containing different concentrations of the tested sample was added, and a blank control group and a positive control group were set; microscope photography was performed at 0 h, 12 h and 24 h, respectively, and then the wound area was planned with Image J software and the healing rate was calculated.

[0080] The test results are shown in Table 1: Figures 1-2 , wherein: *: P < 0.05 compared with the blank group; **: P < 0.01 compared with the blank group; ***: P < 0.001 compared with the blank group; ***: P < 0.0001 compared with the blank group. Figure 1 From left to right are the blank group, 0.0625wt% repair raw material, 0.125wt% repair raw material and the positive control group.

[0081] According to the test results, the sensitive skin repair raw material has a significant increase in the cell scratch healing area at 12 h and 24 h after scratching, and has a significant difference compared with the blank group. Figures 1-2

[0082] In summary, according to the test results of the migration ability of HaCaT, the sensitive skin repair composition of the present application has the comprehensive effects of repairing skin barrier, promoting skin renewal and healing, etc.

[0083] (2) Effect on RAW264.7 cell activity and effect on LPS-induced RAW264.7 cell inflammatory factor mRNA expression.

[0084] A: Effect on RAW264.7 cell activity.

[0085] (I) Experimental conditions Tested sample: sensitive skin repair raw material of Example 1.

[0086] Action time: 24 h.

[0087] Detection method: CCK 8.

[0088] (II) Experimental method: RAW264.7 cells were collected by centrifugation, and the cell density was adjusted to 1×10 5 ​ / mL, inoculated in 96-well culture plates, and placed in a cell culture incubator for overnight culture. After the RAW264.7 cells adhered, the drugs were diluted with serum-free basal medium, and the stimulant LPS was added to a final concentration of 100 ng / mL. After 24 h of culture, CCK8 reagent was added to detect the activity of the cells.

[0089] The test results are shown in Table 1. Figure 3 As shown in Table 1, the results show that the sensitive skin repair raw material has a cell activity of greater than 80% at a concentration of 2.0% or less, and a cell activity close to 80% at a concentration of 2.0%, which can be used as a non-toxic concentration of the sensitive skin repair raw material.

[0090] B: Effect on the mRNA expression of inflammatory factors of LPS-induced RAW264.7 cells.

[0091] (I) Experimental conditions Test sample: sensitive skin repair raw material of Example 1.

[0092] Positive control: 10 μM dexamethasone.

[0093] Cell density: 2.5 x 10 5 / mL; 12-well plate.

[0094] (II) Experimental method: RAW264.7 cells were collected by centrifugation, and the cell density was adjusted to 2.5 x 10 5 / mL. The cells were inoculated in 12-well culture plates and placed in a cell culture incubator for overnight culture. After the RAW264.7 cells adhered, the drugs were diluted with serum-free basal medium, and the stimulant LPS was added to a final concentration of 100 ng / mL. After 6 h of culture, the expression levels of inflammatory factor mRNA in the RAW264.7 cells were detected by qPCR method.

[0095] The test results are shown in Table 2. Figure 4 As shown in Table 2, the sensitive skin repair raw material at 0.125, 0.25, and 1.0 mg / mL can effectively reduce the expression levels of IL-6, IL-1β, and TNF-α mRNA in LPS-induced RAW264.7 cells, and the effect is stronger as the concentration of the sensitive skin repair raw material increases. Thus, the sensitive skin repair raw material of the present application has the effect of anti-inflammation and soothing, and can be used in skin care products for sensitive skin repair.

[0096] In summary, according to the test results of the activity of RAW264.7 cells and the mRNA expression of inflammatory factors of LPS-induced RAW264.7 cells, the sensitive skin repair raw material of the present application has important effects of anti-inflammation and soothing, and is very suitable for sensitive skin care for sensitive, red, and pimple inflammation.

[0097] 4. Application Example The application of the composition for repairing sensitive skin in the essence milk, the mass percentage of each component in the essence milk is shown in Table 1, and the preparation method comprises the following steps: (1) accurately weigh each raw material component according to the formulation amount in Table 1; (2) uniformly stir glycerin, nicotinamide, xanthan gum and deionized water at 50°C to obtain phase A; (3) uniformly stir sodium citrate and polysorbate-20 at 80°C to obtain phase B; (4) add phase B to phase A, emulsify and homogenize, uniformly stir the composition for repairing sensitive skin and jasmine essential oil to obtain the essence milk.

