Preparation method of anti-photoaging camellia extract as well as product and application of anti-photoaging camellia extract

Abstract

The invention relates to the technical field of cosmetic raw materials, in particular to a preparation method of an anti-photoaging camellia extract as well as a product and application of the anti-photoaging camellia extract. The preparation method of the camellia extract comprises the following steps: S1, pretreating camellia; s2, activating the biological enzyme I; s3, carrying out primary enzymolysis; s4, extracting; s5, filtering is carried out; s6, carrying out secondary enzymolysis; and S7, purifying to obtain the camellia extract. The invention aims to make full use of camellia resources, avoid waste of a large amount of camellia as waste, and develop a camellia extract capable of resisting ultraviolet damage and maintaining skin steady state by excavating the synergistic effect between polysaccharide and protein in the camellia, so as to realize more comprehensive and deep development and utilization of the camellia resources.

Description

Technical Field

[0001] The invention relates to the technical field of cosmetic raw materials, and in particular to a preparation method, product and application of an anti-photoaging camellia extract. Background Art

[0002] Skin is the largest organ in the human body, and its aging status directly reflects the degree of aging of the body. Skin aging is mainly divided into two categories: endogenous aging and exogenous aging. Endogenous aging, also known as natural aging, is mainly affected by genetic factors and physical constitution; while exogenous aging, also known as photoaging, mainly refers to aging caused by environmental factors such as ultraviolet radiation, smoking, wind, sun exposure and contact with harmful chemicals. Studies have shown that photoaging accounts for more than 80% of facial aging, among which ultraviolet radiation is the most critical exogenous factor causing skin photoaging. Long-term exposure to ultraviolet rays can cause a series of problems such as abnormal pigmentation, deepening and thickening of wrinkles, dry skin, sagging, atrophy and capillary dilation. The mechanism of skin photoaging mainly involves interference with the immune system, changes in cytokines, increased production of matrix metalloproteinases, reduced mature collagen fibers, increased reactive oxygen species, damage to the extracellular matrix structure, DNA damage and telomere damage. Therefore, how to prevent skin photooxidation or slow down the damage caused by ultraviolet rays to the skin, as well as delay skin aging, has always been highly concerned by people. Traditional physical or chemical sunscreens often have problems such as a heavy feel on the skin, poor safety, and limited protection efficiency, and therefore cannot fully meet the needs of cosmetic applications.

[0003] Existing studies have found that many natural extracts or natural products from plants have certain sun protection and UV radiation damage mitigation effects. They usually achieve the effect of UV protection and photoaging mitigation in the following ways: highly unsaturated structures with benzene rings or conjugated systems that can absorb UV rays; resist the photooxidation process of the skin; regulate the synthesis and decomposition of the extracellular matrix; inhibit the deposition of pigments; relieve skin inflammation; regulate cell autophagy and other mechanisms. For example, curcumin, the main active ingredient of turmeric, can antagonize DNA damage induced by UVR irradiation; silymarin can induce apoptosis of photodamaged cells and affect the cell cycle to facilitate the repair of damaged DNA, so it has UVB protection. Green tea polyphenols, resveratrol, tannic acid and other polyphenolic compounds can significantly reduce intracellular ROS levels and slow down skin aging. Plant-derived polysaccharides, due to their antioxidant properties, show photoaging treatment effects. For example, Dendrobium officinale polysaccharides can upregulate the TGF-β / Smad signaling pathway in fibroblasts, thereby reducing oxidative damage caused by UV irradiation. Green tea seed extract can reduce the expression of MMP-1, MMP-3, and MMP-9 induced by UVB, and increase the production of type I collagen precursors; caffeine can protect the skin from aging induced by oxidative stress by regulating the A2AR / SIRT3 / AMPK pathway to activate cell autophagy. Such natural ingredients are gradually becoming the focus of the cosmetics industry in the field of anti-photoaging and photodamage research due to their higher safety and easier skin absorption.

[0004] Camellia sinensis (L.) O. Kuntze is the flower of the tea tree, a plant of the Theaceae family, the genus Camellia, and the tea group. It is one of the reproductive organs of the tea tree and is known as the "placenta of safe plants" and the "essence of the tea tree". Camellia contains a variety of beneficial ingredients and active substances such as tea polyphenols, tea polysaccharides, proteins, flavonoids, tea saponins, caffeine, amino acids, vitamins, trace elements, superoxide dismutase (SOD) and catalase (CAT), and has certain lipid-lowering and weight-loss, radiation protection, moisturizing, anti-inflammatory and antibacterial, and surfactant effects. Among them, the amount of catechins contained in camellia is similar to that in tea leaves, while the content of polysaccharides and proteins is significantly higher than the average level of tea leaves. These ingredients all show that camellia is a type of plant raw material with high biological activity and great application potential.

[0005] The life span of camellia is usually two days. Once it blooms, if it fails to complete pollination within two days, it will fall off naturally. It has the characteristics of "long flowering period, many flowers, and few fruits". For a long time, people mainly picked fresh buds and leaves of tea trees for tea making, while the flowers were mostly left to wither naturally. Some tea farmers prune branches and apply plant growth regulators to remove flowers and fruits (flowers and fruits need to absorb more than 50% of photosynthetic products of tea trees during growth) to achieve high yields in tea gardens. According to statistics, tea areas across the country can produce 4-8 million tons of fresh flowers each year, and camellia resources are quite rich. It is undoubtedly a huge waste of tea resources to only use fresh tea leaves and abandon camellia. By picking camellia manually, it can ensure that the nutrients of tea trees are more supplied to the growth of tea leaves and buds, and the yield and quality of tea trees will increase by 25%-30% the following year, and bring additional economic benefits to tea farmers.

[0006] In 2013, camellia (tea tree flower) was approved by the National Health Commission as a new food ingredient. However, the current application scope of camellia is relatively narrow, mainly including camellia tea, camellia beverages, camellia wine, camellia essential oil, camellia powder, etc., and these products are mainly used in the fields of food and health care. In the cosmetics industry, the application of camellia extract is still in its initial stage, for example, it is used as an additive for soap and hand sanitizer, but its unique skin care effects such as antioxidant, moisturizing and antibacterial have not been fully and comprehensively developed and utilized. Therefore, the potential application value of camellia extract in the field of cosmetics still needs further in-depth exploration and research.

