A cicada fungus strain and its application as a cosmetic raw material

Abstract

The present invention discloses a cicada fungus strain and its use as a cosmetic raw material. The present invention provides a cicada fungus strain, specifically the cicada fungus (Cordyceps cicadae) ANTI-AGE1, which is registered with the General Microbiology Center of the China Culture Collection Administration under the registration number CGMCC No. 41179. The cicada fungus (Cordyceps cicadae) ANTI-AGE1 provided by the present invention is a new strain with great development prospects as a cosmetic raw material.

Description

Technical Field

[0001] The present invention relates to a edible and medicinal fungus in the field of microorganisms, and in particular to a Cicadae fungi strain and its application as a cosmetic raw material. Background Art

[0002] Isaria cicadae Miquel, commonly known as the giant caterpillar fungus, forms when cicada larvae, about to emerge from the soil, become infected and parasitized by the fungus. As a traditional Chinese herbal medicine, cicada fungus boasts numerous bioactive substances, including cicada polysaccharides, cordycepic acid, nucleosides, sterols, and peptides. It boasts nourishing and strengthening properties, including anti-tumor, immune-regulating, blood sugar-lowering, blood lipid-lowering, antipyretic, analgesic, and renal-protective properties, and possesses valuable medicinal and edible properties.

[0003] The cicada fungus (Cordyceps cicadae) is a filamentous fungus isolated from wild cicada flowers. By simulating the natural formation conditions of wild cicada flowers, artificial cultivation of the fungus has gradually become possible. Research has shown that the content of various bioactive substances in cultivated cicada flowers, such as amino acids, proteins, polysaccharides, nucleosides, cordycepic acid, sterols, and peptides, approaches that of wild cicada flowers.

[0004] In the context of large-scale picking of wild cicada flowers and the special growth environment and seasonal restrictions of wild cicada flowers, which have led to huge restrictions on wild cicada flower production, it is of great significance to realize artificial cultivation of cicada flowers and explore their application value. Summary of the Invention

[0005] The invention provides a Cordyceps sinensis strain and its application as a cosmetic raw material.

[0006] In a first aspect, the present invention claims protection for a cicada fungus strain.

[0007] The cicada fungus strain claimed in the present invention is Cordyceps cicadae ANTI-AGE1, and its registration number in the General Microbiology Center of China Culture Collection Administration of Microorganisms is CGMCC No.41179.

[0008] In the second aspect, the present invention claims protection for the fruiting bodies of the cicada fungus strain described above.

[0009] The fruiting body can be prepared according to a method comprising the following steps: inoculating the cicada fungus (Cordyceps cicadae) ANTI-AGE1 onto a PDA slant and preserving it at 4°C; inoculating the purified strain into a sterilized and cooled seed culture medium through aseptic operation, and obtaining a seed liquid after fermentation for 2-3 days; then transferring the seed liquid to a sterilized and cooled solid fermentation culture medium, culturing at a constant temperature of 26°C for 23 days with a humidity of 50-80% (e.g., 75%), to obtain a solid fermentation culture, i.e., the fruiting body.

[0010] The solid fermentation medium is wheat and nutrient solution at a ratio of 1:1.4 (g / mL); the components (g / L) of the nutrient solution are as follows: tryptone 5, glucose 20, KH2PO4 2, MgSO4·7H2O 1, VB10.02.

[0011] In the solid fermentation culture, the prepared seed liquid was added to the prepared solid culture medium at an inoculum rate of 6% (v / v), and the culture was carried out in a climate box for 23 days. Three parallel experiments were set up for each group of experiments.

[0012] The solid fermentation culture conditions are as follows: the culture medium is placed in a climate chamber at a controlled temperature of 26°C and a humidity of 50-80% (e.g., 75%), incubated in the dark for 5 days. Dark culture can be terminated after mycelium germinates, covers the surface of the culture medium, and grows to the bottom. After stroma formation, supplemental illumination (1000 lx, 24 hours / day) is applied, and several small holes are punched in the sealing film. After 23 days of culture, the culture is terminated, and the fruiting bodies are cut from the stroma, collected, dried at 60°C, and sealed for storage at -80°C.

[0013] In a third aspect, the present invention claims protection for the ethanol extract of the fruiting body of the Cordyceps sinensis strain described above.

[0014] The ethanol extract is a substance obtained by extracting the fruiting body with 10% ethanol by volume.

[0015] Furthermore, the ethanol extract can be prepared according to a method comprising the following steps: taking the dried fruiting body, grinding it and passing it through a 60-mesh sieve to obtain a powder; mixing the powder with 10% ethanol by volume in a ratio of 0.2 (g):6 (mL), and ultrasonicating for 90 minutes; centrifuging (such as centrifuging at 4°C 4500r / min for 30 minutes) and then sterile filtering to obtain the ethanol extract.

[0016] In a fourth aspect, the present invention claims protection for the use of the cicada fungus described in the first aspect above, the fruiting body described in the second aspect above, or the ethanol extract described in the third aspect above in all or part of the following:

[0017] (A1) Repairing sensitive skin or preparing sensitive skin repair products;

[0018] (A2) whitening or preparing whitening products;

[0019] (A3) Antioxidant or preparation of antioxidant products;

[0020] (A4) Anti-inflammatory or preparation of anti-inflammatory products;

[0021] (A5) Anti-ultraviolet damage or preparation of anti-ultraviolet damage products.

[0022] In a fifth aspect, the present invention claims protection for a product having all or part of the following functions (B1) to (B5):

[0023] (B1) Sensitive skin repair;

[0024] (B2) Whitening;

[0025] (B3) Antioxidant;

[0026] (B4) anti-inflammatory;

[0027] (B5) Anti-ultraviolet damage.

[0028] The main component or one of the main components of the product claimed in the present invention is the ethanol extract described in the third aspect above.

[0029] Wherein, the product is a cosmetic or a medicine.

[0030] In a sixth aspect, the present invention claims protection for the use of the cicada fungus described in the first aspect above, the fruiting body described in the second aspect above, the ethanol extract described in the third aspect above, or the product described in the fifth aspect above in any of the following:

[0031] (C1) Reduce the relative activity of intracellular tyrosinase;

[0032] (C2) reduce intracellular melanin content;

[0033] (C3) inhibits tyrosinase activity in vitro;

[0034] (C4) scavenges DPPH free radicals;

[0035] (C5) scavenging reactive oxygen species;

[0036] (C6) Reduce the expression of capsaicin receptor in cells;

[0037] (C7) inhibits abnormal activation of cellular capsaicin receptors;

[0038] (C8) Reduce intracellular IL-6 expression;

[0039] (C9) Improves elastin damage caused by ultraviolet rays.

