Application of milk peptide leavening in regulating skin micro-ecological balance

Abstract

The invention discloses an application of a milk peptide fermentation product in regulating skin micro-ecological balance, and belongs to the technical field of biological fermentation application, and the milk peptide fermentation product selectively regulates and controls the growth of staphylococcus epidermidis and inhibits the growth of staphylococcus aureus and escherichia coli. The milk peptide leavening is found to have a selective bacteriostatic effect on skin microorganisms for the first time, experimental results show that the milk peptide leavening has an inhibiting effect on escherichia coli and staphylococcus aureus, but has no influence on normal growth of epidermis grape germs, and the characteristic is obviously different from that of a common broad-spectrum chemical bacteriostatic agent; the milk peptide leavening is more beneficial to maintenance of skin flora balance.

Description

Technical Field

[0001] The invention belongs to the technical field of biological fermentation application, and specifically relates to the application of milk peptide fermentation in regulating the microecological balance of skin. Background Art

[0002] Skin microecology refers to an ecosystem composed of microorganisms such as bacteria, fungi, viruses, and tissues, cells, various secretions, and microenvironments on the skin surface. These microorganisms interact with the host and the external environment to maintain the balance of skin microecology, which is essential for maintaining skin health.

[0003] The flora on the surface of the skin is usually divided into resident bacteria and transient bacteria. Resident bacteria are a group of microorganisms that settle on healthy skin. Transient bacteria refer to a type of microorganisms acquired through contact with the external environment. Under normal circumstances, the composition and relative abundance of skin microorganisms maintain a dynamic balance within a certain range, which can play a role in skin defense, repair and barrier. It can not only serve as a biological and physical defense line to directly resist the invasion of pathogens, produce chemicals to inhibit the growth of harmful bacteria, but also regulate the human immune response and participate in the establishment of the skin's inherent immunity and adaptive immune barriers, which is essential for maintaining human skin health. If the skin microbiome is unbalanced, the skin barrier function will be reduced, the resistance will be weakened, and harmful bacteria will grow, causing inflammation, itching, infection, repeated acne and other problems on the skin. Therefore, only by balancing the microbial community of the skin can the skin be kept healthy, the skin barrier capacity can be enhanced, and various skin problems can be improved from the source.

[0004] At present, bactericides such as usnic acid, madder, limonene, astragalin, etc. are usually added to skin care products to achieve the effect of killing harmful bacteria.

[0005] However, long-term use of these broad-spectrum bactericides can easily disrupt the balance of skin microecology, thereby causing various skin problems; and existing technology products only focus on the most basic research on microecological balance, such as promoting the growth of probiotics and inhibiting harmful bacteria respectively, and have not confirmed whether the effects of promoting the growth of beneficial bacteria and inhibiting harmful bacteria can be achieved respectively when beneficial bacteria and harmful bacteria grow together.

[0006] Therefore, how to effectively regulate and maintain the balance of skin microecology and verify it through experiments is a technical problem that needs to be solved urgently by technicians in this field. Summary of the invention

[0007] The purpose of this section is to summarize some aspects of embodiments of the present invention and briefly introduce some preferred embodiments. Some simplifications or omissions may be made in this section and the specification abstract and the invention title of this application to avoid blurring the purpose of this section, the specification abstract and the invention title, and such simplifications or omissions cannot be used to limit the scope of the present invention.

[0008] In view of the above problems and / or the problems existing in the prior art, the present invention is proposed.

[0009] Therefore, the purpose of the present invention is to overcome the deficiencies in the prior art and provide an application of milk peptide fermentation in regulating the balance of skin microecology.

[0010] In order to solve the above technical problems, the present invention provides the following technical solutions: an application of milk peptide fermentation product in regulating the balance of skin microecology, wherein the milk peptide fermentation product selectively regulates the growth of Staphylococcus epidermidis and inhibits the growth of Staphylococcus aureus and Escherichia coli;

[0011] The method for preparing milk peptide fermentation product comprises:

[0012] The milk powder is dissolved in deionized water, protease is added, the pH is adjusted to 7-10, and enzymatic hydrolysis is performed to obtain milk peptides;

[0013] The obtained milk peptides are mixed with glucose to obtain a fermentation substrate;

[0014] Inoculating Lactobacillus plantarum into a fermentation substrate, culturing and sterilizing after fermentation, collecting the supernatant by centrifugation, and freeze-drying to obtain a milk peptide fermentation product;

[0015] Wherein, the preservation number of the Lactobacillus plantarum is CCTCC No: M 20232264.

