Extracting composition of camellia oleifera flowers, camellia oleifera shells and camellia oleifera seed shells and application of extracting composition in anti-aging

Abstract

The invention provides an extraction composition of camellia oleifera flowers, camellia oleifera shells and camellia oleifera seed shells and application of the extraction composition in anti-aging. The extraction composition comprises the camellia oleifera flower alcohol extract, the camellia oleifera shell alcohol extract and the camellia oleifera seed shell alcohol extract according to the mass ratio of (1-4): (1-4): 1, hyaluronidase can be remarkably inhibited, the excellent anti-aging effect is achieved, generation of initial-stage product fructosamine and final-stage product AGEs of protein non-enzymatic glycosylation can be remarkably inhibited, and the anti-aging effect is good. The whole-process anti-aging effect from early prevention to later repair is realized. In addition, the extracted composition is analyzed by applying a medium effect principle, and the three components in the extracted composition are found to play a synergistic effect in the anti-aging aspect.

Description

Technical Field

[0001] This invention belongs to the field of cosmetic technology. More specifically, it relates to an extract composition of camellia flowers, camellia fruit shells, and camellia seed shells, and its application in anti-aging. Background Technology

[0002] In the process of skin aging, both non-enzymatic glycosylation of proteins and hyaluronidase have significant effects. In the early stages of non-enzymatic glycosylation, fructosamine is produced. As the reaction progresses, advanced glycation end products (AGEs) gradually accumulate. Both fructosamine and AGEs alter the biomechanical properties and biochemical state of the skin, inducing inflammatory responses. Furthermore, hyaluronidase catalyzes the hydrolysis of hyaluronic acid, leading to the loss of the cellular matrix and consequently causing the skin to lose elasticity and firmness.

[0003] Most anti-aging cosmetics currently contain chemical ingredients, but these ingredients are prone to side effects. Therefore, natural plant extracts with high safety levels are gradually standing out among many anti-aging cosmetics and have become a new type of product with high consumer acceptance. However, there are many plant extracts with anti-aging effects, and the efficacy of different plant extracts varies significantly. The compatibility of these extracts is also unpredictable. Therefore, in order to obtain a plant extract composition with better anti-aging effects, it is necessary to conduct extensive screening of plant extracts and carry out targeted research on their compatibility. Summary of the Invention

[0004] This invention addresses the shortcomings of existing technologies by providing an extraction composition that combines camellia flower alcohol extract, camellia fruit shell alcohol extract, and camellia seed shell alcohol extract to achieve a synergistic effect in anti-aging, thus offering a new option for natural anti-aging cosmetics.

[0005] The first objective of this invention is to provide an extraction composition.

[0006] A second objective of the present invention is to provide a method for preparing the above-described extraction composition.

[0007] A third objective of this invention is to provide the application of the above-described extraction composition in the preparation of cosmetics.

[0008] The above-mentioned objective of this invention is achieved through the following technical solution:

[0009] The present invention provides an extraction composition comprising, by mass ratio of 1-4:1-4:1, camellia flower alcohol extract, camellia fruit shell alcohol extract, and camellia seed shell alcohol extract.

[0010] This invention combines camellia flower ethanol extract, camellia seed shell ethanol extract, and camellia fruit shell ethanol extract. This combination not only significantly inhibits hyaluronidase, exhibiting excellent anti-aging effects, but also significantly inhibits the formation of fructosamine, the initial product of non-enzymatic glycosylation of proteins, and AGEs, the final product. This achieves a comprehensive anti-aging effect from initial prevention to later repair. Furthermore, this invention utilizes the principle of moderate efficacy to analyze the extract composition, discovering that the three components in the extract composition exert a synergistic effect in anti-aging.

[0011] Preferably, the mass ratio of camellia flower alcohol extract, camellia fruit shell alcohol extract and camellia seed shell alcohol extract is 1:1:1 or 4:1:1 or 1:4:1.

[0012] Preferably, the camellia flower alcohol extract is obtained by extracting camellia flowers with n-butanol or an aqueous ethanol solution.

[0013] More preferably, the camellia flowers are pulverized before extraction, such as to a mesh size of 40-200.

[0014] More preferably, the material is dried before pulverization, such as at 40-50°C, most preferably at 45°C.

[0015] More preferably, the ratio of camellia flower to n-butanol is 1g:13-17mL, and most preferably 1g:15mL.