[0098] Table 1: mass percentage of each component in application examples 1-5 is as follows:

[0099] Among them, application examples 1-5 respectively use the sensitive skin repair composition described in examples 1-5.

[0100] Application examples 6-7: replace the sensitive skin repair composition in application example 5 with the sensitive skin repair composition in examples 6-7 respectively, and the others are the same as application example 5.

[0101] Comparative application examples 1-4 Compared with application example 1, in comparative application examples 1-4, the sensitive skin repair composition of example 1 is respectively replaced with the sensitive skin repair composition provided in comparative examples 1-4, and the composition, amount and preparation method of other raw materials are the same as those in application example 1.

[0102] 5. Performance evaluation of essence milk containing the sensitive skin repair composition of the application.

[0103] (1) Stability evaluation Cold and hot resistance stability: 100mL of essence milk is placed in a 250mL transparent glass bottle, and is placed at 50°C for 24h, and then is placed at-10°C for 24h, and the cycle is repeated for 15 times, for a total of 30 days, and the layering and oil-water separation are observed.

[0104] Normal temperature stability: 100mL of essence milk is placed in a 250mL transparent glass bottle, and is placed at 25°C for 30 days, and the layering and oil-water separation are observed.

[0105] Among them, the grade without layering and oil-water separation is A; the grade with a small amount of layering or oil-water separation is B; and the grade with a large amount of layering or oil-water separation is C. The test results are shown in Table 2 below.

[0106] Table 2: stability test results

[0107] From the above test results, the normal temperature stability and cold and heat cycle stability of the essence cream of the present application are good, there is no stratification and no oil and water separation, the stability of comparative application examples 3-4 using only one of octanoyl hydroxamic acid or 1,2-pentanediol at normal temperature and after cold and heat cycle is significantly decreased, and obvious stratification phenomenon occurs. It is proved that the combination of octanoyl hydroxamic acid and 1,2-pentanediol can synergistically improve the stability of the essence cream.

[0108] (2) Transdermal penetration experiment of sample The fermentation product is a complex mixture, and the penetration efficiency of the whole fermentation product is indirectly represented by measuring the transdermal absorption amount of its key functional components. Small molecule peptides (<1000 Da) are one of the core functional components for repair and anti-photoaging. High performance liquid chromatography is used to test small molecule peptides below 1000 Da to represent the penetration rate of the Schizochytrium ferment product.

[0109] Pigskin is used to simulate human epidermal skin, Franz diffusion cell is used as a static diffusion cell, the essence cream to be tested is used as the transdermal release liquid, and the transdermal absorption test is carried out under the condition of constant temperature 37℃ and constant speed 600r / min. High performance liquid chromatography is used to determine the small molecule peptides below 1000 Da in the sample.

[0110] The penetration rate of small molecule peptides below 1000 Da=(the amount of small molecule peptides below 1000 Da in the receiving pool / the amount of small molecule peptides below 1000 Da in the essence liquid)×100%.

[0111] Table 3: Results of in vitro transdermal absorption test

[0112] From the test results in Table 3, it can be found that the in vitro transdermal penetration amount of the essence cream prepared in the present application is significantly higher than that of the comparative examples. And comparative application examples 4-5 use the combination of squalane and ceramide, the penetration of the essence cream in the skin is significantly higher than that of example 1, which indicates that the use of ceramide and squalane in a certain ratio in the present application can synergistically promote the penetration of active ingredients in the Schizochytrium ferment product, which will help to improve the repair performance of the skin. Comparative application example 1 does not use thermophilic thermophilic bacteria fermentation, and comparative application example 2 uses ordinary sucrose as a carbon source, and the small molecule peptide penetration rate decreases.

[0113] (3) Skin patch test Healthy volunteers aged 35-45 years old are selected and randomly divided into 12 groups, 20 people in each group. Using a patch tester, in a closed patch test mode, 0.02g of the essence cream is taken in the patch tester and applied to the back of the test subject. After 24h, the patch tester is removed, and after 60min, the skin appearance of the volunteers is evaluated.

[0114] Evaluation criteria: Grade 0: negative reaction; Grade 1: suspicious reaction, only weak erythema; Grade 2: weak positive reaction, erythema, infiltration, edema, and possibly papules; Grade 3: strong positive reaction, erythema, infiltration, edema, and possibly papules, and the reaction can exceed the test area; Grade 4: extremely strong positive reaction, obvious erythema, severe infiltration, edema, and confluent blisters, and the reaction exceeds the test area.