[0016] Another object of the present invention is to overcome the deficiencies in the prior art and provide a method for preparing a cosmetic having the function of regulating the microecological balance of the skin by using a fermented milk peptide.

[0017] As a preferred embodiment of the application of the present invention, the method for preparing the milk fermentation product comprises:

[0018] The milk powder is dissolved in deionized water, protease is added, the pH is adjusted to 7-10, and enzymatic hydrolysis is performed to obtain milk peptides;

[0019] The obtained milk peptides are mixed with glucose to obtain a fermentation substrate;

[0020] Inoculating Lactobacillus plantarum into a fermentation substrate, culturing and sterilizing after fermentation, collecting the supernatant by centrifugation, and freeze-drying to obtain a milk peptide fermentation product;

[0021] Wherein, the preservation number of the Lactobacillus plantarum is CCTCC No: M 20232264.

[0022] As a preferred embodiment of the application of the present invention, the cosmetics include moisturizing cream, toner, lotion, essence and facial mask.

[0023] As a preferred embodiment of the application of the present invention, the moisturizing cream comprises polydimethylsiloxane, squalane, isopropyl myristate, glyceryl stearate, sorbitan monostearate, glycerin, butylene glycol, disodium edetate, tocopherol, carbomer, milk peptide fermentation, phenoxyethanol, ethylhexylglycerin and water.

[0024] As a preferred solution of the application of the present invention, the percentage of the milk peptide fermentation product in the total mass of the moisturizing cream raw materials is 10-15%.

[0025] As a preferred embodiment of the application of the present invention, the cosmetic has the effects of purifying, conditioning and moisturizing.

[0026] As a preferred embodiment of the application of the present invention, the purification is to inhibit the colonization of harmful bacteria.

[0027] As a preferred embodiment of the application of the present invention, the conditioning is to maintain the balance of skin microecology.

[0028] As a preferred embodiment of the application of the present invention, the cosmetics are used for daily skin care, sensitive skin care, acne skin care, infant skin care or scalp care.

[0029] Beneficial effects of the present invention:

[0030] The present invention discovers for the first time that milk peptide fermentation products have a selective antibacterial effect on skin microorganisms. Experimental results show that milk peptide fermentation products have an inhibitory effect on Escherichia coli and Staphylococcus aureus, but have no effect on the normal growth of Staphylococcus epidermidis. This characteristic is obviously different from the common broad-spectrum chemical antibacterial agents. The use of milk peptide fermentation products is more conducive to maintaining the balance of skin flora. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the following briefly introduces the drawings required for describing the embodiments. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without creative labor. Among them:

[0032] Figure 1 The competitive antibacterial test results after 12 hours of Example 6, Comparative Example 8 and Comparative Example 11 of the present invention are shown in FIG. DETAILED DESCRIPTION

[0033] In order to make the above-mentioned objects, features and advantages of the present invention more obvious and easy to understand, the specific implementation methods of the present invention are described in detail below in conjunction with the embodiments of the specification.

[0034] In the following description, many specific details are set forth to facilitate a full understanding of the present invention, but the present invention may also be implemented in other ways different from those described herein, and those skilled in the art may make similar generalizations without violating the connotation of the present invention. Therefore, the present invention is not limited to the specific embodiments disclosed below.

[0035] Secondly, the term "one embodiment" or "embodiment" as used herein refers to a specific feature, structure, or characteristic that may be included in at least one implementation of the present invention. The term "in one embodiment" that appears in different places in this specification does not necessarily refer to the same embodiment, nor does it refer to a separate or selective embodiment that is mutually exclusive with other embodiments.

[0036] Unless otherwise specified, the chemical reagents, materials, etc. used in the following examples were obtained from commercial sources.

[0037] The Lactobacillus plantarum used to prepare the milk peptide fermentation product in the following examples was deposited in the China Center for Type Culture Collection on November 17, 2023, with a deposit number of CCTCC No: M20232264, the same as the Lactobacillus plantarum in CN118146346A.