[0016] More preferably, the ratio of camellia flowers to ethanol aqueous solution is 1g:13-17mL, and most preferably 1g:15mL.

[0017] More preferably, the concentration of the ethanol aqueous solution is 50% (v / v) to 80% (v / v), and most preferably 70% (v / v).

[0018] More preferably, the extraction is performed at 40–65°C for 40–85 min, and most preferably at 45°C for 45 min.

[0019] More preferably, the extraction is performed simultaneously with ultrasound.

[0020] More preferably, the frequency of the ultrasound is 180-220W, and most preferably 200W.

[0021] More preferably, after extraction, centrifugation is performed, such as centrifuging at 9000-11000×g for 18-22 min, and most preferably centrifuging at 10000×g for 20 min.

[0022] More preferably, the extraction is performed 1 to 2 times.

[0023] More preferably, after extraction, post-processing is performed, such as filtration, concentration, and freeze-drying in sequence.

[0024] More preferably, the concentration is a vacuum evaporation concentration, such as concentration at 40-50°C and -0.1-0 MPa.

[0025] More preferably, the freeze-drying temperature is -40 to -20°C.

[0026] Preferably, the camellia oleifera fruit shell alcohol extract is obtained by extracting camellia oleifera fruit shells with n-butanol or an aqueous ethanol solution.

[0027] More preferably, the camellia fruit shell is further crushed before extraction, such as to 40-200 mesh.

[0028] More preferably, the material is dried before pulverization, such as at 45-55°C, most preferably at 50°C.

[0029] More preferably, the ratio of camellia fruit shell to n-butanol is 1g:13-17mL, and most preferably 1g:15mL.

[0030] More preferably, the ratio of the amount of camellia fruit shell to the ethanol aqueous solution is 1g:13-17mL, and most preferably 1g:15mL.

[0031] More preferably, the concentration of the ethanol aqueous solution is 50% (v / v) to 80% (v / v), and most preferably 70% (v / v).

[0032] More preferably, the extraction is performed at 40–65°C for 40–85 min, and most preferably at 50°C for 50 min.

[0033] More preferably, the extraction is performed simultaneously with ultrasound.

[0034] More preferably, the frequency of the ultrasound is 180-220W, and most preferably 200W.

[0035] More preferably, after extraction, centrifugation is performed, such as centrifuging at 9000-11000×g for 18-22 min, and most preferably centrifuging at 10000×g for 20 min.

[0036] More preferably, the extraction is performed 1 to 2 times.

[0037] More preferably, after extraction, post-processing is performed, such as filtration, concentration, and freeze-drying in sequence.

[0038] More preferably, the concentration is a vacuum evaporation concentration, such as concentration at 40-50°C and -0.1-0 MPa.

[0039] More preferably, the freeze-drying temperature is -40 to -20°C.

[0040] Preferably, the camellia seed shell alcohol extract is obtained by extracting camellia seed shells with n-butanol or an aqueous ethanol solution.

[0041] More preferably, the camellia seed shells are further pulverized before extraction, such as to 40-200 mesh.

[0042] More preferably, the material is dried before pulverization, such as at 45-55°C, most preferably at 50°C.

[0043] More preferably, the ratio of camellia seed shell to n-butanol is 1g:28-32mL, and most preferably 1g:30mL.

[0044] More preferably, the ratio of camellia seed shell to ethanol aqueous solution is 1g:28-32mL, and most preferably 1g:30mL.

[0045] More preferably, the concentration of the ethanol aqueous solution is 50% (v / v) to 80% (v / v), and most preferably 70% (v / v).

[0046] More preferably, the extraction is performed at 40–65°C for 40–85 min, and most preferably at 60°C for 80 min.

[0047] More preferably, the extraction is performed simultaneously with ultrasound.

[0048] More preferably, the frequency of the ultrasound is 180-220W, and most preferably 200W.

[0049] More preferably, after extraction, centrifugation is performed, such as centrifuging at 9000-11000×g for 28-32 min, and most preferably centrifuging at 10000×g for 30 min.

[0050] More preferably, the extraction is performed 1 to 2 times.

[0051] More preferably, after extraction, post-processing is performed, such as filtration, concentration, and freeze-drying in sequence.

[0052] More preferably, the concentration is a vacuum evaporation concentration, such as concentration at 40-50°C and -0.1-0 MPa.

