Bifidobacterium yeast fermentation product filtrate with moisturizing and repairing effects as well as preparation method and application of bifidobacterium yeast fermentation product filtrate

Abstract

The invention belongs to the technical field of microbial fermentation, and particularly discloses a bifidus yeast fermentation product filtrate with moisturizing and repairing effects as well as a preparation method and application of the bifidus yeast fermentation product filtrate. The preparation method of the bifidobacterium fermentation product filtrate comprises the following steps: (1) strain activation: inoculating bifidobacterium adolescentis into a first seed culture medium for anaerobic culture to obtain an activated bacterial solution; (2) seed liquid culture: inoculating the activated bacterial liquid to a second seed culture medium, and continuously transferring anaerobic culture twice to obtain a third-stage seed liquid; (3) fermentation culture: inoculating the third-stage seed solution into a fermentation tank for culture to obtain a fermented product; and (4) separation and purification: filtering and decolorizing the fermentation product to obtain the bifidus yeast fermentation product filtrate. The prepared bifidus yeast fermentation product filtrate is high in active ingredient content and good in stability, and has good moisturizing and repairing effects.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of microbial fermentation, in particular to a bifida ferment filtrate with moisturizing and repairing effects, and a preparation method and application thereof. BACKGROUND

[0002] The bifida ferment filtrate is a filtrate after fermentation of bifidobacterium, does not contain bacteria, and is mainly metabolites such as short-chain fatty acids, extracellular polysaccharides, free amino acids and polypeptides. Among them, polysaccharide substances can form a protective film on the skin surface, not only can effectively lock in moisture, but also can resist the invasion of free radicals, thereby playing excellent moisturizing and antioxidant effects; amino acids and polypeptides not only participate in normal metabolism of the skin, but also can penetrate into the skin bottom layer, repair damaged cells and stimulate the generation of collagen, thereby enhancing the elasticity of the skin. Therefore, the bifida ferment has great market application prospect.

[0003] The existing bifida ferment process has the following problems: low fermentation efficiency, long cycle, high energy consumption, low yield and instability of target active ingredients, and large batch difference. The problems of low fermentation efficiency and insufficient active ingredients in the prior art are solved. SUMMARY

[0004] In view of the above, it is necessary to provide a bifida ferment filtrate with moisturizing and repairing effects, and a preparation method and application thereof. The bifida ferment filtrate prepared by the present application has high active ingredient content, good stability, and good moisturizing and repairing effects.

[0005] To achieve the above-mentioned purpose, the technical scheme adopted by the present application is as follows:

[0006] A preparation method of a bifida ferment filtrate, comprising the following steps:

[0007] (1) Strain activation: inoculate Bifidobacterium adolescentis into a first seed culture medium, and anaerobically culture to obtain an activated bacterial solution;

[0008] (2) Seed liquid culture: inoculate the activated bacterial solution into a second seed culture medium and continuously transfer and anaerobically culture twice to obtain a third-level seed liquid;

[0009] (3) Fermentation culture: inoculate the third-level seed liquid into a fermentation tank containing a fermentation culture medium, and anaerobically ferment and culture to obtain a fermentation product; wherein the anaerobic fermentation culture comprises a first anaerobic fermentation, a second anaerobic fermentation and a third anaerobic fermentation;

[0010] The conditions of the first anaerobic fermentation are as follows: temperature 36-37℃, pH 6.0-6.2, rotation speed 100-110rpm, and fermentation time 8-10h;

[0011] The conditions for the second anaerobic fermentation are: temperature 38.5-39℃, pH 5.8-6.0, rotation speed 110-115 rpm, and fermentation time 10-12 h.

[0012] The conditions for the third anaerobic fermentation are: temperature 35-35.5℃, pH 5.8-6.0, rotation speed 95-100 rpm, and fermentation time 2-4 hours.

[0013] (4) Separation and purification: After filtering and decolorizing the fermentation product, the filtrate of the Bifida ferment lysate can be obtained.

[0014] Furthermore, in step (1), the Bifidobacterium adolescentis is Bifidobacterium adolescentis HH404-1, which is classified as Bifidobacterium adolescentis. It is deposited at the Guangdong Provincial Microbial Culture Collection Center (GDMCC) at the address of Building 59, No. 100 Xianlie Middle Road, Guangzhou, on October 22, 2025, with the accession number GDMCC No: 67151.

[0015] Further, the first seed culture medium comprises the following raw material components by weight percentage: peptone 1.0-1.2%, beef extract 0.8-1.2%, yeast extract 0.4-0.7%, glucose 1.8-2.3%, dipotassium hydrogen phosphate 0.1-0.3%, triammonium citrate 0.1-0.3%, sodium acetate 0.4-0.6%, magnesium sulfate 0.01-0.04%, manganese sulfate 0.003-0.006%, and Tween-80 0.10-0.15%; the second seed culture medium comprises the following raw material components by weight percentage: glucose 2.3-2.7%, peptone 0.8-1.2%, yeast extract 0.3-0.7%, magnesium sulfate 0.01-0.02%, manganese sulfate 0.003-0.006%, sodium acetate 0.4-0.6%, and ammonium citrate 0.13-0.16%.

[0016] Furthermore, in step (3), during the first anaerobic fermentation, a first solution is added; during the second anaerobic fermentation, a second solution is added; and during the third anaerobic fermentation, a third solution is added; the first solution comprises glucose and yeast treatment, the second solution comprises glucose, yeast treatment and glutamine, and the third solution comprises trehalose and serine.

[0017] Further, in step (3), the first solution comprises the following components in percentage by mass: 5-10% of glucose and 0.5-1% of yeast processing product; the second solution comprises the following components in percentage by mass: 5-10% of glucose, 1-2% of yeast processing product and 0.1-0.3% of glutamine; and the third solution comprises the following components in percentage by mass: 0.1-0.3% of trehalose and 0.03-0.05% of serine.

[0018] Further, the feeding amount of the first solution is 2-5% of the mass of the initial fermentation material, the feeding amount of the second solution is 4-7% of the mass of the initial fermentation material, and the feeding amount of the third solution is 1-2% of the mass of the initial fermentation material.

[0019] Further, the preparation method of the yeast processing product is as follows: the Saccharomyces cerevisiae is crushed at 30-40 MPa to obtain a bacterial solution, a compound enzyme is added to the bacterial solution, and the enzyme is hydrolyzed at 45-50℃ and pH 5.0-5.5 for 2-3 hours to obtain the yeast processing product; wherein the compound enzyme is prepared by mixing cellulase and β-glucanase at a mass ratio of 1:2.

[0020] Further, the fermentation medium comprises the following raw material components in percentage by mass: 4.0-5.0% of glucose, 1.5-1.8% of yeast processing product, 0.8-1.2% of peptone, 0.4-0.7% of dipotassium hydrogen phosphate, 0.3-0.5% of glycyrrhiza extract, 0.01-0.02% of manganese sulfate, 0.01-0.03% of magnesium sulfate, 0.4-0.7% of anhydrous sodium acetate, 0.12-0.18% of ammonium citrate, and 0.1-0.3% of cysteine.

