Use of compounds for preparing cosmetics

Abstract

The invention relates to the technical field of cosmetics, in particular to application of a compound Drimendol or a compound Epimendol to preparation of cosmetics. The prepared cosmetic has the effects of moisturizing, repairing, tightening, nourishing, resisting wrinkles and relieving, is mild and non-irritating, is suitable for sensitive skin, has the effects of positively regulating and expressing the acid sphingomyelinase SMPD1 gene and the LOR gene and improving the skin barrier, has the effects of positively regulating and expressing the elastin ELN gene and improving the skin elasticity and compactness, and is suitable for the sensitive skin. The composition has the effects of negative regulation expression of tumor necrosis factor-alpha, interleukin-1alpha and prostaglandin E2 genes, improvement of skin inflammation and relieving of sensitive skin, has negative regulation expression of transient receptor potential vanillin-1 receptor genes, achieves the effects of skin analgesia and anti-inflammation, and has the effect of increasing synthesis of skin cell hyaluronic acid.

Description

Technical Field

[0001] The present invention relates to the technical field of cosmetics, in particular to use of compounds Drimendiol and Epidrimendiol in preparing cosmetics. Background Art

[0002] In 1995, Geoffrey D. Brown first reported that a new compound Drimendiol was isolated from the Chilean plant Drimys winteri (Geoffrey D. Brown, Journal of Essential Oil Research, 1995, 7:6, 705-707); Cristian Paza et al. reported that the compound Drimendiol was isolated from the Chilean plant Drimys winteri, and studied its quorum sensing, and found that Drimendiol could inhibit the formation of quorum effect of Chromobacterium violaceum ATCC 12472, reduce the synthesis of violacein, and reduce the formation of biofilm of Pseudomonas syringae (Cristian Paza et al., Natural Product Communications, 2013, Vol. 8, No. 2, 147-148); in 2013, Marcos Derita et al. studied the structure-activity relationship of natural Drimane Sesquiterpenes and antifungal compounds, and disclosed that the compound Drimendiol has antifungal activity against Candida albicans, Saccharomyces cerevisiae, Cryptococcus neoformans, Aspergillus niger, Aspergillus flavus, Aspergillus fumigatus, Microsporum gypseum, Trichophyton rubrum, and Trichophyton mentagrophytes, and found that its MIC value was above 125 μg / mL (Marcos Derita, Structural Requirements for the Antifungal Activities of Natural Drimane Sesquiterpenes and Analogues, Supported by Conformational and Electronic Studies, Molecules, 2013,18,2029-2051.); 2016, S.Mohanty reported that the compound Drimendiol was isolated from Australian pepper berry, and the antibacterial activity of the compound Drimendiol was extensively studied, which disclosed the compound Drimendiol's antibacterial activity against Alcaligenes faecalis (A.faecalis), Staphylococcus aureus (S.aureus ATCC), Serratia marcescens (S.marcescens), Klebsiella pneumoniae (K.pneumoniae), Escherichia coli (E.coli ATCC), Salmonella typhi (S.typhi), Enterococcus faecalis (E.faecium ATCC), Pseudomonas aeruginosa (P.aeruginosa ATCC), Proteus mirabilis, Streptococcus pyogenes, and Bacillus cereus, and the MIC values ​​for inhibition of various bacteria were disclosed; S. Mohanty proposed that the compound Drimendiol inhibited 8 of the 11 tested bacterial strains and had a relatively broad-spectrum antibacterial activity (S. Mohanty, Bioactive Properties of Australian Native Fruits, Tasmannia lanceolata and Terminalia ferdinandiana: The Characterization of their Active Compounds, 2016, Thesis (PhD Doctorate), Griffith University). However, the researchers of this application tested the MIC value of compound Drimendiol for inhibiting Escherichia coli (E.coli), Pseudomonas aeruginosa (P.aeruginosa) and Staphylococcus aureus (S.aureus), and found that the MIC value of compound Drimendiol for the above three strains was greater than 1000μg / mL, and did not test the strong inhibitory activity of E.coli and Staphylococcus aureus (S.aureus) proposed by S.Mohanty; Purity N.Kipangaa et al. studied the compound Drimendiol isolated from Ugandan camphor (W.ugandensis), and conducted biofilm inhibition tests on Candida albicans, Staphylococcus aureus (S.aureus) and Staphylococcus epidermidis (S.epidermidis), and obtained BIC. 50The values ​​were 25.5±8μg / mL, 65.1±24μg / mL and 67.1±12μg / mL, respectively (Purity N.Kipangaa, Biofilm inhibiting properties of compounds from the leaves of Warburgia ugandensis Sprague subsp ugandensis against Candida and staphylococcal biofilms, Journal of Ethnopharmacology, 2019); In 2020, Cristian Paz et al. reported the compound Drimendiol isolated from the plant Drimys winteri, and tested its inhibitory activity against the wheat take-all pathogen Gaeumannomyces graminis LC 50 is 60μg / mL (Cristian Paz, Applied and Environmental Microbiology, December 2020, Volume 86, Issue 24, e01834-20); in 2022, Nicole Cortez et al. reported that the compound Drimendiol was obtained by reducing polygodial, and the compound was tested for antibacterial effects on three strains of Candida. The minimum inhibitory concentrations for C. parapsilosis, C. krusei and Candida albicans were 25μg / mL, 30μg / mL and 50μg / mL, respectively (Nicole Cortez, Drimane Sesquiterpene Alcohols with Activity against Candida Yeast Obtained by Biotransformation with Cladosporium antarcticum, Int., J. Mol. Sci. 2022, 23, 12995.). However, the researchers of this application tested the minimum inhibitory concentration value of the compound Drimendiol against Candida albicans and found that its MIC value was about 1000 μg / mL, which was 20 times that reported in the literature; Ivan Montenegro et al. studied the cytotoxicity of natural sesquiterpenoid compounds and found that the compound Drimendiol had no cytotoxic activity and its IC value against 4 cell lines was 50All are greater than 200 μM (Ivan Montenegro, Study on the Cytotoxic Activity of DrimaneSesquiterpenes and Nordrimane Compounds against Cancer Cell Lines, Molecules 2014, 19, 18993-19006); Lindon WK Moodie et al. studied the relationship between the structure and anti-fouling performance of water polygonal dialdehyde and its derivatives. The results showed that compared with other derivatives, the compound Drimendiol did not have significant inhibitory activity against the ascidian C. savignyi and the barnacle B. improvisus (Lindon WK Moodie, J. Nat. Prod. 2017, 80, 515-525).

[0003] It can be seen from the above prior art reports that, up to now, most of the reports on the compound Drimendiol are about the antimicrobial activity, cytotoxic activity and anti-fouling performance. There is no report on the cosmetic efficacy and use of the compound Drimendiol.

[0004] In 2013, Marcos Derita et al. studied the structure-activity relationship between natural drimane sesquiterpenes and antifungal compounds, and disclosed that the compound epidrimendiol had antifungal activity against Candida albicans, Saccharomyces cerevisiae, Cryptococcus neoformans, Aspergillus niger, Aspergillus flavus, Aspergillus fumigatus, Microsporum gypseum, Trichophyton rubrum, and Trichophyton mentagrophytes, and found that its MIC value was above 125 μg / mL (Marcos Derita, Structural Requirements for the Antifungal Activities of Natural Drimane Sesquiterpenes and Analogues, Supported by Conformational and Electronic Studies, Molecules, 2013, 18, 2029-2051.); Ivan Montenegro et al. studied the cytotoxicity of natural sesquiterpenoid compounds and found that the compound Epidrimendiol had no cytotoxic activity and its IC 50All are greater than 200μM (Ivan Montenegro, Study on the Cytotoxic Activity of Drimane Sesquiterpenes and Nordrimane Compounds against Cancer Cell Lines, Molecules 2014, 19, 18993-19006); in 2022, Nicole Cortez et al. reported that the compound Epidrimendiol was obtained by reducing isotadeonal (isopolygodial) of Polygonum hydropiper, and the compound was tested for antibacterial activity against three strains of Candida. The minimum inhibitory concentrations for C. parapsilosis, C. krusei and Candida albicans were 25μg / mL, 12.5μg / mL and 15μg / mL, respectively (Nicole Cortez, Drimane Sesquiterpene Alcohols with Activity against Candida Yeast Obtained by Biotransformation with Cladosporium antarcticum, Int., J. Mol. Sci. 2022, 23, 12995.). However, the researchers of the present application conducted a minimum inhibitory concentration (MIC) test on the compound Epidrimendiol to inhibit Escherichia coli (E. coli), Pseudomonas aeruginosa (P. aeruginosa), Staphylococcus aureus (S. aureus), Candida albicans (Candida albicans) and Aspergillus niger (Aspergillus niger), and found that its MIC values ​​were all greater than 1000 μg / mL.

[0005] It can be seen from the contents of the above prior art reports that, up to now, most of the reports on the compound Epidrimendiol are studies on its antimicrobial activity, and there are no reports on the cosmetic efficacy and use of the compound Epidrimendiol. Summary of the invention

[0006] The technical problem to be solved by the present invention is: discovering the use of the compound Drimendiol or the compound Epidrimendiol for preparing cosmetics.

[0007] In order to solve the above technical problems, the technical solution adopted by the present invention is:

[0008] The use of the compound Drimendiol or the compound Epidrimendiol for preparing cosmetics is evaluated by testing the cosmetic human efficacy and cell model of the compound Drimendiol or the compound Epidrimendiol.

[0009] The chemical structural formula of Drimendiol is:

[0010]

[0011] The chemical structural formula of the Epidrimendiol is:

[0012]

[0013] The compound Drimendiol or the compound Epidrimendiol is used for preparing cosmetics with moisturizing, repairing, anti-wrinkle, firming, nourishing and / or soothing effects.

[0014] The compound Drimendiol or the compound Epidrimendiol is used for preparing cosmetics with 72-hour moisturizing effect.

[0015] The compound Drimendiol or the compound Epidrimendiol is used for preparing cosmetics suitable for sensitive skin.

[0016] The compound Drimendiol or the compound Epidrimendiol is used for preparing cosmetics without irritation reaction.

[0017] The compound Drimendiol or the compound Epidrimendiol is used for preparing cosmetics with the effects of moving the apple cheeks upward and / or reducing the angle of the mandibular angle.

[0018] The compound Epidrimendiol is used for preparing cosmetics with exfoliating properties.

[0019] The compound Drimendiol or the compound Epidrimendiol is used for preparing cosmetics which can positively regulate the expression of acid sphingomyelinase SMPD1 gene and loricrin LOR gene and improve the skin barrier.

[0020] The compound Drimendiol or the compound Epidrimendiol is used for preparing cosmetics which can positively regulate the expression of elastin ELN gene and improve the elasticity and firmness of the skin.

[0021] The compound Drimendiol or the compound Epidrimendiol is used for preparing cosmetics that have the functions of negatively regulating the expression of tumor necrosis factor-α, interleukin-1α and prostaglandin E2 genes, improving skin inflammation and soothing sensitive skin.

[0022] The compound Drimendiol or the compound Epidrimendiol is used for preparing cosmetics which have the functions of negatively regulating the expression of transient receptor potential vanilloid-1 receptor gene and achieving skin analgesic and anti-inflammatory soothing effects.

[0023] The compound Drimendiol is used for preparing cosmetics having the effect of increasing the synthesis of hyaluronic acid in skin cells.

[0024] The beneficial effects of the present invention are as follows: a new use of the compound Drimendiol or the compound Epidrimendiol for preparing cosmetics is studied, so that the prepared cosmetics have 72-hour moisturizing, hydrating, repairing, firming, nourishing, anti-wrinkle and soothing effects; and are mild and non-irritating and suitable for sensitive skin; and have the effect of positively regulating the expression of the acid sphingomyelinase SMPD1 gene and the loricrin LOR gene, and improving the skin barrier effect; have the effect of positively regulating the expression of the elastin ELN gene, and improving the skin elasticity and firmness effect; have the effect of negatively regulating the expression of the tumor necrosis factor-α, interleukin-1α and prostaglandin E2 genes, and improving the skin inflammation and sensitive skin soothing effect; have the effect of negatively regulating the expression of the transient receptor potential vanilloid-1 receptor gene, and achieving the skin analgesic and anti-inflammatory soothing effect; and have the effect of increasing the synthesis of hyaluronic acid in skin cells. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 This is a comparison chart of the moisture content of the stratum corneum of the cheek skin of the test subject before and after using the sample in Example 1;

[0026] Figure 2 This is a comparison chart of the moisture content of the stratum corneum of the skin on the arm of the subject in Example 2 before and after the sample is used in the sample area and the blank area;

[0027] Figure 3 This is a comparison chart of the moisture content increment of the stratum corneum of the skin on the arm of the test subject after using the sample in the sample area and the blank area in Example 2;

[0028] Figure 4 This is a comparison chart of the transepidermal water loss rate of the subjects before and after using the samples in Example 3;

[0029] Figure 5 This is a comparison chart of SEr values ​​of cheek skin roughness parameters of the subjects before and after using the samples in Example 4;

[0030] Figure 6This is a comparison chart of the SEsm values ​​of cheek skin smoothness parameters of the subjects before and after using the samples in Example 4;

[0031] Figure 7 This is a comparison chart of SEr values ​​of elbow skin roughness parameters of the subjects before and after using the samples in Example 4;

[0032] Figure 8 This is a comparison chart of the SEsm values ​​of elbow skin smoothness parameters before and after the subjects used the samples in Example 4;

[0033] Fig. 9 This is a comparison chart of the skin glossiness of the subjects in Example 4 before and after using the samples;

[0034] Fig.10 This is a comparison chart of the skin elasticity R2 values ​​of the subjects before and after using the samples in Example 4;

[0035] Fig.11 This is a comparison chart of the thickness of the dermis layer of the skin of the subjects in Example 4 before and after using the samples;

[0036] Fig.12 This is a comparison diagram of the density of the dermis layer of the skin of the test subject in Example 4 before and after using the sample;

[0037] Fig.13 This is a comparison chart of a* values ​​of the skin red area of ​​the subjects before and after using the samples in Example 5;

[0038] Fig.14 This is a comparison chart of the skin color L* values ​​of the subjects in Example 5 before and after using the samples;

[0039] Fig.15 This is a comparison chart of the skin firmness F4 values ​​of the subjects before and after using the samples in Example 6;

[0040] Fig.16 This is a comparison chart of the area ratio of under-eye wrinkles of the subjects before and after using the samples in Example 7;

[0041] Fig.17 This is a comparison diagram of the mandibular angles of the subjects in Example 8 before and after using the samples;

[0042] Fig.18 This is a comparison diagram of the upward movement distance of the apple muscles of the subjects in Example 8 before and after using the samples;

[0043] Fig.19 This is a comparison chart of the subjective skin feeling evaluation (questionnaire on a 10-point scale) of the subjects in Example 10 before and after using the samples;

[0044] Fig. 20 This is a graph showing the self-evaluation results of the subjects in Example 10 after the subjects used the samples for 28 days;

[0045] Fig.21This is a comparison chart of the change in skin smoothness before and after the test subject used the sample in Example 4;

[0046] Fig. 22 This is a rendering effect comparison diagram of the change in skin smoothness before and after the test subject used the sample in Example 4;

[0047] Fig.23 This is a comparison chart of the effect of skin thickness / density changes before and after the subjects used the samples in Example 4;

[0048] Fig.24 This is a comparison chart of the skin color improvement effect of the subjects in Example 5 before and after using the samples;

[0049] Fig.25 This is a comparison chart of the improvement effect of the skin red area before and after the test subject used the sample in Example 5;

[0050] Fig.26 This is a comparison chart of the improvement effect of under-eye wrinkles on the subjects in Example 7 before and after using the samples;

[0051] Fig. 27 This is a comparison diagram of the mandibular angle improvement effect of the subjects in Example 8 before and after using the samples;

[0052] Fig.28 This is a comparison chart of the improvement effect of the upward displacement of the apple muscles before and after the test subjects used the samples in Example 8;

[0053] Fig.29 The relative activity bar graph of each group of cells in the MTT experiment in Example 11;

[0054] Fig.30 It is a bar graph of the relative expression of SMPD1 gene in each group in Example 11;

[0055] Fig.31 The bar graph of the relative expression of LOR genes in each group in Example 11;

[0056] Fig.32 The relative activity bar graph of each group of cells in the MTT experiment in Example 12;

[0057] Fig.33 The bar graph of the relative expression of ELN genes in each group in Example 12;

[0058] Fig.34 The relative activity bar graph of each group of cells in the MTT experiment in Example 13;

[0059] Fig.35 The relative expression level of TNF-α gene in each group in Example 13 is shown in the bar graph;

[0060] Fig.36The bar graph of the relative expression of IL-1α gene in each group in Example 13;

[0061] Fig.37 The relative activity bar graph of each group of cells in the MTT experiment in Example 14;

[0062] Fig.38 The bar graph of the relative expression of TRPV1 gene in each group in Example 14;

[0063] Fig.39 The bar graph of the relative expression of PGE2 gene in each group in Example 14;

[0064] Fig.40 The relative activity bar graph of each group of cells in the MTT experiment in Example 15;

[0065] Fig.41 It is a bar graph of the average HA concentration of each group in Example 15;

[0066] Fig.42 This is a diagram showing the moisture content of the stratum corneum before and after use in Example 16;

[0067] Fig.43 This is an analysis chart of the change in stratum corneum moisture content before and after use in Example 17;

[0068] Fig.44 This is an analysis chart of transepidermal water loss rate before and after use in Example 18;

[0069] Fig.45 This is an analysis diagram of SEsm values ​​before and after use in Example 19;

[0070] Fig.46 This is a gloss analysis diagram before and after use in Example 19;

[0071] Fig.47 This is an analysis diagram of the R0 value before and after use in Example 19;

[0072] Fig.48 This is an analysis diagram of the R2 value before and after use in Example 19;

[0073] Fig.49 This is a graph showing the analysis of dermal thickness before and after use in Example 19;

[0074] Fig.50 This is a diagram of dermal density analysis before and after use in Example 19;

[0075] Fig.51 This is an analysis diagram of a* values ​​before and after use in Example 20;

[0076] Fig.52 This is an analysis chart of the L* value before and after use in Example 20;

[0077] Fig.53 This is an analysis diagram of the ratio of under-eye wrinkle areas before and after use in Example 22;

[0078] Fig.54 This is a volume analysis diagram of the apple muscle sagging before and after use in Example 23;

[0079] Fig.55 This is an analysis diagram of the length of the contour line of the front and rear mandibular drooping parts used in Example 23;

[0080] Fig.56 This is a graph showing the Bowman Questionnaire score analysis before and after use in Example 24;

[0081] Fig.57 This is an analysis diagram of the desquamation index DI value before and after use in Example 25;

[0082] Fig.58 This is a comparison diagram of the volume change of the apple muscle drooping before and after use in Example 23;

[0083] Fig.59 This is a comparison diagram of the contour line changes of the lower jaw drooping part before and after use in Example 23;

[0084] Fig.60 This is a comparison diagram of the changes in under-eye wrinkles before and after use in Example 22;

[0085] Fig.61 This is a comparison chart of skin smoothness changes before and after use in Example 19;

[0086] Fig.62 This is a comparison chart of skin exfoliation changes before and after use in Example 25;

[0087] Fig.63 This is a comparison chart of changes in dermal thickness and dermal density before and after use in Example 19;

[0088] Fig.64 This is a comparison chart of skin redness changes before and after use in Example 20;

[0089] Fig.65 This is a bar graph of the relative activity of each group of cells in the first MTT experiment in Example 27;

[0090] Fig.66 This is a bar graph of the relative activity of each group of cells in the second MTT experiment in Example 27;

[0091] Fig.67 This is a bar graph of the relative expression levels of the SMPD1 gene in each group in the first efficacy test in Example 27;

[0092] Fig.68This is a bar graph of the relative expression levels of the SMPD1 gene in each group in the second efficacy test in Example 27;

[0093] Fig.69 This is a bar graph of the relative expression levels of LOR genes in each group in the first efficacy test in Example 27;

[0094] Fig.70 This is a bar graph of the relative expression levels of LOR genes in each group in the second efficacy test in Example 27;

[0095] Fig.71 This is a bar graph of the relative activity of each group of cells in the first MTT experiment in Example 28;

[0096] Fig.72 This is a bar graph of the relative activity of each group of cells in the second MTT experiment in Example 28;

[0097] Fig.73 This is a bar graph of the relative expression levels of ELN genes in each group in the first efficacy test in Example 28;

[0098] Fig.74 This is a bar graph of the relative expression levels of ELN genes in each group in the second efficacy test in Example 28;

[0099] Fig.75 The relative activity bar graph of each group of cells in the MTT experiment in Example 29;

[0100] Fig.76 The relative expression level of TNF-α gene in each group in Example 29 is shown in the bar graph;

[0101] Fig.77 is a bar graph of the relative expression levels of IL-1α genes in each group in Example 29;

[0102] Fig.78 This is a bar graph of the relative activity of each group of cells in the first MTT experiment in Example 30;

[0103] Fig.79 This is a bar graph of the relative activity of each group of cells in the second MTT experiment in Example 30;

[0104] Fig.80 This is a bar graph of the relative expression levels of TRPV1 genes in each group in the first efficacy test in Example 30;

[0105] Fig.81 This is a bar graph of the relative expression of PGE2 genes in each group in the first efficacy test in Example 30;

[0106] Fig.82 This is a bar graph of the relative expression levels of PGE2 genes in each group in the second efficacy test in Example 30. DETAILED DESCRIPTION

[0107] In order to explain the technical content, achieved objectives and effects of the present invention in detail, the following is an explanation in combination with the implementation modes and the accompanying drawings.