[0007] Chinese patent CN102030834B discloses a method for extracting and preparing camellia polysaccharides from camellia and the use of the obtained camellia polysaccharides, using enzyme preparations for extraction, separating after alcohol precipitation of the extract, and drying the separated precipitate to obtain crude camellia polysaccharides, and decolorizing and deproteinizing the crude camellia polysaccharides by resin or chemical methods, and then purifying them by gel purification or ultrafiltration membrane purification to obtain camellia polysaccharides. This technology has been proven through animal experiments that camellia polysaccharides have the effect of preventing and treating diabetes. Chinese patent application CN105707230A discloses a camellia extract and its extraction method and use. This technology first uses ethanol aqueous solution for extraction, recovers ethanol from the extract, and then uses petroleum ether and ethyl acetate for extraction, and recovers the solvent to obtain a camellia extract. The "camellia" flower extract provided by it has significant antibacterial activity and high safety, and can be used to extend the shelf life of milk by 5-7 days. However, none of these prior arts solve the above technical problems, and the application of camellia extract in the field of skin care products is studied.

[0008] Under such circumstances, there is an urgent need to provide a technology that can effectively develop camellia resources, fully tap its skin care effects, and develop its potential value in the cosmetics field. Summary of the invention

[0009] The present invention aims to make full use of camellia resources that might have been regarded as waste, and to develop a camellia extract that can both resist UV damage and maintain skin homeostasis by exploring the synergistic effect between polysaccharides and proteins in camellia, in order to achieve more comprehensive and in-depth development and utilization of camellia resources.

[0010] The first aspect of the present invention provides a method for preparing an anti-photoaging camellia extract, the preparation steps comprising:

[0011] S1. Pre-treating camellia to obtain camellia powder;

[0012] S2. activating the biological enzyme 1 to obtain an activated enzyme solution;

[0013] S3. Add camellia powder to the activated enzyme solution and perform enzymatic hydrolysis to obtain enzymatic solution 1;

[0014] S4. extracting the enzymatic hydrolyzate 1 to obtain an extract;

[0015] S5. The extract is filtered to obtain liquid 1;

[0016] S6. Mixing the feed solution 1 and the biological enzyme 2, performing secondary enzymolysis, and obtaining enzymolysis solution 2;

[0017] S7. Purify the enzymatic hydrolyzate 2 to obtain the feed solution 2, which is the camellia extract.

[0018] Plant cell walls are mainly composed of cellulose, hemicellulose and other components, and contain pectin and starch between cells. Generally, most solvents have limited swelling capacity for plant cell walls, especially in the extraction process of camellia, where it is difficult to break the cell wall during pollen extraction, which limits the release of active ingredients in cell tissues to a certain extent. The present invention extracts the biological components in camellia to the maximum extent through the action of biological enzymes, so as to give full play to the skin care effect of camellia and realize the full utilization and in-depth development of camellia resources.

[0019] In some preferred embodiments, the step S1 specifically includes: picking fresh camellia flowers, sorting and removing calyx, receptacle and impurities, low-temperature drying, crushing and sieving to obtain camellia fine powder.

[0020] Preferably, the temperature of the low-temperature drying is ≤45° C. The drying method includes at least one of vacuum drying, freeze drying, and oven drying; more preferably, the drying method includes freeze drying or oven drying.

[0021] Preferably, the mesh size of the sieve is 20-200 meshes, more preferably 40-80 meshes.

[0022] In some preferred embodiments, the step S2 specifically comprises: dissolving the biological enzyme 1 in deionized water, activating the enzyme at 30-40° C. for 0.5-2 h, and obtaining an activated enzyme solution.

[0023] Preferably, the biological enzyme 1 is a biological composite enzyme, including at least three of cellulase, pectinase, α-amylase, xylanase, tannase, glucanase, mannanase and glucoamylase.

[0024] More preferably, the biocomplex enzyme includes cellulase, pectinase, α-amylase, xylanase and tannase in a mass ratio of (0.1-3):(0.1-2):(0.1-4):(0.15-2):(0.05-1.5).

[0025] More preferably, the α-amylase is a mesophilic α-amylase.

[0026] Preferably, based on the total mass of camellia powder, the biocomplex enzyme is added according to 0.2%-1.0% cellulase, 0.2%-1.0% pectinase, 0.2%-1.0% α-amylase, 0.2%-1.0% xylanase, and 0.1%-0.5% tannase.

[0027] Preferably, the cellulase activity is (2000-5000) U / g, the pectinase activity is (40000-80000) U / g, the α-amylase activity is (1000-3000) U / g, the xylanase activity is (60000-100000) U / g, and the tannin activity is (100-500) U / g.

[0028] More preferably, the cellulase activity is 3500 U / g, the pectinase activity is 60000 U / g, the medium-temperature α-amylase activity is 1000 U / g, the xylanase activity is 80000 U / g, and the tanninase activity is 300 U / g.

[0029] Preferably, the amount of deionized water added in step S2 is 5-15 times the mass of camellia fine powder.

[0030] Camellia contains active ingredients such as xylan, pectin, tea polyphenols, amino acids, flavonoids, alkaloids, etc. Among them, xylan, as a heterogeneous polysaccharide, is a key component of plant hemicellulose; and pectin is an important substance that constitutes the plant cell wall, and also plays a role in connecting adjacent cell walls. The present invention selects cellulase, pectinase, mesophilic α-amylase, xylanase and tannase as biological composite enzymes, which can efficiently degrade the cell wall of camellia, destroy the cell structure of camellia, and expose and dissolve the effective ingredients from the cells after the cells are broken. The present invention uses biological composite enzyme reaction technology to prepare camellia extract, which can improve the extraction efficiency of active ingredients and maintain their stability as much as possible.

[0031] Furthermore, tea polyphenols, soluble proteins, polysaccharides, caffeine and metal ions contained in camellia are easy to interact with each other to form macromolecular complexes. Among them, ester catechins containing more hydroxyl groups are more likely to combine with other ingredients to form complexes, causing the extract to produce turbid precipitation, which has an adverse effect on the stability of the product and limits its addition in skin care products. The present invention selects cellulase, pectinase, α-amylase, xylanase and tannase in a mass ratio of (0.1-3): (0.1-2): (0.1-4): (0.15-2): (0.05-1.5), which can effectively hydrolyze the corresponding complexes and inhibit the formation of precipitation. It is speculated that the specific amount of biological composite enzyme added in the present invention can catalyze the hydrolysis of gallate catechins to the greatest extent to generate non-ester catechins and gallic acid; gallic acid combines with caffeine to form small molecule water-soluble substances, thereby effectively reducing the formation of precipitates. In addition, the production of gallic acid also enhances the antioxidant properties of camellia extract to a certain extent and improves the comprehensive performance of camellia extract.

[0032] In some preferred embodiments, the step S3 specifically comprises: adding camellia powder to an activated enzyme solution, adjusting the pH to 4-6, and performing enzymolysis at 45-55° C. and 100-200 rpm for 1-3 hours to obtain an enzymolysis solution 1.