[0021] The application further provides a Schizochytrium ferment product filtrate prepared by the preparation method.

[0022] The application further provides an application of the Schizochytrium ferment product filtrate in the preparation of cosmetics.

[0023] Further, the cosmetics are cosmetics with moisturizing and / or repairing effects.

[0024] The application has the following beneficial effects:

[0025] 1. The present application carries out a three-stage fermentation control strategy by adjusting the fermentation parameters during the main fermentation process. In the fast growth stage of the bacteria, the fermentation conditions are controlled at 36-37℃, pH 6.0-6.2, and 100-110 rpm to effectively promote bacterial growth and increase bacterial biomass. When the bacterial growth tends to be stable, the fermentation conditions are controlled at 38.5-39℃, pH 5.8-6.0, and 110-115 rpm, which is beneficial to improve the metabolic rate and promote the accumulation of target metabolites, thereby increasing the content of active ingredients. In the late fermentation stage, by reducing the temperature and speed, the synthesis of by-products can be reduced, and the activity can be stabilized, thereby improving the purity and stability of the product.

[0026] 2. For the three stages of the main fermentation of the present application, specific feed is added to match the nutritional needs of the bacteria at different stages, avoiding substrate deficiency or inhibition, thereby increasing the yield and stability of active ingredients. In the early stage of fermentation, by adding glucose and yeast treatment, the carbon and nitrogen sources are supplemented, which is beneficial to promote bacterial growth and increase bacterial biomass. In the middle stage of fermentation, by adding glucose, yeast treatment and glutamine, product synthesis can be effectively induced, and the accumulation of active metabolites can be promoted. In the late stage of fermentation, by adding trehalose and serine, the activity of bacterial cells can be maintained and the release of products can be promoted.

[0027] 3. The fermentation medium of the present application has comprehensive nutrition, which can provide balanced nutrition for the bacteria and activate related metabolic pathways, improve the fermentation starting force, significantly improve the bacterial density and metabolic activity, thereby effectively improve the fermentation efficiency and effect.

[0028] 4. The Bifidobacterium adolescentis obtained by screening and separation has good anaerobic fermentation effect, large bacterial growth, and high synthesis of metabolic products. The fermentation filtrate obtained by anaerobic fermentation of the Bifidobacterium adolescentis has high active substance content and good stability. When applied to cosmetics, it can effectively play the effects of moisturizing, antioxidant, repair, and brightening. By controlling the reasonable addition amount, it will not cause damage to the organization structure of the cosmetics, and will not bring adverse effects to the stability of the cosmetics, and will not cause discoloration, delamination and other quality problems.

[0029] In summary, the present application optimizes the fermentation medium and combines with the control of fermentation conditions and dynamic feeding strategy, which on the one hand improves the synthesis efficiency of metabolic products, so that the target active ingredient content of the Schizochytrium sp. fermentation product filtrate produced is high, the stability is good, and the product has excellent beauty and skin care effects, especially good repair and moisturizing effects, and is suitable for the development of cosmetics. On the other hand, the controllability of the preparation process is improved, thereby reducing batch differences, improving product consistency and reliability, and facilitating scale-up and production automation. BRIEF DESCRIPTION OF DRAWINGS

[0030] Figure 1It is a colony map of Bifidobacterium adolescentis HH404-1 strain of the application.

[0031] Figure 2 It is a microscope map of Bifidobacterium adolescentis HH404-1 strain of the application.

[0032] Figure 3 It is an environmental temperature map when the efficacy of the essence S1-S7 is determined in the application.

[0033] Figure 4 It is an environmental humidity map when the efficacy of the essence S1-S7 is determined in the application.

[0034] Biological material preservation information

[0035] The strain preserved in the application is Bifidobacterium adolescentis HH404-1, which is classified as Bifidobacterium adolescentis, preserved in Guangdong Microbial Culture Collection Center (GDMCC) located at No. 59 Building, 5th Floor, Guangzhou Xianlie Middle Road 100, with a preservation date of October 22, 2025 and a preservation number of GDMCC No: 67151. DETAILED DESCRIPTION

[0036] In order to make the technical problems, technical solutions and advantages to be solved in the application more clear, specific embodiments will be described in detail below.

[0037] In the application, the reagents and consumables used are purchased from conventional reagent manufacturers in the field, and the experimental methods and technical means used are conventional methods and means in the field, unless otherwise specified.

[0038] Firstly, the application provides a strain of Bifidobacterium adolescentis. The Bifidobacterium adolescentis is isolated and screened from the feces of long-lived old people in Bama Longevity Village, Guangxi Zhuang Autonomous Region, and is identified by biochemical identification and molecular biology identification, and the specific process is shown as follows.

[0039] Isolation and screening of the strain

[0040] The Bifidobacterium adolescentis is isolated from the feces of long-lived old people in Bama Longevity Village, Guangxi Zhuang Autonomous Region, and the specific isolation and screening method is as follows:

[0041] Aseptically take 25 g of feces of long-lived old people, add 225 mL of normal saline, and then dilute the bacterial suspension to 10 -6 , 10 -7 , 10-8 Three gradients, then 0.2 mL was taken to perform Hungate anaerobic rolling tube, placed in 37℃ incubator for 48-72h, then the colony characteristics of smooth, convex, edge neat, white or slightly yellow, soft texture of small colonies inoculated in BS medium, anaerobic culture 24-48h, gram staining and microscopy. Select gram staining observed with the morphological characteristics of Bifidobacterium, surrounded by transparent circle, smooth edge, white or slightly yellowish colonies inoculated in BS medium, in aerobic and anaerobic environment at 37℃ for 24h, while the KOH, hydrogen peroxide enzyme, indole, glucose metabolism test. Select anaerobic conditions, aerobic conditions do not grow, KOH test negative, peroxide enzyme test negative, indole test negative, can metabolize glucose but not gas Bifidobacterium preliminary screening. The preliminary screening of Bifidobacterium inoculated in MRS broth culture, pick up the above medium observation results good colonies repeatedly subcultured to select a strain of Bifidobacterium adolescentis, numbered: HH404-1, placed in -80℃ glycerol preservation.

[0042] 1. Biochemical identification of the strain

[0043] Referring to the "Bergey's Manual of Determinative Bacteriology" (9th edition) to observe the morphology, color and other surface growth of the strain in the MRS medium, its colony morphology is shown in Figure 1 ; pick up young culture, smear, gram staining, observe the morphology, size, gram staining reaction, and the presence or absence of spores, morphology and location of bacteria under a microscope, and the microscopic examination is shown in Figure 2 .

[0044] 3. Molecular biology identification of the strain

[0045] The target strain HH404-1 was taxonomic identified by 16S rDNA sequence analysis.

