Cell energy activator and application thereof

Abstract

The invention relates to the technical field of cosmetics, in particular to a preparation method of a cell energy activator containing nicotinamide, VC ethyl ether and a bacillus fermentation product and application of the cell energy activator in skin oxidation resistance. The preparation method comprises the following steps: preparing a fermentation culture medium containing golden camellia powder and glucose powder, activating bacillus, and inoculating the bacillus into the fermentation culture medium to prepare a first component; inoculating the activated bacillus into a culture medium to prepare a second component; mixing the first component with the second component, and then adding nicotinamide and VC ethyl ether to prepare the cell energy activator. The cell energy activator disclosed by the invention can play a synergistic role, effectively improve the antioxidant activity of the skin and improve the skin barrier.

Description

Technical Field

[0001] The present invention relates to the technical field of cosmetics. Specifically, the present invention relates to a preparation method of a cell energy activator containing microbial metabolites and its application in skin antioxidant and improving the skin barrier function. Background Art

[0002] Free radicals are a natural product of human survival activities. All human activities such as breathing, exercise, and diet will generate free radicals (for example, the human body continuously releases energy through oxidation during breathing, and mitochondria will convert part of the oxygen into reactive oxygen free radicals). In addition, the influence of the external environment will also generate free radicals, such as smoking and drinking, ultraviolet rays, electromagnetic waves, environmental pollutants, drugs, and emotional stress. Cosmetic antioxidant mainly reduces the damage of cell membranes, DNA, lipids, or proteins caused by skin oxidative stress and the damage caused by oxidation to human skin by scavenging free radicals, reducing the ROS level, increasing the activity of antioxidant enzymes, and reducing lipid metabolites. Most antioxidant test methods also use scavenging free radicals, reducing the ROS level, increasing the activity of antioxidant enzymes, etc. as evaluation criteria. With the continuous expansion of the skin care product market, finding natural and more effective anti-aging ingredient bio-skin care formulations is an important direction for current research and development.

[0003] In the field of new cosmetic development, postbiotics, as non-living microbial preparations, are often used to regulate the skin microecology and improve the skin condition, thereby anti-aging. Postbiotics refer to soluble substances secreted by probiotics or released after the decomposition of bacterial cells, mainly bacterial components and bacterial metabolites. Postbiotics can be prepared by fermentation using a carbon source medium and specific strains, and it consists of different metabolites and has important biological significance such as antioxidant and anti-inflammatory effects.

[0004] The main purpose of the present invention is to develop an antioxidant bio-formulation containing postbiotics and use it in cosmetic development. Summary of the Invention

[0005] The purpose of the present invention is to provide a preparation method and application of a skin antioxidant cell energy activator.

[0006] Nicotinamide, also known as niacinamide and nicotinic acid amide, is an amide compound of niacin (vitamin B3). It can be converted into NAD+ or NADP+ in the body and plays a role in transferring hydrogen ions and electrons in many oxidation-reduction reactions in the body. It is mainly used to prevent and treat pellagra, stomatitis, glossitis, sick sinus syndrome, atrioventricular block and other problems. Nicotinamide is a water-soluble vitamin and a member of the vitamin B group. Nicotinamide is an antioxidant that can interact with the skin, stimulate the required coenzymes, and better protect the epidermis from external damage. It can enhance the healing ability of skin cells, improve the structure of different skin layers, ensure optimal skin moisture and regulate sebum secretion.

[0007] VC Ethyl Ether, full name "Vitamin C Ethyl Ether", also known as 3-O-Ethyl Ascorbic Acid, CAS No. 86404-04-8. Due to its obvious skin care effect and good compatibility, VC-Ethyl Ether has been increasingly favored by the domestic and international cosmetics industries. It is reported that vitamin C ethyl ether derivative is the best vitamin C derivative so far. It is not only very stable in chemical properties, but also easily decomposed by biological enzymes after entering the skin to play the role of vitamin C. From a chemical point of view, VC Ethyl Ether is an amphiphilic VC derivative with lipophilic and hydrophilic properties. This makes it extremely convenient to use in formulations and easy to penetrate the stratum corneum into the dermis. After entering the skin, it is easily decomposed by biological enzymes to play the role of VC, improving its bioavailability. It has a strong antioxidant effect and can effectively remove free radicals.