[0108] The "Standards for the Evaluation of Cosmetic Efficacy Claims" (implemented on May 1, 2021) stipulates that the efficacy of cosmetics includes: moisturizing, hair care, freckle removal and whitening, sun protection, anti-hair loss, acne removal, nourishing, repairing, anti-wrinkle, firming, soothing, oil control, exfoliation, anti-hair breakage, anti-dandruff, mild claims (such as non-irritation), specific claims for sensitive skin and tear-free formulas. At the same time, clear requirements are made for the claims of the above cosmetic efficacy. Specifically, cosmetics with freckle removal and whitening, sun protection, anti-hair loss, acne removal, nourishing and repairing efficacy should be evaluated through human efficacy evaluation tests; cosmetics with anti-wrinkle, firming, soothing, oil control, exfoliation, anti-hair breakage and anti-dandruff efficacy, as well as mild claims (such as non-irritation) or quantitative indicators (such as efficacy claim duration, efficacy claim-related statistical data, etc.) should be evaluated through cosmetic efficacy claim evaluation tests, which can be combined with literature or research data analysis results. Cosmetics that make specific claims (such as claiming to be suitable for sensitive skin, claiming to be a tear-free formula) should have their efficacy claims evaluated through human efficacy evaluation trials or consumer use tests.

[0109] It is also stipulated that the test methods used in the evaluation of the efficacy of cosmetics include: methods specified in my country's mandatory national standards and technical specifications for cosmetics; methods specified in other relevant laws and regulations, national standards, and industry standards in my country; methods specified in relevant foreign laws and regulations or technical standards; methods published in technical guidelines of authoritative organizations, technical institutions, and industry associations at home and abroad, methods publicly published in professional academic journals, or methods independently formulated and established. Before conducting efficacy evaluation, the evaluation agency shall complete the necessary transfer, confirmation or verification of the test methods to ensure the scientificity and reliability of the evaluation work.

[0110] In this application, the cosmetic effects of the compounds Drimendiol and Epidrimendiol include 72-hour moisturizing (quantitative indicators), moisturizing, repairing, anti-wrinkle, firming, nourishing, soothing, mild and non-irritating, and suitable for sensitive skin, all of which have passed the human efficacy evaluation, and the conclusions drawn from the evaluation result data were scientifically statistically analyzed through the standard cosmetic efficacy evaluation. The other specific claimed facial lifting effects: lowering of the mandibular angle and upward movement of the apple muscles were measured and statistically analyzed through human efficacy evaluation. In this application, the evaluation methods for efficacy all meet the requirements of the "Specifications for Evaluation of Cosmetic Efficacy Claims", and are specifically described in the embodiments of this application.

[0111] Relationship between SMPD1 gene expression and skin stratum corneum moisturizing

[0112] Sphingomyelin phosphodiesterase 1 (SMPD1), also known as acid sphingomyelinase (ASM), is an enzyme responsible for converting sphingomyelin to ceramide. It is encoded and expressed in the human body through the SMPD1 gene. SMPD1 gene defects can lead to Niemann-Pick disease (NPD), also known as sphingomyelin deposition disease. This disease is a sphingomyelin metabolism disorder caused by the deficiency of sphingomyelinase. In addition, the mutation of leucine to proline at residue 302 of the SMPD1 gene increases the risk of Parkinson's disease.

[0113] Ceramide plays an important role in the integrity of the skin barrier. It is an important component of the phospholipid bilayer of the stratum corneum cell membrane. Patients with SMPD1 deficiency show abnormal permeability of the skin barrier, and the reason is that the content of ceramide in the stratum corneum is abnormally low (Schmuth et al., 2000). By applying SMPD1 inhibitors palmitoyl dihydrosphingosine and desipramine on the skin of nude mice, they found that the content of sphingomyelin in the mouse skin increased and the thickness of the outer layer of the stratum corneum decreased. This type of mouse showed delayed repair of skin barrier damage. In addition, by applying a cream containing ceramide to the damaged area, the symptoms of skin barrier damage caused by the inhibitor to mice can be improved. This finding shows that the loss of the stratum corneum barrier function in patients with SMPD1 deficiency is not caused by the accumulation of sphingomyelin content, but the real reason is due to the imbalance of the ratio of sphingomyelin and ceramide. The important reason for this is that SMPD1 does not play a role in preventing the conversion of sphingomyelin into ceramide.

[0114] In addition, patients with eczema have lower levels of SMPD1 enzyme activity in the affected skin area, which leads to lower ceramide levels and abnormal skin barrier permeability. Obviously, the normal function of the skin depends on the integrity of the epidermal cell membrane, and the role of the SMPD1 enzyme is very critical. When the SMPD1 enzyme activity is lost, the integrity of the epidermal cell membrane is lost, and the moisture of the stratum corneum of the skin is easily evaporated, resulting in dehydration and dryness of the skin. Therefore, the activity level of the SMPD1 enzyme is also closely related to the moisture content of the stratum corneum of the skin. When the SMPD1 enzyme activity is low, ceramide is lost in the stratum corneum of the skin, resulting in a loss of integrity of the stratum corneum of the skin, so the moisture of the stratum corneum of the skin evaporates, showing dry skin. When the SMPD1 enzyme activity is high, the ceramide level in the stratum corneum of the skin is high, thus ensuring the good integrity of the stratum corneum of the skin, and the moisture of the stratum corneum of the skin is easily locked in the stratum corneum, showing good skin moisturizing.

[0115] The compounds of the present application have been found in research to have a positive regulatory effect on the relative expression of the SMPD1 gene. According to the function of the SMPD1 enzyme, it can be inferred that when the expression level of the SMPD1 enzyme increases, the ceramide level in the stratum corneum of the skin is sufficient, the skin barrier integrity is good, and therefore the stratum corneum of the skin is sufficiently hydrated. Therefore, the compounds of the present application can be used for moisturizing purposes of skin SMPD1 gene regulation in cosmetics.

[0116] The relationship between loricrin and skin barrier and moisturizing

[0117] The stratum corneum is located in the outermost layer of the human epidermis and is composed of multiple layers of flat, anucleated keratinocytes. Each keratinocyte is surrounded by a cornified cell envelope (CE), which contains envelope proteins such as involucrin, loricrin, small proline-rich protein (SPR), elastin, keratin filaments, filaggrin, cystatin-A, and desmosomal proteins.

[0118] Loricrin is the main protein that makes up the keratinized membrane, accounting for about 80%. Loricrin is an insoluble polypeptide with a molecular weight of 26kDa. The human loricrin is expressed through the LOR gene. Loricrin mainly plays a role in strengthening the stratum corneum cell membrane and the barrier function of the stratum corneum. Loricrin is expressed in the epidermis of all mammals, and the expression level is highest in tissues with high water content, such as the epidermis of newborns, oral and anal mucosal epithelium, esophagus, foreskin, vagina and sweat ducts.

[0119] The expression of loricrin is affected by various factors, such as cell confluence, calcium ions, vitamin A deficiency, transglutaminase activity, and nectin (Nectin-1). These transcription factors act on loricrin activating protein (AP1), ultimately inducing epidermal differentiation. For example, loricrin expression is negatively regulated by retinoic acid, resulting in the failure to form a corneocyte capsule. Calcium ions affect the transcription level of loricrin. When the calcium ion level is low, such as below 0.10mM, loricrin and corneocyte capsule formation are immature and fragile. When the cell density is high, at a calcium ion level of 0.05mM, loricrin stops expressing. Nectin-1 knockout mice show defective expression of loricrin, resulting in a fragile corneocyte capsule that is easily broken under mechanical force. For example, under the attack of inflammatory factors, the negative regulation expression of filaggrin and loricrin genes in the stratum corneum of eczema patients leads to a significant decrease in barrier function, thereby increasing the water loss rate and making the skin appear drier.

[0120] It can be seen that loricrin is essential for the integrity of the corneocyte capsule. More specifically, from a functional point of view, loricrin is essential for maintaining the corneocyte capsule and the role of corneocytes in the skin barrier. From the perspective of cosmetic efficacy, if the skin barrier function is intact, the stratum corneum will have a strong ability to retain moisture; conversely, if the skin barrier function is imperfect, the stratum corneum will have a weak ability to retain moisture and will be prone to water loss. Whether the skin barrier function is perfect directly determines the moisture content of the stratum corneum. Therefore, to a large extent, the expression level of loricrin in the stratum corneum directly determines the moisture content of the stratum corneum. If the gene expression level of loricrin in the stratum corneum is high, the production of loricrin will increase accordingly, and the moisture content of the stratum corneum will increase. Conversely, if the gene expression level of loricrin in the stratum corneum is low, the amount of loricrin produced will also be low, and the stratum corneum will feel dry.

[0121] During extensive research, the applicant discovered that the compounds of the present application positively regulated the expression of the loricrin gene LOR at a concentration of 2 ppm, and the relative expression level of the LOR gene reached 205%. Specifically, the examples provide a detailed description and summary of the results.

[0122] The relationship between elastin and skin anti-wrinkle and firming

[0123] Skin aging is a complex process, with factors both internal and external to the body. Ultraviolet radiation (UV) is one of the most representative causes of skin damage. After years of exposure to radiation, the skin undergoes photoaging. As we age, skin tissue gradually changes, such as reduced synthesis and degradation of elastin, which causes the skin to lose elasticity.

[0124] Elastin fibers are essential components of skin connective tissue. Elastin fibers are synthesized through extracellular matrix and then secreted into skin fibroblasts. Through the assembly of elastin and a large number of microtubule elements, a fiber mesh structure is formed, which can extend and contract and has elasticity. Mature elastin fibers are mainly composed of elastin. Elastin in the human body is encoded by a single gene ELN. This gene is located on chromosome 7. During development, it differentiates into various cell types and expresses elastin.

[0125] Changes in collagen caused by natural aging and photoaging have been extensively studied. The elastin composition of the skin is poorly studied. The FDA has not approved any elastin substitutes or stimulators of elastin synthesis. Recently, animal studies have shown that a zinc-containing complex can increase skin elastin staining, subcutaneous fat, and epidermal thickness. Histoanatomical studies have shown that zinc complexes can increase the deposition of functional elastin fibers after 10 days of skin application. Clinical trials have reported that the increase in functional elastin promotes the improvement of fine lines and wrinkles (Leslie Baumann and Susan Weinkle, Improving Elasticity: The Science of Aging Skin, 2010).

[0126] Collagen and elastin are fibrous proteins that make up the skin matrix. Cosmetic dermatologists have long used topical medical products, implant fillers, and light- and laser-based treatments to correct the loss of collagen in the skin due to aging. However, to date, there are no fillers or topical agents to address the loss of elastin function, which is also a key feature of photoaging. The decline of collagen, elastin and other skin matrix components, chronological aging and photoaging, leads to the development of wrinkles, sagging and skin fatigue.

[0127] The role of elastin in skin aging

[0128] Elastin is the elastic connective tissue of the extracellular matrix. Elastin fibers are usually distributed around the edges of collagen bundles and elastic skin. The precursor of elastin is the tropoelastin molecule. Elastin is formed by covalent cross-linking. Elastin further forms fibrils by assembling to form microfibrils. The fibrils form the template on which elastin is deposited. The vast majority of elastin production is limited to a very narrow window of growth and development. Elastin is produced in large quantities during the fetal period, labor period and early neonatal period. After maturity, the synthesis of elastin decreases sharply.

[0129] Unlike collagen fibers, elastin fibers exist in various stages of maturity. Oxytalan fibers are the least mature elastin fibers and extend vertically from the dermal-epidermal junction to the top of the reticular dermis. Elaunin fibers are more mature elastin bundles attached to horizontal fiber clusters in the reticular dermis. The most mature elastin fibers are unnamed and are deeper in the top of the dermis.

[0130] This fiber network stretches from the top of the dermis to the bottom of the basement membrane, maintaining the elasticity of young skin. This elastin fiber network deteriorates with age, and the loss or depletion of elastin fibers is an important cause of skin sagging and loss of youthful skin elasticity.

[0131] Initially, photoaging leads to the proliferation of elastin fibers, depending on the level of UV exposure. Then, a degenerative reaction occurs, with the result that the skin loses its elasticity. Under a microscope, degraded elastin appears as an amorphous mass, concentrated in the dermis of the skin. The result is the degeneration of elastic tissue (elastosis), a hallmark of photoaging of the skin, due to the degradation of elastin fibers and the loss of elastin function. Elastin defects and damage can lead to wrinkles, even without exposure to the sun or aging conditions, and children with skin wrinkle syndrome have shown that the main cause is elastin fiber defects. This points to the important role of elastin in maintaining the integrity of the skin.

[0132] Studies have shown that as the skin ages, even in well-protected areas, the amount of elastin decreases. A study in Egypt showed that even in normal skin without UV radiation, the relative amount of elastin dropped from 49% at around 10 years old to 30.4% at 90 years old. Another study of 91 white people on the buttocks of non-UV-exposed skin showed that the amount of elastin decreased by 51% from 20 to 80 years old.

[0133] For a long time, cosmetic skin experts have emphasized the formation of new collagen through collagen supplement injections or topical cosmetics. However, there are fewer products that target the improvement of skin elastin. Although some cosmetics contain collagen and elastin, claiming to increase the level of these protein fibers in the skin, topical collagen and elastin are not convincing. Because collagen and elastin have large molecular weights, it is difficult for them to penetrate into the dermis of the skin.

[0134] The compounds of the present application, in the process of extensive cosmetic activity research, were found to have a significant positive regulatory effect on the ELN gene expression of elastin in fibroblasts. At very low concentrations, for example, 5 to 20 ppm, the relative expression of the ELN gene was 2.8 to 6.5 times higher. Moreover, the compounds of the present application have a small molecular weight and strong permeability, and can easily penetrate into the dermis of the skin. Therefore, they are suitable for use in the preparation of cosmetics that promote the synthesis of skin elastin.

[0135] The synthesis of skin elastin increases, and on human skin, the efficacy is reflected in the anti-wrinkle and firming effect of the skin. In the human efficacy evaluation test, it is also confirmed that the compound of the present application has a significant anti-wrinkle and firming effect on the skin.

[0136] Relationship between inflammatory factor inhibition and skin soothing

[0137] Human keratinocytes (KC) are located in the surface layer of the skin, accounting for more than 80% of epidermal cells, and are the part of the skin that is most directly irradiated by sunlight ultraviolet (UV). Sunlight UV radiation can cause photoaging and photodamage of the skin. Under culture conditions, KC cells were irradiated by UVB, and the morphological changes of cell damage were observed under a microscope, mainly manifested as cell swelling, shedding, increased cell debris, and cells floating on the culture medium. When the UVB dose is too large, it will cause apoptosis of skin cells. At the same time, KC is also an important part of the skin immune system and can secrete a variety of cytokines. Tumor necrosis factor TNF-α is one of the main epidermal cytokines synthesized by it, which participates in the skin inflammatory response process and immune regulation caused by UV, and plays an important role in UV-mediated cell apoptosis. In addition, UV radiation induces skin KC to secrete TNF-a and some other interleukins, such as IL-1α, IL-10, IL-6, PGE2, etc. As the UV irradiation dose increases, the secretion of cytokines such as TNF-α and interleukin secretion also increases, which eventually manifests as apoptosis of KC cells. The above description is about the UV-induced inflammatory process of human keratinocytes and the production of some cytokines.

[0138] In addition to the exposure of the skin's stratum corneum to UV, when the skin comes into contact with toxic, corrosive, and irritating chemicals, exogenous pathogens, and mechanical injuries such as abrasions, these will stimulate the keratinocytes' own immune system to repair the cell tissue damage caused by these factors.

[0139] These skin inflammations are largely mediated by a series of biochemical reactions through epidermal cytokines and lipids. For example, the activation of the kinin and arachidonic acid systems triggers a chain reaction of proteins and lipids, resulting in the expression of pro-inflammatory cytokines.

[0140] Proinflammatory factors that control inflammatory responses include IL-1α, IL-1β, IL-8, IL-6, PGE2, and TNF-α. Among them, IL-1α, IL-1β, and TNF-α are called primary cytokines because they can directly and quickly induce local inflammation of the skin. Primary cytokines can stimulate the production of secondary mediators, such as IL-6 and IL-8. Secondary cytokines play an auxiliary role in the inflammatory response. In order to regulate the inflammatory response process, inflammatory factor receptor antagonists are used to inhibit the inflammatory process.

[0141] Pro-inflammatory cytokines IL-1α, IL-1β, IL-8, IL-6, PGE2 and TNF-α are overexpressed in the skin, which can cause discomfort such as redness, tingling, itching, etc. In cosmetic dermatology, in order to relieve the allergic inflammatory response of the skin, small molecule active substances can be used to inhibit the overexpression of pro-inflammatory factors to relieve the discomfort caused by various skin inflammatory factors.

[0142] The compounds of the present application have an inhibitory effect on the gene expression of inflammatory factors TNF-α, IL-1α and PGE2 at a low concentration of ppm, and can be developed into cosmetics with soothing effects on sensitive skin and sensitive tendencies, with good effects.

[0143] Relationship between TRPV1 and skin soothing

[0144] Transient receptor potential channels play an important role in the process of signal transmission between adjacent cells. So far, six major types of transmembrane receptors have been discovered, with various active mechanisms. Among them, the mechanism of action of TRPV1 (transient receptor potential vanilloid 1) receptor is relatively clear. TRPV1 receptor is a calcium ion channel regulated by temperature. TRPV1 is located on sensory nerve fibers.