[0046] Strain HH404-1 was sent to Shanghai Biosciences Sequencing Identification Department for sequencing identification, and the sequencing results are as follows:

[0047] GCGGCGTTGCTGATCCGCGATTACTAGCGACTCCGCCTTCATGGAGTCGGGTTGCAGACTCCAATCCGAACTGAGACCGGTTTTAAGGGATCCGCTCCACCTCACAGTGTCGCATCCCGTTGTACCGGCCATTGTAGCATGCGTGAAGCCCTGGACGTAAGGGGCATGATGATCTGACGTCATCCCCACCTTCCTCCGAGTTGACCCCGGCGGTCCCCCGTGAGTTCCCACCACGACGTGCTGGCAACACAGGGCGAGGGTTGCGCTCGTTGCGGGACTTAACCCAACATCTCACGACACGAGCTGACGACGACCATGCACCACCTGTGAACCCGCCCCGAAGGGAGACCGTATCTCTACGGCTGTCGGGAACATGTCAAGCCCAGGTAAGGTTCTTCGCGTTGCATCGAATTAATCCGCATGCTCCGCCGCTTGTGCGGGCCCCCGTCAATTTCTTTGAGTTTTAGCCTTGCGGCCGTACTCCCCAGGCGGGATGCTTAACGCGTTGGCTCCGACACGGAGACCGTGGAATGGTCCCCACATCCAGCATCCACCGTTTACGGCGTGGACTACCAGGGTATCTAATCCTGTTCGCTCCCCACGCTTTCGCTCCTCAGCGTCAGTGACGGCCCAGAGACCTGCCTTCGCCATTGGTGTTCTTCCCGATATCTACACATTCCACCGTTACACCGGGAATTCCAGTCTCCCCTACCGCACTCAAGCCCGCCCGTACCCGGCGCGGATCCACCGTTAAGCGATGGACTTTCACACCGGACGCGACGAACCGCCTACGAGCCCTTTACGCCCAATAATTCCGGATAACGCTTGCACCCTACGAATTACCGCGG

[0048] The sequencing result is analyzed by comparison on ribosome database (NCBI). The 16S rDNA sequence analysis result shows that the homology of the strain with Bifidobacterium adolescentis (Bifidobacterium adolescentis) in Bifidobacterium is as high as 100%, that is, the strain is Bifidobacterium adolescentis (Bifidobacterium adolescentis).

[0049] The strain is preserved, and the preservation information is specifically as follows: Bifidobacterium adolescentis HH404-1, the classification name of which is Bifidobacterium adolescentis, is preserved in Guangdong Microbial Culture Collection Center (GDMCC) located at No. 59 Building, 5th Floor, Guangzhou, Guangdong, on October 22, 2025, and the preservation number is GDMCC No: 67151.

[0050] Secondly, the application provides a Schizosaccharomyces ferment product filtrate.

[0051] The preparation method of the Schizosaccharomyces ferment product filtrate comprises the following steps:

[0052] (1) Strain activation: inoculate Bifidobacterium adolescentis into a first seed culture medium, and anaerobically culture to obtain an activated seed liquid;

[0053] (2) Seed liquid culture: inoculate the activated seed liquid into a second seed culture medium and continuously transfer and anaerobically culture twice to obtain a third-level seed liquid;

[0054] (3) Fermentation culture: inoculate the third-level seed liquid into a fermentation tank containing a fermentation culture medium, and anaerobically ferment and culture to obtain a fermentation product; wherein the fermentation culture comprises a first anaerobic fermentation, a second anaerobic fermentation and a third anaerobic fermentation;

[0055] The first anaerobic fermentation is carried out under the conditions of a temperature of 36-37℃, a pH of 6.0-6.2, a rotation speed of 100-110 rpm and a fermentation time of 8-10 h;

[0056] The second anaerobic fermentation is carried out under the conditions of a temperature of 38.5-39℃, a pH of 5.8-6.0, a rotation speed of 110-115 rpm and a fermentation time of 10-12 h;

[0057] The third anaerobic fermentation is carried out under the conditions of a temperature of 35-35.5℃, a pH of 5.8-6.0, a rotation speed of 95-100 rpm and a fermentation time of 2-4 h, and the residual sugar in the tank is <1 g / L at this time;

[0058] (4) Separation and purification: after the fermentation product is filtered and decolorized, the filtrate of the Schizochytrium sp. fermentation product is obtained.

[0059] Preferably, in step (1), the Bifidobacterium adolescentis is Bifidobacterium adolescentis HH404-1, which is classified as Bifidobacterium adolescentis and preserved in the Guangdong Microbial Culture Collection Center (GDMCC) at 59 Building, 5th Floor, NO.100, Martyrs' Avenue, Guangzhou, on October 22, 2025, with the preservation number GDMCC No: 67151.

[0060] Preferably, in step (1), the first seed culture medium comprises the following raw material components in mass percentage: 1.0-1.2% of proteose peptone, 0.8-1.2% of beef extract, 0.4-0.7% of yeast powder, 1.8-2.3% of glucose, 0.1-0.3% of dipotassium hydrogen phosphate, 0.1-0.3% of triammonium citrate, 0.4-0.6% of sodium acetate, 0.01-0.04% of magnesium sulfate, 0.003-0.006% of manganese sulfate, and 0.10-0.15% of Tween-80.

[0061] Exemplarily, in the first seed culture medium, the mass percentage of proteose peptone can be 1.0%, 1.1%, or 1.2%, etc.; the mass percentage of beef extract can be 0.8%, 0.9%, 1.0%, or 1.2%, etc.; the mass percentage of yeast powder can be 0.4%, 0.6%, or 0.7%, etc.; the mass percentage of glucose can be 1.8%, 2.0%, 2.1%, or 2.3%, etc.; the mass percentage of dipotassium hydrogen phosphate can be 0.1%, 0.2%, or 0.3%, etc.; the mass percentage of triammonium citrate can be 0.1%, 0.2%, or 0.3%, etc.; the mass percentage of sodium acetate can be 0.4%, 0.5%, or 0.6%, etc.; the mass percentage of magnesium sulfate can be 0.003%, 0.004%, or 0.006%, etc.; and the mass percentage of Tween-80 can be 0.10%, 0.12%, or 0.15%, etc.

[0062] Preferably, in step (1), the anaerobic culture conditions are 37-38℃ for 16-20h. Exemplarily, the temperature of anaerobic culture can be 37℃ or 38℃, and the culture time can be 16h, 18h, or 20h.

[0063] Preferably, in step (2), the two continuous anaerobic cultures are specifically as follows: inoculating the activated bacteria solution into the second seed culture medium, and carrying out anaerobic culture at 36-38℃ for 12-16h to obtain a first-stage seed solution; inoculating the first-stage seed solution into the first seed culture medium, and carrying out anaerobic culture at 36-38℃, 95-110rpm, pH 6.0-6.2 for 10-12h to obtain a second-stage seed solution; inoculating the second-stage seed solution into the second seed culture medium, and carrying out anaerobic culture at 36-38℃, 100-110rpm, pH 6.0-6.2 for 10-13h to obtain a third-stage seed solution.

[0064] Preferably, in step (2), the second seed culture medium comprises the following raw material components in percentage by mass: glucose 2.3-2.7%, peptone 0.8-1.2%, yeast powder 0.3-0.7%, magnesium sulfate 0.01-0.02%, manganese sulfate 0.003-0.006%, sodium acetate 0.4-0.6%, and ammonium citrate 0.13-0.16%.