[0008] Camellia nitidissima (scientific name: Camellia petelotii) is an evergreen shrub of the genus Camellia in the family Theaceae. Research shows that Camellia nitidissima is rich in various bioactive substances such as tea polysaccharides, tea polyphenols, saponins, and flavonoids. Among them, Camellia nitidissima polysaccharides have attracted much attention due to their unique biological activities. However, when extracting Camellia nitidissima polysaccharides from raw materials such as tea leaves, tea flowers, and tea seeds, they are easily interfered by small molecular compounds such as polyphenols, pigments, proteins, and inorganic salts. These impurities not only affect the biological activities of Camellia nitidissima polysaccharides, but also interfere with their structural quantitative analysis and identification.

[0009] The prior art CN118512341A discloses a cell energy activating composition with anti-aging effects and its application in men's skin care. However, with the in-depth research, protecting the skin by utilizing the antioxidant properties of microorganisms is also one of the research directions with relatively more studies. The prior art CN119950377A discloses an emotional beauty composition for enhancing cell vitality and reducing wrinkles, its preparation method and application, including andrographis paniculata extract, bacillus fermentation product, 4-tert-butylcyclohexanol, and pseudostellaria heterophylla extract. The prior art CN119632893A discloses a composition for anti-pollution repair of acne-prone and sensitive skin containing camellia nitidissima extract and its application. However, it does not disclose the content related to using postbiotics to enhance the anti-aging effect of the skin. The microbial fermentation derivatives in skin care products are fermented under external nutrient conditions from different sources to obtain skin care valuable ingredients. By breaking the cell wall and filtering its cell sap, the effective ingredients can be extracted. Based on this, on the basis of the previous research of the inventors, a combination based on bacillus fermentation product and natural anti-aging substances is further developed to obtain the present invention. The anti-oxidation and anti-aging effects of the present invention are reflected by the determination of relevant indicators. SOD, CAT, and GSH are antioxidant enzyme groups that make up the cell defense system. SOD is an important antioxidant enzyme in the body, which can scavenge the superoxide anion radicals produced by the body and can indirectly reflect the antioxidant capacity of the body. CAT is an antioxidant enzyme present in all living tissues, which can degrade or reduce hydrogen peroxide into water and molecular oxygen. GSH is a peroxidase that maintains the normal immune system function of the body, which can remove lipid peroxides and H2O2, protecting tissues and cells from the attack and damage of free radicals. The total antioxidant capacity (T-AOC) refers to the total antioxidant level composed of various antioxidant enzyme systems and non-enzyme systems, such as antioxidant enzymes like SOD, CAT, and antioxidants like VC, VE, etc., which is used to reflect the total antioxidant capacity of bioactive substances in the body.

[0010] One aspect of the present invention provides a preparation method of a cell energy activator, comprising the following steps:

[0011] 1) Dry and crush camellia nitidissima, sieve it through a 100 - 150 mesh sieve to obtain camellia nitidissima powder; add the camellia nitidissima powder into distilled water at 60°C, mix and stir for 20 min, add glucose powder as a carbon source, stir and homogenize, then place it in a water bath, sterilize at 90 - 100°C for 30 min to obtain a fermentation medium; each liter of the fermentation medium contains 5 - 15 parts of camellia nitidissima powder, 2.5 - 5 parts of glucose powder, and each part is 10 g;

[0012] After activating the bacillus, inoculate it into the fermentation medium, and the inoculation density is 5.0 - 6.0×10 6cfu / mL, ferment at 28 - 33°C for 18 - 24 hours, centrifuge at 6000 rpm for 10 min, take the supernatant, filter through a 0.22 μm filter membrane, collect the filtrate, and concentrate by freeze-drying to prepare the first component;

[0013] 2) Inoculate the activated Bacillus subtilis into a medium for fermentation. After fermentation, homogenize the cells and disrupt them by ultrasonic treatment. The ultrasonic power is 1000 w, the time is 20 min, working for 3 s and intermittent for 3 s. Centrifuge and take the supernatant, and concentrate by freeze-drying to prepare the second component;

[0014] 3) Preparation of cell energy activator: Mix the first component and the second component, and add 3 - 5 parts of nicotinamide and 0.5 - 1.5 parts of VC ethyl ether to prepare the cell energy activator.