[0053] More preferably, the freeze-drying temperature is -40 to -20°C.

[0054] Based on this, the present invention also provides a method for preparing the above-mentioned extraction composition, which is obtained by mixing the camellia flower alcohol extract, the camellia fruit shell alcohol extract, and the camellia seed shell alcohol extract.

[0055] The above-mentioned extract composition not only significantly inhibits hyaluronidase, exhibiting excellent anti-aging effects, but also significantly inhibits the formation of fructosamine, the initial product of non-enzymatic glycosylation of proteins, and AGEs, the final product, achieving a full-process anti-aging effect from initial prevention to later repair. Therefore, the application of the above-mentioned extract composition in the preparation of cosmetics should be within the scope of protection of this invention.

[0056] Preferably, the cosmetic is one of cream, lotion, or solution.

[0057] The present invention has the following beneficial effects:

[0058] 1. This invention combines camellia flower ethanol extract, camellia seed shell ethanol extract and camellia fruit shell ethanol extract, which not only significantly inhibits hyaluronidase and exerts excellent anti-aging effects, but also significantly inhibits the formation of fructosamine, the initial product of protein non-enzymatic glycosylation, and AGEs, the final product, thus achieving a full-process anti-aging effect from initial prevention to later repair.

[0059] 2. This invention uses the principle of moderate efficacy to analyze the extracted composition and finds that the three components in the extracted composition play a synergistic role in anti-aging.

[0060] 3. The extract composition of this invention has high safety and acceptability, and can be used as an excellent natural anti-aging cosmetic.

[0061] 4. The extraction composition of this invention fully utilizes three camellia by-products: camellia flowers, camellia fruit shells, and camellia seed shells. This has positive significance for improving the comprehensive utilization rate of waste, reducing production costs, increasing economic benefits, and reducing environmental pollution. Attached Figure Description

[0062] Figure 1 The inhibition of AGEs by the extraction composition obtained in Example 1 is shown.

[0063] Figure 2 The inhibition rate of the extraction composition obtained in Example 1 on hyaluronidase is shown. Detailed Implementation

[0064] The present invention will be further described below with reference to the accompanying drawings and specific embodiments, but the embodiments do not limit the present invention in any way. Unless otherwise specified, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in this technical field.

[0065] Unless otherwise specified, all reagents and materials used in the following examples are commercially available.

[0066] Example 1: Preparation of the extraction composition

[0067] I. Preparation of Camellia oleifera flower alcohol extract

[0068] S1. After drying the camellia flowers at 45℃, pulverize them and pass them through a 160-mesh sieve to obtain camellia flower powder.

[0069] S2. Add camellia flower powder to n-butanol at a material-to-liquid ratio of 1g:15mL, extract by ultrasonication at 45℃ and 200W for 45min, and then centrifuge at 10000×g for 20min.

[0070] S3. Repeat S2 with the precipitate obtained by centrifugation, mix the resulting supernatant with the supernatant obtained in S2, filter, concentrate by vacuum evaporation at 45℃ and -0.1MPa, and then freeze-dry at -30℃ to obtain the camellia flower alcohol extract, which is stored in a -20℃ refrigerator.

[0071] II. Preparation of Camellia oleifera fruit shell alcohol extract

[0072] S1. After drying the camellia fruit shell at 50℃, crush it and pass it through a 160-mesh sieve to obtain camellia fruit shell powder.

[0073] S2. Add the Camellia oleifera fruit shell powder to a 70% (v / v) ethanol aqueous solution at a material-to-liquid ratio of 1g:15mL, extract by ultrasonication at 50℃ and 200W for 50min, and then centrifuge at 10000×g for 20min.

[0074] S3. Repeat S2 with the precipitate obtained by centrifugation, mix the resulting supernatant with the supernatant obtained in S2, filter, concentrate by vacuum evaporation at 45℃ and -0.1MPa, and then freeze-dry at -30℃ to obtain the camellia fruit shell alcohol extract, which is stored in a -20℃ refrigerator.

[0075] III. Preparation of Camellia oleifera seed shell alcohol extract

[0076] S1. After drying the camellia seed shells at 50℃, crush them and pass them through a 160-mesh sieve to obtain camellia seed shell powder.

[0077] S2. Add the camellia seed shell powder to a 70% (v / v) ethanol aqueous solution at a material-to-liquid ratio of 1g:30mL, extract by ultrasonication at 60℃ and 200W for 80min, and then centrifuge at 10000×g for 30min.