[0065] Illustratively, in the second seed culture medium, the percentage by mass of the glucose can be 2.3%, 2.5% or 2.7%, etc.; the percentage by mass of the peptone can be 0.8%, 1.0% or 1.2%, etc.; the percentage by mass of the yeast powder can be 0.3%, 0.5% or 0.7%, etc.; the percentage by mass of the magnesium sulfate can be 0.01%, 0.015% or 0.02%, etc.; the percentage by mass of the manganese sulfate can be 0.003%, 0.005% or 0.006%, etc.; the percentage by mass of the sodium acetate can be 0.4%, 0.5% or 0.6%, etc.; and the percentage by mass of the ammonium citrate can be 0.13%, 0.15% or 0.16%, etc.

[0066] Illustratively, in step (2), the temperature for anaerobic culture of the activated bacteria solution can be 36℃, 37℃ or 38℃, etc., and the culture time can be 12h, 14h or 16h, etc.; the temperature for anaerobic culture of the first-stage seed solution can be 36℃, 37℃ or 38℃, etc., the rotation speed can be 95rpm, 100rpm or 110rpm, etc., the pH can be 6.0, 6.1 or 6.2, etc., and the culture time can be 10h, 11h or 12h, etc.; the temperature for anaerobic culture of the second-stage seed solution can be 36℃, 37℃ or 38℃, etc., the rotation speed can be 100rpm, 105rpm or 110rpm, etc., the pH can be 6.0, 6.1 or 6.2, etc., and the culture time can be 10h, 12h or 13h, etc.

[0067] Preferably, in step (3), the inoculation amount of the third-stage seed solution is 5-10%. Illustratively, the inoculation amount of the third-stage seed solution can be 5%, 7%, 9% or 10%.

[0068] Preferably, in step (3), the fermentation medium comprises the following raw material components in mass percentage: glucose 4.0-5.0%, yeast extract 1.5-1.8%, peptone 0.8-1.2%, potassium phosphate dibasic 0.4-0.7%, glycyrrhiza extract 0.3-0.5%, manganese sulfate 0.01-0.02%, magnesium sulfate 0.01-0.03%, anhydrous sodium acetate 0.4-0.7%, ammonium citrate 0.12-0.18%, and cysteine 0.1-0.3%.

[0069] For example, in the fermentation medium, the mass percentage of glucose can be 4.0%, 4.3%, 4.7%, 5.0%, etc., the mass percentage of yeast extract can be 1.5%, 1.7%, 1.8%, etc., the mass percentage of peptone can be 1.5%, 1.7%, 1.8%, etc., the mass percentage of potassium phosphate dibasic can be 0.4%, 0.4%, 0.7%, etc., the mass percentage of glycyrrhiza extract can be 0.3%, 0.4%, 0.5%, etc., the mass percentage of manganese sulfate can be 0.01%, 0.015%, 0.02%, etc., the mass percentage of magnesium sulfate can be 0.01%, 0.02%, 0.03%, etc., the mass percentage of anhydrous sodium acetate can be 0.4%, 0.6%, 0.7%, etc., the mass percentage of ammonium citrate can be 0.12%, 0.16%, 0.18%, etc., and the mass percentage of cysteine can be 0.1%, 0.2%, 0.3%, etc.

[0070] For example, in step (3), the temperature of the first anaerobic fermentation can be 36℃, 36.5℃, or 37℃, the pH can be 6.0, 6.1, or 6.2, the rotation speed can be 100rpm, 105rpm, or 110rpm, and the fermentation time can be 8h, 9h, or 10h; the temperature of the second anaerobic fermentation can be 38.5℃, 38.7℃, or 39℃, the pH can be 5.8, 5.9, or 6.0, the rotation speed can be 110rpm, 112rpm, or 115rpm, and the fermentation time can be 10h, 11h, or 12h; the temperature of the third anaerobic fermentation can be 35℃, 35.2℃, or 35.5℃, the pH can be 5.8, 5.9, or 6.0, the rotation speed can be 95rpm, 98rpm, or 100rpm, and the fermentation time can be 2h, 3h, or 4h.

[0071] Preferably, in step (3), during the first fermentation, the first solution is fed; during the second fermentation, the second solution is fed; during the third fermentation, the third solution is fed; the first solution comprises glucose and yeast extract, the second solution comprises glucose, yeast extract and glutamine, and the third solution comprises trehalose and serine. Specifically, the feeding rate is dynamically adjusted according to online dissolved oxygen (DO) and pH feedback, and when DO rises sharply or pH fluctuates abnormally, the feeding rate is reduced; when DO decreases stably, the feeding rate is increased.

[0072] Preferably, the first solution comprises the following components in mass percentage: glucose 5-10% and yeast extract 0.5-1%; the second solution comprises the following components in mass percentage: glucose 5-10%, yeast extract 1-2% and glutamine 0.1-0.3%; and the third solution comprises the following components in mass percentage: trehalose 0.1-0.3% and serine 0.03-0.05%.

[0073] For example, in the first solution, the mass percentage of glucose can be 5%, 7%, 9%, 10%, etc., and the mass percentage of yeast extract can be 0.5%, 0.8%, 1%, etc.; in the second solution, the mass percentage of glucose can be 5%, 7%, 9%, 10%, etc., and the mass percentage of yeast extract can be 1%, 1.5%, 2%, etc.; the mass percentage of glutamine can be 0.1%, 0.2% or 0.3%, etc.; in the third solution, the mass percentage of trehalose can be 0.15%, 0.2%, 0.3%, etc., and the mass percentage of serine can be 0.03%, 0.04%, 0.05%, etc.

[0074] Preferably, the feeding amount of the first solution is 2-5% of the mass of the initial fermentation material, the feeding amount of the second solution is 4-7% of the mass of the initial fermentation material, and the feeding amount of the third solution is 1-2% of the mass of the initial fermentation material.

[0075] For example, the feeding amount of the first solution can be 2%, 4% or 5% of the mass of the initial fermentation material, the feeding amount of the second solution can be 4%, 6% or 7% of the mass of the initial fermentation material, and the feeding amount of the third solution can be 1%, 1.5% or 2% of the mass of the initial fermentation material.

[0076] Preferably, in step (3), the preparation method of the yeast treatment product is as follows: the Saccharomyces cerevisiae cell bodies are broken at 30-40 MPa to obtain a bacterial solution, a composite enzyme is added to the bacterial solution, and the enzyme is hydrolyzed at 45-50℃ and pH 5.0-5.5 for 2-3h to obtain the yeast treatment product; wherein the composite enzyme is prepared by mixing cellulase and β-glucanase at a mass ratio of 1:2.

[0077] Illustratively, the breaking pressure of the Saccharomyces cerevisiae cell bodies can be 30 MPa, 34 MPa or 40 MPa, etc., the enzyme hydrolysis temperature of the bacterial solution is 45℃, 47℃ or 50℃, etc., the enzyme hydrolysis pH is 5.0, 5.3 or 5.5, etc., and the enzyme hydrolysis time is 2h, 2.5h or 3h, etc.