[0145] The receptor is activated under the conditions of temperature greater than 42°C, low pH and chemical activators, and the TRPV1 receptor is in an excited state. For example, capsaicin can activate TRPV1. Inflammatory responses supported by certain metabolites and cytokines, such as arachidonic acid and leukotriene compounds, can activate TRPV1 receptors.

[0146] When TRPV1 receptors are activated, calcium ions flow out of cells, and the activation signal is transferred to nerve factors, which are then transmitted to the brain. When TRPV1 is activated, it causes skin pain, burning, tingling and tightness. These uncomfortable feelings are typical symptoms of sensitive skin. The chemical activator capsaicin can activate TRPV1 in an ion channel activation state, causing a burning sensation in the skin. Interestingly, if capsaicin continuously stimulates TRPV1 receptors, the activation state of TRPV1 receptors will be downregulated, eventually leading to passivation and insensitivity. On the contrary, the application of acidic substances leads to increased TRPV1 expression. Therefore, TRPV1 regulation is regulated by various external changing factors. In people with super sensitive skin, TRPV1 receptors are easily activated, and the number of activated ones is large, making the skin extremely sensitive to changes in the external environment.

[0147] TRPV1 receptor antagonists can control the outflow of calcium ions and have a soothing effect on sensitive skin discomfort. For example, capsaicin, a potent TRPV1 receptor antagonist, has an extremely low micromolar IC 50 , which can inhibit the calcium ion outflow induced by chemical agonists activated by TRPV1. The main reason for this is the complexity of the TRPV1 receptor conformation.

[0148] The compounds of the present application have been found in research to have a negative regulatory effect on the relative expression of the TRPV1 gene. The specific data are detailed in the examples. According to the function of the TRPV1 gene, it can be inferred that it has a certain inhibitory effect on the expression of TRPV1 protein, reduces the expression of TRPV1 receptors in sensitive muscles, and reduces the skin's response to external stimuli. Therefore, the compounds of the present application can be used for skin soothing purposes in cosmetics.

[0149] The chemical structure of Drimendiol is:

[0150]

[0151] Preparation method of Drimendiol:

[0152] 10.0 kg of pepperberry (Tasmannia lanceolata) leaf powder was extracted with 60.0 kg of ethanol: water = 1:1 solvent at 60 ° C for 3 hours under heating and stirring. After filtration, the pepperberry leaf residue was extracted with 30.0 kg of ethanol: water = 1:1 solvent at 60 ° C for 2 hours under heating. After filtration, the two filtrates were combined and concentrated under reduced pressure to obtain 650 g of extract.

[0153] 650g of pepperberry leaf extract was separated by 2kg of 200-300 mesh silica gel column chromatography, and the eluent was gradient eluted with petroleum ether (PE) and PE: ethyl acetate (EA) = 10:1, 5:1 and 3:1, and the eluates of each component were collected respectively. The compound Drimendiol was mainly concentrated in the PE:EA = 5:1 elution component. After the component was concentrated under reduced pressure, a concentrate containing Drimendiol was obtained. Subsequently, it was separated again by 200-300 mesh silica gel column chromatography to remove impurities, and the compound Drimendiol weighed 16g.

[0154] Compound Drimendiol NMR data:

[0155] δ 13C-NMR: 39.1(C-1); 18.4(C-2); 42.0(C-3); 33.0(C-4); 49.4(C-5); 23.6(C-6); 127.7(C-7); 137.3 (C-8); 54.9 (C-9); 35.6 (C-10); 68.0 (C-11); 61.4 (C-12); 33.2 (C-13); 21.9 (C-14); 14.5 (C-15).

[0156] δ 1 H-NMR: 5.81 (H, m); 4.34~4.38 (H, d); 3.96~3.99 (H, d); 3.89~3.92 (H, dd); 3 .65~3.70(H,dd);3.6(2H,br,2OH);2.17(H,m);2.11~2.14(H,m);1.93~2.0 0(H,m); 1.85~1.91(H,m); 1.53~1.61(H,m); 1.48-1.52(H,m); 1.38~1.47(H ,m); 1.25(H,m); 1.13~1.19(2H,m); 0.89(3H,s); 0.88(3H,s); 0.76(3H,s).

[0157] Prepare a 20 ppm Drimendiol solution as a sample: first dissolve 0.2 g Drimendiol in 99.8 g polyol (such as 1,3-butanediol or propylene glycol), stir and heat to dissolve to prepare a 2000 ppm Drimendiol solution, and then dilute the 2000 ppm Drimendiol solution 100 times with pure water to 20 ppm.

[0158] The prepared 20 ppm Drimendiol solution was used as a sample to test the cosmetic uses of various human effects (Examples 1 to 10);

[0159] The prepared 2000 ppm Drimendiol solution was used as a sample to test the cosmetic uses of various gene expression effects (Examples 11 to 15);

[0160] Testing period: July 29, 2024 - August 26, 2024;

[0161] Test environment: Temperature: 21±1℃; Humidity: 50±10%RH;

[0162] 32 subjects with sensitive skin were selected and the Bergman skin typing questionnaire was conducted on the 32 subjects. The results are shown in Table 1:

[0163] Table 1

[0164]

[0165]

[0166] Note: If the score is 34-72, it is very sensitive skin; if the score is 30-33, it is slightly sensitive skin; if the score is 25-29, it is relatively tolerant skin; if the score is 18-24, it is very tolerant skin; in summary, if the score is 30-72, it is sensitive skin (abbreviated as "sensitive" type or S Type), and if the score is 18-29, it is tolerant skin (abbreviated as RType).

[0167] Subject inclusion criteria:

[0168] 1) Chinese women aged 30-55, in good health;

[0169] 2) Sensitive skin (Baumann Questionnaire), visible wrinkles under the eyes (level of wrinkles under the eyes ≥ level 2, refer to the "Skin Aging Atlas"), loose, dry, red facial skin, weak facial skin barrier (TEWL ≥ 15g / h / m 2 )

[0170] 3) Be in good health, without any other chronic diseases or diseases under treatment;

[0171] 4) Voluntarily participate in the evaluation and sign the informed consent form;

[0172] 5) Willing to comply with all evaluation requirements.

[0173] Subject exclusion criteria:

[0174] 1) Those who have used antihistamines in the past week or tried immunosuppressants in the past month;

[0175] 2) Those who have used any anti-inflammatory drugs in the tested area within the past two months;

[0176] 3) Subjects with inflammatory skin diseases that have not been clinically cured;

[0177] 4) Patients with insulin-dependent diabetes;

[0178] 5) Patients who are receiving treatment for asthma or other chronic respiratory diseases;

[0179] 6) Those who have received anti-cancer chemotherapy in the past 6 months;

[0180] 7) Patients with immunodeficiency or autoimmune diseases;

[0181] 8) Lactating or pregnant women;

[0182] 9) Bilateral mastectomy and bilateral axillary lymph node removal;

[0183] 10) Those who are concerned about the test results due to scars, pigmentation, atrophy, port wine stains, uneven skin color, folliculitis or other blemishes on the skin to be tested;

[0184] 11) Participating in other clinical trial researchers;

[0185] 12) People with highly sensitive constitution;

[0186] 13) Non-voluntary participants or those who are unable to complete the required content according to the test requirements.

[0187] Subject withdrawal criteria:

[0188] 1) Not selected (e.g., not meeting the inclusion criteria / or excluded by the exclusion criteria, etc.);

[0189] 2) The subject is determined to be ineligible after being selected (including but not limited to changes in other lifestyle, diet, skin care habits and skin care samples during the evaluation period);

[0190] 3) The subject chooses to withdraw;

[0191] 4) Adverse events or serious adverse events occur;

[0192] 5) Lost to follow-up;

[0193] 6) Force majeure;

[0194] 7) Others.

[0195] Test method based on:

[0196] 1) QT / ZHCA 005-2019 Test method for the effect of cosmetics on skin elasticity

[0197] 2) QB / T 4256-2011 Guidelines for the evaluation of moisturizing efficacy of cosmetics

[0198] 3) T / GDCA 009-2022 Evaluation Method of Cosmetic Repair Efficacy on Human Body

[0199] 4) T / ZHCA 003-2018 Test method for the effect of cosmetics on transepidermal water loss

[0200] 5) T / CNMIA 0015-2020 Clinical Evaluation Standard for Allergy-Relief Functional Skin Care Products

[0201] 6) T / GDCDC 021-2022 Test Method for Soothing Effect of Cosmetics

[0202] 7) Wang Huan, Pan Yao. Evaluation of cosmetic efficacy (V) - Scientific support for soothing efficacy claims[J]. Surfactant Detergent Industry, 2018, 48(05): 247-254

[0203] 8) T / ZHCA 005-2019 Test method for the effect of cosmetics on skin elasticity

[0204] 9) T / GDCDC 032-2023 Test method for cosmetic exfoliation efficacy

[0205] 10) T / ZHCA 006-2019 Test method for anti-wrinkle efficacy of cosmetics

[0206] 11) T / TDCA 003-2021 Test method for firming effect of cosmetics

[0207] 12) T / CAB 0152-2022 Test methods for seven cosmetic effects: anti-wrinkle, firming, moisturizing, oil control, repairing, nourishing, and soothing

[0208] 13) Liu Weiyi, Zhou Lin, Zhao Hua. Evaluation of cosmetic efficacy (ⅩⅢ) - Consumer use test[J]. Surfactant Detergent Industry, 2021, 51(06): 485-490

[0209] 14) "Technical Specifications for Safety of Cosmetics" (2015 Edition)

[0210] 15) Cosmetics classification rules and classification catalogue

[0211] 16) Evaluation Standards for Cosmetic Efficacy Claims

[0212] Testing process:

[0213] D0:

[0214] 1) Subject information is registered, informed consent is understood and signed, and laboratory technicians screen subjects according to inclusion and exclusion criteria;

[0215] 2) The subjects cleaned their entire face with the designated cleansing sample, cleaned the inside of their arms with water, and dried them with a dry paper towel. They then sat quietly in a laboratory at a temperature of 21±1°C and a humidity of 50±10% RH for 30 minutes;

[0216] 3) Mark two measurement areas on the inner side of the subject's forearm. The test area is 3cm*3cm, and the interval between each test area is at least 1cm;

[0217] 4) Before the formal test, sit quietly in a standard room for at least 30 minutes, do not drink water or beverages, expose your forearms, place them in the test position, and stay relaxed;

[0218] 5) The sample areas and blank areas are randomly distributed in the arm calibration area to ensure that the positions of all sample areas and blank areas are statistically balanced;

[0219] 6) Test the basic values ​​of the facial skin of the subjects: Bergman questionnaire, VECTRA H2 image acquisition, VISIA image acquisition, UC22 test of skin dermis density and skin dermis thickness, VC20 image acquisition, Glossymeter test of skin gloss, Corneometer test of skin stratum corneum moisture content, Cutometer test of skin elasticity R2 value and skin firmness F4 value, Tewameter test of skin transepidermal water loss rate;

[0220] 7) Test the basic value of the subject's arm skin: Corneometer tests the moisture content of the skin's stratum corneum;

[0221] 8) Place the test sample on the inside of the subject’s arm at a pressure of (2.0±0.1) mg / cm 2 Apply evenly on the sample area, leaving the blank area untreated;

[0222] 9) Note: Do not wash the inside of your arm during the 72-hour period, and do not apply any cosmetics, medicines, or other products that may affect the test results;

[0223] 10) Laboratory technicians instruct subjects on how to use samples according to sample usage requirements and usage sites, and provide written test precautions and instructions for use;

[0224] 11) The subject receives the sample and leaves the laboratory.

[0225] T72h:

[0226] 1) Skin data value after the test subject used the sample for 72 hours: Corneometer tests the moisture content of the skin stratum corneum;

[0227] 2) The subjects filled in the self-assessment questionnaire.

[0228] D28:

[0229] 1) The subjects cleaned their faces with the designated cleansing samples and sat in a laboratory at a temperature of 21±1°C and a humidity of 50±10% RH for 30 minutes;

[0230] 2) The skin data values ​​of the test subjects after using the samples for 28 days: Bergman questionnaire, VECTRA H2 image acquisition, VISIA image acquisition, UC22 test of skin dermis density and skin dermis thickness, VC20 image acquisition, Glossymeter test of skin gloss, Corneometer test of skin stratum corneum moisture content, Cutometer test of skin elasticity R2 value and skin firmness F4 value, Tewameter test of skin transepidermal water loss rate;

[0231] 3) Subjects fill in self-assessment questionnaires, collect samples and use logs.

[0232] Note: D0 refers to the visit before the subject uses the sample, T72h refers to the visit 72 hours after the subject uses the sample, D7 refers to the visit 7 days after the subject uses the sample, D14 refers to the visit 14 days after the subject uses the sample, D21 refers to the visit 21 days after the subject uses the sample, and D28 refers to the visit 28 days after the subject uses the sample.

[0233] In the following Examples 1-10, statistical analysis software was used to perform statistical analysis of the data. The statistical analysis was a two-tailed test with a significance level of α=0.05.

[0234] Instrument measurement: meet the requirements of normal distribution, and the paired t test is used for comparison before and after, otherwise the two related sample rank sum test is used;

[0235] Instrument measurement (sample area / blank area): Comparison between the test sample area and the blank area, if it meets the normal distribution requirements, use the paired t test, otherwise use the two related sample rank sum test;

[0236] 10-point scale evaluation: Two related sample rank sum tests were used for before-and-after comparison.

[0237] In the following Examples 1 to 10, the significance annotation method of the experimental results is: "ns" indicates no significant difference, P ≥ 0.05; "*" indicates a significant difference, 0.01 ≤ P < 0.05; "**" indicates a significant difference, 0.001 ≤ P < 0.01; "***" indicates a significant difference, P < 0.001.

[0238] Example 1

[0239] Compound Drimediol is used for preparing cosmetics with moisturizing effect

[0240] Test basis: Compared with before use, if the moisture content of the stratum corneum of the skin at any visit point (28 days) after using the sample is significantly improved (P<0.05), the sample is judged to have moisturizing effect, otherwise it is judged that the sample does not have moisturizing effect;

[0241] Test item: moisture content of cheek skin stratum corneum;

[0242] Test instrument / method: Corneometer CM825;

[0243] Test time: D0 / D28;

[0244] Test site: Cheek;

[0245] Parameter description: This instrument is based on the fact that the dielectric constants of water (81) and other substances (<7) vary greatly. According to the different water content, the measuring capacitor will change with the change of the capacitance of the skin, and the capacitance of the skin is within the measurement range, so the moisture content of the skin can be measured. The larger the measured value, the higher the moisture content of the skin stratum corneum.

[0246] The test results are shown in Table 2, Table 3 and Figure 1 ;

[0247] Table 2 Cheek skin stratum corneum moisture content test results (sample size = 32)

[0248]

[0249]

[0250] Note: 1) The larger the measured value, the higher the moisture content of the skin stratum corneum

[0251] 2) Improvement rate (%) = (test value after use - test value before use) / test value before use * 100%

[0252] Table 3 Statistical analysis of cheek skin stratum corneum moisture content test results

[0253]

[0254] Reference Figure 1 After using the sample for 28 days, the moisture content of the stratum corneum of the subjects' cheek skin increased by 45.84%, which was a significant difference.

[0255] Example 2

[0256] Compound Drimediol is used for preparing cosmetics with 72-hour moisturizing effect

[0257] Test basis: 72-hour moisturizing effect: If the moisture content of the stratum corneum at any revisit point (72 hours) after using the sample is significantly improved compared with that before use (P<0.05), or if the increment of the moisture content of the stratum corneum at any revisit point (72 hours) after using the sample is significantly higher than that of the blank area (P<0.05), the sample is judged to have 72-hour moisturizing effect; otherwise, the sample is judged not to have 72-hour moisturizing effect;

[0258] Test item: moisture content of the stratum corneum of the arm skin;

[0259] Test instrument / method: Corneometer CM825;

[0260] Test time point: D0 / T72h;

[0261] Test site: inner side of the arm (sample area / blank area);

[0262] Parameter description: This instrument is based on the fact that the dielectric constants of water (81) and other substances (<7) vary greatly. According to the different water content, the measuring capacitor will change with the change of the capacitance of the skin. The capacitance of the skin is within the measurement range, so the moisture content of the skin can be measured.

[0263] The larger the measured value, the higher the moisture content of the skin's stratum corneum.

[0264] The test results are shown in Table 4, Table 5, Figure 2 and Figure 3 ;

[0265] Table 4 Test results of moisture content of the stratum corneum of the arm skin (sample size = 32)

[0266]

[0267]

[0268] Note: 1) The larger the measured value, the higher the moisture content of the skin stratum corneum

[0269] 2) Improvement rate (%) = (test value after use - test value before use) / test value before use * 100%

[0270] Reference Figure 2 Compared with before use, after using the test sample for 72 hours, the moisture content of the stratum corneum of the arm skin in the sample area increased by 55.59%, which was a significant difference; after 72 hours, the moisture content of the stratum corneum of the arm skin in the blank area increased by 0.70%, which was not a significant difference.

[0271] Table 5 Statistical analysis of the differences in the test results of the moisture content of the stratum corneum of the arm skin

[0272]

[0273] Note: Incremental test value after use - test value before use

[0274] Reference Figure 3 Compared with the blank area, after using the sample for 72 hours, the increase in moisture content in the stratum corneum of the arm skin in the sample area was significantly higher than that in the blank area.

[0275] Example 3

[0276] Compound Drimediol is used to prepare cosmetics with repairing effects

[0277] Test basis: Compared with before use, if the transepidermal water loss rate of the skin at any visit point (28 days) after using the sample is significantly improved (P<0.05), the sample is judged to have a repair effect, otherwise it is judged that the sample does not have a repair effect;

[0278] Test item: Transepidermal water loss rate

[0279] Test instrument / method: Tewameter TM Hex;

[0280] Test time point: D0 / D28;

[0281] Test site: Cheek;

[0282] Parameter description: The Tewameter TM Hex probe contains 30 pairs of relative humidity and temperature sensors. The probe is like a camera that can see the distribution of relative humidity and temperature in the cavity. Through the large amount of data measured each time, the probe can quickly complete the accurate and repeatable test of TEWL value.

[0283] The smaller the measured value, the less transepidermal water loss per unit time and unit cross-sectional area, and the better the skin barrier.

[0284] The test results are shown in Tables 6, 7 and Figure 4 ;

[0285] Table 6 Skin transepidermal water loss rate test results (sample size = 32)

[0286]

[0287] Note: 1) The smaller the measured value, the less transepidermal water loss per unit time and per unit cross-sectional area, indicating a better skin barrier;

[0288] 2) Reduction rate (%) = (test value before use - test value after use) / test value before use * 100%;

[0289] Table 7 Statistical analysis of skin transepidermal water loss rate test results

[0290]

[0291] Reference Figure 4 Compared with before use, after 28 days of using the sample, the transepidermal water loss rate of the skin decreased by 22.25%, and the difference was significant.

[0292] Example 4

[0293] Compound Drimediol is used for preparing cosmetics with nourishing effect

[0294] Test basis: Compared with before use, if two or more parameters at any visit point (28 days) (skin roughness parameter SEr value / skin smoothness parameter SEsm value), skin glossiness, skin elasticity R2 value, and skin dermis density are significantly improved (P<0.05) after using the sample, the sample is judged to have a nourishing effect, otherwise it is judged that the sample does not have a nourishing effect;

[0295] Test Item 1: Cheek skin roughness parameter SEr value, skin smoothness SEsm value; elbow skin roughness parameter SEr value, elbow skin smoothness SEsm value;

[0296] Test instrument / method: VisioScanVC20plus;

[0297] Test time point: D0 / D28;

[0298] Test site: Cheek;

[0299] Parameter description: The instrument uses a dermatoscope consisting of a uniform annular ultraviolet (UVA) illumination source and a black and white high-resolution CCD camera to capture images of the skin surface. The images are transmitted to the host for digital processing and analysis.

