Preparation process of double-bacterium fermented black tea filtrate as well as product and application of double-bacterium fermented black tea filtrate

Abstract

The invention relates to the field of biological fermentation, in particular to a preparation process of double-bacterium fermented black tea filtrate as well as a product and application thereof. The preparation method of the double-bacterium fermented black tea filtrate comprises the following specific preparation steps: S1, preparing black tea leaf powder; s2, preparing a fermentation culture medium; s3, preparing a seed solution; s4, inoculating, fermenting and culturing; s5, collecting filtrate to obtain a fermentation product; the seed solution in the step S3 comprises an acetobacter xylinum seed solution and a saccharomycetes seed solution. The black tea is subjected to double-bacterium fermentation by adopting acetobacter xylinum and saccharomycetes, and substances such as cell walls and cellulose in black tea plant cells can be effectively degraded, so that the intracellular extraction resistance is reduced, the dissolution of active components (such as tea polyphenol and amino acid) in the black tea is greatly improved, and the extraction rate of functional components is increased. The obtained double-bacterium fermented black tea filtrate has excellent anti-wrinkle, firming, anti-aging, moisturizing, soothing and other effects.

Description

Technical Field

[0001] The invention relates to the field of biological fermentation, and in particular to a preparation process of double-bacteria fermented black tea filtrate, and a product and application thereof. Background Art

[0002] With the improvement of national economy and living standards, consumers have higher and higher requirements for cosmetics, but some core cosmetic raw materials in the industry are monopolized by international manufacturers; the development of new cosmetic raw materials and the exploration of their efficacy have become difficult problems in the cosmetics field.

[0003] Relying on China's unique plant resources, plant extracts are a popular way to explore raw materials in the cosmetics field. However, due to the structural peculiarities of plants, plant extraction still has problems such as cumbersome extraction processes, low content of extracted active substances, and poor stability of effective ingredients. In recent years, the development of microbial fermentation technology has provided a new idea for cosmetic raw materials. By using biological enzymes contained in microorganisms, effective substances in plants can be deeply extracted to effectively retain active ingredients. However, the biological activity of microorganisms themselves also determines the inevitable instability of microbial fermentation technology. During the microbial fermentation process, it is necessary to control the stability of the strain and the stability of the extracted ingredients. This is still difficult to achieve at the current technical level, which makes the existing microbial fermentation technology still have problems such as low content of active ingredients, unsignificant effects, insufficient efficacy, and difficulty in mass production.

[0004] Chinese patent CN115569096B discloses a camellia fermentation filtrate and a fermentation process, wherein camellia is fermented and cultured by lactic acid bacteria to finally obtain a camellia fermentation filtrate with significant antioxidant properties. Chinese patent CN116211766A discloses a black tea fermentation product and its application, wherein black tea is fermented by at least one of yeast, acetic acid bacteria, and lactobacillus to obtain a black tea fermentation product with strong repairing effect. Chinese patent CN115105452B discloses a black tea fermentation filtrate and its preparation method and application, to obtain a black tea fermentation filtrate with anti-aging, anti-inflammatory and antioxidant properties, but the technology involves subculture, has extremely high process requirements, and is of poor practicality. None of these prior arts effectively solve the above technical problems.

[0005] Based on this, developing a fermentation raw material that is easy to implement, has significant efficacy, and has good formula compatibility has become an urgent technical issue. Summary of the invention

[0006] The first aspect of the present invention provides a process for preparing a double-bacteria fermented black tea filtrate, wherein the specific steps of preparing the double-bacteria fermented black tea filtrate include: S1. Preparation of black tea powder; S2. preparing fermentation medium; S3. Preparing seed solution; S4. Inoculation and fermentation culture; S5. collecting the filtrate to obtain a fermentation product; Preferably, the seed solution in step S3 includes Acetobacter xylinum seed solution and yeast seed solution.