[0040] In a seventh aspect, the present invention claims protection for the use of the cicada fungus described in the first aspect above, the fruiting body described in the second aspect above, or the ethanol extract described in the third aspect above in any of the following:

[0041] (D1) preparing or acting as a tyrosinase inhibitor;

[0042] (D2) preparing or acting as a DPPH free radical scavenger;

[0043] (D3) preparing or acting as a reactive oxygen species scavenger;

[0044] (D4) Preparation or use as a capsaicin receptor inhibitor.

[0045] The present invention obtains the original strain, i.e., Cordyceps cicadae ANTI-AGE1, from wild cicada fungi isolated from Huzhou, Zhejiang Province, by microbial strain separation and purification technology. On this basis, the present invention also realizes artificial cultivation of the above-mentioned new strain of cicada fungi on barley solid culture medium, and can realize large-scale preparation. The cicada fungi fruiting bodies obtained by artificial culture are extracted using 10% ethanol. After efficacy evaluation, the ethanol extract has the effects of sensitive skin repair, whitening, anti-oxidation, anti-inflammatory, and anti-ultraviolet damage. It is worth mentioning that the extract of the present invention is the edible fruiting body part obtained by solid fermentation from wheat solid culture medium. The cicada fungi fruiting bodies obtained by solid fermentation and the cicada fungi mycelium obtained by liquid fermentation are both from cicada fungi, but the two are very different. The fruiting body is the spore-producing structure of higher fungi, i.e., the fruiting body, which is formed by the specialization of aerial mycelium. Spores can be produced inside or on the fruiting body. The spores have a specific morphology, structure and bioactive substances. The fruiting body is the main edible and medicinal part of edible and medicinal fungi. The mycelium is similar to the root of a plant. It is a structure composed of filamentous fungal cells that are intertwined and interwoven with each other. It is responsible for absorbing water and nutrients and providing nutrition for the fungus. It is usually located in a matrix or on a culture medium. Generally speaking, mycelium is cultivated by liquid fermentation, which usually takes 3-6 days to complete; while the fruiting body is cultivated by solid culture medium, which takes about 30 days. The two have different development and growth times and their metabolites are different. For example, triterpenoids with important biological activity are almost absent in the mycelium. The growth mode of natural Cordyceps cicadae is similar to that of solid fermentation fruiting bodies, rather than liquid fermentation mycelium, so the value of the two to human health is different. The extraction solvent used in the present invention is ethanol, which is basically harmless to the human body, which guarantees its safety to the greatest extent. The above shows that the Cordyceps cicadae ANTI-AGE1 provided by the present invention is a new strain with great development prospects in terms of cosmetic raw materials.

[0046] Preservation Instructions

[0047] Classification and nomenclature: Cordyceps cicadae;

[0048] Reference biological material: ANTI-AGE1;

[0049] Depository: General Microbiology Center of China Culture Collection Administration of Microorganisms;

[0050] Abbreviation of depository institution: CGMCC;

[0051] Address: No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing;

[0052] Deposit date: March 11, 2024;

[0053] The registration number of the CGMCC Collection Center is: CGMCC No.41179. BRIEF DESCRIPTION OF THE DRAWINGS

[0054] Figure 1 This is a photo of the wild cicada fungus collected from Huzhou, Zhejiang Province.

[0055] Figure 2 The similarity between Cordyceps cicadae ANTI-AGE1 and other cicada fungus strains obtained by phylogenetic tree analysis based on ITS sequence sequencing results.

[0056] Figure 3 Flow chart for the artificial cultivation of Cordyceps cicadae ANTI-AGE1 and the preparation of its ethanol extract.

[0057] Figure 4 This is a photo of the fruiting body of cicada fungus cultivated using solid culture medium.

[0058] Figure 5 This study evaluated the whitening activity of an ethanol extract from the fruiting body of the artificially cultivated cicada fungus ANTI-AGE1. A shows the relative tyrosinase activity assay (cell-based assay); B shows the in vitro tyrosinase inhibition assay (biochemical assay); and C shows the relative melanin content assay.

[0059] Figure 6 This study evaluated the antioxidant activity of the ethanol extract from the fruiting body of the artificially cultivated cicada fungus ANTI-AGE1. A shows the DPPH free radical scavenging rate; B shows the relative fluorescence intensity of reactive oxygen species (ROS).

[0060] Figure 7This is a functional evaluation of the ethanol extract of the ANTI-AGE1 fruiting body of artificially cultivated cicada fungus (Cordyceps cicadae) on the repair of sensitive skin. A is the detection result of the ethanol extract of the ANTI-AGE1 fruiting body of artificially cultivated cicada fungus (Cordyceps cicadae) on the expression level of TRPV1 in HaCaT cells (screening of capsaicin modeling conditions); B is the relative gray value result diagram corresponding to the detection result of the ethanol extract of the ANTI-AGE1 fruiting body of artificially cultivated cicada fungus (Cordyceps cicadae) on the expression level of TRPV1 in HaCaT cells under capsaicin modeling conditions; C is the detection result of the ethanol extract of the ANTI-AGE1 fruiting body of artificially cultivated cicada fungus (Cordyceps cicadae) on the expression level of TRPV1 in HaCaT cells under capsaicin modeling conditions; D is the relative gray value result diagram corresponding to the detection result of the ethanol extract of the ANTI-AGE1 fruiting body of artificially cultivated cicada fungus (Cordyceps cicadae) on the expression level of TRPV1 in HaCaT cells; E is the detection result of the ethanol extract of the ANTI-AGE1 fruiting body of artificially cultivated cicada fungus (Cordyceps cicadae) on the expression level of TRPV1 in HaCaT cells under capsaicin modeling conditions; A is a graph showing the inhibitory effect of the ethanol extract of the ANTI-AGE1 fruiting body of cicada fungus (Cordyceps cicadae) on the activation of TRPV1 in HaCaT cells under capsaicin modeling conditions; F is a graph showing the inhibitory effect of the ethanol extract of the ANTI-AGE1 fruiting body of artificially cultivated cicada fungus (Cordyceps cicadae) on the activation of TRPV1 in HaCaT cells under capsaicin modeling conditions in the presence of TRPV1 antagonist (BCTC).

[0061] Figure 8 To evaluate the anti-inflammatory function of the ethanol extract of the ANTI-AGE1 fruiting body of artificially cultivated cicada fungus (Cordyceps cicadae) (IL-6 expression level detection results).