[0033] The present invention adopts water as a solvent, avoids the introduction of organic reagents, reduces environmental pollution and potential health risks. Before the raw materials are pulverized, they are pretreated by low-temperature drying, which can effectively retain the active ingredients in the raw materials and ensure the quality and effect of the final product. The use of oven drying or freeze drying methods can not only help protect heat-sensitive active ingredients, but also reduce energy consumption and improve production efficiency. Through the implementation of the present invention, the application of camellia in the cosmetics industry is further explored, and it is expected to expand the application scope of camellia resources.

[0034] In some preferred embodiments, the step S4 specifically comprises: heating the enzymatic hydrolyzate 1 to 85-95° C., stirring and extracting at 100-200 rpm for 1-3 hours to obtain an extract.

[0035] In some preferred embodiments, the step S5 specifically comprises: filtering the extract obtained in step S4 multiple times to obtain feed solution 1.

[0036] In some preferred embodiments, the step S6 specifically comprises: adjusting the pH of the feed solution to 3-5, adding biological enzyme 2, performing secondary enzymolysis at 45-55° C. and 100-200 rpm for 0.5-2 h, and inactivating the enzyme at 85-92° C. to obtain enzymolysis solution 2;

[0037] Preferably, the biological enzyme 2 includes at least one of protease, amylase, lipase and cellulase; more preferably, it is protease; and further preferably, it is acidic protease.

[0038] Preferably, the added amount of the biological enzyme II is 0.1-3% of the total mass of the camellia powder, and more preferably 0.2-1%.

[0039] In some preferred embodiments, the step S7 specifically includes: adding a clarifier to the enzymatic hydrolyzate 2, standing in a water bath at 50-60° C. for 20-60 min, and then filtering to obtain a feed solution 2, which is a finished camellia extract.

[0040] Preferably, the clarifier includes a combination of one or more of chitosan, ZTC (natural clarifier) ​​type II, ZTC type III, and ZTC 101.

[0041] Preferably, the amount of the clarifier added is 1-5% of the total mass of the liquid 2.

[0042] The filtering means of the present invention may include gauze filtration, porous material filtration, membrane filtration and the like.

[0043] Preferably, gauze filtration, porous material filtration and membrane filtration are used in step S5.

[0044] Preferably, porous material filtration and membrane filtration are used in step S7.

[0045] Preferably, the gauze filtration is 100-400 mesh gauze filtration.

[0046] Preferably, the porous material comprises one or more of diatomaceous earth, activated carbon, zeolite, porous ceramics, and molecular sieves; diatomaceous earth is more preferred.

[0047] Preferably, the particle size of the porous material is 200-400 mesh.

[0048] The membrane filtration may be at least one of polypropylene (PP) membrane, polyethersulfone (PES) membrane, polyvinylidene fluoride (PVDF) membrane, polytetrafluoroethylene (PTFE) membrane, nylon membrane, cellulose acetate membrane (CA), and MCE filter membrane. Preferably, the membrane filtration is polypropylene membrane filtration.

[0049] The second aspect of the present invention provides a camellia extract prepared by the preparation method as described above, wherein the camellia extract contains camellia saccharides and camellia plant peptides.

[0050] Preferably, the total sugar content in the camellia extract is ≥8.0 mg / mL; the total peptide content in the camellia extract is ≥7.0 mg / mL.

[0051] Preferably, the camellia sugars include monosaccharides; the monosaccharides include at least two of glucose, galactose, arabinose, mannose, galacturonic acid, xylose or fructose.

[0052] More preferably, the monosaccharides include glucose, galactose, arabinose, mannose, galacturonic acid and xylose in a mass ratio of (50-80):(5-35):(1-15):(1-10):(0.1-5):(0.1-3).

[0053] Preferably, the weight average molecular weight Mw of the camellia saccharide is 5×10 3 -5×10 4 .

[0054] Preferably, peptide sequence analysis of the camellia extract shows that the number of camellia plant peptides is ≥100.

[0055] Preferably, the camellia plant peptide consists of 2 to 30 amino acids.

[0056] The third aspect of the present invention provides a use of the camellia extract as described above, wherein the camellia extract is used in a skin care composition; the amount of the camellia extract added to the skin care composition is 10-30 wt %.

[0057] In addition to the camellia extract, the skin care composition may also contain functional components such as moisturizers, skin conditioners, thickeners, preservatives, etc. according to the needs of users.

[0058] In some preferred embodiments, the skin care composition comprises, by weight percentage, 20-30% camellia extract, 10%-40% glycerin, 0.1-2.0% 1,2-hexanediol, 0.1%-2.0% p-hydroxyacetophenone, and the balance is water.

[0059] The preparation method of the skin care composition is not particularly limited, for example, the raw materials are mixed and filtered to obtain the skin care composition.

[0060] The camellia extract prepared by the present invention is rich in carbohydrates, protein polypeptides and tea polyphenols. Through the synergistic action of multiple pathways, it can effectively resist the damage caused by ultraviolet rays and maintain the stable state of the skin. After experimental verification, the camellia extract of the present invention has the following effects: ① It has excellent DPPH free radical scavenging ability; ② It can significantly reduce the increase in ROS levels in HaCaT cells caused by ultraviolet irradiation; ③ It can increase the content of C-terminal peptide of type I procollagen and promote type I collagen synthesis; ④ It can reduce the expression of MMP-1 in HSF cells after UVB irradiation and inhibit abnormal extracellular matrix degradation process; ⑤ In clinical applications, after continuous use for 4 weeks, it has the effect of significantly improving the elasticity of facial skin. It can be seen that the implementation of the present invention has achieved in-depth exploration of camellia and excavation of its potential value, and has positive research significance.

[0061] Beneficial effects:

[0062] The present invention provides a camellia extract for resisting photoaging and a preparation method thereof, which has the following advantages:

[0063] (1) The present invention selects cellulase, pectinase, mesophilic α-amylase, xylanase and tannase as biological composite enzymes, which can efficiently degrade the cell wall of camellia, so that the effective ingredients can be exposed and dissolved from the cells, thereby significantly improving the extraction efficiency of the active ingredients and the stability of the camellia extract.

[0064] (2) The present invention optimizes the mass ratio and addition amount of the biological composite enzyme, which can hydrolyze the macromolecular complex generated during the camellia extraction process to the greatest extent and inhibit the formation of precipitation during the extraction process. At the same time, it also enhances the antioxidant properties of the camellia extract to a certain extent and improves the comprehensive properties of the camellia extract.

[0065] (3) In the preferred extraction temperature of step S4 of the present invention, 85-95°C is used, so that the active ingredients in the camellia are fully dissolved under the coordinated action of biological enzyme 1 and biological enzyme 2, and the total sugar content in the final camellia extract is ≥8.0 mg / mL; the total peptide content in the camellia extract is ≥7.0 mg / mL, and the types of camellia plant peptides are ≥100.