[0007] The present invention has been found through research that dual-bacteria fermentation of black tea (Camellia sinensis) using Acetobacter Xylinus (scientific name Gluconacetobacter Xylinus) and yeast (scientific name Saccharomyces) can effectively degrade cell walls, cellulose and other substances in black tea plant cells, thereby reducing intracellular extraction resistance, greatly improving the dissolution of active ingredients in black tea (such as tea polyphenols, amino acids, etc.), and increasing the extraction rate of effective ingredients.

[0008] In some preferred embodiments, the step S1 comprises: crushing black tea leaves, grinding to obtain black tea coarse powder, and sieving the black tea coarse powder to obtain black tea powder.

[0009] Preferably, the sieving in step S1 is specifically through a 20-100 mesh sieve; more preferably, through a 40 mesh sieve.

[0010] In some preferred embodiments, the black tea leaves are derived from at least one of Dian black tea, Keemun black tea, Yingde black tea, Ning black tea, and Sichuan black tea.

[0011] Preferably, the black tea leaves are black tea leaves from ancient trees (a type of Dian black tea).

[0012] The present invention selects ancient tree black tea leaves and cooperates with specific microbial fermentation process technology to effectively retain the active ingredients in the black tea. The final double-bacteria fermented black tea filtrate has excellent moisturizing, soothing, anti-aging and firming effects. The obtained black tea filtrate can be well applied to skin care products to solve the problems of low bioavailability and unsatisfactory skin care effect.

[0013] In some preferred embodiments, the components of the fermentation medium in step S2 include: 10-50 g / L black tea powder, 20-36 g / L glucose, 1-5 g / L peptone, 0.5-3 g / L yeast extract, and the solvent is water; mix, adjust the pH to 6.0-7.0, and sterilize to obtain.

[0014] Further preferably, the components of the fermentation medium in step S2 include: 30 g / L black tea powder, 30 g / L glucose, 2 g / L peptone, 2 g / L yeast extract, and the solvent is water; the mixture is mixed and sterilized to obtain the product.

[0015] In some preferred embodiments, the method for preparing the Acetobacter xylinum seed solution comprises: inoculating the freeze-dried powder of Acetobacter xylinum into a first culture medium, activating and culturing for 2-6 days to obtain an activated Acetobacter xylinum strain; inoculating the activated Acetobacter xylinum strain into a second culture medium, and performing secondary culturing to obtain the Acetobacter xylinum seed solution.

[0016] Preferably, the secondary culture is carried out under the following conditions: 20-35°C, 100-200 rpm for 1 day, and the culture is carried out under aerobic conditions.

[0017] Further preferably, the secondary culture is carried out at 28° C. and 130 rpm for 1 day under aerobic conditions.

[0018] In some preferred embodiments, the components of the first culture medium include, by weight percentage: 0.1-4% yeast extract, 1-5% glucose, 0.8-3% agar, and the balance is supplemented by water; the raw materials are mixed according to the formula, the pH is controlled at 6.6±0.2, and sterilized to obtain the product.

[0019] In some preferred embodiments, the composition of the second culture medium (YGC liquid culture medium) is similar to that of the first culture medium, except that agar is not added to the second culture medium, and the balance is made up with water.

[0020] In some preferred embodiments, the inoculation amount of the Acetobacter xylinum strain is 1-5wt%, preferably 3wt%.

[0021] The Acetobacter xylinum freeze-dried powder can be commercially available, for example, from Beijing Kezhan Biotechnology Co., Ltd.

[0022] In some preferred embodiments, the method for preparing the yeast seed solution comprises: inoculating the yeast into a third culture medium, performing activation culture for 1-3 days, and obtaining an activated yeast strain; inoculating the activated yeast strain into a fourth culture medium, and performing secondary culture to obtain the yeast seed solution.

[0023] Preferably, the secondary culture is carried out under the following conditions: 25-40°C, 100-300 rpm for 1 day, and the culture is carried out under aerobic conditions.

[0024] Preferably, the secondary culture is carried out under the conditions of: culturing at 30° C. and 160 rpm for 1 day under aerobic conditions.