[0078] S3. Repeat S2 with the precipitate obtained by centrifugation, mix the resulting supernatant with the supernatant obtained in S2, filter, concentrate by vacuum evaporation at 45℃ and -0.1MPa, and then freeze-dry at -30℃ to obtain the camellia seed shell alcohol extract, which is stored in a refrigerator at -20℃.

[0079] IV. Preparation of the Extraction Composition

[0080] The extracts are prepared by mixing the camellia flower extract, the camellia fruit shell extract, and the camellia seed shell extract in a mass ratio of 1:1:1, 4:1:1, or 1:4:1, respectively.

[0081] Example 2 Preparation of the extraction composition

[0082] I. Preparation of Camellia oleifera flower alcohol extract

[0083] S1. Dry the camellia flowers at 50℃, then pulverize them and pass them through a 200-mesh sieve to obtain camellia flower powder.

[0084] S2. Add camellia flower powder to n-butanol at a material-to-liquid ratio of 1g:13mL, extract by ultrasonication at 65℃ and 220W for 40min, and then centrifuge at 9000×g for 18min.

[0085] S3. Repeat S2 with the precipitate obtained by centrifugation, mix the resulting supernatant with the supernatant obtained in S2, filter, concentrate by vacuum evaporation at 50℃ and 0MPa, and then freeze-dry at -20℃ to obtain the camellia flower alcohol extract, which is stored in a -20℃ refrigerator.

[0086] II. Preparation of Camellia oleifera fruit shell alcohol extract

[0087] S1. Dry the camellia fruit shell at 55℃, then crush it and pass it through a 200-mesh sieve to obtain camellia fruit shell powder.

[0088] S2. Add the Camellia oleifera fruit shell powder to n-butanol at a material-to-liquid ratio of 1g:13mL, extract by ultrasonication at 65℃ and 220W for 40min, and then centrifuge at 9000×g for 18min.

[0089] S3. Repeat S2 with the precipitate obtained by centrifugation, mix the resulting supernatant with the supernatant obtained in S2, filter, concentrate by vacuum evaporation at 50℃ and 0MPa, and then freeze-dry at -20℃ to obtain the camellia fruit shell alcohol extract, which is stored in a -20℃ refrigerator.

[0090] III. Preparation of Camellia oleifera seed shell alcohol extract

[0091] S1. After drying the camellia seed shells at 55℃, crush them and pass them through a 200-mesh sieve to obtain camellia seed shell powder.

[0092] S2. Add camellia seed shell powder to n-butanol at a material-to-liquid ratio of 1g:28mL, extract by ultrasonication at 65℃ and 220W for 40min, and then centrifuge at 9000×g for 28min.

[0093] S3. Repeat S2 with the precipitate obtained by centrifugation, mix the resulting supernatant with the supernatant obtained in S2, filter, concentrate by vacuum evaporation at 50℃ and 0MPa, and then freeze-dry at -20℃ to obtain the camellia seed shell alcohol extract, which is stored in a -20℃ refrigerator.

[0094] IV. Preparation of the Extraction Composition

[0095] The extracts are prepared by mixing the camellia flower extract, the camellia fruit shell extract, and the camellia seed shell extract in a mass ratio of 1:1:1, 4:1:1, or 1:4:1, respectively.

[0096] Example 3: Preparation of the extraction composition

[0097] I. Preparation of Camellia oleifera flower alcohol extract

[0098] S1. After drying the camellia flowers at 40℃, pulverize them and pass them through a 40-mesh sieve to obtain camellia flower powder.

[0099] S2. Add camellia flower powder to a 50% (v / v) ethanol aqueous solution at a material-to-liquid ratio of 1g:17mL, extract by ultrasonication at 40℃ and 180W for 85min, and then centrifuge at 11000×g for 22min.

[0100] S3. Repeat S2 with the precipitate obtained by centrifugation, mix the resulting supernatant with the supernatant obtained in S2, filter, evaporate and concentrate under reduced pressure at 40℃ and -0.1MPa, and then freeze-dry at -40℃ to obtain the camellia flower alcohol extract, which is stored in a -20℃ refrigerator.

[0101] II. Preparation of Camellia oleifera fruit shell alcohol extract

[0102] S1. After drying the camellia fruit shell at 45℃, crush it and pass it through a 40-mesh sieve to obtain camellia fruit shell powder.