[0066] (4) The extraction method of the present invention uses water as a solvent, avoiding the introduction of organic reagents, reducing environmental pollution and potential health risks. Through the implementation of the present invention, a camellia extract with significant antioxidant, stabilizing and anti-aging effects can be obtained, providing a feasible solution for further exploring the application of camellia in the cosmetics industry.

[0067] (5) The extraction method of the present invention has the advantages of being harmless to health, having no environmental impact, rapid extraction, and efficient output. The extraction process is simple and easy to implement, and can meet the requirements of large-scale batch production.

[0068] (6) The extraction method of the present invention provides a new idea for the application expansion of camellia resources in the field of cosmetics, which is not only conducive to improving the economic value of camellia, but also can promote the development of related industries and create more economic and ecological benefits for society. BRIEF DESCRIPTION OF THE DRAWINGS

[0069] Figure 1 .Glucose standard curve;

[0070] Figure 2 .HaCaT cell ROS fluorescence intensity test results;

[0071] Figure 3 .Test results of type I procollagen C-terminal peptide content in HSF cells;

[0072] Figure 4 .Test results of MMP-1 secretion in HSF cells induced by UVB;

[0073] Figure 5 .Skin elasticity change test results of samples in Example 1 and Comparative Example 2. DETAILED DESCRIPTION

[0074] Note: The fresh camellia used in the examples of the present invention comes from Menghai County, Xishuangbanna Dai Autonomous Prefecture, Yunnan Province; the polypropylene membrane comes from Haining YiBo Filter Equipment Factory, with a pore size of 0.22 μm (φ100 mm); the clarifier ZTCⅡ comes from Beijing Zhengtiancheng Clarification Technology Co., Ltd.; the concentration of concentrated sulfuric acid involved in the test method is 98%.

[0075] Furthermore, unless otherwise specified, the solvents of the solutions involved in the present invention are all water; and the raw materials used are all commercially available.

[0076] Example

[0077] Example 1

[0078] The first aspect of this embodiment provides a method for preparing an anti-photoaging camellia extract, the preparation steps comprising:

[0079] S1. Pre-processing camellia: picking fresh camellia, sorting to remove calyx, receptacle and impurities, drying in a 40°C oven, crushing through a 60-mesh sieve to obtain camellia fine powder.

[0080] S2. Activating the biological enzyme 1: dissolving the biological enzyme 1 in deionized water 10 times the mass of the camellia powder, and activating the enzyme at 35°C for 1 hour to obtain an activated enzyme solution;

[0081] The biological enzyme 1 is a biological composite enzyme, which includes cellulase, pectinase, medium-temperature α-amylase, xylanase and tannase; based on the total mass of camellia powder, the biological composite enzyme is added according to 0.5% of cellulase, 0.5% of pectinase, 0.5% of medium-temperature α-amylase, 0.5% of xylanase and 0.2% of tannase.

[0082] The cellulase activity is 3500U / g, the pectinase activity is 60000U / g, the medium-temperature α-amylase activity is 1000U / g, the xylanase activity is 80000U / g, and the tannin activity is 300U / g; all of them come from Cangzhou Xiasheng Enzyme Biotechnology Co., Ltd.

[0083] S3. Primary enzymatic hydrolysis: Add camellia powder to the activated enzyme solution, adjust the pH to 4.5 with 19% citric acid aqueous solution, and perform enzymatic hydrolysis at 52° C. and 150 rpm for 2 h to obtain enzymatic hydrolyzate 1.

[0084] S4. Extraction: Heat the enzymatic hydrolyzate 1 to 92°C, stir at 150 rpm for 2 h, and obtain an extract.

[0085] S5. Filtration: The extract obtained in step S4 is filtered through 200 mesh gauze, 300 mesh diatomaceous earth and 5 μm polypropylene membrane in sequence to obtain feed solution 1. In this step, 300 mesh diatomaceous earth and 5 μm polypropylene membrane are used simultaneously, and the polypropylene membrane is placed at the bottom and the diatomaceous earth is placed on top of the polypropylene membrane for filtration.

[0086] S6. Secondary enzymatic hydrolysis: Use 19% citric acid aqueous solution to adjust the pH of the feed solution 1 to 4, add biological enzyme 2, and perform secondary enzymatic hydrolysis at 52°C and 150rpm for 1h, and inactivate the enzyme at 90°C for 10min to obtain enzymatic solution 2.

[0087] S7. Purification: Add 2% by weight of a clarifier (ZTCⅡ type clarifier) ​​to the enzymatic hydrolyzate 2, place it in a 55°C water bath for 30 min, and then filter it through 300 mesh diatomaceous earth and 5 μm polypropylene membrane in sequence to obtain liquid 2, which is the finished camellia extract.

[0088] In a second aspect, this embodiment provides a camellia extract prepared by the preparation method described above, wherein the camellia extract contains camellia saccharides and camellia plant peptides.

[0089] The camellia extract was analyzed by HPLC (high performance liquid chromatography), and the camellia sugars therein mainly included monosaccharides with the following mass content ratios: glucose (64.8%), galactose (20.4%), arabinose (8.5%), mannose (3.0%), galacturonic acid (1.4%), and xylose (0.6%).

[0090] The camellia extract was analyzed by GPC (gel permeation chromatography), and the weight average molecular weight Mw of the camellia sugars was 1.288×10 4 .

[0091] Peptide sequence analysis of the camellia extract revealed that there were 111 types of camellia plant peptides.

[0092] The third aspect of this embodiment provides the use of the camellia extract as described above in a skin care composition.

[0093] Calculated by weight percentage, the skin care composition comprises: 20% of the camellia extract, 20% of glycerol, 0.5% of 1,2-hexanediol, 0.5% of p-hydroxyacetophenone, and the balance of water.

[0094] The contents of total sugar and total peptide in the camellia extract are shown in Table 1 below.

[0095] The preparation steps of the skin care composition include: mixing the raw materials according to the formula amount, filtering with a 0.22 μm polypropylene membrane, and obtaining the skin care composition.

[0096] Example 2

[0097] This embodiment provides a preparation method, product and application of camellia extract for resisting photoaging, and the specific implementation method is the same as that of embodiment 1; the difference is that:

[0098] The step S1 is freeze-drying and crushing through an 80-mesh sieve.

[0099] The extraction time in step S4 is 3 hours.

[0100] The camellia extract was analyzed by HPLC (high performance liquid chromatography), and the camellia sugars therein mainly included monosaccharides with the following mass content ratios: glucose (56.1%), galactose (21.6%), arabinose (10.5%), mannose (3.6%), galacturonic acid (1.7%), and xylose (0.7%).