[0025] In some preferred embodiments, the components of the third culture medium include, by weight percentage: 0.1-3% tryptone, 1-5% glucose, 0.1-1.5% yeast extract powder, 0.5-3% agar, and the balance is supplemented by water; the raw materials are mixed according to the formula and sterilized to obtain the product.

[0026] In some preferred embodiments, the composition of the fourth culture medium (YPD liquid culture medium) is similar to that of the third culture medium, except that agar is not added to the fourth culture medium, and the balance is made up with water.

[0027] In some preferred embodiments, the inoculation amount of the yeast strain is 0.5-3wt%, preferably 1wt%.

[0028] The yeast is specifically obtained by single purification from beer residue; the yeast originates from Guangzhou Junyan Biotechnology Co., Ltd., the biological preservation number is GDMCC NO: 64857, the biological preservation date is July 10, 2024, the biological preservation address is Guangdong Provincial Microbiological Culture Collection Center, and the preservation unit is Guangzhou Junyan Biotechnology Co., Ltd.

[0029] The present invention performs single purification and separation of beer residues through specific strain screening, and finally screens out a unique yeast strain. The obtained yeast can cooperate with Acetobacter xylinum to maximize the fermentation culture effect, and further cooperates with specific process parameters to enable the culture process to effectively maintain strain stability and process stability, and finally achieve the stability of the functional components.

[0030] Note: The sterilization conditions in steps S2 and S4 are both: high temperature sterilization at 121°C for 15 minutes.

[0031] In some preferred embodiments, the inoculation and fermentation culture in step S4 is specifically a two-step fermentation, including step (1): inoculating Acetobacter xylinum seed liquid into a fermentation medium for fermentation culture; step (2): inoculating yeast seed liquid into the fermentation culture liquid obtained in step (1) for fermentation culture.

[0032] Preferably, the fermentation conditions of step (1) are: fermentation culture at 20-35°C and 100-200 rpm for 10-24 hours; the fermentation culture is carried out under aerobic conditions.

[0033] In some preferred embodiments, the inoculation amount of Acetobacter xylinum seed solution in step (1) is 1-10wt%, preferably 5wt%.

[0034] Preferably, the fermentation conditions of step (2) are 25-35° C. and 120-240 rpm for 1-10 h.

[0035] In some preferred embodiments, the inoculation amount of the yeast seed solution in step (2) is 0.5-4wt%, preferably 1wt%.

[0036] In some preferred embodiments, the step S5 comprises: centrifuging and filtering the fermentation culture product obtained in step S4, and pasteurizing the product to obtain an Acetobacter xylinum / yeast black tea fermentation filtrate product.

[0037] Preferably, the centrifugation condition in step S5 is 6000-20000 rpm.

[0038] The fermentation method of the present invention is simple and easy to implement; only a short fermentation time is required to obtain a black tea fermentation filtrate with a high content of active ingredients; and the black tea fermentation filtrate can take into account multiple skin care effects such as moisturizing, soothing, firming, anti-wrinkle, anti-aging, etc.; it can achieve large-scale production, which is conducive to promotion in the field of cosmetics.

[0039] The second aspect of the present invention provides a double-bacteria fermented black tea filtrate prepared by the above-mentioned preparation process.

[0040] In some preferred embodiments, the elastase inhibition rate of the double-bacteria fermented black tea filtrate is greater than 18%.

[0041] The third aspect of the present invention provides an application of a double-bacteria fermented black tea filtrate, wherein the double-bacteria fermented black tea filtrate is applied to cosmetics; the cosmetics include any one of skin care water, essence, emulsion, face cream, ointment, facial mask liquid, and facial cleanser.

[0042] In some preferred embodiments, the double-bacteria fermented black tea filtrate is applied to the essence.

[0043] Preferably, in terms of weight percentage, the components of the essence include: 10-60% of double-bacteria fermented black tea filtrate, 5-35% of auxiliary agent, and the balance of water.