[0062] Figure 9 To evaluate the anti-ultraviolet damage function of the ethanol extract of the ANTI-AGE1 fruiting body of artificially cultivated cicada fungus (Cordyceps cicadae) (detection results of human skin elastin ELN expression level). DETAILED DESCRIPTION

[0063] The present invention will be further described in detail below in conjunction with specific embodiments. The examples provided are only for illustrating the present invention and are not intended to limit the scope of the present invention. The examples provided below can serve as a guide for further improvements by those skilled in the art and are not intended to limit the present invention in any way.

[0064] Unless otherwise specified, the experimental methods in the following examples are conventional methods and were performed according to the techniques or conditions described in the literature in the field or according to the product instructions. The materials and reagents used in the following examples, unless otherwise specified, were all commercially available.

[0065] Example 1: Isolation and Identification of Anti-Age 1 from Cordyceps cicadae

[0066] The cicada fungus (Cordyceps cicadae) ANTI-AGE1 of the present invention is an original strain obtained from wild cicada fungus isolated from Huzhou, Zhejiang Province through a microbial strain separation and purification technology.

[0067] The strain of the present invention was collected from wild cicada fungus ( Figure 1 The conidia of the cicada pupa consist of bundles of conidial stalks that emerge from the head of the cicada pupa. The insect's body is brownish-yellow in color, covered with gray or white mycelium. The conidial stalks are 2.1-5.6 cm long and may or may not be branched. The upper fertile portion is 6-9 mm long and 2-3 mm in diameter, and is generally oblong, elliptical, or spindle-shaped, bearing numerous white, powdery conidia.

[0068] 1. Strain isolation and purification

[0069] Cut off the spore stalk of Cicadae and put it into a 15mL centrifuge tube containing 6mL of 0.9% NaCl solution and shake it up and down to release the fungal spores. -1 , 10 -2 , 10 -3 , 10 -4 Dilute and spread 200 μL of the spore suspension onto Red Bengal medium. Incubate in the dark at 26°C. Once a single colony grows, use an inoculating loop to pick the edge of the mycelium and streak it onto Red Bengal medium. Purify 3-5 times to confirm that it is contaminant-free. Transplant onto a PDA slant, incubate at 26°C for 7 days, and then transfer to 4°C for storage to obtain strain ANTI-AGE1. Red Bengal medium (g / L): 5g peptone, 10g glucose, 1g KH2PO4, 0.5g MgSO4, 0.033g Red Bengal, 0.1g chloramphenicol, and 20g agar.

[0070] 2. Identification of strains

[0071] After purification in step 1, strain ANTI-AGE1 was cultured in PDA liquid medium for 3 days. Mycelia were collected by centrifugation at 4500 rpm and 4°C, dried at low temperature (40°C), and ground with liquid nitrogen. Genomic DNA was extracted using an Ezup column-based fungal genomic DNA extraction kit to obtain a genomic DNA solution. ITS sequences were amplified using the universal primers ITS-1F (5'-TCCGTAGGTGAACCTGCGG-3') and ITS-4R (5'-TCCTCCGCTTATTGATATGC-3') for the fungal ribosomal intergenic spacer region. The PCR reaction mixture (50 μl) consisted of 25 μl of PrimeSTAR Max Premix, 2 μl of primers ITS-1F, 2 μl of primers ITS-4R, 1 μl of DNA template, and 20 μl of ddH2O. PCR amplification conditions were: 94°C for 5 min, 94°C for 1 min, 55°C for 1 min, and 72°C for 1 min for 30 cycles, and 72°C for 10 min. The PCR product was directly submitted for sequencing. The resulting ITS sequence is shown in SEQ ID No. 1.

[0072] The ITS sequence sequencing results were subjected to BLAST analysis at https: / / blast.ncbi.nlm.nih.gov / Blast.cgi, and the sequence similarity between the ITS sequence and the Cordyceps cicadae strain was 100%. A phylogenetic tree analysis was performed based on the ITS sequence sequencing results ( Figure 2 ) and found that the isolate shared the highest similarity of 99% with the cicada fungus strain Minfu13 from the cicada fungus Cordyceps cicadae. Furthermore, considering that the isolate was isolated from wild cicada fungi in Huzhou, Zhejiang Province, and was the most dominant fungus in the wild cicada fungus during isolation, further characterization of the strain's conidial morphology revealed that conidiophores were 6-8 × 3-4 μm, bottle-shaped, swollen in the middle, tapering or suddenly narrowing at the end, and often clustered on the spore bundles. Conidia were 5-15 × 1.6-3.6 μm in size and oblong or spindle-shaped. The macroscopic and microscopic characteristics of this fungus were consistent with those described for Cordyceps cicadae, leading to the identification of this strain as a new strain of Cordyceps cicadae and its designation as Cordyceps cicadae ANTI-AGE1. The strain was deposited on March 11, 2024 with the China General Microbiological Culture Collection Center, located at No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences, with the deposit number: CGMCC No.41179.

[0073] Example 2: Artificial Cultivation of Cordyceps cicadae Anti-AGE1 (Preparation of Fruiting Bodies)

[0074] The artificial cultivation (fruiting body preparation) process of Cordyceps cicadae ANTI-AGE1 in the present invention is as follows: Figure 3 It is shown in .

[0075] Take the activated cicada fungus strain ANTI-AGE1 slant and cut 1×1 cm 2 Three different clods of different sizes were inoculated into sterilized seed culture medium and incubated at 26°C, 140 rpm in a shaker for 3 days. The seed culture medium (g / L) consisted of: 20g glucose, 10g yeast powder, 10g peptone, 2g KH2PO4, 0.5g MgSO4, 0.5g ZnSO4; pH was natural.

[0076] Prepare a solid fermentation medium with the following formula: 25g wheat + 35mL nutrient solution (wheat: nutrient solution ratio: 1.4g / mL). The nutrient solution contains (g / L): 5g tryptone, 20g glucose, 2g KH2PO4, 1g MgSO4·7H2O, and 0.02g VB1. Seal the prepared medium, place it in an autoclave, and sterilize it at 115°C for 60 minutes. After the solid fermentation medium cools, bring it to room temperature under sterile conditions. Inoculate 6mL of the seed solution into the solid fermentation medium. Then, incubate the inoculated medium in a climate chamber. Incubate at 26°C, 75% humidity, and incubate in the dark for 5 days. End the dark incubation period when mycelium germinates, covers the surface of the medium, and grows to the bottom. Once stroma is formed, supplement with light (1000lx, 24h / d) and poke several small holes in the sealing film. After 23 days of culture, the culture was stopped, and the fruiting bodies were cut from the fruiting bodies, collected, dried at 60°C, sealed and stored at -80°C. Figure 4 It is the fruiting body of cicada fungus cultivated using solid culture medium.

