Copper peptide high-efficiency soothing composition and application thereof

Abstract

The application provides a copper peptide efficient soothing composition and application thereof, which comprises copper peptide and symbiotic fermentation products of thermus thermophilus and bacillus, can target stimulation of secretion of inflammatory mediators, effectively inhibits secretion of PGE2 and expression of inflammatory factors IL-6, TNF alpha and NO, and has a synergistic effect, especially on secretion of PGE2 and expression of NO, and shows a more significant soothing effect. The composition is used as an efficacy component to prepare skin care products, is particularly suitable for people with sensitive skin, can help the skin to restore a healthy state, and has a good application prospect.

Description

Technical Field

[0001] This invention relates to the field of skincare technology, specifically to a copper peptide highly effective soothing composition and its application. Background Technology

[0002] Sensitive skin is a highly sensitive skin condition where the skin is easily irritated by various external physical and chemical factors, such as climate change, seasonal changes, sun exposure, spicy food consumption, or even without any obvious trigger, resulting in sensations like stinging, burning, tightness, and itching. With societal progress and the increasing pursuit of beauty, more and more people are paying attention to their skin health and quality of life, making sensitive skin a more prominent issue. As a highly prevalent skin problem in modern society, it significantly impacts people's quality of life and has therefore received widespread attention from dermatologists and the general public. Consequently, developing cosmetics suitable for the physiological characteristics of sensitive skin has become a research hotspot.

[0003] Copper peptides (Tripeptide-1 Copper, GHK-Cu) are complexes formed by the combination of small molecule peptides and divalent copper ions. Their composition is glycine-histidine-lysine-copper, and at certain concentrations, they possess multiple functions, including promoting wound healing, anti-inflammation, anti-oxidation, and promoting collagen regeneration. In the cosmetics field, GHK-Cu can be directly used as a ready-made raw material in products such as serums, freeze-dried powders, and creams. However, GHK-Cu complexes are expensive, and their addition in cosmetics is limited by cost. Furthermore, they are prone to hydrolysis under oxidative conditions, and acidic substances can easily release the copper ions from GHK-Cu, significantly reducing the skincare efficacy of the copper peptides. Therefore, providing a highly effective copper peptide composition that balances cost and efficacy has become an urgent problem to be solved.

[0004] The main cause of skin sensitivity and allergies is the secretion and release of inflammatory factors in the skin. Among the many inflammatory cytokines, the main ones that play a role are TNF-α, IL-8, IL-1β, IL-6, PGE2, and NO. Among these, PGE2, IL-6, TNF-α, and NO are most closely related to soothing the skin.

[0005] Therefore, there is an urgent need to find a skincare product that can effectively inhibit inflammatory factors, soothe the skin, and meet the needs of patients with sensitive skin. Summary of the Invention

[0006] To address the above problems, this invention provides a highly effective soothing composition containing copper peptides and its applications. The composition includes copper peptides and symbiotic fermentation products of *Thermophilus thermophilus* and *Bacillus*. This composition can target and stimulate the secretion of inflammatory mediators, effectively inhibiting the secretion of PGE2 and the expression of inflammatory factors IL-6, TNFα, and NO, and exhibits a synergistic effect, particularly in its effect on PGE2 secretion and NO expression, demonstrating a more significant soothing effect. This composition, when used as an active ingredient in skincare products, is especially suitable for people with sensitive skin, helping the skin restore a healthy state and showing promising application prospects.

[0007] The present invention provides a skin-soothing composition comprising copper peptides and symbiotic fermentation products of thermophilic bacteria and Bacillus.

[0008] The thermophilic bacteria and Bacillus co-fermentation product (TIRACLE Pro) is rich in amino acids, polypeptides, polyphenols and trace elements that maintain skin nutrition. Its preparation method is as follows: (1) The thermophilic bacteria are subjected to one or more enrichment fermentations at 65°C and pH 7.8±0.2 to allow them to be in the exponential growth phase; (2) Bacillus is added and secondary fermentation is carried out at pH 7.0-7.2 and temperature 30±2°C. The fermentation liquid is centrifuged to remove bacteria.