[0015] Furthermore, the content of Camellia nitidissima powder is 10 parts, and the content of glucose powder is 2.5 parts.

[0016] Furthermore, the inoculation density of Bacillus subtilis is 5.5×10 6 cfu / mL.

[0017] Furthermore, in step 3), add 3 - 5 parts of nicotinamide and 0.5 - 1.5 parts of VC ethyl ether.

[0018] One aspect of the present invention provides a cell energy activator prepared by the foregoing method.

[0019] One aspect of the present invention provides a skin care product, which is characterized by comprising the foregoing cell energy activator.

[0020] Furthermore, it further comprises at least one of a humectant, an antioxidant, an anti-inflammatory agent, and a chelating agent.

[0021] One aspect of the present invention provides the use of the foregoing cell energy activator in the preparation of a skin antioxidant product.

[0022] Furthermore, the application object is male.

[0023] One aspect of the present invention provides the use of the foregoing cell energy activator in the preparation of a product for enhancing the activity of catalase.

[0024] Among them, the medium for inoculating the activated Bacillus subtilis into the medium for fermentation includes conventional components such as beef extract peptone medium components. It belongs to the general knowledge in the art.

[0025] The beneficial effects of the present invention are as follows:

[0026] (1) The present invention uses a prepared fermentation medium containing camellia nitidissima powder and glucose powder. After activating the bacillus, it is inoculated into the fermentation medium to prepare the first component; the activated bacillus is inoculated into the medium to prepare the second component; the first component and the second component are mixed, and niacinamide and VC ethyl ether are added to prepare a cell energy activator. The cell energy activator of the present invention can effectively enhance the antioxidant activity of the skin.

[0027] (2) The bacillus fermentation method of the present invention can obtain a high-density fermentation product, and its antioxidant effect is significantly superior.

[0028] (3) After being prepared into a skin care product, the cell energy activator of the present invention can effectively increase the contents of CAT, SOD and GSH in the skin. Detailed implementation mode

[0029] The embodiments of the present invention provide a preparation method and application of a cell energy activator containing microbial metabolites. To better illustrate the purpose, technical solution and advantages of the present invention, the present invention will be further described below in conjunction with specific embodiments. Its purpose is to understand the content of the present invention in detail, rather than a limitation to the present invention. All other embodiments obtained by those of ordinary skill in the art without creative efforts fall within the protection scope of the present invention. The experimental reagents and instruments designed in the embodiments and comparative examples of the present invention are all common ordinary reagents and instruments unless otherwise specified, and can be obtained from commercial channels. In the embodiments and comparative examples, the experimental methods used are all conventional methods unless otherwise specified; and unless otherwise specified, the raw materials used in parallel experiments are of the same batch.

[0030] Example 1

[0031] A preparation method of a cell energy activator includes the following steps:

[0032] 1) Dry and crush camellia nitidissima, pass through a 140-mesh sieve to obtain camellia nitidissima powder; add the camellia nitidissima powder to distilled water at 60 °C, mix and stir for 20 min, add glucose powder as a carbon source, stir and homogenize, then put it into a water bath at 90 °C for sterilization for 30 min to obtain a fermentation medium; each liter of the fermentation medium contains 10 parts of camellia nitidissima powder and 2.5 parts of glucose powder, and each part is 10 g;