[0078] Preferably, in step (4), the filtration is ceramic membrane filtration; and the decolorization can be activated carbon decolorization or molecular sieve decolorization.

[0079] The application also provides a use of the above-mentioned Schizochytrium ferment product filtrate in the preparation of cosmetics.

[0080] Preferably, the cosmetic is a cosmetic with moisturizing and / or repairing effects.

[0081] Illustratively, the dosage form of the product includes emulsion, cream, water agent, and gel.

[0082] The application will be further described below in conjunction with specific examples. It should be understood that these examples are only used to illustrate the application, and are not used to limit the scope of the application.

[0083] Example 1

[0084] The present example provides a preparation method of a Schizochytrium ferment product filtrate, which comprises the following steps:

[0085] 1. Strain activation

[0086] (1) Preparation of the first seed culture medium

[0087] The components are weighed according to the following mass percentage: 1.0% of proteose peptone, 1.0% of beef extract, 0.5% of yeast powder, 2.0% of glucose, 0.2% of dipotassium hydrogen phosphate, 0.2% of triammonium citrate, 0.5% of sodium acetate, 0.02% of magnesium sulfate, 0.005% of manganese sulfate, and 0.1% of Tween-80; the first seed culture medium is prepared according to the above formula, and the pH is adjusted to 6.5; the prepared seed culture medium is divided into screw test tubes, sterilized at 115℃ for 30min, and cooled for standby.

[0088] (2) Activation culture

[0089] Bifidobacterium adolescentis HH404-1 was inoculated into the first seed culture medium and placed in an anaerobic workstation for anaerobic culture at 37℃ for 18h to obtain activated bacteria solution.

[0090] 2. Seed expansion culture

[0091] (1) Preparation of the second seed culture medium

[0092] Each component was weighed according to the following mass percentage: glucose 2.5%, peptone 1%, yeast powder 0.5%, magnesium sulfate 0.01%, manganese sulfate 0.005%, anhydrous sodium acetate 0.5%, ammonium citrate 0.15%; the second seed culture medium was prepared according to the above formula, and its pH was adjusted to 6.5; the prepared seed culture medium was divided into screw test tubes, sterilized at 115℃ for 30min, and cooled for standby.

[0093] (2) First-stage seed liquid culture

[0094] The activated bacteria solution was inoculated into a 250mL silk test tube containing 200mL of the second seed culture medium at an inoculation amount of 5%, and placed in an anaerobic workstation for culture at 37℃ for 14h to obtain a first-stage seed liquid.

[0095] (3) Second-stage seed liquid culture

[0096] The first-stage seed liquid was inoculated into a 5L seed tank containing 4L of the second seed culture medium at an inoculation amount of 5%, and after inoculation, high-purity nitrogen gas (99.99%) was introduced into the seed tank for 10min to completely replace the air in the tank, and after the dissolved oxygen dropped to 0, the tank was clamped and pressurized, and cultured at 37℃, 100rpm, pH 6.0 (controlled by adding ammonia water) for 12h to obtain a second-stage seed liquid.

[0097] (3) Third-stage seed liquid culture

[0098] The second-stage seed liquid was inoculated into a 150L seed tank containing 80L of the second seed culture medium at an inoculation amount of 5%, and after inoculation, high-purity nitrogen gas (99.99%) was introduced for 30min to completely replace the air in the tank, and after the dissolved oxygen dropped to 0, the tank was pressurized, and cultured at 37℃, 100rpm, pH 6.0 (controlled by adding ammonia water) for 12h to obtain a third-stage seed liquid.

[0099] 3. Fermentation culture

[0100] (1) Preparation of yeast treatment

[0101] The yeast cell is broken at 30 MPa to obtain a cell liquid, and a composite enzyme prepared by mixing cellulase and β-glucanase at a mass ratio of 1:2 is added to the cell liquid, and then the enzyme is hydrolyzed at 45°C and pH 5.0 for 3h to obtain the yeast treatment product, which is ready for use.

[0102] (2) Preparation of licorice extract

[0103] Licorice powder and ethanol are mixed at a solid-liquid ratio of 1:8, and extracted at 70°C for a total of 3 times, 30 min each time. The extract is concentrated, dried, and the licorice extract is obtained, which is ready for use.

[0104] (3) Preparation of fermentation medium

[0105] The components are weighed according to the following mass percentage: glucose 4.5%, yeast treatment product 1.5%, peptone 1.0%, potassium phosphate dibasic 0.5%, licorice extract 0.4%, manganese sulfate 0.01%, magnesium sulfate 0.01%, anhydrous sodium acetate 0.5%, ammonium citrate 0.15%, and cysteine 0.2%. The glucose is sterilized separately at 115°C for 20 min, and the remaining components are mixed and sterilized at 121°C for 30 min. After sterilization, the fermentation medium is obtained by mixing uniformly, which is ready for use.

[0106] (4) Main fermentation

[0107] The third-stage seed liquid is inoculated into a 1.5-ton fermenter containing 1.05 tons of fermentation medium at an inoculation amount of 7%, and high-purity nitrogen gas (99.99%) is introduced for 50 min after inoculation to completely replace the air in the tank, and then the pressure is maintained after the dissolved oxygen drops, and then anaerobic fermentation is carried out in three times.

[0108] The first anaerobic fermentation condition is: temperature 37°C, pH 6.2, rotation speed 110 rpm, and fermentation time 8h;

[0109] The second anaerobic fermentation condition is: temperature 39°C, pH 6.0, rotation speed 115 rpm, and fermentation time 11h;

[0110] The third anaerobic fermentation condition is: temperature 35°C, pH 5.8, rotation speed 100 rpm, and fermentation time 4h, at this time, the residual sugar in the tank is <1g / L, and the fermentation product is obtained after the tank is stopped.

[0111] 4. Separation and purification

[0112] The ceramic membrane is used to remove the bacteria and insoluble substances in the fermentation product to obtain a clear fermentation liquid, and the activated carbon is used for decolorization treatment of the clear fermentation liquid to obtain the Schizochytrium sp. fermentation product filtrate.

[0113] Example 2

[0114] The present embodiment provides a preparation method of Schizosaccharomyces fermentum fermentation product filtrate, which comprises the following steps:

[0115] 1. Strain activation

[0116] (1) Preparation of first seed medium

[0117] Each component is weighed according to the following mass percentage: 1.0% of proteose peptone, 1.0% of beef extract, 0.5% of yeast powder, 2.0% of glucose, 0.2% of dipotassium hydrogen phosphate, 0.2% of triammonium citrate, 0.5% of sodium acetate, 0.02% of magnesium sulfate, 0.005% of manganese sulfate, and 0.1% of Tween-80. The first seed medium is prepared according to the above formula, and the pH is adjusted to 6.5. The prepared seed medium is divided into screw-neck test tubes, sterilized at 115°C for 30 min, and cooled for standby.

[0118] (2) Activation culture

[0119] Bifidobacterium adolescentis HH404-1 is inoculated into the first seed medium and placed in an anaerobic workstation for anaerobic culture at 37°C for 18 h to obtain an activated bacterial solution.