[0115] Table 4: Results of skin patch test

[0116] According to the skin patch test results in Table 4, the essence cream provided by the embodiments of the present application is mild and non-irritating to the skin; according to the comparison between Application Example 1 and Comparative Application Example 1, the high-temperature fermentation stage of the thermophilic thermus can not only inactivate the bifidobacterium and harmful microorganisms in the first stage and terminate the continuous generation of organic acids, but also utilize the metabolic part of lactic acid residues to reduce the risk of irritation and reduce symptoms such as erythema.

[0117] (4) Long-term moisturizing performance determination Select healthy female volunteers aged 35-55 years old to use the essence cream provided by the application example and the comparative application example, respectively, 6 people in each group, and use a skin moisture tester (Corneometer CM825) to quantitatively detect the water content of facial skin at 0 h, 1 h, and 4 h to evaluate the moisturizing ability of the essence cream. The water content and the moisturizing rate are the average values of each group of subjects. The moisturizing rate after 1 h / % = the water content at 1 h / the water content at 0 h*100; the moisturizing rate after 4 h / % = the water content at 4 h / the water content at 0 h*100.

[0118] Table 5: Moisturizing performance test

[0119] According to the test data in Table 5, the skin moisturizing rates of the essence creams of the application example and the comparative application example after 1 hour of use are similar, but after 4 hours of use, the emulsion of the application example can keep the skin water content stable at more than 61.8%; and the application examples 5-6 use squalane and ceramide combination as a penetrating agent, which is conducive to the penetration of active ingredients in the skin, improving the long-term moisturizing performance of the essence cream, while the moisturizing performance of the essence cream of the comparative application example decreases slightly, especially the moisturizing rate of the comparative application example 1 decreases significantly, which may be that the double fermentation gradually decomposes the large molecular polysaccharides in the radix bletillae extract into small molecular active substances that are more easily penetrated into the skin, thereby improving the long-term moisturizing rate.

[0120] (5) DPPH free radical scavenging activity determination DPPH solution (1,1-diphenyl-2-trinitrophenylhydrazine) was prepared with anhydrous ethanol; 2 g of the essence cream and 2 mL of the DPPH solution were placed in a test tube, and the test tube was placed in a thermostat at 37°C for 3 h; the blank control was anhydrous ethanol; the absorbance A1 of each sample to be tested was measured at 517 nm; and the free radical scavenging rate was calculated according to the following formula: free radical scavenging rate = [1-(A1-A2) / A3] x 100%. Wherein, A3: the absorbance of 2 g of anhydrous ethanol and 2 mL of the DPPH solution; A1: the absorbance of 2 g of the sample to be tested and 2 mL of the DPPH solution; A2: the absorbance of 2 g of the sample to be tested and 2 g of anhydrous ethanol; three sets of parallel samples were measured for each sample to be tested, and the average value was obtained.

[0121] Table 6: Antioxidant performance test

[0122] According to the test results in Table 6, the DPPH scavenging rates of the comparative application examples 1 were lower than that of the application example 1. The comparative application examples 1 did not use the Thermus thermophilus fermentation, and failed to produce heat-stable superoxide dismutase SOD, which reduced the free radical scavenging and antioxidant properties, and thus the DPPH scavenging rate of the comparative application examples 1 was lower than that of the application example 1.

[0123] The above examples are only provided for illustrating the present application, and are not intended to limit the possible embodiments of the present application. Based on the disclosure of the present application, those skilled in the art can make various modifications and adjustments. It is not necessary or possible to list all possible embodiments. Any modification, equivalent replacement or improvement made within the basic principles and scope of the present application shall be considered as falling within the protection scope of the present application.

Claims

1. A method for preparing a composition for repairing sensitive skin, characterized in that, Includes the following steps: S1. Mix Bletilla striata root extract, nitrogen source and inorganic salt to obtain fermentation medium; S2, First step of fermentation: Sterilize the fermentation medium, adjust the pH to 6.5-7.0, inoculate 3-10% of Bifidobacterium inoculum for fermentation, and obtain the primary fermentation broth of Bifida Ferment Lysate; S3. Second fermentation: Heat the primary fermentation broth to 60-80℃, adjust the pH to 7.0-7.5, and inoculate with 0.5-2% of thermophilic bacteria to ferment and obtain the secondary fermentation broth of Bifida ferment lysate. S4. Post-processing: The secondary fermentation broth is homogenized and crushed under high pressure at 80-100MPa, filtered, sterilized, and the filtrate is collected to obtain the final fermentation broth of Bifida ferment lysate. S5. Mix the final fermentation broth of Bifida Ferment Lysate, 1,2-pentanediol, octanoyl hydroxamic acid, 1,3-propanediol, penetration enhancer, and surfactant to obtain the sensitive skin repair composition.