[0033] After activating the bacillus, inoculate it into the fermentation medium, and the inoculation density is 5.5×10 6 cfu / mL, ferment at 30 °C for 20 hours, centrifuge at 6000 rpm for 10 min, take the supernatant, pass through a 0.22-μm filter membrane, collect the filtrate, and freeze-dry and concentrate to prepare the first component;

[0034] 2) Inoculate the activated Bacillus into a culture medium for fermentation. The inoculation density is 5.6×10 6 cfu / mL. After fermentation, homogenize the bacterial cells, ultrasonically disrupt them. The ultrasonic power is 1000 w, the time is 20 min, working for 3 s and intermittent for 3 s. Centrifuge to obtain the supernatant, and lyophilize and concentrate it to prepare the second component;

[0035] 3) Preparation of the cell energy activator: Mix the first component and the second component, and add 3 - 5 parts of nicotinamide and 0.5 - 1.5 parts of VC ethyl ether to prepare the cell energy activator.

[0036] Comparative Example 1 - 1

[0037] Based on Example 1, in step 1), 5 parts of Camellia nitidissima powder and 2.5 parts of glucose powder are used to obtain the fermentation medium, and the rest remains unchanged.

[0038] Comparative Example 1 - 2

[0039] Based on Example 1, in step 1), 15 parts of Camellia nitidissima powder and 2.5 parts of glucose powder are used to obtain the fermentation medium, and the rest remains unchanged.

[0040] Comparative Example 1 - 3

[0041] Based on Example 1, in step 1), 10 parts of Camellia nitidissima powder and 5 parts of glucose powder are used to obtain the fermentation medium, and the rest remains unchanged.

[0042] Comparative Example 1 - 4

[0043] Based on Example 1, in step 1), 15 parts of Camellia nitidissima powder and 5 parts of glucose powder are used to obtain the fermentation medium, and the rest remains unchanged.

[0044] Determine the antioxidant activities of Example 1 and Comparative Examples 1 - 4. The determination method is carried out according to the instructions of the T - AOC determination kit (Nanjing Jiancheng Bioengineering Institute). At 37 °C, when the absorbance (OD) value of the reaction system increases by 0.01 per minute per milliliter of serum (plasma), it is one unit of total antioxidant capacity (U). The determination results are shown in Table 1.

[0045] Table 1 Determination results of antioxidant activities

[0046]

[0047] It can be seen from the results in Table 1 that the total antioxidant activity of the components after fermentation of the fermentation medium prepared in Example 1 is superior to that of Comparative Examples 1 - 1 to 1 - 4. The possible reason is that in the case of a single carbon source, glucose is used as the carbon source for microbial fermentation, but excessive glucose will cause substrate repression. Therefore, the formulation method of Example 1 is selected for subsequent strain inoculation and screening tests.

[0048] Comparative Examples 1-5

[0049] Based on Example 1, in step 1), after activating the Bacillus, it was inoculated into the fermentation medium, and the inoculation density was 5.0×10 6 cfu / mL, and the rest remained unchanged.

[0050] Comparative Examples 1-6

[0051] Based on Example 1, in step 1), after activating the Bacillus, it was inoculated into the fermentation medium, and the inoculation density was 6.0×10 6 cfu / mL, and the rest remained unchanged.

[0052] Comparative Examples 1-7

[0053] Based on Example 1, in step 1), after activating the Bacillus, it was inoculated into the fermentation medium, and the inoculation density was 6.5×10 6 cfu / mL, and the rest remained unchanged.

[0054] Comparative Examples 1-8

[0055] Based on Example 1, in step 1), after activating the Bacillus, it was inoculated into the fermentation medium, and the inoculation density was 7.0×10 6 cfu / mL, and the rest remained unchanged.

[0056] Refer to the foregoing method to further measure the total antioxidant activity of the components after fermentation, and the measurement results are shown in Table 2.

[0057] Table 2 Total Antioxidant Activity

[0058]

[0059] As can be seen from the results in Table 2, the total antioxidant activity of the components after fermentation of the fermentation medium prepared in Example 1 is better than that of Comparative Examples 1-5 to 1-8. Therefore, the preparation method of Example 1 was selected for the single-factor screening test of the subsequent fermentation temperature and fermentation time, and finally the optimal conditions of an inoculation density of 5.5×10 6 cfu / mL and fermentation at 30°C for 20 hours were determined.