[0038] Example 1

[0039] Method of sample antibacterial experiment:

[0040] Staphylococcus epidermidis (BNCC 102555), Escherichia coli (ATCC 8099), and Staphylococcus aureus (ATCC6538) were selected as test bacteria;

[0041] Among them, Staphylococcus epidermidis is a normal parasitic flora on human skin, which can secrete exogenous antimicrobial peptides to strengthen skin immune defense. Escherichia coli and Staphylococcus aureus are common pathogens of the skin, which usually cause papules or pustules on the skin.

[0042] (1) Preparation of bacterial suspension: The strain was cultured in a shaking incubator at 37°C until the logarithmic growth phase, then centrifuged at 3500 rpm for 3 minutes, the original culture medium was discarded and the suspension was resuspended in NB medium until the number of colonies was 10 6 CFU / mL.

[0043] (2) Turbidimetric test: Turbidimetric test for antibacterial effect

[0044] In the sample group, 100 μL of the adjusted bacterial solution and 100 μL of the prepared sample were added to the 96-well plate;

[0045] 100 μL of culture medium and 100 μL of prepared sample were added to the sample blank group.

[0046] To the negative control group, 100 μL of the bacterial solution with adjusted concentration and 100 μL of NB medium were added.

[0047] Add 200 μL of sterile NB culture medium to the negative control blank group.

[0048] The 96-well plate was placed in a 37°C constant temperature incubator for 24 hours, and then the absorbance at a wavelength of 600 nm was detected using an enzyme marker.

[0049] (3) Calculate bacterial survival rate:

[0050] Survival rate (%) = (absorbance value of sample group - absorbance value of sample blank group) / (absorbance value of negative control group - absorbance value of negative control blank group) × 100%

[0051] Example 2

[0052] Preparation method of milk peptide fermentation:

[0053] 1) Preparation of milk peptides:

[0054] Add milk powder to deionized water at a material-liquid ratio of 7:50 g / mL, add protease to deionized water at a material-liquid ratio of 1:200 g / mL, and fully dissolve in a 50°C water bath;

[0055] Milk and protease solution were mixed in a ratio of 6:1 (v / v), pH was adjusted to 8.0, and incubated in a 50°C water bath for 3.5 h;

[0056] After the enzymolysis was completed, the temperature was raised to 90° C. and the mixture was placed in a water bath for 10 minutes to inactivate the enzyme. The mixture was then cooled to room temperature and the pH was adjusted to 6.5 to obtain the milk peptide.

[0057] 2) Preparation of milk peptide fermentation:

[0058] Take 5.6g of glucose and dissolve it in 40mL of deionized water. Place the glucose solution and milk peptide in a high-pressure steam sterilizer respectively, sterilize at 115℃ for 15min, then take 10mL of the sterilized glucose solution into 60mL of milk peptide solution to obtain the fermentation substrate.

[0059] Take 20 mL of Lactobacillus plantarum (CCTCC No: M 20232264) bacterial solution in the logarithmic growth phase in a 50 mL sterile centrifuge tube, centrifuge at 5000 rpm for 5 min, remove the supernatant, resuspend the bacterial cells in 0.9% sterile saline, repeat the above centrifugation and washing steps twice, and resuspend the centrifuged bacterial cells in 20 mL 0.9% saline;

[0060] 3.5 mL of the resuspended bacterial solution was inoculated into the fermentation substrate prepared above, and fermented in a shaker at 37°C and 200 rpm for 48 h. After the fermentation, the fermentation solution was centrifuged at 10,000 rpm for 10 min, the supernatant was collected, and then filtered with a 0.22 μm filter membrane to remove the bacteria and impurities, and freeze-dried to obtain the milk peptide fermentation product.

[0061] Embodiment 3:

[0062] This example uses the milk peptide fermentation product obtained in Example 2, dissolves it in sterile NB culture medium, prepares a sample solution with a concentration of 20% (w / v), and performs antibacterial experiments after filtering and sterilizing with a 0.22 μm water filter membrane.