[0009] Extensive research has demonstrated that the copper peptide soothing composition, combining copper peptide and TIRACLE Pro, effectively inhibits the expression of PGE2, IL-6, TNFα, and NO. Compared to using copper peptide or TIRACLE Pro alone, this highly effective copper peptide soothing composition exhibits a synergistic effect, particularly in inhibiting PGE2 secretion and NO expression, resulting in a more significant soothing effect. Studies have shown that the copper peptide soothing composition has significant soothing efficacy. Using this active ingredient, combined with cosmetically acceptable carriers and prepared using conventional methods, various dosage forms of cosmetics can be formulated to meet the urgent needs of patients with sensitive skin.

[0010] Furthermore, in the composition, the content of copper peptide is 0-0.005%, and the content of symbiotic fermentation products of thermophilic bacteria and Bacillus is 0-5%.

[0011] Furthermore, the content of the copper peptide is 0.005%, and the content of the symbiotic fermentation product of thermophilic bacteria and Bacillus is 5%.

[0012] On the other hand, the present invention provides the use of a composition in the preparation of a skin-soothing skin care product, the composition soothing the skin by inhibiting the expression of any one or more inflammatory factors among PGE2, IL-6, TNFα, or NO; the composition includes a copper peptide and a symbiotic fermentation product of Thermophilic bacteria and Bacillus.

[0013] Furthermore, in the composition, the content of copper peptide is 0-0.005%, and the content of symbiotic fermentation products of thermophilic bacteria and Bacillus is 0-5%.

[0014] Furthermore, the content of the copper peptide is 0.005%, and the content of the symbiotic fermentation product of thermophilic bacteria and Bacillus is 5%.

[0015] In another aspect, the present invention provides the use of copper peptides in the preparation of formulations that inhibit the expression of any one or more inflammatory factors among PGE2, IL-6, TNFα, or NO.

[0016] Furthermore, the content of the copper peptide is 0 to 0.005%.

[0017] Furthermore, the content of the copper peptide is 0.005%.

[0018] In another aspect, the present invention provides the use of the symbiotic fermentation product of thermophilic bacteria and Bacillus in a formulation that inhibits the expression of any one or more inflammatory factors among PGE2, IL-6, TNFα, or NO.

[0019] Furthermore, the content of the symbiotic fermentation product of the thermophilic bacteria and Bacillus is 1-5%.

[0020] In another aspect, the present invention provides the use of the symbiotic fermentation products of thermophilic bacteria and Bacillus in formulations that inhibit the expression of inflammatory factors PGE2 or NO.

[0021] Furthermore, the content of the symbiotic fermentation product of the thermophilic bacteria and Bacillus is 5%.

[0022] In another aspect, the present invention provides the use of the symbiotic fermentation product of Thermophilus thermophilus and Bacillus spp. in the preparation of a formulation that inhibits the expression of inflammatory factors IL-6 or TNFα, wherein the content of the symbiotic fermentation product of Thermophilus thermophilus and Bacillus spp. is 1-5%.

[0023] In another aspect, the present invention provides the use of a composition in the preparation of an agent that inhibits the expression of any one or more inflammatory factors among PGE2, IL-6, TNFα, or NO, said composition comprising a copper peptide and a symbiotic fermentation product of Thermophilus and Bacillus.

[0024] Furthermore, the content of the copper peptide is 0.005%, and the content of the symbiotic fermentation product of thermophilic bacteria and Bacillus is 5%.

[0025] The skin-soothing composition provided by this invention, in addition to including copper peptides, thermophilic bacteria, and Bacillus spp. symbiotic fermentation products, may, without compromising the effectiveness of this invention, be appropriately combined with various ingredients commonly used in skincare and pharmaceutical fields, such as powder components, moisturizers, emulsifiers, light stabilizers, thickeners, solubilizers, metal ion masking agents, pigments, pH adjusters, skin nutrients, vitamins, preservatives, antioxidants, sunscreens, penetration enhancers, liposomes, skin nutrient components, antioxidant auxiliaries, fragrances, perfumes, and pigments, or a mixture of several of these.