[0103] S2. Add the Camellia oleifera fruit shell powder to a 50% (v / v) ethanol aqueous solution at a material-to-liquid ratio of 1g:17mL, extract by ultrasonication at 40℃ and 180W for 85min, and then centrifuge at 11000×g for 22min.

[0104] S3. Repeat S2 with the precipitate obtained by centrifugation, mix the resulting supernatant with the supernatant obtained in S2, filter, concentrate by vacuum evaporation at 40℃ and -0.1MPa, and then freeze-dry at -40℃ to obtain the camellia fruit shell alcohol extract, which is stored in a -20℃ refrigerator.

[0105] III. Preparation of Camellia oleifera seed shell alcohol extract

[0106] S1. After drying the camellia seed shells at 45℃, crush them and pass them through a 40-mesh sieve to obtain camellia seed shell powder.

[0107] S2. Add camellia seed shell powder to a 50% (v / v) ethanol aqueous solution at a material-to-liquid ratio of 1g:32mL, extract by ultrasonication at 40℃ and 180W for 85min, and then centrifuge at 11000×g for 32min.

[0108] S3. Repeat S2 with the precipitate obtained by centrifugation, mix the resulting supernatant with the supernatant obtained in S2, filter, concentrate by vacuum evaporation at 40℃ and -0.1MPa, and then freeze-dry at -40℃ to obtain the camellia seed shell alcohol extract, which is stored in a -20℃ refrigerator.

[0109] IV. Preparation of the Extraction Composition

[0110] The extracts are prepared by mixing the camellia flower extract, the camellia fruit shell extract, and the camellia seed shell extract in a mass ratio of 1:1:1, 4:1:1, or 1:4:1, respectively.

[0111] Example 4: Preparation of the extraction composition

[0112] I. Preparation of Camellia oleifera flower alcohol extract

[0113] Same as Example 1, except that S2 extraction is not performed using ultrasound, as detailed below:

[0114] S1. After drying the camellia flowers at 45℃, pulverize them and pass them through a 160-mesh sieve to obtain camellia flower powder.

[0115] S2. Add camellia flower powder to n-butanol at a material-to-liquid ratio of 1g:15mL, extract at 45℃ for 45min, and then centrifuge at 10000×g for 20min.

[0116] S3. Repeat S2 with the precipitate obtained by centrifugation, mix the resulting supernatant with the supernatant obtained in S2, filter, concentrate by vacuum evaporation at 45℃ and -0.1MPa, and then freeze-dry at -30℃ to obtain the camellia flower alcohol extract, which is stored in a -20℃ refrigerator.

[0117] II. Preparation of Camellia oleifera fruit shell alcohol extract

[0118] Same as Example 1, except that S2 extraction is not performed using ultrasound, as detailed below:

[0119] S1. After drying the camellia fruit shell at 50℃, crush it and pass it through a 160-mesh sieve to obtain camellia fruit shell powder.

[0120] S2. Add the Camellia oleifera fruit shell powder to a 70% (v / v) ethanol aqueous solution at a material-to-liquid ratio of 1g:15mL, extract at 50℃ for 50min, and then centrifuge at 10000×g for 20min.

[0121] S3. Repeat S2 with the precipitate obtained by centrifugation, mix the resulting supernatant with the supernatant obtained in S2, filter, concentrate by vacuum evaporation at 45℃ and -0.1MPa, and then freeze-dry at -30℃ to obtain the camellia fruit shell alcohol extract, which is stored in a -20℃ refrigerator.

[0122] III. Preparation of Camellia oleifera seed shell alcohol extract

[0123] Same as Example 1, except that S2 extraction is not performed using ultrasound, as detailed below:

[0124] S1. After drying the camellia seed shells at 50℃, crush them and pass them through a 160-mesh sieve to obtain camellia seed shell powder.

[0125] S2. Add the camellia seed shell powder to a 70% (v / v) ethanol aqueous solution at a material-to-liquid ratio of 1g:30mL, extract at 60℃ for 80min, and then centrifuge at 10000×g for 30min.

[0126] S3. Repeat S2 with the precipitate obtained by centrifugation, mix the resulting supernatant with the supernatant obtained in S2, filter, concentrate by vacuum evaporation at 45℃ and -0.1MPa, and then freeze-dry at -30℃ to obtain the camellia seed shell alcohol extract, which is stored in a refrigerator at -20℃.