[0120] 2. Seed expansion culture

[0121] (1) Preparation of second seed medium

[0122] Each component is weighed according to the following mass percentage: 2.5% of glucose, 1% of proteose peptone, 0.5% of yeast powder, 0.01% of magnesium sulfate, 0.005% of manganese sulfate, 0.5% of anhydrous sodium acetate, and 0.15% of ammonium citrate. The second seed medium is prepared according to the above formula, and the pH is adjusted to 6.5. The prepared seed medium is divided into screw-neck test tubes, sterilized at 115°C for 30 min, and cooled for standby.

[0123] (2) First-stage seed liquid culture

[0124] The activated bacterial solution is inoculated into a 250 mL screw-neck flask containing 200 mL of the second seed medium at an inoculation amount of 5%, and placed in an anaerobic workstation for culture at 37°C for 14 h to obtain a first-stage seed liquid.

[0125] (3) Second-stage seed liquid culture

[0126] The first seed liquid was transferred to a 5L seed tank containing 4L of second seed culture medium at an inoculation amount of 5%, and after inoculation, high-purity nitrogen gas (99.99%) was introduced into the seed tank for 10 minutes to completely replace the air in the tank. After the dissolved oxygen dropped to 0, the tank was clamped and pressurized, and cultured at 37°C, 100rpm, pH 6.0 (controlled by adding ammonia water) for 12 hours to obtain a second seed liquid.

[0127] (3) Third seed liquid culture

[0128] The second seed liquid was transferred to a 150L seed tank containing 80L of second seed culture medium at an inoculation amount of 5%, and after inoculation, high-purity nitrogen gas (99.99%) was introduced into the seed tank for 30 minutes to completely replace the air in the tank. After the dissolved oxygen dropped to 0, the tank was pressurized, and cultured at 37°C, 100rpm, pH 6.0 (controlled by adding ammonia water) for 12 hours to obtain a third seed liquid.

[0129] 3. Fermentation culture

[0130] (1) Preparation of yeast treatment

[0131] The yeast treatment was prepared by adding a composite enzyme prepared by mixing cellulase and β-glucanase at a mass ratio of 1:2 to the bacterial liquid obtained by crushing the Saccharomyces cerevisiae cells at 30MPa, and enzymatically hydrolyzing at 45°C, pH 5.0 for 3 hours.

[0132] (2) Preparation of licorice extract

[0133] The licorice powder and ethanol were mixed at a solid-liquid ratio of 1:8, and extracted at 70°C. The extraction was repeated for a total of 3 times, with each extraction lasting for 30 minutes. The extraction liquids were combined, concentrated, and dried to obtain the licorice extract.

[0134] (3) Preparation of flow solution

[0135] First solution preparation: 8% glucose and 0.5% yeast treatment were mixed uniformly according to the mass percentage, to obtain the first solution.

[0136] Second solution preparation: 10% glucose, 1.5% yeast treatment, and 0.02% glutamine were mixed uniformly according to the mass percentage, to obtain the second solution.

[0137] Third solution preparation: 0.2% trehalose and 0.03% serine were mixed uniformly according to the mass percentage, to obtain the third solution.

[0138] (4) Preparation of fermentation medium

[0139] The components are weighed according to the following mass percentage: glucose 4.5%, yeast processing product 1.5%, proteose peptone 1.0%, dipotassium hydrogen phosphate 0.5%, glycyrrhiza extract 0.4%, manganese sulfate 0.01%, magnesium sulfate 0.01%, anhydrous sodium acetate 0.5%, ammonium citrate 0.15%, cysteine 0.2%. The glucose is sterilized separately at 115°C for 20 min, and the rest of the components are mixed and sterilized at 121°C for 30 min. After sterilization, the components are mixed uniformly to obtain the fermentation medium, which is ready for use.

[0140] (5) Main fermentation

[0141] The third-stage seed liquid is inoculated into a 1.5-ton fermentation tank containing 1.05 tons of fermentation medium at an inoculation amount of 7%, and after inoculation, high-purity nitrogen gas (99.99%) is introduced for 50 min to completely replace the air in the tank, and then the pressure is maintained after the dissolved oxygen drops. Subsequently, anaerobic fermentation is carried out in three times, wherein:

[0142] The first anaerobic fermentation conditions are: temperature 37°C, pH 6.2, rotation speed 110 rpm, and fermentation time 8 h; and during the fermentation process, the first solution is added, and the addition amount is 3% of the initial fermentation material mass;

[0143] The second anaerobic fermentation conditions are: temperature 39°C, pH 6.0, rotation speed 115 rpm, and fermentation time 11 h; and during the fermentation process, the second solution is added, and the addition amount is 5% of the initial fermentation material mass;

[0144] The third anaerobic fermentation conditions are: temperature 35°C, pH 5.8, rotation speed 100 rpm, and fermentation time 4 h, at this time, the residual sugar in the tank is <1 g / L, and during the fermentation process, the third solution is added, and the addition amount is 1.5% of the initial fermentation material mass. After the tank is stopped, the fermentation product is obtained.

[0145] 4. Separation and purification

[0146] The ceramic membrane is used to remove the bacterial cells and insoluble substances in the fermentation product to obtain a clear fermentation liquid; the activated carbon is used for decolorization treatment of the clear fermentation liquid to obtain the Schizochytrium sp. fermentation product filtrate.

[0147] Comparative Example 1

[0148] The comparative example 1 provides a preparation method of a fermentation product filtrate of a Schizochytrium sp., which is different from the preparation method of the example 1 only in (4) of step 3. The (4) of step 3 of the comparative example 1 is specifically as follows: the third seed liquid is inoculated into a 1.5-ton fermentation tank containing 1.05 tons of fermentation medium at an inoculation amount of 7%, high-purity nitrogen gas (99.99%) is introduced for 50 min after inoculation, the air in the tank is completely replaced, the pressure is maintained after the dissolved oxygen drops to 0, and fermentation is carried out at 37°C, 100 rpm and pH 6.0 for 24 h. At this time, the residual sugar in the tank is <1 g / L, and the fermentation product is obtained after the tank is stopped.

[0149] Comparative example 2

[0150] The comparative example 2 provides a preparation method of a fermentation product filtrate of a Schizochytrium sp., which is different from the preparation method of the example 2 only in the main fermentation of (5) of step 3. The main fermentation of (5) of step 3 of the comparative example 2 is specifically as follows: the third seed liquid is inoculated into a 1.5-ton fermentation tank containing 1.05 tons of fermentation medium at an inoculation amount of 7%, high-purity nitrogen gas (99.99%) is introduced for 50 min after inoculation, the air in the tank is completely replaced, the pressure is maintained after the dissolved oxygen drops to 0, and fermentation is carried out at 37°C, 100 rpm and pH 6.0 for 23 h. At this time, the residual sugar in the tank is <1 g / L, and the fermentation product is obtained after the tank is stopped; and the first solution is added within 1-8 h of fermentation, the addition amount is 3% of the mass of the initial fermentation material; the second solution is added within 8-19 h of fermentation, the addition amount is 5% of the mass of the initial fermentation material; and the third solution is added within 19-24 h of fermentation, the addition amount is 1.5% of the mass of the initial fermentation material, and the fermentation product is obtained after the tank is stopped.