[0300] The larger the SEr value, the smaller the skin roughness.

[0301] The smaller the SEsm value, the better the skin smoothness.

[0302] The test results of cheek skin roughness parameter SEr value and skin smoothness SEsm value are shown in Table 8, Table 9, Figure 5 , Table 10, Table 11 and Figure 6 ;

[0303] Table 8 Cheek skin roughness parameter SEr value test results (sample size = 32)

[0304]

[0305] Note: 1) The larger the SEr value, the smaller the skin roughness

[0306] 2) Improvement rate (%) = (test value after use - test value before use) / test value before use * 100%

[0307] Table 9 Statistical analysis of cheek skin roughness parameter SEr value test results

[0308]

[0309] Please refer to Figure 5 Compared with before use, after 28 days of using the sample, the cheek skin roughness parameter SEr value increased by 56.67%, and the difference was significant.

[0310] Table 10 Cheek skin smoothness parameter SEsm value test results (sample size = 32)

[0311]

[0312] Note: 1) The smaller the SEsm value, the better the skin smoothness

[0313] 2) Reduction rate (%) = (test value before use - test value after use) / test value before use * 100%

[0314] Table 11 Statistical analysis of cheek skin smoothness parameter SEsm value test results

[0315]

[0316] Reference Figure 6 , Fig.21 as well as Fig. 22 Compared with before use, after using the sample for 28 days, the cheek skin smoothness parameter SEsm value decreased by 11.22%, and the difference was significant.

[0317] The test results of elbow skin roughness parameter SEr value are shown in Table 12, Table 13 and Figure 7 ;

[0318] Table 12 Statistical analysis of the test results of SEr value of elbow skin roughness parameter

[0319]

[0320] Note: 1) The larger the SEr value, the smaller the skin roughness

[0321] 2) Improvement rate (%) = (test value after use - test value before use) / test value before use * 100%

[0322] Table 13 Statistical analysis of the test results of SEr value of elbow skin roughness parameter

[0323]

[0324] Please refer to Figure 7 Compared with before use, after 28 days of using the sample, the SEr value of the elbow skin roughness parameter increased by 48.84%, and the difference was significant.

[0325] The test results of elbow skin smoothness SEsm value are shown in Table 14, Table 15 and Figure 8 ;

[0326] Table 14 Elbow skin smoothness parameter SEsm value test results (sample size = 32)

[0327]

[0328] Note: 1) The smaller the SEsm value, the better the skin smoothness

[0329] 2) Reduction rate (%) = (test value before use - test value after use) / test value before use * 100%

[0330] Table 15 Statistical analysis of the test results of elbow skin smoothness parameter SEsm value

[0331]

[0332] Reference Figure 8 Compared with before use, after 28 days of using the sample, the elbow skin smoothness parameter SEsm value decreased by 9.92%, and the difference was significant.

[0333] Test item 2: skin gloss;

[0334] Test instrument / method: Glossymeter GL 200

[0335] Test time point: D0 / D28;

[0336] Test site: Cheek;

[0337] Parameter description: The instrument reflects the direct reflection and scattered reflection of light irradiated to the skin surface.

[0338] The larger the value, the more shiny the skin will be.

[0339] The test results are shown in Tables 16, 17 and Fig. 9 ;

[0340] Table 16 Skin gloss test results (sample size = 32)

[0341]

[0342] Note: 1) The larger the value, the higher the skin glossiness

[0343] 2) Improvement rate (%) = (test value after use - test value before use) / test value before use * 100%

[0344] Table 17 Statistical analysis of skin gloss test results

[0345]

[0346] Reference Fig. 9 Compared with before use, after 28 days of using the sample, the skin glossiness increased by 38.28%, which was a significant difference.

[0347] Test item 3: Skin elasticity R2 value;

[0348] Test instrument / method: Cutometer dual MPA580

[0349] Test time point: D0 / D28;

[0350] Test site: Cheek;

[0351] Parameter description: The testing principle of this instrument is based on the principles of suction and stretching. A negative pressure is generated on the surface of the skin being tested to suck the skin into a specific test probe. The depth of the skin sucked into the test probe is measured by a non-contact optical test system.

[0352] The larger the R2 value, the better the skin elasticity

[0353] The test results are shown in Tables 18, 19 and Fig.10 ;

[0354] Table 18 Skin elasticity R2 value test results (sample size = 32)

[0355]

[0356] Note: 1) The larger the R2 value, the better the skin elasticity

[0357] 2) Improvement rate (%) = (test value after use - test value before use) / test value before use * 100%

[0358] Table 19 Statistical analysis of skin elasticity R2 value test results

[0359]

[0360] Reference Fig.10 Compared with before use, after 28 days of using the sample, the skin elasticity R2 value increased by 13.21%, and the difference was significant.

[0361] Test item 4: Skin dermis thickness, skin dermis density;

[0362] Test instrument / method: Ultrascan UC22

[0363] Test time point: D0 / D28;

[0364] Test site: Cheek;

[0365] Parameter description: The pulse generator generates a short electrical pulse signal, which is converted into an ultrasonic signal with a frequency of 22MHz by piezoelectric means. The ultrasonic signal is then transmitted into the skin, and the uneven substances in the skin, such as cell formations and blood vessels, produce reflections or echoes, which are then received by the sensor and converted into electrical signals. Through the receiver controlled by the motor, these ultrasonic signals are received and processed to generate a cross-sectional image. The ultrasonic signal is digitally processed, stored and analyzed by a computer, and finally an ultrasonic image is formed. By separating and digitally processing the high-frequency ultrasonic signal, a skin ultrasonic image with high resolution and good contrast is finally obtained.

[0366] The larger the value, the thicker the skin dermis.

[0367] The larger the value, the denser the skin dermis.

[0368] The results of the skin dermis thickness test are shown in Table 20, Table 21 and Fig.11 ;

[0369] Table 20 Skin dermis thickness test results (sample size = 32)

[0370]

[0371] Note: 1) The larger the value, the thicker the dermis layer of the skin

[0372] 2) Improvement rate (%) = (test value after use - test value before use) / test value before use * 100%

[0373] Table 21 Statistical analysis of skin dermis thickness test results

[0374]

[0375] Reference Fig.11 and Fig.23 Compared with before use, after 28 days of using the sample, the thickness of the dermis layer increased by 10.89%, which was a significant difference.

[0376] The results of the skin dermis density test are shown in Table 22, Table 23 and Fig.12 ;

[0377] Table 22 Skin dermis density test results (sample size = 32)

[0378]

[0379]

[0380] Note: 1) The larger the value, the greater the density of the dermis layer

[0381] 2) Improvement rate (%) = (test value after use - test value before use) / test value before use * 100%

[0382] Table 23 Statistical analysis of skin dermis density test results

[0383]

[0384] Reference Fig.12 and Fig.23 Compared with before use, after 28 days of using the sample, the density of the dermis layer increased by 24.99%, which was a significant difference.

[0385] Example 5

[0386] Compound Drimediol is used for preparing cosmetics with soothing effect

[0387] Test basis: Compared with before use, if the a* value of the skin red area at any revisit point (28 days) after using the sample is significantly improved (P<0.05), the sample is judged to have a soothing effect, otherwise the sample is judged to have no soothing effect;

[0388] Test Item 1: Skin red zone a* value

[0389] Test instrument / method: IPP;

[0390] Test time point: D0 / D28;

[0391] Test site: Cheek;

[0392] Parameter description: The smaller the value, the less red the skin will be;

[0393] The test results are shown in Table 24, Table 25 and Fig.13 ;

[0394] Table 24 Skin red zone a* value test results (sample size = 32)

[0395]

[0396] Table 25 Statistical analysis of skin red zone a* value test results

[0397]

[0398] Note: 1) The smaller the value, the less red the skin

[0399] 2) Reduction rate (%) = (test value before use - test value after use) / test value before use * 100%

[0400] Reference Fig.13 as well as Fig.25Compared with before use, after 28 days of use, the a* value of the skin red area decreased by 5.46%, which was a significant difference.

[0401] Test Item 2: Skin Color L* Value

[0402] Test instrument / method: IPP;

[0403] Test time point: D0 / D28;

[0404] Test site: Cheek;

[0405] Parameter description: The larger the L* value, the brighter the skin;

[0406] The test results are shown in Table 26, Table 27 and Fig.14 ;

[0407] Table 26 Skin color L* value test results (sample size = 32)

[0408]

[0409] Note: 1) The larger the L* value, the brighter the skin

[0410] 2) Improvement rate (%) = (test value after use - test value before use) / test value before use * 100%

[0411] Table 27 Statistical analysis of skin color L* value test results

[0412]

[0413] Reference Fig.14 as well as Fig.24 Compared with before use, after using the sample for 28 days, the L* value of skin color increased by 1.82%, which is a significant difference.

[0414] Example 6

[0415] Compound Drimediol is used for preparing cosmetics with firming effect

[0416] Test basis: Compared with before use, if the skin elasticity R2 value and firmness F4 value at any visit point (28 days) after using the sample are significantly improved (P<0.05), the sample is judged to have a firming effect, otherwise it is judged that the sample does not have a firming effect;

[0417] Test item: Skin firmness F4 value

[0418] Test instrument / method: Cutometer dual MPA580;

[0419] Test time point: D0 / D28;

[0420] Test site: Cheek;

[0421] Parameter description: The testing principle of this instrument is based on the principles of suction and stretching. A negative pressure is generated on the surface of the skin being tested to suck the skin into a specific test probe. The depth of the skin sucked into the test probe is measured by a non-contact optical test system.

[0422] The smaller the F4 value, the better the skin firmness

[0423] The test results are shown in Table 28, Table 29 and Fig.15 ;

[0424] Table 28 Skin firmness F4 value test results (sample size = 32)

[0425]

[0426] Note: 1) The smaller the F4 value, the better the skin firmness

[0427] 2) Reduction rate (%) = (test value before use - test value after use) / test value before use * 100%

[0428] Table 29 Statistical analysis of skin firmness F4 value test results

[0429]

[0430] Reference Fig.15 Compared with before use, after 28 days of using the sample, the skin firmness F4 value decreased by 15.42%, and the difference was significant.

[0431] Example 7

[0432] Compound Drimediol is used for preparing cosmetics with anti-wrinkle effect

[0433] Test basis: If any wrinkle parameter at any visit point (28 days) after using the sample is significantly improved compared with that before use (P<0.05), the sample is judged to have anti-wrinkle effect; otherwise, the sample is judged to have no anti-wrinkle effect;

[0434] Test item: Area ratio of under-eye wrinkles

[0435] Test instrument / method: IPP;

[0436] Test time point: D0 / D28;

[0437] Test site: Cheek;

[0438] Parameter description: The smaller the value, the smaller the area of ​​under-eye wrinkles;

[0439] The test results are shown in Table 30, Table 31 and Fig.16 ;

[0440] Table 30 Test results of area ratio of under-eye wrinkles (sample size = 32)

[0441]

[0442] Note: 1) The smaller the value, the smaller the area of ​​under-eye wrinkles

[0443] 2) Reduction rate (%) = (test value before use - test value after use) / test value before use * 100%

[0444] Table 31 Statistical analysis of the test results of the area ratio of under-eye wrinkles

[0445]

[0446] Reference Fig.16 and Fig.26 Compared with before use, after 28 days of using the sample, the area ratio of under-eye wrinkles decreased by 15.42%, which was a significant difference.

[0447] Example 8

[0448] Compound Drimediol is used for preparing cosmetics with the effects of increasing the distance of the apple muscles and the angle of the mandibular angle

[0449] Test items: distance of upward movement of apple muscles, angle of mandibular angle

[0450] Test instrument / method: VECTRA H2;

[0451] Test time point: D0 / D28;

[0452] Test site: Cheek;

[0453] Parameter description: This instrument is a portable stereoscopic imaging camera that presents the contours of the face, breasts and body at 360 degrees, analyzes the skin base and the optimal golden ratio;

[0454] The smaller the value, the better the face lifting effect;

[0455] The smaller the value, the better the face lifting effect;

[0456] The test results are shown in Table 32, Table 33, Fig.17 , Table 34, Table 35 and Fig.18 ;

[0457] Table 32 Mandibular angle test results (sample size = 32)

[0458]

[0459] Note: 1) The smaller the value, the better the face lifting effect

[0460] 2) Reduction rate (%) = (test value before use - test value after use) / test value before use * 100%

[0461] Table 33 Statistical analysis of mandibular angle test results

[0462]

[0463] Reference Fig.17 as well as Fig. 27 Compared with before use, after 28 days of using the sample, the mandibular angle reduction rate was 1.53%, and the difference was significant.

[0464] Table 34 Apple muscle upward displacement test results (sample size = 32)

[0465]

[0466] Note: 1) The smaller the value, the better the face lifting effect

[0467] 2) Reduction rate (%) = (test value before use - test value after use) / test value before use * 100%

[0468] Table 35 Statistical analysis of the test results of the upward movement distance of the apple muscle

[0469]

[0470] Reference Fig.18 as well as Fig.28 , compared with before use, after 28 days of using the sample, the upward movement distance of the apple muscles decreased by 2.53%.

[0471] Example 9

[0472] Compound Drimendiol is used to prepare cosmetics suitable for sensitive skin

[0473] Since all the subjects had sensitive skin, no adverse reactions occurred on their skin within 28 days after using the cosmetic samples containing the compound Drimendiol, and the stinging, burning, tightness, itching, overall discomfort and redness of the sensitive skin were greatly improved.

[0474] Improvement rate = (self-assessment score before use - self-assessment score after use) / self-assessment score before use * 100% Questionnaire on a 10-point scale: Skin tingling: the lighter the skin tingling, the lower the score; Skin burning: the lighter the skin burning, the lower the score; Skin tightness: the lighter the skin tightness, the lower the score; Skin itching: the lighter the skin itching, the lower the score; Overall skin discomfort: the lighter the skin discomfort, the lower the score; Skin redness: the lighter the skin redness, the lower the score.

[0475] Reference Fig.19 Compared with before use, after using the sample for 28 days, the subjects' self-rated skin stinging, skin burning, skin tightness, skin itching, skin overall discomfort, and skin redness were significantly improved, with improvement rates of 69.68%, 69.74%, 77.34%, 78.97%, 79.51%, and 79.09%, respectively. It proves that the compound Drimendiol can be used to prepare cosmetics suitable for sensitive skin.

[0476] The self-evaluation results after the subjects used the samples are shown in Tables 36, 37 and Fig.19 ;

[0477] Table 36 Results of the 10-point questionnaire

[0478]

[0479]

[0480] Table 37 Subjective evaluation of skin feeling before and after using samples

[0481] Feelings after using the sample-1 D7 Recognition D14 Recognition D21 Recognition D28 Recognition Did you experience any itching during use? 100.00% No 100.00% No 100.00% No 100.00% No Did you experience any tingling sensation during use? 100.00% No 100.00% No 100.00% No 100.00% No Did you feel any heat during use? 100.00% No 100.00% No 100.00% No 100.00% No Did you experience any redness during use? 100.00% No 100.00% No 100.00% No 100.00% No Did edema occur during use? 100.00% No 100.00% No 100.00% No 100.00% No Do you experience any reactions such as papules / rashes / protrusions during use? 100.00% No 100.00% No 100.00% No 100.00% No Whether there is a blister reaction during use 100.00% No 100.00% No 100.00% No 100.00% No Whether there is any wetting reaction during use 100.00% No 100.00% No 100.00% No 100.00% No

[0482] Evaluation index (feelings after using the sample - 1): 1 point means "yes", 2 points means "no"; the recognition degree is the percentage of "2 points no".

[0483] Example 10

[0484] Use of compound Drimendiol in preparing mild and non-irritating cosmetics

[0485] The adverse reaction feedback of 32 subjects during the use of samples is shown in Tables 38 and 39;

[0486] Table 38 Human trial test results

[0487]

[0488] Table 39 Classification criteria for adverse skin reactions

[0489]

[0490]

[0491] Note: Refer to the 2015 edition of the "Safety Technical Specifications for Cosmetics" for the classification standards for adverse skin reactions in human trials.

[0492] Result description:

[0493] After 72 hours of using the sample, the skin reactions in the observed areas were all grade 0, and no adverse skin reactions occurred;

[0494] After using the samples for 7 days, the skin reactions in the observed areas were all grade 0, and no adverse reactions occurred on the skin;

[0495] After using the samples for 14 days, the skin reactions in the observed areas were all grade 0, and no adverse reactions occurred on the skin;

[0496] After using the samples for 21 days, the skin reactions in the observed areas were all grade 0, and no adverse reactions occurred on the skin;

[0497] After using the samples for 28 days, the skin reactions in the observed areas were all grade 0, and no adverse skin reactions occurred.

[0498] The subjective evaluation of 32 subjects is shown in Table 40 and Fig. 20 ;

[0499] Table 40 Self-evaluation results of subjects after using the samples for 28 days

[0500] Feelings after using the sample-2 T72h Recognition After using the sample, there is a moisturizing effect (face) 100.00％ Skin feels more hydrated after using the sample (face) 100.00％ After using the sample, there is a moisturizing effect (arm) 100.00％ After using the sample, the skin feels more hydrated (arm) 100.00％

[0501] Evaluation index (feelings after using the sample - 2): 1 point means disagreement, 2 points means relatively disagreement, 3 points means neither disagreement nor agreement, 4 points means relatively agreement, and 5 points means agreement; the degree of agreement is the sum of the percentages of "4 points" and "5 points".

[0502] The above human trial test results and the statistical results of adverse skin reactions prove that the compound Drimendiol can be used to prepare mild and non-irritating cosmetics.