[0060] Example 2

[0061] A preparation method of a cell energy activator, comprising the following steps:

[0062] 1) Dry and pulverize the camellia nitidissima, sieve it through a 140-mesh sieve to obtain camellia nitidissima powder; add the camellia nitidissima powder to distilled water at 60 °C, mix and stir for 20 min, add glucose powder as a carbon source, stir and homogenize, then place it in a water bath, sterilize at 90 °C for 30 min to obtain a fermentation medium; each liter of the fermentation medium contains 10 parts of camellia nitidissima powder and 2.5 parts of glucose powder, with each part being 10 g;

[0063] After activating the Bacillus, inoculate it into the fermentation medium, with an inoculation density of 5.6×10 6 cfu / mL, ferment at 30 °C for 20 hours, centrifuge at 6000 rpm for 10 min, take the supernatant, filter it through a 0.22-μm filter membrane, and collect the filtrate, then freeze-dry and concentrate to prepare the first component;

[0064] 2) Inoculate the activated Bacillus into YPD medium for fermentation, with an inoculation density of 5.6×10 6 cfu / mL. After fermentation, homogenize the bacterial cells, ultrasonically disrupt them, with an ultrasonic power of 1000 w, for 20 min, working for 3 s and pausing for 3 s, centrifuge and take the supernatant, then freeze-dry and concentrate to prepare the second component;

[0065] 3) Preparation of the cell energy activator: Mix the first component and the second component, and add 3 - 5 parts of nicotinamide and 0.5 - 1.5 parts of VC ethyl ether to prepare the cell energy activator.

[0066] Comparative Example 2-1

[0067] Based on Example 2, in step 2), the inoculation density is 2.8×10 6 cfu / mL, and the rest remains unchanged.

[0068] Comparative Example 2-2

[0069] Based on Example 2, in step 2), the inoculation density is 5.0×10 6 cfu / mL, and the rest remains unchanged.

[0070] Comparative Example 2-3

[0071] Based on Example 2, in step 2), the inoculation density is 6.0×10 6 cfu / mL, and the rest remains unchanged.

[0072] Comparative Example 2-4

[0073] Based on Example 2, in step 2), the inoculation density is 7.0×10 6 cfu / mL, and the rest remains unchanged.

[0074] Comparative Example 2-5

[0075] On the basis of Example 2, in step 2), the inoculation density is 8.4×10 6 cfu / mL, and the rest remains unchanged.

[0076] The postbiotic yield is expressed by the cell density in the fermentation broth at the end of fermentation. The specific operation method is carried out with reference to the national standard GB4789.15-2016, and the measurement results are shown in Table 3.

[0077] Table 3 Colony counting after Bacillus fermentation

[0078]

[0079] It can be seen from the results in Table 3 that the fermented bacteria obtained from the inoculation density of Bacillus used in Example 1 are better than those in Comparative Examples 2-1 to 2-5. Therefore, the inoculation dose of Example 1 was selected for the single-factor screening test of subsequent cell lysis. Finally, 5.6×10 6 cfu / mL was selected. After fermentation, the cells were homogenized and ultrasonically broken. The ultrasonic power was 1000w, the time was 20min, and the working mode of working for 3s and intermittent for 3s could obtain the most.