[0044] The auxiliary agent can be selected according to the dosage form requirements, and can be listed as a thickener, a skin conditioner, a moisturizer, a preservative, etc.

[0045] The skin conditioning agents include vitamins and their derivatives, bis-PEG-18 methyl ether dimethyl silane, amino acids, polypeptides, polysaccharides, niacinamide, astaxanthin, plant extracts, and the like.

[0046] The moisturizing agents include small molecule alcohols, squalane, hyaluronic acid and its salts, ceramides, plant essential oils and the like.

[0047] Examples of thickeners include, but are not limited to, carbomer and xanthan gum.

[0048] Examples of small molecule alcohols include, but are not limited to, glycerol, butanediol, and 1,2-hexanediol.

[0049] Examples of preservatives include, but are not limited to, p-hydroxyacetophenone.

[0050] The double-bacteria fermented black tea filtrate provided by the present invention has a perfume-like smell and retains the fragrance even after being left to stand for a long time; it has excellent formula compatibility and can be well applied to skin care products such as essences, creams, and face creams; especially when applied to essences, it has the effects of endogenously enhancing the youthfulness of the skin, exogenously building a skin care barrier, and fundamentally nourishing the skin.

[0051] Beneficial effects: The present invention provides a preparation process of a double-bacteria fermented black tea filtrate, a product thereof and an application thereof, which have the following advantages: (1) The present invention uses Acetobacter xylinum and yeast to carry out dual-bacteria fermentation of black tea, which can effectively degrade cell walls, cellulose and other substances in black tea plant cells, thereby reducing intracellular extraction resistance, greatly improving the dissolution of active ingredients in black tea (such as tea polyphenols, amino acids, etc.), and increasing the extraction rate of effective ingredients.

[0052] (2) The present invention selects ancient tree black tea leaves and combines them with specific microbial fermentation process technology to effectively retain the active ingredients in the black tea. The resulting double-bacteria fermented black tea filtrate has excellent moisturizing, soothing, anti-aging and firming effects. The resulting black tea filtrate can be well applied to skin care products to solve the problems of low bioavailability and unsatisfactory skin care effects. It can act on the skin to nourish the skin from the inside out and fundamentally achieve the purpose of skin care.

[0053] (3) The present invention screens specific strains to perform single purification and separation of beer residues, and ultimately screens out a unique yeast strain. The obtained yeast can cooperate with Acetobacter xylinum to maximize the fermentation culture effect. Further, by combining specific process parameters, the culture process can effectively maintain strain stability and process stability, and ultimately achieve the stability of the effective ingredients.

[0054] (4) The fermentation method of the present invention is simple and easy to implement. It only takes a short fermentation time to obtain a black tea fermentation filtrate with a high content of active ingredients. The black tea fermentation filtrate can also have multiple skin care effects such as moisturizing, soothing, firming, anti-wrinkle, and anti-aging. It can be mass-produced, which is conducive to promotion in the cosmetics field.

[0055] (5) The double-bacteria fermented black tea filtrate provided by the present invention has a perfume-like smell and retains the fragrance after being left to stand for a period of time; it also has excellent formula compatibility and can be well applied to skin care products such as essences, creams, and face creams. BRIEF DESCRIPTION OF THE DRAWINGS

[0056] Figure 1 Example 1 Flow chart of the anti-wrinkle and firming efficacy test of the double-bacteria fermented black tea filtrate; Figure 2Example 1 Flow chart of moisturizing detection of double-bacteria fermented black tea filtrate; Figure 3 Example 1 Flow chart of soothing detection of double-bacteria fermented black tea filtrate; Figure 4 . VISIA test results of user 1 using the essence of Example 1; Figure 5 . VISIA test results of user 2 using the essence of Example 1; Figure 6 .VISIA test results of user 3 using the essence of Example 1. DETAILED DESCRIPTION

[0057] Note: There is no particular limitation on the oxygen content of the aerobic environment in the examples.

