Preparation method of hericium erinaceus extract and application thereof in cosmetics
By optimizing the preparation process of Hericium erinaceus extract, Hericenones C was identified as the main active ingredient, solving the problem of unclear composition and achieving the effects of anti-UVB damage and cell growth promotion in cosmetics, thus expanding its application in skin care products.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- WEST ANHUI UNIV
- Filing Date
- 2026-04-21
- Publication Date
- 2026-06-09
AI Technical Summary
The active ingredients of Hericium erinaceus extract are unclear, making accurate component analysis and quality control difficult. Furthermore, there is a lack of systematic research on its anti-UVB damage activity in cosmetics, which limits its development and industrialization in the skincare field.
A purification process combining ethanol reflux extraction, ethyl acetate extraction, and MCI resin column chromatography was employed. The main active ingredient, Hericenones C, was separated by HPLC, resulting in a well-defined Hericium erinaceus extract for use in anti-UVB damage cosmetics.
The main active ingredients of Hericium erinaceus extract have been identified, laying the foundation for cosmetic quality control. It significantly enhances antioxidant capacity and cell proliferation effects, making it suitable for anti-wrinkle, repair, and anti-aging products, which meets market demand.
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Figure CN122167334A_ABST
Abstract
Description
Technical Field
[0001] This invention relates to the field of plant extract technology, and more specifically, to a method for preparing Hericium erinaceus extract and its application in cosmetics. Background Technology
[0002] Hericium erinaceus, a traditional edible and medicinal fungus, has been studied for its extracts in areas such as anti-oxidation and neurological disorders. However, the following technical problems exist: First, the specific active ingredients of Hericium erinaceus extract are unclear, making accurate component analysis and quality control difficult. Second, there are no systematic research reports on the anti-UVB damage activity of Hericium erinaceus extract and its application in cosmetics, limiting its development and application in the skincare field. Third, due to the unclear active ingredients, it is difficult to establish an effective quality standard system for the application of Hericium erinaceus extract in cosmetics, which seriously restricts its industrialization process.
[0003] The lack of systematic preparation methods for Hericium erinaceus extract in the existing technology, especially the lack of extraction and purification processes that can identify active ingredients and have anti-UV damage function, has become the main technical bottleneck restricting the application of Hericium erinaceus extract in the cosmetic industry. Summary of the Invention
[0004] To address the aforementioned technical problems, this invention provides a method for preparing Hericium erinaceus extract and its application in cosmetics.
[0005] A method for preparing Hericium erinaceus extract includes the following steps: Extraction: Take the raw material of Hericium erinaceus, add ethanol at a material-to-liquid ratio of 1:10, and extract by ethanol reflux 3 times, 3 hours each time. Combine the extracts and concentrate under reduced pressure to obtain the extract. Extraction: The extract was subjected to liquid-liquid extraction with ethyl acetate, the ethyl acetate layer was collected, concentrated and freeze-dried to obtain Hericium erinaceus extract; Purification: Hericium erinaceus extract was separated into five fractions (A, B, C, D, and E) using a medium-pressure MCI resin column preparative chromatography system. Fraction B was preferentially selected for the separation of active chemical components. Hericenones C was obtained by preparative HPLC (21.2 × 150 mm, C4, 5 μm, Welch) with 85% methanol elution (flow rate 15 mL / min, detection wavelengths 254 nm and 210 nm).
[0006] The main active ingredient in the Hericium erinaceus extract is Hericenones C.
[0007] Preferably, the temperature for ethanol reflux extraction is 80-90℃.
[0008] Preferably, the conditions for vacuum concentration are 60°C and 0.08 MPa.
[0009] Preferably, the resin column chromatography uses an MCI resin column.
[0010] Preferably, the components of the Hericium erinaceus extract are separated and identified by HPLC.
[0011] A hericium erinaceus extract prepared by the above method.
[0012] The application of the Hericium erinaceus extract in the preparation of cosmetics; the cosmetics are anti-wrinkle products, repair products or anti-aging products.
[0013] The application of the Hericium erinaceus extract in the preparation of anti-UVB damage products.
[0014] A cosmetic composition comprising the Hericium erinaceus extract, wherein the Hericium erinaceus extract has a mass concentration of 31.25-2000 μg / mL.