[0503] In summary, 32 subjects with sensitive skin (Baumann Questionnaire) used the test sample (20ppm Drimendiol) for 28 consecutive days, and the results showed the following effects:

[0504] 1) Compared with before use, after using the sample for 72 hours, the moisture content of the stratum corneum of the arm skin in the sample area increased significantly (P<0.05), with an increase rate of 55.59%; compared with the blank area, after using the sample for 72 hours, the increment of the moisture content of the stratum corneum of the arm skin in the sample area was significantly higher than that in the blank area (P<0.05), indicating that under this test condition, the sample has a 72-hour moisturizing effect;

[0505] 2) Compared with before use, after using the sample for 72 hours, the moisture content of the stratum corneum of the skin on the arm of the sample area increased significantly (P<0.05), with an increase rate of 55.59%; compared with the blank area, after using the sample for 72 hours, the increment of moisture content of the stratum corneum of the skin on the arm of the sample area was significantly higher than that of the blank area (P<0.05); compared with before use, after using the sample for 28 days, the moisture content of the stratum corneum of the skin on the cheek increased significantly (P<0.05), with an increase rate of 45.84%; compared with before use, after using the sample for 28 days, the transepidermal water loss rate of the skin decreased significantly (P<0.05), with a decrease rate of 22.25%, indicating that the sample has moisturizing and repairing effects under this test condition;

[0506] 3) Compared with before use, after using the sample for 28 days, the skin elasticity R2 value was significantly increased (P<0.05), and the increase rate was 13.21%; compared with before use, after using the sample for 28 days, the skin firmness F4 value was significantly decreased (P<0.05), and the decrease rate was 15.42%, indicating that under this test condition, the sample has a firming effect;

[0507] 4) Compared with before use, after using the sample for 28 days, the SEr value of cheek skin roughness parameter increased significantly (P<0.05), with an increase rate of 56.67%; compared with before use, after using the sample for 28 days, the SEsm value of cheek skin smoothness parameter decreased significantly (P<0.05), with a decrease rate of 11.22%; compared with before use, after using the sample for 28 days, the SEr value of elbow skin roughness parameter increased significantly (P<0.05), with an increase rate of 48.84%; compared with before use, after using the sample for 28 days, the SEsm value of elbow skin smoothness parameter decreased significantly (P<0.05), with a decrease rate of 9.92%; compared with before use, after using the sample for 28 days, the skin glossiness increased significantly (P<0.05), with an increase rate of 38.28%;

[0508] Compared with before use, after using the sample for 28 days, the skin elasticity R2 value was significantly increased (P<0.05), and the increase rate was 13.21%; compared with before use, after using the sample for 28 days, the skin firmness F4 value was significantly reduced (P<0.05), and the reduction rate was 15.42%; compared with before use, after using the sample for 28 days, the skin dermis density was significantly increased (P<0.05), and the increase rate was 24.99%, indicating that under the test conditions, the sample has a nourishing effect;

[0509] 5) Compared with before use, after using the sample for 28 days, the area ratio of under-eye wrinkles was significantly reduced (P<0.05), with a reduction rate of 15.42%, indicating that the sample has anti-wrinkle effect under this test condition;

[0510] 6) Compared with before use, after using the sample for 28 days, the a* value of the skin red area was significantly reduced (P<0.05), with a reduction rate of 5.46%, indicating that the sample has a soothing effect under this test condition;

[0511] 7) Compared with before use, after using the sample for 28 days, the skin color L* value increased by 1.82%, and the difference was significant. This indicates that under this test condition, the sample has the effect of brightening the skin;

[0512] 8) Compared with before use, after using the sample for 28 days, the upward movement distance of the apple muscles decreased by 2.53%, and the difference was significant. This indicates that under this test condition, the sample has the effect of upward movement of the apple muscles;

[0513] 9) Compared with before use, after using the sample for 28 days, the mandibular angle reduction rate was 1.53%, and the difference was significant. This indicates that under this test condition, the sample has the effect of improving the mandibular angle;

[0514] 10) After using the sample for 28 days, the subjects with sensitive skin showed no adverse skin reactions related to the sample, 100% of the subjects agreed that the sample was mild and non-irritating, and 100% of the subjects agreed that the sample was suitable for sensitive skin, indicating that under the test conditions, the sample was mild and non-irritating and suitable for sensitive skin.

[0515] Conclusion: Under the test conditions, the sample has 72-hour moisturizing, hydrating, repairing, firming, nourishing, anti-wrinkle, soothing, skin brightening, apple muscle lifting, and mandibular angle improvement effects; it is mild and non-irritating and suitable for sensitive skin.

[0516] Embodiment 11

[0517] Compound Drimendiol is used for preparing cosmetics with positive regulation of the expression of acid sphingomyelinase SMPD1 gene and loricrin LOR gene and the effect of improving skin barrier

[0518] 1. Experimental Principle

[0519] Acid sphingomyelinase (SMPD1) is a key lipid-degrading enzyme that exhibits maximum activity at low pH. Acidic pH also affects the interactions of the lipid bilayer in the stratum corneum, which can significantly improve the integrity and cohesion of the skin. The integrity of the skin barrier is essential for retaining moisture in the skin.

[0520] Loricrin is the main component of the keratinocyte seal, accounting for about 80% of the total keratinocyte seal protein. It is mainly present in the epidermal granular layer and plays an important role in the barrier function of the epidermis. The main functions of loricrin are: (1) strengthening the keratinocyte seal and enhancing the permeability barrier; (2) loricrin interacts with intermediate filaments to increase the flexibility of the keratinocyte seal structure. Loricrin itself does not directly have a moisturizing function, but indirectly affects the moisturizing state of the skin through its role in the skin barrier.

[0521] A method was established with the relative expression of SMPD1 and LOR mRNA detected by fluorescence quantitative RT-qPCR as the detection endpoint. The difference in relative expression of factor mRNA after administration of the blank control and the test substance was compared to evaluate whether the test substance could upregulate moisturizing-related genes.

[0522] 2. Test materials and test samples

[0523] 2.1 Experimental Reagents

[0524] High-glucose DMEM culture medium, fetal bovine serum, trypsin / EDTA solution, PBS, penicillin and streptomycin double antibody mixture, trypan blue dye, thiazolyl blue MTT, total RNA extraction kit, reverse transcription kit, fluorescent quantitative PCR kit, cell line: human immortalized epidermal cell line.

[0525] 2.2 Experimental instruments

[0526] CO2 incubator, biological safety cabinet, analytical balance, inverted microscope, microplate reader, refrigerated centrifuge, fluorescence quantitative PCR instrument, PCR instrument.

[0527] 2.3 Test sample: 2000ppm Drimendiol 1,3-butanediol solution.

[0528] 3. Experimental Procedure

[0529] 3.1 Cell viability test (MTT)

[0530] 3.1.1 Routine cell culture: The cell suspension was inoculated into a 96-well cell culture plate and cultured for 24 hours.

[0531] 3.1.2 Take 300 μL of sample and dissolve it in 1200 μL of culture medium until fully dissolved.

[0532] 3.1.3 Add the same volume and different concentrations of the sample to be tested to each well and return to the incubator for incubation.

[0533] 3.1.4 Add MTT solution to each well and incubate in the incubator.

[0534] 3.1.5 Discard the liquid in the wells, add DMSO to each well, shake evenly, and measure the absorbance using an enzyme-labeled instrument.

[0535] 3.2 Determination of relative gene expression levels of SMPD1 and LOR.

[0536] 3.2.1 Carry out the test according to the test scheme shown in the table below.

[0537] 3.2.2 Routine cell culture: The cell suspension was inoculated into a 6-well cell culture plate and cultured for 24 hours.

[0538] Table 41 is the experimental group processing information table.

[0539] Table 41 Experimental group processing information table

[0540]

[0541] 3.2.3 Aspirate the culture medium from the wells and add different treatment groups to each well according to the table above. Return the 6-well plate to the cell culture incubator and culture for 48 hours. Collect the cells into a centrifuge tube.

[0542] 3.2.4 Extract total RNA using the total RNA extraction kit.

[0543] 3.2.5 Reverse transcribe the extracted total RNA into cDNA according to the reverse transcription kit steps.

[0544] 3.2.6 Prepare primers, probes, and multiplex PCR reaction system and distribute them to each reaction tube, then add cDNA respectively.

[0545] 3.2.7 Open the PCR reaction program, set the appropriate experiment type and mode to start running, and obtain experimental data.

[0546] 4. Data Analysis

[0547] 4.1 Relative cell activity

[0548] Assuming that the cell activity of the blank control group was 100%, the relative activity of cells in each group was calculated.

[0549]

[0550] OD TA : The average value of the optical density of the tested sample group for 3 times

[0551] OD B : The average value of the optical density of the blank control group for 3 times

[0552] 4.2 Gene expression level to be measured

[0553] ΔCT(treatment group) = CT(test gene) - CT(mean value of internal reference gene)

[0554] ΔCT (control group) = CT (test gene) - CT (mean value of internal reference gene)

[0555] ΔΔCT = ΔCT (treated group) - ΔCT (control group)

[0556] Relative expression of the gene to be tested = 2^_△△ CT , take 2^_△△ CT The mean

[0557] 5. Experimental Results

[0558] 5.1 Cell activity test results

[0559] Follow the steps in 3.1 to perform the MTT test. Five sample concentrations were designed for this cell activity test (eg Fig.29 When the sample concentration is ≤0.1%, the cell activity is greater than 90%, and all of them can be used for subsequent experiments. Table 42 shows the relative activity of cells in each group (Mean±SD).

[0560] Table 42 Relative activity of cells in each group (Mean±SD)

[0561] concentration(%) active(%) 0.01 105.99±5.78 0.1 107.63±5.37 1 68.58±3.80 5 19.51±0.57 10 18.52±0.62

[0562] Sample concentrations of 0.025%, 0.05%, and 0.1% were selected as test concentrations for subsequent efficacy experiments.

[0563] 5.2 Relative expression of SMPD1 gene

[0564] According to the sample concentration selected in 5.1, the expression level of SMPD1 was detected. The test results are shown in Table 43 and Fig.30 shown.

[0565] Table 43 Relative expression of SMPD1 in each group (Mean±SD)

[0566]

[0567] Note: "ns" means no significant difference, P ≥ 0.05; * means the difference is statistically significant (0.01 ≤ P < 0.05) compared with the blank control group (B); ** means the difference is extremely significant (P < 0.01) compared with the blank control group (B)

[0568] 5.3 Relative expression of LOR genes

[0569] According to the sample concentration selected in 5.1, LOR expression was detected. The test results are shown in Table 44 and Fig.31 shown.

[0570] Table 44 Relative expression of LOR in each group (Mean±SD)

[0571]

[0572] Note: "ns" means no significant difference, P ≥ 0.05; * means the difference is statistically significant (0.01 ≤ P < 0.05) compared with the blank control group (B); ** means the difference is extremely significant (P < 0.01) compared with the blank control group (B)

[0573] Conclusion: The relative expression levels of SMPD1 gene in samples at concentrations of 0.025% (0.5ppm Drimendiol), 0.05% (1ppm Drimendiol), and 0.1% (2ppm Drimendiol) were 1.61±0.01, 1.54±0.02, and 1.38±0.04, respectively, which were significantly increased compared with the blank control (P<0.01). This indicates that the samples at the above concentrations have the effect of upregulating the SMPD1 gene.

[0574] The relative expression levels of LOR genes in samples at concentrations of 0.025% (0.5ppm Drimendiol), 0.05% (1ppm Drimendiol), and 0.1% (2ppm Drimendiol) were 1.86±0.11, 1.64±0.08, and 2.05±0.13, respectively, which were significantly increased compared with the blank control (P<0.01), indicating that the samples at the above concentrations had the effect of upregulating LOR genes.

[0575] Example 12

[0576] Compound Drimendiol is used for preparing cosmetics with the effect of positively regulating the expression of elastin ELN gene and improving skin elasticity and firmness

[0577] 1. Experimental Principle

[0578] When UVA stimulates human skin fibroblasts, the cells undergo oxidative damage, triggering changes in the expression of aging-related genes and related enzymes, inducing an inflammatory cascade and reducing the expression of elastin, leading to aging phenomena such as sagging and wrinkles in the skin. Elastin (ELN) is an important protein in the skin that gives the skin firmness and elasticity, allowing the skin to stretch and return to its original shape. As we age, the elastin in the skin gradually decreases, causing the skin to lose elasticity and firmness, resulting in wrinkles and sagging.

[0579] This project uses a human dermal fibroblast model and uses fluorescence quantitative RT-qPCR (RT-qPCR) to detect the relative expression of mRNA as the detection endpoint. By comparing the difference in relative expression of factor mRNA after administration of the blank control and the test substance, the gene regulatory effect of the test substance on ELN is evaluated.

[0580] 2. Test materials and test samples

[0581] 2.1 Experimental Reagents

[0582] High-glucose DMEM culture medium, fetal bovine serum, penicillin-streptomycin mixture, trypan blue dye, PBS, thiazolyl blue MTT, RNA extraction kit, reverse transcription kit, real-time quantitative PCR detection kit, cell line: human skin fibroblasts.

[0583] 2.2 Experimental instruments

[0584] Cell culture box, biological safety cabinet, inverted microscope, real-time fluorescence quantitative PCR instrument, centrifuge

[0585] 2.3 Test sample: 2000ppm Drimendiol 1,3-butanediol solution.

[0586] 3. Experimental Procedure

[0587] 3.1 Cell viability test (MTT)

[0588] 3.1.1 Routine cell culture: The cell suspension was inoculated into a 96-well cell culture plate and cultured for 24 hours.

[0589] 3.1.2 Take 300 μL of sample and dissolve it in 1200 μL of culture medium until fully dissolved.

[0590] 3.1.3 Add the same volume and different concentrations of the sample to be tested to each well and return to the incubator for incubation.

[0591] 3.1.4 Add MTT solution to each well and incubate in the incubator.

[0592] 3.1.5 Discard the liquid in the wells, add DMSO to each well, shake evenly, and measure the absorbance using an enzyme-labeled instrument.

[0593] 3.2 Determination of ELN relative gene expression

[0594] 3.2.1 Carry out the test according to the test scheme shown in the table below.

[0595] 3.2.2 Routine cell culture: The cell suspension was inoculated into a 6-well cell culture plate and cultured for 24 hours.

[0596] Table 45 Experimental group processing information table

[0597]

[0598] 3.2.3 Aspirate the culture medium from the wells and add different treatment groups to each well according to the table above. Return the 6-well plate to the cell culture incubator and culture for 24 hours. Collect the cells into a centrifuge tube.

[0599] 3.2.4 RNA extraction: sample lysis—removal of genomic DNA—adsorption of RNA—removal of impurities—elution of RNA—RNA purity and concentration detection.

[0600] 3.2.5 RNA reverse transcription: remove genomic DNA - reverse transcription - obtain cDNA.

[0601] 3.2.6 Prepare primers, probes, and multiplex PCR reaction system and distribute them to each reaction tube, then add cDNA respectively.

[0602] 3.2.7 Open the PCR reaction program, set the appropriate experiment type and mode to start running, and obtain experimental data.

[0603] 4. Data Analysis

[0604] 4.1 Relative cell activity

[0605] Assuming that the cell activity of the blank control group was 100%, the relative activity of cells in each group was calculated.

[0606]

[0607] OD TA : The average value of the optical density of the tested sample group for 3 times

[0608] OD B : The average value of the optical density of the blank control group for 3 times

[0609] 4.2 Gene expression level to be measured

[0610] ΔCT(treatment group) = CT(test gene) - CT(mean value of internal reference gene)

[0611] ΔCT (control group) = CT (test gene) - CT (mean value of internal reference gene)

[0612] ΔΔCT = ΔCT (treated group) - ΔCT (control group)

[0613] Relative expression of the gene to be tested = 2^_△△ CT , take 2^_△△ CT The mean

[0614] 5. Experimental results

[0615] 5.1 Cell activity test results

[0616] Perform the MTT test according to step 3.1. Five sample concentrations were designed for this cell activity test (eg Fig.32 When the sample concentration is ≤1%, the cell activity is greater than 90% and can be used for subsequent experiments.

[0617] Table 46 shows the relative activity of cells in each group (Mean±SD).

[0618] Table 46 Relative activity of cells in each group (Mean±SD)

[0619] concentration(%) active(%) 0.01 96.44±9.66 0.1 102.04±9.47 1 97.48±7.52 5 38.99±4.00 10 37.26±5.83

[0620] Fig.32 In the experiment, sample concentrations of 0.25%, 0.5%, and 1.0% were selected as test concentrations for subsequent efficacy experiments.

[0621] 5.2 Relative expression of ELN genes

[0622] According to the sample concentration selected in 5.1, ELN expression was detected. The test results are shown in Tables 47 and Fig.33 shown.

[0623] Table 47 Relative expression of ELN in each group (Mean±SD)

[0624]

[0625] Note: "ns" means no significant difference, P ≥ 0.05; * means the difference is statistically significant (0.01 ≤ P < 0.05) compared with the blank control group (B); ** means the difference is extremely significant (P < 0.01) compared with the blank control group (B)

[0626] Conclusion: The relative expression of ELN gene in the samples at the concentration of 0.25% (5ppm Drimendiol), 0.5% (10ppm Drimendiol), and 1% (20ppm Drimendiol) were 3.85±0.13, 2.86±0.20, and 6.57±0.37, respectively, which were significantly upregulated compared with the blank control (P<0.01). This indicates that the samples have the effect of upregulating ELN gene at this concentration, which can be used as one of the evidences to support the claim of firming and anti-wrinkle effects.

[0627] Embodiment 13

[0628] Compound Drimendiol is used to prepare cosmetics that have the effects of negatively regulating the expression of tumor necrosis factor-α and interleukin-1α genes, improving skin inflammation, and soothing sensitive skin

[0629] 1. Experimental Principle

[0630] Solar ultraviolet (UVB) radiation can cause photodamage to the skin, thereby causing epidermal HACAT cells to release inflammatory factors and secrete a series of pro-inflammatory cytokines (such as tumor necrosis factor-α (TNF-α), interleukin-1α (IL-1α)). This project uses a UVB-induced HACAT cell inflammatory response model, and uses fluorescence quantitative (RT-qPCR) to detect the relative expression of TNF-α and IL-1α mRNA as the detection endpoint to establish a method. By comparing the difference in the relative expression of the mRNA of the test factor after administration of the negative control and the test substance, the effect of the test substance on inhibiting inflammation is evaluated.

[0631] 2. Test materials and test samples

[0632] 2.1 Experimental Reagents

[0633] High-glucose DMEM medium, fetal bovine serum, penicillin-streptomycin mixture, trypan blue dye, PBS, MTT, dexamethasone, RNA extraction kit, reverse transcription kit, real-time quantitative PCR detection kit, cell line: HACAT.

[0634] 2.2 Experimental instruments

[0635] Carbon dioxide cell culture incubator, biological safety cabinet, inverted microscope, electric constant temperature water bath, microplate reader, real-time fluorescence quantitative PCR instrument, centrifuge, UV cross-linking instrument.

[0636] 2.3 Test sample: 2000ppm Drimendiol 1,3-butanediol solution.

[0637] 3. Experimental Procedure

[0638] 3.1 Cell activity test (MTT method)

[0639] 3.1.1 Routine cell culture: The cell suspension was inoculated into a 96-well cell culture plate and cultured for 24 hours.

[0640] 3.1.2 Take 300 μL of sample and dissolve it in 1200 μL of culture medium until fully dissolved.

[0641] 3.1.3 Add the same volume and different concentrations of the sample to be tested to each well and return to the incubator for incubation.

[0642] 3.1.4 Add MTT solution to each well and incubate in the incubator.

[0643] 3.1.5 Discard the liquid in the wells, add DMSO to each well, shake evenly, and measure the absorbance using an enzyme-labeled instrument.

[0644] 3.2 Determination of relative gene expression of TNF-α and IL-1α

[0645] 3.2.1 Perform the test according to the test scheme shown in Table 48.

[0646] 3.2.2 Routine cell culture: The cell suspension was inoculated into a 6-well cell culture plate and cultured for 24 hours.

[0647] Table 48 is a table of cell grouping and processing information for TNF-α and IL-1α content testing.

[0648] Table 48 TNF-α and IL-1α content test cell grouping processing information table

[0649]

[0650] 3.2.3 Aspirate the culture medium from the wells and treat each well with different treatment groups according to the above table. Return the 6-well plate to the cell culture incubator and culture for 24 hours. Collect the cells into a centrifuge tube.

[0651] 3.2.4 RNA extraction: sample lysis—removal of genomic DNA—adsorption of RNA—removal of impurities—elution of RNA—RNA purity and concentration detection.

[0652] 3.2.5 RNA reverse transcription: remove genomic DNA - reverse transcription - obtain cDNA.

[0653] 3.2.6 Prepare primers, probes, and multiplex PCR reaction system and distribute them to each reaction tube, then add cDNA respectively.

[0654] 3.2.7 Open the PCR reaction program, set the appropriate experiment type and mode to start running, and obtain experimental data.

[0655] 4. Data Analysis

[0656] 4.1 Relative cell activity

[0657] Assuming that the cell activity of the blank control group was 100%, the relative activity of cells in each group was calculated.