[0080] Example 3

[0081] The formulation test of the compound composition was carried out with the components obtained according to the steps described in Example 1

[0082] A preparation method of a cell energy activator includes the following steps:

[0083] 1) Dry and crush Camellia nitidissima, sieve it through a 140-mesh sieve to obtain Camellia nitidissima powder; add the Camellia nitidissima powder to distilled water at 60°C, mix and stir for 20 min, add glucose powder as a carbon source, stir and homogenize, and then put it into a water bath at 90°C for sterilization for 30 min to obtain a fermentation medium; each liter of the fermentation medium contains 10 parts of Camellia nitidissima powder, 2.5 parts of glucose powder, and each part is 10 g;

[0084] After activating Bacillus, inoculate it into the fermentation medium, and the inoculation density is 5.6×10 6 cfu / mL, ferment at 30°C for 20 hours, centrifuge at 6000 rpm for 10 min, take the supernatant, filter it through a 0.22-μm filter membrane, and then collect the filtrate, freeze-dry and concentrate it to prepare the first component;

[0085] 2) Inoculate the activated Bacillus into the YPD medium for fermentation, and the inoculation density is 5.6×10 6 cfu / mL. After fermentation, homogenize the cells, ultrasonically break them, the ultrasonic power is 1000w, the time is 20min, the working mode is working for 3s and intermittent for 3s, centrifuge and take the supernatant, freeze-dry and concentrate it to prepare the second component;

[0086] 3) Preparation of cell energy activator: Mix the first component and the second component, and add 3.5 parts of nicotinamide and 1.0 part of VC ethyl ether to prepare the cell energy activator.

[0087] Comparative Example 3-1

[0088] On the basis of Example 3, in step 3), add 3 parts of nicotinamide and 0.5 part of VC ethyl ether, and the rest remains unchanged.

[0089] Comparative Example 3-2

[0090] On the basis of Example 3, in step 3), add 3 parts of nicotinamide and 1.5 parts of VC ethyl ether, and the rest remains unchanged.

[0091] Comparative Example 3-3

[0092] On the basis of Example 3, in step 3), add 3.5 parts of nicotinamide and 0.5 part of VC ethyl ether, and the rest remains unchanged.

[0093] Comparative Example 3-4

[0094] On the basis of Example 3, in step 3), add 3.5 parts of nicotinamide and 1.0 part of VC ethyl ether, and the rest remains unchanged.

[0095] Comparative Example 3-5

[0096] On the basis of Example 3, in step 3), add 3.5 parts of nicotinamide and 1.5 parts of VC ethyl ether, and the rest remains unchanged.

[0097] Comparative Example 3-6

[0098] On the basis of Example 3, in step 3), add 4.0 parts of nicotinamide and 0.5 part of VC ethyl ether, and the rest remains unchanged.

[0099] Comparative Example 3-7

[0100] On the basis of Example 3, in step 3), add 4.0 parts of nicotinamide and 1.0 part of VC ethyl ether, and the rest remains unchanged.

[0101] Comparative Example 3-8

[0102] On the basis of Example 3, in step 3), add 4.0 parts of nicotinamide and 1.5 parts of VC ethyl ether, and the rest remains unchanged.

[0103] Comparative Example 3-9

[0104] On the basis of Example 3, in step 3), add 0 part of nicotinamide and 1.0 part of VC ethyl ether, and the rest remains unchanged.

[0105] Comparative Example 3-10

[0106] On the basis of Example 3, in step 3), 3.5 parts of niacinamide and 0 part of VC ethyl ether are added, and the rest remains unchanged.

[0107] Anti-glycation assay. High glucose is one of the factors that cause the accumulation of AGEs (Advanced Glycation Endproducts) in cells, thereby inducing glycation damage to cells. High glucose can trigger inflammation, exacerbate the binding of AGE-RAGE and activate the glycation reaction, and promote cell apoptosis. This example is used to evaluate the effect of the compound composition obtained according to the steps described in Example 1 on the glycation of skin cells. The specific steps are as follows:

[0108] Cell preparation: Human skin fibroblast cells (HSF) were purchased from the Kunming Cell Bank of the Chinese Academy of Sciences. 89% DMEM medium + 10% fetal bovine serum + 1% double antibody was used as the medium, and HSFs were cultured with passage every 3 days. They were inoculated into 6-well plates at a density of 1.0×10 5 cells / mL. After 24 hours at 37°C, a blank group, a model group, and an experimental group were set up. Among them, the blank group was replaced with a new medium, and the model group and the experimental group were replaced with a high-glucose medium (a medium containing 30 mM glucose). After the experiment started, PBS was added to the blank group as a control, the high-glucose medium was added to the model group, and the test sample was added to the experimental group, 150 μL each, for 14 days continuously.