[0058] Example 1. A preparation process of a double-bacteria fermented black tea filtrate, wherein the specific preparation steps of the double-bacteria fermented black tea filtrate include: S1. Preparation of black tea powder; S2. preparing fermentation medium; S3. Preparing seed solution; S4. Inoculation and fermentation culture; S5. collecting the filtrate to obtain a fermentation product; The seed solution in step S3 includes Acetobacter xylinum seed solution and yeast seed solution.

[0059] The step S1 comprises: crushing and grinding the sun-dried black tea leaves with a crusher to obtain black tea coarse powder, and then sifting the black tea coarse powder with a 40-mesh tea sieve to obtain black tea powder.

[0060] The black tea leaves are specifically ancient tree black tea leaves (a type of Dian black tea).

[0061] The components of the fermentation medium in step S2 include: 30 g / L black tea powder, 30 g / L glucose, 2 g / L peptone, 2 g / L yeast extract, and the solvent is water; the mixture is mixed, the pH is adjusted to 6.5±0.5, and sterilized to obtain the product.

[0062] The preparation method of the Acetobacter xylinum seed liquid comprises: inoculating the freeze-dried powder of Acetobacter xylinum into a first culture medium, activating and culturing for 4 days to obtain an activated Acetobacter xylinum strain; inoculating the activated Acetobacter xylinum strain into a second culture medium, and performing secondary culturing to obtain the Acetobacter xylinum seed liquid.

[0063] The secondary culture conditions for preparing the Acetobacter xylinum seed solution are: culture at 28° C. and 130 rpm for 1 day, and the culture is carried out under aerobic conditions.

[0064] In terms of weight percentage, the first culture medium comprises: 0.5% yeast extract, 2% glucose, 1.5% agar, and the balance is supplemented by water; the raw materials are mixed according to the formula, the pH is controlled at 6.6±0.2, and sterilized to obtain the product.

[0065] The composition of the second culture medium (YGC liquid culture medium) is similar to that of the first culture medium, except that agar is not added to the second culture medium and the balance is made up with water.

[0066] The inoculation amount of the Acetobacter xylinum strain is 3wt%; the freeze-dried Acetobacter xylinum powder comes from Beijing Kezhan Biotechnology Co., Ltd.

[0067] The method for preparing the yeast seed liquid comprises: inoculating yeast into a third culture medium, performing activation culture for 2 days, and obtaining an activated yeast strain; inoculating the activated yeast strain into a fourth culture medium, performing secondary culture, and obtaining the yeast seed liquid.

[0068] The secondary culture conditions for preparing the yeast seed solution are: culture at 30° C. and 160 rpm for 1 day, and the culture is carried out under aerobic conditions.

[0069] In terms of weight percentage, the third culture medium comprises: 1% tryptone, 2% glucose, 0.5% yeast extract powder, 1.4% agar, and the balance is supplemented by water; the raw materials are mixed according to the formula and sterilized to obtain the third culture medium.

[0070] The composition of the fourth culture medium (YPD liquid culture medium) is similar to that of the third culture medium, except that agar is not added to the fourth culture medium, and the balance is supplemented with water.

[0071] The inoculation amount of the yeast strain is 1 wt %.

[0072] The yeast is specifically obtained by single purification from beer residue; the yeast comes from Guangzhou Junyan Biotechnology Co., Ltd., and the biological preservation number is GDMCC NO: 64857.

[0073] The sterilization conditions in steps S2 and S4 are both: treatment at 121° C. for 15 minutes for high temperature sterilization.

[0074] The inoculation fermentation culture in step S4 is specifically a two-step fermentation, comprising step (1): inoculating the seed liquid of Acetobacter xylinum into the fermentation medium for fermentation culture; and step (2): inoculating the seed liquid of Saccharomyces cerevisiae into the fermentation liquid obtained in step (1) for fermentation culture.

[0075] The fermentation conditions of step (1) are: fermentation at 30° C. and 130 rpm for 16 h; the fermentation is carried out under aerobic conditions.