[0151] Comparative example 3

[0152] The comparative example 3 provides a preparation method of a fermentation product filtrate of a Schizochytrium sp., which is different from the preparation method of the example 2 only in the main fermentation of (5) of step 3. The main fermentation of (5) of step 3 of the comparative example 2 is specifically as follows:

[0153] The third seed liquid is inoculated into a 1.5-ton fermentation tank containing 1.05 tons of fermentation medium at an inoculation amount of 7%, high-purity nitrogen gas (99.99%) is introduced for 50 min after inoculation, the air in the tank is completely replaced, the pressure is maintained after the dissolved oxygen drops to 0, and then anaerobic fermentation is carried out in three times, wherein:

[0154] The first anaerobic fermentation is carried out at a temperature of 37°C, a pH of 6.2, a rotation speed of 110 rpm, and a fermentation time of 8 h; and the first solution is added during the fermentation process, the addition amount is 3% of the mass of the initial fermentation material;

[0155] The second anaerobic fermentation is carried out under the following conditions: temperature 39℃, pH 6.0, rotation speed 115 rpm, fermentation time 11 h; and during the fermentation, the first solution is added at an amount of 5% of the initial fermentation material.

[0156] The third anaerobic fermentation is carried out under the following conditions: temperature 35℃, pH 5.8, rotation speed 100 rpm, fermentation time 4 h, at which time, the residual sugar in the tank is <1 g / L, and during the fermentation, the first solution is added at an amount of 1.5% of the initial fermentation material, and the fermentation material is obtained after the tank is stopped.

[0157] Comparative Example 4

[0158] The comparative example 2 provides a preparation method of the Schizochytrium ferment product filtrate, which is different from the preparation method of example 2 only in the preparation of the fermentation medium in step 3 (4). The preparation of the fermentation medium in step 3 (4) of the comparative example 2 is as follows: each component is weighed according to the following mass percentage: glucose 4.5%, yeast powder 1.5%, peptone 1.0%, potassium phosphate dibasic 0.5%, glycyrrhiza extract 0.4%, manganese sulfate 0.01%, magnesium sulfate 0.01%, anhydrous sodium acetate 0.5%, ammonium citrate 0.15%, and cysteine 0.2%. The glucose is sterilized separately under the sterilization condition of 115℃ for 20 min, and the remaining components are mixed and sterilized under the sterilization condition of 121℃ for 30 min. After sterilization, the fermentation medium is uniformly mixed to obtain the fermentation medium for standby.

[0159] Comparative Example 5

[0160] The comparative example 2 provides a preparation method of the Schizochytrium ferment product filtrate, which is different from the preparation method of example 2 only in the preparation of the fermentation medium in step 3 (4). The preparation of the fermentation medium in step 3 (4) of the comparative example 2 is as follows: each component is weighed according to the following mass percentage: glucose 4.5%, yeast powder 1.5%, peptone 1.0%, potassium phosphate dibasic 0.5%, manganese sulfate 0.01%, magnesium sulfate 0.01%, anhydrous sodium acetate 0.5%, ammonium citrate 0.15%, and cysteine 0.2%. The glucose is sterilized separately under the sterilization condition of 115℃ for 20 min, and the remaining components are mixed and sterilized under the sterilization condition of 121℃ for 30 min. After sterilization, the fermentation medium is uniformly mixed to obtain the fermentation medium for standby.

[0161] The Schizochytrium ferment product filtrate prepared in examples 1-2 and comparative examples 1-5 is applied to the serum to obtain serums S1-S7, respectively. The specific serum formula is shown in Table 1:

[0162] Table 1 Serum formula (% w / w)

[0163]

[0164] The preparation method of the above-mentioned serum S1-S7 is as follows: first, the B phase, the C phase, and the D phase are sequentially added to the A phase, stirred and dispersed, heated to 80°C, soaked and swollen, and stirred and homogenized for 2-3 min to be uniform, to obtain a mixed phase; second, the three raw materials of the E phase are mixed and heated to 60°C, and stirred until the solid is completely dissolved; finally, the mixed phase is stirred and cooled to below 60°C, the pre-dissolved E item is added, and stirred uniformly, to obtain the corresponding serum.

[0165] The moisturizing and repairing effects of the above-mentioned serum S1-S7 are determined, and the determination method is as follows:

[0166] 1. Subjects

[0167] The subjects are selected in combination with the facial conditions (one of the symptoms of impaired skin barrier function such as dryness, desquamation, and redness) and other health conditions of the individual, and 84 subjects are selected, 84 people complete the test, and 0 people drop out, among which 70 are females and 14 are males, with an average age of 28.7±5.6 years, meeting the subject's voluntary selection criteria.

[0168] 2. Test design

[0169] The subjects are evenly divided into 7 groups, each group of 12 people (10 women + 2 men), which are S1-S7 groups (corresponding to the use of serums S1-S7). After the subjects wash their faces, take an appropriate amount of the corresponding product and evenly apply it to the facial skin, gently massage until it is absorbed. Two times a day, for 28 days. The test dates are from June 25, 2025 to July 23, 2025.

[0170] 3. Detection

[0171] The effect evaluation is carried out by self-control before and after.

[0172] (1) Detection basis

[0173] “China Association for Industry-University-Research Cooperation Group Standard Cosmetics Anti-wrinkle, Firming, Moisturizing, Oil Control, Repairing, Nourishing, and Soothing Seven Efficacy Test Methods” T / CAB 0152-2022

[0174] (2) Instruments and equipment

[0175] Skin moisture test probe Corneometer CM825, transdermal water loss tester (CK Tewameter), facial image analyzer VISIA, and each performance index meets the requirements of the determination specification.

[0176] (3) Detection environment

[0177] The instrument test link is carried out in the environment with temperature of 21±1 °C and relative humidity of 50±10% RH, and the subjects need to adapt to the environment conditions for 30 minutes before the evaluation and test. The environmental temperature and humidity during the test are shown in FIG. 3 and FIG. 4, respectively. Figure 4

[0178] (4) Detection process

[0179] ① Cleaning: Before the test, the face needs to be cleaned with a cleaning product (Cetaphil) and dried with a non-lint and non-fluorescent paper towel; the cleaning process is repeated the first operation at each return visit.

[0180] ② The skin basic value measurement of parameters such as VISIA facial image collection, skin stratum corneum moisture content, and skin transepidermal water loss (TEWL) is sequentially carried out by the technical personnel. During the data collection process, the operator needs to make the subject in a comfortable body position to determine the skin parameters of the test area, and the test probes are completely attached to the skin and kept vertical during the test, and the data is recorded. The same test is carried out again after 4 weeks (D28), and all the values are averaged.

[0181] (5) Statistical method

[0182] SPSS software is applied to statistically analyze the data. The measurement data is expressed as: mean ± standard deviation, and normal distribution test is carried out, and the paired t test is adopted for the comparison before and after itself, otherwise the two related sample rank sum test is adopted.

[0183] 4. Test results

[0184] (1) The skin stratum corneum moisture content test results of the subjects are shown in Table 2.