[0109] During the test, the in vitro AGEs generation inhibition rate was determined with reference to the method disclosed by Eze et al. PBS was used as the blank for sample addition. 0.2 mol / L PBS (pH 7.4) was used as the solvent to dissolve BSA and D-fructose, so that the BSA concentration reached 10 mg / mL and the D-fructose concentration was 0.5 mol / L. After filtration and sterilization, it was used as the reaction solution. The sample of the blank group was a mixture of PBS and the reaction solution, and the samples of the model group and the experimental group were mixtures of the test sample and the reaction solution. They were placed in a constant-temperature incubator at 37°C and incubated in the dark for 14 days. At the end of the incubation, the fluorescence intensity was measured with an enzyme-linked immunosorbent assay (ELISA) reader (set the fluorescence intensity at the excitation wavelength of 370 nm and the emission wavelength of 440 nm, slit 5 nm). The inhibition rate calculation formula is as follows:

[0110] Inhibition rate % = 1 - (F1 - F2) / (F3 - F2), where F1 represents the fluorescence value of the experimental group, F2 represents the fluorescence value of the blank group, and F3 represents the fluorescence value of the model control group. The measurement results are shown in Table 4.

[0111] Table 4 AGEs determination inhibition rate

[0112] Group Inhibition rate (%) Example 3 75.2 Comparative Example 3-1 55.1 Comparative Example 3-2 64.0 Comparative Example 3-3 60.4 Comparative Example 3-4 73.3 Comparative Example 3-5 65.6 Comparative Example 3-6 68.9 Comparative Example 3-7 71.4 Comparative Example 3-8 69.5 Comparative Example 3-9 34.1 Comparative Example 3-10 47.5

[0113] As can be seen from the results in Table 4, the metabolites of fermented Camellia nitidissima by Bacillus have different anti-glycation abilities, and after the addition of niacinamide and VC ethyl ether, they have a good synergistic effect. Except for Comparative Example 3-1, Comparative Example 3-9, and Comparative Example 3-10, the inhibition rates of in vitro synergistic inhibition of AGEs generation all reach more than 60%; and when the component ratio of adding 3.5 parts of niacinamide and 1.0 part of VC ethyl ether is used, the in vitro synergistic inhibition of AGEs generation has the best effect.

[0114] Example 4

[0115] A large number of studies have shown that long-term and large-dose injection of D-galactose solution can produce excessive reactive oxygen species (ROS), reduce the activities of antioxidant enzymes in various organs, form more superoxide anions and various oxidation products, etc., cause cell damage, and lead to the decline of the functions of multiple organs and systems in the body. Therefore, in this study, a D-galactose-induced aging mouse model combined with UV light irradiation was used to determine the anti-aging effect of the composition.

[0116] On the basis of the cell energy activator (named M, with glycerol as the solvent) obtained in Example 3, propylene glycol and deionized water were added, and after mixing and preparation, an experimental reagent was obtained. The preparation components are shown in Table 5.

[0117] Table 5 Reagent Preparation Table

[0118]

[0119] Twenty 7-week-old SPF male BALB / c mice (purchased from Guangdong Provincial Medical Experimental Animal Center) were selected and randomly divided into 5 groups (control group, aging group, high-dose experimental group, medium-dose experimental group, low-dose experimental group), with 4 mice in each group. Veet hair removal cream was used to remove the hair on the back skin of the mice. The control group was subcutaneously injected with 0.3 mL of normal saline daily, and the aging group and the experimental groups were subcutaneously injected with D-galactose (100 mg / kg / bw) daily. After continuous feeding for 4 weeks, each group was irradiated on the back of the mice with a UVB lamp with a radiation dose of 80 mJ / cm 2 , and the irradiation was performed every other day for 1 hour each day for 4 weeks. During the experiment, the appearance of the aging and experimental group mice was monitored, and as the experiment progressed, the changes in the back skin of the mice in each group were compared. The specific experimental grouping and treatment methods are shown in Table 6.