[0076] In the step (1), the inoculation amount of the Acetobacter xylinum seed solution is 5 wt %.

[0077] The fermentation culture conditions of step (2) are 28° C. and 160 rpm for 5 hours.

[0078] The inoculation amount of the yeast seed solution in step (2) is 1 wt %.

[0079] The step S5 comprises: centrifuging and filtering the fermentation culture product obtained in the step S4, and pasteurizing (80° C., 40 min) to obtain an Acetobacter xylinum / yeast black tea fermentation filtrate product.

[0080] The centrifugation condition in step S5 is 12000 rpm.

[0081] A double-bacteria fermented black tea filtrate is prepared by the above preparation process.

[0082] An application of a double-bacteria fermented black tea filtrate, wherein the double-bacteria fermented black tea filtrate is applied to an essence; the formula of the essence is shown in Table 1 below in terms of weight percentage.

[0083] Table 1

[0084] The preparation method of the essence comprises: a. Mix Carbomer U20, vitamin B3, 1,3-butylene glycol, p-hydroxyacetophenone and water, raise the temperature to 85°C while stirring, keep warm for 15 minutes, and start cooling after the materials are completely dissolved; b. Cool down to 45°C, add biopolysaccharide gum powder, double bacteria fermented black tea filtrate, L-arginine, bis-PEG-18 methyl ether dimethyl silane, tocopherol, 1,2-hexanediol in sequence while stirring, stir evenly to obtain the essence containing Acetobacter xylinum / yeast ancient tree black tea fermentation filtrate.

[0085] Comparative Example 1 The specific implementation is the same as in Example 1, except that the fermentation culture conditions in step (2) in step S4 are 28° C. and 160 rpm for 7 h.

[0086] Comparative Example 2 Specific implementation method The same as Example 1, except that Acetobacter xylinum + lactic acid bacteria + yeast are used for three-bacteria fermentation. Specifically, the seed liquid in step S3 includes Acetobacter xylinum seed liquid, lactic acid bacteria seed liquid and yeast seed liquid.

[0087] The inoculation fermentation culture in step S4 is specifically a three-step fermentation, including step (1): inoculating the seed liquid of Acetobacter xylinum into the fermentation medium for fermentation culture; inoculating the seed liquid of lactic acid bacteria into the fermentation liquid obtained in step (1) for fermentation culture; step (3): inoculating the seed liquid of yeast into the fermentation liquid obtained in step (2) for fermentation culture.

[0088] The inoculation fermentation culture in step (2) specifically comprises inoculating the lactic acid bacteria seed liquid into the fermentation medium for fermentation culture; the fermentation culture conditions are: fermentation culture at 38° C. and 170 rpm for 16 hours; the fermentation culture is carried out under facultative anaerobic conditions.

[0089] The preparation method of the lactic acid bacteria seed liquid is the same as the preparation method of the xylinum acetobacter seed liquid, except that the lactic acid bacteria freeze-dried powder replaces the xylinum acetobacter. The inoculation amount of the lactic acid bacteria strain is 2wt%.

[0090] The lactic acid bacteria are specifically Lactobacillus casei (Latin name Lactobacillus casei), which comes from Guangzhou Junyan Biotechnology Co., Ltd., with a biological deposit number of GDMCC NO: 64856, a biological deposit date of July 10, 2024, a biological deposit address of Guangdong Provincial Microbiological Culture Collection Center, and a depository unit of Guangzhou Junyan Biotechnology Co., Ltd. .

[0091] Comparative Example 3 The specific implementation method is the same as Example 1, except that the components of the fermentation medium in step S2 include: 30 g / L black tea powder, 30 g / L glucose, 2 g / L peptone, 2 g / L yeast extract, and the solvent is water; mix, adjust the pH to 5.5, and sterilize to obtain.

[0092] Comparative Example 4 The specific implementation is the same as that of Example 1, except that the black tea leaves are specifically Fengqing black tea leaves.