[0185] Table 2 Skin stratum corneum moisture content of subjects in each group

[0186]

[0187] (2) The skin transepidermal water loss (TEWL) test results are shown in Table 3.

[0188] Table 3 Skin transepidermal water loss (TEWL) of subjects in each group

[0189]

[0190] (3) Skin redness index value test results are shown in Table 4.

[0191] Table 4 Skin redness index of subjects in each group value

[0192] ​

[0193] As shown in Tables 2-4, after using the corresponding serum for 28 days, the average skin stratum corneum moisture content of subjects in groups S1 and S2 increased by 71.07% and 77.78% respectively compared to before use (D0). This indicates that the Bifida Ferment Lysate Filtrate prepared in this invention has good moisturizing effects, and the transepidermal water loss (TEWL) and skin redness index are also significantly reduced. The values ​​decreased by at least 8.88% and 6.22% respectively, showing a significant improvement, indicating that the Bifida ferment filtrate prepared by this invention has good repair effects. Specifically, the moisturizing and repairing effects of groups S3 and S4 were significantly lower than those of groups S1 and S2, indicating that the three-stage fermentation control of this invention helps to increase the metabolic rate, promote the accumulation of target metabolites, and thus increase the content of active ingredients, thereby improving its moisturizing and repairing effects. The moisturizing and repairing effects of group S5 were significantly lower than those of group S4, indicating that the addition of specific solutions to the three-stage fermentation process of this invention can match the nutritional and metabolic needs of the cells at different stages, thereby increasing the yield and stability of active ingredients in the fermentation broth, and thus improving the moisturizing and repairing effects. The moisturizing and repairing effects of groups S6 and S7 were also reduced to varying degrees compared to group S2, indicating that the use of a specially treated yeast extract instead of yeast powder in the fermentation medium of this invention not only effectively improves nitrogen source utilization, but also, combined with licorice extract, better meets the growth needs of the cells, induces metabolite synthesis, and thus increases the content of effective active ingredients.

Claims

1. A method of preparing a Schizosaccharomyces fermentat ion product filtrate, characterized by, The method comprises the following steps: (1) strain activation: inoculating Bifidobacterium adolescentis into a first seed culture medium, and carrying out anaerobic culture to obtain an activated bacterial solution; (2) seed liquid culture: inoculating the activated bacterial solution into a second seed culture medium and carrying out continuous twice transfer and anaerobic culture to obtain a third seed liquid; (3) fermentation culture: inoculating the third seed liquid into a fermentation tank containing a fermentation culture medium, and carrying out anaerobic fermentation culture to obtain a fermentation product; wherein the anaerobic fermentation culture comprises a first time anaerobic fermentation, a second time anaerobic fermentation and a third time anaerobic fermentation; the first time anaerobic fermentation is carried out under the conditions of a temperature of 36-37℃, a pH of 6.0-6.2, a rotation speed of 100-110 rpm and a fermentation time of 8-10 h; the second time anaerobic fermentation is carried out under the conditions of a temperature of 38.5-39℃, a pH of 5.8-6.0, a rotation speed of 110-115 rpm and a fermentation time of 10-12 h; the third time anaerobic fermentation is carried out under the conditions of a temperature of 35-35.5℃, a pH of 5.8-6.0, a rotation speed of 95-100 rpm and a fermentation time of 2-4 h; (4) separation and purification: filtering and decolorizing the fermentation product to obtain the Schizosaccharomyces ferment product filtrate.

2. The method of claim 1, wherein the Schizosaccharomyces fermentat ion product filtrate is prepared by, In step (1), the Bifidobacterium adolescentis is Bifidobacterium adolescentis HH404-1, and the preservation number of the Bifidobacterium adolescentis HH404-1 is GDMCC No: 67151.

3. The method of claim 1, wherein the fermentation product filtrate of a Schizosaccharomyces is prepared by the steps of: The first seed culture medium comprises the following raw material components in percentage by mass: 1.0-1.2% of proteose peptone, 0.8-1.2% of beef extract, 0.4-0.7% of yeast powder, 1.8-2.3% of glucose, 0.1-0.3% of dipotassium hydrogen phosphate, 0.1-0.3% of triammonium citrate, 0.4-0.6% of sodium acetate, 0.01-0.04% of magnesium sulfate, 0.003-0.006% of manganese sulfate and 0.10-0.15% of Tween-80; and the second seed culture medium comprises the following raw material components in percentage by mass: 2.3-2.7% of glucose, 0.8-1.2% of proteose peptone, 0.3-0.7% of yeast powder, 0.01-0.02% of magnesium sulfate, 0.003-0.006% of manganese sulfate, 0.4-0.6% of sodium acetate and 0.13-0.16% of ammonium citrate.

4. The method of claim 1, wherein the fermentation product filtrate of a Schizosaccharomyces is prepared by the steps of: In step (3), a first solution is added during the first time anaerobic fermentation, a second solution is added during the second time anaerobic fermentation, and a third solution is added during the third time anaerobic fermentation; the first solution comprises glucose and yeast treatment product, the second solution comprises glucose, yeast treatment product and glutamine, and the third solution comprises trehalose and serine.

5. The method of claim 4, wherein the fermentation product filtrate of a Schizosaccharomyces is prepared by, In step (3), the first solution comprises the following components in mass percentage: glucose 5-10% and yeast processing product 0.5-1%; the second solution comprises the following components in mass percentage: glucose 5-10%, yeast processing product 1-2% and glutamine 0.1-0.3%; and the third solution comprises the following components in mass percentage: trehalose 0.1-0.3% and serine 0.03-0.05%.

6. The method of claim 4, wherein the fermentation product filtrate of a Schizosaccharomyces is prepared by the steps of: The yeast processing product is prepared by crushing the Saccharomyces cerevisiae under 30-40 MPa to obtain a bacterial solution, adding a compound enzyme to the bacterial solution, and enzymatically hydrolyzing at 45-50°C and pH 5.0-5.5 for 2-3 hours to obtain the yeast processing product; wherein the compound enzyme is prepared by mixing cellulase and β-glucanase at a mass ratio of 1:

2.

7. The method for preparing Bifida ferment filtrate according to claim 1, characterized in that, The fermentation medium comprises the following raw material components in mass percentage: glucose 4.0-5.0%, yeast processing product 1.5-1.8%, peptone 0.8-1.2%, dipotassium hydrogen phosphate 0.4-0.7%, glycyrrhiza extract 0.3-0.5%, manganese sulfate 0.01-0.02%, magnesium sulfate 0.01-0.03%, anhydrous sodium acetate 0.4-0.7%, ammonium citrate 0.12-0.18%, and cysteine 0.1-0.3%.

8. The Schizosaccharomyces fermentate filtrate prepared by the preparation method of any one of claims 1-7.

9. The use of the Schizosaccharomyces fermentate filtrate of claim 8 in the preparation of a cosmetic product.

10. The use according to claim 9, wherein the compound is ###0002### The cosmetic product is a cosmetic product having moisturizing and / or repairing effects. The cosmetic product is a cosmetic product having moisturizing and / or repairing effects.

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