[0127] IV. Preparation of the Extraction Composition

[0128] The extracts are prepared by mixing the camellia flower extract, the camellia fruit shell extract, and the camellia seed shell extract in a mass ratio of 1:1:1, 4:1:1, or 1:4:1, respectively.

[0129] Test Example 1 Security Evaluation

[0130] The camellia flower ethanol extract, camellia fruit shell ethanol extract, and camellia seed shell ethanol extract obtained in Example 1 were dissolved in water to obtain final concentrations of 0.01, 0.03, and 0.05 mg / mL, respectively, which were then used as test solutions.

[0131] Human skin fibroblasts were cultured in DMEM medium (containing 200 kU / L penicillin sodium, 100 mg / L streptomycin, and 0.1 mol / L PBS, pH 7.4) with 15% fetal bovine serum. Cells in the logarithmic growth phase (P5-P10 passages) were used for subsequent experiments. The cells were resuspended in the above medium and the concentration was adjusted to 1 × 10⁻⁶. 5Cells / mL. Add 100 μL of cell suspension to a 96-well plate and incubate at 37°C in a CO2 incubator for 24 h. Then, add 100 μL of test solution or water to each well (test solution for the treatment group, water for the blank group) and continue incubation for 48 h. After incubation, carefully aspirate the supernatant and add 110 μL of a mixture of CCK-8 reagent and serum-free DMEM medium (mixing ratio 1:10) to each well. After incubation at 37°C in the dark for 1 h, measure the absorbance (OD value) of each well at 450 nm using a microplate reader. Each experiment has 3 replicates. Finally, calculate the cell viability using the formula: "Cell viability (%) = OD". 处理 / OD 空白 Cell viability was calculated by multiplying the result by 100% to evaluate the safety of the test solution. The results are shown in Table 1.

[0132] Table 1

[0133]

[0134] It is evident that the cell survival rate of the treated groups all reached over 88%, indicating that the camellia flower ethanol extract, camellia fruit shell ethanol extract, and camellia seed shell ethanol extract all have high safety, demonstrating that the extraction composition of the present invention has high safety.

[0135] In addition, the same method was used to test the camellia flower alcohol extract, camellia fruit shell alcohol extract, and camellia seed shell alcohol extract obtained in Examples 2-4. The results were basically consistent with those in Example 1, indicating that the extraction composition of the present invention has high safety.

[0136] Test Example 2: Anti-aging performance test of the extracted composition

[0137] I. Experimental Grouping

[0138] Take 3g each of the camellia flower ethanol extract, camellia fruit shell ethanol extract, camellia seed shell ethanol extract, and the three extraction compositions in the specified mass ratios obtained in Example 1, and then add PBS solution (pH 7.4) to make up to 10mg. These are groups ① to ⑥, and are used as sample solutions for each group. See Table 2 for details.

[0139] Table 2

[0140]

[0141]

[0142] II. Inhibition of fructosamine by the extraction composition

[0143] The ability of the extract composition to inhibit non-enzymatic glycosylation of proteins was evaluated by measuring its inhibitory efficiency on the formation of fructosamine, thereby assessing its anti-aging efficacy.

[0144] The sample solutions from groups ① to ⑥ were diluted with PBS solution (pH 7.4, 0.1 mol / L) to final concentrations of 0.01, 0.02, 0.03, 0.04, and 0.05 mg / mL, respectively, to obtain the test solutions. BSA and fructose were added to the test solutions of different concentrations (to achieve concentrations of 20 mg / mL and 90.1 mg / mL, respectively), and then incubated in a 50℃ water bath for 1 h. 20 μL of the incubation product and 180 μL of NBT reagent (0.5 mmol / L NBT dissolved in carbonate solution, with pH adjusted to 10.8 using NaOH and HCl solutions; the carbonate solution contained sodium carbonate and sodium bicarbonate in a 1:9 mass ratio, maintaining a carbonate ion concentration of 0.1 mol / L) were added to a microplate and incubated at 37℃ for 10 min. The absorbance A was read at 530 nm. 样品 .

[0145] The group without added sample solution was used as the blank group. After processing according to the above procedure, the absorbance A was read at 530 nm. 空白 .