[0077] Example 3: Preparation of ethanol extract from artificially cultivated ANTI-AGE1 fruiting bodies of Cordyceps cicadae

[0078] The process for preparing the ethanol extract of the artificially cultivated ANTI-AGE1 fruiting body of Cordyceps cicadae is as follows: Figure 3 It is shown in .

[0079] Fruiting bodies of the cultivated cicada fungus (Cordyceps cicadae) ANTI-AGE1 were collected and oven-dried at 60°C to constant weight. The mixture was then ground in a mortar and passed through a 60-mesh sieve to obtain a powder. A 0.2 g portion of the powder was added to 6 mL of 10% (v / v) ethanol, sonicated for 90 minutes, and centrifuged at 4500 rpm at 4°C for 30 minutes. The supernatant was filtered through a 0.22 μm filter to obtain a 10% ethanol extract.

[0080] Example 4: Evaluation of the Whitening Function of the Ethanol Extract of the Anti-AGE1 Fruiting Body of Artificially Cultivated Cordyceps cicadae

[0081] 1. Methods

[0082] 1. Melanoma B16F10 cell culture

[0083] According to 5×10 4 Melanoma B16F10 cells were seeded in a 96-well cell culture plate at a cell density of 10 cells / mL, and cultured in DMEM medium containing penicillin, streptomycin, 10% fetal bovine serum (FBS), and 10 μg / mL α-MSH in a cell culture incubator at 37°C and 5% CO2 for 24 hours before use.

[0084] 2. Relative tyrosinase activity assay (cells)

[0085] Melanoma B16F10 cells grown on a 96-well plate were grouped as follows: a blank group (medium only), a solvent control group (an equal volume of extract solvent, i.e., 10% ethanol by volume), a sample group (an 80-fold dilution group, a 40-fold dilution group, and a 20-fold dilution group of the "10% ethanol extract" obtained in Example 3), and a positive control group (arbutin solutions at concentrations of 200, 400, and 800 μg / mL, respectively), with three replicates per group.

[0086] Specific process: Add 10 μL of treatment solution to the cell culture wells (100 μL / well) and continue culturing in a cell culture incubator at 37°C and 5% CO2 for 24 hours. After the incubation period, carefully remove the medium containing the treatment solution, gently wash twice with PBS, add 0.5% TritonX-100, place in a -80°C freezer, lyse for 2 hours, then remove the 96-well plate, thaw naturally at room temperature, add 10 mmol / L levodopa (L-SOPA) solution, react at 37°C for 2.5 hours, and use a microplate reader to measure the absorbance (A) of the cells in each well at 475 nm to calculate the relative intracellular tyrosinase activity.

[0087] Relative tyrosinase activity = (As-Ab) / (Ac-Ab) × 100%;

[0088] Wherein, As is the absorbance A value of the sample group measured at 475 nm; Ac is the absorbance A value of the solvent control group measured at 475 nm; Ab is the absorbance A value of the blank group measured at 475 nm.

[0089] 3. In vitro determination of the inhibition of tyrosinase activity by samples (biochemical method)

[0090] The "10% ethanol extract" obtained in Example 3 was diluted 20-fold, 40-fold, and 80-fold, respectively, and used as test solutions along with positive control arbutin solutions at concentrations of 200, 400, and 800 μg / mL. According to the experimental reaction system in Table 1, 1.66 mmol / L L-tyrosine, phosphate buffer solution (PBS), and the test solution were added to the test tube in sequence and incubated at 37°C for 5 minutes. Then, 900 U / mL of tyrosinase was added, mixed, and incubated at 37°C for another 20 minutes. Finally, the absorbance was measured at 475 nm using a microplate reader.

[0091] Table 1. Tyrosinase activity inhibition test reaction system (unit: μL)

[0092] Components C1 C2 T1 T2 L-Tyrosine 200 200 200 200 Test liquid 0 0 200 200 PBS 150 200 150 200 10% ethanol 200 200 0 0 Tyrosinase 50 0 50 0 Total volume 600 600 600 600

[0093] The inhibition rate of the sample on tyrosinase activity = [(C1-C2)-(T1-T2)] / (C1-C2)×100%

[0094] Where: C1——Absorbance value of blank enzyme system

[0095] C2——Absorbance value of blank enzyme-free system

[0096] T1——Absorbance value of the sample group with enzyme system

[0097] T2——Absorbance value of the sample group without enzyme system

[0098] 4. Cell melanin content determination method

[0099] Melanoma B16F0 cells were cultured at a rate of 5×10 4 The cells were seeded in a 6-well cell culture plate at a density of 10 cells / mL and cultured in DMEM medium containing penicillin, streptomycin, 10% fetal bovine serum (FBS), and 10 μg / mL α-MSH in a cell culture incubator at 37° C. and 5% CO 2 for 24 h before use.

[0100] Melanoma B16F10 cells grown on a 6-well plate were grouped as follows: a blank group (medium only), a solvent control group (an equal volume of extract solvent, i.e., 10% ethanol by volume), a sample group (an 80-fold dilution group, a 40-fold dilution group, and a 20-fold dilution group of the "10% ethanol extract" obtained in Example 3), and a positive control group (arbutin solutions at concentrations of 200, 400, and 800 μg / mL, respectively), with three replicates per group.

[0101] After adding the test samples to the cells of different groups, they were cultured in a cell culture incubator at 37°C and 5% CO2 for 48 h. The cells were then digested with trypsin, collected, and counted. The cell number was adjusted to 2×10 7 Take 0.5 mL of cells and add 0.5 mL of 1 mol / L NaOH solution containing 10% DMSO. Place in an 80°C water bath for 1 h to completely dissolve the cell clumps. Measure the absorbance (A) at 475 nm using a microplate reader and calculate the relative melanin content in the cells.

[0102] Relative melanin content = (As-Ab) / (Ac-Ab) × 100%;

[0103] Wherein, As is the absorbance A value of the sample group measured at 475 nm; Ac is the absorbance A value of the solvent control group measured at 475 nm; Ab is the absorbance A value of the blank group measured at 475 nm.

[0104] 5. Data Analysis

[0105] Statistical analysis was performed using SPSS Statistics 27 software. Experimental data are expressed as mean ± standard deviation, and independent sample t-test was used. Compared with the solvent control group: *p < 0.05, **p < 0.01, ***p < 0.001.