[0026] The beneficial effects of this invention are as follows:

[0027] 1. A composition with a highly effective skin-soothing effect is provided;

[0028] 2. The role of copper peptides in inhibiting inflammatory factors PGE2, IL-6, TNFα, and NO was discovered;

[0029] 3. The role of symbiotic fermentation products of thermophilic bacteria and Bacillus in inhibiting inflammatory factors PGE2, IL-6, TNFα, and NO was discovered;

[0030] 4. A synergistic effect was discovered in the inhibition of inflammatory factors PGE2, IL-6, TNFα, and NO by a combination of copper peptides, thermophilic bacteria, and Bacillus thuringiensis fermentation products.

[0031] 5. Skin care products prepared using this composition are especially suitable for people with sensitive skin, and can help the skin restore its healthy state, with broad application prospects. Attached Figure Description

[0032] Figure 1 This is a schematic diagram showing the effect of copper peptide on the activity of macrophages Raw264.7 in Example 3;

[0033] Figure 2 This is a schematic diagram showing the effect of TIRACLE Pro on the viability of macrophages Raw264.7 in Example 3;

[0034] Figure 3 This is a schematic diagram showing the effect of TIRACLE Pro and the compound composition on the activity of macrophages Raw264.7 in Example 3;

[0035] Figure 4 This is a schematic diagram showing the effect of the test samples in Example 4 on the expression of PEG2 secreted by Raw264.7;

[0036] Figure 5 This is a schematic diagram showing the effects of samples 5, 1, and 3 on the expression of PEG2 secreted by Raw264.7 in Example 4;

[0037] Figure 6 This is a schematic diagram showing the effect of the test samples on the NO expression secreted by Raw264.7 in Example 5;

[0038] Figure 7 This is a schematic diagram showing the effects of samples 5, 1, and 3 in Example 5 on NO expression secreted by Raw264.7.

[0039] Figure 8 This is a schematic diagram showing the effect of the test samples in Example 6 on the expression of IL-6 secreted by Raw264.7.

[0040] Figure 9 This is a schematic diagram showing the effect of the test samples in Example 7 on the expression of TNF-α secreted by Raw264.7. Detailed Implementation

[0041] The present invention will be further described in detail below with reference to the embodiments. It should be noted that the embodiments described below are intended to facilitate the understanding of the present invention and do not limit it in any way.

[0042] Example 1: Preparation of symbiotic fermentation products of thermophilic bacteria and Bacillus.

[0043] The thermophilic bacteria strain used in this embodiment has the accession number BAA-163 and was purchased from the American Type Culture Collection Center; the Bacillus subtilis subsp. Subtilis has the accession number CICC10023 and was purchased from the China Industrial Microbial Culture Collection Center.

[0044] The basic fermentation medium formula is as follows: 10L of medium contains 30g peptone, 2g malt extract, 3.5g anhydrous magnesium sulfate, 1.2g potassium dihydrogen phosphate, 20g ammonium sulfate, 0.5g anhydrous ferric chloride, and 2.5g sodium chloride, with water added to a final volume of 10L. Different volumes of medium can be scaled up or down according to the formula. Seed culture is inoculated into the medium at an inoculum volume of 3-10%.

[0045] All embodiments of this invention use the culture medium formulations with the above proportions as the culture medium for bacterial fermentation. Of course, in actual industrial production, the proportions can be adjusted according to different requirements; these formulations are all common formulations for culturing thermophilic bacteria / Bacillus.

[0046] The symbiotic fermentation culture process is as follows:

[0047] 1. Seed culture of Thermophilic thermophilus

[0048] On a clean bench, 2 ml of cryopreservation solution of Thermophilus thermophilus BAA-163 was inoculated into a 2.5 L Erlenmeyer flask containing 1 L of culture medium with a pH of 7.8±0.2. The flask was incubated at 150 rpm and 65 °C for 12 h to obtain 1 L of Thermophilus thermophilus seed culture in the exponential growth phase.

[0049] 2. Add Bacillus subtilis and carry out secondary fermentation at a pH of 7.0-7.2 and a fermentation temperature of 30±2℃. Centrifuge the fermentation broth to remove bacteria.

[0050] Example 2: Preparation of a combination of copper peptide and TIRACLE Pro

[0051] The fermentation product (TIRACLEPro) of thermophilic bacteria and Bacillus spp. was prepared according to the method provided in Example 1.