[0658]

[0659] OD TA : The average value of the optical density of the tested sample group for 3 times OD C ;

[0660] The average value of the optical density of the tested sample group for 3 times

[0661] OD B : The average value of the optical density of the blank control group for 3 times

[0662] OD C : The average value of the optical density of the negative control group for 3 times

[0663] 4.2 Gene expression level to be measured

[0664] ΔCT(treatment group) = CT(test gene) - CT(mean value of internal reference gene)

[0665] ΔCT (control group) = CT (test gene) - CT (mean value of internal reference gene)

[0666] ΔΔCT = ΔCT (treated group) - ΔCT (control group)

[0667] Relative expression of the gene to be tested = 2^_△△ CT , take 2^_△△ CT The mean

[0668] 5. Experimental results

[0669] 5.1 Cell activity test results

[0670] Perform the MTT test according to step 3.1. Five sample concentrations were designed for this cell activity test (eg Fig.34 When the sample concentration is ≤0.1%, the cell activity is greater than 90% and can be used for subsequent experiments.

[0671] Table 49 shows the relative activity of cells in each group (Mean±SD).

[0672] Table 49 Relative activity of cells in each group (Mean±SD)

[0673] concentration(%) active(%) 0.01 105.99±5.78 0.1 107.63±5.37 1 68.58±3.80 5 19.51±0.57 10 18.52±0.62

[0674] Fig.34 In the experiment, sample concentrations of 0.025%, 0.05%, and 0.1% were selected as test concentrations for subsequent efficacy experiments.

[0675] 5.2 Relative expression of TNF-α gene

[0676] According to the sample concentration selected in 5.1, TNF-α content was detected. The test results are shown in Table 50 and Fig.35 shown.

[0677] Table 50 Relative expression of TNF-α in each group (Mean±SD)

[0678]

[0679]

[0680] Note: "ns" means no significant difference, P ≥ 0.05; * means the difference is statistically significant (0.01 ≤ P < 0.05) compared with the negative control group (C); ** means the difference is extremely significant (P < 0.01) compared with the negative control group (C)

[0681] 5.3 Relative expression of IL-1α gene

[0682] According to the sample concentration selected in 5.1, the IL-1α content was tested. The test results are shown in Table 51 and Fig.36 shown.

[0683] Table 51 Relative expression of IL-1α in each group (Mean±SD)

[0684]

[0685] Note: "ns" means no significant difference, P ≥ 0.05; * means the difference is statistically significant (0.01 ≤ P < 0.05) compared with the negative control group (C); ** means the difference is extremely significant (P < 0.01) compared with the negative control group (C)

[0686] Conclusion: The relative expression of inflammatory factor TNF-α gene in samples at concentrations of 0.025% (0.5ppm Drimendiol), 0.05% (1ppm Drimendiol), and 0.1% (2ppm Drimendiol) were 0.13±0.02, 0.23±0.01, and 0.15±0.04, respectively, which were significantly downregulated compared with the negative control (P<0.01). This indicates that the sample has the effect of inhibiting TNF-α gene at this concentration, which can be used as one of the evidences to support the claim of soothing effect.

[0687] At the concentrations of 0.025% (0.5ppm Drimendiol), 0.05% (1ppm Drimendiol), and 0.1% (2ppm Drimendiol), the relative expression levels of the inflammatory factor IL-1α gene were 0.24±0.04, 0.34±0.00, and 0.35±0.02, respectively, which were significantly downregulated compared with the negative control (P<0.01). This indicates that the sample has the effect of inhibiting the IL-1α gene at this concentration, which can be used as one of the evidences to support the claim of soothing effect.

[0688] Embodiment 14

[0689] Compound Drimendiol is used to prepare cosmetics that negatively regulate the expression of transient receptor potential vanilloid-1 receptor and prostaglandin E2 genes to achieve skin analgesia, anti-inflammatory and soothing effects

[0690] The experiment is as follows:

[0691] 1. Experimental Principle

[0692] Transient receptor potential vanilloid-1 (TRPV1) receptor is a non-selective cation channel with high calcium permeability. Its expression is widely distributed in nociceptive neurons and various non-neuronal cells, including keratinocytes, fibroblasts, and endothelial cells. In keratinocytes, TRPV1 receptors can be activated by a variety of endogenous and exogenous factors, such as high temperature, low pH (protons), and capsaicin. When TRPV1 receptors are activated, Na + and Ca 2+ The permeability / influx of the skin is increased, which in turn activates related signal transduction pathways and produces regulatory effects. Given its important role in skin cells, the TRPV1 pathway has become an important target for the development of anti-inflammatory and analgesic drugs for the treatment of skin diseases.

[0693] This project uses a capsaicin-stimulated cell inflammatory response model and establishes a method using fluorescence quantitative RT-qPCR to detect the relative expression of TRPV1 mRNA as the detection endpoint. By comparing the difference in relative expression of factor mRNA after administration of the negative control and the test substance, the inhibitory effect of the test substance on inflammation is evaluated.

[0694] Prostaglandin E2 (PGE2) is an important mediator that can promote the migration and differentiation of fibroblasts by activating related signaling pathways, thereby accelerating the wound healing process. After the skin nerve fibers are triggered, the immune system receives skin sensitivity, thereby activating keratinocytes, mast cells, antigen presenting cells and T cells in the skin, leading to the release of PGE2.

[0695] 2. Test materials and test samples

[0696] 2.1 Experimental Reagents

[0697] High-glucose DMEM culture medium, fetal bovine serum, penicillin-streptomycin mixture, trypan blue dye, PBS, MTT, capsaicin (CAP), trans-4-tert-butylcyclohexanol (4TB), RNA extraction kit, reverse transcription kit, real-time quantitative PCR detection kit, cell line: HACAT.

[0698] 2.2 Experimental instruments

[0699] Carbon dioxide cell culture incubator, biological safety cabinet, inverted microscope, electric constant temperature water bath, microplate reader, real-time fluorescence quantitative PCR instrument, centrifuge, UV cross-linking instrument.

[0700] 2.3 Test sample: 2000ppm Drimendiol 1,3-butanediol solution.

[0701] 3. Experimental Procedure

[0702] 3.1 Cell viability test (MTT)

[0703] 3.1.1 Routine cell culture: The cell suspension was inoculated into a 96-well cell culture plate and cultured for 24 hours.

[0704] 3.1.2 Take 300 μL of sample and dissolve it in 1200 μL of culture medium until fully dissolved.

[0705] 3.1.3 Add the same volume and different concentrations of the sample to be tested to each well and return to the incubator for incubation.

[0706] 3.1.4 Add MTT solution to each well and incubate in the incubator.

[0707] 3.1.5 Discard the liquid in the wells, add DMSO to each well, shake evenly, and measure the absorbance using an enzyme-labeled instrument.

[0708] 3.2 Determination of relative gene expression of TRPV1

[0709] 3.2.1 Perform the test according to the test plan shown in Table 52.

[0710] 3.2.2 Routine cell culture: The cell suspension was inoculated into a 6-well cell culture plate and cultured for 24 hours.

[0711] Table 52 is a table of TRPV1 content test cell grouping processing information.

[0712] Table 52TRPV1 content test cell grouping processing information table

[0713]

[0714] 3.2.3 Aspirate the culture medium from the wells and add different treatment groups to each well according to the table above. Return the 6-well plate to the cell culture incubator and culture for 24 hours. Collect the cells into a centrifuge tube.

[0715] 3.2.4 RNA extraction: sample lysis - removal of genomic DNA - adsorption of RNA - removal of impurities - elution of RNA - RNA purity and concentration detection

[0716] 3.2.5. RNA reverse transcription: remove genomic DNA - reverse transcription - obtain cDNA

[0717] 3.2.6 Prepare primers, probes, and multiplex PCR reaction system and distribute them to each reaction tube, then add cDNA respectively.

[0718] 3.2.7 Open the PCR reaction program, set the appropriate experiment type and mode to start running, and obtain experimental data.

[0719] 4. Data Analysis

[0720] 4.1 Relative cell activity

[0721] Assuming that the cell activity of the blank control group was 100%, the relative activity of cells in each group was calculated.

[0722]

[0723] OD TA : The average value of the optical density of the tested sample group for 3 times OD C ;

[0724] The average value of the optical density of the tested sample group for 3 times

[0725] OD B : The average value of the optical density of the blank control group for 3 times

[0726] OD C : The average value of the optical density of the negative control group for 3 times

[0727] 4.2 Gene expression level to be measured

[0728] ΔCT(treatment group) = CT(test gene) - CT(mean value of internal reference gene)

[0729] ΔCT (control group) = CT (test gene) - CT (mean value of internal reference gene)

[0730] ΔΔCT = ΔCT (treated group) - ΔCT (control group)

[0731] Relative expression of the gene to be tested = 2^_△△ CT , take 2 ^ _△△ CT The mean

[0732] 5. Experimental results

[0733] 5.1 Carry out MTT experiment according to step 3.1. Five sample concentrations were designed for this cell activity test (such as Fig.37 When the sample concentration is ≤0.1%, the cell activity is greater than 90% and can be used for subsequent experiments.

[0734] Table 53 Relative activity of cells in each group (Mean±SD)

[0735] concentration(%) active(%) 0.01 105.99±5.78 0.1 107.63±5.37 1 68.58±3.80 5 19.51±0.57 10 18.52±0.62

[0736] Fig.37 In the experiment, sample concentrations of 0.025%, 0.05%, and 0.1% were selected as test concentrations for subsequent efficacy experiments.

[0737] 5.2 Relative expression of TRPV1 gene

[0738] According to the sample concentration selected in 5.1, the TRPV1 content was tested. The test results are shown in Table 54 and Fig.38 shown.

[0739] Table 54 Relative expression of TRPV1 in each group (Mean±SD)

[0740]

[0741] Note: "ns" means no significant difference, P ≥ 0.05; * means the difference is statistically significant (0.01 ≤ P < 0.05) compared with the negative control group (C); ** means the difference is extremely significant (P < 0.01) compared with the negative control group (C)

[0742] 5.3 Relative expression of PGE2 gene

[0743] According to the sample concentration selected in 5.1, the PGE2 content was tested. The test results are shown in Table 55 and Fig.39 shown.

[0744] Table 55 Relative expression of PGE2 in each group (Mean±SD)

[0745]

[0746] Note: "ns" means no significant difference, P ≥ 0.05; * means the difference is statistically significant (0.01 ≤ P < 0.05) compared with the negative control group (C); ** means the difference is extremely significant (P < 0.01) compared with the negative control group (C)

[0747] Conclusion: The relative expression of the inflammatory factor TRPV1 gene in the samples at the concentrations of 0.025% (0.5ppm Drimendiol), 0.05% (1ppm Drimendiol), and 0.1% (2ppm Drimendiol) were 0.56±0.01, 0.50±0.02, and 0.48±0.01, respectively, which were significantly downregulated compared with the negative control (P<0.01). This indicates that the samples have the effect of inhibiting the TRPV1 gene at this concentration, which can be used as one of the evidences to support the claim of soothing effect.

[0748] At the concentrations of 0.025% (0.5ppm Drimendiol), 0.05% (1ppm Drimendiol), and 0.1% (2ppm Drimendiol), the relative expression levels of the inflammatory factor PGE2 gene were 0.69±0.02, 0.58±0.01, and 0.66±0.01, respectively, which were significantly downregulated compared with the negative control (P<0.01). This indicates that the sample has the effect of inhibiting the PGE2 gene at this concentration, which can be used as one of the evidences to support the claim of soothing effect.

[0749] Embodiment 15

[0750] Use of compound Drimendiol for preparing cosmetics having the effect of increasing the synthesis of hyaluronic acid in skin cells

[0751] The experiment is as follows:

[0752] 1. Experimental Principle

[0753] Hyaluronic acid, also known as hyaluronic acid, is a disaccharide unit glycosaminoglycan composed of D-glucuronic acid and N-acetylglucosamine. It is synthesized intracellularly by human immortalized epidermal cells (HaCaT) and secreted extracellularly. Hyaluronic acid can increase the gap between skin cells, enhance the barrier function of the skin and reduce water loss. It can promote the communication between skin cells and outside cells, improve skin metabolism, and also increase the skin's ability to absorb water and nutrients. Human immortalized epidermal cells (HaCaT) can be used as a cell model to study the enhancement of hyaluronic acid content in cosmetics. By measuring the increase in hyaluronic acid content after administration of blank control, positive control and test substance, the effectiveness of the test substance in promoting hyaluronic acid synthesis can be evaluated. The hyaluronic acid content was determined by enzyme-linked immunosorbent assay (ELISA). The specific principle is as follows: the capture antibody is coated on the ELISA plate to capture the hyaluronic acid in the sample and the standard. The biotinylated detection antibody is combined with the hyaluronic acid, and the SABC complex is combined with the biotinylated detection antibody to form an immune complex. After adding TMB colorimetric solution, if there is hyaluronic acid in the reaction well, it will appear blue, and it will turn yellow after adding the stop solution. During the detection process, the unbound components are washed away, and the OD value is measured at 450nm with an ELISA instrument. The hyaluronic acid concentration is proportional to the OD value. The concentration of hyaluronic acid in the specimen is calculated by drawing a standard curve, so as to perform qualitative or relative quantitative analysis on the test sample.

[0754] 2. Test materials and test samples

[0755] 2.1 Experimental Reagents

[0756] DMEM cell culture medium, trypsin / EDTA solution, fetal bovine serum, penicillin-streptomycin mixture, PBS, trypan blue dye, thiazolyl blue MTT, Human HA enzyme-linked immunosorbent assay kit, cell line: human immortalized epidermal cells (HaCaT).

[0757] 2.2 Experimental instruments

[0758] Cell culture incubator, biological safety cabinet, inverted microscope, electric thermostatic water bath, analytical balance, and microplate reader.

[0759] 2.3 Test sample: 2000ppm Drimendiol 1,3-butanediol solution.

[0760] 3. Experimental Procedure

[0761] 3.1 Cell viability test (MTT)

[0762] 3.1.1 Routine cell culture: The cell suspension was inoculated into a 96-well cell culture plate and cultured for 24 hours.

[0763] 3.1.2 Weigh 300 μL of sample and dissolve it in 1200 μL of culture medium until fully dissolved.

[0764] 3.1.3 Add the same volume and different concentrations of the sample to be tested to each well and return to the incubator for incubation for 24 hours.

[0765] 3.1.4 Add MTT solution to each well and incubate in the incubator for 4 hours.

[0766] 3.1.5 Discard the liquid in the wells, add DMSO to each well, shake evenly, and measure the absorbance using an enzyme-labeled instrument.

[0767] 3.2 HA content determination

[0768] 3.2.1 Perform the test according to the test scheme shown in Table 56.

[0769] Table 56 Group processing information

[0770]

[0771] 3.2.2 Routine cell culture: The cell suspension was inoculated into a 6-well cell culture plate and cultured for 24 hours.

[0772] 3.2.3 Aspirate the culture medium from the wells and add different control groups to each well. Return the 6-well plate to the cell culture incubator and culture for 24 hours.

[0773] 3.2.4 Detect the HA content of the samples according to the operating instructions of the ELISA kit.

[0774] 4. Data Analysis

[0775] 4.1 Relative cell activity

[0776] Assuming that the cell activity of the blank control group was 100%, the relative activity of cells in each group was calculated.

[0777]

[0778] OD TA : The average value of the optical density of the tested sample group for 3 times

[0779] OD P : The average value of the optical density of the positive control group for 3 times

[0780] OD B : The average value of the optical density of the blank control group for 3 times

[0781] 5. Experimental results

[0782] 5.1 Carry out MTT experiment according to step 3.1. Five sample concentrations were designed for this cell activity test (such as Fig.40 When the sample concentration is ≤0.1%, the cell activity is greater than 90%, and all of them can be used for subsequent experiments. 0.025%, 0.05%, and 0.1% were selected as the test concentrations for subsequent efficacy experiments.

[0783] Table 57 shows the relative activity of cells in each group (Mean±SD).

[0784] Table 57 Relative activity of cells in each group (Mean±SD)

[0785] concentration(%) active(%) 0.01 105.99±5.78 0.1 107.63±5.37 1 68.58±3.80 5 19.51±0.57 10 18.52±0.62

[0786] Fig.40 In the experiment, sample concentrations of 0.025%, 0.05%, and 0.1% were selected as test concentrations for subsequent efficacy experiments.

[0787] 5.2 Calculation results of HA content

[0788] According to the sample concentration selected in 5.1, HA content was tested. The test results are shown in Table 58 and Fig.41 shown.

[0789] Table 58 HA concentration in each group (Mean±SD)

[0790] Culture conditions HA concentration (ng / mL) P-value B Culture medium 1.6826±0.0353 / P Positive control (50 μM EGCG) + culture medium 2.0191±0.0221 0.0019** TA-1 0.025% sample + culture medium 1.7407±0.0276 0.2090 TA-2 0.05% sample + culture medium 1.8716±0.1270 0.1460 TA-3 0.1% sample + culture medium 1.8883±0.0965 0.0430*

[0791] Note: "ns" means no significant difference, P ≥ 0.05; * means the difference is statistically significant (0.01 ≤ P < 0.05) compared with the blank control group (B); ** means the difference is extremely significant (P < 0.01) compared with the blank control group (B)

[0792] Conclusion: At a concentration of 0.1% (2ppm Drimendiol), the HA concentration of the sample was 1.8883±0.0965ng / mL, which was significantly increased compared with the blank control (P<0.05), indicating that the test substance has the ability to promote the synthesis of hyaluronic acid at this test concentration, which can be used as evidence to support the claim of moisturizing effect.

[0793] The chemical structure of Epidrimendiol is:

[0794]

[0795] Preparation of Epidrimendiol:

[0796] 10 kg of pepperberry (Tasmannia lanceolata) leaf powder was extracted with 60 kg of ethanol: water = 1:1 at 60°C, heated and stirred for 3 hours. After filtration, the pepperberry leaf residue was extracted with 30.0 kg of ethanol: water = 1:1 at 60°C for 2 hours. After filtration, the two filtrates were combined and concentrated under reduced pressure to obtain 650 g of waxy extract.

[0797] The waxy extract was dissolved by heating with ethanol: water = 1:1, and extracted with n-hexane to obtain a component containing polygoni hydricarbaldehyde. After separation by silica gel column chromatography, gradient elution with petroleum ether and ethyl acetate was performed, and 60 g of polygoni hydricarbaldehyde compound was obtained by separation and purification.

[0798] The above 60g of Polygonum hydrophilum dialdehyde was dissolved in ethyl acetate, 12g of potassium carbonate was added, and the reaction was stirred at room temperature (25-35°C) for 6h. About 50% of Polygonum hydrophilum dialdehyde was converted into Polygonum hydrophilum isodialdehyde by thin layer chromatography monitoring. After filtration and concentration, 60g of isomeric mixture was obtained. Polygonum hydrophilum dialdehyde and Polygonum hydrophilum isodialdehyde were separated by silica gel column chromatography and gradient elution with petroleum ether and ethyl acetate to obtain 25g of Polygonum hydrophilum isodialdehyde compound.

[0799]

[0800] 25g of Polygonum hydropiper isodialdehyde was dissolved in 1000mL of ethyl acetate, and 20g of NaBH4 was added in batches. The reaction was stirred at room temperature (25-35°C) for 6h. The reaction was monitored by thin layer chromatography. 1.0kg of pure water and 0.9kg of 10% hydrochloric acid were added dropwise, and the pH was adjusted to 2-3. The aqueous phase and the ethyl acetate phase were separated. 0.5kg of ethyl acetate was added to the aqueous phase for extraction 3 times. The organic phases were combined and washed with pure water for 3 times. The organic phases were concentrated to obtain 23g of crude Epidrimendiol. The compound was separated by silica gel column chromatography, and the gradient elution of petroleum ether and ethyl acetate was obtained to obtain 20g of pure Epidrimendiol.