[0106] 2. Results and Analysis

[0107] Tyrosinase activity assay - (cell) results are as follows Figure 5 As shown in Figure A, compared to the 10% ethanol solvent control, 200-800 μg / mL of arbutin significantly reduced tyrosinase activity in melanoma B16F10 cells (p < 0.001 or p < 0.01). The "10% ethanol extract" obtained in Example 3, when applied to B16F10 cells, exhibited a concentration-dependent inhibitory effect on tyrosinase activity. The 10% ethanol extract diluted 20-fold showed the strongest ability to reduce tyrosinase activity in melanoma B16F10 cells (p < 0.01), with an inhibition rate of 32.32%. However, the 10% ethanol extract of Cicadae confusa diluted 80-fold had no significant effect.

[0108] Secondly, the results of the inhibitory effect of 10% ethanol extract of Cicadae confusa on tyrosinase in vitro are as follows Figure 5 As shown in Figure B. Compared with the 10% ethanol solvent control, 200-800 μg / mL arbutin significantly reduced the tyrosinase activity in the in vitro biochemical reaction (p < 0.001). The in vitro inhibition results of the "10% ethanol extract" obtained in Example 3 on tyrosinase were similar to those in Figure 5 The results of inhibition of intracellular tyrosinase activity of medium A were consistent and showed concentration dependence. The 10% ethanol extract diluted 20, 40 and 80 times showed an effect similar to arbutin, with inhibition rates reaching 51.91%, 45.09% and 36.09% respectively, which significantly reduced the activity of tyrosinase in in vitro biochemical reactions (p<0.001).

[0109] Finally, the results of the regulatory effect of 10% ethanol extract of Cicadae confusa on melanin in melanoma B16F10 cells are shown in Figure 2. Figure 5 As shown in Figure C. Similar to 200-800 μg / mL arbutin, the "10% ethanol extract" obtained in Example 3 was diluted 40 times and applied to B16F10 cells. The melanin content in the cells was significantly reduced by 16.51% (p < 0.05) compared with the solvent control group, which was similar to the 200 μg / mL positive control arbutin, which decreased by 23.88% compared with the solvent control group.

[0110] In summary, the "10% ethanol extract" obtained in Example 3 can significantly reduce the melanin content in melanoma B16F10 cells (ie, has a whitening effect), and this down-regulation effect is directly related to its inhibitory effect on tyrosinase.

[0111] Example 5. Evaluation of the antioxidant function of the ethanol extract of the ANTI-AGE1 fruiting body of artificially cultivated cicada fungus (Cordyceps cicadae).

[0112] 1. Methods

[0113] 1. DPPH free radical scavenging ability detection method

[0114] The DPPH free radical, a very stable nitrogen-centered free radical, is one of the important indicators of a sample's antioxidant capacity and is widely used in the research of antioxidant foods, health products, and pharmaceuticals. The DPPH free radical has a single electron, its alcohol solution is purple, and has strong absorption at 515nm. When an antioxidant is present, the DPPH free radical is scavenged, the solution color becomes lighter, and the absorbance at 515nm decreases. Within a certain range, the change in absorbance is proportional to the degree to which the free radical is scavenged. In the present invention, the degree of decrease in absorbance is used to reflect the sample's ability to scavenge DPPH free radicals. 1 mL of different concentrations of the "10% ethanol extract" obtained in Example 3 was taken as the sample solution to be tested, and the DPPH free radical scavenging rate was calculated in strict accordance with the instructions of the DPPH free radical scavenging ability test kit (Solarbio, BC4750).

[0115] 2. Reactive oxygen species (ROS) determination method

[0116] (1) Human skin fibroblast (HSF) cell culture

[0117] Human skin fibroblasts (HSF cells) were cultured in a cell culture incubator at 37°C and 5% CO2 using DMEM medium containing penicillin, streptomycin, and 10% fetal bovine serum (FBS). HSF cells in the logarithmic growth phase were cultured in DMEM medium containing 10% FBS to adjust the cell density to 5×10 4 cell / mL, inoculate into 6-well plates at 2.5 mL / well and culture for 24 h before use.

[0118] (2) Group settings

[0119] Sample treatment groups: the "10% ethanol extract" obtained in Example 3 diluted 20-fold, 40-fold, and 80-fold were added, and H2O2 treatment (40 μmol / L) was performed simultaneously;

[0120] Blank group: added an equal amount of extract solvent, i.e., 10% ethanol by volume, without H2O2 treatment;

[0121] Solvent control group: an equal amount of extract solvent, i.e. 10% by volume ethanol, was added and H2O2 treatment (40 μmol / L) was performed simultaneously.

[0122] Three parallels per group.

[0123] (3) Relative fluorescence intensity detection method of reactive oxygen species (ROS)

[0124] When the cells reached a density of 70%-80% in a 96-well plate, they were treated with different sample concentrations and incubated in a CO2 incubator for 24 hours. After treatment with H2O2, the cells were incubated for an additional 5 hours. The culture medium in each well was aspirated and 200 μL of DCFH-DA (2,7-dichlorofluorescein diacetate, a fluorescent probe for reactive oxygen species (ROS)) solution (dissolved in DMSO, final concentration: 10 μM) was added. The cells were incubated at 37°C for 15 minutes, and the DCFH-DA solution was aspirated. The cells were rinsed three times with serum-free medium to remove any unincorporated DCFH-DA. After rinsing, 100 μL of PBS was added to each well. Fluorescence intensity in each well was measured using a multi-functional microplate reader under the following detection conditions: an excitation wavelength of 485 nm and an emission wavelength of 525 nm. DCFH-DA itself is non-fluorescent and freely crosses the cell membrane. Once inside the cell, it is hydrolyzed by cellular esterases to form DCFH. DCFH, however, cannot penetrate the cell membrane, making it easy for the probe to be loaded into cells. Intracellular reactive oxygen species can oxidize the non-fluorescent DCFH to produce fluorescent DCF. Therefore, detecting the fluorescence of DCF can reflect the level of reactive oxygen species (ROS) in cells.

[0125] 3. Data Analysis

[0126] Statistical analysis was performed using SPSS Statistics 27 software. Experimental data are expressed as mean ± standard deviation, and independent sample t-test was used. Compared with the solvent control group: *p < 0.05, **p < 0.01, ***p < 0.001.