[0093] Performance Testing Physical and chemical test odor: The odor of the black tea filtrate fermented by double bacteria in Example 1 was similar to tea perfume after 24 hours of standing. However, the fermentation time of the black tea fermentation filtrate in Comparative Example 1 was too long, and the odor was sour and unpleasant; Comparative Example 2 used Acetobacter xylinum + lactic acid bacteria + yeast for fermentation, and the odor was very sour after fermentation; Comparative Example 3 selected Fengqing black tea leaves for fermentation, and the tea aroma was very light after fermentation compared with Example 1.

[0094] Appearance: The color of the filtrate of the double-bacteria fermented black tea in Example 1 is dark brown; while the color of the filtrate of the double-bacteria fermented black tea in Comparative Example 3 is light brown.

[0095] Component content detection: Referring to GB / T 8313-2008 and GB / T 8314-2013, the component content of the black tea fermentation filtrate of Example 1 and Comparative Example 2 was detected. The results are shown in Tables 2 and 3 below.

[0096] Table 2. Content of components in the black tea fermentation filtrate of Example 1

[0097] Table 3. Content of components in the black tea fermentation filtrate of Comparative Example 2

[0098] Efficacy test 2.1 Anti-wrinkle and firming efficacy test Test method see Figure 1 , the elastase inhibition rate was selected as the test indicator to measure the anti-wrinkle and firming effects; the specific testing was carried out by Guangdong LiChuang Testing Technology Service Co., Ltd.

[0099] The test results are shown in Table 3 below; wherein the blank control group is a buffer solution without inhibitory components, and the test sample group is the double-bacteria fermented black tea filtrate prepared in Example 1. It can be seen from the data in Table 3 that the elastase inhibition rate (18.84%) of the double-bacteria fermented black tea filtrate prepared by the present invention is significantly higher than that of the blank control group, and the statistical difference P value is <0.05, indicating that the difference is significant, and the test sample has excellent firming and anti-wrinkle effects.

[0100] Table 3

[0101] 2.2 Moisturizing efficacy test Test method see Figure 2 , the moisture change rate is selected as the test indicator to measure the moisturizing effect; the specific testing is carried out by Guangdong LiChuang Testing Technology Service Co., Ltd.

[0102] The test results are shown in Table 4 below; the positive control sample is glycerol, and the test sample is the double-bacteria fermented black tea filtrate prepared in Example 1; the moisture change rate Δm is determined by measuring the sample at Figure 2 The difference in moisture before and after changes under medium temperature and humidity conditions is calculated.

[0103] From the data in Table 4, it can be seen that the moisture change rate Δm of the double-bacteria fermented black tea filtrate prepared by the present invention is always lower than the Δm of the positive control sample; the lower the moisture change rate Δm, the better the moisture retention of the sample, which shows that the double-bacteria fermented black tea filtrate has an excellent moisturizing effect. The reason why the sample Δm at 4h is lower than that at 2h is mainly due to the volatilization and change of normal moisture when placed; at this time, the moisturizing effect of the filtrate is still improving, and at 8h, the moisturizing effect of the filtrate is strong, and Δm is significantly improved.

[0104] Table 4

[0105] 2.3 Soothing effect test Test method see Figure 3 , the hyaluronidase inhibition rate was selected as the test indicator to measure the soothing effect; the specific testing was carried out by Guangdong LiChuang Testing Technology Service Co., Ltd.

[0106] The test results are shown in Table 5 below; wherein the blank control group is a buffer solution without inhibitory components, and the test sample is the double-bacteria fermented black tea filtrate prepared in Example 1.

[0107] It can be seen from the data in Table 5 that the hyaluronidase inhibition rate (44.89%) of the double-bacteria fermented black tea filtrate prepared by the present invention is significantly higher than that of the blank control group, and the statistical difference P value is <0.05, indicating that the difference is significant and the test sample has excellent soothing effect.