2. The method for preparing the sensitive skin repair composition as described in claim 1, characterized in that, The preparation of Bletilla striata root extract in step S1 includes the following steps: take dried Bletilla striata root, crush it through a 30-80 mesh sieve, add 10-20 times the mass of deionized water, reflux extract at 80-90℃ for 2-5 hours, filter and concentrate the filtrate to a solid content of 5-15wt% to obtain Bletilla striata root extract.

3. The method for preparing the sensitive skin repair composition as described in claim 1, characterized in that, The culture medium in step S1 contains 97-99.4 wt% Bletilla striata root extract, 0.5-2 wt% nitrogen source and 0.1-1 wt% inorganic salts; the nitrogen source in step S1 includes at least one of soybean peptone and tryptone; the inorganic salts include at least one of potassium dihydrogen phosphate, magnesium sulfate and zinc sulfate.

4. The method for preparing the sensitive skin repair composition according to any one of claims 1-3, characterized in that, The fermentation temperature in step S2 is 30-40℃, and the fermentation time is 24-72 hours; the concentration of Bifidobacterium is 1×10⁻⁶. 8 ~1×10 10 CFU / mL; the fermentation temperature in step S3 is 50-60℃, and the fermentation time is 24-48h; the concentration of the thermophilic bacteria is 1×10⁻⁶. 7 ~1×10 8 CFU / mL.

5. The method for preparing the sensitive skin repair composition according to any one of claims 1-3, characterized in that, The filtration in step S4 is performed using a 1.5μm to 3.5μm polypropylene microfiltration membrane; the sterilization in step S4 is low-temperature pasteurization, with a sterilization temperature of 80 to 100℃ and a time of 30 to 60 minutes.

6. The method for preparing the sensitive skin repair composition according to any one of claims 1-3, characterized in that, The sensitive skin repair composition in step S5 comprises 80-100 parts of final fermentation broth, 3-10 parts of 1,2-pentanediol, 0.01-0.2 parts of capryloyl hydroxamic acid, 3-7 parts of 1,3-propanediol, 0-10 parts of penetration enhancer, and 1-5 parts of surfactant; preferably, the penetration enhancer is at least one of squalane and ceramide; preferably, the surfactant is at least one of PEG-40 hydrogenated castor oil, Tween-20, Tween-60, and PEG-100 stearate.

7. A composition for repairing sensitive skin, characterized in that, Prepared by the method according to any one of claims 1-6.

8. A skincare product, characterized in that, The skincare product contains the composition for sensitive skin repair of claim 7; preferably, the composition for sensitive skin repair is added to the skincare product in an amount of 0.1-2 wt%; preferably, the skincare product further includes acceptable excipients; the excipients include at least one of chelating agents, moisturizers, thickeners, emulsifiers, and fragrance modifiers.

9. The skincare product as described in claim 8, characterized in that, The excipients, by weight percentage, include at least one of the following: chelating agent 0.05-0.5%, humectant 1-10%, thickener 0.5-2%, emulsifier 1-5%, and fragrance modifier 0.1-0.5%. Preferably, the chelating agent is at least one of disodium ethylenediaminetetraacetate, tetrasodium ethylenediaminetetraacetate, trisodium ethylenediaminedisuccinate, and sodium citrate; the humectant is at least one of glycerin, sodium hyaluronate, nicotinamide, butylene glycol, and sorbitol; the thickener is at least one of sodium polyacrylate, carboxymethyl cellulose, hydroxyethyl cellulose, and xanthan gum; the emulsifier is at least one of cocamidopropyl betaine, polysorbate-20, polysorbate-60, polyglycerol-10 stearate, and potassium lauryl phosphate; and the fragrance modifier is one or a combination of menthol, rose essential oil, jasmine essential oil, carnation essential oil, gardenia essential oil, and violet essential oil.

10. The skincare product according to any one of claims 8-9, characterized in that, The skincare product is at least one of the following: water-based, serum, essence lotion, facial mask liquid, and cream.

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