[0052] A compound composition was obtained by mixing copper peptide (purchased from Jitai Biotechnology) with TIRACLE Pro, wherein the content of copper peptide was 0-0.005% and the content of TIRACLE Pro was 0-5%.

[0053] Example 3: Experiment on the effect of test samples on the viability of Raw264.7 macrophage cells.

[0054] (Raw264.7 mouse macrophage cell line (Guangdong Boxi Biotechnology Co., Ltd.) at 9×10 3 Cells were seeded at a density of cells / well into 96-well plates and incubated overnight in an incubator (37°C, 5% CO2). The experiment included a zeroing group, a solvent control group, a positive control group, and a sample group. Drug administration was performed when the cell deposition rate in the 96-well plates reached 40%–60%. 200 μL of culture medium was added to each well of the solvent control group; 200 μL of culture medium containing 10% DMSO (from Sigma) was added to each well of the positive control group; 200 μL of culture medium containing the corresponding concentration of the sample was added to each well of the sample group; and no cells were seeded in the zeroing group, only 200 μL of cell culture medium was added. Each group was supplemented with copper peptide, TIRACLE Pro (provided in Example 1), or a combination of copper peptide and TIRACLE Pro (provided in Example 2). Eight concentration gradients were set up for each sample, with three replicate wells for each concentration gradient. After drug administration, the 96-well plates were incubated in an incubator (37°C, 5% CO2) for 24 h. After 24 hours of cell incubation, the supernatant was discarded, and MTT working solution (purchased from Sigma) was added. The cells were incubated at 37°C in the dark for 4 hours. After incubation, the supernatant was discarded, and 150 μL of DMSO was added to each well. OD values ​​were read at 490 nm. The results are shown below. Figure 1 , 2 As shown in Figure 3, where Figure 1 This is a schematic diagram showing the effect of copper peptide on the activity of macrophages Raw264.7. Figure 2 A schematic diagram showing the effect of TIRACLE Pro on the viability of macrophages Raw264.7; Figure 3 A schematic diagram showing the effect of TIRACLE Pro and its compound composition on the viability of macrophages Raw264.7.

[0055] Depend on Figures 1-3 As can be seen from the MTT results, the copper peptide sample did not show significant cytotoxicity in the concentration range of 0.08% (g / mL); the TIRACLE Pro sample did not show significant cytotoxicity in the concentration range of 5% (v / v); and the combination of copper peptide and TIRACLE Pro did not show significant cytotoxicity in the concentration ranges of 0.005% (g / mL) copper peptide and 5% (v / v) TIRACLE Pro, respectively.

[0056] Example 4: Testing the expression of the inflammatory cytokine PEG2 secreted by macrophages.

[0057] Raw264.7 mouse macrophage cell line was cultured normally at 37°C in DMEM medium containing 5% CO2 and 10% FBS. When the cells reached 90% confluence, they were cultured at a density of 6.4 × 10⁻⁶ cells / year. 5 Cells were seeded at a density of 1.8 mL / ml in 6-well plates and incubated overnight in an incubator (37°C, 5% CO2). When the cell layering rate reached 40%–60%, the cells were grouped according to Table 1, with 1.8 mL of drug administered to each well. Each group had 3 replicates. The cells were incubated for another 2 hours in an incubator (37°C, 5% CO2). After two hours of incubation, 200 μL of LPS working solution (purchased from Sigma) prepared from the corresponding test substance working solution was added to the drug-treated wells. The wells were shaken left and right to mix the drug, with a final LPS concentration of 1 μg / mL. The cells were incubated for another 22 hours in an incubator (37°C, 5% CO2). After incubation, the cell culture supernatant was collected in EP tubes and detected using a PGE2 ELISA kit. The detection results are shown in Tables 1 and 2. Figure 4 , Figure 5 As shown, where Figure 4 This is a schematic diagram showing the effect of the test sample on the expression of PEG2 secreted by Raw264.7. Figure 5 for Figure 4 A schematic diagram showing the effects of ingredients 5, 1, and 3 on the secretion of PEG2 expression in Raw264.7.