[0101] The camellia extract was analyzed by GPC (gel permeation chromatography), and the weight average molecular weight Mw of camellia sugars was 1.054×10 4 .

[0102] Peptide sequence analysis of the camellia extract revealed that there were 136 types of camellia plant peptides.

[0103] The contents of total sugar and total peptide in the camellia extract are shown in Table 1 below.

[0104] Example 3

[0105] This embodiment provides a preparation method, product and application of camellia extract for resisting photoaging, and the specific implementation method is the same as that of embodiment 1; the difference is that:

[0106] The amount of deionized water added in step S2 is 15 times the mass of camellia powder.

[0107] The skin care composition comprises: 30% of camellia extract, 20% of glycerin, 0.5% of 1,2-hexanediol, 0.5% of p-hydroxyacetophenone, and the balance of water.

[0108] The camellia extract was analyzed by HPLC (high performance liquid chromatography), and the camellia sugars therein mainly included monosaccharides with the following mass content ratios: glucose (69.2%), galactose (16.5%), arabinose (7.7%), mannose (3.3%), galacturonic acid (1.7%), and xylose (0.8%).

[0109] The camellia extract was analyzed by GPC (gel permeation chromatography), and the weight average molecular weight Mw of camellia sugars was 7.658×10 3 .

[0110] Peptide sequence analysis of the camellia extract revealed that there were 151 types of camellia plant peptides.

[0111] The contents of total sugar and total peptide in the camellia extract are shown in Table 1 below.

[0112] Example 4

[0113] This embodiment provides a preparation method, product and application of camellia extract for resisting photoaging, and the specific implementation method is the same as that of embodiment 1; the difference is that:

[0114] The S1 step is freeze drying.

[0115] The temperature in step S4 is raised to 85°C.

[0116] The camellia extract was analyzed by HPLC (high performance liquid chromatography), and the camellia sugars therein mainly included monosaccharides with the following mass content ratios: glucose (62.4%), galactose (20.8%), arabinose (8.9%), mannose (2.7%), galacturonic acid (1.6%), and xylose (0.5%).

[0117] The camellia extract was analyzed by GPC (gel permeation chromatography), and the weight average molecular weight Mw of camellia sugars was 3.792×10 4 .

[0118] Peptide sequence analysis of the camellia extract revealed that there were 103 types of camellia plant peptides.

[0119] The contents of total sugar and total peptide in the camellia extract are shown in Table 1 below.

[0120] Example 5

[0121] This embodiment provides a preparation method, product and application of camellia extract for resisting photoaging, and the specific implementation method is the same as that of embodiment 1; the difference is that:

[0122] Based on the total mass of camellia powder, the biological composite enzyme is added in the following amounts: 0.3% cellulase, 0.3% pectinase, 0.3% mesophilic α-amylase, 0.3% xylanase and 0.2% tannase.

[0123] The camellia extract was analyzed by HPLC (high performance liquid chromatography), and the camellia sugars therein mainly included monosaccharides with the following mass content ratios: glucose (56.3%), galactose (24.5%), arabinose (9.3%), mannose (3.2%), galacturonic acid (1.2%), and xylose (0.4%).

[0124] The camellia extract was analyzed by GPC (gel permeation chromatography), and the weight average molecular weight Mw of camellia sugars was 2.316×10 4 .

[0125] Peptide sequence analysis of the camellia extract revealed that there were 106 types of camellia plant peptides.

[0126] The contents of total sugar and total peptide in the camellia extract are shown in Table 1 below.

[0127] Comparative Example 1

[0128] This comparative example provides a preparation method, product and application of a camellia extract, and the preparation steps include:

[0129] S1. Pre-processing camellia: picking fresh camellia, sorting to remove calyx, receptacle and impurities, drying in a 40°C oven, crushing through a 60-mesh sieve to obtain camellia fine powder.

[0130] S2. Extraction: Mix camellia powder and 10 times the mass of deionized water, heat to 92°C, and stir at 150 rpm for 2 h to obtain an extract.

[0131] S3. Filtration: The extract obtained in step S2 is filtered through 200 mesh gauze, 300 mesh diatomaceous earth and 5 μm polypropylene membrane to obtain feed solution 1. In this step, 300 mesh diatomaceous earth and 5 μm polypropylene membrane are used simultaneously, and the polypropylene membrane is placed at the bottom and the diatomaceous earth is placed on top of the polypropylene membrane for filtration.

[0132] S4. Purification: Add 2% by weight of a clarifier (ZTCⅡ type clarifier) ​​to liquid 1, place in a 55°C water bath for 30 min, and then filter through 300 mesh diatomaceous earth and 5 μm polypropylene membrane in sequence to obtain liquid 2, which is the finished camellia extract.

[0133] The contents of total sugar and total peptide in the obtained camellia extract are shown in Table 1 below.

[0134] The camellia extract is applied to a skin care composition, which comprises, by weight percentage, 20% of the camellia extract, 20% of glycerol, 0.5% of 1,2-hexanediol, 0.5% of p-hydroxyacetophenone, and the balance of water.

[0135] Comparative Example 2

[0136] This comparative example provides a preparation method, product and application of a camellia extract, and the preparation steps include:

[0137] S1. Pre-processing camellia: picking fresh camellia, sorting to remove calyx, receptacle and impurities, drying in a 40°C oven, crushing through a 60-mesh sieve to obtain camellia fine powder.

[0138] S2. Activating the biological enzyme 1: dissolving the biological enzyme 1 in deionized water 10 times the mass of the camellia powder, and activating the enzyme at 35°C for 1 hour to obtain an activated enzyme solution;

[0139] The biological enzyme 1 is a biological composite enzyme, which includes cellulase, pectinase, medium-temperature α-amylase, xylanase and tannase; based on the total mass of camellia powder, the biological composite enzyme is added according to 0.5% of cellulase, 0.5% of pectinase, 0.5% of medium-temperature α-amylase, 0.5% of xylanase and 0.2% of tannase.

[0140] The cellulase activity is 3500U / g, the pectinase activity is 60000U / g, the medium-temperature α-amylase activity is 1000U / g, the xylanase activity is 80000U / g, and the tannin activity is 300U / g; all of them come from Cangzhou Xiasheng Enzyme Biotechnology Co., Ltd.

[0141] S3. Primary enzymatic hydrolysis: Add camellia powder to the activated enzyme solution, adjust the pH to 4.5 with 19% citric acid aqueous solution, and perform enzymatic hydrolysis at 52° C. and 150 rpm for 2 h to obtain enzymatic hydrolyzate 1.

[0142] S4. Extraction: Heat the enzymatic hydrolyzate 1 to 92°C, stir at 150 rpm for 2 h, and obtain an extract.