[0146] According to the formula "fructosamine inhibition rate (%) = (A 空白 -A 样品 ) / A 空白 The inhibition rate of fructosamine was calculated by multiplying the result by 100%, and then the IC50 concentration at which fructosamine achieved half-inhibition was determined for groups ① to ⑥. 50 .

[0147] The combined effect index of compounds ④, ⑤, and ⑥ was calculated using the intermediate-efficiency principle. The calculation formula is CI (Combination Index). 50 = D×(1 / DX1+1 / DX2+1 / DX3), where D represents the IC of ④, ⑤, and ⑥. 50 DX1, DX2, and DX3 represent ICs ①, ②, and ③, respectively. 50 0 <CI 50 ≤0.7 indicates a significant synergistic enhancement of the effects of the three factors; 0.7 <CI 50 <1 indicates a synergistic enhancement of the three effects; CI 50 =1 indicates that the effects of the three elements are additive and enhanced; 1 <CI 50 <10 indicates an antagonistic relationship.

[0148] The results are as follows Figure 1 As shown, CI ① to ⑥ can be seen. 50 The CI values ​​are 0.02453, 0.03373, 0.04740, 0.00826, 0.00756, and 0.00930, respectively, which are the CI values ​​for ④, ⑤, and ⑥. 50The values ​​were 0.76, 0.69, and 0.85, respectively. This indicates that in the extraction composition of the present invention, the camellia flower ethanol extract, the camellia fruit shell ethanol extract, and the camellia seed shell ethanol extract play at least a synergistic enhancing role in inhibiting fructosamine, the initial product of non-enzymatic glycosylation of proteins, especially when the mass ratio of the camellia flower ethanol extract, the camellia fruit shell ethanol extract, and the camellia seed shell ethanol extract is 4:1:1, it plays a significant synergistic enhancing role.

[0149] In addition, the same method was used to test the extraction compositions obtained in Examples 2 to 4 in this test example. The results were basically consistent with those in Example 1, indicating that the extracts of Camellia oleifera flower, Camellia oleifera fruit shell, and Camellia oleifera seed shell played at least a synergistic role in inhibiting fructosamine, the initial product of non-enzymatic glycosylation of proteins, in the extraction compositions of the present invention.

[0150] III. Inhibition of AGEs by the Extraction Composition

[0151] The accumulation of advanced non-enzymatic glycation end products (AGEs) of proteins can lead to yellowing, inflammation, and roughness of the skin. The anti-aging efficacy of the extracted composition can be evaluated by measuring its ability to inhibit AGEs.

[0152] Each sample solution from groups ① to ⑥ was diluted with PBS solution (pH 7.4, 0.1 mol / L) to a final concentration of 0.1 mg / mL to obtain the test solutions. BSA and fructose were added to the test solutions of different concentrations (to achieve concentrations of 20 mg / mL for BSA and 90.1 mg / mL for fructose), and then incubated in a 50°C water bath for 24 hours to obtain the sample groups. The groups without added sample solutions served as the blank groups.

[0153] After incubation, fluorescence spectra of each incubation product were collected at wavelengths of 400–600 nm using a fluorescence spectrometer (with emission and excitation slit widths of 5 nm) under excitation light at 25 °C and 370 nm. The fluorescence intensity at λex / λem = 370 / 450 nm was then measured to analyze the formation of fluorescent AGEs.

[0154] The results are as follows Figure 1 As shown, the fluorescence intensity of groups ④ to ⑥ is significantly lower than that of the blank group and groups ① to ③, indicating that the extraction composition of the present invention can significantly inhibit the generation of AGEs (advanced glycation end products) of proteins, thereby exerting excellent anti-aging effects.

[0155] Based on the aforementioned results of inhibiting fructosamine, it can be seen that the extraction composition of the present invention achieves a full-process anti-aging effect from initial prevention to later repair.

[0156] In addition, the same method was used to test the extract compositions obtained in Examples 2 to 4 in this test example. The results were basically consistent with those in Example 1, indicating that the extract compositions of the present invention can significantly inhibit the formation of protein non-enzymatic glycosylation advanced products (AGEs), thereby exerting excellent anti-aging effects.