[0801]

[0802] Epidrimendiol NMR data:

[0803] δ 13 C-NMR: 36.6(C-1); 18.9(C-2); 42.7(C-3); 33.2(C-4); 54.2(C-5); 24.3(C-6); 127.7(C-7); 137.2 (C-8); 43.4 (C-9); 36.1 (C-10); 67.8 (C-11); 63.2 (C-12); 22.1 (C-13); 21.8 (C-14); 33.1 (C-15).

[0804] δ 1 H-NMR: 5.83 (H, m); 4.11~4.14 (H, d); 3.99~4.01 (H, d); 3.89~3.92 (H, dd); 3.63~3 .66(H,dd);2.23(2H,br,-OH);2.11-2.16(H,m);1.87~1.93(H,m);1.74~1.76(H, m); 1.60~1.66(H,m); 1.57~1.60(H,m); 1.42-1.52(H,m); 1.42~1.51(H,m); 1.48( H,m); 1.31~1.37(H,m); 1.16~1.24(H,m); 0.90(3H,s); 0.89(3H,s); 0.86(3H,s).

[0805] Prepare a 20 ppm Epidrimendiol solution as a sample: first dissolve 0.2 g Epidrimendiol in 99.8 g polyol (such as 1,3-butanediol or propylene glycol), stir and heat to dissolve to prepare a 2000 ppm Epidrimendiol solution, and then dilute the 2000 ppm Epidrimendiol solution 100 times with pure water to 20 ppm.

[0806] The prepared 20 ppm Epidrimendiol was used as a sample to test various human efficacy cosmetic uses (Examples 16 to 26);

[0807] The prepared 2000 ppm Epidrimendiol was used as a sample to test various gene expression efficacy cosmetic uses (Examples 27 to 30);

[0808] Testing period: November 6, 2024 - December 3, 2024;

[0809] Test environment: Temperature: 21±1℃; Humidity: 50±10%RH;

[0810] 32 subjects with sensitive skin were selected and the Bergman skin classification questionnaire was conducted on the 32 subjects. The survey results are shown in Table 59:

[0811] Table 59

[0812]

[0813]

[0814] Inclusion Criteria

[0815] (1) Healthy Chinese women aged 35-60 years old (at the start of the trial);

[0816] (2) sensitive skin according to the Bergman Questionnaire (Bergman Questionnaire sensitivity score range 30-68);

[0817] (3) VAS score of fine lines under the eyes ≥2 points;

[0818] (4) Self-perceived facial sagging;

[0819] (5) The basic value of the moisture meter in the forearm test area is between 15 and 45 (au);

[0820] (6) Be able to use samples well, understand and fill out the project questionnaire and cooperate with the staff;

[0821] (7) Be able to cooperate well with the experimenters and maintain a regular life during the study;

[0822] (8) Be able to read and understand all the contents of the informed consent form and voluntarily sign the informed consent form;

[0823] (9) During the trial, the participants agree not to use any cosmetics, drugs or health products that may affect the results.

[0824] Exclusion criteria

[0825] (1) Those who have allergic constitution, allergic dermatitis, skin diseases or a history of diseases (severe freckles, atopic dermatosis, psoriasis, eczema, severe acne, etc.), and are considered by clinical assessment to be unsuitable for participating in the program;

[0826] (2) The test site has large birthmarks, scratches, white spots, moles, keloids, or other skin manifestations that may affect the test;

[0827] (3) pregnancy, lactation, and menopause;

[0828] (4) Other requirements that affect this test;

[0829] (5) Participated in other clinical trials within the past 2 months;

[0830] (6) In addition, the trial leader determines that the population is not suitable as the subjects of this trial.

[0831] Restrictions

[0832] (1) During the test, treatment that may affect the test area is prohibited;

[0833] (2) During the test, it is prohibited to use other skin care samples or other samples that may affect the test;

[0834] (3) During the test, daily necessities must be the same as those used before the test began and replacement is prohibited.

[0835] Test method based on:

[0836] (1) QT / ZHCA 005-2019 Test method for the effect of cosmetics on skin elasticity

[0837] (2) QB / T 4256-2011 Guidelines for the evaluation of moisturizing efficacy of cosmetics

[0838] (3) T / GDCA 009-2022 Evaluation Methods of Cosmetic Repair Efficacy on Human Body

[0839] (4) T / ZHCA 003-2018 Test method for the effect of cosmetics on transepidermal water loss

[0840] (5) T / CNMIA 0015-2020 Clinical Evaluation Standard for Allergy-Relief Functional Skin Care Products

[0841] (6) T / GDCDC 021-2022 Test method for soothing effect of cosmetics

[0842] (7) Wang Huan, Pan Yao. Evaluation of cosmetic efficacy (V) - Scientific support for soothing efficacy claims[J]. Surfactant Detergent Industry, 2018, 48(05): 247-254

[0843] (8) T / ZHCA 005-2019 Test method for the effect of cosmetics on skin elasticity

[0844] (9) T / GDCDC 032-2023 Test method for cosmetic exfoliation efficacy

[0845] (10) T / ZHCA 006-2019 Test method for anti-wrinkle efficacy of cosmetics

[0846] (11) T / TDCA 003-2021 Test method for firming effect of cosmetics

[0847] (12) T / CAB 0152-2022 Test methods for seven cosmetic effects: anti-wrinkle, firming, moisturizing, oil control, repairing, nourishing, and soothing

[0848] (13) Liu Weiyi, Zhou Lin, Zhao Hua, Evaluation of cosmetic efficacy (ⅩⅢ) - Consumer use test[J]. Surfactant Chemical Industry, 2021, 51(06): 485-490

[0849] (14) "Technical Specifications for Safety of Cosmetics" (2015 edition)

[0850] (15) Cosmetics classification rules and classification catalogue

[0851] (16) Evaluation Standards for Cosmetic Efficacy Claims

[0852] Testing process:

[0853] D0(BL):

[0854] (1) Subjects were recruited as required, visited the laboratory, and laboratory staff explained the test process to the subjects, who then signed a written informed consent form.

[0855] (2) Screen the subjects according to the test requirements.

[0856] (3) The selected subjects cleaned their faces with a standard facial cleanser, dried their faces with a dandruff-free absorbent paper towel, and sat in a constant temperature and humidity chamber with their arms exposed for 30 minutes.

[0857] (4) The test personnel drew a 3cm×3cm square test area on the inner side of both forearms of the subjects who passed the screening. Then they started the test in the following order: VISIA-CR (three sides) → Evaface (three sides) → VC 98 (cheek) → Corneometer (cheek) → Corneometer (left and right arms) → Glossymeter (face) → Tewameter (face) → DermaScan (face).

[0858] (5) The staff will perform a single sample test on the inner side of the subject's forearm.

[0859] (6) After the subjects complete the test, the staff distributes the test products and provides instructions on how to use the products and the dosage to use.

[0860] T72h:

[0861] (7) The subjects returned for a follow-up visit at the prescribed time, exposed the inner forearm test site and sat quietly in a constant temperature and humidity room for 30 minutes.

[0862] (8) Then proceed to the test. The test sequence is as follows: Corneometer (left and right arms) → self-assessment questionnaire.

[0863] D7, D14, D21:

[0864] (9) The subjects filled out the online questionnaire within the corresponding time.

[0865] D28:

[0866] (10) The subjects returned for a follow-up visit. After cleaning their faces with a standard facial cleanser, they sat quietly in a constant temperature and humidity room for 30 minutes.

[0867] (11) Check the subjects’ usage records and weigh the products.

[0868] (12) Then the test was started. The test order was as follows: VISIA-CR (three faces) → Evaface (three faces) → VC 98 (cheeks) → Corneometer (face) → Glossymeter (face) → Tewameter (face) → DermaScan (face) → self-assessment questionnaire (Bauman questionnaire, self-assessment questionnaire).

[0869] (13) The testing process ends.

[0870] Note: D0 refers to the visit before the subject uses the sample, T72h refers to the visit 72 hours after the subject uses the sample, D7 refers to the visit 7 days after the subject uses the sample, D14 refers to the visit 14 days after the subject uses the sample, D21 refers to the visit 21 days after the subject uses the sample, and D28 refers to the visit 28 days after the subject uses the sample.

[0871] In the following examples, EXCEL software was used to perform descriptive statistics on the measured values ​​at each test time point, including the mean and standard error.

[0872] Measurement data: Before and after comparison, SPSS software was used to perform a significance test of the normal distribution of data change values ​​using the Shapiro-Wilk Test. If the data were normally distributed, a paired t test was used, otherwise a two-related sample Wilcoxon rank sum test was used. The significance level α was 0.05.

[0873] Rank data: The comparison before and after use was performed using two related sample Wilcoxon rank sum test, with the significant difference level α set at 0.05.

[0874] Questionnaire survey data: The differences in the results of any scoring criteria of the self-assessment questionnaire are significant, indicating that the samples have corresponding efficacy, and the significance difference level α is 0.05.

[0875] Change rate % = (average value of the test after sample use - average value of the test before sample use) / average value of the test before sample use * 100%

[0876] Note: In the test results of the following Examples 16 to 26, the significance marks are explained as follows: p≥0.05, "ns" means no statistical difference; p<0.05 means significant difference; among them, "*" means 0.01≤p<0.05; "**" means 0.001≤p<0.01; "***" means p<0.001. "*" means the difference before and after use.

[0877] Example 16

[0878] Compound Epidrimendiol is used for preparing cosmetics with moisturizing effect

[0879] The subject's face was tested using a Corneometer;

[0880] Parameter explanation: The larger the parameter value, the higher the moisture content of the skin stratum corneum.

[0881] Result determination: After using the product, the moisture content of the stratum corneum in the test area increased significantly, indicating that the test product has a moisturizing effect.

[0882] After 4 weeks of use, the moisture content of the stratum corneum increased significantly by 69.41% compared to before use.

[0883] The specific test results are shown in Table 60 and Fig.42 ;

[0884] Table 60 Test results of moisture content in stratum corneum (32 people) Unit: CU

[0885]

[0886]

[0887] Embodiment 17

[0888] Use of compound Epidrimendiol for preparing cosmetics with 72-hour moisturizing effect

[0889] The subjects’ arms on both sides were tested using a Corneometer;

[0890] Parameter explanation: The larger the parameter value, the higher the moisture content of the stratum corneum.

[0891] Result determination: After a single use of the product, the increase in moisture content in the stratum corneum in the test area was significantly higher than that in the control group, indicating that the test sample has a short-term moisturizing effect of 72 hours.

[0892] At 72 hours after use, the change in stratum corneum moisture content on the sample side was significantly increased by 12.05% compared with the control side.

[0893] Specific test results are shown in Table 61, Table 62 and Fig.43 ;

[0894] Table 61 Test results of moisture content of stratum corneum (32 people) Unit: CU

[0895]

[0896] Table 62 Test results of moisture content change in stratum corneum (32 people) Unit: CU

[0897]

[0898] Embodiment 18

[0899] Compound Epidrimendiol is used for preparing cosmetics with repairing effect

[0900] The Tewameter was used to test one side of the subject's face for 40 seconds, and the average value was taken for the last 20 seconds. The test was repeated twice and the average value was taken.

[0901] Parameter explanation: The smaller the parameter value, the lower the transepidermal water loss rate of the skin and the better the skin barrier.

[0902] Result determination: After using the product, the parameter values ​​of the test area decreased significantly, indicating that the test product has a repairing effect.

[0903] After 4 weeks of use, the transepidermal water loss rate was significantly reduced by 22.28% compared to before use.

[0904] The specific test results are shown in Table 63 and Fig.44 ;

[0905] Table 63 Skin transepidermal water loss rate test results (32 people) Unit: g / h / m 2

[0906]

[0907] Embodiment 19

[0908] Use of compound Epidrimendiol for preparing cosmetics with nourishing effect

[0909] Test judgment basis: After using the product, at least two of the test area's gloss, smoothness, firmness / elasticity, and dermis thickness / density parameters have significant improvements, indicating that the test product has a nourishing effect.

[0910] Test 1: Smoothness

[0911] The subjects’ cheeks were tested using VC20 and the SEsm values ​​were analyzed;

[0912] Parameter explanation: The smaller the SEsm parameter value, the smoother the skin.

[0913] Result determination: After using the product, the SEsm parameter value in the test area decreased significantly, indicating that the test product has the effect of smoothing the skin.

[0914] After 4 weeks of use, the SEsm value was significantly reduced by 12.13% compared with before use.

[0915] The specific test results are shown in Table 64. Fig.45 and Fig.62 ;

[0916] Table 64 Skin SEsm value test results (32 people) Unit: au

[0917]

[0918] Test 2: Glossiness

[0919] The subjects' cheeks were tested using a Glossymeter.

[0920] Parameter explanation: The larger the test value, the better the skin glossiness.

[0921] Result determination: After using the product, the parameter values ​​in the test area increased significantly, indicating that the test product has the effect of improving skin gloss.

[0922] After 4 weeks of use, the glossiness was significantly increased by 63.62% compared to before use.

[0923] The specific test results are shown in Table 65 and Fig.46 ;

[0924] Table 65 Skin gloss test results (32 people) Unit: au

[0925]

[0926] Test 3: Firmness and elasticity

[0927] The subject's cheek was tested using a Cutometer.

[0928] Parameter explanation: The larger the R2 parameter value, the better the skin elasticity; the smaller the R0 parameter value, the better the skin firmness.

[0929] Result judgment: After using the product, the R2 parameter value of the test area increased significantly, indicating that the test sample has the effect of improving skin elasticity; after using the product, the R0 parameter value of the test area decreased significantly, indicating that the test sample has the effect of improving skin firmness.

[0930] After 4 weeks of use, the R0 value decreased significantly by 42.46% and the R2 value increased significantly by 19.44% compared with before use.

[0931] The specific test results are shown in Table 66. Fig.47 and Fig.48 ;

[0932] Table 66 Firmness and elasticity test results (32 people)

[0933]

[0934] Test 4: Leather Thickness / Density

[0935] The subjects' faces were tested using DermaScan.

[0936] Parameter explanation: The larger the parameter value, the thicker the dermis is and the higher the density is.

[0937] Result determination: After using the product, the parameter values ​​in the test area increased significantly, indicating that the tested product has the effect of increasing the thickness and density of the dermis.

[0938] After 4 weeks of use, compared with before use, the dermis thickness increased significantly by 11.41% and the dermis density increased significantly by 60.22%.

[0939] The specific test results are shown in Table 67. Fig.49 , Fig.50 and Fig.64 ;

[0940] Table 67 Increased skin thickness and density test results (32 people)

[0941]

[0942] Embodiment 20

[0943] Use of compound Epidrimendiol for preparing cosmetics with soothing effect

[0944] Test 1: Use VISIA-CR to take pictures and use IPP software to analyze the skin redness a* parameter value.

[0945] Parameter explanation: The larger the parameter value, the redder the skin.

[0946] Result determination: After using the product, the parameter values ​​in the test area decreased significantly, indicating that the test product has a soothing effect.

[0947] After 4 weeks of use, the a* value decreased significantly by 21.37% compared to before use.

[0948] The specific test results are shown in Table 68. Fig.51 and Fig.65 ;

[0949] Table 68 Skin a* value test results (32 people) Unit: au

[0950]

[0951] Test 2: Photograph the subject's cheek using VISIA-CR and analyze the L* value using IPP.

[0952] Parameter explanation: The larger the L* value, the brighter the skin.

[0953] Result determination: After using the product, the L* value of the test area increased significantly, indicating that the test product has the effect of brightening the skin tone.

[0954] After 4 weeks of use, the L* value increased significantly by 14.35% compared to before use.

[0955] The specific test results are shown in Table 69 and Fig.52 ;

[0956] Table 69 Skin L* value test results (32 people) Unit: au

[0957]

[0958] Embodiment 21

[0959] Use of compound Epidrimendiol for preparing cosmetics with firming effect

[0960] Refer to Table 66 in Example 19, Fig.47 and Fig.48 .

[0961] Embodiment 22

[0962] Use of compound Epidrimendiol for preparing cosmetics with anti-wrinkle effect

[0963] The subjects' faces were photographed using VISIA-CR and the IPP was used to analyze the percentage of wrinkle area under the eyes. Parameter explanation: The smaller the wrinkle area percentage parameter value, the better the anti-wrinkle effect.

[0964] Result determination: After using the product, the parameter values ​​in the test area decreased significantly, indicating that the test product has an anti-wrinkle effect.

[0965] After 4 weeks of use, the area of ​​under-eye wrinkles was significantly reduced by 13.29% compared to before use.

[0966] The specific test results are shown in Table 70. Fig.53 and Fig.60 ;

[0967] Table 70 Test results of area ratio of under-eye wrinkles (32 people) Unit: %

[0968]

[0969] Embodiment 23

[0970] Compound Epidrimendiol is used for preparing cosmetics with the effects of increasing the distance of the apple muscles upward and the angle of the mandibular angle

[0971] Test 1: Improve sagging of apple muscles

[0972] EvaFACE was used to photograph the lower edge of the subjects' apple muscles and analyze the sagging volume of the apple muscles.

[0973] Parameter explanation: The larger the volume of the sagging apple muscles, the more serious the sagging apple muscles are.

[0974] Result determination: After using the product, the volume parameter value of the sagging apple muscles decreased significantly, indicating that the test product has the effect of improving the sagging apple muscles.

[0975] After 4 weeks of use, the volume of sagging apple muscles was significantly reduced by 21.51% compared to before use.

[0976] See Table 71 for specific test results. Fig.54 and Fig.58 ;

[0977] Table 71 Apple muscle sagging volume test results (32 people) Unit: mm 3

[0978]

[0979]

[0980] Test 2: Jawline Lift

[0981] EvaFACE was used to photograph the lower jaw of the subjects and the jawline ptosis was analyzed regardless of the length of the contour line.

[0982] Parameter explanation: The shorter the length of the contour line of the sagging mandibular margin, the more obvious the effect of lifting the mandibular margin is.

[0983] Result judgment: After using the product, the parameter value of the contour line length of the sagging mandibular margin decreased significantly, indicating that the test product has the effect of lifting the mandibular line.

[0984] After 4 weeks of use, the length of the contour line of the mandibular sagging area was significantly reduced by 2.36% compared to before use.

[0985] See Table 72 for specific test results. Fig.55 and Fig.59 ;

[0986] Table 72 Test results of the contour length of the mandibular drooping area (32 people) Unit: mm

[0987]

[0988] Embodiment 24

[0989] Use of the compound Epidrimendiol for preparing cosmetics suitable for sensitive skin

[0990] Improve sensitivity: The subjects were tested using the Bauman sensitivity questionnaire. Parameter explanation: The larger the parameter value, the more sensitive the skin.

[0991] Result judgment: After using the product, the Bauman questionnaire score decreased significantly, indicating that the test product has the effect of improving skin sensitivity.

[0992] After 4 weeks of use, the Bowman Questionnaire score was significantly reduced by 34.60% compared to before use.

[0993] The specific test results are shown in Table 73 and Fig.56 ;

[0994] Table 73 Bauman Questionnaire score test results (32 people) Unit: points

[0995]

[0996] Embodiment 25

[0997] Use of compound Epidrimendiol for preparing cosmetics with exfoliating effect

[0998] The subjects' faces were tested using VC98 and the desquamation index (DI) values ​​were analyzed.

[0999] Parameter explanation: The smaller the desquamation index DI parameter value, the less stratum corneum desquamation.

[1000] Result determination: After using the product, the desquamation index DI parameter value of the test area decreased significantly, indicating that the test product has the effect of exfoliating.

[1001] After 4 weeks of use, the skin desquamation index DI value was significantly reduced by 9.23% compared with before use.