[0143] S5. Filtration: The extract obtained in step S4 is filtered through 200 mesh gauze, 300 mesh diatomaceous earth and 5 μm polypropylene membrane to obtain feed solution 1. In this step, 300 mesh diatomaceous earth and 5 μm polypropylene membrane are used simultaneously, and the polypropylene membrane is placed at the bottom and the diatomaceous earth is placed on top of the polypropylene membrane for filtration.

[0144] S6. Purification: Add 2% of the mass of clarifier (ZTCⅡ clarifier, Beijing Zhengtiancheng Clarification Technology Co., Ltd.) to the feed solution 1, let it stand in a 55°C water bath for 30 minutes, and then filter it through 300 mesh diatomaceous earth and 5 μm polypropylene membrane in sequence to obtain feed solution 2, which is the finished product of camellia extract.

[0145] The contents of total sugar and total peptide in the obtained camellia extract are shown in Table 1 below.

[0146] The camellia extract is applied to a skin care composition, which comprises, by weight percentage, 20% of the camellia extract, 20% of glycerol, 0.5% of 1,2-hexanediol, 0.5% of p-hydroxyacetophenone, and the balance of water.

[0147] Comparative Example 3

[0148] This comparative example provides a preparation method, product and application of a camellia extract, and the specific implementation method is the same as that of Example 1; the difference is that:

[0149] The biological composite enzyme comprises cellulase, pectinase, mesophilic α-amylase and xylanase; based on the total mass of camellia powder, the biological composite enzyme is added in the amount of 0.5% cellulase, 0.5% pectinase, 0.5% mesophilic α-amylase and 0.5% xylanase.

[0150] The contents of total sugar and total peptide in the obtained camellia extract are shown in Table 1 below.

[0151] Comparative Example 4

[0152] This comparative example provides a preparation method, product and application of a camellia extract, and the specific implementation method is the same as that of Example 1; the difference is that:

[0153] The temperature in step S4 is raised to 70°C.

[0154] The contents of total sugar and total peptide in the obtained camellia extract are shown in Table 1 below.

[0155] Performance Testing

[0156] 1. Camellia total sugar content test

[0157] 1. Preparation of standard solution: Accurately weigh 10.5 mg of anhydrous glucose, add distilled water to a 100 mL volumetric flask to make a standard glucose solution with a concentration of 0.105 mg / mL.

[0158] 2. Preparation of phenol solution: Weigh 5 g of phenol, add distilled water to a 100 mL volumetric flask to make a 5% phenol solution.

[0159] 3. Drawing of the standard curve: Accurately measure 0, 0.1, 0.2, 0.4, 0.8, 1.2, 1.6, and 2.0 mL of glucose standard solution respectively and place them in stoppered colorimetric tubes, and add water to make up to 2.0 mL respectively. Add 1 mL of 5% phenol solution and 5 mL of concentrated sulfuric acid respectively, cover with a stoppered lid and shake well, place in a 100°C water bath for 20 min, take out, and quickly cool to room temperature in a cold water bath. Use the corresponding reagent (0 mL of glucose standard solution measured in the previous step) as the blank, and measure the absorbance at a wavelength of 490 nm. Use glucose concentration as the horizontal axis and absorbance as the vertical axis to draw a standard curve as shown below. Figure 1 shown.

[0160] 4. Determination of total sugar content of camellia: Take the camellia extracts prepared in Examples 1 to 5 and Comparative Examples 1 to 4, dilute them 20 times with distilled water, take 0.20 mL and place them in a stoppered test tube, add 1 mL of 5% phenol solution and 5 mL of concentrated sulfuric acid, place them in a 100°C water bath for 20 minutes, take them out, and quickly cool them to room temperature in a cold water bath. Measure the absorbance at a wavelength of 490 nm, and calculate the total sugar content of camellia according to the standard curve. The test results of the total sugar content of camellia in the camellia extracts of each Example and Comparative Example are shown in Table 1.

[0161] 2. Camellia total peptide content test

[0162] 1. Preheat the spectrophotometer for 30 minutes, adjust the wavelength to 540nm, and zero with distilled water.

[0163] 2. Blank tube: Take a glass test tube, add 200μL distilled water and 1000μL biuret reagent, mix well and let stand at room temperature for 15 minutes, compare colors at 540nm, and record it as A blank tube.

[0164] 3. Standard tube: Take a glass test tube, add 200μL standard solution and 1000μL biuret reagent, mix well and let stand at room temperature for 15 minutes, compare colors at 540nm, and record it as A standard tube.

[0165] 4. Test tube: Take the camellia extract prepared in Examples 1 to 5 and Comparative Examples 1 to 4, dilute it 2 times with distilled water to obtain a test solution; take a glass test tube, add 200 μL of the test solution and 1000 μL of biuret reagent, mix well, let it stand at room temperature for 15 minutes, and compare the color at 540 nm, which is recorded as A test tube.

[0166] The formula for calculating the concentration of camellia total peptides is: C to be tested (mg / mL) = C standard tube × (A test tube - A blank tube) ÷ (A standard tube - A blank tube) × D (where C standard: 5 mg / mL; D: dilution multiple)

[0167] The test results of total peptide content in camellia extracts of various embodiments and comparative examples are shown in Table 1.

[0168] 3. Stability Test

[0169] Take 30 g of the camellia extract of Examples 1 to 5 and Comparative Examples 1 to 4 respectively, store them in a sealed clean container, place them in a refrigerator at -18°C for 24 hours, take them out and place them at room temperature for 24 hours, repeat this cycle 7 times and compare and evaluate them with the initial sample to observe whether the sample has abnormal conditions such as color change, turbidity, precipitation, etc.

[0170] The stability test results of each embodiment and comparative example are shown in Table 1.

[0171] Table 1

[0172]

[0173] IV. DPPH free radical scavenging activity test

[0174] 1. Preparation of DPPH solution: Weigh DPPH powder and dissolve it in ethanol to prepare 0.2 mM DPPH ethanol solution.

[0175] 2. Preparation of positive control solution: Weigh L-ascorbic acid powder and dissolve it in distilled water to prepare a 10 mg / mL L-ascorbic acid solution, and dilute it with ethanol to a final experimental concentration of 5 μg / mL.

[0176] 3. Preparation of test sample dilution solution: dilute the test sample (the camellia extracts prepared in Examples 1 to 5 and Comparative Examples 1 to 4) with ethanol to an experimental concentration of 2% to obtain a sample solution.