[0127] 2. Results and Analysis

[0128] 1. DPPH free radical scavenging ability test results

[0129] The results of the "10% ethanol extract" (extract diluted 5 to 320 times) obtained in Example 3 for scavenging DPPH free radicals in vitro are as follows: Figure 6 As shown in Figure A. As can be seen from the figure, at dilution ratios of 5, 10, 20, and 40 times, the DPPH free radical scavenging rates of the "10% ethanol extract" obtained in Example 3 were 82.78±0.95%, 79.53±0.96%, 81.69±0.95%, and 83.88±1.27%, respectively, indicating that it has a strong effect of scavenging DPPH free radicals.

[0130] 2. Reactive oxygen species (ROS) scavenging ability test results

[0131] like Figure 6As shown in Figure B, compared with the blank group, H2O2 treatment in the solvent control group significantly increased the intracellular reactive oxygen species (ROS) content (p < 0.01), indicating that the 40 μmol / L H2O2 model was successfully established. Compared with the solvent control group, the "10% ethanol extract" obtained in Example 3 at 40- and 20-fold dilutions significantly reduced the intracellular ROS content. In particular, at a 20-fold dilution, the "10% ethanol extract" obtained in Example 3 extremely significantly reduced the intracellular ROS content (p < 0.001), reaching 39.87%, indicating its strong ROS scavenging effect.

[0132] Example 6: Functional evaluation of the ethanol extract of the artificially cultivated ANTI-AGE1 fruiting body of Cordyceps cicadae on the repair of sensitive skin

[0133] 1. Methods

[0134] (1) Detection of capsaicin receptor expression in HaCaT keratinocytes

[0135] Sensitive skin (SS) refers specifically to a highly reactive state of the skin under physiological or pathological conditions, mainly occurring on the face. Clinically, the skin is prone to subjective symptoms such as burning, stinging, itching, and tightness when stimulated by physical, chemical, mental and other factors, with or without objective signs such as erythema, scaling, and capillary dilation. As people pay more and more attention to "sensitive skin", "sensitive skin" has become a hot topic. Studies have found that the formation of sensitive skin has a clear correlation with the pain and itch receptor TRPV1. When stimulated by external stimuli, the skin will produce pain, burning and itching. TRPV1 is a non-selective cation channel protein with high permeability to calcium ions. It is mainly distributed in the nervous system and can be activated by capsaicin to release extracellular cations Ca 2+ Influx of TRPV1 into the skin mediates a series of biological responses, such as increasing its own expression, exacerbating itching, high fever, pain, and skin inflammation. Clinical studies have shown that sensitive skin is often accompanied by activation of TRPV1 receptors, the degree of which is directly related to skin sensitivity and can be used as an indicator for evaluating sensitive skin. To characterize whether a 10% ethanol extract of Cicadae cicadae fruiting bodies has a soothing effect on sensitive skin, the function of a 10% ethanol extract of Cicadae cicadae fruiting bodies was evaluated in terms of TRPV1 protein expression levels in keratinocytes HaCaT and TRPV1 channel activity.

[0136] 1. HaCaT cell culture and treatment

[0137] (1) Resuscitate HaCaT cells one week in advance and observe the cell status carefully. The cells should meet the characteristics of rapid growth and strong vitality.

[0138] (2) The cells were digested with trypsin for 5 min, centrifuged and counted, and 5×10 4 1 mL of sample was spotted onto a 24-well plate at a concentration of cells / mL, and the plate was placed in a cell culture incubator at 37°C and 5% CO2. When the cell density reached about 80%, capsaicin at concentrations of 5 μg / mL, 50 μg / mL, and 500 μg / mL (solvent: DMSO) was added, respectively, to determine the concentration of capsaicin for inducing HaCaT cells to construct a skin cell sensitivity model; then, under the screened 5 μg / mL capsaicin treatment condition, the effect of different concentrations of "10% ethanol extract" (20-fold dilution, 40-fold dilution, 80-fold dilution) obtained in Example 3 on the TRPV1 protein expression level was detected after 24 hours of treatment with the model.

[0139] (3) Aspirate the cell culture medium and add 100 μL of 2× loading buffer. Stir continuously to fully lyse the cells. Use a pipette to transfer the cells to a 1.5 mL centrifuge tube and boil them for 10 min. Centrifuge at 12,000 g / min for 10 min. Carefully aspirate the supernatant, avoiding the bottom precipitate. Perform Western Blot hybridization to detect the expression level of the capsaicin receptor TRPV1 protein.

[0140] (4) Use the image processing software ImageJ to perform grayscale value analysis.

[0141] 2. Data Analysis

[0142] Statistical analysis was performed using SPSS Statistics 27 software. Data are presented as mean ± standard deviation, and independent-sample t-tests were performed. Compared with the extract solvent (10% ethanol) control group: *p < 0.05, **p < 0.01, ***p < 0.001; compared with the capsaicin solvent (DMSO) control group: #p < 0.05, ##p < 0.01.

[0143] (II) Detection of TRPV1 channel activity in HaCaT keratinocytes

[0144] The formation of sensitive muscles has a clear correlation with the pain and itch receptor TRPV1, because TRPV1 is a Ca 2+ Highly permeable non-selective cation channel, whose activation can promote the release of Ca2+ from the sarcoplasmic reticulum to the cytoplasm 2+ , so intracellular Ca 2+ The level of Ca can indirectly verify the activation degree of TRPV1, and then reflect a series of itching, high fever and pain problems of the skin. 2+ Probes for detecting intracellular Ca 2+The content of TRPV1 was characterized by its level, and the activation degree and change trend of TRPV1 channel were characterized, thereby judging the inhibitory ability of 10% ethanol extract of Cicadae Convolvulus on TRPV1 activation of sensitive skin cells and characterizing its soothing ability on sensitive skin.

[0145] 1. HaCaT cell culture and treatment

[0146] (1) Resuscitate HaCaT cells one week in advance, and pay attention to the characteristics of the cells, such as rapid growth and strong vitality.

[0147] (2) After the cells adhered, they were divided into a control (equal volume of extract solvent, i.e., 10% ethanol by volume), an 80-fold dilution group, a 40-fold dilution group, and a 20-fold dilution group of the "10% ethanol extract" obtained in Example 3. After 24 h of treatment, 5 μg / mL of capsaicin, 5 μg / mL of capsaicin, and 100 μmol / L of BCTC (TRPV1 antagonist, N-(4-Tertiarybutylphenyl)-4-(3-cholorphyridin-2-yl)tetrahydropyrazine-1(2H)-carboxamide) were added to the cells in each group, and the cells were placed in a cell culture incubator and cultured for another 24 h.