[1002] The specific test results are shown in Table 74. Fig.57 and Fig.62 ;

[1003] Table 74 Skin desquamation index DI value test results (32 people) Unit: au

[1004]

[1005] Embodiment 26

[1006] Compound Epidrimendiol is used for preparing cosmetics without irritation reaction

[1007] The subjects were evaluated for adverse skin reactions.

[1008] Parameter explanation: After the subjects who were screened as sensitive skin by the questionnaire used the samples for 7 days, the skin was evaluated for adverse reactions. The "Human Trial Skin Reaction Grading Standard" in the "Cosmetic Safety Technical Specifications 2015 Edition" was used, refer to Table 75:

[1009] Table 75 Human trial skin reaction classification standards

[1010]

[1011] Result determination: After using the product, the subjects reported no adverse reactions on their faces, indicating that the test product has a mild (non-irritating) effect.

[1012] The number of adverse facial reactions in the subjects was counted.

[1013] Result judgment: After using the product, 96.67% and above of the subjects self-assessed that they had no discomfort, indicating that the tested product is suitable for sensitive skin. It can be used to prepare non-irritating and sensitive skin-suitable cosmetics.

[1014] The specific test results are shown in Table 76;

[1015] Table 76 Adverse reaction evaluation results (32 people)

[1016]

[1017] Subjects completed a self-assessment questionnaire.

[1018] Parameter explanation: The number of people with a statistical score of >3 points is the number of people with beneficial items. Recognition degree = number of people with beneficial items / total number of people*100% Result judgment: The difference in the results of any scoring standard of the self-assessment questionnaire is significant, indicating that the sample has corresponding efficacy.

[1019] The questionnaire test results are shown in Table 77, Table 78, Table 79, Table 80 and Table 81:

[1020] Table 77 Results of the survey on the third day of use (5-point scale) (32 people)

[1021]

[1022] Table 78 Results of the first week of use approval survey (5-point scale) (32 people)

[1023]

[1024]

[1025] Table 79: Results of the second week approval survey (5-point scale) (32 people)

[1026]

[1027] Table 80 Results of the third week of use approval survey (5-point scale) (32 people)

[1028]

[1029]

[1030] Table 81 Results of the 4th week approval survey (5-point scale) (32 people)

[1031]

[1032] Based on Examples 16 to 26, combined with instrument testing, image analysis, visual evaluation and questionnaires, 20 ppm of Epidrimendiol was used as a sample to test the performance and use of cosmetics; the sample was found to be suitable for sensitive skin, mild (non-irritating), soothing, anti-wrinkle, 72-hour moisturizing, moisturizing, repairing, firming, exfoliating, and nourishing; it has the effects of improving skin sensitivity, brightening skin tone, enhancing gloss, smoothing skin, enhancing skin thickness and density, enhancing apple muscles, and tightening jawline.

[1033] Embodiment 27

[1034] Compound Epidrimendiol is used for preparing cosmetics with positive regulation of the expression of acid sphingomyelinase SMPD1 gene and loricrin LOR gene and the effect of improving skin barrier

[1035] 1. The experimental principle, test materials, experimental steps, and data analysis methods are the same as those in Example 11.

[1036] 2. Test sample: 2000ppm Epidimendiol 1,3-butanediol solution.

[1037] 3. Experimental results

[1038] 3.1 Cell activity test results

[1039] Two MTT experiments were performed: The first MTT experiment: 8 sample concentrations were designed for this cell activity test (such as Fig.65 When the sample concentration is ≤0.1%, the cell activity is greater than 90%, and all of them can be used for subsequent experiments. Table 82 shows the relative activity of cells in each group (Mean±SD).

[1040] Table 82 Relative activity of cells in each group (Mean±SD)

[1041] concentration(%) active(%) 10 27.90±0.70 5 27.34±1.90 1 84.39±4.31 0.1 106.82±5.96 0.05 105.58±3.58 0.01 106.81±7.03 0.005 106.04±5.75 0.001 99.11±7.58 C 100.00±7.79

[1042] In the first efficacy test experiment, a sample concentration of 0.1% was selected as the subsequent test concentration for SMPD1; 0.05% and 0.1% were selected as the subsequent test concentrations for LOR;

[1043] In the second MTT experiment, eight sample concentrations were designed for the cell activity test (e.g. Fig.66 When the sample concentration is ≤0.2%, the cell activity is greater than 90%, and all of them can be used for subsequent experiments. Table 83 shows the relative activity of cells in each group (Mean±SD).

[1044] Table 83 Relative activity of cells in each group (Mean±SD)

[1045]

[1046]

[1047] 0.2% was selected as the test concentration for the second efficacy experiment.

[1048] 3.2 Relative expression of SMPD1 gene

[1049] According to the sample concentration selected in 3.1, SMPD1 expression was detected. The test results of the first efficacy test are shown in Table 84 and Fig.67 shown.

[1050] Table 84 Relative expression of SMPD1 in each group (Mean±SD)

[1051]

[1052] Note: "ns" means no significant difference, P ≥ 0.05; * means the difference is statistically significant (0.01 ≤ P < 0.05) compared with the blank control group (B); ** means the difference is extremely significant (P < 0.01) compared with the blank control group (B)

[1053] The test results of the second efficacy test are shown in Tables 85 and Fig.68 shown.

[1054] Table 85 Relative expression of SMPD1 in each group (Mean±SD)

[1055]

[1056] Note: "ns" means no significant difference, P ≥ 0.05; * means the difference is statistically significant (0.01 ≤ P < 0.05) compared with the blank control group (B); ** means the difference is extremely significant (P < 0.01) compared with the blank control group (B)

[1057] According to the sample concentration selected in 3.1, LOR expression was detected. The results of the first efficacy test are shown in Table 86 and Fig.69 shown.

[1058] Table 86 Relative expression of LOR in each group (Mean±SD)

[1059]

[1060] Note: "ns" means no significant difference, P ≥ 0.05; * means the difference is statistically significant (0.01 ≤ P < 0.05) compared with the blank control group (B); ** means the difference is extremely significant (P < 0.01) compared with the blank control group (B)

[1061] The results of the second efficacy test are shown in Tables 87 and Fig.70 shown.

[1062] Table 87 Relative expression of LOR in each group (Mean±SD)

[1063]

[1064] Note: "ns" means no significant difference, P ≥ 0.05; * means the difference is statistically significant (0.01 ≤ P < 0.05) compared with the blank control group (B); ** means the difference is extremely significant (P < 0.01) compared with the blank control group (B)

[1065] Conclusion: At the concentration of 0.1% (2ppm Epidimendiol), the relative expression of SMPD1 gene was 1.52±0.09, which was significantly increased compared with the blank control (P<0.05); at the concentration of 0.2% (4ppm Epidimendiol), the relative expression of SMPD1 gene was 2.44±0.12, which was significantly increased compared with the blank control (P<0.01). This indicates that the samples have the effect of up-regulating SMPD1 gene at the above concentrations. At the concentrations of 0.05% (1ppm Epidimendiol) and 0.1% (2ppm Epidimendiol), the relative expression of LOR gene was 1.50±0.08 and 1.19±0.04, respectively, which were significantly increased compared with the blank control (P<0.05); at the concentration of 0.2%, the relative expression of LOR gene was 2.34±0.17, which was significantly increased compared with the blank control (P<0.01). This indicates that the sample has the effect of upregulating the LOR gene at the above concentration.

[1066] Embodiment 28

[1067] Compound Epidrimendiol is used for preparing cosmetics with the effect of positively regulating the expression of elastin ELN gene and improving skin elasticity and firmness

[1068] 1. The experimental principle, test materials, experimental steps, and data analysis methods are the same as those in Example 12.

[1069] 2. Test sample: 2000ppm Epidimendiol 1,3-butanediol solution.

[1070] 3. Experimental results

[1071] 3.1 Cell activity test results

[1072] The MTT assay was performed twice.

[1073] First MTT experiment: This cell activity test was designed with 8 sample concentrations (such as Fig.71 When the sample concentration is ≤1%, the cell activity is greater than 90% and can be used for subsequent experiments.

[1074] Table 88 shows the relative activity of cells in each group (Mean±SD).

[1075] Table 88 Relative activity of cells in each group (Mean±SD)

[1076] concentration(%) active(%) 5 56.06±4.73 1 90.54±1.58 0.2 93.26±2.71 0.1 98.36±2.81 0.01 101.52±2.43 0.005 100.18±1.30 0.001 99.46±2.58 0.0005 101.42±3.56 C 100.00±1.81

[1077] 0.2% was selected as the test concentration for the first efficacy experiment.

[1078] Second MTT experiment: This cell activity test was designed with 8 sample concentrations (such as Fig.72 When the sample concentration is ≤1%, the cell activity is greater than 90% and can be used for subsequent experiments.

[1079] Table 89 shows the relative activity of cells in each group (Mean±SD).

[1080] Table 89 Relative activity of cells in each group (Mean±SD)

[1081]

[1082]

[1083] 0.5% and 1% were selected as the test concentrations for the subsequent second efficacy experiment.

[1084] 3.2 Relative expression of ELN genes

[1085] According to the sample concentration selected in 3.1, the relative expression of ELN gene was detected. The results of the first efficacy test are shown in Table 90 and Fig.73 shown.

[1086] Table 90 Relative expression of ELN in each group (Mean±SD)

[1087]

[1088] Note: "ns" means no significant difference, P ≥ 0.05; * means the difference is statistically significant (0.01 ≤ P < 0.05) compared with the blank control group (B); ** means the difference is extremely significant (P < 0.01) compared with the blank control group (B)

[1089] The results of the second efficacy test are shown in Tables 91 and Fig.74 shown.

[1090] Table 91 Relative expression of ELN in each group (Mean±SD)

[1091]

[1092] Note: "ns" means no significant difference, P ≥ 0.05; * means the difference is statistically significant (0.01 ≤ P < 0.05) compared with the blank control group (B); ** means the difference is extremely significant (P < 0.01) compared with the blank control group (B)

[1093] in conclusion:

[1094] The relative expression of ELN gene in samples at concentrations of 0.2% (4ppm Epidimendiol), 0.5% (10ppm Epidimendiol), and 1% (20ppm Epidimendiol) were 8.73±1.02, 9.13±0.38, and 5.87±0.57, respectively, which were significantly upregulated compared with the blank control (P<0.01). This indicates that the samples have the effect of upregulating ELN gene at this concentration, which can be used as one of the evidences to support the claim of firming and anti-wrinkle effects.

[1095] Embodiment 29

[1096] Compound Epidrimendiol is used to prepare cosmetics that have the effects of negatively regulating the expression of tumor necrosis factor-α and interleukin-1α genes, improving skin inflammation, and soothing sensitive skin

[1097] 1. The experimental principle, test materials, experimental steps, and data analysis methods are the same as those in Example 13.

[1098] 2. Test sample: 2000ppm Epidimendiol 1,3-butanediol solution.

[1099] 3. Experimental results

[1100] 3.1 Cell activity test results

[1101] Perform MTT assay.

[1102] This cell activity test designed 8 sample concentrations (such as Fig.75 When the sample concentration is ≤0.1%, the cell activity is greater than 90%, and all of them can be used for subsequent experiments. Table 92 shows the relative activity of cells in each group (Mean±SD).

[1103] Table 92 Relative activity of cells in each group (Mean±SD)

[1104] concentration(%) active(%) 10 27.90±0.70 5 27.34±1.90 1 84.39±4.31 0.1 106.82±5.96 0.05 105.58±3.58 0.01 106.81±7.03 0.005 106.04±5.75 0.001 99.11±7.58 C 100％±2.41

[1105] 0.1% was selected as the test concentration for subsequent efficacy experiments.

[1106] 3.2 Relative expression of TNF-α gene

[1107] According to the sample concentration selected in 3.1, TNF-α content was detected. The test results are shown in Table 93 and Fig.76 shown.

[1108] Table 93 Relative expression of TNF-α in each group (Mean±SD)

[1109]

[1110] Note: "ns" means no significant difference, P ≥ 0.05; * means the difference is statistically significant (0.01 ≤ P < 0.05) compared with the negative control group (C); ** means the difference is extremely significant (P < 0.01) compared with the negative control group (C)

[1111] 3.3 Relative expression of IL-1α gene

[1112] According to the sample concentration selected in 3.1, the IL-1α content was tested. The test results are shown in Table 94 and Fig.77 shown.

[1113] Table 94 Relative expression of IL-1α in each group (Mean±SD)

[1114]

[1115] Note: "ns" means no significant difference, P ≥ 0.05; * means the difference is statistically significant (0.01 ≤ P < 0.05) compared with the negative control group (C); ** means the difference is extremely significant (P < 0.01) compared with the negative control group (C)

[1116] Conclusion: At a concentration of 0.1% (2ppm Epidimendiol), the relative expression of the inflammatory factor TNF-α gene was 0.62±0.08, which was significantly downregulated compared with the negative control (P<0.05). This indicates that the sample has the effect of inhibiting the TNF-α gene at this concentration, which can be used as one of the evidences to support the claim of soothing effect.

[1117] At a concentration of 0.1% (2ppm Epidimendiol), the relative expression of the inflammatory factor IL-1α gene was 0.52±0.04, which was significantly downregulated compared with the negative control (P<0.05). This indicates that the sample has the effect of inhibiting the IL-1α gene at this concentration, which can be used as one of the evidences to support the claim of soothing effect.

[1118] Embodiment 30

[1119] The compound Epidrimendiol is used to prepare cosmetics that negatively regulate the expression of transient receptor potential vanilloid-1 receptor and prostaglandin E2 genes to achieve skin analgesia, anti-inflammatory and soothing effects

[1120] 1. The experimental principle, test materials, experimental steps, and data analysis methods are the same as those in Example 14.

[1121] 2. Test sample: 2000ppm Epidimendiol 1,3-butanediol solution.

[1122] 3. Experimental results

[1123] 3.1 Cell activity test results

[1124] The MTT assay was performed twice.

[1125] First MTT experiment: This cell activity test was designed with 8 sample concentrations (such as Fig.78 When the sample concentration is ≤0.1%, the cell activity is greater than 90% and can be used for subsequent experiments.

[1126] Table 95 shows the relative activity of cells in each group (Mean±SD).

[1127] Table 95 Relative activity of cells in each group (Mean±SD)

[1128] concentration(%) active(%) 10 27.90±0.70 5 27.34±1.90 1 84.39±4.31 0.1 106.82±5.96 0.05 105.58±3.58 0.01 106.81±7.03 0.005 106.04±5.75 0.001 99.11±7.58 C 100.00±2.41

[1129] 0.05% and 0.1% were selected as the test concentrations for efficacy experiments.

[1130] Second MTT experiment: This cell activity test was designed with 8 sample concentrations (such as Fig.79 When the sample concentration is ≤0.2%, the cell activity is greater than 90% and can be used for subsequent experiments.

[1131] Table 96 shows the relative activity of cells in each group (Mean±SD).

[1132] Table 96 Relative activity of cells in each group (Mean±SD)

[1133]

[1134]

[1135] 3.2 Relative expression of TRPV1 gene

[1136] According to the sample concentration selected in 3.1, the TRPV1 content was tested. The efficacy test results are shown in Table 97 and Fig.80 shown.

[1137] Table 97 Relative expression of TRPV1 in each group (Mean±SD)

[1138]

[1139] Note: "ns" means no significant difference, P ≥ 0.05; * means the difference is statistically significant (0.01 ≤ P < 0.05) compared with the negative control group (C); ** means the difference is extremely significant (P < 0.01) compared with the negative control group (C)

[1140] 3.3 Relative expression of PGE2 gene

[1141] According to the sample concentration selected in 3.1, PGE2 content detection was performed. The results of the first efficacy test are shown in Table 98 and Fig.81 shown.

[1142] Table 98 Relative expression of PGE2 in each group (Mean±SD)

[1143]

[1144]

[1145] Note: "ns" means no significant difference, P ≥ 0.05; * means the difference is statistically significant (0.01 ≤ P < 0.05) compared with the negative control group (C); ** means the difference is extremely significant (P < 0.01) compared with the negative control group (C)

[1146] The results of the second efficacy test are shown in Tables 99 and Fig.82 shown.

[1147] Table 99 Relative expression of PGE2 in each group (Mean±SD)

[1148]

[1149] Note: "ns" means no significant difference, P ≥ 0.05; * means the difference is statistically significant (0.01 ≤ P < 0.05) compared with the negative control group (C); ** means the difference is extremely significant (P < 0.01) compared with the negative control group (C)

[1150] Conclusion: The relative expression of the inflammatory factor TRPV1 gene in the samples at the concentrations of 0.05% (1ppm Epidimendiol) and 0.1% (2ppm Epidimendiol) were 0.84±0.08 and 0.88±0.02, respectively, which were significantly downregulated compared with the negative control (P<0.05). This indicates that the samples have the effect of inhibiting the TRPV1 gene at this concentration, which can be used as one of the evidences to support the claim of soothing effect.

[1151] The relative expression of the inflammatory factor PGE2 gene in the samples at the concentrations of 0.05% (1ppm Epidimendiol), 0.1% (2ppm Epidimendiol), and 0.2% (4ppm Epidimendiol) were 0.66±0.07, 0.62±0.06, and 0.55±0.07, respectively, which were significantly downregulated compared with the negative control (P<0.05). This indicates that the samples have the effect of inhibiting the PGE2 gene at this concentration, which can be used as one of the evidences to support the claim of soothing effect.

[1152] The above descriptions are merely embodiments of the present invention and are not intended to limit the patent scope of the present invention. Any equivalent transformations made using the contents of the present invention's specification and drawings, or directly or indirectly applied in related technical fields, are also included in the patent protection scope of the present invention.

Claims

1. Use of the compound Drimendiol or the compound Epidrimendiol for preparing cosmetics; The chemical structural formula of Drimendiol is: The chemical structural formula of the Epidrimendiol is:

2. The compound Drimendiol or the compound Epidrimendiol is used for preparing cosmetics with moisturizing, repairing, firming, nourishing, anti-wrinkle and / or soothing effects.

3. Use of the compound Drimendiol or the compound Epidrimendiol for preparing cosmetics with 72-hour moisturizing effect.

4. Use of the compound Drimendiol or the compound Epidrimendiol for preparing cosmetics suitable for sensitive skin.

5. Use of the compound Drimendiol or the compound Epidrimendiol for preparing cosmetics without irritation reaction.

6. Use of the compound Drimendiol or the compound Epidrimendiol for preparing cosmetics having the effect of increasing the upward distance of the apple cheeks and / or the angle of the mandibular angle.

7. The compound Epidrimendiol is used for preparing cosmetics with exfoliating effect.

8. The compound Drimendiol or the compound Epidrimendiol is used for preparing cosmetics that have the effect of positively regulating the expression of acid sphingomyelinase SMPD1 gene and loricrin LOR gene and improving the skin barrier.

9. Use of the compound Drimendiol or the compound Epidrimendiol in preparing cosmetics having the efficacy of positively regulating the expression of elastin ELN gene and improving skin elasticity and firmness.

10. The compound Drimendiol or the compound Epidrimendiol is used for preparing cosmetics that have the effects of negatively regulating the expression of tumor necrosis factor-α, interleukin-1α and prostaglandin E2 genes, improving skin inflammation and soothing sensitive skin.

11. The compound Drimendiol or the compound Epidrimendiol is used for preparing cosmetics that negatively regulate the expression of transient receptor potential vanilloid-1 receptor gene to achieve skin analgesic and anti-inflammatory soothing effects.

12. Use of the compound Drimendiol for preparing synthetic cosmetics having the function of increasing hyaluronic acid in skin cells.

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