[0108] Table 5

[0109] 3. VISIA skin test results of serum Test method: Three female users were randomly selected to try the essence of Example 1, using it once in the morning and once in the evening for a period of time, and the changes in facial skin at the beginning and after use were recorded. User 1 used it from 2023.10.6 to 2023.10.25, user 2 used it from 2023.10.6 to 2023.10.24, and user 3 used it from 2023.10.6 to 2023.10.19; the VISIA results are shown in Table 1. Figure 4-6 .from Figure 4-6 It can be seen that after using the essence containing the double-bacteria fermented black tea filtrate provided by the present invention, the effect of significantly reducing the area of ​​skin wrinkles can be achieved after only two to three weeks of use, and the essence has obvious firming and anti-wrinkle effects.

Claims

1. A process for preparing a double-bacteria fermented black tea filtrate, characterized in that: The specific preparation steps of the double-bacteria fermented black tea filtrate include: S1. Preparation of black tea powder; S2. preparing fermentation medium; S3. preparing seed solution; S4. Inoculation and fermentation culture; S5. collecting the filtrate to obtain a fermentation product; The seed solution in step S3 includes Acetobacter xylinum seed solution and yeast seed solution.

2. The process for preparing the double-bacteria fermented black tea filtrate according to claim 1, characterized in that: The step S1 comprises: crushing black tea leaves, grinding to obtain black tea coarse powder, and sieving the black tea coarse powder to obtain black tea powder; The black tea leaves are at least one of Dian black tea, Keemun black tea, Yingde black tea, Ning black tea and Sichuan black tea.

3. The process for preparing the double-bacteria fermented black tea filtrate according to claim 1, characterized in that: The preparation method of the Acetobacter xylinum seed liquid comprises: inoculating the freeze-dried powder of Acetobacter xylinum into a first culture medium, activating and culturing for 2-6 days to obtain an activated Acetobacter xylinum strain; inoculating the activated Acetobacter xylinum strain into a second culture medium, and performing secondary culturing to obtain the Acetobacter xylinum seed liquid; the inoculation amount of the Acetobacter xylinum strain is 1-5wt%.

4. The process for preparing the double-bacteria fermented black tea filtrate according to claim 1, characterized in that: The preparation method of the yeast seed liquid comprises: inoculating yeast into a third culture medium, performing activation culture for 1-3 days, and obtaining an activated yeast strain; inoculating the activated yeast strain into a fourth culture medium, performing secondary culture, and obtaining a yeast seed liquid; the inoculation amount of the yeast strain is 0.5-3wt%.

5. The process for preparing the double-bacteria fermented black tea filtrate according to claim 4, characterized in that: The yeast is obtained by separation and purification from beer slag.

6. The process for preparing the double-bacteria fermented black tea filtrate according to claim 1, characterized in that: The inoculation and fermentation culture in step S4 is specifically a two-step fermentation, comprising step (1): inoculating the seed liquid of Acetobacter xylinum into the fermentation medium for fermentation culture; and step (2): inoculating the seed liquid of Saccharomyces cerevisiae into the fermentation liquid obtained in step (1) for fermentation culture, thereby obtaining a double-bacteria fermentation black tea filtrate product.

7. The process for preparing the double-bacteria fermented black tea filtrate according to claim 6, characterized in that: The elastase inhibition rate of the double-bacteria fermented black tea filtrate is greater than 18%.

8. A double-bacteria fermented black tea filtrate, characterized in that: The invention is prepared by the preparation process described in any one of claims 1 to 7.

9. An application of the double-bacteria fermented black tea filtrate according to claim 8, characterized in that: The double-bacteria fermented black tea filtrate is applied to cosmetics; the cosmetics include any one of skin care water, essence, emulsion, face cream, ointment, facial mask liquid, and facial cleanser.

10. The use of the double-bacteria fermented black tea filtrate according to claim 9, characterized in that: The double bacteria fermented black tea filtrate is applied to the essence; the addition amount of the double bacteria fermented black tea filtrate in the essence is 10-60wt%.

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