[0058] Table 1. Effects of test samples on PEG2 expression secreted by Raw264.7

[0059]

[0060] Table 2. Effects of test samples on PEG2 expression secreted by Raw264.7

[0061]

[0062] According to Table 1 and Figure 4 The experimental results showed that, compared with the BC group, the secretion of the inflammatory mediator PGE2 by macrophages in the NC group was significantly increased, indicating that the LPS stimulation condition was effective in this experiment. Compared with the NC group, the secretion of the inflammatory mediator PGE2 by macrophages at a drug concentration of 100 μg / mL was significantly reduced, indicating that this experiment was effective.

[0063] Compared with the NC group, sample group 1 showed a slightly weaker inhibitory effect on the secretion of the inflammatory mediator PGE2, indicating that 0.005% copper peptide has a certain inhibitory effect on the secretion of the inflammatory mediator PGE2.

[0064] Compared with the NC group, sample group 2 could not inhibit the secretion of the inflammatory mediator PGE2, but the secretion of the inflammatory mediator PGE2 in sample group 3 was significantly reduced, indicating that 1% TIRACLE PRO was not enough to inhibit the secretion of the inflammatory mediator PGE2, while 5% TIRACLE PRO could significantly and effectively inhibit the secretion of the inflammatory mediator PGE2.

[0065] From Table 2 and Figure 5 A comprehensive comparison of the results of samples 5, 1, and 3 shows that the combination of 0.005% copper peptide and 5% TIRACLE PRO has a synergistic effect on inhibiting the secretion of the inflammatory mediator PGE2, reducing the average concentration of the inflammatory mediator PGE2 to 1309.51 pg / mL, a decrease of 92% compared to sample 1 and a decrease of 52% compared to sample 3, demonstrating a synergistic effect on inhibiting the secretion of the inflammatory mediator PGE2.

[0066] Example 5: Effect of test samples on the expression of the inflammatory cytokine NO secreted by macrophages

[0067] Raw264.7 mouse macrophage cell line was cultured normally at 37°C in DMEM medium containing 5% CO2 and 10% FBS. When the cells reached 90% confluence, they were cultured at a density of 6.4 × 10⁻⁶ cells / year. 5Cells were seeded at a density of 1.8 mL / ml in 6-well plates and incubated overnight in an incubator (37℃, 5% CO2). When the cell layering rate reached 40%–60%, the cells were grouped according to Table 3, with 1.8 mL of drug administered to each well. Each group had 3 replicates. The cells were incubated for another 2 hours in an incubator (37℃, 5% CO2). After two hours of incubation, 200 μL of LPS working solution prepared from the corresponding test substance working solution was added to the drug-treated wells. The wells were shaken left and right to mix the drug. The final LPS concentration was 1 μg / mL. The cells were incubated for another 22 hours in an incubator (37℃, 5% CO2). After incubation, the cell culture supernatant was collected in EP tubes and detected using a NO detection kit. The detection results are shown in Tables 3 and 4. Figure 6 , Figure 7 As shown, where Figure 6 This is a schematic diagram showing the effect of the test sample on NO expression secreted by Raw264.7. Figure 7 for Figure 6 A schematic diagram showing the effects of markers 5, 1, and 3 on NO expression secreted by Raw264.7.

[0068] Table 3. Effects of test samples on NO expression secreted by Raw264.7

[0069]

[0070] Table 4. Effects of test samples on NO secretion expression in Raw264.7

[0071]

[0072]

[0073] According to Table 3 and Figure 6 The experimental results showed that, compared with the BC group, the NO release from macrophages in the NC group was significantly increased, indicating that the LPS stimulation condition was effective in this experiment. Compared with the NC group, the NO release from macrophages at a dexamethasone concentration of 100 μg / mL was significantly reduced, indicating that this experiment was effective.

[0074] Compared with the NC group, sample group 1 could inhibit the release of NO from macrophages, indicating that 0.005% copper peptide has a significant inhibitory effect on the release of the inflammatory factor NO.

[0075] Compared with the NC group, sample group 2 did not inhibit the release of NO from macrophages, but the amount of NO released by macrophages in sample group 3 was significantly reduced, indicating that 1% TIRACLE PRO was not enough to inhibit the release of NO from macrophages, while 5% TIRACLE PRO could significantly and effectively inhibit the release of NO from macrophages.