[0177] 4. DPPH free radical scavenging test: 500 μL of sample solution and positive control solution were added to a 24-well plate (i.e., the final experimental concentration of the sample was 1%), and 500 μL of ethanol was added to the DPPH tube (C) (3 parallels). Two groups were set up for each sample, and 500 μL of 0.2 mM DPPH solution was added to one group, which was recorded as T-sample tube; as a compensation for the sample's own absorbance value, 500 μL of ethanol was added to the other group, which was recorded as T0-sample background. The reaction was carried out at room temperature and in the dark for 30 minutes. The absorbance was measured at a wavelength of 517 nm, and the DPPH scavenging rate of the sample to be tested and the positive control L-ascorbic acid was calculated; DPPH scavenging rate = 1-(T-T0) / C×100%.

[0178] The results of DPPH free radical scavenging rates of various embodiments and comparisons are shown in Table 2; the values ​​in Table 2 are the average values ​​of the measurement results of three parallel samples of each sample.

[0179] Table 2

[0180] name DPPH free radical scavenging rate L-Ascorbic Acid 70.17％ Example 1 91.10％ Example 2 95.28％ Example 3 94.90％ Example 4 85.82％ Example 5 82.07％ Comparative Example 1 24.58％ Comparative Example 2 58.14％ Comparative Example 3 80.94％ Comparative Example 4 40.53％

[0181] 5. HaCaT cell ROS level test

[0182] H2O2, ultraviolet rays, etc. can induce an increase in the level of reactive oxygen species (ROS) in human keratinocytes, activate the mitogen-activated protein kinase (MAPK) signaling pathway, promote the expression of matrix metalloproteinases (MMPs), and inhibit the synthesis of collagen. ROS can oxidize non-fluorescent DCFH to generate fluorescent DCF, and the fluorescence intensity is proportional to the level of intracellular reactive oxygen species.

[0183] Before the ROS level test, the present invention used MTT colorimetry to detect cell viability and determined the safe concentration of the camellia extracts in the embodiment and the comparative example acting on HaCaT cells. HaCaT cells were cultured in a DMEM high-glucose medium containing 10% fetal bovine serum and 1% double antibody in a constant temperature incubator at 37°C and 5% CO2. Cells in the logarithmic growth phase were taken to prepare a cell suspension, and 1×10 5 The cells were inoculated at a density of 100 / well in a 24-well plate and incubated in a CO2 constant temperature incubator. After 24 h of incubation, all wells except the control group were treated with a dose of 15 mJ / cm 2 The cells were irradiated with UVB. After irradiation, a high-sugar culture medium containing 0.156% concentration of the camellia extract samples of the embodiment and the comparative example was added to each well of the sample group, and 500 μL of high-sugar culture medium was added to each well of the control group and the irradiation group. After administration, the 24-well plate was placed at 37°C and 5% CO2 for 24 hours. The culture medium was discarded, the cell well plate was washed twice with serum-free culture medium, 500 μL of DCFH-DA working solution (10 μM) was added, and incubated in a 37°C cell culture incubator for 30 minutes. The cells were washed 3 times with DPBS, observed and photographed under a fluorescence microscope, and the fluorescence intensity was analyzed using Image J. The test results are shown in Figure 2 .

[0184] The level of cellular ROS reflects the degree of cellular oxidative stress. The lower the fluorescence intensity of the test sample, the stronger the effect of the surface sample in alleviating oxidative stress. Figure 2 It can be seen that compared with the control group, the ROS free radicals in the HaCaT cells of the UVB irradiation group increased significantly, indicating that the model was successfully constructed. Compared with the irradiation group, the camellia extract samples of Examples 1-5 can remove the increase of ROS free radicals caused by UVB irradiation to varying degrees, indicating that the camellia extracts of Examples 1-5 can effectively reduce the oxidative stress level of UVB-induced damaged cells. By comparison, it can be seen that Examples 2 and 3 have the best effects, and overall, the effects of the camellia extracts of Examples 1-5 are better than those of Comparative Examples 1-4.

[0185] VI. Test of the C-terminal peptide content of type Ⅰ procollagen in HSF cells

[0186] The type I procollagen synthesized in the cell is secreted outside the cell. Under the action of endopeptidase, the propeptides attached to its amino and carboxyl ends are cut off to form tropocollagen. Tropocollagen molecules polymerize into collagen fibers to form the extracellular matrix. The cut free propeptide is soluble and can be used as a biochemical indicator to reflect the amount of collagen synthesis in the body. The free propeptide detected in this test is the type I procollagen C-terminal peptide (PIP for short).

[0187] Before the PIP test, the cell viability was detected by MTT colorimetry to determine the safe concentration of the camellia extracts in the examples and comparative examples for acting on HSF cells (human skin fibroblasts). HSF cells were cultured in a cell culture incubator at 37°C and 5% CO2 using a high-glucose DMEM medium containing 10% fetal bovine serum and 1% streptomycin / penicillin. The medium was changed and passaged regularly, and cells in the logarithmic growth phase were taken for the experiment. HSF cells were cultured at 8×10 3 The cells were seeded at a density of 0.5 / well in a 96-well plate and cultured for 24 hours. A high-glucose culture medium containing 0.625% concentration of the camellia extract samples of the example and the comparative example was added to each well of the sample group. The positive control group was a high-glucose culture medium containing 100 ng / mL hEGF (human epidermal growth factor). 100 μL of high-glucose culture medium was added to each well of the control group. After the administration was completed, the 96-well plate was placed at 37°C and 5% CO2 and cultured for 48 hours. After the culture was completed, the supernatant was taken, and the content of type I procollagen C-terminal peptide (PIP) was detected using Procollagen type I C-peptide (PIP) EIA Kit (Takara) at 4°C, 12000 rpm, for 10 minutes. The test results are shown in Figure 3 .

[0188] like Figure 3 In the test results of type I procollagen C-terminal peptide (PIP) content, compared with the blank control, the secretion of PIP was significantly increased after the positive drug 100ng / mL hEGF acted on HSF cells. After 48h of the 0.625% concentration of the camellia extract of the example, the PIP content was significantly increased, indicating that it has the effect of promoting the synthesis of type I collagen, among which Examples 2 and 3 have the best effects, and overall, the effects of the camellia extracts of Examples 1-5 are better than those of Comparative Examples 1-4.

[0189] VII. Test of MMP-1 content in UVB-induced HSF cells

[0190] Human skin fibroblasts (HSF) in logarithmic growth phase were selected to prepare cell suspension, and 5×10 4 The cells were inoculated at a density of 100 / well in a 24-well plate. After 24 h of culture, all wells except the control group were irradiated with UVB at a dose of 25 mJ / cm 2After irradiation, a high-glucose medium containing 0.625% of the camellia extract samples of the embodiment and the comparative example was added to each well of the sample group, and 500 μL of high-glucose medium was added to each well of the control group and the irradiation group. After administration, the 24-well plate was placed at 37°C and 5% CO2 for incubation. After 24 hours of incubation, the culture medium was collected, and the MMP-1 content of the culture medium was detected using the Human Total MMP-1 (R&D Systems) kit instructions. The test results are shown in Figure 4 .