[0148] (3) Use HBSS to dilute Fluo-4 AM (Calcium ion fluorescent probe Fluo-4 AM is an acetyl methyl ester derivative of Fluo-4, a fluorescent dye that can penetrate the cell membrane. The fluorescence of Fluo-4 AM is very weak, and its fluorescence does not increase with the increase of calcium ion concentration. After Fluo-4 AM enters the cell, it can be cleaved by intracellular esterase to form Fluo-4, which is then retained in the cell. Fluo-4 can bind to calcium ions and produce strong fluorescence after binding to calcium ions. The maximum excitation wavelength is 494nm and the maximum emission wavelength is 516nm) into a working solution with a concentration of 4μM. Take out the 96-well plate, wash the cells thoroughly 3 times with HBSS, add the prepared working solution to the 96-well plate at a concentration of 200μL / well, and place it in an incubator at 37°C with 5% CO2 for 30 minutes. Take out the 96-well plate and wash the cells 3 times with HBSS. Under the conditions of excitation wavelength of 494nm and emission wavelength of 516nm, the fluorescence intensity of each well of cells was measured using a multifunctional microplate reader. The relative fluorescence intensity was obtained by comparing the absorbance value of each sample well with the absorbance value of the solvent control group containing only capsaicin.

[0149] 2. Data Analysis

[0150] Data were analyzed using SPSS Statistics 27 software. Experimental data are presented as mean ± standard deviation, and independent sample t-tests were performed. The capsaicin-only group, which contained neither the extract nor BCTC, was designated the model group. Comparisons between the sample group and the model group showed: *p ​​< 0.05, **p < 0.01.

[0151] 2. Results and Analysis

[0152] 1. Detection of capsaicin receptor expression

[0153] The effects of different concentrations of capsaicin on the expression of capsaicin receptor TRPV1 are shown in the following table. Figure 7 As shown in A and B. As can be seen from the figure, when the concentration of capsaicin is 5μg / mL, the expression of TRPV1 is significantly increased (p<0.05), increasing by 33.02%. Therefore, 5μg / mL of capsaicin is used as the concentration for modeling sensitive cells. In addition, under the condition of no capsaicin stimulation, different concentrations of dilution (80 times, 40 times, 20 times) of the "10% ethanol extract" obtained in Example 3 all reduced the expression of TRPV1 protein ( Figure 7 Among them, the 20-fold diluted "10% ethanol extract" had the most significant inhibitory effect on TRPV1 protein expression (p<0.001), with an inhibition rate of 64.48%.

[0154] The cells were treated with different dilutions of "10% ethanol extract" and stimulated by adding 5μg / mL capsaicin. The expression of capsaicin receptors in each sample group was as follows: Figure 7 As shown in Figures C and D, compared to the solvent control group treated with 5 μg / mL capsaicin, the "10% ethanol extract" obtained in Example 3, treated with different dilutions at different concentrations, reduced TRPV1 protein expression in cells (5 μg / mL capsaicin was added to the model). Among them, the extract diluted 20 times had the best inhibitory effect on TRPV1 protein (p < 0.001), with an inhibition rate of 33.16%, indicating that the "10% ethanol extract" obtained in Example 3 can inhibit TRPV1 expression in HaCaT cells.

[0155] 2. Detection of TRPV1 channel activation

[0156] In order to explore the regulatory effect of the "10% ethanol extract" obtained in Example 3 on TRPV1 channel activity, the activation degree and change trend of the TRPV1 channel were indirectly characterized by the change in intracellular calcium ion content, and then the soothing effect of the 10% ethanol extract of Cicadae cicadae on sensitive skin was determined. Capsaicin was used as a TRPV1 channel agonist, and the effects of different concentrations of the "10% ethanol extract" obtained in Example 3 on calcium ion content were tested. The results are as follows: Figure 7As shown in E and F. Figure 7 As shown in Figure E, compared with the model group treated with capsaicin, the 40-fold and 20-fold dilutions of the "10% ethanol extract" obtained in Example 3 significantly reduced the calcium ion concentration, especially the 40-fold dilution group and the model group. The difference was significant, reaching 14.83% (p<0.01). At the same time, in order to verify whether the decrease in intracellular calcium ion concentration is directly related to the activation of TRPV1 channels, the TRPV1-specific inhibitor BCTC was further added to detect whether the "10% ethanol extract" dilution obtained in Example 3 has a regulatory effect on TRPV1 channels in the presence of the TRPV1-specific inhibitor BCTC. The results are shown in Figure E. Figure 7 Figure F shows that after adding BCTC, a TRPV1-specific inhibitor, the intracellular calcium ion concentration decreased significantly (p<0.01) compared to the capsaicin model group, reaching 20.13%. This indicates that the intracellular calcium ion concentration stimulated by the capsaicin model group is positively correlated with the TRPV1 channel activity. At the same time, in the presence of BCTC, the change in intracellular calcium ion concentration was not affected by the "10% ethanol extract" obtained in Example 3. This indicates that the "10% ethanol extract" obtained in Example 3 inhibits the TRPV1 channel and thus reduces the increase in intracellular calcium ion concentration caused by capsaicin. Therefore, the "10% ethanol extract" obtained in Example 3 has an inhibitory effect on the activation of TRPV1 channels in keratinocyte HaCaT cells, which means that it has a soothing effect on sensitive skin.

[0157] Example 7: Evaluation of the anti-inflammatory function of the ethanol extract of the artificially cultivated fruiting body of Cordyceps cicadae ANTI-AGE1

[0158] 1. Methods

[0159] 1. HaCaT cell culture and treatment

[0160] (1) Resuscitate HaCaT cells one week in advance, and pay attention to the characteristics of the cells, such as rapid growth and strong vitality.

[0161] (2) After cell attachment, the cells were divided into a control group without 10 μg / mL LPS and an inflammation model group treated with 10 μg / mL LPS. The control group without 10 μg / mL LPS included a blank group (culture medium only), a solvent control group (equal volume of extract solvent, i.e., 10% ethanol by volume), and the "10% ethanol extract" obtained in Example 3 diluted 80-fold, 40-fold, and 20-fold; and the inflammation model group treated with 10 μg / mL LPS included a blank group (culture medium only), a solvent control group (equal volume of extract solvent, i.e., 10% ethanol by volume), and the "10% ethanol extract" obtained in Example 3 diluted 80-fold, 40-fold, and 20-fold.