[0076] From Table 4 and Figure 7 A comprehensive comparison of the results of samples 5, 1, and 3 shows that the combination of 0.005% copper peptide and 5% TIRACLE PRO has a synergistic effect on inhibiting the release of NO from macrophages, reducing the average concentration of the inflammatory factor NO to 3.08 pg / mL, a decrease of 53.2% compared to sample 1 and a decrease of 29% compared to sample 3, demonstrating a synergistic effect on inhibiting the release of the inflammatory factor NO.

[0077] Example 6: Effect of test samples on the expression of the inflammatory cytokine IL-6 secreted by macrophages

[0078] Raw264.7 mouse macrophage cell line was cultured normally at 37°C in DMEM medium containing 5% CO2 and 10% FBS. When the cells reached 90% confluence, they were cultured at a density of 6.4 × 10⁻⁶ cells / year. 5 Cells were seeded at a density of 1.8 mL / ml in 6-well plates and incubated overnight in an incubator (37℃, 5% CO2). When the cell layering rate reached 40%–60%, the cells were grouped according to Table 5, with 1.8 mL of drug administered to each well. Each group had 3 replicates. The cells were incubated for another 2 hours in an incubator (37℃, 5% CO2). After two hours of incubation, 200 μL of LPS working solution prepared from the corresponding test substance working solution was added to the drug-treated wells. The wells were shaken left and right to mix the drug. The final LPS concentration was 1 μg / mL. The cells were incubated for another 22 hours in an incubator (37℃, 5% CO2). After incubation, the cell culture supernatant was collected in EP tubes and detected using a Mouse IL-6 ELISA kit. The detection results are shown in Table 5. Figure 8 As shown, where Figure 8 This is a schematic diagram showing the effect of the test sample on the expression of IL-6 secreted by Raw264.7.

[0079] Table 5. Summary of IL-6 Expression Data

[0080]

[0081]

[0082] According to Table 5 and Figure 8 The experimental results showed that, compared with the BC group, the secretion of the inflammatory cytokine IL-6 by macrophages in the NC group was significantly increased, indicating that the LPS stimulation condition was effective in this experiment. Compared with the NC group, the secretion of the inflammatory cytokine IL-6 by macrophages at a drug concentration of 100 μg / mL was significantly reduced, indicating that this experiment was effective.

[0083] Compared with the NC group, sample group 1 could inhibit the secretion of inflammatory factor IL-6, indicating that 0.005% copper peptide has a significant inhibitory effect on the secretion of inflammatory factor IL-6.

[0084] Compared with the NC group, sample group 2 did not inhibit the secretion of inflammatory factor IL-6, but the secretion of inflammatory factor IL-6 in sample group 3 was significantly reduced, indicating that 1% TIRACLE PRO was not enough to inhibit the secretion of inflammatory factor IL-6, while 5% TIRACLE PRO could significantly inhibit the secretion of inflammatory factor IL-6.

[0085] A comprehensive comparison of the results of samples 1 to 5 shows that the combination of 0.005% copper peptide and 5% TIRACLE PRO (P<0.01) has the best effect on inhibiting the secretion of inflammatory factor IL-6, reducing the average concentration of inflammatory factor IL-6 to 164.15 pg / mL, which is 23% lower than that of sample group 1 and 6% lower than that of sample group 3.

[0086] Example 7: Testing the effect of samples on the expression of the inflammatory cytokine TNF-α secreted by macrophages

[0087] Raw264.7 mouse macrophage cell line was cultured normally at 37°C in DMEM medium containing 5% CO2 and 10% FBS. When the cells reached 90% confluence, they were cultured at a density of 6.4 × 10⁻⁶ cells / year. 5 Cells were seeded at a density of 1.8 mL / ml in 6-well plates and incubated overnight in an incubator (37°C, 5% CO2). When the cell layering rate reached 40%–60%, the cells were grouped according to Table 6, with 1.8 mL of drug administered to each well. Each group had 3 replicates. The cells were incubated for another 2 hours in an incubator (37°C, 5% CO2). After two hours of incubation, 200 μL of LPS working solution prepared from the corresponding test substance working solution was added to the drug-treated wells. The wells were shaken from side to side to mix the drug. The final LPS concentration was 1 μg / mL. The cells were incubated for another 22 hours in an incubator (37°C, 5% CO2). After incubation, the cell culture supernatant was collected in EP tubes and detected using a Mouse TNFα ELISA kit. The detection results are shown in Table 6. Figure 9 As shown, where Figure 9 This is a schematic diagram showing the effect of the test sample on the expression of TNF-α secreted by Raw264.7.