[0191] like Figure 4 In the MMP-1 content detection results shown, compared with the blank control, UVB irradiation significantly increased the secretion of MMP-1 in HSF cells, and the model was successfully constructed. After 24 hours of exposure to 0.625% concentration of the camellia extract in the examples, the MMP-1 content decreased significantly, which has the effect of reducing abnormal degradation of the extracellular matrix, among which Examples 2 and 3 have the best effects, and overall, the effects of the camellia extracts in Examples 1-5 are better than those in Comparative Examples 1-4.

[0192] 8. Skin patch safety test

[0193] Referring to the skin occlusive patch test method in the "Technical Specification for Safety of Cosmetics" (2015 edition), 31 subjects aged 20-55 years old were selected to meet the conditions. The camellia extracts of Example 1 and Example 3 were prepared into sample solutions with mass fractions of 1%, 2%, and 5%, respectively, with distilled water. 2 , a patch test device with a depth of about 1mm. 20μL of sample solution was added to the small chamber of the patch tester, and the blank control group was distilled water of the same dose. The patch tester with the test object was applied to the curved side of the subject's forearm with a low-allergenic tape, and gently pressed with the palm of the hand to evenly apply it to the skin for 24 hours. After removing the test object patch tester, observe the skin reaction according to the standards in Table 3 at 30 minutes (after the indentation disappears), 24 hours and 48 hours, and record the observation results; the skin reaction grading scale of the skin closed patch test is shown in Table 3, and the results of the human closed patch test are shown in Table 4. The results of the skin patch test showed that there was no adverse skin reaction among 31 people, and the camellia extract provided by the present invention has good safety.

[0194] Table 3

[0195]

[0196] Table 4

[0197]

[0198]

[0199] 9. Clinical anti-aging test

[0200] 16 healthy subjects were screened, and after washing their hands and faces, they used paper towels to wipe off the moisture on their skin. After the subjects entered the test environment (constant temperature and humidity environment: temperature 22±2°C, ambient humidity 50±10% RH), they rested and waited for 20 minutes before entering the test. The method of using the samples is to apply 2% of the basic lotion of Example 1 and Comparative Example 2 to the left and right halves of the face respectively (once in the morning and once in the evening) for 4 weeks. Using the ANTSCI Callegari Soft-PLUS elasticity test probe, the skin elasticity of the left and right cheekbones of the subjects was tested on day 0 (D0), day 14 (D14), and day 28 (D28) (the larger the value, the better the surface skin elasticity). The average value was taken after 3 parallel measurements. The test results are shown in the table. Figure 5 (The statistical difference of the test data was analyzed at the same time, compared with D0, *P<0.05). According to weight percentage, the formula of the basic emulsion is: 2% of the camellia extract prepared in Example 1 or Comparative Example 2, 3% of soybean oil, 1.5% of glyceryl stearate (A165), 0.5% of 1618 alcohol, 0.2% of carbomer 940, 3% of butylene glycol, 0.5% of p-hydroxyacetophenone, 0.5% of 1,2-hexanediol, NaOH was used to adjust the pH to 5.5, and water was added to 100%.

[0201] Note: The clinical anti-aging test used a half-face test, that is, 16 subjects used the product containing Example 1 on one side of their face, and used the product containing Comparative Example 2 on the other side. All subject data were averaged, compared with day 0, and significant differences were calculated.

[0202] like Figure 5 As shown, the subjects applied the basic lotion containing 2% of the camellia extract (Example 1) on their faces, and the skin elasticity increased at 14 days and 28 days, with the increase rates being 6.93% and 9.42% respectively, both with significant differences compared to the initial (0 day) (P<0.05). After using the basic lotion of Comparative Example 2, the change in skin elasticity was slightly increased compared to the initial, but there was no significant difference (P>0.05).

Claims

1. A method for preparing an anti-photoaging camellia extract, characterized in that: The preparation steps include: S1. Pre-treating camellia to obtain camellia powder; S2. activating the biological enzyme 1 to obtain an activated enzyme solution; S3. Add camellia powder to the activated enzyme solution and perform enzymatic hydrolysis to obtain enzymatic solution 1; S4. extracting the enzymatic hydrolyzate 1 to obtain an extract; S5. The extract is filtered to obtain liquid 1; S6. Mixing the feed solution 1 and the biological enzyme 2, performing secondary enzymolysis, and obtaining enzymolysis solution 2; S7. Purify the enzymatic hydrolyzate 2 to obtain camellia extract.

2. The method for preparing the anti-photoaging camellia extract according to claim 1, characterized in that: The biological enzyme 1 is a biological composite enzyme, including at least three of cellulase, pectinase, α-amylase, xylanase, tannase, glucanase, mannanase and glucoamylase.

3. The method for preparing the anti-photoaging camellia extract according to claim 2, characterized in that: The biological composite enzyme comprises cellulase, pectinase, α-amylase, xylanase and tannase, and the mass ratio is (0.1-3): (0.1-2): (0.1-4): (0.15-2): (0.05-1.5).

4. The method for preparing the anti-photoaging camellia extract according to claim 3, characterized in that: Based on the total weight of camellia powder, the biological composite enzyme is added according to 0.2%-1.0% of cellulase, 0.2%-1.0% of pectinase, 0.2%-1.0% of alpha-amylase, 0.2%-1.0% of xylanase and 0.1%-0.5% of tannase.

5. The method for preparing the anti-photoaging camellia extract according to claim 1, characterized in that: The step S4 specifically includes: heating the enzymatic hydrolyzate 1 to 85-95° C., stirring and extracting at 100-200 rpm for 1-3 hours to obtain an extract.

6. The method for preparing the anti-photoaging camellia extract according to claim 1, characterized in that: The step S1 specifically includes: picking fresh camellia flowers, sorting and removing calyx and receptacle, low-temperature drying, crushing and sieving to obtain camellia fine powder.

7. The method for preparing the anti-photoaging camellia extract according to claim 6, characterized in that: The drying method includes at least one of vacuum drying, freeze drying and oven drying.

8. The anti-photoaging camellia extract according to claim 7, characterized in that The mesh size of the sieve is 20-200 meshes; Preferably, the mesh size of the sieve is 40-80 meshes.

9. A camellia extract prepared according to the preparation method according to any one of claims 1 to 8, characterized in that: The camellia extract contains camellia sugars and camellia plant peptides; The camellia extract is subjected to peptide sequence analysis, wherein the types of camellia plant peptides are ≥100.

10. A use of the camellia extract according to claim 9, characterized in that: The camellia extract is applied to a skin care composition; the added amount of the camellia extract in the skin care composition is 10-30 wt %.

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