[0063] Embodiment 4:

[0064] This example uses the milk peptide fermentation product obtained in Example 2, dissolves it in sterile NB culture medium, prepares a sample solution with a concentration of 30% (w / v), and performs antibacterial experiments after filtering and sterilizing with a 0.22 μm water filter membrane.

[0065] Comparative Example 1:

[0066] This comparative example uses milk peptides, which differs from Example 2 only in that Lactobacillus plantarum is not used for fermentation. The prepared samples are then dissolved in a solvent sterile NB culture medium to prepare sample solutions with a concentration of 20% (w / v). After filtration and sterilization with a 0.22 μm water filter membrane, an antibacterial experiment is performed.

[0067] Comparative Example 2:

[0068] This comparative example uses the culture supernatant of Lactobacillus plantarum, and the only difference from Example 2 is that milk peptides are not used as fermentation substrates;

[0069] The fermentation substrate comprises 10 g of peptone, 5 g of beef extract, 5 g of sodium chloride, 10 g of glucose, 4 g of yeast extract powder, 2 g of dipotassium hydrogen phosphate, and 1000 mL of deionized water, with a pH of 6.8-7.0;

[0070] The obtained fermentation supernatant was then dissolved in a solvent sterile NB medium to prepare a sample solution with a concentration of 20% (w / v). After sterilization by filtering with a 0.22 μm water filter membrane, an antibacterial experiment was performed.

[0071] Comparative Example 3:

[0072] This comparative example uses direct fermentation of milk powder, and the only difference from Example 2 is that the milk powder is not enzymatically hydrolyzed. The obtained milk fermentation supernatant is then dissolved with a solvent sterile NB medium to prepare a sample solution with a concentration of 20% (w / v), and then sterilized by filtering with a 0.22 μm water filter membrane for antibacterial experiments.

[0073] Comparative Example 4:

[0074] This example uses the milk peptide fermentation product obtained in Example 2, dissolves it in sterile NB culture medium, prepares a sample solution with a concentration of 10% (w / v), and performs an antibacterial experiment after filtering and sterilizing with a 0.22 μm water filter membrane.

[0075] Comparative Example 5:

[0076] This example uses the milk peptide fermentation product obtained in Example 2, dissolves it in sterile NB culture medium, prepares a sample solution with a concentration of 40% (w / v), and performs antibacterial experiments after filtering and sterilizing with a 0.22 μm water filter membrane.

[0077] Comparative Example 6:

[0078] This comparative example uses milk peptides, and the only difference from Example 2 is that Lactobacillus rhamnosus (CICC6141) is used for fermentation. The prepared samples are then dissolved in a solvent sterile NB medium to prepare sample solutions with a concentration of 20% (w / v), and sterilized by filtering with a 0.22 μm water filter membrane before performing an antibacterial experiment.

[0079] Comparative Example 7:

[0080] This comparative example uses milk peptides, which differs from Example 2 only in that Lactobacillus plantarum (CICC24936) is used for fermentation.

[0081] The prepared samples were then dissolved in a solvent sterile NB culture medium to prepare sample solutions with a concentration of 20% (w / v). After sterilization by filtering with a 0.22 μm water filter membrane, an antibacterial experiment was performed.

[0082] Table 1

[0083]

[0084] From the test results in Table 1, it can be seen that the milk peptide fermented products prepared in Examples 3 and 4 can have a significant inhibitory effect on Escherichia coli and Staphylococcus aureus, but have no significant inhibitory effect on the skin symbiotic bacteria Staphylococcus epidermidis. The fermented product of the strain alone (Comparative Example 2) has no obvious inhibitory effect on Staphylococcus epidermidis, but has a proliferative effect on Escherichia coli. At the same time, milk peptide (Comparative Example 1) showed a significant growth-promoting effect on the three strains of bacteria.

[0085] In addition, the unenzymatically hydrolyzed milk fermentation liquid (Comparative Example 3) showed significant inhibitory effects on all three strains. The fermented products fermented by Lactobacillus rhamnosus (Comparative Example 6) and Lactobacillus plantarum (Comparative Example 7) could not significantly promote the growth of Staphylococcus epidermidis, and the inhibition of Staphylococcus aureus and Escherichia coli was not obvious. The above results show that the milk peptide fermented products of Examples 3 and 4 have selective inhibitory effects on common microorganisms and are potential active substances that can maintain the balance of skin microecology.