[0015] The beneficial effects of this invention are as follows: The process is optimized and reasonable: the purification process adopts ethanol reflux extraction combined with ethyl acetate extraction and resin column chromatography. The process route is clear, the operation is simple, and it is easy to implement on an industrial scale. Clear and controllable composition: HPLC analysis identified the main active ingredient of the extract as Hericenones C, laying the foundation for establishing quality control standards and solving the problem of unclear composition in existing technologies. Significant antioxidant activity: the IC50 value of DPPH free radical scavenging ability is 239±7 μg / mL; the IC50 value of hydroxyl radical (·OH) scavenging ability is 547±15 μg / mL; the Trolox equivalent antioxidant capacity (TEAC) of ABTS+· free radical scavenging ability is 477.3±3.1μmol TE / g, indicating that it has strong antioxidant capacity, which is superior to conventional plant extracts.
[0016] The cell proliferation effect was outstanding: at a concentration of 1000 μg / mL, the proliferation rate of HacaT skin cells reached 146.44 ± 1.27%, which significantly promoted cell growth.
[0017] With broad application prospects, it can be used as a natural, safe and efficient active ingredient in cosmetics for anti-wrinkle, repair and anti-aging products, meeting the market demand for natural skin care products and having significant economic value and social benefits. Attached Figure Description
[0018] Figure 1 This is a bar graph showing the effect of Hericium erinaceus fermentation broth extract of the present invention on the proliferation rate (%) of HaCaT cells. Detailed Implementation
[0019] The subject matter described herein will now be discussed with reference to exemplary embodiments. It should be understood that these embodiments are discussed only to enable those skilled in the art to better understand and implement the subject matter described herein, and changes may be made to the function and arrangement of the elements discussed without departing from the scope of this specification. Various processes or components may be omitted, substituted, or added as needed in the examples. Furthermore, some features described in the examples may be combined in other examples. Example 1
[0020] This embodiment presents a method for preparing Hericium erinaceus extract, comprising the following steps: Extraction: Take the raw material of Hericium erinaceus, add ethanol at a material-to-liquid ratio of 1:10, and extract by ethanol reflux 3 times, 3 hours each time. Combine the extracts and concentrate under reduced pressure to obtain the extract. The temperature for ethanol reflux extraction was 85℃; The conditions for vacuum concentration are 60℃ and 0.08MPa.
[0021] Extraction: The extract was subjected to liquid-liquid extraction with ethyl acetate, the ethyl acetate layer was collected, concentrated and freeze-dried to obtain Hericium erinaceus extract; Purification: Hericium erinaceus extract was separated into five fractions (A, B, C, D, and E) using a medium-pressure MCI resin column preparative chromatography system. Fraction B was preferentially selected for the separation of active chemical components. Hericenones C was obtained by preparative HPLC (21.2 × 150 mm, C4, 5 μm, Welch) with 85% methanol (flow rate 15 mL / min, detection wavelengths 254 nm and 210 nm).
[0022] Drying: The collected elution fraction was concentrated and then freeze-dried to obtain Hericium erinaceus extract; The components of the Hericium erinaceus extract were separated and identified by HPLC. The main active ingredient in the Hericium erinaceus extract is Hericenones C.
[0023] Example 2 Assay on HaCaT cell proliferation activity of Hericium erinaceus extract.
[0024] This embodiment proposes the application of the Hericium erinaceus extract described in Example 1 in the preparation of cosmetics; the cosmetics are anti-wrinkle products, repair products, or anti-aging products.
[0025] 1. Materials and Methods (1) Test materials: DMEM medium; fetal bovine serum (FBS); antibiotics; trypsin; PBS buffer (PBS, 0.01 mol / L pH 7.4); CCK-8.
[0026] (2) Test method: HaCaT cells were revived into T25 cell culture flasks, and 5 mL of 10% FBS cell culture medium (90% DMEM medium + 10% fetal bovine serum) was added. The flasks were then incubated at 37°C in a 5% CO2 incubator. When the cells adhered and grew to 80%-90% confluence, they were seeded into plates.