[0088] Table 6. Summary of TNF-α Expression Data

[0089]

[0090] The experimental results in Table 6 show that, compared with the BC group, the secretion of the inflammatory cytokine TNF-α by macrophages in the NC group was significantly increased, indicating that the LPS stimulation condition was effective in this experiment. Compared with the NC group, the secretion of the inflammatory cytokine TNF-α by macrophages at a drug concentration of 100 μg / mL was significantly decreased, indicating that this experiment was effective.

[0091] Compared with the NC group, sample group 1 could inhibit the secretion of inflammatory factor TNF-α, indicating that 0.005% copper peptide has a significant inhibitory effect on the secretion of inflammatory factor IL-6.

[0092] Compared with the NC group, both sample groups 2 and 3 could inhibit the secretion of inflammatory factor TNF-α. Among them, the secretion of inflammatory factor TNF-α in sample group 3 was more significantly affected, indicating that the inhibitory effect on inflammatory factor TNF-α increased with the increase of TIRACLE PRO concentration.

[0093] A comprehensive comparison of the results of samples 1 to 5 shows that the combination of 0.005% copper peptide and 5% TIRACLE PRO has the best effect on inhibiting the secretion of the inflammatory factor TNF-α, reducing the average concentration of TNF-α to 33328.28 pg / mL; compared with sample group 1, it decreased by 36%; compared with sample group 3, it decreased by 11%.

[0094] While the present invention has been disclosed above, it is not limited thereto. Any person skilled in the art can make various modifications and alterations without departing from the spirit and scope of the invention; therefore, the scope of protection of the present invention should be determined by the scope defined in the claims.

Claims

1. A skin-soothing composition, characterized in that, The composition comprises copper peptide, and symbiotic fermentation product of Thermus thermophilus with preservation number BAA-163 and Bacillus with preservation number CICC10023, and the preparation method comprises the following steps: (1) carrying out one or more times of bacteria increasing fermentation on Thermus thermophilus under the condition of 65 DEG C and pH value of 7.8±0.2, and making it in exponential growth phase; (2) adding Bacillus, and carrying out secondary fermentation under the condition of pH value of 7.0-7.2 and temperature of 30±2 DEG C, and centrifuging the fermentation liquor to remove bacteria.

2. The composition of claim 1, wherein The content of copper peptide is 0.005%, and the content of symbiotic fermentation product of Thermus thermophilus with preservation number BAA-163 and Bacillus with preservation number CICC10023 is 5%.

3. Use of a composition for the manufacture of a skin care product for soothing the skin, characterized in that, The composition relieves skin by inhibiting the expression of any one or more of inflammatory factors of PGE2, IL-6, TNF alpha or NO; the composition comprises copper peptide, and symbiotic fermentation product of Thermus thermophilus with preservation number BAA-163 and Bacillus with preservation number CICC10023.

4. The composition of claim 3, wherein The content of copper peptide is 0.005%, and the content of symbiotic fermentation product of Thermus thermophilus with preservation number BAA-163 and Bacillus with preservation number CICC10023 is 5%.

5. Use of a composition for the manufacture of a preparation for soothing the skin by inhibiting the expression of any one or more of the inflammatory factors PGE2, IL-6, TNFα or NO, characterized in that, The composition comprises copper peptide, and symbiotic fermentation product of Thermus thermophilus with preservation number BAA-163 and Bacillus with preservation number CICC10023.

6. Use according to claim 5, characterized in that, The content of copper peptide is 0.005%, and the content of symbiotic fermentation product of Thermus thermophilus with preservation number BAA-163 and Bacillus with preservation number CICC10023 is 5%.

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