[0086] Example 5: Method for competitive inhibition of pathogenic bacteria experiment

[0087] Staphylococcus epidermidis (BNCC 102555) and Escherichia coli (ATCC 8099) or Staphylococcus aureus (ATCC 6538) were selected as the bacterial species combination of resident bacteria and Gram-positive or Gram-negative pathogenic bacteria, and the operation was as follows:

[0088] (1) Preparation of bacterial suspension: The strain was cultured in a shaking incubator at 37°C until the logarithmic growth phase, then centrifuged at 3500 rpm for 3 minutes, the original culture medium was discarded and the suspension was resuspended in NB medium until the number of colonies of resident bacteria was 10. 8 CFU / mL, the colony count of pathogenic bacteria is 10 7 CFU / mL.

[0089] (2) Co-culture: The co-culture medium is NB medium. For Staphylococcus epidermidis and Escherichia coli, Staphylococcus epidermidis, Escherichia coli and the sample are added to the shaking tube at a ratio of 1:1:1 and mixed evenly; the number of colonies of each strain is counted after culturing at 37°C for 0, 6 and 12 hours;

[0090] For S. epidermidis and S. aureus , Transwell insert plates with 0.4 μm permeable membranes were used to establish the co-culture system;

[0091] The sample and Staphylococcus epidermidis were added to the upper chamber at a ratio of 1:2, and the sample and Staphylococcus aureus were added to the lower chamber at a ratio of 1:2;

[0092] All cultures were incubated at 37 °C with gentle shaking to promote diffusion of metabolites between compartments. Dilutions were plated at 0, 6, and 12 h, and the number of colonies was counted.

[0093] The results are shown in Table 3; the control group was NB medium without sample incubated with Staphylococcus epidermidis and Escherichia coli or Staphylococcus aureus.

[0094] Embodiment 6:

[0095] In this example, the milk peptide fermentation product obtained in Example 2 was dissolved in sterile NB medium to prepare a sample solution with a concentration of 30% (w / v). After sterilization by filtration with a 0.22 μm water filter membrane, a competitive antibacterial experiment was performed.

[0096] Comparative Example 8:

[0097] This comparative example uses milk powder peptides, which differs from Example 2 only in that Lactobacillus plantarum is not used for fermentation;

[0098] The prepared samples were then dissolved in a solvent sterile NB culture medium to prepare sample solutions with a concentration of 30% (w / v). After sterilization by filtration with a 0.22 μm water filter membrane, competitive antibacterial experiments were performed.

[0099] Comparative Example 9:

[0100] This comparative example uses the culture supernatant of Lactobacillus plantarum, and the only difference from Example 2 is that milk powder enzymatic hydrolysate is not used as the fermentation substrate;

[0101] The fermentation substrate comprises 10 g of peptone, 5 g of beef extract, 5 g of sodium chloride, 10 g of glucose, 4 g of yeast extract powder, 2 g of dipotassium hydrogen phosphate, and 1000 mL of deionized water, with a pH of 6.8-7.0;

[0102] The obtained fermentation supernatant was then dissolved with a solvent, sterile NB culture medium, and prepared into sample solutions with a concentration of 30% (w / v). After sterilization by filtration with a 0.22 μm water filter membrane, a competitive antibacterial experiment was performed.

[0103] Comparative Example 10:

[0104] This comparative example uses direct fermentation of milk powder, and the only difference from Example 2 is that the milk powder is not enzymatically hydrolyzed. The obtained milk fermentation supernatant is then dissolved with a solvent sterile NB medium to prepare a sample solution with a concentration of 30% (w / v), and then filtered and sterilized with a 0.22 μm water filter membrane for competitive antibacterial experiments.

[0105] Comparative Example 11:

[0106] In this comparative example, sterile NB culture medium was used, and a competitive antibacterial experiment was performed after filtering with a 0.22 μm water filter membrane.

[0107] The results of competitive antibacterial experiments after 12 hours for Example 6, Comparative Example 8 and Comparative Example 11 are shown in Figure 1 .