[0162] (3) When the cell density in the well plate reached 70%-80%, the cells were treated with 10 μg / mL LPS according to the group and cultured for another 12 h. The supernatant was discarded and culture medium containing different concentrations of "10% ethanol extract" was added to the well plate. The well plate was incubated for 24 h, and the cell culture medium was collected for later use. The IL-6 content of the cells in each well was measured according to the instructions of the IL-6 ELISA Kit (Lianke Biotechnology EK106 / 2-96).

[0163] 2. Data Analysis

[0164] Statistical analysis was performed using SPSS Statistics 27 software. Data are presented as mean ± standard deviation, and independent sample t-tests were performed. The model group consisted of the LPS extract solvent (i.e., 10% ethanol by volume). *p < 0.05, **p < 0.01, ***p < 0.001 compared with the model group.

[0165] 2. Results and Analysis

[0166] The effects of different dilution concentrations of the "10% ethanol extract" obtained in Example 3 on the IL-6 content are shown in Figure 3. Figure 8 As shown. As can be seen from the figure, after LPS treatment of cells, the secretion of IL-6 can be significantly increased (p<0.01), reaching 14.68%. Compared with the solvent control group (i.e., the model group) treated with LPS, the different concentrations of dilutions of the "10% ethanol extract" obtained in Example 3 can reduce the content of IL-6, and are concentration-dependent. Among them, when the dilution ratio of the 10% ethanol extract is 20 times, the content of IL-6 is the lowest. And in the 20-fold and 40-fold dilution concentration groups, the content of IL-6 is significantly different from that of the model group (p<0.001), and the inhibition rates are 29.09% and 23.76%, respectively. It shows that the different concentrations of dilutions of the "10% ethanol extract" obtained in Example 3 have a certain anti-inflammatory effect on LPS-induced HaCaT cells.

[0167] Example 8 Evaluation of the Anti-UV Damage Function of the Ethanol Extract of the Anti-AGE1 Fruiting Body of Artificially Cultivated Cordyceps cicadae

[0168] 1. Methods

[0169] 1. Determination of human elastin (ELN) content in UVB-HacaT cell model

[0170] (1) Resuscitate HaCaT cells one week in advance and culture them in DMEM medium containing penicillin, streptomycin and 10% fetal bovine serum (FBS) in a cell culture incubator at 37°C and 5% CO2. Pay attention to the characteristics of the cells, such as rapid growth and strong vitality.

[0171] (2) After the cells adhered to the wall, they were divided into a control group without UV treatment and a UV treatment group (UVB 60mJ / cm 2 The control groups without UV treatment included: blank group (culture medium only), solvent control group (equal volume of extract solvent, i.e., 10% ethanol by volume), 80-fold dilution group, 40-fold dilution group, and 20-fold dilution group of the "10% ethanol extract" obtained in Example 3; and UV treatment (UVB 60mJ / cm 2 The model groups (irradiated for 5 min) included: a blank group (culture medium only), a solvent control group (an equal volume of extract solvent, i.e., 10% ethanol by volume), and the "10% ethanol extract" obtained in Example 3 diluted 80 times, 40 times, and 20 times.

[0172] (3) When the cell density in the well plate reaches 70%-80%, the supernatant is discarded and UVB irradiation is performed (60 mJ / cm2, irradiation for 5 minutes). Culture is continued for 12 hours, the supernatant is discarded, and culture medium containing different concentrations of extracts is added to the well plate. Incubate for 24 hours, and the cell culture fluid is collected for later use. According to the instructions of the Human Elastin ELISA Kit (CUSABIO CSB-EO9338h), the absorbance of the cells in each well at a wavelength of 450 nm is measured using a microplate reader, and the human elastin content in HaCaT cells is calculated based on the drawn standard curve.

[0173] 2. Data Analysis

[0174] Statistical analysis was performed using SPSS Statistics 27 software. Data are presented as mean ± standard deviation, and independent sample t-tests were performed. The model group consisted of the group treated with UVB and containing the extract solvent (i.e., 10% ethanol by volume). *p < 0.05, **p < 0.01, ***p < 0.001 compared with the model group.

[0175] 2. Results and Analysis

[0176] The results of the effect of the "10% ethanol extract" obtained in Example 3 on the secretion of human elastin are as follows: Figure 9 As shown in the figure, UVB treatment significantly reduced ELN secretion (p < 0.01) by 8.81%. Compared to the UVB-treated solvent control group (i.e., the model group), the "10% ethanol extract" obtained in Example 3 at different dilutions increased ELN concentrations in a concentration-dependent manner. The ELN content reached its highest level at a 20-fold dilution, significantly different from the model group (p < 0.01), reaching 14.06%. This indicates that the different dilutions of the "10% ethanol extract" obtained in Example 3 have a protective effect on the UVB-damaged HaCaT cell model.

[0177] The present invention has been described in detail above. For those skilled in the art, without departing from the purpose and scope of the present invention, and without the need to carry out unnecessary experimental conditions, the present invention can be implemented in a wide range under equivalent parameters, concentrations and conditions. Although the present invention provides specific embodiments, it should be understood that further improvements can be made to the present invention. In short, according to the principles of the present invention, this application is intended to include any changes, uses or improvements to the present invention, including changes that depart from the disclosed scope in this application and are made using conventional techniques known in the art.

Claims

1. A cicada fungus strain characterized by: The cicada fungus strain is cicada fungus ( Cordyceps cicadae )ANTI-AGE1, whose registration number in the General Microbiology Center of China Culture Collection Administration is CGMCC No.41179.

2. The fruiting body of the Cordyceps sinensis strain according to claim 1.

3. An ethanol extract of the fruiting body of the cicada fungus strain according to claim 1, wherein the ethanol extract is prepared according to a method comprising the following steps: taking the dried fruiting body, grinding it and passing it through a 60-mesh sieve to obtain a powder; mixing the powder with 10% by volume ethanol at a ratio of 0.2 g:6 mL, and sonicating for 90 minutes; centrifuging and sterile filtering to obtain the ethanol extract.

4. Use of the cicada fungus according to claim 1, the fruiting body according to claim 2, or the ethanol extract according to claim 3 in all or part of the following: (A1) Repairing sensitive skin or preparing sensitive skin repair products; (A2) Whitening or preparation of whitening products; (A3) Preparation of antioxidant products; (A4) Preparation of anti-inflammatory products; (A5) Preparation of UV damage resistant products; The product is a cosmetic.

5. Products that have all or part of the functions listed in (B1) to (B5) below, characterized in that: The main component or one of the main components of the product is the ethanol extract according to claim 3; (B1) Sensitive skin repair; (B2) Whitening; (B3) Antioxidant; (B4) anti-inflammatory; (B5) Anti-ultraviolet damage; The product is a cosmetic.

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