[0157] IV. Inhibition of Hyaluronidase by the Extraction Composition

[0158] Hyaluronic acid is a component in the tissue matrix that restricts the diffusion of water and other extracellular substances. When hydrolyzed by hyaluronidase, it reduces the extracellular matrix, decreases intercellular adhesion, causes cell degranulation and leakage of newly synthesized mediators, exerting biological effects that lead to loss of skin elasticity, thinning, and accelerated skin aging. The anti-aging efficacy of this extracted composition was evaluated by measuring its inhibitory ability on hyaluronidase.

[0159] Group A: 0.1 mL Mix CaCl2 solution (0.25 mmol / L) and 0.5 mL hyaluronidase solution (100 U / mL, in a pH 5.6 acetate buffer solution), incubate at 37 °C for 20 min, then add 0.5 mL of sample solution, incubate for 15 min, then add 0.5 mL of sodium hyaluronate solution (0.5 g / L), react for 30 min, and cool for 5 min. Add 0.1 mL of sodium hydroxide solution (0.4 mol / L) and 0.5 mL of acetylacetone solution (3.5 mL of acetylacetone dissolved in 50 mL of 1.0 mol / L sodium carbonate solution), boil in a water bath for 15 min, then immediately transfer to an ice-water bath for 5 min. Add 1 mL of Ehrlich reagent (0.8 g of p-dimethylaminobenzaldehyde dissolved in a mixture of 15 mL of concentrated hydrochloric acid and 15 mL of anhydrous ethanol), dilute with 3 mL of anhydrous ethanol, incubate at 25 °C for 20 min for color development, and measure the absorbance A at 540 nm.

[0160] Group B: Same as Group A, except that the hyaluronidase solution is replaced with an acetate buffer solution (pH 5.6).

[0161] Group C: Same as Group A, except that the sample solution is replaced with an acetate buffer solution (pH 5.6).

[0162] Group D: Same as Group A, except that both the sample solution and the hyaluronidase solution are replaced with acetate buffer solution (pH 5.6).

[0163] Finally, the hyaluronidase inhibition rate was calculated according to the formula "hyaluronidase inhibition rate (%) = [(CD) - (AB)] / (CD) × 100%", and then the inhibitory ability of the sample solution on hyaluronidase was evaluated.

[0164] The results are as follows Figure 2As shown, the hyaluronidase inhibition rate of groups ④ to ⑥ is significantly higher than that of groups ① to ③, indicating that the extraction composition of the present invention can significantly inhibit hyaluronidase, thereby exerting excellent anti-aging effects.

[0165] In addition, the same method was used to test the extract compositions obtained in Examples 2 to 4 in this test example. The results were basically consistent with those in Example 1, indicating that the extract compositions of the present invention can significantly inhibit hyaluronidase and thus exert excellent anti-aging effects.

[0166] The above embodiments are preferred embodiments of the present invention, but the embodiments of the present invention are not limited to the above embodiments. Any changes, modifications, substitutions, combinations, or simplifications made without departing from the spirit and principle of the present invention shall be considered equivalent substitutions and shall be included within the protection scope of the present invention.

Claims

1. An extraction composition, characterized in that, It contains camellia flower alcohol extract, camellia fruit shell alcohol extract and camellia seed shell alcohol extract in a mass ratio of 1 to 4:1 to 4:

1.

2. The extraction composition according to claim 1, characterized in that, The camellia flower alcohol extract is obtained by extracting camellia flowers with n-butanol or an aqueous ethanol solution.

3. The extraction composition according to claim 2, characterized in that, The extraction was performed at 40–65°C for 40–85 min.

4. The extraction composition according to claim 1, characterized in that, The camellia oleifera fruit shell alcohol extract is obtained by extracting camellia oleifera fruit shells with n-butanol or an aqueous ethanol solution.

5. The extraction composition according to claim 4, characterized in that, The extraction was performed at 40–65°C for 40–85 min.

6. The extraction composition according to claim 1, characterized in that, The camellia seed shell alcohol extract is obtained by extracting camellia seed shells with n-butanol or an aqueous ethanol solution.

7. The extraction composition according to claim 6, characterized in that, The extraction was performed at 40–65°C for 40–85 min.

8. A method for preparing the extraction composition according to any one of claims 1 to 7, characterized in that, The extract is obtained by mixing the ethanol extracts of camellia flowers, camellia fruit shells, and camellia seed shells.

9. The use of the extraction composition according to any one of claims 1 to 7 in the preparation of cosmetics.

10. The application according to claim 9, characterized in that, The cosmetic product is one of the following: cream, lotion, or solution.

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