[0108] Comparative Example 12:

[0109] This comparative example uses milk peptides, which differs from Example 2 only in that Lactobacillus rhamnosus (CICC6141) is used for fermentation, and the obtained sample is then dissolved in a solvent sterile NB medium to prepare a sample solution with a concentration of 30% (w / v). After sterilization by filtration with a 0.22 μm water filter membrane, a competitive antibacterial experiment is performed.

[0110] Comparative Example 13:

[0111] This comparative example uses milk peptides, which differs from Example 2 only in that plant lactobacillus (CICC24936) is used for fermentation; the prepared samples are then dissolved in a solvent sterile NB medium to prepare sample solutions with a concentration of 20% (w / v), and filtered and sterilized with a 0.22 μm water filter membrane before a competitive antibacterial experiment.

[0112] Table 2

[0113]

[0114]

[0115] It can be seen from the data in Table 2 that the milk peptide fermentation product of Example 6 can inhibit Staphylococcus aureus and Escherichia coli in the co-culture system, and promote the growth of skin commensal bacteria Staphylococcus epidermidis.

[0116] However, milk peptides (Comparative Example 8) showed the opposite results. The fermented products of Lactobacillus plantarum (Comparative Example 9) alone, Lactobacillus rhamnosus (Comparative Example 12) and Lactobacillus plantarum (Comparative Example 13) could not significantly promote the growth of Staphylococcus epidermidis, and could not adjust the ratio between harmful bacteria and beneficial bacteria. The unenzymatically hydrolyzed milk powder fermentation liquid (Comparative Example 10) showed an inhibitory effect on the growth of all three strains. Based on the above results, milk peptide fermentation has a selective inhibitory effect on common microorganisms on the skin, which is an efficacy feature that milk peptides and milk fermentation do not have. Unlike known broad-spectrum antibacterial agents, this makes milk peptide fermentation helpful in regulating the balance of skin flora and maintaining healthy homeostasis of the skin.

[0117] Example 7: Method for chicken embryo chorioallantoic membrane irritation test

[0118] The chicken embryo chorioallantoic membrane test can detect whether the product is irritating. The operation is as follows:

[0119] (1) Preparation of chick chorioallantoic membrane (CAM): After incubating SPF eggs to nine days old, the eggs were candled to select eggs rich in blood vessels and to check the position of the air cells in the chick embryos. The shell was gently broken with a hole punch, and the part of the eggshell corresponding to the air cell was peeled off with elbow forceps to expose the white egg membrane and drip 0.9% physiological saline to infiltrate the inner membrane. The inner membrane was carefully removed with forceps to ensure that the vascular membrane was not damaged.

[0120] (2) Test: Use tweezers to gently place the test ring on the surface of the vascular membrane, drop 200 μL of sample into the ring, and start timing at the same time;

[0121] The bleeding time (secH), coagulation time (secC) and blood vessel melting time (secL) of the membrane were observed and recorded within 0, 0.5min, 3min and 5min. Five parallel experiments were performed for each sample.

[0122] Bleeding: bleeding from the blood vessels or capillaries of the CAM.

[0123] Coagulation: Denaturation of proteins inside and outside blood vessels, usually manifested as slower blood flow or thrombosis within the blood vessels, brown-black color of the blood vessels, and turbidity and opacity outside the blood vessels.

[0124] Vascular melting: The walls of small blood vessels on the CAM membrane rupture and the blood vessels melt and disappear.

[0125] (3) Calculation and evaluation of results: The results are expressed as stimulation scores (IS). IS is calculated according to the following formula, and the stimulation intensity is determined according to Table 4.

[0126]

[0127] Table 3

[0128]

[0129] Embodiment 8:

[0130] In this example, the milk peptide fermentation product obtained in Example 2 was used, dissolved in sterile deionized water, and prepared into a sample solution with a mass fraction of 10% for subsequent chicken embryo chorioallantoic membrane irritation experiments.

[0131] Comparative Example 14:

[0132] This comparative example uses milk powder peptides, and the only difference from Example 2 is that Lactobacillus plantarum is not used for fermentation. The prepared samples are then dissolved in sterile deionized water to prepare sample solutions with a mass fraction of 10% for subsequent chicken embryo chorioallantoic membrane irritation experiments.