[0120] Table 6 Experimental Grouping and Treatment Methods

[0121]

[0122] After the experiment, the mice were euthanized. Skin tissues were excised to prepare 10% skin tissue homogenate. The sample extract was placed in a high-speed low-temperature tissue grinder for homogenization to prepare skin tissue homogenate (homogenization frequency: 100 Hz, temperature: -10 °C). The homogenate was centrifuged at 8000 rpm, 4 °C for 30 min, and the supernatant was taken for subsequent determination. The activities of catalase (CAT), superoxide dismutase (SOD), and the content of trace reduced glutathione (GSH) in the skin tissue were determined according to the instructions of the commercial detection kit. The results are shown in Table 7.

[0123] Table 7 Antioxidant determination of skin tissue

[0124]

[0125] As can be seen from the results in Table 7, taking the senescence group as the control, the activities of CAT, SOD, and GSH in the skin of the medium- and high-dose model mice were significantly increased, and each oxidation index was close to that of the control group.

[0126] In this specification, each embodiment is described in a progressive manner. The key point of each embodiment is to illustrate the differences from other embodiments. The same or similar parts among the embodiments can be referred to each other. The above description of the disclosed embodiments enables those skilled in the art to implement or use the present invention. Various modifications to these embodiments will be obvious to those skilled in the art. The general principles defined herein can be implemented in other embodiments without departing from the spirit or scope of the present invention. Therefore, the present invention will not be limited to these embodiments shown herein, but will conform to the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. A method for preparing a cell energy activator, characterized in that It includes the following steps: 1) Dry and crush camellia nitidissima, sieve it through a 100-150 mesh sieve to obtain camellia nitidissima powder; add the camellia nitidissima powder into distilled water at 60 °C, mix and stir for 20 min, add glucose powder as a carbon source, stir and homogenize, then put it into a water bath, sterilize at 90-100 °C for 30 min to obtain a fermentation medium; each liter of the fermentation medium contains 5-15 parts of camellia nitidissima powder, 2.5-5 parts of glucose powder, and each part is 10 g; After activating the Bacillus, inoculate it into the fermentation medium at an inoculation density of 5.0 - 6.0×10 6 cfu / mL, ferment at 28 - 33°C for 18 - 24 hours, centrifuge at 6000 rpm for 10 min, take the supernatant, filter through a 0.22 μm filter membrane, collect the filtrate, and concentrate by freeze-drying to prepare the first component; 2) Inoculate the activated bacillus subtilis into the medium for fermentation. After fermentation, homogenize the thalli, ultrasonically crush them, with an ultrasonic power of 800-1000 w, for a time of 15-20 min, working for 3-5 s and intermittent for 3-5 s, centrifuge to take the supernatant, freeze-dry and concentrate to prepare the second component; 3) Composition preparation: Mix the first component and the second component, and add niacinamide and VC ethyl ether to prepare a cell energy activator.

2. The method according to claim 1, wherein The content of camellia nitidissima powder is 10 parts, and the content of glucose powder is 2.5 parts.

3. The method according to claim 1 or 2, characterized in that, The inoculation density is 5.5×10 6 cfu / mL.

4. The method according to claim 3, characterized in that, In step 3), add 3-5 parts of niacinamide and 0.5-1.5 parts of VC ethyl ether.

5. A cell energy activator prepared by the method according to claim 1 or 2.

6. A skin care product, characterized in that, It includes the cell energy activator according to claim 5.

7. The skin care product according to claim 6, wherein, It also includes at least one of a humectant, an anti-inflammatory agent, and a chelating agent.

8. Use of the cell energy activator according to claim 5 in the preparation of a skin antioxidant product.

9. The use according to claim 8, characterized in that, The application object is male.

10. Use of the cell energy activator according to claim 5 in the preparation of a product for enhancing the activity of catalase.

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