[0027] Logarithmic growth phase cells were seeded into 96-well plates. The CCK8 standard curve group was set with a cell concentration of 1000-10000 cells / well, and the sample group contained approximately 6000 cells per well. After 24 h of culture, the cell culture medium was aspirated. For the CCK8 standard curve group, 200 μL of 2% FBS cell culture medium (98% DMEM medium + 2% fetal bovine serum) was added to each well. For the sample group, a maximum concentration of 1000 μg / mL was used, and the cells were diluted twofold to 500 μg / mL, 250 μg / mL, 125 μg / mL, 62.5 μg / mL, and 31.25 μg / mL, respectively, creating six concentration gradients. 200 μL of Hericium erinaceus extract sample solution was added to each well according to the concentration gradient. After culturing for another 24 hours, the solution in the 96-well plate was aspirated, and 200 μL of DMEM medium containing 10% CCK-8 was added to each well. The plates were incubated at 37°C in a 5% CO2 incubator for 2 hours, and the absorbance was measured at 450 nm using a microplate reader. Three groups were set up: experimental group, control group, and blank group, with three parallel wells in each group. The proliferative activity of the test substance on HaCaT cells was calculated using the following formula.
[0028] Cell viability = [(As-Ab) / (Ac-Ab)] × 100% Where As is the absorbance value of the experimental well (including cells, culture medium, CCK-8 solution and sample solution). Ac is the absorbance value of the control well (including cells, culture medium, and CCK-8 solution, but excluding the sample solution). Ab represents the absorbance value of the blank well (including culture medium and CCK-8 solution, but excluding cells and sample solution).
[0029] 2. Experimental Results The results of the experiment on the proliferation of HaCaT cells by Hericium erinaceus extract are as follows: Figure 1 As shown, the optimal concentration of Hericium erinaceus extract for the proliferation activity of HaCaT cells was 1000 μg / mL, with a proliferation rate of (146.44 ± 1.27)%.
[0030] See Figure 1 Hericium erinaceus extract on HaCaT cell proliferation rate (%).
[0031] The above results indicate that, within a certain concentration range (31.25-1000 μg / mL), the Hericium erinaceus fermentation broth extract had no toxic effect on HaCaT cells and had a positive effect on cell proliferation. Therefore, the extract can be considered an effective active substance for promoting the proliferation of HaCaT skin cells.
[0032] Example 3 Determination of the DPPH free radical scavenging ability of Hericium erinaceus extract Accurately weigh DPPH and dissolve it thoroughly in anhydrous ethanol to prepare a 0.06 mg / mL DPPH solution. Add 100 μL of each concentration of sample solution to a 96-well plate and then add 100 μL of the DPPH solution. Mix well and incubate at 37°C in the dark for 30 min. Set up a positive control group and a blank control group, with three replicates. The positive control group used 1 mg / mL vitamin C solution as the sample. The blank control group used deionized water as the sample. Measure the absorbance at 517 nm using a microplate reader. Calculate the DPPH radical scavenging rate for each sample using the following formula and calculate the half-maximal inhibitory concentration (IC50). 50 value.
[0033] In the formula: A1 is the absorbance value of the sample group; A0 is the absorbance value of the blank control group.
[0034] The IC50 of Hericium erinaceus extract was determined by DPPH free radical scavenging assay. 50 The value was 239±7 μg / mL.
[0035] Example 4 Determination of the ability of Hericium erinaceus extract to scavenge hydroxyl radicals (·OH) Accurately pipette 0.25 mL of Hericium erinaceus ethanol extract sample solutions of different mass concentrations, add 0.25 mL of 0.75 mmol / L o-phenanthroline solution and 0.5 mL of 0.15 mol / L phosphate buffer (pH 7.4) to the sample solution and mix thoroughly. Then add 0.25 mL of 0.75 mmol / L FeSO4 solution and mix thoroughly. Finally, add 0.25 mL of 0.01% H2O2 solution to the reaction system and shake to mix. Incubate at 37℃ for 1.0 h. After the water bath reaction is complete, shake to mix the reaction solution, and use a multichannel pipette or single-channel pipette to transfer 200 μL to the wells of a 96-well plate. Measure the absorbance of the reaction solution at 536 nm. Calculate the ·OH scavenging rate of the sample based on the absorbance using the following formula:
[0036] In the formula: A1 is the absorbance after the sample and H2O2 are added and reacted; AS is the absorbance of the sample without H2O2 and FeSO4 (replaced with buffer solution); A0 is the absorbance of the sample without the sample and replaced with distilled water (with H2O2 added).