[0133] Comparative Example 15:

[0134] This comparative example uses the culture supernatant of Lactobacillus plantarum, and the only difference from Example 2 is that milk peptide is not used as a fermentation substrate. The fermentation substrate composition is 10g of peptone, 5g of beef extract, 5g of sodium chloride, 10g of glucose, 4g of yeast extract powder, 2g of dipotassium hydrogen phosphate, and 1000mL of deionized water, with a pH of 6.8-7.0. The obtained fermentation supernatant is then dissolved with sterile deionized water to prepare a sample solution with a mass fraction of 10% for subsequent chicken embryo chorioallantoic membrane irritation experiments.

[0135] Comparative Example 16:

[0136] This comparative example uses direct fermentation of milk powder, and the only difference from Example 2 is that the milk powder is not enzymatically hydrolyzed. The obtained milk fermentation supernatant is then dissolved with sterile deionized water to prepare sample solutions with a mass fraction of 10% for subsequent chicken embryo chorioallantoic membrane irritation experiments.

[0137] Table 4

[0138]

[0139] Experimental conclusion: This milk peptide fermentation product is not irritating.

[0140] Embodiment 9:

[0141] The preparation method of the moisturizing facial cream specifically comprises the following steps:

[0142] Table 5

[0143]

[0144] 1. Place phase A and phase B in a 85°C water bath and heat until completely dissolved.

[0145] 2. Homogenize phase B with a homogenizer for 3 min (9000-10000 rpm).

[0146] 3. Slowly add phase A into the homogenized phase B and continue homogenizing for 5 minutes (9000-10000 rpm).

[0147] 4. When the temperature of the mixture drops to 45°C, add phase C, cool to room temperature with slow stirring and leave for 24 hours.

[0148] Comparative Example 17:

[0149] The method for preparing the facial cream is different from that in Example 9 in that the milk peptide fermentation product is replaced with milk peptide, and specifically comprises the following steps:

[0150] Table 6:

[0151]

[0152] 1. Place phase A and phase B in a 85°C water bath and heat until completely dissolved.

[0153] 2. Homogenize phase B with a homogenizer for 3 min (9000-10000 rpm).

[0154] 3. Slowly add phase A into the homogenized phase B and continue homogenizing for 5 minutes (9000-10000 rpm).

[0155] 4. When the temperature of the mixture drops to 45°C, add phase C, cool to room temperature with slow stirring and leave for 24 hours.

[0156] Comparative Example 18:

[0157] The method for preparing the facial cream is different from that in Example 9 in that the milk peptide fermentation product is replaced by the culture supernatant of Lactobacillus plantarum, and specifically comprises the following steps:

[0158] Table 7:

[0159]

[0160]

[0161] 1. Place phase A and phase B in a 85°C water bath and heat until completely dissolved.

[0162] 2. Homogenize phase B with a homogenizer for 3 min (9000-10000 rpm).

[0163] 3. Slowly add phase A into the homogenized phase B and continue homogenizing for 5 minutes (9000-10000 rpm).

[0164] 4. When the temperature of the mixture drops to 45°C, add phase C, cool to room temperature with slow stirring and leave for 24 hours.

[0165] Comparative Example 19:

[0166] The method for preparing the facial cream is different from that in Example 9 in that the milk peptide fermentation product is replaced by milk fermentation liquid, and specifically comprises the following steps:

[0167] Table 8:

[0168]

[0169]

[0170] 1. Place phase A and phase B in a 85°C water bath and heat until completely dissolved.

[0171] 2. Homogenize phase B with a homogenizer for 3 min (9000-10000 rpm).

[0172] 3. Slowly add phase A into the homogenized phase B and continue homogenizing for 5 minutes (9000-10000 rpm).

[0173] 4. When the temperature of the mixture drops to 45°C, add phase C, cool to room temperature with slow stirring and leave for 24 hours.

[0174] Comparative Example 20:

[0175] The preparation method of the facial cream is different from that of Example 9 in that the milk peptide fermentation product is replaced with water, and specifically comprises the following steps:

[0176] Table 9:

[0177]

[0178] 1. Place phase A and phase B in a 85°C water bath and heat until completely dissolved.

[0179] 2. Homogenize phase B with a homogenizer for 3 min (9000-10000 rpm).

[0180] 3. Slowly add phase A into the homogenized phase B and continue homogenizing for 5 minutes (9000-10000 rpm).

[0181] 4. When the temperature of the mixture drops to 45°C, add phase C, cool to room temperature with slow stirring and leave for 24 hours.

[0182] Example 10: Experimental method for the effect of samples on skin moisturizing

[0183] The skin moisture retention of 20 volunteers (5 in each group) was tested using a skin moisture tester. A 4cm×4cm test area was marked on the flexor side of the left and right forearms of the subjects, and each sample was applied. The application amount was 3.0mg / cm 2 , gently massage until the sample is absorbed. Measure the skin moisture content 4 hours before and after application, and calculate the skin moisture content increase rate according to the following formula.

[0184] Skin moisture content increase rate (%) = (measured value after application - basic value before application) / basic value before application × 100%.

[0185] The experiment was conducted by adding the cream prepared by Example 9 and Comparative Examples 12-15. The experimental results are shown in Table 10.

[0186] Table 10

[0187]

[0188] It can be seen from Table 10 that, compared with the comparative example, Example 9 can significantly increase the skin moisture content of the test subjects.

[0189] In summary, the milk peptide fermentation product of the present invention can regulate the balance of skin microbiota, has a strong moisturizing effect, is safe and non-irritating, and can alleviate skin problems.

[0190] It should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention rather than to limit it. Although the present invention has been described in detail with reference to the preferred embodiments, those skilled in the art should understand that the technical solutions of the present invention can be modified or replaced by equivalents without departing from the spirit and scope of the technical solutions of the present invention, which should all be included in the scope of the present invention.

Claims

1. An application of milk peptide fermentation product in regulating skin microecological balance, characterized in that: The milk peptide fermentation product selectively regulates the growth of Staphylococcus epidermidis and inhibits the growth of Staphylococcus aureus and Escherichia coli; The method for preparing milk peptide fermentation product comprises: The milk powder is dissolved in deionized water, protease is added, the pH is adjusted to 7-10, and enzymatic hydrolysis is performed to obtain milk peptides; The obtained milk peptides are mixed with glucose to obtain a fermentation substrate; Inoculating Lactobacillus plantarum into a fermentation substrate, culturing and sterilizing after fermentation, collecting the supernatant by centrifugation, and freeze-drying to obtain a milk peptide fermentation product; Wherein, the preservation number of the Lactobacillus plantarum is CCTCC No: M 20232264.

2. Application of milk peptide fermentation product in the preparation of cosmetics that can regulate skin microecological balance.

3. The use according to claim 2, characterized in that: The preparation method of the milk peptide fermentation product comprises: The milk powder is dissolved in deionized water, protease is added, the pH is adjusted to 7-10, and enzymatic hydrolysis is performed to obtain milk peptides; The obtained milk peptides are mixed with glucose to obtain a fermentation substrate; Inoculating Lactobacillus plantarum into a fermentation substrate, culturing and sterilizing after fermentation, collecting the supernatant by centrifugation, and freeze-drying to obtain a milk peptide fermentation product; Wherein, the preservation number of the Lactobacillus plantarum is CCTCC No: M 20232264.

4. The use according to claim 2 or 3, characterized in that: The cosmetics include moisturizing cream, toner, lotion, essence, and facial mask.

5. The use according to claim 4, characterized in that: The moisturizing cream includes dimethicone, squalane, isopropyl myristate, glyceryl stearate, sorbitan monostearate, glycerin, butylene glycol, disodium EDTA, tocopherol, carbomer, milk peptide ferment, phenoxyethanol, ethylhexylglycerin and water.

6. The use according to claim 5, characterized in that: The percentage of the milk peptide fermentation product in the total mass of the moisturizing cream raw materials is 10-15%.

7. The use according to claim 2, characterized in that: The cosmetic has the effects of purifying, conditioning and moisturizing.

8. The use according to claim 7, characterized in that: The purification is to inhibit the colonization of harmful bacteria.

9. The use according to claim 7, characterized in that: The conditioning is to maintain the balance of skin microecology.

10. The use according to claim 2, characterized in that: The cosmetics are used for daily skin care, sensitive skin care, acne skin care, infant skin care or scalp care.

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