[0037] The IC50 of Hericium erinaceus extract was determined by hydroxyl radical (·OH) scavenging assay. 50 The value was 547±15 μg / mL.
[0038] Example 5 Hericium erinaceus extract clears ABTS + • Determination of free radical capacity ABTS solution and oxidant solution were mixed at a volume ratio of 1:1 and allowed to react at room temperature in the dark for 12-16 hours to obtain ABTS. + • Stock solution. ABTS is diluted with the sample solution during experimental testing. + • The stock solution is diluted approximately 30-55 times until its absorbance at 734 nm is within the range of 0.70 ± 0.02. The resulting ABTS is then... + • The assay solution can be used for ABTS samples + • Scavenging capacity assay. Trolox (water-soluble vitamin E) solution was used as the positive control group. The ratio of ABTS working solution to the test sample was 40:1. 7 μL of sample solution and 280 μL of ABTS were added at different mass concentrations. + • Mix the assay solution thoroughly in a 96-well plate, repeating the process in triplicate. Then, allow the mixture to stand at room temperature in the dark for 6 minutes. After the reaction, measure the absorbance of the reaction solution at 734 nm using a microplate reader. Calculate the ABTS of the sample based on the absorbance using the following formula. + • Clearance rate:
[0039] In the formula: Ai represents the amount of sample solution and ABTS added. + • The absorbance of the solution measured after the reaction of the test solution; A0 is the absorbance value of the solution measured after replacing the sample solution with distilled water.
[0040] via ABTS + • In the free radical scavenging experiment, the Trolox equivalent antioxidant capacity (TEAC) of Hericium erinaceus extract was 477.3 ± 3.1 μmol TE / g.
[0041] The above experimental data fully demonstrate the feasibility and benefits of this invention, providing a scientific basis for the application of Hericium erinaceus extract in cosmetics.
[0042] The embodiments of the present invention have been described above. However, the embodiments are not limited to the specific implementation methods described above. The specific implementation methods described above are merely illustrative and not restrictive. Those skilled in the art can make more equivalent embodiments under the guidance of the present embodiments, and all of them are within the protection scope of the present embodiments.
Claims
1. A method for preparing Hericium erinaceus extract, characterized in that, Includes the following steps: Extraction: Take the raw material of Hericium erinaceus, add ethanol at a material-to-liquid ratio of 1:10, and extract by ethanol reflux 3 times, 3 hours each time. Combine the extracts and concentrate under reduced pressure to obtain the extract. Extraction: The extract was subjected to liquid-liquid extraction with ethyl acetate, the ethyl acetate layer was collected, concentrated and freeze-dried to obtain Hericium erinaceus extract; Purification: Hericium erinaceus extract was separated by a medium-pressure MCI resin column preparative chromatograph to obtain five fractions: A, B, C, D, and E. Fraction B was preferentially selected for the separation of active chemical components. Hericene C was obtained by preparative HPLC elution with 85% methanol.
2. The preparation method according to claim 1, characterized in that, The ethanol reflux extraction temperature is 80-90℃.
3. The preparation method according to claim 1, characterized in that, The conditions for vacuum concentration are 60°C and a vacuum degree of -0.08 MPa.
4. The preparation method according to claim 1, characterized in that, The resin column chromatography used was an MCI resin column.
5. The preparation method according to claim 1, characterized in that, The components of the Hericium erinaceus extract were separated and identified by HPLC.
6. Hericium erinaceus extract prepared by the method according to any one of claims 1-5.
7. The application of the Hericium erinaceus extract according to claim 6 in the preparation of cosmetics.
8. The application according to claim 7, characterized in that, The cosmetics mentioned are anti-wrinkle products, repair products, or anti-aging products.
9. The use of the Hericium erinaceus extract according to claim 6 in the preparation of anti-UVB damage products.
10. A cosmetic composition comprising the Hericium erinaceus extract of claim 6, characterized in that, The mass concentration of the Hericium erinaceus extract is 31.25-2000 μg / mL.